Validation of an in-house developed rapid assay for Group B Streptococcus detection in pregnancy

Colonization of Group B Streptococcus (GBS) in pregnant women represents a primary risk factor for neonatal infections and preterm stillbirth, highlighting the necessity for pre-delivery detection. Traditional culture methods and standard molecular assays are limited, highlighting the demand for a rapid, efficient, and straightforward GBS detection technique. This study focused on developing and validating an in-house rapid assay utilizing the CRISPR/Cas12a system, augmented with recombinase polymerase amplification (RPA).

A rapid, sensitive, and specific GBS detection platform was established, employing CRISPR/Cas12a in conjunction with isothermal RPA. Specificity assessments were conducted against GBS as well as other prevalent pathogens of the female reproductive tract. The limit of detection was determined using gradient-diluted GBS-originated plasmids, while clinical swabs from pregnant women were used to evaluate diagnostic efficacy against a commercial quantitative real-time PCR (qPCR) assay.

The GBS rapid assay demonstrated high efficiency, completing the test within one hour, with results available for interpretation through visual examination or a fluorescence reader. This method accurately differentiated GBS from non-target control pathogens, achieving a detection limit of 201.64 copies/µl with a 95% Confidence Interval (95% CI): 66.90-607.75, as assessed by colloidal gold test strips. In a clinical performance evaluation involving 1000 vaginal-rectal swabs from pregnant women, the rapid assay results correlated closely with those from the conventional qPCR method.

An easy-to-operate GBS assay, featuring a short turnaround time, was developed and validated with high sensitivity, specificity, and accuracy, indicating significant potential for clinical application.