Viral infections, often neglected, have been suggested to play an important role in the decline of the panmictic population of the European eel (Anguilla anguilla). Despite the importance of knowledge about pathogenic eel viruses, little is known about their spread in the wild population of this species and only a few pathogenic viruses have been described so far. This knowledge is necessary for the eel broodstock selection from natural populations. Neretva Delta, Croatia, presents an ideal place for possible broodstock selection, in which commercial eel fishing is a tradition that has continued to this day, despite the decline in catches. The aim of this study was to investigate the presence of two most often reported causes of viral diseases in the A. anguilla, anguillid herpesvirus 1 (AngHV-1) and eel virus European X (EVEX).
Asymptomatic eels were collected from the commercial fisherman catch during winter and spring 2021. Three different organs: liver, spleen and kidney were collected and stored in 70% ethanol. A total of 32 samples were used for detection of AngHV-1 by PCR. After chopping individual organs with sterile scissors their total DNAs were extracted using GenElute Mammalian Genomic DNA Miniprep Kit (Sigma), following the manufacturer`s instructions with the overnight lysis step at 55°C. Subsequent nested PCR reactions for amplification of partial DNA polimerase gene of AngHV-1 were performed using EmmeraldAmpMaxHS PCR Master Mix with DNA volumes, primers and cycling conditions. Presence of PCR products from both reactions (509 bp and 312 bp) was checked in 1.7% agarose gels.
To examine the presence of RNA virus EVEX, parts of the L and M genes were amplified using RT-PCR. Isolation of total RNA was performed using TRI reagent (Invitrogen). RT-PCR reactions were performed in one step using the Transcriptor One-Step RT-PCR Kit (Roche) according to the manufacturer’s instructions. The presence of amplified products was checked on a 1.5% agarose gel by gel electrophoresis.
A total of 14 out of 32 fish were positive on AngHV-1 (Fig. 1), while all samples were negative on EVEX (Figure 1.).
These results highlight the importance of broodstock health assessment and biosecurity measures, including quarantine, for eel hatcheries in order to assure higher survival and better quality of larvae.