Introduction

Organochlorine pesticides, including aldrin, dieldrin, endrin, dichloro-diphenyl-trichloroethane (DDT), endosulfan, heptachlor, hexachlorocyclohexanes (HCHs), mirex, and others, are synthetic chemicals, some of which have been used extensively for controlling insect and mite vectors of human and domestic animal diseases and against insect pests that damage agricultural crops. Organochlorine pesticides are bioaccumulative, toxic, and persistent in the environment.

Endosulfan is a broad-spectrum chlorinated cyclodiene insecticide that has been used extensively, on a variety of crops, for over 30 years (Sutherland et al. 2000). It is a mixture of two stereoisomers, α- and β-endosulfan, at a ratio of 7:3. It is extremely toxic to fish and invertebrates and it has increasingly been implicated in mammalian gonadal toxicity, genotoxicity, neurotoxicity (Siddique et al. 2003), and endocrine disruption (US EPA 2002). The estimated acute aquatic toxicity of endosulfan for striped bass “Mornone saxatillis”, water flea “Daphnia magna” and so on ranges from 0.1 to 166 μg/L (US EPA 2002). Endosulfan is still present in soils because it was widely used until relatively recently, and it has a half-life in soil of 60 days for α-endosulfan and 800 days for β-endosulfan (Steward and Cairns 1974). Another report has given estimated half-lives for the combined toxic residues (endosulfan plus endosulfan sulfate) between roughly 9 months and 6 years (US EPA 2002). Endosulfan can be released into the atmosphere through spray drift, post-application volatilization, and wind erosion of soil particles. Endosulfan and endosulfan sulfate were included in the Stockholm Convention list of persistent organic pollutants (POPs) on April 29, 2011.

Microbial degradation is a cost-effective, environmentally sound, and efficient way of remediating contaminated environments. There have been many studies of endosulfan biodegradation. The environmental fate and behavior of endosulfan was discussed in a review article (Weber et al. 2010), which includes important information about environmental endosulfan problems. However, no review articles have been published dealing, in detail, with the biodegradation of endosulfan. In this review, we summarize the aerobic microbial degradation of endosulfan. The review covers: (a) bacterial degradation, (b) fungal degradation, (c) biodegradation of the toxic metabolite endosulfan sulfate, and (d) biodegradation in soil.

Bacterial degradation

One of the problems with endosulfan biodegradation is that endosulfan sulfate, a toxic metabolite, accumulates (Kwon et al. 2002). Indigenous microorganisms in soil seem to oxidize endosulfan to endosulfan sulfate. The majority of reported fungi and bacteria form endosulfan sulfate and endosulfan diol during endosulfan degradation (Fig. 1, see (#1, 2)). It is likely that the toxicities of the endosulfan metabolites, such as endosulfan diol, are related to their water solubilities. In fact, the toxicity of endosulfan diol to Tubifex tubifex was found to be lower than the toxicity of endosulfan (Kumar et al. 2007). Therefore, endosulfan degradation by non-oxidative pathways, without the formation of endosulfan sulfate, is required to minimize its long-term toxicity. For instance, Kwon et al. (2002) isolated 40 bacteria from 24 endosulfan-polluted soil samples, and found that two of the bacteria, KE-1 and R2-3, degraded endosulfan without forming endosulfan sulfate. Klebsiella pneumoniae strain KE-1, in particular, degraded 93.6 % of the endosulfan in 10 days. In another study, Awasthi et al. (2003) performed an endosulfan degradation assay using two co-cultured Bacillus strains and found that the α-isomer was degraded by 94 % and the β-isomer was degraded by 86 %. The metabolite was endosulfan lactone rather than endosulfan sulfate. Proposed soil bacterial metabolic pathways for endosulfan from published studies are shown in Fig. 1. Although endosulfan sulfate is formed as the terminal metabolite in many cases, endosulfan diol, from the hydroxyl pathway, can be further degraded to endosulfan ether (Fig. 1, see (#3)) and endosulfan lactone (Fig. 1, see (#4)). In other pathways, endosulfan hydroxyether was detected (Fig. 1, see (#5)), and was produced from endosulfan through endosulfan monoaldehyde (Sutherland et al. 2000). The dechlorination of endosulfan by microorganisms is still not clearly understood, although Verma et al. (2006) reported the release of the chloride ion from endosulfan. The bacteria that degrade endosulfan and endosulfan sulfate are shown in Table 1.

Fig. 1
figure 1

Proposed pathway of endosulfan biotransformation by soil bacteria. Ese indicates the pathway described by Weir et al. (2006), which involves Ese monooxygenase. The dashed arrow (#6) reaction occurs when Mycobacterium sp. is used to express the ese gene. Esd indicates the pathway described by Sutherland et al. (2002b), which involves Esd monooxygenase, generating endosulfan monoaldehyde or endosulfan hydroxyether from β-endosulfan

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Table 1 Endosulfan-degrading bacteria reported and the metabolites produced
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Many endosulfan-degrading bacteria have been isolated using enrichment cultures. To our knowledge, the enrichment cultures have all used soils with a history of several endosulfan applications (Table 2). It also seems that two types of enrichment culture have mainly been used, with and without using endosulfan as the sole sulfur source. Sulfur-limited conditions have been used in enrichment cultures because many microorganisms can use sulfonates and sulfate esters as a source of sulfur for growth, even when they are unable to use the carbon skeleton of the compounds (Sutherland et al. 2002c). Sulfonates and sulfate esters are widespread in nature, and make up over 95 % of the sulfur content of most aerobic soils (Kertesz 1999). Sutherland et al. (2002a) enriched a growing medium with the endosulfan-degrading bacterium Mycobacterium sp. ESD using endosulfan as the only source of available sulfur. They also demonstrated that the bacterium can use various sulfur sources, including cysteine, dimethyl sulfoxide (DMSO), technical grade endosulfan, α-endosulfan, β-endosulfan, glutathione, MgSO4, methionine, 3-(N-morpholino)-propane sulfonic acid (MOPS), NaSO3, and sulfolane. The ESD strain transformed endosulfan to endosulfan monoaldehyde, endosulfan hydroxyether, and endosulfan sulfate when cysteine, glutathione, DMSO, MOPS, or sulfolane were present, but did not degrade endosulfan when sulfate, sulfite, or methionine was present. Sutherland et al. (2002a) also discussed the likelihood of the presence of sulfate or cysteine repressing the synthesis of the enzyme responsible for the metabolism of endosulfan. The authors found that enrichment in media deficient in sulfur sources seemed to lead to the efficient isolation of endosulfan-degrading bacteria. But it is still not clearly understood whether this was the best enrichment procedure or not, because endosulfan-degrading bacteria could be isolated using enrichment cultures both with and without sulfur sources. Common to almost all of the reports was the use of soil samples with a history of repeated endosulfan applications.

Table 2 Enrichment cultures used for the isolation of endosulfan-degrading bacteria
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Esd and Ese monooxygenase have been isolated from Mycobacterium sp. ESD (Sutherland et al. 2002b) and Arthrobacter sp. KW (Weir et al. 2006), respectively. Ese and Esd monooxygenase belong to the two-component flavin-dependent monooxygenase family. Monooxygenase generally introduces one atom of oxygen to an organic compound, with the other oxygen atom being reduced by electrons, from cofactors, to give water (Joosten and Berkel 2007). Ese monooxygenase is involved in the degradation of both endosulfan isomers to endosulfan monoalcohol and the degradation of endosulfan sulfate to endosulfan hemisulfate (Weir et al. 2006; Fig. 1, see Ese). Interestingly, the recombinant Mycobacterium smegmatis expressing ese gene degraded endosulfan sulfate to endosulfan monoalcohol and endosulfan dimethylene under sulfur-limited conditions (Fig. 1, see (#6)), but did not produce detectable levels of endosulfan sulfate metabolite in sulfur-rich conditions (Weir et al. 2006). This suggests that a sulfur source is one of the key elements in ese gene expression for the degradation of endosulfan or endosulfan sulfate. Esd monooxygenase exhibited no activity with the α-isomer of endosulfan or endosulfan sulfate, but generated endosulfan hydroxyether and endosulfan monoaldehyde from β-endosulfan (Sutherland et al. 2002b; Fig. 1, see Esd).

Fungal degradation

The degradation of endosulfan by fungi has been reported (Table 3). Phanerochaete chrysosporium is a commonly studied white rot fungus that has been shown to degrade and mineralize a wide range of industrial and agricultural pollutants, such as DDT (Bumpus and Aust 1987). P. chrysosporium degraded approximately 90 % of the endosulfan, and generated endosulfan sulfate and endosulfan diol as the principal metabolites (Kullman and Matsumura 1996; Fig. 2, see (#2, 3)). Kullman and Matsumura (1996) showed the proposal pathways that oxidation of the diol to hydroxyether was formed and the dialdehyde and lactone were the end product of metabolism in (Fig. 2, see (#4, 6)). Kamei et al. (2011) also reported that the white rot fungus Trametes hirsuta degraded more than 90 % of the endosulfan and approximately 70 % of the endosulfan sulfate, producing endosulfan diol and endosulfan dimethylene as metabolites (Fig. 2, see (#1)). In other studies, Trichoderma harzianum (Katayama and Matsumura 1993) and Mucor thermohyalospora (Shetty et al. 2000) degraded endosulfan, apparently by both the oxidation of endosulfan to endosulfan sulfate and its hydrolysis to endosulfan diol (Fig. 2, see (#2, 3)). Kataoka et al. (2010) also isolated Mortierella sp. strain W8, which could degrade endosulfan in a liquid culture, hardly generating endosulfan sulfate. This strain degraded more than 70 % and 50 % of α- and β-endosulfan, respectively, in 28 days. This strain also generated endosulfan diol as the first step in the degradation of endosulfan, and then further converted the diol to endosulfan lactone via endosulfan ether (Fig. 2, see (#3, 4, 5)). The pathways that have been proposed up to now are shown in Fig. 2.

Table 3 Endosulfan-degrading fungi reported, and the metabolites produced. Endosulfan is not completely degraded but slightly transformed into sulfur-free chlorinated hydrocarbons
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Fig. 2
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Proposed pathway of endosulfan biotransformation by fungi. #1 and #2 – 6 indicate parts of the pathway described by Kamei et al. (2011) and by Kullman and Matsumura (1996), respectively. Trametes hirsuta produced endosulfan diol and endosulfan dimethylene from the initial endosulfan sulfate substrate (Kamei et al. 2011)

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White rot fungi are able to degrade structurally diverse compounds, and that has generally been attributed to their extracellular oxidative enzymatic system that can degrade wood lignin (Tien and Kirk 1988). Three ligninolytic enzymes, manganese peroxidase (MnP), lignin peroxidase (LiP), and laccase, also catalyzed the degradation of organic pollutants (Mester and Tien 2000). The ligninolytic enzyme systems of the best-studied model organism, P. chrysosporium, have been shown to be triggered in response to N, C, or S limitation (Kotterman et al. 1996), while some basidiomycetes produced ligninolytic enzymes at high N concentrations (Kotterman et al. 1996). Indeed, Kullman and Matsumura (1996) showed the endosulfan-degradation under N limited conditions. This phenomenon seems to be similar with that of Mycobacterium sp. expressing ese gene as described above. On the other hand, endosulfan-degrading activity by T. harzianum was inhibited by di-n-propyl malaoxon, which is a hydrolytic enzyme inhibitor (Katayama and Matsumura 1993). Therefore, it has been suggested that hydrolases such as a sulfatase could hydrolyze endosulfan or endosulfan sulfate to endosulfan diol. Other fungi that produce endosulfan diol probably also possess hydrolyzing enzymes. However, a detailed understanding of the enzyme(s) involved in the formation of endosulfan diol is still lacking. Further study of the involvement of enzymes such as hydrolase and oxygenase on the degradation of endosulfan and endosulfan sulfate is required.

Degradation of endosulfan sulfate

A proportion of the residual endosulfan in soil is oxidized to endosulfan sulfate by indigenous microorganisms. Endosulfan sulfate is a toxic metabolite that is equally persistent to, or more persistent than, the parent endosulfan isomer. It is, therefore, very important that endosulfan sulfate is degraded to achieve the bioremediation of soils that have a history of endosulfan application. There have been quite few reports of the degradation of endosulfan sulfate. Arthrobacter sp. KW degraded endosulfan sulfate to endosulfan monoalcohol or endosulfan dimethylene through endosulfan hemisulfate (Weir et al. 2006). Pseudomonas sp. KS-2P (Lee et al. 2006), Pseudomonas sp. IITR01 (Bajaj et al. 2010), Klebsiella oxytoca KE-8 (Kwon et al. 2005), and Achromobacter xylosoxidans C8B (Singh and Singh 2011) degraded 71 % (7 days), 70 % (9 days), 64 % (6 days), and 80.1 % (15 days) of endosulfan sulfate, respectively. The fungus, T. hirsuta also degraded approximately 70 % of endosulfan sulfate in a 10-day incubation (Kamei et al. 2011). In contrast, Mortierella sp. W8, the endosulfan-degrading fungus that we have isolated (Kataoka et al. 2010), could not degrade endosulfan sulfate. The degradation of endosulfan sulfate seems to be more difficult than the degradation of endosulfan. Therefore, further studies should include the screening of endosulfan sulfate degrading microorganisms.

Biodegradation in soil

There have been reports on residues of total endosulfan, including endosulfan sulfate, in the environment (Weber et al. 2010). The incubation of endosulfan with soil under aerobic conditions resulted in endosulfan sulfate being formed as the major metabolite (Lee et al. 2006). Therefore, all studies of endosulfan concentrations in soil report the sum of the concentrations of α-endosulfan, β-endosulfan, and endosulfan sulfate. For example, maximum total endosulfan concentrations of 19 μg/kg dry soil in China (Jia et al. 2010) and 34.9 ± 52.6 μg/kg dry soil in Argentina (Gonzalez et al. 2010) have been reported. To achieve in situ biodegradation, microorganisms should degrade endosulfan and endosulfan sulfate under field conditions in the same as biodegradability in the laboratory. Mucor alternans, however, has been reported to lose its ability to degrade dieldrin (a structural analog of endosulfan) when added to dieldrin contaminated soil (Anderson et al. 1970). This imply that biodegradation in soil is difficult and is affected by many factors, such as soil microbial diversity, pH, temperature, moisture, organic matter content, and bioavailability. It is, therefore, important to understand characteristics of the microorganism and appropriate environmental conditions to achieve optimal degradation (Matsumoto et al. 2009). Some researchers have demonstrated this using soil microcosms to study the biodegradation of endosulfan. Kumar et al. (2008) showed that Ochrobacterum sp., Arthrobacter sp., and Burkholderia sp. degraded α- and β-endosulfan by over 60 % in soil spiked with endosulfan. Ochrobacterum sp. and Burkholderia sp. generated endosulfan diol as a metabolite, but Arthrobacter sp. oxidized endosulfan to endosulfan sulfate, which was then further metabolized. Arshad et al. (2008) reported that Pseudomonas aeruginosa degraded more than 85 % of α- and β-endosulfan in soil slurry spiked with endosulfan. For fungi, however, Goswami et al. (2009b) reported that Aspergillus sydoni could degrade endosulfan in unsterilized soil spiked with endosulfan. Kataoka et al. (2011) reported that Mortierella sp. W8 degraded α- and β-endosulfan by approximately 80 % and 50 %, respectively, after 28 days incubation in unsterilized soil spiked with endosulfan. Arshad et al. (2008) investigated the optimal conditions for the biodegradation of endosulfan in soil slurry, and found that the addition of organic acids stimulated the growth and activity of P. aeruginosa by acting as a carbon source, and this enhanced the degradation of endosulfan, whereas amino acids suppressed endosulfan degradation. Supplemental amino acids, acting as preferred sources of sulfur and carbon, could suppress the production of enzymes that degrade endosulfan. Goswami et al. (2009a) used different carbon sources, such as cane molasses and dextrose, and found that the initial degradation of endosulfan was more effective than when endosulfan was the sole carbon source, because molasses and dextrose offer a more easily available carbon source than do endosulfan. Mortierella sp. W8 used cane molasses and wheat bran as substrates in soil spiked with endosulfan (Kataoka et al. 2011) and wheat bran was found to be a better substrate than cane molasses for the biodegradation of endosulfan. There have been quite few studies of endosulfan and endosulfan sulfate degradation in soil, as described above, and many have used soils spiked with endosulfan and/or endosulfan sulfate. Endosulfan contaminated (e.g., unspiked) soil in agricultural field is probably more difficult to degrade than endosulfan spiked into soil because the bioavailabilities of endosulfan and endosulfan sulfate are likely to be lower in unspiked soil. Therefore, further work is needed to find which microorganisms degrade endosulfan sulfate in unspiked contaminated soil.

Conclusions

Endosulfan-degrading bacteria and fungi have been screened and their degradation abilities demonstrated. In all of microorganisms reported to date, endosulfan seems not completely to degrade but slightly transformed into sulfur-free chlorinated hydrocarbons (describe the “biodegradation” to “biotransformation” from this section onward). There are two main initial biotransformation pathways, resulting in the formation of endosulfan sulfate (oxidation) and of endosulfan diol (hydrolysis). Many of the efficient endosulfan-degrading microorganisms that have been isolated could metabolize endosulfan via the hydrolysis pathway, whereas indigenous microorganisms could oxidize endosulfan to endosulfan sulfate in soil. Some microorganisms can degrade both endosulfan and endosulfan sulfate. However, to our knowledge, there have been no reports of endosulfan bioremediation on sites polluted with endosulfan and endosulfan sulfate, and there is little information on the toxicity of endosulfan metabolites. More information on the biodegradation or biotransformation of endosulfan and its metabolites in soil, and on the toxicity of endosulfan metabolites is, therefore, still needed. However, two genes, ese and esd, which encode enzymes capable of degrading endosulfan, have been reported (Weir et al. 2006; Sutherland et al. 2002b). Ese monooxygenase, in particular, is involved in the initial oxidation of one of the endosulfan sulfate methylene groups, producing endosulfan hemisulfate, which is a sulfur-containing intermediate. The study of endosulfan and endosulfan sulfate biotransformation is less advanced than the study of the biodegradation of some other pollutants, so further advances in research on the biotransformation and biodegradation of endosulfan sulfate are expected.