The use of homoserine lactone–degrading and poly–β–hydroxybutyrate–accumulating bacteria in crustacean and fish larviculture

Microorganisms play an important role in larviculture. The present study investigated the possibility of improving the microbial management in a hatchery by using bacterial enrichment cultures (ECs) with a double protective function: degradation of quorum sensing molecules (such as acyl homoserine lactone, AHL) and accumulation of a short chain fatty acid–releasing molecule poly–β–hydroxybutyrate (PHB). The isolation of AHL–degrading and PHB–accumulating bacteria was carried out in order to develop the above–described concept. ECs were obtained. The kinetics of AHL degradation by these ECs was established as well as their PHB content. The ECs were characterised through denaturing gradient gel electrophoresis (DGGE) and the main bands were identified by sequence analysis. Their putative probiotic effect was demonstrated in a Vibrio challenge test using Artemia as target organism. Application of probiotics in larviculture needs to be cost–effective. In order to alleviate the cost problem associated with their application, the possibility to produce these putative probionts in a hatchery was evaluated. The ECs can grow at the expense of glycerol which is released by hatching Artemia, can compete with the resident bacteria, and can be boosted through the addition of simple and cheap nutrients. This novel concept is finally applied to marine fish and shrimp larviculture. For these purposes, they are in vivo investigated as biocontrol agent in prawn and Asian sea bass larviculture through their addition to the rearing water at the recommended dosage (106 CFU/mL), and by bio–encapsulation in rotifers and Artemia nauplii. As acyl homoserine lactones (AHLs) molecules have been shown to be responsible for the activation of virulence, this factor is included in the experiments. The supplemented AHL molecules (daily 1mg/mL) had a negative effect on survival and development of these cultured species. These added molecules did not interfere with bacterial growth, but most probably stimulated the production of virulence factors by the resident opportunistic pathogenic bacteria. AHL–degrading ECs can counteract this negative effect and especially enhance larval quality. The increase in both prawn and fish survival due to the presence of an EC correlated to its capacity to degrade the added AHLs. It is not clear to what extent natural AHL concentrations are responsible for negative host microbial concentrations. Hence, the effectiveness of ECs in the absence of AHL addition was investigated. An additional experimental factor, namely green–water is included in this experiment as its use is common in the larval rearing of many marine fish species. The ECs significantly improved L. calcarifer survival in clear–water treatments, but in the green–water treatments, this improvement was only partial. In conclusion, the novel AHL–degrading and PHB–accumulating ECs can be applied into marine fish and shrimp larviculture as a component of the microbial management in a hatchery. The probiotics bacteria can be easily and economically produced at the expense of a C–source available in a hatchery, and be supplied on a daily basis to target organism in sufficiently large quantities. Applying ECs will require a change of practice in the hatcheries, keeping the Artemia hatching water and avoiding the extensive rinsing of Artemia nauplii.