Folia Parasitologica 71:021 (2024) | DOI: 10.14411/fp.2024.021

High frequency of point mutations in the nitroreductase 4 and 6 genes of Trichomonas vaginalis associated with metronidazole resistance

Isabel Iturrieta-González ORCID...1, 2, Pamela Olivares-Ferretti ORCID...1, Alejandro Hidalgo ORCID...1, 2, Fabiola Zambrano ORCID...1, 2, Ximena Ossa ORCID...3, Flery Fonseca-Salamanca ORCID...1, 2, Angélica Melo ORCID...1, 4, *
1 Centre of Excellence in Translational Medicine, Nucleus of Scientific and Technological Bioresources (CEMT-BIOREN), Faculty of Medicine. Universidad de La Frontera, Temuco, Chile;
2 Department of Preclinical Sciences, Faculty of Medicine, Universidad de La Frontera. Temuco, Chile;
3 Public Health Department, Centre of Excellence Training, Research and Management for Evidence-Based Health (CIGES), Universidad de La Frontera, Temuco, Chile;
4 Department of Pathological Anatomy, Faculty of Medicine. Universidad de La Frontera. Temuco, Chile *Address for correspondence: Angélica Melo Angermeyer. Universidad de La Frontera, Faculty of Medicine, Edificio Biociencias, Av. Alemania 0458 Temuco, Chile. E-mail: angelica.melo@ufrontera.cl; ORCID-iD: 0000-0002-3576-1745

Trichomoniasis, a globally distributed sexually transmitted infection, is caused by the urogenital parasite Trichomonas vaginalis Donné, 1836 affecting both women and men. The treatment of choice is metronidazole (MTZ). In the present study, 15 samples of vaginal discharge and urine were analysed by sequencing nitroreductase genes (ntr4 and ntr6). An in silico model was structured to illustrate the location of point mutations (PM) in the protein. The ntr4 gene presented four PMs: G76C (10/10), C213G (9/10), C318A (5/10) and G424A (1/10), while the ntr6 gene had eight PMs; G593A (13/13) the most frequent, G72T and G627C, both in 8/13. The PM C213G and A438T generated a stop codon causing a truncated nitroreductase 4 and 6 protein. Docking analysis demonstrated that some models had a decrease in binding affinity to MTZ (p < 0.0001). A high frequency of mutations was observed in the samples analysed that could be associated with resistance to MTZ in Chile.

Keywords: genital infection, trichomoniasis, refractory treatment, protein polymorphism, dysfunctional enzyme, clinical samples.

Received: July 18, 2023; Revised: August 21, 2024; Accepted: September 17, 2024; Published online: November 7, 2024  Show citation

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Iturrieta-González, I., Olivares-Ferretti, P., Hidalgo, A., Zambrano, F., Ossa, X., Fonseca-Salamanca, F., & Melo, A. (2024). High frequency of point mutations in the nitroreductase 4 and 6 genes of Trichomonas vaginalis associated with metronidazole resistance. Folia Parasitologica, 71, Article 2024.021. https://doi.org/10.14411/fp.2024.021
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    References

    1. Appaneal H.J., Caffrey A.R., LaPlante K.L. 2019: What is the role for metronidazole in the treatment of Clostridium difficile infection? Results from a national cohort study of veterans with initial mild disease. Clin. Infect. Dis. 69: 1288-1295. Go to original source... Go to PubMed...
    2. Armstrong E., Hemmerling A., Miller S., Burke K.E., Newmann S.J., Morris S.R., Reno H., Huibner S., Kulikova M., Liu R., Crawford E.D., Castaneda G.R., Nagelkerke N., Bryan Coburn B., Cohen C.R., Rupert Kaul R. 2022: Metronidazole treatment rapidly reduces genital inflammation through effects on bacterial vaginosis associated bacteria rather than lactobacilli. J. Clin. Invest. 132: e152930. Go to original source... Go to PubMed...
    3. Belfort I.K.P., Cunha A.P.A., Mendes F.P.B., Galvao-Moreira L.V., Lemos R.G., de Lima Costa L.H., Monteiro P., Ferreira M.B., Bastos dos Santos G.R., Costa J.L., de Sa Ferreira A., Brito L.G.O., Brito L.M.O., Vidal F.C.B., Monteiro S.C.M. 2021: Trichomonas vaginalis as a risk factor for human papillomavirus: a study with women undergoing cervical cancer screening in a northeast region of Brazil. BMC Women's Health 21: 174. Go to original source... Go to PubMed...
    4. Bradic M., Carlton J.M. 2018: Does the common sexually transmitted parasite Trichomonas vaginalis have sex? PLoS Pathogens 14: e1006831. Go to original source... Go to PubMed...
    5. Bradic M., Warring S.D., Tooley G.E., Scheid P., Secor W.E., Land K.M., Huang P.J., Chen T.W., Lee C.C., Tang P., Sullivan S.A., Carlton J.M. 2017: Genetic indicators of drug resistance in the highly repetitive genome of Trichomonas vaginalis. Genome Biol. Evol. 9: 1658-1672. Go to original source... Go to PubMed...
    6. Conrad M., Zubacova Z., Dunn L.A., Upcroft J., Sullivan S.A., Tachezy J., Carlton J.M. 2011: Microsatellite polymorphism in the sexually transmitted human pathogen Trichomonas vaginalis indicates a genetically diverse parasite. Mol. Biochem. Parasitol. 175: 30-38. Go to original source... Go to PubMed...
    7. Dingsdag S.A., Hunter N. 2018: Metronidazole: an update on metabolism, structure- cytotoxicity and resistance mechanisms. J. Antimicrob. Chemother. 73: 265-279. Go to original source... Go to PubMed...
    8. Dunne R.L., Dunn L.A., Upcroft P., O'Donoghue P.J., Upcroft J.A. 2003: Drug resistance in the sexually transmitted protozoan Trichomonas vaginalis. Cell Res. 13: 239-249. Go to original source... Go to PubMed...
    9. Eberhardt J., Santos-Martins D., Tillack A.F., Forli S. 2021: AutoDock Vina 1.2.0: New docking methods, expanded force field, and Python bindings. J. Chem. Inf. Model. 61: 3891-3898. Go to original source... Go to PubMed...
    10. Edgar R.C. 2004: MUSCLE: Multiple sequence alignment with high accuracy and high throughput. Nucl. Acids Res. 32: 1792-1797. Go to original source... Go to PubMed...
    11. Edwards T., Burke P., Smalley H., Hobbs G. 2016: Trichomonas vaginalis: clinical relevance, pathogenicity and diagnosis. Crit. Rev. Microbiol. 42: 406-417.
    12. Garber G.E. 2005: The laboratory diagnosis of Trichomonas vaginalis. Can. J. Infect. Dis. Med. Microbiol. 16: 35-38. Go to original source... Go to PubMed...
    13. Gwenin C.D., Kalaji M., Williams P.A., Kay C.M. 2010: A kinetic analysis of three modified novel nitroreductases. Biodegradation 22: 463-474. Go to original source... Go to PubMed...
    14. Halgren T.A. 1996: Merck molecular force field. V. Extension of MMFF94 using experimental data, additional computational data, and empirical rules. J. Comput. Chem. 17: 616-641. Go to original source...
    15. Jumper J., Evans R., Pritzel A., Green T., Figurnov M., Ronneberger O., Tunyasuvunakool K., Bates R., ®idek A., Potapenko A., Bridgland A., Meyer C., Kohl S.A.A., Ballard A.J., Cowie A., Romera-Paredes B., Nikolov S., Jain R., Adler J., Back T., Petersen S., Reiman D., Clancy E., Zielinski M., Steinegger M., Pacholska M., Berghammer T., Bodenstein S., Silver D., Vinyals O., Senior A.W., Kavukcuoglu K., Kohli P., Hassabis D. 2021: Highly accurate protein structure prediction with AlphaFold. Nature 596: 583-589. Go to original source... Go to PubMed...
    16. Kengne P., Veas F., Vidal N., Rey J.L., Cuny G. 1994: Trichomonas vaginalis: repeated DNA target for highly sensitive and specific polymerase chain reaction diagnosis. Cell. Mol. Biol. 40: 819-831.
    17. Kim S., Thiessen P.A., Bolton E.E., Chen J., Fu G., Gindulyte A., Han L., He J., He S., Shoemaker B.A., Wang J., Yu B., Zhang J., Bryant S.H. 2016: PubChem substance and compound databases. Nucl. Acids Res. 44: D1202-D1213. Go to original source... Go to PubMed...
    18. Kirkcaldy R.D., Augostini P., Asbel L.E., Bernstein K.T., Kerani R.P., Mettenbrink C.J., Pathela P., Schwebke J.R., Secor W.E., Workowski K.A., Davis D., Braxton J., Weinstock H.S. 2012: Trichomonas vaginalis antimicrobial drug resistance in 6 US cities, STD surveillance network, 2009-2010. Emerg. Infect. Dis. 18: 939-943.
    19. Kissinger P.J., Gaydos C.A., Sena A.C., Scott McClelland R., Soper D., Secor W.E., Legendre D., Workowski K.A., Muzny C.A. 2022: Diagnosis and management of Trichomonas vaginalis: summary of evidence reviewed for the 2021. Centers For Disease Control and Prevention of Sexually Transmitted Infections Treatment Guidelines. Clin. Infect. Dis. 74: S152-S161.
    20. Kusdian G., Gould S.B. 2014: The biology of Trichomonas vaginalis in the light of urogenital tract infection. Mol. Biochem. Parasitol. 198: 92-99. Go to original source... Go to PubMed...
    21. Liochev S.I., Hausladen A., Fridovich I. 1999: Nitroreductase A is regulated as a member of the soxRS regulon of Escherichia coli. Proc. Natl. Acad. Sci. USA. 96: 3537-3539. Go to original source... Go to PubMed...
    22. Mammen-Tobin A., Wilson J.D. 2005: Management of metronidazole-resistant Trichomonas vaginalis - a new approach. Int. J. STD AIDS 16: 488-490. Go to original source... Go to PubMed...
    23. Marques-Silva M., Lisboa C., Gomes N., Rodrigues A.G. 2021: Trichomonas vaginalis and growing concern over drug resistance: a systematic review. J. Eur. Acad. Dermatol. Venereol. 35: 2007-2021. Go to original source... Go to PubMed...
    24. McClelland R.S., Sangare L., Hassan W.M., Lavreys L., Mandaliya K., Kiarie J., Ndinya-Achola J., Jaoko W., Baeten J.M. 2007: Infection with Trichomonas vaginalis increases the risk of HIV-1 acquisition. J. Infect. Dis. 195: 698-702. Go to original source... Go to PubMed...
    25. Melo A., Ossa X., Fetis G., Lazo L., Bustos L., Fonseca-Salamanca F. 2021: Concordance between clinical and laboratory diagnosis of abnormal vaginal discharge in Chilean women. Rev. Bras. Ginecol. Obstet. 43: 600-607. Go to original source... Go to PubMed...
    26. Mirdita M., Schutze K., Moriwaki Y., Heo L., Ovchinnikov S., Steinegger M. 2022: ColabFold: making protein folding accessible to all. Nat. Methods 19: 679-682. Go to original source... Go to PubMed...
    27. Morikawa E., Mudau M., Olivier D., de Vos L., Joseph Davey D., Price C., McIntyre J.A., Peters R.P., Klausner J.D., Medina-Marino A. 2018: Acceptability and feasibility of integrating point-of-care diagnostic testing of sexually transmitted infections into a South African antenatal care program for HIV-infected pregnant women. Infect. Dis. Obstet. Gynecol. 2018: 3946862. Go to original source... Go to PubMed...
    28. Mtshali A., Ngcapu S., Govender K., Sturm A.W., Moodley P., Joubert B.C. 2022: In vitro effect of 5-nitroimidazole drugs against Trichomonas vaginalis clinical isolates. Microbiol. Spectr. 10: e0091222. Go to original source... Go to PubMed...
    29. Mzenda T. 2021: Identification of mutations in genes associated with metronidazole resistance and susceptibility in Trichomonas vaginalis. PhD thesis, University of KwaZulu-Natal, 68 pages.
    30. Ngobese B., Singh R., Han K.S.S., Tinarwo P., Mabaso N., Abbai N.S. 2022: Detection of metronidazole resistance in Trichomonas vaginalis using uncultured vaginal swabs. Parasitol. Res. 121: 2421-2432. Go to original source... Go to PubMed...
    31. O'Boyle N., Banck M., James C.A., Morley C., Vandermeersch T., Hutchison G.R. 2011: Open Babel: an open chemical toolbox. J. Cheminform. 3: 33. Go to original source... Go to PubMed...
    32. Pal D., Banerjee S., Cui J., Schwartz A., Ghosh S.K., Samuelson J. 2009: Giardia, Entamoeba, and Trichomonas enzymes activate metronidazole (nitroreductases) and inactivate metronidazole (nitroimidazole reductases). Antimicrob. Agents Chemother. 53: 458-464. Go to original source... Go to PubMed...
    33. Paulish-Miller T.E., Augostini P., Schuyler J.A., Smith W.L., Mordechai E., Adelson M.E., Gygax S.E., Secor W.E., Hilbert D.W. 2014: Trichomonas vaginalis metronidazole resistance is associated with single nucleotide polymorphisms in the nitroreductase genes ntr4Tv and ntr6Tv. Antimicrob. Agents Chemother. 58: 2938-2943. Go to original source... Go to PubMed...
    34. Penning T., Su A.L., El-Bayoumy K. 2022: Nitroreduction: a critical metabolic pathway for drugs, environmental pollutants, and explosives. Chem. Res. Toxicol. 35: 1747-1765. Go to original source... Go to PubMed...
    35. Roldan M.D., Perez-Reinado E., Castillo F., Moreno-Vivian C. 2008: Reduction of polynitroaromatic compounds: the bacterial nitroreductases. FEMS Microbiol. Rev. 32: 474-500. Go to original source... Go to PubMed...
    36. Saiki R.K., Gelfand D.H., Stoffel S., Scharf S.J., Higuchi R., Horn G.T., Mullis K.B., Erlich H.A. 1988: Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239: 487-491. Go to original source...
    37. Schilcrot F.F., Cabello E.S., Pérez S.B., Díaz R.B., Pinochet A.H. 2016: Norma de Profilaxis, Diagnóstico y Tratamiento de las Infecciones de Transmisión Sexual (ITS). Ministerio de Salud, Chile, 112 pp.
    38. Schwebke J.R., Barrientes F.J. 2006: Prevalence of Trichomonas vaginalis isolates with resistance to metronidazole and tinidazole. Antimicrob. Agents Chemother. 50: 4209-4210. Go to original source... Go to PubMed...
    39. Sisson G., Goodwin A., Raudonikiene A., Hughes N.J, Mukhopadhyay A.K., Berg D.E., Hoffman P.S. 2002: Enzymes associated with reductive activation and action of nitazoxanide, nitrofurans, and metronidazole in Helicobacter pylori. Antimicrob. Agents Chemother. 46: 2116-2123. Go to original source... Go to PubMed...
    40. Spotin A., Eghtedar S.T., Shahbazi A., Salehpour A., Sarafraz S., Shariatzadeh S.A., Mahami-Oskouei M. 2016: Molecular characterization of Trichomonas vaginalis strains based on identifying their probable variations in asymptomatic patients. Iran J. Parasitol. 11: 507-514.
    41. Tamura K., Stecher G., Peterson D., Filipski A., Kumar S. 2013: MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol. Biol. Evol. 30: 2725-2729. Go to original source... Go to PubMed...
    42. Thomas C., Gwenin C.D. 2021: The role of nitroreductases in resistance to nitroimidazoles. Biology 10: 388. Go to original source... Go to PubMed...
    43. Thompson J.D., Higgins D.G., Gibson T.J. 1994: CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucl. Acids Res. 22: 4673-4680. Go to original source... Go to PubMed...
    44. Van Gerwen O.T., Camino A.F., Sharma J., Kissinger P.J., Muzny C.A. 2021: Epidemiology, natural history, diagnosis, and treatment of Trichomonas vaginalis in men. Clin. Infect. Dis. 73: 1119-1124. Go to original source... Go to PubMed...
    45. Waters L.J., Dave S.S., Deayton J.R., French P.D. 2005: Recalcitrant Trichomonas vaginalis infection - a case series. Int. J. STD AIDS 16: 505-509. Go to original source... Go to PubMed...
    46. Wilkinson S.R., Bot C., Kelly J.M., Hall B.S. 2011: Trypanocidal activity of nitroaromatic prodrugs: current treatments and future perspectives. Curr. Top. Med. Chem. 11: 2072-2084. Go to original source... Go to PubMed...
    47. Yang H.Y., Su R.Y., Chung C.H., Huang K.Y., Lin H.A., Wang J.Y., Chen C.C., Chien W.C., Lin H.C. 2022: Association between trichomoniasis and prostate and bladder diseases: a population-based case - control study. Sci. Rep. 12: 15358. Go to original source... Go to PubMed...
    48. Yarlett N., Yarlett N.C., Lloyd D. 1986: Ferredoxin-dependent reduction of nitroimidazole derivatives in drug-resistant and susceptible strains of Trichomonas vaginalis. Biochem. Pharmacol. 35: 1703-1708. Go to original source... Go to PubMed...
    49. Zhang Z., Kang L., Wang W., Zhao X., Li Y., Xie Q., Wang S., He T., Li H., Xiao T., Chen Y., Zuo S., Kong L., Li P., Li X. 2018: Prevalence and genetic diversity of Trichomonas vaginalis clinical isolates in a targeted population in Xinxiang City, Henan Province, China. Parasit. Vectors 11: 124. Go to original source... Go to PubMed...
    50. Zhang Z., Li Y., Lu H., Li D., Zhang R., Xie X., Guo L., Hao L., Tian X., Yang Z., Wang S., Mei X. 2022: A systematic review of the correlation between Trichomonas vaginalis infection and infertility. Acta Trop. 236: 106693. Go to original source... Go to PubMed...

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