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Wild aquatic birds are considered the natural hosts of 16 HA (H1–H16) and 9 NA (N1–N9) subtypes of influenza A viruses (FLUAV) found in different combinations. H14 FLUAVs are rarely detected in nature. Since 2011, H14 FLUAVs have been consistently detected in Guatemala, leading to the largest collection of this subtype from a single country. All H14 FLUAVs in Guatemala were detected from blue-winged teal samples. In this report, 17 new full-length H14 FLUAV genome sequences detected from 2014 until 2019 were analyzed and compared to all published H14 sequences, including Guatemala, North America, and Eurasia. The H14 FLUAVs identified in Guatemala were mostly associated with the N3 subtype (n = 25), whereas the rest were paired with either N4 (n = 7), N5 (n = 4), N6 (n = 1), and two mixed infections (N3/N5 n = 2, and N2/N3 n = 1). H14 FLUAVs in Guatemala belong to a distinct H14 lineage in the Americas that is evolving independently from the Eurasian H14 lineage. Of note, the ORF of the H14 HA segments showed three distinct motifs at the cleavage site, two of these containing arginine instead of lysine in the first and fourth positions, not previously described in other countries. The effects of these mutations on virus replication, virulence, and/or transmission remain unknown and warrant further studies. [ABSTRACT FROM AUTHOR]

Due to their documented epidemiological relevance as hosts for influenza A viruses (IAV), humans, poultry and pigs in backyard production systems (BPS) within wetlands could be key to the emergence of novel IAV variants able to transmit between humans or animals. To better understand the circulation of IAV at the human–animal interface of BPS within wetlands, we studied IAV in backyard duck flocks and pig herds in the Pacific Coast of Guatemala. From April 2013 to October 2014, we estimated the monthly IAV per cent seropositive and viral positive flocks and herds in two resource‐limited communities. We detected antibodies in sera against the IAV nucleoprotein through ELISA. We also detected IAV viral RNA in respiratory (ducks and pigs) and cloacal (ducks) swabs through rRT‐PCR directed at the matrix gene. We attempted viral isolation in eggs or MDCK cells followed by sequencing from swabs positive for IAV. During our study period, IAV seropositivity in duck flocks was 38%, and viral positivity was 23% (n = 86 BPS sampled). IAV seropositivity in pig herds was 42%, and viral positivity was 20% (n = 90 BPS sampled). Both flocks and herds had detectable antibodies against IAV mostly year‐round, and IAV was detected in several months. We isolated an H3N2 virus from one pig sampled at the end of 2013. Standard nucleotide BLAST searches indicate that the isolated virus was similar to seasonal viruses circulating in humans, suggesting human‐to‐pig transmission. Our data show concurrent circulation of IAV in multiple species of poultry and pigs that were commingled in rudimentary conditions in proximity to humans, but no significant risk factors could be identified. [ABSTRACT FROM AUTHOR]

The greatest diversity of influenza A virus (IAV) is found in wild aquatic birds of the orders Anseriformes and Charadriiformes. In these birds, IAV replication occurs mostly in the intestinal tract. Fecal, cloacal, and/or tracheal swabs are typically collected and tested by real‐time RT‐PCR (rRT‐PCR) and/or by virus isolation in embryonated chicken eggs in order to determine the presence of IAV. Virus isolation may impose bottlenecks that select variant populations that are different from those circulating in nature, and such bottlenecks may result in artifactual representation of subtype diversity and/or underrepresented mixed infections. The advent of next‐generation sequencing (NGS) technologies provides an opportunity to explore to what extent IAV subtype diversity is affected by virus isolation in eggs. In the present work, we evaluated the advantage of sequencing by NGS directly from swab material of IAV rRT‐PCR‐positive swabs collected during the 2013–14 surveillance season in Guatemala and compared to results from NGS after virus isolation. The results highlight the benefit of sequencing IAV genomes directly from swabs to better understand subtype diversity and detection of alternative amino acid motifs that could otherwise escape detection using traditional methods of virus isolation. In addition, NGS sequencing data from swabs revealed reduced presence of defective interfering particles compared to virus isolates. We propose an alternative workflow in which original swab samples positive for IAV by rRT‐PCR are first subjected to NGS before attempting viral isolation. This approach should speed the processing of samples and better capture natural IAV diversity. OPEN RESEARCH BADGES: This article has earned an Open Data Badge for making publicly available the digitally‐shareable data necessary to reproduce the reported results. The data is available at https://doi.org/10.5061/dryad.3h2n106. [ABSTRACT FROM AUTHOR]