Download Perspectives of plant-associated microbes in heavy metal

Biotechnology Advances 30 (2012) 1562–1574
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Biotechnology Advances
journal homepage: www.elsevier.com/locate/biotechadv
Research review paper
Perspectives of plant-associated microbes in heavy metal phytoremediation
M. Rajkumar a,⁎, S. Sandhya a, M.N.V. Prasad b, H. Freitas c
a
b
c
National Environmental Engineering Research Institute (NEERI), CSIR Complex, Taramani, Chennai 600113, India
Department of Plant Sciences, University of Hyderabad, Hyderabad 500046, India
Centre for Functional Ecology, Department of Life Sciences, University of Coimbra, Coimbra, 3000–455, Portugal
a r t i c l e
i n f o
Available online 9 May 2012
Keywords:
Phytoremediation
Heavy metals
Rhizosphere bacteria
Endophytic bacteria
Mycorrhizal fungi
Siderophores
Organic acids
Biosurfactants
1-aminocyclopropane-1-carboxylic acid
a b s t r a c t
"Phytoremediation" know-how to do-how is rapidly expanding and is being commercialized by harnessing
the phyto-microbial diversity. This technology employs biodiversity to remove/contain pollutants from the
air, soil and water. In recent years, there has been a considerable knowledge explosion in understanding
plant-microbes-heavy metals interactions. Novel applications of plant-associated microbes have opened
up promising areas of research in the field of phytoremediation technology. Various metabolites (e.g.,
1-aminocyclopropane-1-carboxylic acid deaminase, indole-3-acetic acid, siderophores, organic acids,
etc.) produced by plant-associated microbes (e.g., plant growth promoting bacteria, mycorrhizae) have
been proposed to be involved in many biogeochemical processes operating in the rhizosphere. The salient
functions include nutrient acquisition, cell elongation, metal detoxification and alleviation of biotic/abiotic
stress in plants. Rhizosphere microbes accelerate metal mobility, or immobilization. Plants and associated
microbes release inorganic and organic compounds possessing acidifying, chelating and/or reductive
power. These functions are implicated to play an essential role in plant metal uptake. Overall the plantassociated beneficial microbes enhance the efficiency of phytoremediation process directly by altering
the metal accumulation in plant tissues and indirectly by promoting the shoot and root biomass production.
The present work aims to provide a comprehensive review of some of the promising processes mediated by
plant-associated microbes and to illustrate how such processes influence heavy metal uptake through various
biogeochemical processes including translocation, transformation, chelation, immobilization, solubilization,
precipitation, volatilization and complexation of heavy metals ultimately facilitating phytoremediation.
© 2012 Elsevier Inc. All rights reserved.
Contents
1.
2.
Microbe assisted phytoremediation . . . . . . . . . . . . . . . . . . . .
Plant-associated microbes improve heavy metal mobilization/immobilization
2.1.
Siderophores . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.2.
Organic acids . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.3.
Biosurfactants . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.4.
Polymeric substances and glycoprotein . . . . . . . . . . . . . . .
2.5.
Metal reduction and oxidization . . . . . . . . . . . . . . . . . .
2.6.
Biosorption . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.
Plant growth promotion by plant-associated microbes . . . . . . . . . . .
4.
Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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1. Microbe assisted phytoremediation
⁎ Corresponding author. Tel.: + 91 44 22541964; fax: + 91 44 22541964.
E-mail address: [email protected] (M. Rajkumar).
0734-9750/$ – see front matter © 2012 Elsevier Inc. All rights reserved.
doi:10.1016/j.biotechadv.2012.04.011
Heavy metal contamination of soils has received considerable attention in the contemporary science. Application of biological processes
for decontaminating the contaminated/polluted sites is a challenging
task because heavy metals cannot be degraded and hence persist in
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
the soil (Kidd et al., 2009; Lebeau et al., 2008; Ma et al., 2011a; Rajkumar
et al., 2010). In order to cleanup the contaminated sites, heavy metals
should be extracted and concentrated by an appropriate technique for
proper disposal in designated secure landfill sites. The established conventional techniques (e.g., thermal processes, physical separation, electrochemical methods, washing, stabilization/solidification and burial)
for clean-up of metal contaminated soils are generally too expensive
and often harmful to soil microbial diversity (Dermont et al., 2008; Ma
et al., 1993; McGrath et al., 1995; Mulligan et al., 2001; Pulford and
Watson, 2003). Plant mediated decontamination/detoxification processes are commonly referred to as “phytoremediation”. It has been proposed as an alternative method to remove pollutants from air, soil and
water or to render pollutants harmless and does not affect soil biological
activity, structure and fertility (Raskin et al., 1997; Salt et al., 1998).
‘Phytoextraction’, is one of the key processes of phytoremediation that
involves the use of metal accumulating (hyperaccumulators) plants to
remove metals from soil by concentrating them in harvestable parts of
the plant. After a certain time period of growth, the plants are harvested
and disposed or incinerated. ‘Phytostabilization’ is the most successful
and well acknowledged process of phytoremediation, in which metal
tolerant plants arrest the leaching of heavy metals through the thick
mat of adventious roots and associated rhizosphere microbes.
The success of phytoremediation is dependent on the potential
of the plants to yield high biomass and withstand the metal stress.
Besides, the metal bioavailability in rhizosphere soil is considered to be
another critical factor that determines the efficiency of metal translocation and phytostabilization process (Ma et al., 2011a). In recent years,
several chemical amendments, such as EDTA, limestone have been
used to enhance either phytoextraction or phytostabilization process
(Barrutia et al., 2010; Wu et al., 2011b). Even though these amendments
increase the efficiency of phytoextracton/phytostabilization, some
chemical amendments (e.g., EDTA) are not only phytotoxic (Evangelou
et al., 2007) but also toxic to beneficial soil microorganisms that play important role in plant growth and development (Mühlbachová, 2009;
Ultra et al., 2005). A promising alternative to chemical amendments
could be the application of microbe-mediated processes, in which the
microbial metabolites/processes in the rhizosphere affect plant metal
uptake by altering the mobility and bioavailabity (Aafi et al., 2012;
Glick, 2010; Ma et al., 2011a; Miransari, 2011; Rajkumar et al., 2010;
Wenzel, 2009; Yang et al., 2012). When considering approaches to
alter heavy metal mobilization, there are several advantages to the
use of beneficial microbes rather than chemical amendments because
the microbial metabolites are biodegradable, less toxic, and it may be
possible to produce them in situ at rhizosphere soils. In addition, plant
growth promoting substances such as siderophores, plant growth
hormones, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase
produced by plant-associated microbes improve the growth of the
plant in metal contaminated soils (Babu and Reddy, 2011; Glick, 2010;
Glick et al., 2007; Kuffner et al., 2010; Lebeau et al., 2008; Luo et al.,
2011; Luo et al., 2012; Ma et al., 2011a,b; Miransari, 2011; Rajkumar et
al., 2010; Wang et al., 2011; Wu et al., 2006). Overall the microbial activities in the root/rhizosphere soils enhance the effectiveness of
phytoremediation processes in metal contaminated soil by two
complementary ways: (i) Direct promotion of phytoremedation in
which plant associated microbes enhance metal translocation (facilitate
phytoextraction) or reduce the mobility/availability of metal contaminants in the rhizosphere (phytostabilization) and (ii) Indirect promotion of phytoremediation in which the microbes confer plant metal
tolerance and/or enhance the plant biomass production in order to
remove/arrest the pollutants.
2. Plant-associated microbes improve heavy metal
mobilization/immobilization
Plants growing in metal contaminated soils harbor a diverse
group of microorganisms (Idris et al., 2004; Zarei et al., 2008, 2010)
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that are capable of tolerating high concentration of metals and providing a number of benefits to both the soil and the plant. Among
the microorganisms involved in heavy metal phytoremediation, the
rhizosphere bacteria deserve special attention because they can
directly improve the phytoremediation process by changing the
metal bioavailability through altering soil pH, release of chelators
(e.g., organic acids, siderophores), oxidation/reduction reactions (Gadd,
2000; Khan et al., 2009; Kidd et al., 2009; Ma et al., 2011a; Rajkumar
et al., 2010; Uroz et al., 2009; Wenzel, 2009). Similarly the metal tolerant
mycorrhizal fungi have also been frequently reported in hyperaccumulators growing in metal polluted soils indicating that these
fungi have evolved a heavy metal-tolerance and that they may play important role in the phytoremediation of the site (Gohre and Paszkowski,
2006; Miransari, 2011; Orłowska et al., 2011; Zarei et al., 2010) (Fig. 1).
Table 1 summarizes the published studies on the effects of microbial
metabolites/actions on heavy metal mobilization/immobilization and/
or its uptake by plants.
2.1. Siderophores
Most plant-associated bacteria and fungi can produce iron chelators called siderophores in response to low iron levels in the rhizosphere. Siderophores are low-molecular mass (400–1,000 Daltons)
compounds with high association constants for complexing iron, but
they can also form stable complexes with other metals, such as
Al, Cd, Cu, Ga, In, Pb and Zn (Glick and Bashan, 1997; Schalk et al.,
2011). Although siderophores contain other functional groups, they
are broadly classified into three main groups based on the chemical nature of the moieties donating the oxygen ligands for Fe(III) coordination,
which are either of the catecholates (enterobactin), hydroxamates
(desferrioxamines), or (α-hydroxy-)carboxylates (aerobactin). Since
siderophores solubilize unavailable forms of heavy metal bearing
minerals by complexation reaction, siderophores producing microbes
that inhabit the rhizosphere soils are believed to play an important
role in heavy metal phytoextraction (Braud et al., 2009b; Dimkpa
et al., 2009a,b; Rajkumar et al., 2010). An example is the production
of pyoverdin and pyochelin by rhizosphere bacteria Pseudomonas
aeruginosa, which increase the concentrations of bioavailable Cr and
Pb in the rhizosphere, thus making them available for maize uptake
(Braud et al., 2009b). Similarly, Dimkpa et al. (2009b) found that
the siderophores produced by Streptomyces tendae F4 significantly enhanced uptake of Cd by sunflower plant. The production of siderophores
has also been demonstrated in some mycorrhizal fungi (Goodell et al.,
1997; Machuca et al., 2007). Machuca et al. (2007) reported that the
ectomycorrhizal fungi (EMF), Scleroderma verrucosum, Suillus luteus
and Rhizopogon luteolus isolated from fruiting bodies of Pinus radiata
were shown to produce catecholates and hydroxamates siderophores
under iron deficient conditions. These studies suggest that by inoculating the plants with siderophore producing microbes, it should be
possible to improve heavy metal uptake in plants. However, the
mechanisms underlying the plant metal uptake through microbial
siderophore-mediated processess remain unknown. Moreover there
are some opposing viewpoints that the presence of siderophore producing microbes reduced the uptake of metals by the plants. For
instance, Sinha and Mukherjee (2008) reported that the inoculation
of siderophore producing P. aeruginosa strain KUCd1 reduced the
Cd uptake in roots and shoots of Cucurbita pepo and Brassica juncea.
Likewise, Tank and Saraf (2009) also observed that the inoculation
with Ni resistant-siderophore producing Pseudomonas increased the
plant growth and reduced Ni uptake in chickpea plants. Some studies
have also shown that siderophore-producing bacteria do not always
lead to an increase in metal uptake by plants (Kuffner et al., 2008,
2010). These contrasting effects may be due to the differences in the
ability of plants to take up heavy metals, which in turn will depend
on metal bioavailability, plant type and their ability to transport
metals from root to shoot. Moreover the plant root activities e.g., the
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M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
Phytoextraction
Phytostabilization
a
Siderophores
H+
Reduce soil pH
Sparingly soluble metals
Immobilized
metals
c
Cr3+
Reduce metal
uptake
Metal reduction
Improve translocation
Metal + chelator
complex
Reduce translocation
Soluble metals
Soluble
metals
b
Glomoline
Cr6+
EPS
Mycorrhization increase root exudation
Organic acids
Exudates
d
Biosurfactants
Exudates induce the rhizosphere
microbial activity
Rhizosphere bacteria
Endomycorrhizae
Ectomycorrhizae
Endophytic bacteria
Root
(Vertical section)
Metal biosorption reduces
plant metal uptake and/or
translocation
Metal polluted soils
Fig. 1. Plant-associated microbes accelerate the phytoremediation process in metal contaminated soils by enhancing metal mobilization/immobilization. (a) Plant-associated microbes
improve plant metal uptake by producing metal mobilizing chelators. Plant associated microbes reduce plant metal uptake and/or translocation through (b) producing metal
immobilizing metabolites, (c) metal reduction and/or (d) metal biosorption. Abbreviations: extracellular polymeric substances (EPS).
release of root exudates, which can be influenced by rhizosphere soil
properties, will affect soil pH and nutrient status and thus alter the diversity and activity of plant-associated microbes (Dakora and Phillips,
2002; Jones et al., 2003).
Although the microbial synthesis of siderophores and their role in
metal transport and tolerance are well documented (Schalk et al.,
2011), the interaction of plant–siderophore producing bacteria–metals
in polluted soils remains poorly understood. In general siderophore
production by microbes is regulated by various factors including
iron availability, pH, nutrient status of soils, type and concentration
of heavy metals, etc. For instance, Braud et al. (2009a) observed that
addition of heavy metals, Al, Cu, Ga, Mn and Ni in iron-limited succinate
medium induced pyoverdine production in the P. aeruginosa. Similarly,
the presence of Cu, Ni and Cr was found to increase pyoverdine production even in the presence of iron (Braud et al., 2010). Synthesis of
siderophores to decrease or increase the detrimental effects of metal
stress in bacteria has also been reported. For example, production of
the siderophores pyoverdine and pyochelin decreased the toxicity of
Al, Co, Cu, Ni, Pb and Zn to the P. aeruginosa (Braud et al., 2010) but
pyochelin increased the toxicity of vanadium to P. aeruginosa (Baysse
et al., 2000). In particular because siderophore producing microbes
depend on the properties of surrounding environment for their nutrition and since these microbes may modulate responses to various environmental factors, further understanding how the plant-associated
siderophore producing microbes influence the heavy metal mobilization and its uptake by plants in multi metal-polluted soils is critical.
2.2. Organic acids
Low molecular weight organic acids (LMWOAs) produced by
plant-associated microbes have received much attention over the
last decades, largely because of their proposed role in heavy metal
solubility and mobilization of mineral nutrients in the rhizosphere.
Organic acids are CHO containing compounds characterized by the
presence of one or more carboxyl groups with a maximum molecular
weight of 300 daltons (Jones, 1998). In general, organic acids can
bind metal ions in soil solution by complexation reaction. However,
the stability of the ligand:metal complexes is dependent on several
factors such as the nature of organic acids (number of carboxylic
groups and their position), the binding form of the heavy metals present as well as the pH of soil solution (Jones, 1998; Ryan et al., 2001).
For instance, a tricarboxylic acid, citrate binds metal ions more strongly
than dicarboxylic acids. Similarly the monocarboxylic acids are still
weaker than dicarboxylic acids. Among the numerous LMWOAs that
can be produced by rhizosphere microbes, gluconic acid, oxalic acid
and citric acid, have received increasing attention as potential compounds to improve the bioavailability of heavy metals. Several reviews
describe organic acid biosynthesis and excretion mechanisms in bacteria and fungi (Ramachandran et al., 2006; Sauer et al., 2008) and these
issues are not addressed in detail here.
Organic acids released by plant-associated microbes play an important role in the complexation of toxic and essential ions and increase
their mobility for plant uptake. A recent study by Saravanan et al.
(2007) demonstrated the Zn solubilizing potential of Gluconacetobacter
diazotrophicus strains under in vitro conditions with ZnO, ZnCO3 or
Zn3(PO4)2 and reported the production of a gluconic acid derivative, 5ketogluconic acid that aid in the solubilization of Zn compounds.
Similar studies on the influence of airborne bacteria isolated from a
tannery air environment on solubilizing insoluble ZnO or Zn3(PO4)2 revealed that the mobilization of Zn compounds strongly depended on the
production of 2-gluconic acid. They found that P. aeruginosa (CMG
823) was able to solubilize large amounts of both ZnO and Zn3(PO4)2
(Fasim et al., 2002). Experiment with rhizosphere bacteria of Cd/Zn
hyperaccumulating plant, Sedum alfredii, also revealed that the inoculation of soils with Cd/Zn resistant bacteria significantly increased the
water soluble Zn and Cd concentrations when compared with the uninoculated controls. In this case, enhanced heavy metal mobilization
could be correlated with the increased production of organic acids such
as formic acid, acetic acid, tartaric acid, succinic acid, and oxalic acid (Li
et al., 2010). Inoculation with Burkholderia caribensis FeGL03 that had
been isolated from Brazilian high-phosphorus iron ore has also been
studied in detail (Delvasto et al., 2009). This bacterium significantly mobilized P and Fe from crushed iron ore. Here, it was suggested that gluconic
acid production together with the production of exopolysaccharides and
the formation of biofilms accounted for the observed P and Fe mobilization from the ore. Wani et al. (2007a) also studied the mobilization of
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
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Table 1
Microbial metabolites/actions characterized for their potential to mobilize/immobilize metals and/or to alter the plant metal uptake.
Microbial metabolites/
reactions
Siderophores
Pyoverdine, pyochelin and
alcaligin E
Desferrioxamine B,
desferrioxamine E and
coelichelin
Pyoverdine
Not reported
Microorganisms
Origin of microorganisms
Microbial effects on metals and/or its
uptake by plants
Reference
Pseudomonas aeruginosa,
Pseudomonas
fluorescens and Ralstonia
metallidurans
Streptomyces tendae F4
Not reported
Enhanced Cr and Pb uptake by plants
through facilitating their mobilization
Braud et al. (2009b)
Uranium mine, Wismut in eastern
Thuringia, Germany
Enhanced Cd and Fe uptake by plants
through facilitating their mobilization
Dimkpa et al.
(2009b)
Pseudomonas putida KNP9
Panki Power Plant, Kanpur,India
Pseudomonas
Chickpea and tomato fields from
around Ahmedabad
Reduced Cd and Pb accumulation in
Phaseolus vulgaris
Reduced Ni accumulation in chickpea
plants
Tripathi et al.
(2005)
Tank and Saraf
(2009)
Organic acids
Oxalic acid, Tartaric acid,
Formic acid Acetic acid
Gluconic and 2-ketogluconic
acid
Not reported
Burkholderia cepacia
Hunan and Zhejiang Provinces, China.
Solubilized ZnO, ZnCO3 and CdCO3
Li et al. (2010)
Pseudomonas aeruginosa
(CMG 823)
Bacillus spp.
Solubilized ZnO and Zn3(PO4)2
Fasim et al. (2002)
Solubilized ZnO and PbCl2.
Wani et al. (2007a)
5-ketogluconic acid
Gluconacetobacter diazotrophicus
Solubilized ZnO, Zn3(PO4)2 and ZnCO3.
Saravanan et al.
(2007)
Not reported
Pseudomonas fluorescens G10
and Microbacterium sp. G16
Pseudomonas fluorescens Pf-5
Tannery air environment, Karachi,
Pakistan
Rhizospheric soils of mustard (Brassica
campestris L.) and tomato (Lycopersicum
esculentum L.)
Culture Collection of Tamil Nadu
Agricultural University, Coimbatore,
India
Tissues of rape roots, Nanjing, China
Increased water-soluble Pb in solution
and in Pb-added soil
Fe-oxide/hydroxide goethite (Goe-P)
Sheng et al.
(2008b)
Hoberg et al.
(2005)
Arwidsson et al.
(2010)
Citric acid
Oxalic acid (Aspergillus niger) A. niger, P. bilaiae,
and citric acid
(Penicillium bilaiae)
Malic and citric acid
Oidiodendron maius 091
Rhizosphere of Gossypium spp.
Solubilised Ni, Cu, Zn, and Pb from the
contaminated soils
Oxalic acid
Beauveria caledonica
Culture collections of Swedish
University of Agricultural Science
and Orebro University
V. angustifolium grown in unpolluted
site, Canada
Calluna vulgaris grown in unpolluted
site, Italy
Lawn soil, United Kingdom
Biosurfactants
Di-rhamnolipid
Pseudomonas aeruginosa BS2
Oily sludge
Mobilized Cdand Pb
Lipopeptide
Bacillus sp. J119
Heavy metal contaminated soils in
Nanjing, China
Rhamnolipids
Pseudomonas aeruginosa
Microbial Type Culture Collection,
Institute of microbial Technology,
Chandigarh, India
Increased in above-ground tissue Cd
content in tomato, maize and rape
plants.
Mobilized Cu
O. maius E
Polymeric substances and glycoprotein
Azotobacter spp.
Extracellular polymeric
substances or cell wall
lipopolysaccharides
Glomalin
Glomus mosseae
Oxidation and reduction reaction
Oxidation
Consortium of sulfur oxidizing
bacteria
Reduction
Cellulosimicrobium cellulans KUCr3
Martino et al.
(2003)
Fomina et al.
(2004)
Juwarkar et al.
(2007)
Sheng et al. (2008a)
Venkatesh and
Vedaraman
(2012)
Manganese mine spoil dump near
Gumgaon, India
Immobilized Cd and Cr and decreased
their uptake by Triticum aestivum.
Joshi and Juwarkar
(2009)
Sorghum
Immobilized Cu, Pb and Cd and
accumulated metals in a non-toxic
form to increase plant fitness and
soil quality.
Gonzalez-Chavez
et al. (2004)
Rhizosphere of rice
Increased bioavailability of Cu
Shi et al. (2011)
Industrial effluents nearby Kolkata,
India
Reduced the mobile and toxic
Cr(VI)to nontoxic and immobile
Cr(III) and decreased Cr uptake
by chilli plants
Reduced soluble and harmful Se(IV)
to insoluble and unavailable Se(0)
and thereby decreased the plant
Se uptake
Reduced As (VI) to As (III) and
enhanced As uptake by Pteris vittata
Chatterjee et al.
(2009)
Di Gregorio et al.
(2005)
Increased the mobility of Cu, Cd,
Hg and Zn
Beolchini et al.
(2009)
Reduction
Stenotrophomonas maltophilia
Reduction
Rhodococcus sp.TS1, Delftia sp.TS33, National Microbiology Laboratory of
Comamonas sp.TS37, Delftia sp.TS41, Huazhong Agricultural University
and Streptomyces lividans sp. PSQ22 (TS1, TS33, TS37, TS41) and Fudan
University (PSQ22)
Consortium of Fe-reducing and Fe/S Acid mine drainage, Bulgaria
oxidizing bacteria (Acidothiobacillus
thiooxidans, At. Ferroxidans,
Leptospirillum ferroxidans)
Oxidation and reduction
Solubilized Zn from both ZnO and
Zn3(PO4)2.
Solubilized Pb from pyromorphite and
accumulated the highest water-soluble
fraction and total Pb concentration in
the mycelium
Rhizosphere of Astragalus bisulcatus
Yang et al. (2012)
(continued on next page)
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Table 1 (continued)
Microbial metabolites/
reactions
Microorganisms
Origin of microorganisms
Microbial effects on metals and/or its
uptake by plants
Reference
Brevibacillus sp B-I
Aautochthonous strain.
Vivas et al. (2006)
Magnaporthe oryzae and
Burkholderia sp
Scleroderma citrinum, Amanita
muscaria and Lactarius rufus
Glomus mosseae
Rice tissues
Decreased the concentration of Zn in
shoot tissues of Trifolium repens
Reduced Ni and Cd uptake and their
translocation to shoots in tomato
Reduced translocation of Zn, Cd or Pb
from roots to shoots in pine seedlings
Reduced metal translocation from root
to shoot in Cajanus ca
Bioaccumulation
Serratia sp.MSMC541
Mycorrhizal roots of pines (Pinus sylvestris
L.)
Department of Microbiology, Indian
Agricultural Research Institute, New Delhi,
India
Nodules of Lupinus consentinii
Pb and Zn by inoculating three common metal resistant Bacillus strains,
PSB 1, PSB 7, and PSB 10 and found that the Bacillus sp. PSB1 solubilizing
high P (via pH reduction) was able to mobilize large amounts of both Pb
and Zn. The mycorrhizal fungi can also excrete organic acids into the soil
to mobilize heavy metals by complexing them and to acidify the rhizosphere. For instance, the ericoid mycorrhizal fungi Oidiodendron maius
have been reported to enhance Zn release from insoluble ZnO and
Zn3(PO4)2 via excretion of Zn chelating citric and malic acid (Martino et
al., 2003). This study also demonstrated a significant role of organic
acids in facilitating the release of adsorbed heavy metals into the soil. In
support of this, Fomina et al. (2005) studied the effect of soil fungi
Beauveria caledonica on solubilization of Zn3(PO4)2 and pyromorphite
and they found that organic acid accelerated Zn3(PO4)2 and pyromorphite dissolution through acidolysis (protonation) reaction. Recent
studies have also been shown that organic acids released by plant associated microbes facilitate plant root absorption of metal ions including
Cd and Zn (Li et al., 2010), Pb (Sheng et al., 2008b) and Cu (Chen et al.,
2005). The metal resistant endophytic bacteria, Pseudomonas fluorescens
G10 and Microbacterium sp. G16 have also been reported to enhance Pb
accumulation in rape via excretion of organic acid (Sheng et al., 2008b).
In yet other study Sayer et al. (1999) found that the organic acid producing fungi, Aspergillus niger was able to mobilize large amounts of Pb and
P from pyromorphite. Furthermore, they observed that A. niger significantly enhanced Pb and P uptake by Lolium perenne. These studies
highlighted the potential of organic acid producing microbes to improve
phytoextraction potential in metal contaminated soils.
Organic acids have been shown to play a role in the mechanisms
underlying heavy metal uptake by roots (Han et al., 2006; Panfili et al.,
2009). For instance, Han et al. (2006) demonstrated the role of organic
acids, acetic and malic acids in stimulating Cd uptake by maize roots and
reported that the organic acid with low stability constant was able to
enhance large amount of Cd accumulation in maize. Furthermore,
maize roots were able to dissociate Cd from Cd-organic acid complex
through root surface mediated process, and thereby led to increased
Cd uptake. Their results indicated that free Cd ions were more easily
available to maize roots than intact Cd-organic ligand complexes.
Panfili et al. (2009) reported a similar effect on the Cd uptake and
distribution among durum wheat roots in the presence of citric acid.
These results suggested that metal-organic acid complexes indirectly
contribute to the metal uptake through metal-complexes dissociation
within the diffusion layer and/or at the root surface thus increasing the
concentrations of free metal ions (Han et al., 2006; Panfili et al., 2009).
On the other hand, some studies show that organic acids can have
either no effect or can negatively affect heavy metal mobilization.
Braud et al. (2006) reported that the inoculation of organic acid producing bacteria Bacillus subtilis in metal contaminated agriculture
soils did not show any significant influence on the mobilization of
Cr and Pb. Park et al. (2011) reported that inoculation of organic
acid producing Pantoea sp. and Enterobacter sp. increased P solubilization and Pb immobilization in soil. This effect was due to an increase
in Pb solubilization and the subsequent immobilization of Pb by
Madhaiyan et al.
(2007)
Krupa and Kozdrój
(2007)
Garg and Aggarwal
(2011)
Reduced translocation of As, Cd, and Cu Aafi et al. (2012)
from roots to shoots in Lupinus luteus
P. Evangelou et al. (2006) revealed that the addition of LMWOAs
(citric, oxalic, and tartaric acid) had no enhancing effect in the uptake
of Pb into the tobacco, although LMWOA treatments showed higher
Pb mobilization capabilities in slurry and soil column experiments.
This effect was probably attributed to biodegradation and increased
rhizosphere soil pH due to the consumption of H + from carboxylic
acids and liberation of OH¯ and CO2. On the other hand several lines of
evidence suggest that the organic acids sorption could increase persistence against microbial degradation (Jones and Edwards, 1998) and
the continues production of organic acids by bacteria could also compensate for degradation in the rhizosphere (Geelhoed et al., 1999).
Taken together, these reports clearly suggest that the degree of metalorganic acid complexation depends on the type of organic acid involved
and the concentration, soil physicochemical properties and mineralogy.
Although many plant-associated microbes have the ability to produce organic acids and to mobilize essential and toxic ions (Fomina
et al., 2005; Martino et al., 2003; Uroz et al., 2009), an important
question that has yet to be adequately resolved is whether they act
as sources and/or sinks of organic acids in the soil solution. Laboratory
studies with soil microbes have explained to some extent, in which
the level of organic acid influx is directly regulated by the external
concentration (Jones et al., 1996). In general the rhizosphere soil
properties such as sorption, buffering capacity, biodegradation and
metal complexation may alter the profile of organic acids, making difficult to predict organic acids behavior (Jones et al., 2003). A detailed
characterization of the factors those control the fate and behavior of
organic acids in soil (e.g., heavy metal/nutrient-mobilization efficiency, concentrations required for metal mobilization, sorption to the
soil's solid phase and biodegradation), are key to identify the precise
mechanisms of microbial organic acids in the metal contaminated
rhizosphere soils. Moreover because the root mediated processes (e.g.,
the release of organic acids/phytosiderophores) could also facilitate
metal uptake by plants (Wenzel, 2009; Wenzel et al., 2003), other
factors affecting metal availability in the rhizosphere soils should also
be taken into account when assessing this issue: for instance, the role
of root-exudates in rhizosphere acidification/metal mobilization and
the contribution of various metabolites other than organic acids produced by plant-associated microbes in mobilizing toxic/essential ions
from sparingly soluble sources. No doubt, the accurate quantification
of organic acids in soils and the complete sequencing of organic acid
producing microbes combined with biomarker tools such as green fluorescent protein-based biosensors, will help address issues such as the
dynamics of organic acid transport between plants, rhizosphere soils
and microorganisms.
2.3. Biosurfactants
Another important metabolite that has potential to improve metal
mobilization and phytoremediation is microbially produced surfactants
(biosurfactants). Biosurfactants are amphiphilic molecules consisting
of a non polar (hydrophobic) tail and a polar/ionic (hydrophilic) head.
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
1567
A hydrophilic group consists of mono-, oligo- or polysaccharides, peptides or proteins and a hydrophobic moiety usually contains saturated,
unsaturated and hydroxylated fatty acids or fatty alcohols. In general
the biosurfactants produced by microbes form complexes with heavy
metals at the soil interface, desorbs metals from soil matrix and thus
increasing metal solubility and bioavailability in the soil solution. Interestingly, there is substantial evidence suggesting that the microorganism producing surfactants increase the heavy metal mobilization in
polluted soils (Juwarkar et al., 2007; Sheng et al., 2008a; Venkatesh and
Vedaraman, 2012). A recent study by Juwarkar et al. (2007), demonstrated the metal mobilization potential of P. aeruginosa BS2 under in vitro
column experiments and reported the production of a biosurfactant, dirhamnolipid that aid in the solubilization of Cd and Pb compounds
from artificially metal contaminated soil. Similarly Venkatesh and
Vedaraman (2012) assessed the potential of rhamnolipids produced by
P. aeruginosa to mobilize Cu in contaminated soils and found that 2%
rhamnolipids removed 71% and 74% of Cu from soil with initial concentrations of 474 and 4,484 mg kg− 1. Moreover, the biosurfactants
produced by plant-associated microorganisms also show promise for
enhancing metal uptake by plants; a desirable parameter for plants to
be used for phytoextraction. For instance Sheng et al. (2008a) assessed
the capability of the biosurfactant-producing bacterial strain Bacillus
sp. J119 to promote Cd uptake by rape, maize, sudangrass and tomato
in soil artificially contaminated with different levels of Cd (0 and
50 mg kg− 1). They observed that the inoculation of live bacterium
Bacillus sp. J119 to soils significantly increased the plant Cd uptake
when compared to dead bacterium-inoculated control. Although the
above studies have demonstrated significant role of microbial biosurfactants in facilitating the release of adsorbed heavy metals and in
enhancing the phytoextraction potential of plants, these observations
were mostly obtained from the plants grown in artificially metal contaminated soils and evidence supporting the biosurfactants enhance
the metal uptake by plant in metal-contaminated field soils is lacking.
Thus, knowing more about the interaction of biosurfactant producing
microbes and plants and their consequences will improve our understanding of the role of biosurfactant producing microbes in heavy
metal phytoremediation.
interesting from a phytoextraction point of view. For instance the
sulfur oxidizing rhizosphere bacteria have been reported to enhance
Cu mobilization in contaminated soils and its uptake in plant tissue
(Shi et al., 2011). This study showed that the sulfur oxidizing bacteria
reduce the rhizosphere soil pH via conversion of reduced sulfur to
sulfates, thus making Cu available for plant uptake. Similarly Chen
and Lin (2001) have also pointed out that Fe/S oxidizing bacteria
have the potential to enhance metal bioavailability in the soils through
acidification reaction.
Plant associated-microbes can also immobilize the heavy metals in
the rhizosphere through metal reduction reactions. For example,
Chatterjee et al. (2009) reported that the inoculation of Cr-resistant
bacteria Cellulosimicrobium cellulans to seeds of green chilli grown
in Cr(VI) contaminated soils decreased Cr uptake into the shoot by
37 and root by 56% compared with uninoculated controls. This
study indicates that bacteria reduced the mobile and toxic Cr(VI) to
nontoxic and immobile Cr(III) in the soil. According to Abou-Shanab
et al. (2007) the lower Cr translocation from root to shoots of water
hyacinth is indicative of a Cr reducing potential of rhizosphere
microbes. In a similar study Di Gregorio et al. (2005) demonstrated
the Se reducing potential of Stenotrophomonas maltophilia isolated
from the rhizosphere of Astragalus bisulcatus. They reported that this
bacterium significantly reduced soluble and harmful Se(IV) to insoluble
and unavailable Se(0) and thereby reducing the plant Se uptake. These
examples illustrate mechanisms, by which metal reducing microbes
immobilize metals within the rhizosphere soil and reflect the suitability
of these microbes for phytostabilization applications.
Besides, the synergistic interaction of metal oxidizing and reducing microbes on heavy metal mobilization in contaminated soils has
also been studied. Beolchini et al. (2009) reported the inoculation
of Fe-reducing bacteria and the Fe/S oxidizing bacteria together
significantly increased the mobility of Cu, Cd, Hg and Zn by 90% and
they attributed this effect to the coupled and synergistic metabolism
of oxidizing and reducing microbes. Though these results open
new perspectives for the bioremediation technology for metal mobilization, further investigations are needed to utilize such bacteria in
phytoextraction practices.
2.4. Polymeric substances and glycoprotein
2.6. Biosorption
The production of extracellular polymeric substances (EPS),
mucopolysaccarides and proteins by plant-associated microbes can
also play an important role in complexing toxic metals and in decreasing their mobility in the soils. For instance, Joshi and Juwarkar
(2009) investigated the immobilization of Cd and Cr after inoculation
of EPS producing Azotobacter spp. and found that these isolates were
able to bind 15.2 mg g - 1 of Cd and 21.9 mg g - 1 of Cr. Moreover they
found that at the addition of Azotobacter to the metal-contaminated
soils decreased Cd (−0.5) and Cr (− 0.4) uptake by Triticum aestivum.
Similarly Gonzalez-Chavez et al. (2004) assessed the ability of arbuscular
mycorrhizal fungi (AMF) produced insoluble glycoprotein, glomalin to
form complexes with heavy metals and found that up to 4.3 mg Cu,
1.1 mg Pb and 0.1 mg Cd per gram of glomalin could be extracted
from metal polluted soils. Since there is a correlation between the
amount of glomalin in the soil and the amount of heavy metal bound,
AMF strains with significant secretion of glomalin should be more suitable for phytostabilization efforts. From this study we know AMF can reduce the mobility of trace element by producing glomalin, but the
complete structure of glomalin and the mechanisms by which they influence plant metal uptake are unidentified.
The plant-associated microbes may also contribute in plant metal
uptake through biosorption mechanism. Biosorption can be defined
as the microbial adsorption of soluble/insoluble organic/inorganic
metals by a metabolism independent, passive, or by a metabolismdependent, active process (Ma et al., 2011a). Several authors have
pointed out that bacterial biosorption mechanism accounted for
reduced plant metal uptake. For instance, Madhaiyan et al. (2007) observed the inoculation of metal binding bacteria Magnaporthe oryzae
and Burkholderia sp reduced Ni and Cd accumulation in roots and
shoots of tomato. These effects of inoculation were reported also by
Vivas et al. (2006), who found that the inoculation of Trifolium repens
with Brevibacillus sp B-I decreased the concentration of Zn in shoot
tissues compared with respective un-inoculated control. This effect
was attributed to the increased Zn biosorption by Brevibacillus sp B-I.
These studies indicate that the metal binding bacteria can reduce the
metal bioavailability and/or restricts its entry into the plant root/shoot.
The mycorrhizal fungi can also act as a filtration barrier against
the translocation of heavy metals from plant roots to shoots. Experiments with pine seedlings revealed that the inoculation with
the EMF Scleroderma citrinum, Amanita muscaria and Lactarius rufus
reduced translocation of Zn, Cd or Pb from roots to shoots compared
with the controls. This effect was attributed to the increased
metal biosorption by outer and inner components of the mycelium
(Krupa and Kozdrój, 2007). Since mycorrhizal fungi have large
surface area, which endows mycorrhizal fungi with a strong capacity
at adsorbing heavy metals from soil. The fungal cell wall components
2.5. Metal reduction and oxidization
Certain plant-associated microorganisms have the potential to
alter the mobility of heavy metals through oxidation or reduction reactions. Metal oxidation by rhizosphere microbes is particularly
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
et al., 2011a; Miransari, 2011; Orłowska et al., 2011; Rajkumar et al.,
2010; Sheng et al., 2008a; Wu et al., 2011a) (Fig. 2). In turn, the plants
supply root borne nutrients including amino acids, sugars, organic
acids etc., which can be metabolized for microbial growth (Dakora
and Phillips, 2002). Since the success of the phytoremediation process
is also dependent on the plant's ability to withstand metal toxicity and
to yield adequate biomass, the plant growth promoting microbes have
attracted much attention from many researchers as rhizosphere/seed
inoculums (Aafi et al., 2012; Kuffner et al., 2010; Ma et al., 2010,
2011b; Maria et al., 2011; Rajkumar and Freitas, 2008; Sheng et al.,
2008a,b). Recent examples of the improved plant growth due to inoculation with plant-associated microbes under heavy metal stress are
summarized in Table 2.
Recent studies investigating the role of plant-associated microbes
in protection against heavy metal stress have demonstrated that the
bacterial colonization often results in increased nutrient uptake and
an increase in plant biomass (Dimkpa et al., 2008; Luo et al., 2011;
Luo et al., 2012; Ma et al., 2010; Maria et al., 2011; Mastretta et al.,
2009). It has been reported that the heavy metals often interfere
with the root uptake of nutrients such as Fe, P, Mg, Ca, Zn and with
metabolic functions of essential nutrients, leading to plant growth retardation (Ouzounidou et al., 2006; Parida et al., 2003). Under such
conditions the plant-associated microbes improve the plant nutrient
acquisition by mobilizing nutrients and making it available to plant
roots. An example is the P solubilizing bacteria, which dissolves
various sparingly soluble P sources such as Ca3(PO4)2 (Rodriguez
et al., 2004) and Zn3(PO4)2 (Saravanan et al., 2007) through lowering
pH of the rhizosphere soil and making P available for plant uptake.
The increased plant growth and P uptake have been reported on the
inoculations of P solubilizing Pseudomonas sp. in wheat (Babana
and Antoun, 2006) Pantoea J49 in peanut (Taurian et al., 2010), and
Psychrobacter sp. SRS8 in Ricinus communis and Helianthus annuus
(Ma et al., 2010). It is also demonstrated that the siderophores
produced by bacteria can solubilize insoluble Fe sources and induce
plant growth and iron uptake in plants (Dimkpa et al., 2009b; Ma
et al., 2010). Similarly mycorrhizal fungi may also improve nutritional
(e.g., chitin, extracellular slime, etc.) and intracellular compounds
(e.g., metallothioneins, P-rich amorphic material) may also immobilize/arrest the metals in the interior of plant roots (Meharg, 2003).
Although these studies suggest that inoculation of plants with
metal binding microbes could be a suitable approach for plant protection against heavy metals and phytostabilization of metal polluted
soils, several authors have pointed out that microbial biosorption/
bioaccumulation mechanism was not solely responsible for the
decreased metal accumulation and translocation in plants (Babu and
Reddy, 2011; Vivas et al., 2003).
All these reports clearly indicate that the plant-associated microorganisms differ in their capacity to alter heavy metal bioavailability
and its uptake by plants through metal mobilizing/immobilizing
metabolites/actions. However, survival and colonization potential of
these microbes greatly influence the quantity of metal accumulation
in plants growing in metal contaminated field soils, because adverse
physico-chemical-biological properties of soils including metal toxicity, indigenous microbial communities, adverse pH, nutrient deficiency
etc., reduce the survival, activity and colonization potential of inoculated microbes and thus potentially lead to alter the metal mobilization and/or immobilization. Since each polluted soil has a specific
profile, the potential of plants to uptake metals can vary to a large
extent, depending on which metal is involved, its concentration, the
microbial partner and their survival and colonization potential, plant
type and its growth conditions.
3. Plant growth promotion by plant-associated microbes
A small alteration in the physico-chemical-biological properties of
rhizosphere soils caused by biotic/abiotic stress can have a dramatic
effect on the plant-microbe interaction. The metal resistant-plantassociated microbes have been reported for potential to stimulate
the acquisition of plant nutrients, reduce metal toxicity, immobilize/
mobilize heavy metals in the soil, recycle the nutrients, improve
plant health and control plant pathogens (Aafi et al., 2012; Glick,
2010; Hayat et al., 2010; Kuffner et al., 2010; Lebeau et al., 2008; Ma
Root
(Transverse section)
Nutrient deficiency
Ethylene
Oxidative
ve
damagee
Atmospheric N2
P N Mg Ca
Less toxic
P + metal complex
Fe
Ca
P
H2O
P
b
Metal biosorption reduces
metal toxicity through
decreasing plant metal uptake
and/or translocation
Rhizosphere bacteria
Endomycorrhizae
Improve nutrient
uptake
P
Mycorrhization increase
root exudation
Fe
Metals
Exudates induce the
rhizosphere microbial
activity
Fungal
hyphae
c
Sparingly soluble
minerals
Siderophores
+ metals
Siderophores
Acidification/
chelation
e
P
Organic acids
Ammonia
& α KB
IAA
Organic acids+metals
Cr6+
Ectomycorrhizae
Endophytic bacteria
ACC
deaminase
Metals
P
Mycorrhization improve root
nutrient and water uptake
ACC
Improve cell elongation and proliferation
a
d
Increase
antioxidative
defense
Toxicity
Mg
ACC
ROS
Less toxic to plants
1568
Cr3+
Metal reduction
Rhizobium bacterioids
Heavy metal
Metal polluted soils
Fig. 2. Schematic overview of plant growth promotion by plant-associated microbes in metal contaminated soils. (a) Plant-associated microbes improve plant nutrients and water
uptake. Microbial metabolites reduce metal toxicity through (b) metal biosorption, (c) metal reduction and complexation reactions. Plant-associated microbes reduce heavy
metal stress in plants through (d) increasing antioxidative defense and/or producing ACC deaminese and (e) improve the plant growth by producing plant growth regulators.
Abbreviations: indole-3-acetic acid (IAA), reactive oxygen species (ROS), 1-aminocyclopropane-1-carboxylate (ACC), α ketobutyrate (α KB).
Table 2
Recent examples of plant growth promotion by plant-associated microbes.
Microorganisms
Microbial effects on plants under metal stress
Mechanisms
Cd stress reduced plants growth and induced
oxidative stress in S. nigrum
Metal stress (Fe, Mn, Zn, and Cd) reduced plant
growth and nutrient acquisition
Improved the growth of S. nigrum L under Cd stress
Improving the antioxidative defense in plants Gao et al. (2010)
Bacterial production of IAA, microbial metal
sequestration, improving plant nutrient
acquisition
Azcón et al. (2010)
Bacillus sp. SLS18
Sweet Sorghum,
P. acinosa, S. nigrum L
Glycine max
Cd stress decreased the aerial part dry weight
of sweet sorghum and S. nigrum L
Mn toxicity; Leaf puckering, necrotic spots, shorter
petioles, and browning of main veins and petioles
Microbial inoculation increased plant biomass,
concomitantly produced the highest symbiotic
(AMF colonisation and nodulation) rates. B. cereus
and AMF improved N, P, and K acquisition in plants.
Increased the aerial parts and root dry weights of all
three plants.
Facilitated plant growth and P uptake and protected
the plant from excessive uptake of Mn and Fe.
Luo et al. (2011)
G. etunicatum,
Calopogonium mucunoides
Reduced the plant growth, nutrient absorption
and ability to produce root nodules
Conferred Pb tolerance to plants, Promoted biomass
production nutrient uptake (P, S and Fe) and
nodulation of plants
Glomus mosseae
Cajanus cajan
Enhanced the number of nodules, their dry weights,
leghemoglobin content, and nitrogenase activity
AMF and Aspergillus
tubingensis
Dendrocalamus
strictus
Cd and Pb toxicity; reduced plant growth and
nodule formation, developed chlorosis and
necrosis, induced the production of PCs
Al and Fe in fly ash reduced plant growth and
nutrient acquisition
Producing IAA, siderophores, and ACC
deaminase
Stimulating the ATP-dependent sequestration
of Mn or Mn-chelates in the vacuoles, or
formation of low-solubility P–Mn complexes
Promoting plant nutrient acquisition,
attenuating the negative effects of Pb on
membranes and contributing to the reduction
of ROS generation
Biosorption (minimizing metal translocation
to the shoots) Dilution effects
Increased the P, K, Ca, and Mg in shoot tissues and
reduced plant Al (50%) and Fe content.
Babu and Reddy
(2011)
Mixture of Glomus clarum, Coffea arabica
Gigaspora margarita and
Acaulospora sp.,
Cellulosimicrobium
Green chilli
cellulans
Agrobacterium radiobacter Populus deltoides
Cu and Zn stress; reduced biomass accumulation
and P content,
Increased plant growth, P uptake, acted as a barrier
for metal translocation to the shoots.
Synergetic effect of A. tubingensis with AMF;
increasing plant nutrient acquisition; diluting
metal effect.
Element uptake by AMF extra-radical hyphae
and improving nutrient acquisition
Andrade et al.
(2010)
Cr(III) stress; reduced shoot length, root length
and the production of chlorophyll
Reduced plant growth
Promoted plant growth and reduced Cr uptake
Reducing Cr(VI) to Cr(III), producing IAA and
solubilizing P
Synthesizing IAA and siderophores
Chatterjee et al.
(2009)
Wang et al. (2011)
Pseudomonas sp.
Ni stress; Increased fresh and dry biomass
production in Alyssum serpyllifolium; Reduced
fresh and dry biomass production in Brassica juncea
Glomus etunicatum or
G. macrocarpum
Test plant
Alyssum serpyllifolium,
Brassica juncea
Enhanced plant height and dry weight of roots, stems
and leaves; Increased the contents of chlorophyll and
soluble sugar, and the activities of superoxide dismutase
and catalase; Decreased the content of malondialdehyde
Improved Ni accumulation in Alyssum serpyllifolium
Producing ACC deaminase, siderophores,
and, biomass production in Brassica juncea
IAA and solubilizing P.
Reference
Nogueira et al.
(2007)
de Souza et al.
(2012)
Garg and Aggarwal
(2011)
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
Metal stress on plants
Paecilomyces
lilacinus Solanum nigrum L
NH1
Bacillus cereus, Candida
Trifolium repens
Parapsilosis and AMF
Ma et al. (2011b)
1569
1570
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
state and growth of their host by acquiring phosphate, micronutrients
and water through the large surface area of their hyphae. Andrade
et al. (2010) recently assessed the influence of AMF on the growth
of coffee seedlings under Cu and Zn stress and found that mycorrhizal
coffee seedlings grew faster, exhibited improved mineral nutrition
(P and K) and had higher yields than non-mycorrhizal seedlings.
This may be due to the presence of extraradical mycelium, increasing
the root surface area through which soluble minerals particularly
P can be taken up. Experiments with Glycine max also revealed that
the inoculation with Glomus etunicatum or Glomus macrocarpum
significantly increased plant growth and P uptake without showing
any symptoms of Mn toxicity compared with the controls. Furthermore, higher P concentrations in plant tissues caused by AMF inoculation led to reduction in internal Mn toxicity through ATP-dependent
sequestration of Mn or formation of low-solubility P–Mn complexes
(Nogueira et al., 2007). Similarly the production of organic acids
such as oxalic acids by EMF (Ahonen-Jonnarth et al., 2000) has been
proposed to play a role in solubilization of P and increase its mobility
for plant uptake. The beneficial effects of mycorrhization on the acquisition of other nutrients, such as N, Ca, Mg, Mn, Cu and Zn, have also
been reported by researchers in various plants (Clark and Zeto,
2000; Orłowska et al., 2011; Smith and Read, 1997).
The symbiotic nitrogen fixing rhizobacterial genera such as
Rhizobium, Bradyrhizobium, Mesorhizobium, etc., also improve the
growth of legumes in metal contaminated soils by providing N to the
plants through N2 fixation (Rai et al., 2004; Wani et al., 2007b, 2008a).
For instance, Rai et al. (2004) reported that the inoculation of Prosopis
juliflora with a fly ash tolerant Rhizobium strain conferred tolerance
for the plant to grow under fly ash stress conditions with more translocation of metals to the above ground parts. Similarly the effects of N2
fixing Mesorhizobium strain RC3 on chickpea grown in Cr contaminated
soils (136 mg Cr kg- 1) was studied (Wani et al., 2008a). The bacterial
strain significantly promoted the dry matter accumulation, number of
nodules, shoot N, seed yield and grain protein by 71, 86, 40, 36 and
16%, respectively, compared to non inoculated plants. Further they
reported that N2 fixation together with Cr reduction and other plant
growth promoting substance (indole-3-acetic acid (IAA), siderophores)
accounted for increased growth of the plants in Cr polluted soils. In
general the phytohormones (e.g., auxin, gibberellins) produced by
rhizosphere microbe enhance the plant growth through directly promoting cell elongation, cell division, root initiation, and/or altering the
expression of specific genes (Davies, 2010; Malhotra and Srivastava,
2009; Taghavi et al., 2009). Since the above ground biomass production
in plants is directly proportional to the total metal extraction from
soils, improvement of biomass production is considered as important
strategy to enhance heavy phytoextraction. An increased metal extraction as a result of increased biomass production in B. juncea, Brassica
napus, Sorghum bicolor, Solanum nigrum was found by the inoculation
of plant-associated microbes (Chen et al., 2010; Dell'Amico et al.,
2008; Luo et al., 2012; Rajkumar and Freitas, 2008; Wu et al., 2006).
In addition to improving plant's nutrient uptake and growth, the
plant-associated microbes alleviate heavy metal toxicity by reducing
stress ethylene production. In general heavy metal stress induces
endogenous ethylene production in plants, which can affect the
root growth and consequently the growth of the whole plant. The
rhizosphere bacteria contain the enzyme, ACC deaminase reduces
the stress ethylene production in plants through metabolizing the
ethylene precursor, ACC into α ketobutyrate and ammonia (Glick et
al., 2007). Under such condition, in order to maintain the equilibrium
between the rhizosphere and root interior ACC levels, the plants
release more ACC through exudation and thus results decrease in the
production of stress ethylene (Adams and Yang, 1979). Recent studies
have revealed that plants inoculated with rhizosphere bacteria
containing ACC were better able to thrive in metal polluted soils
(Rodriguez et al., 2008). Madhaiyan et al. (2007) reported that M. oryzae
strain CBMB20 having ACC deaminase activity increased the growth of
tomato seedlings grown in Ni and Cd polluted soils. The bacterium reduced the production of ethylene, which was otherwise stimulated
when seedlings were challenged with increasing Ni and Cd. Very recently, Zhang et al. (2011) have also confirmed that Pb-resistant and
ACC deaminase-producing endophytic bacteria conferred metal tolerance onto plants by lowering the synthesis of metal-induced stress ethylene and promoted the growth of rape. We have also observed similar
results in the case of Allysusm serpyllifolium and B. juncea growth under
Ni stress in response to inoculation with ACC deaminase producing endophytic bacteria (Ma et al., 2011b).
Microbial production of metal chelating substance such as organic
acids, siderophores, etc., is of special significance because of the
metal complexing properties. These metal chelating substances play
pivotal roles in reducing the detrimental effects of heavy metals and
enhancing the biomass production in plants. Kavita et al. (2008)
demonstrated the potential of a gluconic acid producing Enterobacter
asburiae PSI3 to protect mung bean seedlings from the toxicity of
Cd in high concentrations and reported this effect may due to the
binding of metal ion with the acid ion. Since the organic acids in rhizosphere soil may form complexes with heavy metals and inactivating
and minimizing the cytological impacts of free metal ions (Gao et al.,
2010), the heavy metal:organic acid complex is considered as less
phytotoxic than the free form of heavy metals (Najeeb et al., 2009).
It is also reported that the precipitation of metal oxalates in intercellular spaces of the AMF can reduce metal availability and toxicity
to the host plant. In addition, the mechanisms such as sorption of
metal in fungal cell wall components (chitin, melanin) (Krupa and
Kozdrój, 2007), accumulation of heavy metals in vacuoles of mycorhizal fungi have implications in reducing the toxic effects of heavy
metals in plants (González-Guerrero et al., 2008; Leyval et al., 1997).
The mycorrhizal fungi may also contribute in reducing the metal
phytotoxicity through ‘dilution effect’ (Chen et al., 2007). Orłowska
et al. (2011) have pointed out AMF biosorption/bioaccumulation
mechanism, together with their ability to improve plant mineral nutrition (P, K, Fe, Zn, Mn and Ca) accounted for improved plant growth
and reduced Ni concentrations in aboveground tissues of Berkheya
coddii. They attributed the alleviation of metal toxicity in mycorrhizal
plants to the stronger dilution effect, thus the reducing Ni concentration in plant tissues.
The effects of dual inoculation of mycorrhizal fungi with other
microbes, such as P solubilizing microbes, N fixing bacteria on plant
growth and nutrients uptake has been studied in details. Babu and
Reddy (2011) reported the dual inoculation of AMF along with the
P solubilizing Aspergillus tubingensis significantly increased plant
growth and P, K, Ca, Mg and Na uptake by Dendrocalamus strictus in
fly ash. Similarly Arriagada et al. (2004) found that the combined
inoculation of AMF Glomus deserticola and saprobe fungi Trichoderma
koningii increased the tolerance of eucalyptus to the application of
50 mg l –1 of Cd. Furthermore, the saprobe fungi led to a reduction in
Cd toxicity on AMF through biosorption, thereby increasing AM colonization of plants and enabling them to fulfill their beneficial function
on plant growth. Similarly the rhizosphere bacteria have also been
shown to increase plant growth and heavy metal tolerance of their
host, especially in combination with mycorrhizal fungi (Azcón et al.,
2010). Kozdroj et al. (2007) found that the co-inoculation of EMF
and EMF associated bacteria (Pseudomonas putida or Bacillus cereus)
stimulated the growth of pine seedlings in the presence of Cd(II).
The facts that P. putida or B. cereus can absorb and immobilize
Cd and that can protect EMF against heavy metals may explain
that the combined inoculation of EMF and EMF associated bacteria
increased the tolerance of pine seedling to Cd stress. Similar effects
on plant growth and nutrient uptake have been observed by coinoculation of various microbes such as EMF (S. citrinum, A. muscaria
and L. rufus) with Pseudomonas (Krupa and Kozdrój, 2007), AMF
with B. cereus and Candida parapsilosis (Azcón et al., 2010), Hebeloma
mesophaeum with B. cereus (Hrynkiewicz et al., 2012) Agromyces sp
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
with Streptomyces sp., and Cadophora finlandica (Maria et al., 2011).
These studies suggest that colonization of plant roots by MF improves
the plant growth, health, and productivity in metal polluted soils
by altering the activity and abundance of rhizosphere microbiota or
vice versa.
A large number of studies also confirm the existence of cumulative
effects of microbes such as solubilization of nutrients, production
of phytohormones, alleviation stress ethylene production, metal
biosorption, etc., (Kuffner et al., 2010; Luo et al., 2011; Luo et al.,
2012; Ma et al., 2011b; Orłowska et al., 2011; Rajkumar and Freitas,
2008; Sheng et al., 2008a,b; Wani et al., 2008b; Zhang et al., 2011).
For instance experiments with Cd-hyperaccumulator S. nigrum
revealed that the inoculation with heavy metal-resistant bacterial
endophytes significantly increased plant growth without showing
symptoms of Cd toxicity compared with the controls (Luo et al.,
2011). This effect was attributed to the cumulative effects of endophytes, such as the production of ACC deaminase, siderophores, phytohormones and solubilization of P. These studies suggest that the
inoculation of rhizosphere microbes possessing the ability to withstand
heavy metal stress and the potential to promote plant growth through
various plant growth promoting traits in metal-contaminated soils can
be a potential biotechnological tool for successful phytoremediation
process.
Heavy metals induce oxidative damage in plant cells by generating
reactive oxygen species (ROS) such as superoxide radical (O2-), hydrogen peroxide (H2O2), hydroxyl radical (HO), and singlet oxygen
( 1O2). These ROS can react with nucleic acids, proteins, lipids and
amino acids and in the absence of the protective mechanism; they
can lead irreparable metabolic dysfunction, damage cell structure
and function (Gill and Tuteja, 2010; Gratao et al., 2005; Møller et al.,
2007). In general, plants have mechanisms to protect themselves
from oxidative damage involving antioxidant molecules and enzymes
(Jiang and Zhang, 2002; Yamane et al., 2004). It has been reported
that plants with high concentrations of antioxidants have greater
resistance to oxidative damage (Boojar and Goodarzi, 2007; Gill
and Tuteja, 2010). Using Chenopodium ambrosioides, it has been
shown that Cu accumulation is accompanied by a rapid induction
of antioxidative enzyme activities, superoxide dismutase (SOD) and
catalase (CAT) and found that these enzymes were able to protect
proteins, chlorophyll and lipids of some parts of plants against ROS
attack (Boojar and Goodarzi, 2007). This study demonstrated a significant role of antioxidative enzymes in defense system against oxidative damages in plant and protecting it against toxicity by Cu.
Recent studies also suggested that rhizosphere/seed inoculation
with beneficial microbes helps plants to alleviate heavy metal stress
through enhancing the activities of antioxidant enzymes (Kavita
et al., 2008; Ma et al., 2010; Wang et al., 2011; Wani et al., 2008b).
For instance, Ma et al. (2010) have shown higher activities of antioxidant
enzymes, CAT and peroxidase (POX) in bacterial inoculated R. communis
and H. annuus grown in soils treated with Ni. Similarly Wang et al.
(2011) recently assessed the effect of inoculating an As-resistant and
plant growth–promoting rhizobacterium on As phytoremediation by
Populus deltoides LH05-17 and found Agrobacterium radiobacter significantly increased the plant height, dry weight of roots, stems and leaves,
contents of chlorophyll and soluble sugar, and the activities of SOD
and CAT. Yet in another study, when pea plants inoculated with Niand Zn-tolerant plant growth promoting Rhizobium sp. RP5 grown
in Ni and Zn polluted soils, the bioinoculant significantly enhanced
glutathione reductase (GR) activity in the roots and nodules which detoxifies the H2O2 via the ascorbate-glutathione cycle (Wani et al., 2008b)
The mycorrhizal fungi can also affect physiological and biochemical
basis of plant tolerance to heavy metals by changing the antioxidant
enzyme activities. Azcón et al. (2010) recently studied the effects of
autochthonous microorganisms (AMF and/or plant growth promoting
bacteria) on the antioxidant activities of plants growing in a heavymetal multi-contaminated soil. They found that AMF inoculation
1571
significantly enhanced CAT, ascorbate peroxidase, or GR activities and
helped plants to limit oxidative damage to biomolecules in response
to metal stress. Garg and Aggarwal (2012) also reported that the growth
of AMF inoculated Cajanus cajan were more tolerant to high soil Cd and
Pb contents than non inoculated plants with significant increases in the
activities of SOD, CAT, POX as well as GR. Yet in another study, when
Calopogonium mucunoides plants inoculated with metal tolerant G.
etunicatum was grown in soils exposed to different levels Pb, the
bioinoculant significantly enhanced the plant growth and reduced the
oxidative stress indicator, malondialdehyde (de Souza et al., 2012).
This study shows that AMF can alleviate the damage on the cell membrane caused by Pb stress. Besides, altered regulation of expression
of genes with products putatively involved in the response to heavy
metal stress has also been reported in various plants as a result of
AMF inoculations (Cicatelli et al., 2010; Ouziad et al., 2005). Cicatelli
et al. (2010) observed the improved tolerance of the mycorrhizal
white poplar cuttings to heavy metal stress was associated with higher
expression of metallothioneins and polyamines (membrane stabilizers
and free radical scavenger) biosynthetic genes (PaADC, PaSPDS1 and
PaSPDS2). This study suggests that metallothionin and polyamines
may afford protection against heavy metal-induced stress in plants.
Ouziad et al. (2005) also reported that tomato plants grown on Zn
rich “Breinigerberg” soil and artificially inoculated with AMF Glomus
intraradices showed improved growth relative to uninoculated controls,
reduced the expression of Lemt2 (coding for metallothioneins) and
LeNramp1 (for heavy metal transporter) genes. Thus, the decrease in
the expression of these genes was suggested to be possibly due to the
overall reduction in the toxicity of metals, or reduced metal accumulation in plant tissues caused by mycorrhization to a level insufficient to
induce the expression of these two genes.
Taken together, these reports clearly indicate that the plantassociated microbes with various plant beneficial traits promotes the
plant growth in metal contaminated soils and suggest the potential
usefulness of these microbe inoculation for alleviating metal toxicity
and at the same time improving plant growth and phytoremediation
process. However, research on the effects of co-inoculation of beneficial microbes including beneficial bacteria and mycorrhizal fungi
on heavy metal phytoremediation is not extensive, thus further investigation is needed to better understand the prospects of synergetic
interactions of various plant-associated microbes in such strategies.
4. Conclusion
Since the plant-associated microbes possess the capability of plant
growth promotion and/or metal mobilization/immobilization, there
has been increasing interest in the possibility of manipulating plantmicrobe interactions in metal contaminated soils (Aafi et al., 2012;
Azcón et al., 2010; Braud et al., 2009b; Dimkpa et al., 2008, 2009a,b;
Hrynkiewicz et al., 2012; Kuffner et al., 2010; Luo et al., 2011; Luo et
al., 2012; Maria et al., 2011; Mastretta et al., 2009; Orłowska et al.,
2011; Sheng et al., 2008a,b). Microbial metabolites/processes promote
plant growth and metal mobilization/immobilization in vitro, but are
unable to confer beneficial traits on their host in metal contaminated
soils. Moreover the isolation of various plants associated microbes and
characterization of its beneficial metabolites/processes are time consuming since it requires the analysis of more than thousands of isolates.
Thus strong molecular research effort is required in order to find specific biomarker associated with the beneficial microbes for efficient microbe assisted phytoremediation.
Although promising results have been reported under laboratory
conditions, showing that inoculation of beneficial microbes particularly plant growth promoting bacteria and/or mycorrhizae may stimulate heavy phytoextraction or phytostabilization, only a few studies
have demonstrated the effectiveness of the microbial assisted heavy
metal phytoremediation in field conditions (Brunetti et al., 2011;
Juwarkar and Jambhulkar, 2008; Wu et al., 2011a; Yang et al., 2012).
1572
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
Since the biotic/abiotic stress in multi-metal polluted field soils
greatly influences the activity, composition and function of the inoculated microbes, the microbial mediated processes may be dependent on the nutrient composition and properties of rhizosphere
soils. Moreover, various stimuli in the rhizosphere could be also
associated with production of metabolites (e.g., siderophores, organic
acids) including nutrient deficiency (P, Fe) and exposure to toxic
metals. Thus characterizing the physico-chemical-biological features
of target contaminated soils may be important for making microbeassisted phytoremediation processes successful.
Since the activity of inoculated microbes is necessary to exhibit
beneficial traits for improving the plant growth and overall
phytoremediation process in metal contaminated soils, the colonization and survival in metal stress field environment are considered as
important factors. Thus advancing the knowledge on multiple metal
resistance, survival and compatibility of microbes, may be important
to utilize their potential as inoculants for phytoremediation purpose.
Although significant advances have been made in understanding
the roles of plant associated microbes in metal mobilization/
immoblization and in the application of these processes in heavy
metal phytoremediation (Braud et al., 2009b; Joshi and Juwarkar,
2009; Kuffner et al., 2010; Li et al., 2010; Sheng et al., 2008a; Shi
et al., 2011), additional advances are expected. For example, complete
genome sequences for several environmentally relevant microorganisms, mechanism of microbial chelators-metal complex uptake
in plants, factors influencing the solubility and plant availability of nutrients/heavy metals, signaling processes that occur between plant
roots and microbes, these types of analysis will surely prove useful
for exploring the mechanism of metal-microbes-plant interactions.
Moreover, such knowledge may enable us to improve the performance
and use of beneficial microbes as inoculants for microbial assisted
phytoremediation. We anticipate that manipulating the rhizosphere
processes for example increasing rhizosphere microbial population,
inoculating the microbial strains with various plant growth promoting
features as well as coinoculating ecologically diverse microbes would
yield better results for effective phytoremediation.
Acknowledgements
M. R acknowledges the financial support received in the form
of Ramalingaswami re-entry fellowship from Department of Biotechnology (DBT), Government of India. M. R also acknowledges the kind
support and encouragement extended by Dr. S. R. Wate, Director,
NEERI, Nagpur. Authors are grateful to anonymous referees and editor
for critical reading and improvement of the manuscript.
References
Aafi NE, Brhada F, Dary M, Maltouf AF, Pajuelo E. Rhizostabilization of metals in soils
using Lupinus luteus inoculated with the metal resistant rhizobacterium Serratia
sp. MSMC 541. Int J Phytoremediation 2012;14:261–74.
Abou-Shanab RA, Angle JS, van Berkum P. Chromate-tolerant bacteria for enhanced metal
uptake by Eichhornia crassipes (Mart.). Int J Phytoremediation 2007;9:91-105.
Adams DO, Yang SF. Ethylene biosynthesis: identification of l-aminocyclopropanecarboxylic
acid as an intermediate in the conversion of methionine to ethylene. Proc Natl Acad Sci
U S A 1979;76:170–4.
Ahonen-Jonnarth U, van Hees PAW, Lundström US, Finlay RD. Production of organic
acids by mycorrhizal and nonmycorrhizal Pinus sylvestris L. seedlings exposed to
elevated concentrations of aluminium and heavy metals. New Phytol 2000;146:
557–67.
Andrade SA, Silveira AP, Mazzafera P. Arbuscular mycorrhiza alters metal uptake and
the physiological response of Coffea arabica seedlings to increasing Zn and Cu
concentrations in soil. Sci Total Environ 2010;208:5381–91.
Arriagada CA, Herrera MA, Garcia-romera I, Ocampo JA. Tolerance to Cd of soybean
(Glycine max) and eucalyptus (Eucalyptus globulus) inoculated with arbuscular
mycorrhizal and saprobe fungi. Symbiosis 2004;36:285–99.
Arwidsson Z, Johansson E, von Kronhelm T, Allard B, van Hees P. Remediation of metal
contaminated soil by organic metabolites from fungi—production of organic acids.
Water Air Soil Pollut 2010;205:215–26.
Azcón R, Perálvarez MDC, Roldán A, Barea JM. Arbuscular mycorrhizal fungi, Bacillus
cereus, and Candida parapsilosis from a multicontaminated soil alleviate metal
toxicity in plants. Microb Ecol 2010;59:668–77.
Babana AH, Antoun H. Effect of Tilemsi phosphate rock-solubilizing microorganisms
on phosphorus uptake and yield of field grown wheat (Triticum aestivum L.) in Mali.
Plant Soil 2006;28:51–8.
Babu AG, Reddy S. Dual inoculation of arbuscular mycorrhizal and phosphate solubilizing
fungi contributes in sustainable maintenance of plant health in fly ash ponds. Water
Air Soil Pollut 2011;219:3-10.
Barrutia O, Garbisu C, Hernandez-Allica J, Garcıa-Plazaola JI, Becerril JM. Differences in
EDTA-assisted metal phytoextraction between metallicolous and non-metallicolous
accessions of Rumex acetosa L. Environ Pollut 2010;158:1710–5.
Baysse C, Vos DD, Naudet Y, Vandermonde A, Ochsner U, Meyer JM, et al. Vanadium
interferes with siderophore mediated iron uptake in Pseudomonas aeruginosa.
Microbiol-SGM 2000;146:2425–34.
Beolchini F, Dell'Anno A, Propris LD, Ubaldini S, Cerrone F, Danovaro R. Auto- and
heterotrophic acidophilic bacteria enhance the bioremediation efficiency of sediments contaminated by heavy metals. Chemosphere 2009;74:1321–6.
Boojar MMA, Goodarzi F. The copper tolerance strategies and the role of antioxidative
enzymes in three plant species grown on copper mine. Chemosphere 2007;67:
2138–47.
Braud A, Jézéquel K, Vieille E, Tritter A, Lebeau T. Changes in extractability of Cr and Pb
in a polycontaminated soil after bioaugmentation with microbial producers of biosurfactants, organic acids and siderophores. Water Air Soil Pollut 2006;6:261–79.
Braud A, Hannauer M, Milsin GLA, Schalk IJ. The Pseudomonas aeruginosa pyochelin-iron
uptake pathway and its metal specificity. J Bacteriol 2009a;191:5317–25.
Braud A, Jézéquel K, Bazot S, Lebeau T. Enhanced phytoextraction of an agricultural Cr,
Hg- and Pb-contaminated soil by bioaugmentation with siderophore producing
bacteria. Chemosphere 2009b;74:280–6.
Braud A, Geoffroy V, Hoegy F, Mislin GLA, Schalk IJ. The siderophores pyoverdine
and pyochelin are involved in Pseudomonas aeruginosa resistence against metals:
Another biological function of these two siderophores. Environ Microbiol Report
2010;2:419–25.
Brunetti G, Farrag K, Rovira PS, Nigro F, Senesi N. Greenhouse and field studies on Cr,
Cu, Pb and Zn phytoextraction by Brassica napus from contaminated soils in the
Apulia region, Southern Italy. Geoderma 2011;160:517–23.
Chatterjee S, Sau GB, Mukherjee SK. Plant growth promotion by a hexavalent chromium
reducing bacterial strain, Cellulosimicrobium cellulans KUCr3. World J Microbiol
Biotechnol 2009;25:1829–36.
Chen SY, Lin JG. Effect of substrate concentration on bioleaching of metal-contaminated
sediment. J Hazard Mater 2001;82:77–89.
Chen YX, Wang YP, Lin Q, Luo YM. Effect of copper-tolerant rhizosphere bacteria on
mobility of copper in soil and copper accumulation by Elsholtzia splendens. Environ
Int 2005;31:861–6.
Chen BD, Zhu YG, Duan J, Xiao XY, Smith SE. Effects of the arbuscular mycorrhizal fungus Glomus mosseae on growth and metal uptake by four plant species in copper
mine tailings. Environ Pollut 2007;147:374–80.
Chen L, Luo S, Xiao X, Guo H, Chen J, Wan Y, et al. Application of plant growth-promoting
endophytes (PGPE) isolated from Solanum nigrum L. for phytoextraction of
Cd-polluted soils. Appl Soil Ecol 2010;46:383–9.
Cicatelli A, Lingua G, Todechini V, Biondi S, Torrigiani P, Castiglione S. Arbuscular mycorrhizal fungi restore normal growth in a white poplar clone grown on heavy metalcontaminated soil, and this is associated with up regulation of foliar metallothionein
and polyamine biosynthetic gene expression. Ann Bot (Lond) 2010;106:791–802.
Clark RB, Zeto SK. Mineral acquisition by arbuscular mycorrhizal plants. J Plant Nutr
2000;23:867–902.
Dakora FD, Phillips DA. Root exudates as mediators of mineral acquisition in
low-nutrient environments. Plant Soil 2002;245:35–47.
Davies PJ. The Plant Hormones: Their nature, occurrence and functions. 3rd ed. New
York: Kluwer Academic Publishers; 2010. p. 656.
de Souza LA, de Andrade SAL, De Souza SCR, Schiavinato MA. Arbuscular mycorrhiza confers Pb tolerance in Calopogonium mucunoides. Acta Physiol Plant 2012;34:523–31.
Dell'Amico E, Cavalca L, Andreoni V. Improvement of Brassica napus growth under
cadmium stress by cadmium-resistant rhizobacteria. Soil Biol Biochem 2008;40:
74–84.
Delvasto P, Ballester A, Muñoz JA, González F, Blázquez ML, Igual JM, et al. Mobilization
of phosphorus from iron ore by the bacterium Burkholderia caribensis FeGL03.
Miner Eng 2009;22:1–9.
Dermont G, Bergeron M, Mercier G, Richer-Laflèche M. Soil washing for metal removal:
A review of physical/chemical technologies and field applications. J Hazard Mater
2008;152:1-31.
Di Gregorio S, Lampis S, Vallini G. Selenite precipitation by a rhizospheric strain of
Stenotrophomonas sp isolated from the root system of Astragalus bisulcatus: a biotechnological perspective. Environ Int 2005;31:233–41.
Dimkpa CO, Svatos A, Merten D, Büchel G, Kothe E. Hydroxamate siderophores produced by Streptomyces acidiscabies E13 bind nickel and promote growth in cowpea
(Vigna unguiculata L.) under nickel stress. Can J Microbiol 2008;54:163–72.
Dimkpa CO, Merten D, Svatoš A, Büchel G, Kothe E. Siderophores mediate reduced and
increased uptake of cadmium by Streptomyces tendae F4 and sunflower (Helianthus
annuus), respectively. J Appl Microbiol 2009a;107:1687–96.
Dimkpa CO, Merten D, Svatoš A, Büchel G, Kothe E. Metal-induced oxidative stress
impacting plant growth in contaminated soil is alleviated by microbial siderophores. Soil Biol Biochem 2009b;41:154–62.
Evangelou MWH, Ebel M, Schaeffer A. Evaluation of the effect of small organic acids
on phytoextraction of Cu and Pb from soil with tobacco Nicotiana tabacum. Chemosphere 2006;63:996-1004.
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
Evangelou MWH, Bauer U, Ebel M, Schaeffer A. The influence of EDDS and EDTA on the
uptake of heavy metals of Cd and Cu from soil with tobacco Nicotiana tabacum.
Chemosphere 2007;68:345–53.
Fasim F, Ahmed N, Parsons R, Gadd GM. Solubilization of zinc salts by a bacterium isolated from the air environment of a tannery. FEMS Microbiol Lett 2002;213:1–6.
Fomina MA, Alexander IJ, Hillier S, Gadd GM. Zinc phosphate and pyromorphite solubilization by soil plant-symbiotic fungi. Geomicrobiol J 2004;21:351–66.
Fomina MA, Alexander IJ, Colpaert JV, Gadd GM. Solubilization of toxic metal minerals
and metal tolerance of mycorrhizal fungi. Soil Biol Biochem 2005;37:851–66.
Gadd GM. Bioremedial potential of microbial mechanisms of metal mobilization and
immobilization. Curr Opin Biotechnol 2000;11:271–9.
Gao Y, Miao C, Mao L, Zhou P, Jin Z, Shi W. Improvement of phytoextraction and
antioxidative defense in Solanum nigrum L. under cadmium stress by application
of cadmium-resistant strain and citric acid. J Hazard Mater 2010;181:771–7.
Garg N, Aggarwal N. Effects of interactions between cadmium and lead on growth,
nitrogen fixation, phytochelatin, and glutathione production in Mycorrhizal Cajanus
cajan (L.) Millsp. J Plant Growth Regul 2011;30:286–300.
Garg N, Aggarwal N. Effect of mycorrhizal inoculations on heavy metal uptake and
stress alleviation of Cajanus cajan (L.) Millsp. genotypes grown in cadmium and
lead contaminated soils. Plant Growth Regul 2012;66:9–26.
Geelhoed JS, Van Riemsdjik WH, Findenegg GR. Simulation of the effect of citrate
exudation from roots on the plant availability of phosphate adsorbed on goethite.
Eur J Soil Sci 1999;50:379–90.
Gill SS, Tuteja N. Reactive oxygen species and antioxidant machinery in abiotic stress
tolerance in crop plants. Plant Physiol Biochem 2010;48:909–30.
Glick BR. Using soil bacteria to facilitate phytoremediation. Biotechnol Adv 2010;28:
367–74.
Glick BR, Bashan Y. Genetic manipulation of plant growth-promoting bacteria to
enhance biocontrol of phytopathogens. Biotechnol Adv 1997;15:353–78.
Glick BR, Todorovic B, Czarny J, Cheng Z, Duan J, McConkey B. Promotion of plant
growth by bacterial ACC deaminase. Crit Rev Plant Sci 2007;26:227–42.
Gohre V, Paszkowski U. Contribution of the arbuscular mycorrhizal symbiosis to heavy
metal phytoremediation. Planta 2006;223:1115–22.
Gonzalez-Chavez MC, Carrillo-Gonzalez R, Wright SF, Nichols KA. The role of glomalin,
a protein produced by arbuscular mycorrhizal fungi, in sequestering potentially
toxic elements. Environ Pollut 2004;130:317–23.
González-Guerrero M, Melville LH, Ferrol N, Lott JN, Azcón-Aguilar C, Peterson RL.
Ultrastructural localization of heavy metals in the extraradical mycelium and
spores of the arbuscular mycorrhizal fungus Glomus intraradices. Can J Microbiol
2008;54:103–10.
Goodell B, Jellison J, Liu J, Daniel G, Paszczynski A, Fekete F, et al. Low molecular weight
chelators and phenolic compounds isolated from wood decay fungi and their role
in the fungal biodegradation of wood. J Biotechnol 1997;53:133–62.
Gratao PL, Polle A, Lea PJ, Azevedo RA. Making the life of heavy metal-stressed plants a
little easier. Funct Plant Biol 2005;32:481–94.
Han F, Shan X, Zhang S, Wen B, Owens G. Enhanced cadmium accumulation in maize
roots-the impact of organic acids. Plant Soil 2006;289:355–68.
Hayat R, Ali S, Amara U, Khalid R, Ahmed I. Soil beneficial bacteria and their role in
plant growth promotion: a review. Ann Microbiol 2010;60:579–98.
Hoberg E, Marschner P, Lieberei R. Organic acid exudation and pH changes by Gordonia
sp. and Pseudomonas fluorescens grown with P adsorbed to goethite. Microbiol Res
2005;160:177–87.
Hrynkiewicz K, Dabrowska G, Baum C, Niedojadlo K, Leinweber P. Interactive and single
effects of ectomycorrhiza formation and Bacillus cereus on metallothionein mt1 expression and phytoextraction of Cd and Zn by willows. Water Air Soil Pollut
2012;223:957–68.
Idris R, Trifonova R, Puschenreiter M, Wenzel WW, Sessitsch A. Bacterial communities
associated with flowering plants of the Ni hyperaccumulator Thlaspi goesingense.
Appl Environ Microbiol 2004;70:2667–77.
Jiang M, Zhang J. Water stress-induced abscissic acid accumulation triggers the
increased generation of reactive oxygen species and up-regulates the activities of
antioxidant enzymes in maize-leaves. J Exp Bot 2002;53:2401–10.
Jones DL. Organic acids in the rhizosphere – a critical review. Plant Soil 1998;205:25–44.
Jones DL, Edwards AC. Influence of sorption on the biological utilization of two simple
carbon substrates. Soil Biol Biochem 1998;30:1895–902.
Jones DL, Prabowo AM, Kochian LV. Kinetics of malate transport and decomposition in
acid soils and isolated bacterial populations – the effect of microorganisms on root
exudation of malate under Al stress. Plant Soil 1996;182:239–47.
Jones DL, Dennis PG, Owen AG, van Hees PAW. Organic acid behavior in soils – misconceptions and knowledge gaps. Plant Soil 2003;248:31–41.
Joshi PM, Juwarkar AA. In vivo studies to elucidate the role of extracellular polymeric
substances from Azotobacter in immobilization of heavy metals. Environ Sci
Technol 2009;43:5884–9.
Juwarkar AA, Jambhulkar HP. Phytoremediation of coal mine spoil dump through integrated biotechnological approach. Bioresour Technol 2008;99:4732–41.
Juwarkar AA, Nair A, Dubey KV, Singh SK, Devotta S. Biosurfactant technology for remediation of cadmium and lead contaminated soils. Chemosphere 2007;10:1996–2002.
Kavita B, Shukla S, Naresh Kumar G, Archana G. Amelioration of phytotoxic effects of Cd
on mung bean seedlings by gluconic acid secreting rhizobacterium Enterobacter
asburiae PSI3 and implication of role of organic acid. World J Microbiol Biotechnol
2008;24:2965–72.
Khan MS, Zaidi A, Wani PA, Oves M. Role of plant growth promoting rhizobacteria in
the remediation of metal contaminated soils. Environ Chem Lett 2009;7:1-19.
Kidd P, Barcelo J, Bernal MP, Navari-Izzo F, Poschenrieder C, Shilev S, et al. Trace element behaviour at the root–soil interface: implications in phytoremediation. Environ Exp Bot 2009;67:243–59.
1573
Kozdroj J, Piotrowska-Seget Z, Krupa P. Mycorrhizal fungi and ectomycorrhiza associated bacteria isolated from an industrial desert soil protect pine seedlings against
Cd(II) impact. Ecotoxicology 2007;16:449–56.
Krupa P, Kozdrój J. Ectomycorrhizal fungi and associated bacteria provide protection
against heavy metals in inoculated pine (Pinus sylvestris L.) seedlings. Water Air
Soil Pollut 2007;182:83–90.
Kuffner M, Puschenreiter M, Wieshammer G, Gorfer M, Sessitsch A. Rhizosphere bacteria affect growth and metal uptake of heavy metal accumulating willows. Plant Soil
2008;304:35–44.
Kuffner M, De Maria S, Puschenreiter M, Fallmann K, Wieshammer G, Gorfer M, et al.
Culturable bacteria from Zn- and Cd accumulating Salix caprea with differential
effects on plant growth and heavy metal availability. J Appl Microbiol 2010;108:
1471–84.
Lebeau T, Braud A, Jézéquel K. Performance of bioaugmentation-assisted phytoextraction
applied to metal contaminated soils: a review. Environ Pollut 2008;153:497–522.
Leyval C, Turnau K, Haselwandter K. Effect of heavy metal pollution in mycorrhizal colonization and function: physiological, ecological and applied aspects. Mycorrhiza
1997;7:139–53.
Li WC, Ye ZH, Wong MH. Metal mobilization and production of short-chain organic
acids by rhizosphere bacteria associated with a Cd/Zn hyperaccumulating plant.
Sedum alfredii. Plant Soil 2010;326:453–67.
Luo SL, Chen L, Chen JI, Xiao X, Xu TY, Wan Y, et al. Analysis and characterization
of cultivable heavy metal-resistant bacterial endophytes isolated from
Cd-hyperaccumulator Solanum nigrum L. and their potential use for
phytoremediation. Chemosphere 2011;85:1130–8.
Luo S, Xu T, Chen L, Chen J, Rao C, Xiao X, et al. Endophyte-assisted promotion of biomass
production and metal-uptake of energy crop sweet sorghum by
plant-growth-promoting endophyte Bacillus sp. SLS18. Appl Microbiol Biotechnol
2012;93:1745–53.
Ma QY, Traina SJ, Logan TJ. In situ lead immobilization by apatite. Environ Sci Technol
1993;27:1803–10.
Ma Y, Rajkumar M, Vicente J, Freitas H. Inoculation of Ni-resistant plant growth
promoting bacterium Psychrobacter sp. strain SRS8 for the improvement of nickel
phytoextraction by energy crops. Int J Phytoremediation 2010;13:126–39.
Ma Y, Prasad MNV, Rajkumar M, Freitas H. Plant growth promoting rhizobacteria
and endophytes accelerate phytoremediation of metalliferous soils. Biotechnol
Adv 2011a;29:248–58.
Ma Y, Rajkumar M, Luo Y, Freitas H. Inoculation of endophytic bacteria on host and
non-host plants – Effects on plant growth and Ni uptake. J Hazard Mater
2011b;196:230–7.
Machuca A, Pereira G, Aguiar A, Milagres AMF. Metal-chelating compounds produced
by ectomycorrhizal fungi collected from pine plantations. Lett Appl Microbiol
2007;44:7-12.
Madhaiyan M, Poonguzhali S, Sa T. Metal tolerating methylotrophic bacteria reduces
nickel and cadmium toxicity and promotes plant growth of tomato (Lycopersicon
esculentum L.). Chemosphere 2007;69:220–8.
Malhotra M, Srivastava S. Stress-responsive indole-3-acetic acid biosynthesis by
Azospirillum brasilense SM and its ability to modulate plant growth. Eur J Soil Biol
2009;45:73–80.
Maria SD, Rivelli AR, Kuffner M, Sessitsch A, Wenzel WW, Gorfer M, et al. Interactions
between accumulation of trace elements and macronutrients in Salix caprea after
inoculation with rhizosphere microorganisms. Chemosphere 2011;84:1256–61.
Martino E, Perotto S, Parsons R, Gadd GM. Solubilization of insoluble inorganic zinc compounds by ericoid mycorrhizal fungi derived from heavy metal polluted sites. Soil
Biol Biochem 2003;35:133–41.
Mastretta C, Taghavi S, van der Lelie D, Mengoni A, Galardi F, Gonnelli C, et al.
Endophytic bacteria from seeds of Nicotiana tabacum can reduce cadmium phytotoxicity. Int J Phytoremediation 2009;11:251–67.
McGrath SP, Chaudri AM, Giller KE. Long-term effects of metals in sewage sludge on
soils, microorganisms and plants. J Ind Microbiol 1995;14:94-104.
Meharg AA. The mechanistic basis of interactions between mycorrhizal associations
and toxic metal cations. Mycol Res 2003;107:1253–65.
Miransari M. Hyperaccumulators, arbuscular mycorrhizal fungi and stress of heavy
metals. Biotechnol Adv 2011;29:645–53.
Møller IM, Jensen PE, Hansson A. Oxidative modifications to cellular components in
plants. Annu Rev Plant Biol 2007;58:459–81.
Mühlbachová G. Microbial biomass dynamics after addition of EDTA into heavy metal
contaminated soils. Plant Soil Environ 2009;55:544–50.
Mulligan CN, Yong RN, Gibbs BF. Remediation technologies for metal-contaminated
soils and groundwater: an evaluation. Eng Geol 2001;60:193–207.
Najeeb U, Xu L, Ali S, Jilani G, Gong HJ, Shen WQ, et al. Citric acid enhances the
phytoextraction of manganese and plant growth by alleviating the ultra structural
damages in Juncus effusus L. J Hazard Mater 2009;170:156–63.
Nogueira MA, Nehls U, Hampp R, Poralla K, Cardoso EJBN. Mycorrhiza and soil bacteria
influence extractable iron and manganese in soil and uptake by soybean. Plant Soil
2007;298:273–84.
Orłowska E, Przybyłowicz W, Orlowski D, Turnau K, Mesjasz-Przybyłowicz J. The effect
of mycorrhiza on the growth and elemental composition of Ni-hyperaccumulating
plant Berkheya coddii Roessler. Environ Pollut 2011;159:3730–8.
Ouziad F, Hildebrandt U, Schmelzer E, Bothe H. Differential gene expressions in
arbuscular mycorrhizal-colonized tomato grown under heavy metal stress. J Plant
Physiol 2005;162:634–49.
Ouzounidou G, Moustakas M, Symeonidis L, Karataglis S. Response of wheat seedlings to Ni
stress: effects of supplemental calcium. Arch Environ Contam Toxicol 2006;50:346–52.
Panfili F, Schneider A, Vives A, Perrot F, Hubert P, Pellerin S. Cadmium uptake by durum
wheat in presence of citrate. Plant Soil 2009;316:299–309.
1574
M. Rajkumar et al. / Biotechnology Advances 30 (2012) 1562–1574
Parida BK, Chhibba IM, Nayyar VK. Influence of nickel contaminated soils on fenugreek
(Trigonella corniculata L.) growth and mineral composition. Sci Hortic-Amsterdam
2003;98:113–9.
Park JH, Bolan N, Megharaj M, Naidu R. Isolation of phosphate solubilizing bacteria
and their potential for lead immobilization in soil. J Hazard Mater 2011;185:
829–36.
Pulford ID, Watson C. Phytoremediation of heavy metal contaminated land by trees-a
review. Environ Int 2003;29:529–40.
Rai UN, Pandey K, Sinha S, Singh A, Saxena R, Gupta DK. Revegetating fly ash landfills
with Prosopis juliflora L.: impact of different amendments and Rhizobium inoculation. Environ Int 2004;30:293–300.
Rajkumar M, Freitas H. Effects of inoculation of plant-growth promoting bacteria on Ni
uptake by Indian mustard. Bioresour Technol 2008;99:3491–8.
Rajkumar M, Ae N, Prasad MNV, Freitas H. Potential of siderophore-producing bacteria
for improving heavy metal phytoextraction. Trends Biotechnol 2010;28:142–9.
Ramachandran S, Fontanille P, Pandey A, Larroche C. Gluconic Acid: properties, applications and microbial production. Food Technol Biotechnol 2006;44:185–95.
Raskin I, Smith RD, Salt DE. Phytoremediation of metals: using plants to remove pollutants
from the environment. Curr Opin Biotechnol 1997;8:221–6.
Rodriguez H, Gonzalez T, Goire I, Bashan Y. Gluconic acid production and phosphate
solubilization by the plant growth-promoting bacterium Azospirillum spp.
Naturwissenschaften 2004;91:552–5.
Rodriguez H, Vessely S, Shah S, Glick BR. Effect of a nickel-tolerant ACC
deaminase-producing Pseudomonas strain on growth of nontransformed and transgenic canola plants. Curr Microbiol 2008;57:170–4.
Ryan PR, Delhaize E, Jones DL. Function and mechanism of organic anion exudation
from plant roots. Annu Rev Plant Physiol Plant Mol Biol 2001;52:527–60.
Salt DE, Smith RD, Raskin I. Phytoremediation. Annu Rev Plant Physiol Plant Mol Biol
1998;13:468–74.
Saravanan VS, Madhaiyan M, Thangaraju M. Solubilization of zinc compounds by the
diazotrophic, plant growth promoting bacterium Gluconacetobacter diazotrophicus.
Chemosphere 2007;66:1794–8.
Sauer M, Porro D, Mattanovich D, Branduardi P. Microbial production of organic acids:
expanding the markets. Trends Biotechnol 2008;26:100–8.
Sayer JA, Cotter-Howells JD, Watson C, Hillier S, Gadd GM. Lead mineral transformation
by fungi. Curr Biol 1999;9:691–4.
Schalk IJ, Hannauer M, Braud A. New roles for bacterial siderophores in metal transport
and tolerance. Environ Microbiol 2011;13:2844–54.
Sheng X, He L, Wang Q, Ye H, Jiang C. Effects of inoculation of biosurfactant-producing
Bacillus sp. J119 on plant growth and cadmium uptake in a cadmium-amended soil.
J Hazard Mater 2008a;155:17–22.
Sheng XF, Xia JJ, Jiang CY, He LY, Qian M. Characterization of heavy metal-resistant endophytic bacteria from rape (Brassica napus) roots and their potential in promoting the
growth and lead accumulation of rape. Environ Pollut 2008b;156:1164–70.
Shi JY, Lin HR, Yuan XF, Chen XC, Shen CF, Chen YX. Enhancement of copper availability
and microbial community changes in rice rhizospheres affected by sulfur. Molecules
2011;16:1409–17.
Sinha S, Mukherjee SK. Cadmium-induced siderophore production by a high
Cd-resistant bacterial strain relieved Cd toxicity in plants through root colonization. Curr Microbiol 2008;56:55–60.
Smith SE, Read DJ. Mycorrhizal Symbiosis. 2nd ed. London: Academic Press; 1997.
p. 605.
Taghavi S, Garafola C, Monchy S, Newman L, Hoffman A, Weyens N, et al. Genome
survey and characterization of endophytic bacteria exhibiting a beneficial effect
on growth and development of poplar. Appl Environ Microbiol 2009;75:748–57.
Tank N, Saraf M. Enhancement of plant growth and decontamination of nickel-spiked
soil using PGPR. J Basic Microbiol 2009;49:195–204.
Taurian T, Anzuay MS, Angelini JG, Tonelli ML, Ludueña L, Pena D, et al.
Phosphate-solubilizing peanut associated bacteria: screening for plant
growth-promoting activities. Plant Soil 2010;329:421–31.
Tripathi M, Munot HP, Shouche Y, Meyer JM, Goel R. Isolation and functional characterization of siderophore-producing lead- and cadmium-resistant Pseudomonas
putida KNP9. Curr Microbiol 2005;50:233–7.
Ultra VU, Yano A, Iwasaki K, Tanaka S, Kang YM, Sakurai K. Influence of chelating agent
addition on copper distribution and microbial activity in soil and copper uptake by
brown mustard (Brassica juncea). Soil Sci Plant Nutr 2005;51:193–202.
Uroz S, Calvaruso C, Turpault MP, Frey-Klett P. Mineral weathering by bacteria: ecology,
actors and mechanisms. Trends Microbiol 2009;17:378–87.
Venkatesh NM, Vedaraman N. Remediation of soil contaminated with copper using
rhamnolipids produced from Pseudomonas aeruginosa MTCC 2297 using waste
frying rice bran oil. Ann Microbiol 2012;62:85–91.
Vivas A, Voros I, Biro B, Barea JM, Ruiz-Lozano JM, Azcon R. Beneficial effects of indigenous Cd tolerant and Cd-sensitive Glomus mosseae associated with a Cd-adapted
strain of Brevibacillus sp. in improving plant tolerance to Cd contamination. Appl
Soil Ecol 2003;24:177–86.
Vivas A, Biro B, Ruíz-Lozanoa JM, Azcon R. Two bacterial strains isolated from a
Zn-polluted soil enhance plant growth and mycorrhizal efficiency under Zn toxicity. Chemosphere 2006;52:1523–33.
Wang Q, Xiong D, Zha P, Yu X, Tu B, Wang G. Effect of applying an arsenic-resistant and
plant growth–promoting rhizobacterium to enhance soil arsenic phytoremediation
by Populus deltoides LH05-17. J Appl Microbiol 2011;111:1065–74.
Wani PA, Khan MS, Zaidi A. Chromium reduction, plant growth-promoting potentials,
and metal solubilization by Bacillus sp. isolated from alluvial soil. Curr Microbiol
2007a;54:237–43.
Wani PA, Khan MS, Zaidi A. Effect of metal tolerant plant growth promoting
Bradyrhizobium sp. (vigna) on growth, symbiosis, seed yield and metal uptake by
greengram plants. Chemosphere 2007b;70:36–45.
Wani PA, Khan MS, Zaidi A. Chromium reducing and plant growth promoting
Mesorhizobium improves chickpea growth in chromium amended soil. Biotechnol
Lett 2008a;30:159–63.
Wani PA, Khan MS, Zaidi A. Effect of heavy metal toxicity on growth, symbiosis, seed
yield and metal uptake in pea grown in metal amended soil. Bull Environ Contam
Toxicol 2008b;55:33–42.
Wenzel WW. Rhizosphere processes and management in plant-assisted bioremediation
(phytoremediation) of soils. Plant Soil 2009;321:385–408.
Wenzel WW, Bunkowski M, Puschenreiter M, Horak O. Rhizosphere characteristics of
indigenously growing nickel hyperaccumulator and excluder plants on serpentine
soil. Environ Pollut 2003;123:131–8.
Wu SC, Cheung KC, Luo YM, Wong MH. Effects of inoculation of plant growth-promoting
rhizobacteria on metal uptake by Brassica juncea. Environ Pollut 2006;140:124–35.
Wu SC, Wong CC, Shu WS, Khan AG, Wong MH. Mycorrhizo-remediation of lead/zinc
mine tailings using vetiver: A field study. Int J Phytoremediation 2011a;13:61–74.
Wu Q, Wang S, Thangavel P, Li Q, Zheng H, Bai J, et al. Phytostabilization potential of
Jatropha curcas L. in polymetallic acid mine tailings. Int J Phytoremediation
2011b;13:788–804.
Yamane K, Rahman Md S, Kawasaki M, Taniguchi M, Miyake H. Pretreatment with a
low concentration of methyl viologen decreases the effects of salt stress on chloroplast ultrastructure in rice leaves (Oryza sativa L.). Plant Prod Sci 2004;7:435–41.
Yang Q, Tu S, Wang G, Liao X, Yan X. Effectiveness of applying arsenate reducing bacteria to enhance arsenic removal from polluted soils by Pteris vittata L. Int J
Phytoremediation 2012;14:89–99.
Zarei M, König S, Hempel S, Nekouei MK, Savaghebi G, Buscot F. Community structure
of arbuscular mycorrhizal fungi associated to Veronica rechingeri at the Anguran
zinc and lead mining region. Environ Pollut 2008;156:1277–83.
Zarei M, Wubet T, Schäfer SH, Savaghebi GR, Jouzani GS, Nekouei MK, et al. Molecular
diversity of arbuscular mycorrhizal fungi in relation to soil chemical properties and
heavy metal contamination. Environ Pollut 2010;158:2757–65.
Zhang YF, He LY, Chen ZJ, Zhang WH, Wang QY, Qian M, et al. Characterization of
lead-resistant and ACC deaminase-producing endophytic bacteria and their potential in promoting lead accumulation of rape. J Hazard Mater 2011;186:720–5.