Carnobacterium maltaromaticum vs. Vibrio (Listonella) anguillarum in the midgut of Atlantic cod (Gadus morhua L.): an ex vivo study

The present investigation addresses whether the midgut (MG) of Atlantic cod (Gadus morhua L.) is an infection route for Vibrio (Listonella) anguillarum serotype 02 β and if Carnobacterium maltaromaticum, a probiotic bacterium, can out‐compete the pathogen and modulate the autochthonous MG microbiota. This was investigated by using an ex vivo method the intestinal sac, utilized previously in studies on Atlantic cod and Atlantic salmon (Salmo salar L.). Exposure of the MG to V. (L.) anguillarum did not reveal any cell damage indicating that the MG does not appear to be an infection route for V. (L.) anguillarum in healthy Atlantic cod. This finding together with previous observations on Atlantic cod and Atlantic salmon indicate that intestine as an infection route might vary between these two species. When the MG was exposed to C. maltaromaticum, no cell damage or cellular disruptions were observed. As budding from the apices of microvilli was observed in all treatments exposed to bacteria, we suggest that budding might be involved in the primary barrier against bacterial infection. However, to clarify this hypothesis, further studies are needed. Exposure of the MG to the probiotics and pathogenic bacteria indicated that C. maltaromaticum, to some extent, is able to out‐compete V. (L.) anguillarum but the topic merits further investigation. Analysis of the MG microbiota after sterile saline solution and bacterial exposure indicates that bacteria related to Staphylococcus sciuri belong to the autochthonous gut microbiota in Atlantic cod.     

Antagonistic activity of bacterial isolates from intestinal microbiota of Atlantic cod, Gadus morhua, and an investigation of their immunomodulatory capabilities

In an effort to identify potential probionts, four bacterial strains obtained from the intestinal tract of wild-caught Atlantic cod, Gadus morhua , were tested for their antagonistic activity against Vibrio anguillarum and Aeromonas salmonicida , two of the most common bacterial pathogens in cod aquaculture, using a well-diffusion agar assay at two incubation temperatures: 13 and 20 °C. The tested bacteria exhibited dissimilarity in their inhibitory action against the target pathogens – an enhanced activity was particularly observed for strains GP11 and GS11 at the highest incubation temperature. Based on the 16S ribosomal DNA gene sequence analysis, the strains showed high similarity to Psychrobacter sp. (strain GP11), Shewanella sp. (GS11), Photobacterium sp. (GP31) and Vibrio sp. (GV11). The incubation of Atlantic cod head kidney cells with the heat-inactivated probiotic strains resulted in the differential expression of immune-response genes that are related to bacterial defence and inflammation. Thus, the gut-derived bacterial strains have probiotic potential and their possible immunomodulatory capabilities could be determined under in vitro conditions.     

芽孢杆菌对凡纳对虾生长和消化酶活性的影响

选用豆粕、鱼粉、植物油、淀粉等原料配制基础料为对照组,在基础饲料中分别添加0.5％、1.0％、3.0％、5.0％芽孢杆菌(Bacillus subtilis)制剂为实验组。实验结果显示,凡纳对虾(Pennaus-vannamei)生长的2个阶段,实验组γSGR均大于对照组;生长第1阶段,组1与组2差异显著(P<0.05),与其余3组差异不显著(P>0.05),但各实验组之间差异均不明显(P>0.05);生长第2阶段,组1与组2之间γSGR差异极显著(P<0.01),与其余3组差异不明显(P>0.05),但各实验组间:组2与组4γSGR差异显著(P<0.05),而与组5γSGR差异极显著(P<0.01);对虾生长2个阶段的成活率实验组均略高于对照组,第1阶段的成活率明显高于第2阶段;芽孢杆菌对凡纳对虾2个生长阶段消化酶活性的影响表现出一定的相似性:实验组蛋白酶活性较淀粉酶、脂肪酶和纤维素酶高,且明显高于对照组,但各实验组间变化不明显。表明芽孢杆菌可以提高凡纳对虾消化酶活性和成活率,利于养分的消化吸收,促进生长。     

Screening of gastrointestinal bacteria of Indian major carps for a candidate probiotic species for aquaculture practices

Probotic selection could be detrimental for the success of aquaculture practices. While, in most of the cases emphasis is given to the production of antibacterial compounds, and in vitro inhibitory activities of a bacterium for choosing a probiotic strain, however, other criteria like non‐antibiotic resistant, non‐haemolytic nature and non‐pathogenic potentiality of probiotic strain are often overlooked. We have screened gastrointestinal bacteria isolated from all the three species of Indian major carps viz., Labeo rohita, Catla catla and Cirrhinus mrigala for a suitable probiotic. Bacteria like Aeromonas (Aeromonas species, A. hydrophila), Micrococcus, Corynebacterium, Plesiomonas, Bacillus (Bacillus species, B. subtilis) and Pseudomonas (Pseudomonas species, P. aeruginosa) exhibited antibacterial activities against different pathogens. Among these, B. subtilis was found to grow at a wider range of temperatures, pH and salt concentrations and was non‐haemolytic, non‐antibiotic resistant, non‐pathogenic and inhibited all the tested primary and secondary pathogens. Dietary supplementation of the bacteria at 10⁸ CFU g^?1 feed also enhanced the growth of Indian major carp, L. rohita. Based upon these characteristics, we aimed to develop a suitable probiotic for aquaculture practices.     

Growth and protein utilization in Atlantic salmon (Salmo salar L.) given a protease inhibitor in the diet

In a series of experiments the effects of dietary protease inhibitor inclusions on growth and amino acid absorption rate were investigated in Atlantic salmon ( Salmo salar L.). An optimal inclusion of inhibitor was found by conducting dose–response studies with specific growth rate as the main response variable. No negative effects on feed conversion ratio or nitrogen digestibility were observed at this level. In protein sources studies the addition of potato protease inhibitor to a fishmeal based diet increased specific growth rate by 14%. When a proportion of the fishmeal was replaced with hydrolysed protein (fish silage) the addition of the inhibitor resulted in a 31% increase in specific growth rate. Absorption of amino acids from the gastrointestinal tract into the blood was investigated in two experiments using ¹⁴C-labelled algal ( Synechoccus leopoliensis ) protein. The absorption pattern of ¹⁴C-labelled amino acids was altered by adding potato protease inhibitor when the algal protein was supplied in intact form, but not when the algal protein was hydrolysed. The absorption of amino acids from a hydrolysed protein was significantly faster than from intact protein. The enzymatic activity of pepsin in the stomach and of trypsin, chymotrypsin, carboxypeptidases A and B in the different segments of the intestine changed significantly with increasing inclusion of potato protease inhibitors in the diet.     

Environmental influences on melatonin secretion in Atlantic cod (Gadus morhua L.) and their relevance to commercial culture

This study investigated the effects of photoperiod and temperature on plasma melatonin secretion in Atlantic cod (Gadus morhua L.). Initial work confirmed the presence of a diel profile of melatonin synthesis, with elevated levels during the dark phase. Unusually for fish, the peak in plasma melatonin occurred towards the end of the dark phase, which is indicative of a type `A' melatonin profile. When exposed to 60 hours of continuous darkness a clear endogenous rhythm of melatonin synthesis was observed, which continued for 4 cycles with a periodicity which, approximated to 24 h. When acclimated to varying temperatures (4, 8, 12 or 16 °C) no variation in melatonin production was seen, however, body size appeared to be an important influence, with the smallest fish exhibiting significantly higher levels of dark phase melatonin. Finally, the application of additional night-time illumination to cod maintained in sea cages i.e. without blackout, did not significantly reduce dark phase plasma melatonin, suggesting that cod are less sensitive to photoperiod manipulation in cages than salmonids.     

Characterisation of the microbiota associated with intestine of Atlantic cod (Gadus morhua L.): The effect of fish meal, standard soybean meal and a bioprocessed soybean meal

Populations of aerobic heterotrophic bacteria present in the gastrointestinal (GI) tract of healthy Atlantic cod (Gadus morhua L.) fed three different diets, fish meal, standard or a bioprocessed soybean meal (BPSBM), were estimated using the dilution plate technique. A total of 944 isolates were characterised by biochemical and physiological properties and 425 isolates were identified further by sequencing the 16S rRNA genes. Our results showed that gut microbiota were affected by dietary manipulation. The GI tract of fish fed fish meal was dominated by Gram-positive bacteria of the genera Brochothrix and Carnobacterium. The Gram-negative bacteria Chryseobacterium spp. and Psychrobacter glacincola, and Gram-positive bacteria belonging to Carnobacterium, dominated in the digestive tract of fish fed soybean meal. In contrast to these results, genus Psychrobacter dominated in the GI tract when fish were fed BPSBM.Until recently, it was generally suggested that the gut microbiota of fish were less diverse than in homoeothermic animals. However, the present study identified several “new” bacterial species isolated from the alimentary tract of Atlantic cod. These “new” bacterial species are not normally isolated from the GI tract of fish. Based on our finding we suggest that the GI tract microbiota of fish might not be as simple as believed.Antagonistic activity of carnobacteria regarding inhibition of growth of two fish pathogens (Aeromonas salmonicida ssp. salmonicida and Vibrio anguillarum) was observed. However, some difference in the antibacterial activity of Carnobacterium spp. was observed. Whether this antagonistic activity has any effect in challenge studies will be discussed, especially in relation to the finding that the digestive tract is one of the major infection routes in fish.     

Efficacy of egg surface disinfectants in captive spawning Atlantic cod Gadus morhua L. and haddock Melanogrammus aeglefinus L.

In an effort to optimize the efficiency of high‐density incubation of Atlantic cod Gadus morhua and haddock Melanogrammus aeglefinus eggs, the per cent hatch of eggs treated with four disinfectants (3% hydrogen peroxide, 1% polyvinylpyrrolidone iodine, 0.1% sodium hypochlorite and a 0.005% antibiotic solution – penicillin/streptomycin) was compared in both species. The per cent hatch of eggs of each species was greatest after a 24 h treatment with the antibiotic solution. The hatching success of eggs treated within the different disinfectant treatments depended upon the embryonic developmental stage in both species. The sodium hypochlorite treatment had the lowest % coverage of colony growth after disinfected haddock eggs were plated onto sterile agar media, highest survival rates to the end of the embryonic period, but the lowest per cent hatch.     

Investigation of the temporal effects of spawning season and maternal and paternal differences on egg quality in Atlantic cod Gadus morhua L. broodstock

A better understanding of the parameters affecting egg quality and larval survival is of importance for continued development of cod broodstock and efficient husbandry practices. Decision tree analysis (DTA) was applied to analyse 3 years of egg quality data in an effort to extract the most important variables (i.e. predictors) in explaining differences in egg quality. The effect of three predictors (spawning time, maternal and paternal differences) has been studied on early cleavage pattern parameters, egg diameters, fertilization and hatching rates and has shown that females are the dominant variable and that time has a limited and inconsistent impact on the data. When using maternal, paternal differences and batch number (instead of spawning time) as predictors, the results confirm that no particular relationship is found between batch order (i.e. order in time) and egg quality. Moreover, batches with a higher egg quality show a consistency in the parameters assessed (i.e. batches with higher rates of normality in any parameter tend to be normal for other parameters). This is confirmed by the significant correlations found between cleavage parameters. Our results highlight that spawning time is of less importance than female parent contribution in ensuring high rates of fertilization and larval hatch, and maximizing general egg quality.     

Does different tank bottom colour affect the growth, survival and foraging behaviour of Atlantic cod (Gadus morhua) larvae?

We examined the foraging behaviour, growth and survival of Atlantic cod (Gadus morhua) larvae in two different colours of tank bottom; either beige (light hereafter) or black (dark hereafter) bottomed tanks with black walls. Results showed no significant differences in the growth, foraging behaviour, or survival of Atlantic cod larvae in response to tank bottom colour indicating that larvae could be reared in lighter bottom tanks without any detrimental effect to the larvae. Using a light bottom tank is also of greater benefit to the culturist as they provide better contrast to monitor the behavioural and morphological development of the larvae.     

Is the effect of the NAO on North‐east Arctic cod,Gadus morhua,recruitment stock‐dependent?

There is considerable interest in understanding the effect of the North Atlantic Oscillation (NAO) on recruitment to North Atlantic cod stocks. An earlier analysis of data for the North‐east Arctic cod stock showed that, while there is an empirical relationship between recruitment and the NAO over the period 1973–96, this relationship was absent over the period 1946–72. It has recently been suggested that the effect of the NAO on recruitment depends on the size of the spawning stock. Here, the possibility that this explains the earlier result for North‐east Arctic cod is tested and rejected.     

野生与养殖哲罗鱼消化系统及消化酶的比较研究

采用组织学方法对野生与养殖哲罗鱼（Hucho taimen Pallas）消化系统的形态及组织结构进行比较研究,采用聚丙烯酰胺凝胶电泳方法比较了两个群体哲罗鱼3种消化酶的相对含量。结果表明,野生与养殖哲罗鱼的消化系统在形态上存在一定差异,在组织结构上无明显差异。两个群体哲罗鱼在食道与胃酯酶相对含量上差异不显著（P〉0．05）,在肝脏与幽门盲囊酯酶相对含量上差异显著（P〈0．05）,在肠道酯酶的相对含量上差异极为显著（P〈0．01）。两个群体哲罗鱼在食道与胃淀粉酶的相对含量上差异不显著（P〉0．05）,在肝脏与肠道淀粉酶的相对含量上差异显著（P〈0．05）,在幽门盲囊淀粉酶的相对含量上差异极为显著（P〈0．01）。哲罗鱼的蛋白酶表达存在一定的pH依赖性,在体外酸性条件下（pH3）,两个群体哲罗鱼只有食道与胃有蛋白酶表达;在体外碱性条件下（pH9）,两个群体哲罗鱼只有幽门盲囊与肠道有蛋白酶表达;两种pH条件下,两个群体哲罗鱼各组织中蛋白酶的相对含量差异均不显著（P〉0．05）。两个群体哲罗鱼的消化系统结构及消化酶含量所表现出的差异与其生活环境及食物有关。[中国水产科学,2008,15（2）：330-336]