APPLYING MOLECULAR ANALYTICAL TECHNIQUES FOR ASSESSMENT OF MICROORGANISM COMPONENT AND QUANTITY IN DESIGNED OIL POLLUTED EXPERIMENTS BY BIOREMEDIATION
Abstract
Biorernediation is one of the promising methods to reduce or eliminate toxic pollutants. For making successful application of this method, information about the diversity, population size and activity of environmental microbes is essential. The previous studies have revealed that biostimulation treatment (nutrient addition) can accelerate oil removal from the polluted sites. But that is not always, since little is known about microbiological processes during the biostimulation. Also, it is uncertain how oil pollution and its bioremediation affect marine microbes in the tropical and subtropical regions. So, we designed bioremediation experiments of oil pollution using subtropical seawater collected from the Ishigaki port in November, 2005. Four incubation sets were prepared: C-heavyoil + seawater (CHO + SW), CH + SW + Nitrate(N) + Phosphate(P) + Fe, Tetradecane(TD) + Biphenyl(BP) + Phenanthrene(PHE) + SW, and TD + BP + PHE + SW + N + P + Fe. Total microbial population, microbial diversity and quantitative community changes were monitored by direct counting, 16S rRNA gene sequencing and fluorescence dot blothybridization methods, respectively. The obtained results showed that the microbial growth was remarkably enhanced by the addition of nutrients, 345 and 750 times as much as the non-addition. The seawater sample used was characterized by the abundance of F..-proteobacleria. During the incubation, the community composition dramatically shifted from £-proleobacteria to y-proleobacleria. Among yproleobacteria, the aliphatic HC degrader Alcanivorax borkumensis increased. Especially, Alcanivorax borkumensis was estimated to be 64.5% of total microbes in set 2b at 10th incubation day. Comparing to the TO-added sample, however, the population was not so notable in CHO-added one. Further studies are necessary to detect representative aromatic He degraders.