Cranial Mandibular Fibrosis Syndrome in Adult Farmed Rainbow Trout Oncorhynchus mykiss.

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.

A Seasonal Study of Koi Herpesvirus and Koi Sleepy Disease Outbreaks in the United Kingdom in 2018 Using a Pond-Side Test.

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 Skin Mucosal Parameters during Koi Herpesvirus Shedding in Common Carp, Cyprinus carpio.

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.

Non-lethal loop-mediated isothermal amplification assay as a point-of-care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease.

Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop-mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pemaquidensis and N. branchiphila, or the host DNA. This assay detected 106 copies of the parasite 18S rRNA gene under 13 min and 103 copies under 35 min. Five "fast-and-dirty" DNA extraction methods were compared with a reference method and further validated by TaqMan™ qPCR. Of those, the QuickExtract buffer was selected for field tests. Seventy-one non-lethal gill swabs were analysed from AGD-clinically infected Atlantic salmon. The pathogen was detected under 23 min in fish of gill score >2 and under 39 min for lower gill scores. About 1.6% of the tests were invalid (no amplification of the internal control). 100% of positives were obtained from swabs taken from fish showing gill score ˃3, but only ~50% of positives for lower gill scores. The present LAMP assay could be implemented as a point-of-care test for the on-site identification of N. perurans; however, further work is required to improve its performance for lower scores.

Isolation of a Chinook Salmon Bafinivirus (CSBV) in Imported Goldfish Carassius auratus L. in the United Kingdom and Evaluation of Its Virulence in Resident Fish Species.

This is the first record of a fish nidovirus isolated from a consignment of goldfish at the United Kingdom (UK) border. The full-length viral genome was 25,985 nt, sharing a 97.9% nucleotide identity with the Chinook salmon bafinivirus (CSBV) NIDO with two deletions of 537 and 480 nt on the ORF Ia protein. To assess the potential impact on UK fish species, Atlantic salmon, common carp and goldfish were exposed to the virus via an intraperitoneal (IP) injection and bath challenge. Moribundity was recorded in only 8% of IP-injected goldfish. A high viral load, ≈107 of the CSBV PpIa gene, was measured in the kidney of moribund goldfish. Mild histopathological changes were observed in the kidneys of challenged carps. Ultrastructural observations in renal tubule epithelial cells of goldfish showed cylindrical tubes (≈15 nm in diameter) and tubular structures budding spherical virions (≈200 nm in diameter) with external spike-like structures. Negative staining showed both circular and bacilliform virions. Seroconversion was measured in common carp and goldfish but not in Atlantic salmon. This study reinforces the potential risk of novel and emerging pathogens being introduced to recipient countries via the international ornamental fish trade and the importance of regular full health screens at the border inspection posts to reduce this risk.

Surface water mitigates the anti-metamorphic effects of elevated perchlorate concentrations in New Mexico spadefoot toad larvae (Spea multiplicata).

Perchlorate (ClO4-) has potential to negatively impact amphibian populations by inhibiting thyroid hormone production, and thus metamorphosis in developing larvae. However, variability exists in species sensitivity, and there is evidence suggesting that natural surface waters can mitigate the anti-metamorphic potential of perchlorate. New Mexico spadefoot toad tadpoles, Spea multiplicata, were exposed to natural surface waters spiked with nominal concentrations of 0, 1000, 1350, 1710, 3000, 5110, and 8000 µg/L perchlorate ion for up to 42 days. No consistent dose-response trends were observed in mortality, rate of metamorphosis, Gosner stage, mass, or length. This study suggests that perchlorate exposure to concentrations as high as 8000 µg/L in natural surface waters does not result in adverse effects on New Mexico spadefoot tadpoles and emphasizes the importance of using site-specific conditions and species when evaluating ecological risks in perchlorate-impacted areas.

Surface water mitigates the anti-metamorphic effects of perchlorate in New Mexico spadefoot toads (Spea multiplicata) and African clawed frogs (Xenopus laevis).

Spea multiplicata (New Mexico spadefoot toad) larvae were exposed to 60, 110, and 1000 microg L(-1) perchlorate dissolved in natural surface water to determine risks associated with perchlorate exposure in desert-adapted anurans. Hind- and forelimb development and tail resorption were measured to identify effects of perchlorate exposure. No perchlorate-related effects on snout-vent length, hindlimb length, and proportion metamorphosed were observed in the highest treatment group (positive control; 1000 microg L(-1)) suggesting that either S.multiplicata are not sensitive to the effects of perchlorate at the concentrations tested or that unidentified constituents of natural surface water mitigated perchlorate toxicity. To identify whether surface water mitigated perchlorate toxicity, Xenopuslaevis were exposed to 20 and 60 microg L(-1) perchlorate in surface water and synthetic laboratory prepared water (i.e., FETAX media). X.laevis exposed to perchlorate dissolved in surface water exhibited no perchlorate-related anti-metamorphic effects, whereas X.laevis exposed to perchlorate in FETAX media experienced changes in percent metamorphosing (p<0.001), time to metamorphosis (p<0.001), snout-vent length (p<0.001), and hindlimb length (p<0.001) as compared to FETAX controls. These results suggest that natural surface water can mediate perchlorate effects at concentrations up to 60 microg L(-1) for X.laevis and greater than 1 mg L(-1) for S.multiplicata, potentially due to physicochemical properties of surface water. CAPSULE: This manuscript discusses the effects of perchlorate in natural surface water to S.multiplicata and X.laevis.