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Biomedical subjects

W Donachie

Publications and source records attributed to W Donachie.

At least 37 records · Page 2Linked to original sources
PubMed1996

Moredun Bordetella Medium, an improved selective medium for isolation of Bordetella parapertussis.

Bordetella parapertussis, previously thought to be an obligate human respiratory tract pathogen, has been isolated from sheep. Attempts to assess the prevalence of B. parapertussis in conventionally reared sheep by nasal swabbing proved futile with existing selective media because of extensive overgrowth with Mucor spp. and other nasal commensals. Moredun Bordetella Medium (MBM), which contains cycloheximide and spectinomycin at final concentrations of 0.5 mg/ml and 100 mu g/ml, respectively, was developed as an improved selective medium to isolate B. parapertussis from the nasal cavities of conventionally reared sheep. The selective ability of MBM was evaluated with 200 nasal swabs from conventionally reared sheep, and B. parapertussis was recovered from 31.5% of the samples. MBM facilitated the simple and effective isolation of B. parapertussis from ovine nasal swabs and, in successfully excluding overgrowth with other contaminants, proved superior to other test formulations evaluated and to existing conventional media.

Bordetella↗
PubMed1995

Characterisation of ovine Bordetella parapertussis isolates by analysis of specific endotoxin (lipopolysaccharide) epitopes, filamentous haemagglutinin production, cellular fatty acid composition and antibiotic sensitivity.

Isolates of Bordetella parapertussis, recovered from sheep or man, were characterised by reaction with specific anti-Bordetella lipopolysaccharide monoclonal antibodies, production of filamentous haemagglutinin, fatty acid patterns, and antibiotic sensitivity. Generally, the isolates lay within one of four groups, with separation of the ovine isolates into two groups. Reactions with specific monoclonal antibodies against lipopolysaccharide separated the ovine isolates into these two groupings. Analysis of the cellular fatty acid compositions by cluster analysis differentiated between the human and the ovine strains and also showed variation within the ovine isolates. When the production of filamentous haemagglutinin was analysed in an ELISA system, a similar pattern emerged. Varying concentrations of filamentous haemagglutinin (11-429 ng (mg total protein)-1) were extracted from the human isolates and the one group of ovine isolates with no significant protein detected in the other ovine group. These studies demonstrate variation between and within B. parapertussis isolates recovered from two mammalian sources.

Adhesins, Bacterial↗
PubMed1995

Analysis of the expression and secretion of isotypes of sheep B cell immunoglobulins with a panel of isotype-specific monoclonal antibodies.

Monoclonal antibodies to sheep light chain, IgM and IgG were produced and used to assess total immunoglobulin (Ig) synthesis by sheep B cells in culture and antibodies to specific antigens. By using these antibodies in a dual fluorescence-activated cell sorting analysis of sheep efferent lymph B lymphocytes the percentage change in surface Ig isotype of B lymphoblasts from IgM to IgG after the antigenic stimulation of the local lymph node was measured. An extension of this analysis to paired blood and afferent or efferent lymph B cells made it possible to investigate the recirculation characteristics of B cells expressing different Ig isotypes.

Animals↗
PubMed1995

A native plasmid of Pasteurella haemolytica serotype A1: DNA sequence analysis and investigation of its potential as a vector.

The complete nucleotide sequence of a 4.3 kilobase pair plasmid, pAB2, isolated from a bovine strain of Pasteurella haemolytica serotype A1, was determined. It encodes a Rob-1 type beta-lactamase and a region with homology to the mobilisation (mob) region of the Escherichia coli plasmid, ColE1. An insertion mutant of pAB2 (pTC2/81) carrying a copy of Tn5 was transferred to E coli K12 by conjugation. Subsequently pTC2/81 could be transferred by transformation to E coli HB101, but not to P haemolytica serotypes A1 or A2. However, a derivative of this construct containing only a fragment of the Tn5 insertion sequence was able to transform P haemolytica. A further construct containing a fragment of the P haemolytica A1 leucotoxin A gene, was similarly restricted to transforming E coli. These results demonstrate that the pAB2 plasmid is capable of acting as an E coli/P haemolytica shuttle vector. However, the nature of the cloned DNA sequences are important to transformation.

Base Sequence↗
PubMed1995

Bronchopneumonia in mice caused by Pasteurella haemolytica A2 after predisposition by ovine Bordetella parapertussis.

Initial intranasal inoculation of four to eight-week-old Swiss White mice with 7.5 x 10(6) colony forming units (cfu) of ovine B. parapertussis followed 30 min, three or five days, by intranasal inoculation with 1.4 x 10(5) cfu of Pasteurella haemolytica A2 resulted in a more severe infection pattern than when either agent was administered alone. Histopathological examination showed that inoculation with B. parapertussis alone caused a bronchopneumonia the severity of which was dependant upon the infecting dose. Bacteria were recovered up to 10 days after inoculation. P. haemolytica alone had no apparant pathogenic effect and was cleared from the lungs within 24 h. When both agents were given in combination the lesions were most severe when P. haemolytica was administered three days after B. parapertussis infection. These findings suggest that B. parapertussis predisposes mice to subsequent infection with P. haemolytica and that the timing of the P. haemolytica administration influences the severity of the lung lesions.

Animals↗
PubMed1995

Predisposition of specific pathogen-free lambs to Pasteurella haemolytica pneumonia by Bordetella parapertussis infection.

Three groups of specific pathogen-free (SPF) lambs were inoculated intratracheally with an ovine isolate of Bordetella parapertussis (5.5 x 10(9) colony-forming units) or with B. parapertussis followed 2 or 5 days later with Pasteurella haemolytica serotype A2 (120-180 million colony-forming units). When P. haemolytica A2 was administered 2 days after infection with B. parapertussis all lambs became febrile for at least 72 h. At necropsy their lungs were discoloured, congested and showed large areas of collapse and consolidation which, in one case, covered the entire lung. Histopathological examination confirmed that the combined infection produced a severe acute bronchopneumonia in four of seven lambs. B. parapertussis and P. haemolytica were recovered from all of the lambs in this group. Seven lambs challenged with P. haemolytica 5 days after B. parapertussis and six lambs infected with B. parapertussis alone showed no clinical signs of disease other than mild pyrexia and only mild histopathological changes. B. parapertussis, but not P. haemolytica, was recovered from these lambs. The findings indicated that B. parapertussis predisposed the SPF lambs to P. haemolytica pneumonia. This effect appeared to be dependent upon the time interval between the administration of the two agents.

Animals↗
PubMed1995

Phenotypic characterization of the cells of the inflammatory response in ovine encephalitic listeriosis.

The brainstem (pons cerebri and medulla oblongata) of 22 sheep aged between 6 months and 3 years which had developed clinical signs of central nervous system dysfunction were examined. Histopathological changes characterized by microabscesses, focal gliosis and perivascular cuffing compatible with natural infection with Listeria monocytogenes were present. The brains were examined by lectin histochemistry and immunohistochemistry with markers for T lymphocytes (CD4+ and CD8+ subsets), B lymphocytes, mononuclear phagocytes (including macrophages, ramified microglia, activated microglia and amoeboid microglia), astroglia and L. monocytogenes. These methods allowed semiquantitative analyses of the frequency of the different cell types in the brain lesions. The distribution of listerial antigen in the lesions was variable but always sparse. Mononuclear phagocytes and neutrophils appeared to be the most numerous inflammatory cells in the affected areas of the brainstem. T lymphocytes (CD8+ and CD4+ subsets) and B lymphocytes also played a part in the inflammatory process, in addition to activated astrocytes.

Abscess↗
PubMed1995

The incidence, numbers and types of Listeria monocytogenes isolated from farm bulk tank milks.

Bulk tank milk from 160 producers was tested for Listeria monocytogenes at three monthly intervals over 1 year. Twenty-five producers were positive, most on a single occasion, only seven were positive on three or more of the four samplings. Listeria monocytogenes numbers were low, usually < 1 ml-1, the highest was 35 ml-1. All isolates were serotype 1, the use of multilocus enzyme electrophoresis on representative isolates gave nine different electrophoretic types, two have been associated with listeriosis in humans or animals, a further two had only been isolated from one other source (silage or faeces), while the majority (5) were unique to milk.

Animals↗
PubMed1994

Isolation and characterization of Bordetella parapertussis-like bacteria from ovine lungs.

Bacteria resembling two Bordetella species were isolated from both normal and pneumonic ovine lungs using a selective charcoal agar. Twenty-eight of the 33 isolates showed similarities to stock NCTC B. parapertussis strains in their SDS-PAGE gel protein profiles, in their biochemical reactions and in causing browning on tyrosine agar. Five isolates behaved similarly to stock B. bronchiseptica strains, in being actively motile, in giving identical positive reactions in three out of four biochemical tests and in causing no colour change in tyrosine agar. Multilocus enzyme electrophoresis separated the isolates into two electrophoretic types distinguishable from those of stock B. parapertussis and stock B. bronchiseptica strains.

Animals↗
PubMed1993

Characterization by Multilocus Enzyme Electrophoresis of Listeria monocytogenes Isolates Involved in Ovine Listeriosis Outbreaks in Scotland from 1989 to 1991.

Initial results from a study of five small ovine listeriosis outbreaks in Scotland in 1989 to 1991 are presented. Forty-eight isolates including three from silage were typed at 10 polymorphic enzyme loci by using multilocus enzyme electrophoresis resulting in the identification of 12 electrophoretic types. Phylogenetic analysis partitioned the 12 electrophoretic types into two statistically distinct divisions distinguishing 1/2a serotypes from non-1/2a serotypes.

Journal Article↗
PubMed1992

Characterisation and biological activity of monoclonal antibodies specific for Pasteurella haemolytica A1 capsule and lipopolysaccharide.

Monoclonal antibodies (mAb) against both Pasteurella haemolytica A1 capsule and lipopolysaccharide (LPS) were produced. Anti-capsule mAb reacted with the homologous A1 serotype only, whereas mAb against LPS reacted with P. haemolytica serotypes A2, A5, A8, A12, A14 and A16 but not with 33 bacterial species or rough LPS mutant strains tested. Both capsule and LPS antigens were visualised on the surface of bacteria by immunogold electron microscopy. Neither of the mAbs demonstrated antibody-dependent complement-mediated killing in vitro but both facilitated phagocytosis in vitro.

Animals↗
PubMed1992

Pyrolysis mass spectrometry of Listeria monocytogenes isolates from sheep.

Forty-eight isolates of Listeria monocytogenes from sheep and silage, involved in five small outbreaks of listeriosis, were compared by pyrolysis mass spectrometry (PyMS). The method clustered isolates from single animals, and showed that epidemiologically associated isolates were closely related to each other. PyMS is a simple technique capable of analysing large numbers of samples daily, and its application in veterinary studies should help to elucidate the epidemiology of listeriosis.

Animals↗
PubMed1992

Antigenic analysis of iron-regulated proteins in Pasteurella haemolytica A and T biotypes by immunoblotting reveals biotype-specific epitopes.

The antigenic relationships of the iron-regulated proteins (IRPs) in Pasteurella haemolytica A and T biotype strains were examined by SDS-PAGE and immunoblotting. P. haemolytica cells of the A biotype, grown under conditions of iron-limitation, expressed two IRPs, of 35 and 70 kDa. All T biotype strains expressed IRPs with slightly different molecular masses of 37 and 78 kDa. Immunoblotting of all 16 P. haemolytica serotypes was carried out using a panel of polyclonal and monoclonal antibodies raised against serotype A2 antigens. Polyclonal antibodies revealed inter-serotype cross-reactivity towards the 35 and 70 kDa IRPs within the A biotype but no cross-reactivity against a T biotype protein in the 78 kDa region. Monoclonal antibody against the 35 kDa antigen reacted only with the A biotype 35 kDa IRP. Identical profiles were obtained for 10 field isolates of serotype A2, further emphasizing the antigen conservation within the A biotype. These findings reinforce the view that the A and T biotypes of P. haemolytica should be considered as separate species and suggest that IRPs from single A and T biotype strains incorporated into a vaccine might provide cross-protection against all P. haemolytica serotypable strains. Similar studies on the IRPs of 10 untypable strains revealed some of these to have different antigenic reactivities from those observed within the A and T biotypes.

Antibodies, Bacterial↗
PubMed1992

Purification of listeriolysin O and development of an immunoassay for diagnosis of listeric infections in sheep.

A protein of 58,000-Da molecular mass was purified from the supernatant fluid of a dialysis sac culture of Listeria monocytogenes by cation-exchange chromatography. The purified protein, homogeneous by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and possessing the characteristics of listeriolysin O (LLO), was used to develop an indirect enzyme-linked immunosorbent assay. Anti-LLO antibodies were shown to be consistently produced in sheep after experimental challenge with L. monocytogenes serovar 4b. The assay also successfully detected and measured specific anti-LLO antibodies in the sera of silage-fed sheep among which listeric enteritis and abortions had occurred.

Animals↗
PubMed1991

Clinical and serum antibody responses to lambs to infection by Listeria monocytogenes.

Oral dosing of lambs with 1 x 10(10) colony forming units of Listeria monocytogenes daily for three days produced no clinical signs but protected the animals against bacteraemia following subsequent homologous subcutaneous challenge. Following the subcutaneous injections, comparison with controls revealed significantly lower rectal temperatures and a significant difference in positive blood cultures. In both groups signs of systemic illness were unremarkable. However, two and 10 days after the subcutaneous challenges neurological signs developed in two lambs. L monocytogenes was isolated from the brain of one lamb and histopathological lesions of listeric encephalomyelitis were demonstrated in both. After oral infection antibodies to L monocytogenes whole cell antigen were detectable in serum agglutination tests and by ELISA. Serological responses to flagellin were examined by ELISA and to listeriolysin O by immunoblotting. The responses of the animals to flagellin were weak and inconsistent, but antibodies to listeriolysin O were detectable after both oral and subcutaneous challenge. The subclass of antibody involved in this response was shown to be predominantly IgG1.

Agglutination Tests↗
PubMed1991

Vaccine containing iron-regulated proteins of Pasteurella haemolytica A2 enhances protection against experimental pasteurellosis in lambs.

A vaccine containing sodium salicylate extract (SSE) of Pasteurella haemolytica A2 cells grown in a medium chemically depleted of available iron by the addition of alpha alpha dipyridyl to induce iron-regulated proteins (IRPs) conferred protection to specific pathogen-free (SPF) lambs exposed to an aerosol of P. haemolytica A2. The disease score in these lambs was significantly lower (p less than 0.005) than those in unvaccinated lambs or in lambs immunized with SSE prepared from cells grown in iron-replete medium. Immunoblotting of sera from these SPF lambs against whole cell antigens of P. haemolytica A2 grown under iron-restricted conditions demonstrated that antibodies to IRPs were present only in the sera of animals immunized with SSE-IRP. The antibody profile of sera from the SSE-IRP group was similar to that obtained with serum from a lamb which had recovered from P. haemolytica A2 disease produced experimentally. Negligible levels of cytotoxin-neutralizing and bactericidal antibodies were detectable in the SSE-IRP group and therefore appear not to be involved in the protection observed in this experiment.

Animals↗