Dataset Information

Clinical evaluation of a multi-parameter customized respiratory TaqMan(®) array card compared to conventional methods in immunocompromised patients.

ABSTRACT:

Background

Respiratory viral infections can cause significant morbidity and mortality in immunocompromised patients. Conventional tests routinely available at most institutions are limited by the number of detectable pathogens, by a poor sensitivity and/or a long turnaround time.

Objectives

To compare the performance of routine conventional testing with direct fluorescent antibody assays and viral culture to a customized TaqMan® array card (TAC) real-time PCR method, targeting 24 viruses, 8 bacteria and 2 fungi simultaneously.

Study design

We collected 143 respiratory samples from 120 symptomatic immunocompromised patients. Samples for which conventional and TAC results were discordant underwent further verification testing.

Results

The TAC assay identified viral pathogens in more samples than did conventional testing (77/143 versus 27/143; McNemar P<0.0001), even when TAC results for viruses that could not be detected by conventional testing were excluded from analysis (59/143 versus 26/143; P<0.0001). In addition, the TAC assay identified 18 samples with non-viral pathogens. Verification testing confirmed positive TAC results for 50 out of 55 samples for which conventional testing was negative. Two out of three samples with a positive conventional test but negative TAC result were confirmed positive. A viral and a total pathogen co-infection rate of 5.6% and 11.8% were found, respectively.

Conclusions

The customized TAC assay resulted in a significantly increased identification of respiratory viruses. This study provides a practical real-life assessment of the performance of the TAC assay in a population for whom rapid and accurate diagnosis of viral and atypical pathogens is crucial for appropriate clinical management.

SUBMITTER: Steensels D

PROVIDER: S-EPMC7106552 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

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Publications

Clinical evaluation of a multi-parameter customized respiratory TaqMan(®) array card compared to conventional methods in immunocompromised patients.

Steensels Deborah D Reynders Marijke M Descheemaeker Patrick P Curran Martin D MD Jacobs Frédérique F Denis Olivier O Delforge Marie-Luce ML Montesinos Isabel I

Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 20150903

<h4>Background</h4>Respiratory viral infections can cause significant morbidity and mortality in immunocompromised patients. Conventional tests routinely available at most institutions are limited by the number of detectable pathogens, by a poor sensitivity and/or a long turnaround time.<h4>Objectives</h4>To compare the performance of routine conventional testing with direct fluorescent antibody assays and viral culture to a customized TaqMan® array card (TAC) real-time PCR method, targeting 24 ...[more]

PMID: 26364158

Similar Datasets

Project description:BackgroundMolecular assays for diagnosis of Flu A, Flu B, and RSV with short turn-around-time (TAT) are of considerable clinical importance. In addition, rapid and accurate diagnosis of a large panel of viral and atypical pathogens can be crucial in immunocompromised patients.ObjectivesFirst, to evaluate the performance of the Simplexa™ Direct assay system in comparison with direct fluorescent antibody (DFA) and customized Taqman® Array Card (TAC) testing for RSV, Flu A, and Flu B in immunocompromised patients. Second, to evaluate different algorithms for the detection of respiratory pathogens in terms of cost, turn-around-time (TAT) and diagnostic yield.Study designWe collected 125 nasopharyngeal swabs (NTS) and 25 BAL samples from symptomatic immunocompromised patients. Samples for which Simplexa™ and TAC results were discordant underwent verification testing. The TAC assay is based on singleplex RT-PCR, targeting 24 viruses, 8 bacteria and 2 fungi simultaneously.ResultsThe overall sensitivity was significantly lower for DFA testing than for the two molecular methods (p<0.05). Performance characteristics of Simplexa™ testing were not significantly different compared to TAC testing (p>0.1). For BAL samples only, the sensitivity and specificity of the Simplexa™ assay was 100%. In total, 6.7, 16 and 18% of samples were positive for Flu A, Flu B or RSV by DFA, Simplexa™ and TAC testing, respectively. When considering not only these pathogens but also all results for TAC, the method identified 93 samples with one or more respiratory pathogens (62%). A co-infection rate of 15.3% was found by TAC. The estimated costs and TAT were 8.2€ and 2h for DFA, 31.8€ and 1.5h for Simplexa™ and 55€ and 3h for TAC testing.ConclusionsPerforming the Simplexa™ test 24h a day/7 days a week instead of DFA would considerably improve the overall sensitivity and time-to-result, albeit at a higher cost generated in the laboratory. Performing the TAC would increase the diagnostic yield and detection of co-infections significantly.

Project description:The TaqMan Array Card (TAC) system is a 384-well singleplex real-time PCR format that has been used to detect multiple infection targets. Here we developed an enteric TaqMan Array Card to detect 19 enteropathogens, including viruses (adenovirus, astrovirus, norovirus GII, rotavirus, and sapovirus), bacteria (Campylobacter jejuni/C. coli, Clostridium difficile, Salmonella, Vibrio cholerae, diarrheagenic Escherichia coli strains including enteroaggregative E. coli [EAEC], enterotoxigenic E. coli [ETEC], enteropathogenic E. coli [EPEC], and Shiga-toxigenic E. coli [STEC]), Shigella/enteroinvasive E. coli (EIEC), protozoa (Cryptosporidium, Giardia lamblia, and Entamoeba histolytica), and helminths (Ascaris lumbricoides and Trichuris trichiura), as well as two extrinsic controls to monitor extraction and amplification efficiency (the bacteriophage MS2 and phocine herpesvirus). Primers and probes were newly designed or adapted from published sources and spotted onto microfluidic cards. Fecal samples were spiked with extrinsic controls, and DNA and RNA were extracted using the QiaAmp Stool DNA minikit and the QuickGene RNA Tissue kit, respectively, and then mixed with Ag-Path-ID One Step real-time reverse transcription-PCR (RT-PCR) reagents and loaded into cards. PCR efficiencies were between 90% and 105%, with linearities of 0.988 to 1. The limit of detection of the assays in the TAC was within a 10-fold difference from the cognate assays performed on plates. Precision testing demonstrated a coefficient of variation of below 5% within a run and 14% between runs. Accuracy was evaluated for 109 selected clinical specimens and revealed an average sensitivity and specificity of 85% and 77%, respectively, compared with conventional methods (including microscopy, culture, and immunoassay) and 98% and 96%, respectively, compared with our laboratory-developed PCR-Luminex assays. This TAC allows fast, accurate, and quantitative detection of a broad spectrum of enteropathogens and is well suited for surveillance or clinical purposes.

Dataset Information

Clinical evaluation of a multi-parameter customized respiratory TaqMan(®) array card compared to conventional methods in immunocompromised patients.

Background

Objectives

Study design

Results

Conclusions

Publications

Clinical evaluation of a multi-parameter customized respiratory TaqMan(®) array card compared to conventional methods in immunocompromised patients.

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