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Current Trends in Biotechnology and PharmacyVol. 5 (1) 1011-1020 January 2011. ISSN 0973-8916 (Print), 2230-7303 (Online)1017intranuclear inclusions (Fig. 5a and 5b). TheWSSV infection was reported to have occurredfirst in the stomach, gill and cuticular epidermis,than subsequently got spread systematically toother tissues of mesodermal and ectodermal origin(21). The presence of dense basophilicintranuclear inclusion and chromatin marginationare reported to be characteristic of WSSVinfection (19,2). Cytopathological changes in theinfected gut and gills clearly suggest WSSVinfection in this shrimps. Our findings are inagreement with the cytopathological changes ofdifferent tissues of WSSV infected shrimpsreported by Chang et al. (10) and Yoganandhanet al. (22). In the infected muscles also, severenecrosis, chromatin margination and densebasophilic intranuclear inclusions (Fig. 5d) havebeen identified indicating the damage caused byviral infection. Histopathological analysis ofhepatopancreas of infected shrimp demonstratedsevere pyknosis and karyorrhexis in connectivetissue compartment (Fig. 5c). Prominent changeswere not seen in the tubular epithelial cells of thehepatopancreas. Our observation is in agreementwith Mohan and Shankar (23), who reported thatWSSV does not infect tissues of endodermal originsuch as hepatopancreatic tubular epithelium.Nuclear Pyknosis and karyorrhexis have beenreported to be associated with several viralinfections (21). Pantoja and Lightner (24)observed pyknosis and karyorrhexis in animalsexperimentally infected with WSSV. Presentlyobserved Pyknosis and karyorrhexis in connectivetissue cells of hepatopancreas and also in gutindicate the role of viruses in nuclear damage.The transmission electron microscopyobservations of pellet from the filtrate of diseasedP. monodon cuticular epidermis stained bynegative staining indicated the presence of rodshaped viral particles (Fig. 4). These are similarto the viral particles observed in negative stainingof spontaneously diseased shrimps reported byChou et al. (8). Electron microscopic observationconformed that the rod shaped virus wasconsidered to be the main causative agent in thepresent study. The virus is reported to be highlypathogenic and constitutes a threat to shrimpculture. It is also reported to be transmitted orallyor via water (8). Wongteerasupaya et al. (25)first described the TEM morphology of WSSVby negative staining. The presently observedmorphology of virus described here from SouthGujarat is very similar to the virus previouslyreported from other parts of India (14).In the present study, WSSV infected P.monodon shows differences in the expressionof proteins in muscle and hepatopancreas fromuninfected shrimps. In the infected muscle tissue,four new differently expressed proteins withmolecular weight 79kDa, 58kDa, 36kDa and17kDa have been detected. In the infectedhepatopancreas, three new proteins withmolecular weight 17kDa, 20kDa and 46kDa havebeen detected. A newly expressed protein withmolecular weight 17kDa has been detected inboth the tissues (Fig. 6a and 6b). The variation inthe pattern of expressions of proteins in tissuesof WSSV infected shrimp is possibly related tothe intensity of viral infection, degree ofpathogenicity and tissue specific disintegration(14). Van Hulten et al. (26) have reported thatWSSV virion protein ranges from 18kDa to28kDa. Presently observed expression of 17kDaprotein in infected muscle and hepatopancreasindicates the expression of viral protein andconfirms the presence of WSSV infection in boththe tissues. Expression of new protein, 17kDa ininfected muscle as well as in hepatopancreas isfound to be similar to the expression of newprotein from WSSV infected shrimps reportedby Sathish et al. (27).In terms of morphological observations,histopathological effects, viral morphology andexpression of new proteins in infected tissues,Protein expression in WSSV
Current Trends in Biotechnology and PharmacyVol. 5 (1) 1011-1020 January 2011. ISSN 0973-8916 (Print), 2230-7303 (Online)1018the virus described here from coastal farms ofGujarat is very similar to white spot syndromevirus. This is believed to be the first report ofsystematic study of identification of WSSV fromthe cultured shrimps of South Gujarat region.AcknowledgementDuring the present investigation, the secondauthor DC was supported by a meritoriousfellowship from UGC New Delhi, India. Wegratefully acknowledge the co-operation from theOffice of Assistant Director of Fisheries,Government of Gujarat, Anand, as well as fromthe fish farmer Shree Girishbhai and Shree SushilMahanti from Olapad, Surat District.References1. Wang, C.S., Tang, K.F.J., Kou, G.H. andChen, S.N. (1997). Light and electronmicroscopic evidence of white spot diseasein the giant tiger shrimp, Penaeus monodon(Fabricius), and the kuruma shrimp,Penaeus japonicus (Bate), cultured inTaiwan. J Fish Dis., 20: 323-331.2. Rajendran, K.V., Vijayan, K.K., Santiago,T.C. and Rajan, J.J.S. (2005). White spotsyndrome virus (WSSV) infection in tigershrimp Penaeus monodon: A non-lethalhistopathological rapid diagnostic methodusing paraffin and frozen sections. AquacultInt., 13: 341-349.3. Escobedo-Bonilla, C.M., Alday-Sanz, V.,Wille, M., Sorgeloos, P., Pensaert, M.B. andNauwynck, H.J. (2008). A review on themorphology, molecular characterization,morphogenesis and pathogenesis of whitespot syndrome virus. J Fish Dis., 31: 1-18.4. McColl, K.A., Slater, J., Jeyasekaran, G.,Hyatt, A.D. and Crane, M.S.T.J. (2004).Detection of White Spot Syndrome virus andYellow head virus in prawns imported intoAustralia. Aust. Vet. J., 82: 69-74.5. Ramasamy, P. (1995). Diseases of shrimpin aquaculture systems; Diagnosis andtherapeutic measures. Vanitha Publications,(Eds.) Madras, Chennai, India, pp. 1-99.6. Sahul Hameed, A.S., Xavier Charles, M.,Anilkumar, M. (2000). Tolerance ofMacrobrachium rosenbergii to white spotsyndrome virus. Aquaculture, 183: 207-213.7. Sahul Hameed, A.S., Yoganandhan, K.,Sathish, S., Murugan, V., Rasheed, M., andKunthala, J. (2001). Experimentalpathogenicity of white spot syndrome virus(WSSV) in two freshwater crabs(Partelphusa hydrodomous and P.pulvinata). Aquaculture, 201: 179-186.8. Chou, H.Y., Huang, C.Y., Wang, C.H.,Chiang, H.C. and Chu-Fang, Lo (1995).Pathogenicity of a baculovirus infectioncausing white spot syndrome in culturedpenaied shrimp in Taiwan. Dis Aquat Org.,23: 165-173.9. Lightner, D.V. (1996). A Handbook ofShrimp Pathology and DiagnosticProcedures for Diseases of CulturedPenaeid Shrimp. World Aquaculture Society,Baton Rouge, Lousiana, USA, p.304.10. Chang, P.S., Chu-Fang, Lo, Wang, Y.C. andKou, G.H. (1996). Identification of whitespot syndrome associated baculovirus(WSBV) target organs in shrimp, Penaeusmonodon, by in situ hybridization. DisAquat Org., 27: 131-139.11. Nadala, Jr. E.C.B., and Loh, P.C. (2000).Dot-blot nitrocellulose enzyme immunoassayfor the detection of white-spot virus andyellow head virus of penaeid shrimp. J VirolMethods, 84: 175-179.Bhatt et al
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