Successive Losses of Central Immune Genes Characterize the Gadiformes' Alternate Immunity.

Great genetic variability among teleost immunomes, with gene losses and expansions of central adaptive and innate components, has been discovered through genome sequencing over the last few years. Here, we demonstrate that the innate Myxovirus resistance gene (Mx) is lost from the ancestor of Gadiformes and the closely related Stylephorus chordatus, thus predating the loss of Major Histocompatibility Complex class II (MHCII) in Gadiformes. Although the functional implication of Mx loss is still unknown, we demonstrate that this loss is one of several ancient events appearing in successive order throughout the evolution of teleost immunity. In particular, we find that the loss of Toll-like receptor 5 predates the loss of Mx involving the entire Paracanthopterygii lineage. Using a time-calibrated phylogeny, we show that loss of MHCII and Mx overlap with major paleoclimatic and geological events indicating that these genetic changes were adaptive responses to the changing environment at the time.

Evidence for and characterization of nervous necrosis virus infection in Pacific cod (Gadus macrocephalus).

A mortality rate higher than 90% was observed in a larva-rearing facility for Pacific cod, Gadus macrocephalus, in China. Larvae showing clinical signs of infection were collected. Initial suspicion of nervous necrosis virus (NNV) infection was confirmed by sequencing, absolute quantification real-time PCR (A-qPCR), and electron microscopy. The nucleotide sequence of RNA2 was 1,375 bases long (GenBank no. KM576685), coding for a single ORF corresponding to the capsid protein from residues 21 to 1034. Phylogenetic analysis of the capsid protein sequence showed that PCNNV belongs to the barfin flounder NNV (BFNVV) genotype. An amino acid sequence alignment revealed 39 differences between the cold- and warm-resistant viral groups, suggesting that PCNNV evolved under temperature selection. The 3-D structure of the predicted capsid protein was modeled to identify potential epitopes, and the gene was expressed in Escherichia coli, yielding a protein with a molecular mass of 55 kDa. During PCNNV outbreaks, the viral copy number was found to reach 10(7) per ng of total RNA, which could be considered the lethal copy number of NNV in cod. The gonads, eggs, fertilized eggs and asymptomatic cod fry were all positive for PCNNV, indicating viral vertical transmission as the main source of the viral load. The amount of virus in the apparent healthy fry or survivors seemed to decrease gradually with development. These results might lead to efficient diagnostic methods to help farmers select NNV-free broodfish for cod breeding.

Characterization of susceptibility and carrier status of burbot, Lota lota (L.), to IHNV, IPNV, Flavobacterium psychrophilum, Aeromonas salmonicida and Renibacterium salmoninarum.

In this study, susceptibility and potential carrier status of burbot, Lota lota, were assessed for five important fish pathogens. Burbot demonstrated susceptibility and elevated mortality following challenge with infectious haematopoietic necrosis virus (IHNV) by immersion and to Aeromonas salmonicida by intraperitoneal (i.p.) injection. IHNV persisted in fish for at least 28 days, whereas A. salmonicida was not re-isolated beyond 17 days post-challenge. In contrast, burbot appeared refractory to Flavobacterium psychrophilum following intramuscular (i.m.) injection and to infectious pancreatic necrosis virus (IPNV) by immersion. However, i.p injection of IPNV resulted in re-isolation of virus from fish for the duration of the 28 day challenge. Renibacterium salmoninarum appeared to induce an asymptomatic carrier state in burbot following i.p. injection, but overt manifestation of disease was not apparent. Viable bacteria persisted in fish for at least 41 days, and bacterial DNA isolated by diagnostic polymerase chain reaction was detected from burbot kidney tissue 90 days after initial exposure. This study is the first to investigate susceptibility of burbot to selected fish pathogens, and this information will aid in efforts to culture and manage this species.

Monthly occurrence and prevalence of viral haemorrhagic septicaemia virus (VHSV) in whiting Merlangius merlangus.

A field survey was carried out to determine the occurrence and spread of viral haemorrhagic septicaemia virus (VHSV) in whiting Merlangius merlangus (L.), a key prey species in the southeastern Black Sea region. In March, April, September and December 2007 and monthly in 2008, 70 to 105 M. merlangus captured by gill nets were examined in pools of 7 fish per pool (total number of fish = 1603). After determining the seasonal occurrence of VHSV in whiting, 43, 80 and 88 whiting were sampled individually in January, February and March 2009, respectively, to determine age and size of whiting commonly infected with VHSV. The samples were tested in duplicate using BF-2 and EPC cell lines for virus isolation. Positive cultures (those demonstrating cytopathic effect, [CPE]), were tested further with a commercial ELISA kit to confirm that the observed CPE was caused by VHSV. VHSV was detected in M. merlangus only between February and April following a drop in the surface water temperature to 9 degrees C. All infected fish from the sampled whiting were asymptomatic carriers with low levels of virus. The estimated prevalence from pooled samples ranged between 1.35% (2.5 to 97.5 percentiles; 0.04 to 7.32%) to 3.14% (0.36 to 10.95%), whereas prevalence estimated from individual sampling was 7, 5 and 1% in January, February and March, respectively. Individual sampling also revealed that prevalence was higher among young fish (1 to 1.5 yr old) and decreased with increasing age. This is the first report of VHSV infection in whiting in the Black Sea.