ASSESSING GENETIC RELATEDNESS OF INFECTIOUS HYPODERMAL AND HAEMATOPOIETIC NECROSIS VIRUS (IHHNV) ISOLATES RECENTLY DETECTED IN THE U.S.

Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) is an OIE-listed viral pathogen of penaeid shrimp and is prevalent  in the Americas, Asia and Australia.  The virus  has caused large-scale mortalities in blue shrimp (Penaeus stylirostris ) and is known to cause growth retardation in Pacific white shrimp (P. vannamei) and black tiger (P. monodon ) shrimp. IHHNV is a  non-enveloped,  single-stranded DNA containing virus belonging to the family Parvoviridae .  During June of 2019,  P. vannamei  samples (broodstock and post-larvae originating in shrimp facilities in Texas and Florida) were submitted to Aquaculture Pathology Laboratory for routine screening of a panel of  OIE-listed and non-listed viral, bacterial and fungal pathogens by two commercial facilities in the US.  DNA and RNA were  isolated using a Maxwell 16 cell LEV DNA Purification Kit (Promega, Wisconsin) and a Maxwell 16 cell LEV RNA Purification Kit (Promega, Wisconsin), respectively .  The screening of samples were done by polymerase chain reaction (PCR) and  following OIE-recommended protocols for the OIE-listed pathogens. Fo r the OIE non-listed pathogens, PCR was performed following published protocols for the corresponding pathogens.  For both commercial operations, samples were tested negative for all the pathogens screened except for the IHHNV.  IHHNV PCR screening was done using the primer sets, 389F/R and 309F/R that amplifies  two different regions in the non-structural gene, NS1 in the IHHNV genome.  Both amplicons  (309 bp and 389 bp)  were  sequenced and NCBI database search showed 99-100% similarity to IHHNV isolates from Asia and Latin America. Subsequently,  IHHNV capsid protein (CP) gene (990 bp ) was amplified by PCR and sequenced. The CP gene of the US isolates showed >99 % similarity to  Type II  infectious forms of  IHHNV isolates from Ecuador. Phylogenetic anal yses using Neighbor-Joining and Maximum Likelihood methods showed that the IHHNV isolates from the US clustered with homologous isolates from Ecuador . T he detection of an OIE-listed pathogen in commercial  facilities in the US highlights the need to follow biosecurity in the hatcheries and grow-out ponds to mitigate economic losses.