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Article ; Online: PoMeLo: a systematic computational approach to predicting metabolic loss in pathogen genomes.

Glascock, Abigail / Waltari, Eric / Dudas, Gytis / Wong, Joan / Ahyong, Vida

2024 Volume 25, Issue 1, Page(s) 49

Abstract: Background: Genome streamlining, the process by which genomes become smaller and encode fewer genes over time, is a common phenomenon among pathogenic bacteria. This reduction is driven by selection for minimized energy expenditure in a nutrient-rich ... ...

Abstract	Background: Genome streamlining, the process by which genomes become smaller and encode fewer genes over time, is a common phenomenon among pathogenic bacteria. This reduction is driven by selection for minimized energy expenditure in a nutrient-rich environment. As pathogens evolve to become more reliant on the host, metabolic genes and resulting capabilities are lost in favor of siphoning metabolites from the host. Characterizing genome streamlining, gene loss, and metabolic pathway degradation can be useful in assessing pathogen dependency on host metabolism and identifying potential targets for host-directed therapeutics. Results: PoMeLo (Predictor of Metabolic Loss) is a novel evolutionary genomics-guided computational approach for identifying metabolic gaps in the genomes of pathogenic bacteria. PoMeLo leverages a centralized public database of high-quality genomes and annotations and allows the user to compare an unlimited number of genomes across individual genes and pathways. PoMeLo runs locally using user-friendly prompts in a matter of minutes and generates tabular and visual outputs for users to compare predicted metabolic capacity between groups of bacteria and individual species. Each pathway is assigned a Predicted Metabolic Loss (PML) score to assess the magnitude of genome streamlining. Optionally, PoMeLo places the results in an evolutionary context by including phylogenetic relationships in visual outputs. It can also initially compute phylogenetically-weighted mean genome sizes to identify genome streamlining events. Here, we describe PoMeLo and demonstrate its use in identifying metabolic gaps in genomes of pathogenic Treponema species. Conclusions: PoMeLo represents an advance over existing methods for identifying metabolic gaps in genomic data, allowing comparison across large numbers of genomes and placing the resulting data in a phylogenetic context. PoMeLo is freely available for academic and non-academic use at https://github.com/czbiohub-sf/pomelo .
MeSH term(s)	Phylogeny ; Genome ; Genomics/methods ; Biological Evolution ; Bacteria/genetics ; Software
Language	English
Publishing date	2024-01-30
Publishing country	England
Document type	Journal Article
ZDB-ID	2041484-5
ISSN	1471-2105 ; 1471-2105
ISSN (online)	1471-2105
ISSN	1471-2105
DOI	10.1186/s12859-024-05640-w
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Article ; Online: The antibiotic resistance reservoir of the lung microbiome expands with age in a population of critically ill patients.

Chu, Victoria T / Tsitsiklis, Alexandra / Mick, Eran / Ambroggio, Lilliam / Kalantar, Katrina L / Glascock, Abigail / Osborne, Christina M / Wagner, Brandie D / Matthay, Michael A / DeRisi, Joseph L / Calfee, Carolyn S / Mourani, Peter M / Langelier, Charles R

Nature communications

2024 Volume 15, Issue 1, Page(s) 92

Abstract: Antimicrobial resistant lower respiratory tract infections are an increasing public health threat and an important cause of global mortality. The lung microbiome can influence susceptibility of respiratory tract infections and represents an important ... ...

Abstract	Antimicrobial resistant lower respiratory tract infections are an increasing public health threat and an important cause of global mortality. The lung microbiome can influence susceptibility of respiratory tract infections and represents an important reservoir for exchange of antimicrobial resistance genes. Studies of the gut microbiome have found an association between age and increasing antimicrobial resistance gene burden, however, corollary studies in the lung microbiome remain absent. We performed an observational study of children and adults with acute respiratory failure admitted to the intensive care unit. From tracheal aspirate RNA sequencing data, we evaluated age-related differences in detectable antimicrobial resistance gene expression in the lung microbiome. Using a multivariable logistic regression model, we find that detection of antimicrobial resistance gene expression was significantly higher in adults compared with children after adjusting for demographic and clinical characteristics. This association remained significant after additionally adjusting for lung bacterial microbiome characteristics, and when modeling age as a continuous variable. The proportion of adults expressing beta-lactam, aminoglycoside, and tetracycline antimicrobial resistance genes was higher compared to children. Together, these findings shape our understanding of the lung resistome in critically ill patients across the lifespan, which may have implications for clinical management and global public health.
MeSH term(s)	Adult ; Child ; Humans ; Critical Illness ; Anti-Bacterial Agents/pharmacology ; Anti-Bacterial Agents/therapeutic use ; Microbiota/genetics ; Lung ; Drug Resistance, Microbial/genetics ; Respiratory Tract Infections/drug therapy
Chemical Substances	Anti-Bacterial Agents
Language	English
Publishing date	2024-01-02
Publishing country	England
Document type	Observational Study ; Journal Article
ZDB-ID	2553671-0
ISSN	2041-1723 ; 2041-1723
ISSN (online)	2041-1723
ISSN	2041-1723
DOI	10.1038/s41467-023-44353-1
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Article: Doxycycline post-exposure prophylaxis for sexually transmitted infections impacts the gut antimicrobial resistome.

Langelier, Charles / Chu, Victoria / Glascock, Abigail / Donnell, Deborah / Grabow, Cole / Brown, Clare / Ward, Ryan / Love, Christina / Kalantar, Katrina / Cohen, Stephanie / Cannon, Chase / Woodworth, Michael / Kelley, Colleen / Celum, Connie / Luetkemeyer, Anne

Research square

2024

Abstract: Doxycycline post-exposure prophylaxis (doxy-PEP) reduces bacterial sexually transmitted infections (STIs) among men who have sex with men and transgender women. While poised for widespread clinical implementation, the impact of doxy-PEP on antimicrobial ... ...

Abstract	Doxycycline post-exposure prophylaxis (doxy-PEP) reduces bacterial sexually transmitted infections (STIs) among men who have sex with men and transgender women. While poised for widespread clinical implementation, the impact of doxy-PEP on antimicrobial resistance remains a primary concern as its effects on the gut microbiome and resistome, or the antimicrobial resistance genes (ARGs) present in the gut microbiome, are unknown. To investigate these effects, we studied participants from a randomized clinical trial who either received doxy-PEP as a one-time doxycycline 200 mg taken after condomless sex (DP arm, n = 100) or standard of care treatment (SOC arm, n = 50). From self-collected rectal swabs at enrollment (day-0) and after 6 months (month-6), we performed metagenomic DNA sequencing (DNA-seq) or metatranscriptomic RNA sequencing (RNA-seq). DNA-seq data was analyzable from 127 samples derived from 89 participants, and RNA-seq data from 86 samples derived from 70 participants. We compared the bacterial microbiome and resistome between the two study arms and over time. Tetracycline ARGs were detected in all day-0 DNA-seq samples and 85% of day-0 RNA-seq samples. The proportional mass of tetracycline ARGs in the resistome increased between day-0 and month-6 in DP participants from 46-51% in the metagenome (p = 0.02) and 4-15% in the metatranscriptome (p < 0.01), but no changes in other ARG classes were observed. Exposure to a higher number of doxycycline doses correlated with proportional enrichment of tetracycline ARGs in the metagenome (Spearman's ρ = 0.23, p < 0.01) and metatranscriptome (Spearman's ρ = 0.55, p < 0.01). Bacterial microbiome alpha diversity, beta diversity, and total bacterial mass did not differ between day-0 and month-6 samples from DP participants when assessed by either DNA-seq or RNA-seq. In an abundance-based correlation analysis, we observed an increase over time in the strength of the correlation between tetracycline ARGs and specific bacterial taxa, including some common human pathogens. In sum, doxy-PEP use over a 6-month period was associated with an increase in the proportion of tetracycline ARGs comprising the gut resistome, and an increase in the expression of tetracycline ARGs. Notably, doxy-PEP did not significantly alter alpha diversity or taxonomic composition of the gut microbiome, and did not demonstrate significant increases in non-tetracycline ARG classes. Further studies and population level surveillance are needed to understand the implications of these findings as doxy-PEP is implemented as a public health strategy.
Language	English
Publishing date	2024-04-17
Publishing country	United States
Document type	Preprint
DOI	10.21203/rs.3.rs-4243341/v1
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Article: The antibiotic resistance reservoir of the lung microbiome expands with age.

Research square

2023

Abstract: Antimicrobial resistant lower respiratory tract infections (LRTI) are an increasing public health threat, and an important cause of global mortality. The lung microbiome influences LRTI susceptibility and represents an important reservoir for exchange of ...

Abstract	Antimicrobial resistant lower respiratory tract infections (LRTI) are an increasing public health threat, and an important cause of global mortality. The lung microbiome influences LRTI susceptibility and represents an important reservoir for exchange of antimicrobial resistance genes (ARGs). Studies of the gut microbiome have found an association between age and increasing antimicrobial resistance gene (ARG) burden, however corollary studies in the lung microbiome remain absent, despite the respiratory tract representing one of the most clinically significant sites for drug resistant infections. We performed a prospective, multicenter observational study of 261 children and 88 adults with acute respiratory failure, ranging in age from 31 days to ≥ 89 years, admitted to intensive care units in the United States. We performed RNA sequencing on tracheal aspirates collected within 72 hours of intubation, and evaluated age-related differences in detectable ARG expression in the lung microbiome as a primary outcome. Secondary outcomes included number and classes of ARGs detected, proportion of patients with an ARG class, and composition of the lung microbiome. Multivariable logistic regression models (adults vs children) or continuous age (years) were adjusted for sex, race/ethnicity, LRTI status, and days from intubation to specimen collection. Detection of ARGs was significantly higher in adults compared with children after adjusting for sex, race/ethnicity, LRTI diagnosis, and days from intubation to specimen collection (adjusted odds ratio (aOR): 2.16, 95% confidence interval (CI): 1.10-4.22). A greater proportion of adults compared with children had beta-lactam ARGs (31% (CI: 21-41%) vs 13% (CI: 10-18%)), aminoglycoside ARGs (20% (CI: 13-30%) vs 2% (CI: 0.6-4%)), and tetracycline ARGs (14% (CI: 7-23%) vs 3% (CI: 1-5%)). Adults ≥70 years old had the highest proportion of these three ARG classes. The total bacterial abundance of the lung microbiome increased with age, and microbiome alpha diversity varied with age. Taxonomic composition of the lung microbiome, measured by Bray Curtis dissimilarity index, differed between adults and children (p = 0.003). The association between age and increased ARG detection remained significant after additionally including lung microbiome total bacterial abundance and alpha diversity in the multivariable logistic regression model (aOR: 2.38, (CI: 1.25-4.54)). Furthermore, this association remained robust when modeling age as a continuous variable (aOR: 1.02, (CI: 1.01-1.03) per year of age). Taken together, our results demonstrate that age is an independent risk factor for ARG detection in the lower respiratory tract microbiome. These data shape our understanding of the lung resistome in critically ill patients across the lifespan, which may have implications for clinical management and global public health.
Language	English
Publishing date	2023-09-18
Publishing country	United States
Document type	Preprint
DOI	10.21203/rs.3.rs-3283415/v1
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Article: Simultaneous detection of pathogens and antimicrobial resistance genes with the open source, cloud-based, CZ ID pipeline.

Lu, Dan / Kalantar, Katrina L / Chu, Victoria T / Glascock, Abigail L / Guerrero, Estella S / Bernick, Nina / Butcher, Xochitl / Ewing, Kirsty / Fahsbender, Elizabeth / Holmes, Olivia / Hoops, Erin / Jones, Ann E / Lim, Ryan / McCanny, Suzette / Reynoso, Lucia / Rosario, Karyna / Tang, Jennifer / Valenzuela, Omar / Mourani, Peter M /

Pickering, Amy J / Raphenya, Amogelang R / Alcock, Brian P / McArthur, Andrew G / Langelier, Charles R

bioRxiv : the preprint server for biology

2024

Abstract: Antimicrobial resistant (AMR) pathogens represent urgent threats to human health, and their surveillance is of paramount importance. Metagenomic next generation sequencing (mNGS) has revolutionized such efforts, but remains challenging due to the lack of ...

Abstract	Antimicrobial resistant (AMR) pathogens represent urgent threats to human health, and their surveillance is of paramount importance. Metagenomic next generation sequencing (mNGS) has revolutionized such efforts, but remains challenging due to the lack of open-access bioinformatics tools capable of simultaneously analyzing both microbial and AMR gene sequences. To address this need, we developed the Chan Zuckerberg ID (CZ ID) AMR module, an open-access, cloud-based workflow designed to integrate detection of both microbes and AMR genes in mNGS and whole-genome sequencing (WGS) data. It leverages the Comprehensive Antibiotic Resistance Database and associated Resistance Gene Identifier software, and works synergistically with the CZ ID short-read mNGS module to enable broad detection of both microbes and AMR genes. We highlight diverse applications of the AMR module through analysis of both publicly available and newly generated mNGS and WGS data from four clinical cohort studies and an environmental surveillance project. Through genomic investigations of bacterial sepsis and pneumonia cases, hospital outbreaks, and wastewater surveillance data, we gain a deeper understanding of infectious agents and their resistomes, highlighting the value of integrating microbial identification and AMR profiling for both research and public health. We leverage additional functionalities of the CZ ID mNGS platform to couple resistome profiling with the assessment of phylogenetic relationships between nosocomial pathogens, and further demonstrate the potential to capture the longitudinal dynamics of pathogen and AMR genes in hospital acquired bacterial infections. In sum, the new AMR module advances the capabilities of the open-access CZ ID microbial bioinformatics platform by integrating pathogen detection and AMR profiling from mNGS and WGS data. Its development represents a critical step toward democratizing pathogen genomic analysis and supporting collaborative efforts to combat the growing threat of AMR.
Language	English
Publishing date	2024-04-18
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.04.12.589250
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Article: Building up a genomic surveillance platform for SARS-CoV-2 in the middle of a pandemic: a true North-South collaboration.

Khan, Waqasuddin / Kabir, Furqan / Kanwar, Samiah / Aziz, Fatima / Muneer, Sahrish / Kalam, Adil / Rajab Ali, Mehdia Nadeem / Ansari, Nadia / Vanaerschot, Manu / Ahyong, Vida / Fahsbender, Liz / Kalantar, Katrina / Black, Allison / Glascock, Abigail / Gil, Juliana / Ayscue, Patrick / Tato, Cristina / Jehan, Fyezah / Nisar, Imran

BMJ global health

2023 Volume 8, Issue 11

Abstract: Next-generation sequencing technology has revolutionised pathogen surveillance over the last two decades. However, the benefits are not equitably distributed, with developing countries lagging far behind in acquiring the required technology and ... ...

Abstract	Next-generation sequencing technology has revolutionised pathogen surveillance over the last two decades. However, the benefits are not equitably distributed, with developing countries lagging far behind in acquiring the required technology and analytical capacity. Recent declines in the cost associated with sequencing-equipment and running consumables have created an opportunity for broader adoption. During the COVID-19 pandemic, rapid diagnostics development and DNA sequencing revolutionised the ability to diagnose and sequence SARS-CoV-2 rapidly. Socioeconomic inequalities substantially impact the ability to sequence SARS-CoV-2 strains and undermine a developing country's pandemic preparedness. Low- and middle-income countries face additional challenges in establishing, maintaining and expanding genomic surveillance. We present our experience of establishing a genomic surveillance system at the Aga Khan University, Karachi, Pakistan. Despite being at a leading health sciences research institute in the country, we encountered significant challenges. These were related to collecting standardised contextual data for SARS-CoV-2 samples, procuring sequencing reagents and consumables, and challenges with library preparation, sequencing and submission of high-quality SARS-CoV-2 genomes. Several technical roadblocks ensued during the implementation of the genomic surveillance framework, which were resolved in collaboration with our partners. High-quality genome sequences were then deposited on open-access platforms per the best practices. Subsequently, these efforts culminated in deploying Pakistan's first SARS-CoV-2 phyllo surveillance map as a Nextstrain build. Our experience offers lessons for the successful development of Genomic Surveillance Infrastructure in resource-limited settings struck by a pandemic.
MeSH term(s)	Humans ; SARS-CoV-2/genetics ; Pandemics ; COVID-19 ; Genomics ; Pakistan/epidemiology
Language	English
Publishing date	2023-11-20
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	2059-7908
ISSN	2059-7908
DOI	10.1136/bmjgh-2023-012589
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Article ; Online: Unique roles of vaginal

Glascock, Abigail L / Jimenez, Nicole R / Boundy, Sam / Koparde, Vishal N / Brooks, J Paul / Edwards, David J / Strauss Iii, Jerome F / Jefferson, Kimberly K / Serrano, Myrna G / Buck, Gregory A / Fettweis, Jennifer M

Microbial genomics

2022 Volume 7, Issue 12

Abstract: The composition of the human vaginal microbiome has been extensively studied and is known to influence reproductive health. However, the functional roles of individual taxa and their contributions to negative health outcomes have yet to be well ... ...

Abstract	The composition of the human vaginal microbiome has been extensively studied and is known to influence reproductive health. However, the functional roles of individual taxa and their contributions to negative health outcomes have yet to be well characterized. Here, we examine two vaginal bacterial taxa grouped within the genus
MeSH term(s)	Bacterial Proteins/genetics ; Base Composition ; Case-Control Studies ; Evolution, Molecular ; Female ; Gene Expression Regulation, Bacterial ; Genome Size ; Genome, Bacterial ; High-Throughput Nucleotide Sequencing ; Humans ; Megasphaera/classification ; Megasphaera/genetics ; Megasphaera/isolation & purification ; Megasphaera/metabolism ; Phylogeny ; Pregnancy ; RNA, Ribosomal, 16S/genetics ; Reproductive Health ; Sequence Analysis, DNA/methods ; Vagina/microbiology ; Vaginosis, Bacterial/epidemiology ; Vaginosis, Bacterial/microbiology
Chemical Substances	Bacterial Proteins ; RNA, Ribosomal, 16S
Language	English
Publishing date	2022-01-28
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2835258-0
ISSN	2057-5858 ; 2057-5858
ISSN (online)	2057-5858
ISSN	2057-5858
DOI	10.1099/mgen.0.000526
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Article ; Online: Detection of SARS-CoV-2 RNA using RT-LAMP and molecular beacons.

Sherrill-Mix, Scott / Hwang, Young / Roche, Aoife M / Glascock, Abigail / Weiss, Susan R / Li, Yize / Haddad, Leila / Deraska, Peter / Monahan, Caitlin / Kromer, Andrew / Graham-Wooten, Jevon / Taylor, Louis J / Abella, Benjamin S / Ganguly, Arupa / Collman, Ronald G / Van Duyne, Gregory D / Bushman, Frederic D

Genome biology

2021 Volume 22, Issue 1, Page(s) 169

Abstract: Background: Rapid spread of SARS-CoV-2 has led to a global pandemic, resulting in the need for rapid assays to allow diagnosis and prevention of transmission. Reverse transcription-polymerase chain reaction (RT-PCR) provides a gold standard assay for ... ...

Abstract	Background: Rapid spread of SARS-CoV-2 has led to a global pandemic, resulting in the need for rapid assays to allow diagnosis and prevention of transmission. Reverse transcription-polymerase chain reaction (RT-PCR) provides a gold standard assay for SARS-CoV-2 RNA, but instrument costs are high and supply chains are potentially fragile, motivating interest in additional assay methods. Reverse transcription and loop-mediated isothermal amplification (RT-LAMP) provides an alternative that uses orthogonal and often less expensive reagents without the need for thermocyclers. The presence of SARS-CoV-2 RNA is typically detected using dyes to report bulk amplification of DNA; however, a common artifact is nonspecific DNA amplification, which complicates detection. Results: Here we describe the design and testing of molecular beacons, which allow sequence-specific detection of SARS-CoV-2 genomes with improved discrimination in simple reaction mixtures. To optimize beacons for RT-LAMP, multiple locked nucleic acid monomers were incorporated to elevate melting temperatures. We also show how beacons with different fluorescent labels can allow convenient multiplex detection of several amplicons in "single pot" reactions, including incorporation of a human RNA LAMP-BEAC assay to confirm sample integrity. Comparison of LAMP-BEAC and RT-qPCR on clinical saliva samples showed good concordance between assays. To facilitate implementation, we developed custom polymerases for LAMP-BEAC and inexpensive purification procedures, which also facilitates increasing sensitivity by increasing reaction volumes. Conclusions: LAMP-BEAC thus provides an affordable and simple SARS-CoV-2 RNA assay suitable for population screening; implementation of the assay has allowed robust screening of thousands of saliva samples per week.
MeSH term(s)	COVID-19/diagnosis ; COVID-19 Testing ; Humans ; Molecular Diagnostic Techniques ; Nucleic Acid Amplification Techniques ; Nucleic Acid Probes/genetics ; RNA, Viral/genetics ; SARS-CoV-2/genetics ; SARS-CoV-2/isolation & purification ; Saliva/virology ; Sensitivity and Specificity
Chemical Substances	Nucleic Acid Probes ; RNA, Viral
Language	English
Publishing date	2021-06-03
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2040529-7
ISSN	1474-760X ; 1474-760X
ISSN (online)	1474-760X
ISSN	1474-760X
DOI	10.1186/s13059-021-02387-y
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Article ; Online: Correction for Merenstein et al., "Signatures of COVID-19 Severity and Immune Response in the Respiratory Tract Microbiome".

mBio

2022 Volume 13, Issue 5, Page(s) e0229322

Language	English
Publishing date	2022-08-31
Publishing country	United States
Document type	Journal Article ; Published Erratum
ZDB-ID	2557172-2
ISSN	2150-7511 ; 2161-2129
ISSN (online)	2150-7511
ISSN	2161-2129
DOI	10.1128/mbio.02293-22
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Article: Detection of SARS-CoV-2 RNA using RT-LAMP and molecular beacons

Genome biology. 2021 Dec., v. 22, no. 1

2021

Abstract: BACKGROUND: Rapid spread of SARS-CoV-2 has led to a global pandemic, resulting in the need for rapid assays to allow diagnosis and prevention of transmission. Reverse transcription-polymerase chain reaction (RT-PCR) provides a gold standard assay for ... ...

Abstract	BACKGROUND: Rapid spread of SARS-CoV-2 has led to a global pandemic, resulting in the need for rapid assays to allow diagnosis and prevention of transmission. Reverse transcription-polymerase chain reaction (RT-PCR) provides a gold standard assay for SARS-CoV-2 RNA, but instrument costs are high and supply chains are potentially fragile, motivating interest in additional assay methods. Reverse transcription and loop-mediated isothermal amplification (RT-LAMP) provides an alternative that uses orthogonal and often less expensive reagents without the need for thermocyclers. The presence of SARS-CoV-2 RNA is typically detected using dyes to report bulk amplification of DNA; however, a common artifact is nonspecific DNA amplification, which complicates detection. RESULTS: Here we describe the design and testing of molecular beacons, which allow sequence-specific detection of SARS-CoV-2 genomes with improved discrimination in simple reaction mixtures. To optimize beacons for RT-LAMP, multiple locked nucleic acid monomers were incorporated to elevate melting temperatures. We also show how beacons with different fluorescent labels can allow convenient multiplex detection of several amplicons in “single pot” reactions, including incorporation of a human RNA LAMP-BEAC assay to confirm sample integrity. Comparison of LAMP-BEAC and RT-qPCR on clinical saliva samples showed good concordance between assays. To facilitate implementation, we developed custom polymerases for LAMP-BEAC and inexpensive purification procedures, which also facilitates increasing sensitivity by increasing reaction volumes. CONCLUSIONS: LAMP-BEAC thus provides an affordable and simple SARS-CoV-2 RNA assay suitable for population screening; implementation of the assay has allowed robust screening of thousands of saliva samples per week.
Keywords	DNA ; RNA ; Severe acute respiratory syndrome coronavirus 2 ; fluorescence ; genome ; humans ; pandemic ; reverse transcriptase polymerase chain reaction ; reverse transcription ; reverse transcription loop-mediated isothermal amplification ; saliva
Language	English
Dates of publication	2021-12
Size	p. 169.
Publishing place	BioMed Central
Document type	Article
ZDB-ID	2040529-7
ISSN	1474-760X
ISSN	1474-760X
DOI	10.1186/s13059-021-02387-y
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