Deciphering microbiota of acute upper respiratory infections: a comparative analysis of PCR and mNGS methods for lower respiratory trafficking potential DOI Creative Commons
Sadia Almas, Rob E. Carpenter, Anuradha Singh

и другие.

medRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Янв. 5, 2023

Abstract Although it is clinically important for acute respiratory tract (co)infections to have rapid and accurate diagnosis, critical that medicine understand the advantages of current laboratory methods. In this study we tested nasopharyngeal samples (n=29) with a commercially available PCR assay compared results hybridization capture-based mNGS workflow. Detection criteria positive was Ct<35 >40% target coverage, median depth 1X RPKM>10. A high degree concordance (98.33% PPA 100% NPA) recorded. However, yielded 29 additional microorganisms (23 bacteria, 4 viruses, 2 fungi) beyond PCR. We then characterize each method into three phenotypic categories using IDbyDNA Explify® Platform consideration infectivity trafficking potential lower region. The findings are significant providing comprehensive yet relevant microbiology profile upper infection, especially in immunocompetent cases or where traditional syndromic approaches fail identify pathogenicity. Accordingly, technology can be used supplement syndromic-based tests data quickly effectively phenotypically characterized potential, clinical (co)infection, comorbid consideration—with promise reduce morbidity mortality. Highlights What main findings? there between methodologies, workflow able detect versus Identified were rapidly groups potential. implication finding? provide infection Deciphering microbiota grouping has tool better manage complex cases.

Язык: Английский

Navigating the New Norm: The FDA's Final Rule on Laboratory Developed Tests (LDTs) and Its Impact on Clinical Laboratory Operations DOI Creative Commons
Rob E. Carpenter

Clinical Microbiology Newsletter, Год журнала: 2024, Номер unknown

Опубликована: Сен. 1, 2024

Язык: Английский

Процитировано

3
Deciphering Microbiota of Acute Upper Respiratory Infections: A Comparative Analysis of PCR and mNGS Methods for Lower Respiratory Trafficking Potential DOI Creative Commons
Sadia Almas, Rob E. Carpenter, Anuradha Singh

и другие.

Advances in respiratory medicine, Год журнала: 2023, Номер 91(1), С. 49 - 65

Опубликована: Фев. 2, 2023

Although it is clinically important for acute respiratory tract (co)infections to have a rapid and accurate diagnosis, critical that medicine understands the advantages of current laboratory methods. In this study, we tested nasopharyngeal samples (n = 29) with commercially available PCR assay compared results those hybridization-capture-based mNGS workflow. Detection criteria positive was Ct < 35 >40% target coverage, median depth 1X RPKM > 10. A high degree concordance (98.33% PPA 100% NPA) recorded. However, yielded positively 29 additional microorganisms (23 bacteria, 4 viruses, 2 fungi) beyond PCR. We then characterized each method into three phenotypic categories using IDbyDNA Explify® Platform (Illumina® Inc, San Diego, CA, USA) consideration infectivity trafficking potential lower region. The findings are significant providing comprehensive yet relevant microbiology profile upper infection, especially in immunocompromised or immunocompetent comorbidity cases where traditional syndromic approaches fail identify pathogenicity. Accordingly, technology can be used supplement syndrome-based tests, data quickly effectively phenotypically potential, clinical (co)infection, comorbid consideration-with promise reduce morbidity mortality.

Язык: Английский

Процитировано

7
Evaluation of next generation sequencing approaches for SARS-CoV-2 DOI Creative Commons
Valentina Curini, Massimo Ancora, Lucija Jurišić

и другие.

Heliyon, Год журнала: 2023, Номер 9(11), С. e21101 - e21101

Опубликована: Окт. 19, 2023

Within public health control strategies for SARS-CoV-2, whole genome sequencing (WGS) is essential tracking viral spread and monitoring the emergence of variants which may impair effectiveness vaccines, diagnostic methods, therapeutics. In this manuscript different SARS-CoV-2 WGS including metagenomic shotgun (SG), library enrichment by myBaits® Expert Virus-SARS-CoV-2 (Arbor Biosciences), nCoV-2019 protocol, ampliseq approach Swift Amplicon® Panel kit (Swift Illumina COVIDSeq Test (Illumina Inc.), were evaluated in order to identify best terms results, labour, costs. The analysis revealed that Inc.) choice a cost-effective, time-consuming production consensus sequences.

Язык: Английский

Процитировано

7
SARS-CoV-2 Next Generation Sequencing (NGS) data from clinical isolates from the East Texas Region of the United States DOI Creative Commons
Rob E. Carpenter, Vaibhav K. Tamrakar, Sadia Almas

и другие.

Data in Brief, Год журнала: 2023, Номер 49, С. 109312 - 109312

Опубликована: Июнь 16, 2023

The SARS-CoV-2 virus has evolved throughout the pandemic and is likely to continue evolving into new variants. Some of these variants may affect functional properties, including infectivity, interactions with host immunity, disease severity. And compromised vaccine efficacy an emerging concern every viral variant. Next-generation sequencing (NGS) emerged as tool choice for discovering understanding transmission dynamics SARS-CoV-2. Deciphering genome enabled epidemiological survivance forecast altered etiologically. Clinical presentations infection are influenced by comorbidities such age, immune status, diabetes, infecting Thus, clinical management differ For example, some monoclonal antibody treatments variant-specific, vaccines less efficacious against omicron delta Consequently, determining local outbreaks monitoring Variants Concern (VOC) one primary strategies pandemic's containment. Although next-generation a gold standard genomic surveillance variant discovery, assays not approved diagnosis decision-making. Advanta Genetics, Texas, USA, optimized Illumina COVID-seq protocol reduce cost without compromising accuracy validated COVID-Seq assay Laboratory Developed Test (LDT) according guidelines prescribed College American Pathologists (CAP) Improvement Amendments (CLIA). whole was sequenced in (n = 161) samples from East Texas region using MiniSeq® instrument analyzed baseSpace (https://basespace.illumina.com) bioinformatics pipeline. Briefly, library prepared COVIDSeq research use only (RUO) kit, individual libraries were normalized DNA concentration measured Qubit Flex Fluorometer, pooled on Instrument. application used QC, FASTQ generation, assembly, identification This shotgun project been deposited at GISAID.

Язык: Английский

Процитировано

6
Optimization of the Illumina COVIDSeq™ protocol for decentralized, cost-effective genomic surveillance DOI Creative Commons
Rob E. Carpenter, Vaibhav K. Tamrakar, Sadia Almas

и другие.

Practical Laboratory Medicine, Год журнала: 2023, Номер 34, С. e00311 - e00311

Опубликована: Фев. 1, 2023

A decentralized surveillance system to identify local outbreaks and monitor SARS-CoV-2 Variants of Concern is one the primary strategies for pandemic's containment. Although next-generation sequencing (NGS) a gold standard genomic variant discovery, technology still cost-prohibitive sequencing, particularly in small independent labs with limited resources. We have optimized Illumina COVIDSeq™ protocol MiniSeq instrument reduce cost without compromising accuracy. slashed library preparation by half using 50% recommended reagents at each step normalizing libraries before pooling achieve uniform coverage. Reagent-only (∼$43.27/sample) analysis this normalized input on instruments comparable what achieved high throughput such as NextSeq NovaSeq. Using modified protocol, we tested 153 clinical samples, 90% coverage was 142/153 samples analyzed study. The lineage correctly assigned all (152/153) except one. This can help laboratories constrained resources contribute COVID-19 post-vaccination era.

Язык: Английский

Процитировано

4
Design, Immunogenicity and Preclinical Efficacy of the ChAdOx1.COVconsv12 Pan-Sarbecovirus T-Cell Vaccine DOI Creative Commons

Edmund G. Wee,

Sarah Kempster, David Ferguson

и другие.

Vaccines, Год журнала: 2024, Номер 12(9), С. 965 - 965

Опубликована: Авг. 26, 2024

During the COVID-19 pandemic, antibody-based vaccines targeting SARS-CoV-2 spike glycoprotein were focus for development because neutralizing antibodies associated with protection against infection pre-clinically and in humans. While deploying these spike-based saved millions of lives worldwide, it has become clear that immunological mechanisms severe disease are multifaceted involve non-neutralizing antibody components. Here, we describe a novel pan-sarbecovirus T-cell vaccine, ChAdOx1.COVconsv12, designed to complement broaden vaccines. The vaccine immunogen COVconsv12 employs two regions viral proteome most conserved among sarbecoviruses, which delivered by replication-deficient vector ChAdOx1. It directs T cells towards epitopes shared sarbecoviruses including evolving variants. show ChAdOx1.COVconsv12 induced broad responses BALB/c C57BL/6 mice. In Syrian hamster challenge model, alone did not protect infection, but when co-administered 1/50th ChAdOx1 nCoV-19 protective dose, faster recovery lower oral swab load observed. Induction CD8+ may decrease severity extend response coverage variants match known (and as yet unknown) members β-coronavirus family.

Язык: Английский

Процитировано

1
Evaluation of Illumina® COVIDSeq™ as a tool for Omicron SARS-CoV-2 characterisation DOI Creative Commons
Kym Lowry, Michelle J. Bauer,

Cameron Buckley

и другие.

Journal of Virological Methods, Год журнала: 2023, Номер 322, С. 114827 - 114827

Опубликована: Сен. 30, 2023

The continued emergence and transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants requires ongoing genetic surveillance to support public health responses. expansion reliable next generation sequence (NGS) platforms has enabled the rapid characterisation constant new SARS-CoV-2 using nasopharyngeal swab specimens. Several studies have assessed ability COVIDSeq type earlier strains (pre-Delta) rapidly successfully, however, there is limited data showing suitability against Omicron variants. In present study, we evaluated performance Illumina Assay as a streamlined amplicon-based NGS platform for detection typing Our results demonstrate high sequencing approach, with good repeatability, reproducibility sensitivity samples approaching CT 31. approach was 100% concordant previously characterized by methods. quick library preparation process throughput kit made it ideal reflex testing, total time required analysis approximately two days. This study demonstrates effectiveness versatility method SARS-CoV-2, providing foundation further research development custom-designed amplicon panels targeting different microorganisms.

Язык: Английский

Процитировано

3
Deciphering microbiota of acute upper respiratory infections: a comparative analysis of PCR and mNGS methods for lower respiratory trafficking potential DOI Creative Commons
Sadia Almas, Rob E. Carpenter, Anuradha Singh

и другие.

medRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Янв. 5, 2023

Abstract Although it is clinically important for acute respiratory tract (co)infections to have rapid and accurate diagnosis, critical that medicine understand the advantages of current laboratory methods. In this study we tested nasopharyngeal samples (n=29) with a commercially available PCR assay compared results hybridization capture-based mNGS workflow. Detection criteria positive was Ct<35 >40% target coverage, median depth 1X RPKM>10. A high degree concordance (98.33% PPA 100% NPA) recorded. However, yielded 29 additional microorganisms (23 bacteria, 4 viruses, 2 fungi) beyond PCR. We then characterize each method into three phenotypic categories using IDbyDNA Explify® Platform consideration infectivity trafficking potential lower region. The findings are significant providing comprehensive yet relevant microbiology profile upper infection, especially in immunocompetent cases or where traditional syndromic approaches fail identify pathogenicity. Accordingly, technology can be used supplement syndromic-based tests data quickly effectively phenotypically characterized potential, clinical (co)infection, comorbid consideration—with promise reduce morbidity mortality. Highlights What main findings? there between methodologies, workflow able detect versus Identified were rapidly groups potential. implication finding? provide infection Deciphering microbiota grouping has tool better manage complex cases.

Язык: Английский

Процитировано

1