TRiCit: A High-Throughput Approach to Detect Trichomonas vaginalis from ITS1 Amplicon Sequencing

Trichomoniasis, caused by Trichomonas vaginalis (TV), is the most common non-viral sexually transmitted infection (STI) worldwide, affecting over 174 million people annually and is frequently associated with reproductive co-morbidities. However, its detection can be time-consuming, subjective, and expensive for large cohort studies. This case–control study, conducted at the Mount Sinai Adolescent Health Center in New York City, involved 36 women with prevalent TV infections and 36 controls. The objective was to examine Internal Transcribed Spacer region-1 (ITS1) amplicon-derived communities for the detection of prevalent TV infections with the same precision as clinical microscopy and the independent amplification of the TV-specific TVK3/7 gene. DNA was isolated from clinician-collected cervicovaginal samples and amplified using ITS1 primers in a research laboratory. Results were compared to microscopic wet-mount TV detection of concurrently collected cervicovaginal samples and confirmed against TV-specific TVK3/7 gene PCR. The area under the receiver operating characteristics curve (AUC) for diagnosing TV using ITS1 communities was 0.92. ITS1 amplicons displayed an intra-class correlation coefficient (ICC) of 0.96 (95% CI: 0.93–0.98) compared to TVK3/7 PCR fragment testing. TV cases showed an increased risk of bacterial vaginosis (BV) compared to the TV-negative controls (OR = 8.67, 95% CI: 2.24–48.54, p-value = 0.0011), with no significant differences regarding genital yeast or chlamydia infections. This study presents a bioinformatics approach to ITS1 amplicon next-generation sequencing that is capable of detecting prevalent TV infections. This approach enables high-throughput testing for TV in stored DNA from large-scale epidemiological studies.

[1]  R. Burk,et al.  molBV reveals immune landscape of bacterial vaginosis and predicts human papillomavirus infection natural history , 2022, Nature communications.

[2]  G. Ramirez,et al.  Bacterial Vaginosis and Its Association With Incident Trichomonas vaginalis Infections: A Systematic Review and Meta-Analysis , 2021, Sexually transmitted diseases.

[3]  K. Workowski,et al.  Sexually Transmitted Infections Treatment Guidelines, 2021 , 2021, MMWR. Recommendations and reports : Morbidity and mortality weekly report. Recommendations and reports.

[4]  P. Rappelli,et al.  Impact of Symbiosis Between Trichomonas vaginalis and Mycoplasma hominis on Vaginal Dysbiosis: A Mini Review , 2020, Frontiers in Cellular and Infection Microbiology.

[5]  P. Gajer,et al.  VALENCIA: a nearest centroid classification method for vaginal microbial communities based on composition , 2020, Microbiome.

[6]  Asosiasi Internasional Maha Guru Ching Hai.  Vegan , 2020, Definitions.

[7]  N. Low,et al.  Chlamydia, gonorrhoea, trichomoniasis and syphilis: global prevalence and incidence estimates, 2016 , 2019, Bulletin of the World Health Organization.

[8]  J. Dowd,et al.  Evaluation of Oral Cavity DNA Extraction Methods on Bacterial and Fungal Microbiota , 2019, Scientific Reports.

[9]  R. Burk,et al.  Novel ITS1 Fungal Primers for Characterization of the Mycobiome , 2017, mSphere.

[10]  J. Schwebke,et al.  Pathogenesis of Bacterial Vaginosis: Discussion of Current Hypotheses. , 2016, The Journal of infectious diseases.

[11]  Paul J. McMurdie,et al.  DADA2: High resolution sample inference from Illumina amplicon data , 2016, Nature Methods.

[12]  P. Kissinger.  Epidemiology and Treatment of Trichomoniasis , 2015, Current Infectious Disease Reports.

[13]  Susan Holmes,et al.  phyloseq: An R Package for Reproducible Interactive Analysis and Graphics of Microbiome Census Data , 2013, PloS one.

[14]  I. Schmitt,et al.  Comparison of ITS1 and ITS2 rDNA in 454 sequencing of hyperdiverse fungal communities , 2013 .

[15]  V. Shankar,et al.  Cervical, Anal and Oral HPV in an Adolescent Inner-City Health Clinic Providing Free Vaccinations , 2012, PloS one.

[16]  William A. Walters,et al.  Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms , 2012, The ISME Journal.

[17]  J. Klausner,et al.  Bacterial Vaginosis and Risk for Trichomonas vaginalis Infection: A Longitudinal Analysis , 2011, Sexually transmitted diseases.

[18]  Xavier Robin,et al.  pROC: an open-source package for R and S+ to analyze and compare ROC curves , 2011, BMC Bioinformatics.

[19]  Andy F. S. Taylor,et al.  The UNITE database for molecular identification of fungi--recent updates and future perspectives. , 2010, The New phytologist.

[20]  P. Qian,et al.  Conservative Fragments in Bacterial 16S rRNA Genes and Primer Design for 16S Ribosomal DNA Amplicons in Metagenomic Studies , 2009, PloS one.

[21]  J. Hardin,et al.  Treatment of trichomoniasis in pregnancy and preterm birth: an observational study. , 2009, Journal of women's health.

[22]  D. Mabey,et al.  Global epidemiology and control of Trichomonas vaginalis , 2008, Current opinion in infectious diseases.

[23]  E. Korkmaz,et al.  Association between trichomoniasis and bacterial vaginosis: examination of 600 cervicovaginal smears. , 2005, Central European journal of public health.

[24]  R. Ballard,et al.  Evaluation of Xenostrip-Tv, a Rapid Diagnostic Test for Trichomonas vaginalis Infection , 2004, Journal of Clinical Microbiology.

[25]  T. Quinn,et al.  Diagnosis of Trichomonas vaginalisInfection by PCR Using Vaginal Swab Samples , 1998, Journal of Clinical Microbiology.

[26]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[27]  OUP accepted manuscript , 2022, Clinical Infectious Diseases.

[28]  M. Kuhn.  The caret Package , 2007 .

[29]  P. Legendre,et al.  vegan : Community Ecology Package. R package version 1.8-5 , 2007 .

[30]  G. Garber,et al.  The laboratory diagnosis of Trichomonas vaginalis. , 2005, The Canadian journal of infectious diseases & medical microbiology = Journal canadien des maladies infectieuses et de la microbiologie medicale.

[31]  M. Laga,et al.  Non-ulcerative sexually transmitted diseases as risk factors for HIV-1 transmission in women: results from a cohort study. , 1993, AIDS.