HD-3 Herbal Medicine Against WSSV Infection Andimmune Enzyme Activity Effect On Shrimp Litopenaeus Vannamei

With the deep study in the White Spot Syndrome Virus(WSSV) by dome-stic and foreign researchers, there has been achieved fruitful research results at the molecular level of pathogenic genes, spread of infection and immune protection on prawns, but the effects of prevention and cure of WSS are still not desirable. The present study was designed to investigate the WSSV carrier rate in the farming prawns, to prevent WSSV with Chinese medicinal herbs screen-ed based on cell culture, to determine the optimal days of treatment based on shrimp mortality and immune activity combined with absolute fluorescence qu a-ntitative PCR, histopathological detection, and superfine pathology detection, and to test the efficacy of the production in the pilot test.1. In July 2014--In December 2015,samples of 29 batches were collected in Yangjiabo, Binzhou, Weifang and Jiaonan etc-the major northern vannamei farming areas located in Liaoning, Hebei, Tianjin, and Shandong. Samples from the same farm is defined as a batch and the rate of WSSV carrier as well as breeding technology were investigated. Vannamei WSSV positive rate was determined by nested PCR after extracting the DNAs. Recombinant plasmids for absolute quantitation standards were constructed. The results show that in the North vannamei main producing areas WSSV positive rate is as high as 72.4% by nested PCR and 86.2% by quantitative fluorescence PCR. The detection se-nsitivity of quantitative PCR is better than that of nested PCR. L.vannamei in- traditional pond farming were more sensitive in intensity and quantity than industrial and greenhouse aquaculture.2. L.vannamei were infected with WSSV after feeding anti-WSSV herbs for different days. Then the symptoms and mortality were studied and four immune factors PO, SOD, AKP and ACP were measured.The results showed that: adding Chinese medical herbs 1d、4d、7d、10d、13d、16d and 19 d to the fee d can protect the prawns. At the end of the experimentsthere were survived pr awnsin the groups which were fed with herbs for 13 d, 16 d and 19 d and the immune protective rate of the group of 16 d is the highest, which is 36.7±3.3%. As the infection was aggravated by the virus, the symptoms of prawnswer e consistent with white spot syndrome. 10 d after taking the herbs, the activities PO, ACP and AKP were significantly improved(p <0.05). The activities of PO, SOD and ACP of prawns were broadly in line with the prawns reaction an d mortality. the activities of PO, AKP and ACP(P<0.05) were changed signifi cantly after toxicityattacktest, but that of SOD had a smaller change.3. It was shown that the content of viruses in experiment groups were sig-nificantly lower than positive controls by quantitating the virus in vannamei cheek tissues. And after infection 72 h,the virus contents after feeding Chinese herbs 10 d groups were was lower than positive controls(P< 0.05),only in104 copies/mg. The content of virus in survived shrimps were in low levelabout 103 copies/mg overall.Studying the severity of infection(SOI)by histopathology fo und that the severity was directly proportional to the degree of infection WSS V, which could show the infection progress of prawns. The SOI of 10d-19 d be fore 120 h were significantly lower than that of the other groups, showed good protection effects. In the histopathological analysis of Advanced Micro Device s, the number of cells infected with WSSV and complete WSSV virus within t he gills, stomach and crustaceans epidermis in positive control group were sign ificantly more than the survived prawns. In the experimental groups, the viruse s were mostly shown as un-fully assembled envelopes or nucleocapsid. It is su ggested that herbs could block WSSV infection.4. Test the herbs effect in WSSV-positive farms in the pilot test. The test period was 28 days. There was no outbreak of white spot syndrome of prawn s during the entire breeding cycle. It was shown that in experimental groups, the anti-WSSV herbs significantly improved the rate of weight gain and survival rate(P<0.05). Adding herbs can improve the immune activity to some exten t especially in the high temperature period as PO and ACP were significantly higher. It was revealed that Chinese herbs could reduce viruses slightly in infected prawns through absolute fluorescence quantitative PCR.