Fluorescence real-time LAMP assays were designed for the orf43 gene of CyHV-3 European genotype and the p4a gene of the CEV genogroup I. A third LAMP assay to detect the ef1a gene of the host common carp was designed as an internal control. The limit of detection was 10² and 10³ viral copies under 25 min for CyHV-3 and CEV, respectively. The specificity of the CyHV-3 LAMP assay was 95.6% of 72 fish herpesviruses tested. Sixty-three non-lethal common carp mucus swabs were collected across 16 sites during disease investigations. DNA extractions were performed in under 10 min using the QuickExtract™ digestion buffer. The LAMP amplification of CyHV-3 DNA in mucus swabs from clinical cases was detected from 4 to 13 min in 13 sites, while a co-infection of CyHV-3 and CEV was confirmed by LAMP in a single site. The LAMP results agreed with the results of the reference laboratory. The common carp ef1a was amplified only in 61% of the mucus swabs collected, preventing its use as a robust internal control to distinguish false negatives from invalid tests. After further optimization, these tests could be implemented for border inspection posts surveillance and decentralizing testing during disease outbreaks.

Seroconversion and the mucosal lysozyme G (lysG), complement 3 (c3), and immunoglobulins M (IgMsec) and Z2 (IgZ2) were measured for up to 900 degree days (DD) in skin swabs from common carp exposed to koi herpesvirus (KHV or CyHV-3) at either a non-permissive temperature (12 °C) or permissive temperatures (17 and 22 °C), and in survivors subjected to temperature increase to 22 °C 500 DD after the initial exposure. The survival rate at 22 °C varied from 100% in fish initially exposed at 12 °C, to 20% at 17 °C and 0% at 22 °C. Viral shedding episodes lasted for up to 29 days (493 DD) for fish clinically infected at 17 °C, and up to 57 days (684 DD) for asymptomatic fish held at 12 °C. Up-regulation of lysG transcripts was measured at 17 and 22 °C. Down-regulation of c3 and IgMsec transcripts was measured independent of the water temperature, followed by up-regulation after the temperature increase coinciding with seroconversion and clearance of KHV from the skin mucus. IgZ2 mRNA showed a negative correlation with IgM transcripts. KHV subversion of the complement system at the mucosal site coupled with poor immunoglobulin secretion during the viral replication might contribute to the long window of viral shedding, thus facilitating viral transmission.

An unusual condition affecting market size rainbow trout was investigated. This condition was prevalent for several years at low levels but affected a large proportion of stock during 2018 and 2019. Chronic fibrosis affecting cranial tissues and the jaw was observed in samples collected in 2018. A larger sampling was then conducted in 2019 to investigate the presence of an infectious agent(s). An extensive inflammatory response in the mandibular region was the main finding, however infectious agents in the lesions were not identified through classical virology and bacteriology analysis. Tetracapsuloides bryosalmonae infection, calcinosis, and a Gram-positive bacterial infection of a single fish cardiac tissue was observed, however, a correlation of these pathologies and the cranial mandibular fibrosis (CMF) syndrome was not established. The gene expression of a panel of 16 immune-related genes was studied. Among these, tgf-b, sIgM, il11, hspa, and the antimicrobial peptides lys and cath1 were up-regulated in jaw sections of CMF-affected fish, showing a strong positive correlation with the severity of the lesions. Idiopathic chronic fibrosis with the activation of the Tfg-B pathway and local hyper-immunoglobulaemia was therefore diagnosed. Initiating factors and causative agent(s) (biotic or abiotic) of CMF remain, at present, unclear.