WO2017021959A1

WO2017021959A1 - Indole alkaloids to promote root colonization by beneficial bacteria

Info

Publication number: WO2017021959A1
Application number: PCT/IL2016/050841
Authority: WO
Inventors: Ilana Kolodkin-Gal; Asaph Aharoni
Original assignee: Yeda Research and Development Co Ltd
Current assignee: Yeda Research and Development Co Ltd
Priority date: 2015-08-04
Filing date: 2016-08-02
Publication date: 2017-02-09
Anticipated expiration: 2018-02-04

Abstract

The present invention relates to the use of canthin-6-one based compounds of Formula I and Formula II for inducing, enhancing and maintaining beneficial rhizobacteria biofilm formation on a plant or plant part thereof. Methods of use for stimulating matrix production from the beneficial rhizobacteria and enhanced adhesion of the rhizobacteria to a plant or plant part thereof are also described.

Description

INDOLE ALKALOIDS TO PROMOTE ROOT COLONIZATION BY BENEFICIAL

BACTERIA

FIELD OF INVENTION

[001] Use of canthin-6-one based compounds for inducing, enhancing and maintaining beneficial rhizobacteria biofilm formation on a plant or plant part is disclosed. Methods of use for stimulating matrix production from the beneficial rhizobacteria and enhanced adhesion of the rhizobacteria to a plant or plant part are also disclosed.

BACKGROUND OF THE INVENTION

[002] Despite the widely held view of bacteria as primitive, unicellular organisms that struggle for individual survival in nature, bacteria establish complex communities, referred to as biofilms that are stimulated by chemical communication. A biofilm can be viewed as a differentiated community, where the inhabitant cells are held together by an extracellular matrix. While they can be formed by most, if not all pathogens, biofilms may also be formed by beneficial bacteria. A key- example of beneficial bacteria is the motile Gram-positive, catalase-positive bacteria, Bacillus subtilis (B. subtilis) cells, which is found in soil and the gastrointestinal tract of ruminants and humans. B. subtilis organize themselves into conspicuous multicellular structures that carry out specialized tasks. These orchestrated processes are tightly regulated by multiple forms of cell-cell communication.

[003] Bacterial biofilms are ubiquitous and are of high significance in agricultural, industrial, environmental, and clinical settings. In many instances, they provide beneficial effects to other organisms, as is the case of B. subtilis biofilms that form on the surface of plant roots, thereby preventing the growth of fungal pathogens. Studying the factors that regulate the formation and disassembly of biofilms on top of the root is therefore of great interest from scientific, as well as ecological, and agricultural perspectives.

[004] B. subtilis is a genetically manipulable model organism that can differentiate into a remarkably large number of distinct cell types, including motile cells, rafts of swarmer cells, genetically competent cells, matrix-producing cells, and sporulation cells. The complicated network regulating biofilm formation in B. subtilis is well studied. The main components of the B. subtilis extracellular matrix are exopolysaccharides, synthesized by the epsA-0 operon-encoded genes, and TasA, a functional amyloid, encoded in the three-gene operon yqxM/tapA-sipW-tasA. The master regulator controlling the switch to a biofilm lifestyle is the repressor SinR. In standard laboratory settings, several cues have been associated with biofilm maturation and assembly, including oxygen deprivation, nutrient deprivation, small molecule sensing and physical cues.

[005] Research has primarily focused on intrinsic triggers, resulting from increased cell density or starvation. To date, very little research has focused on an assessment of genetic pathways of biofilm maintenance over the eukaryotic host.

[006] Canthin-6-one is an indole alkaloid that is 6H-indolo[3,2,l-(fe][l ,5]naphthyridine substituted by an oxo group at position 6. So far the effect of this plant-derived secondary metabolite on biofilms of beneficial bacteria, and the Bacilli subgroup remained unknown.

[007] The use of B. subtilis as beneficial bio-control agent may answer an immediate ecological need: the environmental problems caused by the overuse of pesticides. Traditional solutions for fungal and bacterial infections have two cardinal problems: (1) the high toxicity and nonbiodegradable properties of pesticides and (2) the residues in soil, water resources and crops that affect public health. The use of beneficial bacteria such as B. subtilis as a 'protective armor' may be a cutting edge solution to the pesticides problem.

[008] However, there remains a need to induce or maintain biofilm formation while preserving its biological activity. Further, there remains a need to enhance adhesion of beneficial biofilms comprising the bacteria that form these biofilms to plant roots. The present invention addresses this need by providing methods of use of canthin-6-one based compounds for inducing, enhancing, and maintain biofilms produced by beneficial communities, for example on plant roots. The present invention further addresses this need by providing uses of canthin-6-one based compounds to enhance adhesion of bacteria to a plant surface and to stimulate matrix production by the bacteria.

SUMMARY OF THE INVENTION

[009] In one aspect, a method of this invention is for inducing biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to the beneficial rhizobacteria, wherein said canthin-6-one based compound is represented by the structure of Formula I:

wherein,

Ri, R₂, R3, R4, R5, R6, R7, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, N0₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen or its isomer, tautomer, hydrate, salt or a combination thereof, thereby inducing said biofilm formation.

[010] In one aspect, a method of this invention provides a method for enhancing biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to the beneficial rhizobacteria, wherein said canthin-6-one based compound is represented by the structure of Formula I:

(I)

wherein,

Ri, R₂, R₃, R4, R5, R6, R₇, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, N¾, COOH, NO₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen or its isomer, tautomer, hydrate, salt or a combination thereof, thereby enhancing said biofilm formation.

[Oi l] In one aspect, a method of this invention provides a method for maintaining a biofilm formation formed by a beneficial rhizobacteria, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to the beneficial rhizobacteria, wherein said canthin-6-one based compound is represented by the structure of Formula I:

(I)

wherein,

Ri, R₂, R₃, R4, R5, R6, R₇, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, N¾, COOH, NO₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen; or its isomer, tautomer, hydrate, salt or a combination thereof, thereby maintaining said biofilm formation.

[012] In another aspect, a methods of this invention comprises a canthin-6-one based compound is represented by the structure of Formula II:

(II)

or its isomer, tautomer, hydrate, salt or a combination thereof.

[013] In another aspect, a method of this invention increases pathogen resistance in the plant or the part thereof. In another aspect, pathogen resistance is increased in a root or root hair of the plant, or any combination thereof. In another aspect, pathogen resistance comprises systemic resistance in the plant. In another aspect, pathogen comprises a fungus or a bacterium. In another aspect, a bacterium comprises a Pseudomonas or a Xanthomonas. In another aspect a Pseudomonas is Pseudomonas fluroescens PFZ-89. In another aspect, said fungus comprises a Phytophthora infestanson.

[014] In another aspect, a biofilm of this invention comprises a root-associated biofilm or a root hair associated biofilm, or any combination thereof

[015] In another aspect, a composition of this invention comprises a plant extract.

[016] In another aspect, a canthin-6-one based compound is in the form of a semi- synthetic variant or a synthetic variant.

[017] In another aspect, applying a composition of this invention comprises applying the canthin-6-one based compound to the roots of the plant in combination with spores of said rhizobacteria. In another aspect, said canthin-6-one based compound is at a concentration of about 1 - 50 uM.

[018] In another aspect, application comprises applying a composition comprising the canthin- 6-one based compound to the roots of the plant prior to applying spores of said rhizobacteria to the roots of the plant. In another aspect, said canthin-6-one based compound is at a concentration of about 1-50 uM.

[019] In another aspect, application comprises applying a composition comprising the canthin- 6-one based compound to a soil portion around the plant or a part thereof. In another aspect, a part of a plant comprises a root. In another aspect, a soil portion is within about 25 mm of said plant or part thereof. In another aspect, said canthin-6-one based compound is at a concentration of about 1-50 μΜ.

[020] In another aspect, said canthin-6-one based compound is at a concentration of about 1-2 ppm.

[021] In another aspect, methods of this invention are for use with a crop plant. In another aspect, a crop plant is selected from the group comprising corn, tomato, tobacco, cannabis, potato, and cucumber plants.

[022] In another aspect, a beneficial rhizobacteria of this invention comprises a sinl matrix regulator gene pathway. In another aspect, a beneficial rhizobacteria comprises a Bacillus subtilis (B. subtilis) species.

[023] In another aspect, in methods of this invention production of an extracellular matrix by said rhizobacteria is stimulated. In another aspect, methods of this invention enhance adhesion of said rhizobacteria to said plant or plant part thereof.

[024] In another aspect, composition of this invention comprises a solution, a powder, a spray, drops, a tablet, or a paste. In another aspect, a solution is an aqueous solution. In another aspect, an aqueous solution has a pH of about 7.0.

[025] In one aspect, a method of this invention is for stimulating matrix production by a beneficial rhizobacteria, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to said beneficial rhizobacteria, wherein said canthin-6-one based compound is represented by the structure of Formula I:

(I)

wherein,

Ri, R₂, R3, R4, R5, R6, R7, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, NO₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen; or its isomer, tautomer, hydrate, salt or a combination thereof, thereby stimulating said matrix production

[026] In another aspect, stimulating matrix production, increases pathogen resistance in the plant or the part thereof. In another aspect, stimulating matrix induces, enhances, or maintains, or any combination thereof, a biofilm on a plant or part thereof.

[027] In one aspect, a method of this invention enhances adhesion of a beneficial rhizobacteria to a plant surface, said method comprising the step of applying a composition comprising a composition of a canthin-6-one based compound to said rhizobacteria in an amount effective to enhance adhesion of the beneficial rhizobacteria to said plant surface, wherein said canthin-6-one based compound is represented by the structure of Formula I:

(I)

wherein,

Ri, R₂, R3, R4, R5, R₆, R7, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, NO₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen; or its isomer, tautomer, hydrate, salt or a combination thereof. [028] In another aspect, enhancing adhesion, increases pathogen resistance in the plant or the part thereof.

[029] Other features and advantages of the present invention will become apparent from the following detailed description, examples and figures. It should be understood, however, that the detailed description and the specific examples while indicating preferred embodiments of the invention are given by way of illustration only, since various changes and modification within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

[030] The subject matter regarded as the invention is particularly pointed out and distinctly claimed in the concluding portion of the specification. The invention, however, both as to organization and method of operation, together with objects, features, and advantages thereof, may best be understood by reference to the following detailed description when read with the accompanying drawings in which:

[031] Figures 1A-1C show that canthin-6-one specifically enhances biofilm formation and transcription of matrix genes in the beneficial bacteria B. subtilis. Figure 1A presents top-down images of floating biofilms grown under static conditions in liquid MSgg medium for 3 days at 23°C with different concentrations of Canthin-6-one (not treated (NT), ΙμΜ, 5μΜ). Results are a representative of six independent experiments. Figure IB presents data showing the transcription of sinl using luciferase as a reporter and the measured time course of normalized units of luminescence (luminescence divided by OD₆oo)- Luminescence of strains harboring

that were untreated (without canthin-6-one) or in presence of 5 μΜ of Canthin-6-one are presented. Results are a representative of six independent experiments; the SD represents an average of 5 wells. Figure 1C presents growth curves of planktonic growth (single cell growth). Wild-type B. subtilis was grown with shaking in liquid MSgg medium without (untreated) or with (5 μΜ) Canthin-6-one. Results are averages of seven wells within one experiment and their standard deviations. A representative of six independent experiments is shown.

[032] Figures 2A and 2B show the results of quantification of submerged biofilm formation by Pseudomonas species in standardized media (TSB) following 24 hours of incubation in 96 wells plate. Figure 2A shows the average of three independent experiments with the human pathogen Pseudomonas aeruginosa PA01 and Figure 2B shows the average of three independent experiments with the plant pathogen Pseudomonas fluorescens PFZ-89. Biofilms were stained with crystal violet and quantified as done previously (O'Toole and Kolter, 1998)

[033] Figure 3 shows that canthin-6-one enhances adhesion of Bacillus subtilis to tomato root. Shown are the results of quantification of bacterial densities of bacteria eluted from tomato root (microtome) following 10 hours of incubation in plant media in the presence or absence of different concentrations of canthin-6-one. Shown is an average of three independent experiments.

[034] Figure 4 shows that Cantine-6-one improved the bioprotection versus Pseudomonas syringae achieved by Bacillus. Bacillus strain B. subtilis 3610 (B.S), was used. The bacterial strain was maintained on LB plates. 1000 cells were either treated or not-treated (NT) with 5 μΜ of Canthin-6-one, then added to tomato plants grown on agrophonic beads. Plants were incubated at 28 + 2 °C for 5 days P. syringae strain DC3000 pv tomato (P.S) was grown in LB at 37°C to OD₆₀o = 0.2 to 0.3 (3 *10⁸ cells per ml) and diluted to 100 cells, or 10,000 cells, respectively (w/v), and added into the plant media. Plants were further incubated at 28 ± 2 °C for 10 days and the growth of the pathogen was measured and compared with the control. Three replications were maintained for each assay. SD represents the average of three independent experiments.

[035] It will be appreciated that for simplicity and clarity of illustration, elements shown in the figures have not necessarily been drawn to scale. For example, the dimensions of some of the elements may be exaggerated relative to other elements for clarity. Further, where considered appropriate, reference numerals may be repeated among the figures to indicate corresponding or analogous elements. DETAILED DESCRIPTION OF THE PRESENT INVENTION

[036] In the following detailed description, numerous specific details are set forth in order to provide a thorough understanding of the invention. However, it will be understood by those skilled in the art that the present invention may be practiced without these specific details. In other instances, well-known methods, procedures, and components have not been described in detail so as not to obscure the present invention.

Biofilms

[037] Bacteria are able to grow adhered to almost every surface, forming architecturally complex communities called biofilms. Biofilms are communities of cells that settle and proliferate on surfaces and are covered by an exopolymer matrix. The bacteria within a biofilm are slow- growing and many are in the stationary phase of growth. In one embodiment, as used herein the term "biofilm" refers to a multicellular community or communities held together by a self-produced extracellular matrix that may comprise exopolysaccharides (EPSs), proteins, and sometimes DNA. In another embodiment, a biofilm comprises an extracellular matrix comprising exopolysaccharides. In another embodiment, a biofilm comprises an extracellular matrix comprising exopolysaccharides expressed and secreted by the bacteria.

[038] Biofilm development occurs by a series of programmed steps, which include initial attachment to a surface, formation of three-dimensional micro-colonies, and the subsequent development of a mature biofilm. The more deeply a cell is located within a biofilm (such as, the closer the cell is to the solid surface to which the biofilm is attached to, thus being more shielded and protected by the bulk of the biofilm matrix), the more metabolically inactive the cells are. The consequences of this physiologic variation and gradient create a collection of bacterial communities where there is an efficient system established whereby microorganisms have diverse functional traits. A biofilm may be made up of various and diverse non-cellular components and may include, but is not limited to carbohydrates (simple and complex), lipids, proteins (including polypeptides), and lipid complexes of sugars and proteins (lipopolysaccharides and lipoproteins). A biofilm may include an integrated community of two or more bacteria species (polymicrobic biofilms) or predominantly one specific bacterium.

[039] However, it will be understood by those skilled in the art that as biofilms age, nutrients become limiting, waste products accumulate, and it is advantageous for the biofilm-associated bacteria to return to a planktonic existence. Thus, biofilms have a finite lifetime, characterized by eventual disassembly. In certain embodiments, the lifetime of a biofilm is a season. In other embodiments, the lifetime of a biofilm is less than a season, while in an alternated embodiment, the lifetime of a biofilm is more than a season.

[040] Three major types of biofilms can occur in the soil: bacterial, fungal, and fungal-bacterial biofilms. Both bacterial and fungal biofilms are formed on abiotic surfaces, while fungi act as the biotic surface in formation of fungal-bacterial biofilms. The majority of plant-associated bacteria found on roots and in soil are forming biofilms. Therefore, use of beneficial bacterial strains for forming biofilms could be a strategy to protect plants from pathogens.

[041] In one embodiment, biofilms form and adhere to a plant or parts thereof. Biofilms may be found on the aerial surfaces of plants as well as on the vasculature, roots, and root hair surfaces. In another embodiment, a biofilm is formed by a beneficial rhizobacteria, for example a Bacillus subtilis (B. subtilis) bacteria. In another embodiment, a biofilm formed on a plant or a plant surface by an associated beneficial bacteria may confer benefits to the plant. In another embodiment, a biofilm as described herein adheres to a plant root surface. In another embodiment, a biofilm as described herein adheres to a plant root hair surface. In another embodiment, a biofilm as described herein adheres to a plant root and plant root hair surfaces. It will be understood by those skilled in the art that the term "root" may in certain embodiments, include root hairs. In another embodiment, a biofilm as described herein adheres to a surface of a plant or part thereof, increasing pathogen resistance in the plant or part thereof.

[042] In one embodiment, this invention provides methods for inducing, enhancing or maintaining biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound of Formula I to the beneficial rhizobacteria, thereby inducing said biofilm formation. In one embodiment, this invention provides methods for inducing, enhancing or maintaining biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound of Formula II to the beneficial rhizobacteria, thereby inducing said biofilm formation.

[043] In one embodiment, this invention provides methods for stimulating production of a biofilm matrix by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of canthin-6-one based compound of Formula I to the beneficial rhizobacteria, thereby inducing said biofilm formation. In one embodiment, this invention provides methods for stimulating production of a biofilm matrix by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of canthin-6-one based compound of Formula II to the beneficial rhizobacteria, thereby inducing said biofilm formation.

[044] In one embodiment, this invention provides a method of enhancing adhesion of a beneficial rhizobacteria to a plant surface or part thereof by applying a composition comprising an effective amount of a canthin-6-one based compound of Formula I to the beneficial rhizobacteria, thereby inducing said biofilm formation. In one embodiment, this invention provides a method of enhancing adhesion of a beneficial rhizobacteria to a plant surface or part thereof by applying a composition comprising an effective amount of a canthin-6-one based compound of Formula II to the beneficial rhizobacteria, thereby inducing said biofilm formation.

Biofilm-forming Bacteria

[045] Biofilm-forming bacteria may provide a beneficial service to the surface or item to which they are adhered. For example, formation, enhancement or maintenance, or any combination thereof, of a biofilm on a plant surface or part thereof by biofilm-forming bacteria may provide a resistance to plant pathogens that could otherwise harm or kill the plant. Biofilm-forming bacteria are thus termed "beneficial bacteria". Beneficial bacteria include, but are not limited to, rhizobacteria. Rhizobacteria are root-colonizing bacteria that form symbiotic relationships with many plants. Beneficial rhizobacteria may also be known as plant growth-promoting bacteria or plant growth promoting rhizobacteria (PGPR). The term "beneficial rhizobacteria", may be used interchangeably with the terms "plant growth-promoting bacteria", "plant growth promoting rhizobacteria", "PGPR", and "rhizobacteria", having all the same meanings and qualities. In some embodiments, the term "beneficial rhizobacteria" refers to spores of beneficial rhizobacteria.

[046] In one embodiment, a beneficial rhizobacteria used in methods of this invention comprises a Bacillus subtilis (B. subtilis) species. In another embodiment, a beneficial rhizobacteria is a root colonizing bacteria. In another embodiment, the Bacillus subtilis species is selected from the group comprising B. subtilis FB17, B. subtilis NCIB3610, B. subtilis NATTO, Bacillus subtilis CPA-8, B. subtilis RO-FF-1, B. subtilis JH642, B. subtilis GB03, Bacillus subtilis Strain E1R-J. Bacillus amyloliquefaciens FZB24, FZB42, FZB45 and Bacillus subtilis FZB37

[047] B. subtilis, for example, forms architecturally complex communities on semi-solid surfaces and thick pellicles at the air/liquid interface of standing cultures B. subtilis biofilms consist of long chains of cells held together by an extracellular matrix consisting of an exopolysaccharide and amyloid fibers composed of the protein TasA The exopolysaccharide is produced by enzymes encoded by the epsA-0 operon ("eps operon") and the TasA protein is encoded by the promoter- distal gene of the yqxM-sipW-tasA operon ("yqxM operon"). In another embodiment, a beneficial rhizobacteria comprises a sini matrix regulator gene pathway. In another embodiment, the Bacillus subtilis species comprises a sini matrix regulator gene pathway

[048] In one embodiment, a beneficial rhizobacteria, for example a B. subtilis used in the methods of this invention, comprises rhizobacteria spores, for example B. subtilis spores.

Canthin-6-one Based Compounds and Compositions thereof

[049] In one embodiment, this invention is directed to methods comprising applying a composition comprising an effective amount of a canthin-6-one based compound or its isomer, tautomer, hydrate, salt or combination thereof, to a beneficial rhizobacterium, wherein said canthin- 6-one based compound is represented by the structure of Formula I:

(I)

wherein,

Ri, R₂, R₃, R4, R5, R6, R7, Re are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, NO₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen

[050] In another embodiment Ri is hydrogen. In another embodiment R₂ is hydrogen. In another embodiment R₃ is hydrogen. In another embodiment R4 is hydrogen. In another embodiment R₅ is hydrogen. In another embodiment R₆ is hydrogen. In another embodiment R₇ is hydrogen. In another embodiment R₈ is hydrogen. In another embodiment Ri, R₂, R₃, R4, R₅, R6, R7 and R8 are hydrogen.

[051] As used herein, the term "alkyl" can be any linear- or branched-chain alkyl group containing up to about 30 carbons unless otherwise specified. In one embodiment, an alkyl includes C₁-C5 carbons. In another embodiment, an alkyl includes Q-C6 carbons. In another embodiment, an alkyl includes Ci-Cs carbons. In another embodiment, an alkyl includes C₁-C₁₀ carbons. In another embodiment, an alkyl is a Ci-Ci₂ carbons. In another embodiment, an alkyl is a Ci-C₂o carbons. In another embodiment, branched alkyl is an alkyl substituted by alkyl side chains of 1 to 5 carbons. In one embodiment, the alkyl group may be unsubstituted. In another embodiment, the alkyl group may be substituted by a halogen, haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, C0₂H, amino, alkylamino, dialkylamino, carboxyl, thio and/or thioalkyl.

[052] The alkyl group can be a sole substituent or it can be a component of a larger substituent, such as in an alkoxy, alkoxyalkyl, haloalkyl, arylalkyl, alkylamino, dialkylamino, alkylamido, alkylurea, etc. Preferred alkyl groups are methyl, ethyl, and propyl, and thus halomethyl, dihalomethyl, trihalomethyl, haloethyl, dihaloethyl, trihaloethyl, halopropyl, dihalopropyl, trihalopropyl, methoxy, ethoxy, propoxy, arylmethyl, arylethyl, arylpropyl, methylamino, ethylamino, propylamino, dimethylamino, diethylamino, methylamido, acetamido, propylamido, halomethylamido, haloethylamido, halopropylamido, methyl-urea, ethyl-urea, propyl-urea, etc.

[053] As used herein, the term "aryl" refers to any aromatic ring that can be either substituted or unsubstituted. The aryl group can be a sole substituent, or the aryl group can be a component of a larger substituent, such as in an arylalkyl, arylamino, arylamido, etc. Exemplary aryl groups include, without limitation, phenyl, tolyl, xylyl, furanyl, naphthyl, pyridinyl, pyrimidinyl, pyridazinyl, pyrazinyl, triazinyl, thiazolyl, oxazolyl, isooxazolyl, pyrazolyl, imidazolyl, thiophene-yl, pyrrolyl, phenylmethyl, phenylethyl, phenylamino, phenylamido, etc. Substitutions include but are not limited to: F, CI, Br, I, C₁-C5 linear or branched alkyl, C₁-C5 linear or branched haloalkyl, C₁-C5 linear or branched alkoxy, C₁-C5 linear or branched haloalkoxy, CF₃, CN, N0₂, -CH₂CN, NH₂, NH-alkyl, N(alkyl)₂, hydroxyl, -OC(0)CF₃, -OCH₂Ph, -NHCO-alkyl, COOH, -C(0)Ph, C(0)0-alkyl, C(0)H, or -C(0)NH₂.

[054] As used herein, the term "alkoxy" refers to an ether group substituted by an alkyl group as defined above. Alkoxy refers both to linear and to branched alkoxy groups. Nonlimiting examples of alkoxy groups are methoxy, ethoxy, propoxy, wo-propoxy, teri-butoxy.

[055] As used herein, the term "aminoalkyl" refers to an amine group substituted by an alkyl group as defined above. Aminoalkyl refers to monoalkylamine, dialkylamine or trialkylamine. Nonlimiting examples of aminoalkyl groups are -N(Me)₂, -NHMe, -NH₃.

[056] A "haloalkyl" group refers, in another embodiment, to an alkyl group as defined above, which is substituted by one or more halogen atoms, e.g. by F, CI, Br or I. Nonlimiting examples of haloalkyl groups are CF₃, CF₂CF₃, CH₂CF₃.

[057] A "cycloalkyl" group refers, in one embodiment, to a ring structure comprising carbon atoms as ring atoms, which may be either saturated or unsaturated, substituted or unsubstituted. In another embodiment the cycloalkyl is a 3-12 membered ring. In another embodiment the cycloalkyl is a 6 membered ring. In another embodiment the cycloalkyl is a 5-7 membered ring. In another embodiment the cycloalkyl is a 3-8 membered ring. In another embodiment, the cycloalkyl group may be unsubstituted or substituted by a halogen, alkyl, haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, C0₂H, amino, alkylamino, dialkylamino, carboxyl, thio and/or thioalkyl. In another embodiment, the cycloalkyl ring may be fused to another saturated or unsaturated cycloalkyl or heterocyclic 3-8 membered ring. In another embodiment, the cycloalkyl ring is a saturated ring. In another embodiment, the cycloalkyl ring is an unsaturated ring. Non limiting examples of a cycloalkyl group comprise cyclohexyl, cyclohexenyl, cyclopropyl, cyclopropenyl, cyclopentyl, cyclopentenyl, cyclobutyl, cyclobutenyl, cycloctyl, cycloctadienyl (COD), cycloctaene (COE) etc.

[058] A "heterocycle" group refers, in one embodiment, to a ring structure comprising in addition to carbon atoms, sulfur, oxygen, phosphate, nitrogen or any combination thereof, as part of the ring. In another embodiment the heterocycle is a 3- 12 membered ring. In another embodiment the heterocycle is a 6 membered ring. In another embodiment the heterocycle is a 5-7 membered ring. In another embodiment the heterocycle is a 3-8 membered ring. In another embodiment, the heterocycle group may be unsubstituted or substituted by a halogen, alkyl, haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, CO₂H, amino, alkylamino, dialkylamino, carboxyl, thio and/or thioalkyl. In another embodiment, the heterocycle ring may be fused to another saturated or unsaturated cycloalkyl or heterocycle 3-8 membered ring. In another embodiment, the heterocycle is a saturated ring. In another embodiment, the heterocycle ring is an unsaturated ring. Non limiting examples of a heterocyclic rings comprise pyridine, piperidine, morpholine, piperazine, thiophene, pyrrole, benzodioxole, or indole.

[059] In one embodiment, this invention is directed to methods comprising applying a composition comprising an effective amount of a canthin-6-one based compound, or its tautomer, hydrate, isomer, or salt, or combination thereof, to a beneficial rhizobacterium, wherein said canthin-6-one based compound is represented by the structure of Formula II:

[060] In another embodiment, a compound of Formula I comprises a compound of Formula II.

[061] In one embodiment, this invention provides a canthin-6-one based compound of

Formula I or its tautomer, hydrate, isomer, salt thereof or combinations thereof. In another embodiment, this invention provides an isomer of the canthin-6-one based compound of this invention. In another embodiment, this invention provides a salt of the canthin-6-one based compound of this invention. In another embodiment, this invention provides a derivative of the canthin-6-one based compound of this invention. In another embodiment, this invention provides a tautomer of the canthin-6-one based compound of this invention. In another embodiment, this invention provides a hydrate of the canthin-6-one based compound of this invention. In another embodiment, a canthin-6-one based compound comprises a compound of Formula II, or its tautomer, hydrate, isomer, or salt thereof, or a combination thereof.

[062] In another embodiment, this invention provides composition comprising a canthin-6- one based compound of Formula I, as described herein, or, in another embodiment, a combination of the derivative, isomer, salt, tautomer, hydrate, of the canthin-6-one based compound of Formula I. In another embodiment, this invention provides composition comprising a canthin-6-one based compound of Formula II, as described herein, or, in another embodiment, a combination of the derivative, isomer, salt, tautomer, hydrate, of Formula II.

[063] In one embodiment, the term "isomer" includes, but is not limited to, optical isomers and analogs, structural isomers and analogs, conformational isomers and analogs, and the like.

[064] Compounds of the present invention can also be in the form of a hydrate, which means that the compound further includes a stoichiometric or non- stoichiometric amount of water bound by non-covalent intermolecular forces.

[065] Compounds of the present invention may exist in the form of one or more of the possible tautomers and depending on the particular conditions it may be possible to separate some or all of the tautomers into individual and distinct entities. It is to be understood that all of the possible tautomers, including all additional enol and keto tautomers and/or isomers are hereby co se, are included.

[066] The invention includes "salts" of the canthin-6-one based compounds of Formula

I and Formula II of this invention, which may be produced, by reaction of a compound of this invention with an acid or base. Certain compounds, particularly those possessing acid or basic groups, can also be in the form of a salt, The salts are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, and organic acids such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxylic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, mefhanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, N-acetylcysteine and the like. Other salts are known to those of skill in the art and can readily be adapted for use in accordance with the present invention.

[067] Suitable salts of amines of compounds the compounds of this invention may be prepared from an inorganic acid or from an organic acid. In one embodiment, examples of inorganic salts of amines are bisulfates, borates, bromides, chlorides, hemisulfates, hydrobromates, hydrochlorates, 2-hydroxyethylsulfonates (hydroxyethanesulfonates), iodates, iodides, isothionates, nitrates, persulfates, phosphate, sulfates, sulfamates, sulfanilates, sulfonic acids (alkylsulfonates, arylsulfonates, halogen substituted alkylsulfonates, halogen substituted arylsulfonates), sulfonates and thiocyanates.

[068] In one embodiment, examples of organic salts of amines may be selected from aliphatic, cycloaliphatic, aromatic, araliphatic, heterocyclic, carboxylic and sulfonic classes of organic acids, examples of which are acetates, arginines, aspartates, ascorbates, adipates, anthranilates, algenates, alkane carboxylates, substituted alkane carboxylates, alginates, benzenesulfonates, benzoates, bisulfates, butyrates, bicarbonates, bitartrates, citrates, camphorates, camphorsulfonates, cyclohexylsulfamates, cyclopentanepropionates, calcium edetates, camsylates, carbonates, clavulanates, cinnamates, dicarboxylates, digluconates, dodecylsulfonates, dihydrochlorides, decanoates, enanthuates, ethanesulfonates, edetates, edisylates, estolates, esylates, fumarates, formates, fluorides, galacturonates gluconates, glutamates, glycolates, glucorate, glucoheptanoates, glycerophosphates, gluceptates, glycollylarsanilates, glutarates, glutamate, heptanoates, hexanoates, hydroxymaleates, hydroxycarboxlic acids, hexylresorcinates, hydroxybenzoates, hydroxynaphthoates, hydrofluorates, lactates, lactobionates, laurates, malates, maleates, methylenebis(beta- oxynaphthoate), malonates, mandelates, mesylates, methane sulfonates, methylbromides, methylnitrates, methylsulfonates, monopotassium maleates, mucates, monocarboxylates, naphthalenesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, napsylates, N- methylglucamines, oxalates, octanoates, oleates, pamoates, phenylacetates, picrates, phenylbenzoates, pivalates, propionates, phthalates, phenylacetate, pectinates, phenylpropionates, palmitates, pantothenates, polygalacturates, pyruvates, quinates, salicylates, succinates, stearates, sulfanilate, subacetates, tartrates, theophyllineacetates, /?-toluenesulfonates (tosylates), trifluoroacetates, terephthalates, tannates, teoclates, trihaloacetates, triethiodide, tricarboxylates, undecanoates and valerates.

[069] In one embodiment, examples of inorganic salts of carboxylic acids or hydroxyls may be selected from ammonium, alkali metals to include lithium, sodium, potassium, cesium; alkaline earth metals to include calcium, magnesium, aluminium; zinc, barium, cholines, quaternary ammoniums.

[070] In another embodiment, examples of organic salts of carboxylic acids or hydroxyl may be selected from arginine, organic amines to include aliphatic organic amines, alicyclic organic amines, aromatic organic amines, benzathines, i-butylamines, benethamines (N- benzylphenethylamine), dicyclohexylamines, dimethylamines, diethanolamines, ethanolamines, ethylenediamines, hydrabamines, imidazoles, lysines, methylamines, meglamines, N-methyl-D- glucamines, N,N'-dibenzylethylenediamines, nicotinamides, organic amines, ornithines, pyridines, picolies, piperazines, procain, tris(hydroxymethyl)methylamines, triethylamines, triethanolamines, trimethylamines, tromethamines and ureas.

[071] In one embodiment, a canthin-6-one based compound of Formula I may be isolated from a plant or a part thereof. In another embodiment, canthin-6-one based compound of Formula I may be isolated from plants selected from the group comprising: Ailanthus altissima (Simaroubaceae), Brucea antidysenterica (Simaroubaceae), Eurycoma harmandiana (Simaroubaceae), Eurycoma longifolia (Simaroubaceae), Peganum nigellastrum (Zygophyllaceae), Zanthoxylum elephantiasis (Rutaceae), Zanthoxylum chiloperone (Rutaceae), or any combination thereof. In another embodiment, the canthin-6-one based compound of Formula I may be isolated from other plants within the same families or genus from plants selected from the group comprising: Ailanthus altissima (Simaroubaceae), Brucea antidysenterica (Simaroubaceae), Eurycoma harmandiana (Simaroubaceae), Eurycoma longifolia (Simaroubaceae), Peganum nigellastrum (Zygophyllaceae), Zanthoxylum elephantiasis (Rutaceae), Zanthoxylum chiloperone (Rutaceae), or any combination thereof. In another embodiment, the canthin-6-one based compound isolated comprises a compound of Formula II, or its isomer, tautomer, hydrate, or salt, or a combination thereof. Methods for isolating canthin-6-one based compounds are well known for example silica gel chromatography of the CH₂CI₂ extract of air-dried and ground bark material of Zanthoxylum chiloperone var. angustifolium (Soriano- Agaton et al., 2005, Extraction, hemisynthesis, and synthesis of canthin-6- one analogues. Evaluation of their antifungal activities. J Nat Prod. 68(11):1581-7).

[072] In one embodiment, a based compound used in the methods described herein comprises a compound of Formula I, or its isomer, tautomer, hydrate, or salt, or a combination thereof. In another embodiment, a canthin-6-one based compound used in the methods described herein comprises a compound of Formula II, or its isomer, tautomer, hydrate, or salt, or a combination thereof.

[073] In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from Ailanthus altissima (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from Brucea antidysenterica (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from Eurycoma harmandiana (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from, Eurycoma longifolia (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from Peganum nigellastrum (Zygophyllaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is isolated from Zanthoxylum elephantiasis (Rutaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein, is an isolated product. In another embodiment, the canthin-6- one based compound of Formula I is applied as part of a composition. In another embodiment, the canthin-6-one based compound isolated comprises a compound of Formula II or its isomer, tautomer, hydrate, or salt, or a combination thereof.

[074] In one embodiment, a composition comprising canthin-6-one based compound of Formula I is made using a canthin-6-one containing plant extract. In another embodiment, a canthin-6-one based compound of Formula I is extracted from a plant or a part thereof, and used directly or comprised in a composition. In another embodiment, the canthin-6-one based compound of Formula I may be extracted from plants selected from the group comprising: Ailanthus altissima (Simaroubaceae), Brucea antidysenterica (Simaroubaceae), Eurycoma harmandiana (Simaroubaceae), Eurycoma longifolia (Simaroubaceae), Peganum nigellastrum (Zygophyllaceae), Zanthoxylum elephantiasis (Rutaceae), Zanthoxylum chiloperone (Rutaceae), or any combination thereof. In another embodiment, the canthin-6-one based compound of Formula I may be extracted from other plants within the same families or genus from plants selected from the group comprising: Ailanthus altissima (Simaroubaceae), Brucea antidysenterica (Simaroubaceae), Eurycoma harmandiana (Simaroubaceae), Eurycoma longifolia (Simaroubaceae), Peganum nigellastrum (Zygophyllaceae), Zanthoxylum elephantiasis (Rutaceae), Zanthoxylum chiloperone (Rutaceae), or any combination thereof. In another embodiment, the canthin-6-one based compound is a compound of Formula II or its isomer, tautomer, hydrate, or salt, or a combination thereof. Methods for extracting or making extracts comprising canthin-6-one based compounds are well known, for example silica gel chromatography of the CH₂CI₂ extract of air-dried and ground bark material of Zanthoxylum chiloperone var. angustifolium (Soriano-Agaton et al., 2005, ibid.).

[075] In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Ailanthus altissima (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Brucea antidysenterica (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Eurycoma harmandiana (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Eurycoma longifolia (Simaroubaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Peganum nigellastrum (Zygophyllaceae). In another embodiment, the canthin-6-one based compound of Formula I comprised in compositions described herein and applied in methods described herein is extracted from Zanthoxylum elephantiasis (Rutaceae). In another embodiment, the canthin-6-one based compound extracted comprises a compound of Formula II or its isomer, tautomer, hydrate, or salt, or a combination thereof.

[076] As used herein, the term "extract" refers in one embodiment to a preparation obtained by concentrating a solution resulting from the depletion of a plant substance by a solvent. In one embodiment a solvent is water. In another embodiment a solvent is an organic solvent. In some embodiments, an extract of this invention refers to the product of extracting a plant material for a sufficient period of time (typically 1 - 48 hours), temperature (typically 0-100°C) with a suitable solvent, typically an organic solvent, carbon dioxide, and water, or mixtures of such solvents with each other. Typical organic solvents are polyhydric alcohols (e.g. glycerol, polyethylene glycol, propylene glycol), alcohols (e.g. methanol, ethanol, butanol, propanol), hydrocarbons (e.g. hexane, benzene, pentane, toluene, xylene), halogenated hydrocarbons (e.g. chloroform, dichloromethane), ethers (e.g. diethyl ethers, tetrahydrofuran), esters (e.g. ethyl acetate, isopropyl myristate). In another embodiment, an organic extract is a supercritical carbon dioxide extract. Each possibility represents a separate embodiment of the present invention. An extract may be obtained by combinations of these solvents with water (typically at ratio of 0.5 % to 90%).

[077] In one embodiment, a period of extraction is about 1 hour. In another embodiment, a period of extraction is about 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 24 hours, 25 hours, 26 hours, 27 hours, 28 hours, 29 hours, 30 hours, 31 hours, 32 hours, 33 hours, 34 hours, 35 hours, 36 hours, 37 hours, 38 hours, 39 hours, 40 hours, 41 hours, 42 hours, 43 hours, 44 hours, 45 hours, 46 hours, 47 hours, or 48 hours. In another embodiment, a period of extraction is less than 12 hours. In another embodiment, a period of extraction is more than 12 hours. In another embodiment, a period of extraction is less than 24 hours. In another embodiment, a period of extraction is more than 24 hours. In another embodiment, a period of extraction is less than 36 hours. In another embodiment, a period of extraction is more than 36 hours. In another embodiment, a period of extraction is less than 48 hours. In another embodiment, a period of extraction is more than 48 hours.

[078] In one embodiment, a temperature range for extraction is between about 1-100°C. In another embodiment, a temperature range for extraction is between about 1-10°C. In another embodiment, a temperature range for extraction is between about 10-20°C. In another embodiment, a temperature range for extraction is between about 20-30°C. In another embodiment, a temperature range for extraction is between about 30-40X1. In another embodiment, a temperature range for extraction is between about 40-50Χ. In another embodiment, a temperature range for extraction is between about 50-60°C. In another embodiment, a temperature range for extraction is between about 60-70°C. In another embodiment, a temperature range for extraction is between about 70- 80°C. In another embodiment, a temperature range for extraction is between about 80-90°C. In another embodiment, a temperature range for extraction is between about 90-100Χ. In another embodiment, extraction is performed at more than one temperature.

[079] In one embodiment, an extract of this invention comprises a powdered plant material, a seed oil, an essential oil or the product of steam distillation. In each case the extract is derived from the same plant or a part thereof as the plant-derived active ingredient. In one embodiment, the plant extract is a homogeneous material. In another embodiment, an extract of this invention is comprised in a composition. In another embodiment, a composition comprising an extract of this invention is used in methods described herein.

[080] In one embodiment, a plant extract is derived from a part of a plant, including but not limited to a stem, a stem bark, a trunk, a trunk bark, a twig, a tuber, a root, a root bark, a young shoot, a seed, a rhizome, a flower and other reproductive organs, a leaf and other aerial parts, or any combination thereof. In certain embodiments, a plant-derived active ingredient and an extract from the same plant are each obtained from the same part of the plant. In alternate embodiments, a plant- derived active ingredient and an extract from the same plant are each obtained from different parts of the plant. Each possibility represents a separate embodiment of the present invention.

[081] Independent of the method of making an extract, in certain embodiments, an extract of this invention is stable and its synthesis reproducible.

[082] In one embodiment, an extract may be fractionated using means well known in the art, for example column chromatography or high-throughput chromatography in order to separate activities, remove toxic elements, remove undesirable activities, or to concentrate an activity or activities, or any combination thereof. In one embodiment, an extract is a fraction of an extract. As used herein the term "fraction" refers in one embodiment to a group of components or class of structurally similar components having defined parameters such as solubility, molecular weight range, polarity range, adsorption coefficients, binding characteristics, chemical reactivity or selective solubility. In some embodiments, a fractions will be the product of chromatographic separation techniques, i.e., flash chromatography, preparative high performance liquid chromatography (HPLC), preparative gas chromatography, preparative thin layer chromatography, affinity chromatography, size exclusion chromatography, liquid-liquid chromatography e. g., counter-current chromatography or centripetal chromatography. Each possibility represents a separate embodiment of the present invention.

[083] In one embodiment, a canthin-6-one based compound of Formula I may be de novo synthesized or may be a synthetic analogue, or any mixtures thereof. In another embodiment of this invention, canthin-6-one based compound of Formula I may be obtained by synthetic methods, which in certain embodiments provide a flexible and low-cost method of preparation, for example using tryptamines as starting materials. The key step in the published synthesis is to access an appropriately substituted β-carboline precursor promoted by a Bischler-Napieralski cyclization. Exposure of tryptamines to succinic anhydride in CH₂CI₂ at room temperature followed by an esterification of the latter with catalytic Amberlyst H-15 in MeOH afforded methyl esters in almost quantitative yields over the two steps. Treatment of amides in the Bischler-Napieralski conditions using POCI₃ afforded cyclization to produce imines. (Soriano- Agaton et al., 2005, ibid.)

[084] In another embodiment, a canthin-6-one based compound of Formula I comprised in a composition herein and used in the methods described herein may be purchased from a chemical vendor, for example but not limited to ABI chem, Oakwood Products, 4C Pharma Scientific, Angene Chemical, AAA Chemistry, Zinc, Ambintor, eheterocycles LTD, Molport, and Chemfrog. In another embodiment, a de novo synthesized canthin-6-one based compound, or a synthetic analog thereof, or a combination thereof, comprises a compound of Formula I, or its isomer, tautomer, hydrate, or salt, or a combination thereof. In another embodiment, a de novo synthesized canthin-6-one based compound, or a synthetic analog thereof, or a combination thereof, comprises a compound of Formula II, or its isomer, tautomer, hydrate, or salt, or a combination thereof, of Formula II, or its isomer, tautomer, hydrate, or salt, or a combination thereof.

[085] In one embodiment, a canthin-6-one based compound of Formula I may be a semisynthetic variant, a synthetic variant, an isomer, a derivative, a metabolite, an analog, a derivative, a salt, a crystal, an N-oxide, a tautomer, or a hydrate, or any combination thereof. In another embodiment, the canthin-6-one based compound of Formula I, applied in methods described herein comprises a synthetic product. In another embodiment, the canthin-6-one based compound of Formula I, applied in methods described herein, comprises a semi -synthetic product. In another embodiment, the canthin-6-one based compound of Formula I, comprises an isomer, a derivative, a metabolite, an analog, a derivative, a salt, a crystal, an N-oxide, tautomer, or a hydrate, or any combination thereof

[086] In one embodiment, methods of this invention apply a canthin-6-one based compound of formula I to a beneficial rhizobacteria. In another embodiment, methods of this invention apply a compound of Formula II to a beneficial rhizobacteria. In another embodiment, methods of this invention apply a semi-synthetic variant, a synthetic variant, an isomer, a metabolite, a salt, a polymorph, a crystal, an N-oxide, a tautomer, or a hydrate, or any combination thereof of a canthrn- 6-one based compound.

[087] In one embodiment, methods of this invention apply a cnathin-6-one based compound of Formula I to a portion of soil around a plant on which a biofilm may form, is being formed or has formed. In another embodiment, methods of this invention apply a compound of Formula II or its isomer, tautomer, hydrate, or salt, or a combination thereof, to a portion of soil around a plant on which a biofilm may form, is being formed or has formed. In another embodiment, methods of this invention apply a semi- synthetic variant, a synthetic variant, an isomer, a metabolite, a salt, a polymorph, a crystal, an N-oxide, a tautomer or a hydrate, or any combination thereof of a compound of Formula I to apportion of soil around a plant on which a biofilm may be formed. In another embodiment, methods of this invention apply a semi-synthetic variant, a synthetic variant, an isomer, a metabolite, a salt, a polymorph, a crystal, an N-oxide, a tautomer or a hydrate, or any combination thereof of a compound of Formula II to apportion of soil around a plant on which a biofilm may be formed.

[088] In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of a dry powder. In another embodiment, a composition comprising a compound of Formula II is applied in the form of a dry powder. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of a solution. In another embodiment, a composition comprising a compound of Formula II is applied in the form of a solution. In another embodiment, a solution comprises an aqueous solution. In another embodiment, an aqueous solution comprises a PBS (Phosphate buffered Saline) solution. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I, comprises an aqueous solution at pH 7. In another embodiment, a composition comprising a canthin-6-one based compound of Formula II, comprises an aqueous solution at pH 7. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of a spray. In another embodiment, a composition comprising a compound of Formula II is applied in the form of a spray. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of a tablet. In another embodiment, a compound of Formula II is applied in the form of a tablet. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of drops. In another embodiment, a composition comprising a compound of Formula II is applied in the form of drops. In another embodiment, a composition comprising a canthin-6-one based compound of Formula I is applied in the form of a paste. In another embodiment, a composition comprising a compound of Formula II is applied in the form of a paste.

[089] In one embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 ppm - 5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.01 ppm - 2.5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 ppm - 2.0 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 ppm - 2.0 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 1 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 2 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 3 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 4 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 ppm.

[090] In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.01 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 mM-50 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 mM-40 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 mM-30 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 mM- 50 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 20 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 25 mM. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 30 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 35 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 40 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 45 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 50 mM.

[091] In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.0001 uM-1000 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 uM- 1000 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.01 μΜ-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 uM-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 μΜ-10 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 μΜ-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 uM-50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 μΜ-75 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 μΜ-50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 25 μΜ-50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 uM-40 μΜ. In another embodiment, a composition of this invention comprises a canthin-6- one based compound aqueous solution at about 10 uM-30 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 μΜ- 50 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 uM- 50 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 20 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 25 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 30 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 35 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 40 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 45 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 60 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 70 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 80 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 90 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 200 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 250 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 500 uM.

[092] In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.0001 nM-1000 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.001 nM- 1000 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.01 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 0.1 nM-10 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1.0 nM-50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 nM-75 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 25 nM-50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 nM-40 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 nM-30 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 nM- 50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 1 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 5 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 10 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 20 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 25 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 30 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 35 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 40 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 45 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 60 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 70 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 80 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 90 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 200 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 250 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound aqueous solution at about 500 nM.

[093] In one embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 ppm - 5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.01 ppm - 2.5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 ppm - 2.0 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 ppm - 2.0 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.5 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 1 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 2 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 3 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 4 ppm. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 ppm.

[094] In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.01 mM- 100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 mM-100 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 mM-50 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 mM-40 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 mM-30 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 mM- 50 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 20 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 25 mM. In another embodiment, a composition of this invention comprises a canthin-6-one aqueous solution at about 30 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 35 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 40 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 45 mM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 50 mM.

[095] In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.0001 μΜ-1000 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 uM- 1000 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.01 uM-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 uM-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 uM-10 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 uM-100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 μΜ-50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1 μΜ-75 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 25 uM-50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 μΜ-40 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 uM-30 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 μΜ- 50 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 20 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 25 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 30 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 35 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 40 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 45 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 50 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 60 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 70 uM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 80 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 90 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 100 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 200 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 250 μΜ. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 500 μΜ.

[096] In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.0001 nM-1000 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.001 nM- 1000 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.01 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 0.1 nM-10 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 nM-100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1.0 nM-50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1 nM-75 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 25 nM-50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 nM-40 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 nM-30 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 nM- 50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 1 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 5 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 10 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 20 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 25 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 30 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 35 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 40 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 45 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 50 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 60 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 70 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 80 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 90 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 100 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 200 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 250 nM. In another embodiment, a composition of this invention comprises a canthin-6-one based compound at a concentration of about 500 nM.

Applications/Formulations

[097] In one embodiment, application of a composition of this invention comprises applying a composition comprising nanomolar (nM) quantities a canthin-6-one based compound described herein to the roots of a plant, in combination with Bacillus spores. In an alternative embodiment, application of a composition of this invention comprises applying micromolar (μΜ) quantities of a canthin-6-one based compound described herein to the roots of a plant, prior to application of Bacillus spores. In yet a further embodiment, application of a composition of this invention comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound described herein, to a portion of soil surrounding a plant. In another embodiment, application to the portion of soil, promotes natural colonization by Bacillus species.

[098] In another embodiment, application comprises applying a composition comprising nanomolar (nM) quantities of a canthin-6-one based compound of Formula I to the roots of a plant in combination with spores of a beneficial rhizobacteria. In another embodiment, application comprises applying a composition comprising nanomolar (nM) quantities of a canthin-6-one based compound of Formula II to the roots of a plant in combination with spores of a beneficial rhizobacteria. In another embodiment, application comprises applying a composition comprising nanomolar (nM) quantities of a canthin-6-one based compound of Formula I to the roots of a plant in combination with spores of a Bacillus species. In another embodiment, application comprises applying a composition comprising nanomolar (nM) quantities of a canthin-6-one based compound of Formula II to the roots of a plant in combination with spores of a Bacillus species.

[099] In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula I to the roots of a plant prior to application of spores of a beneficial rhizobacteria. In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula II to the roots of a plant prior to application of spores of a beneficial rhizobacteria. In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula I to the roots of a plant prior to application of spores of a Bacillus species. In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula II to the roots of a plant prior to application of spores of a Bacillus species.

[0100] In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula I to an area of soil around a plant. In another embodiment, application comprises applying a composition comprising micromolar (uM) quantities of a canthin-6-one based compound of Formula II to an area of soil around a plant. In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula I to an area of soil around a plant root. In another embodiment, application comprises applying a composition comprising micromolar (μΜ) quantities of a canthin-6-one based compound of Formula II to an area of soil around a plant root. In another embodiment, application to the portion of soil, promotes natural colonization by Bacillus species. It will be understood by those skilled in the art that a composition applied to an area of soil may seep into the soil over time.

[0101] In one embodiment, application is for about 1-24 hours. In another embodiment, application is for about 1-12 hours. In another embodiment, application is for about 12-24 hours. In another embodiment, application is for about 5-10 hours. In another embodiment, application is for about 1 hour. In another embodiment, application is for about 2 hours. In another embodiment, application is for about 3 hours. In another embodiment, application is for about 4 hours. In another embodiment, application is for about 5 hours. In another embodiment, application is for about 6 hours. In another embodiment, application is for about 7 hours. In another embodiment, application is for about 8 hours. In another embodiment, application is for about 9 hours. In another embodiment, application is for about 10 hours. In another embodiment, application is for about 11 hours. In another embodiment, application is for about 12 hours. In another embodiment, application is for about 20 hours. In another embodiment, application is for about 24 hours. In another embodiment, application is for less than 1 hour. In another embodiment, application is for more than 24 hours.

[0102] Compositions containing canthin-6-one based compounds, for example compounds of Formula I, Formula II or their isomer, tautomer, hydrate, or salt, or a combination thereof, may be used to induce, enhance, and enhance biofilm formation on a surface of a plant or a part thereof. Such a surface can be any surface that may be prone to biofilm formation and adhesion of bacteria. Non-limiting examples of surfaces include aerial plant parts, plant vasculature, plant roots, and plant root hairs. In another embodiment, a surface of a plant is a plant root surface. In another embodiment, a surface of a plant is a plant root hair surface. In another embodiment, a surface of a plant is a vasculature surface. In another embodiment, a surface of a plant is an aerial surface, for example a leaf, a flower, or a stem. In another embodiment, a surface of a plant is below ground.

[0103] It will be understood by those skilled in the art that the term "surface" as used herein refers in one embodiment to the outside part or layer of a plant or a part thereof. In one embodiment, a biofilm described herein adheres to a plant surface, for example a root surface. In another embodiment, a biofilm described herein adheres to 100% of a surface, for example adheres to the complete root surface. In another embodiment, a biofilm adheres to less than 100% of a surface. In another embodiment, a biofilm adheres to less than 90% of a surface. In another embodiment, a biofilm adheres to less than 80% of a surface. In another embodiment, a biofilm adheres to less than 70% of a surface. In another embodiment, a biofilm adheres to less than 60% of a surface. In another embodiment, a biofilm adheres to less than 50% of a surface. In another embodiment, a biofilm adheres to less than 40% of a surface. In another embodiment, a biofilm adheres to less than 30% of a surface. In another embodiment, a biofilm adheres to less than 20% of a surface. In another embodiment, a biofilm adheres to less than 10% of a surface.

[0104] In one embodiment, this invention provides a method for inducing biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound described herein to the beneficial rhizobacteria or spores thereof, thereby inducing said biofilm formation. In another embodiment, the beneficial rhizobacteria or spores thereof are adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria or spores thereof are present in soil around the plant.

[0105] Importantly bacterial spores are frequently spread in natural settings. In another embodiment, beneficial rhizobacteria are present in the soil around a plant at about a distance of about up to several mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 100 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 50 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 40 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 30 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 20 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about 10 -100 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 10 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about 0 - 10 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about 0-5 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 5 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria are present in the soil around a plant or a plant root at about a distance of about up to 1 mm from a plant or a plant root.

[0106] In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant at about a distance of about up to several mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 10 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 100 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 50 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 25 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 5 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 1-2 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 0-2 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria spores are present in the soil around a plant or a plant root at about a distance of about up to 0-1 mms from a plant or a plant root. In another embodiment, beneficial rhizobacteria, or spores thereof, or any combination thereof, are present in the soil around a plant root at about a distance of about up to several mms from a plant. In another embodiment, beneficial rhizobacteria, or spores thereof, or any combination thereof, are present in the soil around a plant root at about a distance of about up to 10 mms from a plant.

[0107] In yet another embodiment, the beneficial rhizobacteria is adhered to the plant or part thereof, and is present in soil around the plant or a part thereof, for example around the root of the plant. In another embodiment, the beneficial rhizobacteria is not adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria is not present in soil around the plant. In another embodiment, the beneficial rhizobacteria is cultured in vitro, wherein said composition comprising a canthin-6-one based compound of this invention is applied to the bacteria in vitro. In another embodiment, a method for inducing biofilm formation on a plant or a part thereof, further comprises the step of contacting said plant or part thereof with said beneficial rhizobacteria previously contacted with a composition presented herein. In another embodiment, a method for inducing biofilm formation on a plant or a part thereof, further comprises the step of contacting a soil portion around said plant with said beneficial rhizobacteria previously contacted with a composition presented herein

[0108] As used herein, the term "inducing" refers in one embodiment to causing biofilm formation. In another embodiment, a method of inducing biofilm formation comprises a beneficial rhizobacteria colonizing a plant root and forming a biofilm. In another embodiment, inducing biofilm formation comprises beneficial rhizobacteria first colonizing a plant root and then forming a biofilm. In another embodiment, inducing biofilm formation comprises beneficial rhizobacteria simultaneously colonizing a plant root and forming a biofilm. In another embodiment, a method of inducing a biofilm comprises formation of an extracellular matrix that holds cells together. In another embodiment, the extracellular matrix comprises polysaccharide biopolymers. In another embodiment, a method of inducing biofilm formation comprises inducing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of inducing biofilm formation comprises inducing secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of inducing biofilm formation comprises inducing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of inducing biofilm formation comprises inducing secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of inducing biofilm formation comprises increasing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of inducing biofilm formation comprises increasing secretion by the rhizobacteria of a protein or proteins, for example TasA. In another embodiment, a method of inducing biofilm formation occurs where no biofilm is currently present. In another embodiment, a method of inducing biofilm formation occurs at the site of an already established biofilm. In another embodiment, a method of inducing biofilm formation decreases repression of matrix encoding operons, for example epsA-0 operon and yqxMltapA-sipW-tasA operon. In another embodiment, a method of inducing biofilm formation comprises enhancing the transcription of sinl. In another embodiment, induction of biofilm formation provides a beneficial effect to the plant. In another embodiment, induction of biofilm formation promotes plant growth.

[0109] In another embodiment, a method for inducing biofilm formation on a plant or a part thereof comprises the step of applying an effective amount of a composition comprising a canthin- 6-one based compound of this invention, to a B. subtilis species. In another embodiment, a method for enhancing biofilm formation on a plant or a part thereof, comprises the step of applying a composition comprising an effective amount of a canthin-6-one based compound of this invention, to a B. subtilis species. In another embodiment, a method for maintaining a biofikn formation on a plant or a part thereof comprises the step of applying a composition comprising an effective amount of a canthin-6-one based compound of this invention, to a B. subtilis species. In another embodiment, the B. subtilis species is B. subtilis FB17. In another embodiment, a B. subtilis species is B. subtilis NCIB3610. In yet another embodiment, a B. subtilis species is B. subtilis RO-FF-1. In still another embodiment, a B. subtilis species is B. subtilis JH642.

[0110] In one embodiment, this invention provides a method for enhancing biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound of this invention, to the beneficial rhizobacteria, thereby enhancing biofilm formation. In another embodiment, the beneficial rhizobacteria is adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria is present in soil around the plant. In yet another embodiment, the beneficial rhizobacteria is adhered to the plant or part thereof, and is present in soil around the plant. In another embodiment, the beneficial rhizobacteria is not adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria is not present in soil around the plant. In another embodiment, the beneficial rhizobacteria is cultured in vitro, wherein said composition comprising a canthin-6-one based compound of this invention, is applied to the bacteria in vitro. In another embodiment, a method for enhancing biofilm formation on a plant or a part thereof, further comprises the step of contacting said plant or part thereof with said beneficial rhizobacteria previously contacted with a composition described herein. In another embodiment, a method for enhancing biofilm formation on a plant or a part thereof, further comprises the step of contacting a portion of soil around said plant or part thereof with said beneficial rhizobacteria previously contacted with a composition described herein.

[0111] As used herein, the term "enhancing" refers in one embodiment to increasing formation of a biofilm or improving formation of a biofilm. In another embodiment, a method of enhancing biofilm formation comprises increasing the rate of biofilm formation. In another embodiment, a method of enhancing biofilm formation comprises improving the colonization of a beneficial rhizobacteria associated with a plant, for example with a plant root, a plant root hair, or any combination thereof. In another embodiment, a method of enhancing biofilm formation comprises improving the biofilm formation, for example by improving adhesion of a beneficial rhizobacteria with a plant, for example a plant root, root hair or any combination thereof. In another embodiment, a method of enhancing biofilm formation comprises improving the biofilm formation, for example altering the matrix components. In another embodiment, a method of enhancing biofilm formation comprises improving the properties of a biofilm, for example improved resistance to infection, or improved resistance to pathogens, or any combination thereof.

[0112] In another embodiment, a method of enhancing biofilm formation comprises enhancing beneficial rhizobacteria colonizing a plant root. In another embodiment, enhancing biofilm formation comprises enhancing beneficial rhizobacteria colonizing a plant root and then forming a biofilm. In another embodiment, enhancing biofilm formation comprises enhancing simultaneously beneficial rhizobacteria colonizing a plant root and formation of a biofilm. In another embodiment, a method of enhancing a biofilm comprises enhancing formation of an extracellular matrix that holds cells together. In another embodiment, the extracellular matrix comprises polysaccharide biopolymers. In another embodiment, a method of enhancing biofilm formation comprises inducing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of enhancing biofilm formation comprises inducing secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of enhancing biofilm formation comprises enhancing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of enhancing biofilm formation comprises enhancing secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of enhancing biofilm formation comprises increasing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of enhancing biofilm formation comprises increasing secretion by the rhizobacteria of a protein or proteins, for example TasA. In another embodiment, a method of enhancing biofilm formation occurs where no biofilm is currently present. In another embodiment, a method of enhancing biofilm formation occurs at the site of an already established biofilm. In another embodiment, enhancing biofilm formation provides a beneficial effect to the plant.

[0113] In one embodiment, this invention provides a method for maintaining a biofilm formation on a plant or a part thereof comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound of this invention, to a beneficial rhizobacteria. In another embodiment, the beneficial rhizobacteria is adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria is present in soil around the plant. In yet another embodiment, the beneficial rhizobacteria is adhered to the plant or part thereof, and is present in soil around the plant. In another embodiment, the beneficial rhizobacteria is not adhered to the plant or the part thereof. In another embodiment, the beneficial rhizobacteria is not present in soil around the plant. In another embodiment, the beneficial rhizobacteria is cultured in vitro, wherein said composition comprising an effective amount of canthin-6-one is applied to the bacteria in vitro. In another embodiment, a method for maintaining a biofilm formation on a plant or a part thereof further comprises the step of administering said beneficial rhizobacteria to said plant or part thereof, following applying a composition described herein comprising canthin-6-one to said rhizobacteria. In another embodiment, a method for maintaining a biofilm formation on a plant or a part thereof, further comprises the step of administering said beneficial rhizobacteria to soil around said plant or part thereof, following applying a composition described herein comprising a canthin-6-one based compound of this invention, to said rhizobacteria.

[0114] As used herein, the term "maintaining" refers in one embodiment to keeping a biofilm in existence. In another embodiment, a method of maintaining a biofilm comprises keeping the biofilm in good condition. In another embodiment, method of maintaining a biofilm comprises repairing problems in a biofilm. In another embodiment, a method of maintaining a biofilm comprises keeping a biofilm "as is" without change. In another embodiment, a method of maintaining a biofilm comprises providing nutrients to said biofilm. In another embodiment, a method of maintaining a biofilm comprises keeping beneficial rhizobacteria communities. In another embodiment, a method of maintaining a biofilm comprises attracting beneficial rhizobacteria communities to said plant or part thereof. In another embodiment, method of maintaining a biofilm comprises continued secretion of polysaccharide biopolymers from rhizobacteria. In another embodiment, a method of maintaining biofilm formation comprises inducing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of maintaining biofilm formation comprises inducing secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of maintaining biofilm formation comprises maintaining secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of maintaining biofilm formation comprises maintaining secretion by the rhizobacteria of a protein or proteins, for example TasA In another embodiment, a method of maintaining biofilm formation comprises increasing secretion by the rhizobacteria of polymers, for example polysaccharides EPS, or poly-5-glutamate (PGA), or any combination thereof. In another embodiment, a method of maintaining biofilm formation comprises increasing secretion by the rhizobacteria of a protein or proteins, for example TasA. In another embodiment, a method of maintaining a biofilm formation comprises reforming all or part of a biofilm. In another embodiment, a method of maintaining a biofilm comprises providing nutrients to the biofilm. In another embodiment, a method of maintaining a biofilm comprises removing waste products from a biofilm. [0115] In another embodiment, a method of maintaining biofilm formation comprises maintaining beneficial rhizobacteria colonizing a plant root. In another embodiment, maintaining biofilm formation comprises maintaining beneficial rhizobacteria colonizing a plant root and then forming a biofilm. In another embodiment, maintaining biofilm formation comprises maintaining simultaneously beneficial rhizobacteria colonizing a plant root and formation of a biofilm. In another embodiment, a method of maintaining a biofilm comprises maintaining formation of an extracellular matrix that holds cells together. In another embodiment, the extracellular matrix comprises polysaccharide biopolymers. In another embodiment, a method of maintaining biofilm formation comprises maintaining secretion by the rhizobacteria of at least a protein, for example TasA. In another embodiment, a method of maintaining biofilm formation occurs where no biofilm is currently present. In another embodiment, a method of maintaining biofilm formation occurs at the site of an already established biofilm. In another embodiment, maintaining biofilm formation provides a beneficial effect to the plant.

[0116] In one embodiment, this invention provides a method for stimulating matrix production in a beneficial rhizobacteria for forming or maintaining a biofilm on a plant or a part thereof comprising the step of applying a canthin-6-one based compound of this invention, to said beneficial rhizobacteria.

[0117] As used herein, the term "matrix" may be used interchangeably with the term "biofilm extracellular polymeric substances (EPS)" or "extracellular polymeric substances (EPS)" or "EPS matrix", "biofilm matrix" or "EPS", having all the same qualities and meanings. The EPS matrix may in one embodiment, determine the immediate conditions of life of beneficial rhizobacteria cells living in the biofilm microenvironment by affecting porosity, density, water content, charge, sorption properties, hydrophobicity, and mechanical stability. EPS are biopolymers of microbial origin, for example from the beneficial rhizobacteria, in which these biofilm rhizobacteria are embedded. In another embodiment, an EPS matrix comprises the milieu surrounding the rhizobacteria in a biofilm. Production of exopolysacchrides (EPS) or other exopolymeric material, and consequently the formation of biofilms per se, was shown to enhance bacterial survival and the potential for colonization of roots

[0118] In another embodiment, a beneficial rhizobacteria used in the methods of this invention, produces the biopolymers. In another embodiment, the EPS comprises polysaccharides. In another embodiment, the EPS comprises polysaccharides, a wide variety of proteins, glycoproteins, and glycolipids, and in certain embodiment, extracellular DNA (e-DNA). In another embodiment, EPS biopolymers are highly hydrated. These hydrated EPS biopolymers form the matrix, which keeps the biofilm cells together and retains water. In another embodiment, a matrix interacts with the environment, for example, by attaching biofilms to surfaces. In another embodiment, a matrix interacts with a plant by attaching a biofilm to a surface of the plant or a part thereof.

[0119] As used herein, the term "attaching" or any grammatical form thereof, is interchangeable with the term "adhering" or any grammatical form thereof, having all the same meanings and qualities. In another embodiment, the term "attaching" refers to an association between a biofilm and a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising an indole alkaloid enhance adhesion of rhizobacteria to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a canthin-6-one based compound of this invention, enhance adhesion of rhizobacteria to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a compound of Formula I or its isomer, tautomer, hydrate, salt, or combination thereof, enhance adhesion of B. subtilis to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a compound of Formula II or its isomer, tautomer, hydrate, salt, or combination thereof, enhance adhesion of B. subtilis to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a canthin-6-one based compound enhance adhesion of B. subtilis to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a canthin-6-one based compound enhance adhesion of a biofilm as described herein to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a compound of Formula I or its isomer, tautomer, hydrate, salt, or combination thereof, enhance adhesion of a biofilm as described herein to a surface of a plant or a part thereof. In another embodiment, compositions described herein comprising a compound of Formula II or its isomer, tautomer, hydrate, salt, or combination thereof, enhance adhesion of a biofilm as described herein to a surface of a plant or a part thereof.

[0120] In one embodiment, EPS polymers of a biofilm matrix comprise those polysaccharides whose synthesis is dependent on expression from the epsA-0 operon of B. subtilis. In another embodiment, EPS polymers of a matrix comprise glucose, galactose, poly-N-acetyl glucosamine (PNAG), or levan polysaccharides, or any combination thereof.

[0121] In one embodiment, proteins of a biofilm matrix comprise those proteins expressed from the yqxM/tapA-sipW-tasA operon of B. subtilis. In another embodiment, a biofilm matrix comprises the amyloid proteins TasA, TapA or BslA, or any combination thereof.

[0122] In one embodiment, a method of this invention stimulating matrix product enhances the transcription of sinl in a beneficial rhizobacteria. In another embodiment, a method of this invention stimulating matrix product reduces transcriptional repression of an epsA-0 operon or a yqxM-tapA- sipW-tasA operon, or any combination thereof, in a beneficial rhizobacteria. [0123] In one embodiment, this invention provides a method for enhancing adhesion of a beneficial rhizobacteria to a surface of a plant or a part thereof, the method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound of this invention to said beneficial rhizobacteria, or to said plant or part thereof, or to a soil portion near said plant or part thereof, or any combination thereof, in an amount effective to enhance adhesion of said beneficial rhizobacteria to said plant surface or part thereof.

[0124] In another embodiment, this invention provides a method for enhancing adhesion of a beneficial rhizobacteria to a surface of a plant or a part thereof, the method comprising the step of applying a composition comprising a compound of Formula I or its isomer, tautomer, hydrate, salt, or a combination thereof, to said beneficial rhizobacteria, or to said plant or part thereof, or to a soil portion near said plant or part thereof, or any combination thereof, in an amount effective to enhance adhesion of said beneficial rhizobacteria to said plant surface or part thereof In another embodiment, this invention provides a method for enhancing adhesion of a beneficial rhizobacteria to a surface of a plant or a part thereof, the method comprising the step of applying a composition comprising a compound of Formula II or its isomer, tautomer, hydrate, salt, or a combination thereof, to said beneficial rhizobacteria, or to said plant or part thereof, or to a soil portion near said plant or part thereof, or any combination thereof, in an amount effective to enhance adhesion of said beneficial rhizobacteria to said plant surface or part thereof

[0125] As used herein, the term "adhesion" refers in one embodiment to an act of sticking or attaching a beneficial rhizobacteria to a plant or a part thereof. In another embodiment, adhesion comprises sticking or attaching the biofilm in which rhizobacteria communities are living to the plant or part thereof. In another embodiment, adhesion comprises sticking or attaching the biofilm matrix within which rhizobacteria communities are living to the plant or part thereof. In another embodiment, biofilms as described herein, comprise bacterial communities in which beneficial rhizobacteria communities are embedded in a matrix of extracellular polymeric compounds attached to a surface of a plant or a part thereof.

[0126] In another embodiment, adhesion of a beneficial rhizobacteria to a plant surface is reversible. In an alternative embodiment, adhesion of a beneficial rhizobacteria to a plant surface is not reversible. In another embodiment, adhesion of a beneficial rhizobacteria to a plant surface enhances productivity of the plant. In another embodiment, adhesion of a beneficial rhizobacteria to a plant surface enhances the growth of the plant. In another embodiment, adhesion of a beneficial rhizobacteria to a plant surface enhances the growth of the plant.

[0127] In another embodiment, said plant surface or plant part surface comprises an aerial surface, a root epidermis, a surface of a root hair, or a surface of transport vessels, or any combination thereof. In another embodiment, a plant surface or a surface of a plant part wherein a rhizobacteria may adhere comprises an aerial surface, for example a leaf, a flower, or a stem, or any combination thereof. In another embodiment, a plant surface or a surface of a plant part wherein a rhizobacteria may adhere comprises a root epidermis. In another embodiment, a plant surface or a surface of a plant part wherein a rhizobacteria may adhere comprises a surface of a root hair. In another embodiment, a plant surface or a surface of a plant part wherein a rhizobacteria may adhere comprises a transport vessel. For example, colonization of rhizobacteria on a plant's roots comprises adhesion of the bacteria to the root and/or root hairs. In another embodiment, rhizobacteria may adhere to multiple plant surfaces or surface of plant parts, for example to roots and root hairs.

[0128] In another embodiment, a plant surface or a surface of a plant part wherein a B. subtilis may adhere comprises an aerial surface, for example a leaf, a flower, or a stem, or any combination thereof. In another embodiment, a plant surface or a surface of a plant part wherein a B. subtilis may adhere comprises a root epidermis. In another embodiment, a plant surface or a surface of a plant part wherein a B. subtilis may adhere comprises a surface of a root hair. In another embodiment, a plant surface or a surface of a plant part wherein a B. subtilis may adhere comprises a transport vessel. For example, colonization of B. subtilis on a plant' s roots comprises adhesion of the bacteria to the root and/or root hairs. In another embodiment, B. subtilis may adhere to multiple plant surfaces or surface of plant parts, for example to roots and root hairs.

[0129] In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a rhizobacteria may adhere comprises an aerial surface, for example a leaf, a flower, or a stem, or any combination thereof. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a rhizobacteria may adhere comprises a root epidermis. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a rhizobacteria may adhere comprises a surface of a root hair. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a rhizobacteria may adhere comprises a transport vessel. For example, colonization of a rhizobacteria on a plant's roots comprises adhesion of the bacteria to the root and/or root hairs. In another embodiment, biofilm comprising a rhizobacteria may adhere to multiple plant surfaces or surface of plant parts, for example to roots and root hairs.

[0130] In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a B. subtilis may adhere comprises an aerial surface, for example a leaf, a flower, or a stem, or any combination thereof. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a B. subtilis may adhere comprises a root epidermis. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a B. subtilis may adhere comprises a surface of a root hair. In another embodiment, a plant surface or a surface of a plant part wherein a biofilm comprising a B. subtilis may adhere comprises a transport vessel. For example, colonization of a B. subtilis on a plant' s roots comprises adhesion of the bacteria to the root and/or root hairs. In another embodiment, biofilm comprising a B. subtilis may adhere to multiple plant surfaces or surface of plant parts, for example to roots and root hairs.

[0131] In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects plants from phytopathogens (a process termed biocontrol). In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects plants from infection. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects an aerial part of the plant from phytopathogens. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects an aerial part of the plant from infection. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a root of the plant from phytopathogens. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a root of the plant from infection. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a root hair of the plant from phytopathogens. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a root hair of the plant from infection. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a vasculature part of the plant from phytopathogens. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof protects a vasculature part of of the plant from infection.

[0132] In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects plants from phytopathogens. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects plants from infection. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects an aerial part of the plant from phytopathogens. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects an aerial part of the plant from infection. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a root of the plant from phytopathogens. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a root of the plant from infection. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a root hair of the plant from phytopathogens. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a root hair of the plant from infection. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a vasculature part of the plant from phytopathogens. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof protects a vasculature part of of the plant from infection.

[0133] In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in the plant or in a part thereof. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in an aerial portion of a plant, in a root, in a root hair, or in a vasculature part of a plant, or in any combination thereof. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in an aerial portion of a plant. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in a root of a plant. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in a root hair of a plant. In another embodiment, adhesion of a beneficial rhizobacteria with a surface of a plant or a part thereof increases pathogen resistance in a vasculature portion of a plant.

[0134] In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in the plant or in a part thereof. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in an aerial portion of a plant, in a root, in a root hair, or in a vasculature part of a plant, or in any combination thereof. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in an aerial portion of a plant. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in a root of a plant. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in a root hair of a plant. In another embodiment, adhesion of a B. subtilis with a surface of a plant or a part thereof increases pathogen resistance in a vasculature portion of a plant.

[0135] In another embodiment, a method described herein does not induce or enhance or maintain, or any combination thereof, biofilm formation of a plant pathogen. In another embodiment, a method described herein does not stimulate biofilm matrix production of a plant pathogen. In another embodiment, a method described herein does not enhance adhesion of a plant pathogen.

[0136] Methods of this invention inducing biofilm formation on a plant or part thereof, or enhancing biofilm formation on a plant or part thereof, or maintaining biofilm formation on a plant or part thereof, or enhancing adhesion of a beneficial rhizobacteria to a plant or part thereof, refer in one embodiment, to any plant or part thereof growing a soil portion. A plant may comprise any type of plant. For example, in one embodiment a plant comprises a crop plant. As used herein, the term "crop plant" refers to any plant grown for any commercial purpose, including, but not limited to the following purposes: seed production, hay production, ornamental use, fruit production, berry production, vegetable production, oil production, protein production, forage production, animal grazing, golf courses, lawns, flower production, landscaping, erosion control, green manure, improving soil tilth/health, producing pharmaceutical products/drugs, producing food additives, smoking products, pulp production and wood production. In another embodiment, a crop plant comprises a corn, a tomato, a tobacco, a cannabis, a potato or a cucumber plant.

[0137] A part of a plant may comprise any part of a plant either above ground (aerial part of a plant) or below ground. In another embodiment, an aerial part of a plant comprises leaves, twigs, blossoms, fruits or seeds, or any combination thereof. In another embodiment, below ground parts of a plant comprise roots or root hairs, or any combination thereof.

[0138] In one embodiment, pathogen resistance provided by a method of this invention is systemic in a plant. In another embodiment, methods of this invention increase the health and well- being of a plant. In another embodiment, methods of this invention increase the yield of a plant. In another embodiment, increased yield may comprise a yield of fruits, flowers, oil, seed production, berries, vegetables, a protein product, a forage product, a grazing area, a golf course, a lawn, a drug product, a botanical drug product, a food additive, a smoking product, a pulp product, or any combination thereof. In another embodiment, method of this invention increase size of a plant or plant part. In another embodiment, methods of this invention reduce plant stress, for example reducing stress that can affect growth, survival and crop yields. In another embodiment, reduced stress on a plant reduces stress to any of stress factors selected from the group including light, temperature, water, nutrients, pollution, pathogens, or pests, or any combination thereof.

[0139] In one embodiment, pathogen resistance provided by a method of this invention improves the quality of a product produced by a plant. In another embodiment, methods of this invention increase the quality of a product comprising fruits, flowers, oil, seed production, berries, vegetables, a protein product, a forage product, a grazing area, a golf course, a lawn, a drug product, a botanical drug product, a food additive, a smoking product, a pulp product, or any combination thereof. Increased quality of a product may, for example increase taste, color, texture, yield, fragrance, weight, percent of an active component, or percent protein, or any combination thereof.

[0140] In another embodiment, pathogen resistance provided by a method of this invention improves the life span of a plant. In another embodiment, methods of this invention increase the duration of time a plant produces a product, where said product comprises fruits, flowers, oil, seed production, berries, vegetables, a protein product, a forage product, a grazing area, a golf course, a lawn, a drug product, a botanical drug product, a food additive, a smoking product, a pulp product, or any combination thereof.

[0141] In one embodiment, methods of this invention applying compositions comprising a canfhin-6-one based compound result in less use of non-natural chemical treatments for a plant or a part thereof. In another embodiment, methods of this invention applying compositions comprising a canfhin-6-one based compound result in less use of non-natural chemical treatments for a soil portion around a plant or a plant part. In another embodiment, methods of this invention provide a "green" (bio-natural) solution to providing pathogen resistance for a plant or part thereof.

[0142] In some embodiment, the term "a soil portion" refers to an area around a plant, a zone. In one embodiment, "a soil portion" refers in to the soil directly surrounding the plants roots. A zone encompasses the soil all around a plant, including below the plant. In another embodiment, a soil portion comprises the portion of soil into which the plant's roots extend or partially extend. In another embodiment, a soil portion comprises the portion of soil within about mm distance from a plant's roots. . In another embodiment, a soil portion comprises the portion of soil more than 100 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil within about 100 mm distance from a plant's roots. . In another embodiment, a soil portion comprises the portion of soil more than about 50 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil within about 50 mm distance from a plant' s roots. . In another embodiment, a soil portion comprises the portion of soil more than about 25 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil within about 25 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil more than about 10 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil within aboutlO mm distance from a plant's roots. . In another embodiment, a soil portion comprises the portion of soil more than about 5 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil within about 5 mm distance from a plant's roots. In another embodiment, a soil portion comprises the portion of soil less than 5 mm distance from a plant' s roots.

[0143] In one embodiment, methods of this invention apply an effective amount of a composition comprising a canthin-6-one based compound of this invention to a beneficial rhizobacteria. In another embodiment, application may be directly to the beneficial rhizobacteria. In another embodiment, application may be to a soil portion around the plant on which a biofilm may form or has already formed. In another embodiment, application may be to a plant or part thereof on which a biofilm may form or has already formed. In another embodiment, the part of a plant is a root. In another embodiment, the part of a plant is a root hair. In another embodiment, the part of the plant is a root and a root hair. In another embodiment, application may be to an aerial portion of the plant on which a biofilm may form or has already formed.

[0144] In one embodiment, methods of this invention apply an effective amount of a composition comprising a canthin-6-one based compound to a B. subtilis. In another embodiment, application may be directly to the B. subtilis.. In another embodiment, application may be to a soil portion around the plant on which a biofilm produced by a B. subtilis. may form or has already formed. In another embodiment, application may be to a plant or part thereof on which a biofilm may form or has already formed by a B. subtilis.. In another embodiment, the part of a plant is a root. In another embodiment, the part of a plant is a root hair. In another embodiment, the part of the plant is a root and a root hair. In another embodiment, application may be to an aerial portion of the plant on which a biofilm may form or has already formed.

[0145] The plant growth-promoting bacteria may frequently be found associated with plant roots where they protect plants from infection. In one embodiment, methods of this invention promote plant growth.

[0146] In some embodiment, methods of this invention inducing biofilm formation increases pathogen resistance in a plant or a part thereof. In some embodiment, methods of this invention enhancing biofilm formation increases pathogen resistance in a plant or a part thereof. In some embodiment, methods of this invention maintaining biofilm formation increases pathogen resistance in a plant or a part thereof. In some embodiment, methods of this invention stimulating biofilm matrix production in a beneficial rhizobacteria increases pathogen resistance in a plant or a part thereof. In some embodiment, methods of this invention enhancing adhesion of a beneficial rhizobacteria to a surface of a plant or a surface of a part thereof, increases pathogen resistance in the plant or the part thereof. In some embodiment, methods of this invention stimulating biofilm matrix production in a B. subtilis increases pathogen resistance in a plant or a part thereof. In some embodiment, methods of this invention enhancing adhesion of a B. subtilis to a surface of a plant or a surface of a part thereof, increases pathogen resistance in the plant or the part thereof. In another embodiment, methods of this invention enhancing adhesion of a B. subtilis to a surface of a plant or a surface of a part thereof, increases pathogen resistance in a part of the plant different from the part wherein said B. subtilis adheres. In another embodiment, methods of this invention enhancing adhesion of a B. subtilis to a surface of a plant or a surface of a part thereof, increases pathogen resistance in both the portion of the plant wherein said B. subtilis adheres and a part of the plant different from the part wherein said B. subtilis adheres.

[0147] In one embodiment, pathogen resistance is increased in a part of a plant on which a biofilm forms or is maintained. In another embodiment, pathogen resistance is increased in a part of a plant on which a beneficial rhizobacteria adheres. In another embodiment, pathogen resistance is increased in a part of a plant on which a B. subtilis adheres. In another embodiment, pathogen resistance is increased in a part of a plant on which a beneficial rhizobacteria colonizes. In another embodiment, pathogen resistance is increased in a part of a plant on which a B. subtilis colonizes. In another embodiment, pathogen resistance is increased in a part of a plant on which a beneficial rhizobacteria is not adherent. In another embodiment, pathogen resistance is increased in a part of a plant on which a B. subtilis is not adherent. In another embodiment, pathogen resistance is increased in an aerial part of a plant. In another embodiment, pathogen resistance is increased in a part of a plant bellow the ground. In another embodiment, pathogen resistance is increased in a root, a root hair or any combination thereof. In another embodiment, pathogen resistance is increased in a plant vasculature. In another embodiment, pathogen resistance is increased in multiple portions of a plant, comprising aerial and below ground parts of a plant in any combination thereof. In another embodiment, pathogen resistance is systemic in the plant.

[0148] In certain embodiments, plant pathogens are specific to a species or genus of plants. In one embodiment, a plant pathogen comprises a fungus or a bacteria. In another embodiment, a plant pathogen comprises a Pseudomonas or a Xanthomonas. In another embodiment, a Pseudomonas is a Pseudomonas aeruginosa PA01 or a Pseudomanas fluroescens PFZ-89. In another embodiment, a fungal pathogen is a Phytophthora infestanson.

[0149] In certain embodiments, biofilms of this invention may form on aerial parts of a plant, on vasculature of a plant, or on below ground parts of a plant or any combination thereof. In one embodiment, biofilms of this invention are formed on an aerial part of a plant. In another embodiment, biofilms of this invention are maintained on an aerial part of a plant. In another embodiment, biofilms of this invention are formed on a vasculature of a plant. In another embodiment, biofilms of this invention are maintained on a vasculature of a plant. In another embodiment, biofilms of this invention are formed on a root of a plant. In another embodiment, biofilms of this invention are maintained on a root of a plant. In another embodiment, biofilms of this invention are formed on a root hair of a plant. In another embodiment, biofilms of this invention are maintained on a root hair of a plant. In another embodiment, a biofilm of this invention comprises a root-associated biofilm, a root hair associated biofilm, or any combination thereof.

[0150] In certain embodiments, methods of this invention comprise a step of applying a composition comprising an effective amount of an a canthin-6-one based compound or its isomer, tautomer, hydrate, salt, or a combination thereof. In some embodiments, methods of this invention comprise a step of applying a composition comprising an effective amount of a compound of Formula I or its isomer, tautomer, hydrate, salt, or a combination thereof. In some embodiments, methods of this invention comprise a step of applying a composition comprising an effective amount of a compound of Formula II or its isomer, tautomer, hydrate, salt, or a combination thereof. In one embodiment, an effective amount is determined based on the method of application. For example, application directly to a plant or part thereof, or application to a portion around a plant, or application to a beneficial rhizobacteria such as B. subtilis. In another embodiment, an effective amount is the amount that induces biofilm formation. In another embodiment, an effective amount is the amount that enhances biofilm formation. In another embodiment, an effective amount is the amount that maintains biofilm formation. In another embodiment, an effective amount is the amount that stimulates biofilm matrix production. In another embodiment, an effective amount is the amount that enhances adhesion of a beneficial rhizobacteria. In another embodiment, an effective amount is the amount that enhances adhesion of a B. subtilis.

[0151] In one embodiment, an "effective amount" of a canthin-6-one based compound for a particular result, refers herein to an amount needed (or a sufficient amount) to achieve the desired goal. For example, in one embodiment an effective amount of a canthin-6-one based compound of this invention induces biofilm formation by a beneficial rhizobacteria on a plant or a part thereof. In another embodiment, an effective amount of a canthin-6-one based compound of this invention enhances biofilm formation by a beneficial rhizobacteria on a plant or a part thereof. In another embodiment, an effective amount of a canthin-6-one based compound of this invention maintains a biofilm formed by a beneficial rhizobacteria on a plant or a part thereof. In another embodiment, an effective amount of a canthin-6-one based compound of this invention stimulates matrix production by a beneficial rhizobacteria. In yet another embodiment, an effective amount of a canthin-6-one based compound of this invention enhances adhesion of a beneficial rhizobacteria to a plant surface or a part thereof.

[0152] In one embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose range of about 0.1-4 ppm canthin-6-one (0.1 μΜ - 4μΜ). In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose range of about 0.5-3 ppm canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose range of about 1- 2 ppm canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 1 ppm canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 2 ppm canthin-6-one.

[0153] In one embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose range of about 1-500 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose range of about 10-50 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 10 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 20 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 30 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 40 mM canthin-6-one. In another embodiment, a composition comprising a canthin-6-one based compound of this invention is applied in a dose of about 50 mM canthin-6-one.

[0154] In one embodiment, the term "about", refers to a deviance of between 0.0001-5% from the indicated number or range of numbers. In one embodiment, the term "about", refers to a deviance of between 1 -10% from the indicated number or range of numbers. In one embodiment, the term "about", refers to a deviance of up to 25% from the indicated number or range of numbers.

[0155] In some embodiments, the term "comprise" or grammatical forms thereof, refers to the inclusion of the indicated active agent, such as a canthin-6-one based compound of this invention, as well as inclusion of other active agents, and acceptable carriers, excipients, emollients, stabilizers, etc., as are known in the industry. In some embodiments, the term "consisting essentially of refers to a composition, whose only active ingredient is the indicated active ingredient, however, other compounds may be included which are for stabilizing, preserving, etc. the composition, but are not involved directly in the effect of the indicated active ingredient. In some embodiments, the term "consisting essentially of may refer to components, which exert an effect via a mechanism distinct from that of the indicated active ingredient. In some embodiments, the term "consisting essentially of may refer to components, which exert an effect and belong to a class of compounds distinct from that of the indicated active ingredient. . In some embodiments, the term "consisting essentially of may refer to components, which exert an effect and belong to a class of compounds distinct from that of the indicated active ingredient, by acting via a different mechanism of action. In some embodiments, the term "consisting" refers to a composition, which contains the active ingredient and a acceptable carrier or excipient.

[0156] In one embodiment, the term "a" or "one" or "an" refers to at least one. In one embodiment the phrase "two or more" may be of any denomination, which will suit a particular purpose.

[0157] The following examples are presented in order to more fully illustrate the preferred embodiments of the invention They should in no way be construed, however, as limiting the broad scope of the invention.

Examples

Example 1: Canthin-6-one Enhances Biofilm Formation and Matrix Gene Transcription

[0158] Methods

[0159] Biofilms were grown under static conditions and floated in liquid MSgg medium for 3 days at 23°C with different concentrations of canthin-6-one. The transcription levels of the biofilm activator sini was measured using a luciferase reporter system Luminescence of strains harboring sacA::P_sini-luciferase without or in the presence of canthin-6-one at 1 μΜ or 5 μΜ, was measured. Planktonic growth of wild-type B. subtilis was measure in cultures grown with shaking in liquid MSgg mediaum without (untreated) or with canthin-6-one.

[0160] Results

[0161] B. subtilis, NCBI 3610 (hereafter referred to as "3610"), forms robust and highly structured biofilms in liquid medium. When inoculated into a standing culture of minimal medium, 3610 initially grows planktonically as motile, single cells. The cells then migrate to the air-liquid interface, where they proliferate as long chains of non-motile cells. These chains are highly ordered in parallel patterns and are bound together tightly, presumably by an extracellular matrix (ECM), to form a floating biofilm (pellicle). Growth of the bundled chains continues, giving rise to larger biofilm structures that include the fruiting bodies. Thus, pellicles were considered as a highly robust setting for the study of B. subtilis biofilms.

[0162] The effects of applying canthin-6-one to Bacillus subtilis protective biofilms were evaluated. It was found that a small molecule produced by Eurycoma, induces dramatically the biofilm of the biocontrol agent Bacillus subtilis (Figure 1A) in μΜ range.

[0163] Next, determining the mechanism that enables biofilm induction of Bacillus subtilis by Canthin-6-one was studied. In this bacteria, the commitment to a biofilm state requires the phosphorylation of the master regulator SpoOA. Spo0A-P activates Sini, which represses SinR via direct binding (Bai et al., 1993). SinR is the main transcriptional repressor of two matrix encoding operons (Bai et al., 1993; Kearns et al., 2005), epsA-0 operon, and the yqxM/tapA-sipW-tasA operon, encoding the amyloid protein TasA, the major protein component of the biofilm matrix (Stover and Driks, 1999; Branda et al., 2006; Romero et al, 2010) J. Bacteriology 181 :1664-1672).

[0164] The expression of Sini, the matrix regulator and tapA-sipW-tasA operon were measured in the absence and presence of canthin-6-one.In adition, sini transcription was monitored, sini transcription is a direct readout for the activity of the genetic pathway resulting in matrix production and biofilm formation, sini transcription in biofilm-inducing medium was monitored using the sini promoter fused to luciferase. A minimum biofilm inducing concentration of 5 μΜ canthin-6-one or higher concentrations significantly enhanced the transcription of sinl (Figure IB). Importantly, as the sinl pathway is only conserved among Bacillus subtilis species, it is another indication to the quite specific activity of canthin-6-one on beneficial communities.

[0165] Importantly, pellicle formation, which is an indication of secretion of EPS and biofilm formation, was blocked in concentrations that had no effect on planktonic growth (Figure 1C). The specific induction of biofilm formation without impacting other bacterial behaviors suggests this molecule functions as a signal for enhancing biocontrol-promoting communities.

Example 2: Canthin-6-one Does Not Enhance Biofilm Formation for Pathogens

[0166] Methods

[0167] Quantification of submerged biofilm formation by Pseudomonas species was performed in standardized media (TSB) following 24 hours of incubation in 96 well plates. P. fluorescens is an opportunistic plant pathogen and P. aeruginosa is a human pathogen.

[0168] Results

[0169] Canthin-6-one, a potential enhancer of biocontrol properties of beneficial communities, did not affect biofilm formation in plant pathogens. Strikingly, bioactive concentration of canthin-6- one did not affect biofilm formation of Pseudomonas species (Figures 2A and 2B), which represent a significant group of plant and human pathogens. The specificity of this biofilm-enhancing signal for beneficial biocontrol agents makes it an ideal candidate for agricultural applications. Importantly and consistent with these findings, the sinl pathway, triggering biofilm formation in the beneficial bacteria Bacillus subtilis, is conserved within the species, and not widespread. For example, the sinl pathway is not found in Pseudomonas species..

Example 3: Canthin-6-one Enhances Adhesion of B. subtilis to Tomato Root

[0170] Methods

[0171] Quantification of bacterial densities eluted from tomato root was measured following 10 hours of incubation in plant media in the presence or absences of different concentrations of canthin-6-one.

[0172] Results

[0173] The effect of canthin-6-one on bacterial adhesion to the root of tomato were evaluated. The adhesion tests revealed a dramatic change in B. subtilis adhesion behavior over the range of canthin-6-one concentrations (Figure 3). There were hundreds (10²) of bacteria adhering to the tomato roots in the absence of canthin-6-one. Incubation with canthin-6-one increased the adhesion to millions (10⁶) as reflected by the OD. 0.6 is equivalent to 10⁸ bacteria. It is evident that the adhesion rates increased sharply with an increase in the canthin-6-one concentration from 0 μΜ to 50 μΜ. Therefore, the presence of canthin-6-one has an obvious effect on adhesion of Bacillus subtilis (P < 0.05). The increase in bacterial adhesion properties is consistent with previous findings demonstrating that the sinl pathway induces the synthesis of exopolysaccharides, serving as bacterial adhesins to the root. (Branda et al., 2004, Genes involved in formation of structured multicellular communities by Bacillus subtilis. Journal of bacteriology 186, 3970-3979.)

Example 4: Canthin-6-one Decreased Pathogen Related Mortality of Tomato Plant

[0174] Methods: Bacillus strain B. subtilis 3610 (B.S), was used. The bacterial strain was maintained on LB plates. 1000 cells were treated or not-treated (NT) with 5 μΜ of Canthin-6-one, then added to tomato plants grown on agrophonic beads. Plants were incubated at 28 ± 2 °C for either 5 days or 15 days. The tomato pathogen, P. syringae strain DC3000 pv tomato (P.S) was grown in LB at 37°C to OD600 = 0.2 to 0.3 (3 *108 cells per ml) and diluted to 100 cells, or 10,000 cells, respectively (w/v), and added into the plant media. Plants were further incubated at 28 ± 2 °C for 10 days and the growth of the plants was measured and compared with the control. Three replications were maintained for each assay. SD represents the average of three independent experiments.

[0175] Results: Figure 4 shows that Canthin-6-one improved the bioprotection of the tomato plants versus the tomato pathogen Pseudomonas syringae achieved by Bacillus.

[0176] Conclusion: Canthin-6-one improved significantly the protection achieved by Bacillus subtilis.

[0177] While certain features of the invention have been illustrated and described herein, many modifications, substitutions, changes, and equivalents will now occur to those of ordinary skill in the art. It is, therefore, to be understood that the appended claims are intended to cover all such modifications and changes as fall within the true spirit of the invention.

Claims

CLAIMS What is claimed is:

1. A method for inducing, enhancing, or maintaining biofilm formation by a beneficial rhizobacteria on a plant or a part thereof, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to the beneficial rhizobacteria, wherein said canthin-6-one based compound is represented by the structure of

(I)

wherein,

Ri, R2, R3, R4, R5, R6, R7, Rs are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, N0₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and

R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen, or its isomer, tautomer, hydrate, salt or a combination thereof, thereby inducing, enhancing, or maintaining said biofilm formation.

2. The method according to claim 1 , wherein said canthin-6-one based compound is represented by the structure of Formula II:

or its isomer, tautomer, hydrate, salt or a combination thereof.

3. The method according to any one of claims 1-2, wherein said inducing, enhancing, or maintaining biofilm formation, or any combination thereof, increases pathogen resistance in the plant or the part thereof.

4. The method of claim 3, wherein said pathogen resistance is increased in a root or root hair of the plant, or any combination thereof.

5. The method of claim 3, wherein said pathogen resistance comprises systemic resistance in the plant.

6. The method according to any one of claims 3-5, wherein said pathogen comprises a fungus or a bacterium.

7. The method of claim 6, wherein said bacteria comprises a Pseudomonas or a Xanthomonas and said fungus comprises a Phytophthora infestanson.

8. The method of claim 7, wherein said Pseudomonas comprises Pseudomonas fluroescens PFZ-89.

9. The method according to any one of claims 1-8, wherein said biofilm comprises a root- associated biofilm or a root hair associated biofilm, or any combination thereof

10. The method according to any one of claims 1 -9, wherein said composition comprises a plant extract.

11. The method according to any one of claims 1-9, wherein said canthin-6-one based compound is purified from a plant.

12. The method of any one of claims 10 or 11, wherein said plant is selected from the group comprising an Ailanthus altissima, an Aerva lanata, a Brucea antidysenterica, a Eurycoma harmandiana, a Eurycoma longifolia, a Peganum Nigellastrum, a Zanthoxylum elephantiasis, or any combination thereof.

13. The method of any one of claims 1-9, wherein said canthin-6-one based compound is in the form of a semi-synthetic variant or a synthetic variant.

14. The method according to any one of claims 1-13, wherein said application comprises applying a composition comprising the canthin-6-one based compound (a) to the roots of the plant in combination with spores of said rhizobacteria, (b) to the roots of the plant prior to applying spores of said rhizobacteria to the roots of the plant, (c) to a soil portion around the plant or a part thereof, (d) directly to the rhizobacteria in vitro, followed by contacting said plant roots with said rhizobacteria, or (e) directly to the rhizobacteria in vitro, followed by contacting a soil portion around said plant or part thereof with said rhizobacteria, or any combination thereof.

15. The method according to claim 14, wherein said part thereof comprises a root.

16. The method according to any one of claims 14-15, wherein said soil portion is within about 25 mm of said plant or part thereof.

17. The method according to any one of claims 1-16, wherein said canthin-6-one based compound is at a concentration of about 1 -50 μΜ.

18. The method according to any one of claims 1 -16, wherein said applying is at a dose of about 10-50 mM canthin-6-one based compound.

19. The method according to any one of claims 1 -16, wherein said applying is at a dose of about 1-2 ppm canthin-6-one based compound.

20. The method according to any one of claims 1-19, wherein said plant comprises a crop plant.

21. The method according to claim 20, wherein said crop plant is selected from the group comprising corn, tomato, a tobacco, a cannabis, a potato or a cucumber plants.

22. The method according to any one of claims 1-21, wherein said beneficial rhizobacteria comprises a sinl matrix regulator gene pathway.

23. The method according to any one of claims 1-22, wherein said beneficial rhizobacteria comprises a Bacillus subtilis (B. subtilis) species, a Bacillus amyloliquefaciens FZB24, a Bacillus amyloliquefaciens FZB42, or a Bacillus amyloliquefaciens FZB45..

24. The method of claim 23, wherein said B. subtilis is selected from the group comprising B. subtilis NCBI 3610, B. subtilis FB17, B. subtilis NATTO, B. subtilis CPA-8, B. subtilis RO-FF-1 , B. subtilis JH642, B. subtilis GB03, B. Subtilis E1R-J, and B. subtilis FZB37.

25. The method according to any one of claims 1-24, wherein production of an extracellular matrix by said rhizobacteria is stimulated.

26. The method according to any one of claims 1-25, wherein said method further comprises enhancing adhesion of said rhizobacteria to said plant or plant part thereof.

27. The method according to any one of claims 1-26, wherein said composition comprises a solution, a powder, a spray, drops, a tablet, or a paste.

28. The method of claim 27, wherein said solution is an aqueous solution.

29. The method of claim 28, wherein said aqueous solution has a pH of about 7.0.

30. A method for stimulating matrix production by a beneficial rhizobacteria, said method comprising the step of applying a composition comprising an effective amount of a canthin-6-one based compound to said beneficial rhizobacteria, wherein said canthin-6- one based co a I:

(I)

wherein,

Ri, R₂, R3, R4, R5, R6, R7, Re are each independently hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted, saturated or unsaturated heterocycle, substituted or unsubstituted aryl, halogen, NH₂, COOH, N0₂, CN, OH, alkoxy, haloalkyl, SH, thioalkyl, NO, alkylamine, CONH₂, COOR, S0₂R, N(R)₂, COR, -NHR; and

R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen; or its isomer, tautomer, hydrate, salt or a combination thereof, thereby stimulating said matrix production.

31. The method of claim 31 , wherein said canthin-6-one based compound is represented by the structure of Formula II:

or its isomer, tautomer, hydrate, salt or a combination thereof.

32. The method according to any one of claims 30-31 , wherein said stimulating matrix production reduces transcriptional repression of a matrix encoding operon selected from an epsA-0 operon and yqxM/tapA-sipW-tasA operon, or any combination thereof.

33. The method according to any one of claims, 30-32, wherein said stimulating matrix production, increases pathogen resistance in the plant or the part thereof.

34. The method of claim 33, wherein pathogen resistance is increased in a root or root hair of the plant, or any combination thereof.

35. The method of claim 33, wherein said pathogen resistance comprises a systemic resistance.

36. The method according to any one of claims 30-35, wherein said pathogen comprises a fungus or a bacterium.

37. The method of claim 36, wherein said bacteria comprises a Pseudomonas or a Xanthomonas and fungus comprises a Phytophthora infestanson.

38. The method of claim 37, wherein said Pseudomonas is Pseudomonas fluroescens PFZ- 89.

39. The method according to any one of claims 30-38 wherein said stimulating matrix induces, enhances, or maintains, or any combination thereof, a biofilm on a plant or part thereof.

40. The method of claim 39, wherein said biofilm comprises a root-associated biofilm or a root hair associated biofilm, or any combination thereof.

41. The method according to any one of claims 30-40, wherein said composition comprising the canthin-6-one based compound comprises a plant extract.

42. The method according to any one of claims 30-40, wherein said canthin-6-one based compound is purified from a plant.

43. The method of any one of claim 41-42, wherein said plant is selected from the group comprising an Ailanthus altissima, an Aerva lanata, a Brucea antidysenterica, a Eurycoma harmandiana, a Eurycoma longifolia, a Peganum Nigellastrum, a Zanthoxylum elephantiasis, or any combination thereof.

44. The method of any one of claims 30-40, wherein said canthin-6-one based compound is in the form of a semi-synthetic variant or a synthetic variant.

45. The method according to any one of claims 30-44, wherein said application comprises applying a composition comprising the canthin-6-one based compound (a) to the roots of the plant in combination with spores of said rhizobacteria, (b) to the roots of the plant prior to applying spores of said rhizobacteria to the roots of the plant, (c) to a soil portion around the plant or a part thereof, (d) directly to the rhizobacteria in vitro, followed by contacting said plant roots with said rhizobacteria, or (e) directly to the rhizobacteria in vitro, followed by contacting a soil portion around said plant or part thereof with said rhizobacteria, or any combination thereof..

46. The method according to claim 45, wherein said part thereof comprises a root.

47. The method according to any one of claims 45-46, wherein said soil portion is within about 25 mm of said plant or part thereof.

48. The method according to any one of claims 30-47, wherein said canthin-6-one based compound is at a concentration of about 1 -50 μΜ.

49. The method according to any one of claims 30-47, wherein said applying is at a dose of about 10-50 mM canthin-6-one.

50. The method according to any one of claims 30-47, wherein said applying is at a dose of about 1-2 ppm canthin-6-one.

51. The method according to any one of claims 30-47, wherein said plant comprises a crop plant.

52. The method of claim 51 wherein said crop plant is selected from the group comprising corn, tomato, a tobacco, a cannabis, a potato or a cucumber plants.

53. The method according to any one of claims 30-52, wherein said beneficial rhizobacteria comprises a sinl matrix regulator gene pathway.

54. The method according to any one of claims 30-53, wherein said beneficial rhizobacteria comprises a Bacillus subtilis (B. subtilis) species, a Bacillus amyloliquefaciens FZB24, Bacillus amyloliquefaciens FZB42, or a Bacillus amyloliquefaciens FZB45..

55. The method of claim 54, wherein said B. subtilis is selected from the group comprising B. subtilis NCBI 3610, B. subtilis FB17, B. subtilis NATTO, B. subtilis CPA-8, B. subtilis RO-FF-1 , B. subtilis JH642, B. subtilis GB03, B. Subtilis E1R-J, and B. subtilis FZB37.

56. The method according to any one of claims 30-55, wherein said method further comprises enhancing adhesion of said rhizobacteria to said plant or plant part thereof.

57. The method of claim 56, wherein said part thereof comprises a root.

58. The method according to any one of claims 30-57, wherein said composition comprises a solution, a powder, a spray, drops, a tablet, or a paste.

59. The method of claim 58, wherein said solution is an aqueous solution.

60. The method of claim 59, wherein said aqueous solution is at about pH 7.0.

61. A method of enhancing adhesion of a beneficial rhizobacteria to a plant surface, said method comprising the step of applying a composition comprising a composition of a canthin-6-one based compound to said rhizobacteria in an amount effective to enhance adhesion of the beneficial rhizobacteria to said plant surface, wherein said canthin-6-one based compou

(I)

wherein,

R is hydrogen, substituted or unsubstituted linear or branched alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted aryl or halogen; or its isomer, tautomer, hydrate, salt or a combination thereof.

62. The method of claim 61 , wherein said canthin-6-one based compound is represented by the structure of Formula II:

(Π)

or its isomer, tautomer, hydrate, salt or a combination thereof.

63. The method according to any one of claims 61-62, wherein said enhancing adhesion, increases pathogen resistance in the plant or the part thereof.

64. The method of claim 63, wherein pathogen resistance is increased in a root or root hair of the plant, or any combination thereof.

65. The method of claim 63, wherein said pathogen resistance comprises systemic resistance in said plant.

66. The method according to any one of claims 61-65, wherein said pathogen comprises a fungus or a bacterium.

67. The method of claim 66, wherein said bacteria comprises a Pseudomonas or a Xanthomonas and said fungus comprises a Phytophythora infestanson.

68. The method of claim 67, wherein said Pseudomonas comprises a Pseudomonas fluroescens PFZ-89.

69. The method according to any one of claims 61-68, wherein said enhancing adhesion to a plant surface induces, enhances, or maintains, or any combination thereof, a biofilm on a plant or part thereof.

70. The method of claim 69, wherein said biofilm comprises a root-associated biofilm or a root hair associated biofilm, or any combination thereof.

71. The method according to any one of claims 61-70, wherein said composition comprising said canthin-6-one based compound comprises a plant extract.

72. The method according to any one of claims 61-70, wherein said canthin-6-one based compound is purified from a plant.

73. The method of any one of claims 71-72, wherein said plant is selected from the group comprising an Ailanthus altissima, an Aerva lanata, a Brucea antidysenterica, a Eurycoma harmandiana, a Eurycoma longifolia, a Peganum Nigellastrum, a Zanthoxylum elephantiasis, or any combination thereof.

74. The method of any one of claims 61-70 wherein said canthin-6-one based compound is in the form of a semi-synthetic variant or a synthetic variant.

75. The method according to any one of claims 61-74, wherein said application comprises applying a composition comprising the canthin-6-one based compound (a) to the roots of the plant in combination with spores of said rhizobacteria, (b) to the roots of the plant prior to applying spores of said rhizobacteria to the roots of the plant, (c) to a soil portion around the plant or a part thereof, (d) directly to the rhizobacteria in vitro, followed by contacting said plant roots with said rhizobacteria, or (e) directly to the rhizobacteria in vitro, followed by contacting a soil portion around said plant or part thereof with said rhizobacteria, or any combination thereof.

76. The method according to claim 74, wherein said part thereof comprises a root.

77. The method according to any one of claims 75-76, wherein said soil portion is within about 25 mm of said plant or part thereof.

78. The method according to any one of claims 61-77, wherein said canthin-6-one based compound is at a concentration of about 1 -50 μΜ.

79. The method according to any one of claims 61-77, wherein said applying is at a dose of about 10-50 mM canthin-6-one.

80. The method according to any one of claims 61-77, wherein said applying is at a dose of about 1-2 ppm canthin-6-one.

81. The method according to any one of claims 61-80, wherein said plant comprises a crop plant.

82. The method of claim 81 , wherein said crop plant is selected from the group comprising corn, tomato, a tobacco, a cannabis, a potato or a cucumber plants.

83. The method according to any one of claims 61-82, wherein said beneficial rhizobacteria comprises a sinl matrix regulator gene pathway.

84. The method according to any one of claims 61-83, wherein said beneficial rhizobacteria comprises a Bacillus subtilis (B. subtilis) species, a Bacillus amyloliquefaciens FZB24, Bacillus amyloliquefaciens FZB42, or a Bacillus amyloliquefaciens FZB45. .

85. The method of claim 84, wherein said B. subtilis is selected from the group comprising B. subtilis NCBI 3610, B. subtilis FB17, B. subtilis NATTO, B. subtilis CPA-8, B. subtilis RO-FF-1 , B. subtilis JH642, B. subtilis GB03, B. Subtilis E1R-J, and B. subtilis FZB37.

86. The method according to any one of claims 61-85, wherein said composition comprises a solution, a powder, a spray, drops, a tablet, or a paste.

87. The method of claim 86, wherein said solution is an aqueous solution.

88. The method of claim 87, wherein said aqueous solution has a pH of about 7.0.