Roles and regulation of the AI-2 quorum sensing of vibrio parahaemolyticus in nitrification biofilter of shrimp recirculating system.

Abstract

Thailand is the largest shrimp producing and exporter among Southeast Asia. Up-to-Date, Recirculating Aquaculture System (RAS) have been developed in order to control water quality and enable to conduct intensive cultivation without limitation of land-based cultivation method. Nevertheless, there is concern about pathogenic contamination in Biofilter. Therefore, the role of biofilter in terms of biosecurity is required to study. Recently, shrimp Early Mortality Syndrome (EMS) as known as AHPND/EMS, has spread to shrimp farm in several areas of Thailand. This contributes to severe productivity losses in the shrimp farm industry. It is evident that the causative agent of AHPND disease is bacteria Virbrio parahaemolyticus (VPAHPND), containing specific plasmid for producing bacterial toxins; ToxA and ToxB. however, it is still unclear about the biological mechanism used for toxin production and secretion. Therefore, the objectives of this research are to 1) to study the function of nitrification biological filters and the effects of organic substances on bacteria population in the filters in lab scale 2) to study the role of AI-2 quorum sensing on the toxin secretion related to VPAHPND. The first results found that VPAHPND can grow on biofilm matrix if there was any sufficient organic carbon source. Lack of organic carbon source significant led to elimination of VPAHPND. Moreover, organic substances (glucose) had an effect on changing in the number of bacteria in the biological filters and in the suspended of water. Heterotrophic bacteria were commonly found in Biofilter Unit (over 108 CFU/bead). Moreover, AI-2 like activity, using bioassay, could be observed in Nitrification Biofilter Unit, this was confirmed that AI-2 signal is ubiquitous in marine biofilter. In the second experiment, based on SDS-PAGE and Western Blotting results, the study found that addition of Cell-free supernatant obtained from the mutant Vibiro Campbellii (BB170) into culture medium (TSB+1.5%NaCl) at 1% 5% and 9% v/v could induce ToxA and ToxB production in VPAHPND. There was a strong correlation between the significant change of the toxin production and AI-2 like activity. In contrast, addition of Furanone C-30, a QS inhibitor, at 1 micromolar and 5 micromolar resulted in the significant toxin secretion (P<0.05). From the results aforementioned, this study can be concluded that AI-2 quorum sensing of VPAHPND plays an important role on the AHPND-toxin production. Although the AI-2 like activity in the biological filter had a lower level compared with the culture medium, the accumulation of AI-2 signal molecule may result in the increasing risk of AHPND infection. Therefore, the control and prevention of AHPND/EMS in shrimp cultivation using RAS should concern not only the contaminated VPAHPND, but also the level of QS signals that account for virulence factor of pathogens in the system.