ABSTRACT This study aimed to evaluate 1) the in vitro effect of organic salts on the growth of the probiotic Lactobacillus plantarum and then 2) the combined use of a probiotic with organic salts on the in vitro inhibition of V. alginolyticus, A. hydrophila, E. coli, P. aeruginosa, and S. agalactiae. In vitro tests were performed with eight different organic salts, including butyrate, propionate, succinate, citrate, formate, fumarate, glutamate, and acetate, at two pH values (6.2 and 7.1) to determine their effect on the growth kinetics of L. plantarum. In addition, each organic salt was tested alone and in combination with L. plantarum to evaluate the inhibitory effect against the pathogenic bacteria noted above in either condition. Sodium citrate and formate inhibited the growth of L. plantarum, but sodium glutamate, succinate and fumarate stimulated it. Sodium propionate, butyrate, and acetate did not affect probiotic growth at all. Inhibition against all pathogens was significantly higher in the presence of the probiotic and lower pH. Comparing all organic salts at the two pH values, butyrate, acetate, and propionate exhibited more inhibition against V. alginolyticus than the others, while propionate had higher inhibition against A. hydrophila, E. coli, P. aeruginosa, and fumarate successfully inhibited S. agalactiae. Based on these results, it can be concluded that organic salts showed better in vitro inhibition against the aquaculture pathogenic bacteria tested when combined with the probiotic L. plantarum.

ABSTRACT This study evaluated the performance, immunology, and survival of the Pacific white shrimp Litopenaeus vannamei to experimental challenge to Vibrio alginolyticus based on the use of the probiotic Lactobacillus plantarum and the combined use of probiotic and butyrate. Four different diets resulted from the addition of additives: butyrate, probiotic, butyrate + probiotic, and control (no additives). The attractiveness of the diets was assessed by the percentage of positive choices and rejections, using a dual-choice Y-maze format aquarium. The shrimps were fed during four weeks and performance parameters, intestinal microbiota, and immunological parameters were all evaluated. Subsequently, the shrimps were challenged with V. alginolyticus and after 48 h, survival and immunological parameters were evaluated. The results showed increased attractiveness and intake, but only with diets supplemented with sodium butyrate. However, other diets were not rejected. No difference in performance or immunological parameters was observed among the different diets. Also, among the treatments, no difference in Vibrio spp., or total heterotrophic bacteria counts, was found in the intestinal tract. However, the lactic acid bacteria count was higher in the intestinal tract of shrimps fed diets supplemented with probiotic. After bacterial challenge, shrimp fed all diets had a greater survival when compared with the control group. Lactobacillus plantarum and sodium butyrate increase the resistance of shrimp to infection with V. alginolyticus, but do so without affecting performance, immunological parameters, or Vibrio spp., and total heterotrophic bacteria counts in the intestinal tract.

Resumo O objetivo deste trabalho foi avaliar o efeito de Lactobacillus plantarum sobre a conservação de mexilhões frescos e avaliar a atratividade, o consumo e a microbiota intestinal de reprodutores de camarão-branco-do-pacífico. O experimento avaliou mexilhões estocados com L. plantarum a 4ºC. Os controles foram: mexilhões estocados a -18ºC sem L. plantarum; mexilhões estocados a -18ºC com L. plantarum; e mexilhões estocados a 4ºC sem L. plantarum. As análises microbiológicas dos mexilhões foram realizadas nos dias 1, 7, 15, 30, 45 e 60 após o processamento. Além disso, os mexilhões conservados com L. plantarum e estocados a 4ºC e os mexilhões estocados a -18ºC sem L. plantarum, após 15 dias, foram avaliados quanto à atratividade, ao consumo e à microbiota bacteriana intestinal dos camarões. Mexilhões conservados com L. plantarum tiveram maiores contagens de bactérias ácido-lácticas e menores contagens de Vibrio spp., assim como de bactérias heterotróficas totais, após 60 dias de estocagem. Não foram observadas diferenças significativas quanto à atratividade ou ao consumo entre os tratamentos. A microbiota intestinal dos camarões alimentados com mexilhões conservados com L. plantarum apresentou maior contagem de bactérias ácido-lácticas e menor contagem de Vibrio spp. O uso de L. plantarum inibe Vibrio spp. e conserva o alimento, sem modificar a atratividade ou o consumo pelos camarões.

Abstract The objective of this work was to evaluate the effect of Lactobacillus plantarum on the preservation of fresh mussels and its effect on the attractiveness, consumption, and midgut bacterial microbiota of Pacific white shrimp broodstock. The experiment evaluated mussels stored with L. plantarum at 4ºC. The controls were: mussels stored at -18ºC without L. plantarum; mussels stored at -18ºC with L. plantarum; and mussels stored at 4ºC without L. plantarum. Microbiological analyses on mussels were performed on days 1, 7, 15, 30, 45, and 60 after processing. Additionally, mussels preserved with L. plantarum and stored at 4ºC, and mussels stored at -18ºC without L. plantarum were evaluated after 15 days for attractiveness, consumption, and midgut bacterial microbiota of shrimps. Mussels preserved with L. plantarum showed higher lactic acid bacteria counts and lower counts of Vibrio spp., as well as of total heterotrophic bacteria, after 60 days of storage. No differences were observed for attractiveness or consumption between treatments. The bacterial microbiota of midgut in shrimp fed mussels preserved with L. plantarum showed higher lactic acid bacteria count and lower Vibrio spp. The use of L. plantarum inhibits Vibrio spp. and preserves feed without changing attractiveness or consumption for shrimp.

O objetivo deste trabalho foi avaliar o uso de suplemento prebiótico, probiótico e simbiótico, na dieta de camarões marinhos (Litopenaeus vannamei) e seus efeitos sobre o crescimento, a microbiota intestinal, a resposta imune e a resistência ao desafio experimental com Vibrio alginolyticus. Foram utilizados quatro tratamentos: prebiótico inulina; probiótico Lactobacillus plantarum; simbiótico Lactobacillus plantarum + inulina; e controle. Os camarões foram distribuídos em 16 tanques de dez mil litros de água, povoados com 200 camarões cada, cultivados por seis semanas. Avaliaram-se a microbiologia do trato intestinal dos camarões e a reposta imune, antes e após o desafio com V. alginolyticus. A concentração de Vibrio spp. no trato digestório foi menor em camarões alimentados com dieta suplementada com prebiótico, probiótico e simbiótico, enquanto a concentração de bactérias acidoláticas foi superior somente nos camarões alimentados com probiótico e simbiótico. O título aglutinante do soro contra V. alginolyticus aumentou no grupo probiótico e simbiótico, antes da infecção, e foi maior em todos os tratamentos após infecção com V. alginolyticus, em comparação ao controle. Não foi observada diferença entre os tratamentos quanto aos demais parâmetros avaliados. As dietas probióticas, prebióticas e simbióticas alteram a microbiota intestinal e aumentam o título aglutinante do soro contra V. alginolyticus; contudo, não alteram a resistência ao desafio nem o crescimento dos camarões.

The objective of this work was to evaluate the use of prebiotic, probiotic, and symbiotic supplements, in the diet of shrimp (Litopenaeus vannamei), and their effects on growth, intestinal microbiota, immune response, and resistance to experimental challenge with Vibrio alginolyticus. Four treatments were used: prebiotic (inulin); probiotic (Lactobacillus plantarum); symbiotic (Lactobacillus plantarum + inulin); and a control. Shrimp were distributed in 16 tanks with ten thousand liters of water, stocked with 200 shrimp each, cultivated for six weeks. Microbiology of the intestinal tract and the immune response of shrimp were evaluated before and after the challenge with V. alginolyticus. The concentration of Vibrio spp. in the digestive tract was lower in shrimp fed prebiotic, probiotic, and symbiotic-supplemented diet, while concentration of lactic acid bacteria was higher only in shrimp fed probiotic and symbiotic-supplemented diet. The title of agglutinating serum against V. alginolyticus increased in shrimp fed the probiotic and symbiotic-supplemented diet prior to infection, and was higher in all treatments after infection with V. alginolyticus, in comparison to the control. No difference was observed between treatments for the remaining parameters. Probiotic, prebiotic, and symbiotic diets change intestinal microbiota, and increase the agglutinating serum title against V. alginolyticus; however, they change neither the shrimp resistance to challenge by V. alginolyticus nor shrimp growth.

O objetivo deste trabalho foi determinar o método de coleta de fezes mais adequado para camarões marinhos (Litopenaeus vannamei) na determinação do coeficiente de digestibilidade aparente da proteína bruta e da matéria seca de dietas comerciais, e avaliar o efeito da suplementação de Lactobacillus plantarum na dieta sobre o coeficiente de digestibilidade aparente da proteína, da energia e da matéria seca. Foram testados três sistemas de coletas de fezes: decantação, sifonamento e dissecação dos animais. A coleta por decantação foi o sistema mais adequado para a determinação in vivo dos coeficientes de digestibilidade aparente da proteína bruta e da matéria seca das dietas, por apresentar baixo grau de lixiviação das fezes e por ser de fácil execução. Nos camarões alimentados com a dieta suplementada com L. plantarum, foi observado maior coeficiente de digestibilidade aparente da proteína (84,18±0,35%) e da energia (81,28±0,16%), quando comparado ao grupo controle (82,73±1,01 e 78,63±0,93%, respectivamente). O uso de L. plantarum como aditivo probiótico na ração pode trazer benefícios nutricionais a L. vannamei, com melhor aproveitamento dos nutrientes.

The objective of this work was to determine the most adequate feces collection method for sea shrimp (Litopenaeus vannamei) in the determination of the apparent digestibility coefficients of crude protein and dry matter of commercial diets, and to evaluate the effect of supplementation of Lactobacillus plantarum to the diet on the apparent digestibility coefficient of protein, energy, and dry matter. Three feces collection methods were tested: decanting, siphoning, and animal dissection. Decanting was the method best suited for the in vivo determination of the apparent digestibility coeffcients of crude protein and dry matter of commercial diets, since it promoted lower nutrient leaching from feces and was easy to perform. Shrimp fed the diet supplemented with probiotic showed higher apparent digestibility of crude protein (84.18±0.35%) and energy (81.28±0.16%) when compared to the control group (82.73±1.01 and 78.63±0.93%, respectively). The use of L. plantarum as a probiotic additive in shrimp diets can provide nutritional benefits to L. vannamei, with better nutrient utilization.

The requirement of gross protein and energy and the optimal ratio of these in practical diets for pink shrimp (Farfantepenaeus paulensis) juveniles in different salinities were determined. Shrimp with a mean initial weight of 0.353 g were stocked in 54 plastic tanks at initial densities of 40 ind m-2. The experiment was factorial, with three levels of gross protein (24, 35, 48%), three levels of salinities (10, 20, 30 psu), and two levels of gross energy (3200, 4200 kcal kg-1); the experiment lasted 25 days and considered three replicates. The results showed that salinity and protein had a significant effect (P < 0.05) on weight gain and the specific growth rate, but did not influence on survival. The energy had no significant effect on survival, growth rate, or weight gain. The response of the shrimp to the gross energy and protein level showed different requirements according to the different salinities tested. At salinity of 10 psu, the adequate level was 35% gross protein and 4200 kcal kg-1 (83.3 mg PB kcal-1), whereas at salinities of 20 and 30 psu, levels of 24% gross protein and 3200 kcal kg-1 (75.0 mg PB kcal-1) were appropriate.

Se determinó el requerimiento de proteína (PB) y energía bruta (EB) y su relación óptima en dietas prácticas para juveniles de camarón rosado Farfantepenaeus paulensis en diferentes salinidades. Se sembraron camarones con peso promedio de 0,353 g en 54 estanques de PVC con densidad inicial de 40 ind m-2. El experimento fue factorial, con tres niveles de proteína bruta 24, 35 y 48%, tres salinidades 10, 20 y 30 psu y dos niveles de energía bruta 3200 y 4200 kcal kg-1 de dieta, con tres réplicas, en un período experimental de 25 días. Los resultados indicaron que la salinidad y la proteína tuvieron un efecto significativo (P < 0,05) sobre la ganancia de peso y tasa de crecimiento específico, pero no influyeron en la sobrevivencia. La energía no tuvo efecto significativo (P > 0,05) sobre la sobrevivencia, tasa de crecimiento específica y ganancia en peso. La respuesta de los camarones a las necesidades de proteína y energía bruta en las salinidades probadas fue diferente. En la salinidad de 10 psu los requerimientos fueron 35% PB y 4200 kcal kg-1 (83,3 mg PB kcal-1), mientras que en salinidades de 20 y 30 psu fueron 25% de PB y 3200 kcal kg-1 (75,0 mg PB kcal-1).

The objective of this study was to investigate the growth of juvenile fat snook (Centropomus parallelus) in laboratory conditions when fed a diet supplemented with the probiotic Lactobacillus plantarum. Changes in the intestinal flora, haematological parameters and growth performance were assessed using 180 fishes (54.2 ± 13.4 g each). The fishes were subjected to two treatments divided into six cages: 1) feed supplemented with probiotic, and 2) feed without probiotic (control). The temperature, dissolved oxygen and salinity were maintained at 25 ± 1 ° C, 4.0 mg.L-1 and 33 ‰, respectively. After 10 weeks of culture, the fishes fed probiotic had reduced viable culturable heterotrophic bacteria and Vibrio spp. and increased lactic acid bacteria in the intestinal tract, as well as a higher number of thrombocytes, leukocytes and lymphocytes in the blood. No significant difference was observed in the growth, survival or body composition, but the hepatosomatic index was significantly higher in the fishes fed with probiotic and control.

O objetivo deste trabalho foi avaliar a relação entre parâmetros físico-químicos e biológicos do cultivo de camarão com a enfermidade causada pelo vírus da síndrome da mancha-branca (WSSV). Foram avaliadas a mortalidade, as características físicas e químicas da água e do solo, e foram realizados os exames e as análises microscópicas a fresco, presença de víbrios na hemolinfa, além do diagnóstico histopatológico e molecular (PCR) do WSSV, em viveiros de oito fazendas de Santa Catarina, entre 2008 e 2009. O vírus foi detectado em cinco fazendas com registros da enfermidade em ciclos anteriores. A temperatura da água foi semelhante entre as fazendas com e sem WSSV, e a mortalidade ocorreu a intervalos ascendentes entre 24,6 e 29,3ºC. Não foram encontradas diferenças significativas nos parâmetros de análises microscópicas a fresco, tempo de coagulação e presença de víbrios na hemolinfa e sinais clínicos, entre viveiros com e sem WSSV. As concentrações de nitrito, sílica, fenol e alcalinidade na água e pH do solo apresentaram diferenças significativas entre os viveiros afetados ou não por WSSV. Os valores de nitrito, sílica e pH do solo estavam dentro dos limites recomendados para o cultivo de camarões, no entanto, isto não ocorreu com a alcalinidade e a concentração de fenol, o que sugere uma relação dos últimos com a manifestação da enfermidade da mancha-branca.

The objective of this work was to evaluate the relationship among physicochemical and biological parameters in shrimp cultivation with the disease caused by the white spot syndrome virus (WSSV). The occurrence of mortality, physical and chemical characteristics of water and soil were evaluated, and clinical and microscopic analyses and the presence of vibrio in hemolymph, were performed as well as the histopathological and molecular diagnosis (PCR) of the WSSV in ponds of eight farms in Santa Catarina, Brazil, between 2008 and 2009. The virus was detected in five farms, which had displayed the disease in previous cycles. The temperature of the water was similar among farms independently of the presence of the virus, and mortality occurred at ascendant temperature intervals, between 24.6 and 29.3ºC. No significant differences were found for the parameters of microscopic analyses among ponds either with or without the virus. Nitrite, silica, phenol and water-alkalinity concentrations, as well as soil pH values, showed significant differences among the ponds affected or not by the WSSV. Nitrite, silica and pH values were within the recommended limits for shrimp cultivation, however this did not occur for the alkalinity and phenol concentrations, wich suggests a relationship of the latter parameters with manifestation of the white spot disease.

O objetivo deste trabalho foi avaliar a eficácia da técnica de enriquecimento de náuplios de Artemia franciscana, em emulsões de ácidos graxos poliinsaturados (AGP) para a alimentação de pós-larvas do camarão marinho Farfantepenaeus paulensis. Este procedimento visou o acréscimo dos ácidos eicosapentaenóico (EPA) e docosahexaenóico (DHA), essenciais para organismos marinhos, mas deficientes na composição de náuplios provenientes de ecossistemas aquáticos continentais. Os náuplios foram divididos em cinco tratamentos: um grupo controle, composto por indivíduos recém-eclodidos, e quatro grupos enriquecidos cada um com as emulsões Selco, Super Selco, AlgaMac 2000 e emulsão formulada em laboratório à base de óleo de fígado de bacalhau, e então ofertados às pós-larvas. Foram comparados os efeitos das dietas quanto à taxa de sobrevivência, ao ganho de peso e à resistência ao estresse nos estádios de PL10 e de PL20. Observou-se que as emulsões Super Selco e Selco foram as que proporcionaram aumento mais destacado dos níveis de EPA e DHA nos náuplios. Entretanto, nenhuma das emulsões aplicadas induziu a diferenças significativas para os parâmetros estudados em comparação com as pós-larvas alimentadas com os náuplios do grupo de controle, indicando que o aumento nos níveis de EPA e DHA não exerceu influência na melhora do desenvolvimento de pós-larvas.

The objective of this work was to evaluate the efficiency of Artemia franciscana nauplii, in poliunsturated fatty acid enriched emulsions, for marine shrimp (Farfantepenaeus paulensis) postlarvae feeding. This proceeding aimed the addiction to their diets of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), essential fatty acids for marine organisms, but scarce in nauplii originated in aquatic continental ecosystems. The nauplii were divided in five treatments: a control one with newly-hatched individuals and other four groups, each one enriched with the emulsions: Selco, Super Selco, AlgaMac 2000 and lab made emulsion with cod liver oil, then, offered to the postlarvae. The effect of these diets were compared according to the survival rate, weight gain and stress resistance at the PL10 and PL20 stages. It was observed that Super Selco and Selco emulsions provided the higher increase of the EPA and DHA levels in the nauplii. However, none of the emulsions applied provided significant difference in the observed parameters in comparison to the post-larvae fed with the nauplii from the control group, indicating that the increase of EPA and DHA levels were not critical for the improvement of postlarvae development.