Mycoplasma genitalium is a human bacterial pathogen linked to urethritis and other sexually transmitted diseases. Here, we assessed the incidence of M. genitalium infection in patients attending a sexually transmitted disease clinic in San Antonio, TX, by use of diagnostic real-time PCR. Overall, 16.8% of women and 15.1% of men were found M. genitalium positive. Regions of the mgpB gene, which encodes the MgPa adhesin, were amplified from positive clinical specimens and evaluated for sequence variability, which demonstrated transmission of the pathogen between sexual partners. Follow-up analysis of a subset of patient specimens revealed reinfection by a different strain of M. genitalium, indicating the absence of protective immunity. Eighteen DNA sequence variants were obtained and compared with all other available clinical sequences. Detailed analysis revealed silent mutations of six amino acid residues within the encoded region of the MgPa adhesin in numerous clinical strains. In addition, missense mutations of limited numbers of amino acids were observed. Alignment of putative amino acid sequences revealed the simultaneous occurrence of several mutations and the existence of identical or similar protein variants in strains from different locations.