In a recent study published in PNAS, researchers explored the evolutionary dynamics of swine influenza A virus (swIAV) populations in Asia's southern and eastern regions.
Study: The genomic landscape of swine influenza A viruses in Southeast Asia. Image Credit: LiyaGraphics/Shutterstock.com
Background
Swine in Southeast Asia is a primary source of pandemic influenza A viruses (IAVs), causing zoonotic and reverse-zoonotic transmission. Global swine production and trade increase the risk of swine IAV transmission.
However, the lack of systematic surveillance infrastructure hinders identifying emerging pathogens in pigs and their potential for zoonotic spillover. Effective surveillance is needed to uncover emerging viral strains, but understanding the swIAV genomic landscape remains limited.
About the study
In the present longitudinal study, researchers performed IAV surveillance on 18 slaughterhouse pigs in Cambodia's four provinces (Kandal, Kampong Speu, Takeo, and Phnom Penh) between March 2020 and July 2022.
A total of 4,089 nasopharyngeal swabs from pigs were taken to indicate the long-unnoticed circulation and establishment of several swIAV viruses in Cambodia, including human H1N1/pdm09 and H3N2 viruses transmitted to swine by reverse zoonosis.
The team extracted swIAV ribonucleic acid (RNA) from the samples, and IAV identification was accomplished using quantitative reverse-transcription polymerase chain reaction (RT-qPCR).
Subsequently, positive samples were submitted to next-generation sequencing (SGS), which measured amplified complementary deoxyribonucleic acid (cDNA). Pooled DNA libraries were created and sequenced.
For the evolutionary study, accessible worldwide H1N1 influenza A virus genomes (till September 8, 2022) obtained from swine and human hosts were retrieved from the worldwide Initiative on Sharing All Influenza Data (GISAID) database and the National Center for Biotechnology Information (NCBI) GenBank.
The genetic datasets were subsampled to 766 H3 and 1,008 H1 sequences. Maximum likelihood phylogenies were generated for all genetic segments of swIAV from Cambodia, including hemagglutinin (HA) and neuraminidase (NA) genes, polymerase PB (PB), and polymerase PA (PA), matrix protein (MP), non-structural protein (NS), and nucleoprotein (NP).
In total, 986, 773, 1,009, 766, 915, 923, 924, 927, 927, and 927 segments of the N1-NA, N2-NA, H1-HA, H3-HA, PA, PB1, PB2, MP, NS, and NP, respectively, were analyzed. Phylogeographic analysis was used to investigate the geographical patterns of European avian-like (EA)-swine H1-type viral strains.
The team downloaded all H1 sequences of EA-swine viral organisms from the GISAID and NCBI databases (between 2005 and December 20, 2022), and 7.0 distinct geographical regions (Southeastern Asia, Europe, northern China, eastern China, western China, northeastern China, and southern and central China) were coded.
Results
Swine IAVs were detected and characterized in slaughterhouse pigs in the southern and eastern regions of Asia, with 72 pigs (2.0%) being IAV-positive by RT-PCR. Kandal province had a higher positivity rate of IAV (4.50%) than other provinces.
The study found that China's southern and central regions were the dominant sources of swine lineages spread to other parts of China and Asia's south and eastern areas.
The researchers also discovered nine unique swIAV viruses in Cambodia that split from their nearest progenitors between 2.0 and 15 billion years ago, indicating tremendous undiscovered genomic diversity.
A comparable era of cryptic swIAV circulation occurred for decades preceding the H1N1/2009 outbreak. The findings revealed a complex genomic landscape of swIAV in Southeast Asia, shaped by the repeated introduction and reassortment of virus lineages.
The researchers also unraveled the parental origin and gene constellations of the EA H1N2 reassortant viruses that likely emerged in late 2014, approximately seven years before their initial detection in Cambodia in 2021.
The novel EA H1N2 virus had a distinct genetic constellation comprising a swine-origin EA G4-like H1-HA, an N2-HA originating from human viruses, and internal genes from the triple-reassortant internal gene (TRIG) viruses.
The swine IAV strains circulating in the southern and eastern regions of Asia acquired reassorted genetic segments from multiple geographical origins, facilitated by the intercontinental and intracontinental spread of previously segregated swIAV lineages.
Multiple instances of EA N1 coupled with CS H1 were found. The H1N1/pdm09 viral strain was found to be circulating enzootically in Cambodian swine herds. The team also identified several reverse zoonotic H3 strains among Cambodian pigs that were circulating unidentified for nearly ten years.
The findings emphasized the importance of early identification of novel swIAV lineages. They highlighted concerns surrounding the inter-regional spread of swIAVs as a key driver in the emergence of new virus lineages, especially in the southern and eastern regions of Asia, where surveillance is sporadic.
The researchers found multiple instances of CS H1 coupled with EA N1 and evidence of global intermixing of previously distinct virus populations.
Conclusions
Overall, the study findings highlighted the genomic landscape of swIAVs in Southeast Asia. The results showed an EA H1N2 reassortant genotype in Asia, with China being the leading source of EA swine viruses.
The study revealed nine distinct swIAV HA and NA lineages cocirculating in Cambodian pig populations, highlighting the complex evolutionary processes in the region.
As swine-rearing activities develop, pig surveillance must become more frequent and consistent to discover novel viruses and assess their zoonotic risk.
By monitoring viruses with zoonotic transmission potential, more efficient and continuous surveillance approaches, such as metagenomic monitoring and automated analytical instruments, can enhance animal health and help in human health.
Further research is required to understand the pandemic threat of EA G4-like H1N2 viruses.
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