Isolation of hydrocarbon-degrading extremely halophilic archaea from an uncontaminated hypersaline pond (Camargue, France).

Little information exists about the ability of halophilic archaea present in hypersaline environments to degrade hydrocarbons. In order to identify the potential actors of hydrocarbon degradation in these environments, enrichment cultures were prepared using samples collected from a shallow crystallizer pond with no known contamination history in Camargue, France, with n-alkanes provided as source of carbon and energy. Five alkane-degrading halophilic archaeal strains were isolated: one (strain MSNC 2) was closely related to Haloarcula and three (strains MSNC 4, MSNC 14, and MSNC 16) to Haloferax. Biodegradation assays showed that depending on the strain, 32 to 95% (0.5 g/l) of heptadecane was degraded after 30 days of incubation at 40 degrees C in 225 g/l NaCl artificial medium. One of the strains (MSNC 14) was also able to degrade phenanthrene. This work clearly shows for the first time the potential role of halophilic archaea belonging to the genera Haloarcula and Haloferax in the degradation of hydrocarbons in both pristine and hydrocarbon-contaminated hypersaline environments.

Alkane biodegradation and dynamics of phylogenetic subgroups of sulfate-reducing bacteria in an anoxic coastal marine sediment artificially contaminated with oil.

For 503 days, unoiled control and artificially oiled sediments were incubated in situ at 20m water depth in a Mediterranean coastal area. Degradation of the aliphatic fraction of the oil added was followed by GC-MS. At the same time, terminal restriction fragment length polymorphism (T-RFLP) of 16S rRNA encoding genes was used to detect dynamics in the sulfate-reducing bacteria (SRB) community in response to the oil contamination. Specific polymerase chain reaction (PCR) primer sets for five generic or suprageneric groups of SRB were used for PCR amplification of DNA extracted from sediments. During the experiment, hydrocarbons from C(17) to C(30) were significantly degraded even in strictly anoxic sediment layers. Of the five SRB groups, only two groups were detected in the sediments (control and oiled), namely the Desulfococcus-Desulfonema-Desulfosarcina-like group and the Desulfovibrio-Desulfomicrobium-like group. Statistical analysis of community patterns revealed dynamic changes over time within these two groups following the contamination. Significant differences in community patterns were recorded in artificially oiled compared with control sediments. Cloning and sequencing of 16S rRNA encoding genes performed after 503 days showed that many of the most abundant sequences were closely related to hydrocarbonoclastic SRB which could have played an active role in the observed biodegradation of aliphatic hydrocarbons. Results from the present study provide useful information on the dynamics of dominant SRB in heavily oil-contaminated sediments and their potential for anaerobic biodegradation for the treatment of spilled oil in anoxic marine environments.

Influence of bioturbation by the polychaete Nereis diversicolor on the structure of bacterial communities in oil contaminated coastal sediments.

Patterns of change in the structure of bacterial communities monitored by ribosomal intergenic spacer analysis (RISA) in oil contaminated sediments inhabited or not by the marine polychaete Nereis diversicolor were studied during 45 days under laboratory conditions. Results supported by principal component analysis showed a marked response of the bacterial communities to the oil contamination and to the presence of N. diversicolor. Phylogenetic affiliation of specific RISA bands showed that, in the contaminated sediments, the presence of the marine polychaetes favoured the development of bacteria which may play an active role in natural bioremediation processes of oil polluted environments.

Influence of growth phase on the phospholipidic fatty acid composition of two marine bacterial strains in pure and mixed cultures.

This in vitro study was conducted in order to determine the effects of hydrocarbons and growth phase on the phospholipid ester-linked fatty acid composition of two marine sedimentary hydrocarbon-degrading bacteria. These two strains, namely Corynebacterium sp. and Sphingomonas sp. 2MPII, were cultivated on either a simple soluble substrate (ammonium acetate) or a hydrocarbon (respectively n-eicosane and phenanthrene). The incubations were stopped at different times corresponding to point of lag (2 days), exponential (7 days) and stationary phases (21 and 56 days). The effects of growth phase and hydrophobic substrates were successfully demonstrated by a simple index, given as the sum of saturated fatty acids divided by the sum of unsaturated fatty acids ( summation operatorSFA/ summation operatorMUFA), ranging from 1.4 to 3, 0.3 to 0.6, and 0.5 to 1.0 for Corynebacterium sp., Sphingomonas sp. 2MPII, and mixed cultures, respectively. This result was validated by a principal component analysis. In pure cultures, the phospholipid fatty acid (PLFA) composition was strongly influenced by both the carbon source and the growth phase. Nevertheless, the two strains showed different "behaviors". For 2MPII, the main PLFA composition changes were observed at 2 days while they were progressive as a function of time for Corynebacterium sp. These differences could explain the evolution of PLFAs of mixed cultures.

Effects of spilled oil on bacterial communities of mediterranean coastal anoxic sediments chronically subjected to oil hydrocarbon contamination.

The effects of spilled oil on sedimentary bacterial communities were examined in situ at 20 m water depth in a Mediterranean coastal area. Sediment collected at an experimental site chronically subjected to hydrocarbon inputs was reworked into PVC cores with or without a massive addition of crude Arabian light oil ( approximately 20 g kg(-1) dry weight). Cores were reinserted into the sediment and incubated in situ at the sampling site (20 m water depth) for 135 and 503 days. The massive oil contamination induced significant shifts in the structure of the indigenous bacterial communities as shown by ribosomal intergenic spacer analysis (RISA). The vertical heterogeneity of the bacterial communities within the sediment was more pronounced in the oiled sediments particularly after 503 days of incubation. Response to oil of the deeper depth communities (8-10 cm) was slower than that of superficial depth communities (0-1 and 2-4 cm). Analysis of the oil composition by gas chromatography revealed a typical microbial alteration of n-alkanes during the experiment. Predominant RISA bands in oiled sediments were affiliated to hydrocarbonoclastic bacteria sequences. In particular, a 395-bp RISA band, which was the dominant band in all the oiled sediments for both incubation times, was closely related to hydrocarbonoclastic sulfate-reducing bacteria (SRB). These bacteria may have contributed to the main fingerprint changes and to the observed biodegradation of n-alkanes. This study provides useful information on bacterial dynamics in anoxic contaminated infralittoral sediments and highlights the need to assess more precisely the contribution of SRB to bioremediation in oil anoxic contaminated areas.