Selected nondigestible carbohydrates and prebiotics support the growth of probiotic fish bacteria mono-cultures in vitro.

AIMS: To search for nondigestible but fermentable (NDF) carbohydrates and prebiotics with a potency to promote the growth of selected bacteria in vitro. METHODS AND RESULTS: The growth of three reference bacteria strains Bacillus subtilis LMG 7135(T), Carnobacterium piscicola LMG 9839, Lactobacillus plantarum LMG 9211 and one candidate probiotic bacteria Lactobacillus delbrueckii subsp. lactis was investigated over a minimum period of 48 h in the presence of beta-glucan, xylo-oligosaccharide, arabinoxylo-oligosaccharide, inulin, oligofructose and glucose. Besides the capability to grow on inulin and oligofructose containing media, a distinct high growth in beta-glucan based substrates and a low growth in (arabino)xylooligosaccharide containing media were evident for most bacteria tested. With the exception of B. subtilis and L. plantarum, other bacteria grew equally well or even better on different substrates than on glucose. The fermentation of studied carbohydrates by these micro-organisms was dominated by the production of acetic acid as the main short chain fatty acid. CONCLUSIONS: Selected bacteria are able to ferment and grow on NDF and prebiotic carbohydrates but in a substrate dependent manner. SIGNIFICANCE AND IMPACT OF THE STUDY: This study delivers a first screening of which NDF or prebiotic carbohydrates are the most promising for aquaculture feed supplementations.

Bioenergetics of fish spermatozoa during semen storage.

This mini-review focuses on changes in ATP and creatine phosphate concentrations in fish sperm under storage conditions. The storage of catfish sperm at 4 degrees C leads to ATP depletion and decreased sperm motility. The rate of intracellular ATP depletion can be diminished through the addition of energetic substrates to the sperm storage medium, with lactate + pyruvate being the most efficient substrates for maintaining ATP concentrations in catfish sperm. The decrease in ATP concentration is closely associated with increases in AMP and hypoxanthine content. In contrast to catfish sperm, carp sperm is able to maintain intracellular ATP concentration close to the physiological level during storage. Collectively, these results suggest that fish species differ in terms of the energy metabolism of their spermatozoa and that the semen storage medium must be carefully selected for a particular fish species so as to maintain the ATP concentration and adenylate energy charge close to physiological values as long as possible.

Quality control of refrigerated and cryopreserved semen using computer-assisted sperm analysis (CASA), viable staining and standardized fertilization in African catfish (Clarias gariepinus).

A new integrated approach including computer-assisted sperm analysis (CASA), viability staining and fertilization was used to study the quality of cryodiluents used in fish sperm cryopreservation. As an example the sperm quality of an African catfish, Clarias gariepinus (Burchell, 1822), was assessed by its fertilizing ability, motility and viability at day 0 (fresh), after 2 days' storage at 4 degreesC and after 2 days, 5 months and 10 months frozen at -196 degreesC using solutions containing dimethyl sulphoxide (DMSO) or glycerol as permeating cryoprotectants. Four of the best freezing solutions were used, namely, Steyn's extender (S1, S4) and Mounib's extender (M3, M4) associating 10% hen's egg yolk. Progressive sperm movement measured by CASA and expressed by the straight line velocity (VSL), the average path velocity (VAP) and the curvilinear velocity (VCL) was highly correlated with hatching rates obtained from fertilization using minimal sperm:egg ratios. After 2 days, the motility of spermatozoa frozen with DMSO and 10% egg yolk had deteriorated less than that of spermatozoa stored at 4 degreesC. Post-thaw hatching rates reflected the post-thaw sperm viability, which was cryodiluent dependent: 14.9+/-2.0% (S4), 17.0+/-4.2% (S1), 25.9+/-3.7% (M4) and 52.1+/-3.4% (M3) after 5 months of cryopreservation. The percent motility of 10-months-frozen spermatozoa was high in M3 (70.7+/-11.4%) and M4 (64.0+/-2.0%) cryoprotected sperm when measured between 5 and 20 sec after activation, but decreased rapidly to 24.3+/-8.3% (M3) and 23.0+/-9.0% (M4) between 21 and 35 sec after activation. Mounib's extender (M3, M4) provided the best cryoprotection to the spermatozoa for all post-thaw sperm quality measurements and at all freezing durations. Sperm motility was positively related to fertility. Our method will make it possible to develop even better extenders and cryoprotectants.

Partial and total fish meal replacement by agricultural products in the diets improve sperm quality in African catfish (Clarias gariepinus).

This study investigated the long-term effects of total and partial replacement of dietary fish meal (FM) by a mixture of agricultural products on sperm quality of African catfish Clarias gariepinus. Four isonitrogenous and isoenergetic diets were formulated containing graded levels of either 50% FM and maize meal (diet 1); 25% FM mixed with crude sunflower oil cake (SFOC) and bean meal (BM) (diet 2); 12.5% FM mixed with sunflower oil cake, BM and ground nut oil cake (GOC) (diet 3) and 0% FM mixed with de-hulled sunflower oil cake (SFOCD), BM and ground nut oil cake (diet 4). Gonadosomatic index (GSI), sperm quality, plasma sex steroids (11-keto testosterone [11-KT]; testosterone [T]; estradiol-17beta [E2]) were evaluated on 10 to 24 fish fed on each diet. Sperm quality was assessed using computer-assisted sperm analysis (CASA). Total replacement of fish meal by plant products markedly increased sperm volume, spermatocrit, spermatozoa integrity, and sperm motility. Fish fed diet 3 (12.5% fish meal) provided intermediate results on sperm quality whereas the lowest values were obtained in fish fed diets 1 and 2. In fish fed 0% fish meal (diet 4), androgen levels were higher and estrogen levels were lower than in fish fed fish meal diets. Based on dietary lipid and fatty acid analyses, these results suggest a positive impact of short chain n-6 fatty acids on androgen synthesis and sperm quality. In conclusion, a combination of ground nut oil cake, bean meal and sunflower oil cake (preferably when the sunflower is dehulled) in African catfish diet improves the sperm quality.

Effect of tributyltin on adenylate content and enzyme activities of teleost sperm: a biochemical approach to study the mechanisms of toxicant reduced spermatozoa motility.

The effects of tributyltin (TBT) on the energy metabolism and motility of fish spermatozoa were investigated in vitro in African catfish and common carp. A significant (P<0.05) decrease of the duration and the intensity of motility was observed in catfish spermatozoa exposed to 0.27 microg/l TBT for 24 h. Exposure of catfish spermatozoa to 2.7-27 microg/l TBT caused an instant decrease in ATP content. In the presence of 27 microg/l TBT approximately 55% of the initial ATP concentration in catfish semen was lost after 60 min incubation while AMP concentrations increased and the total adenine nucleotide (TAN) pool remained unchanged. The reduction in sperm ATP levels could not be attributed to cell death since viability decreased only slightly over the period of exposure. In carp by contrast, none of the adenylates concentrations studied (ATP, ADP and AMP) were affected by TBT exposure at any experimental condition. However, carp sperm motility was significantly reduced by exposure to 2.7 microg/l TBT. Among the enzymes investigated only lactate dehydrogenase (LDH) in catfish sperm was significantly (P<0.01) affected by 27 microg/l TBT treatment with a reduction in activity of approximately 75%. Compared with carp sperm before TBT exposure, that of catfish had lower adenylate contents and overall lower enzymatic activities; this explains its slower sperm velocity and shorter duration of movement as measured by computer assisted sperm analysis (CASA). The present in vitro study shows that catfish spermatozoa are more sensitive to TBT exposure (and probably to other toxicants) than those of carp.

Computer-assisted sperm analysis (CASA) as a tool for monitoring sperm quality in fish.

The use of motility as a measure of sperm quality in fish is reviewed. Computer assisted sperm analysis (CASA) provides a simple and rapid quantitative assessment of the quality of fish sperm and may predict its ability to fertilize eggs. It has been used to: monitor the effects of heavy metal pollutants, such as mercury and tributyltin, on sperm quality; to select broodstock; to improve the efficiency of cryopreservation and storage; and to optimise conditions for fertilisation. In combination with CASA, morphological measurements can be used to determine the causes of reduced sperm motility. Technical details for the use of CASA are described.