title: Non-virulence of a recombinant shrimp nidovirus is associated with its non structural gene sequence and not a large structural gene deletion.
abstract: RT-PCR using a commercial kit for yellow head virus (YHV) detection in growth-retarded shrimp yielded an unusual 777 bp amplicon instead of expected amplicons of 277 bp for YHV type-1 (YHV-1) or 406 bp for YHV type-2 (YHV-2). Cloning and sequencing (GenBank EU170438) revealed approximately 80% identity to non-structural (NS) ORF1b sequences of both YHV-1 (GenBank AA083987) and YHV-2 (GenBank AF227196), indicating an atypical YHV type (A-YHV) phylogenetically equidistant from both types. An RT-PCR test specifically designed for A-YHV revealed that it was uncommon and that its occurrence in shrimp culture ponds did not correlate with growth retardation or mortality. By immunohistochemistry with YHV-specific monoclonal antibodies, the A-YHV gave positive reactions for envelope protein gp64 and capsid protein p20, but not for envelope protein gp116, even though gp116 and gp64 originate from a polyprotein of ORF3. Lack of gp116 immunoreactivity correlated with a large ORF3 deletion (GenBank EU123854) in the region of the protein targeted by an MAb against gp116. Transmission electron microscopy of A-YHV-infected shrimp revealed only unenveloped pre-virions. During manuscript revision, information received revealed that typing of YHV isolates based on sequences of ORF1b and ORF3 had yielded several geographical types, including one virulent type (YHV-1b) with an ORF3 deletion sequence that matched the sequence of A-YHV. Using these sequences and an additional A-YHV sequence (EU853170) from the ORF1b typing region, A-YHV potentially represents a recombinant between type 1b and type 5. SDS-PAGE and Western blot analysis revealed that type 1b produced a gp116 deletion protein that did not bind with the MAb or polyclonal Ab to normal gp116. Overall, the information suggested that lack of A-YHV virulence was associated with the NS gene sequence linked to ORF1b rather than the deletion in ORF3.
authors.affiliation: National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Thailand Science Park, Klong Luang Pratumthani, Thailand.
authors.forename: Warachin
authors.initials: W
authors.lastname: Gangnonngiw
authors.forename: Gun
authors.initials: G
authors.lastname: Anantasomboon
authors.forename: Wiwat
authors.initials: W
authors.lastname: Sang-oum
authors.forename: Siriporn
authors.initials: S
authors.lastname: Sriurairatana
authors.forename: Kallaya
authors.initials: K
authors.lastname: Sritunyalucksana
authors.forename: Timothy W
authors.initials: TW
authors.lastname: Flegel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017353:Gene Deletion; D008969:Molecular Sequence Data; D028381:Nidovirales / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D033561:Penaeidae / Q000276:immunology / Q000648:ultrastructure / Q000821:virology*; D016415:Sequence Alignment; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics*; D015678:Viral Structural Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D017361:Viral Nonstructural Proteins; D015678:Viral Structural Proteins