Highly toxic organic compounds have been synthesized and released into the environment for direct or indirect application over a long period of time. Persistent organic pollutants (POPs) such as pesticides, fuels, polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), chlorophenols and dyes are some of these types of compounds [1]. POPs are extremely resistant to biodegradation by native flora [2] compared to the naturally occurring organic compounds that are readily degraded upon introduction into the environment. Therefore, hazardous wastes and chemicals have become one of the greatest worldwide problems of modern society. Among POPs, organochlorine pesticides (OPs) constitute a major environmental problem, because of their high toxicity, persistence in the environment and ability to bioaccumulate in the food chain [3–5]. These compounds reach aquatic environments through effluent release, atmospheric deposition and runoff among other ways [6]. Because of the low water solubility, OPs have a strong affinity for particulate matter, and consequently, sediments can serve as an ultimate sink [7,8].

The organochlorine compound 1,2,3,4,5,6 hexachlorocyclohexane (HCH) is a broad-spectrum pesticide that was used on a large scale worldwide since the 1940s, and is available in two formulations: technical-grade HCH (a mixture of different isomers, mainly α-(60%–70%), β-(5%–12%), γ-(10%–15%) and δ-HCH (6%–10%)) and lindane (almost pure γ-HCH) (Figure 1). This mixture was marketed as an inexpensive insecticide, but since γ-HCH is the only isomer that exhibits strong insecticidal properties, it has been common to refine it from technical-grade HCH and market it under the name “lindane”. As a result, large quantities of byproducts (mixtures of other HCH isomers) were dumped around production facilities leading to the contamination of many sites worldwide: For each ton of lindane produced, 8–12 tons of wastes are generated (generally enriched in the α and β isomers) [9]. Accordingly, there are very few officially recognized OP-contaminated sites in South America, and these are located principally in heavily populated industrial areas, i.e., Sao Paulo (Brazil), Buenos Aires (Argentina), and Santiago and Concepción (Chile). However, these official numbers grossly underestimate the real situation because of the existence of illegally contaminated sites [10]. For instance, the most important known illegal disposal of more than 30 tons of HCHs and other organochlorines such as DDT, took place in the small town of Argentina, in the province of Santiago del Estero [5,10].

All HCH isomers are acutely toxic to mammals, due to their mutagenic, teratogenic and carcinogenic properties [12]. Although nowadays its use is restricted or completely banned in most countries, it continues posing serious environmental and health concerns [13]. This is because γ-HCH was widely used as insecticide, which added to its high environmental persistence, ensuring lindane residues would be found all over the world and reaching aquatic environments through effluent release, atmospheric deposition and runoff, among other ways [6,12]. In this context, residues of γ-HCH and others HCH isomers have now been reported by many countries [13] in samples of air [14,15], water [16,17], soil [17–20], food commodities [21–23], milk [24,25] and even in human blood [26] and adipose tissue [27]. Since HCH spread to remote areas due to migration via various routes, their residues have been detected in Arctic, Antarctic and Pacific Ocean [28], although they had never been used or produced there. The recent inclusion of the α, β and γ-HCH isomers in the Stockholm Convention list of POPs has led to a renewed interest in these contaminants and the remediation of affected sites.

Traditional technologies routinely used for remediation of contaminated environmental soil include excavation, transport to specialized landfills, incineration, stabilization and vitrification. However, bioremediation technologies, which imply microorganisms and/or plants to degrade toxic contaminants in soil into less toxic and/or nontoxic substances, have become the focus of interest [29]. Microbial degradation is regarded as an important process of Ops-removal from sediments, and to date, many researchers have studied this issue [2]. In particular, aerobic and anaerobic bacteria are the most active agents of bioremediation [30], having the ability to interact, both chemically and physically, with substances leading to structural changes or complete degradation at the target molecule, playing a significant role in the transformation and degradation of pesticides. Even the most persistent pesticides can be metabolized to some extent by microbial cultures, either by utilization of the compounds as a source of energy or nutrients, or by cometabolism with other substrates supporting microbial growth. This last process is probably the most widespread mechanism for pesticide biodegradation [31]. Complete mineralization of pesticides or their transformation to nontoxic products is desirable, but it is more likely to be carried out by consortia of microorganisms than by single isolates [32–34].

As a way of not altering natural ecosystems, the trend in the last years has been the search and utilization of native bacteria. Many Gram-negative bacteria have been reported to have metabolic abilities to attack lindane. Several bacterial strains (predominantly Sphingomonas strains) capable of degrading HCH isomers under aerobic conditions have been isolated from contaminated soils [35–38]. On the other hand, among Gram-positive microorganisms, actinobacteria—particularly those belonging to the Streptomyces genus—have a great potential for biodegradation of organic and inorganic toxic compounds [39,40]. Benimeli et al.[41–45], Cuozzo et al.[46,47], Fuentes et al.[5,48] and Saez et al.[34] isolated and selected wild-type Streptomyces spp. which were able to tolerate and remove several OPs from culture media and soils.

Regarding eukaryotic cells, fungi have great capacity to degrade xenobiotic. Among them, white-rot fungi have been widely used to degrade several classes of pesticides (including γ-HCH) due to their ligninolytic potential [49–52]. Likewise, many plants—natural, transgenic, and/or associated to rhizosphere microorganisms—are extraordinarily active in removing or immobilizing pollutants, and particularly HCH isomers [38,53–55]. However, this article is focused on bacteria, although the HCH-bioremediation capacity of certain plants associated with microorganisms is mentioned.

In this review, it is intended to compile and update the information available on bacterial degradation of HCH. A brief account on the persistence and deleterious effects of these pollutant chemicals is also given.

Persistence and mobility of pesticides are defined primarily by their physical and chemical properties [37,56]. Li and Wania [57] compiled selected physicochemical properties, including vapor pressure, water solubility, Henry’s law constant (H), octanol-water partition coefficient (K_OW), and octanol-air partition coefficient (KOA) of major OPs. These properties have been used to screen many chemicals for persistent organic pollutants criteria [58]. For HCH isomers, such properties are quite different from one to another, depending on the chlorine positions in the cyclohexane ring (Figure 1) [12,37]. For instance, the vapor pressure of α-HCH is somewhat less than of γ-HCH. α-HCH has also been shown to be slightly more lipophilic than γ-HCH (log K_ow 3.8 versus 3.6). The Henry’s law constant for α-HCH is about twice as high as that of γ-HCH, so α-HCH is more likely to partition to the air. Another important difference between the isomers is the persistence of the β-isomer. β-HCH is more resistant to environmental degradation, it is also more lipophilic than the other isomers. These properties may result from its significantly smaller molecular volume. Since bonds between H, C, and Cl in the β-HCH at all six positions are equatorial, the molecule is denser and small enough to be stored in the interstices of lipids in animal tissues.

When comparing HCH isomers with other OPs, HCH exhibit relatively higher water solubility and moderately higher vapor pressures. Therefore, HCH is usually present in the environment as a gas in the atmosphere or dissolved in water, and a small percentage is adsorbed onto particles [59]. Brubaker and Hites [60] measured the gas phase reaction kinetics of the hydroxyl radical with α-HCH and γ-HCH and showed that these compounds have fairly long lifetimes in air and therefore can be transported long distances. Despite these approaches, there is controversy in the literature over the persistence of HCHs in soil, water and air, probably due to contrasting data resulting from the complex interactions of environmental factors affecting rates of both abiotic and biotic removal [37].

All the HCH isomers are acutely toxic to mammals. In addition, chronic exposure has been linked to a range of health effects in humans, including immunosuppression and neurological problems, and has been shown to cause liver cancer in rats and mice [12]; HCHs primarily affect the central nervous system. However, among different isomers, α-HCH and β-HCH are classified as probable human carcinogen and possible human carcinogen (respectively) by the United States Environmental Protection Agency [61]. As the most metabolically stable isomer, β-HCH is the predominant isomer accumulating in human tissues. This tendency to bioaccumulate is a cause of concern as some studies indicate that β-HCH may act as an environmental estrogen [12].

Different aerobic and anaerobic bacteria have been found to be capable of using halogenated compounds as a growth substrate. Most of the HCH-bioremediating aerobic bacteria and the isolation countries are: Micromonospora sp. and several Streptomyces spp. strains (Argentina) (see Table 1); Bacillus sp. [62] and Pseudomonas sp. [63] (Canada); Sphingobium sp. MI1205 [64] (China); Rhodanobacter lindaniclasticus RP5557 [65,66] and Sphingobium francense sp. [67] (France); Alcaligenes faecalis S-1, Alcaligenes faecalis S-2 [68], Arthrobacter citreus BI-100, Bacillus circulans, Bacillus brevis[69], Microbacterium sp. ITRC1 [56], Pseudomonas aeruginosa[70], Pseudomonas aeruginosa ITRC-5 [71], Pseudomonas sp. [72], Sphingobium chinhatense IP26 [73], Sphingobium indicum B90A [74], Sphingobium quisquiliarum P25 [75], Sphingobium ummariense RL-3 [76], Sphingobium sp. UM2, Sphingobium sp. HDU05, Sphingobium sp. HDIP04, Sphingobium sp. F2, Sphingomonas sp. UM1 [77] and Xanthomonas sp. ICH12 [78] (India); Sphingobium japonicum UT26 [79], Sphingobium sp. BHC-A [80], Sphingobium sp. SS04-1, Sphingobium sp. SS04-2, Sphingobium sp. SS04-3, Sphingobium sp. SS04-4, and Sphingobium sp. SS04-5 [81] (Japan); Sphingomonas sp. DS2, Sphingomonas sp. DS2-2, Sphingomonas sp. DS3–1 [35], Sphingomonas sp. α1–2, Sphingomonas sp. α4-2, Sphingomonas sp. α4-5, Sphingomonas sp. α16–10, Sphingomonas sp. α16–12, Sphingomonas sp. γ1-7, Sphingomonas sp. γ12-7, Sphingomonas sp. γ16-1 and Sphingomonas sp. γ16-9 [36] (Spain) and Escherichia coli[82] and Pseudomonas putida[83] (USA).

It is important to note that bioremediation involves the intervention aimed at alleviating pollution, where the organism removes the pollutant by different mechanisms (bioaccumulation, biosorption, biodegradation). This section deals only with bacterial biodegradation of HCH, which tackles the biological bases of the metabolism of recalcitrant compounds [30].

Actinobacteria (Gram-positive bacteria with high G+C content of DNA) are morphologically complex bacteria that represent an important component of microbial soil biodiversity. Depending on the taxon, they may produce branched rods, complex mycelial structures and spore bodies, motile or nonmotile rods. The group has a great potential for bioremediation of organic and inorganic toxic compounds [104] and particularly pesticide-degrading strains are not restricted to a particular genus or family. A review by De Schrijver and De Mot [31] showed that the genera Arthrobacter, Brevibacterium, Clavibacter, Corynebacterium, Mycobacterium, Nocardia, Rhodococus and Micromonospora are pesticide degrading actinobacterias. However, in that study, only one strain belonging to the Streptomyces genus [105] is mentioned as able to dechlorinate γ-HCH. Most recent studies have isolated and characterized Streptomyces spp. strains able of degrading OPs (including γ-HCH) and some strains are proposed to be used for soil decontamination (Table 1).

Streptomyces genus is well known for their distinct impacts on mankind since its species are prolific producers of antibiotics [106]. In addition to their potential metabolic diversity, Streptomyces spp. may be well suited for soil inoculation as a consequence of their mycelial growth habit (Figure 2), relatively rapid rates of growth, colonization of semiselective substrates and their ability to be genetically manipulated [107]. For soil bioremediation purposes, one additional advantage is that the vegetative hyphal mass of these microorganisms can differentiate into spores that assist in spreading and persisting. This can be seen as adaptation to living in soil, especially to extreme environmental conditions such as scant nutrients, intense salt load and low pH, as well as contamination [104].

Some phytoremediation techniques, based on the interactions between plants and their associated microorganisms have been proposed as cost-efficient and eco-friendly methods to clean up polluted soils [38,55]. Microbe assisted phytoremediation appears to be particularly effective for organic pollutants, including the more recalcitrant POPs like HCH [115].

Recent studies demonstrate enhanced dissipation and/or mineralization of OPs at the root-soil interface [38]. This rhizosphere effect is generally attributed to an increase in microbial density and/or metabolic activity due to the release of plant root exudates (REs). REs contain water soluble, insoluble, and volatile compounds, including sugars, amino acids, organic acids, nucleotides, flavonoids, phenolic compounds and certain enzymes [116]. Since REs are complex mixtures of substrates, they not only provide a nutrient-rich habitat for pollutant degraders but can also potentially enhance biodegradation in different ways: they may facilitate the cometabolic transformation of pollutants with similar structures, induce catabolic enzymes involved in the degradation process and/or enhance the contaminant bioavailability [38,117]. In addition, REs may directly induce contaminant degradation by root-driven extracellular enzymes [117,118] as was demonstrated by Magee et al.[119], who reported dechlorination of polychlorinated biphenyls (PCBs) by crude extract of nitrate reductase from Medicago sativa and a pure commercial nitrate reductase from maize. Similar results were obtained by Alvarez et al.[54], who detected specific dechlorinase activity (SDA) in maize REs. In that study, SDA explained 42% of γ-HCH degradation in minimal medium supplemented with REs and γ-HCH.

Phytostimulation of OP-degrading microorganisms is therefore likely to be a successful strategy for the remediation of γ-HCH-contaminated environments. However, a careful selection of tolerant plant species and optimizing plant growth are vital since the phytotoxic nature of these contaminants can inhibit plant performance, reducing the overall efficiency of the remediation process [55,120,121]. Some plants are well adapted to acidic conditions as generated during OPs degradation. For instance, Benimeli et al.[45] showed that γ-HCH concentration of 100, 200 and 400 μg kg⁻¹ soil did not affect the germination and vigor index of maize plants seeded in contaminated nonsterile soils. Additionally, maize plants may create particularly good environmental condition for soil microorganisms [122]. Likewise, the leguminous species Cytisus striatus grow spontaneously on HCH-contaminated sites and has been proposed as a candidate species for the cleanup of this type of contaminant [38,123]. Becerra-Castro et al.[55] inoculated substrates seeded with Cytisus striatus with the previously isolated bacteria Rhodococcus erythropolis ET54b and Sphingomonas sp. D4 [55,123]. The authors found that substrates planted with C. striatus showed a higher dissipation of HCH isomers and that both microorganisms protected the plants against the toxic effects of the contaminant. Consequently, they propose that inoculating C. striatus with this combination of bacterial strains could therefore be a promising approach for the remediation of HCH-contaminated sites. The presence of plant-colonizing, naphthalene-degrading bacteria were shown to protect their host plants against the toxic effects of naphthalene exposure [116,124]. Likewise, the trichloroethylene (TCE)-degrading poplar endophyte Pseudomonas putida W619-TCE promoted plant growth and reduced TCE phytotoxicity in inoculated poplar cuttings [121].

Alvarez et al.[54] evaluated the effect of maize REs on growth and δ-HCH removal by Streptomyces sp. A5 and Streptomyces sp. M7 (Table 1). Results showed that both strains were able to grow on minimal medium supplemented with maize REs as sole carbon source, suggesting that these microorganisms are competitive at the rhizosphere level. Furthermore, maize REs markedly influenced the δ-HCH removal by both Streptomyces sp. A5 and M7 since pesticide removal reached approximately 55 and 35%, respectively, when REs are present in the culture medium. On the contrary, no decrease in pesticide concentration was observed by Renzt et al.[125] and Louvel et al.[126], who studied the repression of phenanthrene-degrading activity of Pseudomonas putida in the presence of REs of different plant species. The authors suggest that as REs are a complex mixture of substrates; some of them could be repressing microbial-degrading activity.

On the other hand, fungi growing in symbiotic association with plants have unique enzymatic pathways that help to degrade pesticides that could not be transformed solely by bacteria [2]. For instance, mycorrhizal fungi form symbioses with a broad range of plant species and can contribute to plant growth and survival by reducing stresses associated with toxic wastes. The effects of soil HCH contamination on vegetation and its associated arbuscular mycorrhizas was investigated by Sáinz et al.[127]. The authors found that a preinoculation of four plant species with an isolate of Glomus deserticola obtained from the HCH-contaminated soil resulted in increased growth and fungal colonization of roots, suggesting that the fungus increases the tolerance of plants to the toxic soil environment.

HCH is one of the most extensively used OPs for both agriculture and medical purposes. Currently, its use is being phased out because of its toxicity, environmental persistence and accumulation in the food chain. Though the use of technical mixture containing four isomers was banned in several advanced countries, many developing countries continue using γ-HCH (lindane) for economic reasons. Thus, new sites are continuously being contaminated by γ-HCH and the other HCH-isomers. Considerable research on chemical pollutants now provides the necessary body of knowledge to understand their recalcitrance and toxic nature. A possible pathway for remediation of HCH-contaminated soils is the use of indigenous bacteria. There have been many reports regarding aerobic degradation of HCH by Gram-negative bacteria, especially those belonging to the Sphingomonas genus. The sphingomonads are clearly the group of first choice for further work on strain development for bioremediation purposes, mainly due to the extensive knowledge reached about its metabolic pathways of HCH-degradation. This preponderance of data on Gram-negative bacteria reflects the fact that molecular techniques for this group of bacteria are much more advanced when compared with Gram-positive bacteria, rather than the intrinsic bioremediation potentials of the respective groups. Regarding this approach, Gram-positive bacteria like actinobacteria, are highly promising for HCH-bioremediation purposes. Strains of the Streptomyces genus have been seen to present particularly great potential for remediation of toxic organic and inorganic compounds. In fact, most recent studies have isolated and characterized Streptomyces spp. able to dechlorinate γ-HCH and some strains are proposed to be used for soil decontamination. Furthermore, phytostimulation of γ-HCH-degrading Streptomyces spp. appear to be a successful strategy for the remediation of γ-HCH-contaminated environments.

Work to date indicates that Streptomyces spp. have the requisite capabilities to degrade γ-HCH, although further studies on enzymes characterization, particularly for Lin enzymes, are needed to develop effective bioremediation technologies.