Epidemiology and antibiotic susceptibility patterns of mycoplasma sp. and ureaplasma urealyticum

[摘要] ENGLISH ABSTRACT: Overview: Mycoplasmas and ureaplasmas are not routinely diagnosed andare under researched in South Africa. Prevalence, population shifts especiallyconcerning genital flora and implications in infection or other conditions areunknown. Information pertaining to Mycoplasma pneumoniae in respiratorydisease is similarly lacking. There is little information on antimicrobialsusceptibilities and resistance development against Sexually TransmittedInfections (STI) syndromic management approaches.Aims: a) Elucidate mycoplasmal and ureaplasmal prevalence andcontributing factors concerning cervical colonisation or preterm delivery inconjunction with HIV and Chlamydia trachomatis b) Investigate prevalence ofM. pneumoniae in respiratory infections in conjunction with HIV,Mycobacterium tuberculosis and Pneumocystis jiroveci. c) Determineantimicrobial susceptibilities of mycoplasmas and ureaplasmas and analyseresistance genes. d) Assess the inter-generic transfer potential of resistancegene (tetM) between Ureaplasma spp. and Neisseria gonorrhea.Genital setting: The prevalence of genital mycoplasmas, ureaplasmas andChlamydia on women attending their first prenatal visit, in conjunction withpreterm labour or HIV status was investigated. For preterm labour (2003), 199women were monitored for preterm delivery (<37 weeks); for colonisation andHIV (2005), 219 women were screened. Microbial detection was performed onDNA extracted from endocervical swabs employing PCR techniques.Colonisation was seen to be highest in the 14-20 year group from 2003. Inwomen aged ±21 years, co-colonisation was 13% although there was a shiftfrom co-colonisation with Mycoplasma hominis and Ureaplasma spp. in 2003to other dual/triple combinations in 2005. Overall major trends from bothcollection periods were that the prevalence of Ureaplasma spp. tended to behigher in women ±26 years, whilst prevalence of C. trachomatis and M.hominis were lower. No association was evident between colonisation with M.hominis, U. urealyticum, Ureaplasma parvum and labour outcome. HIV status had no effect on the prevalence/co-colonisation of M. hominis, Ureaplasmaspp. or C. trachomatis.Respiratory setting: Studies were conducted to determine the prevalence ofcommunity acquired atypical pneumonias in adults (M. pneumoniae and P.jiroveci) and neonates (mycoplasmas, ureaplasmas and Chlamydiatrachomatis) in order to improve treatment management programmes in thePort Elizabeth region. Sputum specimens from 102 adult patients presentingwith pneumonia/symptoms of pneumonia admitted to hospitals were assessedby PCR. Details of patient's gender, age, HIV and Mycobacteriumtuberculosis status were provided by the hospitals. Women were seen to be athigh risk for community-acquired P. jiroveci colonisation. Overall, prevalenceof P. jiroveci was 52.9% (54/102 patients). P. jiroveci was mainly associatedwith HIV (25/74) (P. jiroveci and HIV positive patients in patient sample forwhich clinical data and HIV status was available) and co-infection with M.tuberculosis was observed in 12 HIV cases and one HIV negative patient. NoDHPS (20) or DHFR (17) resistance associated mutations were found in P.jiroveci. M. pneumoniae was detected in one patient. For prevalence studies(2007-2008) on atypical pneumonia in neonates, 69 endotracheal aspirateswere obtained. PCR detection of M. hominis, U. urealyticum and C.trachomatis was performed and U. parvum detected in two specimens.Antibiotic susceptibilities and resistance genes: The followinginvestigations on clinical isolates of U. parvum and U. urealyticum wereconducted (i) antibiotic susceptibility profiles, (ii) detection of drug target genemutations, or gene acquisitions and (iii) inter-generic resistance gene transferpotential to Neisseria gonorrhoeae. Culture techniques applied to 132endocervical specimens provided 66 Ureaplasma cultures (35 U. parvum, 9U. urealyticum, 22 U. parvum + U. urealyticum). MIC determinations toofloxacin, erythromycin, tetracycline, doxycycline, azithromycin and josamycinwere performed. Thirty-seven ureaplasma cultures were fully susceptible to allantibiotics tested; 21 showed intermediate resistance to erythromycin,azithromycin and ofloxacin; while seven were resistant to tetracycline, three ofwhich were also resistant to doxycycline and one also resistant to azithromycin. Concerning ofloxacin resistance directed at quinoloneresistance determining regions, a substitution of Ser83Leu in ParC wasdemonstrated in one intermediately-resistant Ureaplasma (MIC 4 µg/ml) whilea triple substitution of Asp112Glu in GyrA along with Ala125Thr andAla136Thr in ParC was found in six further intermediately-resistant strains. Nomutations were found in strains with MICs 1 µg/ml. No mutations weredetected in 23S rRNA operons, L4 or L22 proteins. TetM and int-Tn geneswere found in seven tetracycline-resistant strains. On screening 59tetracycline-susceptible and -intermediate strains, eleven whilst possessingan int-Tn gene lacked a large region of tetM and 48 only contained smallregions of tetM. The tetM genes of the seven tetracycline-resistant strainswere sequenced and comparisons performed against GenBank sequences ofNeisseria gonorrhoeae, Streptococcus pneumoniae and U. urealyticum. Forfive strains tetM was seen to be highly mosaic in structure containing regionsthat were similar to those of the GenBank strains and others that were unique.In the tetM leader region, four hot spot recombination sites were identified thatcould certainly influence the formation of the mosaic structures, upstreaminsertion sequences/open reading frames and transposon regions thatregulate expression. On characterising the int-Tn genes of the seventetracycline-resistant strains, three types were present indicating transposonsfrom different origins had integrated into ureaplasma genomes. Reciprocaltetracycline resistance gene transfer between ureaplasmas and N.gonorrhoeae were unsuccessful. However, low-level tetracycline resistance(MICs 4-8 µg/ml) was transferred to a U. parvum recipient from one U.urealyticum and three U. parvum donors that carried tetM with MICs 16-64µg/ml. On tetM PCR analysis, tetM was not detected in the transformants.Conclusions: The importance of genital mycoplasmas, ureaplasmas and C.trachomatis in long term aetiologies requires further investigations, certainly inrelation with syndromic management regimens that fail to reduce colonisationrates. The high prevalence of P. jiroveci, the presence of M. pneumoniae incases of pneumonia and detection of U. parvum in two cases of neonatalpneumonia investigated emphasises that in the absence of definitivediagnoses, it is crucial to monitor treatment responses carefully, especially when first line antibiotic preferences are ß-lactams, in order to ensureadequate and informed delivery of medical care. The finding of transposonand/or tetM regions in all ureaplasmas investigated with or without fullexpression of tetracycline resistance, in conjunction with tetM gene diversity,certainly places ureaplasmas strongly in the picture for intra- and inter-genericexchange of antibiotic resistance genes.

[发布机构] Stellenbosch University