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 molecular epidemiology of Pasteurella multocida has rarely been studied at the farm level in cattle. The aim of this study was to determine whether single or multiple strains of P. multocida tend to exist within farms. Molecular characterisation was carried out on isolates obtained from nasal swabs from 105 calves from 32 randomly selected beef and dairy farms located throughout Scotland, and from 131 calves from 20 farms in the Mayenne region of France, where sampling occurred in response to respiratory disease outbreaks. P. multocida isolates were characterised by random-amplified polymorphic DNA (RAPD) typing and pulsed-field gel electrophoresis (PFGE) using restriction enzyme ApaI. In addition, isolates representative of each farm/RAPD profile combination were typed by multilocus sequence typing (MLST). Among 105 Scottish isolates, 15 RAPD profiles were distinguished. The majority of farms (27/32) had indistinguishable profiles in all positive animals. Five farms had two profiles. Among 140 French isolates, 23 RAPD profiles were distinguished. More within-farm heterogeneity was observed although 10/20 farms had just one profile (E4) in sampled calves. Profile E4 accounted for 60% (84/140) of French isolates. PFGE was more discriminatory than RAPD but confirmed results with respect to within farm homogeneity or heterogeneity of strains, whereas MLST was not discriminatory enough for farm level epidemiology. As in other host species, either several strains or one dominant strain of P. multocida may exist within farms, with evidence for a role of management factors such as movements onto the farm in the number of strains detected.Veterinary Microbiology 03/2011; 151(3-4):329-35. DOI:10.1016/j.vetmic.2011.03.018 · 2.73 Impact Factor
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ABSTRACT: Three dairy calf-rearing properties experienced high mortality in calves during 2008 and 2009. Affected calves were aged 13-18 weeks (Farm I), 6 months (Farm II), and 2-11 weeks (Farm III), and the mortality rate was 22/175 (13%), 5/80 (6%), and 60/900 (7%), respectively. CLINICAL AND LABORATORY FINDINGS: Affected calves rapidly became moribund, were in respiratory distress, and had a fever (40-41°C). Post-mortem examination of nine calves revealed fibrinopurulent pleuritis, pericarditis, and peritonitis. This was confirmed histopathologically on tissues from three calves, one from each farm; aggregates of small Gram-negative coccobacilli were evident on Gram stain. Pasteurella multocida was cultured from tissues from affected calves on the three farms, and PCR of DNA extracted from tissue samples amplified cap-sular type B-specific DNA. Multi-locus sequence typing (MLST) demonstrated that all capsular type B isolates belonged to the same sequence type (ST), ST62, but did not belong to serotype B:2, the only B serotype classified as causing haemorrhagic septicaemia by the Office International des Epizooties (OIE). Pleuritis and peritonitis due to infection with P. multocida capsular type B strain. Haemorrhagic septicaemia was excluded as a cause of disease from the three farms, however P. multocida was the primary agent in the affected calves. It is possible the agent has been present in New Zealand for some time but not reported, as there had been no transfer of animals between affected farms. Emergence of the syndrome could potentially be a result of factors other than just the presence of the organism, such as changing management. The syndrome described may be of increasing importance in the future.New Zealand veterinary journal 02/2011; 59(1):40-5. DOI:10.1080/00480169.2011.547168 · 1.22 Impact Factor
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ABSTRACT: Pasteurella multocida is a widespread respiratory pathogen in pigs associated with atrophic rhinitis and contributing to aggravation of the pulmonary lesions. The aims of the present study were to characterize isolates of P. multocida from porcine bronchopneumonia by pulsed-field gel electrophoresis (PFGE), PCR based capsular typing and multilocus sequence typing (MLST) and to compare clonal complexes outlined with the type of histological lung lesions to investigate if a correlation between clonal lineages and lesions might exist. Isolates of P. multocida were obtained from cases of cranioventrally located porcine bronchopneumonia. All lung lesions were described and classified according to histological lesions. A total of 139 isolates, from lung (n=111), pericardial sac (n=21) and kidney (n=7) of 111 pigs were described using PFGE with ApaI as the restriction enzyme. Furthermore, 20 and 29 isolates were characterized by capsular serotyping and multilocus sequence typing, respectively. PFGE demonstrated 15 different clusters showing 50% or more similarity. All selected isolates were of capsular serotype A and only three main sequence types (ST) were detected among the isolates. Associations were not found between histopathology and clonal complexes of P. multocida. In conclusion, PFGE demonstrated a high diversity of genotypes of P. multocida associated with porcine bronchopneumonia. However, isolates obtained mainly belonged to few STs, indicating that isolates of P. multocida associated with porcine bronchopneumonia originates from a limited number of clonal lineages and therefore might have adapted to porcine hosts. No correlation was demonstrated between genotypes and types of lesions, and extra-pulmonary spreading was only rarely demonstrated.Veterinary Microbiology 03/2011; 150(3-4):354-61. DOI:10.1016/j.vetmic.2011.02.050 · 2.73 Impact Factor