Eugène Rurangwa

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

Abstract 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 fertilise 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.

Use of computer assisted sperm analysis (CASA) for monitoring the effects of pollution on sperm quality of fish; application to the effects of heavy metals☆

Abstract Computer assisted sperm analysis (CASA) has been used to analyse the effects of cadmium and zinc on the motility of fish sperm treated either for 24 h after partial dilution in an extender, or during the final dilution step on the microscope stage during video recording. The progressive motility of catfish sperm, which was not affected by storage for 24 h, was decreased after exposure to 100 ppm cadmium or 2000 ppm zinc in extender for 24 h. Only 1000 ppm cadmium or 2000 ppm zinc had any effect on motility when added to sperm at the final dilution stage. These results suggest that concentrations of cadmium which have been found to be present in fish gonads as a result of bioaccumulation from the water or diet may decrease the motility of sperm during maturation or storage in the testis. Cadmium toxicity was significantly decreased in the presence of an equal amount of zinc, suggesting an equilibrium between zinc and cadmium at binding sites in the sperm. Computer assisted sperm analysis is shown for the first time to provide a rapid and quantitative measure of the effects of environmental pollutants on the quality of fish sperm. It is suggested that the methodology described may form the basis for a standard toxicity test to assess the potential hazards of environmental pollutants on reproduction.

Prebiotic effects of arabinoxylan oligosaccharides on juvenile Siberian sturgeon (Acipenser baerii) with emphasis on the modulation of the gut microbiota using 454 pyrosequencing

The potential of a novel class of prebiotics, arabinoxylan oligosaccharides (AXOS), was investigated on growth performance and gut microbiota of juvenile Acipenser baerii. Two independent feeding trials of 10 or 12 weeks were performed with basal diets supplemented with 2% or 4% AXOS-32-0.30 (trial 1) and 2% AXOS-32-0.30 or AXOS-3-0.25 (trial 2), respectively. Growth performance was improved by feeding 2% AXOS-32-0.30 in both trials, although not significantly. Microbial community profiles were determined using 454-pyrosequencing with barcoded primers targeting the V3 region of the 16S rRNA gene. AXOS significantly affected the relative abundance of bacteria at the phylum, family, genus and species level. The consumption of 2% AXOS-32-0.30 increased the relative abundance of Eubacteriaceae, Clostridiaceae, Streptococcaceae and Lactobacillaceae, while the abundance of Bacillaceae was greater in response to 4% AXOS-32-0.30 and 2% AXOS-3-0.25. The abundance of Lactobacillus spp. and Lactococcus lactis was greater after 2% AXOS-32-0.30 intake. Redundancy analysis showed a distinct and significant clustering of the gut microbiota of individuals consuming an AXOS diet. In both trials, concentration of acetate, butyrate and total short-chain fatty acids (SCFAs) increased in fish fed 2% AXOS-32-0.30. Our data demonstrate a shift in the hindgut microbiome of fish consuming different preparation of AXOS, with potential application as prebiotics.

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.

Effects of dietary arabinoxylan-oligosaccharides (AXOS) and endogenous probiotics on the growth performance, non-specific immunity and gut microbiota of juvenile Siberian sturgeon (Acipenser baerii)

We investigated the effects of administration of putative endogenous probiotics Lactococcus lactis spp. lactis or Bacillus circulans, alone and in combination with arabinoxylan-oligosaccharides (AXOS), a new class of candidate prebiotics, in juvenile Siberian sturgeon (Acipenser baerii). Eight experimental diets were tested: basal diet (Diet 1), basal diet supplemented with 2% AXOS (Diet 2), or L. lactis ST G81 (Diet 3), L. lactis ST G45 (Diet 4), B. circulans ST M53 (Diet 5), L. lactis ST G81 + 2% AXOS (Diet 6), L. lactis ST G45 + 2% AXOS (Diet 7), B. circulans ST M53 + 2% AXOS (Diet 8). After four weeks, growth performance and feed conversion ratio significantly improved in fish fed diet 7. Innate immune responses of fish were boosted with both AXOS and probiotic diets, however synergistic effects of AXOS and probiotic diets were only observed for phagocytic and alternative complement activity. Phagocytic and respiratory burst activity of fish macrophage increased in fish fed diet 2 and 7, while humoral immune responses only increased in fish fed diet 7. Pyrosequencing analysis (16S rDNA) of the hindgut microbiota demonstrated that AXOS improved the colonization or/and growth capacity of L. lactis, as a higher relative abundance of L. lactis was observed in fish receiving diet 7. However, no observable colonization of B. circulans was found in the hindgut of fish fed diet 5 or 8, containing this bacterium. The dietary L. lactis ST G45 + 2% AXOS caused significant alterations in the intestinal microbiota by significantly decreasing in bacterial diversity, demonstrated by the fall in richness and Shannon diversity, and improved growth performance and boosted immune responses of Siberian sturgeon.

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.

Long-term exposure to environmentally relevant doses of methyl-tert-butyl ether causes significant reproductive dysfunction in the zebrafish (Danio rerio).

Methyl-tert-butyl ether (MTBE), an anthropogenic chemical used as a gasoline additive, is being detected at an increasing frequency in the environment. The acute lethal concentration that kills 50% of the fish test population and the chronic effects of exposure to MTBE were investigated in the zebrafish (Danio rerio). Chronic exposure over three weeks to effective MTBE conceritrations as low as 0.11 mg/L induced a significant increase in the vitellogenin concentration of male fish. The impact of a chronic, eight-week exposure at effective concentrations ranging from 0.44 to 220 mg/L had no significant effect on fecundity, fertilization, or hatch rate but highly significant impacts on sperm motility. Spermatozoa of all MTBE-exposure groups showed a significantly lower straight-line velocity and lower average path velocity compared to those of the nonexposed group. These results suggest that chronic exposure to MTBE negatively affects fish sperm motility at concentrations that are environmentally relevant and several orders of magnitude lower than concentrations inducing acute effects.

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.

Fermentation of Arabinoxylan-Oligosaccharides, Oligofructose and their Monomeric Sugars by Hindgut Bacteria from Siberian Sturgeon and African Catfish in Batch Culture in vitro

The in vitro fermentation of two Non-Digestible Oligosaccharide (NDO) preparations, Arabinoxylan- Oligosaccharides (AXOS) and Oligofructose (OF), and their respective monomeric sugars, xylose and fructose, were investigated by hindgut microbiota of two major aquaculture fish species, Siberian sturgeon (Acipenser baerii) and African catfish (Clarias gariepinus). Inocula from the hindgut of both fish species were incubated for 48 h in bottles containing 1.0% of one of four substrates, i.e. AXOS, OF, xylose or fructose. Amounts and profiles of produced Short-Chain Fatty Acids (SCFAs) differed between the two fish species and substrates. The hindgut microbiota of Siberian sturgeon has a higher fermentation capacity than the microbiota from African catfish. Xylose was much easier fermented than AXOS by microbiota from Siberian sturgeon whereas OF was quicker fermented than fructose with African catfish inoculum. The SCFAs were dominated by acetic acid for both fish species and for all substrates. Fermentation of OF and fructose by hindgut microbiota of Siberian sturgeon also yielded high amounts of butyric and branched-chain fatty acids after 48 h incubation. Results of this study suggest that AXOS, OF, and their monomeric sugars have an impact on microbial fermentation activity of hindgut microbiota from Siberian sturgeon and African catfish in a substrate and species dependent manner.