The Phylogenic Analysis Of H7and H10Subtype Avian Influenza Viruses And The Biological Characteristics Of Duck Tembusu Viruses

Waterfowl is the natural host of various kinds of viruses. Influenza viruses and duck Tembusuviruses(DTMUV) could not only infect waterfowls but also have the possibility to infect other hostscausing new infectious disease. In this research, the genetics and evolution relationships of2H7N3AIVs and3H10subtype AIVs (H10N8, H10N3, H10N7) and14DTMUVs were analyzed. Moreoverthe pathogenicity of DTMUVs was tested in mouse and duck.The whole genomes of A/duck/Zhejiang/690/2009(H7N3)(ZJ690) and A/duck/Zhejiang/766/2009(H7N3)(ZJ766) were sequenced and their genetic characteristic and phylogenetic relationshipswere analyzed. The amino acid residues in the cleavage site of the HA of both viruses were:PETPKGR↓GLFG, a typical feature of low pathogenic avian influenza virus. The amino acid627Eand701D on PB2of both viruses indicate the viruses were difficult to replicate in mammalian cells.The amino acids of277H on NA, and26L,27V,30A,31S on M2on the isolates suggested thesensitivity of these viruses with neuraminidase inhibitors and amantadine. Phylogenetic analysisrevealed that the HA and NA genes of these viruses were closely related to A/duck/Zhejiang/12/2011(H7N3) and A/duck/Jiangsu/10-d4/2011(H11N3), respectively. The PB2, PB1, PA, M, NS genesof these two isolates were closely related to the corresponding genes of H4N2, H11N3, H11N3,H10N8and H10N8, respectively. NP gene of isolate ZJ690was closely related to H11N3,while whichof ZJ766was closely related to H10N8.The whole genomes of A/duck/Shanghai/602/2009(H10N8)(SH602), A/duck/Fujian/1761/2010(H10N3)(FJ1761) and A/duck/Shanxi/3180/2010(H10N7)(SX3180) were sequenced and their geneticcharacteristic and phylogenetic relationships were analyzed.The sequence of HA genes indicated thesethree isolates were low pathogenic in avians. The sequence of PB2genes of these three strains showedthe features of typical avian influenza virus. The sequence of NA and M2indicate the sensitivity ofthese viruses with neuraminidase inhibitors and amantadine. Phylogenetic analysis revealed that theHA genes of SH602and SX3180were closely related to A/wild/bird/Korea/A12/2010(H10N1),while which of FJ1761was closely related to A/mallard/Sweden/65/2002(H10N9). The NA genes ofSH602, FJ1761, SX3180were closely related to A/duck/Thailand/CU-7518C/2010(H3N8),A/duck/Zhejiang/11/2011(H7N3) and A/swan/Slovenia/53/2009(H7N7), respectively. Evolutionaryanalysis showed that the PB2, PB1, PA, NP, M and NS genes of SH602strain were closely related toto the corresponding genes of H12N1, H4N6, H4N6, H6N5, H4N8and H4N2, respectively; the PB2,PB1, PA, NP, M and NS genes of FJ1761strain were closely related to to the corresponding genes ofH4N2, H7N3, H4N2, H10N8, H7N3and H7N7, respectively; the PB2, PB1, PA, NP, M and NS genesof SX3180strain were closely related to to the corresponding genes of H10N1, H10N1, H4N6, H4N6,H6N5and H3N6, respectively.The whole genomes of14duck Tembusu virus isolates were sequenced. Phylogenetic analysisrevealed that14DTMUV strains were divided into5groups. In the evolutionary tree, SD201147,SD201151and SH201001located in one branch; HB201001and HB201002located in one branch; HB201214and HB201217located in one branch; ZJ2011B2, JS201101and JS201102lied in onebranch; HB201101, HB201102and FX2010lied together in one branch; the SD201120liedindependently in another branch. To test the pathogenicity of DTMUV in ducks, FX2010、ZJ2011B2、JS201101、SD201120、SD201147and HB201214were selected to infect4-6weeks ducks intranasally.Real-time PCR was used to test the viral quantitation of duck’s organs. The results showed that all theisolates tested replicated systemically in ducks. To explore the different pathogenicity of DTMUV inmammals, BALB/c mice were inoculated intramuscularly and intranasally, and their spleens, lungs,kidneys, brains and nasal turbinates were colleted to test viral replication by the method of real-timePCR and virus isolation. The results showed that DTMUVs could not replicate in mice inoculatedintramuscularly. However, most viruses could replicate well in nasal turbinates and lungs of miceinoculated intranasally. And interestingly, we found strain HB201214could invade into and replicatein the brains of the mice inoculated intranasally. The results suggested that DTMUVs had a trend togain the infectivities in mammals.