GENOTYPIC AND PHENOTYPIC ANALYSIS OF MYCOPLASMA-FERMENTANS STRAINS ISOLATED FROM DIFFERENT HOST TISSUES

A correlation was found between the expression of a specific Mycoplasm a fermentans surface antigen (Pra, proteinase resistant antigen) and t he site of isolation of the organism from the infected host. Strains w hich expressed Pra were most frequently associated with cells of bone marrow origin, and strains which lacked expression of Pra were most co mmonly isolated from the respiratory tract, genital tract, and arthrit ic joints, i.e., epithelial cell surfaces, Pra was previously shown to be resistant to degradation by proteinases and was hypothesized to pl ay a protective role at the organism surface and perhaps to influence which host tissue site was colonized by the organism. The methods used for this phenotyping scheme required isolation and growth of the myco plasma in quantities sufficient for immunoblot analysis using monoclon al antibodies, We wanted to determine a more rapid and less cumbersome technique to supplement this method for determining the Pra phenotype directly in clinical specimens. Here we describe PCR studies to inves tigate the movement of a previously identified M. fermentans insertion sequence (IS)-like element. These data shelved a correlation between a specific IS genotype and the Pra(+) phenotype. Production of a 160-b p product using a single set of IS-based primers was associated with e xpression of Pra, The genomic IS location resulting in the 160-bp prod uct was determined by using Southern blot analysis and was found to be a stable insertion site characteristic of genotype I strains. Additio nal analyses of sequences within and flanking the IS insertion sites r evealed another pair of PCR primer sites which resulted in the consist ent production of a 450-bp amplicon, The stability of this site was de pendent on the absence of the IS-like element between the primer sites . The production of this 450-bp amplicon correlated with the Pra mutan t phenotype and was characteristic of genotype II strains. The data sh owed that the sequence within the IS may be unstable and that reliable genotyping sequences are more easily found in the stable genomic site s which flank the IS element.