Department of Microbiology (G08), University of Sydney, New South Wales 2006, Australia1
Author for correspondence: Peter R. Reeves. Tel: +61 2 9351 2536. Fax: +61 2 9351 4571. e-mail: reeves{at}angis.usyd.edu.au
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ABSTRACT |
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Keywords: concerted evolution, GDP-D-mannose pathway, gene conversion, lateral transfer, Salmonella enterica
Abbreviations: CA, colanic acid
The GenBank accession numbers for the sequences reported in this paper are AY012160AY012201.
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INTRODUCTION |
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Genes within a given bacterial species generally have the same GC content, which is thought to be due to directional mutation pressure (Sueoka, 1988 , 1992
). The sequencing of several S. enterica O antigen gene clusters has shown that the GC content of this region is variable and atypical in comparison to the mean GC content of S. enterica DNA. We have previously suggested that this indicates that many of the O antigen genes evolved in other species and were later captured by S. enterica via lateral gene transfer (Reeves, 1991
, 1993
).
S. enterica has seven well-defined subspecies, which were initially resolved by biotyping and confirmed by DNA hybridization (Le Minor et al., 1982 , 1986
). Multilocus enzyme electrophoresis and the sequencing of several housekeeping genes have also shown that strains of a given subspecies generally cluster together, indicating low levels of intersubspecies gene transfer (Boyd et al., 1994
; Nelson et al., 1991
; Reeves et al., 1989
). However, most of the 46 O antigens occur in two or more subspecies, indicating extensive intersubspecies transfer of O antigen genes (Reeves, 1997
). Cell surface interactions and niche adaptation are believed to provide the selection pressure for the distribution of O antigen genes amongst subspecies, and the generation of new O antigen gene clusters (Reeves, 1992
, 1997
).
The nucleotide sugar GDP-D-mannose is required for the mannosylation of many bacterial cell surface repeat-unit polysaccharides and acts as the precursor for other nucleotide sugars (GDP-L-fucose, GDP-colitose, GDP-perosamine and GDP-D-rhamnose) involved in polysaccharide biosynthesis. GDP-D-mannose is synthesized from fructose 6-phosphate by products of the manA, manB and manC genes as follows: manA, fructose 6-phosphate to mannose 6-phosphate; manB, mannose 6-phosphate to mannose 1-phosphate; manC, mannose 1-phosphate to GDP-D-mannose. manA encodes type I phosphomannose isomerase (PMI), and the reversible PMI reaction also enables exogenous mannose to be catabolized via the glycolytic pathway (Neidhardt et al., 1996 ). The manA gene is generally present in E. coli and S. enterica due to its role in mannose catabolism, and maps as an individual gene not associated with polysaccharide gene clusters (Neidhardt et al., 1987
). manB and manC encode phosphomannomutase (PMM) and GDP-mannose pyrophosphorylase (GMP), respectively. These genes are exclusively used for GDP-D-mannose synthesis, and are located within relevant polysaccharide gene clusters. In most examples found so far, both genes are transcribed as part of the same operon, proceeding from manC into manB.
Colanic acid (CA) or M antigen is an EPS produced by E. coli, S. enterica and other enteric bacteria. CA contains L-fucose, and the manB and manC genes, required for the production of GDP-L-fucose via GDP-D-mannose, are located within the CA gene cluster (Aoyama et al., 1994 ; Stevenson et al., 1991
), which maps upstream of the O antigen gene cluster. Strains with an O antigen containing mannose and/or sugars with precursors derived from GDP-D-mannose have a separate set of manB and manC genes in their O antigen gene cluster. For the sake of brevity, the location of specific manB and manC genes will be denoted, when necessary, in subscript following the gene name. For example, the manB genes from the O antigen and CA gene clusters will be referred to as manBOAg and manBCA, respectively. In S. enterica, O antigen genes are generally of lower GC content (ranging between 0·30 and 0·50) than the chromosomal mean (0·52), whereas the manBCA and manCCA genes of S. enterica LT2 have a mean GC content of 0·62. Interestingly, the manBOAg genes of S. enterica C1 (Lee et al., 1992
) and E. coli O7 and O157 (Marolda & Valvano, 1993
; Wang & Reeves, 1998
) display a high level of sequence identity to the manBCA genes of S. enterica LT2 and E. coli K-12, respectively, indicating that in these cases the manBOAg gene was derived from a CA manB gene (Lee et al., 1992
; Marolda & Valvano, 1993
; Wang & Reeves, 1998
).
Homologues of the S. enterica manB and manC genes have been identified in a broad range of species and repeat-unit polysaccharide gene clusters. Therefore, the manB and manC genes provide a basis for following lateral gene transfer, as unlike most other genes in these clusters, the distinct homologies of the GDP-mannose pathway genes enable evolutionary relationships to be studied. This initial study focuses on determining the extent of variation within S. enterica and, for the manBOAg gene, the relationships with the CA isogene.
Twenty-one S. enterica O antigen structures are known to contain mannose and/or sugars with precursors derived from GDP-D-mannose, based on a chemotype study (Luderitz et al., 1966 ), which included 37 of the 46 currently known structures (Popoff & Minor, 1997
). The manBOAg and manCOAg genes from 8 of these 21 structures have previously been characterized, many of them being closely related. In this study, we sequenced these genes from representative S. enterica strains for the remaining 13 O antigen gene clusters. Furthermore, manBCA and manCCA genes were also sequenced for comparison.
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METHODS |
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Details of bacterial strains, plasmids and bacteriophages used in this study are given in Table 1. Bacteria were routinely grown in nutrient broth (10 g peptone l-1, Amyl Media; 5 g yeast extract l-1, Amyl Media; and 5 g sodium chloride l-1 in water) and on nutrient agar (15 g bacteriological agar l-1 in nutrient broth), which were supplemented with ampicillin (25 µg ml-1) and chloramphenicol (25 µg ml-1) when required. Antisera against the E. coli O9 antigen was supplied by the Institute of Medical and Veterinary Science (Adelaide, South Australia, Australia).
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Sequencing of the manB and manC genes and adjacent areas revealed that many of the clones were derived from the CA gene cluster. In order to find the manB and manC genes derived from the O antigen gene cluster, we screened clones by using a CA-specific primer, based on sequence immediately upstream of the manCCA gene of LT2, and a degenerate primer, based on manB sequences from a variety of species. If this primer combination did not produce a PCR fragment of 1·62 kb the clone was sequenced. For all of the strains tested, the PCR negative clones were found to be derived from the O antigen gene cluster, on the basis of sequence adjacent to the manB and manC genes.
DNA methods.
Alkaline phosphatase, the Expand Long Template PCR System, restriction endonucleases and T4 DNA ligase were obtained from Roche Molecular Biochemicals. Taq polymerase was obtained from Pharmacia Biotech. Plasmid DNA was isolated by using the Wizard Minipreps DNA Purification System (Promega) and chromosomal DNA by using the method described by Bastin et al. (1991) and the Wizard Genomic DNA Purification Kit (Promega). Oligonucleotide primers were synthesized by Beckman or Auspep, or Life Technologies. PCR was carried out using the method described by Saiki et al. (1988)
and an FTS-960 thermal cycler (Corbett Research). PCR products were purified using the Wizard PCR Purification System (Promega), and DNA sequencing was carried out by the Sydney University and Prince Alfred Macromolecular Analysis Centre (SUPAMAC) (Sydney, New South Wales, Australia).
Computer analysis.
The manB and manC nucleotide sequences were edited and assembled using the Phred, Phrap and Consed programs (Ewing & Green, 1998 ; Ewing et al., 1998
; Gordon et al., 1998
). Sequences were aligned using the PILEUP program (Feng & Doolittle, 1986
), and phylogenetic analysis was carried out using the PHYLIP (Felsenstein, 1993
) and MULTICOMP (Reeves et al., 1994
) programs. These programs were accessed using the Australian National Genomic Information Service (Reisner et al., 1993
).
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RESULTS AND DISCUSSION |
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In general, the manCOAg genes do not display a high level of identity to the LT2 manCCA gene (Stevenson et al., 1991 ). However, the manCOAg genes of M255 (O18), M286 (O41) and M324 (O11) (Fig. 1
) are chimeric in structure, resembling in part both a manCCA gene (3' end) and a manCOAg gene (low GC content) (5' end). We also sequenced the manBCA and manCCA genes from selected S. enterica strains as the basis for a more detailed comparison.
Phylogenetic analysis of the GDP-mannose pathway
To examine the phylogenetic relationships of the derived ManB and ManC sequences, evolutionary trees were constructed using the neighbour-joining method (Saitou & Nei, 1987 ). The trees, shown in Fig. 2(a
, b
), also include ManB and ManC sequences from several other species, and for both trees S. enterica and E. coli sequences predominantly cluster together. In considering this, we also constructed a tree based on the N-terminal end of the ManC sequences, which included the O antigen (not CA-like) segments of the chimeric manCOAg genes from M255 (O18), M286 (O41) and M324 (O11) (Fig. 2c
). It can be seen in Fig. 2(c)
that the O18, O41 and O11 5' gene segments also cluster with S. enterica and E. coli sequences. Furthermore, in addition to the CA and CA-like sequences (discussed in detail later), there are a number of distinct relationships among the S. enterica ManB and ManC sequences which we will discuss.
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S. enterica O35 has the same O antigen structure as E. coli O111 and it is interesting that their manBOAg and manCOAg genes are closely related (Fig. 2a, b
, respectively). The level of divergence between their manBOAg and manCOAg genes (data not shown) is consistent with these genes, and presumably the whole O antigen gene cluster, having been present in the common ancestor and maintained in both extant species. Similarly, S. enterica O50 and O30 have the same O antigen structures as E. coli O55 and O157, respectively. However, in these examples, the manCOAg genes are more divergent and not as closely related (Fig. 2b
). The S. enterica O50 and O30 and E. coli O157 manBOAg genes are CA-like, and are therefore not relevant in discussing common ancestry.
The S. enterica O16 and O39 O antigen structures contain both D-mannose and L-fucose, and their manCOAg and CA-like manBOAg genes are unique in S. enterica, as they are separated by an additional gene. Their manCOAg genes are almost identical (99·9% identity), and are related to those from E. coli O41 and O125 (this study), Yersinia pseudotuberculosis 1b (Skurnik et al., 2000 ) and Yersinia enterocolitica O8 (Zhang et al., 1997
) (Fig. 2b
). Interestingly, in E. coli O41 and O125 and Y. pseudotuberculosis 1b, the manCOAg and manBOAg genes are also separated by an additional gene. In each case, the additional gene encodes a putative glycosyl transferase, which like the manCOAg gene, is related amongst these species (data not shown).
Many of the well-documented O antigens (A, B, D and E) are structurally similar, based on O-unit linkages and sugar composition, and have related gene clusters (discussed above). The individual O antigen groups with related manCmanB gene assemblages, which in some cases include adjacent genes, also have structural elements in common. The relationships within these groups are only based on parts of the O antigen gene cluster; however, they indicate that there are conserved groups of O antigen genes, related to structural elements, which can cross species boundaries.
CA-like manBOAg genes
There is a very distinct group of manBOAg and manCOAg genes with CA-like characteristics, based on sequence similarity and GC content. In addition to the manBCA and manCCA genes, the CA-like O antigen genes also have a GC content ranging from 0·60 to 0·62. In comparison, the GC content of other manBOAg and manCOAg genes (or gene segments) ranges from 0·34 to 0·43. As in the case of the S. enterica C1 CA-like manBOAg gene, the newly sequenced CA-like manBOAg genes display a high level of identity to the LT2 manBCA gene. In addition, the CA-like manBOAg genes of strains M324 (O11), M286 (O41) and M255 (O18) display CA similarity in the upstream intergenic region and/or manC gene (discussed in detail later). However, the 2860 bp segment at the 3' end exhibits a high level of divergence between strains, a low GC content, and variation in the position of stop codons over a range of 15 bp. In the case of strain M255, the divergence extends 360 bp upstream of the 3' end.
CA-like manBOAg genes are subspecies-specific
The phylogenetic relationships of the manBCA genes and the CA-like manBOAg genes are shown in a tree constructed using the neighbour-joining method (Saitou & Nei, 1987 ) (Fig. 3a
). Only CA and CA-like sequences were used for this analysis and it became clear that these genes display subspecies specificity. The strains initially studied were not selected with subspecies in mind and are predominantly subspecies I. Therefore, to provide further data on subspecies specificity of the CA-like manBOAg genes, we sequenced CA and/or O antigen manB genes from additional strains. In particular, we included both the manBCA and CA-like manBOAg isogenes of some strains, and representative O40 strains from subspecies I, II, IIIa, IIIb, IV and V. It can be seen in Fig. 3(a)
that not only do CA and CA-like manB genes display subspecies specificity but that the topology of this tree is generally in accordance with a tree constructed from the combined sequences of five housekeeping genes (Neidhardt et al., 1996
) (Fig. 3b
). The manCCA genes also clustered according to subspecies (data not shown).
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The only other exceptions to the subspecies specificity of the CA-like manBOAg genes were the sequences from strains M1653 (O40, IIIb) and M326 (O45, VI), which clustered with subspecies I sequences. This presumably reflects intersubspecies transfer of O antigen genes, as also inferred from similar observations on the gnd gene (Reeves, 1997 ; Thampapillai et al., 1994
). Thampapillai et al. (1994)
observed that the gnd genes of several S. enterica strains have a chimeric structure, and proposed that the 5' end of these genes had transferred between subspecies in association with O antigen genes, presumably driven by natural selection for antigenic variation.
Origins of the CA-like O antigen sequence
The manBOAg and manCOAg genes identified in this study are distributed at various positions within the O antigen gene cluster (data not shown), and no definitive pattern is evident. Most of these genes are internally located, and in reference to lateral transfer between subspecies, are presumably transferred with the O antigen gene cluster. In considering this, and the subspecies specificity of the CA-like manBOAg genes, we conclude that in general the CA-like manBOAg and manCOAg genes were acquired after transfer to the relevant subspecies. Therefore, it appears that the CA and O antigen manB genes, of individual strains, are evolving in concert via gene conversion events. These gene conversion events appear to be unidirectional, as we have not seen a manBOAg gene (low GC content), in a CA gene cluster.
The CA-like manBOAg genes from strains M277 (O43), M290 (O50), M324 (O11) and M1654 (O40) are not only subspecies-specific but most closely related to their respective CA isogenes (Fig. 3a). This indicates that relatively recent gene conversion events have occurred within these strains, supporting the above explanation for the subspecies specificity of CA-like manBOAg genes. The manB isogenes from strains M264 (O16), M273 (O39), M286 (O41), M1651 (O40) and M1656 (O40) are not as closely related. However, both genes of each pair still cluster within the same subspecies group, and presumably the CA-like manBOAg genes originated from earlier gene conversion events, after transfer of the O antigen gene cluster to the relevant subspecies.
Distribution of polymorphic nucleotide sites within the CA and CA-like manBOAg genes
Among the S. enterica CA and CA-like manB genes (22 and 17 sequences, respectively), there were 362 polymorphic nucleotide sites within the 1307 bp segment studied (codons 1444, excluding the divergent 3' end) (Fig. 4). The mean pairwise difference was 7·65% at the DNA level, with variation predominantly attributable to differences between subspecies.
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The first partition, from nucleotide positions 123 to 825 (5' end of the gene), separates the subspecies V strains from other subspecies, and reflects the fact that the 3' end of the subspecies V genes (both CA and CA-like) is similar to those of subspecies I, as discussed above. The remaining partition, from nucleotide positions 1076 to 1307, separates the M255 (O18) CA-like manBOAg sequence from all others. This region represents the divergent (low GC content) 3' end of the M255 CA-like manBOAg gene (Fig. 4).
Visual inspection of the polymorphic nucleotide sites (Fig. 4) provides evidence for recent intragenic exchange between the manBCA and CA-like manBOAg isogenes from strain M326. M326 is a subspecies VI strain with a subspecies I CA-like manBOAg gene (discussed above). However, for nucleotide positions 403 to 798 the CA and CA-like manB isogenes are identical. This region most closely resembles subspecies VI manBCA sequence, and was presumably obtained by the CA-like manBOAg gene from its CA isogene. This is an example of a gene conversion event, and over time the entire M326 CA-like manBOAg gene may resemble subspecies VI sequence.
Chimeric CA-like manCOAg genes
Strains M255 (O18), M286 (O41) and M324 (O11) possess a CA-like manBOAg gene and a chimeric manC gene with a CA-like 3' end (Fig. 1). These genes (including the intergenic region) were aligned with their respective CA isogenes. Visual inspection of the polymorphic nucleotide sites (Fig. 5
) reveals that, for M286 and M324, the CA-like O antigen manCmanB gene assemblage (nucleotide positions 3762888 and 4662888, respectively) is very similar to that of their own CA gene clusters. It is likely that a relatively recent gene conversion event involved part of the manCCA gene, in addition to the downstream intergenic region and manBCA gene. In contrast, the O antigen gene cluster of M255 appears to have acquired the CA-like part of its manC gene (positions 8081391) in a separate event to the acquisition of the CA-like manBOAg gene.
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In addition, we also sequenced part of the O6, O39, O41 and O125 manBCA genes. In contrast to the situation in S. enterica, we found that the manB isogenes of individual strains were not particularly similar (data not shown), indicating that the CA-like manBOAg genes have been affected by recombination since any gene conversion events. This is not too surprising, as E. coli does not display a defined subspecies structure, which is probably due to a higher rate of recombination than in S. enterica.
Concluding comments
CA-like manBOAg genes have previously been observed in both E. coli and S. enterica; however in this study, a distinct pattern has emerged. The prevalence of subspecies-specific CA-like manBOAg genes in S. enterica indicates that the manB genes of individual strains are evolving in concert, with a strong preference for the acquisition and maintenance of an entire manBCA gene within the O antigen gene cluster.
The efficiency of individual O antigen genes may not be critical with respect to O antigen expression when these genes are first transferred to S. enterica. In contrast, if the O antigen occupies a particular niche, selection pressure may eventually drive replacement of specific genes with better adapted homologues (if available). In particular, if the manBCA genes were better adapted to expression in S. enterica, this could explain the unidirectional gene conversion events (manBCA genes replacing manBOAg genes).
However, the replacement of existing CA-like manBOAg genes with subspecies-specific sequence, derived from the manBCA isogene, is unlikely to confer any selective advantage. This suggests that the direction of gene conversion is not random, and a possible explanation could relate to the chromosomal location of the individual manB isogenes. Abdulkarim & Hughes (1996) observed that for the tufA and tufB genes in S. enterica (which are evolving in concert), the rate of sequence transfer was different depending on which tuf gene was the donor. It was suggested that the distribution and frequency of chromosomal breakpoints and chi sites, in relation to the location of the individual tuf genes, contributed to the biased rates of sequence transfer in one direction (Abdulkarim & Hughes, 1996
). At present we have no specific explanation for the unidirectional gene conversion of manBOAg genes; however, there is a precedent in relation to the tufA and tufB genes.
We embarked on this study with the expectancy that the evolutionary relationships of manB and manC genes would provide information on the origin of O antigen gene clusters. Due to the unexpected finding that homogenization of S. enterica manBOAg genes is widespread, limited information is provided in this regard. However, for both the ManB and ManC trees (Fig. 2a, b
, respectively) there is a general clustering of sequences from E. coli, S. enterica and other related species (Klebsiella pneumoniae, Y. enterocolitica and Y. pseudotuberculosis), which encompasses a number of distinct relationships discussed above. This suggests that in relatively recent times, gene capture from a distant source (as suggested by atypical GC content) has occurred infrequently and that the manB and manC genes, and their associated gene assemblages, are maintained and continue to evolve within these species. It is not surprising that genetic exchange would occur more frequently between closely related species; however, a broad-range study of the GDP-mannose pathway genes (encompassing many bacterial species) is required.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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Aoyama, K. M., Haase, A. M. & Reeves, P. R.(1994). Evidence for effect of random genetic drift on G+C content after lateral transfer of fucose pathway genes to Escherichia coli K-12. Mol Biol Evol 11, 829-838.[Abstract]
Bastin, D. A., Romana, L. K. & Reeves, P. R.(1991). Molecular cloning and expression in Escherichia coli K-12 of the rfb gene cluster determining the O antigen of an E. coli O111 strain. Mol Microbiol 5, 2223-2231.[Medline]
Boyd, E. F., Nelson, K., Wang, F.-S., Whittam, T. S. & Selander, R. K.(1994). Molecular genetic basis of allelic polymorphism in malate dehydrogenase (mdh) in natural populations of Escherichia coli and Salmonella enterica. Proc Natl Acad Sci USA 91, 1280-1284.[Abstract]
Brown, P. K., Romana, L. K. & Reeves, P. R.(1992). Molecular analysis of the rfb gene cluster of Salmonella serovar Muenchen (strain M67): genetic basis of the polymorphism between groups C2 and B. Mol Microbiol 6, 1385-1394.[Medline]
Curd, H., Liu, D. & Reeves, P. R.(1998). Relationships among the O-antigen gene clusters of Salmonella enterica groups B, D1, D2, and D3. J Bacteriol 180, 1002-1007.
Ewing, B. & Green, P.(1998). Base-calling of automated sequencer traces using phred. II. Error probabilities. Genome Res 8, 186-194.
Ewing, B., Hillier, L., Wendl, M. C. & Green, P.(1998). Base-calling of automated sequencer traces using phred. I. Accuracy assessment. Genome Res 8, 175-185.
Felsenstein, J. (1993). PHYLIP, version 3.5. University of Washington, Seattle, USA.
Feng, D. & Doolittle, R. F.(1986). Progressive sequence alignment as a prerequisite to correct phylogenetics trees. J Mol Evol 25, 351-360.
Frick, D. N., Townsend, B. D. & Bessman, M. J.(1995). A novel GDP-mannose mannosyl hydrolase shares homology with the MutT family of enzymes. J Biol Chem 270, 24086-24091.
Gordon, D., Abajian, C. & Green, P.(1998). Consed: a graphical tool for sequence finishing. Genome Res 8, 195-202.
Jayaratne, P., Bronner, D., MacLachlan, P. R., Dodgson, C., Kido, N. & Whitfield, C.(1994). Cloning and analysis of duplicated rfbM and rfbK genes involved in the formation of GDP-mannose in Escherichia coli O9:K30 and participation of rfb genes in the synthesis of the group I K30 capsular polysaccharide. J Bacteriol 176, 3126-3139.[Abstract]
Jiang, X. M., Neal, B., Santiago, F., Lee, S. J., Romana, L. K. & Reeves, P. R.(1991). Structure and sequence of the rfb (O antigen) gene cluster of Salmonella serovar typhimurium (strain LT2). Mol Microbiol 5, 695-713.[Medline]
Kido, N., Torgov, V. I., Sugiyama, T., Uchiya, K., Sugihara, H., Komatsu, T., Kato, N. & Jann, K.(1995). Expression of the O9 polysaccharide of Escherichia coli: sequencing of the E. coli O9 rfb gene cluster, characterization of mannosyl transferases, and evidence for an ATP-binding cassette transport system. J Bacteriol 177, 2178-2187.[Abstract]
Lai, V., Wang, L. & Reeves, P. R.(1998). Escherichia coli clone sonnei (Shigella sonnei) had a chromosomal O antigen gene cluster prior to gaining its current plasmid-borne O antigen genes. J Bacteriol 180, 2983-2986.
Lee, S. J., Romana, L. K. & Reeves, P. R.(1992). Sequence and structural analysis of the rfb (O antigen) gene cluster from a group C1 Salmonella enterica strain. J Gen Microbiol 138, 1843-1855.[Medline]
Le Minor, L., Veron, M. & Popoff, M. (1982). Taxonomie des Salmonella. Ann Microbiol (Inst Pasteur) 133B, 223243.
Le Minor, L., Popoff, M. Y., Laurent, B. & Hermant, D. (1986). Individualisation Dune septieme sous-espece de Salmonella: S. choleraesuis subsp. indica subsp. nov. Ann Inst Pasteur/Microbiol 137B, 211217.
Liu, D., Verma, N. K., Romana, L. K. & Reeves, P. R. (1991). Relationships among the rfb regions of Salmonella serovars A, B, and D. J Bacteriol 173, 48144819.[Medline]
Luderitz, O., Staub, A. M. & Westphal, O.(1966). Immunochemistry of O and R antigens of Salmonella and related Enterobacteriaceae. Bacteriol Rev 30, 192-255.[Medline]
Marolda, C. L. & Valvano, M. A.(1993). Identification, expression, and DNA sequence of the GDP-mannose biosynthesis genes encoded by the O7 rfb gene cluster of strain VW187 (Escherichia coli O7:K1). J Bacteriol 175, 148-158.[Abstract]
Neidhardt, F. C., Ingraham, J. L., Magasanik, B., Schaechter, M. & Umbarger, H. E. (1987). Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology. Washington, DC: American Society for Microbiology.
Neidhardt, F. C., Curtiss, R., III, Ingraham, J. L. & 7 other authors (1996). Escherichia coli and Salmonella: Cellular and Molecular Biology, 2nd edn. Washington, DC: American Society for Microbiology.
Nelson, K., Whittam, T. S. & Selander, R. K.(1991). Nucleotide polymorphism and evolution in the glyceraldehyde-3-phosphate dehydrogenase gene (gapA) in natural populations of Salmonella and Escherichia coli. Proc Natl Acad Sci USA 88, 6667-6671.[Abstract]
Popoff, M. Y. & Minor, L. L. (1997). Antigenic Formulas of the Salmonella Serovars, 7th revision. Paris: WHO Collaborating Centre for Reference and Research on Salmonella. Institut Pasteur.
Reeves, M. W., Evins, G. M., Heiba, A. A., Plikaytis, B. D. & Farmer, J. J.III(1989). Clonal nature of Salmonella typhi and its genetic relatedness to other Salmonellae as shown by multilocus enzyme electrophoresis, and proposal of Salmonella bongori. J Clin Microbiol 27, 313-320.[Medline]
Reeves, P.(1991). The O antigen of Salmonella. Todays Life Sci 3, 30-40.
Reeves, P. R.(1992). Variation in O antigens, niche specific selection and bacterial populations. FEMS Microbiol Lett 100, 509-516.
Reeves, P. R.(1993). Evolution of Salmonella O antigen variation by interspecific gene transfer on a large scale. Trends Genet 9, 17-22.[Medline]
Reeves, P. R.(1997). Specialized clones and lateral transfer in pathogens. In Ecology of Pathogenic Bacteria: Molecular and Evolutionary Aspects , pp. 237-254. Edited by B. A. M. van der Zeijst, W. P. M. Hoekstra, J. D. A. van Embden & A. J. W. van Alphen. Amsterdam:Elsevier.
Reeves, P. R., Farnell, L. & Lan, R.(1994). MULTICOMP: a program for preparing sequence data for phylogenetic analysis. Comput Appl Biosci 10, 281-284.[Abstract]
Reisner, A. H., Bucholtz, C. A., Smelt, J. & McNeil, S. (1993). Australias National Genomic Information Service. In Proceedings of the Twenty-Sixth Annual Hawaii International Conference on Systems Science, pp. 595602.
Saiki, R. K., Gelfand, D. H., Stofell, S., Scharf, S. J., Higuchi, R., Horn, G. T., Mullis, K. B. & Erlich, H. A.(1988). Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239, 487-491.[Medline]
Saitou, N. & Nei, M.(1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4, 406-425.[Abstract]
Skurnik, M., Peippo, A. & Ervelä, E.(2000). Characterization of the O-antigen gene clusters of Yersinia pseudotuberculosis and the cryptic O-antigen gene cluster of Y. pestis shows that the plague bacillus is most closely related to and has evolved from Y. pseudotuberculosis serotype O:1b. Mol Microbiol 37, 316-330.[Medline]
Stephens, J. C.(1985). Statistical methods of DNA sequence analysis: detection of intragenic recombination or gene conversion. Mol Biol Evol 2, 539-556.[Abstract]
Stevenson, G., Lee, S. J., Romana, L. K. & Reeves, P. R.(1991). The cps gene cluster of Salmonella strain LT2 includes a second mannose pathway: sequence of two genes and relationship to genes in the rfb gene cluster. Mol Gen Genet 227, 173-180.[Medline]
Sueoka, N.(1988). Directional mutation pressure and neutral molecular evolution. Proc Natl Acad Sci USA 85, 2653-2657.[Abstract]
Sueoka, N.(1992). Directional mutation pressure, selective constraints, and genetic equilibria. J Mol Evol 34, 95-114.[Medline]
Sugiyama, T., Kido, N., Komatsu, T., Ohta, M., Jann, K., Jann, B., Saeki, A. & Kato, N.(1994). Genetic analysis of Escherichia coli O9 rfb: identification and DNA sequence of phosphomannomutase and GDP-mannose pyrophosphorylase genes. Microbiology 140, 59-71.[Abstract]
Thampapillai, G., Lan, R. & Reeves, P. R.(1994). Molecular variation at the gnd locus of 34 natural isolates of Salmonella enterica: DNA sequence: evidence for probable chi-dependent interallelic recombination. Mol Biol Evol 11, 813-828.[Abstract]
Vieira, J. & Messing, J.(1982). The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene 19, 259-268.[Medline]
Wang, L. & Reeves, P. R.(1998). Organization of Escherichia coli O157 O antigen gene cluster and identification of its specific genes. Infect Immun 66, 3545-3551.
Wang, L., Romana, L. K. & Reeves, P. R.(1992). Molecular analysis of a Salmonella enterica group E1 rfb gene cluster: O antigen and the genetic basis of the major polymorphism. Genetics 130, 429-443.
Xiang, S. H., Hobbs, M. & Reeves, P. R.(1994). Molecular analysis of the rfb gene cluster of a group D2 Salmonella enterica strain: evidence for its origin from an insertion sequence-mediated recombination event between group E and D1 strains. J Bacteriol 176, 4357-4365.[Abstract]
Zhang, L., Radziejewska-Lebrecht, J., Krajewska-Pietrasik, D., Toivanen, P. & Skurnik, M.(1997). Molecular and chemical characterization of the lipopolysaccharide O-antigen and its role in the virulence of Yersinia enterocolitica serotype O8. Mol Microbiol 23, 63-76.[Medline]
Received 9 June 2000;
revised 25 September 2000;
accepted 23 October 2000.