A case of reassortant seasonal influenza A(H1N2) virus, Denmark, April 2019

Trebbien, Ramona; Koch, Anders; Nielsen, Lene; Kur, Dår K; Westerström, Pontus; Krause, Tyra Grove

doi:10.2807/1560-7917.es.2019.24.27.1900406

Cited by 4 publications

(1 citation statement)

References 14 publications

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“…In the human population, H3N2 viruses have cocirculated with H1N1 viruses from 1977 to 2009 and with H1N1p viruses since 2009. Although reassortment readily occurs between H3N2, H1N1, and H1N1p subtypes ( 67 – 69 ), only a few cases of H3N2-gene introduction, i.e., incorporation of the H3N2-NA segment into the H1N1p-background, have been reported ( 70 – 72 ), whilst reassortants harboring the H3N2-(PB1+NA) gene combination in an H1N1p background have not been detected in natural infections. In the H3N2-background, individual incorporations of H1N1p-segments, namely M, NS, NP, and PB1, are the most frequent in our experimental setting, in agreement with data from Phipps et al.…”

Section: Discussionmentioning

confidence: 99%

High-throughput droplet-based analysis of influenza A virus genetic reassortment by single-virus RNA sequencing

Chen

Karuppusamy

O’Neill

et al. 2023

Proc. Natl. Acad. Sci. U.S.A.

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The segmented RNA genome of influenza A viruses (IAVs) enables viral evolution through genetic reassortment after multiple IAVs coinfect the same cell, leading to viruses harboring combinations of eight genomic segments from distinct parental viruses. Existing data indicate that reassortant genotypes are not equiprobable; however, the low throughput of available virology techniques does not allow quantitative analysis. Here, we have developed a high-throughput single-cell droplet microfluidic system allowing encapsulation of IAV-infected cells, each cell being infected by a single progeny virion resulting from a coinfection process. Customized barcoded primers for targeted viral RNA sequencing enabled the analysis of 18,422 viral genotypes resulting from coinfection with two circulating human H1N1pdm09 and H3N2 IAVs. Results were highly reproducible, confirmed that genetic reassortment is far from random, and allowed accurate quantification of reassortants including rare events. In total, 159 out of the 254 possible reassortant genotypes were observed but with widely varied prevalence (from 0.038 to 8.45%). In cells where eight segments were detected, all 112 possible pairwise combinations of segments were observed. The inclusion of data from single cells where less than eight segments were detected allowed analysis of pairwise cosegregation between segments with very high confidence. Direct coupling analysis accurately predicted the fraction of pairwise segments and full genotypes. Overall, our results indicate that a large proportion of reassortant genotypes can emerge upon coinfection and be detected over a wide range of frequencies, highlighting the power of our tool for systematic and exhaustive monitoring of the reassortment potential of IAVs.

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