A-284 Development of a Low Positive Molecular Control for Mycoplasma genitalium and Trichomonas vaginalis Screening
Authors: Huang C , Brandt L , Ruminski Lowe D , Howard R , Truong N , Morreale E , Anekella B 
Published: October 2024

Background

Mycoplasma genitalium (MG) is a sexually transmitted gram -negative bacteria that infects the urinary and genital tracts. Trichomonas vaginalis (TV) is a protozoan parasite and is the most common curable sexually transmitted infection (STI) worldwide. TV/ MG infections often go unrecognized because infected individuals are either asymptomatic or have symptoms similar to other infections of the urogenital tract (cervicitis in females and urethritis in males). Left untreated, TV/MG can lead to severe health complications including pelvic inflammatory disease, adverse pregnancy outcomes such as preterm delivery and miscarriage, increased risk of HIV acquisition, sexually acquired reactive arthritis, and infertility. The rates of STIs are increasing and differentiating among different STIs is critical to ensure patients receive effective treatment. Amplified nucleic acid based tests (NAATs) have emerged as the gold standard for diagnosis of TV/MG infections due to their high sensitivity and high throughput using a variety of clinical samples including male and female urine. This study describes work to develop a low positive quality control material in human urine for TV/MG amplified nucleic acids tests.

Methods

A method was developed to eliminate residual enzymatic activity from raw, pooled human urine from both genders to create a homogeneous diluent that supports analyte stability. The urine diluent was tested for viral pathogens (HIV-1, HCV, HBV) and ensure urine proteins and chemistry analytes were within acceptable limits. TaqMan® assays for TV (targeting RNA polymerase Rpb1 gene) and MG (targeting SMC Chromosome segregation gene) were developed and qualified for digital PCR quantitation of these organisms using the BioRad QX200 system. Cultured Trichomonas vaginalis strain C-1:NIH and Mycoplasma genitalium strain G37 were spiked into the urine diluent along with 0.09% sodium azide as a preservative. The concentration of TV and MG was guided by digital PCR quantitation throughout formulation and the final product was tested on Abbott Alinity m STI Panel, Hologic Aptima® Mycoplasma genitalium Assay, and Hologic Aptima® Trichomonas vaginalis Assay.

Results

Three independent lots were produced that were each detected for both TV and MG on the Abbott Alinity m STI panel. The concentration of the controls targeted a cycle number (CN) of ∼27 for TV and ∼32 for MG and were within acceptable limits. The lots were also positive for TV and MG when tested on the Hologic Aptima TV and MG assays. Accelerated stability studies showed that the product is stable throughout 2 years of shelf life when stored frozen at -20 °C, but is also stable for up to 30 days at 2-8 °C refrigerated storage.

Conclusions

LGC Clinical Diagnostics is developing a stable, whole organism molecular control material for use with in vitro diagnostic test methods that detect TV and MG. The product will fulfill an unmet need for molecular quality controls for STI testing that are formulated in a true patient-like matrix like urine. This study helps ensure the molecular control will be a low positive on leading diagnostic assays, be consistent batch to batch, and have the needed stability to meet the needs of clinical testing labs.