Comparison of the Simplexa FluA/B &amp; RSV Direct Assay and Laboratory-Developed Real-Time PCR Assays for Detection of Respiratory Virus

Woodberry, Mitchell W.; Shankar, Rohit; Cent, Anne; Jerome, Keith R.; Kuypers, Jane

doi:10.1128/jcm.02395-13

Cited by 29 publications

(23 citation statements)

References 8 publications

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“…Using laboratory records, we identified patients who had testing performed for influenza using the laboratory-developed traditional RT-PCR assay in Season 1 and using the rapid PCR-based influenza test in Season 2 according to previously published methods [Woodberry et al, 2013]. The rapid PCR-based influenza assay (Simplexa Flu A/B & RSV Direct kit, Focus Diagnostics, Cypress, CA) combines nucleic acid extraction and real-time RT-PCR for detection of influenza A/B and RSV using primers and probes targeting the conserved regions of the viral genomes.…”

Section: Methodsmentioning

confidence: 99%

“…Influenza reverse-transcriptase polymerase chain reaction (RT-PCR) tests are highly sensitive and specific, but take several hours to days to provide results. Rapid PCR-based influenza tests have been shown to be as sensitive and specific as traditional influenza RT-PCR assays [Woodberry et al, 2013], and have the additional advantage of providing results within 1 hr. These have the potential to be used in settings such as emergency departments and inpatient hospital wards, where reliable and rapid influenza test results in high-risk patients may impact patient care and clinical outcomes.…”

Section: Introductionmentioning

confidence: 99%

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Impact of rapid influenza PCR testing on hospitalization and antiviral use: A retrospective cohort study

et al. 2015

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Rapid PCR-based influenza tests are increasingly used as point-of-care diagnostics in hospitals and clinics. To our knowledge, no prior studies have described clinical outcomes with implementation of rapid PCR-based influenza tests in hospitalized adult inpatients. Electronic medical records were used to assess differences in laboratory testing time and antiviral use among a subset of 175 consecutive adult inpatients tested for influenza in two respiratory seasons before and after implementation of rapid PCR-based influenza testing at an academic medical center. Of the 350 hospitalized inpatients included in this analysis, 96 (27%) were over 65 years of age and 308 (88%) had a comorbid condition. The overall time to result decreased significantly from 25.2 to 1.7 hr (P<0.001) after implementation of rapid PCR-based influenza testing. Among influenza-negative patients, the frequency of oseltamivir initiation remained unchanged (before: 43% vs. after: 45%; P=0.60), though the median duration of oseltamivir was significantly decreased from 1.1 to 0.0 days (P<0.001). By providing an earlier result to clinicians, rapid PCR-based influenza tests may decrease unnecessary antiviral use among adult inpatients who test negative for influenza.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Impact of rapid influenza PCR testing on hospitalization and antiviral use: A retrospective cohort study

et al. 2015

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“…Not surprisingly, the results varied. Two studies using fresh samples found Simplexa direct testing (2 and 50 l) to be similar in sensitivity to their LDT PCRs (8,9). The proportion of samples with low viral loads was not reported in those studies.…”

mentioning

confidence: 99%

“…Several papers were recently published on Simplexa Flu A/B & RSV using one or more of three approaches, extracted nucleic acid (5-7) and/or direct testing of either 2 l of sample in a 96-well disc (6,8) or 50 l of sample in an 8-well disc (7,9). The 50-l option is now available as an in vitro diagnostic (IVD) kit.…”

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confidence: 99%

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Comparison of Simplexa Flu A/B & RSV PCR with Cytospin-Immunofluorescence and Laboratory-Developed TaqMan PCR in Predominantly Adult Hospitalized Patients

Landry

Ferguson

2014

J Clin Microbiol

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To compare Simplexa Flu A/B & RSV PCR with cytospin-immunofluorescence and laboratory-developed TaqMan PCR methods, 445 nasopharyngeal samples were tested. Of these, 199 were positive (46 for respiratory syncytial virus [RSV], 120 for influenza A, and 33 for influenza B) and 246 were negative. The direct fluorescent-antibody assay (DFA) detected 132 (66.3%) positive samples, Simplexa direct detected 162 (81.4%), Simplexa using extracts detected 177 (88.9%), and lab-developed TaqMan PCR reference methods detected 199 (100%). The specificities were 99.6%, 100%, 100%, and 100%, respectively. The two Simplexa methods were more sensitive than the DFA (P ‫؍‬ 0.0001) but less sensitive than the TaqMan reverse transcriptase PCR (P ‫؍‬ 0.0001). Influenza virus and respiratory syncytial virus (RSV) cause a substantial disease burden in people of all ages. A rapid laboratory diagnosis can facilitate early antiviral therapy, discontinuation of antibiotics, and implementation of infection control measures (1). Our laboratory uses the cytospin-enhanced direct fluorescent-antibody assay (DFA) as an initial on-demand screen and laboratory-developed real-time TaqMan (LDT) PCR when greater sensitivity is needed (2). In this report, we evaluated Simplexa Flu A/B & RSV PCR as a potential replacement for DFA and LDT PCR.Nasopharyngeal swab specimens submitted in 3 ml M4 medium (Remel, Lenexa, KS) from February to April 2011 were tested by DFA and LDT PCR as requested by the clinician within 2 to 12 h of collection. The original samples and extracts were then frozen at Ϫ70°C, and a subset of the positive and negative samples were retested with the Simplexa PCR within 2 to 4 months.The cytospin-DFA was performed using SimulFluor reagents (Millipore, Billerica, MA) as previously described (3). For influenza detection and subtyping, the CDC real-time reverse transcriptase (RT) PCR primers and probes were used (2). For RSV, the protocol published by van Elden et al. (4) was used, but the subtypes were not differentiated. Total nucleic acid was extracted from 200 l of sample and eluted in 55 l on an EasyMag instrument (bioMérieux, Durham, NC), and 5 l of extract was added to 20 l AgPath-ID one-step RT-PCR master mix (Life Technologies, Carlsbad, CA) without an internal control (IC), and the manufacturers' cycling parameters (10 min at 45°C, 10 min at 95°C, 15 s at 95°C, 45 s at 60°C) were used. Amplification was performed for 45 cycles using an ABI 7500 instrument (Applied Biosystems, Foster City, CA). Positive samples with cycle threshold (C T ) values of Ͻ38 were accepted as positive. For C T values of Ն38, amplification was repeated in duplicate and accepted if one replicate was positive.For Simplexa Flu A/B & RSV PCR (Focus Diagnostics, Cypress, CA), frozen extracts were thawed, vortexed, and pulse spun. To each well of a 96-well disc was added 5 l of master mix containing primers, an IC, and 5 l of thawed extract or positive or negative control material. Discs were sealed and placed in a 3 M integrated cycler (Focus Diagnostics). Amp...

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Clinical Microbiology

Whitfield¹,

Martínez²,

Wolk³

2016

Manual of Commercial Methods in Clinical Microbiology

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Comparison of the Simplexa FluA/B & RSV Direct Assay and Laboratory-Developed Real-Time PCR Assays for Detection of Respiratory Virus

Cited by 29 publications

References 8 publications

Impact of rapid influenza PCR testing on hospitalization and antiviral use: A retrospective cohort study

Impact of rapid influenza PCR testing on hospitalization and antiviral use: A retrospective cohort study

Comparison of Simplexa Flu A/B & RSV PCR with Cytospin-Immunofluorescence and Laboratory-Developed TaqMan PCR in Predominantly Adult Hospitalized Patients

Clinical Microbiology

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