Accelerating bioinformatics implementation in public health

Libuit, Kevin G.; Doughty, Emma L.; Otieno, James R.; Ambrosio, Frank; Kapsak, Curtis J.; Smith, Emily A.; Wright, Sage M.; Scribner, Michelle R.; Petit III, Robert A.; Mendes, Catarina Inês; Huergo, Marcela; Legacki, Gregory; Loreth, Christine; Park, Daniel J.; Sevinsky, Joel R.

doi:10.1099/mgen.0.001051

Cited by 8 publications

(7 citation statements)

References 71 publications

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“…Modifications to TheiaCoV_Illumina_PE were made to support monkeypox virus genomic characterization; these modifications accommodated the use of a monkeypox virus reference sequence and primer scheme for consensus genome assembly. These updates were made available in the TheiaCov v2.2.0 release [ 33 ]. The Terra platform allows for consistently deployed bioinformatics environments without the need for on-site computing infrastructure and high-level programming capabilities.…”

Section: Methodsmentioning

confidence: 99%

Development of an amplicon-based sequencing approach in response to the global emergence of mpox

et al. 2023

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The 2022 multicountry mpox outbreak concurrent with the ongoing Coronavirus Disease 2019 (COVID-19) pandemic further highlighted the need for genomic surveillance and rapid pathogen whole-genome sequencing. While metagenomic sequencing approaches have been used to sequence many of the early mpox infections, these methods are resource intensive and require samples with high viral DNA concentrations. Given the atypical clinical presentation of cases associated with the outbreak and uncertainty regarding viral load across both the course of infection and anatomical body sites, there was an urgent need for a more sensitive and broadly applicable sequencing approach. Highly multiplexed amplicon-based sequencing (PrimalSeq) was initially developed for sequencing of Zika virus, and later adapted as the main sequencing approach for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Here, we used PrimalScheme to develop a primer scheme for human monkeypox virus that can be used with many sequencing and bioinformatics pipelines implemented in public health laboratories during the COVID-19 pandemic. We sequenced clinical specimens that tested presumptively positive for human monkeypox virus with amplicon-based and metagenomic sequencing approaches. We found notably higher genome coverage across the virus genome, with minimal amplicon drop-outs, in using the amplicon-based sequencing approach, particularly in higher PCR cycle threshold (Ct) (lower DNA titer) samples. Further testing demonstrated that Ct value correlated with the number of sequencing reads and influenced the percent genome coverage. To maximize genome coverage when resources are limited, we recommend selecting samples with a PCR Ct below 31 Ct and generating 1 million sequencing reads per sample. To support national and international public health genomic surveillance efforts, we sent out primer pool aliquots to 10 laboratories across the United States, United Kingdom, Brazil, and Portugal. These public health laboratories successfully implemented the human monkeypox virus primer scheme in various amplicon sequencing workflows and with different sample types across a range of Ct values. Thus, we show that amplicon-based sequencing can provide a rapidly deployable, cost-effective, and flexible approach to pathogen whole-genome sequencing in response to newly emerging pathogens. Importantly, through the implementation of our primer scheme into existing SARS-CoV-2 workflows and across a range of sample types and sequencing platforms, we further demonstrate the potential of this approach for rapid outbreak response.

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Section: Methodsmentioning

confidence: 99%

Development of an amplicon-based sequencing approach in response to the global emergence of mpox

et al. 2023

Self Cite

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“…Detailed protocols and quality control criteria were described previously [ 15 , 16 ]. Illumina sequencing data and Clear Labs consensus genomes were analyzed on Terra.bio using the TheiaCoV_Illumina_PE v2.3.0 and TheiaCoV_FASTA v2.3.0 workflows, respectively, (Theiagen Genomics, Highlands Ranch, CO) [ 17 , 18 , 19 , 20 , 21 ] with Wuhan-Hu-1/2019 (GenBank MN908947) used as the reference genome. Consensus genomes used in this study all had greater than 90% reference genome coverage [ 22 ].…”

Section: Methodsmentioning

confidence: 99%

Transient SARS-CoV-2 RNA-Dependent RNA Polymerase Mutations after Remdesivir Treatment for Chronic COVID-19 in Two Transplant Recipients: Case Report and Intra-Host Viral Genomic Investigation

Yang,

Multani,

Garrigues

et al. 2023

Microorganisms

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Remdesivir is the first FDA-approved drug for treating severe SARS-CoV-2 infection and targets RNA-dependent RNA polymerase (RdRp) that is required for viral replication. To monitor for the development of mutations that may result in remdesivir resistance during prolonged treatment, we sequenced SARS-CoV-2 specimens collected at different treatment time points in two transplant patients with severe COVID-19. In the first patient, an allogeneic hematopoietic stem cell transplant recipient, a transient RdRp catalytic subunit mutation (nsp12:A449V) was observed that has not previously been associated with remdesivir resistance. As no in vitro study had been conducted to elucidate the phenotypic effect of nsp12:A449V, its clinical significance is unclear. In the second patient, two other transient RdRp mutations were detected: one in the catalytic subunit (nsp12:V166A) and the other in an accessory subunit important for processivity (nsp7:D67N). This is the first case report for a potential link between the nsp12:V166A mutation and remdesivir resistance in vivo, which had only been previously described by in vitro studies. The nsp7:D67N mutation has not previously been associated with remdesivir resistance, and whether it has a phenotypic effect is unknown. Our study revealed SARS-CoV-2 genetic dynamics during remdesivir treatment in transplant recipients that involved mutations in the RdRp complex (nsp7 and nsp12), which may be the result of selective pressure. These results suggest that close monitoring for potential resistance during the course of remdesivir treatment in highly vulnerable patient populations may be beneficial. Development and utilization of diagnostic RdRp genotyping tests may be a future direction for improving the management of chronic COVID-19.

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“…Whole-genome sequencing followed by genomic analyses ( 2 , 3 ) were performed on outbreak specimens obtained from 45 patients (14 fully vaccinated, six partially vaccinated, and 25 unvaccinated) as part of LAC’s Monkeypox virus (MPXV) genomic surveillance program; specimens from three patients were sequenced by the California Department of Public Health. Phylogenetic analysis ( 4 ) (data set tag 2023–08–01T12:00:00Z) determined that 32 (71.1%) cases involved MPXV belonging to the B.1.20 lineage of clade IIb, which is currently the dominant lineage identified through surveillance in the United States.…”

Section: Epidemiologic Investigation and Findingsmentioning

confidence: 99%

Mpox Outbreak — Los Angeles County, California, May 4–August 17, 2023

Leonard,

Poortinga,

Nguyen

et al. 2024

MMWR Morb. Mortal. Wkly. Rep.

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Accelerating bioinformatics implementation in public health

Cited by 8 publications

References 71 publications

Development of an amplicon-based sequencing approach in response to the global emergence of mpox

Development of an amplicon-based sequencing approach in response to the global emergence of mpox

Transient SARS-CoV-2 RNA-Dependent RNA Polymerase Mutations after Remdesivir Treatment for Chronic COVID-19 in Two Transplant Recipients: Case Report and Intra-Host Viral Genomic Investigation

Mpox Outbreak — Los Angeles County, California, May 4–August 17, 2023

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