title: Phylogenetic analysis of aerobic freshwater and marine enrichment cultures efficient in hydrocarbon degradation: effect of profiling method.
abstract: Aerobically grown enrichment cultures derived from hydrocarbon-contaminated seawater and freshwater sediments were generated by growth on crude oil as sole carbon source. Both cultures displayed a high rate of degradation for a wide range of hydrocarbon compounds. The bacterial species composition of these cultures was investigated by PCR of the 16S rDNA gene using multiple primer combinations. Near full-length 16S rDNA clone libraries were generated and screened by restriction analysis prior to sequence analysis. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was carried out using two other PCR primer sets targeting either the V3 or V6-V8 regions, and sequences derived from prominent DGGE bands were compared to sequences obtained via cloning. All data sets suggested that the seawater culture was dominated by alpha-subgroup proteobacteria, whereas the freshwater culture was dominated by members of the beta- and gamma-proteobacteria. However, the V6-V8 primer pair was deficient in the recovery of Sphingomonas-like 16S rDNA due to a 3' terminal mismatch with the reverse primer. Most 16S rDNA sequences recovered from the marine enrichment were not closely related to genera containing known oil-degrading organisms, although some were detected. All methods suggested that the freshwater enrichment was dominated by genera containing known hydrocarbon-degrading species.
authors.affiliation: Center for Environmental Biotechnology, University of Tennessee, Knoxville 37932-2575, USA.
authors.forename: Y J
authors.initials: YJ
authors.lastname: Chang
authors.forename: J R
authors.initials: JR
authors.lastname: Stephen
authors.forename: A P
authors.initials: AP
authors.lastname: Richter
authors.forename: A D
authors.initials: AD
authors.lastname: Venosa
authors.forename: J
authors.initials: J
authors.lastname: Brüggemann
authors.forename: S J
authors.initials: SJ
authors.lastname: Macnaughton
authors.forename: G A
authors.initials: GA
authors.lastname: Kowalchuk
authors.forename: J R
authors.initials: JR
authors.lastname: Haines
authors.forename: E
authors.initials: E
authors.lastname: Kline
authors.forename: D C
authors.initials: DC
authors.lastname: White
mesh_terms: D000332:Aerobiosis; D003470:Culture Media; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D004586:Electrophoresis / Q000379:methods; D005618:Fresh Water / Q000382:microbiology; D020459:Genes, rRNA; D019015:Geologic Sediments / Q000382:microbiology*; D006838:Hydrocarbons / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods; D020560:Proteobacteria / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012623:Seawater / Q000382:microbiology; D014871:Water Microbiology; D014874:Water Pollutants, Chemical / Q000378:metabolism
chemical_list: D003470:Culture Media; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D006838:Hydrocarbons; D012336:RNA, Ribosomal, 16S; D014874:Water Pollutants, Chemical