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[Problems in the use of radioactively marked bacteria in animal experiments. 1. Labeling of Pasteurella multocida, Pasteurella haemolytica and Salmonella dublin with eH, 14C, 32P, 59Fe, 99mTc, 125J1].

Several methods are suggested by which to use the radionuclides 3H, 14C, 32P, 59Fe, 99mTc, and 125J for labelling or doublelabelling of Pasteurella multocida, Pasteurella haemolytica, and Salmonella dublin, with particular reference being made to labelling ofr animal experiments. Suitable radioactive substrates for internal labelling in chemically defined or partially defined nutritive media include 3H-thymin, 3H-thymidine, 14C-glucose, 14C-mannose, 14C-aspartic acid, as well as 3H-uracil, 3H-uridine, 3H-orotic acid, 14C-orotic acid, 59Fe-III-citrate or chloride, and Na2H32PO4. The choise of the nuclide and substrate should by governed by the problem at hand.

Isotope Labeling

Bacteremia due to Pasteurella multocida.

Pasteurella multocida should be considered as a possible etiologic agent in any infection that is the result of an animal bite or scratch. Because of its opportunistic capability, it should be included among the possible pathogens in bacteremia, particularly in any patient with immunosuppression or liver cirrhosis, especially if there is a history of animal exposure.

Adolescent

Siderophore production by Pasteurella multocida.

Pasteurella multocida grown under conditions of iron deprivation secreted into the culture medium a growth-enhancing factor which functioned as a siderophore. The siderophore was found to be neither a phenolate nor a hydroxamate by chemical tests and bioassays and was given the trivial name multocidin. Multocidin was partially purified and found to be a highly polar, nonaromatic, and dialyzable compound. This is the first report demonstrating the production of a siderophore by P. multocida.

Culture Media

Tn7 inserts in both orientations at a single chromosomal location and apparently forms cointegrates in Pasteurella multocida.

Pasteurella multocida transconjugants isolated after mating with Escherichia coli strains that carry one or the other of two Tn7-containing suicide plasmids, pRKTV5 and pUW964 (pRKTV5::Tn5), were analysed. These plasmids have the ColE1 replication origin and were thus expected to deliver transposons but not be maintained as free replicons in Pasteurella. Five out of six transconjugants selected for acquisition of Tn7 from E. coli (pRKTV5) had simple insertions of the transposon, in either orientation, at a single chromosomal location, while the sixth had pRKTV5 integrated at the same location. By contrast, all of 27 transconjugants selected for acquisition of either Tn7 or Tn5 from E. coli (pUW964) maintained pUW964. Of seven subsequently examined at the molecular level, all had pUW964 (in one case, a deletion derivative) integrated at the same location as the Tn7 insertions obtained with pRKTV5. A copy of Tn7 was present at each boundary between the integrated plasmids (pRKTV5 or pUW964) and the chromosome in each strain. The two copies of Tn7 at either end of an integrated plasmid were either in the same (six cases) or in opposite (two cases) orientations with respect to each other. These seem to be products of replicative transposition by Tn7 but can also derive from conservative mechanisms.

Chromosomes, Bacterial

In vivo antigen expression by Pasteurella multocida.

Pasteurella multocida was purified from the blood of turkeys affected with acute fowl cholera, and membrane preparations from those bacteria were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and visualized on immunoblots. Antigens were detected in the membranes of these in vivo-propagated bacteria that were not detected in membrane preparations of the same P. multocida strain grown in vitro. The unique antigens were detected in the detergent-insoluble phase and were enriched to various degrees by different detergents.

Animals

Reduced microbicidal activity of peripheral mononuclear phagocytic cells infected with Pasteurella multocida.

Pasteurella multocida inhibits the uptake and killing of Candida albicans and P. multocida by avian mononuclear phagocytic cells. The toxic outer membrane protein of P. multocida, which has been previously described, also inhibited the uptake and killing of C. albicans. Antibody specific for the toxic outer membrane protein reversed this effect resulting not only in an increase in uptake of C. albicans and P. multocida, but also in intracellular killing of P. multocida. This antibody, however, only partially restored killing of C. albicans. These data support the hypothesis that P. multocida is capable of intracellular survival in avian mononuclear phagocytic cells and that the toxic outer membrane protein is totally or partly responsible for this occurrence.

Animals

Sequence analysis of the potent mitogenic toxin of Pasteurella multocida.

Pasteurella multocida toxin is a potent mitogen for cultured Swiss 3T3 cells where it causes an accumulation of inositol phosphates and activation of protein kinase C. The gene sequence described here coded for a 146 kDa protein. The ORF was preceded by a ribosome binding site and followed by a stem loop. There was no evidence for a signal sequence. The gene had a low G + C base ratio which differs from the rest of the Pasteurella genome. There was no significant homology with other known proteins, although a motif found in certain bacterial toxins which are ADP-ribosyl transferases is present. A recombinant expressing only part of the PMT gene was not mitogenic.

Amino Acid Sequence

Solubilization of membrane-associated cross-protection factor(s) of Pasteurella multocida.

Pasteurella multocida harvested from the blood of turkeys dying of experimental fowl cholera were purified by centrifugation and lysed. The soluble and membrane-associated components of the bacteria were separated by centrifugation. Nonionic (octylglucoside) and zwitterionic (3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate; CHAPS) detergents were tested for their abilities to solubilize the cross-protection factor(s) (CPF) from the membrane-associated component. Protection studies in turkeys showed that optimum solubilization was by 1.0% octylglucoside and 0.5% CHAPS. Antibodies from turkeys made against solubilized membrane-associated CPF passively cross-protected poults against challenge. Ion exchange chromatography of detergent-solubilized CPF resulted in elution of two protein-containing peaks, each of which conferred active immune protection.

Animals

[Influence of Pasteurella multocida and Pasteurella-multocida endotoxin on the blood-coagulation-analytic parameter after experimental administration in calves].

Experimental administration to calves of Pasteurella multocida or the application of an endotoxin preparation of the strain produced continuous decline of thrombocytes, disorders in thrombocytic functions, and the development of hypofibrinogenaemia. The changes recorded as well as the clinical symptoms and pathologico-anatomic findings are interpreted as endotoxin shock.

Afibrinogenemia

Multiple drug resistance in Pasteurella multocida and Pasteurella haemolytica from cattle and swine.

The results of antimicrobial susceptibility testing on 262 strains of Pasteurella multocida and 141 strains of Pasteurella haemolytica isolated from cattle and swine from 1971 to 1974 were analyzed for patterns of resistance to streptomycin, penicillin, tetracycline, and chloramphenicol, using a modified Kirby-Bauer procedure. Resistance was recorded for 80.5% of the isolants of P multocida and 92.2% of those of P haemolytica. Resistance to streptomycin was most frequent, followed by resistance to penicillin and tetracycline. Most cultures of P multocida and P haemolytica were susceptible to chloramphenicol. There were 9 patterns of resistance with the aforementioned antibiotics. The combinations, streptomycin and penicillin and streptomycin and tetracycline, each accounted for approximately 10% of the resistance patterns of P multocida. Approximately half of the 14 isolants of P haemolytica were resistant to the combination of streptomycin, penicillin, and tetracycline. These observations underscore the need for antimicrobial susceptibility testing of clinical isolants of P multocida and P haemolytica.

Animals

[Deoxyribonucleic acids from Pasteurella multocida and Pasteurella haemolytica].

Pasteurella multocida and P. haemolytica strains contain between 1.5 and three per cent phosphorus, between nine and 14 per cent nitrogen, between two and four per cent DNA, and between five and 18 per cent RNA, the precise figures depending on culturing conditions. High-molecular DNA may be isolated by means of bacteriolysis, using deoxycholate or dodecylsulphate and the usual steps of purification, with yield and purity differing by strains. DNA with sufficient purity can be obtained from Sepharose 2 B by gel chromatography. The isolated DNA yields were characterised, base values being between 37 and 38 per cent GC for P. haemolytica and between 41 and 48 per cent GC for P. multocida. Highly suitable precursors to DNA synthesis for tritium labelling are 3H-thymidine, which is incorporated in excess of 3H-thymine by a factor of 255, as well as 3H-uracil, with its activity being recovered also from the pyrimidine bases of DNA via pyrimidine biosynthesis.

Chromatography, Gel

Cleavage of immunoglobulin A1, A2 and G by proteases from clinical isolates of Pasteurella multocida.

Several Pasteurella multocida strains were examined for their ability to produce extracellular enzymes that cleave immunoglobulin A and G (Ig A and Ig G) molecules. Two strains isolated from human pulmonary and genital infections produced proteases that cleaved human IgA and IgG, colostral IgA and human myeloma IgA1 and IgA2. Human IgM was not degraded by these enzymes. Examination of cleavage digests showed two main fragments with different electrophoretic mobilities. The two P. multocida strains produced a protease that cleaved IgA and IgG heavy chains outside the hinge region, and differed in this respect from the hinge-cutting proteases of other bacteria. Protease production may be a virulence mechanism for P. multocida strains.

Colostrum

Isolation, characterization, and antibacterial activity of a novel Pasteurella multocida bacteriophage.

Pasteurella multocida is the main pathogen causing fowl cholera and poses a serious threat to the poultry industry. Current clinical control relies on antibiotics, but the prevalence of drug-resistant strains makes it urgent to develop new antibacterial strategies. In this study, a P. multocida-specific bacteriophage vB_PmuS_ZP41 was isolated and identified as a member of the family Siphoviridae by transmission electron microscopy. This phage showed lytic activity against 13 out of 19 clinical isolates of P. multocida (68.4%) and remained stable at 4-50°C and pH 3-9. The optimal multiplicity of infection was 0.01, with a latent period of 10 min and a burst size of approximately 56 PFU/cell. Whole-genome sequencing revealed that the phage genome is a double-stranded DNA of 38,592 bp, containing no virulence genes or antibiotic resistance genes, indicating good safety. In a chick infection model, phage treatment significantly improved the survival rate of infected chicks from 40% to 80%, significantly reduced bacterial loads in blood, lung, liver, and spleen, and decreased serum levels of TNF-α and IL-1β while alleviating histopathological damage. This study systematically characterized the biological properties of phage vB_PmuS_ZP41 and its antibacterial efficacy both in vitro and in vivo, providing an experimental basis and a candidate strain for the future development of phage therapy against avian pasteurellosis.

Biological characteristics

In vitro antibiotic sensitivity of Pasteurella multocida.

Thirty Pasteurella multocida strains were tested against 19 antibiotics using an agar dilution method. Penicillin G was the single most active agent tested. Anti-staphylococcal penicillins were markedly less inhibitory than penicillin G. Other penicillins including phenoxymethyl penicillin, and tetracyclines, chloramphenicol and cephalosporins all inhibited the organisms in low concentrations. Erythromycin and lincomycins had poor activity against several of the strains. The range of M.I.C.s observed for most antibiotics was narrow but wide variation was seen in zone size when antibiotic disk tests were performed.

Anti-Bacterial Agents

Report of 4 cases of Pasteurella multocida septic arthritis.

Pasteurella multocida is frequently responsible for infections in man due to wounds inflicted by animals (generally cats or dogs). However, the development of septic arthritis is a rare complication. We report 4 cases of Pasteurella multocida septic arthritis with demonstration of the organism in the joint in each case. Two cases presented with monoarthritis (sternoclavicular joint and wrist) and 2 cases had polyarticular involvement from the outset.

Aged

Molecular epidemiology of Pasteurella multocida in turkeys.

Pasteurella multocida isolated from turkeys during an outbreak of fowl cholera was characterized by serotype and heterogeneity of genes encoding rRNA (ribotype) to investigate the epidemiology of the organism. Isolates were collected between October 1985 and July 1986. The M9 or Clemson University fowl cholera vaccine-like strain was detected in 17% of the flocks with fowl cholera. One particular strain, isolated only from breeder flocks, was recovered from 7 of the 10 breeder flocks examined in this study. Intracompany transmission appeared to be common, implying a failure in biosecurity. Circumstantial evidence indicated that in the field; the incubation period of P multocida in a turkey flock may be between 2 to 7 weeks. Wildlife did not appear to be an important reservoir of P multocida for turkeys during this study period. Ribotyping results tended to discount several of the possible interflock transmissions, as suggested by examination of serotyping results alone; however, serotyping in combination with ribotyping proved helpful in understanding the epidemiology of P multocida in turkeys.

Animals

A group of indole-negative bovine strains with high deoxyribonucleic acid homology to Pasteurella multocida.

A group of nine bovine Pasteurella strains not producing indole were investigated for their taxonomic relationships with Pasteurella multocida, Pasteurella haemolytica and Pasteurella canis. For all strains, DNA-DNA hybridization has revealed a high genetic relatedness at the species level to P. multocida and significantly lower homologies of only 18-41% towards P. haemolytica and 11-15% towards P. canis. Guanine plus cytosine values of 38.0 to 42.1 mol% and several phenotypic characters have been found to be different from the established pattern for P. multocida subspecies. It is suggested that the strains represent a new taxon, possibly another P. multocida subspecies.

Animals