Diversity of avian Pasteurella multocida strains based on capsular PCR typing and variation of the OmpA and OmpH outer membrane proteins

One hundred avian Pasteurella multocida isolates recovered from cases of fowl cholera and related infections in England and Wales over a 13-year period were characterised by capsular PCR typing and analysis of outer membrane protein (OMP) profiles. Sixty-eight percent of the strains were of capsular type A, 14% were type F, 5% were type D, 4% were type B and 9% were untypable. Nineteen distinct OMP profiles (OMP-types) were identified based mainly on molecular mass heterogeneity of the heat-modifiable (OmpA) and porin (OmpH) proteins. Fifty-six percent of the isolates were represented by 15 OMP-types, whereas 44% of the isolates were associated with four OMP-types. The extensive molecular mass heterogeneity of the OmpA and OmpH proteins supports previous findings that avian P. multocida strains are very diverse. Furthermore, the isolates studied were associated with different clinical symptoms and were recovered from a wide range of lesions and tissues. The high degree of strain diversity together with the wide variety of clinical symptoms suggest that certain avian strains of P. multocida are opportunistic pathogens of relatively low virulence. Strains of capsular types B, D and F, as well as the untypable isolates, were associated exclusively with specific OMP-types and represent distinct and widely disseminated clonal groups. These observations support the view that avian strains of P. multocida have a clonal population structure. Based on previous studies, the molecular mass heterogeneity of the OmpA and OmpH proteins might provide a selective advantage to P. multocida by generating antigenic variation.     

Comparative analyses of Pasteurella multocida strains associated with the ovine respiratory and vaginal tracts

Thirty-five isolates of Pasteurella multocida from the vagina and respiratory tract of sheep were compared by analysing their capsular polysaccharide types and outer membrane protein profiles. The phylogenetic relationships of selected isolates with respect to reference strains of P. multocida were also determined by comparative 16S rRNA sequence analysis. Three capsular types, A, D and F, and three major outer membrane protein types were identified, and there were four different combinations of these characteristics which probably marked four individual clones of P. multocida. Strains representing three of these clones were recovered from cases of ovine pneumonia, whereas isolates of the fourth clone were associated exclusively with the vagina of healthy ewes and the liver of a dead septicaemic lamb on the same farm. Analysis of the 16S rRNA sequences showed that there was 100 per cent identity between representative pneumonic isolates and reference strains of P. multocida subspecies galliseptica and P. multocida subspecies multocida. The 16S rRNA genes of representative vaginal and liver isolates from the same farm were identical but differed from the other strains at one nucleotide position, providing strong evidence that the vaginal and liver isolates represent a distinct subpopulation of P. multocida.     

Pasteurella multocida and bovine respiratory disease

Pasteurella multocida is a pathogenic Gram-negative bacterium that has been classified into three subspecies, five capsular serogroups and 16 serotypes. P. multocida serogroup A isolates are bovine nasopharyngeal commensals, bovine pathogens and common isolates from bovine respiratory disease (BRD), both enzootic calf pneumonia of young dairy calves and shipping fever of weaned, stressed beef cattle. P. multocida A:3 is the most common serotype isolated from BRD, and these isolates have limited heterogeneity based on outer membrane protein (OMP) profiles and ribotyping. Development of P. multocida-induced pneumonia is associated with environmental and stress factors such as shipping, co-mingling, and overcrowding as well as concurrent or predisposing viral or bacterial infections. Lung lesions consist of an acute to subacute bronchopneumonia that may or may not have an associated pleuritis. Numerous virulence or potential virulence factors have been described for bovine respiratory isolates including adherence and colonization factors, iron-regulated and acquisition proteins, extracellular enzymes such as neuraminidase, lipopolysaccharide, polysaccharide capsule and a variety of OMPs. Immunity of cattle against respiratory pasteurellosis is poorly understood; however, high serum antibodies to OMPs appear to be important for enhancing resistance to the bacterium. Currently available P. multocida vaccines for use in cattle are predominately traditional bacterins and a live streptomycin-dependent mutant. The field efficacy of these vaccines is not well documented in the literature.     

Characterization and comparison of antimicrobial susceptibilities and outer membrane protein and plasmid DNA profiles of Pasteurella haemolytica and certain other members of the genus Pasteurella.

The outer membrane protein (OMP), plasmid, and antimicrobial resistance profiles of Pasteurella haemolytica serotypes 1 through 12, a bovine isolate of P multocida, a chicken isolate of P multocida, and an unidentified Pasteurella species of bovine origin were examined. Isolates of P haemolytica serotypes belonging to the same biotype possessed similar OMP profiles. Biotype A isolates contained 2 prominent OMP of 43 kilodaltons (kD) and 29 kD, whereas biotype-T serotypes contained 3 major OMP of 43, 36, and 25 kD. The major OMP profiles of the 2 P multocida isolates and the unidentified Pasteurella species were different from each other and from P haemolytica isolates. Plasmid DNA screening indicated both plasmid-containing and plasmid-free P haemolytica and P multocida isolates. Multiple drug resistance was found in pasteurellae isolates with and without plasmids. However, a relationship between drug resistance and plasmid isolation was found in 3 of 4 haemolytica serotype 1 field isolates, all of which contained a 2.51-megadalton plasmid and had multiple drug resistance for benzylpenicillin, ampicillin, streptomycin, and tetracycline.     

禽多杀性巴氏杆菌血清型与外膜蛋白型相关性研究

为了解禽多杀性巴氏杆菌(Pasteurella multocida,Pm)分离菌株的荚膜血清型、菌体血清型与外膜蛋白型之间的相关性,首先对自行分离的10个菌株采用间接血凝试验和琼脂扩散试验进行鉴定(均为A:1);然后采用超声波破碎、高速离心和十二烷基肌氨酸钠提取外膜蛋白,通过SDS-PAGE电泳的方法对上述10个菌株与C48-1(A:1)、X73(A:1)、P1059(A:3)、CU(A:3,4)等一起进行外膜蛋白(Outer membrane proteins,OMP)分型研究。结果表明:14个禽多杀性巴氏杆菌菌株以2个主要蛋白OmpH和OmpA的差异为依据分为3种主要OMP型;依据次要蛋白的差异,OMP-1,3菌株进一步分为OMP型1.1,1.2和3.1,3.2,其中OMP型1.1有6个菌株,OMP型1.2有5个菌株;OMP型2有1株(YZ7031);P1059为OMP型3.1;CU为OMP型3.2;另外,血清型为A:1的12个菌株中有11个菌株外膜蛋白型均为OMP-1型,血清型为A:3、A:3,4的菌株外膜蛋白型属于3型。说明禽多杀性巴氏杆菌血清型与特定的外膜蛋白型具有很强的相关性。     

Genetic diversity and associated pathology of Pasteurella multocida isolated from porcine pneumonia

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.     

Expression of iron-regulated outer membrane proteins by porcine strains of Pasteurella multocida.

The outer membrane protein (OMP) profiles of two strains of capsular type A Pasteurella multocida isolated from the lungs of pigs with enzootic pneumonia were studied. Sarkosyl extracted OMPs from P. multocida grown under iron-restricted and iron-replete conditions were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis. Results showed that the iron-regulated outer membrane proteins (IROMPs) with molecular masses of 74 kDa, 94 kDa, 99 kDa and 109 kDa were expressed by strain A52, while 74 kDa, 82 kDa, 94 kDa and 99 kDa IROMPs were expressed by strain B80. Swine immune sera, obtained from pigs which were first immunized with a polyvalent P. multocida type A and type D bacterin and subsequently challenged with type A strain of P. multocida, contained antibodies against the IROMPs. These antibodies cross-reacted with the IROMPs expressed by avian strain P1059 of P. multocida. Convalescent-phase serum obtained from turkeys which survived fowl cholera, also cross-reacted with the IROMPs from porcine strains of P. multocida. These results suggested that IROMPs from porcine and avian strains of P. multocida may share common epitopes that were recognized by swine immune serum as well as turkey convalescent-phase serum.     

Molecular epidemiology of Pasteurella multocida in dairy and beef calves

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.     

Random amplification of polymorphic DNA and phenotypic typing of Zimbabwean isolates of Pasteurella multocida

Eighty-one isolates presumptively identified as Pasteurella multocida from a variety of diseases in animals in Zimbabwe were subjected to biochemical characterization, capsular typing and RAPD analysis. The majority of isolates (over 80%) were assigned into named taxa and were predominantly P. multocida subsp. multocida and P. multocida subsp. septica, whilst the remainder were unassigned. Serogroup A was predominant among the three capsular types (A, B and D) of P. multocida detected. Three main RAPD clusters and three subclusters were observed among the majority of isolates (93.8%), whilst the remainder was found to be weakly related. Nine different groups of strains with similar RAPD profiles (100% similarity) were also observed. The reference strain of capsular serogroup F clustered with the reference strain of P. multocida subsp. septica, whilst all other serogroups clustered with reference strains of subsp. multocida and gallicida. Notably, serogroups A and D were observed to be closely related to the reference strain of subsp. multocida. The relationship between biotype, capsular type, host origin and disease manifestation was not clear-cut. However, most pig isolates of subsp. multocida clustered together as did most cattle isolates of subsp. multocida. RAPD tended to separate subsp. multocida from septica.     

OmpA and antigenic diversity of bovine Chlamydophila pecorum strains

Infections with the intracellular bacterium Chlamydophila (C.) pecorum are highly prevalent worldwide in cattle. These infections cause significant diseases such as polyarthritis, pneumonia, enteritis, genital infections and fertility disorders, and occasionally sporadic bovine encephalomyelitis. Subclinical respiratory infections of calves with C. pecorum have been associated with airway obstruction, pulmonary inflammation, and reduced weight gains. This investigation examined four chlamydial strains with biological properties of C. pecorum isolated from feces of clinically normal cattle, from calves with pneumonia, and from bulls with posthitis. The objective was to characterize the evolutionary relationships of these bovine chlamydial isolates to other chlamydiae by genetic analysis of the ompA gene, and by the immunological cross-reactivities in Western immunoblot analysis. PCR typing of the ompA gene identified these isolates as C. pecorum. The OmpA-deduced amino acid dissimilarities between these four strains spanned 10-20%. In phylogenetic analysis, the four isolates clustered with C. pecorum ruminant, porcine, and koala strains of different geographic origins rather than with each other. All four isolates showed different patterns of Western immunoblot reactivity with antiserum against bovine C. pecorum strain LW63, and, interestingly, no cross-reactivity of the OmpA proteins with the anti-LW613 OmpA antibodies. These data underscore the polyphyletic population structure of C. pecorum and suggest that the spectrum of C. pecorum OmpA proteins in a host species can occupy the entire evolutionary bandwidth within C. pecorum. The variant immunoblot reactivities support the notion of considerable genomic plasticity of C. pecorum.     

8株禽多杀性巴氏杆菌外膜蛋白H基因序列分析

为了解国内禽多杀性巴氏杆菌（Pasteurella multocida,Pm）流行株外膜蛋白 H（OmpH）基因的变异情况,参考GenBank中已发表的多杀性巴氏杆菌序列设计1对特异性引物,采用PCR方法对不同来源的8株禽Pm菌株和3个荚膜型参考株（A、B、D）的OmpH基因进行扩增、测序。结果显示,11个菌株的OmpH基因开放阅读框在1002~1071 bp 之间;SignalIP 4.0预测结果表明,信号肽均为N端20个氨基酸残基,成熟蛋白氨基酸残基数量在314~337 aa之间,推测的分子质量在33.76~37.04 ku之间。与GenBank中15个菌株OmpH基因序列比对结果发现,核苷酸同源性在84.9%~100.0%之间;氨基酸同源性在81.5%~100.0%之间;其中C48-1、1010、9003、890920、921012、XJ-e 6个国内禽Pm分离株OmpH序列同源性为100.0%。试验结果表明,国内禽Pm菌株OmpH基因非常保守。     

Molecular characterisation of avian Pasteurella multocida isolates from Australia and Vietnam by REP-PCR and PFGE

A total of 95 isolates of Pasteurella multocida were analysed by pulsed field gel electrophoresis (PFGE) using the enzyme ApaI, including 73 avian isolates from Australia and 22 from Vietnam. The majority of field isolates were capsular Type A, with the predominant somatic serovars of 1, 3, 4 and 3,4. Twenty-one distinct profiles were evident among the Australian isolates, with only 3 profiles observed among the 22 P. multocida strains isolated from Vietnam. Within the Australian isolates, related and unrelated outbreaks could be identified by PFGE. These results correlated well with previously published studies, with greater discrimination shown by PFGE. Repetitive extragenic palindromic sequence PCR (REP-PCR) analysis of representative isolates from PFGE classifications yielded 21 profiles, with most of the subgroups in accordance with PFGE analysis. While REP-PCR was shown to be less discriminating than PFGE, the epidemiological relatedness of strains compared favourably between the techniques. Thus, the ease and rapidity of REP-PCR while maintaining a high level of differentiation, supports the use of REP-PCR as a competent alternative to the more labour-intensive PFGE system for strain identification and epidemiological studies of avian P. multocida.     

Outer membrane protein profiles of Yersinia ruckeri

The outer membrane protein (OMP) profiles of 135 isolates of Yersinia ruckeri, obtained from nine European countries (100 isolates), North America (23 isolates), Australia (six isolates) and South Africa (two isolates), and including four reference strains, were examined by SDS-PAGE. Outer membranes were isolated by selective solubilisation of the cytoplasmic membrane with 0.5% (w/v) sodium N-lauroyl sarcosinate (Sarkosyl). Outer membrane proteins were stable after in vitro passage and there was no variation in OMP profiles due to colony selection. With the exception of a 39.5 kDa peptidoglycan-associated protein there was also no variation at different stages of the growth cycle. The 39.5 kDa protein was not produced during logarithmic growth phase but increased in abundance as the stationary phase progressed. Interstrain variation occurred in the possession of a 36.5 or 38 kDa heat-modifiable protein and in the possession of peptidoglycan-associated proteins in the molecular weight range 36.5 to 40.5 kDa. Based on variation of these proteins five OMP-types, designated OMP-types 1-5, were identified among the 135 isolates examined. Outer membrane protein analysis was demonstrated to be useful in epidemiological studies of Y. ruckeri.     

Characterization of outer membrane protein-enriched extracts from Pasteurella multocida isolated from turkeys

Outer membrane protein (OMP)-enriched extracts of avian strains of Pasteurella multocida were examined by use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Culture medium did not have a significant effect on the OMP profiles of strains of P multocida examined; however, in vivo propagation had an appreciable effect on the OMP profile composition of the reference strain P-1059. Such bacteria, expressed several additional OMP in the 27-kD, 48-kD, 56-kD, 60-kD, 80-kD, and 94-kD molecular mass regions. These OMP were not detected in the electrophorogram of strain P-1059 grown in vitro. The OMP profiles of reference strains of the 16 serotypes of P multocida did not identify any serotype-specific protein markers. Field strains of serotype A:3 had variation in OMP profiles and did not express OMP that all were identical to that expressed by the reference strain P-1059. The live attenuated CU and M9 bacterial vaccine strains expressed strain-specific OMP markers of 48-kD and 45-kD molecular masses, respectively. These strain-specific OMP markers may be used to differentiate these strains from virulent field strains that are of the same serotype and isolated from turkeys that have succumbed to pasteurellosis as a result of vaccine-related reactions or breakdown in immunity.     

Pasteurella multocida and its role in porcine pneumonia

Pasteurella multocida has been recognized as a contributor to debilitating and fatal porcine pneumonia for at least 120 years and there continues to be sustained, unabated high prevalence of the organism in cases submitted for diagnostic work up. Understanding of its role in disease has been limited, in part because of difficulty in reproducing the disease experimentally with capsular type A strains of P. multocida, the predominant type associated with porcine pneumonia. This limitation has stymied the development of improved methods for disease control. In this review, the reports of efforts to reproduce the disease are compared. Reports have indicated induction of pneumonia in combined infections with agents such as hog cholera virus, pseudorabies virus and Mycoplasma hyopneumoniae. Pneumonia has been induced with intratracheal or endobronchial inoculation of anesthetized swine using capsular type A strains. Substantial recent progress in understanding the putative virulence attributes and molecular genetics of P. multocida will likely lead to better understanding of the host-parasite and parasite-parasite interactions in porcine pneumonia associated with this organism. In particular, it seems important to consider the role of biofilm formation in the pathogenesis of this disease. Ultimately, this understanding should provide a foundation for better methods for induction of the experimental disease, development of improved diagnostics, development of better therapeutic/prophylactic pharmaceutical approaches and development of immunoprophylactic products.     

Immunogenicity of iron-regulated outer membrane proteins of Pasteurella multocida A:3 in cattle: molecular characterization of the immunodominant heme acquisition system receptor (HasR) protein

The iron-regulated outer membrane proteins (IROMPs) of Pasteurella multocida A:3 strain 232 (Pm232), a bovine isolate, were investigated as potential immunogens in cattle. We addressed the ability of P. multocida IROMP-enriched fractions to induce antibody responses in cattle by different vaccination strategies and the protective efficacy of these antibodies using a P. multocida-induced pneumonia challenge model. Vaccination of cattle with outer membrane-enriched fractions derived from Pm232 grown on either iron-depleted (IROMPs) or iron-sufficient (OMPs) conditions induced significant antibody responses; however, the correlation with lung lesion scores was not significant (P = 0.01 and P < 0.07, respectively). SDS-PAGE, Western blots and densitometric analyses of Pm232 grown under iron-deficient conditions revealed five major IROMPs including an immunodominant 96 kDa protein band. Mass spectrometry analysis of the 96kDa protein band suggested homology with the heme acquisition system receptor (HasR) of avian P. multocida (strain Pm70) and was confirmed by DNA sequence analysis of the cloned Pm232 hasR gene. Further analyses indicated that Pm232 HasR is a surface-exposed OMP and conserved among most P. multocida isolates investigated. In addition, cattle vaccinated with live Pm232 or IROMPs had significantly higher antibody responses to the 96 kDa protein band and the correlation with lung lesion scores approached significance (P = 0.056). These results indicate that antibody responses in cattle are induced by P. multocida IROMPs, and that the 96 kDa HasR protein is an immunodominant IROMP.     

Relationships among Pasteurellaceae isolated from free ranging chickens and their animal contacts as determined by quantitative phenotyping, ribotyping and REA-typing

One hundred and forty-three Pasteurella spp. strains and 10 unclassified strains obtained from free ranging poultry, dogs and cats were investigated by extended phenotypic characterization. One hundred and forty-nine of these strains were selected for further studies using ribotyping and REA-typing to evaluate the role of dogs and cats in Pasteurella multocida transmission. Seven and six type strains were included for comparison in phenotyping and genotyping, respectively. Eleven clusters and six unclustered strains were revealed by phenotyping. Ribotyping outlined 12 clusters and six unclustered strains. A correlation between clusters obtained by phenotyping and ribotyping was demonstrated which indicated that a genetic basis exists for clusters outlined by quantitative evaluation of phenotypic data. Similarities and differences in hosts, phenotype, ribotype, and zone of isolation were demonstrated among Pasteurella strains investigated. Isolates of P. multocida from ducks were shown to be clonal by both phenotyping and ribotyping. These strains were identical to one of the chickens strains. REA-typing, however, showed that the chicken strain was different underlining that exchange of clones of P. multocida between avian species rarely happens under village conditions. Management practise in the villages suggest the potential for exchange of P. multocida between poultry and animals kept in contact. The present findings, however, did not indicate that clones of P. multocida are widely exchanged between poultry and other animal species, even though close contact exists. In the present investigation exchange of clones of P. multocida was only demonstrated among animals belonging to the same species. Caution is drawn to the use of ribotyping as the sole method for epidemiological typing and tracing of P. multocida. The present results also underline the importance of proper phenotyping in the identification of P. multocida and related species.     

Characterization of nine Pasteurella multocida isolates from avian cholera outbreaks in Indonesia

Avian cholera outbreaks have been identified in Indonesia in recent years. Despite vaccination programs, outbreaks continue to occur. To date, there has been a lack of information on the characteristics of Pasteurella multocida isolates involved in these outbreaks. Hence, the objective of this study was to characterize Indonesian P. multocida isolates in poultry. During 1998-99, 20 field outbreaks were reported in Indonesia. Nine isolates of P. multocida were recovered from these field outbreaks. The isolates were compared with four vaccine strains that were used in Indonesia and designated PM-V1, PM-V2, PM-V3, and PM-V4. The isolates were characterized by biotype, capsular type, somatic serotype, restriction endonuclease analysis, plasmid presence, and antimicrobial susceptibility patterns. Of the nine Indonesian isolates, three were of capsular type A (A:1,3,13; A:1,3; and A:8). One isolate was of type B:2,3 and one isolate was of capsular type F. For three isolates, the capsular serogroup could not be identified. Plasmids the size of 2.3 kbp were present in three of the field isolates and two of the vaccine strains. One plasmid less than 2 kbp was isolated from the vaccine strain PM-V4. Eight distinct DNA profiles were obtained from digestion with the restriction endonuclease EcoRI, and seven distinct DNA profiles were obtained from digestion with the restriction endonuclease HindIII. All of the isolates were resistant to lincomycin and sulfadiazine and were susceptible to ampicillin and trimethoprim. Of the nine isolates, seven (78%) were susceptible to doxycycline and gentamicin and six (67%) were susceptible to enrofloxacin.     

Characterization of Bordetella bronchiseptica strains using phenotypic and genotypic markers

Thirty-five strains of Bordetella bronchiseptica, recovered primarily from pigs, rabbits, dogs, cats and humans, were characterized by phenotypic and genotypic markers. Biochemical typing only showed variation in the ability to reduce nitrate to nitrite. OMP profiles from virulent strains showed variations in the region of 85-95kDa, which lead us to describe five OMP-types alpha, beta, gamma, delta and varepsilon. Genotypic markers included the presence of IS1001, and polymorphisms in the flagellin gene (flaA) and pertussis toxin (PT) promoter region. The IS1001 was detected in 16 isolates (2 from humans and 10 from pigs) but was absent in rabbit isolates. The restriction profiles of the flaA gene allowed us to differentiate the strains into types A-C. The PT types were characterized by an RFLP assay and could be typed through patterns III-V. There was no apparent association between the flaA or PT types and the origin of the isolates. Eleven groups of isolates were identified on the basis of specific combinations of the analyzed markers. The combination of phenotypic and genotypic tests used could be useful in characterizing isolates and differentiating between certain clonal types of B. bronchiseptica.     

Restriction endonuclease analysis of porcine Pasteurella multocida isolates from Quebec

We have used restriction endonuclease analysis (REA) of genomic DNA to classify porcine Pasteurella multocida isolates with similar capsular and somatic serotypes, and to monitor the distribution of isolates from 12 different herds in Quebec. Within herds, P. multocida isolates of similar capsular and somatic serotypes showed similar REA fingerprints. Between herds, some isolates had similar REA fingerprints. However, differences in REA enabled subtyping of many P. multocida isolates with the same antigen types. Our data indicate that REA would enable accurate epidemiological typing of P. multocida in conjunction with classical capsular and somatic typing.