DETECTION AND GENOME RECONSTRUCTION OF TAURA SYNDROME VIRUS OF SHRIMP FROM DAVIDSON’S FIXED PARAFFIN EMBEDDED SHRIMP TISSUE

Taura Syndrome (TS), caused by Taura syndrome virus (TSV),  is an OIE-listed disease that  has caused more than $2 billion losses to shrimp  aquaculture  worldwide  since its emergence in 1992. TS is now prevalent in the Americas and Asia. Aquaculture Pathology Laboratory has an archive of Davidson's fixed paraffin embedded (DFPE) shrimp tissues blocks spanning across decades that  represent a priceless biobank of information, and  provide  a durable and cost-effective  ways of stor ing tissues . The objective of this study was to determine the utility of archived blocks as  biological samples for pathogen detection and genetic studies, using TSV as a model.  DFPE tissues present serious challenge in isolating high quality RNA due to the chemical modification,  RNA hydrolysis by acetic acid,  and cross linking of nucleic acids and proteins.

TSV infected tissue sections (~ 4 µm) (N=29 ) with Grades 3 and 4 levels of infection (o n a  Scale of 0 to 4)  were taken from DFPE blocks from 2005 . Total RNA was isolated using three commercially available kits to determine their suitability to yield high quality RNA ( 29 total x 3 kits = 87 samples).  Upon some modifications, one of the three kits provided good quality RNA (average concentration ~250 ng/µl,  average 260/280 value of 1.91) amenable to PCR amplification . TSV was  successfully  detected by conventional (231 bp ) and real-time (72 bp) RT-PCR in all 29 samples . Sanger sequencing of representative a mplicons  confirmed TSV identity . Using Next Generation Sequencing, the complete genome  of TSV  was reconstructed for three RNA samples representing N=87 .

This  is the first  study  that demonstrates the utility of  archived DFPE histological blocks  for shrimp pathogen d etection using TSV as model.  This shows both health assessment and targeted pathogen screening in shrimp can be done using histological samples . The feasibility to reconstruct viral genome from archived DFPE blocks opens avenues for  the discovery of  novel pathogens in shrimp.  This  will  also enhance our understandin g of the evolution of shrimp pathogens that has a direct implication in disease management in shrimp aquaculture.