Involvement of cholinergic and adenosinergic systems on the branchial immune response of experimentally infected silver catfish with Streptococcus agalactiae

It has been recognized that the cholinergic and adenosinergic systems have an essential role in immune and inflammatory responses during bacterial fish pathogens, such as the enzymes acetylcholinesterase (AChE) and adenosine deaminase (ADA), which are responsible for catalysis of the anti‐inflammatory molecules acetylcholine (ACh) and adenosine (Ado) respectively. Thus, the aim of this study was to investigate the involvement of the cholinergic and adenosinergic systems on the immune response and inflammatory process in gills of experimentally infected Rhamdia quelen with Streptococcus agalactiae. Acetylcholinesterase activity decreased, while ACh levels increased in gills of infected animals compared to uninfected animals. On the other hand, a significant increase in ADA activity with a concomitant decrease in Ado levels was observed in infected animals compared to uninfected animals. Based on this evidence, we concluded that infection by S. agalactiae in silver catfish alters the cholinergic and adenosinergic systems, suggesting the involvement of AChE and ADA activities on immune and inflammatory responses, regulating the ACh and Ado levels. In summary, the downregulation of AChE activity exerts an anti‐inflammatory profile in an attempt to reduce or prevent the tissue damage, while the upregulation of ADA activity exerts a pro‐inflammatory profile, contributing to disease pathophysiology.     

Serum adenosine deaminase and xanthine oxidase activities in silver catfish naturally infected with Ichthyophthirius multifiliis: The influence of these enzymes on inflammatory and oxidative status

Adenosine deaminase (ADA) activity, through adenosine (Ado) levels, as well as xanthine oxidase (XO) activity through uric acid levels exerts an essential role on immune and inflammatory responses during infectious diseases. Thus, the aim of this study was to evaluate the involvement of seric ADA and XO activities in the inflammatory and oxidative status of silver catfish naturally infected with Ichthyophthirius multifiliis. Seric ADA activity decreased, while Ado levels increased in infected animals compared to uninfected animals. Moreover, the seric XO activity increased in infected animals compared to uninfected animals, alongside the seric levels of uric acid, metabolites of nitric oxide (NOx) and reactive oxygen species (ROS). Based on this evidence, the downregulation of seric ADA activity exerts an anti‐inflammatory profile, contributing to restricting the inflammatory process. The most important finding is that upregulation of seric XO activity leads to an excessive formation of uric acid, which contributes to oxidative and inflammatory processes. Moreover, uric acid induces the release of pro‐inflammatory and pro‐oxidative mediators, such NOx and ROS, which contribute directly to disease pathogenesis. In summary, the upregulation of XO activity may be considered a pathway involved in NOx and ROS production in silver catfish infected with I. multifiliis.     

Aeromonas caviae inhibits hepatic enzymes of the phosphotransfer network in experimentally infected silver catfish: Impairment on bioenergetics

Several studies have been demonstrated that phosphotransfer network, through the adenylate kinase (AK) and pyruvate kinase (PK) activities, allows for new perspectives leading to understanding of disease conditions associated with disturbances in energy metabolism, metabolic monitoring and signalling. In this sense, the aim of this study was to evaluate whether experimental infection by Aeromonas caviae alters hepatic AK and PK activities of silver catfish Rhamdia quelen. Hepatic AK and PK activities decreased in infected animals compared to uninfected animals, as well as the hepatic adenosine triphosphate (ATP) levels. Also, a severe hepatic damage was observed in the infected animals due to the presence of dilation and congestion of vessels, degeneration of hepatocytes and loss of liver parenchyma architecture and sinusoidal structure. Therefore, we have demonstrated, for the first time, that experimental infection by A. caviae inhibits key enzymes linked to the communication between sites of ATP generation and ATP utilization. Moreover, the absence of a reciprocal compensatory mechanism between these enzymes contributes directly to hepatic damage and for a severe energetic imbalance, which may contribute to disease pathophysiology.     

Xylanase supplementation in plant protein‐enriched diets improves growth performance by optimizing the intestinal microflora and enhancing the intestinal immune function in grass carp grow‐out (Ctenopharyngodon idella)

This study investigated the effect of xylanase supplementation of a plant protein‐enriched diet on the growth performance and intestinal immune function of grass carp grow‐out (Ctenopharyngodon idella). Five hundred forty healthy grass carp grow‐out (232.74 ± 0.12 g) were fed diets containing graded levels of xylanase for 60 days. The fish were then challenged with Aeromonas hydrophila for 14 days. The results indicated that xylanase supplementation (a) improved fish growth performance and optimized the intestinal microflora; (b) increased lysozyme and acid phosphatase activities, complement and immunoglobulin M levels and antibacterial peptide hepcidin mRNA levels, thereby enhancing fish intestinal immune function; and (c) attenuated intestinal inflammation associated with the inhibition of the NF‐κB signalling pathway and activation of the TOR signalling pathway, resulting in the down‐regulation of pro‐inflammatory cytokines [except IL‐1β and IL‐12p40 in the distal intestine (DI) and IL‐6 in the proximal intestine (PI)] and up‐regulation of anti‐inflammatory cytokines [except IL‐4/13B and TGF‐β1 in the mid intestine (MI) and DI]. Finally, on the premise of a basal diet containing 45.8 g/kg araboxylans (AXs), the appropriate supplementation levels based on the ability to improve per cent weight gain (PWG) and prevent enteritis were estimated to be 1,527 and 1,608 U/kg, respectively.     

Histopathological changes induced by the digenean intestinal parasite Masenia nkomatiensis Dumbo,Dos Santos, & Avenant‐Oldewage, 2019 of the catfish Clarias gariepinus (Burchell) from Incomati Basin,Mozambique

The intestines of 154 Clarias gariepinus were examined of which 29 were naturally infected with Masenia nkomatiensis, and of these, seven (intensity ranging from 8 to 231) were examined for pathology. Destruction of the epithelium covering the villi, detachment of epithelial cells and parts of villi were observed. Excessive mucus secretion occurred in the vicinity of the worm and catarrh was observed, indicative of an inflammatory response. The number of mucous and mast cells was higher at the attachment site than at an area 5,000 µm away and in uninfected individuals, suggesting that the parasite triggered a localized innate immune response. The number of neutrophils, basophils and lymphocytes in infected tissue was not significantly different from uninfected tissue confirming that no acquired immune response was produced against the maseniid. The caecae in the anterior part of the parasites' intestine consisted of convoluted epithelium forming invaginations or “crypts.” Contraction of the thick layer of circular muscle fibres of the caeca facilitates the movement of digested material. Observation of digested host cells and cell debris within the caecae provides further evidence that M. nkomatiensis is consuming host cells.     

Synergistic osmoregulatory dysfunction during salmon lice (Lepeophtheirus salmonis) and infectious hematopoietic necrosis virus co‐infection in sockeye salmon (Oncorhynchus nerka) smolts

While co‐infections are common in both wild and cultured fish, knowledge of the interactive effects of multiple pathogens on host physiology, gene expression and immune response is limited. To evaluate the impact of co‐infection on host survival, physiology and gene expression, sockeye salmon Oncorhynchus nerka smolts were infected with the salmon louse Lepeophtheirus salmonis (V?/SL+), infectious hematopoietic necrosis virus (IHNV; V+/SL?), both (V+/SL+), or neither (V?/SL?). Survival in the V+/SL+ group was significantly lower than the V?/SL? and V?/SL+ groups (p = 0.024). Co‐infected salmon had elevated osmoregulatory indicators and lowered haematocrit values as compared to the uninfected control. Expression of 12 genes associated with the host immune response was analysed in anterior kidney and skin. The only evidence of L. salmonis‐induced modulation of the host antiviral response was down‐regulation of mhc I although the possibility of modulation cannot be ruled out for mx‐1 and rsad2. Co‐infection did not influence the expression of genes associated with the host response to L. salmonis. Therefore, we conclude that the reduced survival in co‐infected sockeye salmon resulted from the osmoregulatory consequences of the sea lice infections which were amplified due to infection with IHNV.     

ATP,ADP and AMP dephosphorylation in membrane fractions of <Emphasis Type="Italic">Rhamdia quelen</Emphasis> exposed to different temperatures

The silver catfish Rhamdia quelen is a teleost species from South America that can resist cold winters and grow quickly in the summer, representing an important species for aquaculture in both temperate and subtropical climates. NTPDase and 5′-nucleotidase are enzymes responsible for the sequence of ATP dephosphorylation to adenosine in a variety of organs. This enzymatic cascade permits the resultant adenosine to be taken up by the cells. In the present study, membrane fractions of brain, liver and kidney from fish Rhamdia quelen exposed to 15, 23 and 31 °C for 21 days (chronic) or 12 h (acute) were incubated with ATP, ADP and AMP as substrates. In the 21 day experiment the results showed that ATP hydrolysis was enhanced by the increase in temperature in all tissues, except for 31 °C in the brain. NTPDase hydrolysis using ADP as substrate showed a behavior similar to that observed for ATP hydrolysis, except for 31 °C, when the behavior was similar to that observed at 15 °C in the brain. 5′-Nucleotidase activity using AMP as substrate raised with increasing temperature in the three tissues studied. In the 12 h experiment the results obtained for the three temperatures did not differ significantly from each other for any of the tissues. We conclude that after a period of 21 days temperature may affect the activity of NTPDase and 5′-nucleotidase, important enzymes implicated in nucleotide degradation.     

Potential vector species of carp edema virus (CEV)

During a PCR‐based CEV survey in Poland in 2015–2017, the virus was detected in many farms both in clinical and asymptomatic cases and in common as well as in koi carp (Cyprinus carpio). In order to evaluate the potential carrier role of fish species that share the same habitats with carp, an experimental trial was performed. Investigations carried out on specimens of bleak (Alburnus alburnus), crucian carp (Carassius carassius), European perch (Perca fluviatilis), Prussian carp (Carassius gibelio), roach (Rutilus rutilus) and tench (Tinca tinca) cohabited with CEV‐infected carp yielded positive results. These species of fish were experimentally cohabited with CEV‐infected common carp at a temperature of 16°C ± 1. Material from the brain, gills, spleen, kidneys, intestine and skin was investigated for the presence of CEV DNA. Similar investigations were performed with uninfected fish designated controls. Samples were tested for CEV by qPCR.     

The response of intestinal mucous cells to the presence of enteric helminths: their distribution,histochemistry and fine structure

Histochemical and ultrastructural investigations were conducted on the mucous cells of the intestine of brown trout, Salmo trutta L., naturally infected with the cestode Cyathocephalus truncatus (Pallas, 1781) and the acanthocephalan Echinorhynchus truttae Shrank, 1788. A subpopulation of 45 S. trutta were examined of which 15 specimens harboured E. truttae, 15 of which were infected with C. truncatus and 15 fish, the control group, were uninfected. In histological sections, hyperplasia and hypertrophy of the mucous cells were evident at the site of parasite infection. Enhanced mucus secretion was also recorded in infected fish. The number of mucous cells close to the site of parasite attachment within the intestine was significantly higher than the number detected in uninfected individuals and in infected individuals at sites 1 cm or greater from the point of parasite attachment. There were no significant differences between the number of mucous cells found at the latter two sites. Alcian blue and periodic acid‐Schiff’s staining of representative histological sections revealed a significant increase in the number of mucous cells staining positively for acid glycoconjugates compared to the number of cells found in the intestines of uninfected S. trutta. In transmission electron microscopy sections, each mucous cell typically possessed an elongated, basally positioned nucleus. The cytoplasm was observed to possess numerous electron dense and lucent vesicles, in addition to well‐developed rough endoplasmic reticulum, Golgi apparatus and a few round mitochondria.     

Dietary 5‐aminolevulinic acid enhances adenosine triphosphate production,ecdysis and immune response in Pacific white shrimp,Litopenaeus vannamei (Boone)

Previous studies have shown that 5‐aminolevulinic acid (5‐ALA), a precursor of haem, can enhance haem synthesis and the activity of haemoproteins. Here, we investigated the effects of dietary 5‐ALA on Pacific white shrimp, Litopenaeus vannamei (Boone). Dietary groups included basal diet (BD, control) and BD plus 15, 30 and 60 ppm 5‐ALA (ALA15, ALA30 and ALA60 respectively). Hepatopancreas adenosine triphosphate (ATP) levels increased with increasing 5‐ALA concentration (ALA60 p < 0.05) after 2 weeks of feeding. 5‐aminolevulinic acid diets significantly increased the expression of ecdysis‐related genes: nuclear receptor E75 and chitinase 4 (ALA15, ALA30, ALA60), cytochrome P450 Shade (ALA60), chitinase 1 (ALA60) and chitinase 3 (ALA15, ALA60). Catalase (CAT) and prophenoloxidase gene expression levels were also significantly higher in ALA60 after 12 weeks of feeding. Six hours after L. vannamei were exposed to Vibrio parahaemolyticus, total haemocyte count (ALA60) and gene expression levels of CAT (ALA30, ALA60) were significantly higher in 5‐ALA groups compared to the control. 5‐aminolevulinic acid diets also increased survival of L. vannamei following V. parahaemolyticus immersion challenge. These results suggest that supplementing L. vannamei diets with 5‐ALA can enhance ATP production, immune response against V. parahaemolyticus, total haemocyte count and expression of some immune‐related genes. 5‐aminolevulinic acid can also induce ecdysis‐related gene expressions, without adversely affecting growth.     

A natural infection by the red sea bream iridovirus‐type Megalocytivirus in the golden mandarin fish Siniperca scherzeri

An outbreak of a Megalocytivirus infection was found in the golden mandarin fish Siniperca scherzeri during September and October 2016, in Korea. Phylogeny and genetic diversity based on the major capsid protein (MCP) and adenosine triphosphatase (ATPase) genes showed a new strain. Designated as GMIV, this strain derived from the golden mandarin fish was suggested to belong to the red sea bream iridovirus (RSIV)‐subgroup I. Additionally, this train clustered with the ehime‐1 strain from red sea bream Pagrus major in Japan and was distinguished from circulating isolates (RSIV‐type subgroup II and turbot reddish body iridovirus [TRBIV] type) in Korea. The infection level, evaluated by qPCR, ranged from 8.18 × 10² to 7.95 × 10⁶ copies/mg of tissue individually, suggesting that the infected fish were in the disease‐transmitting stage. The diseased fish showed degenerative changes associated with cytomegaly in the spleen as general sign of Megalocytivirus infection. The results confirm that the RSIV‐type Megalocytivirus might have crossed the environmental and species barriers to cause widespread infection in freshwater fish.     

Molecular isolation and characterization of a haemocyanin of Macrobrachium rosenbergii reveal its antibacterial activities

Haemocyanin is a multi‐subunit protein complex found in the haemolymph and is involved in the immune system of crustaceans. In this study, a haemocyanin gene of Macrobrachium rosenbergii, designated MrHc, was successfully isolated. The MrHc gene contained an open reading frame (ORF) of 1,992 nucleotides, encoding a protein of 663 amino acid residues with a molecular mass of 76.5 kDa. The deduced amino acid sequence contained distinct structural motifs of the haemocyanin superfamily, including an all‐alpha domain, a copper‐containing domain and an immunoglobulin‐like domain. Based on the phylogenetic analysis, the MrHC protein demonstrated a close relationship with the haemocyanins of Palaemon carinicauda and Macrobrachium nipponense. The MrHc gene was expressed in various shrimp tissues, including the hepatopancreas, gill, haemocytes, stomach and muscle. After Macrobrachium rosenbergii nodavirus (MrNV) challenge tests, the MrHc gene was up‐regulated 237‐fold at day 2. A recombinant protein of the MrHc immunoglobulin‐like domain exhibited antibacterial activity against Vibrio vulnificus, V. parahaemolyticus, Aeromonas caviae, A. veronii, A. hydrophila and Bacillus cereus. This study suggested that MrHc may play important roles in the shrimp innate immune response to MrNV infection and bacterial infection.     

The immune response in rohu,Labeo rohita (Actinopterygii: Cyprinidae) to Argulus siamensis (Branchiura: Argulidae) infection: kinetics of immune gene expression and innate immune response

Argulus siamensis is a major pathogen in freshwater aquaculture. The immune responses of Indian major carp, Labeo rohita to experimental infection of A. siamensis was evaluated by quantitation of immune‐relevant gene expression in head kidney and skin, and serum innate immune parameters through the course of infection. In skin of infected fish, antioxidant genes like natural killer cell enhancing factor (NKEF‐B) and superoxide dismutase (MnSOD) were significantly up‐regulated in addition to lysozyme G and β₂ microglobulin (β₂M). Both tumour necrosis factor α (TNFα) and toll‐like receptor 22 (TLR22) genes were significantly down‐regulated in skin during early phases of the infection. Most of the genes exhibited significant down‐regulation in head kidney; immunoglobulin (IgM) and β₂M genes being the exceptions which were significantly up‐regulated at 12 h and 3 days post infection. Most of the innate immune parameters like serum complement activity and ceruloplasmin levels showed significant reduction in infected fish. The observed results are indicative of A. siamensis modulating the immune response of rohu by down‐regulation of many immune factors which may explain the susceptibility of rohu to A. siamensis infection. The interaction of this parasite with the host need to be further explored to understand its pathogenesis.