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The arsRBC operon encodes a three-protein arsenic resistance system. ArsR regulates the transcription of the operon, while ArsB and ArsC are involved in exporting trivalent arsenic and reducing pentavalent arsenic, respectively. Previous research into Agrobacterium tumefaciens 5A has demonstrated that ArsR has regulatory control over a wide range of metal-related proteins and metabolic pathways. We hypothesized that ArsR has broad regulatory control in other Gram-negative bacteria and set out to test this. Here, we use differential proteomics to investigate changes caused by the presence of the arsR gene in human microbiome-relevant Escherichia coli during arsenite (AsIII) exposure. We show that ArsR has broad-ranging impacts such as the expression of TCA cycle enzymes during AsIII stress. Additionally, we found that the Isc [Fe-S] cluster and molybdenum cofactor assembly proteins are upregulated regardless of the presence of ArsR under these same conditions. An important finding from this differential proteomics analysis was the identification of response mechanisms that were strain-, ArsR-, and arsenic-specific, providing new clarity to this complex regulon. Given the widespread occurrence of the arsRBC operon, these findings should have broad applicability across microbial genera, including sensitive environments such as the human gastrointestinal tract. [ABSTRACT FROM AUTHOR]

'Thermobaculum terrenum' Botero et al. 2004 is the sole species within the proposed genus 'Thermobaculum'. Strain YNP1(T) is the only cultivated member of an acid tolerant, extremely thermophilic species belonging to a phylogenetically isolated environmental clone group within the phylum Chloroflexi. At present, the name 'Thermobaculum terrenum' is not yet validly published as it contravenes Rule 30 (3a) of the Bacteriological Code. The bacterium was isolated from a slightly acidic extreme thermal soil in Yellowstone National Park, Wyoming (USA). Depending on its final taxonomic allocation, this is likely to be the third completed genome sequence of a member of the class Thermomicrobia and the seventh type strain genome from the phylum Chloroflexi. The 3,101,581 bp long genome with its 2,872 protein-coding and 58 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

Arsenic (As) and mercury (Hg) were examined in the Yellowstone Lake food chain, focusing on two lake locations separated by approximately 20 km and differing in lake floor hydrothermal vent activity. Sampling spanned from femtoplankton to the main fish species, Yellowstone cutthroat trout and the apex predator lake trout. Mercury bioaccumulated in muscle and liver of both trout species, biomagnifying with age, whereas As decreased in older fish, which indicates differential exposure routes for these metal(loid)s. Mercury and As concentrations were higher in all food chain filter fractions (0.1‐, 0.8‐, and 3.0‐μm filters) at the vent‐associated Inflated Plain site, illustrating the impact of localized hydrothermal inputs. Femtoplankton and picoplankton size biomass (0.1‐ and 0.8‐μm filters) accounted for 30%–70% of total Hg or As at both locations. By contrast, only approximately 4% of As and <1% of Hg were found in the 0.1‐μm filtrate, indicating that comparatively little As or Hg actually exists as an ionic form or intercalated with humic compounds, a frequent assumption in freshwaters and marine waters. Ribosomal RNA (18S) gene sequencing of DNA derived from the 0.1‐, 0.8‐, and 3.0‐μm filters showed significant eukaryote biomass in these fractions, providing a novel view of the femtoplankton and picoplankton size biomass, which assists in explaining why these fractions may contain such significant Hg and As. These results infer that femtoplankton and picoplankton metal(loid) loads represent aquatic food chain entry points that need to be accounted for and that are important for better understanding Hg and As biochemistry in aquatic systems. Environ Toxicol Chem 2023;42:225–241. © 2022 SETAC [ABSTRACT FROM AUTHOR]

Transposon Tn5-B22 mutagenesis was used to identify genetic determinants required for arsenite [As(III)] oxidation in an Agrobacterium tumefaciens soil isolate, strain 5A. In one mutant, the transposon interrupted modB, which codes for the permease component of a high-affinity molybdate transporter. In a second mutant, the transposon insertion occurred in mrpB, which is part of a seven-gene operon encoding an Mrp-type [Na.sup.+]:[H.sup.+] antiporter complex. Complementation experiments with mod and mrp operons PCR cloned from the genomesequenced A. tumefaciens strain C58 resulted in complementation back to an As(III)-oxidizing phenotype, confirming that these genes encode activities essential for As(III) oxidation in this strain of A. tumefaciens. As expected, the mrp mutant was extremely sensitive to NaCI and LiCI, indicating that the Mrp complex in A. tumefaciens is involved in [Na.sup.+] circulation across the membrane. Gene expression studies (lacZ reporter and reverse transcriptase PCR experiments) failed to show evidence of transcriptional regulation of the mrp operon in response to As(III) exposure, whereas expression of the mod operon was found to be up-regulated by As(III) exposure. In each mutant, the loss of As(III)-oxidizing capacity resulted in conversion to an arsenate [As(V)]-reducing phenotype. Neither mutant was more sensitive to As(III) than the parental strain.

Seminal regulatory controls of microbial arsenite [As(III)] oxidation are described in this study. Transposon mutagenesis of Agrobacterium tumefaciens identified genes essential for As(III) oxidation, including those coding for a two-component signal transduction pair. The transposon interrupted a response regulator gene (referred to as aoxR), which encodes an ntrC-like protein and is immediately downstream of a gene (aoxS) encoding a protein with primary structural features found in sensor histidine kinases. The structural genes for As(III) oxidase (aoxAB), a c-type cytochrome ([cytc.sub.2]), and molybdopterin biosynthesis (chlE) were downstream of aoxR. The mutant could not be complemented by aoxSR in trans but was complemented by a clone containing aoxS-aoxR-aoxA-aoxB-[cytc.sub.2] and consistent with reverse transcriptase (RT) PCR experiments, which demonstrated these genes are cotranscribed as an operon. Expression of aoxAB was monitored by RT-PCR and found to be up-regulated by the addition of As(III) to cell cultures. Expression of aoxAB was also controlled in a fashion consistent with quorum sensing in that (i) expression of aoxAB was absent in As(III)-unexposed early-log-phase cells but was observed in As(III)-unexposed, late-log-phase cells and (ii) treating As(III)-unexposed, early-log-phase cells with ethyl acetate extracts of As(III)-unexposed, late-log-phase culture supernatants also resulted in aoxAB induction. Under inducing conditions, aoxS expression was readily observed in the wild-type strain but significantly reduced in the mutant, indicating that AoxR is autoregulatory and at least partially controls the expression of the aox operon. In summary, regulation of A. tumefaciens As(III) oxidation is complex, apparently being controlled by As(III) exposure, a two-component signal transduction system, and quorum sensing.

The impact of arsenite [As(III)] on several levels of cellular metabolism and gene regulation was examined in Pseudomonas aeruginosa. P. aeruginosa isogenic mutants devoid of antioxidant enzymes or defective in various metabolic pathways, DNA repair systems, metal storage proteins, global regulators, or quorum sensing circuitry were examined for their sensitivity to As(III). Mutants lacking the As(III) translocator (ArsB), superoxide dismutase (SOD), catabolite repression control protein (Crc), or glutathione reductase (Gor) were more sensitive to As(III) than wild-type bacteria. The MICs of As(III) under aerobic conditions were 0.2, 0.3, 0.8, and 1.9 mM for arsB, sodA sodB, ctc, and gor mutants, respectively, and were 1.5- to 13-fold less than the MIC for the wild-type strain. A two-dimensional gel/matrix-assisted laser desorption ionization--time of flight analysis of As(III)-treated wild-type bacteria showed significantly (>40-fold) increased levels of a heat shock protein (IbpA) and a putative allo-threonine aldolase (GlyI). Smaller increases (up to 3.1-fold) in expression were observed for acetyl-coenzyme A acetyltransferase (AtoB), a probable aldehyde dehydrogenase (KauB), ribosomal protein L25 (RpIY), and the probable DNA-binding stress protein (PA0962). In contrast, decreased levels of a heme oxygenase (HemO/PigA) were found upon As(III) treatment. Isogenic mutants were successfully constructed for six of the eight genes encoding the aforementioned proteins. When treated with sublethal concentrations of As(III), each mutant revealed a marginal to significant lag period prior to resumption of apparent normal growth compared to that observed in the wild-type strain. Our results suggest that As(III) exposure results in an oxidative stress-like response in P. aeruginosa, although activities of classic oxidative stress enzymes are not increased. Instead, relief from As(III)-based oxidative stress is accomplished from the collective activities of ArsB, glutathione reductase, and the global regulator Crc. SOD appears to be involved, but its function may be in the protection of superoxide-sensitive sulfhydryl groups.

A protocol that couples poly(A) polymerase with a multifunctional reverse transcriptase (RT) for targeted cloning of environmental prokaryotic cDNA is described. The reverse transcriptase PCR (RT-PCR) cDNA synthesis kit demonstrates the feasibility of directly accessing prokaryote RNA (rRNA and/or mRNA) in environmental samples, but the results also show potentially important problems.

Water samples from the Norris Geyser Basin in Yellowstone National Park are taken at regular intervals from the spring's mouth, and measured for levels of arsenic, arsenite, and arsenate. The role of three microbial communities are studied in relation to arsenic oxidation, and graphed.

Research demonstrates that the oxidative stress response in pseudomonas aeruginosa is not only influenced by iron availability but is also controlled by quorum sensing regulatory network. Data indicate that quorum sensing gene expression is governed by nutritional status and it is dynamic in biofilms.

Iron seems to be a requirement for the production of catalase in Pseudomonas aeruginosa biofilms but not in planktonic cells. When bacteria are grown in culture media they are planktonic, but research shows the most bacteria prefer to grow in biofilms.

Results indicate that Lon protease in Sinorhizobium meliloti regulates the turnover of constitutively expressed proteins. Data also suggest that such a regulation is essential as otherwise nodule formation and growth of the bacteria are affected.

Rhizobium tropici contains two phosphate transport systems, a low-affinity system and a high-affinity system. Both are induced by phosphate starvation, inhibited by ATPase inhibitors, and are unidirectional.

Isolation of the gltA gene that encodes Sinorhizobium meliloti has been carried out. The results of the liquid chromatography analysis of the succinylated succinoglycan are discussed.

Research was conducted to determine the role of catalase enzymes in protecting bacterial biofilms against hydrogen peroxide biocide. Results indicate besides the protective action of the catalases, other mechanisms also appear to influence resistance of biofilms to hydrogen peroxide.

A research was conducted to study the responses of Rhizobium meliloti to phosphate stress and to determine the phosphate level at which R. meliloti induces its P stress response. Transposon Tn5 was conjugated from Escherichia coli S17-1 into R. meliloti via the suicide vector pSUP2021. Alfalfa seedlings in sterile growth box units were utilized to determine the symbiosis phenotype of the phoB mutant. Results indicated that the loss of a functional PhoB did not disturb the symbiotic function of R. meliloti regardless of the available phosphate.