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Molecular Assays for Diagnosis of Vaginitis

Review the latest on clinical testing for vaginitis with Ashley Fuller, MD, FACOG, NCMP

Dr. Fuller is a gynecologist at Swedish Medical Group in Seattle, WA.  She has a background in microbiology and immunology, with previous work in both academic and industry research.  Her practice focuses on vulvovaginal diseases and sexual health.

SUMMARY:

Molecular diagnostics and specifically, Nucleic Acid Amplification Tests (NAATs) are becoming more popular for the diagnosis of Bacterial Vaginosis, Vulvovaginal Candidiasis, and Trichomoniasis.  Although many diagnostic methods exist, NAATs show distinct advantages in accuracy over both in-clinic tests and DNA probe testing. Other considerations include pricing, variable insurance coverage and turnaround time.  

Learning Objectives

  • To provide an overview of the molecular assays available for diagnosis of vaginitis including Direct DNA probe assays and Nucleic Acid Amplification Tests
  • To explain the testing methods and how they are performed
  • To evaluate the benefits and limitations of each test including result time and cost
  • To demonstrate that NAATs can increase the accuracy of vaginitis diagnosis and ultimately decrease health care costs

MOLECULAR ASSAY BASICS 

  • A molecular assay is a laboratory test that uses nucleotides (DNA or RNA) to identify a specific organism or pathogen, or a group of similar organisms
  • Specific DNA or RNA probes are created that bind to a precise part of the matching DNA or RNA found in the organism, a process known as ‘hybridization’
  • These tests generally use Polymerase Chain Reaction (PCR) technology to optimize detection and efficiency
    • PCR: Technique that selectively amplifies a target sequence of DNA generating millions of additional copies : Allows for selective detection of minute amounts of target DNA
    • Multiplex PCR: Refers to the amplification of multiple sequences at the same time rather than performing several separate amplification procedures

MOLECULAR ASSAYS FOR VAGINITIS

DNA DIRECT PROBE ASSAY (BD Affirm)

  • Specific labelled DNA probes bind to a unique genetic sequence of the target organisms
  • The test is designed to detect the following infections
    • Gardnerella vaginalis | Detects at 2 x 105 Colony Forming Units (CFU)
    • Candida sp | Detects at 1 x 104 CFU
    • Trichomonas vaginalis | Detects at 5 x 103 CFU
  • Sensitivity/Specificity
    • Bacterial vaginosis: 90.1%/67.6%
    • Candida: 58.1%/100%
    • Trichomonas: 46.3%/100%
  • Turn around time: Test takes 45 min but if send-out lab required, may take >24 hours
  • Limitations
    • BV: Diagnosis of BV is complicated as it is a polymicrobial clinical syndrome that is not just defined by the presence of G vaginalis but also the lack of healthy lactobacilli | G vaginalis can also be found in vaginal microbiome of women without BV which is what leads to the lower specificity
    • Performance: Low sensitivity for Trichomonas and not recommended for diagnosis
    • Testing for gonorrhea and chlamydia: Separate swab needed to test for GC/CT

NUCLEIC ACID AMPLIFICATION TESTS (NAAT):

  • Multiple NAATs available | NuSwab (LapCorp), SureSwab (Quest), Aptima (Hologic), BD Max Vaginal panel (Becton Dickinson), OneSwab (Medical Diagnostic Laboratory)
  • These assays involve 3 steps
    • Target capture (typically RNA)
    • Target amplification
    • Detection of amplification products using nucleic acid probes | Typically using fluorescent signal to indicate the presence of target of interest
  • Theoretically capable of detecting as little as one organism in a sample
  • Can detect any combination of the following tests from one swab

What Organisms are Detected with NAAT?

  • Bacterial vaginosis
    • Atopobium vaginae
    • Fastidious bacteria termed BV-associated bacterium (BVAB2)
    • Megasphera type 1
    • Gardnerella vaginalis (some assays don’t include this organism)
    • Lactobacillus crispatus and L Jensenii -negative predictors of BV
  • Candida
    • Candida albicans
    • Candida glabrata
  • Expanded Candida panel
    • Candida parapsilosis
    • Candida krusei
    • Candida tropicalis
    • Candida lusitaniae
    • Candida dubliniensis (in some assays)
  • Trichomonas vaginalis
  • Chlamydia trachomatis
  • Neisseria gonorrhoeae

NAAT Sensitivity and Specificity

  • Bacterial vaginosis
    • Scoring system that shows negative, intermediate, and positive results based on presence of different types of bacteria
    • Evaluates for multiple species of both causative bacteria and protective bacteria allowing for a more accurate diagnosis
    • Better evaluation of the polymicrobial variations of the vaginal microbiome and how they contribute to the diagnosis of BV
    • Sensitivity 90.5 to 96.9% | Specificity 85.8 to 92.6% (dependent on which assay performed)
  • Candida
    • Has option for larger multi species candida test which is helpful if recurrent or nonresolving yeast
    • C. albicans and C. glabrata only
      • Sensitivity: 97.7%
      • Specificity: 93.2%
    • Candida group (C albicans, C tropicalis, C parapsilosis, and C dubliniensis)
      • Sensitivity: 90.9%
      • Specificity: 94.1%
  • Trichomonas
    • Sensitivity 93 to 98.1%
    • Specificity 99%

Additional Important NAAT Notes 

  • NAAT can better detect co-infections or multiple infections at the same time than other testing methods
  • Acceptable for patient collected swabs
  • Should never be used for screening asymptomatic patients as some of these organisms can be found in normal vaginal flora
  • Average results can take >24 hours if a send-out lab is required

ECONOMIC OVERVIEW:

  • In clinic ‘point of care’ testing (saline microscopy, Amsel’s criteria, pH)
    • Very inexpensive
    • However, widely unavailable to clinicians due to lack of training, time, or availability of microscope
  • DNA Probe testing
    • More expensive than ‘point of care’ clinical testing
    • Insurance coverage available but variable
  • Nucleic Acid Amplification Tests
    • More sensitive and cover more microbes allowing for more accurate diagnosis than other tests currently available (clinical point of care and direct DNA Probe testing)
    • Cost varies according to number of tests ordered and can run up costs billed to insurance  
    • Insurance coverage is variable
      • Most cover trichomonas NAATs as this is the recommended diagnostic method
      • May not cover multiplex panels
      • Better coverage more likely if (1) Symptomatic diagnosis code is appropriately provided (2) Appeal/discussion between healthcare professional and payor  

Cost/Benefit Discussion

  • Clinical diagnosis using in-office diagnostic methods (Amsel’s criteria, microscopy) are subjective
    • Approximately half of all symptomatic women evaluated will not be accurately diagnosed with these methods
    • Due to low sensitivity and specificity, DNA probe testing, in general, is predicted to give incorrect diagnosis about 30% of the time
  • There are consequences to incorrect diagnosis of vaginitis
    • Increase in health care costs
      • Multiple return visits (Ob/Gyn, primary care, ED, ID)
      • Failed medical treatments with multiple prescriptions
      • Medical expenditures
      • Hospitalizations
    • Increase in pregnancy/perinatal complications
      • Preterm labor
      • Preterm delivery
      • Uterine infection after delivery
      • Miscarriage
    • Increase in post surgical complications
      • Post-hysterectomy cuff infection
      • Pelvic inflammatory disease
      • Endometritis
    • Increased risk of acquisition of STIs
    • Increased risk of azole resistant yeast strains
  • NAAT has been shown to have 20 to 25% higher diagnostic accuracy over other diagnostic methods
  • Use of NAAT leads to more accurate diagnosis, better patient management, and improved health for our patients
    • While upfront cost is higher, it ultimately results in cost-savings to the commercial payer and can reduce overall health care costs
    • It has been suggested that professional guidelines from ACOG and CDC should align and recognize NAATs as a new standard reference | This would, indeed, reduce confusion and improve diagnosis
    • Payer policies must evolve to cover these more accurate tests
  • NAAT allows the option of self-collected swabs accurately
    • As we shift office visits to telehealth visits, this could offer a reliable and accurate diagnosis

THE WRAP UP:

  • NAATs have promising data with high sensitivity and specificity and do provide clinical benefit with increased accuracy over other methods
  • Diagnosis of Trichomonas should always be done by NAAT
  • NAATs are especially helpful
    • For species identification in more difficult cases like recurrent infections or multiple simultaneous infections
    • To provide accurate diagnosis in places where microscopy and in clinic tests are not available
  • Cost and insurance coverage
    • NAATs are more expensive and have variable insurance coverage
    • Increased adoption of NAAT for diagnosis of vaginitis would considerably increase the success rate in treatment of vaginitis
  • Our patients deserve accurate diagnostics for the identification of these infections which is attainable in NAATs
    • Hopefully, commercial payers will see the benefit and how they can ultimately result in cost savings and better patient care

References

ACOG Practice Bulletin 215: Vaginitis in Nonpregnant Patients

CDC 2015 Sexually Transmitted Diseases Treatment Guidelines: BV

CDC 2015 Sexually Transmitted Diseases Treatment Guidelines: Trichomoniasis

CDC 2015 Sexually Transmitted Diseases Treatment Guidelines: Candidiasis

Clinical validation of the Aptima Bacterial Vaginosis and Aptima Candida/Trichomonas Vaginitis Assays: Results from a Prospective Multicenter Clinical Study (Schwebke et al. JCM, 2020)

Diagnostic Performance of Molecular test vs clinician assessment of vaginitis (Schwebke et al. JCM, 2018)

Diagnosis and Treatment of Vaginal Discharge Syndromes in Community Practice Settings (Hillier et al. Clinical Infectious Diseases, 2020) 

Syndromic Treatment of Women with Vulvovaginal Symptoms in the United States: A call to Action! (Sobel, Jack. Clinical Infectious Diseases, 2020)

Molecular Diagnosis of Bacterial Vaginosis: An Update (Coleman et al, JCM, 2018)

Comparison of Nucleic Acid Amplification Assays with BD Affirm VPIII and Diagnosis of Vaginitis in Symptomatic Women (Cartwright et al. JCM, 2013)  

Clinical Validation of a Test for the Diagnosis of Vaginitis (Gaydos et al, Obstetrics and Gynecology, 2017)

Health Care Utilization and Costs Following Amplified versus Non-amplified Molecular Probe Testing for Symptomatic Patients with Suspected Vulvovagintis: A US Commercial Payer Population.  (Ackerman et al, Clinical Economics and Outcomes Research 2019)

Improving the Diagnosis of Vulvovaginitis: Perspectives to Align Practice, Guidelines, and Awareness (Brown and Drexler, Population Health Management, 2020)

Commercial Support

This educational activity is supported by Hologic

Faculty Disclosures

Dr. Fuller reports that she has no relevant financial relationships to disclose

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Comments (2) Leave a Comment

  1. Candace Courtney

    I have had Candida since 2009 when I had a hysterectomy. Done topical Rx’s, pills that only temporarily stop it only for it to return. I have read many published papers on it and it seems that taking a culture from the gut by an imunoligist has to be done. Please advise me on on a specialist in my area that can help me. Zip code 93063. Thank you🌹

  2. Marrianne Aldrich

    Interested in it

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