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Papers by William Kelly

Journal of Applied Microbiology, 2004

Aims: To characterize a group of closely related Lactococcus lactis subsp. lactis casein starter ... more Aims: To characterize a group of closely related Lactococcus lactis subsp. lactis casein starter strains used commercially, which differ in their sensitivity to bacteriophages isolated from the same industrial environment. Methods and Results: Nine strains of L. lactis, six of which had been used as starter cultures for lactic casein manufacture, were shown to be closely related by pulsed-field gel electrophoresis and total DNA profiles. Nineteen phages which propagated on one or more of these starter strains were isolated from industrial casein whey samples. The phages were all small isometric-headed and could be divided into five groups on the basis of host range on the nine strains. Most of the phages did not give a PCR product with primers designed to detect the two most common lactococcal small isometric phage species (936 and P335). The hosts could be divided into six groups depending on their phage sensitivity. Plasmids encoding genes for the cell envelope associated PI-type proteinase, lactose metabolism and specificity subunits of a type I restriction/modification system were identified. Conclusions: This work demonstrates how isolates of the same starter strain may come to be regarded as separate cultures because of their different origins, and how these closely related strains may differ in some of their industrially relevant characteristics. Significance and Impact of the Study: This situation may be very common among lactococci used as dairy starter cultures, and implies that the dairy industry worldwide depends on a small number of different strains.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2002

Two species of dairy streptococci, Streptococcus waius and Streptococcus macedonicus, were origin... more Two species of dairy streptococci, Streptococcus waius and Streptococcus macedonicus, were originally characterized by 16S-23S intergenic spacer sequence analysis, random amplified polymorphic DNA fingerprinting, PFGE analysis and DNA-DNA reassociation experiments. All genetic data suggested that S. waius strains belong to the previously described species S. macedonicus. Likewise, the phenotypic characterization showed that strains of S. macedonicus and S. waius were highly related and easily differentiated from the closest phylogenetic neighbour, Streptococcus bovis, principally by their failure to produce a blackening reaction in medium containing aesculin. The utilization of maltose and cellobiose by S. macedonicus/S. waius strains allowed their differentiation from the most studied dairy species, Streptococcus thermophilus. On the basis of genetic and phenotypic data S. macedonicus and S. waius species should be considered synonyms and S. macedonicus has the priority.

Applied Microbiology and Biotechnology, 1989

Cellulase genes of the ruminant micro-organism Ruminococcus flavefaciens strain 186 have been clo... more Cellulase genes of the ruminant micro-organism Ruminococcus flavefaciens strain 186 have been cloned and expressed in Escherichia coli using the bacteriophage vector ?NM1149. Twenty-six clones showed expression of endo-ß-1,4-d-glucanases and were divided into four groups according to their insert sizes of approximately 2, 3, 4 or 9 kilobases (kb). Two of the clones with 4 kb inserts also showed exo-ß-1,4-d

Applied and Environmental Microbiology, 2007

The diversity of a collection of 102 lactococcus isolates including 91 Lactococcus lactis isolate... more The diversity of a collection of 102 lactococcus isolates including 91 Lactococcus lactis isolates of dairy and nondairy origin was explored using partial small subunit rRNA gene sequence analysis and limited phenotypic analyses. A subset of 89 strains of L. lactis subsp. cremoris and L. lactis subsp. lactis isolates was further analyzed by (GTG) 5 -PCR fingerprinting and a novel multilocus sequence analysis (MLSA) scheme. Two major genomic lineages within L. lactis were found. The L. lactis subsp. cremoris type-strain-like genotype lineage included both L. lactis subsp. cremoris and L. lactis subsp. lactis isolates. The other major lineage, with a L. lactis subsp. lactis type-strain-like genotype, comprised L. lactis subsp. lactis isolates only. A novel third genomic lineage represented two L. lactis subsp. lactis isolates of nondairy origin. The genomic lineages deviate from the subspecific classification of L. lactis that is based on a few phenotypic traits only. MLSA of six partial genes (atpA, encoding ATP synthase alpha subunit; pheS, encoding phenylalanine tRNA synthetase; rpoA, encoding RNA polymerase alpha chain; bcaT, encoding branched chain amino acid aminotransferase; pepN, encoding aminopeptidase N; and pepX, encoding X-prolyl dipeptidyl peptidase) revealed 363 polymorphic sites (total length, 1,970 bases) among 89 L. lactis subsp. cremoris and L. lactis subsp. lactis isolates with unique sequence types for most isolates. This allowed high-resolution cluster analysis in which dairy isolates form subclusters of limited diversity within the genomic lineages. The pheS DNA sequence analysis yielded two genetic groups dissimilar to the other genotyping analysis-based lineages, indicating a disparate acquisition route for this gene.

Current Opinion in Biotechnology, 2006

Frontiers in Microbiology, 2013

Phages of the P335 species infect Lactococcus lactis and have been particularly studied because o... more Phages of the P335 species infect Lactococcus lactis and have been particularly studied because of their association with strains of L. lactis subsp. cremoris used as dairy starter cultures. Unlike other lactococcal phages, those of the P335 species may have a temperate or lytic lifestyle, and are believed to originate from the starter cultures themselves. We have sequenced the genome of L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it contains an integrated P335 species prophage. This 41 kb prophage (Φ KW2) has a mosaic structure with functional modules that are highly similar to several other phages of the P335 species associated with dairy starter cultures. Comparison of the genomes of 26 phages of the P335 species, with either a lytic or temperate lifestyle, shows that they can be divided into three groups and that the morphogenesis gene region is the most conserved. Analysis of these phage genomes in conjunction with the genomes of several L. lactis strains shows that prophage insertion is site specific and occurs at seven different chromosomal locations. Exactly how induced or lytic phages of the P335 species interact with carbohydrate cell surface receptors in the host cell envelope remains to be determined. Genes for the biosynthesis of a variable cell surface polysaccharide and for lipoteichoic acids (LTAs) are found in L. lactis and are the main candidates for phage receptors, as the genes for other cell surface carbohydrates have been lost from dairy starter strains. Overall, phages of the P335 species appear to have had only a minor role in the adaptation of L. lactis subsp. cremoris strains to the dairy environment, and instead they appear to be an integral part of the L. lactis chromosome. There remains a great deal to be discovered about their role, and their contribution to the evolution of the bacterial genome.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2008

Proteins-structure Function and Bioinformatics, 2010

PLoS ONE, 2010

Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 2... more Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 21 times that of carbon dioxide (CO 2 ). Methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are lacking. Ruminant methane is formed by the action of methanogenic archaea typified by Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify genes and proteins that can be targeted to reduce methane production, we have sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be completed.

Proteins-structure Function and Bioinformatics, 2010

Frontiers in Microbiology, 2013

Phages of the P335 species infect Lactococcus lactis and have been particularly studied because o... more Phages of the P335 species infect Lactococcus lactis and have been particularly studied because of their association with strains of L. lactis subsp. cremoris used as dairy starter cultures. Unlike other lactococcal phages, those of the P335 species may have a temperate or lytic lifestyle, and are believed to originate from the starter cultures themselves. We have sequenced the genome of L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it contains an integrated P335 species prophage. This 41 kb prophage (Φ KW2) has a mosaic structure with functional modules that are highly similar to several other phages of the P335 species associated with dairy starter cultures. Comparison of the genomes of 26 phages of the P335 species, with either a lytic or temperate lifestyle, shows that they can be divided into three groups and that the morphogenesis gene region is the most conserved. Analysis of these phage genomes in conjunction with the genomes of several L. lactis strains shows that prophage insertion is site specific and occurs at seven different chromosomal locations. Exactly how induced or lytic phages of the P335 species interact with carbohydrate cell surface receptors in the host cell envelope remains to be determined. Genes for the biosynthesis of a variable cell surface polysaccharide and for lipoteichoic acids (LTAs) are found in L. lactis and are the main candidates for phage receptors, as the genes for other cell surface carbohydrates have been lost from dairy starter strains. Overall, phages of the P335 species appear to have had only a minor role in the adaptation of L. lactis subsp. cremoris strains to the dairy environment, and instead they appear to be an integral part of the L. lactis chromosome. There remains a great deal to be discovered about their role, and their contribution to the evolution of the bacterial genome.

Standards in Genomic Sciences, 2014

Methanobacterium formicicum BRM9 was isolated from the rumen of a New Zealand Friesan cow grazing... more Methanobacterium formicicum BRM9 was isolated from the rumen of a New Zealand Friesan cow grazing a ryegrass/ clover pasture, and its genome has been sequenced to provide information on the phylogenetic diversity of rumen methanogens with a view to developing technologies for methane mitigation. The 2.45 Mb BRM9 chromosome has an average G + C content of 41%, and encodes 2,352 protein-coding genes. The genes involved in methanogenesis are comparable to those found in other members of the Methanobacteriaceae with the exception that there is no [Fe]-hydrogenase dehydrogenase (Hmd) which links the methenyl-H4MPT reduction directly with the oxidation of H 2 . Compared to the rumen Methanobrevibacter strains, BRM9 has a much larger complement of genes involved in determining oxidative stress response, signal transduction and nitrogen fixation. BRM9 also has genes for the biosynthesis of the compatible solute ectoine that has not been reported to be produced by methanogens. The BRM9 genome has a prophage and two CRISPR repeat regions. Comparison to the genomes of other Methanobacterium strains shows a core genome of~1,350 coding sequences and 190 strain-specific genes in BRM9, most of which are hypothetical proteins or prophage related.

PLoS ONE, 2010

Determining the role of rumen microbes and their enzymes in plant polysaccharide breakdown is fun... more Determining the role of rumen microbes and their enzymes in plant polysaccharide breakdown is fundamental to understanding digestion and maximising productivity in ruminant animals. Butyrivibrio proteoclasticus B316 T is a Grampositive, butyrate-forming rumen bacterium with a key role in plant polysaccharide degradation. The 4.4Mb genome consists of 4 replicons; a chromosome, a chromid and two megaplasmids. The chromid is the smallest reported for all bacteria, and the first identified from the phylum Firmicutes. B316 devotes a large proportion of its genome to the breakdown and reassembly of complex polysaccharides and has a highly developed glycobiome when compared to other sequenced bacteria. The secretion of a range of polysaccharide-degrading enzymes which initiate the breakdown of pectin, starch and xylan, a subtilisin family protease active against plant proteins, and diverse intracellular enzymes to break down oligosaccharides constitute the degradative capability of this organism. A prominent feature of the genome is the presence of multiple gene clusters predicted to be involved in polysaccharide biosynthesis. Metabolic reconstruction reveals the absence of an identifiable gene for enolase, a conserved enzyme of the glycolytic pathway. To our knowledge this is the first report of an organism lacking an enolase. Our analysis of the B316 genome shows how one organism can contribute to the multi-organism complex that rapidly breaks down plant material in the rumen. It can be concluded that B316, and similar organisms with broad polysaccharide-degrading capability, are well suited to being early colonizers and degraders of plant polysaccharides in the rumen environment. Citation: Kelly WJ, Leahy SC, Altermann E, Yeoman CJ, Dunne JC, et al. (2010) The Glycobiome of the Rumen Bacterium Butyrivibrio proteoclasticus B316 T Highlights Adaptation to a Polysaccharide-Rich Environment. PLoS ONE 5(8): e11942.

PLoS ONE, 2010

Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 2... more Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 21 times that of carbon dioxide (CO 2 ). Methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are lacking. Ruminant methane is formed by the action of methanogenic archaea typified by Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify genes and proteins that can be targeted to reduce methane production, we have sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be completed.

Journal of Biotechnology, 2008

The development of high-throughput DNA sequencing techniques has enabled the sequencing of severa... more The development of high-throughput DNA sequencing techniques has enabled the sequencing of several hundred bacterial genomes. However, the major step towards understanding the molecular basis of an organism will be the determination of all gene functions in its genome. Current gene assignments by sequence homology generate numerous hints to putative or unknown functions. Even hits with good homology are often not specific enough to describe the in vivo biochemical functions and the underlying biological roles. In this work we applied metabolic footprinting analysis to characterize Tn916-inserted mutants of a hemicellulose-degrading rumen bacterium grown on complex culture medium. Interestingly, the most distinctive phenotypic difference was observed in a mutant with a transposon insertion in a non-coding region of the genome, while disruption of a gene with high homology to a known alpha-glucosidase/xylosidase showed no distinctive phenotypic effect. Our results demonstrate that extracellular metabolomics data coupled to genome information is a powerful and low-cost approach to rapidly screen and characterize microbial mutants with single gene deletions. However, metabolomics as a stand-alone technique is unlikely to give a complete answer to define gene functions, and, therefore, is an approach to be used to generate hypotheses and direct new experiments to confirm gene function.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2008

International Journal of Food Microbiology, 1998

Lactic acid bacteria isolated from minimally processed fresh fruit and vegetable products were id... more Lactic acid bacteria isolated from minimally processed fresh fruit and vegetable products were identified as Lactococcus lactis subsp. lactis on the basis of phenotypic tests, presence of lactococcal IS elements, and partial sequence analysis of the 16S rRNA gene. Isolated bacteria were differentiated using pulsed-field gel electrophoresis of SmaI digests of genomic DNA. Sprouted seeds were the best source of strains, and lactococci appear to be the dominant microflora on these products during the period they are intended to be eaten. Although these plant strains showed many similarities to strains of L. lactis used as dairy starter cultures, their carbohydrate fermentation patterns were unusual and probably reflect their environmental origin. Most strains fermented sucrose and xylose, and some also fermented raffinose and melibiose. Most of the bacteriocin-producing strains produced nisin, and nisin genes could also be detected in strains that showed no bacteriocin activity, or that produced a different bacteriocin with a narrow spectrum of activity. One strain produced nisin but was unable to ferment sucrose, properties that have been generally regarded as linked. These strains may have uses as biopreservatives for minimally processed plant products.

FEMS Microbiology Letters, 1998

Lactococcus lactis subsp. lactis strains isolated from various sprouted seed products were able t... more Lactococcus lactis subsp. lactis strains isolated from various sprouted seed products were able to transfer the ability to ferment raffinose in conjugation experiments at frequencies between 10 3R and 10 3U per donor cell. There was no evidence of plasmid transfer, but pulsed-field gel electrophoresis analysis showed that all transconjugants had acquired large chromosomal insertions indicative of conjugative transposons. Raffinose transconjugants contained inserts of 45 or 60 kb at one of two chromosomal sites, and these inserts contained two copies of an element related to the lactococcal insertion sequence ISS1. z

FEMS Microbiology Letters, 2000

Lactococcus lactis strains isolated from vegetable products transferred the ability to ferment su... more Lactococcus lactis strains isolated from vegetable products transferred the ability to ferment sucrose in conjugation experiments with the recipient strain L. lactis MG1614. Nisin production and sucrose fermentation were transferred together from two strains, but transfer also occurred from several other strains which did not produce nisin. Pulsed-field gel electrophoresis analysis showed that all transconjugants had acquired large chromosomal insertions at two main sites. Nisin^sucrose transconjugants had gained inserts of 70 kb, while those that fermented sucrose without nisin production contained inserts of between 50 and 110 kb. Transconjugants from one donor had acquired a separate insertion of 55 kb which correlated with enhanced bacteriophage resistance, but contained neither nisin nor sucrose fermentation genes. ß

Journal of Applied Microbiology, 2004

Aims: To characterize a group of closely related Lactococcus lactis subsp. lactis casein starter ... more Aims: To characterize a group of closely related Lactococcus lactis subsp. lactis casein starter strains used commercially, which differ in their sensitivity to bacteriophages isolated from the same industrial environment. Methods and Results: Nine strains of L. lactis, six of which had been used as starter cultures for lactic casein manufacture, were shown to be closely related by pulsed-field gel electrophoresis and total DNA profiles. Nineteen phages which propagated on one or more of these starter strains were isolated from industrial casein whey samples. The phages were all small isometric-headed and could be divided into five groups on the basis of host range on the nine strains. Most of the phages did not give a PCR product with primers designed to detect the two most common lactococcal small isometric phage species (936 and P335). The hosts could be divided into six groups depending on their phage sensitivity. Plasmids encoding genes for the cell envelope associated PI-type proteinase, lactose metabolism and specificity subunits of a type I restriction/modification system were identified. Conclusions: This work demonstrates how isolates of the same starter strain may come to be regarded as separate cultures because of their different origins, and how these closely related strains may differ in some of their industrially relevant characteristics. Significance and Impact of the Study: This situation may be very common among lactococci used as dairy starter cultures, and implies that the dairy industry worldwide depends on a small number of different strains.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2002

Two species of dairy streptococci, Streptococcus waius and Streptococcus macedonicus, were origin... more Two species of dairy streptococci, Streptococcus waius and Streptococcus macedonicus, were originally characterized by 16S-23S intergenic spacer sequence analysis, random amplified polymorphic DNA fingerprinting, PFGE analysis and DNA-DNA reassociation experiments. All genetic data suggested that S. waius strains belong to the previously described species S. macedonicus. Likewise, the phenotypic characterization showed that strains of S. macedonicus and S. waius were highly related and easily differentiated from the closest phylogenetic neighbour, Streptococcus bovis, principally by their failure to produce a blackening reaction in medium containing aesculin. The utilization of maltose and cellobiose by S. macedonicus/S. waius strains allowed their differentiation from the most studied dairy species, Streptococcus thermophilus. On the basis of genetic and phenotypic data S. macedonicus and S. waius species should be considered synonyms and S. macedonicus has the priority.

Applied Microbiology and Biotechnology, 1989

Cellulase genes of the ruminant micro-organism Ruminococcus flavefaciens strain 186 have been clo... more Cellulase genes of the ruminant micro-organism Ruminococcus flavefaciens strain 186 have been cloned and expressed in Escherichia coli using the bacteriophage vector ?NM1149. Twenty-six clones showed expression of endo-ß-1,4-d-glucanases and were divided into four groups according to their insert sizes of approximately 2, 3, 4 or 9 kilobases (kb). Two of the clones with 4 kb inserts also showed exo-ß-1,4-d

Applied and Environmental Microbiology, 2007

The diversity of a collection of 102 lactococcus isolates including 91 Lactococcus lactis isolate... more The diversity of a collection of 102 lactococcus isolates including 91 Lactococcus lactis isolates of dairy and nondairy origin was explored using partial small subunit rRNA gene sequence analysis and limited phenotypic analyses. A subset of 89 strains of L. lactis subsp. cremoris and L. lactis subsp. lactis isolates was further analyzed by (GTG) 5 -PCR fingerprinting and a novel multilocus sequence analysis (MLSA) scheme. Two major genomic lineages within L. lactis were found. The L. lactis subsp. cremoris type-strain-like genotype lineage included both L. lactis subsp. cremoris and L. lactis subsp. lactis isolates. The other major lineage, with a L. lactis subsp. lactis type-strain-like genotype, comprised L. lactis subsp. lactis isolates only. A novel third genomic lineage represented two L. lactis subsp. lactis isolates of nondairy origin. The genomic lineages deviate from the subspecific classification of L. lactis that is based on a few phenotypic traits only. MLSA of six partial genes (atpA, encoding ATP synthase alpha subunit; pheS, encoding phenylalanine tRNA synthetase; rpoA, encoding RNA polymerase alpha chain; bcaT, encoding branched chain amino acid aminotransferase; pepN, encoding aminopeptidase N; and pepX, encoding X-prolyl dipeptidyl peptidase) revealed 363 polymorphic sites (total length, 1,970 bases) among 89 L. lactis subsp. cremoris and L. lactis subsp. lactis isolates with unique sequence types for most isolates. This allowed high-resolution cluster analysis in which dairy isolates form subclusters of limited diversity within the genomic lineages. The pheS DNA sequence analysis yielded two genetic groups dissimilar to the other genotyping analysis-based lineages, indicating a disparate acquisition route for this gene.

Current Opinion in Biotechnology, 2006

Frontiers in Microbiology, 2013

Phages of the P335 species infect Lactococcus lactis and have been particularly studied because o... more Phages of the P335 species infect Lactococcus lactis and have been particularly studied because of their association with strains of L. lactis subsp. cremoris used as dairy starter cultures. Unlike other lactococcal phages, those of the P335 species may have a temperate or lytic lifestyle, and are believed to originate from the starter cultures themselves. We have sequenced the genome of L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it contains an integrated P335 species prophage. This 41 kb prophage (Φ KW2) has a mosaic structure with functional modules that are highly similar to several other phages of the P335 species associated with dairy starter cultures. Comparison of the genomes of 26 phages of the P335 species, with either a lytic or temperate lifestyle, shows that they can be divided into three groups and that the morphogenesis gene region is the most conserved. Analysis of these phage genomes in conjunction with the genomes of several L. lactis strains shows that prophage insertion is site specific and occurs at seven different chromosomal locations. Exactly how induced or lytic phages of the P335 species interact with carbohydrate cell surface receptors in the host cell envelope remains to be determined. Genes for the biosynthesis of a variable cell surface polysaccharide and for lipoteichoic acids (LTAs) are found in L. lactis and are the main candidates for phage receptors, as the genes for other cell surface carbohydrates have been lost from dairy starter strains. Overall, phages of the P335 species appear to have had only a minor role in the adaptation of L. lactis subsp. cremoris strains to the dairy environment, and instead they appear to be an integral part of the L. lactis chromosome. There remains a great deal to be discovered about their role, and their contribution to the evolution of the bacterial genome.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2008

Proteins-structure Function and Bioinformatics, 2010

PLoS ONE, 2010

Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 2... more Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 21 times that of carbon dioxide (CO 2 ). Methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are lacking. Ruminant methane is formed by the action of methanogenic archaea typified by Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify genes and proteins that can be targeted to reduce methane production, we have sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be completed.

Proteins-structure Function and Bioinformatics, 2010

Frontiers in Microbiology, 2013

Phages of the P335 species infect Lactococcus lactis and have been particularly studied because o... more Phages of the P335 species infect Lactococcus lactis and have been particularly studied because of their association with strains of L. lactis subsp. cremoris used as dairy starter cultures. Unlike other lactococcal phages, those of the P335 species may have a temperate or lytic lifestyle, and are believed to originate from the starter cultures themselves. We have sequenced the genome of L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it contains an integrated P335 species prophage. This 41 kb prophage (Φ KW2) has a mosaic structure with functional modules that are highly similar to several other phages of the P335 species associated with dairy starter cultures. Comparison of the genomes of 26 phages of the P335 species, with either a lytic or temperate lifestyle, shows that they can be divided into three groups and that the morphogenesis gene region is the most conserved. Analysis of these phage genomes in conjunction with the genomes of several L. lactis strains shows that prophage insertion is site specific and occurs at seven different chromosomal locations. Exactly how induced or lytic phages of the P335 species interact with carbohydrate cell surface receptors in the host cell envelope remains to be determined. Genes for the biosynthesis of a variable cell surface polysaccharide and for lipoteichoic acids (LTAs) are found in L. lactis and are the main candidates for phage receptors, as the genes for other cell surface carbohydrates have been lost from dairy starter strains. Overall, phages of the P335 species appear to have had only a minor role in the adaptation of L. lactis subsp. cremoris strains to the dairy environment, and instead they appear to be an integral part of the L. lactis chromosome. There remains a great deal to be discovered about their role, and their contribution to the evolution of the bacterial genome.

Standards in Genomic Sciences, 2014

Methanobacterium formicicum BRM9 was isolated from the rumen of a New Zealand Friesan cow grazing... more Methanobacterium formicicum BRM9 was isolated from the rumen of a New Zealand Friesan cow grazing a ryegrass/ clover pasture, and its genome has been sequenced to provide information on the phylogenetic diversity of rumen methanogens with a view to developing technologies for methane mitigation. The 2.45 Mb BRM9 chromosome has an average G + C content of 41%, and encodes 2,352 protein-coding genes. The genes involved in methanogenesis are comparable to those found in other members of the Methanobacteriaceae with the exception that there is no [Fe]-hydrogenase dehydrogenase (Hmd) which links the methenyl-H4MPT reduction directly with the oxidation of H 2 . Compared to the rumen Methanobrevibacter strains, BRM9 has a much larger complement of genes involved in determining oxidative stress response, signal transduction and nitrogen fixation. BRM9 also has genes for the biosynthesis of the compatible solute ectoine that has not been reported to be produced by methanogens. The BRM9 genome has a prophage and two CRISPR repeat regions. Comparison to the genomes of other Methanobacterium strains shows a core genome of~1,350 coding sequences and 190 strain-specific genes in BRM9, most of which are hypothetical proteins or prophage related.

PLoS ONE, 2010

Determining the role of rumen microbes and their enzymes in plant polysaccharide breakdown is fun... more Determining the role of rumen microbes and their enzymes in plant polysaccharide breakdown is fundamental to understanding digestion and maximising productivity in ruminant animals. Butyrivibrio proteoclasticus B316 T is a Grampositive, butyrate-forming rumen bacterium with a key role in plant polysaccharide degradation. The 4.4Mb genome consists of 4 replicons; a chromosome, a chromid and two megaplasmids. The chromid is the smallest reported for all bacteria, and the first identified from the phylum Firmicutes. B316 devotes a large proportion of its genome to the breakdown and reassembly of complex polysaccharides and has a highly developed glycobiome when compared to other sequenced bacteria. The secretion of a range of polysaccharide-degrading enzymes which initiate the breakdown of pectin, starch and xylan, a subtilisin family protease active against plant proteins, and diverse intracellular enzymes to break down oligosaccharides constitute the degradative capability of this organism. A prominent feature of the genome is the presence of multiple gene clusters predicted to be involved in polysaccharide biosynthesis. Metabolic reconstruction reveals the absence of an identifiable gene for enolase, a conserved enzyme of the glycolytic pathway. To our knowledge this is the first report of an organism lacking an enolase. Our analysis of the B316 genome shows how one organism can contribute to the multi-organism complex that rapidly breaks down plant material in the rumen. It can be concluded that B316, and similar organisms with broad polysaccharide-degrading capability, are well suited to being early colonizers and degraders of plant polysaccharides in the rumen environment. Citation: Kelly WJ, Leahy SC, Altermann E, Yeoman CJ, Dunne JC, et al. (2010) The Glycobiome of the Rumen Bacterium Butyrivibrio proteoclasticus B316 T Highlights Adaptation to a Polysaccharide-Rich Environment. PLoS ONE 5(8): e11942.

PLoS ONE, 2010

Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 2... more Background: Methane (CH 4 ) is a potent greenhouse gas (GHG), having a global warming potential 21 times that of carbon dioxide (CO 2 ). Methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. Technologies to reduce these emissions are lacking. Ruminant methane is formed by the action of methanogenic archaea typified by Methanobrevibacter ruminantium, which is present in ruminants fed a wide variety of diets worldwide. To gain more insight into the lifestyle of a rumen methanogen, and to identify genes and proteins that can be targeted to reduce methane production, we have sequenced the 2.93 Mb genome of M. ruminantium M1, the first rumen methanogen genome to be completed.

Journal of Biotechnology, 2008

The development of high-throughput DNA sequencing techniques has enabled the sequencing of severa... more The development of high-throughput DNA sequencing techniques has enabled the sequencing of several hundred bacterial genomes. However, the major step towards understanding the molecular basis of an organism will be the determination of all gene functions in its genome. Current gene assignments by sequence homology generate numerous hints to putative or unknown functions. Even hits with good homology are often not specific enough to describe the in vivo biochemical functions and the underlying biological roles. In this work we applied metabolic footprinting analysis to characterize Tn916-inserted mutants of a hemicellulose-degrading rumen bacterium grown on complex culture medium. Interestingly, the most distinctive phenotypic difference was observed in a mutant with a transposon insertion in a non-coding region of the genome, while disruption of a gene with high homology to a known alpha-glucosidase/xylosidase showed no distinctive phenotypic effect. Our results demonstrate that extracellular metabolomics data coupled to genome information is a powerful and low-cost approach to rapidly screen and characterize microbial mutants with single gene deletions. However, metabolomics as a stand-alone technique is unlikely to give a complete answer to define gene functions, and, therefore, is an approach to be used to generate hypotheses and direct new experiments to confirm gene function.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, 2008

International Journal of Food Microbiology, 1998

Lactic acid bacteria isolated from minimally processed fresh fruit and vegetable products were id... more Lactic acid bacteria isolated from minimally processed fresh fruit and vegetable products were identified as Lactococcus lactis subsp. lactis on the basis of phenotypic tests, presence of lactococcal IS elements, and partial sequence analysis of the 16S rRNA gene. Isolated bacteria were differentiated using pulsed-field gel electrophoresis of SmaI digests of genomic DNA. Sprouted seeds were the best source of strains, and lactococci appear to be the dominant microflora on these products during the period they are intended to be eaten. Although these plant strains showed many similarities to strains of L. lactis used as dairy starter cultures, their carbohydrate fermentation patterns were unusual and probably reflect their environmental origin. Most strains fermented sucrose and xylose, and some also fermented raffinose and melibiose. Most of the bacteriocin-producing strains produced nisin, and nisin genes could also be detected in strains that showed no bacteriocin activity, or that produced a different bacteriocin with a narrow spectrum of activity. One strain produced nisin but was unable to ferment sucrose, properties that have been generally regarded as linked. These strains may have uses as biopreservatives for minimally processed plant products.

FEMS Microbiology Letters, 1998

Lactococcus lactis subsp. lactis strains isolated from various sprouted seed products were able t... more Lactococcus lactis subsp. lactis strains isolated from various sprouted seed products were able to transfer the ability to ferment raffinose in conjugation experiments at frequencies between 10 3R and 10 3U per donor cell. There was no evidence of plasmid transfer, but pulsed-field gel electrophoresis analysis showed that all transconjugants had acquired large chromosomal insertions indicative of conjugative transposons. Raffinose transconjugants contained inserts of 45 or 60 kb at one of two chromosomal sites, and these inserts contained two copies of an element related to the lactococcal insertion sequence ISS1. z

FEMS Microbiology Letters, 2000

Lactococcus lactis strains isolated from vegetable products transferred the ability to ferment su... more Lactococcus lactis strains isolated from vegetable products transferred the ability to ferment sucrose in conjugation experiments with the recipient strain L. lactis MG1614. Nisin production and sucrose fermentation were transferred together from two strains, but transfer also occurred from several other strains which did not produce nisin. Pulsed-field gel electrophoresis analysis showed that all transconjugants had acquired large chromosomal insertions at two main sites. Nisin^sucrose transconjugants had gained inserts of 70 kb, while those that fermented sucrose without nisin production contained inserts of between 50 and 110 kb. Transconjugants from one donor had acquired a separate insertion of 55 kb which correlated with enhanced bacteriophage resistance, but contained neither nisin nor sucrose fermentation genes. ß