Detection of Clade 2.3.4.4b Avian Influenza A(H5N8) Virus in Cambodia, 2021

Edwards, Kimberly M.; Siegers, Jurre Y.; Wei, Xiaoman; Aziz, Ammar; Deng, Yi‐Mo; Yann, Sokhoun; Bun, Chan; Bunnary, Seng; Izzard, Leonard; Hak, Makara; Thielen, Peter; Tum, Sothyra; Wong, Frank; Lewis, Nicola S.; James, Joe; Claes, Filip; Barr, Ian G.; Vijaykrishna, Dhanasekaran; Karlsson, Erik A.

doi:10.3201/eid2901.220934

Cited by 4 publications

(7 citation statements)

References 8 publications

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“…Viral RNA was extracted using the QIAamp Viral RNA Mini Kit (Qiagen, Maryland, USA) according to the manufacturer’s protocol. All samples were screened by real-time reverse transcription PCR with M gene, as previously described [ 17 , 23 ]. Samples with a cycle threshold (Ct) of less than 40 were deemed positive.…”

Section: Methodsmentioning

confidence: 99%

Detection and phylogenetic analysis of contemporary H14N2 Avian influenza A virus in domestic ducks in Southeast Asia (Cambodia)

Siegers,

Wille,

Yann

et al. 2024

Emerging Microbes & Infections

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Avian influenza virus (AIV) in Asia is a complex system with numerous subtypes and a highly porous wild birds-poultry interface. Certain AIV subtypes, such as H14, are underrepresented in current surveillance efforts, leaving gaps in our understanding of their ecology and evolution. The detection of rare subtype H14 in domestic ducks in Southeast Asia comprises a geographic region and domestic bird population previously unassociated with this subtype. These H14 viruses have a complex evolutionary history involving gene reassortment events. They share sequence similarity to AIVs endemic in Cambodian ducks, and Eurasian low pathogenicity and high pathogenicity H5Nx AIVs. The detection of these H14 viruses in Southeast Asian domestic poultry further advances our knowledge of the ecology and evolution of this subtype and reinforces the need for continued, longitudinal, active surveillance in domestic and wild birds. Additionally, in vivo and in vitro risk assessment should encompass rare AIV subtypes, as they have the potential to establish in poultry systems.

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Section: Methodsmentioning

confidence: 99%

Detection and phylogenetic analysis of contemporary H14N2 Avian influenza A virus in domestic ducks in Southeast Asia (Cambodia)

Siegers,

Wille,

Yann

et al. 2024

Emerging Microbes & Infections

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“…The presence of influenza virus in the supernatant and allantoic fluid was tested using a hemagglutination (HA) assay 43 . Both original samples and isolates were subtyped using influenza RT-PCR assays to test for human seasonal and avian subtypes 28,41,44 . In total, 45 unique isolates were sequenced for this study, with seasonal human influenza used as positive controls (Table S1).…”

Section: Methodsmentioning

confidence: 99%

“…Nanopore Sequencing Whole genomes were amplified using a modified multi-segment RT-PCR approach that incorporates integrated molecular indices, with a degenerate base at position four of the Uni-12 primer (5'AGCRAAAGCAGG) 28,46 . Resulting indexed RT-PCR products were pooled and prepared for sequencing using ligation sequencing kit SQK-LSK109 (R9 chemistry) or SQK-LSK114 (R10 chemistry), and sequenced on the GridION platform (Oxford Nanopore Technologies, Oxford, UK).…”

Section: Sanger Sequencingmentioning

confidence: 99%

“…ONT sequencing utility has been demonstrated for whole genome sequencing from cultured isolates and clinical specimens. [19][20][21][22][23][24] In addition to human seasonal surveillance, these techniques have been important in influenza surveillance and responseincluding in field deployable versions 25,26 -in avian species 27,28 , swine 26,29 , and AIV spillover into humans 17 . However, the hemagglutinin (HA) cleavage site of avian influenza, a major virulence determinant, is composed of basic amino acids encoded by a low-complexity A/G-rich portion of the genome.…”

Section: Introductionmentioning

confidence: 99%

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Improved Resolution of Highly Pathogenic Avian Influenza Virus Haemagglutinin Cleavage Site Using Oxford Nanopore R10 Sequencing Chemistry

Ratcliff,

Merritt,

Gooden

et al. 2023

Preprint

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Highly pathogenic avian influenza viruses continue to pose global risks to One Health, including agriculture, public, and animal health. Rapid and accurate genomic surveillance is critical for monitoring viral mutations, tracing transmission, and guiding interventions in near real-time. Oxford Nanopore sequencing holds promise for real-time influenza genotyping, but data quality from R9 chemistry has limited its adoption due to challenges resolving low-complexity regions such as the biologically critical hemagglutinin cleavage site, a homopolymer of basic amino acids that distinguish highly pathogenic strains. In this study, human and avian influenza isolates (n=45) from Cambodia were sequenced using both R9.4.1 and R10.4.1 flow cells and chemistries to evaluate performance between approaches. Overall, R10.4.1 yielded increased data output with higher average quality compared to R9.4.1, producing improved consensus sequences using a reference-based bioinformatics approach. R10.4.1 had significantly lower minor population insertion and deletion frequencies, driven by improved performance in low sequence complexity regions prone to insertion and deletion errors, such as homopolymers. Within the hemagglutinin cleavage site, R10.4.1 resolved the correct motif in 90% of genomes compared to only 60% with R9.4.1. Further examination showed reduced frameshift mutations in consensus sequences generated with R10.4.1 that could result in incorrectly classified virulence. Improved consensus genome quality from nanopore sequencing approaches, especially across biologically important low-complexity regions, is critical to reduce subjective hand-curation and will improve local and global genomic surveillance responses.

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“…As a result, significant reductions in the circulation of all common respiratory viruses have been reported globally since the pandemic onset (20,21,22,23,24,25), and winter epidemics were notably absent in 2020 and 2021. However, intermittent outbreaks of influenza (26), RSV (27), and HRV (23,28,29) have occurred in locations where control measures were relaxed intermittently or completely.…”

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confidence: 99%

Molecular epidemiology of Human Rhinoviruses in children in Hong Kong, 2020-2021

Gurung

Edwards

Chan

et al. 2023

Preprint

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Background COVID-19 related public health and social measures (PHSMs) worldwide have influenced respiratory virus epidemiology. In Hong Kong, paediatric hospitalisations associated with non-COVID-19 respiratory viruses declined in 2020. As PHSMs eased, rhinoviruses/enteroviruses became the primary detected respiratory viruses. This study examines the genetic diversity in resurgent human rhinovirus (HRV) cases. Methods We sequenced rhinovirus/enterovirus samples from children at Queen Mary Hospital, Hong Kong, between August 2020 and October 2021 to estimate changes in HRV genotypes and describe their epidemic characteristics. Whole genome sequencing was performed on the three most prevalent HRV genotypes to infer patterns of virus introduction and persistence. Results Despite reduced respiratory virus circulation, HRV type A and C infections persisted in children, with sporadic detection of HRV B and other respiratory viruses. A resurgence of HRV A cases in November 2020, dominated by genotypes A47 and A101, was observed during the relaxation of PHSMs between the third and fourth waves of COVID-19. Strict PHSMs implemented during the fourth wave, including school closures, substantially reduced respiratory virus circulation, though overall diversity increased due to heightened vigilance. HRV genotype A49 became predominant in May 2021 upon relaxation of control measures, with phylogenetic analysis suggesting persistence of multiple transmission lineages despite strict PHSMs. Genotypes A49 and A47 were frequently associated with upper respiratory tract infections, highlighting their epidemic potential. Conclusion This study underscores the impact of control measures on HRV genetic diversity and highlights the need for continuous surveillance and sequencing to inform public health interventions.

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Detection of Clade 2.3.4.4b Avian Influenza A(H5N8) Virus in Cambodia, 2021

Cited by 4 publications

References 8 publications

Detection and phylogenetic analysis of contemporary H14N2 Avian influenza A virus in domestic ducks in Southeast Asia (Cambodia)

Detection and phylogenetic analysis of contemporary H14N2 Avian influenza A virus in domestic ducks in Southeast Asia (Cambodia)

Improved Resolution of Highly Pathogenic Avian Influenza Virus Haemagglutinin Cleavage Site Using Oxford Nanopore R10 Sequencing Chemistry

Molecular epidemiology of Human Rhinoviruses in children in Hong Kong, 2020-2021

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