Evaluation of Protective Effect of Poly-β-Hydroxybutyrate Producing Halophilic Microorganisms on Vibrio-Challenged Artemia

Abstract

Infectious diseases pose a significant obstacle to expanding aquaculture production, prompting the prolonged use of antibiotics for prophylactic and therapeutic purposes. However, concerns about the adverse effects of antibiotics have sparked interest in alternative approaches. Probiotics and poly-β-hydroxybutyrate (PHB) have emerged as potential biocontrol agents. This study explores the protective effects of PHB-producing halophilic microorganisms in gnotobiotic Artemia franciscana during a Vibrio campbellii challenge, representing the first investigation of its kind. Various halophilic bacterial and archaeal species were screened for PHB production qualitatively (using Sudan Black B and safranin staining technique) and quantitatively (using a spectrophotometric method), revealing Halomonas ventosae, Halomonas sp., Halomonas boliviensis, Haloferax mediterranei, and Haloferax volcanii as capable PHB accumulators. H. ventosae and Hfx. mediterranei stood out, achieving 46% and 31% of cell dry weight (CDW) when growing on complex media with 2% glucose. Then, the gnotobiotic Artemia challenge test was used to evaluate the protective effect of selected PHB-producing halophiles. In these vivo challenge tests, supplementing H. ventosae and Hfx. mediterranei in culture water significantly increased the survival of Vibrio-challenged gnotobiotic Artemia compared to the challenged control. Survival increased significantly with increasing cell density of halophiles. Moreover, the survival of the challenged group supplemen-ted with halophiles was comparable to that of the unchallenged control at the halophile cell concentration of 5 × 107 cells ml-1. Notably, amorphous PHB concentration estimates indi-cating complete protection in Artemia were 2.5 mg l-1 for H. ventosae and 3.5 mg l-1 for Hfx. mediterranei. However, higher amorphous PHB content in the halophiles did not result in higher survival, indicating that cellular components of halophiles other than PHB might be responsible for the protective effect. While these findings suggest that halophile supplementation protects Artemia against Vibrio campbellii, the study did not conclusively attribute this protection solely to the PHB content in the halophiles.