Objective:To study the astragalus polysaccharides ( APS) effect on immune induced by influenza A virus HA2 eu-karyotic expression vector.Methods: The HA2 encoded by the DNA vaccine vector was efficiently expressed in CHO cells, as determined by reverse transcription polymerase chain reaction ( RT-PCR) and fluorescence analysis.60 rats were divided into six groups randomly,which were immunized with normal saline,pEGFP-N1,pHA2/EGFP+different dose of APS by intramuscular injection.The control sera were collected before injection.After injected the 36th day, sera were collected to analyzing IFN-γ, IL-4 and IgG level.Results:IFN-γ,IL-4 and IgG level of pHA2/EGFP+mAPS group was different from that of pEGFP-N1 group or pHA2/EGFP+lAPS group( P<0.05 ).Conclusion: Middle dose of APS could increase immune induced by influenza A virus HA2 eukaryotic expression vector.

Objective:To study whether high dose astragalus polysaccharides( APS) could affect the expression of pNS1/EGFP that included influenza A virus(IAV) non-structure protein 1(NS1) gene in the tissue.Methods:pNS1/EGFP was constructured with NS1 of IAV.Sixty Kunming mice were divided into three groups randomly.Each group of mice was injected separately with one of the following:pEGFP-N1, pNS1/EGFP and pNS1/EGFP+APS in intraperitoneal injection.The mice were injected by intramuscular injection twice with a 3-week interval between injections.The serum samples and muscle samples were obtained on day 14 and day 28 after the booster injection.Sera IL-4,sera IFN-γ,muscle caspase-3 and muscle NS1 expression were measured in ELISA,Western blot or RT-PCR.Results:There were no significant difference among the different groups in day 14 expect that IFN-γof pNS1/EGFP+APS were lower(P<0.05).IFN-γlevel or IL-4 level of pNS1/EGFP+APS were lower compared with other groups in day 28.caspase-3 of pNS1/EGFP+APS were lower compared with other groups in day 28.Conclusion:APS could increase the expression of pNS1/EGFP by decreasing the inflammation and apoptosis.

A new variant of concern for SARS-CoV-2,Omicron(B.1.1.529),was designated by the World Health Organization on November 26,2021.This study analyzed the viral genome sequenc-ing data of 108 samples collected from patients infected with Omicron.First,we found that the enrichment efficiency of viral nucleic acids was reduced due to mutations in the region where the primers anneal to.Second,the Omicron variant possesses an excessive number of mutations compared to other variants circulating at the same time(median:62 vs.45),especially in the Spike gene.Mutations in the Spike gene confer alterations in 32 amino acid residues,more than those observed in other SARS-CoV-2 variants.Moreover,a large number of nonsynonymous mutations occur in the codons for the amino acid residues located on the surface of the Spike protein,which could potentially affect the replication,infectivity,and antigenicity of SARS-CoV-2.Third,there are 53 mutations between the Omicron variant and its closest sequences available in public databases.Many of these mutations were rarely observed in public databases and had a low muta-tion rate.In addition,the linkage disequilibrium between these mutations was low,with a limited number of mutations concurrently observed in the same genome,suggesting that the Omicron vari-ant would be in a different evolutionary branch from the currently prevalent variants.To improve our ability to detect and track the source of new variants rapidly,it is imperative to further strengthen genomic surveillance and data sharing globally in a timely manner.

COVID-19 has swept globally and Pakistan is no exception.To investigate the initial introductions and transmissions of the SARS-CoV-2 in Pakistan,we performed the largest genomic epidemiology study of COVID-19 in Pakistan and generated 150 complete SARS-CoV-2 genome sequences from samples collected from March 16 to June 1,2020.We identified a total of 347 mutated positions,31 of which were over-represented in Pakistan.Meanwhile,we found over 1000 intra-host single-nucleotide variants(iSNVs).Several of them occurred concurrently,indicating possible interactions among them or coevolution.Some of the high-frequency iSNVs in Pakistan were not observed in the global population,suggesting strong purifying selections.The genomic epidemiology revealed five distinctive spreading clusters.The largest cluster consisted of 74 viruses which were derived from different geographic locations of Pakistan and formed a deep hierarchical structure,indicating an extensive and persistent nation-wide transmission of the virus that was probably attributed to a signature mutation(G8371T in ORF 1ab)of this cluster.Further-more,28 putative international introductions were identified,several of which are consistent with the epidemiological investigations.In all,this study has inferred the possible pathways of introduc-tions and transmissions of SARS-CoV-2 in Pakistan,which could aid ongoing and future viral surveillance and COVID-19 control.