Metagenomic profiling of halites from the Atacama Desert: an extreme environment with natural perchlorate does not promote high diversity of perchlorate reducing microorganisms.

We surveyed the presence of perchlorate-reducing microorganisms in available metagenomic data of halite environments from the Atacama Desert, an extreme environment characterized by high perchlorate concentrations, intense ultraviolet radiation, saline and oxidizing soils, and severe desiccation. While the presence of perchlorate might suggest a broad community of perchlorate reducers or a high abundance of a dominant taxa, our search reveals a scarce presence. In fact, we identified only one halophilic species, Salinibacter sp003022435, carrying the pcrA and pcrC genes, represented in low abundance. Moreover, we also discovered some napA genes and organisms carrying the nitrate reductase nasB gene, which hints at the possibility of cryptic perchlorate reduction occurring in these ecosystems. Our findings contribute with the knowledge of perchlorate reduction metabolism potentially occurring in halites from Atacama Desert and point towards promising future research into the perchlorate-reducing mechanism in Salinibacter, a common halophilic bacterium found in hypersaline ecosystems, whose metabolic potential remains largely unknown.

Effects of temperature, chloride and perchlorate salt concentration on the metabolic activity of Deinococcus radiodurans.

The extremophile bacterium Deinococcus radiodurans is characterized by its ability to survive and sustain its activity at high levels of radiation and is considered an organism that might survive in extraterrestrial environments. In the present work, we studied the combined effects of temperature and chlorine-containing salts, with focus on perchlorate salts which have been detected at high concentrations in Martian regolith, on D. radiodurans activity (CO2 production rates) and viability after incubation in liquid cultures for up to 30 days. Reduced CO2 production capacity and viability was observed at high perchlorate concentrations (up to 10% w/v) during incubation at 0 or 25 °C. Both the metabolic activity and viability were reduced as the perchlorate and chloride salt concentration increased and temperature decreased, and an interactive effect of temperature and salt concentration on the metabolic activity was found. These results indicate the ability of D. radiodurans to remain metabolically active and survive in low temperature environments rich in perchlorate.

Biogenic Palladium Improved Perchlorate Reduction during Nitrate Co-Reduction by Diverting Electron Flow in a Hydrogenotrophic Biofilm.

Microbial reduction of perchlorate (ClO4-) is emerging as a cost-effective strategy for groundwater remediation. However, the effectiveness of perchlorate reduction can be suppressed by the common co-contamination of nitrate (NO3-). We propose a means to overcome the limitation of ClO4- reduction: depositing palladium nanoparticles (Pd0NPs) within the matrix of a hydrogenotrophic biofilm. Two H2-based membrane biofilm reactors (MBfRs) were operated in parallel in long-term continuous and batch modes: one system had only a biofilm (bio-MBfR), while the other incorporated biogenic Pd0NPs in the biofilm matrix (bioPd-MBfR). For long-term co-reduction, bioPd-MBfR had a distinct advantage of oxyanion reduction fluxes, and it particularly alleviated the competitive advantage of NO3- reduction over ClO4- reduction. Batch tests also demonstrated that bioPd-MBfR gave more rapid reduction rates for ClO4- and ClO3- compared to those of bio-MBfR. Both biofilm communities were dominated by bacteria known to be perchlorate and nitrate reducers. Functional-gene abundances reflecting the intracellular electron flow from H2 to NADH to the reductases were supplanted by extracellular electron flow with the addition of Pd0NPs.

Insights into the chaotropic tolerance of the desert cyanobacterium Chroococcidiopsis sp. 029 (Chroococcidiopsales, Cyanobacteria).

The mechanism of perchlorate resistance of the desert cyanobacterium Chroococcidiopsis sp. CCMEE 029 was investigated by assessing whether the pathways associated with its desiccation tolerance might play a role against the destabilizing effects of this chaotropic agent. During 3 weeks of growth in the presence of 2.4 mM perchlorate, an upregulation of trehalose and sucrose biosynthetic pathways was detected. This suggested that in response to the water stress triggered by perchlorate salts, these two compatible solutes play a role in the stabilization of macromolecules and membranes as they do in response to dehydration. During the perchlorate exposure, the production of oxidizing species was observed by using an oxidant-sensing fluorochrome and determining the expression of the antioxidant defense genes, namely superoxide dismutases and catalases, while the presence of oxidative DNA damage was highlighted by the over-expression of genes of the base excision repair. The involvement of desiccation-tolerance mechanisms in the perchlorate resistance of this desert cyanobacterium is interesting since, so far, chaotropic-tolerant bacteria have been identified among halophiles. Hence, it is anticipated that desert microorganisms might possess an unrevealed capability of adapting to perchlorate concentrations exceeding those naturally occurring in dry environments. Furthermore, in the endeavor of supporting future human outposts on Mars, the identified mechanisms might contribute to enhance the perchlorate resistance of microorganisms relevant for biologically driven utilization of the perchlorate-rich soil of the red planet.

Bioprospecting of extremophilic perchlorate-reducing bacteria: report of promising Bacillus spp. isolated from sediments of the bay of Cartagena, Colombia.

Three extremophile bacterial strains (BBCOL-009, BBCOL-014 and BBCOL-015), capable of degrading high concentrations of perchlorate at a range of pH (6.5 to 10.0), were isolated from Colombian Caribbean Coast sediments. Morphological features included Gram negative strain bacilli with sizes averaged of 1.75 × 0.95, 2.32 × 0.65 and 3.08 × 0.70 µm, respectively. The reported strains tolerate a wide range of pH (6.5 to 10.0); concentrations of NaCl (3.5 to 7.5% w/v) and KClO4- (250 to 10000 mg/L), reduction of KClO4- from 10 to 25%. LB broth with NaCl (3.5-30% w/v) and KClO4- (250-10000 mg/L) were used in independent trials to evaluate susceptibility to salinity and perchlorate, respectively. Isolates increased their biomass at 7.5 % (w/v) NaCl with optimal development at 3.5 % NaCl. Subsequently, ClO4- reduction was assessed using LB medium with 3.5% NaCl and 10000 mg/L ClO4-. BBCOL-009, BBCOL-014 and BBCOL-015 achieved 10%, 17%, and 25% reduction of ClO4-, respectively. The 16 S rRNA gene sequence grouped them as Bacillus flexus T6186-2, Bacillus marisflavi TF-11 (T), and Bacillus vietnamensis 15 - 1 (T) respectively, with < 97.5% homology. In addition, antimicrobial resistance to ertapenem, vancomycine, amoxicillin clavulanate, penicillin, and erythromycin was present in all the isolates, indicating their high adaptability to stressful environments. The isolated strains from marine sediments in Cartagena Bay, Colombia are suitable candidates to reduce perchlorate contamination in different environments. Although the primary focus of the study of perchlorate-reducing and resistant bacteria is in the ecological and agricultural realms, from an astrobiological perspective, perchlorate-resistant bacteria serve as models for astrobiological investigations.

Coremoval of Energetics and Oxyanions via the In Situ Coupling of Catalytic and Enzymatic Destructions: A Solution to Ammunition Wastewater Treatment.

Ammunition wastewater contains toxic nitrated explosives like RDX and oxyanions like nitrate and perchlorate. Its treatment is challenged by low efficiency due to contaminant recalcitrance and high cost due to multiple processes needed for separately removing different contaminant types. This paper reports a H2-based low-energy strategy featuring the treatment of explosives via catalytic denitration followed by microbial mineralization coupled with oxyanion reduction. After a nitrate- and perchlorate-reducing biofilm incapable of RDX biodegradation was coated with palladium nanoparticles (Pd0NPs), RDX was rapidly denitrated with a specific catalytic activity of 8.7 gcat-1 min-1, while biological reductions of nitrate and perchlorate remained efficient. In the subsequent 30-day continuous test, >99% of RDX, nitrate, and perchlorate were coremoved, and their effluent concentrations were below their respective regulation levels. Detected intermediates and shallow metagenome analysis suggest that the intermediates after Pd-catalytic denitration of RDX ultimately were enzymatically utilized by the nitrate- and perchlorate-reducing bacteria as additional electron donor sources.

Impacts of co-contaminants and dilution on perchlorate biodegradation using various carbon sources.

This research investigates the biodegradation of perchlorate in the presence of the co-contaminants nitrate and chlorate using soluble and slow-release carbon sources. In addition, the impact of bio-augmentation and dilution, which results in lower total dissolved salts (TDS) and contaminant levels, is examined. Laboratory microcosms were conducted using actual groundwater and soils from a contaminated aquifer. The results revealed that both soluble and slow-release carbon sources support biodegradation of contaminants in the sequence nitrate > chlorate > perchlorate. Degradation rates, including and excluding lag times, revealed that the overall impact of the presence of co-contaminants depends on degradation kinetics and the relative concentrations of the contaminants. When the lag time caused by the presence of the co-contaminants is considered, the degradation rates for chlorate and perchlorate were two to three times slower. The results also show that dilution causes lower initial contaminant concentrations, and consequently, slower degradation rates, which is not desirable. On the other hand, the dilution resulting from the injection of amendments to support remediation promotes desirably lower salinity levels. However, the salinity associated with the presence of sulfate does not inhibit biodegradation. The naturally occurring bacteria were able to support the degradation of all contaminants. Bio-augmentation was effective only in diluted microcosms. Proteobacteria and Firmicutes were the dominant phyla identified in the microcosms.

Perchlorate-specific proteomic stress responses of Debaryomyces hansenii could enable microbial survival in Martian brines.

If life exists on Mars, it would face several challenges including the presence of perchlorates, which destabilize biomacromolecules by inducing chaotropic stress. However, little is known about perchlorate toxicity for microorganisms on the cellular level. Here, we present the first proteomic investigation on the perchlorate-specific stress responses of the halotolerant yeast Debaryomyces hansenii and compare these to generally known salt stress adaptations. We found that the responses to NaCl and NaClO4 -induced stresses share many common metabolic features, for example, signalling pathways, elevated energy metabolism, or osmolyte biosynthesis. Nevertheless, several new perchlorate-specific stress responses could be identified, such as protein glycosylation and cell wall remodulations, presumably in order to stabilize protein structures and the cell envelope. These stress responses would also be relevant for putative life on Mars, which-given the environmental conditions-likely developed chaotropic defence strategies such as stabilized confirmations of biomacromolecules or the formation of cell clusters.

11C-Labeled Radiotracer for Noninvasive and Quantitative Assessment of the Thiocyanate Efflux System in the Brain.

Thiocyanate (SCN-) alters the potency of certain agonists for the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, and dysfunctions in AMPA receptor signaling are considered to underlie a number of neurological diseases. While humans may be exposed to SCN- from the environment, including food sources, a carrier-mediated system transports SCN- from the brain into the blood and is an important regulator of SCN- distribution in the central nervous system. The assessment of this SCN- efflux system in the brain would thus be useful for understanding the mechanisms underlying the neurotoxicity of SCN- and for elucidating the relationship between the efflux system and brain diseases. However, the currently available technique for studying SCN- efflux is severely limited by its invasiveness. Here, we describe the development of a SCN- protracer, 9-pentyl-6-[11C]thiocyanatopurine ([11C]1), to overcome this limitation. [11C]1 was synthesized by the reaction of the iodo-precursor and [11C]SCN- or the reaction of the disulfide precursor with [11C]NH4CN. The protracer [11C]1 entered the brain after intravenous injection into mice and was rapidly metabolized to [11C]SCN-, which was then eliminated from the brain. The efflux of [11C]SCN- was dose-dependently inhibited by perchlorate, a monovalent anion, and the highest dose caused an 82% reduction in the efflux rate. Our findings demonstrate that [11C]1 can be used for the noninvasive and quantitative assessment of the SCN- efflux system in the brain.

Impact of Acetate in Reduction of Perchlorate by Mixed Microbial Culture under the Influence of Nitrate and Sulfate.

The biological reduction of slow degradation contaminants such as perchlorate (ClO4-) is considered to be a promising water treatment technology. The process is based on the ability of a specific mixed microbial culture to use perchlorate as an electron acceptor in the absence of oxygen. In this study, batch experiments were conducted to investigate the effect of nitrate on perchlorate reduction, the kinetic parameters of the Monod equation and the optimal ratio of acetate to perchlorate for the perchlorate reducing bacterial consortium. The results of this study suggest that acclimated microbial cultures can be applied to treat wastewater containing high concentrations of perchlorate. Reactor experiments were carried out with different hydraulic retention times (HRTs) to determine the optimal operating conditions. A fixed optimal HRT and the effect of nitrate on perchlorate reduction were investigated with various concentrations of the electron donor. The results showed that perchlorate reduction occurred after nitrate removal. Moreover, the presence of sulfate in wastewater had no effect on the perchlorate reduction. However, it had little effect on biomass concentration in the presence of nitrate during exposure to a mixed microbial culture, considering the nitrate as the inhibitor of perchlorate reduction by reducing the degradation rate. The batch scale experiment results illustrated that for efficient operation of perchlorate reduction, the optimal acetate to perchlorate ratio of 1.4:1.0 would be enough. Moreover, these experiments found the following results: the kinetic parameters equivalent to Y = 0.281 mg biomass/mg perchlorate, Ks = 37.619 mg/L and qmax = 0.042 mg perchlorate/mg biomass/h. In addition, anoxic-aerobic experimental reactor results verify the optimal HRT of 6 h for continuous application. Furthermore, it also illustrated that using 600 mg/L of acetate as a carbon source is responsible for 100% of nitrate reduction with less than 50% of the perchlorate reduction, whereas at 1000 mg/L acetate, approximately 100% reduction was recorded.

(Per)chlorate in Biology on Earth and Beyond.

Respiration of perchlorate and chlorate [collectively, (per)chlorate] was only recognized in the last 20 years, yet substantial advances have been made in our understanding of the underlying metabolisms. Although it was once considered solely anthropogenic, pervasive natural sources, both terrestrial and extraterrestrial, indicate an ancient (per)chlorate presence across our solar system. These discoveries stimulated interest in (per)chlorate microbiology, and the application of advanced approaches highlights exciting new facets. Forward and reverse genetics revealed new information regarding underlying molecular biology and associated regulatory mechanisms. Structural and functional analysis characterized core enzymes and identified novel reaction sequences. Comparative genomics elucidated evolutionary aspects, and stress analysis identified novel response mechanisms to reactive chlorine species. Finally, systems biology identified unique metabolic versatility and novel mechanisms of (per)chlorate respiration, including symbiosis and a hybrid enzymatic-abiotic metabolism. While many published studies focus on (per)chlorate and their basic metabolism, this review highlights seminal advances made over the last decade and identifies new directions and potential novel applications.

High-strength wastewater treatment using microbial biofilm reactor: a critical review.

Biofilm reactors retain microbial cells in the form of biofilm which is attached to free moving or fixed carrying materials, thus providing a high active biomass concentration and automatic liquid and solid separation. Nowadays, microbial biofilm reactors have been widely used in high-strength wastewater treatment where very high pollutant removal efficiency is required, which usually requires excessive space and aeration energy for conventional activated sludge-based treatment. This paper provides an overview of microbial biofilm reactors developed over the last half-century, including moving bed biofilm reactor (MBBR), trickling filter (TF) reactor, rotating biological contactor (RBC), membrane biofilm reactor (MBfR), passive aeration simultaneous nitrification and denitrification (PASND) biofilm reactor, for their applications in high-strength wastewater treatment of not only removing carbon, nitrogen, sulphur but also a variety of oxidized contaminants including perchlorate and bromate. Despite the advance of biofilm reactor that exhibits high resistance to excessive pollutants loading, its drawbacks both from engineering and microbiological point of view are reviewed. The future prospects of biofilm reactor are also discussed in this review paper.

Identification of a perchloric acid-soluble protein (PSP)-like ribonuclease from Trichomonas vaginalis.

A perchloric acid-soluble protein (PSP), named here tv-psp1, was identified in Trichomonas vaginalis. It is expressed under normal culture conditions according to expressed sequence tag (EST) analysis. On the other hand, Tv-PSP1 protein was identified by mass spectrometry with a 40% of identity to human PSP (p14.1). Polyclonal antibodies against recombinant Tv-PSP1 (rTv-PSP1) recognized a single band at 13.5 kDa in total protein parasite extract by SDS-PAGE and a high molecular weight band analyzed by native PAGE. Structural analysis of Tv-PSP1, using dynamic light scattering, size exclusion chromatography, and circular dichroism spectroscopy, showed a trimeric structure stable at 7 M urea with 38% α-helix and 14% ß-sheet in solution and a molecular weight of 40.5 kD. Tv-PSP1 models were used to perform dynamic simulations over 100 ns suggesting a stable homotrimeric structure. Tv-PSP1 was located in the nucleus, cytoplasm, and hydrogenosomes of T. vaginalis, and the in silico analysis by Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) showed interactions with RNA binding proteins. The preliminary results of RNA degradation analysis with the recombinant Tv-PSP1 showed RNA partial deterioration suggesting a possible putative ribonuclease function.

Perchlorate Reductase Is Distinguished by Active Site Aromatic Gate Residues.

Perchlorate is an important ion on both Earth and Mars. Perchlorate reductase (PcrAB), a specialized member of the dimethylsulfoxide reductase superfamily, catalyzes the first step of microbial perchlorate respiration, but little is known about the biochemistry, specificity, structure, and mechanism of PcrAB. Here we characterize the biophysics and phylogeny of this enzyme and report the 1.86-Å resolution PcrAB complex crystal structure. Biochemical analysis revealed a relatively high perchlorate affinity (Km = 6 µm) and a characteristic substrate inhibition compared with the highly similar respiratory nitrate reductase NarGHI, which has a relatively much lower affinity for perchlorate (Km = 1.1 mm) and no substrate inhibition. Structural analysis of oxidized and reduced PcrAB with and without the substrate analog SeO3 (2-) bound to the active site identified key residues in the positively charged and funnel-shaped substrate access tunnel that gated substrate entrance and product release while trapping transiently produced chlorate. The structures suggest gating was associated with shifts of a Phe residue between open and closed conformations plus an Asp residue carboxylate shift between monodentate and bidentate coordination to the active site molybdenum atom. Taken together, structural and mutational analyses of gate residues suggest key roles of these gate residues for substrate entrance and product release. Our combined results provide the first detailed structural insight into the mechanism of biological perchlorate reduction, a critical component of the chlorine redox cycle on Earth.

Anion inhibitors of the ß-carbonic anhydrase from the pathogenic bacterium responsible of tularemia, Francisella tularensis.

A ß-class carbonic anhydrase (CA, EC 4.2.1.1) from the pathogenic bacterium Francisella tularensis (FtußCA) was cloned and purified, and the anion inhibition profile was investigated. Based on the measured kinetic parameters for the enzyme catalyzed CO2 hydration reaction (kcat of 9.8×105s-1 and a kcat/KM of 8.9×107M-1s-1), FtußCA is a highly effective enzyme. The activity of FtußCA was not inhibited by a range of anions that do not typically coordinate Zn(II) effectively, including perchlorate, tetrafluoroborate, and hexafluorophosphate. Surprisingly, some anions which generally complex well with many cations, including Zn(II), also did not effectively inhibit FtußCA, e.g., fluoride, cyanide, azide, nitrite, bisulphite, sulfate, tellurate, perrhenate, perrhuthenate, and peroxydisulfate. However, the most effective inhibitors were in the range of 90-94µM (sulfamide, sulfamic acid, phenylarsonic and phenylboronic acid). N,N-Diethyldithiocarbamate (KI of 0.31mM) was a moderately potent inhibitor. As Francisella tularensis is the causative agent of tularemia, the discovery of compounds that can interfere with the life cycle of this pathogen may result in novel opportunities to fight antibiotic drug resistance.

A Multiple Reaction Modelling Framework for Microbial Electrochemical Technologies.

A mathematical model for the theoretical evaluation of microbial electrochemical technologies (METs) is presented that incorporates a detailed physico-chemical framework, includes multiple reactions (both at the electrodes and in the bulk phase) and involves a variety of microbial functional groups. The model is applied to two theoretical case studies: (i) A microbial electrolysis cell (MEC) for continuous anodic volatile fatty acids (VFA) oxidation and cathodic VFA reduction to alcohols, for which the theoretical system response to changes in applied voltage and VFA feed ratio (anode-to-cathode) as well as membrane type are investigated. This case involves multiple parallel electrode reactions in both anode and cathode compartments; (ii) A microbial fuel cell (MFC) for cathodic perchlorate reduction, in which the theoretical impact of feed flow rates and concentrations on the overall system performance are investigated. This case involves multiple electrode reactions in series in the cathode compartment. The model structure captures interactions between important system variables based on first principles and provides a platform for the dynamic description of METs involving electrode reactions both in parallel and in series and in both MFC and MEC configurations. Such a theoretical modelling approach, largely based on first principles, appears promising in the development and testing of MET control and optimization strategies.

Genetic dissection of chlorate respiration in Pseudomonas stutzeri†PDA reveals syntrophic (per)chlorate reduction.

Genes important for growth of Pseudomonas stutzeri PDA on chlorate were identified using a randomly DNA bar-coded transposon mutant library. During chlorate reduction, mutations in genes encoding the chlorate reductase clrABC, predicted molybdopterin cofactor chaperon clrD, molybdopterin biosynthesis and two genes of unknown function (clrE, clrF) had fitness defects in pooled mutant assays (Bar-seq). Markerless in-frame deletions confirmed that clrA, clrB and clrC were essential for chlorate reduction, while clrD, clrE and clrF had less severe growth defects. Interestingly, the key detoxification gene cld was essential for chlorate reduction in isogenic pure culture experiments, but showed only minor fitness defects in Bar-seq experiments. We hypothesized this was enabled through chlorite dismutation by the community, as most strains in the Bar-seq library contained an intact cld. In support of this, Δcld grew with wild-type PDA or ΔclrA, and purified Cld also restored growth to the Δcld mutant. Expanding on this, wild-type PDA and a Δcld mutant of the perchlorate reducer Azospira suillum PS grew on perchlorate in co-culture, but not individually. These results demonstrate that co-occurrence of cld and a chloroxyanion reductase within a single organism is not necessary and raises the possibility of syntrophic (per)chlorate respiration in the environment.

Concurrent microbial reduction of high concentrations of nitrate and perchlorate in an ion exchange membrane bioreactor.

We investigated effective simultaneous removal of high loads of nitrate and perchlorate from synthetic groundwater using an ion exchange membrane bioreactor (IEMB). The aim of this research was to characterize both transport aspects and biodegradation mechanisms involved in the treatment process of high loads of the two anions. Biodegradation process was proven to be efficient with over 99% efficiency of both perchlorate and nitrate, regardless of their load. The maximum biodegradation rates were 18.3 (mmol m(-2) h(-1) ) and 5.5 (mmol m(-2) h(-1) ) for nitrate and perchlorate, respectively. The presence of a biofilm on the bio-side of the membrane only slightly increased the nitrate and perchlorate transmembrane flux as compared to the measured flux during a Donnan dialysis experiment where there is no biodegradation of perchlorate and nitrate in the bio-compartment. The nitrate flux in presence of a biofilm was 18.3 (±1.9) (mmole m(-2) h(-1) ), while without the biofilm, the flux was 16.9 (±1.5) (mmole m(-2) h(-1) ) for the same feed inlet nitrate concentration of 4 mM. The perchlorate transmembrane flux increased similarly by an average of 5%. Samples of membrane biofilm and suspended bacteria from the bio-reactor were analyzed for diversity and abundance of the perchlorate and nitrate reducing bacteria. Klebsiella oxytoca, known as a glycerol fermenter, accounted for 70% of the suspended bacteria. In contrast, perchlorate and nitrate reducing bacteria predominated in the biofilm present on the membrane. These results are consistent with our proposed two stage biodegradation mechanism where glycerol is first fermented in the suspended phase of the bio-reactor and the fermentation products drive perchlorate and nitrate bio-reduction in the biofilm attached to the membrane. These results suggest that the niche exclusion of microbial populations in between the reactor and membrane is controlled by the fluxes of the electron donors and acceptors. Such a mechanism has important implications for controlling the bio-reduction reaction in the IEMB when using glycerol as a carbon source and allowing treating a complex contamination of high concentrations of perchlorate and nitrating in groundwater and successfully biodegrading them to non-hazardous components. Biotechnol. Bioeng. 2016;113: 1881-1891. © 2016 Wiley Periodicals, Inc.

A novel perchlorate- and nitrate-reducing bacterium, Azospira sp. PMJ.

A novel perchlorate-reducing bacterium (PCRB), PMJ, was isolated from the mixed liquor suspended solids in the aerobic tank of a wastewater treatment plant. The 16S ribosomal RNA (rRNA), perchlorate reductase, and chlorite dismutase gene sequences revealed that PMJ belonged to the genus Azospira. PMJ was removed high-strength (700 mg/L) perchlorate and also removed low-strength (≤50 mg/L) perchlorate below the detection limit (2 µg/L) when acetate was used as a sole and carbon source. The maximum specific perchlorate utilization rate, q max, was 0.96 mg ClO4 (-)/mg dry cell weight day, and the half-saturation constant, K S , was lower than 0.002 mg ClO4 (-)/L. PMJ also utilized inorganic electron donors [(H2, S(0), and Fe(II)] with perchlorate as an electron acceptor. Perchlorate reduction by PMJ was completely inhibited by oxygen and chlorate but was not inhibited by nitrate. In the presence of similar concentrations (100∼140 mg/L) of nitrate and perchlorate, PMJ simultaneously removed both electron acceptors. Therefore, it was concluded that the strains PMJ might possess separate pathways for perchlorate and nitrate reduction. These results indicated that Azospira sp. PMJ could be efficiently used for treating perchlorate-contaminated groundwater and wastewater because many of these water bodies are known to contain both perchlorate and nitrate. In addition, low K S value and autotrophic perchlorate reduction of PMJ might be useful to design the biological treatment systems.

Characterization of an anaerobic marine microbial community exposed to combined fluxes of perchlorate and salinity.

The recent recognition of the environmental prevalence of perchlorate and its discovery on Mars, Earth's moon, and in meteorites, in addition to its novel application to controlling oil reservoir sulfidogenesis, has resulted in a renewed interest in this exotic ion and its associated microbiology. However, while plentiful data exists on freshwater perchlorate respiring organisms, information on their halophilic counterparts and microbial communities is scarce. Here, we investigated the temporal evolving structure of perchlorate respiring communities under a range of NaCl concentrations (1, 3, 5, 7, and 10 % wt/vol) using marine sediment amended with acetate and perchlorate. In general, perchlorate consumption rates were inversely proportional to NaCl concentration with the most rapid rate observed at 1 % NaCl. At 10 % NaCl, no perchlorate removal was observed. Transcriptional analysis of the 16S rRNA gene indicated that salinity impacted microbial community structure and the most active members were in families Rhodocyclaceae (1 and 3 % NaCl), Pseudomonadaceae (1 NaCl), Campylobacteraceae (1, 5, and 7 % NaCl), Sedimenticolaceae (3 % NaCl), Desulfuromonadaceae (5 and 7 % NaCl), Pelobacteraceae (5 % NaCl), Helicobacteraceae (5 and 7 % NaCl), and V1B07b93 (7 %). Novel isolates of genera Sedimenticola, Marinobacter, Denitromonas, Azoarcus, and Pseudomonas were obtained and their perchlorate respiring capacity confirmed. Although the obligate anaerobic, sulfur-reducing Desulfuromonadaceae species were dominant at 5 and 7 % NaCl, their enrichment may result from biological sulfur cycling, ensuing from the innate ability of DPRB to oxidize sulfide. Additionally, our results demonstrated enrichment of an archaeon of phylum Parvarchaeota at 5 % NaCl. To date, this phylum has only been described in metagenomic experiments of acid mine drainage and is unexpected in a marine community. These studies identify the intrinsic capacity of marine systems to respire perchlorate and significantly expand the known diversity of organisms capable of this novel metabolism.