Interaction of tetracycline and copper co-intake in inducing antibiotic resistance genes and potential pathogens in mouse gut.

The widespread use of copper and tetracycline as growth promoters in the breeding industry poses a potential threat to environmental health. Nevertheless, to the best of our knowledge, the potential adverse effects of copper and tetracycline on the gut microbiota remain unknown. Herein, mice were fed different concentrations of copper and/or tetracycline for 6 weeks to simulate real life-like exposure in the breeding industry. Following the exposure, antibiotic resistance genes (ARGs), potential pathogens, and other pathogenic factors were analyzed in mouse feces. The co-exposure of copper with tetracycline significantly increased the abundance of ARGs and enriched more potential pathogens in the gut of the co-treated mice. Copper and/or tetracycline exposure increased the abundance of bacteria carrying either ARGs, metal resistance genes, or virulence factors, contributing to the widespread dissemination of potentially harmful genes posing a severe risk to public health. Our study provides insights into the effects of copper and tetracycline exposure on the gut resistome and potential pathogens, and our findings can help reduce the risks associated with antibiotic resistance under the One Health framework.

Migration of phosphorus in pig manure during pyrolysis process and slow-release mechanism of biochar in hydroponic application.

Pyrolysis is an effective method for treating of livestock and poultry manure developed in recent years. It can completely decompose pathogens and antibiotics, stabilize heavy metals, and enrich phosphorus (P) in biochar. To elucidate the P migration mechanism under different pig manure pyrolysis temperatures, sequential fractionation, solution 31P nuclear magnetic resonance, X-ray photoelectron spectroscopy, X-ray diffraction, and K-edge X-ray absorption near-edge structure techniques were used to analyze the P species in pig manure biochar (PMB). The results indicated that most of the organic P in the pig manure was converted to inorganic P during pyrolysis. Moreover, the transformation to different P groups pathways was clarified. The phase transition from amorphous to crystalline calcium phosphate was promoted when the temperature was above 600 °C. The content of P extracted by hydrochloric acid, which was the long-term available P for plant uptake, increased significantly. PMB pyrolyzed at 600 °C can be used as a highly effective substitute for P source. It provides the necessary P species (e.g. water-soluble P.) and metal elements for the growth of water spinach plants, and which are slow-release comparing with the Hogland nutrient solution.

Chronic Exposure to Drinking Water As, Pb, and Cd at Provisional Guideline Values Reduces Weight Gain in Male Mice via Gut Microflora Alterations and Intestinal Inflammation.

Few studies have investigated the long-term effect of exposure to arsenic (As), lead (Pb), and cadmium (Cd) via drinking water at the provisional guideline values on gut microflora. In this study, male and female mice were exposed to water As, Pb, or Cd at 10, 10, or 5 µg L-1 for 6 months. At the end of the exposure, the net weight gain of male mice exposed to As and Pb (9.91 ± 1.35 and 11.2 ± 1.50 g) was significantly (p < 0.05) lower compared to unexposed control mice (14.1 ± 3.24 g), while this was not observed for female mice. Relative abundance of Akkermansia, a protective gut bacterium against intestinal inflammation, was reduced from 29.7% to 3.20%, 4.83%, and 17.0% after As, Pb, and Cd exposure in male mice, which likely caused chronic intestinal inflammation, as suggested by 2.81- to 9.60-fold higher mRNA levels of pro-inflammatory factors in ileal enterocytes of male mice. These results indicate that long-term exposure to drinking water As, Pb, and Cd at concentrations equivalent to the China provisional guideline values can cause loss of protective bacteria and lead to chronic intestinal inflammation, thereby affecting body weight gain in male mice.

Identification and characterization of a phosphinothricin N-acetyltransferase from Enterobacter LSJC7.

Phosphinothricin (PPT) is a widely used and non-selective herbicide. PPT-resistance genes, especially PPT N-acetyltransferase genes, have been used in the development of transgenic PPT-resistant crops. However, there are only a limited number of available PPT-resistance genes for use in plant biotechnology. In this study, we found that Enterobacter LSJC7 is highly resistant to PPT and can acetylate PPT to N-acetyl phosphinothricin (Ac-PPT). Furthermore, a novel PPT N-acetyltransferase gene, named LsarsN, was identified from LSJC7. When LsarsN was expressed in E. coli AW3110, it confered resistance to PPT. Ac-PPT was detected in both the culture medium and cells of AW3110 expressing the LsarsN-pET22b plasmid. The purified LsArsN protein also showed strong N-acetylation ability in vitro, and its enzymatic kinetic curve was fitted with the Michaelis-Mentan equation. Compared with wild-type LsArsN, both R72A and R74A mutants showed significantly lower PPT N-acetylation ability. In summary, our results systematically characterized LsArsN with strong ability for PPT N-acetylation, which lays the groundwork for future research into the use of this novel gene, LsarsN, to create PPT-resistant crops.

Gut Microbiota Control the Bioavailability and Metabolism of Organoarsenicals of Seaweeds in Mice after Oral Ingestion.

Edible seaweed consumption is an essential route of human exposure to complex organoarsenicals, including arsenosugars and arsenosugar phospholipids. However, the effects of gut microbiota on the metabolism and bioavailability of arsenosugars in vivo are unknown. Herein, two nori and two kelp samples with phosphate arsenosugar and sulfonate arsenosugar, respectively, as the predominant arsenic species, were administered to normal mice and gut microbiota-disrupted mice treated with the broad-spectrum antibiotic cefoperazone for 4 weeks. Following exposure, the community structures of the gut microbiota, total arsenic concentrations, and arsenic species in excreta and tissues were analyzed. Total arsenic excreted in feces and urine did not differ significantly between normal and antibiotic-treated mice fed with kelp samples. However, the total urinary arsenic of normal mice fed with nori samples was significantly higher (p < 0.05) (urinary arsenic excretion factor, 34-38 vs 5-7%), and the fecal total arsenic was significantly lower than in antibiotic-treated mice. Arsenic speciation analysis revealed that most phosphate arsenosugars in nori were converted to arsenobetaine (53.5-74.5%) when passing through the gastrointestinal tract, whereas a large portion of sulfonate arsenosugar in kelp was resistant to speciation changes and was excreted in feces intact (64.1-64.5%). Normal mice exhibited greater oral bioavailability of phosphate arsenosugar from nori than sulfonate arsenosugar from kelp (34-38 vs 6-9%). Our work provides insights into organoarsenical metabolism and their bioavailability in the mammalian gut.

Ca Minerals and Oral Bioavailability of Pb, Cd, and As from Indoor Dust in Mice: Mechanisms and Health Implications.

BACKGROUND: Elevating dietary calcium (Ca) intake can reduce metal(loid)oral bioavailability. However, the ability of a range of Ca minerals to reduce oral bioavailability of lead (Pb), cadmium (Cd), and arsenic (As) from indoor dust remains unclear. OBJECTIVES: This study evaluated the ability of Ca minerals to reduce Pb, Cd, and As oral bioavailability from indoor dust and associated mechanisms. METHODS: A mouse bioassay was conducted to assess Pb, Cd, and As relative bioavailability (RBA) in three indoor dust samples, which were amended into mouse chow without and with addition of CaHPO4, CaCO3, Ca gluconate, Ca lactate, Ca aspartate, and Ca citrate at 200-5,000µg/g Ca. The mRNA expression of Ca and phosphate (P) transporters involved in transcellular Pb, Cd and As transport in the duodenum of mice was quantified using real-time polymerase chain reaction. Serum 1,25-Dihydroxyvitamin D3 [1,25(OH)2D3], parathyroid hormone (PTH), and renal CYP27B1 activity controlling 1,25(OH)2D3 synthesis were measured using ELISA kits. Metal(loid) speciation in the feces of mice was characterized using X-ray absorption near-edge structure (XANES) spectroscopy. RESULTS: In general, mice exposed to each of the Ca minerals exhibited lower Pb-, Cd-, and As-RBA for three dusts. However, RBAs with the different Ca minerals varied. Among minerals, mice fed dietary CaHPO4 did not exhibit lower duodenal mRNA expression of Ca transporters but did have the lowest Pb and Cd oral bioavailability at the highest Ca concentration (5,000µg/g Ca; 51%-95% and 52%-74% lower in comparison with the control). Lead phosphate precipitates (e.g., chloropyromorphite) were observed in feces of mice fed dietary CaHPO4. In comparison, mice fed organic Ca minerals (Ca gluconate, Ca lactate, Ca aspartate, and Ca citrate) had lower duodenal mRNA expression of Ca transporters, but Pb and Cd oral bioavailability was higher than in mice fed CaHPO4. In terms of As, mice fed Ca aspartate exhibited the lowest As oral bioavailability at the highest Ca concentration (5,000µg/g Ca; 41%-72% lower) and the lowest duodenal expression of P transporter (88% lower). The presence of aspartate was not associated with higher As solubility in the intestine. DISCUSSION: Our study used a mouse model of exposure to household dust with various concentrations and species of Ca to determine whether different Ca minerals can reduce bioavailability of Pb, Cd, and As in mice and elucidate the mechanism(s) involved. This study can contribute to the practical application of optimal Ca minerals to protect humans from Pb, Cd, and As coexposure in the environment. https://doi.org/10.1289/EHP11730.

Arsenic bioaccumulation in the soil fauna alters its gut microbiome and microbial arsenic biotransformation capacity.

The biotransformation of arsenic mediated by microorganisms plays an important role in the arsenic biogeochemical cycle. However, the fate and biotransformation of arsenic in different soil fauna gut microbiota are largely unknown. Herein the effects of arsenic contamination on five types of soil fauna were compared by examining variations in arsenic bioaccumulation, gut microbiota, and arsenic biotransformation genes (ABGs). Significant difference was observed in the arsenic bioaccumulation across several fauna body tissues, and Metaphire californica had the highest arsenic bioaccumulation, with a value of 107 ± 1.41 mg kg-1. Arsenic exposure significantly altered overall patterns of ABGs; however, dominant genes involved in arsenic redox and other genes involved in arsenic methylation and demethylation were not significantly changed across animals. Except for M. californica, the abundance of ABGs in other animal guts firstly increased and then decreased with increasing arsenic concentrations. In addition, exposure of soil fauna to arsenic led to shifts in the unique gut-associated bacterial community, but the magnitude of these changes varied significantly across ecological groups of soil fauna. A good correlation between the gut bacterial communities and ABG profiles was observed, suggesting that gut microbiota plays important roles in the biotransformation of arsenic. Overall, these results provide a universal profiling of a microbial community capable of arsenic biotransformation in different fauna guts. Considering the global distribution of soil fauna in the terrestrial ecosystem, this finding broadens our understanding of the hidden role of soil fauna in the arsenic bioaccumulation and biogeochemical cycle.

Antibiotic exposure decreases soil arsenic oral bioavailability in mice by disrupting ileal microbiota and metabolic profile.

Oral bioavailability of arsenic (As) determines levels of As exposure via ingestion of As-contaminated soil, however, the role of gut microbiota in As bioavailability has not evaluated in vivo although some in vitro studies have investigated this. Here, we made a comparison in As relative bioavailability (RBA) estimates for a contaminated soil (3913 mg As kg-1) using a mouse model with and without penicillin perturbing gut microbiota and metabolites. Compared to soil exposure alone (2% w/w soil in diets), addition of penicillin (100 or 1000 mg kg-1) reduced probiotic Lactobacillus and sulfate-reducing bacteria Desulfovibrio, enriched penicillin-resistant Enterobacter and Bacteroides, and decreased amino acid concentrations in ileum. With perturbed gut microbiota and metabolic profile, penicillin and soil co-exposed mice accumulated 2.81-3.81-fold less As in kidneys, excreted 1.02-1.35-fold less As in urine, and showed lower As-RBA (25.7-29.0%) compared to mice receiving diets amended with soil alone (56 ± 9.63%). One mechanism accounted for this is the decreased concentrations of amino acids arising from the gut microbiota shift which resulted in elevated iron (Fe) and As co-precipitation, leading to reduced As solubilization in the intestine. Another mechanism was conversion of bioavailable inorganic As to less bioavailable monomethylarsonic acid (MMAV) and dimethylarsinic acid (DMAV) by the antibiotic perturbed microflora. Based on in vivo mouse model, we demonstrated the important role of gut microbiota and gut metabolites in participating soil As solubilization and speciation transformation then affecting As oral bioavailability. Results are useful to better understand the role of gut bacteria in affecting As metabolism and the health risks of As-contaminated soils.

The response of Pyropia haitanensis to inorganic arsenic under laboratory culture.

Up to now, complicated organoarsenicals were mainly identified in marine organisms, suggesting that these organisms play a critical role in arsenic biogeochemical cycling because of low phosphate and relatively high arsenic concentration in the marine environment. However, the response of marine macroalgae to inorganic arsenic remains unknown. In this study, Pyropia haitanensis were exposed to arsenate [As(V)] (0.1, 1, 10, 100 µM) or arsenite [As(III)] (0.1, 1, 10 µM) under laboratory conditions for 3 d. The species of water-soluble arsenic, the total concentration of lipid-soluble and cell residue arsenic of the algae cells was analyzed. As(V) was mainly transformed into oxo-arsenosugar-phosphate, with other arsenic compounds such as monomethylated, As(III), demethylated arsenic and oxo-arsenosugar-glycerol being likely the intermediates of arsenosugar synthesis. When high concentration of As(III) was toxic to P. haitanensis, As(III) entered into the cells and was transformed into less toxic organoarsenicals and As(V). Transcriptome results showed genes involved in DNA replication, mismatch repair, base excision repair, and nucleotide excision repair were up-regulated in the algae cells exposed to 10 µM As(V), and multiple genes involved in glutathione metabolism and photosynthetic were up-regulated by 1 µM As(III). A large number of ABC transporters were down-regulated by As(V) while ten genes related to ABC transporters were up-regulated by As(III), indicating that ABC transporters were involved in transporting As(III) to vacuoles in algae cells. These results indicated that P. haitanensis detoxifies inorganic arsenic via transforming them into organoarsenicals and enhancing the isolation of highly toxic As(III) in vacuoles.

Arsenic transformation mediated by gut microbiota affects the fecundity of Caenorhabditis elegans.

Arsenic biotransformation has been discovered in guts of soil invertebrates. Reproduction of invertebrates is sensitive to arsenic contamination in soils. However, little is known about the impact of gut microbe-mediated arsenic biotransformation on the fecundity of invertebrates. Here, Caenorhabditis elegans was firstly pre-fed with Escherichia coli BL21 possessing the capability of reducing arsenate [As(V)] or BL21M having the ability to reduce As(V) and methylate arsenite [As(III)], then inoculated worms were transferred to inactive E. coli AW3110 (harboring no arsenic transformation gene)-seeded plates treated with As(V) at different concentrations. Quantification of gut microbes showed that both E. coli BL21 and BL21M stably colonized in the guts after worms were cultured on inactive E. coli AW3110-seeded plates for 72 h. The analysis of arsenic species indicated that there was As(III) in C. elegans guts colonized with E. coli BL21, As(III) and dimethylarsinic acid [DMAs(V)] in C. elegans guts with E. coli BL21M exposed to As(V) for 6 h. After treatment of 100 µM As(V), decrease in brood sizes was observed for worms that were colonized with E. coli BL21 or BL21M compared to that with AW3110 in the guts. The levels of vitellogenin (VTG), glutathione S-transferases (GST) and superoxide dismutase (SOD), closely linked to reproduction and antioxidation-linked indicators, were the highest in worms whose guts colonized with E. coli BL21, followed by worms colonized with E. coli BL21M and worms colonized with inactive E. coli AW3110 exposed to As(V). Our results indicated the toxic impact of As(III) and DMAs(V) produced by gut microbes on reproduction of C. elegans. The work provides novel insight into the interplay between arsenic biotransformation mediated by gut microbes and the host fecundity in soils.

Arsenic and Sulfamethoxazole Increase the Incidence of Antibiotic Resistance Genes in the Gut of Earthworm.

Combinations of metal(loid) contamination and antibiotics are considered to increase the abundance of resistance genes in the environment, whereas the combined effect of metal(loid)s and antibiotics on microbial communities and antibiotic resistance genes (ARGs) in the gut of soil fauna remains unknown. We investigated herein the alteration of ARGs and the gut microbial communities after the earthworm Metaphire sieboldi was exposed to arsenate and/or sulfamethoxazole using high-throughput quantitative PCR and Illumina sequencing analysis. Arsenic accumulation in the body tissues of arsenic-exposed earthworms exerted a significant inhibition on growth and survival. The synergistic interactions of arsenic and sulfamethoxazole increased significantly the incidence of ARGs and mobile genetic elements in the earthworm gut microbiota. In addition, co-exposure to arsenic and sulfamethoxazole altered the structure of the gut microbial communities, and the changes correlated with ARG profiles of the gut microbiota. Our results indicate that the gut of soil fauna is a neglected hotspot of antibiotic resistance.

Exposure to microplastics lowers arsenic accumulation and alters gut bacterial communities of earthworm Metaphire californica.

Ubiquitous contamination of microplastics and arsenic in soil ecosystems can induce many health issues to nontarget soil organisms, and will also cause many potential threats to the gut bacterial communities of soil fauna. However, the changes in the gut bacterial communities of soil fauna after exposure to both microplastics and arsenic remain unknown. In this study, the toxicity and effects on the gut microbiota of earthworm Metaphire californica caused by the combined exposure of microplastics and arsenic were examined by using arsenic species analysis and high throughput sequencing of gut microbiota. Results showed that total arsenic and arsenic species in the earthworm gut and body tissues after exposure to combination of microplastics with arsenate (As(V)) were significantly different from that treated with As(V) alone. Microplastics lessened the accumulation of total arsenic and the transformation rate of As(V) to arsenite (As(III)). Microplastics alleviated the effect of arsenic on the gut microbiota possibly via adsorbing/binding As(V) and lowering arsenic bioavailability, thus prevented the reduction of As(V) and accumulation of total arsenic in the gut which resulted in a lower toxicity on the earthworm. The study broadens our understanding of the ecotoxicity of microplastics with other pollutants on the soil animals and on their gut microbiota.

Effects of Arsenic on Gut Microbiota and Its Biotransformation Genes in Earthworm Metaphire sieboldi.

Arsenic biotransformation mediated by gut microbiota can affect arsenic bioavailability and microbial community. Arsenic species, arsenic biotransformation genes (ABGs), and the composition of gut microbial community were characterized after the earthworm Metaphire sieboldi was cultured in soils spiked with different arsenic concentrations. Arsenite (As(III)) was the major component in the earthworm gut, whereas arsenate (As(V)) was predominant in the soil. A total of 16 ABGs were quantified by high-throughput quantitative polymerase chain reaction (HT-qPCR). Genes involved in arsenic redox and efflux were predominant in all samples, and the abundance of ABGs involved in arsenic methylation and demethylation in the gut was very low. These results reveal that the earthworm gut can be a reservoir of microbes with the capability of reducing As(V) and extruding As(III) but with little methylation of arsenic. Moreover, gut microbial communities were dominated by Actinobacteria, Firmicutes, and Proteobacteria at the phylum level and were considerably different from those in the surrounding soil. Our work demonstrates that exposure to As(V) disturbs the gut microbiota of earthworms and provides some insights into arsenic biotransformation in the earthworm gut.

Nitrogen inputs are more important than denitrifier abundances in controlling denitrification-derived N2O emission from both urban and agricultural soils.

Cities are increasingly being recognized as important contributors in global warming, for example by increasing atmospheric nitrous oxide (N2O). However, urban ecosystems remain poorly understood due to their functional complexity. Further, few studies have documented the microbial processes governing the N2O emissions from urban soils. Here, a field study was performed to assess in situ N2O emissions in an urban and agricultural soil located in Xiamen, China. The mechanisms underlying the difference in N2O emission patterns in both soils were further explored in an incubation experiment. Field investigations showed that N2O emission (3.5-19.0 µg N2O-N m-2 h-1) from the urban soil was significantly lower than that from the agricultural soil (25.4-18,502.3 µg N2O-N m-2 h-1). Incubation experiments showed that the urban soil initially emitted lower denitrification-derived N2O because of the lower nirS (encoding nitrite reductases) abundances, whereas overall N2O accumulation during the incubation was mainly controlled by the initial nitrate content in soil. Nitrate addition in a short period (5 days) did not change the total bacterial and denitrifier abundances or the soil bacterial community composition, but significantly altered the relative distribution of some key genera capable of denitrification. Although the urban soil exhibited lower N2O emission than its agricultural counterpart in this study, the expanding urban green areas should be taken into account when building N2O emission reduction targets.

Identification of Steps in the Pathway of Arsenosugar Biosynthesis.

Arsenosugars are arsenic-containing ribosides that play a substantial role in arsenic biogeochemical cycles. Arsenosugars were identified more than 30 years ago, and yet their mechanism of biosynthesis remains unknown. In this study we report identification of the arsS gene from the cyanobacterium Synechocystis sp. PCC 6803 and show that it is involved in arsenosugar biosynthesis. In the Synechocystis sp. PCC 6803 ars operon, arsS is adjacent to the arsM gene that encodes an As(III) S-adenosylmethionine (SAM) methyltransferase. The gene product, ArsS, contains a characteristic CX3CX2C motif which is typical for the radical SAM superfamily. The function of ArsS was identified from a combination of arsS disruption in Synechocystis sp. PCC 6803 and heterologous expression of arsM and arsS in Escherichia coli. Both genes are necessary, indicating a multistep pathway of arsenosugar biosynthesis. In addition, we demonstrate that ArsS orthologs from three other freshwater cyanobacteria and one picocyanobacterium are involved in arsenosugar biosynthesis in those microbes. This study represents the identification of the first two steps in the pathway of arsenosugar biosynthesis. Our discovery expands the catalytic repertoire of the diverse radical SAM enzyme superfamily and provides a basis for studying the biogeochemistry of complex organoarsenicals.

Linking Genes to Microbial Biogeochemical Cycling: Lessons from Arsenic.

The biotransformation of arsenic is highly relevant to the arsenic biogeochemical cycle. Identification of the molecular details of microbial pathways of arsenic biotransformation coupled with analyses of microbial communities by meta-omics can provide insights into detailed aspects of the complexities of this biocycle. Arsenic transformations couple to other biogeochemical cycles, and to the fate of both nutrients and other toxic environmental contaminants. Microbial redox metabolism of iron, carbon, sulfur, and nitrogen affects the redox and bioavailability of arsenic species. In this critical review we illustrate the biogeochemical processes and genes involved in arsenic biotransformations. We discuss how current and future metagenomic-, metatranscriptomic-, metaproteomic-, and metabolomic-based methods will help to decipher individual microbial arsenic transformation processes, and their connections to other biogeochemical cycle. These insights will allow future use of microbial metabolic capabilities for new biotechnological solutions to environmental problems. To understand the complex nature of inorganic and organic arsenic species and the fate of environmental arsenic will require integrating systematic approaches with biogeochemical modeling. Finally, from the lessons learned from these studies of arsenic biogeochemistry, we will be able to predict how the environment changes arsenic, and, in response, how arsenic biotransformations change the environment.

Arsenic biotransformation by a cyanobacterium Nostoc sp. PCC 7120.

Nostoc sp. PCC 7120 (Nostoc), a typical filamentous cyanobacterium ubiquitous in aquatic system, is recognized as a model organism to study prokaryotic cell differentiation and nitrogen fixation. In this study, Nostoc cells incubated with arsenite (As(III)) for two weeks were extracted with dichloromethane/methanol (DCM/MeOH) and the extract was partitioned between water and DCM. Arsenic species in aqueous and DCM layers were determined using high performance liquid chromatography - inductively coupled plasma mass spectrometer/electrospray tandem mass spectrometry (HPLC-ICPMS/ESIMSMS). In addition to inorganic arsenic (iAs), the aqueous layer also contained monomethylarsonate (MAs(V)), dimethylarsinate (DMAs(V)), and the two arsenosugars, namely a glycerol arsenosugar (Oxo-Gly) and a phosphate arsenosugar (Oxo-PO4). Two major arsenosugar phospholipids (AsSugPL982 and AsSugPL984) were detected in DCM fraction. Arsenic in the growth medium was also investigated by HPLC/ICPMS and shown to be present mainly as the inorganic forms As(III) and As(V) accounting for 29%-38% and 29%-57% of the total arsenic respectively. The total arsenic of methylated arsenic, arsenosugars, and arsenosugar phospholipids in Nostoc cells with increasing As(III) exposure were not markedly different, indicating that the transformation to organoarsenic in Nostoc was not dependent on As(III) concentration in the medium. Our results provide new insights into the role of cyanobacteria in the biogeochemical cycling of arsenic.

Co-expression of Cyanobacterial Genes for Arsenic Methylation and Demethylation in Escherichia coli Offers Insights into Arsenic Resistance.

Arsenite [As(III)] and methylarsenite [MAs(III)] are the most toxic inorganic and methylated arsenicals, respectively. As(III) and MAs(III) can be interconverted in the unicellular cyanobacterium Nostoc sp. PCC 7120 (Nostoc), which has both the arsM gene (NsarsM), which is responsible for arsenic methylation, and the arsI gene (NsarsI), which is responsible for MAs(III) demethylation. It is not clear how the cells prevent a futile cycle of methylation and demethylation. To investigate the relationship between arsenic methylation and demethylation, we constructed strains of Escherichia coli AW3110 (ΔarsRBC) expressing NsarsM or/and NsarsI. Expression of NsarsI conferred MAs(III) resistance through MAs(III) demethylation. Compared to NsArsI, NsArsM conferred higher resistance to As(III) and lower resistance to MAs(III) by methylating both As(III) and MAs(III). The major species found in solution was dimethylarsenate [DMAs(V)]. Co-expression of NsarsM and NsarsI conferred As(III) resistance at levels similar to that with NsarsM alone, although the main species found in solution after As(III) biotransformation was methylarsenate [MAs(V)] rather than DMAs(V). Co-expression of NsarsM and NsarsI conferred a higher level of resistance to MAs(III) than found with expression of NsarsM alone but lower than expression of only NsarsI. Cells co-expressing both genes converted MAs(III) to a mixture of As(III) and DMAs(V). In Nostoc NsarsM is constitutively expressed, while NsarsI is inducible by either As(III) or MAs(III). Thus, our results suggest that at low concentrations of arsenic, NsArsM activity predominates, while NsArsI activity predominates at high concentrations. We propose that coexistence of arsM and arsI genes in Nostoc could be advantageous for several reasons. First, it confers a broader spectrum of resistance to both As(III) and MAs(III). Second, at low concentrations of arsenic, the MAs(III) produced by NsArsM will possibly have antibiotic-like properties and give the organism a competitive advantage. Finally, these results shed light on the role of cyanobacteria in the arsenic biogeochemical cycle.

Arsenic Methyltransferase is Involved in Arsenosugar Biosynthesis by Providing DMA.

Arsenic is an ubiquitous toxic element in the environment, and organisms have evolved different arsenic detoxification strategies. Studies on arsenic biotransformation mechanisms have mainly focused on arsenate (As(V)) reduction, arsenite (As(III)) oxidation, and arsenic methylation; little is known, however, about the pathway for the biosynthesis of arsenosugars, which are significant arsenic transformation products. Here, the involvement of As(III) S-Adenosylmethionine methyltransferase (ArsM) in arsenosugar synthesis is demonstrated for the first time. Synechocystis sp. PCC 6803 incubated with As(III) or monomethylarsonic acid (MMA(V)) produced dimethylarsinic acid (DMA(V)) and arsenosugars, as determined by high performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC/ICPMS). Arsenosugars were also detected in the cells when they were exposed to DMA(V). A mutant strain Synechocystis ΔarsM was constructed by disrupting arsM in Synechocystis sp. PCC 6803. Methylation of arsenic species was not observed in the mutant strain after exposure to arsenite or MMA(V); when Synechocystis ΔarsM was incubated with DMA(V), arsenosugars were detected in the cells. These results suggest that ArsM is a required enzyme for the methylation of inorganic arsenicals, but not required for the synthesis of arsenosugars from DMA, and that DMA is the precursor of arsenosugar biosynthesis. The findings will stimulate more studies on the biosynthesis of complex organoarsenicals, and lead to a better understanding of the bioavailability and function of the organoarsenicals in biological systems.

Biochar decreases nitrogen oxide and enhances methane emissions via altering microbial community composition of anaerobic paddy soil.

Biochar application to agricultural soil is an appealing approach to mitigate nitrous oxide (N2O) and methane (CH4) emissions. However, the underlying microbial mechanisms are unclear. In this study, a paddy soil slurry was incubated anaerobically for 14d with biochar amendments produced from rice straw at 300, 500, or 700°C (B300, B500, and B700) to study their influences on greenhouse gas emissions. Illumina sequencing was used to characterize shift of soil bacterial and archaeal community composition. After peaking at day 1, N2O emission then sharply decreased to low levels while CH4 started to emit at day 3 then continually increased with incubation. Compared to control soil (57.9mgkg-1 soil), B300, B500, and B700 amendments decreased N2O peak emission to 17.9, 1.28, and 0.59mgkg-1, mainly due to increased soil pH. In contrast, the amendments enhanced CH4 production from 58.2 to 93.4, 62.6, and 63.4mgkg-1 at day 14 due to increased soil dissolved organic carbon. Abundance of denitrifying bacteria (e.g., Bacilli, 7.07-13.6 vs. 16.9%) was reduced with biochar amendments, especially with B500 and B700, contributing to the decreased N2O emissions. However, larger pore size of B500 and B700 (surface area of 68.1 and 161m2g-1) than B300 (4.40m2g-1) favored electron transfer between bacteria and iron minerals, leading to increased abundance of iron-reducing bacteria, (e.g., Clostridia, 48.2-50.6 vs. 33.3%), which competed with methanogens to produce CH4, thereby leading to lower increase in CH4 emission. Biochar amendments with high pH and surface area might be effective to mitigate emission of both N2O and CH4 from paddy soil.