Performance characteristics of an automated, high-throughput RT-PCR assay for the detection of Candida auris on 3-point and nasal swabs

Abstract

Candida auris is a multidrug-resistant yeast responsible for invasive infections with high mortality rates, primarily spread through prolonged colonization on biotic and abiotic surfaces and traveling. Effective control necessitates comprehensive screening protocols, as recommended by the Centers for Disease Control and Prevention, which endorses a real-time polymerase chain reaction-based assay for C. auris screening. This study evaluates the performance of this assay on the Hologic Panther Fusion System using nasal and 3-point swab specimens (nares/axilla/groin) compared with culture. The assay was assessed for accuracy, sensitivity, specificity, and reproducibility, alongside an evaluation of probe primer reagent (PPR) onboard stability over 30 working days. Analytical sensitivity studies determined limits of detection of 1.95 Log CFU/mL for nasal swabs and 2.18 Log CFU/mL for 3-point swabs, both with >95.0% confidence intervals. The assay demonstrated 100% specificity (n = 25), with no false positives from genetically similar or clinically relevant species, and no significant interference from co-infecting microbes on cycle threshold (CT) values. Both swab types exhibited high intra- and inter-reproducibility, with low coefficients of variation (1.79% and 1.06%, respectively). The assay detected C. auris in 100% of 108 spiked-positive samples across both swab types. Clinical analysis showed 100% concordance with culture for 3-point swabs. Additionally, the nasal swab method showed 96.0% overall agreement, with 86.2% sensitivity and 100.0% specificity. The PPR mixes remained stable over 30 working days, with no significant CT value changes. This study confirms the assay's robust sensitivity, specificity, reproducibility, and accuracy for both nasal and 3-point screening swabs.

Importance: Candida auris is a multidrug-resistant yeast responsible for severe infections with high mortality rates. Rapid and accurate detection is critical for preventing the spread of this pathogen in healthcare settings. This study assesses the performance of an automated real-time PCR screening assay for detecting C. auris using nasal and 3-point swabs. The findings demonstrate the assay's high sensitivity, specificity, and reproducibility, making it a valuable tool for infection control. By providing a reliable and efficient screening method, this assay can significantly enhance efforts to control C. auris outbreaks, ultimately improving patient outcomes and reducing the spread of this dangerous pathogen.

Keywords: Candida auris; PCR; diagnostic; screening.

Fig 1

Linearity was established across the range of C. auris concentrations from 2.22 to 7.22 Lo g CFU/mL in (A) 3-point ESwab and (B) nasal-alone ESwab.

Fig 2

Probit analysis at multiple levels of low concentration with the lowest at 1.18 Log CFU/mL. (A) 3-point Eswab: Probit analysis determined the LoD to be 1.88 Log CFU/mL with a mean CT value of 36.77 ± 1.32. (B) Nasal-alone Eswab: Probit analysis determined the LoD to be 1.65 Log CFU/mL with a mean CT value of 35.20 ± 1.50.

Fig 3

Stability of onboard PPR reagents. CT values of the same C. auris 3-point and nasal-alone (nares) specimens tested by onboard PPR reagents left at ambient temperature over the span of 30 days (Mean ± SD = 25.7 ± 0.5 for 3-point, 23.7 ± 0.5 for nasal).