Integration of choline geranate into electrospun protein scaffolds affords antimicrobial activity to biomaterials used for cutaneous wound healing.

Wound healing attempts to maintain homeostasis in the wound while minimizing the risk of infection to the tissue by foreign agents, such as opportunistic bacterial pathogens. Biofilms established by these pathogens are a common cause of chronic infections that slow the healing process. Preparation of skin wound healing devices comprised of electrospun proteins associated with skin have been shown to accelerate the healing process relative to conventional wound dressings. In this work, we have developed electrospinning methods to incorporate the antimicrobial ionic liquid/deep eutectic solvent choline geranate (CAGE) into these devices. Integration of CAGE into the dressing material was verified via 1 H nuclear magnetic resonance spectrometry, and the effect on the material property of the resultant devices were assessed using scanning electron microscopy. CAGE-containing devices demonstrate a concentration-dependent inactivation of exogenously applied solutions of both gram-positive and gram-negative pathogens (Enterococcus sp and Pseudomonas aeruginosa, respectively), but maintain their ability to serve as a compatible platform for proliferation of human dermal neonatal fibroblasts.

Scope and efficacy of the broad-spectrum topical antiseptic choline geranate.

Choline geranate (also described as Choline And GEranic acid, or CAGE) has been developed as a novel biocompatible antiseptic material capable of penetrating skin and aiding the transdermal delivery of co-administered antibiotics. The antibacterial properties of CAGE were analyzed against 24 and 72 hour old biofilms of 11 clinically isolated ESKAPE pathogens (defined as Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanii, Pseudomonas aeruginosa, and Enterobacter sp, respectively), including multidrug resistant (MDR) isolates. CAGE was observed to eradicate in vitro biofilms at concentrations as low as 3.56 mM (0.156% v:v) in as little as 2 hours, which represents both an improved potency and rate of biofilm eradication relative to that reported for most common standard-of-care topical antiseptics in current use. In vitro time-kill studies on 24 hour old Staphylococcus aureus biofilms indicate that CAGE exerts its antibacterial effect upon contact and a 0.1% v:v solution reduced biofilm viability by over three orders of magnitude (a 3log10 reduction) in 15 minutes. Furthermore, disruption of the protective layer of exopolymeric substances in mature biofilms of Staphylococcus aureus by CAGE (0.1% v:v) was observed in 120 minutes. Insight into the mechanism of action of CAGE was provided with molecular modeling studies alongside in vitro antibiofilm assays. The geranate ion and geranic acid components of CAGE are predicted to act in concert to integrate into bacterial membranes, affect membrane thinning and perturb membrane homeostasis. Taken together, our results show that CAGE demonstrates all properties required of an effective topical antiseptic and the data also provides insight into how its observed antibiofilm properties may manifest.

Enamide Prodrugs of Acetyl Phosphonate Deoxy-d-xylulose-5-phosphate Synthase Inhibitors as Potent Antibacterial Agents.

To fight the growing threat of antibiotic resistance, new antibiotics are required that target essential bacterial processes other than protein, DNA/RNA, and cell wall synthesis, which constitute the majority of currently used antibiotics. 1-Deoxy-d-xylulose-5-phosphate (DXP) synthase is a vital enzyme in bacterial central metabolism, feeding into the de novo synthesis of thiamine diphosphate, pyridoxal phosphate, and essential isoprenoid precursors isopentenyl diphosphate and dimethylallyl diphosphate. While potent and selective inhibitors of DXP synthase in vitro activity have been discovered, their antibacterial activity is modest. To improve the antibacterial activity of selective alkyl acetylphosphonate (alkylAP) inhibitors of DXP synthase, we synthesized peptidic enamide prodrugs of alkylAPs inspired by the natural product dehydrophos, a prodrug of methyl acetylphosphonate. This prodrug strategy achieves dramatic increases in activity against Gram-negative pathogens for two alkylAPs, butyl acetylphosphonate and homopropargyl acetylphosphonate, decreasing minimum inhibitory concentrations against Escherichia coli by 33- and nearly 2000-fold, respectively. Antimicrobial studies and LC-MS/MS analysis of alkylAP-treated E. coli establish that the increased potency of prodrugs is due to increased accumulation of alkylAP inhibitors of DXP synthase via transport of the prodrug through the OppA peptide permease and subsequent amide hydrolysis. This work demonstrates the promise of targeting DXP synthase for the development of novel antibacterial agents.

Growth medium-dependent antimicrobial activity of early stage MEP pathway inhibitors.

The in vivo microenvironment of bacterial pathogens is often characterized by nutrient limitation. Consequently, conventional rich in vitro culture conditions used widely to evaluate antibacterial agents are often poorly predictive of in vivo activity, especially for agents targeting metabolic pathways. In one such pathway, the methylerythritol phosphate (MEP) pathway, which is essential for production of isoprenoids in bacterial pathogens, relatively little is known about the influence of growth environment on antibacterial properties of inhibitors targeting enzymes in this pathway. The early steps of the MEP pathway are catalyzed by 1-deoxy-d-xylulose 5-phosphate (DXP) synthase and reductoisomerase (IspC). The in vitro antibacterial efficacy of the DXP synthase inhibitor butylacetylphosphonate (BAP) was recently reported to be strongly dependent upon growth medium, with high potency observed under nutrient limitation and exceedingly weak activity in nutrient-rich conditions. In contrast, the well-known IspC inhibitor fosmidomycin has potent antibacterial activity in nutrient-rich conditions, but to date, its efficacy had not been explored under more relevant nutrient-limited conditions. The goal of this work was to thoroughly characterize the effects of BAP and fosmidomycin on bacterial cells under varied growth conditions. In this work, we show that activities of both inhibitors, alone and in combination, are strongly dependent upon growth medium, with differences in cellular uptake contributing to variance in potency of both agents. Fosmidomycin is dissimilar to BAP in that it displays relatively weaker activity in nutrient-limited compared to nutrient-rich conditions. Interestingly, while it has been generally accepted that fosmidomycin activity depends upon expression of the GlpT transporter, our results indicate for the first time that fosmidomycin can enter cells by an alternative mechanism under nutrient limitation. Finally, we show that the potency and relationship of the BAP-fosmidomycin combination also depends upon the growth medium, revealing a striking loss of BAP-fosmidomycin synergy under nutrient limitation. This change in BAP-fosmidomycin relationship suggests a shift in the metabolic and/or regulatory networks surrounding DXP accompanying the change in growth medium, the understanding of which could significantly impact targeting strategies against this pathway. More generally, our findings emphasize the importance of considering physiologically relevant growth conditions for predicting the antibacterial potential MEP pathway inhibitors and for studies of their intracellular targets.

Challenges and Hallmarks of Establishing Alkylacetylphosphonates as Probes of Bacterial 1-Deoxy-d-xylulose 5-Phosphate Synthase.

1-Deoxy-d-xylulose 5-phosphate (DXP) synthase catalyzes the thiamin diphosphate (ThDP)-dependent formation of DXP from pyruvate and d-glyceraldehyde 3-phosphate. DXP is at a metabolic branch point in bacteria, feeding into the methylerythritol phosphate pathway to indispensable isoprenoids and acting as a precursor for biosynthesis of essential cofactors in central metabolism, pyridoxal phosphate and ThDP, the latter of which is also required for DXP synthase catalysis. DXP synthase follows a unique random sequential mechanism and possesses an unusually large active site. These features have guided the design of sterically demanding alkylacetylphosphonates (alkylAPs) toward the development of selective DXP synthase inhibitors. alkylAPs studied here display selective, low µM inhibitory activity against DXP synthase. They are weak inhibitors of bacterial growth in standard nutrient rich conditions. However, bacteria are significantly sensitized to most alkylAPs in defined minimal growth medium, with minimal inhibitory concentrations (MICs) ranging from low µM to low mM and influenced by alkyl-chain length. The longest analog (C8) displays the weakest antimicrobial activity and is a substrate for efflux via AcrAB-TolC. The dependence of inhibitor potency on growth environment emphasizes the need for antimicrobial screening conditions that are relevant to the in vivo microbial microenvironment during infection. DXP synthase expression and thiamin supplementation studies offer support for DXP synthase as an intracellular target for some alkylAPs and reveal both the challenges and intriguing aspects of these approaches to study target engagement.

Draft Nuclear Genome Sequence of the Liquid Hydrocarbon-Accumulating Green Microalga Botryococcus braunii Race B (Showa).

Botryococcus braunii has long been known as a prodigious producer of liquid hydrocarbon oils that can be converted into combustion engine fuels. This draft genome for the B race of B. braunii will allow researchers to unravel important hydrocarbon biosynthetic pathways and identify possible regulatory networks controlling this unusual metabolism.

Rapid Thermostabilization of Bacillus thuringiensis Serovar Konkukian 97-27 Dehydroshikimate Dehydratase through a Structure-Based Enzyme Design and Whole Cell Activity Assay.

Thermostabilization of an enzyme with complete retention of catalytic efficiency was demonstrated on recombinant 3-dehydroshikimate dehydratase (DHSase or wtAsbF) from Bacillus thuringiensis serovar konkukian 97-27 (hereafter, B. thuringiensis 97-27). The wtAsbF is relatively unstable at 37 °C, in vitro (t1/237 = 15 min), in the absence of divalent metal. We adopted a structure-based design to identify stabilizing mutations and created a combinatorial library based upon predicted mutations at specific locations on the enzyme surface. A diversified asbF library (∼2000 variants) was expressed in E. coli harboring a green fluorescent protein (GFP) reporter system linked to the product of wtAsbF activity (3,4-dihydroxybenzoate, DHB). Mutations detrimental to DHSase function were rapidly eliminated using a high throughput fluorescence activated cell sorting (FACS) approach. After a single sorting round and heat screen at 50 °C, a triple AsbF mutant (Mut1), T61N, H135Y, and H257P, was isolated and characterized. The half-life of Mut1 at 37 °C was >10-fold higher than the wtAsbF (t1/237 = 169 min). Further, the second-order rate constants for both wtAsbF and Mut1 were approximately equal (9.9 × 105 M-1 s-1, 7.8 × 105 M-1 s-1, respectively), thus demonstrating protein thermostability did not come at the expense of enzyme thermophilicity. In addition, in vivo overexpression of Mut1 in E. coli resulted in a ∼60-fold increase in functional enzyme when compared to the wild-type enzyme under the identical expression conditions. Finally, overexpression of the thermostable AsbF resulted in an approximate 80-120% increase in DHB accumulation in the media relative to the wild-type enzyme.

Choline and Geranate Deep Eutectic Solvent as a Broad-Spectrum Antiseptic Agent for Preventive and Therapeutic Applications.

Antiseptic agents are the primary arsenal to disinfect skin and prevent pathogens spreading within the host as well as into the surroundings; however the Food and Drug Administration published a report in 2015 requiring additional validation of nearly all current antiseptic agents before their continued use can be allowed. This vulnerable position calls for urgent identification of novel antiseptic agents. Recently, the ability of a deep eutectic, Choline And Geranate (CAGE), to treat biofilms of Pseudomonas aeruginosa and Salmonella enterica was demonstrated. Here it is reported that CAGE exhibits broad-spectrum antimicrobial activity against a number of drug-resistant bacteria, fungi, and viruses including clinical isolates of Mycobacterium tuberculosis, Staphylococcus aureus, and Candida albicans as well as laboratory strains of Herpes Simplex Virus. Studies in human keratinocytes and mice show that CAGE affords negligible local or systemic toxicity, and an ≈180-14 000-fold improved efficacy/toxicity ratio over currently used antiseptic agents. Further, CAGE penetrates deep into the dermis and treats pathogens located in deep skin layers as confirmed by the ability of CAGE in vivo to treat Propionibacterium acnes infection. In combination, the results clearly demonstrate CAGE holds promise as a transformative platform antiseptic agent for preventive as well as therapeutic applications.

Ionic liquids as a class of materials for transdermal delivery and pathogen neutralization.

Biofilm-protected microbial infections in skin are a serious health risk that remains to be adequately addressed. The lack of progress in developing effective treatment strategies is largely due to the transport barriers posed by the stratum corneum of the skin and the biofilm. In this work, we report on the use of Ionic Liquids (ILs) for biofilm disruption and enhanced antibiotic delivery across skin layers. We outline the syntheses of ILs, analysis of relevant physicochemical properties, and subsequent neutralization effects on two biofilm-forming pathogens: Pseudomonas aeruginosa and Salmonella enterica. Further, the ILs were also examined for cytotoxicity, skin irritation, delivery of antibiotics through the skin, and treatment of biofilms in a wound model. Of the materials examined, choline-geranate emerged as a multipurpose IL with excellent antimicrobial activity, minimal toxicity to epithelial cells as well as skin, and effective permeation enhancement for drug delivery. Specifically, choline-geranate was comparable with, or more effective than, bleach treatment against established biofilms of S. enterica and P. aeruginosa, respectively. In addition, choline-geranate increased delivery of cefadroxil, an antibiotic, by >16-fold into the deep tissue layers of the skin without inducing skin irritation. The in vivo efficacy of choline-geranate was validated using a biofilm-infected wound model (>95% bacterial death after 2-h treatment). This work establishes the use of ILs for simultaneous enhancement of topical drug delivery and antibiotic activity.

Synthetic Routes to Methylerythritol Phosphate Pathway Intermediates and Downstream Isoprenoids.

Isoprenoids constitute the largest class of natural products with greater than 55,000 identified members. They play essential roles in maintaining proper cellular function leading to maintenance of human health, plant defense mechanisms against predators, and are often exploited for their beneficial properties in the pharmaceutical and nutraceutical industries. Most impressively, all known isoprenoids are derived from one of two C5-precursors, isopentenyl diphosphate (IPP) or dimethylallyl diphosphate (DMAPP). In order to study the enzyme transformations leading to the extensive structural diversity found within this class of compounds there must be access to the substrates. Sometimes, intermediates within a biological pathway can be isolated and used directly to study enzyme/pathway function. However, the primary route to most of the isoprenoid intermediates is through chemical catalysis. As such, this review provides the first exhaustive examination of synthetic routes to isoprenoid and isoprenoid precursors with particular emphasis on the syntheses of intermediates found as part of the 2C-methylerythritol 4-phosphate (MEP) pathway. In addition, representative syntheses are presented for the monoterpenes (C10), sesquiterpenes (C15), diterpenes (C20), triterpenes (C30) and tetraterpenes (C40). Finally, in some instances, the synthetic routes to substrate analogs found both within the MEP pathway and downstream isoprenoids are examined.

Sub-inhibitory fosmidomycin exposures elicits oxidative stress in Salmonella enterica serovar Typhimurium LT2.

Fosmidomycin is a time-dependent nanomolar inhibitor of methylerythritol phosphate (MEP) synthase, which is the enzyme that catalyzes the first committed step in the MEP pathway to isoprenoids. Importantly, fosmidomycin is one of only a few MEP pathway-specific inhibitors that exhibits antimicrobial activity. Most inhibitors identified to date only exhibit activity against isolated pathway enzymes. The MEP pathway is the sole route to isoprenoids in many bacteria, yet has no human homologs. The development of inhibitors of this pathway holds promise as novel antimicrobial agents. Similarly, analyses of the bacterial response toward MEP pathway inhibitors provides valuable information toward the understanding of how emergent resistance may ultimately develop to this class of antibiotics. We have examined the transcriptional response of Salmonella enterica serovar typhimurium LT2 to sub-inhibitory concentrations of fosmidomycin via cDNA microarray and RT-PCR. Within the regulated genes identified by microarray were a number of genes encoding enzymes associated with the mediation of reactive oxygen species (ROS). Regulation of a panel of genes implicated in the response of cells to oxidative stress (including genes for catalases, superoxide dismutases, and alkylhydrogen peroxide reductases) was investigated and mild upregulation in some members was observed as a function of fosmidomycin exposure over time. The extent of regulation of these genes was similar to that observed for comparable exposures to kanamycin, but differed significantly from tetracycline. Furthermore, S. typhimurium exposed to sub-inhibitory concentrations of fosmidomycin displayed an increased sensitivity to exogenous H2O2 relative to either untreated controls or kanamycin-treated cells. Our results suggest that endogenous oxidative stress is one consequence of exposures to fosmidomycin, likely through the temporal depletion of intracellular isoprenoids themselves, rather than other mechanisms that have been proposed to facilitate ROS accumulation in bacteria (e.g. cell death processes or the ability of the antibiotic to redox cycle).

Structural characterization of polysaccharides expressed by Burkholderia oklahomensis E0147.

Burkholderia oklahomensis E0147 is a US isolated bacterium believed to express a similar O-antigen to type A structure of the highly pathogenic species, Burkholderia pseudomallei. Both species are genetically closely related. Lipopolysaccharide was collected from E0147 and structurally characterized to test this hypothesis. Glycosyl composition and linkage analyses in conjunction with 1D and 2D (1)H and (13)C NMR spectroscopy showed that the O-antigen was a repeating disaccharide with the following structure: [3)-ß-D-Glcp-(1→3)-2OAc-α-L-6dTalp-(1→]n NMR spectroscopy also revealed the presence of a co-extracted exopolysaccharide previously described in B. pseudomallei, with the structure: [3)-2OAc-ß-D-Galp-(1→4)-α-D-Galp-(1→3)-ß-D-Galp-(1→5)-ß-D-Kdop-(2→]n.

Targeting DXP synthase in human pathogens: enzyme inhibition and antimicrobial activity of butylacetylphosphonate.

The unique methylerythritol phosphate pathway for isoprenoid biosynthesis is essential in most bacterial pathogens. The first enzyme in this pathway, 1-deoxy-D-xylulose 5-phosphate (DXP) synthase, catalyzes a distinct thiamin diphosphate (ThDP)-dependent reaction to form DXP from D-glyceraldehyde 3-phosphate (D-GAP) and pyruvate and represents a potential anti-infective drug target. We have previously demonstrated that the unnatural bisubstrate analog, butylacetylphosphonate (BAP), exhibits selective inhibition of Escherichia coli DXP synthase over mammalian ThDP-dependent enzymes. Here, we report the selective inhibition by BAP against recombinant DXP synthase homologs from Mycobacterium tuberculosis, Yersinia pestis and Salmonella enterica. We also demonstrate antimicrobial activity of BAP against both Gram-negative and Gram-positive strains (including E. coli, S. enterica and Bacillus anthracis), and several clinically isolated pathogens. Our results suggest a mechanism of action involving inhibition of DXP synthase and show that BAP acts synergistically with established antimicrobial agents, highlighting a potential strategy to combat emerging resistance in bacterial pathogens.

Single-pot extraction-analysis of dyed wool fibers with ionic liquids.

Analytical capabilities to identify dyes associated with structurally robust wool fibers would critically assist crime-scene and explosion-scene forensics. Nondestructive separation of dyes from wool, removal of contaminants, and dye analysis by MALDI- or ESI-MS, were achieved in a single-pot, ionic liquid-based method. Ionic liquids (ILs) that readily denature the wool α-keratin structure have been identified and are conducive to small volume, high-throughput analysis for accelerated threat-response times. Wool dyed with commercial or natural, plant-based dyes have unique signatures that allow classification and matching of samples and identification of dyestuffs. Wool released 0.005 mg of dye per mg of dyed wool into the IL, allowing for analysis of single-thread sample sizes. The IL + dye mixture promotes sufficient ionization in MALDI-MS: addition of common MALDI matrices does not improve analysis of anionic wool dyes. An inexpensive, commercially available tetrabutylphosponium chloride IL was discovered to be capable of denaturing wool and was determined to be the most effective for this readily fieldable method.

Tetraalkylphosphonium-based ionic liquids for a single-step dye extraction/MALDI MS analysis platform.

Room temperature ionic liquids, or RTILs, based on tetraalkylphosphonium (PR(4)(+)) cations were used as the basis of a platform that enables separation of dyes from textiles, extraction of dyes from aqueous solution, and identification of the dyes by MALDI-MS in a single experimental step for forensic purposes. Ionic liquids were formed with PR(4)(+) cations and ferulate (FA), α-cyano-4-hydroxycinnamate (CHCA), and 2,5-dihydroxybenzoate (DHB) anions. The use of tetraalkylphosphonium-based ionic liquids in MALDI-MS allowed detection of small molecule dyes without addition of a traditional solid MALDI matrix.

Siderophore-mediated iron acquisition in Bacillus anthracis and related strains.

Recent observations have shed light on some of the endogenous iron-acquisition mechanisms of members of the Bacillus cereus sensu lato group. In particular, pathogens in the B. cereus group use siderophores with both unique chemical structures and biological roles. This review will focus on recent discoveries in siderophore biosynthesis and biology in this group, which contains numerous human pathogens, most notably the causative agent of anthrax, Bacillus anthracis.

The missing link in petrobactin biosynthesis: asbF encodes a (-)-3-dehydroshikimate dehydratase.

The siderophore petrobactin harbors unique 3,4-dihydroxybenzoyl iron-liganding groups. These moieties are known to be synthesized from shikimate pathway precursors, but no reports of the biosynthetic enzymes responsible for this conversion have been published. The gene encoding AsbF from Bacillus thuringiensis 97-27 was overexpressed in an Escherichia coli host. AsbF rapidly and efficiently transforms (-)-3-dehydroshikimate (DHS) into 3,4-dihydroxybenzoate (k(cat)(DHS) = 217 +/- 10 min(-1); K(m)(DHS) = 125 +/- 14 microM) at 37 degrees C and has an absolute requirement for divalent metal. Finally, the pH versus k(cat)(DHS) profile revealed two ionizable groups (pK(a1) = 7.9 +/- 0.1, and pK(a2) = 9.3 +/- 0.1).

Biosynthesis of the 3,4-dihydroxybenzoate moieties of petrobactin by Bacillus anthracis.

The biosynthesis of the 3,4-dihydroxybenzoate moieties of the siderophore petrobactin, produced by B. anthracis str. Sterne, was probed by isotopic feeding experiments in iron-deficient media with a mixture of unlabeled and D-[(13)C6]glucose at a ratio of 5:1 (w/w). After isolation of the labeled siderophore, analysis of the isotopomers was conducted via one-dimensional (1)H and (13)C NMR spectroscopy, as well as (13)C-(13)C DQFCOSY spectroscopy. Isotopic enrichment and (13)C-(13)C coupling constants in the aromatic ring of the isolated siderophore suggested the predominant route for the construction of the carbon backbone of 3,4-DHB (1) involved phosphoenol pyruvate and erythrose-4-phosphate as ultimate precursors. This observation is consistent with that expected if the shikimate pathway is involved in the biosynthesis of these moieties. Enrichment attributable to phosphoenol pyruvate precursors was observed at C1 and C6 of the aromatic ring, as well as into the carboxylate group, while scrambling of the label into C2 was not. This pattern suggests 1 was biosynthesized from early intermediates of the shikimate pathway and not through later shikimate intermediates or aromatic amino acid precursors.

Petrobactin is produced by both pathogenic and non-pathogenic isolates of the Bacillus cereus group of bacteria.

Petrobactin is the primary siderophore synthesized by Bacillus anthracis str Sterne and is required for virulence of this organism in a mouse model. The siderophore's biosynthetic machinery was recently defined and gene homologues of this operon exist in several other Bacillus strains known to be mammalian pathogens, but are absent in several known to be harmless such as B. subtilis and B. lichenformis. Thus, a common hypothesis regarding siderophore production in Bacillus species is that petrobactin production is exclusive to pathogenic isolates. In order to test this hypothesis, siderophores produced by 106 strains of an in-house library of the Bacillus cereus sensu lato group were isolated and identified using a MALDI-TOF-MS assay. Strains were selected from a previously defined phylogenetic tree of this group in order to include both known pathogens and innocuous strains. Petrobactin is produced by pathogenic strains and innocuous isolates alike, and thus is not itself indicative of virulence.

Purification and characterization of rhodobactin: a mixed ligand siderophore from Rhodococcus rhodochrous strain OFS.

The siderophore produced by Rhodococcus rhodochrous strain OFS, rhodobactin, was isolated from iron-deficient cultures and purified by a combination of XAD-7 absorptive/partition resin column and semi-preparative HPLC. The siderophore structure was characterized using 1D and 2D (1)H, (13)C and (15)N NMR techniques (DQFCOSY, TOCSY, NOESY, HSQC and LR-HSQC) and was confirmed using ESI-MS and MS/MS experiments. The structural characterization revealed that the siderophore, rhodobactin, is a mixed ligand hexadentate siderophore with two catecholate and one hydroxamate moieties for iron chelation. We further investigated the effects of Fe concentrations on siderophore production and found that Fe limiting conditions (Fe concentrations from 0.1 microM to 2.0 microM) facilitated siderophore excretion. Our interests lie in the role that siderophores may have in binding metals at mixed contamination sites (containing metals/radionuclides and organics). Given the broad metabolic capacity of this microbe and its Fe scavenging ability, R. rhodochrous OFS may have a competitive advantage over other organisms employed in bioremediation.