Preliminary assessment of in vivo safety of potentially probiotic lactic acid bacteria for American bullfrog culture

Abstract

The effect of the administration of beneficial lactic acid bacteria (LAB): Lactococcus lactis CRL 1584, L. lactis CRL 1827, Enterococcus gallinarum CRL 1826 and combined CRL 1584+ CRL 1826 on the development of bullfrog embryos from the hatching stage until 31 days (tadpole) was evaluated. In vitro assays indicated that 103, 106 and 109 CFU mL−1 single LAB strains remained viable until 24 h in 10% Ringer. Around 109 CFU mL−1 LAB (individually and combined) were used in an experimental design built to evaluate their effect when administrated at different intervals (three 7-day cycles with 5-day rest periods in between) to embryos until day 31. The highest potentially beneficial population (LAB) numbers were detected in the LAB-treated groups. All the LAB-treatments increased it and were significantly higher than the controls. Although the highest, potentially pathogenic, population (Red-Leg Syndrome-RLS-related pathogens) numbers were detected in the control and the lowest in the CRL 1584+ CRL 1826-treated group, they did not differ significantly. Stereoscopic studies showed no malformations in any LAB-treated group and all the specimens reached the same stage of their biological cycle with a survival >94%. The histological structure of target organs for RLS-associated pathogens (intestine and skin) and stomach was not affected and the spleen was developed. Only the LAB-treated groups showed microorganisms associated with the intestinal mucus, without inflammatory reaction in the lamina propria. This is the first report on the evaluation of the absence of adverse effects after LAB administration to bullfrog embryos using indigenous microorganisms.