Accumulation Of Poly-?-hydroxybutyrate(Phb)in Halomonas And Its Function In Artemia Culture

Poly-?-hydroxybutyrate(PHB)is a polymer synthesised intracellularly and stored as carbon and energy reservoir in microorganisms,usually when cellular growth is restricted due to limitaiton of nurtrinents such as O,P,N,S,etc.,and in the presence of excess carbon source.PHB can be used as potential biodegradable plastic and biocontrol agents in acquaculture.As a moderate halophile,Halomonas is able to accumulate PHB at salt contentration of 30-150 g/L and use glucose,sucrose,starch,etc.as carbon source.In this study,orthogonal experimental design and response surface method were used to design different factors such as different concentrations of salt,C,N and P,to study the best condition for PHB accumulation in Halomoas,isolated form Hanggu Saltworks,Tianjin,China.Under the best condition,5-L and 50-L fermentor were conducted to culture Halomonas.During the fermentation,temperature,pH,DO,stirring rate and carbon source supplement were controled to increase the PHB production.An open and unsterile prosess was developed based on halophilic characteristic of Halomonas and the fesibility was analyzed by PCR and DGGE.In gnotobiotic system,Artemia were fed with Halomonas,Halomonas containing higher amount of PHB,PHB-degrading bacterium Comamonas testosteroni and PHB particles,and were challenged with Vibrio anguillarium and Vibrio harveyi,to study the possibility of bacterial PHB application in aquaculture.The result shows that Halomonas could accumulate PHB up to 56.74%wt.under the condition of salinity 30 g/L,glucose 30 g/L,NH4Cl 2 g/L and KH2PO4 0.5 g/L in flask culture.The PHB production was 44.26%wt.at salinity 30 g/L and pH 7.0 in a 5-L fermentor,whilist it reached 54.90%wt.at 30 g/L,pH 7.0 and DO 60%-80%in a 50-L fermentor.In addition,Halomonas could also produce PHB in an open and unsterile condition at salinity 50-150 g/L and glucose 20 g/L,and the maximum PHB production in Halomonas reached 67.00%wt.at salinity 100 g/L.The molecular weight of PHB was between 300-800 kDa in condition of salinity 30 g/L,pH 7.0 and stirring rate 300-600 r/min.The gnotobiotic culture showed that bacterial PHB could promote the growth of Artemia and protect Artemia from Vibrio.The above results have provided some basic data for industrial application of halophilic organisms.The protective effect of bacterial PHB to Artemia indicated the possibility of using bacterial PHB in aquaculture.