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Mucosal barriers are gatekeepers of health and exhibit homeostatic variation in relation to habitat and disease. Mucosal Mapping technology provides an in‐depth examination of the dynamic mucous cells (MCs) in fish mucosal barriers on tangential sections, about 90° from the view of traditional histology. The method was originally developed and standardized in academia prior to the establishment of QuantiDoc AS to apply mucosal mapping, now trademarked as Veribarr™ for the analysis of skin, gills and gastrointestinal tracts. Veribarr™ uses design‐based stereology for the selection and measurement of cell area (size) (μm2), the volumetric density of MCs in the epithelium (MCD, amount of the epithelia occupied by MCs, in %) and the calculated abundance of the MCs (barrier status or defence activity). MC production was mapped across the skin and gill epithelia in 12 species, discovering that gills consistently have two distinct groups of MCs, one on the lamellae where MCs are few and small and one on the filament where MCs are larger and more abundant. MCs were usually much larger in the skin than in the gills, with the latter requiring fewer and smaller cells for adequate respiration. The difference observed between MCs in gill lamella and gill filament is likely a result of functional demands. In addition, our findings also highlight a variation in the mucosal parameters between the species skin, which cannot be explained by the weight differences, and a potential link between MC distribution and species‐specific lifestyles in the gill lamella. This diversity necessitates the development of species and tissue site‐specific reference intervals for mucosal health evaluation. Mucosal bivariate reference intervals were developed for MC production, including size (trophy) and calculated defence activity (plasia) in the skin and gills of Atlantic salmon, to contrast new measurements against historical data patterns. The application of mucosal reference intervals demonstrates that stress from parasites and treatments can manifest as changes in mucosal architecture, as evidenced by MC hypertrophy and hyperplasia within the gill lamellae. These reference intervals also facilitate comparisons with wild Atlantic salmon, revealing a somewhat higher MC level in farmed salmon gill lamellae. These findings suggest that MC hyperplasia and hypertrophy in the gills are stress/environmental responses in aquaculture. They also advocate for developing specific mucosal bivariate homeostatic reference intervals in aquaculture to improve fish health and welfare across all farmed species. [ABSTRACT FROM AUTHOR]

ABSTRACT Myxobolus Bütschli, 1882 is a highly important genus of fish myxozoans, consisting of many species that cause serious diseases in ornamental cyprinid fish species in different geographic areas by infecting a wide range of organs. This study describes a new Myxobolus species infecting the gill lamellae, gill rakers and skin epidermis of goldfish (Carassius auratus) from Mohammadshahr, Iran, based on detailed morphological, histological and molecular analysis. Mature spores were small and spherical in shape, consisting of two symmetrical shell valves in frontal view, measuring 10.3 (9.8–10.9) μm in length and 10.5 (10.3–11.2) μm in width. Two equal‐sized relatively pyriform polar capsules were 5.4 (3.9–6.0) μm long and 4.5 (3.7–5.2) μm wide. The polar tubules were coiled with 4–6 turns. A comparative analysis of the morphological and molecular data confirmed that this species is distinct from other identified myxosporeans. Plasmodia in the cutaneous tissue were found in the epidermis, causing extensive hyperplasia of Malpighian cells and mild inflammatory infiltration. In the gill tissue, plasmodia affected only one side of the lamellae, leading to hyperplasia of the lamellar epithelium, edema, and lymphoplasmacytic branchitis. A novel myxozoan species, Myxobolus branchioepidermis n. sp., has been proposed. The study provides a comprehensive analysis of the morphological, histological, and molecular characteristics of this parasite, which infects the gill lamellae and epidermal layer of the skin in oranda goldfish (C. auratus). The identification of this new species adds to the growing body of research on myxozoan parasites and highlights its potential impact on ornamental fish aquaculture. [ABSTRACT FROM AUTHOR]

Amyloodinium ocellatum is among the most devastating protozoan parasites, causing huge economic losses in the mariculture industry. However, the pathogenesis of amyloodiniosis remains unknown, hindering the development of targeted anti‐parasitic drugs. The A. ocellatum in vitro model is an indispensable tool for investigating the pathogenic mechanism of amyloodiniosis at the cellular and molecular levels. The present work developed a new cell line, ALG, from the gill of yellowfin seabream (Acanthopagrus latus). The cell line was routinely cultured at 28°C in Dulbecco's modified Eagle medium (DMEM) supplemented with 15% fetal bovine serum (FBS). ALG cells were adherent and exhibited an epithelioid morphology; the cells were stably passed over 30 generations and successfully cryopreserved. The cell line derived from A. latus was identified based on partial sequence amplification and sequencing of cytochrome B (Cyt b). The ALG was seeded onto transwell inserts and found to be a platform for in vitro infection of A. ocellatum, with a 37.23 ± 5.75% infection rate. Furthermore, scanning electron microscopy (SEM) revealed that A. ocellatum parasitizes cell monolayers via rhizoids. A. ocellatum infection increased the expression of apoptosis and inflammation‐related genes, including caspase 3 (Casp 3), interleukin 1 (IL‐1), interleukin 10 (IL‐10), tumour necrosis factor‐alpha (TNF‐α), in vivo or in vitro. These results demonstrated that the in vitro gill cell monolayer successfully recapitulated in vivo A. latus host responses to A. ocellatum infection. The ALG cell line holds great promise as a valuable tool for investigating parasite–host interactions in vitro. [ABSTRACT FROM AUTHOR]

Nodular gill disease (NGD) is an infectious condition characterized by proliferative gill lesions leading to respiratory problems, oxygen deficiency and mortality in fish. Globally, NGD primarily impacts freshwater salmonids in intensive aquaculture systems. In recent years, numerous outbreaks of severe gill disease have affected more than half of the larger rainbow trout (Oncorhynchus mykiss) farms in Switzerland, mainly during spring and early summer. Mortality has reached up to 50% in cases where no treatment was administered. Freshwater amoeba are the presumed aetiologic agent of NGD. The gross gill score (GS) categorising severity of gill pathology is a valuable first‐line diagnostic tool aiding fish farmers in identifying and quantifying amoebic gill disease (AGD) in farmed marine salmonids. In this study, the GS was adapted to the NGD outbreak in farmed trout in Switzerland. In addition to scoring disease severity, gill swabs from NGD‐affected rainbow trout were sampled and amoeba were cultured from these swabs. Morphologic and molecular methods identified six amoeba strains: Cochliopodium sp., Naegleria sp., Vannella sp., Ripella sp., Saccamoeba sp. and Mycamoeba sp. However, the importance of the different amoeba species for the onset and progression of NGD still has to be evaluated. This paper presents the first description of NGD with associated amoeba infection in farmed rainbow trout in Switzerland. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate histologic post‐mortem autolytic changes in farmed Atlantic salmon. The fish were either stored at room temperature (RT, 21°C), refrigerated (4°C) or frozen (−20°C), while fish necropsy was performed at 0, 1, 4, 24 and 48 h post‐storage (hps). In addition, gills were sampled at 0, 5, 10, 15, 30 and 45 min post‐storage (mps) at room temperature (RT). The haematoxylin and eosin‐stained tissue slides were evaluated and scored by using a semi‐quantitative scoring system. Our findings demonstrated gills and pyloric caeca/pancreas as the most severely autolysed organs while heart and skeletal musculature were least affected. Generally, moderate to severe autolysis appeared first at 4 hps, while severe changes were seen at 24 hps. Gills demonstrated autolytic changes as early as 10 mps and pyloric caeca/pancreas at 1 hps. Freezing did not prevent the autolysis and even contributed to freezing artefacts, which may lead to misdiagnosis. Keeping organs refrigerated slowed the autolytic progress within the first 4 hps marginally. This study recommends gills and pyloric caeca/pancreas should be sampled as early as possible, at least within 10 min post‐necropsy. [ABSTRACT FROM AUTHOR]

Understanding disease aetiology and pathologic mechanisms is essential for fish health evaluation. Carp edema virus (CEV) is the causative agent of a disease (CEVD) responsible for high mortality rates in both wild and cultured common carp Cyprinus carpio. Inspection of two carp specimens from a pond with high fish mortality revealed CEV infection in both the host and its ectoparasite (Argulus foliaceus). In addition to flavobacteria, well known to be associated with gill lesions, we found that free‐living eukaryotes (amoebae and ciliates) and a temporary parasite (Ichthyobodo spp.) colonizing the gills may also contribute to alterations in gill structure and/or function, either directly, through firm (Ichthyobodo) or weak (amoebae) attachment of trophozoites to the gill epithelium, or indirectly, through carriage of pathogenic bacteria. Bacterial assemblages rich in families and genera, with predominance of Cetobacterium spp. in low‐intensity alteration of the gill tissue and of Flavobacterium spp. in gills with extensive necrotic lesions, were detected in gills and within the cytoplasm of associated amoebae using high‐throughput sequencing. Quantitative PCR indicated F. swingsii as the prevailing flavobacterial species within amoebae from less affected gills and F. psychrophilum within amoebae from extensively affected gills. This case study suggests that eukaryotic organisms as part of the gill pathobiome may also contribute to irreversible gill lesions seen in CEVD. Emphasizing the complexity of mutual relationships between bacterial assemblages and eukaryotic co‐pathogens, further studies regarding factors that trigger pathology and influence severity in the CEV‐positive carp are needed. [ABSTRACT FROM AUTHOR]

Gill disease is an important cause of economic losses, fish mortality and reduced animal welfare in salmonid farming. We performed a prospective cohort study, following groups of Atlantic salmon in Western Norway with repeated sampling and data collection from the hatchery phase and throughout the 1st year at sea. The objective was to determine if variation in pathogen prevalence and load, and zoo‐ and phytoplankton levels had an impact on gill health. Further to describe the temporal development of pathogen prevalence and load, and gill pathology, and how these relate to each other. Neoparamoeba perurans appeared to be the most important cause of gill pathology. No consistent covariation and no or weak associations between the extent of gill pathology and prevalence and load of SGPV, Ca. B. cysticola and D. lepeophtherii were observed. At sea, D. lepeophtherii and Ca. B. cysticola persistently infected all fish groups. Fish groups negative for SGPV at sea transfer were infected at sea and fish groups tested negative before again testing positive. This is suggestive of horizontal transmission of infection at sea and may indicate that previous SGPV infection does not protect against reinfection. Coinfections with three or more putative gill pathogens were found in all fish groups and appear to be the norm in sea‐farmed Atlantic salmon in Western Norway. [ABSTRACT FROM AUTHOR]

Recently, the culture of American eels (Anguilla rostrate) in China has been impacted by emergence of a disease with signs of haemorrhagic gill necrosis. The gills of diseased eels are covered with petecchia and they bleed when the operculum is pressed. In this study, a novel American eel adomavirus (AEAdoV) was isolated from the diseased eels using the eel ovary cell line (EO). The virus proliferated in the EO cells with a maximum TCID50/ml of 106.29 ± 0.23 at 6 days post‐infection. The virions were non‐enveloped with a diameter of 75–85 nm and shown to be a DNA virus upon 5‐iodo‐2'‐deoxyuridine (IDU) treatment. PCR assays showed that AEAdoV encodes a superfamily 3 helicases (S3H) replicase and shared high similarities with Anguilla marmorata adomavirus (MEAdoV). Although no clinical signs or mortality was observed among the eels injected with AEAdoV, the virus was reisolated from livers, kidneys and gills of injected eels at 35 days post‐injection. Our results suggested that AEAdoV exhibited a latent infection in A. rostrata. The pathogenicity of the AEAdoV needs to be confirmed further. [ABSTRACT FROM AUTHOR]

Amoebic gill disease (AGD) is caused by the marine amoeba Neoparamoeba perurans, a facultative parasite. Despite the significant impact this disease has on production of Atlantic salmon worldwide, the mechanisms involved in host–parasite interaction remains unknown. Excessive gill mucus secretion is reported as a host defence mechanism to prevent microbial colonization in the gill epithelium. Despite this response, N. perurans still attaches and proliferates. The present study aimed to investigate the interaction between N. perurans and mucin, the most abundant component in mucus. An in vitro adhesion assay using bovine submaxillary mucin (BSM) demonstrated that amoeba binding to mucin‐coated substrate was significantly higher than to the BSA control. This binding interaction is likely glycan‐mediated as pre‐incubation with galactose, galactosamine, N‐acetylgalactosamine and fucose reduced mucin adhesion to control levels. The ability of N. perurans to secrete proteases that target mucin was also investigated. Protease activity was detected in the amoeba culture media in the presence of BSM, but not when protease inhibitor was added. Mucin degradation was visually assessed on protein gels. This study provides preliminary evidence that N. perurans has developed mechanisms to interact with and evade mucus by binding to mucin glycan receptors and secreting proteases with mucolytic activity. [ABSTRACT FROM AUTHOR]

Cryptocaryonosis is the greatest threat to most teleost species among all parasitic diseases, causing mass loss to the marine aquaculture industry. Epidemiological investigation of teleost susceptibility to Cryptocaryon irritans infection revealed that yellow drum (Nibea albiflora) is highly resistant. In order to further understand the activation of the immune system in the gill, which is one of the main mucosal‐associated lymphoid tissues and a target of parasites, transcriptome analysis of the yellow drum gill was performed. Gill samples were collected from fish challenged after 24 hr and 72 hr with theronts at a median death rate (2050 theronts per gram fish). Gene expression profiles showed that TLR5 was the only receptor that activated the downstream immune response. The infection activated complement cascade through alternative pathway and increased the expression of C5a anaphylatoxin chemotactic receptor 1. In addition, possible antimicrobial molecules, including lipoprotein and haptoglobin, which are responsible for trypanolysis in humans, were among the top significantly upregulated genes at 24 hr. After 72 hr, the expression of secreted immunoglobulin T‐related genes was induced. These results suggested a rapid innate and adaptive immune response at the mucosal level. In conclusion, the results provide new perspectives on mucosal immune resistance in yellow drum against cryptocaryonosis and provide the possibility of mining resistance genes for future therapy. [ABSTRACT FROM AUTHOR]

During the conservation aquaculture of the freshwater mussel Margaritifera margaritifera, fish health has become a concern due to the need of mussel larvae (glochidia) to parasitize the salmonid gills and metamorphose into juveniles. However, there is a lack of information about the impact on fish during the juvenile detachment and the subsequent gill healing. To evaluate the morphopathological changes and gill recovery after the parasitism of M. margaritifera, 51 Atlantic salmon fry (Salmo salar), infested with around 22 larvae/fish g, were necropsied during the synchronized detachment of the mussel juveniles, and gills were assessed by stereomicroscopy and by light and scanning electron microscopy. Salmon showed no clinical signs during the trial and gills recovered their normal morphology almost completely in a short time, suggesting a minimal impact on fish health after glochidiosis. In this sense, the non‐erosive droplet detachment and the goblet cell hyperplasia favoured an effective gill remodelling mediated by apoptosis, polarization and cell shedding of the gill epithelia, providing insights to the defence, clearing and healing mechanisms of the gill. These morphopathological techniques could also be implemented to preserve fish welfare and to optimize the artificial breeding programmes of endangered freshwater mussels. [ABSTRACT FROM AUTHOR]

Amoebic gill disease (AGD) is a significant issue in Atlantic salmon mariculture. Research on the development of treatments or vaccines uses experimental challenges where salmon is exposed to amoebae concentrations ranging from 500 to 5,000/L. However, the water concentrations of N. perurans on affected salmon farms are much lower. The lowest concentration of N. perurans previously reported to cause AGD was 10/L. Here, we report that concentrations as low as 0.1/L of N. perurans can cause AGD. We propose that concentrations of N. perurans that reflect those measured on salmon farms should be used for future experimental challenges. [ABSTRACT FROM AUTHOR]

Paramoeba perurans causes amoebic gill disease (AGD), which is a major problem in aquaculture worldwide. The parasite can be cultured in vitro, but to this date, no method for long‐term storage of the clones exists. In this study, we describe a method for cryopreservation of Paramoeba perurans. The method was successfully employed on four out the five clones we tested. The thawing success rate, that is the percentage of successfully thawed vials relative to the total number of vials that were thawed, differed for the clones and ranged from 25% to 100%. The age of the clones seemed to have a negative impact on the ability to survive cryopreservation. [ABSTRACT FROM AUTHOR]

Takifugu rubripes and Dicentrarchus labrax are important commercial fish in China that are under serious threat from Cryptocaryon irritans. C. irritans is a ciliated obligate parasite that causes marine white spot disease and leads to heavy economic losses. We analysed the transcriptome in the gills of T. rubripes and D. labrax to compare differentially expressed genes (DEGs) and pathways during infection with C. irritans. In total, we identified 6,901 and 35,736 DEGs from T. rubripes and D. labrax, respectively. All DEGs were annotated into GO terms; 6,901 DEGs from T. rubripes were assigned into 991 sub‐categories, and 35,736 DEGs from D. labrax were assigned into 8,517 sub‐categories. We mapped DEGs to the KEGG database and obtained 153 and 350 KEGG signalling pathways from T. rubripes and D. labrax, respectively. Immune‐related categories included Toll‐like receptors, MAPK, lysosome, C‐type lectin receptor and NOD‐like receptor signalling pathways were significantly enriched pathways. In immune‐related signalling pathways, we found that AP‐1, P38, IL‐1β, HSP90 and PLA were significantly up‐regulated DEGs in T. rubripes, but P38 and PLA were significantly down‐regulated in D. labrax. In this study, transcriptome was used to analyse the difference between scaly and non‐scaly fish infection by C. irritans, which not only provided a theoretical basis for the infection mechanism of C. irritans, but also laid a foundation for effectively inhibiting the occurrence of this disease. Our work provides further insight into the immune response of host resistance to C. irritans. [ABSTRACT FROM AUTHOR]

Amoebic gill disease (AGD) is one of the main health issues impacting farmed Atlantic salmon. Neoparamoeba perurans causes AGD; however, a diversity of other amoeba species colonizes the gills and there is little understanding of whether they are commensal or potentially involved in different stages of gill disease development. Here, we conduct in vivo challenges of naïve Atlantic salmon with cultured Nolandella sp. and Pseudoparamoeba sp. to investigate their pathogenicity to Atlantic salmon gills. Additionally, we assessed whether the presence of Nolandella sp. and Pseudoparamoeba sp. influences the onset and/or severity of N. perurans‐induced AGD. All three strains attached and multiplied on the gills according to qPCR analysis. Furthermore, minor gross gill lesions and histological changes were observed post‐exposure. While N. perurans was found associated with classical AGD lesions, Nolandella sp. and Pseudoparamoeba sp. were not found associated with lesion sites and these lesions did not meet the expected composite of histopathological changes for AGD. Moreover, the presence of these non‐N. perurans species did not significantly increase the severity of AGD. This trial provides evidence that cultured Nolandella sp. and Pseudoparamoeba sp. do not induce AGD and do not influence the severity of AGD during the early stages of development. [ABSTRACT FROM AUTHOR]

Fish gills are heavily exposed to the external milieu and may react against irritants with different cellular responses. We describe variations in mucous cell counts in gills from healthy Atlantic salmon (Salmo salar) presmolts in five recirculating aquaculture system (RAS) farms and one flow‐through farm. Based on certain criteria, mucous cells were histologically quantified in a defined lamellar region of the gills and the counts were analysed. Immunohistochemistry (IHC) was used to investigate epithelial responses. The median number of total mucous cells in the defined region was 59 per fish. Between the farms, the medians varied from 31 to 101 with the lowest in the flow‐through farm. A regression model was fitted with "total mucous cells" as the dependent variable and with "fish length" and "fish farm" as independent variables. The proportion of variation in mucous cell counts explained by the model was twice as high when "fish farm" was included compared to only "fish length." IHC revealed proliferative responses in coherence with high mucous cell numbers. Conclusively, the variation in mucous cell counts depends on combined farm‐related factors. Establishing a baseline for mucous cell counts is fundamental in the development of high‐throughput monitoring programmes of gill health in farmed fish. [ABSTRACT FROM AUTHOR]

Routine gill swabbing is a non‐destructive sampling method used for the downstream qPCR detection and quantitation of the pathogen Neoparamoeba perurans, a causative agent of amoebic gill disease (AGD). Three commercially available swabs were compared aiming their application for timelier AGD diagnosis (Calgiswab® (calcium alginate fibre‐tipped), Isohelix® DNA buccal and cotton wool‐tipped). Calcium alginate is soluble in most sodium salts, which potentially allows the total recovery of biological material, hence a better extraction of target organisms' DNA. Thus, this study consisted of (a) an in vitro assessment involving spiking of the swabs with known amounts of amoebae and additional assessment of retrieval efficiency of amoebae from agar plates; (b) in vivo testing by swabbing of gill arches (second, third and fourth) of AGD‐infected fish. Both in vitro and in vivo experiments identified an enhanced amoeba retrieval with Calgiswab® and Isohelix® swabs in comparison with cotton swabs. Additionally, the third and fourth gill arches presented significantly higher amoebic loads compared to the second gill arch. Results suggest that limiting routine gill swabbing to one or two arches, instead of all, could likely lead to reduced stress‐related effects incurred by handling and sampling and a timelier diagnosis of AGD. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Gill parasitic infections challenge farming of rainbow trout (Oncorhynchus mykiss, Walbaum) in freshwater facilities. Apart from flagellates (Ichthyobodo, (Pinto) and ciliates (Ichthyophthirius (Fouquet), Ambiphrya (Raabe), Apiosoma (Blanchard), Trichodinella (Sramek‐Husek) and Trichodina (Ehrenberg)), we have shown that amoebae are prevalent in Danish trout farms. Gills were isolated from farmed rainbow trout in six fish farms (conventional and organic earth pond and recirculated systems) and placed on non‐nutrient agar (NNA) moistened with modified Neff's amoeba saline (AS) (15°C). Gill amoebae from all examined fish colonized the agar and were identified based on morphological criteria showing species within the genera Trinema (Dujardin) (family Trinematidae), Vannella (Bovee) (family Vannellidae). In addition, hartmannellid amoebae were recorded. We established a monoculture of Vannella sp., confirmed the genus identity by PCR and sequencing and performed an in vitro determination of antiparasitic effects (dose–response studies) of various compounds including sodium chloride (NaCl), hydrogen peroxide, peracetic acid, formalin, aqueous garlic and oregano extracts and a Pseudomonas H6 surfactant. All amoebae were killed in concentrations of 16.90 mg/ml (garlic), 17.90 mg/ml (oregano), NaCl (7.5 mg/ml), hydrogen peroxide (100 µg/ml), peracetic acid (0.03 µg/ml), formaldehyde (25 µg/ml) and the Pseudomonas H6 surfactant (250 µg/ml). [ABSTRACT FROM AUTHOR]

In June 2019, massive mortalities of cultured Penaeus vannamei occurred in a local farm in Hainan Province, China. The diseased shrimp displayed evident black gills. Three bacterial strains 20190611001, 20190611007 and 20190611022 were isolated from hepatopancreas and gills of the diseased shrimp and identified as Photobacterium damselae subsp. damselae based on the sequence analysis of 16S rRNA and toxR genes. These three isolates showed haemolytic activities. Of them, strain 20190611022 isolated from hepatopancreas was selected and processed for pathogenic analysis. The calculated median lethal dose (LD50) was 9.75 ± 4.29 × 105CFU/g (body weight) by challenging P. vannameivia reverse gavage. The diseased shrimp displayed enlarged hepatopancreatic tubules and sloughing of epithelial cells in tubular lumens. The strain 20190611022 was also characterized by the testing of API 20NE systems and antibiotic susceptibility. The results of disc diffusion test showed that strain 20190611022 was sensitive to chloramphenicol, compound sulfamethoxazole, cefoperazone, ceftriaxone, ceftazidime and cefuroxime. To our knowledge, this is the first report of isolation and characterization of Photobacterium damselae subsp. damselae from natural diseased P. vannamei. Our findings can serve as a basis for further studies of its pathogenicity and provide technological support for disease controlling in shrimp aquaculture. [ABSTRACT FROM AUTHOR]

Histopathological changes of Cichlidogyrus philander Douëllou, 1993 on the gills of Pseudocrenilabrus philander (Weber, 1897) were studied using light and scanning electron microscopy. Observations revealed that C. philander attaches to its host by alternating the prohaptor (for temporary attachment or feeding) or haptor (using haptoral parts for firm and secured attachment). The sharp terminal ends of the anchors are inserted basally into the gill lamella, between two adjacent secondary gill lamellae and the marginal hooklets assist by superficially penetrating, holding and lifting epithelial tissue in the proximal region of the secondary gill lamella. The attachment of C. philander resulted in compression, rupturing of the interlamellar epithelium, change in the organization of epithelial cells in both primary and secondary gill lamella, displacement of the extracellular cartilaginous matrix, occasional rupturing of blood vessels and erythrocytes and some cells becoming ill‐defined. At the site of attachment, the host response comprises of hyperplasia, increase in the number of mucous cells and infiltration with neutrophils. It was concluded that the effect of C. philander is mild in natural conditions. [ABSTRACT FROM AUTHOR]

Hydrogen peroxide (H2O2) is a commonly used treatment for a range of parasitic diseases of marine finfish, including amoebic gill disease (AGD). While this treatment is partially effective at reducing parasite load, H2O2 can have detrimental effects on the host under certain conditions. Treatment temperature and dose concentration are two factors that are known to influence the toxicity of H2O2; however, their impact on the outcome of AGD treatment remains unclear. Here, we investigated the effects of treatment temperature (8, 12 or 16°C) and dose concentration (750, 1,000, 1,250 mg/L) on the efficacy of H2O2 to treat AGD. We demonstrated that a 20‐min bath treatment of H2O2 at all doses reduced both parasite load and gross gill score significantly. Parasite load and gross gill score were lowest in the 1,000 mg/L treatment performed at 12°C. At the high dose and temperature combinations, H2O2 caused moderate gill damage and a significant increase in the plasma concentration of electrolytes (sodium, chloride and potassium). Taken together, our study demonstrates that higher H2O2 treatment temperatures can adversely affect the host and do not improve the effectiveness of the treatment. [ABSTRACT FROM AUTHOR]

A relationship between increasing water temperature and amoebic gill disease (AGD) prevalence in Atlantic salmon (Salmo salar) has been noted at fish farms in numerous countries. In Scotland (UK), temperatures above 12°C are considered to be an important risk factor for AGD outbreaks. Thus, the purpose of this study was to test for the presence of an association between temperature and variation in the severity of AGD in Atlantic salmon at 10 and 15°C. The results showed an association between temperature and variation in AGD severity in salmon from analysis of histopathology and Paramoeba perurans load, reflecting an earlier and stronger infection post‐amoebae exposure at the higher temperature. While no significant difference between the two temperature treatment groups was found in plasma cortisol levels, both glucose and lactate levels increased when gill pathology was evident at both temperatures. Expression analysis of immune‐ and stress‐related genes showed more modulation in gills than in head kidney, revealing an organ‐specific response and an interplay between temperature and infection. In conclusion, temperature may not only affect the host response, but perhaps also favour higher attachment/growth capacity of the amoebae as seen with the earlier and stronger P. perurans infection at 15°C. [ABSTRACT FROM AUTHOR]

Two aqueous fixation methods (modified Davidson's solution and modified Davidson's solution with 2% (w/v) Alcian blue) were compared against two non‐aqueous fixation methods (methacarn solution and methacarn solution with 2% (w/v) Alcian blue) along with the standard buffered formalin fixation method to (a) improve preservation of the mucous coat on Atlantic salmon, Salmo salar L., gills and (b) to examine the interaction between the amoebae and mucus on the gill during an infection with amoebic gill disease. Aqueous fixatives demonstrated excellent cytological preservation but failed to deliver the preservation of the mucus when compared to the non‐aqueous‐based fixatives; qualitative and semi‐quantitative analysis revealed a greater preservation of the gill mucus using the non‐aqueous methacarn solution. A combination of this fixation method and an Alcian blue/Periodic acid–Schiff staining was tested in gills of Atlantic salmon infected with amoebic gill disease; lectin labelling was also used to confirm the mucus preservation in the methacarn‐fixed tissue. Amoebae were observed closely associated with the mucus demonstrating that the techniques employed for preservation of the mucous coat can indeed avoid the loss of potential mucus‐embedded parasites, thus providing a better understanding of the relationship between the mucus and parasite. [ABSTRACT FROM AUTHOR]

Autophagy is primarily an adaptive response to provide nutrients and energy following exposure to stress and starvation but can also regulate muscle mass and impact infectious disease susceptibility. Expression of 10 autophagy‐related (Atg) genes in rainbow trout was monitored throughout the autophagosome formation cycle. The Atg gene sequences of rainbow trout were compared to other species to identify highly conserved regions and to generate primers. Phylogeny trees created with rainbow trout and 14 other species demonstrate that rainbow trout Atg gene sequences have greatest similarity to Atlantic salmon and other fish species. RTgill‐W1 cells were subjected to nutrient restriction and compared to cells in normal nutrient conditions using quantitative reverse transcriptase polymerase chain reaction to assess changes in Atg gene expression. Nutrient restriction had a direct impact on Atg gene expression, with atg4, atg9, atg12, lc3, gabarap and becn1 undergoing the greatest differential expression (p < 0.05), most dramatically on Day 3. This was corroborated by Western blot detection of LC3, which also showed a peak of autophagy activity at Day 3 post‐nutrient restriction. Atg gene expression revealed autophagy flux in RTgill‐W1, as well as, those genes that were most significantly altered by nutrient restriction. [ABSTRACT FROM AUTHOR]

The article discusses a research which described a novel salmon gill poxvirus (SGPV) variant among Atlantic salmon in Eastern Canada. It attempts to isolate and identify the variant. It discusses the results of disease testing, quantitative polymerase chain reaction and transmission electron microscopy.

Shortnose Sucker (Chasimistes brevirostris) and Lost River Sucker (Deltistes luxatus) are endemic to the Upper Klamath Basin of Southern Oregon and Northern California, and their populations are in decline. We used histopathology and external examination of 140 and external examination only of 310 underyearling suckers collected in 2013, 2015 and 2016 to document pathological changes, particularly those relating to parasites. The most severe infection was caused by a Contracaecum sp., infecting the atrium of 8%–33% of Shortnose Suckers. The most prevalent infections were caused by Bolbophorus sp. metacercariae in the muscle of Shortnose Suckers (21%–63%) and Lernaea cyprinacea in the skin and muscle of Lost River Suckers (30%–81%). Histology detected Bolbophorus in only 5% of cases where it was not seen externally. Three myxozoans were observed; a Parvicapsula sp. in the renal tubules (10%), a Myxobolus sp. in the intestinal mucosa (2%) and an unusual multicellular, presporogonic myxozoan in the intestinal lumen of one sucker. Severe gill epithelial hyperplasia was observed in several fish collected in 2016. Trichodinids and Ichthyobodo sp. were observed on some of the gills, but absent in many of the fish with severe lesions. A histiocytic sarcoma was observed in sucker. [ABSTRACT FROM AUTHOR]

Koi sleepy disease (KSD) is a disease with increasing importance in global common carp aquaculture. Despite the fact that carp edema virus (CEV) is most likely the causative agent of KSD, the disease often presents itself as multifactorial with several parasites and bacteria species present on gills, skin or in internal organs. Therefore, in this study, we analysed and presented initial results on an interaction of flavobacteria and CEV in the development of clinical KSD in carp suffering from proliferative gill disease. We examined selected field samples from Germany and Hungary and confirmed the presence of CEV and flavobacteria co‐infections in subset of the samples. In several infection experiments, we studied the transfer and dynamics of both infections. Furthermore, we analysed which Flavobacterium species could be isolated from KSD‐affected fish and concluded that Flavobacterium branchiophilum is a possible copathogen. Antibiotic treatment experiments showed that CEV seems to be the primary pathogen causing an insult to the gills of carp and by these enabling other pathogens, including F. branchiophilum, to establish co‐infections. Despite the fact that F. branchiophilum co‐infection is not required for the development of clinical KSD, it could contribute to the pathological changes recorded during the outbreaks. [ABSTRACT FROM AUTHOR]

Gill diseases are a complex and multifactorial challenge for marine farmed Atlantic salmon. Co-infections with putative pathogens are common on farms; however, there is a lack of knowledge in relation to the potential effect co-infections may have on pathology. The objective of this study was to determine the prevalence and potential effects of Neoparamoeba perurans, Desmozoon lepeophtherii, Candidatus Branchiomonas cysticola, Tenacibaculum maritimum and salmon gill poxvirus (SGPV) during a longitudinal study on a marine Atlantic salmon farm. Real-time PCR was used to determine the presence and sequential infection patterns of these pathogens on gill samples collected from stocking until harvest. A number of multilevel models were used to determine the effect of these putative pathogens on gill health (measured as gill histopathology score), while adjusting for the effect of water temperature and time since the last freshwater treatment. Results indicate that between 12 and 16 weeks post-seawater transfer (wpst), colonization of the gills by all pathogens had commenced and by week 16 of marine production each of the pathogens had been detected. D. lepeophtherii and Candidatus B. cysticola were by far the most prevalent of the potential pathogens detected during this study. Detections of T. maritimum were found to be significantly correlated with temperature showing distinct seasonality. Salmon gill poxvirus was found to be highly sporadic and detected in the first sampling point, suggesting a carryover from the freshwater stage of production. Finally, the model results indicated no clear effect between any of the pathogens. Additionally, the models showed that the only variable which had a consistent effect on the histology score was N. perurans. [ABSTRACT FROM AUTHOR]

Abstract: Jellyfish‐induced gill pathology relies upon occasional diagnostic observations yet the extent and impact of jellyfish blooms on aquaculture may be significant. Idiopathic gill lesions are often observed in apparently healthy fish. This study exposed Atlantic salmon (Salmo salar L.) smolts to macerated Cyanea capillata at 2.5 and 5 g/L for 2 hr under controlled laboratory conditions. Blood chemistry and gill histopathology were examined over a subsequent 4‐week period. Fish showed an acute response to the presence of jellyfish, including characteristic external “whiplash” discoloration of the skin and acute increases in blood electrolytes and CO2 concentration; however, these were resolved within 4 days after exposure. Histopathologically, gills showed first an acute oedema with epithelial separation followed by focal haemorrhage and thrombus formation, and then progressive inflammatory epithelial hyperplasia that progressively resolved over the 4 weeks post‐exposure. Results were consistent with the envenomation of gills with cytotoxic neurotoxins and haemolysins known to be produced by C. capillata. This study suggests that many focal hyperplastic lesions on gills, especially those involving focal thrombi, may be the result of jellyfish stings. Thus, the presence of jellyfish and their impact may be severe and understated in terms of marine fish aquaculture and fish welfare. [ABSTRACT FROM AUTHOR]

The article offers information on a study according to which designed primer pair and polymerase chain reaction (PCR) methods are effective in detecting Tenacibaculum dicentrarchi which is a fish pathogen responsible for high mortalities in farmed Atlantic salmon. Topics include use of Insta Gene Matrix for extracting Chromosomal DNA from bacteria; conduction of triplicate reactions in independent DNA extracts; and mentions that PCRs were performed on gill, kidney, skin and lesion samples.

Abstract: Chlamydial infections are recognised as causative agent of epitheliocystis, reported from over 90 fish species. In the present study, the farmed striped catfish Pangasianodon hypophthalmus (14–15 cm, 70–90 g) with a history of cumulative mortality of about 23% during June and July 2015, were brought to the laboratory. The histopathological examination of gills from the affected fish revealed presence of granular basophilic intracellular inclusions, mostly at the base of the interlamellar region and in gill filaments. A concurrent infection with Trichodina spp., Ichthyobodo spp. and Dactylogyrus spp. was observed in the gills. The presence of chlamydial DNA in the gills of affected fish was confirmed by amplification and sequencing of 16S rRNA gene. BLAST‐n analysis of these amplicons revealed maximum similarity (96%) with Candidatus Actinochlamydia clariae. On the basis of phylogenetic analysis, it was inferred that the epitheliocystis agents from striped catfish were novel and belonged to the taxon Ca. Actinochlamydia. It is proposed that epitheliocystis agents from striped catfish will be named as Ca. Actinochlamydia pangasiae. The 16S rRNA gene amplicons from novel chlamydiae were labelled and linked to inclusions by in situ hybridisation. This is the first report of epitheliocystis from India in a new fish host P. hypophthalmus. [ABSTRACT FROM AUTHOR]

In a parasitology survey of Hungarian fishes, heavy infections of parasitic copepods Lamproglena pulchella and a Lamproglena sp. were found in the gills of the asp and the European catfish, respectively. Individuals of both fish species were emaciated and infected with hundreds of Lamproglena. Copepods located close to the tip of gill filaments and formed a depression at the attachment sites. In histological sections, cell degenerations and local haemorrhages were present adjacent to the maxillipeds and where the maxillary claws pierced the gill tissue. Around maxillae and in the midgut of the Lamproglena, damaged piscine blood cells and remains of the gill tissue were observed. Host reaction was expressed by proliferation of epithelioid cells, increase in both number and size of goblet and mast cells and formation of giant cells. [ABSTRACT FROM AUTHOR]

Gill diseases cause considerable losses in Norwegian salmon farming. In 2015, we characterized salmon gill poxvirus ( SGPV) and associated gill disease. Using newly developed diagnostic tools, we show here that SGPV infection is more widely distributed than previously assumed. We present seven cases of complex gill disease in Atlantic salmon farmed in seawater and freshwater from different parts of Norway. Apoptosis, the hallmark of acute SGPV infection, was not easily observed in these cases, and qPCR analysis was critical for identification of the presence of SGPV. Several other agents including Costia-like parasites, gill amoebas, Saprolegnia spp. and bacteria were observed. The studied populations experienced significant mortalities, which increased to extreme levels when severe SGPV infections coincided with smoltification. SGPV infection appears to affect the smoltification process directly by affecting the gills and chloride cells in particular. SGPV may be considered a primary pathogen as it was often found prior to identification of complex gill disease. It is hypothesized that SGPV-induced gill damage may impair innate immunity and allow invasion of secondary invaders. The distinct possibility that SGPV has been widely overlooked as a primary pathogen calls for extended use of SGPV qPCR in Atlantic salmon gill health management. [ABSTRACT FROM AUTHOR]

Kynurenic acid ( KYNA) is an endogenous substance produced on the kynurenine pathway which is primarily known for its neuroactive properties. Recently, it has been proven that KYNA is a selective ligand for G protein-coupled receptor ( GPR 35), presented on immunocompetent cells such as T lymphocytes. This opens up new possibilities of its application as an immunostimulating substance in aquaculture. Thus far, no histopathological investigations in fish have been completed to evaluate influence of KYNA supplementation in feed. This study has been undertaken to determine the effect of feed supplementation with KYNA (2.5, 25, 250 mg kg−1 of feed) for 28 days on the liver, gills and kidney in healthy fish and experimentally infected with Yersinia ruckeri. In a control group were observed a fatty liver, which is natural for this fish species in the autumn and winter season. As the dose of the supplement was increased, the fat liver changed, it decreased or completely disappeared. Additionally, inflammatory changes occurred in all the analysed organs, and their intensification was dose dependent. In the fish experimentally infected, KYNA caused aggravation of the signs in the liver, kidneys and gills, and the effect was dose dependent. The results implicate that KYNA may be a stressor for fish. [ABSTRACT FROM AUTHOR]

The article presents a study which aims to determine the seasonal variation of prevalence and mean intensity of the Amyloodinium ocellatum in Solea senegalensis from Bizerte lagoon in Tunisia. Information regarding the measurement of water temperature and salinity, the presence of several subspherical brown trophonts at the infested gills under light microscopy, and the use of Analysis of Variance (ANOVA) to determine the seasonal variations, are mentioned.