Project description:We analyzed the transcriptional response of the actinomycete Rhodococcus aetherivorans I24 to biphenyl and polychlorinated biphenyls (PCBs). This species has not been extensively exposed to PCBs, as it was first isolated from a toluene contaminated aquifer, rather than a site contaminated with polychlorinated hydrocarbons. Using a microarray targeting 3524 genes, we assessed gene expression in minimal medium supplemented with various substrates (e.g. PCBs) and in both PCB-contaminated and non-contaminated sediment slurries. Relative to the reference condition (minimal medium supplemented with glucose), 408 genes were up-regulated in the various treatments. In medium and in sediment, PCBs elicited the up-regulation of a common set of 100 genes, including chaperones (groEL), a superoxide dismutase (sodA), alkyl hydroperoxide reductase protein C (ahpC), and a catalase/peroxidase (katG). Analysis of the R. aetherivorans I24 genome sequence identified orthologs of many of the genes in the canonical biphenyl pathway, but very few of these genes were up-regulated in response to PCBs or biphenyl. This study is one of the first which utilizes microarrays to assess the transcriptional response of a soil bacterium to a pollutant under conditions which more closely resemble the natural environment. Our results indicate that the transcriptional response of R. aetherivorans I24 to PCBs, in both medium and sediment, is primarily directed towards reducing oxidative stress, rather than catabolism. In addition, the identification of numerous genes expressed in contaminated soil specifically may have implications for the development of biosensors. Finally, comparative genomic and transcriptomic analyses suggest that the mere presence of orthologs of the required enzymes may not be sufficient to confer a vigorous biphenyl/PCB metabolism. RNA was isolated from cells incubated in the following: sediment from a PCB-contaminated industrial site, uncontaminated sediment from a comparable site, and defined media supplemented with glucose (3 g/L), glucose and biphenyl (3 g/L, 4.5 Î¼M), or glucose and PCBs (3 g/L, 5 mg/L Aroclor 1254). In all cases, there were 3 biological replicates and 2 technical replicates (repeat hybridizations). A total of 3524 genes are represented on the arrays; of these, 41 and 176 are found on the plasmids pRA2 and pRA3, respectively. On average, there are 3 distinct 24nt probes per gene.

Project description:Fire disturbances are becoming more common, more intense, and further-reaching across the globe, with consequences for ecosystem functioning. Importantly, fire can have strong effects on the soil microbiome, including community and functional changes after fire, but surprisingly little is known regarding the role of soil fire legacy in shaping responses to recent fire. To address this gap, we conducted a manipulative field experiment administering fire across 32 soils with varying fire legacies, including combinations of 1-7 historic fires and 1-33 years since most recent fire. We analyzed soil metatranscriptomes, determining for the first time how fire and fire legacy interactively affect metabolically-active soil taxa, the microbial regulation of important carbon (C), nitrogen (N) and phosphorus (P) cycling, expression of carbohydrate-cycling enzyme pathways, and functional gene co-expression networks. Experimental fire strongly downregulated fungal activity while upregulating many bacterial and archaeal phyla. Further, fire decreased soil capacity for microbial C and N cycling and P transport, and drastically rewired functional gene co-expression. Perhaps most importantly, we highlight a novel role of soil fire legacy in regulation of microbial C, N, and P responses to recent fire. We observed a greater number of functional genes responsive to the interactive effects of fire and fire legacy than those affected solely by recent fire, indicating that many functional genes respond to fire only under certain fire legacy contexts. Therefore, without incorporating fire legacy of soils, studies will miss important ways that fire shapes microbial roles in ecosystem functioning. Finally, we showed that fire caused significant downregulation of carbon metabolism and nutrient cycling genes in microbiomes under abnormal soil fire histories, producing a novel warning for the future: human manipulation of fire legacies, either indirectly through global change-induced fire intensification or directly through fire suppression, can negatively impact soil microbiome functional responses to new fires.

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