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Vaccine-related fowl cholera must be considered when flock mortality increases after use of a live Pasteurella multocida vaccine product. All registered live vaccines serotype as Heddleston 3,4; however, in some regions this is also the most common serotype of outbreak isolates in broiler breeders and turkeys. Therefore, serotyping may not be useful for diagnosing vaccine-related fowl cholera. This project sought to apply a vaccine-specific test to differentiate vaccine-related disease from naturally occurring outbreaks. Results indicate that vaccine strains were commonly isolated from broiler breeders exhibiting signs of fowl cholera postvaccination, but some of these isolates exhibited only serotype 4 antigenicity. The isolates' lipopolysaccharides, the target antigen for serotyping, contained compositional changes that may explain the varying serotype results and virulence of the commercial preparations. These results suggest that vaccine-related disease may be common in broiler breeders, and live commercial vaccine preparations need to be assessed for serotype and titer prior to use in order to reduce vaccine-related fowl cholera.

The objective of this research was to evaluate the safety of the 6/85 strain vaccine strain of Mycoplasma gallisepticum in turkeys by backpassing the vaccine strain up to 10 times by contact infection in turkeys and challenging turkeys with the resulting backpassaged strain. The vaccine strain, however, did not spread to in-contact turkeys, and it was necessary to reisolate the organism before challenging turkeys for the next passage. The challenge strain, therefore, was one that had been backpassaged four times in turkeys, with a total in vivo time in turkeys of 66 days. The backpassaged 6/85 vaccine strain was no different in pathogenicity than the original vaccine strain, except that at 10 days postchallenge, it was isolated in higher numbers from air sacs. Both the original 6/85 vaccine strain and the backpassaged strain were apathogenic in turkeys, except for a slightly increased diameter of the tracheal mucosa at 10 days postchallenge; at 20 days postchallenge the tracheal mucosal thickness was no different from that of controls.

An outbreak of Mycoplasma gallisepticum (MG) in commercial turkeys involving very mild clinical signs was difficult to confirm by routine methods. In the first part of this study (trial A), we conducted a bioassay to increase the likelihood of detecting MG. Susceptible turkeys were inoculated with sinus exudates from four different affected commercial turkey flocks. Turkeys were evaluated for clinical signs, as well as by serology and culture of tracheal swabs, at 21 and 42 days postchallenge. An MG isolate from one of the sinus exudates used for inoculation, designated K5054, was very similar to isolates from house finches when characterized by random amplified polymorphic DNA analysis as well as DNA sequence analysis of portions of the phase-variable putative adhesin protein (pvpA) gene, a lipoprotein gene, and the cytadhesin gapA/mgc1 gene. The turkeys inoculated with the K5054 sinus exudate seroconverted in the absence of severe clinical signs. There was a single reisolation of K5054 from these turkeys 42 days postchallenge. Susceptible contact turkeys were commingled with the K5054-inoculated turkeys at 49 days postchallenge. We found no evidence of transmission of MG to the contacts by culture or serology at 7, 21, or 35 days after commingling. In the second part of this study (trial B), we challenged the contacts and K5054 sinus exudate-inoculated turkeys from trial A with virulent R strain 88 days after the K5054 sinus exudate inoculation. On necropsy 10 days postchallenge, the evaluation of gross and microscopic lesions, serology, and culture showed that the turkeys previously inoculated with K5054 sinus exudate were protected against disease and reinfection.

A total of 67 Mycoplasma gallisepticum field isolates from the USA, Israel and Australia, and 10 reference strains, were characterized by gene-targeted sequencing (GTS) analysis of portions of the putative cytadhesin pvpA gene, the cytadhesin gapA gene, the cytadhesin mgc2 gene, and an uncharacterized hypothetical surface lipoprotein-encoding gene designated genome coding DNA sequence (CDS) MGA_0319. The regions of the surface-protein-encoding genes targeted in this analysis were found to be stable within a strain, after sequencing different in vitro passages of M. gallisepticum reference strains. Gene sequences were first analysed on the basis of gene size polymorphism. The pvpA and mgc2 genes are characterized by the presence of different nucleotide insertions/deletions. However, differentiation of isolates based solely on pvpA/mgc2 PCR size polymorphism was not found to be a reliable method to differentiate among M. gallisepticum isolates. On the other hand, GTS analysis based on the nucleotide sequence identities of individual and multiple genes correlated with epidemiologically linked isolates and with random amplified polymorphic DNA (RAPD) analysis. GTS analysis of individual genes, gapA, MGA_0319, mgc2 and pvpA, identified 17, 16, 20 and 22 sequence types, respectively. GTS analysis using multiple gene sequences mgc2/pvpa and gapA/MGA_0319/mgc2/pvpA identified 38 and 40 sequence types, respectively. GTS of multiple surface-protein-encoding genes showed better discriminatory power than RAPD analysis, which identified 36 pattern types from the same panel of M. gallisepticum strains. These results are believed to provide the first evidence that typing of M. gallisepticum isolates by GTS analysis of surface-protein genes is a sensitive and reproducible typing method and will allow rapid global comparisons between laboratories.

A Mycoplasma gallisepticum (MG) isolate from an atypically mild outbreak in turkey breeders was found to be similar to house finch isolates by DNA analyses. A preliminary study in turkeys showed that this isolate (K5054) caused very mild lesions and protected turkeys against subsequent challenge with a virulent MG strain. In this study, K5054 was further evaluated as a potential vaccine strain in commercial layer-type chickens and turkeys. The safety of K5054 was evaluated by aerosol challenge followed by evaluation of gross and histopathologic lesions as well as serologic reactions and isolation of MG from the trachea and air sacs. Infection of chickens (trial 1) and turkeys (trial 2) with K5054 resulted in little evidence of MG lesions. There was weak seroconversion, and K5054 was consistently reisolated from the tracheas of chickens and turkeys. The efficacy of K5054 as a vaccine was evaluated by aerosol challenge of vaccinated chickens (trial 3) and turkeys (trial 4) with virulent R strain. There was evidence of protection from lesions associated with MG.