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Hepatitis C virus (HCV) elimination is an important global public health goal. However, the United States is not on track to meet the World Health Organization's 2030 targets for HCV elimination. Recently, the White House proposed an HCV elimination plan that includes point-of-care (POC) HCV RNA testing, which is currently in use in many countries but is not approved in the United States. POC HCV RNA testing is crucial for implementing community-based testing and for enabling test-and-treat programs, assessing cure, and monitoring for reinfection. Here, we review the status of POC HCV RNA testing in the United States, discuss factors that are needed for successful implementation, and issue specific public health and policy recommendations that would allow for the use of POC HCV RNA testing to support HCV elimination. [ABSTRACT FROM AUTHOR]

Thyroid nodules remain a common diagnosis encountered in general and endocrine surgical practice, and thyroid cancer appears to be increasing in incidence in both the United States and worldwide. The understanding of the complex genetics surrounding thyroid neoplasia has increased substantially in recent years and, consequently, has become a consideration in risk stratification, diagnosis, prognosis, and treatment. Molecular genetic analysis of thyroid nodules is now a readily available technology for diagnostic purposes via analysis of fine-needle aspiration biopsy (FNAB) specimens. Although their routine use is controversial, they may aid in selecting which patients require surgery and those who may be safely observed. In the following review, the genetics of the tumorigenesis of thyroid cancer are reviewed, focusing on the most common and clinically relevant of the literally hundreds of known mutations. Following this, the current status of the use of genetic analysis and molecular diagnostics in the workup of thyroid nodules and the diagnosis of differentiated thyroid cancer is explored. Finally, evolving concepts relating to the use of thyroid cancer genetics in individualizing treatment planning, follow-up, and management of recurrent disease is discussed. The goal is to provide the general surgeon with a working knowledge of the most common genetic alterations present in differentiated thyroid cancer, their relevance in clinical practice, and how they impact prognosis and treatment. [ABSTRACT FROM AUTHOR]

Tick-borne diseases (TBDs) have become a significant public health concern in the United States over the past few decades. The increasing incidence and geographical spread of these diseases have prompted the implementation of robust surveillance systems to monitor their prevalence, distribution, and impact on human health. This comprehensive review describes key disease features with the geographical distribution of all known tick-borne pathogens in the United States, along with examining disease surveillance efforts, focusing on strategies, challenges, and advancements. Surveillance methods include passive and active surveillance, laboratory-based surveillance, sentinel surveillance, and a One Health approach. Key surveillance systems, such as the National Notifiable Diseases Surveillance System (NNDSS), TickNET, and the Tick-Borne Disease Laboratory Network (TBDLN), are discussed. Data collection and reporting challenges, such as underreporting and misdiagnosis, are highlighted. The review addresses challenges, including lack of standardization, surveillance in non-human hosts, and data integration. Innovations encompass molecular techniques, syndromic surveillance, and tick surveillance programs. Implications for public health cover prevention strategies, early detection, treatment, and public education. Future directions emphasize enhanced surveillance networks, integrated vector management, research priorities, and policy implications. This review enhances understanding of TBD surveillance, aiding in informed decision-making for effective disease prevention and control. By understanding the current surveillance landscape, public health officials, researchers, and policymakers can make informed decisions to mitigate the burden of (TBDs). [ABSTRACT FROM AUTHOR]

Spotted-wing Drosophila, Drosophila suzukii, is an economically important pest of small fruits worldwide. Currently, the timing of management strategies relies on detection of adult flies captured in baited monitoring traps; however, identifying D. suzukii in trap catch based on morphology can be challenging for growers. DNA-based diagnostic methods such as loop-mediated isothermal amplification (LAMP) have the potential to improve D. suzukii detection. This study evaluated a LAMP assay as a diagnostic tool to discriminate between D. suzukii and closely related drosophilid species found commonly in monitoring traps in the Midwestern United States. Targeting the mitochondrial cytochrome oxidase I (COI) gene, we found the LAMP assay accurately detected D. suzukii with as little as 0.1 ng/µl of DNA at 63 °C for 50 min. Under these optimal incubation conditions, D. suzukii could be discriminated from D. affinis and D. simulans consistently, when specimens collected from liquid monitoring traps were tested independently. Compared to other DNA-based diagnostic tools for D. suzukii, LAMP offers unique benefits: DNA extraction is not required, testing occurs at one temperature in less than 1 h, and positive results are visible as a colorimetric change from pink to yellow. The LAMP assay for D. suzukii can reduce reliance on morphological identification, enhance the adoption of monitoring tools, and improve accuracy of detection. Further optimization can be conducted to evaluate the accuracy and sensitivity of results when a mixture of DNA from both D. suzukii and congener flies are tested in a single LAMP reaction. [ABSTRACT FROM AUTHOR]

Molecular advances facilitate fruit export by improving rapid pest diagnosis by polymerase chain reaction (PCR) and advanced sequencing technology. Improved pest detection provides timely certification of the quarantine pest-free status in the commodity being exported, avoiding unnecessary commodity treatment. The U.S.–Japan Systems Approach to export fresh cherries from the Western United States that targets the codling moth, Cydia pomonella (Linnaeus) (Lepidoptera: Tortricidae), is used as an example. Suspect codling moth larvae interdicted at cherry packing houses are distinguished by PCR from other internal fruit moth larvae such as the oriental fruit moth, Grapholita molesta (Busck) (Lepidoptera: Tortricidae); lesser appleworm, G. prunivora (Walsh) (Lepidoptera: Tortricidae); cherry fruitworm, G. packardi (Zeller) (Lepidoptera: Tortricidae); and filbertworm, Cydia latiferreana (Walsingham) (Lepidoptera: Tortricidae). Identification is confirmed by sequencing the amplicon of a 301 bp region of the COI gene produced by PCR of the DNA from a suspect moth and comparing this sequence of COI gene sequences of other internal fruit feeders of pome fruit. This sequence comparison results in unambiguous pest identification. These findings are discussed in the context of systems approach research to meet evolving needs of phytosanitary requirements for global export of fruits. [ABSTRACT FROM AUTHOR]

Objective: In this study, the performance of a commercially available malaria LAMP assay (Alethia® Malaria Plus LAMP) was evaluated using retrospective clinical samples obtained from travelers returning to the United States of America (USA). Recently, several laboratories in non-malaria endemic countries evaluated the use of the loop mediated isothermal amplification (LAMP) assays for the diagnosis of imported malaria cases. These tests are simpler than polymerase-chain reaction (PCR)-based assays and were shown to have high sensitivity. Much of malaria diagnoses in the USA, is undertaken at the state level using mainly microscopy and rapid diagnostic tests (RDTs). However, molecular tools offer greater sensitivity over microscopy and RDTs. A reliable, easy to perform molecular assay can provide a test of choice for the accurate detection of malaria parasites in places where expert microscopy is lacking and/or for the detection of low-parasite density infections. Results: The Alethia® Malaria Plus LAMP assay was easy to use, had similar test performances as the real-time PCR reference test and results were obtained faster (within 1 h) than the reference test. The sensitivity of the assay was 100% with a kappa score of 1 when compared to the reference PET-PCR assay. [ABSTRACT FROM AUTHOR]

The current recommended testing algorithm for assessing the alloimmunized pregnancy utilized by many obstetricians in the United States (US) fails to consider the most recent evidence, placing fetuses, and mothers at unnecessary risk of poor outcome or death. This narrative review of the current landscape of fetal red blood cell (RBC) antigen testing evaluates the history of hemolytic disease of the fetus and newborn (HDFN) and how its discovery has continued to influence practices in the US today. We compare current US-based HDFN practice guidelines with those in Europe. We also provide transfusion medicine and hematology perspectives and recommendations addressing the limitations of US practice, particularly regarding paternal RBC antigen testing, and discuss the most valuable alternatives based on decades of data and evidence-based recommendations from Europe., Competing Interests: Declaration of competing interest The authors have no conflict of interest to disclose in relation to the submitted review., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Objectives: At the onset of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic in the United States, testing was limited to the Centers for Disease Control and Prevention-developed reverse transcription polymerase chain reaction assay. The urgent and massive demand for testing prompted swift development of assays to detect SARS-CoV-2. The objective of this study was to assess the accuracy of these newly developed tests.Methods: The American Proficiency Institute sent 2 test samples to 346 clinical laboratories in order to assess the accuracy of SARS-CoV-2 assays. The positive sample, containing 5,175 viral copies/mL, was fully extractable with SARS-CoV-2 viral capsid protein and RNA. The negative sample, with 3,951 viral copies/mL, contained recombinant virus particles with sequences for targeting human RNAase P gene sequences.Results: Of the laboratories submitting results, 97.4% (302/310) correctly detected the virus when present and 98.3% (296/301) correctly indicated when the virus was not present. Among incorrect results reported in this proficiency challenge, 76.9% (10/13) were likely related to clerical error. This accounts for 1.6% (10/611) of all reported results.Conclusions: Overall performance in this SARS-CoV-2 RNA detection challenge was excellent, providing confidence in the results of these new molecular tests and assurance for the clinical and public health decisions based on these test results. [ABSTRACT FROM AUTHOR]

Urine culture and sensitivity (C&S) remains the gold standard diagnostic for urinary tract infections (UTI). To reduce the use of inaccurate or broad-spectrum empiric antimicrobials, rapid identification and quantification (IDQ) and antimicrobial susceptibility testing (AST) with results within 30 and 150 min, respectively, are under development. To assess the impact of rapid diagnostics, five UTI vignettes were constructed, and ninety-one United States physicians were surveyed regarding their diagnostic, management, and antimicrobial choices before and after IDQ and AST results. Rapid diagnostics increased the postponement of antimicrobial therapy pending AST results from 16% to 38% and 5% to 54% in Vignettes 1 and 2 and reduced the use of ineffective antibiotics from 41% to 0% and 69% to 0% in Vignettes 2 and 4. Rapid diagnostics increased the use of narrow spectrum agents in the five vignettes, indicating its potential to revitalize physician responsibility in antimicrobial stewardship. [ABSTRACT FROM AUTHOR]

Faecal pollution of water and the resulting potential presence of human enteric pathogens is a predominant threat to public health. Microbiological water quality can be assessed by the detection of standard faecal indicator bacteria (SFIB) such as E. coli or certain Enterococcus species. In recent years, isothermal amplification methods have become a useful alternative to polymerase chain reaction (PCR), allowing molecular diagnostics with simple or no instrumentation. In this study, a novel screening method for the molecular detection of Enterococcus spp. by loop-mediated isothermal amplification (LAMP) is described. A set of six specific LAMP primers was designed to amplify a diagnostic fragment of the Enterococcus 23S rRNA gene, which is present in several enterococcal species targeted by quantitative PCR (qPCR), which is the standard technique recommended by the US Environmental Protection Agency. Sensitivity and specificity tests were performed using a set of 30 Enterococcus and non-target bacterial reference strains. It is shown that LAMP is equally sensitive and even more specific than the qPCR assay. A dilution series of Enterococcus faecalis DNA revealed that the LAMP method can reliably detect 130 DNA target copies per reaction within 45 min. Additionally, enterococci isolated from Austrian surface waterbodies, as well as a set of DNA extracts from environmental waters, were tested. Contingency analysis demonstrated a highly significant correlation between the results of the developed LAMP assay and the reference qPCR method. Furthermore, a simple staining procedure with a fluorescence dye demonstrated the identification of amplified products by eye. In conclusion, this method is an important component for the efficient screening and testing of water samples in low-resource settings lacking sophisticated laboratory equipment and highly trained personnel, requiring only a simple heating block. [ABSTRACT FROM AUTHOR]

Thyroid nodules are quite common, and the determination of a nodule of concern is complex, involving serum testing, radiology and, in some cases, pathological evaluation. For those nodules that raise clinical concern of neoplasia, fine needle aspiration biopsy is the gold standard for evaluation; however, in up to 30% of cases, results are indeterminate for malignancy, and further testing is needed. Advances in molecular testing have shown it to be of benefit for both diagnostic and prognostic purposes, and its use has become an integral part of thyroid cancer management in the United States and in several global nations. After The Cancer Genome Atlas (TCGA) consortium published its molecular landscape of papillary thyroid carcinoma (PTC) and reduced the "black matter" in PTC from 25% to 3.5%, further work ensued to clarify the remaining fraction not neatly attributed to the BRAF V600E -like or RAS -like phenotypes of the TCGA. Over the past decade, commercial molecular platforms have been refined as data accrues, and they increasingly cover most genetic variants of thyroid carcinomas. Molecular reporting focuses on the nodule tested, including related clinical information for that nodule (size of nodule, Bethesda category, etc.). This results in a comprehensive report to physicians that may also include patient-directed, clear language that facilitates conversations about nodule management. In cases of advanced or recurrent disease, molecular testing may become essential for devising an individual therapeutic plan. In this review, we focus on the evolution of integrated molecular testing in thyroid nodules, and how our understanding of tumor genetics, combined with histopathology, is driving the next generation of rational patient management, particularly in the context of emerging small, targetable therapeutics.

While the sensitivity of detection of pneumococcal carriage can be improved by testing respiratory tract samples with quantitative PCR (qPCR), concerns have been raised regarding the specificity of this approach. We therefore investigated the reliability of the widely used lytA qPCR assay when applied to saliva samples from older adults in relation to a more specific qPCR assay ( piaB ). During the autumn/winter seasons of 2018/2019 and 2019/2020, saliva was collected at multiple time points from 103 healthy adults aged 21 to 39 ( n = 34) and >64 ( n = 69) years ( n = 344 total samples). Following culture enrichment, extracted DNA was tested using qPCR for piaB and lytA . By sequencing the variable region of rpsB (S2 typing), we identified the species of bacteria isolated from samples testing lytA -positive only. While 30 of 344 (8.7%) saliva samples (16.5% individuals) tested qPCR-positive for both piaB and lytA , 52 (15.1%) samples tested lytA -positive only. No samples tested piaB -positive only. Through extensive reculture attempts of the lytA -positive samples collected in 2018/2019, we isolated 23 strains (in 8 samples from 5 individuals) that were also qPCR-positive for only lytA . Sequencing determined that Streptococcus mitis and Streptococcus infantis were predominantly responsible for this lytA -positive qPCR signal. We identified a comparatively large proportion of samples generating positive signals with the widely used lytA qPCR and identified nonpneumococcal Streptococcus species responsible for this signal. This highlights the importance of testing for the presence of multiple gene targets in tandem for reliable and specific detection of pneumococcus in polymicrobial respiratory tract samples. IMPORTANCE Testing saliva samples with quantitative PCR (qPCR) improves the sensitivity of detection of pneumococcal carriage. The qPCR assay targeting lytA , the gene encoding the major pneumococcal autolysin, has become widely accepted for the identification of pneumococcus and is even considered the "gold standard" by many. However, when applying this approach to investigate the prevalence of pneumococcal carriage in adults in New Haven, CT, USA, we identified nonpneumococcal Streptococcus spp. that generate positive signals in this widely used assay. By testing also for piaB (encoding the iron acquisition ABC transporter lipoprotein, PiaB), our findings demonstrate the importance of testing for the presence of multiple gene targets in tandem for reliable molecular detection of pneumococcus in respiratory tract samples; targeting only lytA may lead to an overestimation of true carriage rates., Competing Interests: The authors declare a conflict of interest. A.L.W has received consulting and/or advisory board fees from Pfizer, Diasorin, PPS Health, Co-Diagnostics, and Global Diagnostic Systems for work unrelated to this project, and is Principal Investigator on research grants from Pfizer, Merck, and Flambeau Diagnostics to Yale University. D.M.W has received consulting fees from Pfizer, Merck, GSK, Affinivax, and Matrivax for work unrelated to this project and is Principal Investigator on research grants and contracts with Pfizer and Merck to Yale University. All other co-authors declare no potential conflict of interest.

This paper is prompted by a series of recent high-profile decisions emanating from the apex courts in the US and Australia – namely,MayoandMyriad– on the patent eligibility of gene-based inventions. Adopting a comparative approach, this paper critically examines whether isolated gene sequences and diagnostic methods qualify as patentable subject matter in several leading jurisdictions, including the US, Australia, Europe and the UK. An attempt will also be made, after carefully considering various arguments on both sides of the policy debate, at predicting what the future might hold (or, perhaps more accurately, at suggesting what the future ought to be) for gene patenting and the patenting of diagnostic methods in the global biotechnology landscape. [ABSTRACT FROM PUBLISHER]

Background: Treatment of Neisseria gonorrhoeae infection is empirical and based on population-wide susceptibilities. Increasing antimicrobial resistance underscores the potential importance of rapid diagnostic tests, including sequence-based tests, to guide therapy. However, the usefulness of sequence-based diagnostic tests depends on the prevalence and dynamics of the resistance mechanisms.Methods: We define the prevalence and dynamics of resistance markers to extended-spectrum cephalosporins, macrolides, and fluoroquinolones in 1102 resistant and susceptible clinical N. gonorrhoeae isolates collected from 2000 to 2013 via the Centers for Disease Control and Prevention's Gonococcal Isolate Surveillance Project.Results: Reduced extended-spectrum cephalosporin susceptibility is predominantly clonal and associated with the mosaic penA XXXIV allele and derivatives (sensitivity 98% for cefixime and 91% for ceftriaxone), but alternative resistance mechanisms have sporadically emerged. Reduced azithromycin susceptibility has arisen through multiple mechanisms and shows limited clonal spread; the basis for resistance in 36% of isolates with reduced azithromycin susceptibility is unclear. Quinolone-resistant N. gonorrhoeae has arisen multiple times, with extensive clonal spread.Conclusions: Quinolone-resistant N. gonorrhoeae and reduced cefixime susceptibility appear amenable to development of sequence-based diagnostic tests, whereas the undefined mechanisms of resistance to ceftriaxone and azithromycin underscore the importance of phenotypic surveillance. The identification of multidrug-resistant isolates highlights the need for additional measures to respond to the threat of untreatable gonorrhea. [ABSTRACT FROM AUTHOR]

Reticulitermes termites play key roles in dead wood decomposition and nutrient cycling in forests. They also damage man-made structures, resulting in considerable economic loss. In the eastern United States, five species (R. flavipes, R. virginicus, R. nelsonae, R. hageni and R. malletei) have overlapping ranges and are difficult to distinguish morphologically. Here we present a molecular tool for species identification. It is based on polymerase chain reaction (PCR) amplification of a section of the mitochondrial cytochrome oxidase subunit II gene, followed by a three-enzyme restriction fragment length polymorphism (RFLP) assay, with banding patterns resolved via agarose gel electrophoresis. The assay was designed using a large set of training data obtained from a public DNA sequence database, then evaluated using an independent test panel of Reticulitermes from the Southern Appalachian Mountains, for which species assignments were determined via phylogenetic comparison to reference sequences. After refining the interpretive framework, the PCR-RFLP assay was shown to provide accurate identification of four co-occurring species (the fifth species, R. hageni, was absent from the test panel, so accuracy cannot yet be extended to training data). The assay is cost- and time-efficient, and will help improve knowledge of Reticulitermes species distributions. [ABSTRACT FROM AUTHOR]

Molecular diagnostics are fast becoming a big business, with the promise of personalized medicine fueling the growth of "blockbuster" tests with high expectations for health system impact and commercial success. We investigate the polycentric regulatory regime for molecular diagnostics in the US, drawing attention to the prominent role of coverage and reimbursement systems in setting regulatory standards for this industry. We hone in on the private consultants who assist molecular diagnostics companies to gain broad clinical uptake of their products. Through a web-based search of consulting companies, analysis of their online materials, and 13 qualitative interviews with consultants, we describe the role of these actors in the coverage and reimbursement of novel diagnostics and highlight the production of evidence as a critical part of the process. We argue that consultants operate as regulatory intermediaries, helping to develop the evidentiary standards for payment decisions that ultimately benefit their clients, the manufacturers. We suggest that public policy discussions over how best to realize the promise of personalized medicine should be re-oriented to consider whose interests are represented in the regulatory regime governing access to these technologies. • Consultants help molecular diagnostic manufacturers commercialize their products. • Diagnostics can bypass statutory regulation but require reimbursement for uptake. • Consultants assemble actors to produce the evidence needed for reimbursement. • Consultants act as regulatory intermediaries to shape the regulatory regime. [ABSTRACT FROM AUTHOR]

The article offers information on a study related to the development of a molecular diagnostic marker for the identification of Scirthrips dorsalis (S. dorsalis), an invasive pest. It further informs that Scirthrips pose a potential threat to the U.S. agriculture and trade with an estimated loss of 5.98 billion dollar upon chilli thrips infestation. It mentions that several molecular techniques have been developed for species diagnosis due to their minute size and cryptic behavior.

Abstract: The diagnostic potential of RT-PCR for detection of bluetongue virus (BTV) ribonucleic acid (RNA) sequence in cell culture and tissue samples from infected ruminants from United States, Sudan, South Africa and Senegal, was evaluated. The non structural protein 1 (NS1) gene of North American BTV serotype 11 was targeted for PCR amplification. The United States BTV serotypes 2, 10, 11, 13 and 17 and the Sudanese BTV serotypes 1, 2, 4 and 16 and BTV serotype 4 from South Africa and BTV serotype 2 from Senegal were studied. RNAs from all BTV field isolates used in this study, propagated in cell cultures, were detected by the described RT-PCR-based assay. The first specific 790bp BTV PCR products were amplified using a pair of outer primers (BTV1 and BTV2). Specificity of the PCR products was confirmed by a nested amplification of a 520bp PCR product using a pair of internal (nested) primers (BTV3 and BTV4). The BTV PCR products were visualized on ethidium bromide-stained agarose gels. Amplification products were not detected when the RT-PCR-based assay was applied to RNAs from closely related orbiviruses including, epizootic hemorrhagic disease virus (EHDV) prototypes serotypes 1, 2, 4; RNA from Sudanese isolate of palyam orbiviruses serogroup and total nucleic acid extracts from uninfected Vero cells. Application of the nested BTV RT-PCR to clinical samples resulted in amplification of BTV RNA from blood and serum samples from goats experimentally infected with BTV4 and from naturally infected sheep, goats, cattle and deer. The results of this study indicated that this RT-PCR assay could be applied for rapid detection of BTV, in cell culture and clinical samples from susceptible ruminants during an outbreak of the disease, in the United States and African. [Copyright &y& Elsevier]

Acute and chronic diarrheal illness secondary to gastrointestinal infection is a significant cause of morbidity and mortality around the world. A cornerstone of management includes prompt diagnosis and appropriate treatment of culprit pathogens. Timely diagnosis can improve patient care, assist in infection control, and prevent disease outbreaks. Historical methods of diagnosis include traditional culture methods and stool analysis. These are limited by long turnaround time and inability to simultaneously assess multiple pathogens. The advent of multiplexed nucleic acid amplification tests first began with the Food and Drug Administration-approved respiratory virus multiplex polymerase chain reaction (PCR) panel in 2009, followed by gastrointestinal infections in 2013, and neurological infections in 2014. We conducted a review of current literature pertaining to the clinical utility of a gastrointestinal multiplex PCR in management of acute and chronic diarrhea in patients. To date, seven platforms approved by the US Food and Drug Administration are used in detection of various bacterial, viral, and parasitic causative organisms for diagnosis of gastrointestinal infections. The sensitivity and specificity of each assay vary depending on the tested organism. Interpretation of a positive result has to be tailored to the clinical context. Further studies are required to establish the utility of gastrointestinal multiplex PCR from a cost-based perspective, whether specific enteropathogens such as Clostridioides difficile are better assessed with toxin gene detection and whether new parameters such as cycle threshold values can improve clinical application of test results., (© 2021 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

The 2019 coronavirus disease (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 virus, caused a worldwide pandemic in 2020 and is the most urgent health issue worldwide. In this review, we highlight the details of Food and Drug Administration-Emergency Use Authorizations approved diagnostics kits, focusing on the similarities and differences. It is essential to understand the currently available options and the advantages and disadvantages each provides to select the appropriate products that maximize the testing efficiency. We believe this work will provide a holistic evaluation of the current COVID-19 diagnostic resources, including variations across the countries, and guide developing novel diagnostic techniques to improve and optimize the current testing options.

Objectives: To identify factors associated with COVID-19 test positivity and assess viral and antibody test concordance., Design: Observational retrospective cohort study., Setting: Optum de-identified electronic health records including over 700 hospitals and 7000 clinics in the USA., Participants: There were 891 754 patients who had a COVID-19 test identified in their electronic health record between 20 February 2020 and 10 July 2020., Primary and Secondary Outcome Measures: Per cent of viral and antibody tests positive for COVID-19 ('positivity rate'); adjusted ORs for factors associated with COVID-19 viral and antibody test positivity; and per cent concordance between positive viral and subsequent antibody test results., Results: Overall positivity rate was 9% (70 472 of 771 278) and 12% (11 094 of 91 741) for viral and antibody tests, respectively. Positivity rate was inversely associated with the number of individuals tested and decreased over time across regions and race/ethnicities. Antibody test concordance among patients with an initial positive viral test was 91% (71%-95% depending on time between tests). Among tests separated by at least 2 weeks, discordant results occurred in 7% of patients and 9% of immunocompromised patients. Factors associated with increased odds of viral and antibody positivity in multivariable models included: male sex, Hispanic or non-Hispanic black or Asian race/ethnicity, uninsured or Medicaid insurance and Northeast residence. We identified a negative dose effect between the number of comorbidities and viral and antibody test positivity. Paediatric patients had reduced odds (OR=0.60, 95% CI 0.57 to 0.64) of a positive viral test but increased odds (OR=1.90, 95% CI 1.62 to 2.23) of a positive antibody test compared with those aged 18-34 years old., Conclusions: This study identified sociodemographic and clinical factors associated with COVID-19 test positivity and provided real-world evidence demonstrating high antibody test concordance among viral-positive patients., Competing Interests: Competing interests: LL, MHS, SR, DSK and LT are employed by and hold shares in Genentech. FY is employed by and holds shares in Roche Diagnostics., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

The COVID-19 pandemic in the United States created a unique situation where multiple molecular SARS-CoV-2 diagnostic assays rapidly received Emergency Use Authorization by the FDA and were validated by laboratories and utilized clinically, all within a period of a few weeks. We compared the performance of four of these assays that were evaluated for use at our institution: Abbott RealTime m2000 SARS-CoV-2 Assay, DiaSorin Simplexa COVID-19 Direct, Cepheid Xpert Xpress SARS-CoV-2, and Abbott ID NOW COVID-19. Nasopharyngeal and nasal specimens were collected from 88 ED and hospital-admitted patients and tested by the four methods in parallel to compare performance. ID NOW performance stood out as significantly worse than the other 3 assays despite demonstrating comparable analytic sensitivity. Further study determined that the use of a nasal swab compared to a nylon flocked nasopharyngeal swab, as well as use in a population chronically vs. acutely positive for SARS-CoV-2, were substantial factors. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in December 2019 and has quickly become a worldwide pandemic. In response, many diagnostic manufacturers have developed molecular assays for SARS-CoV-2 under the Food and Drug Administration (FDA) Emergency Use Authorization (EUA) pathway. This study compared three of these assays, the Hologic Panther Fusion SARS-CoV-2 assay (Fusion), the Hologic Aptima SARS-CoV-2 assay (Aptima), and the BioFire Defense COVID-19 test (BioFire), to determine analytical and clinical performance as well as workflow. All three assays showed similar limits of detection (LODs) using inactivated virus, with 100% detection, ranging from 500 to 1,000 genome equivalents/ml, whereas use of a quantified RNA transcript standard showed the same trend but had values ranging from 62.5 to 125 copies/ml, confirming variability in absolute quantification of reference standards. The clinical correlation found that the Fusion and BioFire assays had a positive percent agreement (PPA) of 98.7%, followed by the Aptima assay at 94.7%, compared to the consensus result. All three assays exhibited 100% negative percent agreement (NPA). Analysis of discordant results revealed that all four samples missed by the Aptima assay had cycle threshold ( C t ) values of >37 by the Fusion assay. In conclusion, while all three assays showed similar relative LODs, we showed differences in absolute LODs depending on which standard was employed. In addition, the Fusion and BioFire assays showed better clinical performance, while the Aptima assay showed a modest decrease in overall PPA. These findings should be kept in mind when making platform testing decisions., (Copyright © 2020 American Society for Microbiology.)

Coronavirus disease 2019 (COVID-19), caused by novel enveloped single stranded RNA coronavirus (SARS-CoV-2), is responsible for an ongoing global pandemic. While other countries deployed widespread testing as an early mitigation strategy, the U.S. experienced delays in development and deployment of organism identification assays. As such, there is uncertainty surrounding disease burden and community spread, severely hampering containment efforts. COVID-19 illuminates the need for a tiered diagnostic approach to rapidly identify clinically significant infections and reduce disease spread. Without the ability to efficiently screen patients, hospitals are overwhelmed, potentially delaying treatment for other emergencies. A multi-tiered, diagnostic strategy incorporating a rapid host immune response assay as a screening test, molecular confirmatory testing and rapid IgM/IgG testing to assess benefit from quarantine/further testing and provide information on population exposure/herd immunity would efficiently evaluate potential COVID-19 patients. Triaging patients within minutes with a fingerstick rather than hours/days after an invasive swab is critical to pandemic response as reliance on the existing strategy is limited by assay accuracy, time to results, and testing capacity. Early screening and triage is achievable from the outset of a pandemic with point-of-care host immune response testing which will improve response time to clinical and public health actions.Key messagesDelayed testing deployment has led to uncertainty surrounding overall disease burden and community spread, severely hampering public health containment and healthcare system preparation efforts.A multi-tiered testing strategy incorporating rapid, host immune point-of-care tests can be used now and for future pandemic planning by effectively identifying patients at risk of disease thereby facilitating quarantine earlier in the progression of the outbreak during the weeks and months it can take for pathogen specific confirmatory tests to be developed, validated and manufactured in sufficient quantities.The ability to triage patients at the point of care and support the guidance of medical and therapeutic decisions, for viral isolation or confirmatory testing or for appropriate treatment of COVID-19 and/or bacterial infections, is a critical component to our national pandemic response and there is an urgent need to implement the proposed strategy to combat the current outbreak.

In pediatric practice it is common for infants under 2 months of age to undergo evaluation for sepsis when they are ill, often including lumbar puncture to assess for central nervous system (CNS) infection. The FilmArray Meningitis/Encephalitis (ME) panel is a newly approved test for rapid identification of CNS pathogens. Our objective was to study the epidemiology of CNS infection in young infants and the potential impact of rapid multiplex PCR on their care. A performance evaluation of the FilmArray ME panel was conducted from February 2014 to September 2014 at 11 sites. FilmArray ME panel results were compared to reference standards but not shared with providers. In our study, medical records for infants (aged 1 to 60 days) enrolled at three sites were reviewed for clinical, laboratory, and outcome data. A total of 145 infants were reviewed. The median age was 25 days. Most of the infants were hospitalized (134/145 [92%]) and received antibiotics (123/145 [85%]), and almost half (71/145 [49%]) received acyclovir. One infant had a bacterial pathogen, likely false positive, identified by the FilmArray ME panel. Thirty-six infants (25%) had a viral pathogen detected, including 21 enteroviruses. All infants with enteroviral meningitis detected by the FilmArray ME panel and conventional PCR were hospitalized, but 20% were discharged in less than 24 h when conventional PCR results became available. The FilmArray ME panel may play a role in the evaluation of young infants for CNS infection. Results may be used to guide management, possibly resulting in a decreased length of stay and less antimicrobial exposure for infants with low-risk viral infection detected., (Copyright © 2018 Blaschke et al.)

Cattle lymph nodes can harbor Salmonella and potentially contaminate beef products. We have developed and validated a new real-time PCR (qPCR) assay for the detection and quantification of Salmonella enterica in cattle lymph nodes. The assay targets both the invA and pagC genes, the most conserved molecular targets in Salmonella enterica. An 18S rRNA gene assay that amplifies from cattle and other animal species was also included as an internal control. Available DNA sequences for invA, pagC and 18S rRNA genes were used for primer and probe selections. Three Salmonella serotypes, S. Typhimurium, S. Anatum, and S. Montevideo, were used to assess the assay's analytical sensitivity. Correlation coefficients of standard curves generated for each target and for all three serotypes were >99% and qPCR amplification efficiencies were between 93% and 110%. Assay sensitivity was also determined using standard curve data generated from Salmonella-negative cattle lymph nodes spiked with 10-fold dilutions of the three Salmonella serotypes. Assay specificity was determined using Salmonella culture method, and qPCR testing on 36 Salmonella strains representing 33 serotypes, 38 Salmonella strains of unknown serotypes, 252 E. coli strains representing 40 serogroups, and 31 other bacterial strains representing 18 different species. A collection of 647 cattle lymph node samples from steers procured from the Midwest region of the US were tested by the qPCR, and compared to culture-method of detection. Salmonella prevalence by qPCR for pre-enriched and enriched lymph nodes was 19.8% (128/647) and 94.9% (614/647), respectively. A majority of qPCR positive pre-enriched samples (105/128) were at concentrations between 10 4 and 10 5  CFU/mL. Culture method detected Salmonella in 7.7% (50/647) and 80.7% (522/647) of pre- and post-enriched samples, respectively; 96.0% (48/50) of pre-enriched and 99.4% (519/522) of post-enriched culture-positive samples were also positive by qPCR. More samples tested positive by qPCR than by culture method, indicating that the real-time PCR assay was more sensitive. Our data indicate that this triplex qPCR can be used to accurately detect and quantify Salmonella enterica strains from cattle lymph node samples. The assay may serve as a useful tool to monitor the prevalence of Salmonella in beef production systems., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: Health care-associated infections caused by antibiotic-resistant gram-negative bacteria, such as Pseudomonas aeruginosa, are an emerging and increasingly important public health threat. However, there are very few studies that examine the relationships between antimicrobial resistance strategies and interventions and the prevalence of antibiotic-resistant and multidrug-resistant strains of Pseudomonas., Methods: The membership of the Association for Professionals in Infection Control and Epidemiology was invited to participate in a Web-based survey of U.S. acute care hospital infection preventionists., Results: A final sample of 225 completed responses was analyzed using correlational techniques. Resistance to gentamicin (r = 0.17, P = .01), imipenem (r = 0.18, P = .02), ceftazidime (r = 0.20, P = .03), and ciprofloxacin (r = 0.15, P = .03) all showed significant direct associations with epidemiologic investigation using molecular testing. A significant inverse relationship was found between resistance to ceftazidime and effective efforts in the identification and isolation of infected patients (r = -0.18, P = .02)., Conclusions: The finding of significant direct relationships between antimicrobial resistance and epidemiologic investigations using molecular testing suggests that the increased burden of resistance is associated with molecular testing rather than the intervention driving down rates. Effective identification and isolation of infected patients appeared to be associated with lower resistance rates. Further research is needed to uncover causal relationships., (Copyright © 2016 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.)

Enteric pathogens that cause gastroenteritis remain a major global health concern. The goal of this study was to develop a multiplex PCR/ligation detection reaction (LDR) assay for the detection of all NIAID category B bacterial food and water-borne pathogens directly from stool specimens. To validate the PCR/LDR assay, clinical isolates of Campylobacter spp., Vibrio spp., Shigella spp., Salmonella spp., Listeria monocytogenes, Yersinia enterocolitica, and diarrheagenic Escherichia coli were tested. The sensitivity and specificity of the assay were assessed using a large number of seeded culture-negative stool specimens and a smaller set of clinical specimens from Haiti. The overall sensitivity ranged from 91% to 100% (median 100%) depending on the species. For the majority of organisms, the sensitivity was 100%. The overall specificity based on initial testing ranged from 98% to 100% depending on the species. After additional testing of discordant samples, the lowest specificity was 99.4%. PCR/LDR detected additional category B agents (particularly diarrheagenic E. coli) in 11/40 specimens from Haiti that were culture-positive for V. cholerae and in approximately 1% of routine culture-negative stool specimens from a hospital in New York. This study demonstrated the ability of the PCR/LDR assay to detect a large comprehensive panel of category B enteric bacterial pathogens as well as mixed infections. This type of assay has the potential to provide earlier warnings of possible public health threats and more accurate surveillance of food and water-borne pathogens., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Recent advances in the understanding of the molecular basis of cancer and the development of molecular diagnostics based on this knowledge have done much to progress the fields of oncology and pathology. Technological developments such as Next Generation Sequencing (NGS) and multiplex assays have made feasible the widespread adoption of molecular diagnostics for clinical use. While these developments and advances carry much promise, there are pitfalls to implementing this testing. Choosing appropriate biomarkers is a vital first step for clinical use and being able to understand the complex relationship between predictive and prognostic biomarkers is a crucial component of this. Testing for standard of care biomarkers is not straightforward, one must choose carefully between clinical trial assays, assays that analyse the same biological phenomenon or surrogate biomarkers. Sample heterogeneity and population specific difference is assays may skew results and must be controlled for at the assay design stage. At a technical level, NGS has the potential to revolutionise laboratory practice and approaches to cancer treatment. However, use of this technology requires careful planning and implementation if one is to avoid technical and ethical quagmires. Finally, with FDA regulation of companion diagnostics one may be limited to therapy specific assays., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)