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Acidithiobacillus ferrooxidans is one of the most widely used microorganisms in bioleaching operations to recover copper from low-grade copper sulfide ores. This work aimed to investigate the relative expression of genes related to the iron uptake system when A. ferrooxidans LR was maintained in contact with chalcopyrite or bornite as the sole energy source. Real-time quantitative PCR analysis revealed that the presence of bornite had no effect on the expression of seven genes related to the siderophore-mediated Fe(III) uptake system, while in the presence of chalcopyrite the expression of the genes was up-regulated. Bioinformatic analysis of the genomic region where these genes were found revealed the existence of three new putative DNA-binding sequences for the ferric iron uptake transcriptional regulator (Fur). Electrophoretic mobility shift assays demonstrated that a purified A. ferrooxidans His-tagged Fur protein was able to bind in vitro to each of these putative Fur boxes, suggesting that Fur regulated the expression of these genes. The expression of fur and two known Fur-regulated genes, mntH and dsrK, was also investigated in the presence of chalcopyrite. While the expression of fur and mntH was up-regulated, the expression of dsrK was down-regulated. The low amount of ferrous iron in the medium was probably responsible for the up-regulation of fur and the genes related to the siderophore-mediated Fe(III) uptake system when A. ferrooxidans LR was kept in the presence of chalcopyrite. A homology model of the A. ferrooxidans Fur was constructed and revealed that the putative DNA-binding surface presents conserved positively charged residues, supporting a previously suggested mode of interaction with DNA. The up-regulation of fur and the siderophore-mediated Fe(III) uptake genes, and the down-regulation of dsrK suggest that in the presence of chalcopyrite Fur acts as a transcription inducer and repressor.

Biodegradation may result in physicochemical changes in crude oil and natural gas properties, being responsible for the decrease of saturated hydrocarbons and yielding heavy oil with low economic value. Studies on the diversity of microbial catabolic genes in oil reservoirs are scarce and could help to predict the potential of a petroleum sample to be biodegraded. The aim of this study was to evaluate the diversity of genes involved in hydrocarbon degradation in Brazilian petroleum samples (biodegraded and non-biodegraded) through the construction and analysis of gene libraries (alkane monooxygenase-alk, dioxygenase-ARHDs and 6-oxocyclohex-1-ene-1-carbonyl-CoA hydrolase-bamA). The results showed a differential distribution of catabolic genes between the sites, being the biodegraded oil more diverse for the alk and bamA genes. Sequences were similar to the alkB genes from Geobacillus thermoleovorans and several species of Acinetobacter, to ARHD genes from Pseudomonas spp. and two species of Burkholderia, and to bamA genes from deltaproteobacteria. Interestingly, most of the catabolic sequences recovered from both petroleum reservoirs grouped together forming distinct clusters in the phylogenetic tree reconstruction and may correspond to potentially new genes, possibly harbored by yet uncultivated microorganisms. This is the first report on the detection of alk, ARHD and bamA genes in petroleum reservoir environments, demonstrating the genetic potential of such microbial communities to biodegrade the oil.

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