Peter C. Scott

Genetic organization of the ovine MHC class II region.

To study the class II genes of the major histocompatibility region of the sheep genome, human HLA class II genes corresponding to the known subregions in man (DR, DQ, DP, DO, and DZ) were used for Southern hybridization analysis of sheep DNA and to probe a sheep genomic library. Hybridizing bands were noted for all probes except DPα. DQ α and β and DRβ appear to be present as multicopy genes, while DRα-, DZα-, , and DOβ-like genes appear to be single copy. All bands detected with the DPβ probe were also detectable with other β chain probes. From eight λ-bacteriophage clones of a sheep genomic library nine distinct class II genes were identified. These genes were characterized by differential hybridization analysis and restriction mapping. Two genes were DRβ-like, three DQα-like and four DQβ-like. The extensive cross-hybridization observed with β chain probes was not seen with α chain probes. The results of this study suggest that the major histocompatibility complex class II region of the sheep has a similar genetic organization to that of man, with the provisional exception of the DP subregion.

Nucleotide sequence, polymorphism, and evolution of ovine MHC class II DQA genes

The nucleotide sequence of all exons and introns, excluding exon 1, of the ovine major histocompatibility complex (MhcOvar) genes analogous to the HLA-DQA1 and -DQA2 genes has been determined and the gene structure found to be similar to that reported for other species. The predicted amino acid sequences of the Ovar-DQA genes have been compared with the equivalent DQA genes in man, mouse, rat, rabbit, and cattle and used to determine the evolutionary relationships of the sheep class II genes to these other species. Northern blot analysis of sheep mRNA using exon specific probes for each of the two Ovar-DQA genes show that both genes are transcribed, whereas in humans there is no evidence that HLA-DQA2 is transcriptionally active. Restriction fragment length polymorphisms (RFLPs) have been used to define a polymorphic series of alleles in both Ovar-DQA genes and have indicated that the number of DQA genes is not constant in sheep as it is in humans, but varies with the haplotype.

Safety and Efficacy of Two Live Pasteurella multocida aro-A Mutant Vaccines in Chickens

Two auxotrophic aro-A mutants of Pasteurella multocida designated PMP1 (serotype 1) and PMP3 (serotype 3) were tested as vaccine candidates to protect chickens against fowl cholera. A reliable intratracheal challenge method was established that resulted in > or = 75% mortality in both specific-pathogen-free chickens and commercial broiler breeders 24 hr after challenge. Dose protection studies indicated that at least 10(6) colony-forming units (CFU) of PMP1 and 10(8) CFU of PMP3 were required to provide complete protection against challenge in all birds. Although high doses of 10(9) CFU of the vaccine strains produced some endotoxinlike reactions, lower but protective dose levels produced no clinical sign or lesion in any chicken. Both vaccine strains provided cross-protection with a heterologous challenge strain PM206 (serotype 4). Future studies will examine the duration of protective immunity induced by the two vaccine candidates, PMP1 and PMP3, and cross-protection against other serovars.

Field Evaluation of the Safety and Efficacy of a Temperature-sensitive Mycoplasma synoviae Live Vaccine

Mycoplasma synoviae (MS) strain MS-H was used in three separate commercial flocks for large-scale evaluation of the safety and efficacy of the vaccine under commercial conditions. MS-H successfully colonized meat and layer-breeders vaccinated by eyedrop and persisted for up to 55 wk after vaccination. Restriction fragment length polymorphism analysis showed that MS-H was the only strain isolated from two vaccinated flocks. In a third flock, challenge with a wild-type MS occurred, and this strain was isolated from both vaccinated and unvaccinated birds. Vertical transmission of MS-H was investigated by culturing pipped embryos and testing broiler progeny for MS antibody at processing (56 days old). No evidence of vertical transmission was detected. Lateral transmission of MS-H strain from vaccinated to unvaccinated birds occurred in one of the commercial flocks. Forty-one of 50 isolates of MS-H obtained from vaccinated flocks maintained their temperature-sensitive phenotype, but nine isolates showed a nontemperature-sensitive phenotype.

Chronological analysis of gross and histological lesions induced by field strains of fowl adenovirus serotypes 1, 8b and 11 in one-day-old chickens

Fowl adenoviruses (FAdVs) cause diseases in domestic chickens, including inclusion body hepatitis (IBH), with immunosuppression believed to play a role in their pathogenesis. To gain a better understanding of the pathogenesis and chronology of disease caused by FAdVs, the gross pathology, histopathology and dissemination of virus were examined at several different time points, after inoculation of one-day-old specific pathogen-free chickens with FAdV-1, FAdV-8b or FAdV-11 via the ocular route. FAdV-8b had a slightly greater virulence than FAdV-11, but both were primary pathogens. The presence and severity of hepatic lesions were used to define the three stages of the disease: incubation (1–3 days post-inoculation, PI), degeneration (4–7 days PI) and convalescence (14 days PI). Both viruses were detected in the liver, kidney, bursa, thymus and gizzard of most birds during the degenerative stage, and persisted in the gizzard into convalescence. The FAdV-1 isolate was found to be apathogenic, but virus was detected in the bursa and/or gizzard of several birds between 2 and 7 days PI. This is the first study examining the chronology of gross and microscopic lesions of pathogenic and apathogenic FAdVs in association with viral presence in multiple tissues. It was concluded that both FAdV-8b and FAdV-11 are primary pathogens, and that these strains may play a role in immunosuppression.

Behavioural responses of poultry during kosher slaughter and their implications for the birds' welfare

Measurements were made during Shechita (kosher) slaughter of 692 meat chickens, including the behaviour of the birds during the procedure and the times from their removal from the crate, to neck cutting, bleed-out and shackling. Four of 100 birds showed a mild physical response to neck cutting but the others showed no response. Approximately 60 per cent of the birds showed a physical response to touching the eye or eyelid at up to 5 seconds after neck cutting, but by 15 seconds none showed this response. The birds became unable to retain their posture and suffered involuntary muscular contractions at 12 to 15 seconds after neck cutting and had lost approximately 40 per cent of their total blood volume by 30 seconds after neck cutting.

The presence of viral subpopulations in an infectious bronchitis virus vaccine with differing pathogenicity--a preliminary study.

Abstract There are currently four commercially available vaccines in Australia to protect chickens against infectious bronchitis virus (IBV). Predominantly, IBV causes clinical signs associated with respiratory or kidney disease, which subsequently cause an increase in mortality rate. Three of the current vaccines belong to the same subgroup (subgroup 1), however, the VicS vaccine has been reported to cause an increased vaccinal reaction compared to the other subgroup 1 vaccines. Molecular anomalies detected in VicS suggested the presence of two major subspecies, VicS-v and VicS-del, present in the commercial preparation of VicS. The most notable anomaly is the absence of a 40 bp sequence in the 3′UTR of VicS-del. In this investigation, the two subspecies were isolated and shown to grow independently and to similar titres in embryonated chicken eggs. An in vivo investigation involved 5 groups of 20 chickens each and found that VicS-del grew to a significantly lesser extent in the chicken tissues collected than did VicS-v. The group inoculated with an even ratio of the isolated subspecies scored the most severe clinical signs, with the longest duration. These results indicate the potential for a cooperative, instead of an expected competitive, relationship between VicS-v and VicS-del to infect a host, which is reminiscent of RNA viral quasi-species.

Onset of Immunity with Mycoplasma synoviae: Comparison of the Live Attenuated Vaccine MS-H (Vaxsafe MS) with Its Wild-Type Parent Strain (86079/7NS)

Abstract The duration of protective immunity elicited by the MS-H vaccine was evaluated by experimental challenge of chickens at 15 and 40 wk after eyedrop vaccination. Immunity induced by the parent strain of the vaccine, 86079/7NS, was also investigated for comparison. A serological response to Mycoplasma synoviae was detected in 89% to 100% of MS-H vaccinates and 86079/7NS inoculates at 15, 27, 30, 35, and 40 wk after inoculation. A significantly lower incidence of air-sac lesions and lower air-sac lesion severity were observed in both the MS-H vaccinated and the 86079/7NS inoculated groups, as compared to the unvaccinated controls, after both challenge points. Tracheal mucosal thicknesses in MS-H vaccinates was significantly lower in the upper, lower, and total trachea at 40 wk after vaccination, as compared to the controls. It was demonstrated in this experiment that protective immunity, as determined by protection against experimental challenge, was maintained to at least 40 wk after vaccination.

Determination of the Effective Dose of the Live Mycoplasma synoviae Vaccine, Vaxsafe MS (Strain MS-H) by Protection Against Experimental Challenge

Abstract The minimum effective dose of the Mycoplasma synoviae-H (MS-H) vaccine was determined through protection against experimental challenge. Chickens were vaccinated by eyedrop with the following doses of a vaccine: 1.2 × 105, 2.4 × 105, 4.8 × 105, 9.6 × 105, 1.92 × 106, and 3.84 × 106 color change units (CCU), then challenged 6 wk after vaccination. Rapid serum agglutination results indicated that 100% of birds receiving an MS-H dose of ≥4.8 × 105 CCU had antibodies to MS and enzyme-linked immunosorbent assay results showed that 60% of birds receiving a dose of 4.8 × 105 or 9.6 × 105 CCU and 100% of birds receiving a dose of 1.92 × 106 or 3.84 × 106 had antibodies to MS. At postmortem after challenge, the following parameters were significantly lower in birds vaccinated with an MS-H dose of ≥4.8 × 105 CCU: air sac (AS) lesion severity; incidence of AS lesions; mucosal thicknesses in the upper trachea, middle trachea, and lower trachea (LT); and MS colonization of the LT and AS. It was concluded that an MS-H dose of 4.8 × 105 CCU was sufficient to elicit an antibody response in birds, prevent MS colonization in the LT and AS, and protect against AS lesions caused by an experimental MS and infectious bronchitis virus challenge.

Biochemical and molecular analysis of sheep MHC class II molecules.

A panel of monoclonal antibodies was used for structural and immunodepletion analysis of sheep MHC class II molecules. The results indicate the antibodies recognize molecules of molecular weight 32-34,000 (alpha chain) and 26-28,000 (beta chain). Immunodepletion analysis indicates that the antibodies may recognize up to four distinct class II molecules some of which are structurally distinguishable using SDS-PAGE. Southern blot analysis using HLA-D region DR, DQ, DP, DO and DZ cDNA probes showed that a number of the cDNA probes hybridized specifically to sheep DNA indicating the presence of closely related genes in sheep. Together the results suggest that the sheep MHC class II region contains distinct MHC class II genes similar to those found in man.