Jacobs, N. J. | bioRxiv Science

bioRxiv2019

Pseudomonas aeruginosa ethanol oxidation by AdhA in low oxygen environments

Pseudomonas aeruginosa has a broad metabolic repertoire that facilitates its co-existence with different microbes. Many microbes secrete products that P. aeruginosa can then catabolize, including ethanol, a common fermentation product. Here, we show that under oxygen limiting conditions P. aeruginosa utilizes AdhA, an NAD-linked alcohol dehydrogenase, as a previously undescribed means for ethanol catabolism. In a rich medium containing ethanol, AdhA, but not the previously described PQQ-linked alcohol dehydrogenase, ExaA, oxidizes ethanol and leads to the accumulation of acetate in culture supernatants. AdhA-dependent acetate accumulation, and the accompanying decrease in pH, promotes P. aeruginosa survival in LB-grown stationary phase cultures. The transcription of adhA is elevated by hypoxia and in anoxic conditions, and we show that it is regulated by the Anr transcription factor. We have shown that lasR mutants have higher levels of Anr-regulated transcripts in low oxygen conditions compared to their wild type counterparts. Here, we show that a lasR mutant, when grown with ethanol, has an even larger decrease in pH than WT that is dependent on both anr and adhA. The large increase in AdhA activity similar to that of a strain expressing a hyperactive Anr-D149A variant. Ethanol catabolism in P. aeruginosa by AdhA supports growth on ethanol as a sole carbon source and electron donor in oxygen-limited settings and in cells growing by denitrification in anoxic conditions. This is the first demonstration of a physiological role for AdhA in ethanol oxidation in P. aeruginosa.\n\nImportanceEthanol is a common product of microbial fermentation, and the Pseudomonas aeruginosa response to and utilization of ethanol is relevant to our understanding of its role in microbial communities. Here, we report that the putative alcohol dehydrogenase, AdhA, is responsible for ethanol catabolism and acetate accumulation in low oxygen conditions and that it is regulated by Anr.

microbiology↗

bioRxiv2018

Profiling of bacterial and fungal microbial communities in cystic fibrosis sputum using RNA

Here, we report an approach to detect diverse bacterial and fungal taxa in complex samples by direct analysis of community RNA in one step using NanoString probe sets. We designed rRNA-targeting probe sets to detect forty two bacterial and fungal genera or species common in cystic fibrosis (CF) sputum, and demonstrated taxon-specificity of these probes as well as a linear response over more than three logs of input RNA. Culture-based analyses correlated qualitatively with relative abundance data on bacterial and fungal taxa obtained by NanoString and the analysis of serial samples demonstrated the use of this method to simultaneously detect bacteria and fungi and to detect microbes at low abundance without an amplification step. The relative abundances of bacterial taxa detected by analysis of RNA correlated with the relative abundances of the same taxa as measured by sequencing of the V4V5 region of the 16S rRNA gene amplified from community DNA from the same sample. We propose that this method may complement other methods designed to understand dynamic microbial communities, may provide information on bacteria and fungi in the same sample with a single assay, and, with further development, may provide quick and easily-interpreted diagnostic information on diverse bacteria and fungi at the genus or species level.\n\nImportanceHere we demonstrate the use of an RNA-based analysis of specific taxa of interest, including bacteria and fungi, within microbial communities. This multiplex method may be useful as a means to identify samples with specific combinations of taxa and to gain information on how specific populations vary over time and space or in response to perturbation. A rapid means to measure bacterial and fungal populations may aid in the study of host response to changes in microbial communities.

microbiology↗