Bordetella parapertussis from lambs

Extract

Madam:—An organism identified as Bordetella parapertussis has been isolated in significant numbers from nasal swabs, bronchial washings and the lungs of 6–10 month old lambs showing signs of pneumonia. It has also been isolated from normal lambs but attempts to recover it from adult sheep have so far been unsuccessful (Table I).     

Pneumonia in lambs inoculated with Bordetella parapertussis: clinical and pathological studies

Eight one-week-old, colostrum-deprived lambs were inoculated intratracheally with ovine isolates of Bordetella parapertussis. After inoculation, the lambs showed slight depression and anorexia. The total circulating leucocyte counts in these animals rose gradually to a peak five days post-inoculation. Neutrophil counts also increased and were highest at Day 3. Lesions grossly and histologically similar to those of naturally-occurring ovine chronic non-progressive pneumonia were seen in the majority of infected animals. Grossly, they consisted of many small areas of collapse and dull-red consolidation. Histologically, B. parapertussis caused mild acute tracheobronchitis, severe alveolar collapse and acute bronchopneumonia, which was most severe from Days 1 to 3. Pure cultures of B. parapertussis were consistently recovered from nasal swabs of infected lambs throughout the study. Viable bacterial counts of bronchoalveolar lavage fluid showed a rapid elimination of this organism from the lower respiratory tract between Days 1 to 5. Bordetella parapertussis infection in the respiratory tract of lambs has the potential to compromise pulmonary defence mechanisms and allow other pathogenic organisms to become established in the lower respiratory tract.     

Pneumonia in lambs inoculated with Bordetella parapertussis: Clinical and pathological studies

Eight one-week-old, colostrum-deprived lambs were inoculated intratracheally with ovine isolates of Bordetella parapertussis. After inoculation, the lambs showed slight depression and anorexia. The total circulating leucocyte counts in these animals rose gradually to a peak five days post-inoculation. Neutrophil counts also increased and were highest at Day 3. Lesions grossly and histologically similar to those of naturally-occurring ovine chronic non-progressive pneumonia were seen in the majority of infected animals. Grossly, they consisted of many small areas of collapse and dull-red consolidation. Histologically, B. parapertussis caused mild acute tracheobronchitis, severe alveolar collapse and acute bronchopneumonia, which was most severe from Days 1 to 3. Pure cultures of B. parapertussis were consistently recovered from nasal swabs of infected lambs throughout the study. Viable bacterial counts of bronchoalveolar lavage fluid showed a rapid elimination of this organism from the lower respiratory tract: between Days 1 to 5. Bordetella parapertussis infection in the respiratory tract of lambs has the potential to compromise pulmonary defence mechanisms and allow other pathogenic organisms to become established in the lower respiratory tract.     

Pneumonia in mice produced by cell-free extract of cultures of Bordetella parapertussis

A single dose of culture fluid of Bordetella parapertussis freed from cells (CFCF) given intranasally to four-week-old mice free from intercurrent respiratory disease produced a subacute bronchopneumonia, which was similar to that induced by whole cells of ovine isolates of B parapertussis, except that the lesions were less severe and less extensive. From eight hours to 17 days after inoculation, the mice exhibited marked infiltration of neutrophils and macrophages into the alveolar septa, bronchiolar and alveolar spaces, and hyperplasia of peribronchiolar and perivascular lymphoid tissue. Electron microscopy showed damage to ciliated cells, type 1 pneumocytes and alveolar macrophages. These results suggest that extracellular toxic substance(s) produced by ovine isolates of B parapertussis might be involved in the initiation and development of lesions in ovine chronic non-progressive pneumonia.     

Pneumonia in lambs inoculated with Bordetella parapertussis: bronchoalveolar lavage and ultrastructural studies

Eight colostrum-deprived lambs were inoculated intratracheally with ovine isolates of Bordetella parapertussis. Fluids obtained by bronchoalveolar lavage had a large increase in total cell counts 24 hours after inoculation; up to 93% of cells were neutrophils. From 3 days after inoculation, the number of alveolar macrophages in lavage samples was markedly increased. From 5 days onwards, many alveolar macrophages had moderate to severe cytoplasmic vacuolation. Topographically, tracheal and bronchial epithelium was covered by a large amount of inflammatory exudate 24 hours after inoculation. Later, the tracheobronchial epithelium showed focal extrusions from ciliated cells, which were occasionally associated with B. parapertussis organisms. Ultrastructurally, cytopathological changes associated with B. parapertussis infection were mild focal degeneration of airway epithelium with slight loss of cilia, moderate to severe degeneration of type I and type II alveolar epithelial cells, and focal inflammation in the lungs. These results suggest that the primary targets of B. parapertussis infection are alveolar macrophages and the epithelial cells of bronchioles and alveoli.     

Clearance of bacteria in turkeys with Bordetella avium-induced tracheitis

Quantitative clearance of aerosolized Escherichia coli from the trachea, lung, and air sacs was measured in turkeys infected with Bordetella avium. Clearance of E. coli in turkeys with B. avium-induced tracheitis was minimally affected early in infection. Sixteen to 23 days after infection with B. avium, sporadic, mild depressions in clearance of E. coli were observed in the tracheas, which had large areas of deciliated tracheal epithelium or replacement of normal epithelium by immature hyperplastic epithelium or metaplastic squamous epithelium. Clearance of E. coli from the lung and air sacs was minimally affected in turkeys infected with B. avium.     

Antimicrobial sensitivity testing of Australian isolates of Bordetella avium and the Bordetella avium-like organism

SUMMARY: The in-vitro sensitivity of 16 Australian isolates of Bordetella avium and 15 isolates of B avium -like organism to 11 antimicrobial agents or combinations of agents was determined using a microtitre plate system to establish minimal inhibitory concentrations. All the B avium isolates were sensitive to ampicillin but resistant to erythromycin, lincomycin, spectinomycin, sulphamethoxazole, trimethoprim, and lincomycin + spectinomycin. Most of the B avium isolates were sensitive to tetracycline and resistant to streptomycin and sulphadiazine. All the B avium -like isolates were resistant to ampicillin, erythromycin, lincomycin, spectinomycin, streptomycin, tetracycline, trimethoprim, and lincomycin + spectinomycin. Most B avium -like isolates were sensitive to sulphadiazine, sulphamethoxazole and trimethoprim-sulphamethoxazole.     

Antibiotic susceptibility of canine Bordetella bronchiseptica isolates

The antimicrobial sensitivities of 78 recent (1995-1998) canine isolates of Bordetella bronchiseptica from 13 separate sources were determined. Minimum inhibitory concentrations were assessed using the E-test method or by agar dilution. All 78 isolates were sensitive to tetracycline, doxycycline, enrofloxacin, and amoxycillin/clavulanic acid; the majority were sensitive to ampicillin (63/78; 81%), trimethoprim (57/78; 73%), and sulphadiazine (63/78; 81%). Plasmids were detected in 14 out of the 24 isolates tested. There was no correlation between the presence of plasmids and antibiotic resistance, but there was some correlation between the presence of plasmids and the origin of the isolates. Three sizes of plasmid were found: 20, 14, and 5.5 kb. Eight of the isolates contained all three plasmids, the remainder one or two, Thirteen isolates demonstrated beta-haemolysis, of which six produced a soluble haemolysin. Except for one isolate, haemolysin production correlated with plasmid carriage. Pulsed-field gel electrophoresis showed that all except one isolate could be grouped in the same genotype. Within this genotype isolates could be divided into three subtypes, generally corresponding to their place of origin.     

Immunodominant antigens of zoospores from ovine isolates of Dermatophilus congolensis.

Zoospores of Dermatophilus congolensis were analysed by SDS-PAGE and western blotting. The electrophoretic profiles of zoospores from 13 isolates of D. congolensis were similar but not identical when stained with Coomassie blue or silver. Immunodominant polypeptides with apparent molecular masses of 76 and 31 kDa were identified in western blots of 13 of 13 and 12 of 13 isolates respectively of D. congolensis reacted with hyperimmune, ovine, antizoospore sera. Identical immunodominant polypeptides were observed in western blots reacted with sera obtained from naturally infected sheep. Initial characterisation of the 76 and 31 kDa polypeptides indicated that they were probably surface exposed because (i) antibodies eluted from the surface of live zoospores after adsorption of hyperimmune antizoospore serum, reacted principally against the 76 and 31 kDa subunit polypeptides in western blots, (ii) adsorption of hyperimmune antizoospore serum with live zoospores resulted in significant diminution of reactivity against both the 76 and 31 kDa polypeptides in western blots, (iii) indirect fluorescent immunostaining of zoospores with antiserum prepared against gel-purified 76 kDa polypeptide, resulted in intense staining of the zoospore outer coat. Immuno-gold electron microscopy of negatively stained zoospores with antiserum prepared against gel-purified 31 kDa polypeptide identified this antigen as a flagella subunit.     

Polymorphism of pertactin gene repeat regions in Bordetella bronchiseptica isolates from pigs

Bordetella bronchiseptica pertactin (prn) is an outer membrane protein which has been implicated as both an adhesin and a protective antigen that induces immunity against atrophic rhinitis in pigs. Previous studies demonstrated extensive heterogeneity of the prn sequence within two distinct regions of amino acid repeats for B. bronchiseptica isolated from the United States and Europe. By deducing the amino acid sequences of the repeat regions of the prn gene from recent isolates from Korea, two region 1 variants and five region 2 variants were identified. Five pertactin types were distinguished based on combinations of variants of both regions. Interestingly, none of the field isolates have the same pertactin type as the B. bronchiseptica P4 strain widely used to vaccinate pigs.     

Serotypes and electrophoretic protein profiles of Pasteurella haemolytica isolated from pneumonic ovine lungs.

Serotypes and SDS-PAGE protein profiles of P. haemolytica isolated from pneumonic ovine lungs were investigated. Of 268 P. haemolytica isolates, 232 (86.6%) were serotypable. A total of 12 serotypes were recognized in 20 different geographic origins of central Turkey. The most common serotype was A2, followed by A7, A1 and T4. Serotypes A13, A14, A16 and T15 could not be detected. In SDS-PAGE, marked differences between major bands of biotype A and T strains were found. In numerical analysis of protein profiles, biotype A and T strains were separated at 58% similarity level. Biotype A isolates produced a cluster at 80% similarity level, and biotype T isolates at 92% similarity level. No single cut off level was able to discriminate between each serotype studied and isolates could not be clustered on the basis of their geographic origins.     

Visual assessment of pneumonic area in ovine lungs

Twelve veterinarians were asked to estimate the percentage of lung surface that appeared consolidated in eleven projected colour transparencies of pneumonic ovine lungs. The estimates were compared to the pneumonic area measured by image analysis. Although all observers had some experience in veterinary pathology there was considerable variation between individuals in the accuracy of their estimates. Large lesions were underestimated and small lesions overestimated by most observers.     

Serologic classification of two ovine adenovirus isolates from the central United States

Two ovine adenovirus (OAV) strains (RTS-42 and RTS-151), isolated from lambs in the central United States, were compared using 2-way cross-neutralization tests with the 6 recognized OAV species, 9 bovine adenovirus species, and 4 porcine species. Virus RTS-42 was identified as OAV type 5, confirming previous results. Virus RTS-151 was identified as OAV type 6, although the serologic crossing was largely one-sided.     

Distinguishing between ovine abortion and ovine arthritis Chlamydia psittaci isolates with specific monoclonal antibodies

Monoclonal antibodies to Chlamydia psittaci were prepared by both in vivo and in vitro immunization methods, using an abortion strain of C psittaci as the immunizing antigen. Seven of the 8 monoclonal antibodies produced were genus-specific by the enzyme-linked immunosorbent assay and immunofluorescence test. The genus-specific antibodies were reactive with a protease-resistant, periodate-sensitive antigen of less than 14 kilodaltons. The remaining monoclonal antibody, 10D7, was specific for ovine abortion strains of C psittaci and nonreactive with 2 strains isolated from the joints of lambs with polyarthritis. The type-specific antigen was protease sensitive, but could not be detected in the immunoblot assay.     

Influence of Bordetella avium infection on association of Escherichia coli with turkey trachea

Four-week-old Bordetella avium-infected and B avium-free turkeys were inoculated intratracheally with a suspension of fimbriated or nonfimbriated Escherichia coli. Numbers of E coli associated with tracheal sections were determined at postinoculation hour (PIH) 1 or 6. Significantly (P less than 0.05) greater numbers of E coli were isolated from the tracheas of B avium-infected turkeys compared with numbers in B avium-free turkeys. In B avium-free turkeys, tracheal associated E coli were 90% less at PIH 6 compared with that at PIH 1. However, in B avium-infected turkeys, numbers of E coli were not affected by postinoculation time. Seemingly, B avium-infected turkeys had reduced capacity to clear E coli from the trachea.     

猪萎缩性鼻炎支气管败血波氏杆菌PCR检测方法的建立

对表现猪萎缩性鼻炎临床症状的猪群中分离得到的34株支气管败血波氏杆菌(Bordetella bronchiseptica,Bb),采用针对Bb flagellum gene的一对引物进行PCR扩增,结果所有分离物均能扩增出237bp特异性DNA条带,与传统生化鉴定结果相一致,且其最小检出量为0.64pg;而猪鼻腔和肺组织中常见的多杀性巴氏杆菌、金黄色葡萄球菌、枯草芽胞杆菌、铜绿假单胞菌、变形杆菌及大肠埃希氏菌均未能出现任何DNA条带。这表明,本试验建立的PCR方法具有特异性强、灵敏度高、可靠性好等特点,可用于猪萎缩性鼻炎的临床诊断。     

Absence of mouse-lethal toxins in Australian isolates of Bordetella avium

A total of 18 Australian isolates of Bordetella avium and seven reference B. avium strains from Europe and the USA were tested for the presence of a mouse-lethal toxin. Five of the seven reference strains of B. avium, but none of the 18 Australian isolates, produced the toxin.     

Antimicrobial susceptibility of Bordetella bronchiseptica isolates from cats and a comparison of the agar dilution and E-test methods

One hundred and fifty-two predominantly feline isolates of Bordetella bronchiseptica were tested for their susceptibility to seven antimicrobial agents using an agar dilution method. The majority of isolates tested by the agar dilution method were resistant to trimethoprim (MIC₉₀ 500 μg/ml) and ampicillin (MIC₉₀ > 32 μg/ml) but sensitive to tetracycline, doxycycline and enrofloxacin (MIC₉₀ 2 μg/ml for all three agents). The isolates showed a spectrum of susceptibility to sulphadiazine and clavulanate potentiated amoxycillin. The MIC's of twenty-nine of the 152 isolates were then compared for five of the antimicrobial agents using the E-test (AB Biodisk, Sweden), a recently introduced method for measuring the MIC's of antimicrobial agents based on the diffusion of a pre-defined antibiotic gradient from a plastic strip. Comparisons with the E-test demonstrated an overall agreement (±1 log₂ dilution) with the agar dilution method of 79.4% and an agreement within ±2 log₂ dilutions of 96.2%.     

Diversity of swine Bordetella bronchiseptica isolates evaluated by RAPD analysis and PFGE

The degree of genetic diversity in 45 Bordetella (B.) bronchiseptica strains comprised of a vaccine strain (N = 1), reference strains (N = 3) and field isolates (N = 41) was evaluated using random amplified polymorphic DNA (RAPD) fingerprinting and pulsed-field gel electrophoresis (PFGE). Three candidate primers were selected for RAPD analysis after screening 20 random decamer oligonucleotides for their discriminatory abilities. The OPA-07, OPA-08 and OPA-18 primers yielded 10, 10, and 6 distinct fingerprint patterns, respectively. The most common identical RAPD pattern was produced by OPA-07 which was shared by 32 isolates (71.1%), the pattern produced by OPA-08 was shared by 26 isolates (57.8%), and the pattern produced by OPA-18 was shared by 40 isolates (88.9%). The RAPD patterns of the vaccine strain and the 3 reference strains did not match any of the patterns produced by the field isolates when primers OPA-07 and OPA-08 were used. PFGE using the restriction endonuclease XbaI produced a total of 15 patterns consisting of 4 PFGE types (A, B, B1 and C, differing by ≥ 4 bands) and 11 A subtypes (differing by ≤ 3 bands). Most of the field isolates exhibited identical type A and B patterns, suggesting that they were related. The vaccine strain and the three reference strains showed different PFGE patterns as compared to the identical type A strains.