Candida auris is an emerging multidrug-resistant yeast causing invasive health care-associated infection with high mortality worldwide. Rapid identification of C. auris is of primary importance for the implementation of public health measures to control the spread of infection. To achieve these goals, we developed and validated a TaqMan-based real-time PCR assay targeting the internal transcribed spacer 2 (ITS2) region of the ribosomal gene. The assay was highly specific, reproducible, and sensitive, with the detection limit of 1 C. auris CFU/PCR. The performance of the C. auris real-time PCR assay was evaluated by using 623 surveillance samples, including 365 patient swabs and 258 environmental sponges. Real-time PCR yielded positive results from 49 swab and 58 sponge samples, with 89% and 100% clinical sensitivity with regard to their respective culture-positive results. The real-time PCR also detected C. auris DNA from 1% and 12% of swab and sponge samples with culture-negative results, indicating the presence of dead or culture-impaired C. auris. The real-time PCR yielded results within 4 h of sample processing, compared to 4 to 14 days for culture, reducing turnaround time significantly. The new real-time PCR assay allows for accurate and rapid screening of C. auris and can increase effective control and prevention of this emerging multidrug-resistant fungal pathogen in health care facilities.
Candida auris is a multidrug-resistant yeast which has emerged in health care facilities worldwide; however, little is known about identification methods, patient colonization, environmental survival, spread, and drug resistance. Colonization on both biotic (patients) and abiotic (health care objects) surfaces, along with travel, appear to be the major factors for the spread of this pathogen across the globe. In this investigation, we present laboratory findings from an ongoing C. auris outbreak in New York (NY) from August 2016 through 2018. A total of 540 clinical isolates, 11,035 patient surveillance specimens, and 3,672 environmental surveillance samples were analyzed. Laboratory methods included matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) for yeast isolate identification, real-time PCR for rapid surveillance sample screening, culture on selective/nonselective media for recovery of C. auris and other yeasts from surveillance samples, antifungal susceptibility testing to determine the C. auris resistance profile, and Sanger sequencing of the internal transcribed spacer (ITS) and D1/D2 regions of the ribosomal gene for C. auris genotyping. Results included (a) identification and confirmation of C. auris in 413 clinical isolates and 931 patient surveillance isolates as well as identification of 277 clinical cases and 350 colonized cases from 151 health care facilities, including 59 hospitals, 92 nursing homes, 1 long-term acute care hospital (LTACH), and 2 hospices, (b) successful utilization of an in-house developed C. auris real-time PCR assay for the rapid screening of patient and environmental surveillance samples, (c) demonstration of relatively heavier colonization of C. auris in nares than in the axilla/groin, and (d) predominance of the South Asia clade I with intrinsic resistance to fluconazole and elevated MIC to voriconazole (81%), amphotericin B (61%), flucytosine (5FC) (3%), and echinocandins (1%). These findings reflect greater regional prevalence and incidence of C. auris and the deployment of better detection tools in an unprecedented outbreak.
The multidrug-resistant yeast pathogen Candida auris continues to cause outbreaks and clusters of clinical cases worldwide. Previously, we developed a real-time PCR assay for the detection of C. auris from surveillance samples (L. Leach, Y. Zhu, and S. Chaturvedi, J Clin Microbiol 56:e01223-17, 2018, https://doi.org/10.1128/ JCM.01223-17). The assay played a crucial role in the ongoing investigations of the C. auris outbreak in New York City. To ease the implementation of the assay in other laboratories, we developed an automated sample-to-result real-time C. auris PCR assay using the BD Max open system. We optimized sample extraction at three different temperatures and four incubation periods. Sensitivity was determined using eight pools of patient samples, and specificity was calculated using four clades of C. auris and closely and distantly related yeasts. Three independent extractions and testing of two patient sample pools in quadruplicate yielded assay precision. BD Max optimum assay conditions were as follows: DNA extraction at 75°C for 20 min and the use of PerfeCTa multiplex quantitative PCR (qPCR) ToughMix. The limit of detection (LOD) of the assay was one C. auris CFU/PCR. We detected all four clades of C. auris without cross-reactivity to other yeasts. Of the 110 patient surveillance samples tested, 50 were positive for C. auris using the BD Max system with 96% clinical sensitivity and 94% accuracy compared to the results of the manual assay. The BD Max assay allows high-throughput C. auris screening of 180 surveillance samples in a 12-h workday. C andida auris, an emerging multidrug-resistant yeast pathogen, continues to cause outbreaks and clusters of clinical cases worldwide (1, 2). There are ongoing efforts to devise better diagnostic approaches for the rapid detection of C. auris in clinical and surveillance samples (3-5). Previously, we developed and validated a manual real-time PCR assay for the direct detection of C. auris from surveillance samples at the New York State Department of Health (NYSDOH) Mycology Laboratory (3). The laboratorydeveloped test (LDT) enabled NYSDOH laboratory scientists and epidemiologists to carry out unprecedented surveillance and testing in hospitals and health care facilities in New York City. To date, over 13,000 clinical samples from 151 health care facilities and over 1,000 C. auris isolates were processed (S. Chaturvedi, unpublished data). The Candida auris LDT, standard operating procedures, and validation results were shared extensively with hospital, commercial, and public health laboratories. However, the C. auris LDT is not amenable to automation and high-throughput screening, and adoption of the LDT has progressed slowly while the affected facilities and sample numbers continue to grow. Therefore, we developed and validated a real-time PCR assay for C. Citation Leach L, Russell A, Zhu Y, Chaturvedi S, Chaturvedi V. 2019. A rapid and automated sample-to-result Candida auris real-time PCR assay for high-throughput testing of surveillance samples with the B...
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