Survey of Victorian small ruminant herds for mycoplasmas associated with contagious agalactia and molecular characterisation of Mycoplasma mycoides subspecies capri isolates from one herd.

OBJECTIVE: Regarded as one of the most expensive production diseases of dairy sheep and goats, contagious agalactia (CA) is caused by any of four agents: Mycoplasma agalactiae, M. mycoides subspecies capri (Mmc), M. capricolum subspecies capricolum (Mcc) and M. putrefaciens. Although CA is worldwide in distribution, it has not been reported in Australia, even though studies between the 1950s and 1980s isolated each agent from sheep or goats without any clinical signs associated with it. The aim of this study was to examine sheep and goats in Victoria, Australia, for the presence of CA-associated mycoplasmas and to investigate the evolutionary relationships of these isolates by comparing their genetic differences with their counterparts from other parts of the world. METHODS: A 3-year epidemiological survey of small ruminant populations in Victoria, Australia, was conducted for the presence of CA-associated mycoplasmas and the isolates obtained were genotyped by multilocus sequence typing (MLST). RESULTS: Mmc was the only CA-associated agent isolated from the 1358 samples analysed in the study, but was not associated with CA on the property where it was found. MLST analyses of Mmc strains revealed a distinct clustering of Australian isolates into a novel clade, with the closest relatives being strains from Europe. The distinct clustering is consistent with the absence of clinical disease in Australia. CONCLUSION: The isolation of Mmc indicates that this subspecies persists in Australian small ruminant populations. However, full genome sequencing and in vitro animal experimentation are needed to unequivocally demonstrate the avirulence of Australian strains.

Induction of enzootic pneumonia in pigs by the administration of an aerosol of in vitro-cultured Mycoplasma hyopneumoniae.

The aim of this study was to establish whether enzootic pneumonia could be induced reliably in piglets by administering an aerosolised culture of Mycoplasma hyopneumoniae. Groups of five M hyopneumoniaefree Landrace x Large White piglets weaned at 11 to 14 days of age were exposed to aerosols of in vitro cultures of a virulent strain of M hyopneumoniae. In three separate trials, 14 of 15 pigs exposed to the bacteria developed pneumonia, but pigs exposed to the culture medium alone did not develop the disease. Lung pathology, both gross and histological, indicated acute disease. Ten of the pigs were tested for seroconversion by Western blot and they were all positive. The growth rates of the infected pigs were significantly reduced and the water consumption of the infected groups was also depressed. M hyopneumoniae was recovered from eight of the 15 infected pigs.

A novel mechanism for control of antigenic variation in the haemagglutinin gene family of mycoplasma synoviae.

High-frequency phase and antigenic variation of homologous lipoprotein haemagglutinins has been seen in both the major avian mycoplasma pathogens, Mycoplasma synoviae and Mycoplasma gallisepticum. The expression and, hence, antigenic variation of the pMGA gene family (encoding these lipoproteins in M. gallisepticum) is controlled by variation in the length of a trinucleotide repeat motif 5' to the promoter of each gene. However, such a mechanism was not detected in preliminary observations on M. synoviae. Thus, the basis for control of variation in the vlhA gene family (which encodes the homologous haemagglutinin in M. synoviae) was investigated to enable comparison with its homologue in M. gallisepticum and with other lipoprotein gene families in mycoplasmas. The start point of transcription was identified 119 bp upstream of the initiation codon, but features associated with control of transcription in other mycoplasma lipoprotein genes were not seen. Comparison of three copies of vlhA revealed considerable sequence divergence at the 3' end of the gene, but conservation of the 5' end. Southern blot analysis of M. synoviae genomic DNA revealed that the promoter region and part of the conserved 5' coding sequence occurred as a single copy, whereas the remainder of the coding sequence occurred as multiple copies. A 9.7 kb fragment of the genome was found to contain eight tandemly repeated regions partially homologous to vlhA, all lacking the putative promoter region and the single-copy 5' end of vlhA, but extending over one of four distinct overlapping regions of the 3' coding sequence. Examination of sequential clones of M. synoviae established that unidirectional recombination occurs between the pseudogenes and the expressed vlhA, with duplication of pseudogene sequence and loss of the corresponding region previously seen in the expressed gene. Expression of the 5' end of two variants of the vlhA gene showed that they differed in their reaction with monoclonal antibodies specific for this region. These data suggest that the control of vlhA antigenic variation in M. synoviae is achieved by multiple gene conversion events using a repertoire of coding sequences to generate a chimeric expressed gene, with the greatest potential for variation generated in the region encoding the haemagglutinin. Thus, completely distinct mechanisms have been adopted to control antigenic variation in homologous gene families.