Vetiver grass as an Ideal Phytosymbiont for Glomalian Fungi for…

 Vetiver grass as an Ideal Phytosymbiont for Glomalian Fungi for
Ecological Restoration of Heavy Metal Contaminated Derelict Land

                                         Abdul G. Khan
              School of Science, Food and Horticulture, University of Western Sydney
              Locked Bag 1797, Penrith South DC 1797. New South Wales, Australia

Abstract: Pollution of the soil environment with toxic materials from fossil burning, mining and smelting
of metalliferous ores, disposal of sewage, fertilizers and pesticides, etc. has increased dramatically since
the onset of industrial revolution. Various strategies including bioremediation and phytoremediation are
employed to remove heavy metals from such soils and making them available for agricultural purposes
and urban developments. Role of plants as phytosymbionts and their associated arbuscular Glomalian
mycorrhizal fungi as mycosymbionts are discussed as an alternative (mycorrhizo-remediation) strategy
for safe and efficient decontamination of such soils. Prospects of using vetiver grass as an ideal
phytosymbiont due its fast growth rate and root morphology and Glomalian mycorrhizal fungi as
mycosymbionts for enhanced uptake of heavy metals is discussed.
Key words: phytosymbionts, mycosymbionts, arbuscular Glomalian mycorrhiza, heavy metals, vetiver
               grass, Vetiveria zizanioides, mycorrhizo-remediation, phytoremediation.
Email contact: Abdul G. Khan 


1     GLOBAL SOIL CONTAMINATION

       Due to ever increasing industrial, agricultural, and mining activities worldwide, heavy metal
pollution of land and water is becoming globally important environmental, health, economic, and
planning issue. There is an increase in world population, and unpleasant disposal of industrial effluents,
especially in the third world countries, is causing soil pollution. Utilization of these lands for agricultural
purposes and urban developments requires a safe and efficient decontamination process. With the
increasing use of agrochemicals to maintain and improve soil fertility, unwanted elements such as
cadmium into soils due to contaminated sources of fertilizers, especially in developing countries, are
being introduced into agricultural soils, which, poses a potential threat to the food chain (Chaney and
Oliver, 1996). Mining and industrial operations also lead to significant challenges for the management of
the natural environments during and after these activities. Increased public awareness of the
environmental impact of such activities is demanding an interdisciplinary, inter-organizational, and
international effort.

2     REMEDIATION TECHNIQUES

2.1    Physical and Chemical Techniques
       Various physical and chemical techniques to de-contaminated soils have been undertaken during
the last 25 years (Salomons et al. 1995; Wise and Trantolo, 1995; Rao et al. 1996; Burns et al. 1996) and
millions of dollars being spent by governments all over the world on preventive measures. However, all
of them are labor intensive and costly, and cannot be applied to thousands of hectares of land
contaminated with inorganic heavy metals (Burns et al. 1996). These technologies results in rendering the
soil biologically dead and useless for plant growth as they remove all flora, fauna and microbes including
useful nitrogen fixing bacteria and P-enhancing mycorrhizal fungi.

2.2    Bioremediation Techniques
       Microbial bioremediation technology, well known for decontamination of organics (Flathman et
al., 1994), is not available for large-scale biodegradation of inorganic heavy metals. The health hazards
caused by the accumulation of toxic metals in the environment together with the high cost of removal and
replacement of metal-polluted soil have prompted efforts to develop alternative and cheaper techniques to
recover the degraded land.

2.3     Phytoremediation
        Restoration of derelict land by establishing a plant cover is important before it poses serious health
hazard by transferring the trace metals into the surroundings. Current research in this area now includes
utilization of plants to remediate polluted soils and to facilitate improvement of soils structure in cases of
severe erosion, the innovative technique being known as phytoremediation (Chaudhry et al., 1998; Khan
et al., 2000; Brooks, 1997).
        Recently Reeves (2003) have reviewed tropical hyperaccumulator of heavy metal plants and
concluded that there is a lack of investigation for the occurrence of hyperaccumulator plant species. No
botanical or biogeochemical exploration of trace metal tolerant and/or accumulating plant species has yet
taken place in many parts of the world. Many plant species, which can accumulate high concentrations of
trace elements, have been known for over a century. Renewed interest in the role of these hyper-
accumulating plants in phytoremediation has stimulated research in this area (Brooks, 1997). Several
plant species or ecotypes, associated with heavy metal enriched soils, accumulate metals in the shoots.
These plants can be used to clean up heavy metal contaminated sites by extracting metals from soils and
accumulating them in aboveground biomass (Chaudhry et al., 1999; Khan et al., 2000). The metal
enriched biomass can be harvested and smelted to recover the metal (phytomining).
2.3.1 Plants for Phytoremediation:
        Plants that are used to extract heavy metals from contaminated soils have to be the most suitable
for the purpose, i.e. tolerant to specific heavy metal, adapted to soil and climate, capable of high uptake of
heavy metal(s), etc. Plants either take up one or two specific metals in high concentrations into their
tissues (hyperaccumulators) with low biomass (Chaudhry et al., 1998)), or extract low to average heavy
metal (not metal specific) concentrations in their shoots with high biomass. Low biomass
hyperaccumulators, generally, have a restricted root system (Ernst, 1996). In contrast, non-accumulators,
high biomass producing and tolerant plants have physiological adaptation mechanisms, which allow them
to grow in contaminated soils better than others (Palazzo and Lee, 1997). The tolerance and specific
behaviour at the root level must be taken into consideration while selecting plants for phytoremediation
(Keller et al., 2003). Root system morphology allows some plants to be more efficient than others in
nutrient uptake in infertile soil or stressed soil conditions (Fitter and Stickland, 1991).
2.3.2 Vetiver grass as an ideal plant for phytoremediation:
        Recently vetiver grass, due to its ecofriendly nature, found a new use for phytoremediation of
contaminated sites. Vetiver grass is both a xerophyte and a hydrophyte and, once established, is not
affected by droughts or floods (Greenfield, 1988). It is highly tolerant to droughts and water logging,
frost, heat, extreme soil pH, sodicity, salinity, Al and Mn toxicity (Truong and Claridge, 1996). It is also
highly tolerant to a range of trace elements such as As, Cd, Cu, Cr, and Ni (Truong and Claridge, 1996).
It is suitable for the stabilization and rehabilitation and reclaiming of acid sulfate and trace metals
contaminated soils, i.e. phytoremediation. In Australia vetiver has been successfully used to stabilize
mining overburden and highly saline, sodic, magnesic, and alkaline or acidic tailings of coal and gold
mines (Truong, 1999). This grass has been extensively used for land protection by mitigating soil erosion
and water conservation, especially on very steep slopes, due to its faster root growth, i.e. root length may
reach up to 3 m just in one year (Lavania and Lavania, 2000; Walle and Sims, 1998). Vetiver grass is
regarded as a tool for environmental engineering (Mucciarelli et al., 1998) and as one of the most
versatile crops of the third millennium (Maffei, 2002). Truong (1999) furnished field observations
relating to high tolerance levels of vetiver grass to adverse soil conditions, trace metal toxicities, and
agrochemicals. Chen et al. (2000) made a comparative study of the effects of chemical methods on the
growth and uptake of trace elements by many plants including vetiver grass and found this perennial grass
having a greater ability to remove Cd, Pb, and Zn from soil, the values of Cd accumulation close to those
of hyperaccumulator Thlaspi caerulescenes. The authors discussed the effectiveness of phytoremediation
with this grass with great biomass and concluded that `VGT is an effective, low-cost, and environmentally
friendly technology to clean Cd contaminated soils'. The authors suggested to develop a genetically
modified vetiver grass incorporating genes of hypoaccumulator. Recently Shu et al. (2002) reported
enhanced trace metal extraction in field experiments using vetiver grasses for re-vegetation of Pb / Zn
mine tailing. VGT is emerging as an alternative technology for rehabilitation of degraded, saline, or trace
metal contaminated soils, and for purification of water polluted with trace elements, agrochemicals, and
industrial- effluent disposals.
        The success of phytoremedial efforts is not only dependant upon the choice of plant species but
also their method of establishment (Bradshaw, 1987). Among the plants involved in phytoremedial
measures, Vetiver grass {Vetiveria zizanioides L. (Nash)}, should receive special attention. It is a densely
tufted, awnless, wiry and glabrous plant occurring in large clumps as hydrophyte or xerophyte on
vertisols through to red alfisols. It can grow on both acidic (pH 3) and alkaline (pH 11) soils, and is
tolerant to high levels of various trace metals such as arsenic, cadmium, copper, chromium and nickel
(Truong and Claridge, 1996; Truong and Baker, 1998; Truong, 1999). It is one of those few plants which
possess both economical and ecological capabilities, i.e. volatile essential oil distilled from its roots in
over 70 countries (Akhila and Rani, 2002) and its conservation properties, such as up to 2 m high plant
with a strong dense and mainly vertical root system often measuring > 3 m, useful in soil erosion control
(Greenfield, 1988, 1989, 1993, 1995; Truong, 2002). It is propagated vegetatively and is non-invasive
(National Research Council, USA, 1993)). It is extremely resistant to insect pests and diseases (Zisong,
1991) and is widely used worldwide for soil and moisture conservation and soil restoration. It is immune
to flooding, grazing, fires, and other hazards (Grimshaw and Helfer, 1995). Maffei (2002) stated that this
plant is one of the most versatile crops of the third millennium. On the request of the World Bank, the US
Agency for International Development, and the US Conservation Services, the National Research Council
of USA, evaluated the usefulness of the vetiver system and its implications. China is among the nations
most active in studying vetiver system and massive projects have been started in several-conservation
areas, as soil and moisture conservation are China's national priorities. Vetiver was introduced in China
in 1950's as a source of aromatic oil and when the oil prices dropped, the plant was abandoned. It was
only in 1988, that its importance in soil and moisture conservation was realized and massive vetiver trial
projects started by China's Ministry of Water Resources in many provinces (Vetiver Information
Network, 1993). Chinese scientists and researchers also started their own vetiver networks to exchange
their experiences and results of vetiver trials. Hill and Peart (1999) reviewed the methodology of vetiver
trials and results of these trials in Hong Kong and Guizhou, China, for soil erosion control.
3     SOIL MICROBES AND PLANT RELATIONSHIPS

3.1   Soil Microbes
       Soil contains a great variety of microbial populations, the properties and behavior of which affect
the function of the soil ecosystem in many crucial ways, especially microbial re- mediation of C, N, S, P,
Fe, Mn, Hg and Se (Khan, 2002a). Microorganisms in soil are not only affected by the physical and
chemical properties of the soil in general, but also by the moisture, temperature, pH and organic matter
released from the roots in the rhizospheres of plants, all of which alter the microbial diversity and activity
(Lynch, 1982).
      Nevertheless, relatively few studies have focused on the ef fects of rhizosphere microorganisms on
the soil erosion and metal remediation efforts, despite the important role that these microorganisms play
in plant interactions with soil environment (for toxic metals in particular). Added to this, the effects of
phytoremediation practices on microbial communities to the remediated sites have also been largely
ignored, and these microbes may be essential for re-vegetation efforts following the removal of excessive
soil metals (Pawlowska et al., 2000; Khan 2003b).

3.2    Arbuscular Mycorrhizal Fungi
       Among the rhizosphere microorganisms involved in plant interactions with soil, the arbuscular
mycorrhizal (AM) fungi, which belong to Family Glomales, Class Zygomycetes, should receive special
attention. This is because over 80 % of the world's plant species are mycorrhizal and potentially benefit
from AM fungus-mediated mineral nutrition (Smith and Read, 1997). Recently, plants capable of forming
association with AM fungi have been shown to accumulate a considerable amount of trace metals (Burke
et al., 2000; Karagiannidis and Nikolaou, 2000). Unfortunately, the role that AM fungi play in plant
interactions with soil metals is not fully understood (Pawlowska et al., 2000; Weissenhorn and Leyval,
1996), and little is known about mycorrhiza functioning under conditions imposed by trace metal
remediation protocols and also the effects of phytoextraction on AM fungi. Arbuscular mycorrhizal fungi
(AMF) should be considered as an essential component of soil microflora and as a potential tool for re-
establishment of plant cover and population diversity during ecosystem restoration (Turnau and
Haselwandter, 2002). The rate of reclaiming derelict land may be increased by AMF inoculation of
plants used for re-vegetation, i.e. mycorrhizo-remediation (Jamal et al., 2002).
       Arbuscular mycorrhizal (AM) fungi, which coevolved with plant roots, form symbiotic
associations with around 82 % of angiosperms (Brundrett, 2002). These fungi are well known to improve
plant growth on nutrient-poor soils and enhance the uptake of P, Cu, Ni, Pb, and Zn (Khan et al., 2000).
Despite the important role that AM fungi play in plant interactions with the soil environment in general,
and trace elements in particular, relatively few studies have focused on their effect on the metal
remediation efforts. Early phytoremediation studies have focused on the predominantly non-mycorrhizal
plant families, e.g. Brassicaceae or Caryophyllaceae, so AM have not been considered as important
component of phytoremediation practices (Pawlowska et al., 2000).
       Term `mycorrhizo-remediation' is coined by Jamal et al. (2002) to use mycorrhizal plants in
phytoremediation of heavy metal contaminated soils. The interaction between arbuscular mycorrhizae
(AM) and minerals other than phosphorus, particularly trace elements, has been the subject of many
recent studies because of the possibility of the beneficial effect of mycorrhizae in improving the tolerance
of plants against toxicity (Khan et al., 2000). The uptake of trace elements by mycorrhizal plants depends
on several factors such as physicochemical properties of the soil, particularly its fertility level, pH, the
host plants and the fungi involved, and above all, the concentration of the trace elements in soil. Under
deficiency conditions, most studies point to an increase in trace element uptake by mycorrhizal plants. For
example, increased Zn uptake by maize inoculated with mycorrhizal fungus, Glomus mosseae, has been
reported compared to the non-mycorrhizal plants at low Zn levels (Kothari et al., 1991). Another study
revealed an increase in Cu uptake by white clover inoculated with G. mosseae under low level of Cu - a
limiting factor (Li et al., 1991). Jamal et al. (2002) found AM fungi enhancing the uptake of zinc and
nickel from contaminated soils by soybean and lentils. Using indigenous AM fungal populations as
mycorrhizal treatment, it has been observed that there is a decrease in foliar concentration of Pb in
mycorrhizal Anthyllis cytisoides plants growing in Pb polluted soil (Diaz et al.,1996). However, when the
soil contained potentially toxic amounts of trace elements, mycorrhizal formation usually induces lower
concentrations of these elements in the aerial part of the host plant, thus consequently lead to a beneficial
effect on plant growth. This has been reported for Zn when growing white clover in Zn contaminated soils
(Zhu et al., 2001). Reduction in both Zn and Pb contents in shoot when growing Lygeum spartum and
Anthyllis cytisoides, inoculated with AMF in different concentrations of Pb and Zn amended soils, has
also been observed (Diaz et al., 1996). Davies et al. (2002) reported higher chromium accumulation in
sunflower plants inoculated with AM fungus Glomus intraradices as compared to the non-mycorrhizal
plants. The authors showed that the mycorrhizal fungi helped to partially alleviate Cr toxicity and
enhanced plant growth. They found greatest Cr concentrations in the roots, intermediate in the shoots and
lowest in leaves. Khan (2001) studied the relationships between Cr III magnification ratio, accumulation
factor, and AM in three tree species growing on tannery effluent-polluted soil, reported differing capacity
of Cr III uptake, and found AM encouraged mineral nutrition, including Cr.

4     VETIVER; AM MYCORRHIZAE & PHYTOREMEDIATION

4.1     Historical Background to Vetiver Grass
       Vetiver grass (Vetiveria zizanioides (L.) Nash) has been known to be a useful plant for thousands of
years and has been cultivated for the production of scented oil produced by its roots as well as for its
ability to retain soil and prevent erosion (Maffei, 2002). It is a perennial grass belonging to the family
Poaceae (Gramineae), originally from India, growing wild or cultivated. The name derives from the Tamil
`vetti' (khus-khus) and `ver' (root), referring to aromatic roots. It is mentioned in the ancient Sanskrit
writings and it is a part of Hindu mythology. Since late last century, it is being used by the sugar industry
in West Indies, Fiji, and eastern African islands like Mauritius for its soil conservation properties.

4.2    Historical Background to Glomalian Fungi
       Glomales are one of the oldest group of fungi, older than land plants. First land plants, Bryophytes,
appeared 476-430 million years ago in early Silurian periods. No fossil records are available for the
rootless fresh water Charophycean alage, which were the probable ancestors of land plants, to show if
they were mycorrhizal. Fossil evidence of Glomales in the rhizomes of early vascular plants like
Sphenophytes, Lycopodophytes, and Pteridophytes, suggests that the origin of vascular terrestrial plants
is likely from their Bryophyte-like ancestors(Edwards et al. 1998). Both living and Triassic fossil cycads
had glomalian arbuscular mycorrhizal fungi in their roots. AM associations are ubiquous in the living
angiosperms which probably arose in the early Cretaceous (Khan, 2003). The phylogenetic relationship
between origin and diversification of AM fungi and coincidence with vascular land plants was
investigated by Simon et al. (1993) by sequencing rDNA genes as a molecular clock to infer dates. The
authors estimated the origin of AM-like fungi of 353-462 Myr ago, which is consistent with the
hypothesis that AM were instrumental in the colonization of land by ancient plants. This hypothesis is
also supported by the observation that AM can now be found worldwide in the angiosperms,
gymnosperms as well as ferns, suggesting that its nature is ancestral (Brundrett, 2002).

4.3    Vetiver as Phytosymbiont and Glomalian Fungi as Mycosymbionts for Mycorrhizo-
       remediation of Heavy Metal Contaminated Land
       Although vetiver grass is regarded to be suitable candidate for phytoremediation (Chen et al. 2000),
no record of its mycorrhizal status exists in literature. V. zizanioides showed good growth performance on
Pb/Zn mine tailings in Shu et al's (2002) revegetation field experiment. Again no mention of its
mycorrhizal status was made by the authors. Vietmeyer (2002), while discussing VGT beyond the hedge
against soil erosion and essential oil secretions, identified many areas requiring investigations including
symbiosis of vetiver grass roots with mycorrhizal fungi and N-fixing bacteria. Wong (2003) found the
roots of vetiver grass, growing in the South China Botanical Gardens in Guangzhou, RPC, in soil
containing moderate amounts of basic nutrients and trace elements, to be mycorrhizal. As far as we are
aware of, this is the first published record of occurrence of AM in vetiver grass. McGee et al. (2003)
studied the role of mycorrhizae in revegetation of a waste disposal area south of Sydney, Australia, and
reported establishment of seedlings of native plants and mycorrhizal fungi in these extremely disturbed
conditions. Thus it is possible to return the degraded site to the original condition, providing we know the
nature of plants and their mycorrhizal associates (McGee et al., 2003). The authors also found that
mycorrhizal fungi were associated with stabilization of the soil, especially between widely spaced plants.
Soil stabilization, in turn, will reduce erosion and assist in revegetation extremely impoverished soils.

5     REFERENCES
Akhila A, and Rani M. 2002. Chemical constituents and essential oil biogenesis in Vetiveria zizanioides.
   In: Maffei A, ed. Vetiveria ­ The genus Vetiveria. London: Taylor and Francis
Bradshaw AD. 1987. Reclamation of land and ecology of ecosystems. In: RJ William, ME Gilpin, and JD
   Aber, (eds.). Restoration Ecology. Cambridge Uni Press, Cambridge. 53-74
Brazi F, Naidu R, and McLaughlin MJ. 1996. in R Naidu, RS Kookana, DP Olivers, S Rogers and MJ
   McLaughlin (eds.),Contaminants and the Soil Environment in the Australia Pacific Region, Kluwer
   Academic Publishers, Netherlands. 451
Brooks RR. 1997. Plants That Accumulate Heavy Metals. CAB International, Wallingford, Oxon, UK
Brundrett MC. 2002. Coevolution of roots and mycorrhizas of land plants. New Phytologist,154: 275-304
Burke SC, Angle JS, Chaney RL, et al. 2000. Arbuscular mycorrhizae effects on heavy metal
   uptake by Corn. Internatioan Jour Phytoremediation, 2: 23-29
Burns RG, Rogers S, and McGhee I. 1996. Remediation of inorganics and organics in industrial and
   urban contaminated soils. in R Naidu, RS Kookana, DP Olivers, S Rogers and MJ McLaughlin (eds.),
   Contaminants and the Soil Environment in the Australia Pacific Region, Kluwer Academic
   Publishers, Netherlands. 361-410
Channey, Oliver Chaudhry TM, Hayes WJ, et al., 1998. Phytoremediation ­ Focusing on accumulator
   plants that remediation metal-contaminated soils. Australian Journal of Ectoxicology 41: 37-51
Chaudhry TM, Hill L, Khan, AG, et al., 1999. Colonization of iron and zinc-contaminated dumped filter-
   cake waste by microbe, plants and associated mycorrhizae In: Wong MH., Wong JWC, Baker AJM,
   eds, Remediation and Management of Degraded Land. CRC Press LLC, Boca Raton, Chap, 27: 275-
   283
Davies FT, Puryear JD, Newton RJ, et al. 2002. Mycorrhizal fungi increase chromium uptake by
     sunflower plants: Influence on tissue mineral concentration, growth, and gas exchange. Jour Pl Nutri.,
     25: 2389-2407
Diaz G, Azcon-Aguilar C, and Honrubia M. 1996. Influence of arbuscular mycorrhizae on heavy metal
     (Zn and Pb) uptake and growth of Lygeum spartum and Anthyllis cytisoides. Plant and Soil, 180:
     241-249
Edwards D, Wellman CH, and Axe L. 1998. The fossil record of early land plants and relationships
     between primitive embryophytes: too little and too late? In Bryology for the Twenty First- Century.
     JW Bates, NS Ashton and JG Duckett (eds.). Maney Publishing and British Bryological Society,
     Leeds, UK. 15-43
Ernst WHO. 1996. Bioavailability of heavy metals and decontamination of soils by plants. Appl.
     Geochem. 11: 163-167
Fitter AH, and Stickland TR. 1991. Architectural analysis of plant root system. 2. Influence of nutrient
     supply on architecture in contrasting plant species. New Phytol., 118: 383-389
Flathman PE, Jerger DE, and Exner JH. 1994. Bioremediation Field Experiences. CRC Press, Boca
     Raton, Florida. 548
Greenfield JC. 1995. Vetiver grass (Vetiveria spp.): the ideal plant for vegetative soil and moisture
     conservation. In: Grimshaw RG, Helfer L, Eds. Vetiver grass for soil and water conservation, land
     rehabilitation, and embankment stabilization. Washington, DC: The World Bank. 3-38
Greenfield JC. 1993. Vetiver grass: The hedge against erosion. 4th ed. Washington DC: The World Bank
Greenfield JC. 1989. Vetiver grass (Vetiveria zizanioides): the ideal plant for vegetative soil and water
     conservation. Washington DC: The World Bank
Greenfield JC. 1988. Vetiver grass (Vetiveria zizanioides): A method for soil and water conservation. PR
     Press Services Pvt. Ltd. New Delhi, India. 72
Grimshaw RG, and Helfer L (eds.) 1995. Vetiver grass for soil and water conservation, land
     rehabilitation, and embankment stabilization. World Bank Technical Paper no. 273. Washington DC:
     The World Bank
Hill RD, and Peart MR, 1999. Vetiver grass for erosion control: Hong Kong and Guizhou, China. In: MH
     Wong, JWS Wong, AJM Baker, Eds. Remediation and Management of Degraded Land. CRC Press
     LLC, Boca Raton, Chap, 29: 295-304
Jamal A, Ayub N, Usman M, et al., 2002. Arbuscular mycorrhizal fungi enhance zinc and nickel uptake
     from contaminated soil by soybean and lentil. International Journal Phytoremediation, 4(3): 205-221
Karagiannidis N, and Nikolaou N. 2000. Influence of arbuscular mycorrhizae on heavy metal (Pb and Cd)
     uptake, growth, and chemical composition of Vitis vinifera L. (cv. Razaki). American Society for
     Ecology and Viticulture, 51(3): 269-275
Keller C, Hammer D, Kayser A, et al. 2003. Root development and heavy metal phytoextraction
     comparison of different species in the field. Plant and Soil, 249: 67-81
Khan AG. 2001. Relationship between chromium biomagnification ratio, accumulation factor, and
     mycorrhizae in plants growing on tannery effluent-polluted soil. Environment International 26: 417-
     423
Khan AG. 2002a. The significance of microbes. In: Wong MH, Bradshaw AD, eds. The restoration and
     management of derelict land ­ Modern Approaches, Singapore: World Scientific. 80-92
Khan AG. 2002b. The handling of microbes. In: Wong MH, and Bradshaw AD, (eds.), The restoration
     and management of derelict land ­ Modern Approaches, Singapore: World Scientific. 149-160
Khan AG. 2003. Mycotrophy and its significance in wetland ecology and wetland management. In
    Proceedings Croucher Foundation Study Institute: Wetland Ecosystems in Asia ­ Function and
    Management, 11-15 March, 2003. Hong Kong
Khan AG, Kuek C, Chaudhry, TM, et al., 2000. Role of plants, mycorrhizae and phytochelators in heavy
    metal contaminated land remediation. Chemosphere, 41: 197-207
Kothari SK, Marschner H, and Romheld V. 1991. Contribution of VA mycorrhizal hyphae in acquisition
    of phosphorus and zinc by maize grown in a calcareous soil. Plant Soil, 131: 177-185
Lavania UC, and Lavania S. 2002. Vetiver grass technology for environmental technology and
    sustainable development. Current Science, 78(8): 944-946
Li XL, George DE, and Marschner H. 1991. Extention of the phosphorus depletion zone in VA-
    mycorrhizal white clover in a calcareous soil. Plant Soil, 136: 41-48
Lynch JM. 1982. The Rhizosphere, in RG Burns and JH Slater (Eds.): Experimental
    Microbial Ecology. Pp 395-411. Blackwell, Oxford, UK
McGee P, Pattinson G, and Markovina A. 2003. Mycorrhizas and revegetation. Australia Microbiology
    24(3): 32-33
Maffei, M. ed. 2002. Vetiveria: The genus Vetiveria. London: Taylor & Francis
Mucciarelli M, Bertea CM, Scannerini S, et al., 1998. Vetiveria zizanioides as a tool for environmental
    engineering. Acta Horticulturae, 23: 337-347
National Research Council, USA, 1993. Vetiver grass, a thin green line against erosion. Washington:
    National Academy Press
Palazzo AJ, and Lee CR. 1997. Root growth and metal uptake of plants grown on zinc-contaminated soils
    as influenced by soil treatment and plant species. In Extended abstracts of the 4th International
    Conference on the Biogeochemistry of Trace Elements, pp 441-442. 23-26 June 1997. Berkeley, CA
Pawlowska TE, Chaney RL, Chin, M, et al., 2000. Effects of metal phytoextraction practices on the
    indigenous community of arbuscular mycorrhizal fungi at a metal-contaminated landfill. Applied and
    Environmental microbiology, 66 (6): 2526-2530
Rao PSC, Davis GB, and Johnston CD. 1996. Technologies for enhanced remediation of contaminated
    soils and aquifers: an overview, analysis and case studies. in R Naidu, RS Kookana, DP Olivers, S
    Rogers and MJ McLaughlin (eds.),Contaminants and the Soil Environment in the Australia Pacific
    Region, Kluwer Academic Publishers, Netherlands. 629-646
Reeves R. 2003. Tropical hyperaccumulators of metals and their potential for phytoextraction. Planr Soil
    249: 57-65
Shu WS, Xia HP, Zhang ZQ, et al., 2002. Use of vetiver and three other grasses for revegetation of
    Pb/Zn mine tailings: Field experiment. International J. Phytoremediation, 4(1): 47-57
Simmon L, Bousquet J, Levesque RC, et al., 1993. Origin and diversification of endomycorrhizal fungi
    and coincidence with vascular land plants. Nature 363: 67-69
Smith SE, and Read DJ. 1997. Mycorrhizal symbiosis, 2nd edition. London: Academic Press
Salomons W, Forstner U, and Mader P. 1995. Heavy Metals: Problems and Solutions. Springer, Berlin.
    412
Truong P. 2002. Vetiver grass technology. In: Maffei A, (ed.), Vetiveria ­ The genus Vetiveria. London:
    Taylor and Francis. 114-132
Truong P. 1999. Vetiver Grass Technology for Mine Rehabilitation. Tech. Bull. No. 1999/2, PRVN /
    ORDPB, Bangkok, Thailand
Truong P, and Baker D. 1998. Vetiver Grass System for Environmental Protection. Tech. Bull. No.
    1998/1, PRVN / ORDPB, Bangkok, Thailand
Truong P, and Claridge J. 1996. Effect of heavy metals toxicities on vetiver growth. Bangkok, Thailand:
    Vetiver Newsletter, Number 15
Turnau K, and Haselwandter K. 2002. Arbuscular mycorrhizal fungi: an essential component of soil
    microflora in ecosystem restoration. In: S. Gianinazi, H. schuepp, JM Barea, K. Haselwandter, (eds.),
    Mycorrhizal technology in agriculture ­ from genes to bioproducts. Berlin: Birkhauser Verlag. 137-
    149
Vetiver Information Network (VIN). 1993. Vetiver Grass: Technical Information Package, 2 vols. USA
    National Research Council, National Academy Press, Washington, DC
Vietmeyer N. 2002. Beyond the vetiver hedge: Organizing vetiver's next steps to global acceptance. In:
    Maffei A, ed. Vetiveria ­ The genus Vetiveria. London: Taylor and Francis. 176-186
Walle RJ, and Sims BG. 1998. Natural terrace formation through vegetative barriers on
    hillside forms in Honduras. Amer J Alternative Agric, 13: 79-82
Weissenhorn I, and Leyval C. 1996. Spore germination of arbuscular mycorrhizal fungi in soil differing
    in heavy metal content and other parameters. European Journal of Soil Biology 32: 165-172
Wise DL, and Trantolo DJ. 1994. Remediation of Hazardous Wastes Contaminated Soils. Marcel Dekker,
    NY. 929
Wong CC. 2003. The Riole of Mycorrhizae associated with Vetiveria zizanioides and Cyperus
    polystachyos in the remediation of metals (lead and zinc) contaminated soils. M. Phil Thesis. Hong
    Kong Baptist University, Hong Kong
Zhu YG, Christie P, and Laidlaw AS. 2001. Uptake of Zn by arbuscular mycorrhizal white
    clover from Zn-contaminated soil. Chemosphere 42: 193-199
Zisong W. 1991. Excerpts from the experiments and popularization of Vetiver grass, Nanping prefecture,
    Fujian Province, China. Vetiver Newsletter, Number 20.