title: A new RNA virus found in black tiger shrimp Penaeus monodon from Thailand.
abstract: A new, apparently innocuous virus was found while investigating the cause of monodon slow growth syndrome (MSGS) in cultured black tiger shrimp (Penaeus monodon). It was identified via plasmid vector clones of E. coli containing randomly amplified cDNA fragments produced from total nucleic acid extracts of hemolymph from MSGS shrimp. Of 421 clones, 30 that failed to give positive dot blot hybridization with a digoxigenin (DIG)-labeled shrimp DNA probe were sequenced and compared to GenBank records. Of these, 22 corresponded to known shrimp DNA records. Of eight that did not, one (20A) showed significant deduced amino acid sequence similarity to RNA-dependent RNA polymerases (RdRp) of the viruses in the family Luteoviridae and alignment revealed commonly conserved amino acids including a GDD motif believed to be at the enzyme active site. However, phylogenetic analysis showed that the virus sequence did not cluster with the Luteoviridae or other known RNA virus sequences. Thus, in accordance with frequent practice, it was named according to the area where it was first collected as Laem-Singh virus (LSNV). In situ hybridization with a DIG-labeled 20A insert revealed strong cytoplasmic staining confined to the lymphoid organ (LO), the heart and hepatopancreatic connective tissue in both normal and MSGS shrimp. RT-PCR assays based on the 20A clone sequence also gave positive results with both normal and MSGS shrimp. Transmission electron microscopy (TEM) of LO tissue revealed viral-like particles of approximately 27 nm diameter (within the Luteoviridae size range) in locations that matched those of positive in situ hybridization reactions in parallel samples. Although not directly associated with MSGS in Penaeus monodon, the presence or effect of this virus with other crustacean species is presently unknown.
authors.affiliation: Centex shrimp, Faculty of Science, Mahidol University and National Center for Genetic Engineering and Biotechnology, BIOTEC, National Science and Technology Development Agency, NSTDA, Klong Luang, Pathumthani 12120, Thailand.
authors.forename: Kallaya
authors.initials: K
authors.lastname: Sritunyalucksana
authors.forename: Somjai
authors.initials: S
authors.lastname: Apisawetakan
authors.forename: Anutara
authors.initials: A
authors.lastname: Boon-Nat
authors.forename: Boonsirm
authors.initials: B
authors.lastname: Withyachumnarnkul
authors.forename: Timothy W
authors.initials: TW
authors.lastname: Flegel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015723:Gene Library; D006458:Hemolymph / Q000821:virology; D017403:In Situ Hybridization; D017793:Luteovirus / Q000235:genetics; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D033561:Penaeidae / Q000821:virology*; D010802:Phylogeny; D012328:RNA Viruses / Q000145:classification* / Q000302:isolation & purification; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013785:Thailand
chemical_list: D012324:RNA-Dependent RNA Polymerase