Evaluating methods for Avian avulavirus-1 whole genome sequencing

Tal, Saar; Izhak, Meirav Ben; Wachtel, Chaim; Wiseman, Anat; Braun, Tzipi; Yechezkel, Elinor; Golan, Einav; Hadas, Ruth; Turjeman, Adi; Banet-Noach, Caroline; Bronstein, Michal; Lublin, Avishai; Berman, Elyakum; Raviv, Ziv; Pirak, M.; Klement, Eyal; Louzoun, Yoram

doi:10.1016/j.gene.2019.100004

Cited by 1 publication

(1 citation statement)

References 52 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In recent years, untargeted next-generation sequencing (NGS) technology has been increasingly employed for the detection and characterization of RNA viruses, particularly in the context of avian diseases ( Chen et al, 2013 ; Tang et al, 2015 ; Goraichuk et al, 2016 , 2017 , 2019 , 2021a , b ; Croville et al, 2018 ; Ferreri et al, 2019 ; Tal et al, 2019 ; Patzina-Mehling et al, 2020 ; Crossley et al, 2021 ; Chrzastek et al, 2022 ; Damir et al, 2023 ; Ip et al, 2023 ; Kariithi et al, 2023 ; Techera et al, 2023 ). Avian viruses, including highly pathogenic avian influenza virus (AIV) and Newcastle disease virus (NDV), pose significant threats to poultry populations and public health ( Alexander, 1998 ; Suarez, 2017 ; Swayne et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

The 28S rRNA RT-qPCR assay for host depletion evaluation to enhance avian virus detection in Illumina and Nanopore sequencing

Goraichuk,

Harden,

Spackman

et al. 2024

Front. Microbiol.

View full text Add to dashboard Cite

Abundant host and bacterial sequences can obscure the detection of less prevalent viruses in untargeted next-generation sequencing (NGS). Efficient removal of these non-targeted sequences is vital for accurate viral detection. This study presents a novel 28S ribosomal RNA (rRNA) RT-qPCR assay designed to assess the efficiency of avian rRNA depletion before conducting costly NGS for the detection of avian RNA viruses. The comprehensive evaluation of this 28S-test focuses on substituting DNase I with alternative DNases in our established depletion protocols and finetuning essential parameters for reliable host rRNA depletion. To validate the effectiveness of the 28S-test, we compared its performance with NGS results obtained from both Illumina and Nanopore sequencing platforms. This evaluation utilized swab samples from chickens infected with highly pathogenic avian influenza virus, subjected to established and modified depletion protocols. Both methods significantly reduced host rRNA levels, but using the alternative DNase had superior performance. Additionally, utilizing the 28S-test, we explored cost- and time-effective strategies, such as reduced probe concentrations and other alternative DNase usage, assessed the impact of filtration pre-treatment, and evaluated various experimental parameters to further optimize the depletion protocol. Our findings underscore the value of the 28S-test in optimizing depletion methods for advancing improvements in avian disease research through NGS.

show abstract