Validation of an automated, end-to-end metagenomic sequencing assay for agnostic detection of respiratory viruses.

BACKGROUND: Current molecular diagnostics are limited in the number and type of detectable pathogens. Metagenomic next generation sequencing (mNGS) is an emerging, and increasingly feasible, pathogen-agnostic diagnostic approach. Translational barriers prohibit the widespread adoption of this technology in clinical laboratories. We validate an end-to-end mNGS assay for detection of respiratory viruses. Our assay is optimized to reduce turnaround time, lower cost-per-sample, increase throughput, and deploy secure and actionable bioinformatic results. METHODS: We validated our assay using residual nasopharyngeal swab specimens from Vancouver General Hospital (n = 359), RT-PCR-positive, or negative for Influenza, SARS-CoV-2, and RSV. We quantified sample stability, assay precision, the effect of background nucleic acid levels, and analytical limits of detection. Diagnostic performance metrics were estimated. RESULTS: We report that our mNGS assay is highly precise, semi-quantitative, with analytical limits of detection ranging from 103-104 copies/mL. Our assay is highly specific (100%) and sensitive (61.9% Overall: 86.8%; RT-PCR Ct < 30). Multiplexing capabilities enable processing of up to 55-specimens simultaneously on an Oxford Nanopore GridION device, with results reported within 12-hours. CONCLUSIONS: This study outlines the diagnostic performance and feasibility of mNGS for respiratory viral diagnostics, infection control, and public health surveillance. We addressed translational barriers to widespread mNGS adoption.

Retrospective validation of MetaSystems' deep-learning-based digital microscopy platform with assistance compared to manual fluorescence microscopy for detection of mycobacteria.

This study aimed to validate Metasystems' automated acid-fast bacilli (AFB) smear microscopy scanning and deep-learning-based image analysis module (Neon Metafer) with assistance on respiratory and pleural samples, compared to conventional manual fluorescence microscopy (MM). Analytical parameters were assessed first, followed by a retrospective validation study. In all, 320 archived auramine-O-stained slides selected non-consecutively [85 originally reported as AFB-smear-positive, 235 AFB-smear-negative slides; with an overall mycobacterial culture positivity rate of 24.1% (77/320)] underwent whole-slide imaging and were analyzed by the Metafer Neon AFB Module (version 4.3.130) using a predetermined probability threshold (PT) for AFB detection of 96%. Digital slides were then examined by a trained reviewer blinded to previous AFB smear and culture results, for the final interpretation of assisted digital microscopy (a-DM). Paired results from both microscopic methods were compared to mycobacterial culture. A scanning failure rate of 10.6% (34/320) was observed, leaving 286 slides for analysis. After discrepant analysis, concordance, positive and negative agreements were 95.5% (95%CI, 92.4%-97.6%), 96.2% (95%CI, 89.2%-99.2%), and 95.2% (95%CI, 91.3%-97.7%), respectively. Using mycobacterial culture as reference standard, a-DM and MM had comparable sensitivities: 90.7% (95%CI, 81.7%-96.2%) versus 92.0% (95%CI, 83.4%-97.0%) (P-value = 1.00); while their specificities differed 91.9% (95%CI, 87.4%-95.2%) versus 95.7% (95%CI, 92.1%-98.0%), respectively (P-value = 0.03). Using a PT of 96%, MetaSystems' platform shows acceptable performance. With a national laboratory staff shortage and a local low mycobacterial infection rate, this instrument when combined with culture, can reliably triage-negative AFB-smear respiratory slides and identify positive slides requiring manual confirmation and semi-quantification. IMPORTANCE: This manuscript presents a full validation of MetaSystems' automated acid-fast bacilli (AFB) smear microscopy scanning and deep-learning-based image analysis module using a probability threshold of 96% including accuracy, precision studies, and evaluation of limit of AFB detection on respiratory samples when the technology is used with assistance. This study is complementary to the conversation started by Tomasello et al. on the use of image analysis artificial intelligence software in routine mycobacterial diagnostic activities within the context of high-throughput laboratories with low incidence of tuberculosis.

In vitro assessment of antibacterial and antiviral activity of three copper products after 200 rounds of simulated use.

Copper has well-documented antibacterial effects but few have evaluated it after prolonged use and against bacteria and viruses. Coupons from three copper formulations (solid, thermal coating, and decal applications) and carbon steel controls were subjected to 200 rounds simulated cleaning using a Wiperator™ and either an accelerated hydrogen peroxide, quaternary ammonium, or artificial sweat products. Antibacterial activity against S. aureus and P. aeruginosa was then evaluated using a modified Environmental Protection Agency protocol. Antiviral activity against coronavirus (229E) and norovirus (MNV-1) surrogates was assessed using the TCID50 method. Results were compared to untreated control coupons. One hour after inoculation, S. aureus exhibited a difference in log kill of 1.16 to 4.87 and P. aeruginosa a log kill difference of 3.39-5.23 (dependent upon copper product and disinfectant) compared to carbon steel. MNV-1 demonstrated an 87-99% reduction on each copper surfaces at 1 h and 99% reduction at 2 h compared to carbon steel. Similarly, coronavirus 229E exhibited a 97-99% reduction after 1 h and 90-99% after 2 h. Simulated use with artificial sweat did not hinder the antiviral nor the antibacterial activity of Cu surfaces. Self-sanitizing copper surfaces maintained antibacterial and antiviral activity after 200 rounds of simulated cleaning.

Approach to Assessment of New Swabs and Viral Transport Media for SARS-CoV-2 Testing.

In light of the present pandemic of novel coronavirus disease 2019 (COVID-19) and the unprecedented high demand for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing worldwide, there are shortages of established specimen collection devices for respiratory viral testing for diagnostic microbiology laboratories. This creates the need to validate unverified collection devices from manufacturers that may not be a registered supplier for medical devices. As clinical laboratories do not routinely perform quality control of established collection devices, there is a need to have a systematic, robust approach to the assessment of substitute unregistered collection swabs and viral transport media (VTM). A discussion of the aspects requiring consideration when determining the suitability and implementation of new collection devices is presented. These specific assessment criteria include an inspection of device integrity, determination of swab and VTM sterility and in vitro performance, VTM stability, and examination of the clinical performance of the device. This method was used in a front-line medical microbiology laboratory on swabs and VTM from an unregistered manufacturer, with suboptimal results that precluded implementation. As the pandemic continues, it will be important for diagnostic laboratories to adopt a flexible and streamlined approach to maintaining adequate supply chains for testing reagents and materials.

Self-Collected Saline Gargle Samples as an Alternative to Health Care Worker-Collected Nasopharyngeal Swabs for COVID-19 Diagnosis in Outpatients.

We assessed the performance, stability, and user acceptability of swab-independent self-collected saliva and saline mouth rinse/gargle sample types for the molecular detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in adults and school-aged children. Outpatients who had recently been diagnosed with COVID-19 or were presenting with suspected COVID-19 were asked to have a nasopharyngeal (NP) swab collected and provide at least one self-collected sample type. Participants were also asked about sample acceptability using a five-point Likert scale. For those previously diagnosed with COVID-19, all samples underwent real-time PCR testing using a lab-developed assay, and the majority were also tested using an FDA-authorized assay. For those presenting with suspected COVID-19, only those with a positive nasopharyngeal swab sample went on to have other samples tested. Saline mouth rinse/gargle and saliva samples were tested daily at time zero, day 1, and day 2 to assess nucleic acid stability at room temperature. Fifty participants (aged 4 to 71 years) were included; of these, 40 had at least one positive sample and were included in the primary sample yield analysis. Saline mouth rinse/gargle samples had a sensitivity of 98% (39/40), while saliva samples had a sensitivity of 79% (26/33). Both saline mouth rinse/gargle and saliva samples showed stable viral RNA detection after 2 days of room temperature storage. Mouth rinse/gargle samples had the highest (mean, 4.9) and health care worker (HCW)-collected NP swabs had the lowest acceptability scores (mean, 3.1). In conclusion, saline mouth rinse/gargle samples demonstrated higher combined user acceptability ratings and analytical performance than saliva and HCW-collected NP swabs. This sample type is a promising swab-independent option, particularly for outpatient self-collection in adults and school-aged children.

One-year trial evaluating the durability and antimicrobial efficacy of copper in public transportation systems.

Surfaces on transit vehicles are frequently touched and could potentially act as reservoirs for micro-organism transmission. Regular cleaning and disinfection to minimize the spread of micro-organisms is operationally challenging due to the need to keep vehicles in circulation. The application of copper (Cu) alloys to high- touch surfaces could help reduce the risk of cross-contamination, however, little is known about the durability and efficacy of engineered copper surfaces after prolonged use. Three Cu products (decal, thermal fabrication, and alloy covers) were assessed over a 12-month period. These Cu products were randomly installed on 110 stanchions on three buses and four train (SkyTrain) cars in Vancouver and three buses, two subway cars, and two streetcars in Toronto with mirrored control surfaces directly opposite. Bacterial counts (Colony forming units, CFU) and ATP bioluminescence (ATPB) were measured every two months after peak morning routes. Durability of the Cu products were assessed monthly through visual inspection and colorimetry assays or by ex-situ microscopy. Cu products on stanchions reduced the mean colony forming units (CFU) of all vehicles by 42.7% in the mean CFU (0.573 (CI 95% 0.453-0.726), p-value < 0.001) compared to control surfaces. The three Cu products exhibited an overall 87.1% reduction in the mean ATPB readings (0.129 (CI 95% 0.059-0.285, p-value < 0.001) compared to controls. Surface Cu concentration for all three products was consistent throughout the 12-month period. Electron microscopy (SEM) and Energy-dispersive X-ray Spectroscopy (EDS) cross-sectional analysis showed no change in thickness or dealloying of Cu products, however SEM top-down analysis revealed substantial carbon accumulation on all surfaces. Cu products installed on transit vehicles maintained antimicrobial efficacy and durability after 12 months of use.

The science behind the nose: correlating volatile organic compound characterisation with canine biodetection of COVID-19.

Background: The SARS-CoV-2 pandemic stimulated the advancement and research in the field of canine scent detection of COVID-19 and volatile organic compound (VOC) breath sampling. It remains unclear which VOCs are associated with positive canine alerts. This study aimed to confirm that the training aids used for COVID-19 canine scent detection were indeed releasing discriminant COVID-19 VOCs detectable and identifiable by gas chromatography (GC-MS). Methods: Inexperienced dogs (two Labradors and one English Springer Spaniel) were trained over 19 weeks to discriminate between COVID-19 infected and uninfected individuals and then independently validated. Getxent tubes, impregnated with the odours from clinical gargle samples, used during the canines' maintenance training process were also analysed using GC-MS. Results: Three dogs were successfully trained to detect COVID-19. A principal components analysis model was created and confirmed the ability to discriminate between VOCs from positive and negative COVID-19 Getxent tubes with a sensitivity of 78% and a specificity of 77%. Two VOCs were found to be very predictive of positive COVID-19 cases. When comparing the dogs with GC-MS, F1 and Matthew's correlation coefficient, correlation scores of 0.69 and 0.37 were observed, respectively, demonstrating good concordance between the two methods. Interpretation: This study provides analytical confirmation that canine training aids can be safely and reliably produced with good discrimination between positive samples and negative controls. It is also a further step towards better understanding of canine odour discrimination of COVID-19 as the scent of interest and defining what VOC elements the canines interpret as "essential".

Legionella micdadei brain abscess.

We describe an immunocompromised patient who developed a large frontal brain abscess caused by Legionella micdadei. This is, to our knowledge, a rare case of culture-proven Legionella central nervous system infection.

Evaluation of the Aptima BV and CV/TV assays compared to conventional laboratory based testing methods for the diagnosis of vaginitis.

PURPOSE: Vaginitis is caused by bacterial vaginosis (BV), Candida vaginitis (CV) and Trichomonas vaginalis (TV). This retrospective study evaluates the performance of the Aptima CV/TV, and BV assays on the automated Panther system. METHODS: Two hundred forty-two multitest swabs were tested on the CV/TV assay and 422 on the BV assay. Positive and negative percent agreement (PPA, NPA) of the Candida glabrata (CG), Candida species group (CSG), TV and BV targets were calculated using a modified gold standard, with review of Gram smear and the usage of the Allplex Vaginitis Screening Assay to resolve discrepancies. RESULTS: The PPA and NPA were respectively 98.4% and 95.9% for BV, 100% and 95.4% for CSG, 100% and 99% for CG, and 100% and 100% for TV, and when compared to consensus results. CONCLUSION: The CV/TV and BV assays surpassed the acceptance criteria threshold of 95%, and proved to be an excellent alternative to conventional testing.

Quick versus Quantitative: Evaluation of Two Commercial Real-Time PCR Assays for the Detection of Pneumocystis jirovecii from Bronchoalveolar Lavage Fluids.

Two commercial real-time PCR assays for the detection of Pneumocystis jirovecii were compared, the quantitative RealStar P. jirovecii assay and the qualitative DiaSorin P. jirovecii assay, the latter of which can be used without nucleic acid extraction. Archived bronchoalveolar lavage (BAL) specimens (n = 66), previously tested by molecular methods, were tested by both assays, and the results were compared to the respective original result. The RealStar P. jirovecii assay demonstrated good positive percent agreement (PPA) (90% [95% confidence interval (CI), 72 to 97%]; 27/30) and negative percent agreement (NPA) (100% [95% CI, 88 to 100%]; 36/36) with the reference method. The DiaSorin P. jirovecii assay concordantly detected P. jirovecii in 19 of 24 positive BAL samples (PPA = 73% [95% CI, 52 to 88%]). All negative BAL samples gave concordant results (NPA = 100% [95% CI, 87 to 100%]; 34/34). Discordant results occurred mostly in samples with low fungal loads. In conclusion, the RealStar assay demonstrated good concordance with reference results, and the DiaSorin P. jirovecii assay performed well for negative BAL and positive BAL samples with P. jirovecii concentrations of greater than 260 copies/mL. IMPORTANCE Pneumonia, caused by the opportunistic fungus Pneumocystis jirovecii, poses a significant risk for immunocompromised individuals. Laboratory testing for P. jirovecii is progressively shifting toward the use of molecular tests such as real-time PCR; however, this is often performed at reference laboratories. Many frontline laboratories are looking into improving their service and reducing turnaround times for obtaining P. jirovecii results by bringing molecular P. jirovecii testing in-house. We evaluated and compared two commercial real-time PCR assays with different workflows for the detection of P. jirovecii from bronchoalveolar lavage specimens. The RealStar P. jirovecii assay requires nucleic acid extraction and provides a quantification of fungal load for positive samples. The DiaSorin P. jirovecii assay offers a simple workflow without nucleic extraction from patient samples and qualitative results. Results from this study provide valuable information on performance and workflow considerations for laboratories that wish to implement P. jirovecii molecular testing.

A nested cohort 5-year Canadian surveillance of Gram-negative antimicrobial resistance for optimized antimicrobial therapy.

We analyzed 5 years (2016-2020) of nested Canadian data from the Study for Monitoring Antimicrobial Resistance Trends (SMART) to identify pathogen predominance and antimicrobial resistance (AMR) patterns of adult Gram-negative infections in Canadian health care and to complement other public surveillance programs and studies in Canada. A total of 6853 isolates were analyzed from medical (44%), surgical (18%), intensive care (22%) and emergency units (15%) and from respiratory tract (36%), intra-abdominal (25%), urinary tract (24%) and bloodstream (15%) infections. Overall, E. coli (36%), P. aeruginosa (18%) and K. pneumoniae (12%) were the most frequent isolates and P. aeruginosa was the most common respiratory pathogen. 18% of Enterobacterales species were ESBL positive. Collective susceptibility profiles showed that P. aeruginosa isolates were highly susceptible (> 95%) to ceftolozane/tazobactam and colistin, though markedly less susceptible (58-74%) to other antimicrobials tested. Multi-drug resistance (MDR) was present in 10% of P. aeruginosa isolates and was more frequent in those from respiratory infections and from ICU than non-ICU locations. Of P. aeruginosa isolates that were resistant to combinations of ceftazidime, piperacillin/tazobactam and meropenem, 73-96% were susceptible to ceftolozane/tazobactam over the period of the study. These national data can now be combined with clinical prediction rules and genomic data to enable expert antimicrobial stewardship applications and guide treatment policies to optimize adult patient care.

Abbott ID NOW™ COVID-19 assay: do not discard the swab.

We compared the performance of ID NOW™ COVID-19 assay nasal swabs with RT-PCR of nasopharyngeal swabs for SARS-CoV-2 in an outbreak setting, determining whether addition of RT-PCR of residual nasal swabs (rNS) (post ID NOW™ elution) would increase overall analytic sensitivity. Devices were placed at 2 long term and 1 acute care sites and 51 participants were recruited. Prospective paired nasopharyngeal and nasal samples were collected for RT-PCR and ID NOW™.  ID NOW™ had a positive and negative categorical agreement of 86% and 93% compared to RT-PCR of nasopharyngeal swabs. Sensitivity and specificity of the ID NOW™ was 86% and 100%, positive and negative predictive value was 100% and 95% (COVID-19 positivity rate: 8%). Addition of rNS RT-PCR increased the positive and negative categorical agreement to 93% and 97%. Based on these results, we propose an alternative workflow which includes complementary testing of rNS on a secondary assay.

Alterations in the nasopharyngeal microbiome associated with SARS-CoV-2 infection status and disease severity.

OBJECTIVES: The COVID-19 pandemic and ensuing public health emergency has emphasized the need to study SARS-CoV-2 pathogenesis. The human microbiome has been shown to regulate the host immune system and may influence host susceptibility to viral infection, as well as disease severity. Several studies have assessed whether compositional alterations in the nasopharyngeal microbiota are associated with SARS-CoV-2 infection. However, the results of these studies were varied, and many did not account for disease severity. This study aims to examine whether compositional differences in the nasopharyngeal microbiota are associated with SARS-CoV-2 infection status and disease severity. METHODS: We performed Nanopore full-length 16S rRNA sequencing on 194 nasopharyngeal swab specimens from hospitalized and community-dwelling SARS-CoV-2-infected and uninfected individuals. Sequence data analysis was performed using the BugSeq 16S analysis pipeline. RESULTS: We found significant beta (PERMANOVA p < 0.05), but not alpha (Kruskal-Wallis p > 0.05) diversity differences in the nasopharyngeal microbiota among our study groups. We identified several differentially abundant taxa associated with SARS-CoV-2 infection status and disease severity using ALDEx2. Finally, we observed a trend towards higher abundance of Enterobacteriaceae in specimens from hospitalized SARS-CoV-2-infected patients. CONCLUSIONS: This study identified several alterations in the nasopharyngeal microbiome associated with SARS-CoV-2 infection status and disease severity. Understanding the role of the microbiome in infection susceptibility and severity may open new avenues of research for disease prevention and treatment.

Automated 16S Sequencing Using an R-Based Analysis Module for Bacterial Identification.

Sanger sequencing of the 16S rRNA gene is routinely used for the identification of bacterial isolates. However, this method is still performed mostly in more-specialized reference laboratories, and traditional protocols can be labor intensive. In this study, 99 clinical bacterial isolates were used to validate a fast, simplified, and largely automated protocol for 16S sequencing. The workflow combines real-time PCR of the first 500 bp of the bacterial 16S rRNA gene and amplicon sequencing on an automated, cartridge-based sequence analyzer. Sequence analysis, NCBI BLAST search, and result interpretation were performed using an automated R-based script. The automated workflow and R analysis described here produced results equal to those of manual sequence analysis. Of the 96 sequences with adequate quality, 90 were concordantly identified to the genus (n = 62) or species level (n = 28) compared with routine laboratory identification of the organism. One organism identification was discordant, and 5 resulted in an inconclusive identification. For sequences that gave a valid result, the overall accuracy of identification to at least the genus level was 98.9%. This simplified sequencing protocol provides a standardized approach to clinical 16S sequencing, analysis, and quality control that would be suited to frontline clinical microbiology laboratories with minimal experience. IMPORTANCE Sanger sequencing of the 16S rRNA gene is widely used as a diagnostic tool for bacterial identification, especially in cases where routine diagnostic methods fail to provide an identification, for organisms that are difficult to culture, or from specimens where cultures remain negative. Our simplified protocol is tailored toward use in frontline laboratories with little to no experience with sequencing. It provides a highly automated workflow that can deliver fast results with little hands-on time. Implementing 16S sequencing in-house saves additional time that is otherwise required to send out isolates/specimens for identification to reference laboratories. This makes results available much faster to physicians who can in turn initiate or adjust patient treatment accordingly.

In search of COVID-19: The ability of biodetection canines to detect COVID-19 odours from clinical samples.

BACKGROUND: COVID-19 continues to be a public health concern and the demand for fast and reliable screening tests remains. SARS-CoV-2 infection in humans generates a specific volatile organic compound signature; this 'volatilome' could be used to deploy highly trained canine scent detection teams if they could reliably detect odours from infected individuals. METHODS: Two dogs were trained over 19 weeks to discriminate between the odours produced by breath, sweat, and gargle specimens collected from SARS-CoV-2 infected and uninfected individuals. Third party validation was conducted in a randomized double-blinded controlled manner using fresh odours obtained from different patients within 10 days of their first positive SARS-CoV-2 molecular result. RESULTS: Cumulatively, the dogs completed 299 training sessions on odours from 108 unique participants. Validation was conducted over 2 days with 120 new odours. Twenty-four were odours collected from SARS-CoV-2 positive individuals (8 gargle, 8 sweat, and 8 breath); 21 were from SARS-CoV-2 negative individuals (5 gargle, 8 sweat, and 8 breath) and the remaining 75 were odours that the dogs could have associated with the target odour during training. The dogs were able to identify odours from positive specimens with an overall sensitivity of 100% and a specificity of 87.5%. Considering a community prevalence of 10%, the combined negative predictive value of the dogs was 100% and the positive predictive value was 47.1%. CONCLUSIONS: Multiple dogs can be trained to accurately detect SARS-CoV-2 positive individuals. Future research is required to determine how and when canine scent detection teams should be deployed.

HISTORIQUE: La COVID-19 continue d'être une préoccupation sanitaire, et la demande de tests de dépistage rapides et fiables se maintient. L'infection par le SRAS-CoV-2 chez les humains produit une signature composée organique volatile bien précise. Ce « volatilome ¼ pourrait être utilisé pour déployer des équipes canines hautement formées et spécialisées dans la détection des odeurs afin d'établir si elles peuvent détecter les odeurs des personnes infectées avec fiabilité. MÉTHODOLOGIE: Deux chiens ont été formés pendant 19 semaines pour distinguer les odeurs émanantes des échantillons d'haleine, de sueur et de gargarisme prélevés chez des personnes infectées et non infectées par le SRAS-CoV-2. Les chercheurs ont effectué une validation par des tiers dans le cadre d'une étude contrôlée randomisée à double insu au moyen d'odeurs fraîches obtenues auprès de divers patients dans les dix jours suivant le premier résultat moléculaire positif au SRAS-CoV-2. RÉSULTATS: Dans l'ensemble, les chiens ont effectué 299 séances de formation sur les odeurs de 108 participants uniques. La validation a eu lieu sur deux jours à partir de 120 nouvelles odeurs. Ainsi, 24 odeurs provenaient de personnes positives au SRAS-CoV-2 (8 échantillons de gargarisme, 8 de sueur et 8 d'haleine); 21 provenaient de personnes négatives au SRAS-CoV-2 (5 échantillons de gargarisme, 8 de sueur et 8 d'haleine) et les 75 autres étaient des odeurs que les chiens avaient pu associer à l'odeur cible pendant la formation. Les chiens ont été en mesure de dépister les odeurs des échantillons positifs selon une sensibilité globale de 100 % et une spécificité de 87,5 %. Étant donné une prévalence communautaire de 10 %, la valeur prédictive négative combinée des chiens s'élevait à 100 % et la valeur prédictive positive, à 47,1 %. CONCLUSIONS: De nombreux chiens peuvent être formés pour dépister avec exactitude les personnes positives au SRAS-CoV-2. De futures recherches devront être réalisées pour déterminer quand et comment déployer ces équipes canines spécialisées en biodétection.

BugSplit enables genome-resolved metagenomics through highly accurate taxonomic binning of metagenomic assemblies.

A large gap remains between sequencing a microbial community and characterizing all of the organisms inside of it. Here we develop a novel method to taxonomically bin metagenomic assemblies through alignment of contigs against a reference database. We show that this workflow, BugSplit, bins metagenome-assembled contigs to species with a 33% absolute improvement in F1-score when compared to alternative tools. We perform nanopore mNGS on patients with COVID-19, and using a reference database predating COVID-19, demonstrate that BugSplit's taxonomic binning enables sensitive and specific detection of a novel coronavirus not possible with other approaches. When applied to nanopore mNGS data from cases of Klebsiella pneumoniae and Neisseria gonorrhoeae infection, BugSplit's taxonomic binning accurately separates pathogen sequences from those of the host and microbiota, and unlocks the possibility of sequence typing, in silico serotyping, and antimicrobial resistance prediction of each organism within a sample. BugSplit is available at https://bugseq.com/academic .

Evaluation of the BioFire® COVID-19 test and Respiratory Panel 2.1 for rapid identification of SARS-CoV-2 in nasopharyngeal swab samples.

The BioFire® COVID-19 Test and Respiratory Panel 2.1 (RP2.1) are rapid, fully automated assays for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in nasopharyngeal swabs. In the case of the RP2.1, an additional 21 viral and bacterial pathogens can be detected. Both tests have received emergency use authorization from the U.S. Food & Drug Administration and Interim Order authorization from Health Canada for use in clinical laboratories. We evaluated the performance characteristics of these tests in comparison to a laboratory-developed real-time PCR assay targeting the viral RNA-dependent RNA polymerase and E genes. A total of 78 tests were performed using the BioFire COVID-19 Test, including 30 clinical specimens and 48 tests in a limit of detection study; 57 tests were performed using the RP2.1 for evaluation of SARS-CoV-2 detection, including 30 clinical specimens and 27 tests for limit of detection. Results showed 100% concordance between the BioFire assays and the laboratory-developed test for all clinical samples tested, and acceptable performance of both BioFire assays at their stated limits of detection. Conclusively, the BioFire COVID-19 Test and RP2.1 are highly sensitive assays that can be effectively used in the clinical laboratory for rapid SARS-CoV-2 testing.