Isolation And Identification Of PAHs-degrading Bacteria From Bei Bu Bay And Synergy Effect Between Co-cultured Strains

Polycyclic aromatic hydrocarbons are organic molecules composed of two ormore fused aromatic rings, which are persistent organic pollutants with ubiquitousdistribution in marine environment. Their cytotoxic, genotoxic, and carcinogenicpotential can lead to deleterious effects on the marine ecotope and human health.Previous studies showed that microbial degradation played a predominate role inPAHs removal in marine environment. Therefore, it is significant to screen andinvestigate PAHs-degrading bacteria from the marine environment.Therefore, it issignificant to screen and investigate PAHs-degrading bacteria from the marineenvironment.The present thesis is composed of the following two sections.Section one is responsible to explain the isolation，identification and biodiversityof PAHs-degrading bacteria from Beibu Bay: with phenanthrene and mixture ofphenanthrene and pyrene as selective carbon source, four different consortia havebeen enriched from Beibu Bay sediments. And then, to obtain the pure cultures of thePAHs-degradating bacteria, naphthalene vapor was used as the sole carbon and energysource for their growth through plate spread method. Then the resulting bacteria weresubjected to taxonomy analysis based on partial or whole16S ribosomal DNAsequences. Among the59resulting PAHs-degrading bacteria strains,35have beenidentified. The genera and species are as follows:21strains belonging to threeMarinobacter species, nine strains belonging to one Halomonas species, onePlanococcus strain, three Vibrio strains and one Bacillus strains. Marinobacterhydrocarbonoclasticus strains occupied54%of all the isolations; In section two, we describe the characteristics of three strains of genusCycloclasticus and the synergy effect between Cycloclasticus and Marinobacter:Three Cycloclasticus strains NY93E, PY97M and PY97N were isolated from YellowSea sediments. The PAHs degradation range of the three bacteria were determined byusing the sole carbon and energy source method, and their metabolic characteristicswere verified. We co-cultured Cycloclasticus sp. PY97M and Marinobacternanhaiticus D15-8WTto construct defined consortia, investigated its synergic effectin high molecular weight polycyclic aromatic hydrocarbons (pyrene and fluorantheneas representative compounds) biodegradation. To evaluate the consorium’s capabilityin degrading HMW-PAHs, comprehensive assessment have been implemented. Thedegradation rates of pyrene and fluoranthene and the end-product’s biotoxicity ofPAHs were measured by gas chromatography-mass spectrometry (GC-MS) andphotobacterium test respectively. Three Cycloclasticus strains were capable of using avariety of carbon sources, including the simple carbon source, PAHs intermediates,mononuclear aromatics and PAHs.Pyrene and fluoranthene, each at an initial concentration of0.1g/L, wassupplemented as sole source of carbon and energy for the growth of the definedconsortia throughout a21days’ incubation. The degradation rates by the consortiumcomposed of Cycloclasticus sp. PY97M and Marinobacter nanhaiticus D15-8WTcould reach67.40%and62.79%for pyrene and fluoranthene respectively，which was20.30%and20.29%higher than treatment of the PY97M pure culture. While theconsortium could utilize71.05%of pyrene and67.36%of fluoranthene respectivelythroughout a14days’ incubation, in which the mixed carbon source composed ofpyrene and fluoranthene was each at an initial concentration of0.1g/L. The results ofthe biotoxicity test also indicated that a significant reduction of the acute toxicity andgeno-toxicity after HMW-PAHs biodegradation was observed compared with theparent chemicals. The prominent synergic effect of Cycloclasticus strains and Marinobacter nanhaiticus strains suggested a great application potential in marineHMW-PAHs bioremediation.In conclusion, the thesis obtained numerous PAHs-degrading bacteria fromBeibu Bay and investigated their diversity, which is valuable for the research ofgenetic and biodiversity of the microbial resources in this sea aera. The study ofCycloclasticus strains’ metabolic characteristics in degrading HMW-PAHs wasdetermined through identifying their PAHs degradation range. The synergic effectbetween Cycloclasticus and co-cultured Marinobacter was also studied to explore themechanism of HMW-PAHs degradation.