One-step and sequential SARSCOV-2 polymerase chain reaction tests would not work every time.

INTRODUCTION: RT-PCR is widely used as a diagnostic test for the detection of SARS-CoV-2. In this study, we aim to describe the clinical utility of serial PCR testing in the final detection of COVID-19. METHOD: We collected multiple nasopharyngeal swab samples from patients who had negative RT-PCR test on the first day after hospitalization. RT-PCR tests were performed on the second day for all patients with initial negative result. For the patients with secondary negative results on day 2, tertiary RT-PCR tests were performed on day 3 after hospitalization. RESULT: Among 68 patients with initial negative test results, at the end of follow-up, the mortality number was 20 (29.4%). About 33.8% of patients had subsequent positive PCR test results for the second time and 17.4% of the patients who performed third PCR test had positive result. CONCLUSION: Based on this study, serial RT-PCR testing is unlikely to yield additional information.

Use of point-of-care molecular tests reduces hospitalization and oseltamivir administration in children presenting with influenza-like illness.

Influenza is associated with increased morbidity, healthcare costs, hospitalization rates, and mortality in children. Rapid immunochromatography assay (ICA), a test with low sensitivity, is often used as point-of-care (POC) test. Recently, the rapid syndromic molecular test FilmArray has become available. This observational study aims to evaluate whether the use of FilmArray would decrease the use of antivirals and hospitalization rates among children presenting to the emergency room (ER) with influenza-like illness (ILI) symptoms. Nasopharyngeal swabs were prospectively collected from children, aged 0-16 years, presenting with ILI at the ER of a tertiary hospital during the peak endemic period. Patients were allocated to be tested by either FilmArray or ICA. The use of antivirals and hospitalization rates were noted. Logistic regression models were used to investigate the impact of testing methods on decision-making. Overall, 80 children were included (mean age: 5 years). Admissions were more likely to occur if an ICA test was performed (OR, 3.16; 95% CI, 1.01-9.82; p = .046). Oseltamivir administration was more likely among children who had undergone the ICA test (OR, 4.67; 95% CI, 1.06-20.43; p = .041). The implementation of rapid molecular test had no impact on complementary diagnostic testing or antibacterial prescription. The use of FilmArray significantly reduced both hospitalization and oseltamivir administration in children. Further knowledge on the use of POC tests is required to improve current management of children presenting with ILI and decrease associated healthcare costs.

False negative rate of COVID-19 PCR testing: a discordant testing analysis.

BACKGROUND: COVID-19 is diagnosed via detection of SARS-CoV-2 RNA using real time reverse-transcriptase polymerase chain reaction (rtRT-PCR). Performance of many SARS-CoV-2 rtRT-PCR assays is not entirely known due to the lack of a gold standard. We sought to evaluate the false negative rate (FNR) and sensitivity of our laboratory-developed SARS-CoV-2 rtRT-PCR targeting the envelope (E) and RNA-dependent RNA-polymerase (RdRp) genes. METHODS: SARS-CoV-2 rtRT-PCR results at the Public Health Laboratory (Alberta, Canada) from January 21 to April 18, 2020 were reviewed to identify patients with an initial negative rtRT-PCR followed by a positive result on repeat testing within 14 days (defined as discordant results). Negative samples from these discordant specimens were re-tested using three alternate rtRT-PCR assays (targeting the E gene and N1/N2 regions of the nucleocapsid genes) to assess for false negative (FN) results. RESULTS: During the time period specified, 95,919 patients (100,001 samples) were tested for SARS-CoV-2. Of these, 49 patients were found to have discordant results including 49 positive and 52 negative swabs. Repeat testing of 52 negative swabs found five FNs (from five separate patients). Assuming 100% specificity of the diagnostic assay, the FNR and sensitivity in this group of patients with discordant testing was 9.3% (95% CI 1.5-17.0%) and 90.7% (95% CI 82.6-98.9%) respectively. CONCLUSIONS: Studies to understand the FNR of routinely used assays are important to confirm adequate clinical performance. In this study, most FN results were due to low amounts of SARS-CoV-2 virus concentrations in patients with multiple specimens collected during different stages of infection. Post-test clinical evaluation of each patient is advised to ensure that rtRT-PCR results are not the only factor in excluding COVID-19.

Performance of rapid diagnostic tests, microscopy, loop-mediated isothermal amplification (LAMP) and PCR for malaria diagnosis in Ethiopia: a systematic review and meta-analysis.

BACKGROUND: Rapid accurate diagnosis followed by effective treatment is very important for malaria control. Light microscopy remains the "golden standard" method for malaria diagnosis. Diagnostic test method must have sufficient level of accuracy for detecting malaria parasites. Therefore, this study aimed to investigate the diagnostic accuracy of rapid diagnostic tests (RDTs), microscopy, loop-mediated isothermal amplification (LAMP) and/or polymerase chain reaction (PCR) for the malaria diagnosis in Ethiopia. METHODS: Data bases such as PubMed, PubMed central, Science direct databases, Google scholar, and Scopus were searched from September to October, 2020 for studies assessing the diagnostic accuracy of RDTs, microscopy, LAMP and PCR methods for malaria diagnosis. RESULTS: A total of 29 studies published between 2001 and 2020 were analysed using review manager, Midas (Stata) and Meta-disc. The sensitivity and specificity of studies comparing RDT with microscopy varies from 79%-100% to 80%-100%, respectively. The sensitivity of LAMP (731 tests) was 100% and its specificity was varies from 85 to 99% when compared with microscopy and PCR. Considerable heterogeneity was observed between studies included in this meta-analysis. Meta-regression showed that blinding status and target antigens were the major sources of heterogeneity (P < 0.05). RDT had an excellent diagnostic accuracy (Area under the ROC Curve = 0.99) when compared with microscopy. Its specificity was quite good (93%-100%) except for one outlier (28%), but lower "sensitivity" was observed when PCR is a reference test. This indicates RDT had a good diagnostic accuracy (AUC = 0.83). Microscopy showed a very good diagnostic accuracy when compared with PCR. CONCLUSIONS: The present study showed that microscopy and RDTs had high efficiency for diagnosing febrile malaria patients. The diagnostic accuracy of RDT was excellent when compared with microscopy. This indicates RDTs have acceptable sensitivities and specificities to be used in resource poor settings as an alternative for microscopy. In this study, LAMP showed an excellent sensitivities and specificities. Furthermore, the need of minimum equipment and relatively short time for obtaining results can made LAMP one of the best alternatives especially for accurate diagnosis of asymptomatic malaria.

Integrating Xpert MTB/RIF for TB diagnosis in the private sector: evidence from large-scale pilots in Patna and Mumbai, India.

BACKGROUND: Xpert MTB/RIF (Xpert) has been recommended by WHO as the initial diagnostic test for TB and rifampicin-resistance detection. Existing evidence regarding its uptake is limited to public health systems and corresponding resource and infrastructure challenges. It cannot be readily extended to private providers, who treat more than half of India's TB cases and demonstrate complex diagnostic behavior. METHODS: We used routine program data collected from November 2014 to April 2017 from large-scale private sector engagement pilots in Mumbai and Patna. It included diagnostic vouchers issued to approximately 150,000 patients by about 1400 providers, aggregated to 18,890 provider-month observations. We constructed three metrics to capture provider behavior with regards to adoption of Xpert and studied their longitudinal variation: (i) Uptake (ordering of test), (ii) Utilization for TB diagnosis, and (iii) Non-adherence to negative results. We estimated multivariate linear regression models to assess heterogeneity in provider behavior based on providers' prior experience and Xpert testing volumes. RESULTS: Uptake of Xpert increased considerably in both Mumbai (from 36 to 60.4%) and Patna (from 12.2 to 45.1%). However, utilization of Xpert for TB diagnosis and non-adherence to negative Xpert results did not show systematic trends over time. In regression models, cumulative number of Xpert tests ordered was significantly associated with Xpert uptake in Patna and utilization for diagnosis in Mumbai (p-value< 0.01). Uptake of Xpert and its utilization for diagnosis was predicted to be higher in high-volume providers compared to low-volume providers and this gap was predicted to widen over time. CONCLUSIONS: Private sector engagement led to substantial increase in uptake of Xpert, especially among high-volume providers, but did not show strong evidence of Xpert results being integrated with TB diagnosis. Increasing availability and affordability of a technically superior diagnostic tool may not be sufficient to fundamentally change diagnosis and treatment of TB in the private sector. Behavioral interventions, specifically aimed at, integrating Xpert results into clinical decision making of private providers may be required to impact patient-level outcomes.

Improved outcomes and staging in non-small-cell lung cancer guided by a molecular assay.

There remains a critical need for improved staging of non-small-cell lung cancer, as recurrence and mortality due to undetectable metastases at the time of surgery remain high even after complete resection of tumors currently categorized as 'early stage.' A 14-gene quantitative PCR-based expression profile has been extensively validated to better identify patients at high-risk of 5-year mortality after surgical resection than conventional staging - mortality that almost always results from previously undetectable metastases. Furthermore, prospective studies now suggest a predictive benefit in disease-free survival when the assay is used to guide adjuvant chemotherapy decisions in early-stage non-small-cell lung cancer patients.

Lay abstract There is a need for improvement in the way early-stage non-small-cell lung cancers are staged and treated because many patients with 'early-stage' disease suffer high rates of cancer recurrence after surgery. In recent years, a specialized test has been developed to allow better characterization of a tumor's risk of recurrence based on the genes being expressed by tumor cells. Use of this test, in conjunction with standard staging methods, is better able to identify patients at high risk of cancer recurrence after surgery. Evidence suggests that giving chemotherapy to patients at high risk of recurrence after surgery reduces recurrence rates and improves long-term patient survival.

Prenatal molecular testing and diagnosis of Beckwith-Wiedemann syndrome.

OBJECTIVE: The objective of this study was to describe molecular findings and phenotypic features among individuals referred for prenatal Beckwith-Wiedemann syndrome (BWS) testing. METHODS: Molecular diagnostic testing was performed using a sensitive quantitative real-time PCR-based assay capable of detecting mosaic methylation to the level of 3% at IC1 and IC2. Sanger sequencing of CDKN1C was performed in cases with normal methylation. RESULTS: Of the 94 patients tested, a molecular diagnosis was identified for 25.5% of cases; 70.9% of diagnosed cases had loss of methylation at IC2, 4.2% had gain of methylation at IC1, 12.5% had paternal uniparental isodisomy, and 12.5% had CDKN1C loss-of-function variants. Methylation level changes in prenatal cases were significantly greater than changes identified in cases tested after birth. Cases with a prenatal molecular diagnosis had a significantly greater number of BWS-associated phenotypic features. The presence of either macroglossia or placentomegaly was most predictive of a BWS diagnosis. CONCLUSION: Our results support the consensus statement advocating BWS molecular testing for all patients with one or more BWS-associated prenatal features and suggest that low-level mosaic methylation changes may be uncommon among prenatal BWS diagnoses.

Rapid Detection of mrp, epf, and sly Genes by Loop-Mediated Isothermal Amplification in Streptococcus suis.

Streptococcus suis remains a serious threat to the worldwide swine industry and human health. In this study, rapid assays for the detection of three common virulence-related factors (mrp, epf, and sly) were developed, evaluated, and applied. Loop-mediated isothermal amplification (LAMP) primers were designed using Primer Explorer V5 software. The sensitivity and specificity of the LAMP assays were determined based on sample turbidity. For all three genes, LAMP assays were performed at 62°C with a reaction time of 60 min. The detection limit of conventional polymerase chain reaction (PCR) was 1 ng/µL, 10 pg/µL, and 100 fg/µL for the epf, sly, and mrp genes, respectively. For the LAMP assays, the detection limits were 10 pg/µL, 10 fg/µL, and 100 fg/µL for epf, sly, and mrp, respectively, representing sensitivities 100-1000 times higher than those of the PCR assay. Furthermore, when the LAMP assays were applied to clinical strains, the results were consistent with those of the PCR assay, confirming the LAMP assays as rapid and reliable detection techniques. In conclusion, the LAMP assays described in this study have the potential to become standard methods to detect the virulence factors mrp, epf, and sly. To the best of our knowledge, this is the first study to report the application of LAMP to detect the mrp, epf, and sly genes.

Direct Comparison of SARS-CoV-2 Analytical Limits of Detection across Seven Molecular Assays.

Analytical sensitivity for SARS-CoV-2 detection is a key performance metric for the evaluation of viral detection assays. We determined analytical limits of detection for seven SARS-CoV-2 assays using serial dilutions of pooled patient material quantified with droplet digital PCR. Limits of detection ranged from ≤10 to 74 copies/ml for commercial high-throughput laboratory analyzers (Roche Cobas, Abbott m2000, and Hologic Panther Fusion) and 167 to 511 copies/ml for sample-to-answer (DiaSorin Simplexa, GenMark ePlex) and point-of-care instruments (Abbott ID NOW). The CDC assay yielded limits of detection ranging from 85 to 499 copies/ml, depending on the extraction method and thermocycler used. These results can help to inform the assay choice for testing approaches to manage the current COVID-19 outbreak.

Active case detection of malaria in pregnancy using loop-mediated amplification (LAMP): a pilot outcomes study in South West Ethiopia.

BACKGROUND: 125 million women are pregnant each year in malaria endemic areas and are, therefore, at risk of Malaria in Pregnancy (MiP). MiP is the direct consequence of Plasmodium infection during pregnancy. The sequestration of Plasmodium falciparum parasites in the placenta adversely affects fetal development and impacts newborn birth weight. Importantly, women presenting with MiP commonly develop anaemia. In Ethiopia, the Ministry of Health recommends screening symptomatic women only at antenatal care visits with no formal intermittent preventive therapy. Since MiP can display low-level parasitaemia, current tests which include microscopy and RDT are challenged to detect these cases. Loop mediated isothermal Amplification (LAMP) technology is a highly sensitive technique for DNA detection and is field compatible. This study aims to evaluate the impact of active malaria case detection during pregnancy using LAMP technology in terms of birth outcomes. METHODS: A longitudinal study was conducted in two health centres of the Kafa zone, South West Ethiopia. Both symptomatic and asymptomatic pregnant women were enrolled in the first or second trimester and allocated to either Standard of Care (SOC-microscopy and RDT) or LAMP (LAMP, microscopy and RDT). Women completed at least three visits prior to delivery, and the patient was referred for treatment if Plasmodium infection was detected by any of the testing methods. The primary outcome was to measure absolute birth weight, proportion of low birth weight, and maternal/neonatal haemoglobin in each arm. Secondary outcomes were to assess the performance of microscopy and RDT versus LAMP conducted in the field. RESULTS: One hundred and ninety-nine women were included and assigned to either LAMP or SOC. Six were lost to follow up. In this cohort, 66.8% of women did not display any clinical symptoms and 70.9% were multi-parous. A reduced proportion of low birth weight newborns was observed in the LAMP group (0%) compared to standard of care (14%) (p <0.001). Improved neonatal haemoglobin was observed in the LAMP (13.1 g/dL) versus the SOC (12.8 g/dL) (p = 0.024) arm. RDT and microscopy had an analytical sensitivity of 66.7% and 55.6% compared to LAMP as a reference standard. CONCLUSIONS: These results support the use of highly sensitive tools for rapid on-site active case detection of MiP which may improve birth outcomes in the absence of IPT. However, further large-scale studies are required to confirm this finding.

Impact of Rapid Molecular Diagnostic Testing of Respiratory Viruses on Outcomes of Adults Hospitalized with Respiratory Illness: a Multicenter Quasi-experimental Study.

A standard multiplex PCR offers comprehensive testing for respiratory viruses. However, it has traditionally been performed in a referral laboratory with a lengthy turnaround time, which can reduce patient flow through the hospital. We aimed to determine whether the introduction of a rapid PCR, but with limited targets (Cepheid Xpert Flu/RSV XC), was associated with improved outcomes for adults hospitalized with respiratory illness. A controlled quasi-experimental study was conducted across three hospitals in New South Wales, Australia. Intervention groups received standard multiplex PCR during the preimplementation, July to December 2016 (n = 953), and rapid PCR during the postimplementation, July to December 2017 (n = 1,209). Control groups (preimplementation, n = 937, and postimplementation, n = 1,102) were randomly selected from adults hospitalized with respiratory illness during the same periods. The outcomes were hospital length of stay (LOS) and microbiology test utilization (blood culture, urine culture, sputum culture, and respiratory bacterial and virus serologies). The introduction of rapid PCR was associated with a nonsignificant 8.9-h reduction in median LOS (95% confidence interval [CI], -21.5 h to 3.7 h; P = 0.17) for all patients and a significant 21.5-h reduction in median LOS (95% CI, -36.8 h to -6.2 h; P < 0.01) among patients with positive test results in an adjusted difference-in-differences analysis. For patients receiving test results before disposition, rapid PCR use was associated with a significant reduction in LOS, irrespective of test results. Compared with standard PCR testing, rapid PCR use was significantly associated with fewer blood culture (adjusted odds ratio [aOR], 0.67; 95% CI, 0.5 to 0.82; P < 0.001), sputum culture (aOR, 0.56; 95% CI, 0.47 to 0.68, P < 0.001), bacterial serology (aOR, 0.44; 95% CI, 0.35 to 0.55, P < 0.001) and viral serology (aOR, 0.42; 95% CI, 0.33 to 0.53, P < 0.001) tests, but not with fewer urine culture tests (aOR, 0.94; 95% CI, 0.78 to 1.12, P = 0.48). Rapid PCR testing of adults hospitalized with respiratory illnesses can deliver benefits to patients and reduce resource utilization. Future research should consider a formal economic analysis and assess its potential impacts on clinical decision making.

Correlates and determinants of Early Infant Diagnosis outcomes in North-Central Nigeria.

BACKGROUND: A negative status following confirmatory Early Infant Diagnosis (EID) is the desired pediatric outcome of prevention of Mother to Child Transmission (PMTCT) programs. EID impacts epidemic control by confirming non-infected HIV-exposed infants (HEIs) and prompting timely initiation of ART in HIV-infected babies which improves treatment outcomes. OBJECTIVES: We explored factors associated with EID outcomes among HEI in North-Central Nigeria. METHOD: This is a cross-sectional study using EID data of PMTCT-enrollees matched with results of HEI's dried blood samples (DBS), processed for DNA-PCR from January 2015 through July 2017. Statistical analyses were done using SPSS version 20.0 to generate frequencies and examine associations, including binomial logistic regression with p < 0.05 being statistically significant. RESULTS: Of 14,448 HEI in this analysis, 51.8% were female and 95% (n = 12,801) were breastfed. The median age of the infants at sample collection was 8 weeks (IQR 6-20), compared to HEI tested after 20 weeks of age, those tested earlier had significantly greater odds of a negative HIV result (≤ 6 weeks: OR = 3.8; 6-8 weeks: OR = 2.1; 8-20 weeks: OR = 1.5) with evidence of a significant linear trend (p < 0.001). Similarly, HEI whose mothers received combination antiretroviral therapy (cART) before (OR = 11.8) or during the index pregnancy (OR = 8.4) had significantly higher odds as compared to those whose mothers did not receive cART. In addition, HEI not breastfed had greater odds of negative HIV result as compared to those breastfed (OR = 1.9). CONCLUSIONS: cART prior to and during pregnancy, earlier age of HEI at EID testing and alternative feeding other than breastfeeding were associated with an increased likelihood of being HIV-negative on EID. Therefore, strategies to scale-up PMTCT services are needed to mitigate the burden of HIV among children.

Evaluation and statistical optimization of a method for methylated cell-free fetal DNA extraction from maternal plasma.

PURPOSE: Methylated cell-free fetal DNA (cffDNA) in maternal plasma can potentially be used as a biomarker for accurate noninvasive prenatal testing (NIPT) of fetal disorders. Recovery and purification of cffDNA are key steps for downstream applications. In this study, we aimed to developed and evaluated different aspects of an optimized method and compared its efficiency with common methods used for extraction of methylated cffDNA. METHODS: Single factor experiments, Plackett-Burman (PB) design, and response surface methodology (RSM) were conducted for conventional Triton/Heat/Phenol (cTHP) method optimization. The total cell-free DNA (cfDNA) was extracted from pooled maternal plasma using the optimized method called the Triton/Heat/Phenol/Glycogen (THPG), cTHP method, a column-based kit, and a magnetic bead-based kit. In the next step, methylated cfDNA from the extracted total cfDNA was enriched using a methylated DNA immunoprecipitation (MeDIP) kit. Real-time quantitative polymerase chain reaction was performed on the RASSF1 gene and hyper region to determine the genomic equivalents per milliliter (GEq/ml) values of the methylated cfDNA and cffDNA, respectively. RESULTS: The optimum values of the significant factors affecting cfDNA extraction from 200 µl of plasma were 3% SDS, 1% Triton X-100, 0.9 µg/µl glycogen, and 0.3 M sodium acetate. The GEq/ml values of methylated cffDNA extracted using the THPG method were significantly higher than for the tested extraction methods (p < 0.001). CONCLUSIONS: Our results indicate that the THPG method is more efficient than the other tested methods for extraction of low copy number methylated cffDNA from a small volume of maternal plasma.

Low Yield of FilmArray GI Panel in Hospitalized Patients with Diarrhea: an Opportunity for Diagnostic Stewardship Intervention.

The FilmArray GI panel (BioFire Diagnostics, Salt Lake City, UT) is a multiplex, on-demand, sample-to-answer, real-time PCR assay for the syndromic diagnosis of infectious gastroenteritis that has become widely adopted and, in some instances, has replaced conventional stool culture and parasite exams. Conventional testing has historically been restricted among hospitalized patients due to low diagnostic yield, but it is not known whether use of the FilmArray GI panel should be circumscribed. Cary-Blair stool samples submitted for FilmArray GI panel in adult patients admitted to an academic hospital from August 2015 to January 2017 were included in this study. Of 481 tests performed >72 h after admission, 29 (6.0%) were positive, all for a single target, excluding Clostridium difficile When follow-up tests beyond the first positive per hospitalization were excluded, 20 (4.8%) of 414 tests were positive. There was no difference in yield by immune status. Most targets detected were viral (79% of all positives [n = 23] and 70% in unique patients [n = 14]). All four cases positive for a bacterial target could not be confirmed and presentation was atypical, suggesting possible false positives. After removing potential false positives and chronic viral shedders, the yield was 3.0% (12/406). Repeat testing performed >72 h after admission and following a negative result within the first 72 h was done in 19 patients and 100% (22/22) remained negative. The FilmArray GI panel has low yield in adult patients hospitalized for >72 h, similar to conventional stool microbiology tests, and it is reasonable to restrict its use in this population.

Quantitative Thresholds Enable Accurate Identification of Clostridium difficile Infection by the Luminex xTAG Gastrointestinal Pathogen Panel.

Clostridium difficile colonizes the gastrointestinal (GI) tract, resulting in either asymptomatic carriage or a spectrum of diarrheal illness. If clinical suspicion for C. difficile is low, stool samples are often submitted for analysis by multiplex molecular assays capable of detecting multiple GI pathogens, and some institutions do not report this organism due to concerns for high false-positive rates. Since clinical disease correlates with organism burden and molecular assays yield quantitative data, we hypothesized that numerical cutoffs could be utilized to improve the specificity of the Luminex xTAG GI pathogen panel (GPP) for C. difficile infection. Analysis of cotested liquid stool samples (n = 1,105) identified a GPP median fluorescence intensity (MFI) value cutoff of ≥1,200 to be predictive of two-step algorithm (2-SA; 96.4% concordance) and toxin enzyme immunoassay (EIA) positivity. Application of this cutoff to a second cotested data set (n = 1,428) yielded 96.5% concordance. To determine test performance characteristics, concordant results were deemed positive or negative, and discordant results were adjudicated via chart review. Test performance characteristics for the MFI cutoff of ≥150 (standard), MFI cutoff of ≥1,200, and 2-SA were as follows (respectively): concordance, 95, 96, and 97%; sensitivity, 93, 78, and 90%; specificity, 95, 98, and 98%; positive predictive value, 67, 82, and 81%;, and negative predictive value, 99, 98, and 99%. To capture the high sensitivity for organism detection (MFI of ≥150) and high specificity for active infection (MFI of ≥1,200), we developed and applied a reporting algorithm to interpret GPP data from patients (n = 563) with clinician orders only for syndromic panel testing, thus enabling accurate reporting of C. difficile for 95% of samples (514 negative and 5 true positives) irrespective of initial clinical suspicion and without the need for additional testing.

The Drift in Molecular Testing for Influenza: Mutations Affecting Assay Performance.

Influenza is associated with rapid evolution due to lack of RNA polymerase proofreading, immunogenic selection, and frequent rearrangement of gene segments. Evolutionary changes affecting the performance of diagnostic testing have long been recognized. Hence, it is not surprising that such challenges apply to nucleic acid amplification tests, even though they are designed to target highly conserved regions. Initially, case reports involved single isolates of A(H1N1)pdm09. Over the past 4 years, subtype H3N2 viruses evolved to viral clades with mutations in the WHO-recommended target region, such that almost all isolates worldwide have significantly reduced sensitivities with many commercial reverse transcription-PCR tests.

Point-Counterpoint: Meningitis/Encephalitis Syndromic Testing in the Clinical Laboratory.

INTRODUCTIONSyndromic panels were first FDA cleared for detection of respiratory pathogens in 2008. Since then, other panels have been approved by the FDA, and most recently, the FilmArray meningitis/encephalitis panel (BioFire, Salt Lake City, UT) has become available. This assay detects 14 targets within 1 h and includes pathogens that typically cause different manifestations of infection, although they infect the same organ system. Several studies have reported both false-positive and false-negative results with this test, and all agree that the cost is significant. As with other panels, health care systems have adopted different strategies for offering this assay. Some have implemented strategies to limit the use of the test to certain patient populations, others have elected not to offer the test, and others have elected not to offer the test and instead request that providers order specific PCRs for the pathogens that best fit the patient's symptoms. In this Point-Counterpoint, Jennifer Dien Bard of the Department of Pathology and Laboratory Medicine, Children's Hospital Los Angeles, and of the Keck School of Medicine at the University of Southern California explains why laboratories should offer these assays without restriction. Kevin Alby of the University of Pennsylvania explains the concerns about the use of these assays as first-line tests and why some limitations on their use might be appropriate.

Clinical evaluation of a novel and simple-to-use molecular platform for diagnosis of respiratory syncytial virus.

Rapid molecular diagnostic testing for respiratory infections can improve patient care and minimize unnecessary prescriptions of antibiotics. We present the preliminary clinical evaluation of Orion GenRead® RSV, a novel, rapid, and easy-to-use molecular test for the diagnosis of respiratory syncytial virus (RSV) infection. The sensitivity and specificity of Orion GenRead RSV were 99% and 100%, respectively. Orion GenRead RSV detected RSV-positive specimens within 15 min. The performance of Orion GenRead RSV was similar to that of the reference method and this test could rapidly detect RSV within minutes. Orion GenRead RSV is applicable for near-patient testing.

Impact of hepatitis E virus testing on the safety of blood components in Germany - results of a simulation study.

Hepatitis E virus (HEV) infections may be acquired through transfusion of blood components. As transfusion-transmitted infections mostly affect vulnerable individuals, measures to ensure the supply of safe blood components are under discussion. On the basis of the epidemiological situation in Germany, different testing strategy scenarios were investigated through simulation studies. Testing for HEV RNA by nucleic acid amplification technique (NAT) assays with a pool size of 96, and a 95% LoD of 20 IU/ml will result in an 80% reduction in expected HEV transmissions as well as of consequent chronic infections with subsequent severe complications.