Characterization of bacterial consortia for its use in bioremediation of gas-oil contaminated Antarctic soils

RUBERTO L.; VAZQUEZ S.; MESTRE MC; NOGALES B; CHRISTIE-OLEZA J; BOSCH R; MAC CORMACK W.

Palma de Mallorca, Islas Baleares, España

Congreso; Third Meeting on Environmental Biotechnology and Engineering (3 IMEBE); 2008

Success of bioaugmentation of chronically-contaminated soils is controversial, mainly because the inocula are frequently unable to establish in the matrix under bioremediation. In Antarctica, the environmental conditions and the restriction for the introduction of non-autochthonous organisms (imposed by the Antarctic Treaty) prevent inoculation with foreign bacteria. For these reasons, our aim was to isolate and characterize hydrocarbon-degrading Antarctic bacterial consortia, totest their capacity to enhance soil bioremediation and to evaluate the effect of two different inoculum sizes on remediation rate. Bacterial consortia M10 and J13 were isolated by enrichment cultures from contaminated Antarctic soils. Their degradative capacity on hydrocarbons was evaluated in-vitro and by detection (using PCR and blotting) of genes (alkB, nahAc, xylE) involved in hydrocarbons catabolic pathways. Consortia showed similar T-RFLP profiles, suggesting the presence of the same phylotypes. Although composition and degradative profiles were similar, field trials proved that consortia differed in their ability to establish in the soil, J13 being less efficient to remove hydrocarbons. Thus, M10 was selected for further characterization. Total 16S rDNA was amplified by PCR and cloned. Sixty clones were screened and grouped by similar AluI ARDRA profile, sequencing one representative of each phylotype. T-RFLP of clones showed that all the peaks from the entire consortia were recovered. To avoid the problems of reproducibility in preparation of the inoculum from M10 stock, a culture-dependent approach was also used to isolate its components for further preparation of a mixed-culture with similar degradative performance than M10. Analizing phenotypic and molecular traits, Pseudomonas,Stenotrophomonas and Sphingobacterium/Pedobacter were detected. By cloning, also Brevundimonas was found as minority representative. Effect of bioaugmentation with M10 on remediation of sterile and non-sterile hydrocarboncontaminated biostimulated soils was analyzed on site using microcosms inoculated at two different levels (10E6 and 10E9 UFC/g). An only-biostimulated and an untreated soil were also tested. Total and degrading counts and hydrocarbon concentration were measured. Results showed that, in the presence of the natural microbiota, only bioaugmentation with a bacterial load several orders of magnitude higher than the number of autochthonous bacteria will improve hydrocarbon removal, shortening the length of the bioremediation process.