ABSTRACT The objective of this study was to evaluate the use of a diet supplemented with Lactobacillus plantarum on performance, immunological parameters, and bacterial microbiota of the digestive tract of white leg shrimp (Litopenaeus vannamei). We used six earthen ponds (1.2 ha) stocked with 14 shrimp m−2 (3 g mean weight). Three ponds received a diet supplemented with L. plantarum and three were maintained as control. The survival and feed efficiency of shrimp fed the supplemented diet were greater than that observed in shrimp fed the control diet, with survival values of 83.02±6.12% and 74.65±9.07% and feed efficiencies of 117.97±4.45% and 104.46±7.30%, respectively. However, we observed no differences in weight gain. The intestines of shrimp fed probiotics had lower counts of Vibrio spp. and higher counts of lactic acid bacteria, compared with those of control shrimp. Diets supplemented with L. plantarum alter the intestinal bacterial microbiota of shrimp, resulting in increased survival and feed efficiency.

Aiming at assessing the cryopreservation potential of Litopenaeus vannamei sperm cells, 74 spermatophores were manually extracted from the sexually mature individuals. After the toxicity test to define the cryoprotectant concentration, suspensions of spermatic cells were cryopreserved in the groups in freezing solutions comprising different cryoprotectants such as dimethyl sulfoxide (DMSO) and ethylene glycol (EG) at 10% concentration. Each treatment was divided in subgroups for storage in liquid nitrogen during 0, 30, 60 and 90 days, in triplicate. After thawing at 25ºC for 40 seconds, cell viability in the suspensions was analyzed under the microscope in eosin-nigrosin stain and flow cytometry. There were no significant differences between the cryoprotectants used. For all the treatments, lower and higher mortalities occurred in the 0 and 90 days, respectively. By applying the eosin-nigrosin technique, lower and higher mortalities were 23.17 and 82.11% for DMSO and 29.94 and 83.72% for EG, while the flow cytometry registered mortalities of 2.42 and 55.13% for DMSO and 0.90 and 55.56% for EG. The Spearman correlation coefficient indicated a positive correlation (R=0.91) between the two techniques used. It was concluded that there was a decrease in cell viability within a longer cryopreservation time.

O objetivo deste trabalho foi isolar cepas de bactérias ácido-lácticas, com potencial probiótico do trato digestório de camarões marinhos e realizar seleção in vitro baseada em múltiplos caracteres. O ideótipo (cepa ideal proposta) foi definido por meio das maiores médias para os caracteres velocidade máxima de crescimento, contagem final de células viáveis e halo de inibição contra nove patógenos de origem continental e marinha, e pelas menores médias para os caracteres tempo de duplicação e tolerância das cepas a NaCl (1,5 e 3%), pH (6, 8 e 9) e sais biliares (5%). Foram estimadas as distâncias de Mahalanobis (D²) entre as cepas avaliadas, e as melhores foram aquelas que apresentaram menor distância do ideótipo. Foram isoladas dez cepas de bactérias identificadas bioquimicamente como Lactobacillus plantarum (3), L. brevis (3), Weissella confusa (2), Lactococcus lactis (1) e L. delbrueckii (1). As cepas de L. plantarum apresentaram amplo espectro de ação e os maiores halos de inibição, tanto para patógenos Gram-positivos quanto negativos, alta taxa de crescimento e tolerância a todos os parâmetros avaliados. Em relação ao ideótipo, L. plantarum apresentou a menor distância de Mahalanobis (D²), seguida pelas cepas de W. confusa, L. brevis, L. lactis e L. delbrueckii. Entre as cepas bacterianas avaliadas, as de L. plantarum apresentam o maior potencial para uso como probiótico para camarões marinhos.

The objective of this work was to isolate strains of lactic acid bacteria with probiotic potential from the digestive tract of marine shrimp (Litopenaeus vannamei), and to carry out in vitro selection based on multiple characters. The ideotype (ideal proposed strain) was defined by the highest averages for the traits maximum growth velocity, final count of viable cells, and inhibition halo against nine freshwater and marine pathogens, and by the lowest averages for the traits duplication time and resistance of strains to NaCl (1.5 and 3%), pH (6, 8, and 9), and biliary salts (5%). Mahalanobis distance (D²) was estimated among the evaluated strains, and the best ones were those with the shortest distances to the ideotype. Ten bacterial strains were isolated and biochemically identified as Lactobacillus plantarum (3), L. brevis (3), Weissella confusa (2), Lactococcus lactis (1), and L. delbrueckii (1). Lactobacillus plantarum strains showed a wide spectrum of action and the largest inhibition halos against pathogens, both Gram-positive and negative, high growth rate, and tolerance to all evaluated parameters. In relation to ideotype, L. plantarum showed the lowest Mahalanobis (D²) distance, followed by the strains of W. confusa, L. brevis, L. lactis, and L. delbrueckii. Among the analyzed bacterial strains, those of Lactobacillus plantarum have the greatest potential for use as a probiotic for marine shrimp.

O objetivo deste trabalho foi avaliar a substituição de dois níveis de inclusão de farinha de peixe por outros ingredientes, nas dietas de Litopenaeus vannamei cultivados em sistema bioflocos. Foram avaliadas dietas sem uso de farinha de peixe (dieta A, 100% de substituição), com inclusão de 12,5% de farinha de peixe (dieta B, 40% de substituição) e direta controle com inclusão de 21% (dieta C, 0% de substituição). Nas dietas A e B, o farelo de soja e as farinhas de carne e vísceras foram os principais substitutos proteicos. Foram analisados os índices de desempenho dos camarões e os parâmetros físicos e químicos da água de cultivo. Os camarões alimentados com a dieta B apresentaram maior peso final (11,63±1,38 g), em comparação aos camarões alimentados com a dieta A (peso final, 9,39±0,31 g) e com a dieta C (peso final, 10,20±1,10 g). Os demais parâmetros de desempenho como produtividade, conversão alimentar e sobrevivência não apresentaram diferenças entre os tratamentos. A redução de até 40,0% da farinha de peixe pode ser feita em cultivos superintensivos de L. vannamei com bioflocos, sem interferir em seu desempenho zootécnico e na qualidade de água do cultivo.

The objective of this work was to evaluate the substitution of two fish meal inclusion levels by other ingredients, in the diets of Litopenaeus vannamei reared in a biofloc system. The following diets were evaluated: without fish meal (diet A, 100% replacent), with addition of 12.5% of fish meal (diet B, 40% replacent), and a control diet with addition of 21% (diet C, 0% replacent). In diets A and B, soybean meal and meat and viscera meals were the main protein replacement. The performance indexes of shrimps and the physical and chemical parameters of water were analyzed. Shrimp fed diet B had higher final weight (11.63±1.38 g), compared to shrimp fed a diet A (9.39±0.31 g final weight) or C (10.20±1.10 g final weight). The other parameters of performance such as yield, feed conversion and survival did not differ between treatments. The reduction of fish meal by 40.0% can be done in a L. vannamei superintensive biofloc system without interfering on their growth performance or on the water quality of the culture.

O objetivo deste trabalho foi avaliar o tempo de atuação de Lactobacillus plantarum, na microbiota bacteriana intestinal de Litopenaeus vannamei, e sua relação com a contagem total de hemócitos e a atividade da fenoloxidase após a exposição a Vibrio harveyi. Os camarões foram alimentados com dieta suplementada com probióticos por oito dias e, depois, substituída por dieta comercial. Os camarões alimentados com dieta comercial constituíram o controle. As avaliações foram realizadas no oitavo dia de experimento e repetidas dois, quatro, seis e oito dias depois. A contagem total de bactérias lácticas no intestino foi maior até o quarto dia de avaliação, no grupo alimentado com probióticos. A contagem de Vibrio spp. foi superior no grupo controle nos dias zero e dois. Até o quarto dia, a contagem total de hemócitos dos camarões após a inoculação de V. harveyi foi superior no grupo alimentado com probiótico em relação ao grupo controle. Não foi observada diferença na atividade da fenoloxidase. No sexto dia, após a substituição da dieta suplementada com probióticos por dieta controle, todos os parâmetros avaliados foram iguais nos dois grupos, o que indica que o tempo de atuação de L. plantarum em camarões é curto.

The objective of this work was to assess the time-related action of probiotic Lactobacillus plantarum in the bacterial microbiota of the digestive tract of Litopenaeus vannamei, and the relation of total haemocyte count and serum phenol oxidase activity of shrimp challenged with Vibrio harveyi. Shrimps were fed with a probiotic-supplemented diet, for eight days, then shifted to a commercial diet. Shrimps fed only with the commercial diet served as control. Evaluations were made on the 8th day of experiment and repeated two, four, six and eight days later. Total lactic bacteria in the digestive tract was higher until the 4th day of evaluation in the probiotic-supplemented group. Vibrio spp. counts were higher in the control at days zero and two. Until the 4th day of evaluation, the total haemocyte counts in shrimps after challenge with V. harveyi were higher in probiotic-supplemented group than in control group. Significant difference was not observed in phenol oxidase activity. On the 6th day after shifting from supplemented to control diet, all parameters were equal in both groups, suggesting that the time-related action of L. plantarum in shrimp is short.