Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Detection of sexually transmitted disease–causing pathogens from direct clinical specimens with the multiplex PCR-based STD Direct Flow Chip Kit

  • 141 Accesses


Pathogens causing sexually transmitted diseases (STDs) include viruses, bacteria, and parasites. The ability to rapidly and efficiently detect these pathogens in a single reaction still remains a health challenge. The aim of this study was to evaluate the clinical reliability and accuracy of the STD Direct Flow Chip Kit (Vitro, IVD-EC approved), which can simultaneously detect up to 9 different species of STD pathogens at once. This kit enables direct analysis—direct-PCR—of clinical specimens (urine, semen, endocervical, urethral, nasopharyngeal, and perianal swabs) without DNA purification for the following pathogens: Chlamydia trachomatis (serovars A-K and L1-L3), Haemophilus ducreyi, Herpes Simplex Virus (Types I and II), Mycoplasma genitalium, Mycoplasma hominis, Neisseria gonorrhoeae, Treponema pallidum, Trichomonas vaginalis, and Ureaplasma. The Anyplex™ II STI-7 Detection Kit (Seegene, IVD-EC) was used as the reference’s method. Existing discordances were resolved using either a third molecular assay or DNA sequencing. Clinical performance was evaluated at two different stages: (i) from purified DNA of three hundred and fifty-eight clinical specimens with a diagnostic sensitivity (SE) and specificity (SP) of 99.4% and 100%, respectively, and an agreement of 99% (kappa index, κ = 0.97) with the reference’s method and; (ii) by direct-PCR from six hundred and thirty-three specimens rendering SE, SP, and agreement values of 98.4%, 99.9%, and 98.0% (κ = 0.95), respectively. The STD Direct Flow Chip Kit constitutes a promising alternative to routine procedures in diagnostic, allowing direct analysis of specimens and enabling the detection of a broad panel of pathogens.

This is a preview of subscription content, log in to check access.


  1. 1.

    World Health Organization (2016) Report on global sexually transmitted infection surveillance 2015. WHO. Available in: http://www.who.int/iris/handle/10665/249553. Accessed May 2018

  2. 2.

    Gewirtzman A, Bobrick L, Conner K, Tyring SK (2011) Epidemiology of sexually transmitted infections. In: Gorss G, Tyring SK (eds) Sexually transmitted infections and sexually transmitted diseases. Springer, New York

  3. 3.

    Global incidence and prevalence of selected curable sexually transmitted infections 2008. World Health Organization, Geneva, 2012

  4. 4.

    Taylor-Robinson D, Jensen JS (2011) Mycoplasma genitalium: from chrysalis to multicolored butterfly. Clin Microbiol Rev 24:498–514

  5. 5.

    Sethi S, Singh G, Samanta P, Sharma M (2012) Mycoplasma genitalium: an emerging sexually transmitted pathogen. Indian J Med Res 136:942–955

  6. 6.

    Larsen B, Hwang J (2010) Mycoplasma, Ureaplasma and adverse pregnancy outcomes: a fresh look. Infect Dis Obstet Gynecol 2010:1–7

  7. 7.

    Levett PN, Brandt K, Olenius K, Brown C, Montgomery K, Horsman GB (2008) Evaluation of three automated nucleic acid amplification Systems for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae in first-void urine specimens. J Clin Microbiol 46:2109–2111

  8. 8.

    Cheng A, Qian Q, Kirby JE (2011) Evaluation of the Abbott RealTime CT/NG assay in comparison to the Roche Cobas Amplicor CT/NG assay. J Clin Microbiol 49:1294–1300

  9. 9.

    Shipitsyna E, Zolotoverkhaya E, Chen CY, Chi KH, Grigoryev A, Savicheva A et al (2013) Evaluation of polymerase chain reaction assays for the diagnosis of Trichomonas vaginalis infection in Russia. J Eur Acad Dermatol Venereol 27:e217–e223

  10. 10.

    Choe HS, Lee DS, Lee S-J, Hong S-H, Park DC, Lee M-K et al (2013) Performance of Anyplex™ II multiplex real-time PCR for the diagnosis of seven sexually transmitted infections: comparison with currently available methods. Int J Infect Dis 17:1134–1140

  11. 11.

    Berçot B, Amarsy R, Goubard A, Aparicio C, Loeung HU, Segouin C et al (2015) Assessment of co-infection of sexually transmitted pathogen microbes by use of the Anyplex™ II STI-7 Molecular Kit. J Clin Microbiol 53:991–993

  12. 12.

    Fernández G, Martró E, González V, Saludes V, Bascuñana E, Marcó C et al (2016) Usefulness of a novel multiplex real-time PCR assay for the diagnosis of sexually-transmitted infections. Enferm Infecc Microbiol Clin 34:471–476

  13. 13.

    Wu HN, Nakura Y, Motooka D, Nakamura S, Nishiumi F, Ishino S et al (2014) Complete genome sequence of Ureaplasma parvum Serovar 3 strain SV3F4, isolated in Japan. Genome Announc 2:e00256–e00214

  14. 14.

    Mao M, Liu HL (2015) Genetic diversity of Trichomonas vaginalis clinical isolates from Henan province in Central China. Pathog Glob Health 109:242–246

  15. 15.

    Pillay A, Katz SS, Abrams AJ, Ballard RC, Simpson SV, Taleo F et al (2016) Complete genome sequences of 11 Haemophilus ducreyi isolates from children with cutaneous lesions in Vanuatu and Ghana. Genome Announc 4:e00459–e00416

  16. 16.

    Cohen J (1960) A coefficient of agreement for nominal scales. Educ Psychol Meas 20:37–46

  17. 17.

    Bergeri I, Michel R, Boutin J-B (2002) Pour tout savoir ou presque sur le coefficient Kappa. Med Trop 62:634–636

  18. 18.

    Ghazi Saeedi K, Fateminasab F, Vatani SH, Anzimi Y, Bakhshandenosrat S, Mohamadi M (2008) Compare two methods prostatic massage and urine initial drop sample in isolates of Mycoplasma hominis and Ureaplasma urealyticum in urinary tract. J Lab Med 2:15–18

  19. 19.

    Kong F, Ma Z, James G, Gordon S, Gilbert GL (2000) Species identification and subtyping of Ureaplasma parvum and Ureaplasma urealyticum using PCR-based assays. J Clin Microbiol 38:1175–1179

  20. 20.

    Ollikainen J, Heiskanen-Kosma T, Korppi M, Katila ML, Heinonen K (1998) Clinical relevance of Ureaplasma urealyticum colonization in preterm infants. Acta Paediatr 87:1075–1078

  21. 21.

    Beeton ML, Spiller OB (2017) Antibiotic resistance among Ureaplasma spp. isolates: cause for concern? J Antimicrob Chemother 72:330–337

  22. 22.

    Horner P, Donders G, Cusini M, Gomberg M, Jensen JS, Unemo M (2018) Should we be testing for urogenital Mycoplasma hominis, Ureaplasma parvum and Ureaplasma urealyticum in men and women? – a position statement from the European STI Guidelines Editorial Board. J Eur Acad Dermatol Venereol 32:1845–1851

  23. 23.

    González-Beiras C, Marks M, Chen CY, Roberts S, Mitjà O (2016) Epidemiology of Haemophilus ducreyi infections. Emerg Infect Dis 22:1–8

  24. 24.

    Bruisten SM, Cairo I, Fennema H, Pijl A, Buimer M, Peerbooms PG et al (2001) Diagnosing genital ulcer disease in a clinic for sexually transmitted diseases in Amsterdam, the Netherlands. J Clin Microbiol 39:601–605

  25. 25.

    Herring A, Ballard R, Mabey D, Peeling RW (2006) Evaluation of rapid diagnostic tests: chlamydia and gonorrhoea. Nat Rev Microbiol 4:S41–S48

  26. 26.

    Banoo S, Bell D, Bossuyt P, Herring A, Mabey D, Poole F et al (2010) Evaluation of diagnostic tests for infectious diseases: general principles. Nat Rev Microbiol 8:S17–S12

Download references


The results published here are part of the thesis´s work, of the PhD candidate Adolfo de Salazar, in the Biomedicine Doctoral Program of the University of Granada.

Author information

Correspondence to Federico Garcia.

Ethics declarations

Vitro provided kits and reagents for testing. The Ethics Committee of the Hospital San Cecilio approved the study protocol. All the clinical specimens were received in the Microbiology laboratory for routine diagnostics and anonymized prior testing.

Conflict of interest

The authors declare that they have no conflicts of interest.

Additional information

Publisher's note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Barrientos-Durán, A., de Salazar, A., Alvarez-Estévez, M. et al. Detection of sexually transmitted disease–causing pathogens from direct clinical specimens with the multiplex PCR-based STD Direct Flow Chip Kit. Eur J Clin Microbiol Infect Dis 39, 235–241 (2020). https://doi.org/10.1007/s10096-019-03686-w

Download citation


  • Clinical specimens
  • Direct analysis
  • DNA: DNA hybridization
  • Multiplex-PCR based
  • Sexually transmitted diseases