| The shrimp farming industry has been a pillar industry in many countries. High density, continuous deterioration of the ecological environment and physical decline of the shrimp in the culturing process have led to widespread disease to the shrimp farming. Viruses that infect shrimp have the characteristics of rapid onset, fast spread and causing high mortality. Viral diseases that cause huge economic losses to the shrimp farmers have hindered the development of the shrimp farming industry. Shrimp White Spot Syndrom Virus(WSSV) and Infectious Hypodermal and Hematopoietic Necrosis Virus(IHHNV) are the two of the most important DNA viruses in shrimp. They have caused the outbreak of epidemics worldwide, and become the main topics of the shrimp viral disease researches in late years. Due to the lack of specific medicine and the vulnerable immune system in shrimp, the effective prevention and treatment for IHHNV and WSSV is difficult.Thus, early prevention and breeding disease-resistant shrimp strain are significant for the shrimp industry.The traditional PCR method was used in the experiment to detect the samples from Nanjing, Nantong, Yancheng, Nanchang and Dafeng. We amplified the complete genome of Sheyang strain for genetic analysis of the evolutionary tree. The detection of494shrimps for epidemiological investigation of IHHNV showed that the average positive rate was27.94%(138/494). The shrimps that were collected from Sheyang Newport farm (62/62) and Dafeng farm (12/12) were all positive. The positive rate of Nantong and Xiamen were also higher than average rate were37.5%(12/32) and38.64%(17/44) respectively. High detection rates of this investigation appeared in the months from June to August, which were69.44%(25/36),65%(26/40) and53.57%(15/28). Since a certain degree of physical redness and small size existed in shrimp from Sheyang farm, We amplified the whole genome of Sheyang strain and built the phylogenetic tree. The results showed that the strain of Sheyang had high homology with the strains of Vietnam, Korea and Africa yet the phylogenetic trees that based on ORF1and ORF3were different. The result of positive rate in epidemiological investigation of WSSV was36.84%(182/494). Shrimps that were collected from Sheyang Newport farm (62/62) and Dafeng farm (12/12) showed a positive rate of100%which means t hat IHHNV and WSSV had co-infection. The positive rates of other locations were Nantong71.88%(23/32), Xiamen34.09%(15/44) and Weigang market22.70%(69/304). The positive rate of crayfish Procambarus was only2.5%(1/40) col lected from Nanchang. Higher detection rates of this investigation appeared in t he month July, August and October, which were73.53%(25/34),64.58%(31/48),43.75%(7/16) respectively. In this experiment, we amplified the variation s equence of Weigang strain and Sheyang strain and build the phylogenetic tree. The results showed that Sheyang, Taiwan isolate had high homology, while the highest homology Weigang strain has with Sheyang strain was only44.8%.As a novel detection method real-time PCR has good specificity, high sensitivity and can fix the quantity.Compared to real-time PCR the traditional PCR is only for qualitative analysis and the sensitivity is100-1000times lower than quantitative PCR. In this experiment we amplified a size of189bp of the conserved region of the IHHNV genome.The detection limit of real-time PCR was1.2×102plasmid copies per μL. Compared with conventional PCR it has a very high sensitivity. The melting curve showed one specific peak and no amplification of unrelated DNA samples. This method also has good reproducibility. Thus a new detection method was established for IHHNV epidemiological investigation. |
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