Arsenic and phosphorus are group 15 elements with similar chemical properties. Is it possible that arsenate could replace phosphate in some of the chemicals that are required for life? Phosphate esters are ubiquitous in biomolecules and are essential for life, from the sugar phosphates of intermediary metabolism to ATP to phospholipids to the phosphate backbone of DNA and RNA. Some enzymes that form phosphate esters catalyze the formation of arsenate esters. Arsenate esters hydrolyze very rapidly in aqueous solution, which makes it improbable that phosphorous could be completely replaced with arsenic to support life. Studies of bacterial growth at high arsenic:phosphorus ratios demonstrate that relatively high arsenic concentrations can be tolerated, and that arsenic can become involved in vital functions in the cell, though likely much less efficiently than phosphorus. Recently Wolfe-Simon et al.  reported the isolation of a microorganism that they maintain uses arsenic in place of phosphorus for growth. Here, we examine and evaluate their data and conclusions.
arsenate; arsenic life; ester hydrolysis; phosphate
A strain of Halomonas bacteria, GFAJ-1, has been reported to be able to use arsenate as a nutrient when phosphate is limiting, and to specifically incorporate arsenic into its DNA in place of phosphorus. However, we have found that arsenate does not contribute to growth of GFAJ-1 when phosphate is limiting and that DNA purified from cells grown with limiting phosphate and abundant arsenate does not exhibit the spontaneous hydrolysis expected of arsenate ester bonds. Furthermore, mass spectrometry showed that this DNA contains only trace amounts of free arsenate and no detectable covalently bound arsenate.
A recent finding of a bacterial strain (GFAJ-1) that
can rely on
arsenic instead of phosphorus raised the questions of if and how arsenate
can replace phosphate in biomolecules that are essential to sustain
cell life. Apart from questions related to chemical stability, there
are those of the structural and functional consequences of phosphate-arsenate
substitutions in vital nucleotides in GFAJ1-like cells. In this study
we selected three types of molecules (ATP/ADP as energy source and
replication regulation; DNA–protein complexes for DNA replication
and transcription initiation; and a tRNA–protein complex and
ribosome for protein synthesis) to computationally probe if arsenate
nucleotides can retain the structural and functional features of phosphate
nucleotides. Hydrolysis of adenosine triarsenate provides 2–3
kcal/mol less energy than ATP hydrolysis. Arsenate DNA/RNA interacts
with proteins slightly less strongly than phosphate DNA/RNA, mainly
due to the weaker electrostatic interactions of arsenate. We observed
that the weaker arsenate RNA–protein interactions may hamper
rRNA assembly into a functional ribosome. We further compared the
experimental EXAFS spectra of the arsenic bacteria with theoretical
EXAFS spectra for arsenate DNA and rRNA. Our results demonstrate that
while it is possible that dried GFAJ-1 cells contain linear arsenate
DNA, the arsenate 70S ribosome does not contribute to the main arsenate
depository in the GFAJ-1 cell. Our study indicates that evolution
has optimized the inter-relationship between proteins and DNA/RNA,
which requires overall changes at the molecular and systems biology
levels when replacing phosphate by arsenate.
A genome-wide association study identifies the enzyme in plants that transforms arsenate into arsenite, allowing its extrusion into the soil and thereby controlling arsenic accumulation.
Inorganic arsenic is a carcinogen, and its ingestion through foods such as rice presents a significant risk to human health. Plants chemically reduce arsenate to arsenite. Using genome-wide association (GWA) mapping of loci controlling natural variation in arsenic accumulation in Arabidopsis thaliana allowed us to identify the arsenate reductase required for this reduction, which we named High Arsenic Content 1 (HAC1). Complementation verified the identity of HAC1, and expression in Escherichia coli lacking a functional arsenate reductase confirmed the arsenate reductase activity of HAC1. The HAC1 protein accumulates in the epidermis, the outer cell layer of the root, and also in the pericycle cells surrounding the central vascular tissue. Plants lacking HAC1 lose their ability to efflux arsenite from roots, leading to both increased transport of arsenic into the central vascular tissue and on into the shoot. HAC1 therefore functions to reduce arsenate to arsenite in the outer cell layer of the root, facilitating efflux of arsenic as arsenite back into the soil to limit both its accumulation in the root and transport to the shoot. Arsenate reduction by HAC1 in the pericycle may play a role in limiting arsenic loading into the xylem. Loss of HAC1-encoded arsenic reduction leads to a significant increase in arsenic accumulation in shoots, causing an increased sensitivity to arsenate toxicity. We also confirmed the previous observation that the ACR2 arsenate reductase in A. thaliana plays no detectable role in arsenic metabolism. Furthermore, ACR2 does not interact epistatically with HAC1, since arsenic metabolism in the acr2 hac1 double mutant is disrupted in an identical manner to that described for the hac1 single mutant. Our identification of HAC1 and its associated natural variation provides an important new resource for the development of low arsenic-containing food such as rice.
Arsenic is a human carcinogen that accumulates from soil into many different food crops, where it presents a significantly increased cancer risk when foods derived from these crops are consumed. Plants naturally control the amount of arsenic they accumulate by first chemically converting arsenate into arsenite, which is then extruded from the roots back into the soil. Because arsenate is a chemical analogue of phosphate, conversion of arsenate in the root to arsenite may also prevent arsenic being efficiently transported to the shoots via the phosphate transport system. The chemical reduction of arsenate to generate arsenite is therefore clearly a key component of a plant's detoxification strategy. Here, we use genetic methods to identify the enzyme responsible for this crucial reaction—HAC1. We show that HAC1 is responsible for arsenate reductase activity in both the outer layer of the root (epidermis) and the inner layer adjacent to the xylem (pericycle). In its absence, the roots return less arsenic to the soil and the shoots accumulate up to 300 times more arsenic. This knowledge creates new opportunities to limit arsenic accumulation in food crops, thereby helping to reduce the cancer risk from this food-chain contaminant.
The recent paper by Wolfe-Simon et al.1 reporting a bacterial strain, which is able to grow in high concentrations of arsenate, apparently in the absence of phosphate, and claims that in this strain arsenate is substituting for phosphate, e.g. in nucleic acids (Figure 1), was highly profiled, attracted broad attention, and almost immediately resulted in heavy scientific criticism (see e.g. 2–7).
Arsenate; DNA; evolution; origin of life; bacteria
A newly identified bacterial strain that can grow in the presence of arsenate, and possibly in the absence of phosphate, has raised much interest, but also fueled an active debate. Can arsenate substitute for phosphate in some, or possibly in most, of the absolutely essential phosphate-based biomolecules, including DNA? If so, then the possibility of alternative, arsenic-based life forms must be considered. The physicochemical similarity of these two oxyanions speaks in favor of this idea. However, arsenate-esters, and arsenate-diesters in particular, are extremely unstable in aqueous media. Here we explore the potential of arsenate to be used as substrate by phosphate-utilizing enzymes. We review the existing literature on arsenate enzymology, that intriguingly, dates back to the 1930s, and Otto Warburg. We address the issue of how and to what degree proteins can distinguish between arsenate and phosphate, and what is known in general about oxyanion specificity. And, we also discuss how phosphate-arsenate promiscuity may affect evolutionary transitions between phosphate and arsenate based biochemistry. Finally, we highlight potential applications of arsenate as a structural and mechanistic probe of enzymes whose catalyzed reactions involve the making or breaking of phosphoester bonds.
Phytoplankton plays an important role in arsenic speciation, distribution, and cycling in freshwater environments. Little information, however, is available on arsenic efflux from the cyanobacteria Microcystis aeruginosa under different phosphate regimes. This study investigated M. aeruginosa arsenic efflux and speciation by pre-exposing it to 10 µM arsenate or arsenite for 24 h during limited (12 h) and extended (13 d) depuration periods under phosphate enriched (+P) and phosphate depleted (−P) treatments. Arsenate was the predominant species detected in algal cells throughout the depuration period while arsenite only accounted for no greater than 45% of intracellular arsenic. During the limited depuration period, arsenic efflux occurred rapidly and only arsenate was detected in solutions. During the extended depuration period, however, arsenate and dimethylarsinic acid (DMA) were found to be the two predominant arsenic species detected in solutions under −P treatments, but arsenate was the only species detected under +P treatments. Experimental results also suggest that phosphorus has a significant effect in accelerating arsenic efflux and promoting arsenite bio-oxidation in M. aeruginosa. Furthermore, phosphorus depletion can reduce arsenic efflux from algal cells as well as accelerate arsenic reduction and methylation. These findings can contribute to our understanding of arsenic biogeochemistry in aquatic environments and its potential environmental risks under different phosphorus levels.
Background: Diabetes affects an estimated 346 million persons globally, and total deaths from diabetes are projected to increase > 50% in the next decade. Understanding the role of environmental chemicals in the development or progression of diabetes is an emerging issue in environmental health. In 2011, the National Toxicology Program (NTP) organized a workshop to assess the literature for evidence of associations between certain chemicals, including inorganic arsenic, and diabetes and/or obesity to help develop a focused research agenda. This review is derived from discussions at that workshop.
Objectives: Our objectives were to assess the consistency, strength/weaknesses, and biological plausibility of findings in the scientific literature regarding arsenic and diabetes and to identify data gaps and areas for future evaluation or research. The extent of the existing literature was insufficient to consider obesity as an outcome.
Data Sources, Extraction, and Synthesis: Studies related to arsenic and diabetes or obesity were identified through PubMed and supplemented with relevant studies identified by reviewing the reference lists in the primary literature or review articles.
Conclusions: Existing human data provide limited to sufficient support for an association between arsenic and diabetes in populations with relatively high exposure levels (≥ 150 µg arsenic/L in drinking water). The evidence is insufficient to conclude that arsenic is associated with diabetes in lower exposure (< 150 µg arsenic/L drinking water), although recent studies with better measures of outcome and exposure support an association. The animal literature as a whole was inconclusive; however, studies using better measures of diabetes-relevant end points support a link between arsenic and diabetes.
animal; arsenic toxicity; cell line; chemically induced/epidemiology; cultured cell; diabetes; environmental epidemiology; glucose; insulin; metabolism; obesity
Chronic arsenic exposure is a worldwide health problem. Although arsenic-induced cancer has been widely studied, comparatively little attention has been paid to arsenic-induced vascular disease. Epidemiological studies have shown that chronic arsenic exposure is associated with increased morbidity and mortality from cardiovascular disease. In addition, studies suggest that susceptibility to arsenic-induced vascular disease may be modified by nutritional factors in addition to genetic factors. Recently, animal models for arsenic-induced atherosclerosis and liver sinusoidal endothelial cell dysfunction have been developed. Initial studies in these models show that arsenic exposure accelerates and exacerbates atherosclerosis in apolipoprotein E–knockout mice. Microarray studies of liver mRNA and micro-RNA abundance in mice exposed in utero suggest that a permanent state of stress is induced by the arsenic exposure. Furthermore, the livers of the arsenic-exposed mice have activated pathways involved in immune responses suggesting a pro-hyperinflammatory state. Arsenic exposure of mice after weaning shows a clear dose-response in the extent of disease exacerbation. In addition, increased inflammation in arterial wall is evident. In response to arsenic-stimulated oxidative signaling, liver sinusoidal endothelium differentiates into a continuous endothelium that limits nutrient exchange and waste elimination. Data suggest that nicotinamide adenine dinucleotide phosphate oxidase–derived superoxide or its derivatives are essential second messengers in the signaling pathway for arsenic-stimulated vessel remodeling. The recent findings provide future directions for research into the cardiovascular effects of arsenic exposure.
arsenic; inflammation; oxidative signaling; vascular disease; nutrition; microarray
Arsenic exposure induces overproduction of reactive nitrogen species (RNS) in brain tissue and results in nucleic acid damage to the nerve cells. The 8-nitroguanine is one of the major products formed by the reaction of guanine, and ONOO-, and has been used as a popular biomarker of nucleic acid damage due to RNS attacking. In the present study, we examined whether the administration of taurine can protect against nucleic acid damage of brain neurons by arsenic-induced RNS.
Materials and methods
Sixty mice (30 male and 30 female) weighing 19.5 ± 1.5 g were divided into 3 groups: (1) control group, (2) experimental group that received arsenic (As2O3), and (3) antagonistic group that received taurine with arsenic. Arsenic was administered for 60 days. 8-Nitroguanine expressions in brain neurons of mice were examined by the immunohistochemical method. Histopathological changes in brain tissues of mice were observed under light microscope and the immunohistochemistry method was used to investigate 8-nitroguanine expressions in cerebrum and cerebellum of mice.
In the control group, no abnormal histopathological changes were observed in brain tissue of the mice. In brain tissue of the mice exposed to arsenic, histopathological results showed swells, evident vacuolar degeneration in cytoplasm, karyorrhexis and karyolysis. Relatively light pathological changes were observed in brain of the mice co-administered arsenic and taurine. Little or no expression of 8-nitroguanine in brain tissue was observed in controls. However, intensive expression of 8-nitroguanine was found in brain tissue of mice exposed to arsenic and it was mainly distributed in nucleus neighbouring the nuclear membrane, but a little in cytoplasm. A weak expression of 8-nitroguanine was observed in brain cells of mice co-administered arsenic and taurine.
The brain neurons may be the major target cells of arsenic neurotoxicity. Co-administration of arsenic and taurine can alleviate DNA damage of brain neurons caused by arsenic through the RNS signal pathway.
Biomolecules are composed primarily of the elements carbon, nitrogen, hydrogen, oxygen, sulfur, and phosphorus. The structured assembly of these elements forms the basis for proteins, nucleic acids and lipids. However, the recent discovery of a new bacterium, strain GFAJ-1 of the Halomonadaceae, has shaken the classic paradigms for the architecture of life. Mounting evidence supports the claim that these bacteria substitute arsenic for phosphorus in macromolecules. Herein, we provide a brief commentary and fuel the debate related to what may be a most unusual organism.
arsenic; GFAJ-1; biomolecules; scientific debate; elemental substitution
While epidemiological methods have grown in sophistication during the 20th century, their application in historical occupational (and environmental) health research has also led to a corresponding growth in uncertainty in the validity and reliability of the attribution of risk in the resulting studies, particularly where study periods extend back in time to the immediate postwar era (1945–70) when exposure measurements were sporadic, unsystematically collected and primitive in technique; and, more so, to the pre-WWII era (when exposure data were essentially non-existent). These uncertainties propagate with animal studies that are designed to confirm the carcinogenicity by inhalation exposure of a chemical putatively responsible for historical workplace cancers since exact exposure conditions were never well characterized. In this report, we present a weight of scientific evidence examination of the human and toxicological evidence to show that soluble nickel is not carcinogenic; and, furthermore, that the carcinogenic potencies previously assigned by regulators to sulphidic and oxidic nickel compounds for the purposes of developing occupational exposure limits have likely been overestimated.
Published, file and archival evidence covering the pertinent epidemiology, biostatistics, confounding factors, toxicology, industrial hygiene and exposure factors, and other risky exposures were examined to evaluate the soluble nickel carcinogenicity hypothesis; and the likely contribution of a competing workplace carcinogen (arsenic) on sulphidic and oxidic nickel risk estimates.
Sharp contrasts in available land area and topography, and consequent intensity of production and refinery process layouts, likely account for differences in nickel species exposures in the Kristiansand (KNR) and Port Colborne (PCNR) refineries. These differences indicate mixed sulphidic and oxidic nickel and arsenic exposures in KNR's historical electrolysis department that were previously overlooked in favour of only soluble nickel exposure; and the absence of comparable insoluble nickel exposures in PCNR's tankhouse, a finding that is consistent with the absence of respiratory cancer risk there. The most recent KNR evidence linking soluble nickel with lung cancer risk arose in a reconfiguration of KNR's historical exposures. But the resulting job exposure matrix lacks an objective, protocol-driven rationale that could provide a valid and reliable basis for analyzing the relationship of KNR lung cancer risk with any nickel species. Evidence of significant arsenic exposure during the processing step in the Clydach refinery's hydrometallurgy department in the 1902–1934 time period likely accounts for most of the elevated respiratory cancer risk observed at that time. An understanding of the mechanism for nickel carcinogenicity remains an elusive goal of toxicological research; as does its capacity to confirm the human health evidence on this subject with animal studies.
Epidemiological methods have failed to accurately identify the source(s) of observed lung cancer risk in at least one nickel refinery (KNR). This failure, together with the negative long-term animal inhalation studies on soluble nickel and other toxicological evidence, strongly suggest that the designation of soluble nickel as carcinogenic should be reconsidered, and that the true causes of historical lung cancer risk at certain nickel refineries lie in other exposures, including insoluble nickel compounds, arsenic, sulphuric acid mists and smoking.
Inorganic arsenic is clearly a human carcinogen causing tumors of the skin, lung, urinary bladder, and possibly liver (IARC, 2004). At the time of construction of this monograph, the evidence for arsenic as a hepatocarcinogen in humans was considered controversial and in rodents considered insufficient. However, recent data has accumulated indicating hepatocarcinogenicity of arsenic. This forum reevaluates epidemiology studies, rodent studies together with in vitro models, and focuses on the liver as a target organ of arsenic toxicity and carcinogenesis. Hepatocellular carcinoma and hepatic angiosarcoma, have been frequently associated with environmental or medicinal exposure to arsenicals. Preneoplastic lesions, including hepatomegaly, hepatoportal sclerosis, fibrosis, and cirrhosis often occur after chronic arsenic exposure. Recent work in mice clearly shows that exposure to inorganic arsenic during gestation induces tumors, including hepatocellular adenoma and carcinoma, in offspring when they reach adulthood. In rats, the methylated arsenicals, dimethylarsinic acid promotes diethylnitrosamine-initiated liver tumors, whereas trimethylarsine oxide induces liver adenomas. Chronic exposure of rat liver epithelial cells to low concentrations of inorganic arsenic induces malignant transformation, producing aggressive, undifferentiated epithelial tumors when inoculated into the Nude mice. There are a variety of potential mechanisms for arsenical-induced hepatocarcinogenesis, such as oxidative DNA damage, impaired DNA damage repair, acquired apoptotic tolerance, hyperproliferation, altered DNA methylation, and aberrant estrogen signaling. Some of these mechanisms may be liver specific/selective. Overall, accumulating evidence clearly indicates that the liver could be an important target of arsenic carcinogenesis.
arsenic; liver; carcinogenesis; epidemiology; animal models; mechanisms
The aim of this study was to detect arsenic concentrations in feed, well-water for drinking, eggs, and excreta of laying hens in arsenic-prone areas of Bangladesh and to assess the effect of arsenic-containing feed and well-water on the accumulation of arsenic in eggs and excreta of the same subject. One egg from each laying hen (n=248) and its excreta, feed, and well-water for drinking were collected. Total arsenic concentrations were determined by atomic absorption spectrophotometer, coupled with hydride generator. Effects of arsenic-containing feed and drinking-water on the accumulation of arsenic in eggs and excreta were analyzed by multivariate regression model, using Stata software. Mean arsenic concentrations in drinking-water, feed (dry weight [DW]), egg (wet weight [WW]), and excreta (DW) of hens were 77.3, 176.6, 19.2, and 1,439.9 ppb respectively. Significant (p<0.01) positive correlations were found between the arsenic contents in eggs and drinking-water (r=0.602), drinking-water and excreta (r=0.716), feed and excreta (r=0.402) as well as between the arsenic content in eggs and the age of the layer (r=0.243). On an average, 55% and 82% of the total variation in arsenic contents of eggs and excreta respectively could be attributed to the variation in the geographic area, age, feed type, and arsenic contents of drinking-water and feed. For each week's increase in age of hens, arsenic content in eggs increased by 0.94%. For every 1% elevation of arsenic in drinking-water, arsenic in eggs and excreta increased by 0.41% and 0.44% respectively whereas for a 1% rise of arsenic in feed, arsenic in eggs and excreta increased by 0.40% and 0.52% respectively. These results provide evidence that, although high arsenic level prevails in well-water for drinking in Bangladesh, the arsenic shows low biological transmission capability from body to eggs and, thus, the value was below the maximum tolerable limit for humans. However, arsenic in drinking-water and/or feed makes a significant contribution to the arsenic accumulations in eggs and excreta of laying hens.
Arsenic; Drinking-water; Egg; Excreta; Feed; Laying hen; Bangladesh
Halomonas strain GFAJ-1 was reported in Science magazine to be a remarkable microbe for which there was “arsenate in macromolecules that normally contain phosphate, most notably nucleic acids.” The draft genome of the bacterium was determined (NCBI accession numbers AHBC01000001 through AHBC01000103). It appears to be a typical gamma proteobacterium.
The effect of arsenate on strains dependent on the two major inorganic phosphate (Pi) transport systems in Escherichia coli was examined in cells grown in 1 mM phosphate medium. The development of arsenate-resistant Pi uptake in a strain dependent upon the Pst (phosphate specific transport) system was examined. The growth rate of Pst-dependent cells in arsenate-containing medium was a function of the arsenate-to-Pi ratio. Growth in arsenate-containing medium was not due to detoxification of the arsenate. Kinetic studies revealed that cells grown with a 10-fold excess of arsenate to Pi have almost a twofold increase in capacity (Vmax) for Pi, but maintained the same affinity (Km). Pi accumulation in the Pst-dependent strain was still sensitive to changes in the arsenate-to-Pi ratio, and a Ki (arsenate) for Pi transport of 39 microM arsenate was determined. The Pst-dependent strain did not accumulate radioactive arsenate, and showed only a transient decrease in intracellular adenosine triphosphate levels after arsenate was added to the medium. The Pi transport-dependent strain ceased growth in arsenate-containing media. This strain accumulated 74As-arsenate, and intracellular adenosine triphosphate pools were almost completely depleted after the addition of arsenate to the medium. Arsenate accumulation required a metabolizable energy source and was inhibited by N-ethylmaleimide. Previously accumulated arsenate could exchange with arsenate or Pi in the medium.
Exposure to arsenic, an established human carcinogen, through consumption of highly contaminated drinking water is a worldwide public health concern. Several mechanisms by which arsenical compounds induce tumorigenesis have been proposed, including oxidative stress, genotoxic damage, and chromosomal abnormalities. Recent studies have suggested that epigenetic mechanisms may also mediate toxicity and carcinogenicity resulting from arsenic exposure.
We examined the evidence supporting the roles of the three major epigenetic mechanisms—DNA methylation, histone modification, and microRNA (miRNA) expression—in arsenic toxicity and, in particular, carcinogenicity. We also investigated future research directions necessary to clarify epigenetic and other mechanisms in humans.
Data sources and synthesis
We conducted a PubMed search of arsenic exposure and epigenetic modification through April 2010 and summarized the in vitro and in vivo research findings, from both our group and others, on arsenic-associated epigenetic alteration and its potential role in toxicity and carcinogenicity.
Arsenic exposure has been shown to alter methylation levels of both global DNA and gene promoters; histone acetylation, methylation, and phosphorylation; and miRNA expression, in studies analyzing mainly a limited number of epigenetic end points. Systematic epigenomic studies in human populations exposed to arsenic or in patients with arsenic-associated cancer have not yet been performed. Such studies would help to elucidate the relationship between arsenic exposure, epigenetic dysregulation, and carcinogenesis and are becoming feasible because of recent technological advancements.
arsenic carcinogenesis; arsenical compounds; DNA methylation; epigenetics; histone modification; microRNA
Inorganic arsenicals have been used in agriculture as pesticides or defoliants for many years and, in localized areas, oxides of arsenic have contaminated soils as a result of fallout from ore-smelting operations and coal-fired power plants. Use of inorganic arsenicals is no longer permitted in most agricultural operations, and recent air pollution controls have markedly reduced contamination from smelters. Thus, this paper will concentrate on the effect of past applications on arsenic accumulation in soil, phytotoxicity to and uptake by plants as influenced by soil properties, and alleviation of the deleterious effects of arsenic.
Once incorporated into the soil, inorganic arsenical pesticides and arsenic oxides revert to arsenates, except where the soil is under reducing conditions. The arsenate ion has properties similar to that of orthophosphate, and is readily sorbed by iron and aluminum components. This reaction greatly restricts the downward movement (leaching) of arsenic in soils and the availability of arsenic to plants.
Several methods of estimating plant available arsenic in soils have been developed. They involve extraction of the soil with reagents used to estimate phosphorus availability. This extractable arsenic is reasonably well correlated with reduced plant growth by, and plant uptake of arsenic. For most plants, levels of arsenic in the edible portion of the plant are well below the critical concentration for animal or human consumption, even when severe phytotoxicity occurs.
Alleviation of arsenic phytotoxicity has been attempted by increasing the soil pH, by use of iron or aluminum sulfate, by desorbing arsenate with phosphate and subsequent leaching, and by cultural practices such as deep plowing. Only limited benefits have accrued from these procedures the cost of which is often prohibitively high. Since attempts to reduce arsenic toxicity have not been very successful, its excessive accumulation in soils should be avoided.
In the late twentieth century, emergence of high rates of treatment failure with antimonial compounds (SSG) for visceral leishmaniasis (VL) caused a public health crisis in Bihar, India. We hypothesize that exposure to arsenic through drinking contaminated groundwater may be associated with SSG treatment failure due to the development of antimony-resistant parasites.
A retrospective cohort design was employed, as antimony treatment is no longer in routine use. The study was performed on patients treated with SSG between 2006 and 2010. Outcomes of treatment were assessed through a field questionnaire and treatment failure used as a proxy for parasite resistance. Arsenic exposure was quantified through analysis of 5 water samples from within and surrounding the patient’s home. A logistic regression model was used to evaluate the association between arsenic exposure and treatment failure. In a secondary analysis survival curves and Cox regression models were applied to assess the risk of mortality in VL patients exposed to arsenic.
One hundred and ten VL patients treated with SSG were analysed. The failure rate with SSG was 59%. Patients with high mean local arsenic level had a non-statistically significant higher risk of treatment failure (OR = 1.78, 95% CI: 0.7–4.6, p = 0.23) than patients using wells with arsenic concentration <10 μg/L. Twenty one patients died in our cohort, 16 directly as a result of VL. Arsenic levels ≥ 10 μg/L increased the risk of all-cause (HR 3.27; 95% CI: 1.4–8.1) and VL related (HR 2.65; 95% CI: 0.96–7.65) deaths. This was time dependent: 3 months post VL symptom development, elevated risks of all-cause mortality (HR 8.56; 95% CI: 2.5–29.1) and of VL related mortality (HR 9.27; 95% CI: 1.8–49.0) were detected.
This study indicates a trend towards increased treatment failure in arsenic exposed patients. The limitations of the retrospective study design may have masked a strong association between arsenic exposure and selection for antimonial resistance in the field. The unanticipated strong correlation between arsenic exposure and VL mortality warrants further investigation.
The parasitic disease visceral leishmaniasis (VL) causes a significant burden of illness and death in India. The main drug used to treat VL, which is based on the chemical element antimony, stopped working well in about half of all patients in the late twentieth century. We hypothesised that arsenic exposure of the Indian population, through contaminated groundwater, was contributing to treatment failure with antimony based drugs. Arsenic and antimony are similar chemical elements and exposure of the parasite to arsenic within the liver of arsenic-exposed patients could allow the parasite to become resistant to treatment with antimony. Using a field-based questionnaire study we retrospectively evaluated whether arsenic exposure was linked to antimonial treatment failure in a cohort of 110 antimonial treated patients. No significant association was found, although this may be because the number of patients in the study was low as antimony use was officially discontinued in 2005 due to high rates of treatment failure. However, arsenic exposure was found to increase risk of mortality from VL particularly if death occurred more than 3 months after the symptoms of VL developed. More research into the relationship between arsenic exposure and mortality in VL is warranted.
The toxic metalloid arsenic is widely distributed in food, water, and soil. While inorganic arsenic enters the environment primarily from geochemical sources, methylarsenicals either result from microbial biotransformation of inorganic arsenic or are introduced anthropogenically. Methylarsenicals such as monosodium methylarsonic acid (MSMA) have been extensively utilized as herbicides, and aromatic arsenicals such as roxarsone (Rox) are used as growth promoters for poultry and swine. Organoarsenicals are degraded to inorganic arsenic. The toxicological effects of arsenicals depend on their oxidation state, chemical composition, and bioavailability. Here we report that the active forms are the trivalent arsenic-containing species. We constructed a whole-cell biosensor utilizing a modified ArsR repressor that is highly selective toward trivalent methyl and aromatic arsenicals, with essentially no response to inorganic arsenic. The biosensor was adapted for in vitro detection of organoarsenicals using fluorescence anisotropy of ArsR-DNA interactions. It detects bacterial biomethylation of inorganic arsenite both in vivo and in vitro with detection limits of 10−7 M and linearity to 10−6 M for phenylarsenite and 5×10−6 M for methylarsenite. The biosensor detects reduced forms of MSMA and roxarsone and offers a practical, low cost method for detecting activate forms and breakdown products of organoarsenical herbicides and growth promoters.
Previous research demonstrated that 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment increased the number of skin papillomas in v-Ha-ras transgenic (Tg.AC) mice that had received sodium arsenite [(As(III)] in drinking water, indicating that this model is useful for studying the toxic effects of arsenic in vivo. Because the liver is a known target of arsenic, we examined the pathophysiologic and molecular effects of inorganic and organic arsenical exposure on Tg.AC mouse liver in this study. Tg.AC mice were provided drinking water containing As(III), sodium arsenate [As(V)], monomethylarsonic acid [(MMA(V)], and 1,000 ppm dimethylarsinic acid [DMA(V)] at dosages of 150, 200, 1,500, or 1,000 ppm as arsenic, respectively, for 17 weeks. Control mice received unaltered water. Four weeks after initiation of arsenic treatment, TPA at a dose of 1.25 μg/200 μL acetone was applied twice a week for 2 weeks to the shaved dorsal skin of all mice, including the controls not receiving arsenic. In some cases arsenic exposure reduced body weight gain and caused mortality (including moribundity). Arsenical exposure resulted in a dose-dependent accumulation of arsenic in the liver that was unexpectedly independent of chemical species and produced hepatic global DNA hypomethylation. cDNA microarray and reverse transcriptase–polymerase chain reaction analysis revealed that all arsenicals altered the expression of numerous genes associated with toxicity and cancer. However, organic arsenicals [MMA(V) and DMA(V)] induced a pattern of gene expression dissimilar to that of inorganic arsenicals. In summary, subchronic exposure of Tg.AC mice to inorganic or organic arsenicals resulted in toxic manifestations, hepatic arsenic accumulation, global DNA hypomethylation, and numerous gene expression changes. These effects may play a role in arsenic-induced hepatotoxicity and carcinogenesis and may be of particular toxicologic relevance.
arsenicals (arsenic forms); gene expression; mouse liver; subchronic toxicity; toxicokinetics
Arsenate is a pentavalent form of arsenic that shares similar chemical properties to phosphate. It has been shown to be taken up by phosphate transporters in both eukaryotic and prokaryotic microbes such as yeast and Escherichia coli. Recently, the arsenate uptake in vertebrate cells was reported to be facilitated by mammalian type II sodium/phosphate transporter with different affinities. As arsenate is the most common form of arsenic exposure in aquatic system, identifying the uptake pathway of arsenate into aquatic animals is a crucial step in the elucidation of the entire metabolic pathway of arsenic. In this study, the ability of a zebrafish phosphate transporter, NaPi-IIb1 (SLC34a2a), to transport arsenate was examined. Our results demonstrate that a type II phosphate transporter in zebrafish, NaPi-IIb1, can transport arsenate in vitro when expressed in Xenopus laevis oocytes. NaPi-IIb1 mediates a high-affinity arsenate transport, with a Km of 0.22 mM. The natural substrate of NaPi-IIb1, dibasic phosphate, inhibits arsenate transport. Arsenate transport via NaPi-IIb1 is coupled with Na+ and exhibits sigmoidal kinetics with a Hill coefficient of 3.24 ± 0.19. Consistent with these in vitro studies, significant arsenate accumulation is observed in all examined zebrafish tissues where NaPi-IIb1 is expressed, particularly intestine, kidney, and eye, indicating that zebrafish NaPi-IIb1 is likely the transport protein that is responsible for arsenic accumulation in vivo.
Chronic exposures to arsenic and estrogen are known risk factors for prostate cancer. Though the evidence suggests that exposure to arsenic or estrogens can disrupt normal DNA methylation patterns and histone modifications, the mechanisms by which these chemicals induce epigenetic changes are not fully understood. Moreover, the epigenetic effects of co-exposure to these two chemicals are not known. Therefore, the objective of this study was to evaluate the effects of chronic exposure to arsenic and estrogen, both alone and in combination, on the expression of epigenetic regulatory genes, their consequences on DNA methylation, and histone modifications. Human prostate epithelial cells, RWPE-1, chronically exposed to arsenic and estrogen alone and in combination were used for analysis of epigenetic regulatory genes expression, global DNA methylation changes, and histone modifications at protein level. The result of this study revealed that exposure to arsenic, estrogen, and their combination alters the expression of epigenetic regulatory genes and changes global DNA methylation and histone modification patterns in RWPE-1 cells. These changes were significantly greater in arsenic and estrogen combination treated group than individually treated group. The findings of this study will help explain the epigenetic mechanism of arsenic- and/or estrogen-induced prostate carcinogenesis.
The pink yeast Rhodotorula rubra of marine origin was found to be capable of extended growth at very low phosphate concentrations (K0.5 = 10.8 nm). Average intracellular phosphate concentrations, based on isotope exchange techniques, were 15 to 200 nm, giving concentration gradients across the cell envelope of about 106. Sensitivity to metabolic inhibitors occurred at micromolar concentrations. Inability of the phosphate transport system, Ks = 0.5 to 2.8 μm, Vmax = 55 μmoles per g of cells per min, to discriminate against arsenate transport led to arsenate toxicity at 1 to 10 nm, whereas environmental arsenate levels are reportedly much higher. Phosphate competitively prevented arsenate toxicity. The Ki for phosphate inhibition of arsenate uptake was 0.7 to 1.2 μm. Phosphate uptake experiments showed that maximal growth rates could be achieved with approximately 4% of the total phosphate-arsenate transport system. Organisms adapted to a range both of concentration of NaCl and of pH. Maximal affinity for phosphate occurred at pH 4 and at low concentrations of NaCl; however, Vmax for phosphate transport was little affected. Maximal specific growth rates on minimal medium were consistent in batch culture but gradually increased to the much higher rates found with yeast extract media when the population was subjected to long-term continuous culture with gradually increasing dilution rates. Phosphate initial uptake rates that were in agreement with the steady-state flux in continuous culture were obtained by using organisms and medium directly from continuous culture. This procedure resulted in rates about 500 times greater than one in which harvested batch-grown cells were used. Discrepancies between values found and those reported in the literature for other organisms were even larger. Growth could not be sustained below a threshold phosphate concentration of 3.4 nm. Such thresholds are explained in terms of a system where growth rate is set by intracellular nutrient concentrations. Threshold concentrations occur in response to nutrient sinks not related to growth, such as efflux and endogenous metabolism. Equations are presented for evaluation of growth rate-limiting substrate concentrations in the presence of background substrate and for evaluating low inhibitor concentration inhibition mechanisms by substrate prevention of inhibitor flux.
Microbial arsenate respiration can enhance arsenic release from arsenic-bearing minerals—a process that can cause arsenic contamination of water. In Shewanella sp. strain ANA-3, the arsenate respiration genes (arrAB) are induced under anaerobic conditions with arsenate and arsenite. Here we report how genes that encode anaerobic regulator (arcA and etrA [fnr homolog]) and carbon catabolite repression (crp and cya) proteins affect arsenate respiration in ANA-3. Transcription of arcA, etrA, and crp in ANA-3 was similar in cells grown on arsenate and cells grown under aerobic conditions. ANA-3 strains lacking arcA and etrA showed minor to moderate growth defects, respectively, with arsenate. However, crp was essential for growth on arsenate. In contrast to the wild-type strain, arrA was not induced in the crp mutant in cultures shifted from aerobic to anaerobic conditions containing arsenate. This indicated that cyclic AMP (cAMP)-cyclic AMP receptor (CRP) activates arr operon transcription. Computation analysis for genome-wide CRP binding motifs identified a putative binding motif within the arr promoter region. This was verified by electrophoretic mobility shift assays with cAMP-CRP and several DNA probes. Lastly, four putative adenylate cyclase (cya) genes were identified in the genome. One particular cya-like gene was differentially expressed under aerobic versus arsenate respiration conditions. Moreover, a double mutant lacking two of the cya-like genes could not grow with arsenate as a terminal electron acceptor; exogenous cAMP could complement growth of the double cya mutant. It is concluded that the components of the carbon catabolite repression system are essential to regulating arsenate respiratory reduction in Shewanella sp. strain ANA-3.