Folia Microbiol (2014) 59:321–332

DOI 10.1007/s12223-014-0304-8

Bacterial mechanisms for Cr(VI) resistance and reduction:

an overview and recent advances
Munees Ahemad

Received: 23 July 2013 / Accepted: 12 January 2014 / Published online: 29 January 2014
# Institute of Microbiology, Academy of Sciences of the Czech Republic, v.v.i. 2014

Abstract Chromium pollution is increasing incessantly due (Viti 2006; Ahemad et al. 2009). The stability and sustainabil-
to continuing industrialization. Of various oxidation states, ity of the environment is severely affected by different types of
Cr6+ is very toxic due to its carcinogenic and mutagenic pollution like water, air, soil, noise, pesticides, heavy metal
nature. It also has deleterious effects on different microorgan- pollution, etc. because they have a long-term detrimental
isms as well as on plants. Many species of bacteria thriving in impact on the functional activities of microorganisms by
the Cr6+-contaminated environments have evolved novel affecting their growth and metabolism (Ahemad and Malik
strategies to cope with Cr6+ toxicity. Generally, decreased 2011; Ahemad and Khan 2012; Ahemad 2012). Of them,
uptake or exclusion of Cr6+ compounds through the mem- chromium (Cr) pollution is increasing rapidly and ceaselessly
branes, biosorption, and the upregulation of genes associated owing to mine tailing, corrosion control and effluents released
with oxidative stress response are some of the resistance from non-ferrous metal industries, the continuing incorrect
mechanisms in bacterial cells to overcome the Cr6+ stress. In disposal of industrial by-products and wastes and an extensive
addition, bacterial Cr6+ reduction into Cr3+ is also a mecha- application of chromium in electroplating industries, pulp and
nism of specific significance as it transforms toxic and mobile paper production, manufacturing resistant alloy products, air-
chromium derivatives into reduced species which are innocu- crafts, and nuclear reactor vessels, negative and film making,
ous and immobile. Ecologically, the bacterial trait of reductive wood preservation, pyrotechnics, electronics, leather tanning,
immobilization of Cr6+ derivatives is of great advantage in cement producing plants, and dye productions (Baral et al.
bioremediation. The present review is an effort to underline 2006; Chirwa and Molokwane 2011; Alam and Ahmad 2012;
the bacterial resistance and reducing mechanisms to Cr6+ Lu et al. 2013).
compounds with recent development in order to garner a Chromium, the seventh most plentiful element in the
broad perspective. earth’s crust (0.1–0.3 mg/g), is a redox active 3d transition
metal from group VI-B of the periodic table with diverse
oxidation states (Fig. 1) wherein the trivalent Cr3+ species
and hexavalent Cr6+ forms are ecologically significant, since
Introduction
these are the most stable oxidation states in the natural envi-
ronment (Smith et al. 2002; Barceloux 1999). Moreover,
Natural environment is tremendously diverse and exhibits a
physic-chemical properties and biological reactivity of triva-
wide array of microorganisms which reflect the nature of the
lent and hexavalent species vary substantially. Cr3+ exhibits
habitat and the ability of individual members to coexist suc-
greater affinity for organics and consequently forms insoluble
cessfully by maintaining the dynamics of a given ecosystem
complexes which precipitate generally in the form of hydrox-
ides, oxides, and sulphates (Nickens et al. 2010; Chirwa and
M. Ahemad (*)
Department of Agricultural Microbiology, Faculty of Agricultural Molokwane 2011). In contrast, Cr6+ is exceedingly persistent
Sciences, Aligarh Muslim University, Aligarh 202002, U.P., India into the environment due to considerable solubility and is
e-mail: [email protected] mainly the result of anthropogenic input, and only 0.001 %
is reported to be produced by geological processes (Barceloux
M. Ahemad
Department of Biology, College of Science, Bahir Dar University, 1999; Chirwa and Molokwane 2011). Despite this, organic
Bahir Dar, Ethiopia matter reduces Cr6+ into Cr3+; high concentration of Cr6+,
322 Folia Microbiol (2014) 59:321–332

however, can overcome the reducing capacity of an environ- Cr5+/Cr4+ and free radicals are also the principal sources of
ment, and consequently, Cr6+ persists as a pollutant because of Cr6+ mediated carcinogenesis (Salnikow and Zhitkovich
its comparatively higher biological availability (Cheung and 2008) as well as apoptosis (Ye et al. 1999). Also, Cr5+ in
Gu 2007; Nickens et al. 2010). Microorganisms in such Cr6+- chemical solutions generates free radicals to a large extent, for
laden environments have evolved specific resistance systems instance, hydroxyl radicals and superoxide which are collec-
to survive by evading metal stress through efflux or minimiz- tively referred to as reactive oxygen species (ROS). During
ing uptake and detoxifying the various Cr6+ species including the redox cycle, Cr6+ is regenerated to be reduced again
chromates and dichromates via reductive immobilization through the cellular constituents and one-electron reducers,
(Dhal et al. 2010; Nguema and Luo 2012; Tharannum et al. thus producing considerable ROS and, in turn, substantial
2012; Garg et al. 2012). A number of Cr6+-resistant and Cr6+- oxidative stress (Ackerley et al. 2004a). On the other hand,
reducing bacterial species belonging to diverse genera have intracellular cationic Cr3+ as well as Cr4+ complexes electro-
been isolated (Riaz et al. 2010; Alam and Ahmad 2012; Ge statically interact with negatively charged phosphate in DNA
et al. 2012; Murugavelh and Mohanty 2013; Shi et al. 2012). and inhibit DNA replication, consequently increasing the
This review is intended to highlight different resistance strat- errors during RNA transcription (Plaper et al. 2002;
egies and reducing mechanisms in bacteria with current up- Bencheikh-Latmani et al. 2007; Cervantes and Campos-
dates and development. García 2007; Ramírez-Díaz et al. 2008). Furthermore, Cr3+
species react with the carboxyl and thiol groups in enzymes
thus affect their enzymatic efficiency by altering the structure
Chromium toxicity (Cervantes et al. 2001).
As environmental contaminants, Cr6+ compounds are re-
Whether chromium is really an essential element for organ- sponsible for the changes in the structure and diversity of
isms is questionable as a low concentration of chromium different microbial communities in soil ecosystem (Zhou
(Cr3+) in the diet displayed no toxicity in current studies, et al. 2002; Turpeinen et al. 2004; Viti 2006). For instance,
and no essential metabolic role is known till date (Di Bona bacterial growth rate declines, or lag phase is extended with
et al. 2011). However, a minor increment in chromium con- uncoupling of energy, as the chromate concentration is pro-
centration results into several environmental and health haz- gressively increased (Garbisu et al. 1998; Nepple et al. 2000;
ards (MAK 2012). The oxidation states of chromium princi- Chardin et al. 2002). Moreover, Cr6+ toxicity to bacteria is
pally are the sole determiner of the biological toxicity in also evidenced in the form of morphological changes like
addition to its reactivity and solubility as well as absorption filamentous growth (Ackerley et al. 2006; Chourey et al.
capacity (Daulton et al. 2007; MAK 2012). Among different 2006), altered gene expression (Chourey et al. 2006;
oxidation states, Cr6+ species being strong oxidants are more Ackerley et al. 2006), activation of the SOS response to
toxic (comparatively, 1,000 times more toxic than Cr3+ forms) counteract the oxidative stress, and induction of prophage-
and causes oxidative damage because of its mutagenic and related genes (Ackerley et al. 2006). Under Cr6+ stress, bac-
carcinogenic nature (Chirwa and Molokwane 2011). More- terial genes involved in DNA metabolism, cell division, bio-
over, the United States Environmental Protection Agency synthesis and degradation of murein and membrane response
(USEPA) has included Cr6+ among 17 chemicals which are as well as environmental stress protection have been reported
considered the most hazardous to human health (Marsh and to be upregulated, whilst genes related with chemotaxis and
McInerney 2001). In contrast, Cr3+ species are comparatively protein transport were repressed by Cr6+ (Chourey et al.
less toxic because they generally fail to cross the membranes 2006).
and enter cells owing to their low solubility and low tendency Compared to bacteria, fungi are generally less susceptible
to be adsorbed in organic carbon and mineral surfaces to metal toxicity. Most of the fungal species are resistant to
(Cervantes et al. 2001; Codd et al. 2001; Daulton et al. chromate toxicity due to the decreased uptake of Cr6+, defect
2007). After entering the cell, Cr6+ can subsequently react in the sulphate transport system, and participation of vacuolar
with several reducing compounds (such as NAD(P)H, structures and acid phosphatase enzymes in addition to reduc-
FADH2, pentoses, cysteines, and antioxidants like ascorbate ing mechanisms for Cr6+-containing compounds (Ware 2003).
and glutathione as well as one-electron reducers such as Unlike studies of bacterial and fungal interactions with chro-
glutathione reductase) producing unstable intermediates such mium derivatives, interactions in the case of algae are least
as Cr5+ and Cr4+ as well as free radicals. The resultant inter- studied. However, noxious effects of Cr6+ on algal motility,
mediates can cause oxidative damage to proteins and DNA by growth rate, respiration, and photosynthesis are scarcely avail-
forming a range of DNA lesions, together with Cr-DNA able (Cervantes et al. 2001).
adducts, DNA-DNA crosslinks, DNA-protein crosslinks In plant systems, Cr6+ not only adversely affects the
(Cervantes and Campos-García 2007; Cheung and Gu 2007; growth, photosynthesis, and transpiration (Dey et al. 2009)
Nickens et al. 2010). Moreover, short-lived intermediates but also affects the activities of different physiologically
Folia Microbiol (2014) 59:321–332 323

Fig. 1 Different chemical species

of chromium and their Naturally absent,
characteristics derived from CrO43-, half-life long, relatively Cr5+
few stable compounds containing Cr5+

Naturally absent,
Naturally absent Cr0 Cr4+
short half-life

CHROMIUM (Cr)
SPECIES

Most stable state, Forms unstable

rarely natural Cr6+ Cr3+ compounds, naturally
but anthropogenic origin occurs in ores

Relatively unstable,
Cr2+
readily oxidized in Cr3+

important enzymes like, superoxide dismutase, catalase, biological membranes are almost impervious to Cr3+ owing to
ascorbate peroxidase, and glutathione reductase the insolubility of Cr3+ compounds (Nickens et al. 2010).
(Subrahmanyam 2008). For instance, Riaz et al. (2010) re-
ported that growth, yield, and biochemical parameters of
chickpea (Cicer arietinum var NM-88) plants were severely Bacterial Cr(VI) resistance
decreased in the presence of two chromium salts CrCl3 and
K2CrO4 (300 μg/mL). Moreover, substantial chromium con- Mostly, decreased uptake or expulsion of Cr6+ compounds
tents were detected in roots (1.912 mg Cr/g dry mass), and through the transmembrane sulphate shuttle in membranes,
even smaller quantities were also transferred into shoots biosorption, and the upregulation of genes associated with
(0.086 mg Cr/g dry mass) and leaves (0.074 mg Cr/g dry oxidative stress response is considered as different modes of
mass). It is also reported that even no seed was formed at resistance in bacterial cells to overcome the Cr6+ stress
1.0 mM concentration (Sharma et al. 1995). (Brown et al. 2006; Cheung and Gu 2007; Congeevaram
et al. 2007; Ramírez-Díaz et al. 2008; Alam and Ahmad
2013). In most cases, metal resistance mechanisms have been
attributed to plasmid genes and are believed to have evolved
Chromium transport by horizontal gene transfer in response to the selective pres-
sure in the contaminated environment (Francisco et al. 2002).
To exert toxicity or to participate in physiological functions, Plasmid-mediated Cr6+ resistance has been reported in bacte-
heavy metal ions firstly have to traverse the cell through the rial species of Pseudomonas (Mondaca et al. 1998),
membranes (Nies 1999). Cr6+, which largely occurs as Alcaligenes (Collard et al. 1994), Salmonella (Ghosh et al.
oxyanions, is unable to bind by the anionic components of 2000), Bacillus (Kamala-Kannan and Lee 2008; Verma et al.
bacterial membranes, while the cationic Cr3+ derivates bind 2009), Escherichia coli (Verma et al. 2002), Shewanella
tightly to different components of bacterial envelopes (Aguilar-Barajas et al. 2008), etc. In fact, Cr6+ resistance is
(Cervantes et al. 2001; Neal et al. 2002). Due to the structural independent from Cr6+ reduction which was initially sug-
similarity of Cr6+ forms [like chromate (CrO42−) and dichro- gested as a mechanism for bacterial Cr6+ resistance by de-
mate (Cr2O72−) oxyanions] to SO42−/PO43− anions, they can creasing the intracellular Cr6+ concentration (Nies et al. 1989;
easily be transported across the biological membranes via Cervantes and Campos-García 2007). Evidently, the degree of
active sulphate transporters in both prokaryotes and eukary- Cr6+-reducing activity was found similar in both sensitive
otes (Daulton et al. 2007; Collins et al. 2010). On the contrary, (wild-type) and resistant (mutant) strains of Pseudomonas
324 Folia Microbiol (2014) 59:321–332

fluorescens LB300 under sublethal concentrations of Cr6+ et al. 2005; Brown et al. 2006; Chourey et al. 2006; Ramírez-
(Bopp and Ehrlich 1988). Díaz et al. 2008). For instance, expression of the chrB, chrC,
In addition to the abovementioned mechanisms, bacteria or chrF genes for Cr6+ resistance in O. tritici strain 5bvl1 was
also employ other resistance strategies to counter the Cr6+ related to increased resistance to superoxide-generating agents
stress as reviewed by Chirwa and Molokwane (2011), as (Morais et al. 2011).
follows: (1) offsetting Cr6+-induced oxidative stress by acti-
vating ROS scavenging enzymes (e.g., catalase, superoxide
dismutase), (2) DNA repair by SOS response enzymes (RecA,
RecG, RuvAB) to counter DNA damage, and (3) regulation of Bacterial Cr(VI) reduction
iron uptake to prevent the production of hydroxyl radicals
through the Fenton reaction (Table 1; Fig. 2). Since the first report of anaerobic Cr 6+ reduction by
Chromate transporters (Chr) which act as efflux pumps are Romanenko and Koren’Kov (1977) in uncharacterized
believed to be responsible for Cr6+ resistance (Díaz-Pérez Pseudomonas sp., worldwide researchers have isolated both
et al. 2007; Ramírez-Díaz et al. 2008). These have been aerobic and anaerobic Cr6+ reducing bacteria belonging to a
identified by cloning and sequencing of the chrA genes which range of genera from diverse environments (Opperman et al.
encode hydrophobic proteins (ChrA) of 416 and 401 amino 2008; He et al. 2009; Alam and Ahmad 2012; Batool et al.
acid residues in Pseudomonas aeruginosa and Cupriavidus 2012; Ge et al. 2012; Narayani and Shetty 2012; Nguema and
metallidurans, respectively, sharing amino acid identity ap- Luo 2012; Shi et al. 2012). Evidently, bacteria catalyzing the
proximately 29 % (Nies et al. 1990; Cervantes et al. 1990). reduction of Cr6+ to Cr3+ are ubiquitous in Cr6+ contaminated
Hydropathic profiles revealed that both ChrA from as well as uncontaminated water bodies, soils, and sediments
Pseudomonas and Cupriavidus are the membrane proteins (Table 2). These reduce Cr6+ either aerobically or anaerobi-
(Díaz-Pérez et al. 2007; Ramírez-Díaz et al. 2008). Lately, cally or under both conditions depending on the microbial
ChrA of Pseudomonas has been found to have a 13- species (Cervantes et al. 2001; Cervantes and Campos-García
transmembrane segment that came into existence by the du- 2007) and various factors affect their reducing efficiency
plication of a six-transmembrane segment ancestral protein; in (Lowe et al. 2003; Alam and Malik 2008; Xu et al. 2013).
this manner, the two halves of ChrA could carry out distinct Generally, Cr6+-reducing aerobes utilize NADH and endoge-
functions in the Cr6+ transportation (Cervantes and Campos- nous cell reserves, and Cr6+-reducing anaerobes use electron
García 2007; Jiménez-Mejía et al. 2006). Further, the reduced transport systems containing cytochromes to reduce Cr6+ de-
uptake of Cr6+ by ChrA of Pseudomonas is based on an rivatives (Zhu et al. 2008; Dey and Paul 2013) Redox condi-
energy-dependent chemiosmotic efflux system of Cr6+ from tions employed by different Cr6+-reducing bacteria have been
the cytoplasm (Pimentel et al. 2002; Ramírez-Díaz et al. displayed in Table 2.
2008). Notwithstanding, ChrA proteins have not been dem- Biologically and ecologically, mechanisms of bacterial
onstrated to be involved in sulphate transportation; the efflux Cr6+ reduction are of specific significance as they transform
of chromate has been found to be inhibited by sulphate. noxious and mobile chromium derivatives into reduced spe-
However, Nies et al. (1998) proposed that ChrA might func- cies which are innocuous and immobile (Daulton et al. 2007;
tion as chromate/sulphate antiporters. In addition to the Soni et al. 2013). In fact, the bacterial enzymes which reduce
plasmid-encoded chrA1 gene, a ChrA homolog (chrA2) is Cr6+ species are not primarily the Cr6+ reductases; they also
too found in Cupriavidus chromosome, which is also respon- perform other biological functions in addition to Cr6+ reduc-
sible for Cr6+ resistance (Juhnke et al. 2002). Moreover, tion (Ishibashi et al. 1990; Cervantes and Campos-García
Ochrobactrum tritici strain 5bvl1 contains the transposon- 2007). For instance, they may principally act as iron reduc-
located (TnOtChr) genes chrB, chrA, chrC, and chrF for tases, nitroreductases, glutathione reductase, lipoyl reductase,
Cr6+ resistance. Of them, the chrB and chrA genes were found ferredoxin-NADP+ reductase, or other metal reductases
to be essential for the establishment of high resistance (Morais (Kwak et al. 2003; Mazoch et al. 2004; Cervantes and
et al. 2011). Campos-García 2007; Opperman et al. 2008).
Generally, amino acid sequences in different Chr proteins Generally, three underlying mechanisms of Cr6+ reduction
are not remarkably conserved and these proteins differ in have been identified (Fig. 2) (Chueng and Gu 2007; Ramírez-
topological orientations and genomic sequences (Díaz-Pérez Díaz et al. 2008; Ngwenya and Chirwa 2011; Chirwa and
et al. 2007). Regarding Cr6+ resistance mediated by the up- Molokwane 2011):
regulation of oxidative stress response genes, bacterial genes
encoding for superoxide dismutase, catalase, thioredoxin, 1. Under anaerobic conditions, numerous components of the
glutaredoxin, and different regulatory proteins related to cell’s protoplasm such as amino acids, nucleotides, car-
DNA repair system together with DNA helicases and endo- bohydrates, vitamins, organic acids, glutathione, hydro-
nucleases are also upregulated under chromate stress (Miranda gen NADH (NADPH in some species), flavoproteins, and
Folia Microbiol (2014) 59:321–332 325

Table 1 Bacterial mechanisms for Cr(VI) resistance

Enzyme/system Bacterial species Function Reference

Transport
ChrA transporter Pseudomonas. aeruginosa Efflux of cytoplasmic chromate Alvarez et al. (1999)
Cys operon products Shewanella oneidensis Sulphate transport Brown et al. (2006)
TonB receptor, hemin transporter Shewanella oneidensis Iron transport Brown et al. (2006)
Reduction
chrB, chrC/chrF Ochrobactrum tritici 5bvl1 Resistance to superoxide- Morais et al. (2011),
generating agents Branco et al. (2008)
SOD, catalase Escherichia coli Combat of oxidative stress Ackerley et al. (2004a)
Outer membrane proteins Caulobacter crescentus General stress response Hu et al. (2005)
DNA repair
RuvB Ochrobactrum tritici 5bvl1 Repair of DNA damage Morais et al. (2011)
RecG and RuvB DNA helicases Pseudomonas aeruginosa Repair of DNA damage Miranda et al. (2005)
SO0368, UvrD, and HrpA helicases Shewanella oneidensis Repair of DNA damage Chourey et al. (2006)
Other mechanisms
Cys operon products Shewanella oneidensis Sulphur metabolism Brown et al. (2006)
Adenylyl sulphate kinase Shewanella oneidensis Sulphur metabolism Brown et al. (2006)
Ferritin Shewanella oneidensis Iron binding Brown et al. (2006)

Modified from Das (2009)

hemeproteins which act as electron donors reduce Cr6+ Amphibacillus sp. KSUCr3 employs membrane cell frac-
which serves as terminal electron acceptor. tions for Cr6+ reduction which are expressed constitutive-
2. Soluble reductases: under aerobic conditions, NAD(P)H- ly (Blake et al. 1993; Cheung et al. 2006; Ibrahim et al.
dependent extracellular soluble reductases are produced 2012b). Glucose was the best external electron donor,
purposely by the cell to reduce Cr6+ to Cr3+ that is re- showing enhancement of the enzyme activity by about
moved by reacting with functional groups present on cell 3.5-fold (Ibrahim et al. 2012b).
surface. For instance, Pseudomonas putida PRS2000,
Pseudomonas ambigua G-1, Desulfovibrio vulgaris, and Very few bacterial Cr6+ reductases have been purified and
E. coli ATCC 33456 have been reported to produce sol- characterized with regard to their gene sequence to illuminate
uble Cr6+ reductases (Ishibashi et al. 1990; Suzuki et al. their functional aspect. Most of the information regarding the
1992; Shen and Wang 1993; Lovley and Phillips 1994; biochemistry of Cr6+ reductases has come from the studies on
Chen and Hao 1998) which utilize diverse electron donors chromate reductases of P. putida (ChrR) and E. coli (YieF)
and can be located either inside or outside the bacterial which are cytoplasmic flavoproteins that completely reduce
cell (Chen and Hao 1998). Since reduction mediated by Cr6+ into Cr3+ (Park et al. 2000; Gonzalez et al. 2003;
such reductases is an energy-requiring and highly regu- Ackerley et al. 2004b; Gonzalez et al. 2005). These
lated process, these enzymes are produced constitutively. NAD(P)H-dependent homodimeric enzymes belonging to
Due to independence from transport mechanisms for the NADH-dh2 family possess non-covalently bound flavin
Cr6+/Cr3+ intake and expulsion, extracellular Cr6+ reduc- mononucleotide (FMN). In addition, ChrR of P. putida cata-
tion is advantageous for bacterial cell as it protects the cell lyzes the reduction of Cr6+ into Cr3+ optimally at 70 °C
from Cr6+/Cr3+-induced DNA damage. transiently producing Cr5+ as an intermediate, while YieF of
3. Membrane-associated reductases: anaerobic Cr6+ reduc- E. coli directly reduces Cr6+ by transferring four electrons at
tion generally involves membrane-associated reductases optimal temperature 35 °C wherein three electrons are used up
which sometimes require H2 or glucose as electron donors in reducing Cr6+ and the rest one is transferred to oxygen
(Ibrahim et al. 2012a). Moreover, anaerobic Cr6+ reduc- (Ackerley et al. 2004b). Both ChrR and YieF also reduce
tion wherein Cr6+ acts as an electron acceptor in the quinones, potassium ferricyanide, and 2,6-
electron transport chain is also reported (Wang 2000; dichloroindophenol. In contrast, YieF also reduces cyto-
QuiIntana et al. 2001). As mentioned above, aerobic chrome c including some high-valence metals, for instance,
Cr6+-reducing bacteria utilize reductases soluble in their V5+ and Mo6+. It is speculated on the basis of sequence
cytosols. Exceptionally, Pseudomonas maltophilia strain similarities with quinone reductases that bacterial ChrR and
O-2, Bacillus megaterium strain TKW3, and YieF might be equivalent to mammalian DT-diaphorase that
326 Folia Microbiol (2014) 59:321–332

Fig. 2 Schematic depiction of chromium resistance and toxicology in reductase which is encoded by chromosomal DNA reduces Cr6+ anaero-
bacterial cell: (1) Chromate due to the structural similarity with sulphate bically in the presence of electron donors. (5) Cr5+ produced during the
enters the bacterial cell through sulphate transporter encoded by the redox cycle of Cr6+ produces oxidative stress by the production of
chromosomal DNA. (2) Plasmid DNA encoded efflux systems are used reactive oxygen species (ROS). (6) To combat the ROS-generated oxi-
to expel the intracellular chromates outside the bacterial cell to resist the dative stress, protective metabolic enzymes, superoxide dismutase,
chromate toxicity. (3) Aerobic Cr6+ reduction into Cr3+ involves soluble catalase, and glutathione are secreted. Some outer membrane proteins
reductase which require NAD(P)H as an electron donor while anaerobic are also involved to counter the oxidative stress. (7) Cr6+ and principally
Cr6+ reduction occurs in the electron transport pathway by cytochrome b Cr3+ not only negatively affect DNA replication and RNA transcription
(cyt b) or cytochrome c (cyt c) along the respiratory chains in the inner by damaging DNA but also alter gene expression. In addition, Cr3+ also
membrane; Cr3+ cannot pass the bacterial cell membranes due to the damages proteins by impairing their functions. (8) DNA repair system is
insolubility of Cr3+ derivatives. (4) Membrane-embedded chromate activated in order to repair the damaged DNA

detoxify the quinonoid compounds. Further, both bacterial harveyi KCTC 2720 exhibiting Cr6+-reducing potential
Cr6+-reducing enzymes also provide protection against Cr6+ (Kwak et al. 2003) in addition to reducing nitrocompounds
toxicity, probably by diminishing the concentration of reactive and quinones (Ackerley et al. 2004a). The soluble homodi-
oxygen species (ROS) produced in response to other meta- meric FerB with non-covalently attached FAD in Paracoccus
bolic reactions. However, YieF-mediated Cr6+ reduction is denitrificans also exhibits high sequence similarity with the
considered far superior to that carried out by ChrR because Cr6+ reductase from P. putida and flavin reductases (Fre) from
of the minimal production of ROS (Park et al. 2000). In Pseudomonas syringae and has Cr6+ as well as quinone-
addition, a membrane-embedded Cr6+ reductase which uti- reducing capacity but cannot reduce free flavins (Mazoch
lizes NADH as an electron donor is also reported in et al. 2004; Sedláček and Kučera 2010). Rather than reducing
B. megaterium TKW3 (Cheung et al. 2006). Recently, Cr6+ Cr6+ directly, FerB protects the bacterial cells against the
reductase ChrR, like that of Pseudomonas putida, is also oxidative stress accompanying Cr6+ reduction (Sedláček and
reported in E. coli. This enzyme with bound FMN is actually Kučera 2010).
an obligatory two-electron quinone reductase which also re- Flavin reductase (Fre) from E. coli also reduces Cr6+ to a
duces Cr6+ species and belongs to the flavodoxin superfamily soluble Cr3+-NAD+ complex through highly reduced and
(Eswaramoorthy et al. 2012). unbound reactive flavins (Puzon et al. 2005). Moreover,
Furthermore, NAD(P)H-dependent Cr6+ reductase of nitroreductases (NfsA) and flavin reductases share a very
P. ambigua G-1 is also a homodimeric flavoprotein with close homology as demonstrated by Zenno et al. (1998) by
non-covalently bound FMN, which utilized NAD(P)H as transforming the NfsA from E. coli from a nitroreductase into
electron donor, generating a Cr5+ as an intermediate in reduc- a flavin reductase through a single amino acid substitution in
ing Cr6+ (Suzuki et al. 1992). As well, the gene encoding the the active site. As well, some NADPH-dependent
Cr6+ reductase of P. ambigua G-1 was 58 % homologous in flavoenzymes, for instance, lipoyl dehydrogenase, glutathione
nucleotide sequence to a nitroreductase (NfsA) of Vibrio reductase, and ferredoxin-NADP + oxidoreductase also
Folia Microbiol (2014) 59:321–332 327

Table 2 Chromium (Cr6+)-reducing bacteria and their isolating sites

Bacterial species Niche/redox condition Reference

Pseudomonas aeruginosa CRM100 Wastewater treatment plant/anaerobic Salamanca et al. (2013)

Pelosinus sp. strain HCF1 Aquifer sediment/anaerobic Beller et al. (2013)
Stenotrophomonas maltophilia ZA-6, Tannery effluent/aerobic Alam and Ahmad (2012)
Staphylococcus gallinarum W-61,
Pantoea sp. KS-2, Aeromonas sp. KS-14
Ochrobactrum intermedium Rb-2 Tannery effluent/aerobic Batool et al. (2012)
Clostridium sp. SS1 Activated sludge/aerobic Nguema and Luo (2012)
Bacillus sp. strain KSUCr9a, Salt lake/aerobic Ibrahim et al. (2012a),
Amphibacillus sp. KSUCr3 Ibrahim et al. (2012b)
Staphylococcus arlettae strain Cr11 Tannery effluent/aerobic Sagar et al. (2012)
Ochrobactrum intermedium BCR400 Soil/aerobic Kavita and Keharia (2012)
Bacillus sp. MRKV Tannery effluent/aerobic Chandhuru et al. (2012)
Acinetobacter junii VITSUKMW2 Chromite mine/aerobic Pulimi et al. (2012)
Acinetobacter sp. Cr-B2 Activated sludge/aerobic Narayani and Shetty (2012)
Bacillus sp. Tannery effluent/aerobic Smrithi and Usha (2012)
Arthrobacter viscosus –/aerobic Silva et al. (2012)
Pseudomonas sp. TIE2 Soil/aerobic Jayalakshmi and Rao (2012)
Planococcus maritimus VITP21 Marine water/aerobic Subramanian et al. (2012)
Bacillus sp. SUCR44, Microbacterium sp. Tannery effluent irrigated soil/aerobic Soni et al. (2013)
SUCR140, Bacillus thuringiensis SUCR186,
Bacillus subtilis SUCR188
Bacillus subtilis PESA Electroplating effluent/aerobic Tharannum et al. (2012)
Pseudomonas putida SKG-1 MTCC (1,050) -/aerobic Garg et al. (2012)
Pannonibacter phragmitetus Slag/aerobic Shi et al. (2012)
Leucobacter sp. G161 Soil/aerobic and anaerobic Ge et al. (2012)
Enterococcus gallinarum Soil/aerobic Sayel et al. (2012)
Pseudomonas putida strain KI, Serratia Tannery wastewater and sludge/aerobic Mahmood et al. (2012)
proteamaculans strain SL14
Staphylococcus aureus IFR-2, Tannery effluents/aerobic Ilias et al. (2011)
Pediococcus pentosaceus IFR-3
Bacillus sp. CSB-4 Chromite mine soils/aerobic Dhal et al. (2010)
Ochrobactrum sp. strain CSCr-3 Chromium landfill/aerobic He et al. (2009)
Bacillus sp. ev3 Wastewater/aerobic Rehman et al. (2008)
Thermus scotoductus strain SA-01 Gold mine/aerobic Opperman and van Heerden (2007),
Opperman and van Heerden (2008),
Opperman et al. (2008)
Bacillus sp. KCH2, Bacillus sp. Chromate contaminated soil/aerobic Sarangi and Krishnant (2008)
KCH3, Leucobacter sp. KCH4,
Exiguobacterium sp. KCH5
Exiguobacterium sp. ZM-2 Soil irrigated with tannery effluent/aerobic Alam and Malik (2008)
Brucella sp. Chromate contaminated sites/aerobic Thacker et al. (2007)
Cellulomonas spp. Subsurface soils/aerobic Viamajala et al. (2007)
Bacillus megaterium TKW3 Marine sediments/aerobic Cheung et al. (2006)
Bacillus sphaericus AND 303 Soil/aerobic Pal et al. (2005)
Serratia marcescens Tannery effluent/aerobic Mondaca et al. (2002)

catalyze Cr6+ reduction through consecutive one-electron nitroreductase activity but utilizes NAD(P)H as electron donor
transfers (Shi and Dalal 1990a; Shi and Dalal 1990b). Con- to reduce Cr6+. Recently, Opperman et al. (2008) have iden-
trary to Cr6+ reductases described above, cytoplasmic homo- tified a membrane-associated (Opperman and van Heerden
dimeric Cr6+ reductase, which was identified by Bae et al. 2008) homodimeric NAD(P)H-dependent Cr6+ reductase con-
(2005) in E. coli, neither exhibits flavin-cofactor nor has any taining a non-covalently bound flavin mononucleotide
328 Folia Microbiol (2014) 59:321–332

cofactor in Thermus scotoductus SA-01 isolated from African contaminated sites. In addition, expected results of bioreme-
gold mine. This oxidoreductase protein functions under both diation through Cr6+ reduction may not be achieved until the
aerobic and anaerobic conditions requires Ca2+ or Mg2+ for produced Cr 3+ species are precipitated or efficiently
catalysis. In addition, the catalytic efficiency of this Cr6+ immobilized as Cr3+ coordinating ligands on the bacterial
reductase was found to be 50 times greater than that of the surface have stronger affinity compared to those present on
quinone reductases and 180 times higher than that of the cell debris in media (Cheng et al. 2006). Developing new
nitroreductases. Sequencing of T. scotoductus revealed an desired traits in chromate-reducing bacteria by genetic engi-
open reading frame of 1,386 bp which was homologous neering to overcome the limitations in their detoxifying effi-
(84 %) to the dihydrolipoamide dehydrogenase gene of ciency would pave the way to make them more effective
Thermus thermophilus HB8 (Opperman and van Heerden bioremediating agents. For instance, by providing bacteria
2008). In another study, a homotetrameric and NADH- with a biochemical defence (e.g., engineering bacteria to
dependent Cr6+ reductase bound with flavin mononucleotide produce compatible solutes like trehalose) against the chro-
is reported in Gluconacetobacter hansenii which catalyzes the mate stress may be a prolific advantage in view of the toxic
reduction of chromate, ferricyanide, and uranyl anions under and damaging effects of high concentration of Cr6+ (Frederick
aerobic conditions (Jin et al. 2012). et al. 2013).
Bacterial Cr6+ reduction can either be plasmid borne as in
several Pseudomonas species or may be located on the chro-
mosomal DNA as in a number of Bacilli and Enterobacteria-
ceae members (Chirwa and Molokwane 2011). For instance,
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