Purpose.Chlamydiatrachomatis is responsible for trachoma-associated blindness as well as the most common sexually transmitted bacterial infection worldwide, although the genovars for the former are typically A–C, whilst for the latter they are D–K and for the uncommon infection lymphogranuloma venereum they are L1–3. Nucleotide variations within the ompA gene facilitate the identification of C. trachomatis genovars. This study describes a colorimetric multiplex PCR/RLB typing assay (mPCR-RLB) directed to the VD2 region of the ompA gene for general C. trachomatis positivity and the identification of 14 individual C. trachomatis genovars.
Methodology. The assay was validated by analysing 40 blinded samples that included reference strains of C. trachomatis genovars and other non-chlamydial micro-organisms that had been analysed previously using quantitative PCR (qPCR). Ninety clinical samples that had previously been found to be C. trachomatis-positive by qPCR were also evaluated using the mPCR-RLB assay.
Results. The mPCR-RLB assay showed 100 % agreement with the qPCR in the detection of C. trachomatis reference strains and no cross-reaction of non-chlamydial micro-organisms was observed. In the analysis of the chlamydial clinical samples, 97.8 % were C. trachomatis-positive by mPCR/RLB assay and there was a 96.6 % concordance with the qPCR at the group identification level and a 92.2 % concordance at the genovar level.
Conclusion. The mPCR-RLB assay is a rapid and sensitive methodology for the identification of C. trachomatis genovars associated with urogenital infections, trachoma or lymphogranuloma venereum diseases that can be implemented in clinical settings, helping to identify reinfections and treatment failures and establish the appropriate treatment course.
World Health Organization Global incidence and prevalence of selected curable sexually transmitted infections-2008; 2012www.who.int/iris/handle/10665/75181
ListerNA, FairleyCK, TabriziSN, GarlandS, SmithA. Chlamydia trachomatis serovars causing urogenital infections in women in Melbourne, Australia. J Clin Microbiol2005; 43:2546–2547 [View Article][PubMed]
PiñeiroL, LekuonaA, CillaG, LasaI, Martinez-GallardoLP et al. Prevalence of Chlamydia trachomatis infection in parturient women in Gipuzkoa, Northern Spain. Springerplus2016; 5:566 [View Article][PubMed]
SharmaA, SatpathyG, NayakN, TandonR, SharmaN et al. Ocular Chlamydia trachomatis infections in patients attending a tertiary eye care hospital in north India: a twelve year study. Indian J Med Res2012; 136:1004–1010[PubMed]
OssewaardeJM, RieffeM, de VriesA, Derksen-NawrockiRP, HooftHJ et al. Comparison of two panels of monoclonal antibodies for determination of Chlamydia trachomatis serovars. J Clin Microbiol1994; 32:2968–2974[PubMed]
YuanY, ZhangYX, WatkinsNG, CaldwellHD. Nucleotide and deduced amino acid sequences for the four variable domains of the major outer membrane proteins of the 15 Chlamydia trachomatis serovars. Infect Immun1989; 57:1040–1049[PubMed]
LanjouwE, OuburgS, de VriesHJ, StaryA, RadcliffeK et al. 2015 European guideline on the management of Chlamydia trachomatis infections. Int J STD AIDS2016; 27:333–348 [View Article][PubMed]
van der PolB, LiesenfeldO, WilliamsJA, TaylorSN, LillisRA et al. Performance of the cobas CT/NG test compared to the Aptima AC2 and Viper CTQ/GCQ assays for detection of Chlamydia trachomatis and Neisseria gonorrhoeae. J Clin Microbiol2012; 50:2244–2249 [View Article][PubMed]
DizeL, WestS, WilliamsJA, van der PolB, QuinnTC et al. Comparison of the Abbott m2000 RealTime CT assay and the Cepheid GeneXpert CT/NG assay to the Roche Amplicor CT assay for detection of Chlamydia trachomatis in ocular samples from Tanzania. J Clin Microbiol2013; 51:1611–1613 [View Article][PubMed]
GeislerWM, BlackCM, BandeaCI, MorrisonSG. Chlamydia trachomatis OmpA genotyping as a tool for studying the natural history of genital chlamydial infection. Sex Transm Infect2008; 84:541–544 [View Article][PubMed]
StevensMP, TwinJ, FairleyCK, DonovanB, TanSE et al. Development and evaluation of an ompA quantitative real-time PCR assay for Chlamydia trachomatis serovar determination. J Clin Microbiol2010; 48:2060–2065 [View Article][PubMed]
KapilR, PressCG, HwangML, BrownL, GeislerWM. Investigating the epidemiology of repeat Chlamydia trachomatis detection after treatment by using C. trachomatis OmpA genotyping. J Clin Microbiol2015; 53:546–549 [View Article][PubMed]
Gallo VauletL, EntrocassiC, PortuAI, CastroE, di BartolomeoS et al. High frequency of Chlamydia trachomatis mixed infections detected by microarray assay in South American samples. PLoS One2016; 11:e0153511 [View Article][PubMed]
PannekoekY, MorelliG, KusecekB, MorréSA, OssewaardeJM et al. Multi locus sequence typing of Chlamydiales: clonal groupings within the obligate intracellular bacteria Chlamydia trachomatis. BMC Microbiol2008; 8:42 [View Article][PubMed]
HerrmannB, IsakssonJ, RybergM, TångrotJ, SalehI et al. Global multilocus sequence type analysis of Chlamydia trachomatis strains from 16 countries. J Clin Microbiol2015; 53:2172–2179 [View Article]
VersteegB, BruistenSM, van der EndeA, PannekoekY. Does typing of Chlamydia trachomatis using housekeeping multilocus sequence typing reveal different sexual networks among heterosexuals and men who have sex with men?. BMC Infect Dis2016; 16:162 [View Article][PubMed]
PeuchantO, Le RoyC, HerrmannB, ClercM, BébéarC et al. MLVA subtyping of genovar E Chlamydia trachomatis individualizes the Swedish variant and anorectal isolates from men who have sex with men. PLoS One2012; 7:e31538 [View Article][PubMed]
MolanoM, MeijerCJ, MorréSA, PolR, van den BruleAJ. Combination of PCR targeting the VD2 of omp1 and reverse line blot analysis for typing of urogenital Chlamydia trachomatis serovars in cervical scrape specimens. J Clin Microbiol2004; 42:2935–2939 [View Article][PubMed]
SmithKS, HockingJS, ChenM, FairleyCK, McNultyA et al. Rationale and design of REACT: a randomised controlled trial assessing the effectiveness of home-collection to increase chlamydia retesting and detect repeat positive tests. BMC Infect Dis2014; 14:223 [View Article][PubMed]
QuintKD, van DoornLJ, KleterB, de KoningMN, van den MunckhofHA et al. A highly sensitive, multiplex broad-spectrum PCR-DNA-enzyme immunoassay and reverse hybridization assay for rapid detection and identification of Chlamydia trachomatis serovars. J Mol Diagn2007; 9:631–638 [View Article][PubMed]
XiongL, KongF, ZhouH, GilbertGL. Use of PCR and reverse line blot hybridization assay for rapid simultaneous detection and serovar identification of Chlamydia trachomatis. J Clin Microbiol2006; 44:1413–1418 [View Article][PubMed]
TwinJ, MooreEE, GarlandSM, StevensMP, FairleyCK et al.Chlamydia trachomatis genotypes among men who have sex with men in Australia. Sex Transm Dis2011; 38:279–285 [View Article][PubMed]
HerrmannB, IsakssonJ, RybergM, TångrotJ, SalehI et al. Global multilocus sequence type analysis of Chlamydia trachomatis strains from 16 Countries. J Clin Microbiol2015; 53:2172–2179 [View Article][PubMed]
GharsallahH, Frikha-GargouriO, SellamiH, BesbesF, ZnazenA et al.Chlamydia trachomatis genovar distribution in clinical urogenital specimens from Tunisian patients: high prevalence of C. trachomatis genovar E and mixed infections. BMC Infect Dis2012; 12:333 [View Article][PubMed]
ZhengHP, JiangLF, FangDY, XueYH, WuYA et al. Application of an oligonucleotide array assay for rapid detecting and genotyping of Chlamydia trachomatis from urogenital specimens. Diagn Microbiol Infect Dis2007; 57:1–6 [View Article][PubMed]
DanielewskiJA, PhillipsS, KongFYS, SmithKS, HockingJS et al. A snapshot of Chlamydia trachomatis genetic diversity using multilocus sequence type analysis in an Australian metropolitan setting. Eur J Clin Microbiol Infect Dis2017; 36:1297–1303 [View Article][PubMed]
de VriezeNH, van RooijenM, Schim van der LoeffMF, de VriesHJ. Anorectal and inguinal lymphogranuloma venereum among men who have sex with men in Amsterdam, The Netherlands: trends over time, symptomatology and concurrent infections. Sex Transm Infect2013; 89:548–552 [View Article][PubMed]
MachadoAC, BandeaCI, AlvesMF, JosephK, IgietsemeJ et al. Distribution of Chlamydia trachomatis genovars among youths and adults in Brazil. J Med Microbiol2011; 60:472–476 [View Article][PubMed]
HanY, YinYP, ShiMQ, ZhongMY, ChenSC et al. Difference in distribution of Chlamydia trachomatis genotypes among different provinces: a pilot study from four provinces in China. Jpn J Infect Dis2013; 66:69–71 [View Article][PubMed]
AnderssonP, HarrisSR, Seth SmithHM, HadfieldJ, O'NeillC et al.Chlamydia trachomatis from Australian Aboriginal people with trachoma are polyphyletic composed of multiple distinctive lineages. Nat Commun2016; 7:10688 [View Article][PubMed]
QinX, ZhengH, XueY, RenX, YangB et al. Prevalence of Chlamydia trachomatis genotypes in men who have sex with men and men who have sex with women using multilocus VNTR analysis-ompA typing in Guangzhou, China. PLoS One2016; 11:e0159658 [View Article][PubMed]
AnnanNT, SullivanAK, NoriA, NaydenovaP, AlexanderS et al. Rectal chlamydia–a reservoir of undiagnosed infection in men who have sex with men. Sex Transm Infect2009; 85:176–179 [View Article][PubMed]
Centers for Disease Control and Prevention Recommendations for the laboratory-based detection of Chlamydia trachomatis and Neisseria gonorrhoeae-2014. MMWR Recomm Rep2014; 63:1–19
MillmanK, BlackCM, JohnsonRE, StammWE, JonesRB et al. Population-based genetic and evolutionary analysis of Chlamydia trachomatis urogenital strain variation in the United States. J Bacteriol2004; 186:2457–2465 [View Article][PubMed]