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Thesis (MSc (Med)(Microbiology))--University of Limpopo (Medunsa Campus), 2010. / Mycoplasma genitalium, a human mycoplasma species has been established as a cause of nongonococcal urethritis (NGU) in men, particularly in Chlamydia trachomatis-negative patients. It was also shown to play a role in cervicitis and pelvic inflammatory disease (PID) in women. Due to difficulty in culturing, and the lack of routine molecular diagnostic tests, many M. genitalium infections are undetected. The purpose of this study was to evaluate three nucleic acid amplification tests (NAATs) i.e. a recently developed Gen-Probe research only transcription mediated amplification (TMA) assay, a conventional polymerase chain reaction (PCR) assay and a real-time PCR (q-PCR) assay for the detection of M. genitalium in urine specimens of men with symptoms of urethritis. To evaluate the three assays, 300 urine specimens were collected between June 2007 and July 2008 from sexually active male patients presenting with discharge (N=94) and/or burning on micturition (N=206) to a private medical practitioner in Silverton, Pretoria. A specimen was considered positive by extension of the gold standard i.e. if any two of the three assays were positive. This was used to calculate the sensitivity and specificity of each method. TMA detected M. genitalium in 62 (21%), PCR in 43 (14%) and q-PCR in 48 (16%) of the 300 patients. The sensitivities of the assays were 100% (TMA), 92% (q-PCR) and 78% (PCR), with specificities of 90% (TMA), 95% (q-PCR) and 97% (PCR). The sensitivity of the TMA assay was higher than that of the q-PCR and PCR assays. The lower sensitivity obtained by the q-PCR assay might have been due to inhibition and limitations in the amount of the DNA template. However, the q-PCR assay was easy to perform as it combines amplification and detection thus eliminating further handling of PCR products. The PCR, although with a higher specificity, was the least desirable in terms of testing time and problems with subjectivity when reading agarose gels. v We concluded that the Gen-Probe TMA assay is a highly sensitive method for detection of M. genitalium in urine specimens of men. The use of Gen-Probe TMA and the q-PCR assay, will increase the detection of M. genitalium in clinical specimens at this catchment area.

This work further elucidates processes involved in promoting and sustaining evolutionary diversification within the gall-inducing thrips that specialise on Australian Acacia. A phylogenetic approach was taken to determine modes of diversification available to these insects. The extension and revision of the gall-thrips phylogeny is central to the work and primarily focuses on cryptic populations of the Kladothrips rugosus and Kladothrips waterhousei species complexes. Parallel diversification, where the radiation of the K. rugosus and K. waterhousei lineages broadly mirror one another, offered a rare opportunity to test hypotheses of coevolution between gall-thrips and their Acacia hosts. In the absence of a reliable host Acacia phylogeny, indirect inference of insect/plant cospeciation can be arrived at as these two complexes share the same set of host species. The expectation is that if the phylogenies for the gall-thrips complexes show a significant level of concordance, then cospeciation between insect and host-plant can be inferred. Results indicate that the K. rugosus species complex comprise populations at species level. A significant level of phylogenetic concordance between the two species complexes is consistent with gall-thrips lineages tracking the diversification of their Acacia hosts. Given the less than strict form of insect/host cospeciation, factors impacting host diversification become important to gall-thrips diversification. Gall-thrips radiated over a period during the expansion of the Australian arid-zone. Cycles of host range expansion and fragmentation during the Quaternary could have played a major role in gall-thrips diversity. An interesting feature of resourse sharing amongst the K. rugosus and K. waterhousei complex members is the apparent absence of competitive exclusion between them. The persistence of this sympatry over millions of years is an unusual feature and merits further investigation.