# Comprehensive genomic analysis of a novel Bacillus cereus decomposing toluene potentially applicable in bioremediation

## TL;DR

A new strain of Bacillus cereus was found to effectively break down toluene, a common pollutant, making it useful for bioremediation of industrial wastewater.

## Contribution

The study identifies and characterizes a novel Bacillus cereus strain capable of toluene degradation with optimized environmental conditions and genomic insights.

## Key findings

- A novel Bacillus cereus strain was identified that efficiently degrades toluene under specific environmental conditions.
- Genomic analysis revealed key genes involved in toluene degradation pathways, including catechol 2,3-dioxygenase and flavin-dependent oxidoreductase.
- The strain achieved a 38.15% reduction in chemical oxygen demand when applied to petrochemical effluent.

## Abstract

Environmental pollution treatment is a worldwide challenge. Biological treatments are preferred to remedy the pollutants owing to higher efficiency and biocompatibility. Here, 18 indigenous bacterial strains with the usability of toluene as the sole carbon and energy source were screened, resulting in the selection of one isolate that was most capable of removing toluene. This isolate was characterized as a rod-shaped, gram-positive, spore-forming, halotolerant, catalase-positive, motile bacterium. The optimal temperature (40°C), pH (8), and NaCl concentration (50 g/L) for toluene removal were determined. Then, the experiment with four variables (petrochemical effluent dilution, carbon source, minerals, and amount of municipal wastewater) was designed using a Taguchi L16 orthogonal array. Ultimately, a 38.15% chemical oxygen demand decrease, as the best removal efficiency, was achieved. Gas chromatography–mass spectrometry analysis of bacterial metabolites assisted in recognizing the toluene degradation products, such as acetate and 2-propanone, indicating a meta-pathway of toluene decomposition. Next-generation sequencing strategy followed by core/pan-genome analysis indicated that the genome of the strain encompasses an assembly size of 5,154,622 bp with a content of 35.34% comprising 3,437 core and 2,258 accessory genes. Phylogenetic and multi-locus sequence typing studies confirmed that the strain belongs to the genus Bacillus and the species cereus. All the annotated and anticipated coding sequences were allocated to Kyoto Encyclopedia of Genes and Genomes routes. Accordingly, genes contributing to the toluene degradation pathway, including genes coding catechol 2, 3-dioxygenase, 4-oxalocrotonate isomerase, 2-hydroxymuconate-semialdehyde hydrolase, 2-keto-4-pentenoate hydratase, acetaldehyde dehydrogenase, alcohol dehydrogenase, flavocytochrome P450, anthranilate 3-monooxygenase (flavin adenine dinucleotide [FAD]), and flavin-dependent oxidoreductase, were ascertained. Herein, a general view of the GYRND102 strain is presented, which led to providing further insight into the decomposition mechanisms of aromatic pollutants by the strain for its effective application at the industrial level. Therefore, using this strain for pollutant removal will be cost-effective due to the simplicity of the method and the fact that it requires no special facilities or equipment.

The purpose of the present study was to develop a feasible process for the treatment of heavily contaminated Mahshahr petrochemical effluent. Since regular treatment of this effluent was inefficient, isolation of an indigenous bacterium with the ability to decompose toluene (the main pollutant of the effluent) and use it as a biological material to treat wastewater containing aromatic compounds on an industrial scale was very important.