Microbial ecology of arsenotrophic genes in Bangladesh environment: a possible genetic control on arsenic mobilization

Abstract

Arsenic (As) contamination is a severe health hazard in Southeast Asia, notably in Bangladesh. An ecologically sustainable biological arsenite oxidation technology is preferred due to pollution created by chemical methods. The hypothesis of our research work- arsenic-contaminated aquifers and soils contain arsenotrophic bacteria capable of transforming highly toxic arsenite (III) to less toxic arsenate (V), which play a critical role in the development of a sustainable, eco-friendly bioremediation model on a laboratory and pilot scale. Therefore, this study was designed to identify potential candidates that could significantly contribute to arsenic detoxification, accumulation, and immobilization while also providing a scientific foundation for future electrochemical sensor development. We applied both cultivation-dependent and independent (metagenomic) approaches for the study. 403 isolates were retrieved from fourteen As containing (0.01-0.5 mg/L) groundwater (GW) and twelve soil samples from arsenicprone areas- Munshiganj, Chandpur, and Bogura districts in Bangladesh. 29 GW isolates

were screened as arsenite transforming bacteria. Based on the 16S rRNA gene sequence,

five

taxonomic classes (α, β, γ, Firmicutes, Actinobacteria) were identified in heterotrophic and three (β, γ, Actinobacteria) in autotrophic GW bacteria. γ proteobacteria dominated the cultivated isolates. Common genera Lysinibacillus, Pseudomonas, Acinetobacter, Stenotrophomonas, Delftia, Enterobacter, Achromobacter, Bacillus, Staphylococcus, Paraburkholderia, Burkholderia, Comamonas, Klebsiella were found. We also identified some unique genera Ponticoccus, Kluyvera, Janibacter, Microbacterium, and Brevundimonas in As-contaminated water, especially in Bangladesh. Arsenic metabolizing genes arsenite efflux pumps (arsB) with high abundance and arsenite oxidase (aioA) genes were detected in cultured isolates, confirming their role in As resistance and biotransformation. They also revealed a wide range of MICarsenite concentrations ranging from 2 to 32 mM. We also assessed the arsenite transformation efficiency of arsenite oxidizing bacteria. As-affected groundwater microbiomes were identified, along with their interactions with arsenotrophic genes, virulence factor-associated genes (VFGs), antibiotic resistance genes (AGRs), and metabolic functional potentials. There was considerable heterogeneity in species richness and microbial community structure. Phyla proteobacteria (γ -proteobacteria), firmicutes, and acidobacteria dominate these diversities between culture-independent and dependent methods. The cultureindependent approach revealed considerable parallels with the culture-dependent method at the genus level. Pseudomonas, Acinetobacter, Stenotrophomonas, Delftia, Enterobacter, Achromobacter, Paraburkholderia, Burkholderia, Comamonas, and Klebsiella were detected using both techniques, proving their complementarity in detecting native population bacteria in As containing GW. MR pipeline explored the presence of arsenotrophic (arsB, acr3, arsD, arsH, arsR) arsenate reductase, etc.) and other associated functional genes in the metagenomes of both districts. Most of these genes were arsenical pump-specific, as indicated in our culture-dependent study. The soil microbiome was strongly linked to the GW microbiome based on bacterial abundance, diversity, and arsenotrophic genes distribution. The present study selected and explored highly arsenite-resistant novel bacteria Achromobacter xylosoxidans BHW-15 with good As (III) transformation capability for electrochemical As species detection and bioremediation. Scanning Electron Microscopy (SEM) analysis evidenced the intracellular As absorption capability of A. xylosoxidans BHW15 and established a substantial correlation with its MIC value. Arsenite oxidase (aioA) gene expression was also assessed to observe the As (III) oxidation efficiency. Additionally, the immobilized whole-cell demonstrated As (III) conversion throughout 18 days. We developed a modified GCE/P-Arg/ErGO-AuNPs electrode that effectively sensed and evaluated the conversion of As (III) to As (V) by electron acceptance revealing the existence of a functioning As oxidase enzyme in the cells. We reported the electrochemical Astransformation

in Achromobacter sp. for the first time. Our study found promising arsenotrophic bacteriomes whose genetic profile will be helpful to develop arsenic detoxification strategies. The data from this investigation may enable the future development of a cost-effective, environmentally friendly biosensor for arsenic species detection.