Opportunities and pitfalls of molecular testing for detecting sexually transmitted pathogens

Trembizki, Ella; Costa, Anna‐Maria; Tabrizi, Sepehr N.; Whiley, David M.; Twin, Jimmy

doi:10.1097/pat.0000000000000239

Cited by 22 publications

(19 citation statements)

References 96 publications

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“…However, NAATs have shortcomings, recently reviewed by Trembizki et al . [8], to be kept in mind when interpreting test results. One of the main disadvantages of NAAT assays is that the available target DNA is amplified without discriminating between DNA originating from viable or non-viable CT.…”

Section: Introductionmentioning

confidence: 99%

Viability-PCR Shows That NAAT Detects a High Proportion of DNA from Non-Viable Chlamydia trachomatis

et al. 2016

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ObjectivesAccording to the current guidelines for laboratory diagnosis of sexually transmitted infections (STIs), nucleic acid amplification tests (NAATs) are the preferred diagnostic method for Chlamydia trachomatis (CT) infections. However, NAATs amplify the available target DNA without discriminating between DNA originating from viable or non-viable CT. Assessing CT viability will provide more insights in the clinical and public health relevance of a CT positive test result. The aim of this study was to technically validate and implement viability-PCR (V-PCR) to asses CT viability.MethodsTechnical validation of V-PCR was performed by the assessment of predefined viability ratios of CT. Samples were subjected to V-PCR which consisted of propidium monoazide (PMA) treatment prior to DNA extraction followed by quantitative PCR (qPCR) targeting the ompA gene for the detection of CT DNA. Finally, V-PCR was applied to vaginal swabs of 50 CT positive patients, as indicated by routine NAAT, collected at our outpatient STD clinics before antimicrobial treatment.ResultsTechnical validation of V-PCR showed that PMA treatment of heat-inactivated CT culture resulted in an almost complete loss of qPCR signal. PMA treated samples of the fresh viable CT culture showed no marked reduction of PCR signal, indicating that all DNA from viable CT could be detected. Applying V-PCR to clinical samples showed that in 36% of samples (18/50) less than 1% of CT DNA originated from viable bacteria.ConclusionsV-PCR showed to be a fast and easy method to assess CT viability in clinical samples, without the need of traditional challenging cell culture methods. Furthermore, V-PCR results of clinical samples have indicated that a substantial amount of the amplified CT DNA originated from non-viable cells. Although results might be influenced by cell death during transport, this study suggests that there is a potential overestimation of quantitative CT positivity by currently used NAATs.

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Section: Introductionmentioning

confidence: 99%

Viability-PCR Shows That NAAT Detects a High Proportion of DNA from Non-Viable Chlamydia trachomatis

et al. 2016

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“…The increased sensitivity of NAATs makes them particularly suitable for screening, enabling accurate diagnosis of both symptomatic and asymptomatic gonococcal infections, which is critical to the control of the disease [24]. Second, specimens collected for NAATs do not require the organism to be viable for detection and so require less stringent transport conditions compared with those collected for bacterial culture.…”

Section: Molecular Detection Methodsmentioning

confidence: 99%

Diagnosis And Antimicrobial Treatment Of Bacterial Of Neisseria Gonorrhea Infections: Update Review Article

Shehabi¹,

Hamzé²

2017

int. arab. j antimicrob. agents

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Sexually transmitted infections (STIs) are caused by a wide spectrum of bacteria, viruses and parasites. These agents can be easily transmitted by direct genital or oral sexual contact. Recently, the World Health Organization (WHO), reported that more than 1 million STIs are acquired every day worldwide. Each year, there are an estimated 357 million new infections with one of the following four STIs: chlamydia, gonorrhea, syphilis and trichomoniasis. There are few studies and official reports published on the prevalence of STIs in most Arab countries. However, a few recent studies showed indicate an increased prevalence of certain STIs in some Arab countries, and few data is available on their antimicrobial susceptibility.

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“…4,5 Conventional diagnostic algorithms for STIs include cell culture, microscopic examination, enzyme immunoassay, and other serological investigation techniques. [6][7][8] The diagnosis of STIs by culture or serology remains a challenge. 9 Especially for herpes simplex virus and Treponema pallidum detection, cell culture shows low-sensitivity and long turnaround time, making this assay clinically useless.…”

Section: Introductionmentioning

confidence: 99%

<p>Simultaneous detection of eleven sexually transmitted agents using multiplexed PCR coupled with MALDI-TOF analysis</p>

Xiu¹,

Zhang²,

Li³

et al. 2019

IDR

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PurposeSexually transmitted infections (STIs), representing a major global health problem, are caused by different microbes, including bacteria, viruses, and protozoa. Unfortunately, infections of different sexually transmitted pathogens often present similar clinical symptoms, so it is almost impossible to distinguish them clinically. Therefore, the aim of the current study was to develop a sensitive, multitarget, and high-throughput method that can detect various agents responsible for STIs.MethodsWe developed and tested a 23-plex PCR coupled with matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) assay (sexually transmitted infection-mass spectrometry, STI-MS) that simultaneously targets 11 different agents, including 8 most common clinical pathogens related to STIs (HSV-1, HSV-2, Neisseria gonorrhoeae, Chlamydia trachomatis, Treponema pallidum, Trichomonas vaginalis, Mycoplasma genitalium, and Haemophilus ducreyi) and 3 controversial microorganisms as pathogens (Mycoplasma hominis, Ureaplasma urealyticum, and Ureaplasma parvum).ResultsThe results showed that the STI-MS approach can accurately detect the expected agents, without cross-reaction with other organisms. The limit of detection of each STI-MS assay was ranged from 1.739 to 10.009 copies/reaction, using probit analyses. The verification rate for each target organism of the STI-MS ranged from a minimum of 89.3% to a maximum of 100%, using conventional assays and ultrasensitive digital PCR to confirm the STI-MS-positive results. To further evaluate the clinical performance of this assay, 241 clinical specimens (124 urethral/cervical swabs and 117 urine) were tested in parallel using the STI-MS assay and monoplex real-time PCR for each agent. The overall validation parameters of STI-MS were extremely high including sensitivity (from 85.7% to 100%), specificity (from 92.3% to 100%), PPV (from 50% to 100%), and NPV (from 99.1% to 100%) for each target.ConclusionSTI-MS is a useful high-throughput screening tool for detecting mixed infections of STIs and has great potential for application in large-scale epidemiological programs for specific microorganisms of STI.

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Opportunities and pitfalls of molecular testing for detecting sexually transmitted pathogens

Cited by 22 publications

References 96 publications

Viability-PCR Shows That NAAT Detects a High Proportion of DNA from Non-Viable Chlamydia trachomatis

Viability-PCR Shows That NAAT Detects a High Proportion of DNA from Non-Viable Chlamydia trachomatis

Diagnosis And Antimicrobial Treatment Of Bacterial Of Neisseria Gonorrhea Infections: Update Review Article

<p>Simultaneous detection of eleven sexually transmitted agents using multiplexed PCR coupled with MALDI-TOF analysis</p>

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