Complete Genome Sequence Analysis And PCR-Based Detection Methods Of Guangxi Muscovy Duck Parvovirus And Duck Hepatitis A Virus Type 3

The muscovy duck parvovirus disease is a poisonous infection caused by the muscovy duck parvovirus(MDPV),which is characterized by high morbidity within 3 weeks old muscovy ducklings. The clinical symptom are enteritis, diarrhea and pedal flaccidity, it has caused heavy losses in muscovy duck dustry. At present, this disease is widely popular in Fujian, Guangdong, Guangxi and other major muscovy duck breeding area.Duck virus A hepatitis (DVAH) is a poisonous infection disease caused by duck hepatitis A virus (DHAV), which is characterized by high morbidity and mortality within 3 weeks old ducklings. According to the serum type, DHAV can be divided into DHAV-1 type, DHAV-2 type and DHAV-3 type, corresponding to the original type DHAV-1, the type isolated in Taiwan (DHAV-2) and the isolated in South Korea (DHAV-3). As the duck industry prosperity, the incidence of DHAV is rising. DVAH has caused heavy losses in duck industry. For the past few years, DVAH caused by DHAV-1 and DHAV-3 are widely popular in China.In order to understand the genetic regularity of MDPV and DHAV-3, we got the whole genome of GX2011-5 and GXLZ07 strain and analyzed them. Sequence analysis showed that GX2011-5 strain had the highest similarity with Shanghai SAAS-SHNH strain and the lowest similarity with Hungary FM strain, and the nucleotide similarity with other MDPV strains published in GeneBank is 90.5% to 94.3%. While compared with genome sequences with GPV strains, GX2011-5 strain had the highest similarity with Chongqin DY strain and the lowest similarity with Hungary Virulent B strain, and the nucleotide similarity was 81.4% to 90.4%.GXLZ07 strain had the highest similarity with Fujian G strain and the lowest similarity with Beijing B63 strain, and the nucleotide homology with other DHAV-3 strains published in GeneBank was 95.1% to 99.3%.In order to develop a duplex two-temperature RT-PCR assay for detecting of DHAV-1 and DHAV-3 simultaneous, we designed two pairs of specific primers according to the sequences of 3C gene of DHAV-A and VPO gene of DHAV-C available in GeneBank. It has good specificity, other duck pathogens can not be amplified using this method. And the detection limit was 4.9pg RNA in DHAV-1 and 7.5pg RNA in DHAV-3. The assay was successfully used to detect 53 suspected clinical samples, the positive rate was 5.6%for DHAV-1, 3.7% for DHAV-3.In order to develop a multiplex PCR assay for detecting of DHAV-1? DHAV-3 and MDPV simultaneous, we designed three sets of specific primers according to the sequences of 3C gene of DHAV-1, VPO gene of DHAV-3 and NS gene available in GeneBank. It has good specificity, other duck pathogens can not be amplified using this method. And the detection limit was 5.2pg RNA in DHAV-1,7.9pg RNA in DHAV-3 and 5.8pg DNA in MDPV. The assay was successfully used to detect 53 suspected clinical samples, the positive rate was 5.6%for DHAV-1,3.7% for DHAV-3 and 9.4%for MDPV.