Luiz Carlos Kreutz

Nursery rearing of jundiá, Rhamdia quelen (Quoy & Gaimard) in cages: cage type, stocking density and stress response to confinement

Abstract The use of net cages allows the exploration of some water bodies, without usual aquacultural techniques for earthen ponds. Several advantages are proposed for the use of cages, however some disadvantages, as the higher possibility of stress and diseases are known. Growth parameters, survival rate and stress response of hatchery-bred Jundia ( Rhamdia quelen , Quoy & Gaimard) fry reared at different cage types and densities in net cages suspended in pond were evaluated. The first experiment compared fry reared at the circular and cubic-shaped cages at a density of 100 fry/m 3 . Fish held in cubic cages were heavier than the fish held in circular cages, presented higher weight gain and daily weight gain and a better food conversion. The survival rates were similar between fish raised in both types of cages. In the second experiment, cubic cages were stocked with 100, 200 and 300 fry/m 3 . Fish held in cages with lower density were heavier than the ones held at higher densities, presented higher weight gain and daily weight gain. Food conversion and survival rates were similar between different fish densities. According to the results presented in this study, the growth of jundia in cubic-shaped cages was density dependent. In terms of weight parameters, the most effective stocking density of jundia was 100 fry/m 3 that reaches 63.74±3.69 g of body weight. However, if the desirable weight was from 30 to 40 g, the density of 300 fry/m 3 is most effective because the fingerling production increases three times. In experiment 3, in all sample days, the serum cortisol levels were higher than the pre-stock and basal levels. The 80 days of cage confinement were not enough for adaptation of fingerlings to cage environment. The maintenance of high cortisol concentrations after 80 days of experimental period suggests the presence of typical chronic stress response that might have some detrimental effects over fish growth. Taken together, the data presented herein suggests that the improvement of rearing conditions (e.g. water quality, food quality, cage design, culture management, etc.) capable to minimize the stress response, may affect positively the growth performance in jundia during the nursery period.

Assessment of oxidative stress in Rhamdia quelen exposed to agrichemicals.

Due to the proximity of crop and fish culture areas, some agrichemicals that could be harmful for fish could enter into fishponds by different ways, such as by leaching through rain. Rhamdia quelen (Teleostei) were exposed to sublethal concentrations of methyl parathion (MP), a glyphosate based herbicide (Gly), and tebuconazole (Teb). The liver of R. quelen exposed to MP and Teb showed enhanced levels of thiobarbituric acid reactive substances (TBARS), higher than in the control fish (56% and 59%, respectively). In contrast, Gly did not alter the TBARS generation. The protein carbonyl content increased only in fish exposed to Teb. Fish exposed to the three agrichemicals showed a significant decrease of catalase activity (52%, 48%, and 67%, respectively) and increased glutathione-S-transferase (57%, 46%, and 160%, respectively) activity. Fish exposed to MP, Gly, and Teb showed higher reduced glutathione (151%, 472%, and 130%, respectively, when compared with the control levels) and ascorbic acid concentrations (121%, 102%, and 184%, respectively),while the non-protein thiol content increased only in R. quelen exposed to tebuconazole. Fish exposed to MP and Teb showed several pathological changes in the liver, including hepatocyte degeneration and bile stagnation. The present work reports for the first time the toxicity of the pesticide MP and the fungicide Teb in R. quelen, and as in other works, suggests the relatively lower liver toxicity of Gly for fish. The data presented herein demonstrate that sublethal concentrations of MP and Teb cause changes in oxidative stress parameters as well as hepatic cell injuries in R. quelen, and that these parameters have the potential to be developed as bioindicators of exposure to these agrichemicals.

Chronic exposure to sub-lethal concentration of a glyphosate-based herbicide alters hormone profiles and affects reproduction of female Jundiá (Rhamdia quelen).

This work was carried out to verify the effect of a glyphosate-based herbicide on Jundiá hormones (cortisol, 17β-estradiol and testosterone), oocyte and swim-up fry production. Earthen ponds containing Jundiá females were contaminated with glyphosate (3.6mg/L); blood samples were collected from eight females from each treatment immediately before, or at 1, 10, 20, 30 and 40 days following contamination. A typical post-stress rise in cortisol levels was observed at the 20th and 40th days following exposure to glyphosate. At the 40th day, 17β-estradiol was decreased in the exposed females. A similar number of oocytes were stripped out from females from both groups; however, a lower number of viable swim-up fry were obtained from the herbicide exposed females, which also had a higher liver-somatic index (LSI). The results indicate that the presence of glyphosate in water was deleterious to Rhamdia quelen reproduction, altering steroid profiles and egg viability.

Acute toxicity test of agricultural pesticides on silver catfish (Rhamdia quelen) fingerlings

Toxicity risks of agricultural pesticides to fishes are pivotal. Currently, many questions remain unsolved regarding to the toxicity of commonly used pesticides to silver catfish (Rhamdia quelen), a South American catfish. The present studies have been designed to investigate the acute toxicity and the lethal concentration (LC50) of four herbicides, two fungicides and two insecticides to silver catfish fingerlings. All experiments were carried out in triplicates, in a static bioassay system, using commercially available pesticides. The data was analyzed through the Trimmed Spearman-Karber method available from the Environmental Protection Agency. The 96hLC50 and 95% lower and upper confidence limits, respectively, for the following pesticides were determined: glyphosate (7.3mg L-1; 6.5-8.3), atrazine (10.2mg L-1; 9.1-11.5), atrazine+simazine (10.5mg L-1; 8.9-12.4), mesotrione (532.0mg L-1; 476.5-594), tebuconazole (5.3mg L-1; 4.9-5.7), methylparathion (4.8mg L-1; 4.3-5.3), strobulurin and triazol (9.9mg L-1; 8.7-11.2). Diflubenzuron was also tested and caused no fish mortality up to 1g L-1. The toxic concentration of these pesticides to silver catfish fingerlings fell above the concentration used for application in the field and, except following accidental application or misplacing of empty recipients, it should not cause fish mortality. Nonetheless, the data obtained will be useful to study the long-term effect of these products on the hematological, biochemical, hormonal and immunological parameters of silver catfish and related fish species in South Brazil.

Altered hematological and immunological parameters in silver catfish (Rhamdia quelen) following short term exposure to sublethal concentration of glyphosate

Using agrichemicals to control unwanted species has become a necessary and common worldwide practice to improve crop production. Although most currently used agrichemicals are considered relatively safe, continuous usage contributes for soil and water contamination and collateral toxic effects on aquatic species. Few studies correlated the presence of agrichemicals on fish blood cells and natural immune system. Thus, in this study, silver catfish (Rhamdia quelen) were exposed to sublethal concentrations (10% of the LC(50-96 h)) of a glyphosate based herbicide and hematological and natural immune system parameters were evaluated. Silver catfish fingerlings exposed to glyphosate for 96 h had a significant reduction on blood erythrocytes, thrombocytes, lymphocytes and total leukocytes in contrast to a significant increase in the number of immature circulating cells. The effect of glyphosate on natural immune system was evaluated after 24h or 10 days exposure by measuring the phagocytic index of coelomic cells, and lysozyme, total peroxidase, bacteria agglutination, bactericidal activity and natural complement hemolytic activity in the serum of fingerlings. A significant reduction on phagocytic index, serum bacteria agglutination and total peroxidase was observed only after 24h exposure to glyphosate. In contrast, fingerlings exposed to glyphosate for 10 days had a significant lower serum bacteria agglutination and lysozyme activity. Glyphosate had no effect on serum bactericidal and complement natural hemolytic activity after 24h or 10 days exposure. Nonetheless, the information obtained in this study indicates that glyphosate contaminated water contributes to alter blood cells parameters and to reduce the activity of natural immune components important to mediate fish resistance to infecting microorganisms.

Cortisol response to acute stress in jundiá Rhamdia quelen acutely exposed to sub-lethal concentrations of agrichemicals

Exposure to agrichemicals can have deleterious effects on fish, such as disruption of the hypothalamus-pituitary-inter-renal axis (HPI) that could impair the ability of fish to respond to stressors. In this study, fingerlings of the teleost jundiá (Rhamdia quelen) were used to investigate the effects of the commonly used agrichemicals on the fish response to stress. Five common agrichemicals were tested: the fungicide - tebuconazole, the insecticide - methyl-parathion, and the herbicides - atrazine, atrazine+simazine, and glyphosate. Control fishes were not exposed to agrichemicals and standard stressors. In treatments 2-4, the fishes were exposed to sub-lethal concentrations (16.6%, 33.3%, and 50% of the LC(50)) of each agrichemical for 96 h, and at the end of this period, were subjected to an acute stress-handling stimulus by chasing them with a pen net. In treatments 5-7 (16.6%, 33.3%, and 50% of the LC(50)), the fishes were exposed to the same concentrations of the agrichemicals without stress stimulus. Treatment 8 consisted of jundiás not exposed to agrichemicals, but was subjected to an acute stress-handling stimulus. Jundiás exposed to methyl-parathion, atrazine+simazine, and glyphosate presented a decreased capacity in exhibiting an adequate response to cope with stress and in maintaining the homeostasis, with cortisol level lower than that in the control fish (P<0.01). In conclusion, the results of this study clearly demonstrate that the acute exposure to sub-lethal concentrations of methyl-parathion, atrazine+simazine, and glyphosate exert a deleterious effect on the cortisol response to an additional acute stressor in the jundiá fingerlings.

Isolation of bovine herpesvirus 1 from preputial swabs and semen of bulls with balanoposthitis.

gone REA pattern alterations following only 1 passage through the host species. Some of the atypical WT-PRV identified in this study may have arisen in a similar manner. The similarities between the REA pattern of No. 89 and those of Nos. 90-92 suggests a possible iatrogenic V-PRV origin for these viruses. There is 1 report from Germany of a bovine origin PRV isolate with a pattern of split MAb reactivity similar to that found with isolate No. 95. In that study, the molecular basis of the unusual MAb reactivity was associated with a truncated form of gI. The simple phenotypic screening system described here will help identify those unusual isolates requiring further study to determine their possible origins and veterinary diagnostic significance.

Exposure to sublethal concentration of glyphosate or atrazine-based herbicides alters the phagocytic function and increases the susceptibility of silver catfish fingerlings (Rhamdia quelen) to Aeromonas hydrophila challenge.

The resistance of fish to microorganisms challenge depends mainly on the efficacy of the immune response. Most studies on the natural immune response of fish have focused on the effect of diets and immunostimulants. Few studies correlated the presence of commonly used agrichemical and susceptibility to infection by aquatic microorganism. Thus, this study aimed to investigate the effect of glyphosate and atrazine-based herbicides on immune cell phagocytosis and susceptibility of silver catfish to Aeromonas hydrophila infection. Following exposure to sublethal concentrations of glyphosate or atrazine (10% of the LC(50-96 h)), a significant decrease in the number of intracelomatic cells and phagocytic index could be observed. In addition, silver catfish fingerlings exposed to glyphosate or atrazine were more susceptible to intracelomatic challenge with pathogenic A. hydrophila. Thus, the presence of these herbicides on the water alters the natural immune response to bacterial and possibly to other aquatic microorganism.

Responsiveness of the interrenal tissue of Jundiá (Rhamdia quelen) to an in vivo ACTH test following acute exposure to sublethal concentrations of agrichemicals.

As in many aquatic environments, pollution is a widespread problem in Southern Brazil. In our previous work, we demonstrated that sublethal contamination with some agrichemicals impairs the capacity of fishes to elevate cortisol levels in response to an additional acute stressor. In earlier experiments, the experimental design did not allow us to conclude where this effect occurs. In the present work, we used the adrenocorticotropic hormone (ACTH) challenge test to help us identify if the impairment occur in the interrenal tissue. For this purpose, five experiments were conducted, each with one specific agrichemical (methyl-parathion, atrazine+simazine, atrazine, tebuconazole, and glyphosate) in sublethal concentrations of 16.6% of the LC(50-96h), as previously determined. Fish were subjected to the ACTH challenge test protocol as follows: group 1, were non-injected and maintained as the specific control group; group 2 received an injection of the vehicle alone (the saline group); and group 3 receive an injection of ACTH. One hour later, blood samples were taken from the caudal plexus, using sterile syringes. In all specific control groups, the injection of ACTH induced a strong rise in plasma cortisol, compared with the fish injected only with the vehicle and the non-injected group. Fish exposed to methyl-parathion and tebuconazole did not elevate cortisol in response to the ACTH injection, with values significantly lower than the control fish. Fish exposed to sublethal concentrations of atrazine+simazine, atrazine, and glyphosate showed a rise in plasma cortisol very similar to the control fish. We conclude that the ACTH challenge test revealed that R. quelen exposed to sublethal concentrations of tebuconazole and methyl-parathion had a reduced ability to elevate plasma cortisol in response to an intraperitoneal (i.p.) injection of exogenous ACTH, indicating that the interrenal tissue is the site of the impairment within the HPI axis. These ACTH challenge tests also revealed that the impairment of the cortisol response verified in fish exposed to atrazine+simazine and glyphosate, as shown in our previous work, seems to be related to steps of cortisol secretion in higher levels within the HPI axis.

Embryonic and larval development of Jundiá (Rhamdia quelen, Quoy & Gaimard, 1824, Pisces, Teleostei), a South American Catfish

The jundiá (Rhamdia quelen, Quoy and Gaimard) is an endemic South American fish species. Because this species supports cold winters and grows faster during warm months, it has begun to be viewed as an ideal species for fish production in southern South America. In the present study, jundiá oocytes used were obtained by extrusion from females after hormone injection. Soon after hydration, the eggs were transferred to 50 L conic glass incubators, with constant and controlled water influx. Samples of fertilized eggs were transferred to Petri dishes and, examined under a stereoscopic microscope, were spherical, demersal, and non-adhesive with defined perivitelline space and resistant chorion. Cleavage stages occurred during the first 3.5 h. After hatching, larvae were transferred to 200 L glass fiber incubators. First signs of embryo movement were observed 21 h after fertilization; larval eclosion occurred 30.5 h after fertilization. Present findings may provide a basis for studies aimed at determining the complete ontogeny of jundiá and may be useful in eco-toxicological studies.