Intrastrain Heterogeneity of the mgpB Gene in Mycoplasma genitalium Is Extensive In Vitro and In Vivo and Suggests that Variation Is Generated via Recombination with Repetitive Chromosomal Sequences

Abstract

Mycoplasma genitalium is associated with reproductive tract disease in women and may persist in the lower genital tract for months, potentially increasing the risk of upper tract infection and transmission to uninfected partners. Despite its exceptionally small genome (580 kb), approximately 4% is composed of repeated elements known as MgPar sequences (MgPa repeats) based on their homology to the mgpB gene that encodes the immunodominant MgPa adhesin protein. The presence of these MgPar sequences, as well as mgpB variability between M. genitalium strains, suggests that mgpB and MgPar sequences recombine to produce variant MgPa proteins. To examine the extent and generation of diversity within single strains of the organism, we examined mgpB variation within M. genitalium strain G-37 and observed sequence heterogeneity that could be explained by recombination between the mgpB expression site and putative donor MgPar sequences. Similarly, we analyzed mgpB sequences from cervical specimens from a persistently infected woman (21 months) and identified 17 different mgpB variants within a single infecting M. genitalium strain, confirming that mgpB heterogeneity occurs over the course of a natural infection. These observations support the hypothesis that recombination occurs between the mgpB gene and MgPar sequences and that the resulting antigenically distinct MgPa variants may contribute to immune evasion and persistence of infection.Mycoplasma genitalium has been identified as a possible cause of several reproductive tract disease syndromes including urethritis in men and mucopurulent cervicitis, endometritis, pelvic inflammatory disease, and tubal factor infertility in women (3,6,7,18,23,28,30,34,35,41,48,52,55,56). When untreated (or inappropriately treated), this organism persists at infected sites as demonstrated by its association with chronic nongonococcal urethritis in men (22, 53) and its persistence for months, if not years, in the lower genital tract of women (C. R. Cohen, M. Nosek, A. Meier, S. G. Astete, S. Iverson-Cabral, N. R. Mugo, and P. A. Totten, submitted for publication). The longevity of M. genitalium infection in the human reproductive tract suggests that this pathogen may have mechanism(s) to modulate or evade the host immune response.At 580 kb, the M. genitalium G-37 T genome is one of the smallest of any self-replicating cellular organism (9, 17, 49), yet 4% of the fully sequenced genome consists of repeated elements (43) designated as MgPa repeats or MgPar sequences (17). These repeat sequences are so named because they are composed of partial, incomplete copies of the mgpB gene, which encodes the MgPa protein that mediates attachment to various cell types, including the ciliated epithelium of human fallopian tubes (8). Although there is only a single mgpB expression site, there are nine distinct MgPar sequences found throughout the genome (17). The MgPa protein is antigenic and elicits an immune response in animal models (25, 37) and in women with tubal factor infertility (6). The presence...

Analysis Identifying Common and Distinct Sequences among Texas Clinical Strains of Mycoplasma genitalium

“…Reference strain G37 and 73 distinct clinical sequences obtained in this study or previously (14,17,21) were aligned by CLUSTALW. The sequence of the corresponding region of the M. pneumoniae adhesin P1 gene (MPN141) was included as an outlier.…”

“…Alignment of MgPa-13 sequences corroborated transmission between partners and colonization of different anatomical sites by the same strain. Comparison of newly recovered sequences with all currently available clinical data (14,17,21) revealed not only the presence of several common variants but also distinct sequences. Analyses of encoded amino acids uncovered mutations that determine common features among MgPa-13 region variants.…”

“…Recently, sequence variability of the rRNA operon and tandem repeats in the MG309 locus have been evaluated with clinical specimens (28). However, the majority of M. genitalium clinical sequence data available today is based upon the gene mgpB (locus MG191 of sequenced reference strain G37 [11]) encoding the M. genitalium adhesin MgPa (14,17,21). An M. genitaliumspecific diagnostic PCR assay was designed to target the proximal unique region of this gene (using primers MgPa-1 and MgPa-3 [22] and designated here "MgPa-13").…”

Genetic Variation inMycoplasma genitalium

“…3) [31,39,40]. The MgPar DNA sequences have a partial similarity with the B, EF, and G areas in the MgPa gene, suggesting the possibility of genetic recombination [31,[39][40][41][42].…”

Molecular Diagnostics of Mycoplasmas: Perspectives from the Microbiology Standpoint

“…The RAPD / AP-PCR method involves PCR amplification with a single arbitrary primer at low stringency, while AFLP method selectively amplifies restriction fragments from whole genome. These PCR-based genotyping techniques have allowed faster and reproducible typing of mycoplasmas for epidemiologic studies [Cousin-Allery et al, 2000;Geary & Forsyth, 1996;Grattard et al, 1995;Iverson-Cabral, et al, 2006;Kokotovic et al, 1999;Rawadi, 1998;Schwartz et al, 2009].…”