Biological processes as bioremediation hold a promising alternative to overcome pollution accumulation worldwide. There are commercial products available for bioremediation, most based on strains from few species of Bacillus and Pseudomonas. In the present work, both direct isolation and adaptive evolution experiments (AEx) were set-up to screen new and improved microbial biodegraders towards fat, oils and grease (FOG) and polycyclic aromatic hydrocarbon (PAH) contaminants.
By the direct isolation, 196 microorganisms were screened and fingerprinted by PCR with primers csM13 and PH, allowing the selection of two bacterial strains, identified as Aeromonas sp. and Staphylococcus sp., with good biodegradation capacity for FOG residues. The AEx focused on PAH pollution and insoluble triglycerides, and were conducted for 110 cycles, creating a collection of 455 microorganisms (bacteria and yeast) by selective solid media isolation. All isolates were analysed by PCR-fingerprint with multiple primers (csM13, PH and [GTG]5), to assess genomic diversity and detect indistinguishable isolates. The microbial population changes during AEx were also followed by third generation sequencing, to assess limitations and bias of the isolation strategy. A total of 19 microorganisms were selected and identified as belonging to Acinetobacter, Brevundimonas, Paraburkholderia, Pigmentiphaga, Pseudomonas, Sphingobacterium and Yarrowia, genera with known biodegradative ability. Methods were developed for effective pollutant removal quantification to compare the selected isolates. Concerning FOG residues, a gravimetric approach showed that the two best isolates removed 90% and 55% of FOG, after 5 days assay. For PAH, a HPLC quantification method revealed that the two best isolates were capable to remove 40% of anthracene and 60% phenanthrene, after 5 days assay.
These results pave the way for the integration of the selected strains into new products, with high added value and effectiveness as an ecological bioremediation alternative, to deal with recalcitrant pollutants as FOG and PAH
