Comparative genetic analysis of porcine mycoplasmas

by Blank, Walter Arnold

The members of the class Molliciltes, also known as the mycoplasmas, are among the simplest self replicating organisms. These prokaryotes are characterized by the lack of a ce11 wall, small genomes, and a low G+C DNA content. Three species isolated from the respiratory tracts of pigs, Mycoplasma hyopneumoniae,Mycoplasmafloccuiare. and Mycoplasma hyorhinis are known to be closely related. In order to determine whether two other porcine mycoplasma. Mvcoplasma hyosynoviae and M~copiusma hyopharyngis, are also related to these organisms, 16s rRNA phylogenetic studies were performed. It was found that M. hyosynoviae and M. hyopharyngis were not specificdly related either to each other nor to the three other pig mycoplasmas. Low resolution genetic maps of the genomes of M. hyopneumoniae, the causative agent of enzootic pneumonia in pigs (EPP), and M. flocciilare, a commensal organisrn, were constructed by pulsed-field electrophoresis (PFGE) and hybridization. It was found that since these species diverged, at least three chromosomal inversions had occurred which differentiate the genomes of these organisrns. One of these inversions involved a region in M. hyopneumoniae which contains a ciliary adhesin gene (believed to be a pathogenic factor) which is not found in M. flocculare and several genes whose products are highly immunogenic. Three different types of repeated elements were also found in M. hyopneitmoniae, two of which could not be found by hybridization in M.jZocculare but do not appear to have been involved in the chromosomal remangements. The third repeated element, present in one copy in M.fIocculare, had both of its copies in M. hyopneumoniae located near the ends of one of the inverted regions and could potentidly have mediated a recombination event. Assembly of the M. hyopneumoniae genomic map was facilitated and complemented by the simultaneous construction of an ordered cosrnid libraq. Five contigs of overlapping clones were assembled, which together represent coverage of approximately 67%of its genome. as calculated by comparing the total sizes of EcoRI fragments from the cosmid library to the genome size estimated by PFGE. This library could be useful in discovering genes as either vaccination or thenpeutic targets against EPP.