Vibrio parahaemolyticus is a rapidly evolving pathogen of both human and seawater animals. Depending on the diversity of horizontally acquired pathogenicity factors, V. parahaemolyticus may cause a broad range of diseases, including food poisoning in humans and Early Mortality Syndrome (EMS) or Hepatopancreatic Acute Necrosis Syndrome in shrimp. In shrimp farms, EMS-caused mortality may reach 100% in the first 30 days after infection leading to substantial business losses for farmers.
The control of bacterial pathogens without use of antibiotics or harsh chemicals is a challenge in modern aquaculture where animal densities are often very high. In many bacteria quorum sensing molecules (QS autoinducers) regulate many pathogen traits including biofilm and toxin production. Thus, the elimination of QS molecules or quorum quenching (QQ) may be used to manage bacterial infections and virulence. Here we test the hypothesis that probiotic bacterial strains engineered to reduce QS concentrations may be an alternative approach to conventional antibacterial treatments for the control of EMS in shrimp.
As an enhanced probiotic, we selected Enterobacter Ag1, which is salt-resistant and capable of colonization of shrimp intestines. To create Ag1 quorum-quenching derivatives, we cloned and expressed a number of lactone-degrading enzymes under the control of a strong gene promoter. Lactonase AidH expressing Ag1 was selected as the optimal construct due to its ability to quench QS regulated luminescence in light-emitting V. harveii strain. To test Ag1-AidH ability to manage EMS in shrimp, we supplemented shrimp food with bacteria and monitored shrimp mortality. We showed that the experimental shrimp group had reduced mortality (25 to 55%) relative to controls. Additionally, we found that expression of the pir-like toxin, the primary pathogenicity factor in V. parahaemolyticus infected shrimp intestines, was reduced by more than 2-fold by QQ bacteria.
The level pirA expression increases more than 10,000 fold during bacterial growth associated with the production of QS molecules. Co-cultivation of quorum quenching Ag1 with Vibrio lead to a 2- to 3-fold reduction in pirA expression. We propose that reduction in shrimp mortality is, in part, associated with reduced toxin production mediated by QQ bacteria. QQ may provide sustainable solution to control Vibrio-related shrimp diseases and mortality.