WO2010028215A1 - Antimicrobial fish and shrimp feed - Google Patents

Abstract

The present invention relates to modified "aqua feed," or fish and shrimp feed, having enhanced antimicrobial properties effective at suppressing and controlling infection. The modified feed composition includes an acidulant, such as ACS or HAMO, as well as optionally a clay material and a desiccant. The ACS is preferably at a normality of 5-10 N. The clay material has been shown to be effective at adsorption of aflatoxins. The pH of the feed composition is preferably about 4.5 to about 5.0. Other acids and additives can also be added to the feed composition.

Description

ANTIMICROBIAL FISH AND SHRIMP FEED

BACKGROUND

[0001] This application claims priority to U.S. Provisional Patent Application Serial No. 61/094.592 entitled "Antimicrobial Fish and Shrimp Feed,^"' filed on September 5, 2008, the entire content of which is hereby incorporated by reference.

[0002] This invention pertains to fish and shrimp feed and particularly to modifying and acidifying fish and shrimp feed with selected acidulants and additives in order to enhance its antimicrobial properties.

[0003] Aquaculture is the fastest growing sector of food production in the world. The Food and Agriculture Organization ("FAO") of the United Nations reports that aquaculture accounts for almost 50% of the world's food fish. Aquaculture has the greatest potential to meet the growing demand of aquatic food that will arise from an expected population increase of 1.5 billion people in 2020. In order to maintain the current per capita consumption, an additional 40 million tons of aquatic food will be required by 2030. Mounting scientific evidence indicates that dramatic declines in many natural fish stocks are occurring. Capture fisheries are not capable of providing fish to an additional 1.5 billion people, considering FAO already lists 89% of the ocean's wild fish stocks as moderately, fully or over-exploited.

[0004] In addition to meeting global consumption needs, aquaculture. particularly shrimp farming, has become an increasingly important economic activity. In recent years, high value and market demand by mainly affluent consumers in developed countries has lead to rapid expansion of shrimp aquaculture throughout Asia and Latin America. In 1999, shrimp aquaculture only represented about 2.6% of total global aquaculture, but accounted for 16.5% of the total revenue at a value of about $6.7 billion. Considerable private and public sector investment has induced an annual average increase in cultured shrimp production of about 5- 10% since the 1990's.

[0005] Shrimp farming is one ofthe most profitable and fastest-growing segments of the aquacuiture industry. Global farmed shrimp production has grown more than 100- fold (by weight) in less than two decades; from under 10.000 metric tons (MT) produced by fewer than a dozen countries in the early 1970s to over 1 million MT (MMT) by the late 1990s. Worldwide sale of shrimp is estimated to be a 10 billion dollar industry. In 2006, aquafeeds used 68 percent of the world's supply of fish meal (3.724 million metric tons) and 88.5 percent offish oil (835,000 million metric tons). Aquafeeds production is expected to double within 10 years. Researchers began looking for fish meal substitutes in earnest after an El Nino event in the 1970s drastically reduced fish meal production and sent prices to historic highs. Protecting against catastrophic events was one goal, but so was protecting fish populations by not harvesting as many for fish meal. While fish meal prices have remained relatively stable, prices are about 50 percent higher than just a few years ago. At the same time, the prices of corn, soybeans, wheat and rice have soared

[0006] Vibriosis is the term used to refer to a multitude of infections caused by bacteria belonging to the genus Vibrio. The direct economic implications of vibriosis can be devastating, particularly when acute outbreaks occur. The major species causing vibriosis in shrimp are Vibrio parahemolyticus, V. vulnificus, V. al ginυly tic us anά V. hαrveyi. If vibriosis occurs in the hatchery, the loss is 100%. Vibriosis during grow-out precipitates chronic losses either directly, or following immunosuppression and consequential secondary infection.

[0007] Probiotic bacteria that produce acidic metabolites when added to shrimp feed have been shown to suppress vibriosis. The mechanism is not fully defined but the microbial gut content of the shrimp continuously fed with a diet supplemented with the probiotic, had a lower microbial content, i.e., the number of vibrio organisms was reduced. There were also fewer vibrio organisms in the shrimps heamolymph.

[0008] Necrotizing Hepatopancreatitis ("NHP") is a severe bacterial disease affecting penaeid shrimp aquaculture. NHP was first reported in 1985 from shrimp ponds in Texas and resulted in significant mortalities and devastating losses to shrimp crops. NHP has since been observed in penaeid shrimp aquaculture in Central and South American countries and is possibly in the Eastern Hemisphere as well. Elevated salinity and temperature appear to be factors associated with NHP outbreaks. Reported hosts are Litopenueus vαnnαmei, L. setiferiis, L. stylirostris. Fαrfαntepenαeus αzfecus and F. cαlifomiensis. The N HP-bacterium ("NHPB") is a gram-negative, pleomorphic, obligate intracellular bacterium that is non- culturable through established cell lines or traditional bacteriological methods. Therefore, laboratory research of NI JP is dependent on maintaining the disease agent in live animals. Gross signs of NHP include reduced feed intake, empty gut, lethargy, anorexia, and a pale and atrophied hepatopancreas.

[0009] The beneficial application of organic acid salts for shrimp has been reported. Inactivated lactobacilli along with 5 kg/MT sodium citrate has been used to boost growth of Komura shrimp {Penaeus japonicus). In addition, Luckstadt used a dosage of 2.5 kg/MT of calcium formate to show that organic salts can enhance survival of brackish water shrimp during grow-out (Luckstadt 2006).

[0010] Diseases in fish tend to be similar to those found in shrimp. In particular, they are primarily caused by bacteria. Intracellular rickettsia is a major disease affecting fish, and antibiotic treatments so far have failed.

[0011] Microorganism and aflatoxin contamination of animal feeds also constitutes a serious threat to the health of humans and animals. Generally, microorganisms are considered to be any microscopic or ultramicroscopic animal, plant, bacterium, virus, etc., and aflatoxins (Afs) are harmful by-products of mold growth, which both are potentially fatal to humans and animals. Furthermore, such contamination in animal feeds also can lead to severe economic losses.

[0012] Aflatoxins are produced by Aspergillus flams or A. parasiticus. There are at least four naturally occurring aflatoxins, namely AlBl, AfB2, AfGl and AfG2, as shown in Figure 1. Many aflatoxins occur as natural contaminants in a variety of foods and feeds, such as corn, wheat, barley, beans, sorghum, moldy peanuts, mixed feed, and some coffee beans. They are also found as residues in liver, kidneys of pigs. Many of these aflatoxins have been known to be strong carcinogens, thus causing cancers, in humans and they are also capable of eliciting other toxic effects, such as teratogenesis.

[0013] Many strategics have been developed to inactivate aflatoxins in feeds and the strategies give varying degrees of success. The strategies include processing the food and feed, biocontrol and microbial inactivation, chemical treatment after structural degradation, dietary modification of toxicity and absorption method to reduce bioavailable aflatoxin. One popular method is to add an non-nutritive adsorbents in contaminated feeds to inactivate the aflatoxin. Various adsorbents have been used, such as aluminas, zeolites, silicas, phyllosilicates, bentonite, activated charcoal, and montmorillonite. [0014] As mentioned above, feeds are very often contaminated with both harmful microorganisms and toxic aflatoxins. The methods currently available to solve these problems usually are those that will either kill harmful microorganisms or deactivate toxic aflatoxins, but not both. Thus, it is desirable to have a method that can both kill the harmful microorganisms and simultaneously deactivate toxic aflatoxins.

SUMMARY

[0015] The present invention pertains to the suppression and control of microbial infection of fish and shrimp through the use of modified and acidified fish and shrimp feed.

[0016] Acidification of fish and shrimp feed assists in the reduction of bacterial growth and infection, as well as the production of histamines. Carboxylic acids in general are useful feed acidifiers. Even more useful are salts, particularly calcium salts, of organic acids. When these salts become dissociated, the organic acid is regenerated and can then freely enter microbial cells. Organic acids can freely enter microbial cells if they are protonated, which is the case when they are acidulated.

[0017] An acidulant such as any acid, or an acidic, or low pH, solution of sparingly-soluble Group lIA-complexes ("AGIlS"), or preferably acidic calcium sulphate (''ACS"), added to fish and shrimp feed is effective at suppressing and controlling infection by bacteria, including Vibrio, Pseuάomonas, and NHP-bacteria, and rickettsia. Use of an acidulant, including AGIIS (or ACS), in fish and shrimp feed both prevents infection and decreases recovery time for infected organisms. This reduces stress on the population and leads to normal feed consumption and growth patterns, as well as higher yield at harvest. Thus, a greater return on investment is achieved. Furthermore, the use of an acidulant such as ACS rather than antibiotics means that there is no chance of contributing to antibiotic resistance in bacteria, nor is there any antibiotic-related effects on the food supply.

[0018] Other acidifiers may accomplish the same purpose, such as organic acids or highly acidic metalated organic acid ("HAMO"). In addition, adding clay material and a desiccant in addition to the acidifier is preferable. The clay material may be any suitable clay, including an attapulgite type clay, a montmorillonite clay, preferably a hydrated sodium calcium aluminosilicate ("HSCAS") clay, an amorphous opaline silica clay, including silica hydrated, or crystalline silica clay, including quartz.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

[0019] One aspect of the present invention is a modified composition for treating "aqua feed," that is, fish and shrimp feed, to suppress and control the infection of fish and shrimp by bacteria, including Vibrio, Pseudomotias, NHP-bacteria, and rickettsia. The composition comprises an acidulant, such as acidic calcium sulfate ("ACS"), added to conventional aqua feed, or fish and shrimp feed. In preferred embodiments, the composition can also comprise clay material, other acids or acidulants, desiccants, dyes, or combinations thereof.

[0020] One preferred embodiment of the modified composition comprises fish or shrimp feed material, an acidulant, and a clay material. Another preferred embodiment is fish or shrimp feed material combined with an acidulant.

[0021] Preferably, the acidulant is ACS or HAMO. "ACS" is also referred to as an acidic, or low pH, solution of sparingly-soluble Group IlA-complexes ("AGIΪS") (See, U.S. Patent No.6,902,753, "Acidic Solution of Sparingly-Soluble Group HA Complex"; see also, U.S. Patent No. 6,436,891, "Adduct Having An Acidic Solution of Sparingly-Soluble Group HA Complexes"; the entire content of each of the two is hereby incorporated by reference). Preferred strengths of ACS include 5 N - 10 N ACS, or stronger, such as ACS-50 or ACS-100 (Mionix Corporation, Rocklin, CA). The term "complex," as used herein, denotes a composition wherein individual constituents are associated. "Associated" means constituents are bound to one another either covalently or non-covalently, the latter as a result of hydrogen bonding or other inter-molecular forces. The constituents may be present in ionic, non-ionic, hydrated or other forms.

[0022] The acidic solution of sparingly-soluble Group II A-complex salt (AGIlS), or ACS, can be prepared in several ways. Some of the methods involve the use of Group IA hydroxide but some of syntheses are devoid of the use of any added Group IA hydroxide, although it is possible that a small amount of Group IA metal may be present as "impurities." The preferred way of manufacturing AGIIS (or ACS) is not to add Group IA hydroxide to the mixture. As the phrase implies, AGIIS is highly acidic, ionic, with a pH of below about 2.

[0023] The preferred method of preparing AGIlS (or ACS) involves mixing a mineral acid with a Group HA hydroxide, or with a Group UA salt of a dibasic acid, or with a mixture of the two Group HA materials. In the mixing, a salt of Group HA is also formed. Preferably, the starting Group HA material or materials selected will give rise to, and form, the Group HA salt or salts that are sparingly soluble in water. The preferred mineral acid is sulfuric acid, the preferred Group HA hydroxide is calcium hydroxide, and the prefer Group HA salt of a dibasic acid is calcium sulfate. Other examples of Group HA salt include calcium oxide, calcium carbonate, and "calcium bicarbonate."

[0024] AGIIS (or ACS) is preferably prepared by mixing calcium hydroxide with concentrated sulfuric acid, with or without an optional Group HA salt of a dibasic acid (such as calcium sulfate) added to the sulfuric acid. For every mole of concentrated acid, such as sulfuric acid, the amount, in moles, of calcium hydroxide used is application specific and ranges from about 0.1 to about 1. The optional calcium sulfate can be added to the concentrated sulfuric acid prior to the introduction of calcium hydroxide into the blending mixture. The addition of calcium sulfate to the concentrated sulfuric acid appears to reduce the amount of calcium hydroxide needed for the preparation of AGIIS (or ACS). For every mole of concentrated acid, such as sulfuric acid, the amount, in moles, of calcium carbonate ranges from about 0.001 to about 0.2, depending on the amount of calcium hydroxide used. Other optional reactants include calcium carbonate and gaseous carbon dioxide being bubbled into the mixture. Regardless of the use of any optional reactants, the use of calcium hydroxide is desirable.

[0025] The following procedure may be used to make 1.2 - 1.5 N AGIlS (or ACS). An amount of 1055 ml ( 19.2 moles, after purity adjustment and taking into account the amount of acid neutralized by base) of concentrated sulfuric acid (FCC Grade, 95-98% purity) is slowly added with stirring, to 16.868 L of RO/DI water in each of reaction flasks a, b, c, e, and f. The amount of water is adjusted to allow for the volume of acid and the calcium hydroxide slurry. The mixture in each flask is mixed thoroughly. Each of the reaction flasks is chilled in an ice bath until the temperature of the mixture in the reaction flask is about 8- 12°C. The mixture is continuously stirred at a rate of about 700 rpm.

[0026] Separately, a slurry is made by adding RO/DI water to 4 kg of calcium hydroxide (FCC Grace, 98% purity) making a final volume of 8 L. The mole ratio of calcium hydroxide to concentrated sulfuric acid is 0.45 to 1. The slurry is a 50% (W/V) mixture of calcium hydroxide in water. The slurry is mixed well with a high-shear-force mixer until the slurry appears uniform. The slurry is then chilled to about 8-12⁰C in an ice bath and continuously stirred at about 700 rpm.

[0027] To each of the reaction flasks is added 150 ml of the calcium hydroxide slurry every 20 minutes until 1.276 L (i.e. 638 g dry weight, 8.61 moles, of calcium hydroxide) of the slurry has been added to each reaction vessel. The addition is again accompanied by mixing at about 700 rpm. After the completion of the addition of the calcium hydroxide to the reaction mixture in each reaction vessel, the mixture is filtered through a 5- micron filter. The filtrate is allowed to sit for 12 hours, then the clear solution is decanted to discard any precipitate formed. The resulting product is AGIIS (or ACS) having an acid normality of 1 .2-1.5.

[0028 ] Other acids or acidulants can preferably be organic acids or salts, or formic acid, calcium formate, diformate, acetic acid, diacetate, calcium acetate, lactic acid, calcium lactate, propionic acid, calcium propionate, citric acid, calcium citrate, fumaric acid, calcium fumarate, malic acid, calcium malate, or products of lactobacillus fermentation. Group 1 and II salts are suitable, but calcium is the most preferred. Another example of a preferred acidulant is highly acidic metalated organic acid ("HAMO") as it is described in U.S. Patent Nos. 6,881 ,424, 6,808,730, and 6,572,908, all of which are hereby incorporated by reference.

[0029] Broadly, one way HAMO can be prepared is by mixing the following ingredients: (1) at least one regenerating acid; (2) at least one metal base; and (3) at least one organic acid, wherein the equivalent amount of the regenerating acid is in excess of the equivalent amount of the metal base. The equivalent amount of the metal base should be about equal to that of the organic acid. Instead of using a metal base and an organic acid, a metal salt of the organic acid can be used in place of the metal base and the organic acid. The insoluble solid is removed by any conventional method, such as sedimentation, filtration, or centrifugation.

[0030] Generally, HAMO can be prepared by blending or mixing the necessary ingredients in at least the following manners:

1. Regenerating acid + (metal base + organic acid);

2. Regenerating acid + (metal base + salt of organic acid);

3. (Regenerating acid + salt of organic acid) + base; and 4. Regenerating acid + salt of organic acid.

[0031 ] The parenthesis in the above scheme denotes "pre-mixing" the two ingredients recited in the parenthesis. Normally, the regenerating acid is added last to generate the HAMO. Although each of the reagents is listed as a single reagent, optionally, more than one single reagent, such as more than one regenerating acid or organic acid, can be used in the current invention. The number of equivalents of the regenerating acid must be larger than the number of equivalents of the metal base, or those of the metal salt of the organic acid. When the organic acid is an amino acid, which, by definition contains at least one amino group, then the number of equivalents of the regenerating acid must be larger than the total number of equivalents of the metal base, or metal salt of the organic acid, and the "base" amino group of the amino acid. Thus, the resultant highly acidic metalated organic acid is different from, and not, a buffer.

[0032] Λs used herein, a regenerating acid is an acid that will "re-generate" the organic acid from its salt. Examples of a regenerating acid include a strong binary acid, a strong oxyacid, and others. A binary acid is an acid in which protons are directly bound to a central atom, that is (central atom)-H. Examples of a binary acid include HF, HCI, HBr, HI, H₂S and HN3. An oxyacid is an acid in which the acidic protons are bound to oxygen, which in turn is bound to a central atom, that is (central atom)-O-H. Examples of oxyacid include acids having Cl, Br, Cr, As, Ge, Te, P, B, As, I, S, Se, Sn, Te, N, Mo, W, or Mn as the central atom. Some examples include H₂SO₄, HNO₃, H₂SeO₄, HClO₄, H₃PO₄, and HMnO₄. Some of the acids (e.g. HMnO₄)cannot actually be isolated as such, but occur only in the form of their dilute solutions, anions, and salts. A "strong oxyacid" is an oxyacid which at a concentration of 1 molar in water gives a concentration Of H₃O⁺ greater than about 0.8 molar.

[0033] The regenerating acid can also be an acidic solution of sparingly-soluble Group II A complexes ("AGI IS'^').

[0034] The clay material can be any suitable clay, including an attapulgite type clay, such as Fullers Earth clay (CAS No. 8031 -18-3), at about 80-100%. Other preferred clays include montmorillonite clay, such as bentonite clay (CAS No. 1302-78-9), or a hydrated sodium calcium aluminosilicate ("HSCAS") clay, which is preferably about 300 MESH. Additional clays include an amorphous opaline silica clay, including silica hydrated, or silica sand (CAS No. 7631-86-9) at about 80-100%, or crystalline silica clay, including quartz (CAS No. 14808-60-7) at about 0-20%. Preferred clays include phyllosilicate, Florisil®, bayerite, pseudoboehmite, alumina, silica gel, aluminum oxides, gibbisite, boehmite, and bauxite. Mixtures of different clays can also be used.

[0035] The use of clay material is also effective for control of mycotoxins such as aflatoxins. Clay has been shown to absorb mycotoxins when added to animal feed. When adding an acidified clay material to feed for the purpose of controlling mycotoxins, it should preferably be added at about 1-2%. A preferred clay is a HSCAS clay such as NovaSil Plus™ (hydrated sodium calcium aluminosilicate produced by Engelhard Corporation and available from Trouw Nutrition, USA) that can be used to "absorb" and inactivate aflatoxin. See, U.S. Patent No. 5, 178,832 to Timothy D. Phillips, et al.; U.S. Patent No. 5, 165,946 to Dennis R. Taylor, et al.; and K. Pimpukdee, Feed & Livestock, pages 40-43, December 2003/January 2004, the content of each of which is hereby incorporated by reference.

[0036] The appearance of NovaSil Plus™ is off white to tan colored and is a free flowing powder. The free moisture content is about 9%. The loose bulk density is 0.64 g/cc; the packed bulk density is about 0.80 g/cc; and the particle size distribution is about 5% of +100 mesh, 18% of +200 mesh, and 60% of -325 mesh. Chemical analysis showed that %CaO is between 3.2-4.8; % MgO is between 4.0-5.4; %Fe₂O₃ is between 5.4-6.5; % K₂O is between 0.50-0.90: %Na₂O is between 0.10-0.30; %MnO is between 0.01 -0.03; % Al₂O₃ is between 14.8-18.2; and %SiO₂ is between 62.4-73.5. Content of traces of heavy metals is as follows: Pb. 6.0-6.5 ppm; As, 0.5-0.7 ppm; Cd. 0.2-0.4 ppm; Cr, 5.5-6.0 ppm, and Hg, less than 0.1 ppm. The clay is substantially free from dioxins (dioxin as used here refers to the toxic contaminant 2,3,7,8-tetrachlorodibenzodioxin ("TCDD") which is used as an index of the presence of dioxins in food ingredient) in NovaSil Plus above the detection limit of 0.33 parts per trillion ("ppt").

[0037] A preferred desiccant is silicon dioxide or silica gel (CAS # 1 12926-00-8). Any suitable clay desiccant, including montmorillonite clay and silica clay, can also be used. The clay material of the composition itself functions as a desiccant.

[0038] A preferred mixture of acidulant blended with clay material contains about 25% by weight of 10 N ACS and about 5% by weight of a clay material mixture including

Fullers earth clay (80- 100%), montmorillonite clay (80- 100%), silica hydrate clay (80- 100%), and quartz clay (0-20%) (VITROXAL, Mionix Corporation, Round Rock. TX), Another preferred mixture of acidulant blended with clay material contains about 50% by weight of 5 N ACS, about 35% by weight of the desiccant silicon dioxide, about 10% by weight of montmorillonite clay (preferably of Ecuador), and about 5% by weight of other optional ingredients, such as dyes (VITROX, Mionix Corporation, Round Rock, TX).

[0039] It is preferable to use the acidifier, such as ACS or HAMO, to lower the pH of the feed to a pH of about 4.5 to about 5.0. Preventing histamine production as a result of bacterial replication can require acidulating water in a holding tank to a pH of about 4.5 and adjusting it depending on the number of fish to be held. ACS or another acidifier can accomplish this task.

[0040] In some examples, the feed material, or fish or shrimp feed, might comprise 25% fishmeal, 14% bran, 26% corn, and 35% soybean, by weight. If dry ACS is added in an amount of about 4 kilograms per ton, or 200 g per 40 kg bag, the pH of the feed will be approximately 4.8 - 5.0. Fish feed is usually an extruded feed compared to pelleted feed for shrimp. Fish feed must float because fish raised in ponds or jails have to eat at the surface of the water. Shrimp usually eat from the middle to the bottom of the pool.

[0041] The modified feed composition is prepared by first slowly blending a preferred acidulant with a preferred clay. The weight ratio of the acid to the clay can vary widely, depending on the type of clay and acid used. The normal ratio can vary from 50 : 1 to 1 : 5. Preferably, the weight ratio of the acid to the clay ranges from 1 : I to 1 : 1.25. In some instances, the resultant acidified clay appears as lumps of solid, which can then be ground to powder form. After the completion of blending, the resultant acidified clay is allowed to dry, preferably at ambient temperature.

[0042] The acidified clays should preferably be in the form of substantially free flowing powder, and of relatively uniform fine or small particle sizes, and hence capable of being uniformly applied to, and mixed with, foods or feeds. Thus, the clay, because of its structure, has selective affinities to various mycotoxins, such as aflatoxins. The acid used to acidify the acidified clay should not significantly alter or destroy the structure of the clay to cause the clay to lose its selective affinities to various mycotoxins. The relatively fine particle size clay also function as a carrier, or medium, for the acid absorbed or adsorbed therein. The desiccant can be added as needed to the modified feed composition. [0043] The acidulant can be added directly to the shrimp or fish feed, in the absence of clay, by direct injection of the acidulant into a cooker or extruder device used during the preparation of the feed. This method allows the pH of the feed to be lowered directly to about 4.5 to 5.0. If clay is also included, the acidulant is first blended with the clay as described above and then mixed with the feed material. For example, the acidified clay plus the shrimp or fish feed can be extruded by running the mixture through a cooker or extruder device, or pelletized by running the mixture through a pellet mill.

EXAMPLE l. CLAY MATERIAL

|0044] Blending clay material into the modified aqua feed composition is effective at not only suppressing and controlling microbial growth, but may also assist with adsorption of aflatoxins. Adsorbent clay minerals have been reported to bind aflatoxin Bi in liquids. In the first enterosorbent study with aflatoxins, a calcium montmorillonite clay that is commonly used as an anti-caking additive for animal feeds has been shown to significantly sorb aflatoxin B₁ with high affinity and high capacity in aqueous solutions and to protect broiler and Leghorn chicks from the toxic effects of 7,500 ppb aflatoxins in the diet. Since this initial study, calcium montmorillonite clay and other similar montmorillonite clays have been reported to diminish the toxic effects of aflatoxins in a variety of young animals including rodents, chicks, turkey poults, ducklings, lambs, mink and pigs. Clay in the diet has also been shown to diminish levels of aflatoxin Mi in milk. More recently, urinary biomarkers of AfBi exposure in dogs were reduced by the inclusion of calcium montmorillonite clay. Thus, CAS at 0.5% (w/w) in the feed protected against the adverse effects of 7,500 ppb aflatoxins in the same feed. This high aflatoxin level would not normally be found as a contaminant of food or feed and (as such) represents a "worst case scenario." Aflatoxin protection can be shown at levels as low as 0.25% (w/w) in the diet. Extrapolating to the human, an approximate 3g dose of CAS in a capsule would approximate the 0.25% level in food based on food intake data in Ghana and a body weight of 70 Kg.

[0045] GRAS Status and Safety Studies for in vivo Use of Clay. One of ordinary skill in the art would be aware that scientific publications support the use of calcium montmorillonite clay as an aflatoxin binding agent in animal feeds. A compilation of various in vivo studies involving calcium montmorillonite clay in multiple animal species is described herein (see Figure 2). For example, hydrated sodium calcium aluminosilicate is generally recognized as safe for use in feeds at a level not exceeding 2 percent in accordance with good manufacturing or feeding practice.

[0046] In animal studies with calcium montmorillonite clay, no adverse effects from clay treatment, at levels up to 2.0% in the diet, have been reported. In recent studies in rodents, montmorillonite clays were evaluated for potential toxicity and trace metal bioavailability in pregnant Sprague-Dawley rats throughout the period of gestation following high level exposure in the diet (2.0% w/w). Clays were supplemented in the balanced diet of Sprague-Dawley rats during pregnancy at a level of 2.0% (w/w). Evaluations of toxicity were performed on gestation day 16 and included maternal body weights, maternal feed intakes, litter weights, in addition to embryonic resorptions. Liver and, kidneys, tibia, brain, uterus, pooled placental, and pooled embryonic mass were collected and weighed. Tissue were lyophilized and neutron activation analysis (NAA) was then performed. Elements considered by NAA included: Al, Br, Ca, Ce, Co, Cr, Cs, Cu, Dy, Eu, Fe, Hf K, La, Lu, Mg, Mn, Na, Nd, Ni, Rb, S, Sb, Sc, Se, Sm, Sr, Ta, Th, Te, Th, Ti, Tl, U, V, Yb, Zn, and Zr. Inductively coupled plasma-mass spectroscopy further confined that Al was below detection limits (0.5ppm) in the brain, indicating no significant bioavailability of this metal from clay interactions in the GI tract. Animals supplemented with either clay were similar to controls with respect to toxicity evaluations and metal analysis, with the exception of decreased brain Rb following clay supplementation. Overall, the results of this study suggest that neither clay at high dietary concentrations, result in overt toxicity or influence mineral uptake or utilization in the pregnant rat. In some embodiments, clay was selected for testing due to its GRAS status and its purity priority trace metals and dioxin levels, see Figure 3.

[0047] Other studies in rodents and subjects have confirmed the safety calcium montmorillonite clay for application in human diets. In the rodent study, rats were fed rations containing about 0, 0.25, 0.5, 1 .0, and 2.0% levels of calcium montmorillonite clay. Body weights, body weight gain, organ weights, histopathology, plasma biochemistry, serum vitamins A and E and micronutrients (Fe and Zn) were measured, standardized and compared to determine toxicity and any interactions of clay with critical nutrients at the end of the study. After 6 months exposure to clay, no morbidity or mortality was observed among treatment groups. There were no changes in the major organs, serum biochemistry or micronutrient levels. The ratios of organ weight to final body weight for the liver, kidneys, lungs, heart, brain, spleen, and tibia among the treatment groups in each sex were not significantly different histopathological analysis of the liver and kidneys indicated no differences between controls and clay treatments. These results suggest that inclusion of clay at levels less than 2.0 % (w/w) in the diet should not result in overt toxicity and can be used safely to reduce exposure aflatoxins in the gastrointestinal tract. In the human study, Calcium montmorillonite clay was initially tested for trace metals and dioxin content in order to confirm the composition of matter and ensure low levels of contamination.

[0048J Calcium montmorillonite clay was then heat sterilized and packed into capsules for use in the study. The study design was based on 2 treatment groups: 1) low dose- 3 x 500 mg capsules x 3 times/day for a total of 2 weeks, and 2) high dose- 3 x 1,000 mg capsules x 3 times/day for a total of 2 weeks. The 2-week trial consisted of 50 healthy adults, age 22-40 selected by initial physical exams, laboratory analysis of biological fluids and questionnaire. One of ordinary skill in the art would be able to make capsules that are modified from the above description, that varied in dose, see Remmington's Pharmaceutical Sciences 17^th Edition. Participants were then given clay capsules before meals with a bottle of spring water. Medical personnel were onsite to monitor any complaints or adverse effects. Blood and urine samples were taken at the end of the 2 week period and laboratory analysis and physical examinations were administered again. Any adverse events were reported according to NlH guidelines. Compliance with the dosing protocol reached 100% over the two-week study period. Analysis of clinical and biochemical data for side effects monitoring, blood, and urine parameters for liver and kidney function did not show any specific adverse effects.

[0049] Mode of Action and Mechanistic Studies. Several in vitro studies have assessed the sorption of aflatoxins onto the surface of hydrated sodium calcium aluminosilicate clay (HSCAS). HSCAS, in aqueous solution, has been shown to tightly and preferentially sorb AfB] and similar analogs of aflatoxin B₁ (AfBι) that contain an intact β- dicarbonyl system in their molecular structure. Isothermal analysis of AfBi sorption to HSCAS indicated both high affinity and high capacity characteristics and also suggested that different sites and/or mechanisms of action may be involved in AfBj sorption at clay surfaces. The enthalpy of AfBi sorption (near -40 kJ/mol) showed some variation, suggesting multiple sites on HSCAS with dissimilar thermodynamic properties. These findings indicate that multiple sites on the surface of HSCASs may act to chemisorb AfBi and that the optimal orientation of the AfBi molecule is most likely planar on interlaycr clay surfaces. Functional groups on aflatoxin analogs may sterically hinder sorption at the surface of HSCAS or may block sorption by interacting across the interlayer region. Other mechanisms of AfB] sorption to HSCAS surfaces may involve the potential chelation of predominant interlayer cations such as calcium and various other edge-site metals.

[0050] Ingredient Description and Profile. Calcium aluminosilicate clay (CAS) has a different composition from hydrated sodium calcium aluminosilicate (HSCAS) clay, which has a dark tan color. The CAS has the appearance of an offwhite to gray-greenish colored free flowing powder. The CAS is odorless having a specific gravity of about 2.4. The isolated CAS is negligibly soluble in water and has a pH in the range of about 5-9. Due to the silica and aluminum silicate components, the isolated CAS may have some adverse effects if dry particles are inhaled, but no adverse health effects are suspected from ingestion. The typical values are as follows:

Typical Physical Properties:

Free Moisture (LOD) 9%

Loose Bulk Density 0.64 g/cc 40 Ibs/ft3

Packed Bulk Density 0.80 g/cc 50 lbs/ft-3

Particle Size Distribution: 5% +100 mesh

18% +200 mesh

60% +325 mesh

Typical Chemical Analysis:

Chemical Analysis by %CaO 3.2 - 4.8

X-Ray Fractionation (XRF) %MgO 4.0 - 5.4

Spectroscopy (weight %): %Fe₂0₃ 5.4 - 6.5

%K₂0 0.50 - 0.90

%Na₂0 0.10 - 0.30 %MnO 0.01 - 0.03

%A1₂O₃ 14.8 - 18.2

%SiO₂ 62.4 - 73.5

[0051] Additionally, testing of the processed clay products from Engelhard's, Jackson, MS plant have confirmed low levels of TCDD in CAS (< 0.33 parts per trillion, ppt). TCDD is given in Engelhard specifications as an index of the presence of dioxins in food ingredients.

[0052] Analytical Procedures and Methods for Isothermal Adsorption Analysis. Isothermal Adsorption analysis was performed using a stock solution of aflatoxin Bi (AfBi) is prepared by dissolving pure AfBi crystals (Sigma Chemical Co., St. Louis, MO) in acetonitrile. A volume of the stock solution is then injected into purified (deionized) water, yielding an 8 μg/mL working solution of AfBi. The working solution's concentration is then verified with a UV-vis spectrophotometer (λ_max ⁼ 362 nm; ε = 21.865). The batch isotherm procedure entails the exposure of samples containing 100 μg of sorbent to an increasing concentration of solute (AfBj)(OA 0.8, 1 .6, 2.4, 3.2, 4, 4.8, 6, 6.4, 7.2, and 8 μg/mL). This study uses three replicates at each solute concentration. The solute concentration is achieved by adding an appropriate amount of working AfBj solution to sterile 17 x 100 mm polypropylene centrifuge test tubes and then adding a complementary amount of purified water to bring the total volume to 5 mL/tube. Approximately 10 mg of sorbent is weighed in a 16 x 125 mm disposable borosilicate glass test tube, and purified water is added to the sorbent to make a 2 mg/mL suspension. This sorbent/water suspension is vortexed for 3s before each 50 μL transfer to each replicate by an autopipetter. The mixing is repeated before each transfer. Along with the samples, there are three controls consisting of 5 mL of purified water, 5 mL of AfBi working solution without sorbent, and 5 mL of the lowest concentration of AfBi without sorbent. The samples and controls are capped and placed on an electric orbital shaker at 1,000 rpm for 24 h in an incubator at either 15, 25, or 37 ⁰C. After shaking, the samples are centrifuged at 10,000 rpm for 15 min at the same temperature that the shaking occurred. The UV-vis absorption of AfBj remaining in the supernatant from the samples and controls is measured with a spectrophotometer. At the highest AfB] concentration level, the supernatant is saved for analysis by HPLC to check for any degradation compounds since the adsorption calculations are dependent on a different calculation.

[00531 Data Calculations and Curve Fitting. The UV-vis absorption data are used to calculate the amount of AfB| left in solution and the amount adsorbed for each data point. Using TableCurve 2D software (Systat Software Inc., Richmond, CA) these data are fit to the Langmuir isotherm equation:

[0055] where q is the amount of AfB] adsorbed, Q_max is the maximum amount of AfBi adsorbed, C_w is the equilibrium concentration of AfBi in solution and K_d is the distribution constant. The Langmuir equation is entered into the TableCurve 2D program as a user-defined function and has limits and first approximations for variable parameters. The parameter limits for Q_max are positive numbers ranging from 0 to a maximum of 1 mol/kg. Parameter limits for Kj range from 0 to 1 x 10²⁵. Starting estimates for the parameters Q_max and Kd are determined by TableCurve 2D. After these values are entered into the Langmuir user-defined function in TableCurve 2D, the data is fit, and theoretical values for Q_max and K_d are obtained. The AH_ad_X (enthalpy of adsorption) is calculated by comparing the individual Kj values at 15, 25, and 37 ⁰C by the equation:

ΔHads =

(1/Γ₂) - (1/Γ,)

The definition of Kt is derived by solving for K_d from the Langmuir equation giving:

K_d = q/(Q_mm - q)C_w

The Q_max is taken from the fit of Langmuir equation to the adsorption data at 15, 25, and 37°C.

[0056] Methods for COLE Index. A measure of expansive properties, the coefficient of linear extensibility (COLE) index is the ratio of the volume of a soil after wetting to the volume of soil before wetting minus one. COLE = (volume of clay after wetting/volume of clay before wetting) - 1 COLE index values greater than 0.03 indicate that significant smectite (swelling clay) is present in the sample. The general procedure can be summarized as follows:

1. Λdd 5 niL (5 cm ) of dry clay to a 25 rnL graduated cylinder.

2. Add distilled water to the clay bringing the total volume to 25 mL.

3. Shake or stir suspension vigorously to ensure thorough wetting of clay.

4. Allow suspension to stand for 24 hr. at room temperature.

5. Measure the expanded volume of settled clay.

[0057] Shrink-swell potential correlates closely with the kind and amount of clay. The greatest shrink-swell potential occurs in soils that have high amounts of 2: 1 lattice clays, such as smectites. Illitic clays are intermediate, and kaolinitic clays are least affected by volume change as the content in moisture changes. Adsorption isotheπns of regular vs. collapsed HSCAS at 25⁰C are shown in Figure 4.

EXAMPLE 2. COMPARISON OF ANTIMICROBIAL EFFECTS OF ACS AND BENZYLKONIUM CHLORIDE/UREA

[0058] A study was conducted to determine if ACS added to shrimp feed could effectively suppress and control infection of shrimp by Vibrio spp. and Pseudomonas spp.

[0059] To conduct the experiment, two premixes were prepared. The first premix, or "Premix A" contained one kg of ACS 50 (Mionix Corporation), which was blended with 0.7 kilograms of 300 MESH hydrated sodium calcium aluminosilicate (HSCAS) clay, i.e., a montmorillonite clay, and 0.3 kg of silica gel (CAS #1 12926-00-8). "Premix B" contained one kg of Timsen™ (N-Alkyl dimethyl benzyl ammonium chloride, or ADBAC), which is a quaternary ammonium salt mix comprised of 40% benzylkonium chloride and 60% urea and is also considered to be GRAS (i.e., general recognized as safe).

[0060] Shrimp feed was then prepared and mixed with the premixes. One metric ton of shrimp feed comprised of 25% fishmeal, 14% bran. 26% corn and 35% soybean was blended with 3 kg of the "Premix A" and molasses was added to facilitate pellet formation. Additionally, one metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 4 kg of "Premix B." [0061] Six shrimp ponds stocked with Peneaus vannamei (white shrimp) that were beginning to exhibit signs of Vibrio and Pseudomonas spp. infection were tested. Under normal circumstances the ponds were treated with shrimp feed blended with Premix B. Feed was added three times per day: 30% in the early morning, 50% at noon and 20% before the evening, respectively. The benzylkonium chloride/urea salt mix (Timsen^1M) product was added to the feed in the amount of 4 kg per ton during the disease stage (i.e., "Premix B").

[0062] For trial purposes one of the six ponds was set aside and treated with shrimp feed containing the ACS 50 premix, i.e. "Premix A." The five remaining ponds were treated with shrimp feed containing "Premix B," as defined above.

[0063] Previous studies showed treatment with feed supplemented with "Premix B" would bring vibriosis under control by seven days during shrimp grow-out. However, circumstances being what they were during this trial vibriosis was suppressed in all ponds by day 5 post-treatment. In other words, the ACS 50 was equally effective as the benzylkonium chloride/urea salt mix in bringing about control of vibrio and/or pseudomonas infection.

EXAMPLE 3. COMPARISON OF ANTIMICROBIAL EFFECTS OF ACS AND

OXYTETRACYCLINE

[0064] A study was conducted to determine if ACS added to shrimp feed could effectively suppress and control infection of shrimp by Vibrio spp. as compared to oxytetracycline, a widely used antibiotic.

[0065] To conduct the experiment, two premixes were prepared. The first premix, or "Premix A" contained one kg of ACS 50 (Mionix Corporation), which was blended with 0.7 kilograms of 300 MESH hydrated sodium calcium aluminosilicate (HSCAS) clay, i.e., a montmorillonite clay, and 0.3 kg of silica gel (CAS #1 12926-00-8). "Premix B" contained one kg of Oxytetracycline chlorohydrate (HCl) formula C₂₂H₂₄N₂O₉J ICl (CAS #79-57-2), a broad-spectrum antibiotic at 50% active ingredient in dry form, having the structure shown below:

[0066] Shrimp feed was then prepared and mixed with the premixes. One metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 4 kg of '"Premix A" and fish oil was added to facilitate pellet formation and binding. Additionally, one metric ton of shrimp feed comprised of 25% fishmeal. 14% bran, 26% corn and 35% soybean was blended with Oxytetracycline HCl (50% active) in the amount of 7.5 kg of Premix B and fish oil was added to facilitate pellet binding.

[0067] Eight ponds stocked with Peneaus vannamei (white shrimp) that were 45 days into the grow-out cycle when signs of vibriosis were detected were utilized in this experiment. Shrimp feed supplemented with ACS 50 ("Premix A") was applied to two ponds and shrimp feed blended with Oxytetracycline ("Premix B") was applied to six ponds, respectively.

[0068] After two days of application of treated shrimp feed, further development of vibriosis ceased in all ponds. However, notably, recovery was delayed in the oxytetracycline ponds. Importantly, shrimp recovery in the ACS 50 treated ponds occurred on day 5 post-treatment, whereas recovery did not occur until day 7 post-treatment in ponds treated with oxytetracycline.

[0069] In addition to more rapid recovery from infection with vibrio bacteria, it was noted that shrimp in the ACS 50 treated ponds initiated feed consumption more rapidly than the shrimp in ponds treated with Oxytetracycline ponds. The ACS-50 treated shrimp also exhibited reduced erratic movements consistent with disease stress due to bacterial infection and immune suppression. By day 10 after medical treatments, both groups were healthy and a normal weekly growth and feed pattern was observed.

[0070] The results show that ACS 50 added to shrimp feed is more effective at suppressing vibriosis in a timely manner compared to standard treatment of shrimp with oxytetracyline. Shrimp recovered two days earlier in ACS-50 treated ponds and returned to normal feed consumption and growth patterns more rapidly. Less erratic movements due to bacterial infection stress was also noted. The beneficial aspects are several: (1) Higher shrimp yield at harvest, (2) Greater return on investment, and (3) No development of antibiotic resistance due to selection of oxytetracycline resistant Vibrio spp.

EXAMPLE 4. COMPARISON OF ANTIMICROBIAL EFFECTS OF ACS AND

BENZYLKONIUM CHLORIDE

[0071] A study was conducted to determine if ACS added to shrimp feed could effectively suppress and control infection of shrimp by Vibrio spp.

[0072] To conduct the experiment, two premixes were prepared. The first premix. or "Premix A" contained one kg of ACS 50 (Mionix Corporation), which was blended with 0.7 kilograms of 300 MESH hydrated sodium calcium aluminosilicate (HSCAS) clay, i.e., a montmorillonite clay, and 0.3 kg of silica gel (CAS #1 12926-00-8). "Premix B'' contained a quaternary ammonium salt mix comprised of 60% benzylkonium chloride (N-Alkyl dimethyl benzyl ammonium chloride, or ADBAC).

[0073] Shrimp feed was then prepared and mixed with the premixes. One metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 3.75 kg of "Premix A" and molasses was added to facilitate pellet formation. Additionally, one metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 3.75 kg of "Premix B."

[0074] Four shrimp ponds stocked with Peneaus vannamei (white shrimp) that were beginning to exhibit signs of vibriosis infection 30 days into grow-out were utilized in this study. Under normal circumstances the ponds were treated with shrimp feed blended with Premix B. Feed was added three times per day: 30% in the early morning, 50% at noon and 20% before evening, respectively. For this study one of four ponds was elected for treatment with shrimp feed blended with "Premix A." The three remaining ponds were treated with shrimp feed blended with "Premix B."

[0075] A positive response to treatment was noted on day three post-treatment for the pond treated with ACS-50, whereas the first signs of infection control for ponds treated with the quaternary ammonium salt were not detected until day seven post-treatment. At day 10 post-treatment both groups were healthier and showed a complete disease recovery. [0076] The results indicate that ACS 50 is more effective than commonly used quaternary ammonium salts for the control of vibriosis.

EXAMPLE 5. PREPARATION OF FEED FORTIFIED WITH FERMENTATION

PRODUCTS

[0077] Fish and shrimp feed can also be fortified with fermentation products to control and suppress infection by Vibrio spp. and Pseudomonas spp. Shrimp feed fortified with naturally fermented products can effectively suppress and control infection of shrimp by Vibrio spp. and Pseudomonas spp. during the 120-day grow-out cycle.

[0078] To prepare the fortified shrimp feed, the fortification culture media was first prepared. 1,000 liters of pond water was filtered (50 microns) into a light opaque tank. 30 kg of cane molasses and 2 kg of soy meal (FCC grade) were added, then the mixture was blended. 200 g of microbial culture, consisting of a mixture of Lactobacillus spp., bacillus spp. and yeast, was then added. Then, to culture, the mixture was fermented under anaerobic conditions, controlling exposure to sunlight and insects for 48 to 72 hours at ambient temperature. When the pH of the fermentation product reached 4 to 4.5, it was considered ready to use.

[0079] To fortify the shrimp feed with the fermentation product, one metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 175 liters of the fortification supplement and molasses to facilitate pellet formation.

EXAMPLE 6. PREPARATION OF FEED FORTIFIED WITH FERMENTATION

PRODUCTS AND pH ADJUSTED

[0080] Fish and shrimp feed can be fortified with fermentation products and pH adjusted to control and suppress infection by Vibrio spp. and Pseudomonas spp.

[0081] To prepare the fortified shrimp feed, the fortification culture media was first prepared. 1.000 liters of pond water was filtered (50 microns) into a light opaque tank. 30 kg of cane molasses and 2 kg of soy meal (FCC grade) were added, then the mixture was blended. 200 g of microbial culture, consisting of a mixture of lactobacillus spp., bacillus spp. and yeast, was then added. Then, to culture, the mixture was fermented under anaerobic conditions, controlling exposure to sunlight and insects for 48 to 72 hours at ambient temperature. When the pH of the fermentation product reached 4 to 4.5, it was considered ready to use.

[0082] The fermentation product was then pH adjusted. 200 liters of fortification supplement was transferred to a polypropylene or HDPE container. 2 liters of ACS-50 was added to the 200 liters of fortification supplement and blended accordingly. The pH was then checked and adjusted to a desirable level. Preferably, to protect the pH probe, 1 ml of the blend should be diluted into 100 ml of water before inserting probe. If the pH is 4.0, then the real pH is 2.0. ACS-50 can be added in small increments and the pH checked accordingly until the desired pH is reached.

[0083] To fortify the shrimp feed with the pH adjusted fermentation product, one metric ton of shrimp feed comprised of 25% fishmeal, 14% bran, 26% corn and 35% soybean was blended with 175 liters of the fortification supplement and molasses to facilitate pellet formation.

EXAMPLE 7. COMPARISON OF EFFICACY RELATIVE TO OXYTETRACYCLINE

[0084] The comparative effects of treatment with oxytetracyline ("OTC") and treatment with an example of the composition, both used to modify shrimp feed, was performed. The composition was a mixture of acidulant blended with clay material that contained about 50% by weight of 5 N ACS, about 35% by weight of the desiccant silicon dioxide, about 10% by weight of montmorillonite clay (preferably of Ecuador), and about 5% by weight of other optional ingredients, such as dyes (VITROX, Mionix Corporation, Round Rock, TX). Ten ponds on a farm were selected for this assessment. Each of the ten test ponds was stocked at the same time with the same number of shrimp and were evenly divided between the two treatments. Either OTC at about 0.8 - 1 % of the total weight of the feed or the composition ('"COMP") at about 1 - 1.2% of the total weight of the feed were added to shrimp feed and fed to shrimp at least four times daily for 60 days. At least three distinct pathogen types were noted as being present in the ponds, including vibrio bacteria, hepatopancreatic rickettsia, and unicellular gregarine gut parasites. The average weight of the shrimp in each pond was measured as well as the percent survival rate. Finally, the days to the onset of disease caused by one or more of the pathogen types present in the pond, as well as the days to recovery from the onset of disease, were measured. The results are shown in Table 1 below.

Table 1. Comparison of the effects of treatment with OTC vs. COMP on shrimp

Test Parameter OTC COMP

Average weight/shrimp (g) 9.2 9.7

Average % survival 70 75

Days to onset of disease 28-32 38-42

Days to recovery from onset of disease 10 8

[0085] From a gross perspective, shrimp present in ponds treated with OTC versus the composition exhibited similar disease and recovery patterns. However, laboratory analysis showed there was a distinct difference in terms of recovery. Namely, the shrimp hepatopancreas and gi tract recovered more rapidly in the shrimp treated with the composition. These results are reflected in the parameters defined in Table 1. The shrimp on average grew to a larger size, survival was increased significantly, disease onset was delayed and recovery from infection was accelerated. The composition is a superior product to OTC and leaves no antibiotic residue. Moreover, from the producer's perspective, survivability and increased weight gain mean higher yields and therefore a significantly greater return on investment.

EXAMPLE 8. COMPARISON OF EFFICACY RELATIVE TO OXYTETRACYCLINE

[0086] In an additional case study, six ponds on a different faπn were also selected for a study of the comparative effects of treatment with oxytetracycline (OTC) versus an example of the current composition. The composition was a mixture of acidulant blended with clay material that contained about 50% by weight of 5 N ACS, about 35% by weight of the desiccant silicon dioxide, about 10% by weight of montmorillonite clay (preferably of Ecuador), and about 5% by weight of other optional ingredients, such as dyes (VITROX, Mionix Corporation, Round Rock, TX). The farm was totally separate and under the control of a different farm group from that presented in Example 7. The ponds were stocked at the same time with the same number of shrimp and were evenly divided between the two treatments. Oxytetracycline at about 1 % of the total weight of the feed and the composition at about 1.2% of the total weight of the feed were added to shrimp feed and fed to shrimp at least four times daily for 60 days. The survival percentage, weight, and average weight per shrimp were measured and the results are shown in Tables 2 and 3 below. Table 2. Ponds Treated with Composition

[0087] As can be seen from Tables 2 and 3, there is a very significant difference in the survivability of shrimp in ponds treated with the composition versus OTC. On average, survivability was increased by about 16% in ponds treated with the composition. The difference in weight gain was insignificant. Due to the approximately 16% improvement in survivability, the shrimp treated with the current composition had an overall net gain in total shrimp weight approaching 20%. This is a very significant result.

REFERENCES CITED

The entire content of each of the following documents is hereby incorporated by reference.

U.S. PATENT DOCUMENTS

U.S. Patents

U.S. Patent No. 5165946 Taylor, et al.

U.S. Patent No. 5178832 Phillips, et ai.

U.S. Patent No. 6436891 Kemp et al.

U.S. Patent No. 6572908 Kemp et al.

U.S. Patent No. 6808730 Kemp et al.

U.S. Patent No. 6881424 Kemp et al.

U.S. Patent No. 6902753 Kemp et al.

OTHER PUBLICATIONS

Luckstadt, C, "Acidifiers in Aqua Feeds: A Solution for Antibiotic-Free Feeding of Fish and Shrimp," Aqua Feeds: Formulation & Beyond, Vol. 3, No. 4, 2006, pp. 4-6.

Luckstadt, C, "Acidifiers in aquaculture prove beneficial," Feed Mix, Vol. 14, No. 3, 2006, pp. 1 1-12.

Pimpukdee, K., Feed & Livestock, pages 40-43, December 2003/January 2004.

Claims

What is claimed is:

1. A modified fish or shrimp feed composition to suppress and control infection offish or shrimp by bacteria, comprising: a fish or shrimp feed material; an acidulant; and a clay material.

2. The composition of claim 1, wherein the acidulant is ACS.

3. The composition of claim 2, wherein the ACS has a noπnality of about 5-10 N.

4. The composition of claim 1 , further comprising a desiccant.

5. The composition of claim 1, wherein the clay material is attapulgite type clay, montmorillonite clay, amorphous opaline silica clay, crystalline silica clay, or mixtures thereof.

6. The composition of claim 1, wherein the composition has a pH of about 4.5 to about

5.

7. A modified fish or shrimp feed composition to suppress and control infection offish or shrimp by bacteria, comprising: a fish or shrimp feed material; and an acidulant, wherein the acidulant is ACS, wherein the ACS has a normality of about 5-10 N, and wherein the composition has a pH of about 4.5 to about 5.

8. A modified fish or shrimp feed composition to suppress and control infection offish or shrimp by bacteria, comprising: a fish or shrimp feed material; an acidulant, wherein the acidulant is ACS or HAMO; and a clay material.

9. The composition of claim 8, further comprising a desiccant.

10. The composition of claim 8, wherein the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide, or a calcium salt, or a mixture of the two, wherein when the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide then the mole ratio of calcium hydroxide to sulfuric acid ranges from about 0.1 to about 0.5, and wherein the ACS has a pH of less than about 2.

1 1. The composition of claim 8, wherein the acidulant is HAMO, and wherein the HAMO is prepared by mixing at least one regenerating acid, at least one metal base, and at least one organic acid, wherein an equivalent amount of the regenerating acid is in excess of an equivalent amount of the metal base.

12. The composition of claim 8, wherein the clay material is attapulgite type clay, montmorillonite clay, amorphous opaline silica clay, crystalline silica clay, or mixtures thereof.

13. The composition of claim 8, wherein the clay material is hydrated sodium calcium aluminosilicate ("HSCAS") clay.

14. The composition of claim 8. wherein the clay material is hydrated sodium calcium aluminosilicate ("HSCAS") clay, and wherein the clay material has the following properties:

(a) a free moisture content of about 9%,

(b) a loose bulk density of about 0.64 g/cc,

(c) a packed bulk density of about 0.80 g/cc,

(d) a particle size distribution of about 5% of +100 mesh, 18% of +200 mesh, and 60% of -325 mesh,

(e) a chemical analysis showing %CaO between about 3.2-4.8, % MgO between about 4.0-5.4, %Fe₂θ3 between about 5.4-6.5, % K₂O between about 0.50-0.90, %Na₂O between about 0.10-0.30, %MnO between about 0.01-0.03, % Al₂O₃ between about 14.8-18.2, and %SiO₂ between about 62.4-73.5, (f) a content of heavy metals of Pb at about 6.0-6.5 ppm, As at about 0.5-0.7 ppm; Cd at about 0.2-0.4 ppm, Cr at about 5.5-6.0 ppm, and Hg less than 0.1 ppm, and

(g) a 2,3,7,8-tetrachlorodibenzodioxin ("TCDD^'') amount of less than 0.33 parts per trillion ("ppt").

15. The composition of claim 8, wherein the fish or shrimp feed material comprises 25% fishmeal. 14% bran, 26% corn, and 35% soybean by weight.

16. The composition of claim 8, further comprising organic acids or Group I and Group II salts of organic acids.

17. The composition of claim 8, further comprising formic acid, calcium formate, diformate, acetic acid, diacetate, calcium acetate, lactic acid, calcium lactate, propionic acid, calcium propionate, citric acid, calcium citrate, fumaric acid, calcium fumarate, malic acid, calcium malate, or mixures thereof.

18. The composition of claim 8, further comprising products of lactobacillus fermentation.

19. The composition of claim 8, wherein the desiccant is silicon dioxide.

20. The composition of claim 8, wherein the acidulant adjusts the pH of the composition to about 4.5 to about 5.0.

21. A method for preparing a modified fish or shrimp feed composition having antimicrobial properties, comprising: blending an acidulant and a clay material, wherein the acidulant is ACS or

HAMO, to form an acidified clay material; and mixing the acidified clay material and a fish or shrimp feed material.

22. The method of claim 21 , wherein the acidulant is ACS, and wherein the ACS has a normality of about 5-10 N.

23. The method of claim 21, wherein the acidulant is ACS, and wherein the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide, or a calcium salt, or a mixture of the two, wherein when the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide then the mole ratio of calcium hydroxide to sulfuric acid ranges from about 0.1 to about 0.5, and wherein the ACS has a pH of less than about 2.

24. The method of claim 21 , wherein the acidulant is HAMO, and wherein the HAMO is prepared by mixing at least one regenerating acid, at least one metal base, and at least one organic acid, wherein an equivalent amount of the regenerating acid is in excess of an equivalent amount of the metal base.

25. The method of claim 21, wherein the clay material is attapulgite type clay, montmorillonite clay, amorphous opaline silica clay, crystalline silica clay, or mixtures thereof.

26. The method of claim 21, wherein the clay material is hydrated sodium calcium aluminosilicate ("HSCAS") clay.

27. The method of claim 21, wherein the clay material is hydrated sodium calcium aluminosilicate ("HSCAS") clay, and wherein the clay material has the following properties:

(a) a free moisture content of about 9%,

(b) a loose bulk density of about 0.64 g/cc.

(c) a packed bulk density of about 0.80 g/cc,

(d) a particle size distribution of about 5% of +100 mesh, 18% of +200 mesh, and 60% of -325 mesh,

(e) a chemical analysis showing %CaO between about 3.2-4.8, % MgO between about 4.0-5.4, %Fe₂O₃ between about 5.4-6.5, % K₂O between about 0.50-0.90, %Na₂O between about 0.10-0.30, %MnO between about 0.01-0.03, % Al₂O₃ between about 14.8- 18.2, and %SiO₂ between about 62.4-73.5, (f) a content of heavy metals of Pb at about 6.0-6.5 ppm, As at about 0.5-0.7 ppm; Cd at about 0.2-0.4 ppm, Cr at about 5.5-6.0 ppm, and Hg less than 0.1 ppm, and

(g) a 2,3,7,8-tetrachlorodibenzodioxin ("TCDD") amount of less than 0.33 parts per trillion ("ppt").

28. The method of claim 21, wherein the fish or shrimp feed material comprises 25% fishmeal, 14% bran, 26% corn, and 35% soybean by weight.

29. The method of claim 21, wherein the mixture further comprises formic acid, calcium formate, diformate, acetic acid, diacetate, calcium acetate, lactic acid, calcium lactate, propionic acid, calcium propionate, citric acid, calcium citrate, fumaric acid, calcium fumarate, malic acid, calcium malate, products of lactobacillus fermentation, or mixtures thereof.

30. The method of claim 21 , wherein the mixture further comprises a desiccant.

31. The method of claim 21. further comprising the step of blending molasses or fish oil in the mixture.

32. The method of claim 21, wherein the mixing step further comprises the step of extruding or pelletizing the acidified clay material and the fish or shrimp feed material.

33. The method of claim 21 , further comprising the step of adjusting the pH of the mixture to about 4.5 to 5.0.

34. A method for preparing a modified fish or shrimp feed composition having antimicrobial properties, comprising: mixing an acidulant and a fish or shrimp feed material to form a mixture, wherein the acidulant is ACS, and wherein the ACS has a normality of about 5-10 N.

35. The method of claim 34, wherein the mixing step further comprises the step of extruding the fish or shrimp feed material, wherein the acidulant is added to the fish or shrimp feed material during the extruding.

36. The method of claim 34, further comprising the step of adjusting the pH of the mixture to about 4.5 to 5.0.

37. The method of claim 34, wherein the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide, or a calcium salt, or a mixture of the two, wherein when the ACS is isolated from a mixture comprising sulfuric acid and calcium hydroxide then the mole ratio of calcium hydroxide to sulfuric acid ranges from about 0.1 to about 0.5, and wherein the ACS has a pH of less than about 2.