Objective To conduct a pathogenic identification and genetic characteristics analysis on the samples from the first reported case of a clustered fever outbreak at a kindergarten in Fangshan District, Beijing in September 2021.

Methods The epidemiological information and throat swab specimens of the cases of the outbreak were collected. After extracting the viral nucleic acid of the specimen, some common respiratory viruses were screened. Human parainfluenza virus type 3 (HPIV3) nucleic acid-positive specimens were inoculated into Hep-2 cells for virus isolation and culture. Reverse transcription - polymerase chain reaction was used to amplify the coding sequences of hemagglutinin-neuraminidase protein (HN) and fusion protein (F) protein genes in HPIV3. BioEdit 7.0.9 software was used to analyze nucleotide and amino acid variations of the HN and F genes of the virus. Mega 6.0 software was used to construct phylogenetic analysis of HN and F genes, and the glycosylation sites of HN and F protein of isolated virus strains in this HPIV3 outbreak were analyzed online. Recombinant analysis was performed using Simplot 3.5.1,followed by reconfirmation of the recombination results using RDP4 software.

Results The multiple respiratory pathogens screening test showed that in total of 9 specimens , 8 were HPIV3 nucleic acid positive , of which 6 were HPIV3 single positive, 1 was co-infected with mycoplasma pneumoniae, the other co-infected with OC43 coronavirus. Subsequently, HN and F protein gene of 7 HPIV3 virus strains isolated successfully were sequenced. The homology between the HN and F protein gene sequences of the outbreak strains and the foreign Wash/47885/57 nucleotide sequences is 91.21%−91.28% and 93.33%, respectively. The homology with the 2020 Shenyang strain in China (GenBank only retrieved its HN protein gene sequence) is over 99.73%. Phylogenetic analysis showed the sequence of the outbreak strains was located in the C3a. 1 subgroup. Furthermore, both its HN and F proteins have potential N-glycosylation sites and O-glycosylation sites. The recombination analysis results showed that the HN protein genes of the HPIV3 outbreak strains had recombination events.

Conclusion The this outbreak of acute febrile respiratory illness was caused by HPIV3 in Beijing, which was first reported in Beijing and belongs to the C3a.1 subgroup. We should strengthen the monitoring and research of HPIVs mutations to enhance our response and early warning capabilities.