Catabolic and Genetic Diversity of Degradative Bacteria from Fuel-Hydrocarbon Contaminated Aquifers

A BSTRACTSubsurface sediments were recovered from two aquifers contaminated with petroleum hydrocarbons in order to isolate and characterize indigenous microorganisms capable of biodegrading fuel-related compounds. These sediments had been previously studied using catabolic gene probes providing putative demonstration of significant biodegradation potential. Based on 16S rDNA sequence analysis, the isolates were phylogenetically similar to common soil microorganisms, including members of the genera Pseudomonas, Ralstonia, Burkholderia, Sphingomonas, Flavobacterium, and Bacillus. A total of 26 isolates were obtained using a vapor-plate enrichment technique with the volatile aromatic hydrocarbons toluene, ethylbenzene, p-xylene, naphthalene, and JP-4 jet fuel. JP-4, toluene, and ethylbenzene served as growth substrates for more than 80% of the isolates, while less than 10% of the organisms grew on the aromatic compounds benzene and o-xylene. Subsequent assays monitoring the evolution of (14)CO(2) indicated that only seven isolates were able to completely mineralize target compounds. One isolate, designated CAFB-naph-5, was able to completely mineralize the monoaromatic compounds salicylate and toluene, as well as the polyaromatic compound naphthalene. Molecular probing of the isolates showed four isolates hybridized with DNA probes targeting genes commonly associated with hydrocarbon-degrading bacteria. The isolates also demonstrated broad ability to grow in the presence of the antimicrobial agents streptomycin, tetracycline rifampicin, carbenicillin, nalidixic acid, kanamycin, and ampicillin. The results of the study demonstrate the biochemical and biodegradative capabilities of microorganisms isolated from contaminated aquifer systems and provide closure for indirect molecular monitoring of degradative potential in contaminated environments.