Protocol Of A Ranavirus

In 2006, epizootic haematopoietic necrosis disease (EHN) was included on list of exotic fish diseases in European Union (EU) legislation. The criteria for listing a disease as exotic are that disease i) is not established in Community and pathogen is not known to be present and ii) it has potential for either significant economic impact if introduced or detrimental environmental impact on wild aquatic animal species (Anonymous, 2006). EHN is caused by epizootic haematopoietic necrosis virus (EHNV), which has previously induced high mortalities in redfin perch (Perca fluviatilis) and morbidity in rainbow trout (Oncorhynchus mykiss) in Australia ([Langdon et al., 1986] and [Langdon et al., 1988]). These two species are listed as susceptible to EHN in current legislation (Anonymous, 2006), and surveillance programmes are in place, targeting EHNV infection in these species.

. However, other important European fish species can be susceptible to EHNV, as has been demonstrated for black bullhead (Ameiurus melas) and pike (Esox lucius) ([Bang Jensen et al., 2009] and [Gobbo et al., 2010]).

EHNV is a member of genus Ranavirus (family Iridoviridae), which contains viral isolates obtained from fish, amphibians and reptiles from all over world. EHNV has not been isolated in EU. A ranavirus isolate (pike-perch iridovirus, PPIV) has also been isolated from apparently healthy farmed pike-perch (Sander lucioperca) (Tapiovaara et al., 1998), another (Rmax) from clinically healthy turbot fry (Psetta maxima) and another (CodV) from free-living cod (Gadus morhua) with an ulcus-syndrome ([Jensen et al., 1979] and [Ariel et al., 2010]). Short-finned eel virus (SERV) was found in non-symptomatic short-finned eels (Anguilla australis) imported to Italy from New Zealand (Bang Jensen et al., 2009).

Previous challenge studies have shown that ECV, ESV, PPIV and SERV can be pathogenic to European stocks of pike, and that ECV and frog virus 3(FV3), type species of genus, can infect pike without causing significant mortality (Bang Jensen et al., 2009). Even though PPIV was isolated from pike-perch, we have no knowledge of pathogenicity of ranaviruses to this species. The aim of study presented here was to supplement knowledge on host range of ranavirus by determining susceptibility of pike-perch to different ranavirus isolates, and to confirm its susceptibility to PPIV under experimental settings. Another aim was to investigate whether those isolates .

In challenge III, five fish were injected intraperitoneally (i.p.) with 100 µL of viral suspension, and cohabitated with five naïve fish from same batch for each of four viral isolates at 16 °C (± 2 °C). The number of fish in challenge was limited to five because of limited resources and justified by precedence of Langdon (1989) who used small numbers of fish to successfully demonstrate transmission of EHNV between infected and cohabitated fish. To distinguish between exposed and naïve fish, latter had caudal dorsal fin clipped. The four viral isolates were (final titre in brackets): EHNV (4 × 106 TCID50/mL), ESV (5.9 × 106 TCID50/mL), PPIV (4 × 107 TCID50/mL) and ECV-24 (4.8 × 106 TCID50/mL). In negative control treatment, five fish were injected with phosphate buffered saline (PBS) and cohabitated with five naïve fish for control. The fish were observed and sampled daily as described for bath challenges. After 36 days any surviving fish were euthanized, and fish from each tank were pooled and frozen for subsequent virus re-isolation.

From challenge I, up to seven samples of pooled ...