Development of a multi-locus sequence typing scheme for avian isolates of Pasteurella multocida.
ABSTRACT A total of 63 isolates of Pasteurella multocida from Australian poultry, all associated with fowl cholera outbreaks, and three international reference strains, representing the three subspecies within P. multocida were used to develop a multi-locus sequence typing scheme. Primers were designed for conserved regions of seven house-keeping enzymes -adk, est, gdh, mdh, pgi, pmi and zwf - and internal fragments of 570-784 bp were sequenced for all isolates and strains. The number of alleles at the different loci ranged from 11 to 20 and a total of 29 allelic profiles or sequence types were recognised amongst the 66 strains. There was a strong concordance between the MLST data and the existing multi-locus enzyme electrophoresis and ribotyping data. When used to study a sub-set of isolates with a known detailed epidemiological history, the MLST data matched the results given by restriction endonuclease analysis, pulsed-field gel electrophoresis, ribotyping and REP-PCR. The MLST scheme provides a high level of resolution and is an excellent tool for studying the population structure and epidemiology of P. multocida.
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ABSTRACT: The population biology of Streptococcus pneumoniae is poorly understood. Most of the important issues could be addressed by the molecular characterization of large, well sampled populations from carriage and from the different manifestations of pneumococcal disease. The authors have therefore developed a pneumococcal multilocus sequence typing scheme and database by sequencing approximately 450 bp fragments of seven housekeeping loci from 295 isolates. The combination of alleles at the seven loci provided an allelic profile, or sequence type (ST), and the relatedness between isolates was obtained by constructing a dendrogram from the matrix of pairwise differences between STs. The typing scheme was validated using pneumococci of known genetic relatedness and could resolve >6 billion STs. Among 274 isolates from recent cases of invasive pneumococcal disease in eight countries, 143 STs were resolved, but 12 STs contained at least five isolates (range 5-21 isolates). The repeated recovery of indistinguishable isolates from invasive disease in different countries implies that these STs define strains with an increased capacity to cause invasive disease. The relationship between STs and serotypes suggested that, in the longer term, capsular genes have been distributed horizontally within the pneumococcal population, but in the short term, expansion of clones occurs with only occasional changes of serotype. The multilocus sequence typing scheme provides a powerful new approach to the characterization of pneumococci, since it provides molecular typing data that are electronically portable between laboratories, and which can be used to probe aspects of the population and evolutionary biology of these organisms. A Web site for the molecular characterization of pneumococci by MLST is available (http ://mlst.zoo.ox.ac.uk).Microbiology 12/1998; 144 ( Pt 11):3049-60. · 2.85 Impact Factor
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ABSTRACT: Multilocus sequence typing (MLST) has been applied to 266 Campylobacter jejuni isolates, mainly from veterinary sources, including cattle, sheep, poultry, pigs, pets, and the environment, as well as isolates from human cases of campylobacteriosis. The populations of veterinary and human isolates overlap, suggesting that most veterinary sources should be considered reservoirs of pathogenic campylobacters. There were some associations between source and sequence type complex, indicating that host or source adaptation may exist. The pig isolates formed a distinct group by MLST and may well represent a potential pig-adapted clone of C. jejuni. A subset (n = 82) of isolates was reanalyzed with a second MLST scheme which provided a unique set of isolates that had been analyzed at a total of 12 loci. The distribution of isolates among the complexes in each of the two schemes was similar but not identical. In addition to isolates from human outbreaks, one group of isolates that were not epidemiologically linked was also identical at all 12 loci. This group of isolates is believed to represent another stable strain of C. jejuni.Applied and Environmental Microbiology 12/2003; 69(11):6370-9. · 3.68 Impact Factor
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ABSTRACT: One hundred and fifty-three bovine Pasteurella multocida strains recovered primarily from cases of pneumonia and mastitis in England and Wales over an 11-year period were characterised by capsular PCR typing, comparison of outer membrane protein (OMP) profiles, and multilocus sequence analysis. All of the strains were of capsular type A with the exception of a single capsular type F isolate. Thirteen distinct OMP profiles (OMP-types) were identified based mainly on molecular mass heterogeneity of the heat-modifiable (OmpA) and porin (OmpH) proteins. However, 85% of the isolates were represented by just five OMP-types and 39% of the strains were of a single OMP-type. Multilocus sequence analysis revealed a limited degree of genetic diversity among bovine P. multocida isolates; strains of the same OMP-type have identical genetic backgrounds and represent distinct clones. Analysis of OMP variation was more discriminating than multilocus sequence analysis because strains of different OMP-types had the same, or similar, genetic backgrounds. The association of a small number of clones with the majority of cases of bovine pneumonia suggests that these clones have an increased capacity to cause disease compared to less frequently recovered clones. Molecular mass heterogeneity of OmpA and OmpH, in strains of the same or similar genetic background, suggests that these proteins are subject to diversifying selection within the host and might play important roles in host-pathogen interactions. Comparison of the OMP profiles of bovine isolates with those of avian, ovine and porcine strains showed that a high proportion of the respiratory tract infections in each of these species are caused by different strains of P. multocida. However, the presence of small numbers of closely related strains in more than one host species suggests that transmission of bacteria between different host species is also a factor in the population biology of P. multocida.Veterinary Microbiology 05/2004; 99(2):145-58. · 3.13 Impact Factor