Isolation of bacteria able to degrade poly-hydroxybutyrate-co-hydroxyhexanoate, and the inhibitory effects of the degradation products on shrimp pathogen Vibrio penaeicida

Abstract

Poly-hydroxybutyrate-co-hydroxyhexanoate (PHBH) is a biodegradable, water-insoluble polymer produced by specific bacteria. The monomers of PHBH are the hydroxyalkanoic acids 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HH). Previously, we reported that 3HB and 3HH showed marked antibacterial activities against the shrimp pathogenic bacterium Vibrio penaeicida, and that addition of 5% (w/w) PHBH to the standard aquaculture diet significantly increased survival rate in kuruma shrimp (Marsupenaeus japonicus) after challenge by V. penaeicida, which we attributed to the degradation of PHBH to its monomers in the shrimp gut. In the present study, we isolated four strains of bacteria with high PHBH-degrading activity and evaluated their inhibitory effects on V. penaeicida with PHBH: one strain from shrimp gut contents (E1; Pseudoalteromonas shioyasakiensis/P. mariniglutinosa), two strains from coastal surface seawater (F1; P. shioyasakiensis/P. mariniglutinosa, and F5; Alcanivorax dieselolei/A. xenomutans), and one strain that was a contaminant in commercial PHBH powder (Y1; Bacillus pseudofirmus). Strains E1, F1, and Y1 showed strong PHBH-degrading activity within 24 h of inoculation to PHBH-containing agar plates. Although none of the isolates alone had any effect on the growth of V. penaeicida, when cultured with E1 or F1 and PHBH, the growth of V. penaeicida was markedly suppressed. Incubation with E1 and PHBH resulted in a gradual reduction in the concentration of V. penaeicida from 2 days after the start of incubation until the concentration was 1.2% of that in the control (V. penaeicida alone). Incubation with F1 and PHBH resulted in a rapid reduction in the concentration of V. penaeicida from 2 days after the start of incubation until the concentration was only 0.32% of that of the control. Compared with strains E1 and F1, Y1 showed similar PHBH-degrading activity but did not show any suppressive effect on the growth of V. penaeicida until 5 days after the start of incubation. In addition, this suppressive effect was relatively weak compared with that of the other two strains, suggesting that Y1 can quickly degrade PHBH but that it takes several days to produce monomers. Together, these results suggest that addition to the aquaculture diet of PHBH and PHBH-degrading bacteria that rapidly degrade PHBH to its monomers may speed up degradation of PHBH to its monomers in the shrimp gut, and that it would increase resistance to infection mortality by V. penaeicida in kuruma shrimp.

Keywords: Marsupenaeus japonicus; PHBH-Degrading bacteria; Pseudoalteromonas shioyasakiensis; Shrimp pathogen resistance; Vibrio penaeicida.