COMPOSITIONAL CHANGES FOR HYDROCARBON-DEGRADING MICROORGANISMS IN MARSH SEDIMENT FROM GRAND ISLE, LOUISIANA, 2011-2014

Some microorganisms, including Alcanivorax spp. and Pseudomonas spp., have the ability to degrade hydrocarbons, like normal (n)-alkanes, by specific enzyme pathways, such as alkane hydroxylase. Salt marshes along the Louisiana coast were significantly impacted by the Deepwater Horizon oil spill in 2010, in which 779 million liters of oil were released into the Gulf of Mexico. This study surveyed bacterial community compositions from a salt marsh at Grand Isle, Louisiana, from 2011-2014, and evaluated taxonomic changes in alkane hydroxylase genes. Total n-alkanes measured at Grand Isle varied from pre-spill concentrations at 1 mg/kg to over 25 mg/kg in October 2013. Sediment samples were collected in the spring and fall seasons, from 0-1 cm and 4-5 cm depths. Total environmental DNA was extracted and used for 454 tag pyrosequencing of 16S rRNA and the alkB gene, an alkane hydroxylase gene that can be used to assess alkane-degrading communities. Bacterial diversity from 16S rRNA genes was determined using the mothur computer pipeline, and translated amino acid sequences from alkB genes were done using the mothur and FunGene pipelines, and Blast2GO bioinformatics platform. The hypothesis of this study was that the alkane-degrading community would vary through time, and with sediment depth, because of temporal changes in n-alkanes. The taxonomic affiliations of alkB genes retrieved from surface sediment were more diverse than those from the deeper sediments. Alphaproteobacteria, Deltaproteobacteria, and Gammaproteobacteria dominated the surface, and only Gammaproteobacteria, belonging to the orders Oceanospirillales and Pseudomonadales, dominated at depth. These results did not match 16S rRNA data, where Deltaproteobacteria were abundant at all depths, although the relative abundances of Alphaproteobacteria at the surface increased through time, and Gammaproteobacteria increased with depth. Canonical correspondence analyses indicated that sediment depth explained 90% of the variance among alkane-degrading community compositions and total n-alkane concentrations explained 9% of the variance. These results suggest that the capacity for bacterial n-alkane degradation may change through time, and that microbial responses to hydrocarbon contamination will vary with sediment depth.