PURPOSE OF THE REVIEWLaboratory-developed tests (LDTs) are essential for the clinical care of immunocompromised individuals. These patients often require specialized testing not available from ...commercial manufacturers and are therefore dependent on the laboratory to create, validate, and perform these assays. Recent paradigm-shifting legislation could alter the way that LDTs are operationalized and regulated.
RECENT FINDINGSOn March 5th, 2020 the Verifying Accurate and Leading-Edge In-Vitro Clinical Tests Development Act (VALID) was introduced in the US Congress. This statute would overhaul existing regulatory framework by unifying the oversight of LDTs and commercial in-vitro diagnostic tests (IVDs) through the FDA. If enacted, LDTs would be subject to regulatory requirements like those found in commercial submissions for market review. Stakeholders continue to discuss the details and scope of the proposed legislation in the setting of the Severe Acute Respiratory Syndrome Coronavirus 2 pandemic, where LDTs are integral to the national COVID-19 response.
SUMMARYCongressional lawmakers have introduced legislation to alter the regulatory framework governing LDTs. Moving forward, a balance must be struck to ensure the availability of safe and accurate testing without delays or overregulation that could be harmful to patients. The downstream implications of how VALID and other legislation will impact laboratories, clinicians, and patients warrant close examination.
Parkinson's disease (PD) is a common movement disorder marked by the loss of dopaminergic (DA) neurons in the brain stem and the presence of intraneuronal inclusions designated as Lewy bodies (LB). ...The cause of neurodegeneration in PD is not clear, but it has been suggested that protein misfolding and aggregation contribute significantly to the development of the disease. Misfolded and aggregated proteins are cleared by ubiquitin proteasomal system (UPS) and autophagy lysosomal pathway (ALP). Recent studies suggested that different types of ubiquitin linkages can modulate these two pathways in the process of protein degradation. In this study, we found that co-expression of ubiquitin can rescue neurons from α-syn-induced neurotoxicity in a Drosophila model of PD. This neuroprotection is dependent on the formation of lysine 48 polyubiquitin linkage which is known to target protein degradation via the proteasome. Consistent with our results that we observed in vivo, we found that ubiquitin co-expression in the cell can facilitate cellular protein degradation by the proteasome in a lysine 48 polyubiquitin-dependent manner. Taken together, these results suggest that facilitation of proteasomal protein degradation can be a potential therapeutic approach for PD.
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged into a world of maturing pathogen genomics, with >2 million genomes sequenced at this writing. The rise of more ...transmissible variants of concern that affect vaccine and therapeutic effectiveness has led to widespread interest in SARS-CoV-2 evolution. Clinicians are also eager to take advantage of the information provided by SARS-CoV-2 genotyping beyond surveillance purposes. Here, we review the potential role of SARS-CoV-2 genotyping in clinical care. The review covers clinical use cases for SARS-CoV-2 genotyping, methods of SARS-CoV-2 genotyping, assay validation and regulatory requirements, clinical reporting for laboratories, and emerging issues in clinical SARS-CoV-2 sequencing. While clinical uses of SARS-CoV-2 genotyping are currently limited, rapid technological change along with a growing ability to interpret variants in real time foretell a growing role for SARS-CoV-2 genotyping in clinical care as continuing data emerge on vaccine and therapeutic efficacy.
This article reviews the potential role of SARS-CoV-2 genotyping in clinical care. Rapid technological change along with a growing ability to interpret variants in real-time foretell a growing role for SARS-CoV-2 genotyping in clinical care.
Abstract
Urinary tract infections (UTIs) are among the most common bacterial infections in the United States and are a major driver of antibiotic use, both appropriate and inappropriate, across ...healthcare settings. Novel UTI diagnostics are a strategy that might enable better UTI treatment. Members of the Antibacterial Resistance Leadership Group Laboratory Center and the Infectious Diseases Society of America Diagnostics Committee convened to envision ideal future UTI diagnostics, with a view towards improving delivery of healthcare, patient outcomes and experiences, and antibiotic use, addressing which types of UTI diagnostics are needed and how companies might approach development of novel UTI diagnostics.
Urinary tract infections are among the most common bacterial infections in the United States and are a major driver of antibiotic use, both appropriate and inappropriate, across healthcare settings.
Abstract
Background
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge, and effective tracking requires rapid return of results. Surveillance of variants is ...typically performed by whole genome sequencing (WGS), which can be financially prohibitive and requires specialized equipment and bioinformatic expertise. Genotyping approaches are rapid methods for monitoring SARS-CoV-2 variants but require continuous adaptation. Fragment analysis may represent an approach for improved SARS-CoV-2 variant detection.
Methods
A multiplex fragment analysis approach (CoVarScan) was validated using PCR targeting variants by size and fluorescent color. Eight SARS-CoV-2 mutational hot spots in variants of concern (VOCs) were targeted. Three primer pairs (recurrently deleted region RDR 1, RDR2, and RDR3–4) flank RDRs in the S-gene. Three allele-specific primers target recurrent spike receptor binding domain mutants. Lastly, 2 primer pairs target recurrent deletions or insertions in ORF1A and ORF8. Fragments were resolved and analyzed by capillary electrophoresis (ABI 3730XL), and mutational signatures were compared to WGS results.
Results
We validated CoVarScan using 3544 clinical respiratory specimens. The assay exhibited 96% sensitivity and 99% specificity compared to WGS. The limit of detection for the core targets (RDR1, RDR2, and ORF1A) was 5 copies/reaction. Variants were identified in 95% of samples with cycle threshold (CT) <30 and 75% of samples with a CT 34 to 35. Assay design was frozen April 2021, but all subsequent VOCs have been detected including Delta (n = 2820), Mu, (n = 6), Lambda (n = 6), and Omicron (n = 309). Genotyping results are available in as little as 4 h.
Conclusions
Multiplex fragment analysis is adaptable and rapid and has similar accuracy to WGS to classify SARS-CoV-2 variants.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged into a world of maturing pathogen genomics, with more than 2 million genomes sequenced at the time of writing. The rise of more ...transmissible variants of concern that impact vaccine and therapeutic effectiveness has led to widespread interest in SARS-CoV-2 evolution. Clinicians are also eager to take advantage of the information provided by SARS-CoV-2 genotyping beyond surveillance purposes. Here, we review the potential role of SARS-CoV-2 genotyping in clinical care. The review covers clinical use cases for SARS-CoV-2 genotyping, methods of SARS-CoV-2 genotyping, assay validation and regulatory requirements, and clinical reporting for laboratories, as well as emerging issues in clinical SARS-CoV-2 sequencing. While clinical uses of SARS-CoV-2 genotyping are currently limited, rapid technological change along with a growing ability to interpret variants in real time foretells a growing role for SARS-CoV-2 genotyping in clinical care as continuing data emerge on vaccine and therapeutic efficacy.
The persisting Coronavirus disease 2019 (COVID-19) pandemic and limited vaccine supply has led to a shift in global health priorities to expand vaccine coverage. Relying on severe acute respiratory ...syndrome coronavirus 2 (SARS-CoV-2) molecular testing alone cannot reveal the infection proportion, which could play a critical role in vaccination prioritization. We evaluated the utility of a combination orthogonal serological testing (COST) algorithm alongside RT-PCR to quantify prevalence with the aim of identifying candidate patient clusters to receive single and/or delayed vaccination.
We utilized 108,505 patients with suspected COVID-19 in a retrospective analysis of SARS-CoV-2 RT-PCR vs. IgG-nucleocapsid (IgG
) antibody testing coverage in routine practice for the estimation of prevalence. Prospectively, an independent cohort of 21,388 subjects was simultaneously tested by SARS-CoV-2 RT-PCR and IgG
to determine the prevalence. We used 614 prospective study subjects to assess the utility of COST (IgG
, IgM-spike (IgM
), and IgG-spike (IgG
)) in establishing the infection proportion to identify a single-dose vaccination cohort.
Retrospectively, we observed a 6.3% (6871/108,505) positivity for SARS-CoV-2 RT-PCR, and only 2.3% (2533/108,505) of cases had paired IgG
serology performed. Prospectively, IgG
serology identified twice the number of COVID-positive cases in relation to RT-PCR alone. COST further increased the number of detected positive cases: IgG
+ or IgM
+ (18.0%); IgG
+ or IgG
+ (23.5%); IgM
+ or IgG
+ (23.8%); and IgG
+ or IgM
+ or IgG
+ (141/584 = 24.1%).
COST may be an effective tool for the evaluation of infection proportion and thus could define a cohort for a single dose and/or delayed vaccination.
Diagnostics and laboratory role in outbreaks Lee, Francesca
Current opinion in infectious diseases,
2017-August, 2017-Aug, 2017-08-00, 20170801, Letnik:
30, Številka:
4
Journal Article
Recenzirano
PURPOSE OF REVIEWThe review describes the investigative benefits of traditional and novel molecular epidemiology techniques, while acknowledging the limitations faced by clinical laboratories seeking ...to implement these methods.
RECENT FINDINGSPulse-field gel electrophoresis and other traditional techniques remain powerful tools in outbreak investigations and continue to be used by multiple groups. Newer techniques such as matrix-assisted laser desorption/ionization-time of flight mass-spectrometry and whole genome sequencing show great promise. However, there is a lack of standardization regarding definitions for genetic relatedness, nor are there established criteria for accuracy and reproducibility. There are also challenges regarding availability of trained bioinformatics staff, and concerns regarding reimbursement.
SUMMARYThere are many tools available for molecular epidemiologic investigation. Epidemiologists and clinical laboratorians should work together to determine which testing methods are best for each institution.
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