The nervous necrosis virus (NNV; genus Betanodavirus) is an aquatic pathogen that is responsible for a neurological disease affecting marine fish. Despite its almost worldwide distribution, global warming could favor the spread of NNV to new areas, highlighting the importance of conducting epidemiological surveys on both wild and farmed marine fish species. In this study, we assessed NNV prevalence in wild fish caught along the Galician Atlantic coast.
In total, 1277 fish were analyzed by reverse transcription real-time polymerase chain reaction.
Twenty two (1.72%) of those fish tested positive for NNV, including two species in which the pathogen had not yet been reported.
The reassortant RGNNV/SJNNV (red-spotted grouper NNV/striped jack NNV) was detected in 55% of NNV-positive individuals, while the remaining 45% harbored the SJNNV-type genome. Moreover, from European Pilchard Sardina pilchardus and Atlantic Mackerel Scomber scombrus, we isolated four reassortant strains that carried amino acid mutations at key sites related to NNV–host interaction.
The first objective of the study aimed to detect the presence of Lactococcus petauri, L. garvieae, and L. formosensis in fish (n = 359) and environmental (n = 161) samples from four lakes near an affected fish farm in California during an outbreak in 2020. The second objective was to compare the virulence of the Lactococcus spp. in Rainbow Trout Oncorhynchus mykiss and Largemouth Bass Micropterus salmoides.
Standard bacterial culture methods were used to isolate Lactococcus spp. from brain and posterior kidney of sampled fish from the four lakes. Quantitative PCR (qPCR) was utilized to detect Lactococcus spp. DNA in fish tissues and environmental samples from the four lakes. Laboratory controlled challenges were conducted by injecting fish intracoelomically with representative isolates of L. petauri (n = 17), L. garvieae (n = 2), or L. formosensis (n = 4), and monitored for 14 days postchallenge (dpc).
Lactococcus garvieae was isolated from the brains of two Largemouth Bass in one of the lakes. Lactococcus spp. were detected in 14 fish (8 Bluegills Lepomis macrochirus and 6 Largemouth Bass) from 3 out of the 4 lakes using a qPCR assay. Of the collected environmental samples, all 4 lakes tested positive for Lactococcus spp. in the soil samples, while 2 of the 4 lakes tested positive in the water samples through qPCR. Challenged Largemouth Bass did not show any signs of infection postinjection throughout the challenge period. Rainbow Trout infected with L. petauri showed clinical signs within 3 dpc and presented a significantly higher cumulative mortality (62.4%; p < 0.0001) at 14 dpc when compared to L. garvieae (0%) and L. formosensis (7.5%) treatments.
The study suggests that qPCR can be used for environmental DNA monitoring of Lactococcus spp. and demonstrates virulence diversity between the etiological agents of piscine lactococcosis.
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