Caracterização genômica e da capacidade de degradar componentes de petróleo de uma linhagem de Burkholderia gladioli isolada do Lago de Coari (Coari-Amazonas)

Abstract

The petroleum is the main motor force that drives industry and transportation, it is a dark liquid, with variable composition and rich in hydrocarbons. A sample of oil can contain more than twenty thousand different substances, some of them toxic to organisms, to the environment and recalcitrant. The industrial necessity forces the countries to explore regions with hight biodiversity, exposing the environments to the risks involved during the extraction and transport to refine the crude oil. Thus, low-cost and environmentally-friendly technologies wich can reduce the petroleum spill impacts are critical. The Petroleum Province of Urucu is the largest terrestrial site of oil extraction in Brazil, belonging to the macro region of Coari, Amazonas. The objective of this work was to investigate whether the resistance to Urucu oil from the Burkholderia gladioli Coa14 strain isolated from Lake Coari may be indicative of its potential for use in bioremediation. For this, chemical analyzes, genomics, functional genomics and comparative genomics were performed. Initially, to test the action of the bacterium on the oil, an experiment was carried out where the lineage was inoculated in Erlenmeyers containing Bushell-Hass medium and 5% (V/V) petroleum as carbon and energy source, in a final volume of 100 ml. They were kept in shaker at 150 RPM and 37o C for 21 days. At the end of the period, the media were centrifuged for removal of the cells and extraction of the resulting oil. This was performed with hexane and the organic phases directly analyzed by GC-MS (SIM mode) for selective monitoring of n-alkanes. As control, flasks and procedures were used in the same conditions, without the inoculum. The loss by manipulation was estimated by extracting the oil from flasks in the condition of the controls at zero time. Growth assessment was estimated by measuring the optical density of cells obtained from 10 mL of the medium and resuspended in 0.85% NaCl. Then, Illumina sequencing and assembly of the genome to the draf level was performed. The assembly was followed by the annotation and investigation of the genes present related to the degradation of crude oil, as well as the reconstruction of the metabolic pathways that can be used by the microorganism for this purpose. As a final analysis, comparisons were made between the genome of the species in question with others of the same species, as well as genomes of bacteria of different species related to oil bioremediation and derivatives deposited publicly in NCBI. The strain reduced the volume of n-alkanes by 48% while its optical density increased from 1.0 to 2.2 in 21 days, indicating the possibility of using it in bioremediation strategies. It is resistant to three antibiotics in the cephalosporin group and susceptible to nine other related tests for Gram Negative bacteria. Sequencing revealed a genome of 8.4 Mb in size and several genes / pathways related to petroleum degradation. Comparative analyzes revealed similarity in genes for the initiation of the recalcitrant substances degradation regarding to bacteria of the same species and superiority in relation to other known bioremediators.