Rapid differentiation between fluconazole-sensitive and -resistant species of Candida directly from positive blood-culture bottles by real-time PCR

In view of both the delay in obtaining identification by conventional methods following blood-culture positivity in patients with candidaemia and the close relationship between species and fluconazole (FLC) susceptibility, early speciation of positive blood cultures has the potential to influence therapeutic decisions. The aim was to develop a rapid test to differentiate FLC-resistant from FLC-sensitive Candida species. Three TaqMan-based real-time PCR assays were developed to identify up to six Candida species directly from BacT/Alert blood-culture bottles that showed yeast cells on Gram staining at the time of initial positivity. Target sequences in the rRNA gene complex were amplified, using a consensus two-step PCR protocol, to identify Candida albicans, Candida parapsilosis, Candida tropicalis, Candida dubliniensis, Candida glabrata and Candida krusei; these are the most commonly encountered Candida species in blood cultures. The first four of these (the characteristically FLC-sensitive group) were identified in a single reaction tube using one fluorescent TaqMan probe targeting 1 8S rRNA sequences conserved in the four species. The FLC-resistant species C. krusei and C. glabrata were detected in two further reactions, each with species-specific probes. This method was validated with clinical specimens (blood cultures) positive for yeast (n=33 sets) and the results were 100% concordant with those of phenotypic identification carried out concomitantly. The reported assay significantly reduces the time required to identify the presence of C. glabrata and C. krusei in comparison with a conventional phenotypic method, from ~72 to

Arsenic and selenium mobilisation from organic matter treated mine spoil with and without inorganic fertilisation

Organic matter amendments are applied to contaminated soil to provide a better habitat for revegetation and remediation, and olive mill waste compost (OMWC) has been described as a promising material for this aim. We report here the results of an incubation experiment carried out in flooded conditions to study its influence in As and metal solubility in a trace elements contaminated soil. NPK fertilisation and especially organic amendment application resulted in increased As, Se and Cu concentrations in pore water. Independent of the amendment, dimethylarsenic acid (DMA) was the most abundant As species in solution. The application of OMWC increased pore water dissolved organic-carbon (DOC) concentrations, which may explain the observed mobilisation of As, Cu and Se; phosphate added in NPK could also be in part responsible of the mobilisation caused in As. Therefore, the application of soil amendments in mine soils may be particularly problematic in flooded systems. (C) 2012 Elsevier Ltd. All rights reserved.

The dynamics of arsenic in four paddy fields in the Bengal delta

Irrigation with arsenic contaminated groundwater in the Bengal Delta may lead to As accumulation in the soil and rice grain. The dynamics of As concentration and speciation in paddy fields during dry season (boro) rice cultivation were investigated at 4 sites in Bangladesh and West Bengal, India. Three sites which were irrigated with high As groundwater had elevated As concentrations in the soils, showing a significant gradient from the irrigation inlet across the field. Arsenic concentration and speciation in soil pore water varied temporally and spatially; higher As concentrations were associated with an increasing percentage of arsenite, indicating a reductive mobilization. Concentrations of As in rice grain varied by 2-7 fold within individual fields and were poorly related with the soil As concentration. A field site employing alternating flooded-dry irrigation produced the lowest range of grain As concentration, suggesting a lower soil As availability caused by periodic aerobic conditions.

The impact of a rice based diet on urinary arsenic

Rice is elevated in arsenic (As) compared to other staple grains. The Bangladeshi community living in the United Kingdom (UK) has a ca. 30-fold higher consumption of rice than white Caucasians. In order to assess the impact of this difference in rice consumption, urinary arsenicals of 49 volunteers in the UK (Bangladeshi n = 37; white Caucasians n = 12) were monitored along with dietary habits. Total urinary arsenic (As(t)) and speciation analysis for dimethylarsinic acid (DMA), monomethylarsonic acid (MA) and inorganic arsenic (iAs) was conducted. Although no significant difference was found for As(t) (median: Bangladeshis 28.4 µg L(-1)) and white Caucasians (20.6 µg L(-1)), the sum of medians of DMA, MA and iAs for the Bangladeshi group was found to be over 3-fold higher (17.9 µg L(-1)) than for the Caucasians (3.50 µg L(-1)). Urinary DMA was significantly higher (p <0.001) in the UK Bangladeshis (median: 16.9 µg DMA L(-1)) than in the white Caucasians (3.16 µg DMA L(-1)) as well as iAs (p <0.001) with a median of 0.630 µg iAs L(-1) for Bangladeshi and 0.250 µg iAs L(-1) for Caucasians. Cationic compounds were significantly lower in the Bangladeshis (2.93 µg L(-1)) than in Caucasians (14.9 µg L(-1)). The higher DMA and iAs levels in the Bangladeshis are mainly the result of higher rice consumption: arsenic is speciated in rice as both iAs and DMA, and iAs can be metabolized, through MA, to DMA by humans. This study shows that a higher dietary intake of DMA alters the DMA/MA ratio in urine. Consequently, DMA/MA ratio as an indication of methylation capacity in populations consuming large quantities of rice should be applied with caution since variation in the quantity and type of rice eaten may alter this ratio.

Accumulation or production of arsenobetaine in humans?

Arsenobetaine has always been referred to as a non-toxic but readily bioavailable compound and the available data would suggest that it is neither metabolised by nor accumulated in humans. Here this study investigates the urine of five volunteers on an arsenobetaine exclusive diet for twelve days and shows that arsenobetaine was consistently excreted by three of the five volunteers. From the expected elimination pattern of arsenobetaine in rodents, no significant amount of arsenobetaine should have been detectable after 5 days of the trial period. The arsenobetaine concentration found in the urine was constant after 5 days and varied between 0.2 and 12.2 microg As per L for three of the volunteers. Contrary to the established belief that arsenobetaine is neither accumulated nor generated by humans, the presented results would suggest that either accumulated arsenobetaine in the tissues is slowly released over time or that arsenobetaine is a human metabolite of dimethylarsinic acid or inorganic arsenic from the trial food, or both. Either possibility is intriguing and raises fundamental questions about human arsenic metabolism and the toxicological and environmental inertness of arsenobetaine.

Arsenic shoot-grain relationships in field grown rice cultivars

Arsenic (As) accumulation in rice grains is a risk to human health. The mechanism of transfer of As from the shoot into the grain during grain filling is unknown at present. In this study As speciation in the shoot and grains at maturity were examined, and the relationships between phosphorus (P) and As, and silicon (Si) and As were established in a wide range of cultivars grown in As contaminated field trials in Bangladesh and China. No correlations were observed between shoot and grain speciation, with the inorganic form comprising 93.0-97.0% of As in the shoot and 63.0-83.7% in the grains. The percentage of dimethylarsinic acid (DMA) was between 1.4 and 6.6% in the shoot and 14.6 and 37.0% in the grains; however, the concentrations were comparable, ranging from 0.07 to 0.26 mg kg(-1) in the shoots and 0.03 to 0.25 mg kg(-1) in the grains. A positive correlation was observed between shoot As and shoot Si, however, no correlation was observed between shoot Si and grain As. A significant negative correlation was observed between shoot P and grain As concentrations. These results suggest that the translocation of As into the grain from the shoots is potentially using P rather than Si transport mechanisms. The findings also indicate that inorganic As and DMA translocation to the grain differ considerably.

Arsenic accumulation and phosphorus status in two rice (Oryza sativa L.) cultivars surveyed from fields in South China

The consumption of paddy rice (Oryza sativa L.) is a major inorganic arsenic exposure pathway in S.E. Asia. A multi-location survey was undertaken in Guangdong Province, South China to assess arsenic accumulation and speciation in 2 rice cultivars, one an Indica and the other a hybrid Indica. The results showed that arsenic concentrations in rice tissue increased in the order grain <husk <straw <root. Rice grain arsenic content of 2 rice cultivars was significant different and correlated with phosphorus concentration and molar ratio of P/As in shoot, being higher for the Indica cultivar than for the hybrid Indica, which suggests altering shoot phosphorus status as a promising route for breeding rice cultivars with reduced grain arsenic. Speciation of grain arsenic, performed using HPLC-ICP-MS, identified inorganic arsenic as the dominant arsenic species present in the rice grain.

Quantitative and qualitative trapping of volatile methylated selenium species entrained through nitric acid

Quantification and speciation of volatile selenium (Se) fluxes in remote areas has not been feasible previously, due to the absence of a simple and easily transportable trapping technique that preserves speciation. This paper presents a chemo-trapping method with nitric acid (HNO3) for volatile Se species, which preserves speciation of trapped compounds. The recovery and speciation of dimethylselenide (DMSe) and dimethyl diselenide (DMDSe) entrained through both concentrated nitric acid and hydrogen peroxide (H2O2) were compared by HPLC-ICP-MS and HPLC-HG-AFS analyses. It was demonstrated that trap reproducibility was better for nitric acid and a recovery of 65.2 +/- 1.9% for DMSe and 81.3 +/- 3.9% for DMDSe was found in nitric acid traps. HPLC-ES-MS identified dimethyl selenoxide (DMSeO) as the trapped product of DMSe. Methylseleninic acid (MSA) was identified to be the single product of DMDSe trapping. These oxidized derivatives have a high stability and low volatility, which makes nitric acid a highly attractive trapping liquid for volatile Se species and enables reconstruction of the speciation of those species. The presented trapping method is simple, quantifiable, reproducible, and robust and can potentially be applied to qualitatively and quantitatively study Se volatilization in a wide range of natural environments.

Grain unloading of arsenic species in rice

Rice (Oryza sativa) is the staple food for over half the world's population yet may represent a significant dietary source of inorganic arsenic (As), a nonthreshold, class 1 human carcinogen. Rice grain As is dominated by the inorganic species, and the organic species dimethylarsinic acid (DMA). To investigate how As species are unloaded into grain rice, panicles were excised during grain filling and hydroponically pulsed with arsenite, arsenate, glutathione-complexed As, or DMA. Total As concentrations in flag leaf, grain, and husk, were quantified by inductively coupled plasma mass spectroscopy and As speciation in the fresh grain was determined by x-ray absorption near-edge spectroscopy. The roles of phloem and xylem transport were investigated by applying a +/- stem-girdling treatment to a second set of panicles, limiting phloem transport to the grain in panicles pulsed with arsenite or DMA. The results demonstrate that DMA is translocated to the rice grain with over an order magnitude greater efficiency than inorganic species and is more mobile than arsenite in both the phloem and the xylem. Phloem transport accounted for 90% of arsenite, and 55% of DMA, transport to the grain. Synchrotron x-ray fluorescence mapping and fluorescence microtomography revealed marked differences in the pattern of As unloading into the grain between DMA and arsenite-challenged grain. Arsenite was retained in the ovular vascular trace and DMA dispersed throughout the external grain parts and into the endosperm. This study also demonstrates that DMA speciation is altered in planta, potentially through complexation with thiols.

Selenium Characterization in the Global Rice Supply Chain

For up to 1 billion people worldwide, insufficient dietary intake of selenium (Se) is a serious health constraint Cereals are the dominant Se source for those on low protein diets, as typified by the global malnourished population. With crop Se content constrained largely by underlying geology, regional soil Se variations are often mirrored by their locally grown staples. Despite this, the Se concentrations of much of the world's rice, the mainstay of so many, is poorly characterized, for both total Se content and Se speciation. In this study, 1092 samples of market sourced polished rice were obtained. The sampled rice encompassed dominant rice producing and exporting countries. Rice from the U.S. and India were found to be the most enriched, while mean average levels were lowest in Egyptian rice: similar to 32-fold less than their North American equivalents. By weighting country averages by contribution to either global production or export, modeled baseline values for both were produced. Based on a daily rice consumption of 300 g day(-1), around 75% of the grains from the production and export pools would fail to provide 70% of daily recommended Se intakes. Furthermore, Se localization and speciation characterization using X-ray fluorescence (mu-XRF) and X-ray absorption near edge structure (mu-XANES) techniques were investigated in a Se-rich sample. The results revealed that the large majority of Se in the endosperm was present in organic forms.

Geographical variation in total and inorganic arsenic content of polished (white) rice

An extensive data set of total arsenic analysis for 901 polished (white) grain samples, originating from 10 countries from 4 continents, was compiled. The samples represented the baseline (i.e., notspecifically collected from arsenic contaminated areas), and all were for market sale in major conurbations. Median total arsenic contents of rice varied 7-fold, with Egypt (0.04 mg/kg) and India (0.07 mg/kg) having the lowest arsenic content while the U.S. (0.25 mg/kg) and France (0.28 mg/kg) had the highest content. Global distribution of total arsenic in rice was modeled by weighting each country's arsenic distribution by that country's contribution to global production. A subset of 63 samples from Bangladesh, China, India, Italy, and the U.S. was analyzed for arsenic species. The relationship between inorganic arsenic contentversus total arsenic contentsignificantly differed among countries, with Bangladesh and India having the steepest slope in linear regression, and the U.S. having the shallowest slope. Using country-specific rice consumption data, daily intake of inorganic arsenic was estimated and the associated internal cancer risk was calculated using the U.S. Environmental Protection Agency (EPA) cancer slope. Median excess internal cancer risks posed by inorganic arsenic ranged 30-fold for the 5 countries examined, being 0.7 per 10,000 for Italians to 22 per 10,000 for Bangladeshis, when a 60 kg person was considered.

Occurrence and Partitioning of Cadmium, Arsenic and Lead in Mine Impacted Paddy Rice: Hunan, China

Paddy rice has been likened to nictiana sp in its ability to scavenge cadmium (Cd) from soil, whereas arsenic (As) accumulation is commonly an order of magnitude higher than in other cereal crops. In areas such as those found in parts of Hunan province in south central China, base-metal mining activities and rice farming coexist. Therefore there is a considerable likelihood that lead (Pb), in addition to Cd and As, will accumulate in rice grown in parts of this region above levels suitable for human consumption. To test this hypothesis, a widespread provincial survey of rice from mine spoilt paddies (n = 100), in addition to a follow-up market grain survey (n = 122) conducted in mine impacted areas was undertaken to determine the safety of local rice supply networks. Furthermore, a specific Cd, As, and Pb biogeochemical survey of paddy soil and rice was conducted within southern China, targeting sites impacted by mining of varying intensities to calibrate rice metal(loid) transfer models and transfer factors that can be used to predict tissue loading. Results revealed a number of highly significant correlations between shoot, husk, bran, and endosperm rice tissue fractions and that rice from mining areas was enriched in Cd, As, and Pb. Sixty-five, 50, and 34% of all the mine-impacted field rice was predicted to fail national food standards for Cd, As, and Pb, respectively. Although, not as elevated as the grains from the mine-impacted field survey, it was demonstrated that metal(loid) tainted rice was entering food supply chains intended for direct human consumption.

Cooking rice in a high water to rice ratio reduces inorganic arsenic content

Total arsenic and arsenic speciation was performed on different rice types (basmati, long-grain, polished ([white] and wholegrain [brown]) that had undergone various forms of cooking. The effect of rinse washing, low volume (2.5 : 1 water : rice) and high volume (6 : 1 water : rice) cooking, as well as steaming, were investigated. Rinse washing was effective at removing circa. 10% of the total and inorganic arsenic from basmati rice, but was less effective for other rice types. While steaming reduced total and inorganic arsenic rice content, it did not do so consistently across all rice types investigated. Low volume water cooking did not remove arsenic. High volume water : rice cooking did effectively remove both total and inorganic arsenic for the long-grain and basmati rice (parboiled was not investigated in high volume cooking water experiment), by 35% and 45% for total and inorganic arsenic content, respectively, compared to uncooked (raw) rice. To reduce arsenic content of cooked rice, specifically the inorganic component, rinse washing and high volume of cooking water are effective.

Inorganic arsenic in rice bran and its products are an order of magnitude higher than in bulk grain

Rice is more elevated in arsenic than all other grain crops tested to date, with whole grain (brown) rice having higher arsenic levels than polished (white). It is reported here that rice bran, both commercially purchased and specifically milled for this study, have levels of inorganic arsenic, a nonthreshold, class 1 carcinogen, reaching concentrations of approximately 1 mg/kg dry weight, around 10-20 fold higher than concentrations found in bulk grain. Although pure rice bran is used as a health food supplement, perhaps of more concern is rice bran solubles, which are marketed as a superfood and as a supplement to malnourished children in international aid programs. Five rice bran solubles products were tested, sourced from the United States and Japan, and were found to have 0.61-1.9 mg/kg inorganic arsenic. Manufactures recommend approximately 20 g servings of the rice bran solubles per day, which equates to a 0.012-0.038 mg intake of inorganic arsenic. There are no maximum concentration levels (MCLs) set for arsenic or its species in food stuffs. EU and U.S. water regulations, set at 0.01 mg/L total or inorganic arsenic, respectively, are based on the assumption that 1 L of water per day is consumed, i.e., 0.01 mg of arsenic/ day. At the manufacturers recommended rice bran solubles consumption rate, inorganic arsenic intake exceeds 0.01 mg/ day, remembering that rice bran solubles are targeted at malnourished children and that actual risk is based on mg kg(-1) day(-1) intake.

Growing rice aerobically markedly decreases arsenic accumulation

Arsenic (As) exposure from consumption of rice can be substantial, particularly for the population on a subsistence rice diet in South Asia. Paddy rice has a much enhanced As accumulation compared with other cereal crops, and practical measures are urgently needed to decrease As transfer from soil to grain. We investigated the dynamics of As speciation in the soil solution under both flooded and aerobic conditions and compared As accumulation in rice shoot and grain in a greenhouse experiment. Flooding of soil led to a rapid mobilization of As, mainly as arsenite, in the soil solution. Arsenic concentrations in the soil solution were 7-16 and 4-13 times higher under the flooded than under the aerobic conditions in the control without As addition and in the +As treatments (10 mg As kg(-1) as arsenite or arsenate), respectively. Arsenate was the main As species in the aerobic soil. Arsenic accumulation in rice shoots and grain was markedly increased under flooded conditions; grain As concentrations were 10-15-fold higher in flooded than in aerobically grown rice. With increasing total As concentrations in grain, the proportion of inorganic As decreased, while that of dimethylarsinic acid (DMA) increased. The concentration of inorganic As was 2.6-2.9 fold higher in the grain from the flooded treatment than in that from the aerobic treatment. The results demonstrate that a greatly increased bioavailability of As under the flooded conditions is the main reason for an enhanced As accumulation by flooded rice, and growing rice aerobically can dramatically decrease the As transfer from soil to grain.