Meta-Transcriptomes From Microcosms From a Cr Impacted Soil Provides Insights Into the Metabolic Response of the Microbial Populations to Acetate Stimulation

Abstract

Environmental contamination by Cr(VI) leaching from chromite ore processing residue (COPR) legacy disposal sites can pose a threat to human health. Under iron-reducing conditions, microbial activity can convert mobile and toxic Cr(VI) to less mobile and less toxic Cr(III); however, COPR waste is a very hostile environment for microbial life. Microcosms using soil from beneath a COPR disposal site were challenged with Cr(VI) with and without acetate to stimulate microbial metabolism. Geochemistry showed that when the microbial populations were reducing iron, Cr(VI) was also reduced, and 16S rRNA gene sequencing showed that the community composition evolved over the course of the experiment. Meta-transcriptome data revealed ~3% of transcripts were differentially regulated (p = 0.01) between the acetate amended and unamended systems, with twice as many transcripts downregulated by acetate. Gene ontology (GO) terms for processes involving the cell wall, cell periphery, plasma membrane and encapsulating structures as well as catabolic processes, especially carbohydrate metabolism, were significantly enriched in the unamended microcosm meta-transcriptome. Transcripts for alternative sigma (σ) factors and anti-σ factors were prominent among the differentially regulated genes. The study provides insight into how the provision of acetate shapes metabolic processes and life history strategies in an alkaline Cr(VI) impacted environment.

Keywords: COPR; acetate; alkaline environment; bioremediation; chromium; meta‐transcriptome.

FIGURE 1

Geochemical conditions in the unamended and acetate‐amended microcosms (A) Cr(VI) concentration, (B) nitrate concentration, (C) nitrite concentration, (D) percentage of the acid extractable Fe as Fe(II), and (E) sulphate concentration. Error bars represent one standard deviation of triplicates. Panels on right‐hand side show an expanded view of Days 0–6.

FIGURE 2

Phylogenetic diversity in the original soil and at the final sampling point in the unamended and acetate‐amended microcosm experiments determined by 16S rRNA gene sequencing: (A) distribution by phylum, and distribution by class within the (B) Proteobacteria, (C) Firmicutes and (D) Bacteroidetes.

FIGURE 3

Differential expression analysis of transcripts found in both systems (acetate‐amended system B/unamended system A). Negative expression coefficient means upregulation in the unamended system, whereas positive ones mean upregulation in the acetate‐amended system. Red and blue dots show the 1534 differentially expressed genes (DEGs) where p value < 0.01. The red dots identify the subset of those DEGs where the |coefficient| is also > 2 (1127 of 1534 DEGs). Yellow and grey dots show transcripts where differential expression was not significant (51,485 transcripts).

FIGURE 4

Unamended and acetate‐amended systems' gene expression profile. (A) Normalised expression (counts per million—CPM) heatmap of all 1534 DEGs depicting two clear sample clusters (columns' dendrogram) through unsupervised hierarchical clustering (HC) assessment. (B) Unsupervised multi‐dimensional scaling (MDS) plot of the ReadCounts of all 53 k genes across the six samples, corroborating the presence of both sample clusters observed from HC in panel A under a 44% explained variance on the first dimension. Replicates from the unamended system are A1, A2 and A3; replicates from the acetate‐amended system are B1, B2 and B3.

FIGURE 5

GO‐ and KEGG‐based gene enrichment analyses on the cluster of orthologous groups (COGs) derived from DEGs. (A) GO terms enriched in the unamended system, (B) GO term enriched in the acetate‐amended system, (C) KEGG pathways and orthologies enriched in the unamended system, (D) KEGG orthology enriched in the acetate‐amended system.

FIGURE 6

Heatmap of genes assigned to the phosphoserine phosphatase rsbU/P K07315 KEGG Orthology group showing differential expression according to taxa. Replicates from the unamended system are A1, A2 and A3; replicates from the acetate‐amended system are B1, B2 and B3. See text for details.

FIGURE 7

Transcripts associated with acetate metabolism in bacteria. (A) schematic diagram showing pathways of acetate assimilation/dissimilation. Transcripts of genes shown in red are displayed in the graph in B. (B) changes in transcript abundance between the acetate amended and unamended microcosms. Only transcripts showing significant differential expression (p < 0.01) are shown. Negative coefficients mean lower transcript abundance in the acetate‐amended microcosm compared to unamended microcosm. Created in BioRender. Baker (2025) https://BioRender.com.