Characterisation of Pasteurella dagmatis-like isolates recovered from the feline oral cavity.

Three Pasteurella dagmatis-like strains recovered from the feline oral cavity were analysed by traditional biochemical tests, the Biolog Microstation™ ID System, and 16S rRNA and sodA gene sequence analysis. The molecular biological methods revealed that these strains differ from P. dagmatis, forming a new genomospecies in the genus Pasteurella sensu stricto. Furthermore, sequence analysis and multiple alignments of 16S rRNA and the sodA gene established that the P. pneumotropica NCTC10827 and the P. dagmatis-like strains described here possess high genetic similarity.

Pasteurella multocida sialic acid aldolase: a promising biocatalyst.

Sialic acid aldolases or N-acetylneuraminate lyases (NanAs) catalyze the reversible aldol cleavage of N-acetylneuraminic acid (Neu5Ac) to form pyruvate and N-acetyl-D: -mannosamine (ManNAc). A capillary electrophoresis assay was developed to directly characterize the activities of NanAs in both Neu5Ac cleavage and Neu5Ac synthesis directions. The assay was used to obtain the pH profile and the kinetic data of a NanA cloned from Pasteurella multocida P-1059 (PmNanA) and a previously reported recombinant Escherichia coli K12 NanA (EcNanA). Both enzymes are active in a broad pH range of 6.0-9.0 in both reaction directions and have similar kinetic parameters. Substrates specificity studies showed that 5-O-methyl-ManNAc, a ManNAc derivative, can be used efficiently as a substrate by PmNanA, but not efficiently by EcNanA, for the synthesis of 8-O-methyl Neu5Ac. In addition, PmNanA (250 mg l(-1) culture) has a higher expression level (2.5-fold) than EcNanA (94 mg l(-1) culture). The higher expression level and a broader substrate tolerance make PmNanA a better catalyst than EcNanA for the chemoenzymatic synthesis of sialic acids and their derivatives.

Acceptor specificity of the Pasteurella hyaluronan and chondroitin synthases and production of chimeric glycosaminoglycans.

The hyaluronan (HA) synthase, PmHAS, and the chondroitin synthase, PmCS, from the Gram-negative bacterium Pasteurella multocida polymerize the glycosaminoglycan (GAG) sugar chains HA or chondroitin, respectively. The recombinant Escherichia coli-derived enzymes were shown previously to elongate exogenously supplied oligosaccharides of their cognate GAG (e.g. HA elongated by PmHAS). Here we show that oligosaccharides and polysaccharides of certain noncognate GAGs (including sulfated and iduronic acid-containing forms) are elongated by PmHAS (e.g. chondroitin elongated by PmHAS) or PmCS. Various acceptors were tested in assays where the synthase extended the molecule with either a single monosaccharide or a long chain (approximately 10(2-4) sugars). Certain GAGs were very poor acceptors in comparison to the cognate molecules, but elongated products were detected nonetheless. Overall, these findings suggest that for the interaction between the acceptor and the enzyme (a) the orientation of the hydroxyl at the C-4 position of the hexosamine is not critical, (b) the conformation of C-5 of the hexuronic acid (glucuronic versus iduronic) is not crucial, and (c) additional negative sulfate groups are well tolerated in certain cases, such as on C-6 of the hexosamine, but others, including C-4 sulfates, were not or were poorly tolerated. In vivo, the bacterial enzymes only process unsulfated polymers; thus it is not expected that the PmCS and PmHAS catalysts would exhibit such relative relaxed sugar specificity by acting on a variety of animal-derived sulfated or epimerized GAGs. However, this feature allows the chemoenzymatic synthesis of a variety of chimeric GAG polymers, including mimics of proteoglycan complexes.

Distribution of RTX toxin genes in strains of [Actinobacillus] rossii and [Pasteurella] mairii.

Strains of [Actinobacillus] rossii, [Pasteurella] mairii and [Pasteurella] aerogenes can be isolated from abortion in swine. The RTX toxin Pax has previously been found only in those [P.] aerogenes strains isolated from abortion. Nothing is known about RTX toxins in field isolates of the other two species. To gain insight into the distribution of selected RTX toxin genes and their association with abortion, PCR screening for the pax, apxII and apxIII operons on 21 [A.] rossii and seven [P.] mairii isolates was done. Since species can be phenotypically misidentified, the study was backed up by a phylogenetic analysis of all strains based on 16S rRNA, rpoB and infB genes. The pax gene was detected in all [P.] mairii but not in [A.] rossii strains. No apx genes were found in [P.] mairii but different gene combinations for apx were detected in [A.] rossii strains. Most of these strains were positive for apxIII, either alone or in combination with apxII. Whereas pax was found to be associated to strains from abortion no such indication could be found with apx in [A.] rossii strains. Phylogenetically [A.] rossii strains formed a heterogeneous cluster separated from Actinobacillus sensu stricto. [P.] mairii strains clustered with [P.] aerogenes but forming a separate branch. The fact that [P.] aerogenes, [P.] mairii and [A.] rossii can phylogenetically clearly be identified and might contain distinct RTX toxin genes allows their proper diagnosis and will further help to investigate their role as pathogens.

Genetic relationships among avian isolates classified as Pasteurella haemolytica, 'Actinobacillus salpingitidis' or Pasteurella anatis with proposal of Gallibacterium anatis gen. nov., comb. nov. and description of additional genomospecies within Gallibacterium gen. nov.

Bacteria of the avian [Pasteurella haemolytica]-'Actinobacillus salpingitidis' complex have been associated with different pathological conditions in birds, among which salpingitis and peritonitis in chickens of layer type seem to dominate. The aim of this study was to classify these bacteria by comparison of 37 strains tentatively classified as biovars of the avian [P. haemolytica]-'A. salpingitidis' complex or as Pasteurella anatis. PFGE, AFLP and plasmid profiling showed that strains representing different biovars were genotypically different. Phylogenetic analysis of 22 strains characterized by 16S rRNA gene sequence comparison showed that strains classified as biovars 5, 8 and 9 were closely related to the suggested type strain of 'A. salpingitidis' (98.4-99.9% similarity), whereas the remaining strains classified in 12 biovars or as P. anatis were closely related to the type strain of P. anatis (98.1-100% similarity). The two groups were related at 95.7-97.1% similarity. The closest similarity outside this group was 94.6%, between biovar 15 and Bisgaard taxon 3. DNA-DNA hybridization was performed with 34 strains and showed binding above 85% for strains of biovars 5 and 8, including the suggested type strain of 'A. salpingitidis'. Two strains of P. anatis (F 149T and F 279) were closely related at 79% DNA binding to 27 strains of biovars 1,3, 4, 11, 12, 17-20, 22 and 24. A new genus, Gallibacterium gen. nov., is proposed to include the avian [P. haemolytica]-'A. salpingitidis'-P. anatis complex, since these taxa form a monophyletic unit with similarities above 95% on the basis of 16S rRNA sequence comparison and they are unrelated to other genera of the family Pasteurellaceae Pohl 1981. The new genus consists of Gram-negative, non-motile, rod-shaped or pleomorphic bacteria. The bacteria are catalase-, oxidase- and phosphatase-positive. Nitrate is reduced and acid is produced without gas formation from glycerol, (-)D-ribose, (+)D-xylose, (-)D-mannitol, (-)D-fructose, (+)D-galactose, (+)D-glucose, (+)D-mannose, sucrose and raffinose. The genus Gallibacterium can be separated from other genera of Pasteurellaceae by differences in catalass, symbiotic growth, haemolysis, urease, indole, acid production from (+)D-xylose, (-)D-mannitol, (-)D-sorbitol, (+)D-mannose, maltose, raffinose and dextrin and ONPG and PNPG tests. Pasteurella anatis Mutters et al. 1985 is transferred to the new genus as Gallibacterium anatis gen. nov., comb. nov. Genomospecies 1 of Gallibacterium is proposed to include the former biovars 5 and 8 of the avian [P. haemolytica]-'A. salpingitidis' complex. The type strain of Gallibacterium anatis is F 149T (=ATCC 43329T = NCTC 11413T) and the reference strain of Gallibacterium genomospecies 1 is CCM 5974.

Sharing of Pasteurella spp. between free-ranging bighorn sheep and feral goats.

Pasteurella spp. were isolated from feral goats and free-ranging bighorn sheep (Ovis canadensis canadensis) in the Hells Canyon National Recreation Area bordering Idaho, Oregon, and Washington (USA). Biovariant 1 Pasteurella haemolytica organisms were isolated from one goat and one of two bighorn sheep found in close association. Both isolates produced leukotoxin and had identical electrophoretic patterns of DNA fragments following cutting with restriction endonuclease HaeIII. Similarly Pasteurella multocida multocida a isolates cultured from the goat and one of the bighorn sheep had D type capsules, serotype 4 somatic antigens, produced dermonecrotoxin and had identical HaeIII electrophoretic profiles. A biovariant U(beta) P.haemolytica strain isolated from two other feral goats, not known to have been closely associated with bighorn sheep, did not produce leukotoxin but had biochemical utilization and HaeIII electrophoretic profiles identical to those of isolates from bighorn sheep. It was concluded that identical Pasteurella strains were shared by the goats and bighorn sheep. Although the direction of transmission could not be established, evidence suggests transmission of strains from goats to bighorn sheep. Goats may serve as a reservoir of Pasteurella strains that may be virulent in bighorn sheep; therefore, goats in bighorn sheep habitat should be managed to prevent contact with bighorn sheep. Bighorn sheep which have nose-to-nose contact with goats should be removed from the habitat.

Mannheimia haemolytica serotype 1 and Pasteurella trehalosi serotype 10 culture supernatants contain fibrinogen-binding proteins.

Fibrinogen-binding proteins were found in the culture supernatants of Mannheimia haemolytica serotype 1 (ATCC 43270) and Pasteurella trehalosi serotype 10 (ECO-100). Sheep fibrinogen was biotinylated and shown to bind to proteins in the culture supernatants by modified western blot. Fibrinogen-binding proteins in the culture supernatant may be important virulence factors leading to the characteristic fibrinous pneumonia caused by these organisms and may be critical antigenic targets for immune prophylaxis.

Studies on time-kill kinetics of different classes of antibiotics against veterinary pathogenic bacteria including Pasteurella, Actinobacillus and Escherichia coli.

A systematic analysis of the bacteriostatic/bactericidal effect of several antibiotics used in veterinary medicine was carried out by time-kill kinetic analysis using P. haemolytica, P. multocida, A. pleuropneumoniae, and E. coli. The antibiotics tested were enrofloxacin, danofloxacin, erythromycin, tilmicosin, penicillin G, ceftiofur and tetracycline. Unexpectedly, the antibiotics well characterized as bacteriostatic agents against human pathogens such as tetracycline and macrolides, showed bactericidal activity against P. haemolytica and A. pleuropneumoniae. In contrast, tetracycline and erythromycin were bacteriostatic and tilmicosin was bactericidal against P. multocida. In addition, P. multocida was killed by fluoroquinolones at a slower rate than the other bacteria. Spectrum analysis revealed that ceftiofur and tilmicosin were good substrates of the universal efflux pump, AcrA/B, but penicillin and tetracycline were not. The fluoroquinolones were modest substrates for AcrA/B.

Degradation of fluoranthene by Pasteurella sp. IFA and Mycobacterium sp. PYR-1:isolation and identification of metabolites.

The findings from a biodegradability study of fluoranthene using two pure bacterial strains, Pasteurella sp. IFA (B-2) and Mycobacterium sp. PYR-1 (AM) are reported. Of total fluoranthene, 24% (B-2) and 46% (AM) was biodegraded in an aqueous medium during 14 d of incubation at room temperature. During this period the bacteria were capable of mineralizing approximately two-thirds (B-2) and four-fifths (AM) of biodegraded fluoranthene to CO2, while one-third (B-2) and one-fifth (AM) of the original fluoranthene remained as stable metabolic products. These metabolites were isolated using liquid-liquid extraction and identified using gas chromatography-mass spectrometry (GC-MS) and derivatization techniques. Two metabolites (9-fluorenone-1-carboxylic acid and 9-fluorenone) were identified by GC-MS directly, while the metabolites 9-fluorenone-1-carboxylic acid, 9-hydroxyfluorene, 9-hydroxy-1-fluorene-carboxylic acid, 2-carboxybenzaldehyde, benzoic acid and phenylacetic acid were determined in their derivatized forms. From the identified metabolites, a fluoranthene biodegradation pathway was proposed for Pasteurella sp. IFA.

Further studies of the relationships among strains classified as taxon 15, taxon 18, taxon 20, (Pasteurella) granulomatis or the (Pasteurella) haemolytica-complex in ruminants using quantitative evaluation of phenotypic data.

Ninety-three trehalose-negative (P.) haemolytica-like strains of ruminant, porcine and leprine origin were investigated. A quantitative evaluation of phenotypic tests was used and the results obtained were compared with those from 246 previously investigated ruminant strains. Cluster analysis of the results obtained displayed most of the taxa as distinct groups which could be related to differences in key characters. Although only minor phenotypic differences were observed between the taxa investigated and the taxa were internally heterogeneous for many of the tests, it was possible to identify characters separating most groups. However, in three instances, taxa isolated from different species could not be separated by any of the tests used or by quantitative evaluation of all 79 tests--the only difference being the species of animals from which they had been isolated. Taxa which could not be separated by phenotypic tests included the ruminant biogroup 6 of (P.) haemolytica and the porcine taxon 15/biovar 1, the ruminant biogroup 7 of (P.) haemolytica and the porcine taxon 15/biovar 2, and ruminant biogroup 31 of (P.) haemolytica and the leprine taxon 20/biovar 1.

Biodegradation studies of polyaromatic hydrocarbons in aqueous media.

Sixteen bacterial strains isolated from an activated sludge and Mycobacterium ssp. PYR-1 were tested for their ability to degrade polyaromatic hydrocarbons (PAHs). The bacterial strains Pasteurella ssp. (B-2) and Mycobacterium ssp. PYR-1 (AM) showed a high biodegradation potential of three- and four-ring PAHs. Bacterial strain AM was able to degrade up to 80% of three and four-ring PAHs (phenanthrene, fluoranthene and pyrene) within the first month of incubation, while the bacterial strain B-2 achieved the same biodegradation in 2 months. The metabolic pathway of PAH degradation was studied using fluoranthene and the bacterial strain AM. Ninety per cent of fluoranthene was biodegraded within the first 9 d of incubation when applied as a single substrate. Retention factor values from thin-layer chromatography studies, gas chromatography with mass selective detection and tandem mass spectrometry identified 9-fluorenone-1-carboxylic acid as one of the stable metabolic products and from this a fluoranthene biodegradation pathway is proposed.

Species selectivity of new siderophore-drug conjugates that use specific iron uptake for entry into bacteria.

Siderophores selectively bind ferric iron and are involved in receptor-specific iron transport into bacteria. Several types of siderophores were synthesized, and growth-promoting or inhibitory activities when they were conjugated to carbacephalosporin, erythromycylamine, or nalidixic acid were investigated. Overall, 11 types of siderophores and 21 drug conjugates were tested against seven different bacterial species: Escherichia coli, Bordetella bronchiseptica, Pasteurella multocida, Pasteurella haemolytica, Streptococcus suis, Staphylococcus aureus, and Staphylococcus epidermidis. In some species, the inhibitory activities of the drug conjugates were associated with the ability of the bacteria to use the siderophore portion of the molecules for growth promotion in disc diffusion tests (0.04 mumol of conjugate or siderophore per disc). E. coli used catechol-based siderophore portions as well as hydroxamate-based tri-delta-OH-N-OH-delta-N-acetyl-L-ornithine ferric iron ligands for growth under iron-restricted conditions achieved by supplemental ethylenediamine di (O-hydroxyphenylacetic acid) (100 micrograms/ml) and was sensitive to carbacephalosporin conjugated to these siderophore types (up to a 34-mm-diameter inhibition zone). B. bronchiseptica used desferrioxamine B and an isocyanurate-based or trihydroxamate in addition to catechol-based siderophore portions for promotion but was not inhibited by beta-lactam conjugates partly because of the presence of beta-lactamase. P. multocida and P. haemolytica did not use any of the synthetic siderophores for growth promotion, and the inhibitory activities of some conjugates seemed partly linked to their ability to withhold iron from these bacteria, since individual siderophore portions showed some antibacterial effects. Individual siderophores did not promote S. suis growth in restrictive conditions, but the type of ferric iron ligands attached to beta-lactams affected inhibitory activities. The antibacterial activities of the intracellular-acting agents erythromycylamine and nalidixic acid were reduced or lost, even against S. aureus and S. epidermidis, when the agents were conjugated to siderophores. Conjugate-resistant E. coli mutants showed the absence of some iron-regulated outer membrane proteins in gel electrophoresis profiles and in specific phage or colicin sensitivity tests, implying that the drugs used outer membrane receptors of ferric complexes to get into cells.

Intra-specific diversity within Pasteurella trehalosi based on variation of capsular polysaccharide, lipopolysaccharide and outer-membrane proteins.

Intra-specific diversity within Pasteurella trehalosi was investigated by analysis of variation of capsular polysaccharide, and lipopolysaccharide (LPS) and outer-membrane protein (OMP) profiles. Sixty isolates of P. trehalosi, from diverse geographical locations within the UK, were examined. Capsular polysaccharide serotypes were determined by indirect haemagglutination assay; LPS and OMP profiles were compared by SDS-PAGE analysis. Capsular serotyping identified three isolates of serotype T3, 18 isolates each of serotypes T4, T10 and T15, and three untypable (UT) isolates. Analysis of LPS and OMP profiles identified six smooth LPS types and four OMP types among the 60 isolates. Forty-five (75%) of the isolates belonged to a single OMP type whereas 52 (87%) of the isolates possessed one of three LPS types. Each typing method, by itself, was not very discriminating but when the data from the three methods were combined, the 60 isolates could be separated into 14 distinct subgroups containing from one to 16 isolates as follows: serotype T3, two subgroups; serotype T4, four subgroups; serotype T10, two subgroups; serotype T15, five subgroups; UT isolates, one subgroup. Certain subgroups were associated with only one serotype whereas other subgroups were common to two or more serotypes. The subgroupings were capable of differentiating between isolates of the same serotype from the same and different geographical origins. Based on their LPS and OMP profiles, isolates of serotypes T4 and T15 were more closely related to each other than to isolates of serotype T10; serotype T4 and T15 isolates were also more heterogeneous than those of serotype T10. Certain isolates of serotype T10, recovered from a wide geographical area, were characterized by the possession of a unique capsule/LPS/OMP combination and represented a single clonal group which was responsible for a large proportion (31%) of recent disease outbreaks. Overall, a combination of capsular serotyping, and LPS and OMP typing, was found to be extremely useful for assessing diversity within P. trehalosi and should be of value for epidemiological and virulence studies.

Small-subunit rRNA sequences and whole DNA relatedness concur for the reassignment of Pasteurella piscicida (Snieszko et al.) Janssen and Surgalla to the genus Photobacterium as Photobacterium damsela subsp. piscicida comb. nov.

The taxonomic status of Pasteurella piscicida (strain NCIMB 2058T [T = type strain] and a strain isolated from the environment) was investigated by performing phylogenetic analyses of small-subunit rRNA sequences, DNA-DNA hybridization analyses, and biochemical characterization analyses. The results of the phylogenetic analyses and the levels of DNA-DNA complementarity demonstrated conclusively that Pasteurella piscicida is extremely closely related to Photobacterium damsela ATCC 33539T. Since the two taxa exhibited a level of DNA-DNA relatedness of 80%, they are members of the same species. The high level of DNA relatedness and the presence of specific morphological and biochemical characteristics support the hypothesis that two subspecies should be recognized. On the basis of its phylogenetic position, we concluded that Pasteurella piscicida should be renamed Photobacterium damsela subsp. piscicida comb. nov.

Accumulation of selenium in a model freshwater microbial food web.

The transfer of selenium between bacteria and the ciliated protozoan, Paramecium putrinum, was examined in laboratory cultures. The population growth of the ciliate was not inhibited in the presence of the highest concentrations of dissolved selenite or selenate tested (10(3) micrograms liter-1). Experiments with radioactive 75selenite or 75selenate indicated that accumulation of selenium by ciliates through time was low when feeding and metabolism were reduced by incubating at 0 degrees C. However, selenium accumulated in ciliate biomass during incubation with dissolved 75Se and bacteria at 24 degrees C and also when bacteria prelabeled with 75Se were offered as food in the absence of dissolved selenium. When 75Se-labeled bacterial food was diluted by the addition of nonradioactive bacteria, the amount of selenite and selenate in ciliates decreased over time, indicating depuration by the ciliates. In longer-term (> 5-day) fed-batch incubations with 75selenite-labeled bacteria, the selenium concentration in ciliates equilibrated at approximately 1.4 micrograms of Se g (dry weight)-1. The selenium content of ciliates was similar to that of their bacterial food on a dry-weight basis. These data indicate that selenium uptake by this ciliate occurred primarily during feeding and that biomagnification of selenium did not occur in this simple food chain.

Iron uptake by Pasteurella piscicida and its role in pathogenicity for fish.

We evaluated the iron uptake mechanisms in Pasteurella piscicida strains as well as the effect of iron overload on the virulence of these strains for fish. With this aim, the capacity of the strains to obtain iron from transferrin and heme compounds as well as their ability to overcome the inhibitory activity of fish serum was analyzed. All the P. piscicida strains grew in the presence of the iron chelator ethylene-diamine-di (O-hydroxyphenyl acetic acid) or of human transferrin, which was used by a siderophore-mediated mechanism. The chemical tests and cross-feeding assays showed that P. piscicida produced a siderophore which was neither a phenolate nor a hydroxamate. Cross-feeding assays as well as preliminary chromatographic analysis suggest that this siderophore may be chemically related to multocidin. All the P. piscicida isolates utilized hemin and hemoglobin as an iron source, since the virulence of the strains increased when the fish were preinoculated with these compounds. This effect was stronger in the avirulent strains (50% lethal dose was reduced by 4 logs when fish were pretreated with hemin or hemoglobin). Only the pathogenic P. piscicida isolates were resistant to the bactericidal action of the fresh fish serum. The nonpathogenic strains grew in fish serum only when it was heat-inactivated or when it was supplemented with ferric ammonium citrate, hemin, or hemoglobin. In all the strains, at least three iron-regulated outer membrane proteins (IROMPs) (105, 118, and 145 kDa) were increased when the strains were cultured in iron-restricted medium.(ABSTRACT TRUNCATED AT 250 WORDS)

Phenotypical characters and ribotyping of Pasteurella aerogenes from different sources.

On the occasion of five Danish human Pasteurella aerogenes from pig bite lesions, a comparison was made between 6 isolates from man and 15 animal isolates, mainly from pigs. The strains originated from 6 different countries (USA, Canada, Czechoslovakia, France, Belgium and Denmark). The 21 isolates were characterized by conventional biochemical tests, antibiogram and the API 20 NE kit; finally ribotyping was carried out by hybridizing EcoRI-digested chromosomal DNA with a probe derived from E. coli ribosomal RNA. By ribotyping, 19 of the 21 strains clustered at a similarity level of 81% or more; both phenotypical tests and ribotyping indicated that the remaining two strains did not belong to the species P. aerogenes. In conclusion, despite minor differences our P. aerogenes isolates constituted a well-defined group and they could not be subdivided on basis of animal or geographical origin.

Characterization and distribution of Pasteurella species recovered from infected humans.

During a 3-year period, all Pasteurella strains recovered at the Clinical Microbiological Laboratory, Lund, Sweden, were studied biochemically with respect to their relationship to the recently described taxa of this genus. Of 159 strains recovered from 146 infected humans, 95 were identified as Pasteurella multocida subsp. multocida, 21 as Pasteurella multocida subsp. septica, 28 as Pasteurella canis, 10 as Pasteurella stomatis, and 5 as Pasteurella dagmatis. The homology within and between the Pasteurella species regarding cellular fatty acids and enzymatic activities was also studied. Strains of the different Pasteurella species were indistinguishable from each other regarding fatty acid composition; all strains contained major amounts of C14:0, C16:1, C16:0, and 3-OH-C14:0 acids and minor amounts of C18:2, C18:1, and C18:0 acids. Neither did the enzymatic activities distinguish between strains belonging to different species. In addition, of 56 strains examined, toxin production was demonstrated only in 1 strain each of P. multocida subsp. multocida and P. canis. Except for one severe case of necrotizing cellulitis involving P. dagmatis, P. multocida subsp. multocida or P. multocida subsp. septica was recovered in the more serious cases of infection. Except for P. canis, which in all cases was associated with dog bites, most Pasteurella strains were recovered in cases of infection associated with cat bites or scratches. Pasteurella strains occurred in four infected patients without evident connections with animals.

Distribution of indole-producing urease-negative pasteurellas in animals.

Three hundred fifty-six animal isolates of indole-positive urease-negative cultures of Pasteurella, which would formerly have been classified as P. multocida, were examined with respect to their relationship to the recently described P. multocida subspecies (ssp.) multocida, septica, and gallicida and P. canis, P. stomatis/Taxon 16, and Pasteurella sp. B. Two hundred sixty-three (73.9%) of the cultures could be identified with one of these taxa, and 93 isolates (26.1%), representing 17 different biotypes, were unassignable. Pasteurella multocida ssp. multocida was the predominant taxon throughout and in most of the 25 animal species from which isolations were made. In dogs, P. canis was the most frequent. Different degrees of host predilection were observed also in P. multocida ssp. septica for cats, P. canis for sheep, and 2 of the unassignable biotypes for cattle and dogs, respectively. Overall, the respiratory tract was the most frequent source of isolates, but a propensity of P. multocida ssp. septica for localization in the central nervous system of cats was noted.

Detection and enumeration of toxin-producing Pasteurella multocida with a colony-blot assay.

Colonies of toxin-producing Pasteurella multocida were detected with peroxidase-labeled monoclonal antibodies by a membrane assay. Examination of the specificity of the assay with 29 P. multocida cultures representing various geographic origins, hosts, and serotypes indicated that the test was specific for toxin-producing strains. No cross-reactions were observed with Bordetella species that can be associated with P. multocida in producing diseases in animals. A single membrane could be used to assay several isolated strains for toxin production or to enumerate toxin-producing colonies in mixed cultures. Toxin-producing P. multocida colonies were detected in primary cultures; hence, the assay appears to have good potential for widespread application with clinical samples.