Use of Diet Crossover to Determine the Effects of β‐glucan Supplementation on Immunity and Growth of Nile Tilapia,Oreochromis niloticus

Juvenile Nile tilapia were fed either a basal (control) diet (n = 6 aquaria) or a diet supplemented with 1 g/kg β‐glucan (n = 24 aquaria) for 4 wk. At the end of this period, fish receiving β‐glucan were continued on the same diet (n = 12 aquaria) or switched to the control diet (n = 12 aquaria) for 2 wk. After 6 wk, tilapia continuously fed the β‐glucan supplemented diets had improved weight gain and feed efficiency than those fed the control diet uninterrupted or switched from the β‐glucan diet to the control after 4 wk. Feeding tilapia β‐glucan for 4 wk and then switching to the basal diet for 2 wk caused a significant increase in the respiratory burst of polymorphonuclear lymphocytes (17.77 × 10³ units/1000 white blood cells [WBC]) compared to catfish fed the control diet (13.50 × 10³ units/1000 WBC) or the β‐glucan diet continuously (13.57 × 10³ units/1000 WBC), but other immune parameters were unaffected. Tilapia were then challenged with Streptococcus iniae. The two groups were divided again (n = 6 aquaria) postchallenge and continued on the same diet or switched to the other diet (β‐glucan or control) for another 3 wk. No differences in survival to S. iniae infection occurred between dietary groups.     

Lymphocyte blastogenic responses of calves experimentally infected with Ostertagia ostertagi

Twelve 9-week-old calves were divided into four groups; two groups were maintained helminth-free as controls and the other groups were given Ostertagia ostertagi infective-stage larvae (L3) orally. One group received 100,000 L3 as a single inoculum and the other group received L3 in increasing dosages at weekly intervals for 8 consecutive weeks. The blastogenic responses to concanavalin A (Con A), phytohemagglutinin (PHA), and a soluble larval antigen from O. ostertagi (SLA) of peripheral blood lymphocytes were evaluated using tritiated thymidine incorporation into DNA as a measure of blastogenesis. The responses to Con A of all infected calves were significantly depressed while the responses to PHA were not. SLA, at concentrations of 4 micrograms ml-1 and above, caused blastogenic activity in lymphocytes from uninfected calves. Using SLA at 1 microgram ml-1 in lymphocyte cultures supplemented with autologous serum, an antigen-induced blastogenic response was detected in calves receiving serial inoculations of L3.     

Molecular typing of Streptococcus agalactiae isolates from fish

The genetic variability among Streptococcus agalactiae isolates recovered from fish was characterized using single-stranded conformation polymorphism (SSCP) analysis of the intergenic spacer region (ISR), and amplified fragment length polymorphism (AFLP) fingerprinting. A total of 46 S. agalactiae cultures isolated from different fish species and geographic origins as well as related reference strains were included in the study. ISR-SSCP divided the S. agalactiae isolates analysed into five distinct genotypes. Genotype 1 grouped all Kuwait isolates while genotype 4 clustered the majority of non-Kuwait isolates (USA, Brazil and Honduras). AFLP analysis offered a higher resolution level by dividing the isolates into 13 different genotypes. Two different AFLP profiles were identified within the Kuwait isolates. When data from both ISR-SSCP and AFLP were combined through a multidimensional analysis (MDS), a good correlation between geographical origin and genotypes was observed. Both AFLP and ISR-SSCP revealed genetic differences between S. agalactiae isolates from fish. While AFLP offered a higher resolution, ISR-SSCP also provided valid information being a simpler and faster method.     

Evaluation of the link between gyrodactylosis and streptococcosis of Nile tilapia, Oreochromis niloticus (L.)

Streptococcus iniae and Gyrodactylus niloticus are two common pathogens of cultured Nile tilapia, Oreochromis niloticus. We studied concurrent infection of tilapia by G. niloticus and S. iniae and evaluated whether parasitism in tilapia with Gyrodactylus increased susceptibility and mortality following immersion infection with S. iniae. Results showed that death mainly occurred in fish with G. niloticus and challenged with S. iniae (G-S group). The accumulative mortality (42.2%) was significantly higher in the G-S group than in fish not infected by the parasite (6.7%), but exposed to S. iniae. Bacteriological examination revealed S. iniae from > or =92% of dead or moribund fish challenged with S. iniae. Gyrodactylus not only damaged fish epithelium and provided entry for invasive bacteria but also was found to harbour viable cells of S. iniae for 24 and 72 h. Streptococcus iniae was isolated from 60% and 40% of G. niloticus collected from fish infected by intraperitoneal injection or immersion, respectively, at 24 h post-challenge. The present study confirms that parasitism of tilapia by G. niloticus increased host mortality following exposure to the bacterial pathogen S. iniae.     

Vaccination of Mixed and Full-Sib Families of Channel Catfish Ictalurus punctatus Against Enteric Septicemia of Catfish With a Live Attenuated Edwardsiella ictaluri Isolate (RE-33)

These studies were conducted to evaluate the efficacy of a live attenuated Edwardsiella ictaluri vaccine against enteric septicemia of catfish. In one study channel catfish fingerlings (72 d of age post hatch) were immersed for 30 min in water containing E. ictaluri RE-33 at dosages of 1 × 10⁶, 1 × 10⁷ and 2 × 10⁷ CFU/ML of water. No mortalities were observed following vaccination. Following exposure to virulent Edwardsiella ictaluri the cumulative mortality of fish vaccinated with dosages of at least 1 × 10⁷ CFU/mL were significantly lower than that of non-vacccinated fish in both laboratory and field challenges. Vaccination with 1 × 10⁶ CFU RE-33mL provided some protection during the laboratory challenge but failed to protect fish under field conditions. In a second study, vaccination of 6 full-sib families of channel catfish at a vaccine dosage of 1 × 10⁷ CFU/mL resulted in a relative percent survival among families ranging from 67.1 to 100%. Significant differences in mortality were found among families and between vaccinated and unvaccinated groups, but there was no family by vaccine interaction. Families with the highest mortality after vaccination were also shown to have the highest mortality without vaccination (r = 0.82; P = 0.04).     

Intra- and interspecific phenotypic characteristics of fish-pathogenic Edwardsiella ictaluri and E. tarda

Intra‐ and interspecific characteristics of fish‐pathogenic Edwardsiella ictaluri, and E. tarda were determined by numerical analysis of gel electrophoresed protein profiles, fatty acid methyl esters (FAMEs) and immunoblotting. The 18 E. ictaluri isolates revealed a high degree of homogeneity (70% similarity or higher) in their protein profiles and 95% similarity in their FAME, while the nine E. tarda isolates revealed 30% similarity in their protein profiles and 95% similarity in their FAME. Immunoblots probed for antigenic epitopes with goat antiserum produced against E. ictaluri and E. tarda, respectively, revealed that E. ictaluri were more homogeneous compared with the E. tarda isolates. Overall, there was a considerable degree of relatedness between the two species. Our findings suggest that phenotypically E. ictaluri represents a clonal bacterial population structure compared with the less monomorphic E. tarda.     

Feed deprivation of channel catfish, Ictalurus punctatus (Rafinesque), influences organosomatic indices, chemical composition and susceptibility to Flavobacterium columnare

Withholding feed has been suggested as a strategy to manage infectious disease of channel catfish, Ictalurus punctatus (Rafinesque). In a previous study, we demonstrated that deprivation of feed for as little as 7 days reduced innate resistance of catfish to Flavobacterium columnare. This study was conducted to evaluate the effect of feeding regimens [no feeding (NF), fed once every other day to satiation (FEOD) and fed once daily to satiation (FD)] on organosomatic indices, physiological changes and susceptibility of channel catfish to F. columnare. Fish that were not fed for 2 and 4 weeks had a significant increase (P < 0.05) in gutted weight:-wet weight ratio and decrease in other organosomatic indices [gut index (GI), mesenteric fat index (MFI) and hepatosomatic index (HSI)]. Haematology was not effected by feeding regimen except at week 4, when a significantly higher haemoglobin level was observed in the NF fish. Serum protein did not differ at week 2, but the level at week 4 of the NF fish (35.91 mg mL(-1)) was significantly lower than that of the FD fish (41.77 mg mL(-1)). Significantly lower (P < 0.05) blood glucose (39.5 and 40.3 mg dL(-1)) and liver glycogen (1.7 and 1.8 mg g(-1)) were seen in the NF fish at weeks 2 and 4, respectively, as compared with blood glucose and liver glycogen levels of FD fish (67.5 and 92.8 mg dL(-1) and 46.5 and 52.6 mg g(-1) at weeks 2 and 4, respectively) and FEOD (82.8 and 85.5 mg dL(-1) and 45.1 and 51.4 mg g(-1) at weeks 2 and 4, respectively). Mortality in the NF fish caused by F. columnare (78%) was significantly higher (P < 0.05) than mortality in the FD and FEOD treatments (0.0 and 1.7%, respectively). Blood glucose and liver glycogen showed the same trend of low values for NF fish following challenge (week 6). Blood glucose, liver glycogen, GI and HSI are sensitive indicators for channel catfish deprived of feed (NF) for 4 weeks. Blood glucose and liver glycogen levels around 40 mg dL(-1) and 2 mg g(-1), respectively, are indicative of starvation in juvenile channel catfish. Moreover, NF fish were susceptible to F. columnare infection. Thus, it is suggested that in the absence of natural food, juvenile channel catfish should be fed at least once every other day to apparent satiation to maintain normal physiological function and improve resistance to F. columnare.     

Influence of Dietary Levels of Magnesium on Growth, Tissue Mineral Content, and Resistance of Channel Catfish Ictalurus punctatus Challenged with Edwardsiella ictaluri

Juvenile channel catfish were fed purified diets supplemented with magnesium (Mg) from Mg sulfate at levels of 0, 200, 400, 600, 800, and 1,000 mg/kg and 0, 200, 400, 600, and 800 mg/kg in two separate feeding studies. In study I, the effect of dietary levels of Mg on growth response, vertebral mineral content, and macrophage chemotaxis were evaluated. Study II had similar objectives except that whole body mineral content was measured, and resistance of channel catfish to Edwardsiella ictaluri challenge was also determined. Fish with an average weight of 10.89 g were stocked at a rate of 50 fish/110‐L aquarium (study I). In study II, fish with an average weight of 4.14 g were stocked at rates of 40 fish/110‐L aquarium. Prior to stocking, each batch of fish was acclimated to laboratory conditions and fed the basal diet for 2 wk. The concentration of Mg in rearing water was 1.8 mg/L. Each diet was fed to fish in quadruplicate and triplicate aquaria to apparent satiation for 10 wk for studies I and II, respectively. Fish fed the basal diet started to die as early as 3 d after the study began (17 d of feeding the diet without Mg supplementation). In both studies, weight gain, survival, and feed efficiency were lowest for fish fed the basal diet but increased with increasing dietary levels of Mg. However, the differences between the values of each of these parameters for fish fed diets containing supplemental Mg were not always significant. Magnesium‐deficiency signs observed were anorexia, sluggishness, convulsions, deformed snout, vertebral curvature, muscle flaccidity, and high mortality. Vertebral and whole body ash concentrations were high, but Mg content was low for fish fed the basal and the 200‐mg Mg diets. Bone Ca content did not differ among fish fed different diets (study I), but whole body Ca tended to increase for fish fed the basal diet, suggesting the possibility of calcification of soft tissues. Macrophage chemotaxis in the presence of exoantigen was highest for fish fed diets supplemented with Mg at 400 and 200 mgkg for studies I and II, respectively. When expressed in terms of chemotaxis index, however, maximum or near maximum value was observed at a dietary Mg level of 400 mg/kg. Thus, a dietary level of Mg of 400 mg/kg from Mg sulfate was required for optimum growth and survival, maintaining high tissue levels of Mg, prevention of muscle flaccidity and skeletal deformity, and stimulating macrophage chemotaxis. Dietary levels of Mg had no effect on the resistance of juvenile channel catfish to Edwarsiella. ictaluri challenge.     

Molecular cloning and sequencing of hemoglobin-β gene of channel catfish, Ictalurus punctatus Rafinesque

The hemoglobin-β gene of channel catfish, Ictalurus punctatus, was cloned and sequenced. Total RNA from head kidneys was isolated, reverse transcribed and amplified. The sequence of the channel catfish hemoglobin-β gene consists of 600 nucleotides. Analysis of the nucleotide sequence reveals one open reading frame and 5′- as well as 3′-untranslated regions. The open reading frame of the sequence potentially encodes 148 amino acids with a calculated molecular mass of 16.3 kDa. The pI and charge at pH 7.0 of the deduced hemoglobin-β protein were 7.28 and 0.47, respectively. Overall, 22 amino acid residues were conserved throughout the sequences, including His64 and His93, the sites for heme-binding. Unlike the counterpart of other common cultured fish such as Salmo salar, Oncorhynchus nerka, Oncorhynchus mykiss, Cyprinus carpio and Ctenopharyngodon idella, the hemoglobin-β of channel catfish did not have cysteine. The amino acid sequence of channel catfish hemoglobin-β shows 84% homology with that of Silurus asotus (both are in the order Siluriformes). However, comparison with those of other fish species shows homology ranging from 53 to 68%. Structural analysis by the 3D-PSSM program displays that channel catfish hemoglobin-β has eight α-helices, A–H.