Arsenic Oxidizing Microorganisms

Project Report, 2017
120 Pages, Grade: 5



1.1 Background of the Research

Arsenic (As) contamination of groundwater has remained a critical open issue in numerous world areas, especially in Uttar Pradesh in India, in light of its intense and ceaseless harmfulness and cancer-causing properties (Srivastava and Sharma 2013). In 1993, the World Health Organization set an arsenic permissible level of 10g/L in its drinking water rules (WHO, 2011). Practical procedures to expel arsenic from ground water must be produced to make water fit for drinking (Duan et al., 2013). Previous studies have shown that this region of India is characterized by extensive As pollution of rivers and ground waters by both geologic and anthropogenic activities. The various other states, including, Jharkhand and Bihar are at biggest risk, because of the flood plain of the Ganga River (Ahamed et al., 2006; Srivastava and Sharma 2013). In many foods a trace amount of arsenic (1g g-1 ) is essential for good health. However, the excess of this element (above permissible limit) causes cellular damage in biological system, like gastrointestinal and respiratory disorder. Skin, liver and bladder cancer (Kongkea et al., 2010; Shankar et al., 2014). Both inorganic arsenate As (V) and arsenite As (III) exist together in natural environment. Arsenite is more toxic than arsenate. As (III) frequently prevails as 74 to 98% of aggregate arsenic. Arsenic (III) is harder to evacuate than As (V). Utilizing ordinary methods, for example, precipitation, adsorption, particle trade and layer filtration and coagulation were used to remove arsenic from contaminated waters. Ahmed, 2003; Ghosh et al., 2014). All these methods are expansive and a source of pollution. Microbial As (III) oxidation has been viewed as an appealing option in light of its particular response for As (III) (Shivaji et al., 2006; Corsini et al., 2013; Uttiya et al., 2016). The biological mechanism of arsenic removal (arsenite oxidation) is possible because of presence of arsenite oxidase and transporter genes studied in various microorganisms like Alkalilimnicola ehrlichii strain MLHE-1 (Zargar et al., 2010). Alcaligenes faecalis (Ellis et al., 2001). Rhizobium sp. strain NT-26 and Hydrogenophaga sp. strain NT- 14 (Santini and vanden Hoven 2004). Different heterotrophic and chemolithoautotrophic (CAOs) As (III) oxidizing microscopic organisms have been separated from distinctive situations. Chemolithoautotrophic As (III) oxidizing microorganisms (CAOs) can use As (III) as an electron benefactor and As (III) oxidation can bolster their development, while the oxidation of As (III) by heterotrophic As (III) oxidizing microbes (HAOs) is by and large thought to be a detoxification instrument. Since CAOs by and large demonstrate a higher particular As (III) oxidizing rate than HAOs (Hamamura et al., 2013). CAOs are more appealing in the light of common sense remediated applications. The confirmation that CAOs can likewise become under heterotrophic conditions is another point of preference of the use of CAOs for arsenic bioremediation. Nonetheless, most As (III) oxidizing microorganisms disengaged so far were heterotrophs, and just restricted quantities of CAOs have been disconnected and portrayed on their As (III) oxidation capacity in point of interest. Also, few reports have portrayed arsenic evacuation utilizing CAOs. Thus, more studies must be embraced to set up a bioremediation technique for arsenic-contaminated groundwater utilizing CAOs (Jean et al., 2014; Corsini et al., 2014).

1.2 Aims and Objectives of the Research

The main aim of the research is the Isolation and Characterization of Arsenite Oxidizing Bacteria from Arsenic Contaminated Ground & Surface water in five Districts of Uttar Pradesh, India. This will be achieved through following objectives:

- To examine the arsenic contamination of surface and ground water.
- Isolation and Molecular characterization of arsenite oxidizing bacteria.

1.3 Research Purpose

Arsenic (As), a known human cancer-causing agent, is circulated broadly in water, soil, and air. In larger part of the cases, As sullying of groundwater is gotten normally from As-rich aquifer residue. Common groundwater containing As utilized for drinking, watering system, or agriculture purposes causes genuine wellbeing issues for people (Hamamura et al., 2014). Consequently, the World Health Organization has decreased the worthy edge for As in drinking water from 50 to 10 μg/l. Nations have embraced higher values as national measures, for example, 7 μg/l for Australia and New Zealand and 50 μg/l for India. Therefore, research on improving or developing effective treatment technologies like biological oxidation of arsenite for removing As from contaminated waters is becoming more important. On the basis of these facts, this study is conducted in five Districts of Uttar Pradesh, India. To isolate and characterize the Arsenite Oxidizing Bacteria from Arsenic Contaminated Ground & Surface water.


2.0 Overview of Arsenic

Arsenic (As) is a semi-metal (non-metal) that is found naturally in the earth's crust and is present in over 200 minerals (Jones, 2007). It is obtained as a by-product of the treatment of copper, lead, cobalt and gold ores. The only stable isotope of Arsenic discovered is As 75. It is 47th most abundant element among the 88 elements which naturally occur in the world. The average crustal abundance of arsenic is 1.5 mg kg -1 (1.5 ppm). Shales, Slates and Mudstones have the highest concentrations among the common rocks, although extremely high concentrations can be found in some coals (Kao et al., 2013). The concentration of Arsenic is generally very low in natural rain water and sea water as compared to river water. It has sometimes very high quantity of Arsenic. The two most important natural sources of arsenic are:

- Regions with high concentrations of As-Inorganic in the water
- Soil containing arsenic mines; with metal intake in the form of particles (Bahar et al., 2013).

The two main natural sources of arsenic are volatilization releasing, approximately 26,000 tonnes of arsenic per year and volcanic activity, releasing to the atmosphere around 17,000 tonnes for the year (Smedley et al., 2002). The element is found in hundreds of minerals, of which 60% are arsenates, 20 % are arsenosulfetos with metals such as Fe, Pb, Cu, Ag and Ti and the remainder consists of arsenites, oxides, elemental arsenic and arsenides. The most common mineral is arsenopyrite (FeAsS). The earth's crust, rocks and sediments have differentiated arsenic levels in their composition, but most of the arsenic is associated with pyrite.

The processes rocks refining, including the merger of copper and lead are the biggest sources of release of arsenic. In the past, arsenic compounds were widely used in agriculture and forestry as insecticide or herbicide, especially in cotton crops. Currently, the inorganic compounds may be used in agriculture. Disodium and monosodium metilarsenato - are still used as pesticides. In agriculture, arsenic also has an application to increase its resistance to decay. In the field of veterinary medicine, arsenic is used as an antiparasiticagent and as an additive in animal feed to accelerate growth (Achal et al., 2012). Elemental arsenic is used in the production of glass and as an additive in the manufacture of non-ferrous alloys. The arsine gas (AsH3) is used in the microelectronics industry, and semiconductors (Andres et al., 2013). Anthropogenic sources of Arsenic are industrial wastes including mining, smelting (gold, lead, copper and nickel), iron and steel production, and coal combustion (Khianngam et al., 2014). Leaching of abandoned mines of gold, decades and centuries ago, are still significant source of arsenic pollution in aquatic systems. Arsenic is released into the atmosphere by natural phenomena and anthropogenic sources in ratio of 60:40, and returns to the earth's surface through dry deposition or wet (Bachate et al., 2013). Arsenic is a very famous poisonous element. Today, when human is more intelligent and sophisticated, he tries to eliminate all the factors that may affect the life of human beings or try to eliminate all the factors that serve as a source which may affect environment of human being negatively (Jean et al., 2014). The reason of doing such efforts is only to increase the life expectancy and quality of living of peoples especially living in areas that are located near the industrial zones. Industrial zones are considered as an important source that contaminates the world with this poisonous element.

2.1.1 Chemical forms of Arsenic

The metallic arsenic is used in the production of non-ferrous alloys and other compounds for the manufacture of semiconductors, diodes including light emission, lasers, integrated circuits and solar cells. Arsenic trioxide acid and arsenic are used for bleaching and dispersing air bubbles in production of glass bottles and glassware (Das et al., 2013).

Calcium arsenate (Ca3 (AsO4) 2), calcium arsenite (Ca (AsO2) 2), arsenic trioxide and arsenic acid (H3AsO4) are soluble in water, where as sodium arsenate (Na3AsO4) and sodium arsenite (NaAsO2),- have higher solubility in water. The arsenic trisulfide (As2S3) is practically insoluble in water. When heated it undergoes decomposition, emitting toxic fumes. The arsine (AsH3) is a colourless gas with garlic odour and slightly soluble in water, alcohol and alkali (Datta, 2015).

2.1.2 Arsenic Oxides

Arsenic occurs in the environment in four oxidation states: As (III)-, As (0), As (III)+, and As (V). Arsenic common species found in most of the natural waters are the trivalent arsenite, inorganic oxyanions and pentavalent arsenate (Lami et al., 2013). Trivalent arsenite includes H3AsO3, H2AsO3 - and HAsO32 - while, pentavalent arsenate includes H2AsO4, HAsO4 and H3AsO4. In addition, there are a variety of organic forms of arsenic such as methane arsonic acid and dimethyl arsenic acid. As (III) is the more toxic form of the species. The main abiotic factors that control the speciation of arsenic are redox potential (Eh) and pH (Vink, 1996).

2.2.0 Phases of Arsenic

2.2.1 Solution Phase

Two states of arsenic which are oxidized, As (III) and As envy, prevail in near surface and surface environment. As for solution, arsenic exists basically as acid of oxy anionic; arsenite has 2.2 pKa’s, 11.5 and 6.9 while arsenite has 9.2 pKa’s 13.4 and 12.1. Therefore, at pH circumnetural, H2AsO4, H3AsO3 and HAsO4 are dominating species. Microbial and plant activity may methylate As (III) or As (envy) forming MMAA (monomethylarsonous acid) and DMAA (dimethylarsenic acid) (Petrick et al., 2000; Megharaj et al., 2012).

2.2.2 Precipitation of Arsenic Phases

Both As (III) and As (envy) may precipitate within soils and sediments. Under acidic and alkaline conditions Arsenic precipitate with hard cations and multivalent ions just like phosphate Sulphates may also be replaced by Arsenate or Phosphatein minerals due to charge and smaller size characteristics, many alkali earth metals and heavy metals also have the capability for precipitating with arsenate (Osborne et al., 2015).

2.3 Arsenic Toxicity

In natural water, arsenic is present mainly in the form of inorganic compounds, which has the valencies 3+ and 5+ (Islam et al., 2013: Hassan et al., 2015). It has been established that compounds of As (III) + may be up to 10 times more toxic than the As (V)+ (Pous et al., 2015).

The toxicity of various species of arsenic decreases in the following order: inorganic compounds As3+ the compounds of inorganic 5+ the organic compounds 3+ the 5+ organic compounds. Inorganic arsenic is considered carcinogenic.

Arsenic is absorbed by the human body mainly by inhalation. Trivalent inorganic arsenic As (III) +, interacts strongly with group’s sulfhydryl organic molecules. Several enzymes are affected with it, causing damage in several cell systems. A dose of 140 milligrams of trivalent inorganic arsenic is enough to cause the death of an adult human. It damages the cellular respiration in few hours or days (Paul et al., 2014). The arsenic can induce the production of metallothionein a protein which binds to the metal and also cadmium, mercury and many essential metals. It is assumed that this is one of the mechanisms of adaptation that leads to relative tolerance to the toxicity of arsenic in multicellular organisms (Petrick et al., 2000).

In many foods a trace amount of this element is essential for good health. However, the excess of this element causes skin and liver cancer and perhaps cancer of bladder and kidneys (Wang, et al., 2012; Davolos and Pietrangeli, 2013). Most of the compounds, organic or inorganic arsenic, are white powder having no odour or special flavour. Since arsenic is found naturally in the environment, poisoning occurs through the industrial occupation, the food intake or drink contaminated by breathing, by skin contact with water or soil containing arsenic or due to intentional acts (Zhu et al., 2014). Arsenite is considered the most soluble, mobile and more toxic species than arsenate compounds. The toxicity is directly related to the mobility in water and in biological fluids. The toxicity is in thefollowing descending order: arsine inorganic arsenates compounds organic trivalent inorganic arsenatespentavalent compounds Organic compounds arsonicos the elementary (Yamamura et al., 2013).

The species arsenobetaína, arsenocolina are considered non-toxic. The effects caused by exposure to arsenic depend upon the species, dose, duration and route of exposure. Other factors to consider are age, sex, family habits and health status of the person exposed. The arsenic in food occurs as a mixture of species inorganic and organic, including arsenobetaína. Generally the oganic compounds represent 60-99% of total arsenic in food (Figure 21). Studies indicate that in aquatic species, the inorganic arsenic levels are, usually less than 1%, but in other foods such as meat, poultry and cereals, inorganic arsenic content is increased (Yamamura and Amachi, 2014). The food containing high organic arsenic are fish and crustaceans, usually in the form of arsenobetaína (Petrick et al., 2000).

In food, a limit considered tolerable to arsenic consumption is of the order of 1 ug g -1 as arsenic trioxide. When being exposed to oral doses between 3 and 30 mg per kg of the body mass, the most apparent symptoms are stomach or intestinal irritation, causing pain, nausea, vomiting and diarrhea. Moreover, contamination by inorganic arsenic may result in decreased production of red and white cells blood causing fatigue, changes in heart beat or nerve function (Tsai et al., 2003).

When it comes to oral presentations for prolonged periods, inorganic arsenic poisoning can cause skin changes, such as darkening and appearance of small warts or calluses on the palm of hand, foot, or dorsum plant, which are often associated with changes in blood vessels in the skin (Abhyankar et al., 2012). As some of these calli may result in cancer, the inorganic arsenic is considered carcinogenic to humans by various public health institutions. Inhalation of low concentrations of inorganic arsenic for long periods can produce effects on the skin and circulatory problems. Exposure to high concentrations of arsenic can cause neck pain and irritation in the lungs, increasing considerably the risk of lung cancer. Besides, there is data to suggest that inorganic arsenic inhalation may interfere with the fetal development. However, through animal studies, it was found that the exposure to high concentrations can lead to the appearance of certain symptoms, such as stomach irritation and nerve problems (Xie et al., 2014). Arsenic compounds appear to be oxidized in vivo from trivalent to pentavalent; however, the opposite does not occur. In animals, arsenic poisoning at low concentrations, may cause inflammation of the membranes of the upper respiratory tract mucosa, diarrhoea, eczema and lack of motor coordination. However, arsenic poisoning in high concentrations causes the death of the animal within a few days (Jack 2005; Wu et al., 2014). Symptoms of inorganic arsenic poisoning vary with the amount and form of administration and include abdominal cramping, vomiting, diarrhoea, strong depression and dermatitis, usually due to increased capillary permeability and cell necrosis. In horses, the signs in acute cases of poisoning include brain trauma, caused by successive beating of his head against the wall and signs of severe pain. However, animals can survive a single oral dose. In this case, nearly all the arsenic compounds administered are excreted within a few days (Vishnoi et al., 2014). The use of compounds with inorganic arsenic was banned as a pesticide. The organic compound in use containing arsenic includes Cacodylic acid, disodium metilarsenato (DSMA) and metilarsenato monosodium (MSMA). These compounds also are used as additives in animal feed (Petrick et al., 2000; Poirel et al., 2013). Once ingested, arsenic are consumed by the gastrointestinal tract and retained after the mixes are disseminated to organs and tissues. In liver cells, methylation procedure may lead to the creation of two metabolites, methylarsonic corrosive (MEAs) and corrosive dimethylarsinic (ADMA).

In drinking water arsenic present as arsenite it is effectively absorbed by the gastrointestinal tract. The impact of continual exposure to arsenic, may lead to the following (Tsai et al., 2003; Jack 2005; Kongkea et al., 2010).

- Consumption of arsenic contaminated drinking water increases the risk of lung, kidney or bladder cancer, and skin lesions, the latter being the first symptom of chronic exposure, while cancer can take years to appear.  There is a very clear correlation between exposure and health effects. The various symptoms include hypertension, cardiovascular disease, diabetes, including effects on reproduction, all associated with chronic exposure Arsenic.

- The disease known as "black foot", is typical of chronic arsenic exposure. This disease affects the circulatory system, resulting in gangrene.

- Signs and symptoms differ between individuals and geographic areas, therefore, there is no universal description this fact is a problem in weighing the influence of arsenic on health. In the same way, there is no method to distinguish between a cancer caused by chronic arsenic exposure and cancer caused by other factors.

2.4 Geogenic Arsenic Contamination

Surface water and groundwater are two main sources of drinking water. River, lakes, and reservoirs are the main source of surface water and aquifers are the source of groundwater. Such water can be contaminated with improperly disposed chemicals, animal wastes, pesticides, human waste, and naturally-occurring substances (Corsini et al., 2013). To treat the water contaminants, a centralized water monitoring system is used in many parts of India like Uttar Pradesh, Chandigarh, Delhi and Bihar, etc. These Indian states like Bihar, Uttar Pradesh, Delhi and Chandigarh etc., lack modern water quality monitoring facilities (Rahman et al., 2014). A cost-effective removal process is needed in developing countries that lack centralized treatment facilities to fight against the deadly contaminants. Throughout the deltaic regions of Southeast Asia the primary source of drinking water is groundwater because surface waters contain high levels of human pathogens due to runoff from agricultural fields and household waste. In fact, since the 1960's residents in Uttar Pradesh have drilled shallow tube wells in order to access an alternative, presumably safer, drinking water source (Xie, et al., 2014).

Unfortunately, residents began to develop a variety of ailments, including skin lesions, which were eventually traced to arsenic (As) poisoning. According to the World Health Organization regulatory guideline, more than a 100 million people in Uttar Pradesh and West Bengal consume groundwater containing unsafe levels of As (10 µg/l) (Seth, 2014). The regular consumption of unsafe levels of As causes serious illnesses such as lung, bladder and skin cancer, vascular disease, diabetes, and negatively affects the intellectual development of children (Wang et al., 2012). Therefore, understanding the processes that promote As mobilization in groundwater aquifers is vital for managing water supplies and protecting human health (Rahman et al., 2014).

2.5 The origin of arsenic in Southeast Asia

The As within the shallow (<100 m) unconfined aquifers of Southeast Asia originates from As- rich metal-sulphide minerals in the Himalayas, such as arsenopyrite, which were weathered and transported through fluvial networks during the Holocene glacial retreat. The weathered material has been deposited within the Ganges-Meghna-Brahmaputra Delta Plain (GMBD), and Macedonia (MKD) throughout the millennia, and today, the As is sorbed to iron (Fe)-oxide minerals which coat deltaic aquifer sediments (Polizzotto et al., 2008; Shrivastava et al., 2015). Research throughout the past decade has demonstrated that a suite of hydrologic and biogeochemical processes contributes to the release of As from Feoxide mineral surfaces in high As aquifers (Smedley and Kinniburgh, 2002; Corsini et al., 2013). In general, the scientific research on groundwater As mobilization has been conducted in the GMBD since geogenic As contamination in Southeast Asia was first recognized (Vishnoi et al., 2014).

2.6 Geochemistry and Mobilization of Arsenic in India

The presence of arsenic in groundwater has been accounted for from numerous parts of the world, especially in the Uttar Pradesh of India. Arsenic pollution in India is very much recorded particularly in Ganga-Meghna- Brahmaputra plain (Majumder et al., 2013). The WHO rule and additionally the Bureau of Indian Standard (BIS) passable breaking points for arsenic in drinking water is 10 μg/L. An interval standard of 50 μg/L is accepted by the Ministry of Rural Development (MHRD) in India (Singh et al., 2014; Lami et al., 2013).

Activation of arsenic happens in nature through a blend of few unpredictable characteristic procedures. Despite the fact that there are a few reports on different activation courses of arsenic to the earth, the absolute most broadly acknowledged speculations are definite underneath (Leleyter, et al., 1999; Majumder et al., 2013).  Reductive disintegration: In this mechanism, the decrease of As-rich Fe oxides, because of covered peat and other natural stores, prompts arsenic discharge to the aquifer.

- Alkali desorption: At pH 8, the retention of arsenic from metal oxides, particularly Fe and Mn, can prompt a high arsenic fixation in the groundwater.

- Sulphide oxidation: Oxidation of arsenical pyrite in the alluvial silt as aquifer draw down licenses environmental oxygen to attack the aquifer. This mechanism is normally alluded to as the "oxidation theory."  Geothermal action: Blending of geothermal arrangements and crisp groundwater can prompt high arsenic fixations in few areas.

In the condition of Uttar Pradesh, India, couple of examinations have been completed on the conveyance of arsenic sullying. Arsenic assembly mechanism may change with the area relying upon hydrogeological conditions (Figure 1). Subsequently, it is vital to study the relationship of arsenic with different species including nitrate, sulphate, iron, bicarbonate, phosphate, and so forth, for evaluating the imaginable mechanism of arsenic (Leleyter et al., 1999; Mukhopadhyay et al., 2002).

Abbildung in dieser Leseprobe nicht enthalten

Figure-1: The Geocycle of Arsenic (Mukhopadhyay et al., 2002).

2.6.1 Hydrologic Processes Influencing Groundwater Arsenic Mobilization

The deltaic aquifers in Southeast Asia are recharged from regional and local sources. Regional groundwater recharge rates are primarily influenced by topography, and there is very little relief in the regions surrounding these large river deltas. For example, hydraulic gradients within the recharge zone of the GMBD is ~10-4 cm/yr and thus regional recharge rates are relatively slow, with estimates ranging from 400-1800 cm/yr. Local recharge rates vary between 0.05 to 1m/yr within the GMBD, and the high degree of variability is likely due to differing rates of groundwater extraction (Shakya et al., 2012).

The primary local sources of groundwater recharge are adjacent rivers, man- made ponds, and seasonal floods from monsoonal rains. In general, As concentrations in surface water recharge sources, which are oxygenated, are below 10 µg/l. However, there is a strong relationship between groundwater age and groundwater As concentrations, suggesting that there is an accumulation of As with lateral groundwater flow, and a transport of As-laden groundwater to depth (Stute et al., 2008) It appears that the factors influencing lateral and vertical flow and recharge rates, such as sediment grain size and human activities such as irrigation pumping, are also related to groundwater As concentrations in the Southeast Asia deltaic aquifers (Shakya et al., 2012). For example, groundwater flow rates are slower in fine-grained, salty sediments, which are less permeable than sandy sediments, and pore water As concentrations are also higher within fine-grained sediments (Seth, 2014). By contrast, groundwater pumping for irrigation increases the rate of vertical groundwater flow, and promotes the flow of organic-rich pond water into the aquifer, which may increase rates of groundwater As mobilization. Research suggests that patchiness in sediment grain size, and hydrologic perturbations due to irrigation pumping, are partially responsible for the considerable spatial heterogeneity in groundwater As concentrations observed in high As aquifers in Southeast Asia (Acharya et al., 1999; Abedin et al., 2002; Ahmed et al., 2004).

2.6.2 Fractionation of Arsenic Implications for Mobility

Sequential extraction schemes to identify concentrations of As associated with various chemical fractions in whole soil systems have been used by numerous researchers. Limitations of sequential extraction techniques have been noted by some researchers, including the lack of selectivity and efficiency of the various steps in the procedures, and the possible re-adsorption of extracted elements (Leleyter and Probst 1999; Ahnstrom and Parker 1999). Arsenic associated with Fe oxides have been extracted using solutions containing NH4 oxalate because it dissolves Fe oxides (Krysiak and Karczewska, 2007). Hydrogen peroxide and NaOCl have been used to extract metals associated with organic matter because both chemicals strongly oxidize organic matter (Ahnstrom and Parker, 1999). Strong acid digestions involving hydrofluoric acid (HF), nitric acid (HNO3), and/or hydrochloric acid (HCl) have been used to extract metals from the residual fraction as they provide rapid and reproducible recovery of many elements (Ahnstrom and Parker, 1999). Concentrations of As considered dissolved, soluble, or labile generally accounted for a small fraction of total extractable metals (Girling, 1978; Laren et al., 1998; Casado et al., 2007; Krysiak and Karczewska, 2007 and Baeza et al., 2010). Less than 10% of arsenic is freely exchangeable, and similarly small proportions are readily labile (Acharya et al., 1999) The water soluble fractions of As are generally considered as the mobile fraction of Sb in contaminated soils and sediments (Casado et al., 2007; Baeza et al., 2010). The highest fractions of As were typically associated with Fe, Al, or Mn oxides. The majority of As and about half of the Sb was associated with extractable Fe and Al compounds, while in contaminated samples, less than 20% of As was associated with these fractions. Laren et al., (1998) found that the largest proportion of As (about 60% of total As) was in the Fe and Al oxide-associated fraction. About 23 to 96% of total As was associated with hydrous Fe and Al oxides (Krysiak and Karczewska, 2007). It is inferred that the variation in the percent association of As with metal oxides is due to differing metal content and chemical conditions, such as soil pH. pH is a controlling variable in As sorption/desorption to metal oxides (Masscheleyn et al., 1991). Additionally, redox conditions can cause desorption or reductive dissolution of the oxyhydroxide soil phase (Krysiak and Karczewska, 2007). Thus, As may become mobile under appropriate pH and redox conditions although As associated with crystalline or non-crystalline oxides and hydroxides are often assumed to be immobile (Krysiak and Karczewska, 2007; Baeza et al., 2010 and Wilson et al., 2010).

2.6.3 Organic Matter Associations

Organic matter-associated fractions of As are also relatively low. Crecelius et al.,(1975) found that less than 10% of As was associated with the oxidizable organic matter in the sediment where as Baeza et al., (2010) reported 6.2 to 11.4% of As bound to organic matter. Sorption/desorption and specific conditions of water chemistry can affect the fractions of As bound to organic matter (Baeza et al., 2010). Thus, fractions of As associated with organic matter in soils and sediments may become mobile under appropriate conditions of pH and ionic strength. Significant, and sometimes the highest, concentrations of As were found in the residual or recalcitrant fractions. Fractions of As associated with residual or recalcitrant phases in sediments and soils are widely considered to be stable (Tighe and Lockwood, 2007; Wilson et al., 2010). Though fractions of As associated with this phase may be high in some studies, it has been noted that the relative proportions of As in this phase are dependent on the As source (Tighe et al., 2007; Wilson et al., 2010). Tailing samples have shown higher residual fractions of As than more mineralized samples (Wilson et al., 2010).

2.6.4 Release of Arsenic from Sediments and Soils

Mok and Wai (1990) studied the release of As from contaminated sediment taken from the Coeur d' Alene River in northern Idaho. In these leaching experiments, 60 g of sediment was equilibrated with 800 mL of distilled deionized water, and samples were taken over time for about 12 days. Researchers indicated that pH varied little over the time scale, with an average pH around 6.9. The predominant As species released was As (III). As release occurred mainly in the initial leaching period (24 hours) and appeared to approach asymptotic limits (around 175 ng As/g of sediment). Wilson et al., (2010) performed three experiments to test the short-term mobility of As. They tested As dissolution from solid samples grey (unoxidized) sand, brown (oxidized) sand, and coarse tailings collected at a historic antimony smelter site located in New Zealand. As release from the unoxidized sand tailings was initially 411 mg/kg, with released concentration increasing over time to 800 mg/kg As.

2.6.5 Effect of Water Chemistry on As Release from Streambeds

Effect of pH on As Release from Sediments pH has a significant effect on anion solubility (Masscheleyn et al., 1991; Smedley, and Edmunds, 2002). As is soluble at low and high pH values, which is typical of anions in aqueous solutions (Mok and Wai, 1990). Arsenic in ground water exists in multiple valencies. In oxidizing environment the arsenic acid will be the dominant form with a valency state of As (V). while as in reducing environments the dominant form is arsenious acid with a valency of As (III). Various authors have studied the effect of pH on arsenic mobilization from contaminated waters and soils (Mok and wai 1990; Masscheleyn et al., 1991; Casado et al., 2007; Baeza et al., 2010). In 2-11 pH range the release of arsenic from sediments to overlying water follows a pattern of substantial release with decreasing pH with a sharp increase in release of asrsenic also noted at high pH (Mok and wai 1994).

2.6.6 Effect of Ionic Strength on As Release from Sediments

The effect of ionic strength on anion desorption from sediment is dependent on the point of zero charge of the sediment and whether the anion is adsorbed through a specific (inner sphere complexing) or non-specific adsorption mechanism (outer-sphere complexing) (Wilson et al., 2010). Some studies have noted that ionic strength has less effect on specific adsorption, or inner-sphere complexing (Tighe and Lockwood 2007). When considering outer-sphere complexing, increased ionic strength decreases negative surface charge at high pH thus increasing adsorption, while adsorption is decreased below the point of zero charge because increased ionic strength decreases electrostatic potential at the solid surface (Wilson et al., 2010). Based on this logic, it is inferred that at increased ionic strength and high pH, desorption would be low, but below the point of zero charge, desorption would be increased.

2.6.7 Effect of Redox Conditions on Arsenic Release from Sediments

As is highly sensitive to oxidation-reduction (redox) conditions (Masscheleyn et al., 1991), and as such, redox conditions in the hyporheic zone are expected to have a significant effect on As mobility in river systems. Redox conditions control the oxidation state of both As. Subsequently, the oxidation state of both metals controls their reactivity, toxicity, mobility, and transport. In soils, As (V) and As (III) are the primary oxidation states of As. At most environmentally relevant pH values (4-9), H2AsO4 - and HAsO42 - are the predominant As forms in the As-water system. The most predominant As (III) species is the neutral species As(OH)3. (Masscheleyn et al., 1991) studied the effect of redox potential on arsenic speciation and solubility in contaminated soil collected from an abandoned As dipping vat near Kolin, Louisiana. Batch experiments were conducted under varying pH-redox conditions, and with approximately 200 g of contaminated soil and de- ionized distilled water at a soil:water ratio of 1:6. Redox conditions investigated were -200, 0, 200, and 500 mV. Samples were collected for 24 days. Results showed that As(V) was the major dissolved As species at redox potentials of 200 and 500 mV, and accounted for more than 95% of total soluble As. Under reduced conditions of redox potentials at 0 and -200 mV, As (III) was the dominant dissolved As species. It is predicted that As will exist as As (V) under oxidizing conditions. When conditions are moderately reducing and across the pH scale, As will exist as the neutral species As(OH)3. When Fe is considered in the system, As is expected to co- precipitate with Fe oxy-hydroxides (Mok and Wai, 1990; Wilson et al., 2010). The Fe sulfides will scavenge As at lower As concentrations (Wilson et al., 2010).

2.7 Arsenic mobilization

A suite of biogeochemical processes promotes groundwater As mobilization in the deltaic aquifers in India. The current paradigm is that the native Fe-reducing bacteria in aquifer sediments cause the reductive dissolution of Fe-oxide minerals, which results in the release of sorbed As under anaerobic, reducing conditions (Bang and Meng, 2004; Rowland et al., 2007). In addition, research throughout the past decade has demonstrated that the rates of bacterial Fe-reduction and groundwater As mobilization are largely influenced by the availability of organic matter (Drewniak, and Sklodowska 2013). First, heterotrophic bacteria in the near-surface depths of the aquifer use organic matter sources for carbon and nitrogen to synthesize biomass and for electron donors during the respiratory reduction of oxygen (Stolz and Oremland 1999). Heterotrophic activity rapidly consumes oxygen and results in anaerobic, reducing conditions in the saturated zone of the aquifer. Under reducing conditions, Fe-reducing bacteria pair the oxidation of labile fractions of dissolved organic matter (DOM), such as low-molecular weight organic acids, to the reduction of Fe (III) within Fe-oxide minerals. Also, it is likely that native bacteria utilize labile DOM as an electron donor during dissimilatory As (V)-reduction in high As aquifer in Southeast Asia (Valenzuela et al., 2015). More recently, research has indicated that bacteria may use more recalcitrant fractions of DOM while mediating groundwater As cycling (Drewniak, and Sklodowska, 2013). In anaerobic aquatic environments, a diverse group of bacteria generate energy by pairing the oxidation of labile DOM to the reduction of redox-active Quinone moieties within humic substances. It is likely that Quinone-reduction is phylogenetically widespread amongst bacteria in neutral pH and anaerobic aquatic sediments because other electron acceptors are insoluble, and Quinone-reduction may yield more energy than the reduction of metal-oxide minerals (Bang and Meng 2004; Lafferty and Loeppert, 2005). Bacterial Quinone reduction promotes an electron cascade in which reduced Quinone moieties abiotically transfer electrons to Fe(III), and cause the reductive dissolution of Fe-oxide minerals. Actually, evidence from laboratory experiments suggest that abiotic Fe (III)-reduction via reduced Quinone’s is significantly faster than bacterial Fe-reduction reduction (Jiang & Kappler, 2008). Thus, in the high As aquifers of Southeast Asia, bacterial Quinone reduction could be responsible for accelerating rates of Fe-oxide dissolution and subsequently the release of As into groundwater (Zeng, 2004). In fact, reduced Quinone moieties have been discovered in high As groundwater in the GMBD.

2.8 The Microbial Ecology of High Arsenic Aquifers

The presence of dissolved organic matter (DOM) influence the activity of bacterial communities in aquifers contaminated with arsenic (Ghosh and Sar, 2013). Previous studies have shown that the bacterial communities in aquifer sediments are diverse, for example, Beta Proteobacteria were the dominant members of sediments collected from aquifers in the Ganges-Meghna-Brahmaputra delta plain (GMBD), while Gamma proteobacteria, Alpha proteobacteria, Bacteriodetes, and Firmicutes were also relatively abundant bacterial communities from a number of aquifers affected by geogenic As contamination in Southeast Asia (Corsini et al., 2014). Delta proteobacteria are less abundant as compared to other taxa (Ghosh et al., 2014). Geobacter genus is known as Fe- and As-reducers (Yamamura et al., 2013) and genus Pseudomonas in Gamma Proteobacteria (Vicente et al., 1990) Thus, availability of DOM influences the abundance of bacterial taxa such as Delta proteobacteria, in addition to other subphyla within the Proteobacteria that may promote groundwater As mobilization in high As aquifers (Guo et al., 2015).

2.8.1 Arsenite oxidizing microorganisms

Arsenic As (III) oxidation by microorganisms is a potential detoxification process that allows microorganism to tolerate higher As (III) levels (Cai et al., 2009). As (III) may serve as an electron donor for microbial respiration in combination with O2 under anoxic and oxic environments (Cai et al., 2009; Luo et al., 2013; Dong et al., 2014). Microorganisms, Alcaligenes faecalis, Hydrogenophaga sp., A. ferrooxidans, T. aquaticus, T. thermophiles are known for arsenite oxidation (Gihring et al., 2001; Oremland and Stolz, 2004). Various genera responsible for arsenite oxidation and reduction are listed in (Table 1).

Table-1: Arsenite Oxidizing and reducing bacteria

Abbildung in dieser Leseprobe nicht enthalten

The transformation of As (III) to As (V) using microorganisms is to reduce As mobility in the environment as the affinity of As (V) to mineral solids is usually higher than that of As(III) (Smedley and Kinniburgh, 2002). As (III) removal from arsenic contaminated waters using bacteria (microbial oxidation) has been proposed by various researchers (Inskeep et al., 2007; Ito et al., 2012; Cavalca et al., 2013; Duan et al., 2013). Immobilization of arsenic can be enhanced by simultaneous microbial oxidation of Fe (II) and As (III) and is considered as a potential bioremediation alternative for arsenic in anoxic environments. Formation of Fe (III) hydroxides, which adsorb As (V) formed from oxidation of As (III) (Sun, 2004). So far not much work has been done to remove arsenic from contaminated waters by biological methods. Microorganisms such as Thiobacillus ferrooxidans, Lepto spirillum ferrooxdans (Das et al., 2013). and fungi like Scopulariopsis previcaulus, Penicillum sp., Gliocladium roseum, and yeast Candida humicola (Pokhrel and Viraraghavan, 2006) haven been shown to oxidize As (III) to less toxic form As (V). Iron oxidation by microorganisms such as Gallionella ferruginea and Leptothrixo chracea have been effectively used for the removal of arsenic from waters by As (III) oxidation. It has been shown that it increases the removal of arsenic up to 95% from 200 μg/L As-contaminated water. The As (V) is also removed by same way as mentioned above and hence bring the limit of As with in the permissible limits (Katsoyiannis and Zouboulis, 2004). Methylation by certain microorganisms like bacteria, Archea and fungi is an effective strategy of arsenic detoxification. The production of monomethylarsonic (MMA) or acid of dimethylarsinic (DMA) react with arsenic and makes it less toxic (Paul et al., 2014).The soil composition, pH and redox environments play an important role in the oxidation of As (III) to As (V) and vice versa. The microbes and pH of the environment affect the arsenic speciation. The exact mechanism of arsenite oxidation by microbes for removal of arsenic from contaminated waters is not still well understood. The study of 16S rDNA and chromosomal ars gene using PCR will be helpful for confirming the ars gene sequences in microbes. In Pseudomonas aeruginosa the ars operon detoxification possess three ORFs encode proteins which are very similar to the ars operon of E.coli (arsR, arsC and arsB). arsB gene sequence encode the presence of efflux pump system. The molecular geomicrobiology of arsenite-oxidizing bacteria can be applied for bioremediation of arsenic contaminated waters. The chromosomal ars operon homolog in Pseudomonas aeruginosa was not present in different strains of Pseudomonas for example, ars operon in Pseudomonas putida, was not always the same as of Pseudomonas aeruginosa sp. even though the strains belong to same genera. Therefore, versatility should be taken into consideration when ars operon are observed when using PCR.


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Arsenic Oxidizing Microorganisms
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Arsenic contamination, groundwater, chemics, Microorganism, Oxidizing
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PhD Bilal Ahmad Tantry (Author), 2017, Arsenic Oxidizing Microorganisms, Munich, GRIN Verlag,


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