Study: Natural Reassortment of Eurasian Avian-like Swine H1N1 and Avian H9N2 Influenza Viruses in Pigs, China. Image Credit: Kateryna Kon/Shutterstock

Researchers report detection of swine influenza A virus with avian H9N2 gene segments

In a recent study published in Emerging infectious diseasesresearchers have reported a natural reassortment of influenza viruses in pigs.

Study: Natural reassortment of Eurasian swine H1N1 and avian H9N2 influenza viruses in pigs, China. Image Credit: Kateryna Kon/Shutterstock


Pigs are considered vessels for reassortment of influenza A viruses (IAVs). Porcine, human and avian IAVs could co-infect pigs and create new reassortant IAVs that could cause zoonoses or pandemics. One study provided evidence of bird flu virus infections in pigs. Avian IAVs linked to human infections have been revealed to contain internal genes from avian H9N2 viruses, implying that viral H9N2 genes may facilitate zoonotic transmission.

About the study

In the current study, researchers detected a porcine IAV with matrix (M) and basic polymerase 1 (PB1) gene segments from H9N2 avian influenza. The research team collected nasal swab samples from imported pigs at a local abattoir in Hong Kong as part of the monthly influenza monitoring program, which resumed in April 2021 following disruptions due to the coronavirus disease 2019 (COVID-19) outbreak.

IAVs were isolated from swab specimens using Madin-Darby Canine Kidney (MDCK) cells. Cultures exhibiting a cytopathic effect (CPE) were identified and tested by a standard hemagglutination test using turkey red blood cells. Cultures positive for hemagglutination were subjected to a reverse transcription-polymerase chain reaction (RT-PCR) test, specific for M segments. Next-generation sequencing (NGS) was performed on samples positive for PCR.

Results and conclusion

The researchers collected 829 nasal swab specimens from April 2021 to February 2022. They isolated eight IAVs, seven from August 2021 and one from September 2021. The authors reported that the inferred viral sequences had been made available on the Global Avian Influenza Data Sharing Initiative. (GISAID). These sequences were compared to references.

All of the August 2021 IAVs were H3N2 viruses, and their neuraminidase (NA) and hemagglutinin (HA) segments were related to human-like porcine IAV H3N2. However, the inner segments of the gene were derived from the pandemic H1N1 (pH1N1) influenza virus. Although these IAVs are not genetically identical, they share important similarities. The pigs from which the IAVs were isolated were imported from two provinces in southern China.

Since the imported animals were slaughtered within 24 hours, the researchers speculated that flu transmission could have occurred between pigs outside of Hong Kong prior to importation. Previously, this genotype (H3N2) was found in pigs raised in Guangxi, China. From the September 2021 samples, the team isolated a porcine H1N1 IAV and noted reassortments of several porcine IAV lineages. The M and PB1 segments were of H9N2 avian origin.

In addition, it also contained acid polymerase, PB2 and NA segments from the pH1N1 lineage. The NA and HA gene segments belonged to the Eurasian avian-like H1N1 lineage, and the non-structural gene segment showed triple lineage reassortment. This virus isolate was further purified on plaque assays, and the resulting plaque-purified clones were genotypically identical, excluding the possibility of mixed infection.

It also included an M gene segment from the G1 type H9N2 line and a PB1 gene segment from the SH/F/98 type line. Such similar sequences have been detected in zoonotic IAVs in humans. However, both sequences did not contain known mutations for adaptation in mammalian hosts. Additionally, the encoded PB1 and M proteins had amino acid sequences rarely found in avian and mammalian IAVs.

A recent study reported multiple reassortments of the Eurasian avian-like H1N1 swine flu; of these, a reassortment class (genotype 4) had a similar genotype to the IAV isolated in September 2021. However, its M and PB1 segments were of pH1N1 lineage, unlike that reported in the present study. The authors postulate that the Eurasian avian-type and genotype 4 H1N1 viruses may have originated from a common ancestor. Nevertheless, further evaluation of its pandemic potential is needed.


In conclusion, various zoonotic IAVs in humans exhibit genes derived from H9N2 viruses. These results indicated that H9N2 IAVs infect pigs and reassort with porcine IAVs, highlighting the continued need to monitor porcine IAVs.

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