NaCl promotes antibiotic resistance by reducing redox states in Vibrio alginolyticus.

The development of antibiotic resistance in Vibrio alginolyticus represents a threat to human health and fish farming. Environmental NaCl regulation of bacterial physiology is well documented, but whether the regulation contributes to antibiotic resistance remains unknown. To explore this, we compared minimum inhibitory concentration (MIC) of V. alginolyticus cultured in different media with 0.5%-10% NaCl, and found that the MIC increased as the NaCl concentration increased, especially for aminoglycoside antibiotics. Consistent with this finding, internal NaCl also increased, while intracellular gentamicin level decreased. GC-MS-based metabolomics showed different distributions of pyruvate cycle intermediates among 0.5%, 4% and 10% NaCl. Differential activity of enzymes in the pyruvate cycle and altered expression of Na(+)-NQR led to a reducing redox state, characterized by decreased levels of NADH, proton motive force (PMF) and ATP. Meanwhile, NaCl negatively regulated PMF as a consequence of the reducing redox state. These together are responsible for the decreased intracellular gentamicin level with the increased external level of NaCl. Our study reveals a previously unknown redox state-dependent mechanism regulated by NaCl in V. alginolyticus that impacts antibiotic resistance.

Boosted TCA cycle enhances survival of zebrafish to Vibrio alginolyticus infection.

Vibrio alginolyticus is a waterborne pathogen that infects a wide variety of hosts including fish and human, and the outbreak of this pathogen can cause a huge economic loss in aquaculture. Thus, enhancing host's capability to survive from V. alginolyticus infection is key to fighting infection and this remains still unexplored. In the present study, we established a V. alginolyticus-zebrafish interaction model by which we explored how zebrafish survived from V. alginolyticus infection. We used GC-MS based metabolomic approaches to characterize differential metabolomes between survival and dying zebrafish upon infection. Pattern recognition analysis identified the TCA cycle as the most impacted pathway. The metabolites in the TCA cycle were decreased in the dying host, whereas the metabolites were increased in the survival host. Furthermore, the enzymatic activities of the TCA cycle including pyruvate dehydrogenase (PDH), α-ketoglutaric dehydrogenase (KGDH) and succinate dehydrogenase (SDH) also supported this conclusion. Among the increased metabolites in the TCA cycle, malic acid was the most crucial biomarker for fish survival. Indeed, exogenous malate promoted zebrafish survival in a dose-dependent manner. The corresponding activities of KGDH and SDH were also increased. These results indicate that the TCA cycle is a key pathway responsible for the survival or death in response to infection caused by V. alginolyticus, and highlight the way on development of metabolic modulation to control the infection.

Glucose enhances tilapia against Edwardsiella tarda infection through metabolome reprogramming.

We have recently reported that the survival of tilapia, Oreochromis niloticus, during Edwardsiella tarda infection is tightly associated with their metabolome, where the survived O. niloticus has distinct metabolomic profile to dying O. niloticus. Glucose is the key metabolite to distinguish the survival- and dying-metabolome. More importantly, exogenous administration of glucose to the fish greatly enhances their survival for the infection, indicating the functional roles of glucose in metabolome repurposing, known as reprogramming metabolomics. However, the underlying information for the reprogramming is not yet available. Here, GC/MS based metabolomics is used to understand the mechanisms by which how exogenous glucose elevates O. niloticus, anti-infectious ability to E. tarda. Results showed that exogenous glucose promotes stearic acid and palmitic acid biosynthesis but attenuates TCA cycle to potentiate O. niloticus against bacterial infection, which is confirmed by the fact that exogenous stearic acid increases immune protection in O. niloticus against E. tarda infection in a manner of Mx protein. These results indicate that exogenous glucose reprograms O. niloticus anti-infective metabolome that characterizes elevation of stearic acid and palmitic acid and attenuation of the TCA cycle. Therefore, our results proposed a novel mechanism that glucose promotes unsaturated fatty acid biosynthesis to cope with infection, thereby highlighting a potential way of enhancing fish immunity in aquaculture.

Construction and immune protection evaluation of recombinant polyvalent OmpAs derived from genetically divergent ompA by DNA shuffling.

A wide variety of bacterial infections is a major challenge in aquaculture. Development of polyvalent vaccines that can fight against as many pathogens as possible is especially necessary. The present study uses DNA shuffling to create a new hybrid OmpA with improved cross-protection against Vibrio alginolyticus and Edwardsiella tarda through the recombination of six OmpA genes from Vibrio parahaemolyticus, V. alginolyticus, E. tarda and Escherichia coli. Out of the 43 recombinant chimeras genes constructed using VA0764 primers, EompAs-19 was demonstrated as an ideal polyvalent vaccine against infections caused V. alginolyticus and E. tarda. Compared with VA0764, OmpAs-19 had three mutations, which may be a molecular basis of EompAs-19 as an efficient polyvalent vaccine against both V. alginolyticus and E. tarda infections. These results develop a polyvalent vaccine that prevents the infections caused by extracellular and intracellular bacteria. Thus, the present study highlights the way to develop polyvalent vaccines against microbial infections by DNA shuffling.