Resistance of the European catfish (Silurus glanis) to channel catfish virus

European catfish (Silurus glanis) fingerlings (2 to 4 g each) were tested for susceptibility to channel catfish virus (CCV). They had supported CCV replication at 2 days after intraperitoneal injection with 0.1 ml of saline containing 10⁵ TCID₅₀. Homogenized visceral organs (liver, kidney and spleen) contained 10⁴ TCID₅₀/0.1 ml at 2 days post inoculation (PI) but at 4 days the titer decreased to 10¹ TCID₅₀. Bathing European catfish in CCV yielded only one positive sample with à titer of 10^0.83 TCID₅₀ per 0.1 ml of tissue. No clinical signs of CCV developed and no virus related deaths occurred.     

Blood flow distribution and tissue allometry in channel catfish

Blood flow (as percentage of cardiac output) in fasted channel catfish acclimated to 21°C was directed primarily to white muscle (72%) followed by head kidney (5·7%), red muscle (5·5%), trunk kidney (3·1%), liver (2·2%), swim bladder (1·4%) and skin (1·1%). The stomach, intestines, pyloric caeca, gonads, brain, abdominal fat and spleen contained <0·5% of blood flow. There was considerable interfish variation among blood flow distribution to visceral organs with substantial spatial heterogeneity of blood flow to white muscle. The spatial heterogeneity of flow to muscle prevented accurate estimation of total flow to this tissue based on the microsphere deposition of a few sub-samples. Instead, a novel approach, based on the whole animal counting of the eviscerated carcass was used to measure blood flow to white muscle. The scaling relationships for tissue mass in catfish (63–1873 g) followed the allometric equation (aW^b) and tended to exhibit negative allometry, with organ weight decreasing in proportion to body weight. The b values for most tissues ranged between 0·83 and 1·0. The relative mass of the brain showed the greatest decline and with a b value of 0·32. The results, together with previous data on cardiac output, permitted calculation of organ blood flow rates in channel catfish. © 1999 The Fisheries Society of the British Isles     

Pathology associated with Edwardsiella ictaluri in catfish, Ictalurus punctatus Rafinesque, and Danio devario (Hamilton-Buchanan, 1822)

The pathology associated with infections of Edwardsiella ictaluri in a new host, the danio, is described and compared to that observed in the channel catfish. In catfish the acute stage was characterized by petechial haemorrhage of the jaw, ventral body surface, and at the base of the fins. In chronic cases a characteristic finding was an erosion of skin and muscle overlying the skull, exposing bone and occasionally the brain. Histologically there was a diffuse granulomatous inflammation of the olfactory bulb and the telencephalon. In the catfish the infection was often systemic, involving intestine, liver, spleen, and occasionally kidney. In the danio no gross lesions were observed. Infected fish exhibited an erratic, spinning-type swimming behaviour. Histologically, lesions usually were confined to the brain. They consisted of an acute, primarily granulocytic inflammation of the medulla oblongata (rhomboencephalon). Only one fish showed evidence of a systemic infection.     

Cloning and characterisation of a channel catfish (Ictalurus punctatus) Mx gene

A 2.5 kb full-length cDNA clone of a channel catfish (Ictalurus punctatus) Mx gene was obtained using RACE (rapid amplification of cDNA ends) polymerase chain reaction (PCR) from RNA extracted from the liver of poly I:C stimulated channel catfish. The gene consists of an open reading frame of 1905 nucleotides encoding a 635 amino acid protein. The predicted protein is 72.5 kDa and contains the dynamin family signature, a tripartite GTP binding motif and a leucine zipper, characteristic of all known Mx proteins. The catfish Mx protein exhibited 79% identity with perch Mx and between 71% and 74% identity with the three Atlantic salmon and the three rainbow trout Mx proteins. Mx mRNA was constitutively expressed in channel catfish ovary (CCO) cells, but in higher quantities in response to poly I:C treatment. Mx was induced in channel catfish following injection with channel catfish virus (CCV) and poly I:C.     

Pathological changes in cultured channel catfish Ictalurus punctatus spontaneously infected with Streptococcus iniae

The pathological changes present in channel catfish Ictalurus punctatus spontaneously infected by Streptococcus iniae are described. The most consistent gross findings were marked petechial hemorrhages of the skin and congestion of internal organs, particularly the liver, spleen and kidneys. Other features included color fading at the edge of fin rays, enteritis and ascites. Histological examination showed oedema, degeneration and necrotic changes in many organs. Further, hepatitis, splenitis, interstitial nephritis, and meningitis with numerous monocyte and neutrocyte infiltrates were evident. Intact S. iniae cells were seen in macrophages. Apparently, spontaneous S. iniae infection caused acute septicaemia in channel catfish. This is the first histopathological report on channel catfish naturally infected with S. iniae.     

Immune hypersensitivity in the channel catfish, Ictalurus punctatus (Rafinesque)

Channel catfish, Ictalurus punctatus , exhibited passive cutaneous anaphylaxis (PCA) when sensitized with Flexibacter columnaris antiserum produced in channel catfish (the first record of PCA in any fish), but not when sensitized with homologous antiserum against channel catfish virus. Channel catfish did not give delayed skin hypersensitivity responses after immunization with either channel catfish virus or F. columnaris .     

Histology of cultured channel catfish, Ictalurus punctatus (Rafinesque)

Dissolved oxygen and un-ionized ammonia concentrations were monitored in 12 0.04 ha earthen ponds stocked with 10 000 channel catfish, Ictalurus punctatus ,/ha. Gill, liver and trunk kidney tissue samples were removed periodically for histological examination. Total ammonia and dissolved oxygen levels were in the ranges reported for catfish culture at this level of intensity. Average un-ionized ammonia nitrogen concentrations ranged from 20 to 67 μgh⁻¹ and average daily maxima ranged from 63 to 183 μgh⁻¹. Gill lesions characteristic of un-ionized ammonia exposure were common in fish from all ponds.     

Alternative complement pathway of channel catfish (Ictalurus punctatus): molecular characterization, mapping and expression analysis of factors Bf/C2 and Df

The complement system is important in both innate and adaptive host defense against microbial infection in vertebrates. It contains three pathways: the classical, alternative, and lectin pathways. Complement component factors B and D are two crucial proteases in the alternative pathway. In this study, the genes of complement factors Bf/C2 and Df from channel catfish, Ictalurus punctatus were identified and characterized. Two complement factor B-related genes, Bf/C2A and Bf/C2B, and factor D gene Df were identified. Phylogenetic analysis suggested that Bf/C2A and Bf/C2B is likely orthologous to factor B and factor C2, respectively. Southern blot results suggested that these three genes are all single-copy genes in the catfish genome. The catfish Bf/C2A, Bf/C2B and Df genes were genetically mapped on linkage group 3, 20 and 29, respectively. Bf/C2A and Bf/C2B are highly expressed in liver and kidney, while Df is highly expressed in gill and spleen. After infection with Edwardsiella ictaluri, the expression of Bf/C2A, Bf/C2B and Df genes were found to be remarkably induced in the gill, liver, spleen and kidney at some sampling times, indicating that these three complement factors play a pivotal role in immune responses after the bacterial infection in catfish.     

Effects of 2-methylisoborneol (MIB), and ethanol on the expression and activity of cytochrome P450s from the channel catfish

Injection of 1 mg kg⁻¹ of 2-methylisoborneol (MIB) caused an increase in a CYP1A-like protein of approximately 56 kDa from the kidneys of juvenile channel catfish as determined by western blot analysis using antibodies raised against trout CYP1A1. Ethoxyresorufin O-deethylase activity from kidneys of MIB-injected fish was significantly elevated as compared to ethanol-injected controls. Static exposure of juvenile channel catfish to 10 ppm MIB caused a statistically significant induction of a CYP2K-like protein of approximately 53 kDa from the livers of treated catfish as determined by western blot analysis using antibodies raised against trout CYP2K1. Treatment of juvenile channel catfish with ethanol (1 ml kg⁻¹) reduced the expression of one kidney and two constitutive liver P450s, while increasing another kidney form. There was no difference in carbon tetrachloride-induced lipid peroxidation in livers after ethanol treatment. Thus, MIB and ethanol affect the expression of at least three P450 isoforms in channel catfish tissues.     

Cloning and expression analysis of c-type and g-type lysozymes in yellow catfish (<Emphasis Type="Italic">Pelteobagrus fulvidraco</Emphasis>)

Lysozymes have important roles in innate immune system. Here, a c-type and a g-type lysozyme were identified from yellow catfish (Pelteobagrus fulvidraco). The deduced amino acid sequences of both lysozymes were conserved in catalytic sites and structural features as compared to their counterparts from other species. It was interesting that the g-type lysozyme possessed a signal peptide. The c-type and g-type lysozymes had the highest identity 89.4 and 76.2 % with that from channel catfish respectively. Phylogenetic analysis showed that the two lysozymes had a closely relationship with that from channel catfish and Astyanax mexicanus. Lysozymes from one order could form more than one clade in the phylogenetic tree, which indicated the gene duplications in evolution. Expression analysis with real time quantitative PCR revealed that the two lysozyme genes were constitutively expressed in all the tested tissues. The highest expression of c-type lysozyme was observed in liver, followed by spleen, head kidney, and trunk kidney, while the g-type lysozyme had highest expression in intestine, followed by spleen, head kidney, and trunk kidney. The mRNA levels of both genes were all up-regulated after challenging with Aeromonas hydrophila. However, there were differences in tissues and time points when the mRNA levels reached its peak between the two lysozymes. It indicated the diversity in regulation mechanisms and detailed functions among lysozymes. Taking together, these results will benefit the understanding of yellow catfish lysozymes.     

Susceptibility of European catfish (Silurus gianis) to Edwardsiella ictaluri

European catfish (Silurus glanis) were tested for their susceptibility to the bacterium Edwardsiella ictaluri. The LD₅₀ of E. ictaluri when injected into European catfish was 5.4 × 10⁶ compared to 7.1 times; 10⁴ for channel catfish (Ictalurus punctatus). E. ictaluri was isolated from dead and moribund European catfish and the bacterium was also detected in kidney smears by an indirect fluorescent antibody (FA) technique. The bacterium was not isolated or detected by FA from surviving fish 15 days after injection. No clinical signs of E. ictaluri infection were noted in European catfish, but these were prevalent in the channel catfish. These experiments indicate that under experimental conditions European catfish are not as susceptible to E. ictaluri as channel catfish.     

Morphogenesis of blood cell lineages in channel catfish

The morphogenesis of blood cell lineages in channel catfish Ictalurus punctatus , from head and trunk kidney and spleen imprints as well as from blood smears of bled and control fish, showed that early maturation stages resembled those in higher vertebrates. The erythroid lineage consisted of the proerythroblast, erythroblasts (basophilic, polychromatic, orthochromic), young erythrocyte and erythrocyte. The rare bilobed erythrocyte seemed to be a cell in apoptosis while old erythrocytes and erythroplastids represented remnants of this process. Maturation stages of neutrophils and basophils encompassed the granuloblast, young progranulocyte, progranulocyte and metagranulocyte. The basophilic lineage was regularly present in kidneys, rare in spleen and absent from blood. It contained large Sudan Black and PAS-negative, water soluble granules and small PAS-positive ones. Lymphocytes with azurophilic granulation occured regularly in kidneys and spleen. Monoblasts and promonocytes in kidneys preceded monocytes. A phagocytic lineage devouring apoptotic blood cell remnants was present in kidneys and spleen. Its youngest identified stage (promacrophage) resembled more a granuloid cell without granules than a monocytoid one. The larger, young macrophages contained a few to several ingestions and the very large mature macrophages were loaded with them. The latter two stages corresponded to cells in melano-macrophage centres (macrophage aggregates). Precursor stages of the thrombocyte were not identified.     

BIRC7 gene in channel catfish (Ictalurus punctatus): identification and expression analysis in response to Edwardsiella tarda, Streptococcus iniae and Channel catfish Hemorrhage Reovirus

A family member of inhibitor of apoptosis protein (IAP) termed baculoviral IAP repeat-containing 7 (BIRC7) from channel catfish (Ictalurus punctatus) was identified, the full length cDNA sequence of channel catfish BIRC7 (CcBIRC7) was 1686?bp, containing a 5'UTR of 93?bp, a 3'UTR of 399?bp with a poly (A) tail and an ORF of 1194?bp encoding a putative protein of 398 amino acids. The putative CcBIRC7 protein contains two BIR super-family conservative domains and a C-terminal RING finger motif. Phylogenetic analysis showed that catfish CcBIRC7 was moderately conserved with other BIRC7. Quantitative real-time PCR was conducted to examine the expression profiles of CcBIRC7 in healthy tissues and responding to different pathogens (Edwardsiella tarda, Streptococcus iniae and Channel catfish Hemorrhage Reovirus (CCRV)). CcBIRC7 was widely expressed in healthy tissues of channel catfish and with the highest 37.28-fold expression in blood. E.?tarda and S.?iniae could induce CcBIRC7 gene expression drastically in head kidney, liver and spleen, which the peak value reached 31.6-fold, 613.9-fold and 34.4-fold increase by E.?tarda infection, and 248.3-fold, 1540.3-fold and 120.4-fold increase post S.?iniae challenge, respectively. While, CCRV virus could slightly induce CcBIRC7 expression in head kidney and liver but reduce it in spleen. The result suggested BIRC7 may play a potential role in channel catfish innate immune system against bacterial and virus infections, especially as the anti-bacteria immune gene. This is the first report of BIRC7 gene identification and its expression in fish.     

Molecular characterization of the insulin-like growth factor-I (IGF-I) gene in channel catfish (Ictalurus punctatus)

The insulin-like growth factor I (IGF-I) gene was characterized in channel catfish. Partial cDNA sequence, missing exon 1 and part of exon 2, was obtained in 5'- and 3'-RACE experiments. Direct sequencing of two bacterial artificial chromosome clones revealed gene structure and provided sequence from 640 bp upstream of the initiator methionine to 136 bp beyond the polyadenylation site. Genomic sequence contained a putative TATA box 506 bp upstream of the initiator methionine. The 477-bp reading frame within five exons encoded a 159-amino acid (aa) pre-propeptide highly similar to IGF-I in higher vertebrates. The sequence encoding the signal peptide was unique in catfish and contained 70% G+C content with the potential for a stable stem-loop structure. Full-length cDNA was only maintained in recombination-deficient (DH10B) strain E. coli. Levels of IGF-I mRNA were highest in liver, followed by brain and muscle, then heart and kidney (P<0.05). A CT/GA dinucleotide microsatellite in intron 1 was highly polymorphic in commercial channel catfish, and permitted placement of the IGF-I gene on the catfish genetic map. However, specific IGF-I alleles were not correlated with differences in growth rate from 100 to 130 days post-hatch in USDA103 line catfish.     

Response of toll-like receptors, lysozyme, and IGF-I in back-cross hybrid (F1 male (blue x channel) x female channel) catfish challenged with virulent Edwardsiella ictaluri

Responses of toll-like receptors (TLR3 and TLR5), lysozyme, and insulin-like growth factor-I (IGF-I) to experimental challenge with virulent Edwardsiella ictaluri were measured in back-cross hybrid (F1 male (blue x channel) x female channel) catfish. The resistance levels to E. ictaluri and host response mechanisms of back-cross hybrids are unknown. Fish were challenged with virulent E. ictaluri and sampled pre-challenge, 2 h and 2, 5, 8, 14, and 21 days post-challenge. Levels of mRNA expression of two toll-like receptors (TLR3 and TLR5) in liver, kidney, spleen, and stomach, plasma lysozyme activity, and circulating IGF-I levels were measured at each timepoint. Throughout challenge, TLR3 was expressed at higher levels than TLR5 in liver (P=0.0011) and kidney (P=0.0007) whereas TLR5 was more highly expressed than TLR3 in stomach (P=0.0032). TLR3 was upregulated in comparison to non-exposed controls in liver (P=0.0015) and stomach (P<0.0001) on day 14 and TLR5 was upregulated in liver (P=0.0175) on days 2 through 8. Plasma lysozyme activity peaked on day 5 (P<0.001) and IGF-I levels significantly decreased on days 2 through 14 (P<0.0001). TLR expression patterns suggest that both TLR3 and TLR5 may play a role in host response to bacterial challenge. Plasma lysozyme activity also increased and circulating IGF-I decreased in response to the presence of the pathogen.     

Virulence of Aeromonas hydrophila to channel catfish Ictaluras punctatus fingerlings in the presence and absence of bacterial extracellular products

We investigated the virulence of three 2009 west Alabama isolates of Aeromonas hydrophila (AL09-71, AL09-72 and AL09-73) to channel catfish Ictalurus punctatus fingerlings (4.6 +/- 1.3 g) in the presence and absence of extracellular products (ECPs) from overnight bacterial culture using both bath immersion and intraperitoneal injection routes. At a concentration of 1.65 x 10(8) colony-forming units (CFU) ml(-1), AL09-73 without its ECPs killed 100% of the catfish fingerlings within 2 h by bath immersion. However, at a similar concentration, AL09-73 in the presence of its ECPs killed only 23 +/- 6% catfish fingerlings. The absence of ECPs in the bath immersion experiment also significantly (p < 0.05) increased the virulence of AL09-71, AL09-72, and AL98-C1B, a 1998 Alabama strain of A. hydrophila, suggesting that the virulence of the 4 A. hydrophila isolates was mainly due to bacterial cells, not to their overnight ECPs. Filter-sterilized ECPs failed to kill any catfish by bath immersion or injection. The virulence order of the 4 A. hydrophila isolates, by both bath immersion and intraperitoneal injection, was: AL09-73 > or = AL09-71 > AL09-72 > or = AL98-C1B. At 2 h post bath immersion, all 4 isolates of A. hydrophila were found in all tissues studied (skin, intestine, liver, spleen, kidney, gill and brain), with the highest bacteria count being in the gill and kidney.