Problematic transfer of viruses amongst penaeid shrimp Center of Excellence for Shrimp Molecular Biology and Biotechnology National Center for Genetic Engineering and Biotechnology Tim Flegel Centex Shrimp, Chalermprakiat Building, Fac. Science Mahidol University, Rama 6 Road, Bangkok 10400, Thailand

Overall viewpoint  Live aquatic animals present far the highest risk for viral disease transfer  All well studied cases of shrimp viral disease transfer have occurred by this route  Most cases have involved shrimp for aquaculture  No proven case of transfer by importation and use of fresh frozen shrimp for human consumption  Disease imported with crustaceans in ballast and on hulls has not been properly assessed

Intentional import of live shrimp  All major cases of shrimp viral transfer have occurred by importation of brooders and PL  This includes importation of exotic species and resident species from other countries  Three well documented cases: –Infectious hypodermal and hematopoeitic necrosis virus (IHHNV) from Asia to the Americas –White spot syndrome virus (WSSV) initially from China to Japan but then to Asia and the Americas –Taura syndrome virus (TSV) initially around the Americas but then to Taiwan and Asia

Most exotic introductions intentional Source: FAO website

Importations by group Source: FAO website

Shrimp viruses a particular danger  Viruses alone are responsible for the most severe losses in shrimp aquaculture  We know that shrimp react to viruses in a way different from vertebrates (from fish to man)  Shrimp and other crustaceans are characterized by persistent viral infections  These infections often produce no gross signs of disease and no mortality  Many of these “ hidden ” or “ cryptic ” viruses are still unknown

Dual and multiple viral infections  Dual, triple and multiple viral infections are often seen in shrimp, but rarely reported  We rarely examine “ healthy ” shrimp  This applies also to other crustaceans and arthropods such as insects  Viruses with the same name may be quite different genetically (e.g., HPV variants)  A cryptic virus tolerated in one host or location may not be tolerated in another host or location

Crustaceans a particular threat Grossly healthy shrimp A study of 6 ponds in Thailand (Flegel et al. 2001)

Triple viral infection

Example of dangerous exchanges YHV WSSV IHHNV MOV? LOVV? TSV? BP? Reo-like virus? P. monodonP. vannamei

LOVV-like virus in P. monodon  Dr. Lightner found this in LO of P. monodon brooders from the Andaman Sea in late 2002 Virions 25 nm

Test with P. monodon  Aj. Boonsirm took slow growing P. monodon  Homogenized LO and filtered out bacteria  Injected into SPF P. monodon and cultured with un-injected control shrimp  Result after 2 months –Un-injected shrimp ………. 8 g –LO injected shrimp ………. 4 g  LO samples sent for microscopy

Light microscopy of LO

Electron microscopy of LO Virions 25 nm

Probe urgently needed  For a non-destructive detection method to screen broodstock and PL  To identify possible carriers  We now have a tentative probe that gives positive results with test shrimp and negative with control shrimp  Its sensitivity, specificity and applicability are being tested

Risks of viral transfer

Important lessons  Crustaceans can carry unknown viral pathogens as innocuous, active infections  Many carriers (including PL) are grossly normal, so PCR needed for detection  These viruses may be deadly to other species or the same species at distant locations  Greater geographical separation = greater danger  By far, the most common route of transfer is with live broodstock or PL for aquaculture  No proven transfer from processing plants or from shrimp for human consumption

Conclusions  Careless movement of live stocks has been the cause of most shrimp disease transfers  Live introductions from ballast water and hull fouling may also be a source of disease  The evidence in hand suggests that frozen shrimp for human consumption does not pose a significant transfer risk  Any risks from frozen shrimp can be decreased substantially by simple risk reduction measures  PCR is recommended for screening live shrimp but is inappropriate for frozen shrimp