WO2016203246A1

WO2016203246A1 - Method

Info

Publication number: WO2016203246A1
Application number: PCT/GB2016/051802
Authority: WO
Inventors: James Bray; Keith Jolley; Martin Maiden
Original assignee: Oxford University Innovation Ltd
Current assignee: Oxford University Innovation Ltd
Priority date: 2015-06-17
Filing date: 2016-06-16
Publication date: 2016-12-22
Anticipated expiration: 2017-12-17
Also published as: GB201510649D0

Abstract

The invention relates to identifying genomic sequences which are characteristic of an organism. The invention also relates to using the genomic sequences to distinguish the organism from other organisms.

Description

METHOD

Field of the Invention

Background of the Invention

It is important to be able to rapidly identify organisms and rapidly differentiate them from other organisms. For instance, infections with microorganisms, occurring primarily in hospitalised patients, can delay recovery and in some instances can be life threatening. It has been established that a decrease in the time before diagnosis of microorganism infection improves patient outcome. The ability to rapidly identify a microorganism is therefore important for improved patient treatment and survival.

Summary of the Invention

The inventors have surprisingly shown that is possible to identify in the genome of an organism one or more polynucleotide sequences (one or more k-mers) which are capable of distinguishing the organism from one or more different organisms. In particular, the inventors have surprisingly shown that it is possible to identify one or more k-mers which are present in all known versions of the genome of an organism, but which are not present in the genomes of one or more different organisms or all different organisms. Such k-mers allow the organism to be distinguished from the one or more different organisms. Most surprisingly of all, the inventors have shown that it is possible to identify one k-mer in all known versions of the genome of an organism which is capable of distinguishing the organism from one or more different organisms. This makes it possible to rapidly and efficiently identify the organism.

The invention provides a method for identifying one or more nucleotide k-mers in the genome of an organism which are capable of distinguishing the organism from one or more different organisms, comprising:

(a) extracting all nucleotide k-mers from the genome of the organism; and

(b) comparing the nucleotide k-mers extracted in step (a) with the genome(s) of the one or more different organisms and identifying those nucleotide k-mers which do not appear in the genome(s) of the one or more different organisms.

The invention also provides:

- a computer program configured to carry out a method of the invention; a computer program medium comprising a computer program of the invention;

a computer for carrying out a method of the invention or programmed with a computer program of the invention;

one or more nucleotide k-mers identified using a method of the invention or a computer program of the invention;

and oligonucleotide probe which comprises a sequence which is complementary to a nucleotide k-mer of the invention;

a plurality of oligonucleotide probes each of which comprises a sequence which is complementary to one of the nucleotide k-mers in a population of two or more nucleotide k-mers of the invention;

a support having attached thereto an oligonucleotide probe of the invention or a plurality of oligonucleotide probes of the invention;

a method for detecting the presence or absence of an organism in a sample, comprising detecting the presence or absence in the sample of one or more nucleotide k-mers of the invention, wherein the presence of the one or more nucleotide k-mers is indicative of the presence of the organism in the sample and the wherein the absence of the one or more nucleotide k-mers is indicative of the absence of the organism from the sample;

a nucleotide k-mer capable of distinguishing an organism from one or more different organisms, wherein the k-mer comprises about 10 or more consecutive nucleotides from any one of the sequences shown in SEQ ID NOs: 1 to 213;

an oligonucleotide probe which comprises a sequence which is complementary to a nucleotide k-mer of the invention and derived from any one of SEQ ID NOs: 1 to 213; a support having attached thereto a oligonucleotide probe of the invention and derived from any one of SEQ ID NOs: 1 to 213; and

- a method for detecting the presence or absence of an organism in a sample, comprising detecting the presence or absence in the sample of a nucleotide k-mer of the invention and derived from any one of SEQ ID NOs: 1 to 213, wherein the presence of the nucleotide k-mer is indicative of the presence of the organism in the sample and the wherein the absence of the nucleotide k-mer is indicative of the absence of the organism from the sample.

Description of the Figures

Figure 1 shows a flowchart describing the four computational stages required to generate a small single species k-mer library from protein-encoding ribosomal gene DNA. Description of the Sequences

SEQ ID NOs: 1 to 5 show k-mer cluster sequences from Aggregatibacter

actinomycetemcomitans.

SEQ ID NOs: 6 to 10 show k-mer cluster sequences Bacillus anthracis.

SEQ ID NOs: 11 to 15 show k-mer cluster sequences from Bacillus licheniformis.

SEQ ID NOs: 16 to 20 show k-mer cluster sequences from Bacteroides fragilis.

SEQ ID NOs: 21 to 25 to 5 show k-mer cluster sequences from Bartonella henselae.

SEQ ID NOs: 26 to 30 show k-mer cluster sequences from Bordetella pertussis.

SEQ ID NOs: 31 to 35 show k-mer cluster sequences from Borrelia burgdorferi.

SEQ ID NOs: 36 to 40 show k-mer cluster sequences from Brucella abortus.

SEQ ID NOs: 41 to 44 show k-mer cluster sequences from Campylobacter jejuni.

SEQ ID NOs: 45 to 49 show k-mer cluster sequences from Chlamydia trachomatis.

SEQ ID NOs: 50 to 54 show k-mer cluster sequences from Chlamydophila pneumonia.

SEQ ID NOs: 55 to 59 show k-mer cluster sequences from Clostridium difficile.

SEQ ID NOs: 60 to 64 show k-mer cluster sequences from Clostridium perfringens.

SEQ ID NOs: 65 to 69 show k-mer cluster sequences from from Enterobacter aerogenes.

SEQ ID NOs: 70 to 74 show k-mer cluster sequences from from Enterococcus faecalis.

SEQ ID NOs: 75 to 79 show k-mer cluster sequences from from Enterococcus faecium.

SEQ ID NOs: 80 to 84 show k-mer cluster sequences from Francisella tularensis.

SEQ ID NO: 85 shows a k-mer cluster sequence from Haemophilus influenza.

SEQ ID NOs: 86 and 87 show k-mer cluster sequences from Helicobacter pylori.

SEQ ID NOs: 88 to 92 show k-mer cluster sequences from Klebsiella oxytoca

SEQ ID NOs: 93 to 97 show k-mer cluster sequences from Legionella pneumophila.

SEQ ID NOs: 98 to 102 show k-mer cluster sequences from Listeria monocytogenes.

SEQ ID NOs: 103 to 107 show k-mer cluster sequences from Moraxella catarrhalis.

SEQ ID NOs: 108 to 112 show k-mer cluster sequences from Mycobacterium avium.

SEQ ID NOs: 113 to 116 show k-mer cluster sequences from Mycobacterium bovis.

SEQ ID NOs: 117 to 121 show k-mer cluster sequences from Mycoplasma genitalium.

SEQ ID NOs: 122 to 126 show k-mer cluster sequences from Mycoplasma pneumonia.

SEQ ID NOs: 127 to 131 show k-mer cluster sequences from Neisseria gonorrhoeae.

SEQ ID NOs: 132 to 136 show k-mer cluster sequences from Neisseria meningitides.

SEQ ID NOs: 137 to 141 show k-mer cluster sequences from Porphyromonas gingivalis.

SEQ ID NOs: 142 to 146 show k-mer cluster sequences from Proteus mirabilis.

SEQ ID NOs: 147 to 151 show k-mer cluster sequences from Pseudomonas aeruginosa.

SEQ ID NOs: 152 to 156 show k-mer cluster sequences from Salmonella enterica. SEQ ID NOs: 157 to 161 show k-mer cluster sequences from Serratia marcescens. SEQ ID NOs: 162 to 166 show k-mer cluster sequences from Staphylococcus aureus. SEQ ID NOs: 167 to 171 show k-mer cluster sequences from Staphylococcus

epidermidis.

SEQ ID NOs: 172 to 176 show k-mer cluster sequences from Staphylococcus

haemolyticus.

SEQ ID NO: 177 shows a k-mer cluster sequence from Stenotrophomonas maltophilia.

SEQ ID NOs: 178 to 182 show k-mer cluster sequences from Streptococcus mutans.

SEQ ID NOs: 183 to 187 show k-mer cluster sequences from Streptococcus pyogenes. SEQ ID NOs: 188 to 192 show k-mer cluster sequences from Streptococcus salivarius.

SEQ ID NOs: 193 to 197 show k-mer cluster sequences from Streptococcus sanguinis.

SEQ ID NOs: 198 to 202 show k-mer cluster sequences from Treponema pallidum.

SEQ ID NO: 203 shows a k-mer cluster sequence from Vibrio cholera.

SEQ ID NOs: 204 to 208 show k-mer cluster sequences from Vibrio parahaemolyticus. SEQ ID NOs: 209 to 213 show k-mer cluster sequences from Yersinia enter ocolitica.

Detailed Description of the Invention

It is to be understood that different applications of the disclosed methods may be tailored to the specific needs in the art. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments of the invention only, and is not intended to be limiting.

In addition, as used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to "an organism" includes two or more organisms, reference to "a k-mer" includes two or more k-mers, reference to "a probe" includes two or more such probes and the like.

All publications, patents and patent applications cited herein are hereby incorporated by reference in their entirety. Methods of identifying k-mers

The invention concerns identifying one or more nucleotide k-mers in the genome of an organism. A nucleotide k-mer is a nucleotide sequence containing a whole number value, k, of nucleotides. Suitable lengths of the k-mers are discussed in more detail below.

The one or more k-mers in the genome of the organism are capable of distinguishing the organism from one or more different organisms. The one or more k-mers in the genome of the organism are characteristic of the organism. The one or more k-mers in the genome of the organism are specific for the organism.

The one or more k-mers in the genome of the organism are capable of distinguishing the organism from any number of different organisms. The one or more k-mers in the genome of the organism are preferably capable of distinguishing the organism from at least about 2, at least about 3, at least about 4, at least about 5, at least about 10, at least about 20, at least about 30, at least about 50, at least about 100, at least about 150, at least about 200, at least about 250, at least about 500, at least about 1000, at least about 5000 or at least about 10,000 different organisms or more. The one or more k-mers in the genome of the organism are preferably capable of distinguishing the organism from all different organisms.

An organism is typically different from the organism in which the one or more k-mers are identified (i.e. the organism of interest) if it belongs to a different species.

Organisms may be distinguished at any level of taxonomy. The one or more k-mers in the genome of an organism in a particular kingdom are preferably capable of distinguishing the organism from one or more different organisms in a different kingdom or different kingdoms, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all organisms in different kingdoms. For instance, the one or more k-mers in the genome of a bacterium are preferably capable of distinguishing the bacterium from one or more different fungi, such as any number of fungi as described above, and vice versa. The one or more k-mers in the genome of a bacterium are more preferably capable of distinguishing the bacterium from all different fungi and vice versa.

The one or more k-mers in the genome of an organism in a particular kingdom are preferably capable of distinguishing the organism from one or more different organisms in the same kingdom, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all organisms in the same kingdom. For instance, the one or more k-mers in the genome of a bacterium are preferably capable of distinguishing the bacterium from one or more different bacteria, such as any number of bacteria as described above. The one or more k-mers in the genome of a bacterium are more preferably capable of distinguishing the bacterium from all different bacteria. The one or more k-mers in the genome of a fungus are preferably capable of distinguishing the fungus from one or more different fungi, such as any number of fungi as described above. The one or more k-mers in the genome of a fungus are more preferably capable of distinguishing the fungus from all different fungi. The one or more k-mers in the genome of an organism in a particular phylum are preferably capable of distinguishing the organism from one or more organisms in a different phylum or different phyla, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all organisms in different phyla.

Organisms within the same phylum may be distinguished in accordance with the invention. The one or more k-mers in the genome of an organism in a particular phylum are preferably capable of distinguishing the organism from one or more different organisms in the same phylum, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all different organisms in the same phylum.

The one or more k-mers in the genome of an organism in a particular family are preferably capable of distinguishing the organism from one or more organisms in a different family or different families, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all organisms in different families.

Organisms within the same family may be distinguished in accordance with the invention. The one or more k-mers in the genome of an organism in a particular family are preferably capable of distinguishing the organism from one or more different organisms in the same family, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all different organisms in the same family.

The one or more k-mers in the genome of an organism in a particular genus are preferably capable of distinguishing the organism from one or more organisms in a different genus or different genera, such as any number of organisms as described above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all organisms in different genera.

Organisms within the same genus may be distinguished in accordance with the invention. The one or more k-mers in the genome of the organism are preferably capable of distinguishing the organism from one or more different species in the same genus, such as any number of different species as discussed above. The one or more k-mers in the genome of the organism are more preferably capable of distinguishing the organism from all of the different species in the same genus.

The one or more k-mers may be identified in the genome of any organism. The organism may be eukaryotic. The organism may be an animal or a plant. The organism may be human or another mammalian animal, such as a commercially farmed animal, such as a horse, a cow, a sheep, a fish, a chicken or a pig, a laboratory animal, such as a mouse or a rat, or a pet, such as a guinea pig, a hamster, a rabbit, a cat or a dog. The organism may be a plant, such a cereal, legume, fruit or vegetable, for example wheat, barley, oats, canola, maize, soya, rice, rhubarb, bananas, apples, tomatoes, potatoes, grapes, tobacco, beans, lentils, sugar cane, cocoa or cotton. The organism may be a fungus.

The organism may be prokaryotic, such a bacterium or archaeon.

Preferably, the organism is a microorganism, a fungus or a virus and the one or more different organisms are one or more different microorganisms, fungi or viruses.

The organism is preferably a bacterium. The bacterium may be Gram negative or Gram positive. The Gram positive bacterium is preferably from the genus Bacillus, Clostridium, Enterococcus, Mycobacterium, Staphylococcus or Streptococcus. The Gram positive bacterium may be from the genus Pasteurella or Nocardia.

The Gram negative bacterium is preferably from the genus Aggregatibacter, Bacteroides,

Bartonella, Brucella, Campylobacter, Chylamidia, Enterbacter, Francisella, Haemophilus, Heliobacter, Klebsiella, Legionella, Moraxella, Neisseria, Porphyromonas, Pseudomonas, Salmonella, Serratia, Stenotrophomonas, Vibrio or Yersinia. The Gram negative bacterium may be from the genus Escherichia or Pseudomonas.

The bacterium may be from the genus Borrelia, Chlamydophila, Listeria, Mycoplasma,

Proteus or Treponema,

The bacterium is preferably Aggregatibacter actinomycetemcomitans, Bacillus anthracis, Bacillus licheniformis, Bacteroides fragilis, Bartonella henselae, Bordetella pertussis, Borrelia burgdorferi, Brucella abortus, Campylobacter jejuni, Chlamydia trachomatis, Chlamydophila pneumoniae, Clostridium difficile, Clostridium perfringens, Enterobacter aerogenes,

Enterococcus faecalis, Enterococcus faecium, Francisella tularensis, Haemophilus influenzae, Helicobacter pylori, Klebsiella oxytoca, Legionella pneumophila, Listeria monocytogenes, Moraxella catarrhalis, Mycobacterium avium, Mycobacterium bovis, Mycoplasma genitalium, Mycoplasma pneumoniae, Neisseria gonorrhoeae, Neisseria meningitidis, Porphyromonas gingivalis, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella enter ica, Serratia marcescens, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Stenotrophomonas maltophilia, Streptococcus mutans, Streptococcus pyogenes, Streptococcus salivarius, Streptococcus sanguinis, Treponema pallidum, Vibrio cholera, Vibrio

parahaemolyticus or Yersinia enter ocolitica. Specific and preferred k-mers present in these bacteria are discussed below. Other specific examples of bacteria include, but are not limited, to Mycobacterium tuberculosis, Mycobacterium intracellilare, Mycobacterium kansaii, Mycobacterium gordonae, Streptococcus agalactiae, Streptococcus viridans group, Streptococcus faecalis, Streptococcus bovis, Streptococcus pneumoniae, Corynebacterium diptheriae, Erysipelothrix rhusiopathie, Clostridium tetani, Klebsiella pneumoniae, Pasteurella multocida, Fusobacterium nucleatum, Streptobacillus moniliformis, Treponema pertenue and Actinomyces israelii.

Preferably, the organism is a bacterium and the one or more different organisms are one or more different bacteria. There may be any number of different bacteria as discussed above. Preferably, the organism is a bacterium and the one or more different organisms are all different bacteria.

Preferably, the organism is a bacterium and the one or more different organisms are one or more bacteria from one or more different genera of bacteria, such as all different genera of bacterium. For instance, (a) the organism may be a bacterium from the genus Bacillus and (b) the one or more different organisms may be one or more bacteria from one or more of, or all of, Aggregatibacter, Bacteroides, Bartonella, Bordetella, Borrelia, Brucella, Campylobacter, Chlamydia, Chlamydophila, Clostridium, Enterobacter, Enterococcus, Francisella,

Haemophilus, Helicobacter, Klebsiella, Legionella, Listeria, Moraxella, Mycobacterium, Mycoplasma, Neisseria, Porphyromonas, Proteus, Pseudomonas, Salmonella, Serratia,

Staphylococcus, Stenotrophomonas, Streptococcus, Treponema, Vibrio and Yersinia.

Alternatively, the organism in (a) may be a bacterium from one of the genera in list (b) and the one or more different organisms may be one or more bacteria from one or more of, or all of, Bacillus and the remaining genera in list (b).

More preferably, (i) the organism is a bacterium and (ii) the one or more different organisms are one or more different species of bacteria, (i) is preferably selected from the list Aggregatibacter actinomycetemcomitans, Bacillus anthracis, Bacillus licheniformis, Bacteroides fragilis, Bartonella henselae, Bordetella pertussis, Borrelia burgdorferi, Brucella abortus, Campylobacter jejuni, Chlamydia trachomatis, Chlamydophila pneumoniae, Clostridium difficile, Clostridium perfringens, Enterobacter aerogenes, Enterococcus faecalis, Enterococcus faecium, Francisella tularensis, Haemophilus influenzae, Helicobacter pylori, Klebsiella oxytoca, Legionella pneumophila, Listeria monocytogenes, Moraxella catarrhalis,

Mycobacterium avium, Mycobacterium bovis, Mycoplasma genitalium, Mycoplasma

pneumoniae, Neisseria gonorrhoeae, Neisseria meningitidis, Porphyromonas gingivalis, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella enterica, Serratia marcescens, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Stenotrophomonas maltophilia, Streptococcus mutans, Streptococcus pyogenes, Streptococcus salivarius, Streptococcus sanguinis, Treponema pallidum, Vibrio cholera, Vibrio parahaemolyticus or

Yersinia enterocolitica and the one or more species in (ii) are preferably the species remaining in the list.

The organism is preferably a fungus. Preferably, the organism is a fungus and the one or more different organisms are one or more different fungi. The fungus is preferably from the genus Absidia, Acremonium, Aspergillus, Aureobasidium, Basidiobolus, Blastomyces,

Blastoschizomyces, Candida, Cladosporium, Coccidioides, Cryptococcus, Cunninghamella, Curvularia, Debaryomyces, Exophiala, Exserohilum, Fonsecea, Fusarium, Geotrichum,

Histoplasma, Issatchenkia, Kluyveromyces, Malezzesia, Mucor, Paracoccidioides,

Paecilomyces, Penicillium, Pichia, Pneumocystis, Rhizomucor, Rhizopus, Rhodotorula,

Saccharomyces, Scedosporium, Schizophyllum, Scopulariopsis, Sporothrix, Trichoderma;

Trichophyton or Trichosporon.

The organism is preferably Aspergillus fumigatus, Aspergillus flavus, Aspergillus lentulus, Aspergillus terreus, Aspergillus nidulans, Aspergillus oryzae, Aspergillus niger, Candida albicans, Candida caribbica {Candida fermentati), Candida dubliniensis, Candida famata (Debaryomyces hansenii), Candida fukuyamaensis {Candida xestobii or Candida carpophila), Candida guilliermondii, Candida kefyr (Kluyveromyces marxianus), Candida krusei (Issatchenkia orientalis), Candida metapsilosis, Candida orthopsilosis, Candida parapsilosis, Candida parapsilosis, Candida pelliculosa, Candida psychrophila, Candida rugosa, Candida smithsonii, Candida tropicalis, Candida utilis, Coccidioides immitis , Cryptococcus

bacillisporus; Cryptococcus gattii; Cryptococcus grubii; Cryptococcus neoformans,

Debaryomyces coudertii, Debaryomyces maramus, Debaryomyces nepalensis, Debaryomyces prosopidis, Debaryomyces robertsiae, Debaryomyces udenii, Histoplasma capsulatum,

Kluyveromyces lactis, Pichia cecembensis, Rhodotorula araucariae, Rhodotorula babjevae, Rhodotorula dairensis, Rhodotorula diobovatum, Rhodotorula glutinis, Rhodotorula

kratochvilovae, Rhodotorula paludigenum, Rhodotorula sphaerocarpum, Rhodotorula toruloides, Rhodotorula mucliaginosa, Saccharomyces 'sensu stricto ', Saccharomyces bayanus, Saccharomyces boulardii, Saccharomyces cariocanus, Saccharomyces kudiavzevii,

Saccharomyces mikatae, Saccharomyces paradioxus, Saccharomyces pastorianus,

Saccharomyces uvarum, Saccharomyces cerevisiae and Tsuchiyaea wingfieldii.

Preferably, the organism is a fungus and the one or more different organisms are one or more different fungi. There may be any number of different fungi as discussed above.

Preferably, the organism is a fungus and the one or more different organisms are all different fungi. Preferably, the organism is a fungi and the one or more different organisms are one or more fungi from one or more different genera of fungi, such as all different genera of fungi. For instance, (a) the organism may be a fungus from the genus Candida and (b) the one or more different organisms may be one or more fungi from one or more of, or all of, Absidia,

Acremonium, Aspergillus, Aureobasidium, Basidiobolus, Blastomyces, Blastoschizomyces, Candida, Cladosporium, Coccidioides, Cryptococcus, Cunninghamella, Curvularia,

Debaryomyces, Exophiala, Exserohilum, Fonsecea, Fusarium, Geotrichum, Histoplasma, Issatchenkia, Kluyveromyces, Malezzesia, Mucor, Paracoccidioides, Paecilomyces, Penicillium, Pichia, Pneumocystis, Rhizomucor, Rhizopus, Rhodotorula, Saccharomyces, Scedosporium, Schizophyllum, Scopulariopsis, Sporothrix, Trichoderma; Trichophyton and Trichosporon. Alternatively, the organism in (a) may be a fungus from one of the genera in list (b) (such as Aspergillus) and the one or more different organisms may be one or more bacteria from one or more of, or all of, Candida and the remaining genera in list (b) (such as everything except Aspergillus).

More preferably, (i) the organism is a fungus and (ii) the one or more different organisms are one or more different species of fungus, (i) is preferably selected from the list of fungal species above and the one or more species in (ii) are preferably the species remaining in the list.

The organism is preferably a virus. The virus may belong to the family Retroviridae, such as human deficiency viruses, such as HIV-I (also referred to as HTLV- III), HIV-II, LAC, IDLV-III/LAV, HIV-III or other isolates such as HIV-LP, the family Picornaviridae, such as poliovirus, hepatitis A, enteroviruses, human Coxsackie viruses, rhinoviruses, echoviruses, the family Calciviridae, such as viruses that cause gastroenteritis, the family Togaviridae, such as equine encephalitis viruses and rubella viruses, the family Flaviviridae, such as dengue viruses, encephalitis viruses and yellow fever viruses, the family Coronaviridae, such as coronaviruses, the family Rhabdoviridae, such as vesicular stomata viruses and rabies viruses, the family

Filoviridae, such as Ebola viruses, the family Paramyxoviridae, such as parainfluenza viruses, mumps viruses, measles virus and respiratory syncytial virus, the family Orthomyxoviridae, such as influenza viruses, the family Bungaviridae, such as Hataan viruses, bunga viruses, phleoboviruses and Nairo viruses, the family Arena viridae, such as hemorrhagic fever viruses, the family Reoviridae, such as reoviruses, orbiviruses and rotaviruses, the family Bimaviridae, the family Hepadnaviridae, such as hepatitis B virus, the family Parvoviridae, such as parvoviruses, the Papovaviridae, such as papilloma viruses and polyoma viruses, the family Adenoviridae, such as adenoviruses, the family Herpesviridae, such as herpes simplex virus (HSV) I and II, varicella zoster virus and pox viruses, or the family Iridoviridae, such as African swine fever virus). The virus may be an unclassified virus, such as the etiologic agents of Spongiform encephalopathies, the agent of delta hepatitis, the agents of non-A, non-B hepatitis (class 1 enterally transmitted; class 2 parenterally transmitted such as Hepatitis C); Norwalk and related viruses and astroviruses.

Preferably, the organism is a virus and the one or more different organisms are one or more different virus. There may be any number of different viruses as discussed above.

Preferably, the organism is a virus and the one or more different organisms are all different viruses.

Step (a)

The method comprises extracting all nucleotide k-mers from the genome of the organism. The genome of the organism is typically publically available, for instance on GenBank. A computer is capable of extracting all nucleotide k-mers from the genome.

A nucleotide typically contains a nucleobase, a sugar and at least one linking group, such as a phosphate, 2'0-methyl, 2' methoxy-ethyl, phosphoramidate, methylphosphonate or phosphorothioate group. The nucleobase is typically heterocyclic. Nucleobases include, but are not limited to, purines and pyrimidines and more specifically adenine (A), guanine (G), thymine (T), uracil (U) and cytosine (C). The sugar is typically a pentose sugar. Nucleotide sugars include, but are not limited to, ribose and deoxyribose.

The nucleotides in the genome of the organism are typically ribonucleotides or deoxyribonucleotides. The nucleotides typically contain a monophosphate, diphosphate or triphosphate. Phosphates may be attached on the 5' or 3' side of a nucleotide.

Nucleotides include, but are not limited to, adenosine monophosphate (AMP), adenosine diphosphate (ADP), adenosine triphosphate (ATP), guanosine monophosphate (GMP), guanosine diphosphate (GDP), guanosine triphosphate (GTP), thymidine monophosphate (TMP), thymidine diphosphate (TDP), thymidine triphosphate (TTP), uridine monophosphate (UMP), uridine diphosphate (UDP), uridine triphosphate (UTP), cytidine monophosphate (CMP), cytidine diphosphate (CDP), cytidine triphosphate (CTP), 5-methylcytidine monophosphate, 5- methylcytidine diphosphate, 5-methylcytidine triphosphate, 5-hydroxymethylcytidine

monophosphate, 5-hydroxymethylcytidine diphosphate, 5-hydroxymethylcytidine triphosphate, cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP), deoxyadenosine monophosphate (dAMP), deoxyadenosine diphosphate (dADP),

deoxyadenosine triphosphate (dATP), deoxyguanosine monophosphate (dGMP),

deoxyguanosine diphosphate (dGDP), deoxyguanosine triphosphate (dGTP), deoxythymidine monophosphate (dTMP), deoxythymidine diphosphate (dTDP), deoxythymidine triphosphate (dTTP), deoxyuridine monophosphate (dUMP), deoxyuridine diphosphate (dUDP), deoxyuridine triphosphate (dUTP), deoxycytidine monophosphate (dCMP), deoxycytidine diphosphate (dCDP) and deoxycytidine triphosphate (dCTP), 5-methyl-2' -deoxycytidine monophosphate, 5- methyl-2' -deoxycytidine diphosphate, 5 -methyl-2' -deoxycytidine triphosphate, 5- hydroxymethyl-2' -deoxycytidine monophosphate, 5 -hydroxymethyl-2' -deoxycytidine diphosphate and 5 -hydroxymethyl-2 '-deoxycytidine triphosphate. The nucleotides are preferably selected from AMP, TMP, GMP, UMP, dAMP, dTMP, dGMP or dCMP.

The organism preferably comprises a genome which is based on deoxyribonucleic acid (DNA) and which comprises dAMP, dTMP, dGMP and dCMP. If the organism's genome is based on DNA, k-mers which comprise nucleotides other than dAMP, dTMP, dGMP and dCMP are preferably be excluded from the comparison in step (b). The organism, such as a virus, may comprise a genome which is based on ribonucleic acid (RNA) and comprises AMP, TMP, GMP and UMP. If the organism's genome is based on RNA, k-mers which comprise one or more nucleotides other than AMP, TMP, GMP and UMP are preferably excluded from the comparison in step (b).

The nucleotide k-mers may be any length. Step (a) preferably comprises defining a length for the one or more nucleotide k-mers and extracting all nucleotide k-mers having that length from the genome of the organism. Step (a) more preferably comprises defining a length of from about 10 to about 40 nucleotides for the one or more nucleotide k-mers and extracting all nucleotide k-mers having that length from the genome of the organism. The method may comprise defining a length of from about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, or about 19 nucleotides to about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38 or about 39 nucleotides.

Preferred k-mer lengths include, but are not limited to, about 17, about 20, about 25 and about 30 nucleotides.

Certain nucleotide k-mers may appear more than once in the genome of the organism. All instances of theses nucleotide k-mers may be extracted and compared in step (b). A nucleotide k-mer which appears more than once in the organism's genome is preferably extracted only once in step (a) and only compared once with the genome(s) of the one or more different organisms in step (b).

Several different versions of the genome of the organism may be publically available. The ability to distinguish the organism from one or more different organisms may be improved if two or more different versions of the genome are analysed using the method of the invention. Step (a) preferably comprises extracting all nucleotide k-mers from two or more different versions of the genome of the organism. Step (a) more preferably comprises extracting all nucleotide k-mers from (i) two hundred or more or (ii) two thousand or more different versions of the genome of the organism. Any number of different versions of the genome may be analysed such as about five or more, about ten or more, about fifty or more, about one hundred or more, about five hundred or more, about one thousand or more, about five thousand or more or even more.

Step (a) most preferably comprises extracting all nucleotide k-mers from all known versions of the genome of the organism.

In some instances, the nucleotide k-mers may be extracted from certain parts or fragments of the genome. For instance, the method may be focussed on those parts or fragments of the genome which are expected to be conserved amongst different versions of the genome of the organism and/or not conserved in the one or more different organisms. Thus, the method may comprise using only a subset of sequences from the genome of the organism. This may advantageously reduce complexity and input data for genome comparisons. The person skilled in the art is capable of identifying such parts or fragments of the genomes. Step (a) preferably comprises extracting all nucleotide k-mers from a predefined or specific part or fragment of the genome of the organism. The part or fragment may be any part or fragment. The nucleotide k- mers are preferably extracted from the parts or fragments of the genome which encode the ribosomal protein subunits. This is disclosed in the Example. The nucleotide k-mers are preferably extracted from the parts or fragments of the genome which are involved in DNA replication, such as a part or fragment of the genome which encode a polymerase, a helicase and/or a topoisomerases, such as a gyrase.

Step (b)

The method also comprises comparing the nucleotide k-mers extracted in step (a) with the genome(s) of the one or more different organisms, such as all different organisms, and identifying those nucleotide k-mers which do not appear in the genome(s) of the one or more different organisms. This is straightforward using a computer. Once the k-mers have been extracted, the computer can investigate the genome(s) of the one or more different organisms to determine whether or not the k-mers are present. Those k-mers which do not appear in (i.e. are not present in) the genome(s) of the one or more different organisms may be used to distinguish the organism from the one or more different organisms. These k-mers may be used in the subsequent optional steps of the method of the invention. Those k-mers which appear in (i.e. are present in) the genome(s) of the one or more different organisms may not be used to distinguish the organism from the one or more different organisms. These k-mers are typically discarded and are not used in the subsequent optional steps of the method of the invention. Step (b) preferably comprises comparing the nucleotide k-mers extracted in step (a) with two or more versions of the genome of each the one or more different organisms, such as all different organisms, and identifying those nucleotide k-mers which do not appear in the genomes of the one or more different organisms. Step (b) more preferably comprises comparing the nucleotide k-mers extracted in step (a) with about two hundred or more, about two thousand or more or all known versions of the genome of each the one or more different organisms, such as all different organisms, and identifying those nucleotide k-mers which do not appear in the genomes of the one or more different organisms. Any number of versions of the one or more different organsims may be investigated as discussed above for the organism of interest.

Optional step (c)

The method of the invention may also involve removing those k-mers which may not be helpful in distinguishing the organism from the one or more different organisms. In particular, the method preferably comprises (c) discounting any nucleotide k-mers identified in step (b) which have an undesirable property. The undesirable property is preferably (i) its presence in the human genome, (ii) a homopolymer or repetitive sequence, (iii) the ability to form secondary structure or (iv) its presence in a contaminating organism.

Optional step (d)

At the end of step (b) or step (c), there may be hundreds, thousands or millions of unique k-mers identified in the organism's genome. This may be acceptable for computational applications, but a smaller collection of k-mers (e.g. tens or hundreds) may be desirable in some experimental contexts, such as microarrays or genotyping by species-specific primers.

The method preferably further comprises (d) analysing the nucleotide k-mers identified in step (b) or step (c) and reducing their number by removing at least some sequence redundancy. This can be done using know method, such as set covering. A preferred method is disclosed in the example. Step (d) preferably comprises minimising the number of nucleotide k-mers by removing all sequence redundancy. Numbers of identified k-mers

The method of the invention may comprise identifying any number of k-mers. The method preferably comprises identifying about 25 or fewer k-mers which are capable of distinguishing the organism from the one or more different organisms such as all different organisms, such as about 24 or fewer k-mers, about 23 or fewer k-mers, about 22 or fewer k- mers, about 21 or fewer k-mers, about 20 or fewer k-mers, about 19 or fewer k-mers, about 18 or fewer k-mers, about 17 or fewer k-mers, about 16 or fewer k-mers, about 15 or fewer k-mers, about 14 or fewer k-mers, about 13 or fewer k-mers, about 12 or fewer k-mers, about 11 or fewer k-mers, about 10 or fewer k-mers, about 9 or fewer k-mers, about 8 or fewer k-mers, about 7 or fewer k-mers, about 6 or fewer k-mers, about 5 or fewer k-mers, about 4 or fewer k-mers, about 3 or fewer k-mers, or about 2 or fewer k-mers. The method may identify two or more nucleotide k-mers each having the same length. Alternatively, the method may identify two or more nucleotide k-mers having different lengths. The method may identify a population of k-mers in which two or more k-mers have the same length and two or more k-mers have different lengths.

The method preferably identifies only one k-mer which is capable of distinguishing the organism from the one or more different organisms, such as all different organisms.

As explained above, step (a) may involve extracting one or nucleotide k-mers from all of the different versions in the genome of the organism. The method is preferably for identifying about 25 or fewer nucleotide k-mers (or any of the numbers listed above) which are present in all of the different versions of the genome of the organism and which are capable of distinguishing the organism from the one or more different organisms, such as all different organisms.

The method is most preferably for identifying only one nucleotide k-mer which is present in all of the different versions of the genome of the organism and which is capable of

distinguishing the organism from the one or more different organisms, such as all different organisms.

The method of the invention may comprise identifying a cluster of k-mer sequences in the genome of the organism of interest, wherein more than one k-mer in the cluster is capable of distinguishing the organism from one or more different organisms. The k-mer sequences in the cluster which are capable of distinguishing the organism may be overlapping. Each nucleotide in the cluster is typically found in at least one k-mer which is capable of distinguishing the organism. The method may comprise identification of a cluster of k-mer sequences in the genome of the organism that has the highest number of overlapping k-mer sequences which are capable of distinguishing the organism. The cluster of k-mer sequences may comprise a k-mer that is not capable of distinguishing the organism from one or more different organisms, in addition to the k-mer sequences which are capable of distinguishing the organism. Examples of k-mer clusters identified according to the invention include SEQ ID NOs: 1 to 213, as further described below.

Phenotype The invention may also be used to identifying one or more nucleotide k-mers in the genome of an organism which can be associated with a phenotype of the organism. Any of the embodiments apply to this method.

Step (a) comprises extracting all nucleotide k-mers from the genome of the organism as described above. K-mers may be extracted from any number of different versions of the genome of the organism as discussed above.

Step (b) comprises comparing the nucleotide k-mers extracted in step (a) with the genome(s) of one or more different organisms, such as all different organisms, which do not display the phenotype and identifying those nucleotide k-mers which do not appear in the genome(s) of the one or more different organisms which do not display the phenotype. Any number of different versions of the genomes of the one or more different organisms, such as all different organisms, which do not display the phenotype may be investigated as discussed above.

Those k-mers which do not appear in (i.e. are not present in) the genome(s) of the one or more different organisms may be associated with a phenotype of the organism. These k-mers may be used in the subsequent optional steps of the method of the invention. Those k-mers which appear in (i.e. are present in) the genome(s) of the one or more different organisms may not be associated with a phenotype of the organism. These k-mers are typically discarded and are not used in the subsequent optional steps of the method of the invention.

The phenotype may be any phenotype of the organism. Preferred phenotypes include, but are not limited to, pathogenicity, resistance to antibiotics, host specificity, tissue specificity, transmissibility, virulence, antigenicity and biochemical properties. The one or more different organisms are typically from the same phylum, family or genus as the organism of interest.

Computer program

The invention also provides a computer program configured to carry out the method of the invention. The program can be written using routine methods.

The invention also provides a computer program medium, such as hard drive, USD or disk, comprising the computer program of the invention.

The invention further provides a computer for carrying out the method of the invention programmed with the computer program of the invention.

K-mers and probes

The invention also provides one or more nucleotide k-mers identified using the method of the invention or the computer program of the invention. The one or more nucleotide k-mers may have any length as discussed above. There may be any number of nucleotide k-mers as discussed above. The one or more k-mers can preferably be associated with a phenotype of the organism. The invention further provides one or more nucleotide k-mers capable of

distinguishing an organism from one ore more different organism, wherein the nucleotide k- mer(s) are from the parts of the organism's genome which encode the ribosomal protein subunits, and wherein the nucleotide k-mer(s) are from about 10 to about 40 nucleotides in length. The nucleotide k-mer(s) may be of a preferred number, length or other characteristic as described above.

The invention also provides an oligonucleotide probe which comprises a sequence which is complementary to a nucleotide k-mer of the invention. The invention also provides an oligonucleotide probe which comprises a sequence which is the reverse complement of a nucleotide k-mer of the invention. The invention also provides a plurality of oligonucleotide probes each of which comprises a sequence which is complementary to one of the nucleotide k- mers in a population of two or more nucleotide k-mers of the invention. The invention also provides a plurality of oligonucleotide probes each of which comprises a sequence which is the reverse complement of one of the nucleotide k-mers in a population of two or more nucleotide k- mers of the invention.

Oligonucleotides are short nucleotide polymers which typically have about 50 or fewer nucleotides, such about 40 or fewer, about 30 or fewer or about 20 or fewer nucleotides. The sequence in the oligonucleotide probe/probes which is complementary to or the reverse complement of a nucleotide k-mer is typically the same length as the k-mer, such as from about 10 to about 40 nucleotides. The sequence may have a length of from about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, or about 19 nucleotides to about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38 or about 39 nucleotides.

Preferred sequence lengths include, but are not limited to, about 17, about 20, about 25 and about 30 nucleotides.

The oligonucleotide probe/probes may comprise any of the nucleotide discussed above. The nucleotides are preferably selected from AMP, TMP, GMP, UMP, dAMP, dTMP, dGMP or dCMP.

The nucleotides may contain additional modifications. In particular, suitable modified nucleotides include, but are not limited to, 2'amino pyrimidines (such as 2'-amino cytidine and 2'-amino uridine), 2'-hyrdroxyl purines (such as , 2'-fluoro pyrimidines (such as 2'- fluorocytidine and 2'fluoro uridine), hydroxyl pyrimidines (such as 5'-a-P-borano uridine), 2'- O-methyl nucleotides (such as 2'-0-methyl adenosine, 2'-0-methyl guanosine, 2'-0-methyl cytidine and 2'-0-methyl uridine), 4'-thio pyrimidines (such as 4'-thio uridine and 4'-thio cytidine) and nucleotides have modifications of the nucleobase (such as 5-pentynyl-2'-deoxy uridine, 5-(3-aminopropyl)-uridine and l,6-diaminohexyl-N-5-carbamoylmethyl uridine).

One or more nucleotides in the oligonucleotide probe/probes can be oxidized or methylated.

The nucleotides in the oligonucleotide probe/probes may be attached to each other in any manner. The nucleotides may be linked by phosphate, 2'0-methyl, 2' methoxy-ethyl, phosphoramidate, methylphosphonate or phosphorothioate linkages. The nucleotides are typically attached by their sugar and phosphate groups as in nucleic acids. The nucleotides may be connected via their nucleobases as in pyrimidine dimers.

The oligonucleotide probe/probes can be a nucleic acid, such as deoxyribonucleic acid (DNA) or a ribonucleic acid (RNA). The polynucleotide may be any synthetic nucleic acid known in the art, such as peptide nucleic acid (PNA), glycerol nucleic acid (GNA), threose nucleic acid (TNA), locked nucleic acid (LNA), morpholino nucleic acid or other synthetic polymers with nucleotide side chains.

The oligonucleotide probe/probes may be single stranded. The oligonucleotide probe/probes may be double stranded. The oligonucleotide probe/probes may compirse a hairpin. Oligonucleotides may be synthesised using standard techniques known in the art.

The oligonucleotide probe/probes preferably comprise a sequence which complementary to or the reverse complement of the k-mer/k-mers through Watson and Crick base pairing. If the nucleotide k-mer/k-mers is/are derived from an organism whose genome is based on

deoxyribonucleic acid (DNA), the probe/probes preferably comprise(s) dAMP, dTMP, dGMP and dCMP. If the nucleotide k-mer/k-mers are derived from an organism whose genome is based on ribonucleic acid (RNA), the probe/probes preferably comprise(s) AMP, TMP, GMP and UMP.

Preferably, the probe or probes of the invention is/are detectably-labelled. The detectable label preferably allows the presence or absence of a hybridization product formed by specific hybridization between the probe and the complementary nucleotide k-mer (and thereby the presence or absence of the nucleotide k-mer) to be determined. Any label can be used. Suitable labels include, but are not limited to, fluorescent molecules, radioisotopes, e.g. ¹²⁵1, ³⁵S, enzymes, antibodies and linkers such as biotin.

The probe can be a molecular beacon probe comprising a fluroescent label at one end and a quenching molecule at the other. In the absence of the complementary nucleotide k-mer, the probe forms a hairpin loop and the quenching molecule is brought into close proximity with the fluorescent label so that no signal can be detected. Upon hybridization of the probe to the nucleotide k-mer, the loop unzips and the fluorescent molecule is separated from the quencher such that a signal can be detected. Suitable fluorescent molecules and quencher for use in molecular beacons are known in the art. These include, but are not limited to, the fluorophores carboxyfluorsecein (FAM) and HEX and the quenchers dabcyl, DDQ1 and DDQ2.

The probe can be a scorpion probe, which is a probe linked to primer. The primer part of the probe can be designed to amplify the nucleotide k-mer to be detected and the probe part can be designed to detect the amplified nucleotide k-mer. Scorpion probes are well-known in the art. They are described in, for example, Whitcombe et al. (Nat. Biotechnol., 1994; 17: 804- 807).

The probe/probes may be attached to or immobilised on a support using any technology which is known in the art. Suitable solid supports are well-known in the art and include plates, such as multi-well plates, filters, membranes, beads, chips, pins, array, dipsticks, nanoparticles and porous carriers. The probe may be used as part of an array-based detection method.

The invention also provides a support having attached thereto or immobilised thereon a oligonucleotide probe of the invention or a plurality of oligonucleotide probes of the invention. The support preferably comprises a chip, pin, array or dipstick.

Detection method

The invention also provides a method for detecting the presence or absence of an organism in a sample. The method comprises detecting the presence or absence in the sample of one or more nucleotide k-mers which are capable of distinguishing the organism from one or more different organisms, such as all different organisms. The one or more k-mers are characteristic of the organism. The one or more k-mers are specific for the organism.

The presence of the one or more nucleotide k-mers is indicative of the presence of the organism in the sample. The absence of the one or more nucleotide k-mers is indicative of the absence of the organism from the sample. The method is preferably for detecting the presence or absence of an organism with a phenotype in a sample and the method comprises detecting the presence or absence in the sample of one or more nucleotide k-mers which are associated with the phenotype. The method preferably comprises the rapid detection of the presence or absence in the sample of one or more nucleotide k-mers.

The method may be carried out on any suitable sample. The method is typically carried out on a sample that is known to contain or suspected to contain the organism. Alternatively, the invention may be carried out on a sample to confirm the presence of an organism whose presence in the sample is known or expected.

The sample may be a biological sample. The method may be carried out in vitro using a sample obtained from or extracted from any organism or microorganism. The organism or microorganism is typically archaeal, prokaryotic or eukaryotic and typically belongs to one of the five kingdoms: plantae, animalia, fungi, monera and protista. The invention may be carried out in vitro on a sample obtained from or extracted from any virus. The sample is preferably a fluid sample. The sample typically comprises a body fluid of the patient. The sample may be urine, lymph, saliva, mucus or amniotic fluid but is preferably blood, plasma or serum.

Typically, the sample is human in origin, but alternatively it may be from another mammal animal such as from commercially farmed animals such as horses, cattle, sheep, fish, chickens or pigs or may alternatively be pets such as cats or dogs. Alternatively, the sample may be of plant origin, such as a sample obtained from a commercial crop, such as a cereal, legume, fruit or vegetable, for example wheat, barley, oats, canola, maize, soya, rice, rhubarb, bananas, apples, tomatoes, potatoes, grapes, tobacco, beans, lentils, sugar cane, cocoa, cotton.

The sample may be a non-biological sample. The non-biological sample is preferably a fluid sample. Examples of non-biological samples include surgical fluids, water such as drinking water, sea water or river water, and reagents for laboratory tests.

The sample is typically processed prior to being used in the invention, for example by centrifugation or by passage through a membrane that filters out unwanted molecules or cells, such as red blood cells. The sample may be measured immediately upon being taken. The sample may also be typically stored prior to assay, preferably below -70°C.

The one or more nucleotide k-mers are present in the genome of the organism. The method typically comprises extracting the genomic material, such as DNA or RNA, from the sample before the presence or absence of the one or more nucleotide k-mers is detected. The genomic material can be extracted using routine methods known in the art. For instance, commercially available extraction kits may be used such as MycXtra (Myconostica, UK), QIAamp Blood mini kit (Qiagen, Germany), QIAamp DNA mini kit (Qiagen, Germany) and BiOstic Bacteremia DNA Isolation kit (MoBio, USA). Suitable methods of extracting RNA are disclosed in the art such as the commercially available RNeasy mini kit (Qiagen, Germany).

The method preferably comprises the step of amplifying the one or more nucleotide k- mers. In one embodiment, the one or more nucleotide k-mers are amplified before their presence or absence is detected. In another embodiment, the one or more nucleotide k-mers are amplified in real time as their presence is determined. Real-time methods are described in the art.

In one embodiment, only the one or more nucleotide k-mers to be detected are amplified. In other embodiments, the one or more nucleotide k-mers to be detected is amplified as part of a much larger length of genomic material, such as DNA or RNA. Sequences of genomic material, such as DNA or RNA, having at least about 50, at least about 70, at least about 90, at least about 110, at least about 120, at least about 130, at least about 140, at least about 150, at least about 160, at least about 170, at least about 180, at least about 190, at least about 200, at least about 250, at least about 300, at least about 400 or at least about 500 nucleotides and comprising the region to be detected can be amplified. For example, sequences having from 10 to 2000, from 20 to 1500, from 50 to 1000 or from 100 to 500 nucleotides can be amplified.

The one or more nucleotide k-mers can be amplified using routine methods that are known in the art. The amplification of DNA is preferably carried out using polymerase chain reaction (PCR), nucleic acid sequence based analysis (NASBA) or Loop-mediated isothermal amplification (LAMP). RNA can be amplified using routine methods in the art, such as reverse transcription-PCR.

The one or more nucleotide k-mers can be detected using any method known in the art.

The one or more k-mers may be detected using next generation sequencing such as nanopore sequencing disclosed in WO 2012/107778, WO 2013/014451, WO 2013/121224 and WO 2013/153359.

The one or more nucleotide k-mers can be detected using TaqMan PCR or TaqMan realtime PCR. These techniques are well-known in the art.

The one or more nucleotide k-mers are preferably detected using the probe or probes of the invention. Typically, the detecting comprises contacting the probe or probes with the sample under conditions in which the probe hybridizes to the one or more nucleotide k-mers, if present, and determining the presence or absence of the hybridization product. The presence of the hybridization product indicates the presence of the one or more nucleotide k-mers. Conversely, the absence of the hybridization product indicates the absence of the one or more nucleotide k- mers. Conditions that permit the hybridization are well-known in the art (for example,

Sambrook et al., 2001, Molecular Cloning: a laboratory manual, 3rd edition, Cold Spring Harbour Laboratory Press; and Current Protocols in Molecular Biology, Chapter 2, Ausubel et al., Eds., Greene Publishing and Wiley-lnterscience, New York (1995)). The method of the invention can be carried out under low stringency conditions, for example in the presence of a buffered solution of 30 to 35% formamide, 1 M NaCl and 1 % SDS (sodium dodecyl sulfate) at 37°C followed by a wash in from IX (0.1650 M NaCl) to 2X (0.33 M NaCl) SSC (salt sodium citrate) at 50°C. The method of the invention can be carried out under moderate stringency conditions, for example in the presence of a buffer solution of 40 to 45% formamide, 1 M NaCl, and 1 % SDS at 37°C, followed by a wash in from 0.5X (0.0825 M NaCl) to IX (0.1650 M NaCl) SSC at 55°C. The method of the invention can be carried out under high stringency conditions, for example in the presence of a buffered solution of 50% formamide, 1 M NaCl, 1% SDS at 37°C, followed by a wash in 0.1X (0.0165 M NaCl) SSC at 60°C. If more than one of nucleotide k-mers are being detected simultaneously, the different probes used to detect the different nucleotide k-mers may be labelled with different labels.

Probes having different labels are preferable when different nucleotide k-mers are being detected simultaneously in the same volume of sample. When the two or more different nucleotide k- mers are being detected in the same volume of sample, it must be possible to distinguish between the different labels and hence detect the different nucleotide k-mers. For instance, fluorescent molecules that emit different wavelengths of light can be used. A suitable group of fluorescent labels, each of which can be simultaneously detected, is HEX hexachloro fluorescein

phosphoramidite (HEX), carboxyfluorescein (FAM), Cy(R)5 and Texas Red(R). Other suitable groups of labels are known in the art.

Any of the probes and supports of the invention may be used in the detection method. The presence or absence of multiple different organisms may be simultaneously detected using array-based methods. Specific nucleotide k-mers

The inventors have also identified specific k-mer clusters in various organisms, as described in Example 1. These are shown as SEQ ID NOs: 1 to 213 in the sequence listing. Each cluster comprises various k-mers which may be used to distinguish the relevant bacterium from one or more different organisms, such as one or more different bacteria.

The invention therefore provides a nucleotide k-mer capable of distinguishing an organism from one or more different organisms, wherein the k-mer comprises (or consists of) about 10 or more consecutive nucleotides from any one of the sequences shown in SEQ ID NOs: 1 to 213. The k-mer may be any length. The k-mer may comprise at least about 15, at least about 20, at least about 25, at least about 30, or at least about 35 consecutive nucleotides from any one of the sequences shown in SEQ ID NOs: 1 to 213.

The k-mer preferably comprises (or consists of) from about 10 to about 40 consecutive nucleotides from any of the sequences shown in SEQ ID NOs: 1 to 213. The k-mer is preferably from about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, or about 19 nucleotides to about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38 or about 39 consecutive nucleotides from any one of SEQ ID NOs: 1 to 213.

Preferred k-mer lengths include, but are not limited to, about 17, about 20, about 25 and about 30 consecutive nucleotides from any one of the sequences shown in SEQ ID NOs: 1 to 213. The k-mer preferably comprises (or consists of) any of the consecutive nucleotides listed in the last column in Table 3. The locations of the preferred k-mers are listed in this column as SEQ ID NO:/start nucleotide-end nucleotide. For instance, the first preferred k-mer is located at 1/18-34. This means it corresponds to nucleotides 18 to 34 of SEQ ID NO: 1. The second preferred k-mer is located at 2/9-25. This means it corresponds to nucleotides 9 to 25 of SEQ ID NO: 2 and so on.

The nucleotide k-mer of the invention can be associated with a phenotype of the organism. This is discussed in detail above.

Preferably, the k-mer is from one of SEQ ID NOs: 1 to 5 and is capable of distinguishing Aggregatibacter actinomycetemcomitans from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 6 to 10 and is capable of distibuising Bacillus anthracis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 11 to 15 and is capable of

distinguishing Bacillus licheniformis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 16 to 20 and is capable of

distinguishing Bacteroides fragilis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 21 to 25 to 5 and is capable of distinguishing Bartonella henselae from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 26 to 30 and is capable of

distinguishing Bordetella pertussis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 31 to 35 and is capable of

distinguishing Borrelia burgdorferi from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 36 to 40 and is capable of

distinguishing Brucella abortus from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 41 to 44 and is capable of

distinguishing Campylobacter jejuni from one or more different organisms, such as all different organisms. Preferably, the k-mer is from one of SEQ ID NOs: 45 to 49 and is capable of distinguishing Chlamydia trachomatis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 50 to 54 and is capable of distinguishing Chlamydophila pneumonia from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 55 to 59 and is capable of distinguishing Clostridium difficile from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 60 to 64 and is capable of distinguishing Clostridium perfringens from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 65 to 69 and is capable of distinguishing from Enterobacter aerogenes from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 70 to 74 and is capable of distinguishing from Enter -ococcus faecalis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 75 to 79 and is capable of distinguishing from Ente -ococcus faecium from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 80 to 84 and is capable of distinguishing Francisella tularensis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from SEQ ID NO: 85 and is capable of distinguishing

Haemophilus influenza from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 86 and 87 and is capable of distinguishing Helicobacter pylori from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 88 to 92 and is capable of distinguishing Klebsiella oxytoca

Preferably, the k-mer is from one of SEQ ID NOs: 93 to 97 and is capable of distinguishing Legionella pneumophila from one or more different organisms, such as all different organisms. Preferably, the k-mer is from one of SEQ ID NOs: 98 to 102 and is capable of distinguishing Listeria monocytogenes from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 103 to 107 and is capable of distinguishing Moraxella catarrhalis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 108 to 112 and is capable of distinguishing Mycobacterium avium from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 113 to 116 and is capable of distinguishing Mycobacterium bovis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 117 to 121 and is capable of distinguishing Mycoplasma genitalium from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 122 to 126 and is capable of distinguishing Mycoplasma pneumonia from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 127 to 131 and is capable of distinguishing Neisseria gonorrhoeae from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 132 to 136 and is capable of distinguishing Neisseria meningitides from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 137 to 141 and is capable of distinguishing Porphyromonas gingivalis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 142 to 146 and is capable of distinguishing Proteus mirabilis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 147 to 151 and is capable of distinguishing Pseudomonas aeruginosa from one or more different organisms, such as all different organisms. Preferably, the k-mer is from one of SEQ ID NOs: 152 to 156 and is capable of distinguishing Salmonella enterica from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 157 to 161 and is capable of distinguishing Serratia marcescens from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 162 to 166 and is capable of distinguishing Staphylococcus aureus from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 167 to 171 and is capable of distinguishing Staphylococcus epidermidis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 172 to 176 and is capable of distinguishing Staphylococcus haemolyticus from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from SEQ ID NO: 177 and is capable of distinguishing

Stenotrophomonas maltophilia from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 178 to 182 and is capable of distinguishing Streptococcus mutans from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 183 to 187 and is capable of distinguishing Streptococcus pyogenes from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 188 to 192 and is capable of distinguishing Streptococcus salivarius from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 193 to 197 and is capable of distinguishing Streptococcus sanguinis from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from one of SEQ ID NOs: 198 to 202 and is capable of distinguishing Treponema pallidum from one or more different organisms, such as all different organisms.

Preferably, the k-mer is from SEQ ID NO: 203 and is capable of distinguishing Vibrio cholera from one or more different organisms, such as all different organisms. Preferably, the k-mer is from one of SEQ ID NOs: 204 to 208 and is capable of distinguishing Vibrio parahaemolyticus from one or more different organisms, such as all different organisms, or

Preferably, the k-mer is from one of SEQ ID NOs: 209 to 213 and is capable of distinguishing Yersinia enterocolitica from one or more different organisms, such as all different organisms.

The preferred k-mers are preferably capable of distinguishing the bacterium of interest {i.e. the bacterium from which they derived) from one or more different bacteria, such as all different bacteria. The preferred k-mers are preferably capable of distinguishing the bacterium of interest {i.e. the bacterium from which they are derived) from one or more of the different bacteria in Table 3, such as all the different bacteria in Table 3.

The invention also provides an oligonucleotide probe which comprises (or consists of) a sequence which is complementary to a nucleotide k-mer of the invention. Probes are discussed in more detail above. The probe preferably comprises (or consists of) a sequence which is complementary to one of the k-mers listed in the last column in Table 3.

The probe is preferably detectably labelled and/or is a molecular beacon probe.

The invention also provides a support having attached thereto an oligonucleotide probe of the invention. The support comprises a chip, pin, array or dipstick.

The invention also provides a method for detecting the presence or absence of an organism in a sample, comprising detecting the presence or absence in the sample of a nucleotide k-mer of the invention. The nucleotide k-mer is capable of distinguishing the organism from one or more different organisms, such as all different organisms. The nucleotide k-mer is characteristic of the organism. The nucleotide k-mer is specific for the organism. Any of the combinations of specific organisms and specific k-mers disclosed above may be used in the method.

The presence of the nucleotide k-mer is indicative of the presence of the organism in the sample. The absence of the nucleotide k-mer is indicative of the absence of the organism from the sample. The method may comprise detecting two or more k-mers of the invention, such as three or more, four or more or five or more k-mers of the invention. The organism is typically one of the organisms shown in Table 3 and the k-mer is typically derived from one of the cluster sequences in this organism. The k-mer is preferably one of the k-mers listed in the last column in Table 3.

The method is preferably for detecting the presence or absence of the organism with a particular phenotype in a sample and the method comprises detecting the presence or absence in the sample of a nucleotide k-mer of the invention which is associated with the phenotype. Detection methods are discussed in more detail above and any of the embodiments discussed above equally apply to the method of using the specific k-mers of the invention.

Example 1

Overview

This Example describes the computational methodology to efficiently identify collections of short regions of DNA (k-mers) from protein-encoding ribosomal genes that can be used to identify bacterial species. Initial findings show that it is possible to identify a relatively small number of k-mers from these collections that are both highly species specific and have high coverage in terms of the number of isolates within the species that can be reliably identified. The results have implications for practical applications such as rapid diagnostic tests for pathogenic bacteria. Methods

DNA sequence data for bacterial isolates

The whole genome sequence data for bacterial isolates were downloaded from the NCBI Nucleotide database (2,312 isolates) and the NCBI Assembly database (13,307 isolates) and entered into the BIGSdb rMLST isolate database. Plasmid sequences were also included where appropriate. A further 71,348 genomes were derived from assembling short-read sequencing data for isolate entries in the European Nucleotide Archive Sequence Read Archive (ENA-SRA) using the Velvet assembly algorithm. In total, 86,967 isolates with associated whole genome data were used in this study, representing 3,062 different species. The species annotation of each isolate was validated using an analysis of the protein-encoding ribosomal genes to a set of reliable isolate references.

Identification of protein-encoding ribosomal genes

The protein-encoding ribosomal genes within the isolates in the dataset were identified through a series of BLAST sequence searches against the BIGSdb rMLST sequence definition database. This is an iterative process known colloquially as scanning and tagging. The sequence definition database is based on 'seed' alleles from the ribosomal genes of reference genomes of many different species and has now grown to include over 6,000 alleles for each ribosomal gene. The genome sequence of each isolate is searched against the rMLST sequence definition database. DNA regions with identical matches to existing alleles and with valid start and stop codons are automatically assigned an allele identifier and the start/stop positions of the allele are recorded. Non-identical matches can be identified by manual inspection, helped by an algorithm that looks for start and stop codons within 18 nucleotides of the ends of the alignment with the closest matching allele. Allele matches with a high sequence identity (at least 95%) and at least 90% overlap to an existing allele are added to the sequence definition database and can therefore be automatically identified in subsequent scans. Allele matches below these thresholds require additional validation to confirm that the gene in question has been correctly identified. The protein-encoding ribosomal gene sequences of the 86,967 isolate dataset were extracted based on these allele definitions and used to create a dataset called the 'protein-encoding ribosomal DNA sequence dataset' . The whole genomes of the 86,967 isolates were extracted and form a dataset called 'whole genome DNA sequence dataset'.

Single species k-mer library generation

In order to identify short regions of DNA (k-mers) from protein-encoding ribosomal gene sequences that uniquely identify a particular species of bacteria, four computational stages were defined: 1) k-mer extraction, 2) k-mer scanning and species filtering, 3) k-mer property filtering and 4) final k-mer selection. Each stage is described in more detail below and shown in Figure 1.

Figure 1 : A flowchart describing the four computational stages required to generate a small single species k-mer library from protein-encoding ribosomal gene DNA.

Stage 1: K-mer Extraction

K-mers of a particular size, N (e.g. 20 nucleotides) were extracted from the protein- encoding ribosomal DNA sequence dataset by sliding a window of N bases along the DNA sequence of each protein-encoding gene and recording each k-mer the first time it was observed. K-mers containing non-standard nucleotides (non-ATGC) were not included. This creates a unique list of 'protein-encoding ribosomal DNA k-mers' .

Stage 2: K-mer Scanning and Species Filtering

The isolate sequences from the whole genome DNA sequence dataset were scanned one after another against the list of protein-encoding ribosomal DNA k-mers. This was achieved by sliding along each contiguous sequence with a window of N bases (the same size as the list of k- mers) and looking to see if the exact k-mer existed in the input k-mer list. Presence of a k-mer resulted in the internal recording of the k-mer and the species of the isolate in question. If the number of species observed for a particular k-mer exceeded one, the k-mer was removed from the ribosomal k-mer list. This process creates a library of 'single species k-mers' with each k- mer in the library associated with a single species definition.

Stage 3: K-mer Property Filtering (optional)

Filtering steps can now be applied to the single species k-mer library to remove sequences that have undesirable properties in an experimental context, for example long runs of the same nucleotide that may be difficult to sequence accurately. Another example would be to remove any k-mers that are present in the human genome as these may cause false positive matches in a diagnostic library used to analyse human samples. These steps are optional but should take place before final k-mer selection.

Stage 4: Final K-mer Selection

At this stage there can be millions of unique k-mers in the single species k-mer library. This may be acceptable for computational applications, but a smaller collection of k-mers (e.g. hundreds or thousands) may be desirable in some experimental contexts e.g. microarrays or genotyping by species-specific primers.

(4 A) Total Coverage Single Species K-mers

To reduce the redundancy in the single species k-mer library, the first step is to identify the k-mers that have total coverage of the isolates in the species. If these are present, the next step is to map the total coverage k-mers back onto the original protein-encoding ribosomal genes for a representative isolate for each species. Any contiguous region of ribosomal sequence containing at least one k-mer is defined as a k-mer cluster. A limited number of clusters can then be selected (for example 5 per species) and a single representative k-mer can be selected from each cluster.

Using the above 86,967 isolate dataset, we performed stages 1 and 2 for k-mer lengths 17, 20, 25 and 30 and then filtered out any k-mer that matched human DNA in stage 3. Total coverage single species k-mers were then identified in stage 4. Summary statistics for each stage of the process are show in Table 1.

K-mer Unique K-mers Unique Single Unique Single Unique Single Species Length from K-mer Species Species K-mers K-mers with Total extraction K-mers Filtered to Remove Coverage of the Isolate (Stage 1) (Stage 2) Human DNA Population (Stage 4)

(Stage 3)

17 32,479,274 4,794,410 4,098,981 3,343,710 20 37,446,505 25,962,711 25,759,215 20,878,341

25 42,727,248 35,034,225 35,032,769 27,858,947

30 46,063,946 39,036,215 39,036, 166 30,432,337

Table 1. Number of k-mers at each stage of the process for four different k-mer lengths.

The total coverage non-human k-mers for the four k-mer lengths were combined and mapped onto a set of protein-encoding ribosomal DNA sequences from representative isolates (one per species). The cluster density of each k-mer cluster was calculated according to the definition below and the number of k-mer lengths present was recorded. The clusters were ranked by 1) number of k-mer lengths present in the cluster (descending) 2) cluster density (descending). The 5 highest ranked k-mer clusters on different ribosomal genes for each species were selected.

Cluster Density Definition: The number of observed k-mers in each cluster divided by the maximum number of possible k-mers for the cluster length (expressed as a percentage).

Example: For the 4 k-mer lengths used in the study (17, 20, 25, 30), a cluster with length 30 can have 1 x 30nt, 6 x 25nt, 11 x 20nt and 14 x 17nt. This equals a maximum of 32 possible k-mers for this cluster length. If all 32 possible k-mers were present the cluster density would be 100%. If 20 different k-mers were found in the cluster, the cluster density would be 20/32 x 100% = 62.5%.

Identifying a representative k-mer within each k-mer cluster for each k-mer length was performed as follows: Each nucleotide position in the k-mer cluster was assigned a number equal to the total number of k-mers mapping to that position (across all sampled lengths). Each k-mer in the cluster was scored based on the sum of the nucleotide position scores. The representative k-mer for each k-mer length was the single species k-mer with the highest total score for that length.

(4B) Partial Coverage Single Species K-mers

If total coverage k-mers are not present in the single species k-mer library for a particular species, then the minimal number of partial coverage k-mers for that species can be identified by formalising the scenario as a 'set covering problem' and this can be solved by implementing a greedy algorithm. For example, in a simple scenario with 5 isolates in a particular species, k-mer A is found in isolates 1, 2, 3, k-mer B matches isolates 2 and 4 and k-mer C matches isolates 3 and 4 and k-mer D matches isolates 4 and 5. k-mers A and D would be the minimum set of k- mers to achieve 100% coverage of the isolates in the species (5 isolates). The greedy algorithm selects sets according to the rule that at each stage, choose the set/k-mer that contains the largest number of uncovered elements (matched isolates). In Computer Science terms (as described in Wikipedia, http://en.wikipedia.org/wiki/Set_coverj3roblem): Universe = { 1,2,3,4,5}, Sets S = { { 1,2,3 }, {2,4}, {3,4}, {4,5} }. We can cover all the elements of the universe with two sets, { { 1,2,3 }, {4,5} }.

Results

Species Summary (Table 2)

Summary of the results for each of the pathogenic species (46) including external database references of a representative genome (NCBI) and the number of selected k-mers at each k-mer length.

Species lilililiTot NCBI NCBI NCBI Nucleotide me me me me al Nucleotid Genbank Entry

rs rs rs rs k- e Identifier Description w/ w/ w/ w/ mer Accession

17n 20n 25n 30n s

t t t t

Aggregatibacter 5 5 5 5 20 CP001733 GL261412 Aggregatibacter actinomycetemco .1 053 actinomycetemco mitans mitans Dl l S-1, complete genome

Bacillus anthracis 5 5 5 5 20 AE01733 GL500829 Bacillus anthracis

4.2 67 str. 'Ames

Ancestor', complete genome

Bacillus 5 5 5 5 20 NC 0063 GL527838 Bacillus licheniformis 22.1 55 licheniformis

DSM 13 = ATCC 14580, complete genome

Bacteroides 5 5 5 5 20 NC 0063 GL537112 Bacteroides fragilis 47.1 91 fragilis YCH46

DNA, complete genome

Bartonella 5 5 5 5 20 NC 0059 GL494748 Bartonella henselae 56.1 31 henselae str.

Houston- 1 chromosome, complete genome

Bordetella 5 5 5 5 20 BX47024 GL335910 Bordetella pertussis 8.1 69 pertussis strain

Tohama I, complete genome

Borrelia 5 5 5 5 20 NC 0117 GI:218249 Borrelia burgdorferi 28.1 165 burgdorferi ZS7, complete genome

Brucella abortus 4 5 5 5 19 AM04026 GI: 826150 Brucella

4.1 33 melitensis biovar

Abortus 2308 chromosome I, complete sequence, strain 2308

Campylobacter 2 3 2 2 9 AL11116 GL304071 Campylobacter jejuni 8.1 39 jejuni subsp.

jejuni NCTC 11168 complete genome

Chlamydia 5 5 5 5 20 AM88417 GI: 165929 Chlamydia trachomatis 6.1 961 trachomatis strain

L2/434/Bu complete genome

Chlamydophila 5 5 5 5 20 AE00216 GI: 120572 Chlamydophila pneumoniae 1.1 10 pneumoniae

AR39, complete genome

Clostridium 5 5 5 5 20 NC 0090 GI: 126697 Peptoclostridium difficile 89.1 566 difficile 630, complete genome

Clostridium 5 5 5 5 20 NC 0082 GI: 110798 Clostridium perfringens 61.1 562 perfringens

ATCC 13124, complete genome

Enterobacter 5 5 5 5 20 NC 0156 GL336246 Enterobacter aerogenes 63.1 508 aerogenes KCTC

2190

chromosome, complete genome

Enterococcus 5 5 5 5 20 AE01683 GL293501 Enterococcus faecalis 0.1 90 faecalis V583, complete genome

Enterococcus 5 5 5 5 20 NC 0179 GL389867 Enterococcus faecium 60.1 183 faecium DO

chromosome, complete genome

Francisella 5 5 5 5 20 CP000803 GI: 156251 Francisella tularensis .1 972 tularensis subsp.

holarctica FTNF002-00, complete genome

Haemophilus 1 1 1 1 4 CP000057 GI: 156617 Haemophilus influenzae .2 157 influenzae 86- 028NP, complete genome

Helicobacter 1 1 0 0 2 CP001173 GL208431 Helicobacter pylori .1 905 pylori G27, complete genome

Klebsiella 5 5 5 5 20 NC 0166 GL375256 Klebsiella oxytoca 12.1 816 oxytoca KCTC

1686, complete genome

Legionella 5 5 5 5 20 NC 0094 GL295815 Legionella pneumophila 94.2 281 pneumophila str.

Corby, complete genome

Listeria 5 5 5 5 20 FM24271 GL225875 Listeria monocytogenes 1.1 101 monocytogenes

Clip80459 serotype 4b complete genome

Moraxella 5 5 5 5 20 CP008804 GL672768 Moraxella catarrhalis .1 103 catarrhalis strain

25240, complete genome

Mycobacterium 5 5 5 5 20 CP000479 GL 118163 Mycobacterium avium .1 506 avium 104,

complete genome

Mycobacterium 2 4 4 4 14 BX24833 GI:317425 Mycobacterium bovis 3.1 09 bovis subsp. bovis

AF2122/97 complete genome

Mycoplasma 5 5 5 5 20 L43967.2 GI: 846261 Mycoplasma genitalium 23 genitalium G37, complete genome

Mycoplasma 5 5 5 5 20 NC 0009 GI: 135077 Mycoplasma pneumoniae 12.1 39 pneumoniae

M129

chromosome, complete genome

Neisseria 5 5 5 5 20 AE00496 GI: 597173 Neisseria gonorrhoeae 9.1 68 gonorrhoeae FA

1090, complete genome

Neisseria 3 5 5 5 18 AM42180 GI: 120865 Neisseria meningitidis 8.1 607 meningitidis serogroup C FAM18 complete genome

Porphyromonas 5 5 5 5 20 NC 0107 GI: 188993 Porphyromonas gingivalis 29.1 864 gingivalis ATCC

33277, complete genome

Proteus mirabilis 5 5 5 5 20 NC 0105 GI: 197283 Proteus mirabilis

54.1 915 strain HI4320, complete genome

Pseudomonas 5 5 5 5 20 NC 0117 GL218888 Pseudomonas aeruginosa 70.1 746 aeruginosa

LESB58 complete genome sequence

Salmonella 3 5 4 1 13 NC 0031 GI: 167589 Salmonella enterica 98.1 93 enterica subsp.

enterica serovar Typhi str. CT18, complete genome

Serratia 5 5 5 5 20 NC 0200 GL440228 Serratia marcescens 64.1 796 marcescens

FGI94, complete genome

Staphylococcus 2 5 5 4 16 NC 0076 GI: 827497 Staphylococcus aureus 22.1 77 aureus RF 122 complete genome

Staphylococcus 5 5 5 5 20 NC 0044 GL274669 Staphylococcus epidermidis 61.1 18 epidermidis

ATCC 12228 chromosome, complete genome

Staphylococcus 5 5 5 5 20 AP006716 GL684457 Staphylococcus haemolyticus .1 25 haemolyticus

JC SCI 435 DNA, complete genome

Stenotrophomona 0 1 1 1 3 NC 0109 GI: 190572 Stenotrophomona s maltophilia 43.1 091 s maltophilia

K279a complete genome, strain K279a

Streptococcus 5 5 5 5 20 AP010655 GL254996 Streptococcus mutans .1 425 mutans NN2025

DNA, complete genome

Streptococcus 5 5 5 5 20 NC 0189 GL409913 Streptococcus pyogenes 36.1 960 pyogenes A20, complete genome

Streptococcus 5 5 5 5 20 NC 0157 GL340397 Streptococcus salivarius 60.1 867 salivarius

CCHSS3 complete genome

Streptococcus 5 5 5 5 20 CP000387 GI: 125496 Streptococcus sanguinis .1 804 sanguinis SK36, complete genome

Treponema 5 5 5 5 20 NC 0168 GL378974 Treponema pallidum 44.1 633 pallidum subsp.

pallidum DAL-1, complete genome

Vibrio cholerae 0 1 1 0 2 NC 0125 GL227080 Vibrio cholerae

78.1 / 237 / M66-2

NC 0125 GL227811 chromosome I, 80.1 634 complete

sequence / Vibrio cholerae M66-2 chromosome II, complete sequence

Vibrio 5 5 5 5 20 NC 0046 GL288967 Vibrio parahaemolyticus 03.1 74 parahaemolyticus

RIMD 2210633 chromosome 1, complete sequence

Yersinia 5 5 5 5 20 AM28641 GI: 122087 Yersinia enter ocolitica 5.1 364 enterocolitica subsp.

enterocolitica 8081 complete genome

Grand Total 198 211 208 203 820 K-mer Cluster Summary

Each k-mer cluster is listed in the sequence listing (SEQ ID NOs 1 to 213). Preferred K-mer Summary (Table 3)

Preferred k-mers are listed in the Table below with their sequence length, the k-mer cluster sequence from which they are derived and their location within that sequence. For instance, the first preferred k-mer is located at 1/18-34. This means it corresponds to nucleotides 18 to 34 of SEQ ID NO: 1. The second preferred k-mer is located at 2/9-25. This means it corresponds to nucleotides 9 to 25 of SEQ ID NO: 2 and so on.

Borrelia burgdorferi 17 31 31/12-28

Borrelia burgdorferi 17 32 32/10-26

Borrelia burgdorferi 17 33 33/28-44

Borrelia burgdorferi 17 34 34/16-32

Borrelia burgdorferi 17 35 35/34-50

Brucella abortus 17 36 36/28-44

Brucella abortus 17 37 37/23-39

Brucella abortus 17 38 38/19-35

Brucella abortus 17 40 40/22-38

Campylobacter jejuni 17 41 41/4-20

Campylobacter jejuni 17 43 43/7-23

Chlamydia trachomatis 17 45 45/16-32

Chlamydia trachomatis 17 46 46/9-25

Chlamydia trachomatis 17 47 47/13-29

Chlamydia trachomatis 17 48 48/73-89

Chlamydia trachomatis 17 49 49/40-56

Chlamydophila pneumoniae 17 50 50/31-47

Chlamydophila pneumoniae 17 51 51/28-44

Chlamydophila pneumoniae 17 52 52/120-136

Chlamydophila pneumoniae 17 53 53/17-33

Chlamydophila pneumoniae 17 54 54/91-107

Clostridium difficile 17 55 55/12-28

Clostridium difficile 17 56 56/43-59

Clostridium difficile 17 57 57/24-40

Clostridium difficile 17 58 58/9-25

Clostridium difficile 17 59 59/28-44

Clostridium perfringens 17 60 60/17-33

Clostridium perfringens 17 61 61/48-64

Clostridium perfringens 17 62 62/162-178

Clostridium perfringens 17 63 63/23-39

Clostridium perfringens 17 64 64/66-82

Enterobacter aerogenes 17 65 65/24-40

Enterobacter aerogenes 17 66 66/23-39

Enterobacter aerogenes 17 67 67/27-43

Enterobacter aerogenes 17 68 68/24-40

Enterobacter aerogenes 17 69 69/23-39

Enterococcus faecalis 17 70 70/11-27

Enterococcus faecalis 17 71 71/12-28

Enterococcus faecalis 17 72 72/18-34

Enterococcus faecalis 17 73 73/25-41

Enterococcus faecalis 17 74 74/26-42

Enterococcus faecium 17 75 75/9-25

Enterococcus faecium 17 76 76/32-48

Enterococcus faecium 17 77 77/11-27

Enterococcus faecium 17 78 78/6-22

Enterococcus faecium 17 79 79/19-35 Francisella tularensis 17 80 80/16-32

Francisella tularensis 17 81 81/11-27

Francisella tularensis 17 82 82/9-25

Francisella tularensis 17 83 83/15-31

Francisella tularensis 17 84 84/17-33

Haemophilus influenzae 17 85 85/30-46

Helicobacter pylori 17 87 87/1-17

Klebsiella oxytoca 17 88 88/14-30

Klebsiella oxytoca 17 89 89/10-26

Klebsiella oxytoca 17 90 90/23-39

Klebsiella oxytoca 17 91 91/28-44

Klebsiella oxytoca 17 92 92/17-33

Legionella pneumophila 17 93 93/7-23

Legionella pneumophila 17 94 94/14-30

Legionella pneumophila 17 95 95/30-46

Legionella pneumophila 17 96 96/11-27

Legionella pneumophila 17 97 97/14-30

Listeria monocytogenes 17 98 98/7-23

Listeria monocytogenes 17 99 99/8-24

Listeria monocytogenes 17 100 100/22-38

Listeria monocytogenes 17 101 101/9-25

Listeria monocytogenes 17 102 102/22-38

Moraxella catarrhalis 17 103 103/31-47

Moraxella catarrhalis 17 104 104/37-53

Moraxella catarrhalis 17 105 105/95-111

Moraxella catarrhalis 17 106 106/62-78

Moraxella catarrhalis 17 107 107/22-38

Mycobacterium avium 17 108 108/17-33

Mycobacterium avium 17 109 109/19-35

Mycobacterium avium 17 110 110/6-22

Mycobacterium avium 17 111 111/50-66

Mycobacterium avium 17 112 112/22-38

Mycobacterium bovis 17 115 115/23-39

Mycobacterium bovis 17 116 116/21-37

Mycoplasma genitalium 17 117 117/8-24

Mycoplasma genitalium 17 118 118/17-33

Mycoplasma genitalium 17 119 119/110-126

Mycoplasma genitalium 17 120 120/33-49

Mycoplasma genitalium 17 121 121/132-148

Mycoplasma pneumoniae 17 122 122/26-42

Mycoplasma pneumoniae 17 123 123/38-54

Mycoplasma pneumoniae 17 124 124/56-72

Mycoplasma pneumoniae 17 125 125/70-86

Mycoplasma pneumoniae 17 126 126/14-30

Neisseria gonorrhoeae 17 127 127/12-28

Neisseria gonorrhoeae 17 128 128/19-35 Neisseria gonorrhoeae 17 129 129/18-34

Neisseria gonorrhoeae 17 130 130/12-28

Neisseria gonorrhoeae 17 131 131/14-30

Neisseria meningitidis 17 134 134/36-52

Neisseria meningitidis 17 135 135/21-37

Neisseria meningitidis 17 136 136/16-32

Porphyromonas gingivalis 17 137 137/73-89

Porphyromonas gingivalis 17 138 138/18-34

Porphyromonas gingivalis 17 139 139/8-24

Porphyromonas gingivalis 17 140 140/97-113

Porphyromonas gingivalis 17 141 141/57-73

Proteus mirabilis 17 142 142/16-32

Proteus mirabilis 17 143 143/53-69

Proteus mirabilis 17 144 144/61-77

Proteus mirabilis 17 145 145/25-41

Proteus mirabilis 17 146 146/26-42

Pseudomonas aeruginosa 17 147 147/9-25

Pseudomonas aeruginosa 17 148 148/13-29

Pseudomonas aeruginosa 17 149 149/18-34

Pseudomonas aeruginosa 17 150 150/13-29

Pseudomonas aeruginosa 17 151 151/23-39

Salmonella enterica 17 153 153/3-19

Salmonella enterica 17 154 154/2-18

Salmonella enterica 17 156 156/9-25

Serratia marcescens 17 157 157/11-27

Serratia marcescens 17 158 158/17-33

Serratia marcescens 17 159 159/14-30

Serratia marcescens 17 160 160/10-26

Serratia marcescens 17 161 161/28-44

Staphylococcus aureus 17 163 163/7-23

Staphylococcus aureus 17 165 165/7-23

Staphylococcus epidermidis 17 167 167/13-29

Staphylococcus epidermidis 17 168 168/21-37

Staphylococcus epidermidis 17 169 169/28-44

Staphylococcus epidermidis 17 170 170/10-26

Staphylococcus epidermidis 17 171 171/23-39

Staphylococcus haemolyticus 17 172 172/18-34

Staphylococcus haemolyticus 17 173 173/9-25

Staphylococcus haemolyticus 17 174 174/23-39

Staphylococcus haemolyticus 17 175 175/19-35

Staphylococcus haemolyticus 17 176 176/6-22

Streptococcus mutans 17 178 178/37-53

Streptococcus mutans 17 179 179/7-23

Streptococcus mutans 17 180 180/10-26

Streptococcus mutans 17 181 181/13-29

Streptococcus mutans 17 182 182/16-32 Streptococcus pyogenes 17 183 183/18-34

Streptococcus pyogenes 17 184 184/13-29

Streptococcus pyogenes 17 185 185/9-25

Streptococcus pyogenes 17 186 186/63-79

Streptococcus pyogenes 17 187 187/9-25

Streptococcus salivarius 17 188 188/11-27

Streptococcus salivarius 17 189 189/27-43

Streptococcus salivarius 17 190 190/19-35

Streptococcus salivarius 17 191 191/13-29

Streptococcus salivarius 17 192 192/5-21

Streptococcus sanguinis 17 193 193/8-24

Streptococcus sanguinis 17 194 194/9-25

Streptococcus sanguinis 17 195 195/37-53

Streptococcus sanguinis 17 196 196/14-30

Streptococcus sanguinis 17 197 197/20-36

Treponema pallidum 17 198 198/45-61

Treponema pallidum 17 199 199/34-50

Treponema pallidum 17 200 200/30-46

Treponema pallidum 17 201 201/28-44

Treponema pallidum 17 202 202/22-38

Vibrio parahaemolyticus 17 204 204/25-41

Vibrio parahaemolyticus 17 205 205/56-72

Vibrio parahaemolyticus 17 206 206/18-34

Vibrio parahaemolyticus 17 207 207/14-30

Vibrio parahaemolyticus 17 208 208/23-39

Yersinia enterocolitica 17 209 209/15-31

Yersinia enterocolitica 17 210 210/25-41

Yersinia enterocolitica 17 211 211/10-26

Yersinia enterocolitica 17 212 212/14-30

Yersinia enterocolitica 17 213 213/19-35

Aggregatibacter actinomycetemcomitans 20 1 1/15-34

Aggregatibacter actinomycetemcomitans 20 2 2/8-27

Aggregatibacter actinomycetemcomitans 20 3 3/26-45

Aggregatibacter actinomycetemcomitans 20 4 4/11-30

Aggregatibacter actinomycetemcomitans 20 5 5/11-30

Bacillus anthracis 20 6 6/19-38

Bacillus anthracis 20 7 7/20-39

Bacillus anthracis 20 8 8/20-39

Bacillus anthracis 20 9 9/21-40

Bacillus anthracis 20 10 10/11-30

Bacillus licheniformis 20 11 11/17-36

Bacillus licheniformis 20 12 12/10-29

Bacillus licheniformis 20 13 13/13-32

Bacillus licheniformis 20 14 14/18-37

Bacillus licheniformis 20 15 15/38-57

Bacteroides fragilis 20 16 16/27-46 Bacteroides fragilis 20 17 17/23-42

Bacteroides fragilis 20 18 18/11-30

Bacteroides fragilis 20 19 19/27-46

Bacteroides fragilis 20 20 20/24-43

Bartonella henselae 20 21 21/131-150

Bartonella henselae 20 22 22/21-40

Bartonella henselae 20 23 23/15-34

Bartonella henselae 20 24 24/106-125

Bartonella henselae 20 25 25/7-26

Bordetella pertussis 20 26 26/21-40

Bordetella pertussis 20 27 27/21-40

Bordetella pertussis 20 28 28/17-36

Bordetella pertussis 20 29 29/21-40

Bordetella pertussis 20 30 30/21-40

Borrelia burgdorferi 20 31 31/13-32

Borrelia burgdorferi 20 32 32/8-27

Borrelia burgdorferi 20 33 33/26-45

Borrelia burgdorferi 20 34 34/20-39

Borrelia burgdorferi 20 35 35/33-52

Brucella abortus 20 36 36/17-36

Brucella abortus 20 37 37/21-40

Brucella abortus 20 38 38/20-39

Brucella abortus 20 39 39/20-39

Brucella abortus 20 40 40/21-40

Campylobacter jejuni 20 41 41/2-21

Campylobacter jejuni 20 43 43/3-22

Campylobacter jejuni 20 44 44/16-35

Chlamydia trachomatis 20 45 45/14-33

Chlamydia trachomatis 20 46 46/7-26

Chlamydia trachomatis 20 47 47/11-30

Chlamydia trachomatis 20 48 48/71-90

Chlamydia trachomatis 20 49 49/39-58

Chlamydophila pneumoniae 20 50 50/29-48

Chlamydophila pneumoniae 20 51 51/28-47

Chlamydophila pneumoniae 20 52 52/118-137

Chlamydophila pneumoniae 20 53 53/16-35

Chlamydophila pneumoniae 20 54 54/89-108

Clostridium difficile 20 55 55/11-30

Clostridium difficile 20 56 56/37-56

Clostridium difficile 20 57 57/21-40

Clostridium difficile 20 58 58/7-26

Clostridium difficile 20 59 59/25-44

Clostridium perfringens 20 60 60/15-34

Clostridium perfringens 20 61 61/47-66

Clostridium perfringens 20 62 62/161-180

Clostridium perfringens 20 63 63/14-33 Clostridium perfringens 20 64 64/64-83

Enterobacter aerogenes 20 65 65/24-43

Enterobacter aerogenes 20 66 66/21-40

Enterobacter aerogenes 20 67 67/40-59

Enterobacter aerogenes 20 68 68/23-42

Enterobacter aerogenes 20 69 69/21-40

Enterococcus faecalis 20 70 70/9-28

Enterococcus faecalis 20 71 71/9-28

Enterococcus faecalis 20 72 72/17-36

Enterococcus faecalis 20 73 73/19-38

Enterococcus faecalis 20 74 74/26-45

Enterococcus faecium 20 75 75/7-26

Enterococcus faecium 20 76 76/30-49

Enterococcus faecium 20 77 77/10-29

Enterococcus faecium 20 78 78/5-24

Enterococcus faecium 20 79 79/17-36

Francisella tularensis 20 80 80/17-36

Francisella tularensis 20 81 81/9-28

Francisella tularensis 20 82 82/7-26

Francisella tularensis 20 83 83/14-33

Francisella tularensis 20 84 84/15-34

Haemophilus influenzae 20 85 85/22-41

Helicobacter pylori 20 86 86/1-20

Klebsiella oxytoca 20 88 88/13-32

Klebsiella oxytoca 20 89 89/11-30

Klebsiella oxytoca 20 90 90/22-41

Klebsiella oxytoca 20 91 91/21-40

Klebsiella oxytoca 20 92 92/13-32

Legionella pneumophila 20 93 93/6-25

Legionella pneumophila 20 94 94/12-31

Legionella pneumophila 20 95 95/29-48

Legionella pneumophila 20 96 96/8-27

Legionella pneumophila 20 97 97/10-29

Listeria monocytogenes 20 98 98/8-27

Listeria monocytogenes 20 99 99/7-26

Listeria monocytogenes 20 100 100/20-39

Listeria monocytogenes 20 101 101/8-27

Listeria monocytogenes 20 102 102/20-39

Moraxella catarrhalis 20 103 103/30-49

Moraxella catarrhalis 20 104 104/35-54

Moraxella catarrhalis 20 105 105/93-112

Moraxella catarrhalis 20 106 106/59-78

Moraxella catarrhalis 20 107 107/14-33

Mycobacterium avium 20 108 108/17-36

Mycobacterium avium 20 109 109/14-33

Mycobacterium avium 20 110 110/7-26 Mycobacterium avium 20 111 111/27-46

Mycobacterium avium 20 112 112/18-37

Mycobacterium bovis 20 113 113/21-40

Mycobacterium bovis 20 114 114/21-40

Mycobacterium bovis 20 115 115/21-40

Mycobacterium bovis 20 116 116/20-39

Mycoplasma genitalium 20 117 117/6-25

Mycoplasma genitalium 20 118 118/15-34

Mycoplasma genitalium 20 119 119/109-128

Mycoplasma genitalium 20 120 120/28-47

Mycoplasma genitalium 20 121 121/128-147

Mycoplasma pneumoniae 20 122 122/23-42

Mycoplasma pneumoniae 20 123 123/36-55

Mycoplasma pneumoniae 20 124 124/54-73

Mycoplasma pneumoniae 20 125 125/69-88

Mycoplasma pneumoniae 20 126 126/15-34

Neisseria gonorrhoeae 20 127 127/8-27

Neisseria gonorrhoeae 20 128 128/18-37

Neisseria gonorrhoeae 20 129 129/16-35

Neisseria gonorrhoeae 20 130 130/14-33

Neisseria gonorrhoeae 20 131 131/13-32

Neisseria meningitidis 20 132 132/18-37

Neisseria meningitidis 20 133 133/13-32

Neisseria meningitidis 20 134 134/32-51

Neisseria meningitidis 20 135 135/19-38

Neisseria meningitidis 20 136 136/16-35

Porphyromonas gingivalis 20 137 137/71-90

Porphyromonas gingivalis 20 138 138/17-36

Porphyromonas gingivalis 20 139 139/7-26

Porphyromonas gingivalis 20 140 140/96-115

Porphyromonas gingivalis 20 141 141/56-75

Proteus mirabilis 20 142 142/15-34

Proteus mirabilis 20 143 143/51-70

Proteus mirabilis 20 144 144/55-74

Proteus mirabilis 20 145 145/24-43

Proteus mirabilis 20 146 146/24-43

Pseudomonas aeruginosa 20 147 147/7-26

Pseudomonas aeruginosa 20 148 148/10-29

Pseudomonas aeruginosa 20 149 149/16-35

Pseudomonas aeruginosa 20 150 150/12-31

Pseudomonas aeruginosa 20 151 151/23-42

Salmonella enterica 20 152 152/11-30

Salmonella enterica 20 153 153/1-20

Salmonella enterica 20 154 154/2-21

Salmonella enterica 20 155 155/6-25

Salmonella enterica 20 156 156/6-25 Serratia marcescens 20 157 157/15-34

Serratia marcescens 20 158 158/15-34

Serratia marcescens 20 159 159/21-40

Serratia marcescens 20 160 160/11-30

Serratia marcescens 20 161 161/21-40

Staphylococcus aureus 20 162 162/8-27

Staphylococcus aureus 20 163 163/6-25

Staphylococcus aureus 20 164 164/4-23

Staphylococcus aureus 20 165 165/6-25

Staphylococcus aureus 20 166 166/8-27

Staphylococcus epidermidis 20 167 167/11-30

Staphylococcus epidermidis 20 168 168/20-39

Staphylococcus epidermidis 20 169 169/26-45

Staphylococcus epidermidis 20 170 170/8-27

Staphylococcus epidermidis 20 171 171/21-40

Staphylococcus haemolyticus 20 172 172/16-35

Staphylococcus haemolyticus 20 173 173/12-31

Staphylococcus haemolyticus 20 174 174/18-37

Staphylococcus haemolyticus 20 175 175/20-39

Staphylococcus haemolyticus 20 176 176/6-25

Stenotrophomonas maltophilia 20 177 177/21-40

Streptococcus mutans 20 178 178/36-55

Streptococcus mutans 20 179 179/5-24

Streptococcus mutans 20 180 180/8-27

Streptococcus mutans 20 181 181/11-30

Streptococcus mutans 20 182 182/14-33

Streptococcus pyogenes 20 183 183/17-36

Streptococcus pyogenes 20 184 184/15-34

Streptococcus pyogenes 20 185 185/7-26

Streptococcus pyogenes 20 186 186/52-71

Streptococcus pyogenes 20 187 187/10-29

Streptococcus salivarius 20 188 188/9-28

Streptococcus salivarius 20 189 189/17-36

Streptococcus salivarius 20 190 190/16-35

Streptococcus salivarius 20 191 191/11-30

Streptococcus salivarius 20 192 192/6-25

Streptococcus sanguinis 20 193 193/7-26

Streptococcus sanguinis 20 194 194/7-26

Streptococcus sanguinis 20 195 195/38-57

Streptococcus sanguinis 20 196 196/15-34

Streptococcus sanguinis 20 197 197/16-35

Treponema pallidum 20 198 198/38-57

Treponema pallidum 20 199 199/29-48

Treponema pallidum 20 200 200/30-49

Treponema pallidum 20 201 201/26-45

Treponema pallidum 20 202 202/20-39 Vibrio cholerae 20 203 203/6-25

Vibrio parahaemolyticus 20 204 204/20-39

Vibrio parahaemolyticus 20 205 205/47-66

Vibrio parahaemolyticus 20 206 206/14-33

Vibrio parahaemolyticus 20 207 207/13-32

Vibrio parahaemolyticus 20 208 208/16-35

Yersinia enter ocolitica 20 209 209/12-31

Yersinia enterocolitica 20 210 210/23-42

Yersinia enterocolitica 20 211 211/12-31

Yersinia enterocolitica 20 212 212/16-35

Yersinia enterocolitica 20 213 213/22-41

Aggregatibacter actinomycetemcomitans 25 1 1/12-36

Aggregatibacter actinomycetemcomitans 25 2 2/5-29

Aggregatibacter actinomycetemcomitans 25 3 3/25-49

Aggregatibacter actinomycetemcomitans 25 4 4/8-32

Aggregatibacter actinomycetemcomitans 25 5 5/9-33

Bacillus anthracis 25 6 6/17-41

Bacillus anthracis 25 7 7/17-41

Bacillus anthracis 25 8 8/18-42

Bacillus anthracis 25 9 9/18-42

Bacillus anthracis 25 10 10/8-32

Bacillus licheniformis 25 11 11/15-39

Bacillus licheniformis 25 12 12/7-31

Bacillus licheniformis 25 13 13/11-35

Bacillus licheniformis 25 14 14/16-40

Bacillus licheniformis 25 15 15/34-58

Bacteroides fragilis 25 16 16/27-51

Bacteroides fragilis 25 17 17/25-49

Bacteroides fragilis 25 18 18/9-33

Bacteroides fragilis 25 19 19/24-48

Bacteroides fragilis 25 20 20/21-45

Bartonella henselae 25 21 21/128-152

Bartonella henselae 25 22 22/18-42

Bartonella henselae 25 23 23/12-36

Bartonella henselae 25 24 24/99-123

Bartonella henselae 25 25 25/4-28

Bordetella pertussis 25 26 26/18-42

Bordetella pertussis 25 27 27/18-42

Bordetella pertussis 25 28 28/15-39

Bordetella pertussis 25 29 29/18-42

Bordetella pertussis 25 30 30/18-42

Borrelia burgdorferi 25 31 31/11-35

Borrelia burgdorferi 25 32 32/5-29

Borrelia burgdorferi 25 33 33/23-47

Borrelia burgdorferi 25 34 34/19-43

Borrelia burgdorferi 25 35 35/32-56 Brucella abortus 25 36 36/15-39

Brucella abortus 25 37 37/18-42

Brucella abortus 25 38 38/17-41

Brucella abortus 25 39 39/18-42

Brucella abortus 25 40 40/18-42

Campylobacter jejuni 25 42 42/6-30

Campylobacter jejuni 25 44 44/14-38

Chlamydia trachomatis 25 45 45/11-35

Chlamydia trachomatis 25 46 46/5-29

Chlamydia trachomatis 25 47 47/9-33

Chlamydia trachomatis 25 48 48/70-94

Chlamydia trachomatis 25 49 49/37-61

Chlamydophila pneumoniae 25 50 50/28-52

Chlamydophila pneumoniae 25 51 51/28-52

Chlamydophila pneumoniae 25 52 52/116-140

Chlamydophila pneumoniae 25 53 53/14-38

Chlamydophila pneumoniae 25 54 54/85-109

Clostridium difficile 25 55 55/8-32

Clostridium difficile 25 56 56/33-57

Clostridium difficile 25 57 57/18-42

Clostridium difficile 25 58 58/5-29

Clostridium difficile 25 59 59/22-46

Clostridium perfringens 25 60 60/13-37

Clostridium perfringens 25 61 61/42-66

Clostridium perfringens 25 62 62/158-182

Clostridium perfringens 25 63 63/11-35

Clostridium perfringens 25 64 64/60-84

Enterobacter aerogenes 25 65 65/25-49

Enterobacter aerogenes 25 66 66/18-42

Enterobacter aerogenes 25 67 67/34-58

Enterobacter aerogenes 25 68 68/21-45

Enterobacter aerogenes 25 69 69/18-42

Enterococcus faecalis 25 70 70/6-30

Enterococcus faecalis 25 71 71/6-30

Enterococcus faecalis 25 72 72/15-39

Enterococcus faecalis 25 73 73/17-41

Enterococcus faecalis 25 74 74/25-49

Enterococcus faecium 25 75 75/5-29

Enterococcus faecium 25 76 76/28-52

Enterococcus faecium 25 77 77/7-31

Enterococcus faecium 25 78 78/3-27

Enterococcus faecium 25 79 79/14-38

Francisella tularensis 25 80 80/15-39

Francisella tularensis 25 81 81/6-30

Francisella tularensis 25 82 82/5-29

Francisella tularensis 25 83 83/11-35 Francisella tularensis 25 84 84/13-37

Haemophilus influenzae 25 85 85/17-41

Klebsiella oxytoca 25 88 88/11-35

Klebsiella oxytoca 25 89 89/8-32

Klebsiella oxytoca 25 90 90/19-43

Klebsiella oxytoca 25 91 91/19-43

Klebsiella oxytoca 25 92 92/10-34

Legionella pneumophila 25 93 93/4-28

Legionella pneumophila 25 94 94/9-33

Legionella pneumophila 25 95 95/27-51

Legionella pneumophila 25 96 96/6-30

Legionella pneumophila 25 97 97/7-31

Listeria monocytogenes 25 98 98/6-30

Listeria monocytogenes 25 99 99/5-29

Listeria monocytogenes 25 100 100/18-42

Listeria monocytogenes 25 101 101/6-30

Listeria monocytogenes 25 102 102/18-42

Moraxella catarrhalis 25 103 103/29-53

Moraxella catarrhalis 25 104 104/32-56

Moraxella catarrhalis 25 105 105/89-113

Moraxella catarrhalis 25 106 106/56-80

Moraxella catarrhalis 25 107 107/12-36

Mycobacterium avium 25 108 108/14-38

Mycobacterium avium 25 109 109/11-35

Mycobacterium avium 25 110 110/4-28

Mycobacterium avium 25 111 111/25-49

Mycobacterium avium 25 112 112/16-40

Mycobacterium bovis 25 113 113/19-43

Mycobacterium bovis 25 114 114/19-43

Mycobacterium bovis 25 115 115/18-42

Mycobacterium bovis 25 116 116/18-42

Mycoplasma genitalium 25 117 117/4-28

Mycoplasma genitalium 25 118 118/12-36

Mycoplasma genitalium 25 119 119/105-129

Mycoplasma genitalium 25 120 120/23-47

Mycoplasma genitalium 25 121 121/121-145

Mycoplasma pneumoniae 25 122 122/21-45

Mycoplasma pneumoniae 25 123 123/31-55

Mycoplasma pneumoniae 25 124 124/50-74

Mycoplasma pneumoniae 25 125 125/70-94

Mycoplasma pneumoniae 25 126 126/13-37

Neisseria gonorrhoeae 25 127 127/5-29

Neisseria gonorrhoeae 25 128 128/16-40

Neisseria gonorrhoeae 25 129 129/14-38

Neisseria gonorrhoeae 25 130 130/12-36

Neisseria gonorrhoeae 25 131 131/10-34 Neisseria meningitidis 25 132 132/15-39

Neisseria meningitidis 25 133 133/11-35

Neisseria meningitidis 25 134 134/29-53

Neisseria meningitidis 25 135 135/17-41

Neisseria meningitidis 25 136 136/13-37

Porphyromonas gingivalis 25 137 137/70-94

Porphyromonas gingivalis 25 138 138/14-38

Porphyromonas gingivalis 25 139 139/4-28

Porphyromonas gingivalis 25 140 140/94-118

Porphyromonas gingivalis 25 141 141/58-82

Proteus mirabilis 25 142 142/12-36

Proteus mirabilis 25 143 143/45-69

Proteus mirabilis 25 144 144/53-77

Proteus mirabilis 25 145 145/22-46

Proteus mirabilis 25 146 146/21-45

Pseudomonas aeruginosa 25 147 147/5-29

Pseudomonas aeruginosa 25 148 148/6-30

Pseudomonas aeruginosa 25 149 149/14-38

Pseudomonas aeruginosa 25 150 150/9-33

Pseudomonas aeruginosa 25 151 151/21-45

Salmonella enterica 25 152 152/6-30

Salmonella enterica 25 154 154/1-25

Salmonella enterica 25 155 155/1-25

Salmonella enterica 25 156 156/1-25

Serratia marcescens 25 157 157/12-36

Serratia marcescens 25 158 158/11-35

Serratia marcescens 25 159 159/18-42

Serratia marcescens 25 160 160/9-33

Serratia marcescens 25 161 161/19-43

Staphylococcus aureus 25 162 162/5-29

Staphylococcus aureus 25 163 163/3-27

Staphylococcus aureus 25 164 164/2-26

Staphylococcus aureus 25 165 165/4-28

Staphylococcus aureus 25 166 166/6-30

Staphylococcus epidermidis 25 167 167/8-32

Staphylococcus epidermidis 25 168 168/17-41

Staphylococcus epidermidis 25 169 169/23-47

Staphylococcus epidermidis 25 170 170/6-30

Staphylococcus epidermidis 25 171 171/18-42

Staphylococcus haemolyticus 25 172 172/14-38

Staphylococcus haemolyticus 25 173 173/9-33

Staphylococcus haemolyticus 25 174 174/15-39

Staphylococcus haemolyticus 25 175 175/23-47

Staphylococcus haemolyticus 25 176 176/5-29

Stenotrophomonas maltophilia 25 177 177/18-42

Streptococcus mutans 25 178 178/32-56 Streptococcus mutans 25 179 179/3-27

Streptococcus mutans 25 180 180/6-30

Streptococcus mutans 25 181 181/8-32

Streptococcus mutans 25 182 182/11-35

Streptococcus pyogenes 25 183 183/14-38

Streptococcus pyogenes 25 184 184/12-36

Streptococcus pyogenes 25 185 185/5-29

Streptococcus pyogenes 25 186 186/50-74

Streptococcus pyogenes 25 187 187/8-32

Streptococcus salivarius 25 188 188/5-29

Streptococcus salivarius 25 189 189/15-39

Streptococcus salivarius 25 190 190/13-37

Streptococcus salivarius 25 191 191/8-32

Streptococcus salivarius 25 192 192/5-29

Streptococcus sanguinis 25 193 193/5-29

Streptococcus sanguinis 25 194 194/4-28

Streptococcus sanguinis 25 195 195/33-57

Streptococcus sanguinis 25 196 196/12-36

Streptococcus sanguinis 25 197 197/14-38

Treponema pallidum 25 198 198/32-56

Treponema pallidum 25 199 199/27-51

Treponema pallidum 25 200 200/30-54

Treponema pallidum 25 201 201/24-48

Treponema pallidum 25 202 202/18-42

Vibrio cholerae 25 203 203/2-26

Vibrio parahaemolyticus 25 204 204/18-42

Vibrio parahaemolyticus 25 205 205/42-66

Vibrio parahaemolyticus 25 206 206/11-35

Vibrio parahaemolyticus 25 207 207/10-34

Vibrio parahaemolyticus 25 208 208/14-38

Yersinia enterocolitica 25 209 209/9-33

Yersinia enterocolitica 25 210 210/20-44

Yersinia enterocolitica 25 211 211/12-36

Yersinia enterocolitica 25 212 212/13-37

Yersinia enterocolitica 25 213 213/20-44

Aggregatibacter actinomycetemcomitans 30 1 1/10-39

Aggregatibacter actinomycetemcomitans 30 2 2/2-31

Aggregatibacter actinomycetemcomitans 30 3 3/23-52

Aggregatibacter actinomycetemcomitans 30 4 4/6-35

Aggregatibacter actinomycetemcomitans 30 5 5/6-35

Bacillus anthracis 30 6 6/15-44

Bacillus anthracis 30 7 7/15-44

Bacillus anthracis 30 8 8/15-44

Bacillus anthracis 30 9 9/16-45

Bacillus anthracis 30 10 10/5-34

Bacillus licheniformis 30 11 11/12-41 Bacillus licheniformis 30 12 12/5-34

Bacillus licheniformis 30 13 13/8-37

Bacillus licheniformis 30 14 14/13-42

Bacillus licheniformis 30 15 15/28-57

Bacteroides fragilis 30 16 16/25-54

Bacteroides fragilis 30 17 17/23-52

Bacteroides fragilis 30 18 18/6-35

Bacteroides fragilis 30 19 19/22-51

Bacteroides fragilis 30 20 20/19-48

Bartonella henselae 30 21 21/125-154

Bartonella henselae 30 22 22/15-44

Bartonella henselae 30 23 23/10-39

Bartonella henselae 30 24 24/98-127

Bartonella henselae 30 25 25/2-31

Bordetella pertussis 30 26 26/16-45

Bordetella pertussis 30 27 27/15-44

Bordetella pertussis 30 28 28/12-41

Bordetella pertussis 30 29 29/15-44

Bordetella pertussis 30 30 30/15-44

Borrelia burgdorferi 30 31 31/9-38

Borrelia burgdorferi 30 32 32/3-32

Borrelia burgdorferi 30 33 33/20-49

Borrelia burgdorferi 30 34 34/16-45

Borrelia burgdorferi 30 35 35/30-59

Brucella abortus 30 36 36/12-41

Brucella abortus 30 37 37/16-45

Brucella abortus 30 38 38/15-44

Brucella abortus 30 39 39/15-44

Brucella abortus 30 40 40/15-44

Campylobacter jejuni 30 42 42/3-32

Campylobacter jejuni 30 44 44/11-40

Chlamydia trachomatis 30 45 45/8-37

Chlamydia trachomatis 30 46 46/3-32

Chlamydia trachomatis 30 47 47/6-35

Chlamydia trachomatis 30 48 48/68-97

Chlamydia trachomatis 30 49 49/33-62

Chlamydophila pneumoniae 30 50 50/26-55

Chlamydophila pneumoniae 30 51 51/28-57

Chlamydophila pneumoniae 30 52 52/111-140

Chlamydophila pneumoniae 30 53 53/12-41

Chlamydophila pneumoniae 30 54 54/80-109

Clostridium difficile 30 55 55/5-34

Clostridium difficile 30 56 56/30-59

Clostridium difficile 30 57 57/15-44

Clostridium difficile 30 58 58/3-32

Clostridium difficile 30 59 59/19-48 Clostridium perfringens 30 60 60/10-39

Clostridium perfringens 30 61 61/35-64

Clostridium perfringens 30 62 62/153-182

Clostridium perfringens 30 63 63/9-38

Clostridium perfringens 30 64 64/55-84

Enterobacter aerogenes 30 65 65/26-55

Enterobacter aerogenes 30 66 66/16-45

Enterobacter aerogenes 30 67 67/27-56

Enterobacter aerogenes 30 68 68/18-47

Enterobacter aerogenes 30 69 69/16-45

Enterococcus faecalis 30 70 70/4-33

Enterococcus faecalis 30 71 71/4-33

Enterococcus faecalis 30 72 72/12-41

Enterococcus faecalis 30 73 73/14-43

Enterococcus faecalis 30 74 74/23-52

Enterococcus faecium 30 75 75/3-32

Enterococcus faecium 30 76 76/25-54

Enterococcus faecium 30 77 77/4-33

Enterococcus faecium 30 78 78/1-30

Enterococcus faecium 30 79 79/11-40

Francisella tularensis 30 80 80/12-41

Francisella tularensis 30 81 81/3-32

Francisella tularensis 30 82 82/2-31

Francisella tularensis 30 83 83/9-38

Francisella tularensis 30 84 84/10-39

Haemophilus influenzae 30 85 85/15-44

Klebsiella oxytoca 30 88 88/8-37

Klebsiella oxytoca 30 89 89/6-35

Klebsiella oxytoca 30 90 90/17-46

Klebsiella oxytoca 30 91 91/16-45

Klebsiella oxytoca 30 92 92/8-37

Legionella pneumophila 30 93 93/2-31

Legionella pneumophila 30 94 94/6-35

Legionella pneumophila 30 95 95/27-56

Legionella pneumophila 30 96 96/3-32

Legionella pneumophila 30 97 97/5-34

Listeria monocytogenes 30 98 98/4-33

Listeria monocytogenes 30 99 99/3-32

Listeria monocytogenes 30 100 100/15-44

Listeria monocytogenes 30 101 101/4-33

Listeria monocytogenes 30 102 102/15-44

Moraxella catarrhalis 30 103 103/29-58

Moraxella catarrhalis 30 104 104/30-59

Moraxella catarrhalis 30 105 105/85-114

Moraxella catarrhalis 30 106 106/54-83

Moraxella catarrhalis 30 107 107/9-38 Mycobacterium avium 30 108 108/12-41

Mycobacterium avium 30 109 109/8-37

Mycobacterium avium 30 110 110/2-31

Mycobacterium avium 30 111 111/22-51

Mycobacterium avium 30 112 112/14-43

Mycobacterium bovis 30 113 113/16-45

Mycobacterium bovis 30 114 114/16-45

Mycobacterium bovis 30 115 115/15-44

Mycobacterium bovis 30 116 116/15-44

Mycoplasma genitalium 30 117 117/2-31

Mycoplasma genitalium 30 118 118/10-39

Mycoplasma genitalium 30 119 119/101-130

Mycoplasma genitalium 30 120 120/20-49

Mycoplasma genitalium 30 121 121/118-147

Mycoplasma pneumoniae 30 122 122/19-48

Mycoplasma pneumoniae 30 123 123/28-57

Mycoplasma pneumoniae 30 124 124/47-76

Mycoplasma pneumoniae 30 125 125/65-94

Mycoplasma pneumoniae 30 126 126/10-39

Neisseria gonorrhoeae 30 127 127/2-31

Neisseria gonorrhoeae 30 128 128/14-43

Neisseria gonorrhoeae 30 129 129/12-41

Neisseria gonorrhoeae 30 130 130/11-40

Neisseria gonorrhoeae 30 131 131/8-37

Neisseria meningitidis 30 132 132/12-41

Neisseria meningitidis 30 133 133/8-37

Neisseria meningitidis 30 134 134/26-55

Neisseria meningitidis 30 135 135/15-44

Neisseria meningitidis 30 136 136/11-40

Porphyromonas gingivalis 30 137 137/65-94

Porphyromonas gingivalis 30 138 138/12-41

Porphyromonas gingivalis 30 139 139/2-31

Porphyromonas gingivalis 30 140 140/92-121

Porphyromonas gingivalis 30 141 141/56-85

Proteus mirabilis 30 142 142/10-39

Proteus mirabilis 30 143 143/40-69

Proteus mirabilis 30 144 144/51-80

Proteus mirabilis 30 145 145/19-48

Proteus mirabilis 30 146 146/18-47

Pseudomonas aeruginosa 30 147 147/2-31

Pseudomonas aeruginosa 30 148 148/4-33

Pseudomonas aeruginosa 30 149 149/12-41

Pseudomonas aeruginosa 30 150 150/7-36

Pseudomonas aeruginosa 30 151 151/19-48

Salmonella enterica 30 152 152/2-31

Serratia marcescens 30 157 157/10-39 Serratia marcescens 30 158 158/8-37

Serratia marcescens 30 159 159/16-45

Serratia marcescens 30 160 160/6-35

Serratia marcescens 30 161 161/15-44

Staphylococcus aureus 30 162 162/3-32

Staphylococcus aureus 30 164 164/1-30

Staphylococcus aureus 30 165 165/2-31

Staphylococcus aureus 30 166 166/3-32

Staphylococcus epidermidis 30 167 167/6-35

Staphylococcus epidermidis 30 168 168/15-44

Staphylococcus epidermidis 30 169 169/20-49

Staphylococcus epidermidis 30 170 170/3-32

Staphylococcus epidermidis 30 171 171/16-45

Staphylococcus haemolyticus 30 172 172/11-40

Staphylococcus haemolyticus 30 173 173/7-36

Staphylococcus haemolyticus 30 174 174/13-42

Staphylococcus haemolyticus 30 175 175/21-50

Staphylococcus haemolyticus 30 176 176/2-31

Stenotrophomonas maltophilia 30 177 177/16-45

Streptococcus mutans 30 178 178/28-57

Streptococcus mutans 30 179 179/1-30

Streptococcus mutans 30 180 180/3-32

Streptococcus mutans 30 181 181/6-35

Streptococcus mutans 30 182 182/8-37

Streptococcus pyogenes 30 183 183/11-40

Streptococcus pyogenes 30 184 184/10-39

Streptococcus pyogenes 30 185 185/3-32

Streptococcus pyogenes 30 186 186/47-76

Streptococcus pyogenes 30 187 187/6-35

Streptococcus salivarius 30 188 188/2-31

Streptococcus salivarius 30 189 189/12-41

Streptococcus salivarius 30 190 190/10-39

Streptococcus salivarius 30 191 191/5-34

Streptococcus salivarius 30 192 192/3-32

Streptococcus sanguinis 30 193 193/2-31

Streptococcus sanguinis 30 194 194/2-31

Streptococcus sanguinis 30 195 195/28-57

Streptococcus sanguinis 30 196 196/10-39

Streptococcus sanguinis 30 197 197/11-40

Treponema pallidum 30 198 198/29-58

Treponema pallidum 30 199 199/26-55

Treponema pallidum 30 200 200/29-58

Treponema pallidum 30 201 201/22-51

Treponema pallidum 30 202 202/15-44

Vibrio parahaemolyticus 30 204 204/15-44

Vibrio parahaemolyticus 30 205 205/37-66 Vibrio parahaemolyticus 30 206 206/7-36

Vibrio parahaemolyticus 30 207 207/7-36

Vibrio parahaemolyticus 30 208 208/11-40

Yersinia enterocolitica 30 209 209/7-36

Yersinia enterocolitica 30 210 210/18-47

Yersinia enterocolitica 30 211 211/9-38

Yersinia enterocolitica 30 212 212/1 1-40

Yersinia enterocolitica 30 213 213/18-47

Claims

1. A method for identifying one or more nucleotide k-mers in the genome of an organism which are capable of distinguishing the organism from one or more different organisms, comprising:

(a) extracting all nucleotide k-mers from the genome of the organism or from one or more parts of said genome; and

2. A method according to any one of the preceding claims, wherein the nucleotide k- mers are extracted from the parts of the genome which encode the ribosomal protein subunits.

3. A method according to claim 1 or 2, wherein step (a) comprises defining a length for the one or more nucleotide k-mers and extracting all nucleotide k-mers having that length from the genome of the organism.

4. A method according to claim 3, wherein step (a) comprises defining a length of from about 10 to about 40 nucleotides for the one or more nucleotide k-mers and extracting all nucleotide k-mers having that length from the genome of the organism.

5. A method according to any one of the preceding claims, wherein the method further comprises (c) discounting any nucleotide k-mers identified in step (b) which have an undesirable property.

6. A method according to claim 5, wherein the undesirable property is (i) its presence in the human genome, (ii) a homopolymer or repetitive sequence, (iii) the ability to form secondary structure, or (iv) its presence in a contaminating organism.

7. A method according to any one of the preceding claims, wherein the method further comprises (d) analysing the nucleotide k-mers identified in step (b) or step (c) and reducing their number by removing at least some sequence redundancy.

8. A method according to claim 7, wherein step (d) comprises minimising the number of nucleotide k-mers by removing all sequence redundancy.

9. A method according to any one of the preceding claims, wherein the organism is a microorganism, a fungus or a virus and the one or more different organisms are one or more different microorganisms, fungi or viruses.

10. A method according to claim 9, wherein the organism is a bacterium and the one or more different organisms are one or more different bacteria.

11. A method according to claim 10, wherein the organism is a bacterium and the one or more different organisms are one or more bacteria from one or more different genera of bacteria.

12. A method according to claim 10 or 11, wherein the organism is a bacterium and the one or more different organisms are one or more different species of bacteria.

13. A method according to any one of the preceding claims, wherein step (a) comprises extracting all nucleotide k-mers from about two or more different versions of the genome of the organism.

14. A method according to any one of the preceding claims, wherein step (a) comprises extracting all nucleotide k-mers from (i) about two hundred or more or (ii) about two thousand or more different versions of the genome of the organism.

15. A method according to any one of the preceding claims, wherein step (a) comprises extracting all nucleotide k-mers from all known versions of the genome of the organism.

16. A method according to any one of claims 13 to 15, wherein the method is for identifying about 25 or fewer nucleotide k-mers which are present in all of the different versions of the genome of the organism and which are capable of distinguishing the organism from the one or more different organisms.

17. A method according to any one of claims 13 to 15, wherein the method is for identifying only one nucleotide k-mer which is present in all of the different versions of the genome of the organism and which is capable of distinguishing the organism from the one or more different organisms.

18. A method according to any one of the preceding claims, wherein step (b) comprises comparing the nucleotide k-mers extracted in step (a) with two or more versions of the genome of each the one or more different organisms and identifying those nucleotide k- mers which do not appear in the genomes of the one or more different organisms

19. A method according to any one of claims 1 to 17, wherein step (b) comprises comparing the nucleotide k-mers extracted in step (a) with two hundred or more, two thousand or more or all known versions of the genome of each the one or more different organisms and identifying those nucleotide k-mers which do not appear in the genomes of the one or more different organisms.

20. A method according to any one of the preceding claims, wherein the method is for identifying one or more nucleotide k-mers in the genome of an organism which can be associated with a phenotype of the organism, comprising:

(a) extracting all nucleotide k-mers from the genome of the organism; and

(b) comparing the nucleotide k-mers extracted in step (a) with the genome(s) of one or more different organisms which do not display the phenotype and identifying those nucleotide k-mers which do not appear in the genome(s) of the one or more different organisms which do not display the phenotype.

21. A method according to claim 20, wherein the phenotype is pathogenicity, resistance to antibiotics, host specificity, tissue specificity, transmissibility, virulence, antigenicity or biochemical properties.

22. A computer program configured to carry out a method according to any one of the preceding claims.

23. A computer program medium comprising a computer program according to claim 22.

24. A computer for carrying out a method according to any one of claims 1 to 21 or programmed with a computer program according to claim 22.

25. One or more nucleotide k-mers identified using a method according to any one of claims 1 to 21 or a computer program according to claim 22.

26. One more nucleotide k-mers according to claim 25, which are identified from the parts of the genome which encode the ribosomal protein subunits.

27. One or more nucleotide k-mers according to claim 25 or 26, wherein the one or more k-mers can be associated with a phenotype of the organism.

28. An oligonucleotide probe which comprises a sequence which is complementary to a nucleotide k-mer according to any one of claims 25 to 27.

29. A plurality of oligonucleotide probes each of which comprises a sequence which is complementary to one of the nucleotide k-mers in a population of two or more nucleotide k-mers according to any one of claims 25 to 27.

30. An oligonucleotide probe according to claim 28 or a plurality of oligonucleotide probes according to claim 29, wherein a said nucleotide k-mer to which a sequence in a probe is complementary is identified from the parts of the genome which encode the ribosomal protein subunits.

31. An oligonucleotide probe according to claim 28 or 30 or a plurality of

oligonucleotide probes according to claim 29 or 30, wherein the probe is or the probes are detectably labelled.

32. An oligonucleotide probe according to any one of claims 28, 30 or 31 or a plurality of oligonucleotide probes according to any one of claims 29 to 31, wherein the probe is a molecular beacon probe or the probes are molecular beacon probes.

33. A support having attached thereto an oligonucleotide probe according to any one of claims 28 or 30 to 32 or a plurality of oligonucleotide probes according to any one of claims 29 to 32.

34. A support according to claim 33, wherein the support comprises a chip, pin, array or dipstick.

35. A method for detecting the presence or absence of an organism in a sample, comprising detecting the presence or absence in the sample of one or more nucleotide k- mers according to claim 25, wherein the presence of the one or more nucleotide k-mers is indicative of the presence of the organism in the sample and the wherein the absence of the one or more nucleotide k-mers is indicative of the absence of the organism from the sample.

36. A method according to claim 35, wherein the nucleotide k-mers are extracted from the parts of the genome which encode the ribosomal protein subunits.

37. A method according to claim 35 or 36, wherein the method is for detecting the presence or absence of an organism with a phenotype in a sample and the method comprises detecting the presence or absence in the sample of one or more nucleotide k- mers according to claim 24.

38. A nucleotide k-mer capable of distinguishing an organism from one or more different organisms, wherein the k-mer comprises about 10 or more consecutive

nucleotides from any one of the sequences shown in SEQ ID NOs: 1 to 213.

39. A nucleotide k-mer according to claim 38, wherein the k-mer comprises (i) from about 10 to about 40 consecutive nucleotides from any of the sequences shown in SEQ ID NOs: 1 to 213 or (ii) any of the consecutive nucleotides listed in the last column in Table 3.

40. A nucleotide k-mer according to claim 38 or 39, wherein the k-mer can be associated with a phenotype of the organism.

41. A nucleotide k-mer according to any one of claims 38 to 40, wherein the k-mer is from one of SEQ ID NOs: 1 to 5 and the organism is Aggregatibacter actinomycetemcomitans;

the k-mer is from one of SEQ ID NOs: 6 to 10 and the organism is Bacillus anthracis;

the k-mer is from one of SEQ ID NOs: 11 to 15 and the organism is Bacillus licheniformis;

the k-mer is from one of SEQ ID NOs: 16 to 20 and the organism is Bacteroides fragilis;

the k-mer is from one of SEQ ID NOs: 21 to 25 to 5 and the organism is Bartonella henselae;

the k-mer is from one of SEQ ID NOs: 26 to 30 and the organism is Bordetella pertussis;

the k-mer is from one of SEQ ID NOs: 31 to 35 and the organism is Borrelia burgdorferi;

the k-mer is from one of SEQ ID NOs: 36 to 40 and the organism is Brucella abortus;

the k-mer is from one of SEQ ID NOs: 41 to 44 and the organism is Campylobacter jejuni;

the k-mer is from one of SEQ ID NOs: 45 to 49 and the organism is Chlamydia trachomatis;

the k-mer is from one of SEQ ID NOs: 50 to 54 and the organism is Chlamydophila pneumonia;

the k-mer is from one of SEQ ID NOs: 55 to 59 and the organism is Clostridium difficile;

the k-mer is from one of SEQ ID NOs: 60 to 64 and the organism is Clostridium perfringens;

the k-mer is from one of SEQ ID NOs: 65 to 69 and the organism is from

Enterobacter aerogenes;

the k-mer is from one of SEQ ID NOs: 70 to 74 and the organism is from

Enterococcus faecalis;

the k-mer is from one of SEQ ID NOs: 75 to 79 and the organism is from

Enterococcus faecium;

the k-mer is from one of SEQ ID NOs: 80 to 84 and the organism is Francisella tularensis; the k-mer is from SEQ ID NO: 85 and the organism is Haemophilus influenza; the k-mer is from one of SEQ ID NOs: 86 and 87 and the organism is Helicobacter pylori;

the k-mer is from one of SEQ ID NOs: 88 to 92 and the organism is Klebsiella oxytoca

the k-mer is from one of SEQ ID NOs: 93 to 97 and the organism is Legionella pneumophila;

the k-mer is from one of SEQ ID NOs: 98 to 102 and the organism is Listeria monocytogenes;

the k-mer is from one of SEQ ID NOs: 103 to 107 and the organism is Moraxella catarrhalis;

the k-mer is from one of SEQ ID NOs: 108 to 112 and the organism is

Mycobacterium avium;

the k-mer is from one of SEQ ID NOs: 113 to 116 and the organism is

Mycobacterium bovis;

the k-mer is from one of SEQ ID NOs: 117 to 121 and the organism is Mycoplasma genitalium;

the k-mer is from one of SEQ ID NOs: 122 to 126 and the organism is Mycoplasma pneumonia;

the k-mer is from one of SEQ ID NOs: 127 to 131 and the organism is Neisseria gonorrhoeae;

the k-mer is from one of SEQ ID NOs: 132 to 136 and the organism is Neisseria meningitides;

the k-mer is from one of SEQ ID NOs: 137 to 141 and the organism is

Porphyromonas gingivalis;

the k-mer is from one of SEQ ID NOs: 142 to 146 and the organism is Proteus mirabilis;

the k-mer is from one of SEQ ID NOs: 147 to 151 and the organism is

Pseudomonas aeruginosa;

the k-mer is from one of SEQ ID NOs: 152 to 156 and the organism is Salmonella enterica;

the k-mer is from one of SEQ ID NOs: 157 to 161 and the organism is Serratia marcescens; the k-mer is from one of SEQ ID NOs: 162 to 166 and the organism is

Staphylococcus aureus;

the k-mer is from one of SEQ ID NOs: 167 to 171 and the organism is

Staphylococcus epidermidis;

the k-mer is from one of SEQ ID NOs: 172 to 176 and the organism is

Staphylococcus haemolyticus;

the k-mer is from SEQ ID NO: 177 and the organism is Stenotrophomonas maltophilia;

the k-mer is from one of SEQ ID NOs: 178 to 182 and the organism is

Streptococcus mutans;

the k-mer is from one of SEQ ID NOs: 183 to 187 and the organism is

Streptococcus pyogenes;

the k-mer is from one of SEQ ID NOs: 188 to 192 and the organism is

Streptococcus salivarius;

the k-mer is from one of SEQ ID NOs: 193 to 197 and the organism is

Streptococcus sanguinis;

the k-mer is from one of SEQ ID NOs: 198 to 202 and the organism is Treponema pallidum;

the k-mer is from SEQ ID NO: 203 and the organism is Vibrio cholera;

the k-mer is from one of SEQ ID NOs: 204 to 208 and the organism is Vibrio parahaemolyticus; or

the k-mer is from one of SEQ ID NOs: 209 to 213 and the organism is Yersinia enter ocolitica;

42. An oligonucleotide probe which comprises a sequence which is complementary to a nucleotide k-mer according to any one of claims 38 to 41.

43. An oligonucleotide probe according to claim 42, wherein the probe is detectably labelled and/or is a molecular beacon probe.

44. A support having attached thereto a oligonucleotide probe according to claim 42 or 43.

45. A support according to claim 44, wherein the support comprises a chip, pin, array or dipstick.

46. A method for detecting the presence or absence of an organism in a sample, comprising detecting the presence or absence in the sample of a nucleotide k-mer according to any one of claims 38 to 41, wherein the presence of the nucleotide k-mer is indicative of the presence of the organism in the sample and the wherein the absence of the nucleotide k-mer is indicative of the absence of the organism from the sample.

47. A method according to claim 46, wherein the method is for detecting the presence or absence of the organism with a particular phenotype in a sample and the method comprises detecting the presence or absence in the sample of a nucleotide k-mer according to claim 40.