**Meet the editors**

Dr. Hemanth KN. Vasanthaiah graduated from University of Agricultural Sciences, Bangalore, India. He is presently working as a Research Scientist at Plant Biotechnology and Biochemistry Laboratory, Florida A & M University, USA. The overall goal of his investigation is to determine biochemical and molecular factors involved in disease tolerance, drought, nutraceutical and

enological characteristics of grapes, raspberry and peanuts. The distinction of his research is association with Muscadine and Florida Hybrid Bunch grapes, which are unique to southeastern United States where the cultivation of European grapes is limited due to the prevalence of pest and diseases. Dr. Vasanthaiah has accomplished significant progress and achievements to address some of the issues in plant science and the outcomes have been published in leading scientific journals. He has also received meritorious awards and quite a few grants as Co-PI and collaborator. Dr. Vasanthaiah also has a computer science degree to compliment with this research work.

Dr. Devaiah Kambiranda expertise is in plant biotechnology and biochemistry. He obtained PhD in biotechnology at Bangalore University, India, where he worked on identification of genes involved in drought tolerance in peanut plants. He did his Postdoctoral fellowship at Gyeongsang National University, South Korea, where he gained expertise in protein expression and purification.

He has also worked in the Indian Institute of Ayurvedic medicine where he developed molecular markers for several medicinal plants. Currently he is working as a Research Associate in Florida A&M University where he is working on identification of genes and proteins involved in carbohydrate metabolism, disease tolerance and berry ripening in Florida hybrid and Muscadine grapes. He has authored about thirteen publications in peer reviewed journals and has presented in several national and International conferences.

Contents

**Preface IX** 

Chapter 1 **Enhancing Phytoremediation Efficiency in** 

Mohsen Soleimani, Samira Akbar and Mohammad Ali Hajabbasi

Chapter 2 **Morphophysiological Investigations in** 

Chapter 3 **Alteration of Abiotic Stress Responsive** 

**Jasmonic Acid Elicitation 49** 

SM Chabregas, WL Burnquist, AM Benko-Iseppon and EA Kido

Chapter 5 **Effect of UV Light on Secondary Metabolite** 

Chapter 6 **Drought Tolerance and Stress Hormones:** 

Chapter 7 **Iron Stress in Citrus 165** 

Aimar D., Calafat M., Andrade A.M., Carassay L., Abdala G.I. and Molas M.L.

Chapter 4 **Transcriptomics of Sugarcane** 

**Response to Environmental Pollution Stress 1** 

**Some Dominant Alien Invasive Weeds 15**  Nivedita Ghayal and Kondiram Dhumal

Ismanizan Ismail, Mian-Chee Gor, Zeti-Azura Mohamed-Hussein,

Lorena Almagro, Ana Belén Sabater-Jara, Sarai Belchí-Navarro, Francisco Fernández-Pérez, Roque Bru and María A. Pedreño

**Genes in** *Polygonum minus* **Roots by** 

Zamri Zainal and Normah Mohd Noor

**Osmoprotectants Under Drought 89**  RLO Silva, JRC Ferreira Neto, V Pandolfi,

**Biosynthesis in Plant Cell Cultures Elicited with Cyclodextrins and Methyl Jasmonate 115** 

**From Model Organisms to Forage Crops 137** 

Maria Angeles Forner-Giner and Gema Ancillo

### Contents

### **Preface IX**



Devaiah M. Kambiranda, Hemanth KN. Vasanthaiah, Ramesh Katam Athony Ananga, Sheikh M. Basha and Karamthotsivasankar Naik

### Preface

The purpose of this book is to discuss intricate problems associated with plant performance under hostile conditions. Plants are sessile organisms and are frequently exposed to extreme stresses. Therefore, they have to be considerably more adaptable to stressful environments and must acquire greater tolerance to multiple stresses. Individual plant performance varies with their magnitude of tolerance. Among the stresses, abiotic stress is known to repeatedly limit the growth and productivity of plants, which negatively affects the yield, quality and other characteristics of the crops. As plants do not posses immune system, they have evolved a complex and dynamic defense system for their adaptation in response to stress. Drought and temperature are the major abiotic stresses that affect the plants, along with light, salinity and minerals. Prolonged abiotic stress may cause irreversible damage to plant function or development.

The possibilities for increasing tolerance to abiotic stresses are enormous. Progress has been made worldwide to utilize advanced tools and techniques from all branches of science in order to understand how plants respond to abiotic stresses with the aim of helping to manipulate plant performance that will be better suited to withstand these stresses. Though several projects have been developed to tackle effect of stress, many questions still remain unanswered. Therefore, an effort is made through this publication to show some of the current research and critical roles of genetics, physiology, biochemistry, biotechnology, and other related science in crop improvement for the benefit of fellow researchers. It is necessary to constantly get acquainted with the past and present occurrences in order to learn lessons that could help in the acquisition of new knowledge or the further development of appropriate technology ensuing from it. Among the high-priority research areas identified in plant sciences, developing crops with high yielding characteristics under adverse conditions is gaining attention in view of the future predictable earth's environment and also for possible establishments of plants on other terrestrial bodies in our universe.

In this book an attempt has been made to unfold some of the new findings and achievements that will enhance ongoing research and help solve some of the pressing issues in plant science.

#### X Preface

This book presents the holistic view of the affect of various abiotic stresses on plants and their response to it. Chapters offer a critical discussion on the available literature and major problems , as well as understanding of plant stress responses and mechanisms of their tolerance in detail. I honestly trust that this book will be of great help to students, researchers and professionals in various fields of science, both in academic and industrial sectors.

I would like to express my deep sense of gratitude and indebtedness to all the authors for their valuable contributions and also to the researchers who actually performed experiments and reported their findings. I must confess that it had been a rare privilege for me to be associated with InTech publishers. Thanks is the least word to offer to Ms. Viktorija Zgela, Ms. Natalia Reinic and Ms. Dragana Manestar, Intech Publishing Process Managers, yet I shall avail this opportunity to extend my sincere gratitude for their help and co-operation at various phases of book publication. I would like to express my sincere thanks to Devaiah Kambiranda for helping me in editing some of the chapters. Last but not least I express my sincere thanks and affection to my wife Roopashri Puttaramu and my daughter Sahitya H. Setty for their sensible co-operation and cheerful encouragement. I hope the information available in this book will have a greater impact on the scientific world working towards crop improvement, which will pay off one day in the survival of life form on this beautiful planet, the Earth.

> **Dr. Hemanth KN. Vasanthaiah, M.C.A.** Center for Viticulture and Small Fruit Research Florida Agricultural & Mechanical University United States of America

X Preface

academic and industrial sectors.

planet, the Earth.

This book presents the holistic view of the affect of various abiotic stresses on plants and their response to it. Chapters offer a critical discussion on the available literature and major problems , as well as understanding of plant stress responses and mechanisms of their tolerance in detail. I honestly trust that this book will be of great help to students, researchers and professionals in various fields of science, both in

I would like to express my deep sense of gratitude and indebtedness to all the authors for their valuable contributions and also to the researchers who actually performed experiments and reported their findings. I must confess that it had been a rare privilege for me to be associated with InTech publishers. Thanks is the least word to offer to Ms. Viktorija Zgela, Ms. Natalia Reinic and Ms. Dragana Manestar, Intech Publishing Process Managers, yet I shall avail this opportunity to extend my sincere gratitude for their help and co-operation at various phases of book publication. I would like to express my sincere thanks to Devaiah Kambiranda for helping me in editing some of the chapters. Last but not least I express my sincere thanks and affection to my wife Roopashri Puttaramu and my daughter Sahitya H. Setty for their sensible co-operation and cheerful encouragement. I hope the information available in this book will have a greater impact on the scientific world working towards crop improvement, which will pay off one day in the survival of life form on this beautiful

> **Dr. Hemanth KN. Vasanthaiah, M.C.A.** Center for Viticulture and Small Fruit Research Florida Agricultural & Mechanical University

> > United States of America

**1** 

*Iran* 

**Enhancing Phytoremediation Efficiency in** 

Mohsen Soleimani, Samira Akbar and Mohammad Ali Hajabbasi

*University of Guilan, Department of Soil Science,* 

*Isfahan University of Technology, Department of Soil Science,* 

**Response to Environmental Pollution Stress** 

Environmental pollution, particularly contamination of soil and water resources, has been accelerated as a result of global industrialization and so is considered as a major risk for human communities throughout the world. Due to the adverse effects of organic and inorganic pollutants on human health and environmental safety, it is necessary to be removed in order to minimize the entry of these potentially toxics into the food chain. There are several methods to remove the soil pollutants which are categorized into 3 main parts including chemical, physical and biological methods. While conventional methods of soil clean-up including solidification, vitrification, electrokinetic, excavation, soil washing and flushing, oxidation and reduction etc. have shown to be effective in small areas, they need special equipments and are labor intensive. However, due to the side effects and highly costs of physical and chemical techniques, the biological methods especially phytoremediation, seem to be promising remedial strategies and so are highlighted as alternative techniques to traditional methodologies. Although phytoremediation as a "green technology" has shown many encouraging results, there have also been numerous inconclusive and unsuccessful attempts, especially in the field conditions, mostly because of biotic and abiotic stresses. "Abiotic stress is defined as the negative impact of non-living factors on the living organisms in a specific environment" (http://en.wikipedia.org). Abiotic stressors as the plant stress factors including high concentration of organic and inorganic pollutants, salinity, drought, flooding etc**.** could be considered as the main general themes adversely affect phytoremediation efficiency. Therefore, decrease of abiotic stresses is considered as a promising approach to introduce phytoremediation technique more applicable even though in the present of environmental stressors which significantly affect plant growth and

development (Dimkpa et al., 2009; Gerhardt et al., 2009; Weyens et al., 2009).

In this chapter we have discussed phytoremediation and its various types, as well as the plant response to abiotic stresses and the mechanisms which could be efficient to enhance phytoremediation efficiency regarding to abiotic stresses, especially considering

Phytoremediation is defined as an environmental friendly, cheap and large scale method which uses plants and their associated microorganisms to degrade, stabilize, reduce and/or

**1. Introduction** 

environmental pollutants.

**2. Phytoremediation** 

## **Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress**

Mohsen Soleimani, Samira Akbar and Mohammad Ali Hajabbasi *University of Guilan, Department of Soil Science, Isfahan University of Technology, Department of Soil Science, Iran* 

### **1. Introduction**

Environmental pollution, particularly contamination of soil and water resources, has been accelerated as a result of global industrialization and so is considered as a major risk for human communities throughout the world. Due to the adverse effects of organic and inorganic pollutants on human health and environmental safety, it is necessary to be removed in order to minimize the entry of these potentially toxics into the food chain. There are several methods to remove the soil pollutants which are categorized into 3 main parts including chemical, physical and biological methods. While conventional methods of soil clean-up including solidification, vitrification, electrokinetic, excavation, soil washing and flushing, oxidation and reduction etc. have shown to be effective in small areas, they need special equipments and are labor intensive. However, due to the side effects and highly costs of physical and chemical techniques, the biological methods especially phytoremediation, seem to be promising remedial strategies and so are highlighted as alternative techniques to traditional methodologies. Although phytoremediation as a "green technology" has shown many encouraging results, there have also been numerous inconclusive and unsuccessful attempts, especially in the field conditions, mostly because of biotic and abiotic stresses. "Abiotic stress is defined as the negative impact of non-living factors on the living organisms in a specific environment" (http://en.wikipedia.org). Abiotic stressors as the plant stress factors including high concentration of organic and inorganic pollutants, salinity, drought, flooding etc**.** could be considered as the main general themes adversely affect phytoremediation efficiency. Therefore, decrease of abiotic stresses is considered as a promising approach to introduce phytoremediation technique more applicable even though in the present of environmental stressors which significantly affect plant growth and development (Dimkpa et al., 2009; Gerhardt et al., 2009; Weyens et al., 2009).

In this chapter we have discussed phytoremediation and its various types, as well as the plant response to abiotic stresses and the mechanisms which could be efficient to enhance phytoremediation efficiency regarding to abiotic stresses, especially considering environmental pollutants.

### **2. Phytoremediation**

Phytoremediation is defined as an environmental friendly, cheap and large scale method which uses plants and their associated microorganisms to degrade, stabilize, reduce and/or

Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress 3

Plants can also remove toxic substances from soil through phytovolatization. In this process, the soluble contaminants are taken up by the roots, transported to the leaves, and volatized into the atmosphere through the stomata (Vara Prasad & Freitas, 2003; Pilon-Smits, 2005). Some heavy metals such as Hg, As and Se are inactivated when they are translocated from

Phytorestoration involves the complete remediation of contaminated soils to fully functioning soils which is an attempt to return the land to its natural state (Bradshaw, 1997).

Plants react to environmental stresses on various levels including biochemical, cellular and morphological scales depending on type of species or population (Mulder & Breure, 2003). These mechanisms include production of reactive oxygen species by autoxidation and Fenton reaction (for Fe and Cu), blocking of essential functional groups in biomolecules (for Cd and Hg), and displacement of essential metal ions from biomolecules for different kind of heavy metals (Schützendübe & Polle, 2002). Malondialdehyde is a major cytotoxic product of lipid peroxidation and acts as an indicator of free radicals which its production together with chlorophyll, caretonoids, as stress markers, increases in response to metal stress (Ben Ghnaya et al., 2009). Basically, reaction of plants to abiotic stresses depends on type of plant species having fundamental differences in development and anatomy as well as environmental limiting factors (i.e. stressors) (Tester & Bacic, 2005). For example, while a

When plants are faced to metal stress (such as Cd) the abundance of stress-related proteins, like heat shock proteins, proteinases and pathogenesis-related proteins could be changed in leaves and roots. Since roots are not photosynthetical tissues, whereas metal stress could adversely affect CO2 uptake, electron transport in chloroplasts by damaging photosystem I and II in leaves, proteomic changes in plant tissues upon an abiotic stress exposure would be different (Kieffer et al. 2009). Hence, plant exposed to high level of heavy metals causes reduction in photosynthesis, water and nutrient uptake, growth inhibition and finally death (Yadav, 2010; Soleimani et al., 2010a; Kieffer et al. 2009). The biosynthesis of ethylene as a gaseous plant hormone could be induced in response to environmental stressors which affect germination, growth and development of plant species as well as defence and

Although, one abiotic stress can usually decrease the ability of plant to resist a second stress (Tester & Bacic, 2005), interaction of various environmental stresses might decrease or increase plant tolerance to the growth limiting factors. For example; though soil salinity usually increases Cd bioavailability in heavy metal polluted soils and subsequently induces their toxicity, chloride salinity increased tolerance of an halophyte species (*Atriplex halimus* L.) to Cd toxicity both by decreasing the absorption of heavy metal and by improving plant tolerance through an increase in the synthesis of osmoprotective compounds in its tissues (Lefevre et al., 2009). The opposite trends were reported in the case of wheat which salinity increased Cd absorption and translocation by plants exposed to the metal in a nutrient solution (Mühling & Läuchli, 2003 In Lefevre et al., 2009; Liu et al., 2007) and for *Elodea canadensis* (Michx.) and *Potamogeton natans* (L.) in the presence of Cd, Cu and Zn (Fritioff et

flood may kill most plants in a certain area, but rice would be thrived there.

the soil into the atmosphere by bonding to free radicals in the air (Pilon-Smits, 2005).

**2.5 Phytovolatization** 

**2.6 Phytorestoration** 

**3. Plant response to abiotic stresses** 

resistance (Glick, 2004; Kang et al., 2010).

remove organic and inorganic pollutants from the environment (Pilon-Smits, 2005). It can be achieved in several ways including phytoextraction, phytostabilization, phytodegradation, phytovolatilization, phytorestoration, phytomining, rhizosphere-enhanced degradation and rhizofiltration (Vara Prasad & Freitas 2003; Pilon-Smits, 2005).

### **2.1 Phytoextraction and phytomining**

Phytoextraction is the removal of heavy metals and metalloids by plant roots with subsequent transport to shoots (Vara Prasad & Freitas 2003). Generally, plants which can grow in heavy metal contaminated soils and waters are categorized to "tolerant", "indicators" and "hyperaccumulators" (Bert et al., 2003). A tolerant species can grow in contaminated soils while other plants have not this ability. For indicator species, there is a linear correlation between metal concentration in growth media and plant tissues. While both indicator and hyperaccumulator species are also tolerant, however since tolerant species could prevent entering metals to roots, they are not necessarily hyperaccumulators or indicators (Bert et al., 2003). Hyperaccumulators have a high potential to uptake and accumulate heavy metals or metalloids which could be more than 100 fold in comparison with common plants. A hyperaccumulator species has i) the ability to accumulate more than 100 μg g-1 dry weight Cd, 1000 μg g-1 dry weight Ni, Cu, Co, Pb, Se, As and 10000 μg g-1 dry weight Zn and Mn, and ii) the bioconcentration factor (i.e. the ratio of metal concentration in plant to soil) and translocation factor (i.e. the ratio of metal concentration in shoots to roots) greater than 1.0 (Sun et al, 2008). "The technique of phytomining involves growing a hyperaccumulator plant species, harvesting the biomass and burning it to produce a bioore" (Anderson et al., 1999).

### **2.2 Rhizofiltration**

Rhizofiltration is a promising technique for removal of heavy metals from aquatic environments using suitable plants which could accumulate metals in their roots and shoots (Vara Prasad & Freitas, 2003).

### **2.3 Phytostabilization**

Phytostabilization, where plants are used to stabilize rather than clean organic and inorganic pollutants in contaminated soils to prevent their movement to surface and groundwater and/or to prevent translocation of pollutants from plant roots to shoots. The latter would be important for prevention of pollutant's transport to the upper levels of food chain (Pilon-Smits, 2005; Vara Prasad & Freitas, 2003). Additionally, in phytostabilization plants accumulate pollutants in their roots or immobilize, precipitate and reduce soil contaminants. Phytostabilization could also be important for reduction of wind and water erosion (Vara Prasad & Freitas, 2003).

### **2.4 Phytodegradation and rhizosphere-enhanced degradation**

Degradation of organic pollutants which are easily entered into the plant tissues or in the rhizosphere through the plant enzymes called phytodegradation. If this phenomenon occurs in plant rhizosphere by enhancing the activity of degrading microorganisms through the release of root exudates, named rhizosphere-enhanced degradation, which in fact is achieved by microbial enzymes rather than plant enzymes (Vara Prasad & Freitas, 2003; Pilon-Smits, 2005).

### **2.5 Phytovolatization**

2 Plants and Environment

remove organic and inorganic pollutants from the environment (Pilon-Smits, 2005). It can be achieved in several ways including phytoextraction, phytostabilization, phytodegradation, phytovolatilization, phytorestoration, phytomining, rhizosphere-enhanced degradation and

Phytoextraction is the removal of heavy metals and metalloids by plant roots with subsequent transport to shoots (Vara Prasad & Freitas 2003). Generally, plants which can grow in heavy metal contaminated soils and waters are categorized to "tolerant", "indicators" and "hyperaccumulators" (Bert et al., 2003). A tolerant species can grow in contaminated soils while other plants have not this ability. For indicator species, there is a linear correlation between metal concentration in growth media and plant tissues. While both indicator and hyperaccumulator species are also tolerant, however since tolerant species could prevent entering metals to roots, they are not necessarily hyperaccumulators or indicators (Bert et al., 2003). Hyperaccumulators have a high potential to uptake and accumulate heavy metals or metalloids which could be more than 100 fold in comparison with common plants. A hyperaccumulator species has i) the ability to accumulate more than 100 μg g-1 dry weight Cd, 1000 μg g-1 dry weight Ni, Cu, Co, Pb, Se, As and 10000 μg g-1 dry weight Zn and Mn, and ii) the bioconcentration factor (i.e. the ratio of metal concentration in plant to soil) and translocation factor (i.e. the ratio of metal concentration in shoots to roots) greater than 1.0 (Sun et al, 2008). "The technique of phytomining involves growing a hyperaccumulator plant species, harvesting the biomass and burning it to produce a bio-

Rhizofiltration is a promising technique for removal of heavy metals from aquatic environments using suitable plants which could accumulate metals in their roots and shoots

Phytostabilization, where plants are used to stabilize rather than clean organic and inorganic pollutants in contaminated soils to prevent their movement to surface and groundwater and/or to prevent translocation of pollutants from plant roots to shoots. The latter would be important for prevention of pollutant's transport to the upper levels of food chain (Pilon-Smits, 2005; Vara Prasad & Freitas, 2003). Additionally, in phytostabilization plants accumulate pollutants in their roots or immobilize, precipitate and reduce soil contaminants. Phytostabilization could also be important for reduction of wind and water

Degradation of organic pollutants which are easily entered into the plant tissues or in the rhizosphere through the plant enzymes called phytodegradation. If this phenomenon occurs in plant rhizosphere by enhancing the activity of degrading microorganisms through the release of root exudates, named rhizosphere-enhanced degradation, which in fact is achieved by microbial enzymes rather than plant enzymes (Vara Prasad & Freitas, 2003;

rhizofiltration (Vara Prasad & Freitas 2003; Pilon-Smits, 2005).

**2.1 Phytoextraction and phytomining** 

ore" (Anderson et al., 1999).

(Vara Prasad & Freitas, 2003).

erosion (Vara Prasad & Freitas, 2003).

**2.4 Phytodegradation and rhizosphere-enhanced degradation** 

**2.3 Phytostabilization** 

Pilon-Smits, 2005).

**2.2 Rhizofiltration** 

Plants can also remove toxic substances from soil through phytovolatization. In this process, the soluble contaminants are taken up by the roots, transported to the leaves, and volatized into the atmosphere through the stomata (Vara Prasad & Freitas, 2003; Pilon-Smits, 2005). Some heavy metals such as Hg, As and Se are inactivated when they are translocated from the soil into the atmosphere by bonding to free radicals in the air (Pilon-Smits, 2005).

### **2.6 Phytorestoration**

Phytorestoration involves the complete remediation of contaminated soils to fully functioning soils which is an attempt to return the land to its natural state (Bradshaw, 1997).

### **3. Plant response to abiotic stresses**

Plants react to environmental stresses on various levels including biochemical, cellular and morphological scales depending on type of species or population (Mulder & Breure, 2003). These mechanisms include production of reactive oxygen species by autoxidation and Fenton reaction (for Fe and Cu), blocking of essential functional groups in biomolecules (for Cd and Hg), and displacement of essential metal ions from biomolecules for different kind of heavy metals (Schützendübe & Polle, 2002). Malondialdehyde is a major cytotoxic product of lipid peroxidation and acts as an indicator of free radicals which its production together with chlorophyll, caretonoids, as stress markers, increases in response to metal stress (Ben Ghnaya et al., 2009). Basically, reaction of plants to abiotic stresses depends on type of plant species having fundamental differences in development and anatomy as well as environmental limiting factors (i.e. stressors) (Tester & Bacic, 2005). For example, while a flood may kill most plants in a certain area, but rice would be thrived there.

When plants are faced to metal stress (such as Cd) the abundance of stress-related proteins, like heat shock proteins, proteinases and pathogenesis-related proteins could be changed in leaves and roots. Since roots are not photosynthetical tissues, whereas metal stress could adversely affect CO2 uptake, electron transport in chloroplasts by damaging photosystem I and II in leaves, proteomic changes in plant tissues upon an abiotic stress exposure would be different (Kieffer et al. 2009). Hence, plant exposed to high level of heavy metals causes reduction in photosynthesis, water and nutrient uptake, growth inhibition and finally death (Yadav, 2010; Soleimani et al., 2010a; Kieffer et al. 2009). The biosynthesis of ethylene as a gaseous plant hormone could be induced in response to environmental stressors which affect germination, growth and development of plant species as well as defence and resistance (Glick, 2004; Kang et al., 2010).

Although, one abiotic stress can usually decrease the ability of plant to resist a second stress (Tester & Bacic, 2005), interaction of various environmental stresses might decrease or increase plant tolerance to the growth limiting factors. For example; though soil salinity usually increases Cd bioavailability in heavy metal polluted soils and subsequently induces their toxicity, chloride salinity increased tolerance of an halophyte species (*Atriplex halimus* L.) to Cd toxicity both by decreasing the absorption of heavy metal and by improving plant tolerance through an increase in the synthesis of osmoprotective compounds in its tissues (Lefevre et al., 2009). The opposite trends were reported in the case of wheat which salinity increased Cd absorption and translocation by plants exposed to the metal in a nutrient solution (Mühling & Läuchli, 2003 In Lefevre et al., 2009; Liu et al., 2007) and for *Elodea canadensis* (Michx.) and *Potamogeton natans* (L.) in the presence of Cd, Cu and Zn (Fritioff et

Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress 5

Understanding the plant response to abiotic stresses, mainly due to excess environmental pollutants, can be important in selecting a suitable approach to prevent decreasing

Due to limitations of phytoremediation such as low biomass of hyperaccumulator species, plant sensitivity to high concentrations of environmental pollutants as well as other abiotic stresses and less efficiency of ions and compounds which have low bioavailability to uptake by plants, several approaches have been mentioned in recent decays to boost the efficiency of this technology. Although there are some chemicals (e.g. surfactants and ligands) which may increase phytoextraction, phytodegradation or phytostimulation of pollutants through the enhancement of bioavailability of organic and in-organic compounds in media, naturebased methods like using plant-microorganisms symbiosis not only seem to be more acceptable due to having less side-effects by protection of food chain but could also be efficient in remediation process by increasing plant biomass (Weyens et al., 2009). In the following, we mainly discuss several approaches including plant symbiosis with fungi and bacteria as well as plant genetic engineering which have revealed improvement of

phytoremediation efficiency of various environmental pollutants consequently.

abiotic stresses in comparison to rhizospheric bacteria (Rajkumar et al., 2009).

reduce their related stress to plants in contaminated soils (Glick, 2010).

**4.1.1 Mechanisms underlying abiotic stress tolerance within the plant** 

Generally there are several bacterial species in the rhizosphere called rhizobacteria. Root zone bacteria which have shown beneficial effects on various plants are named plant growth-promoting rhizobacteria (PGPR) and categorized into 2 main parts; extracellular and intracellular PGPR (Dimkpa et al., 2009). The latter group includes bacteria which are capable of entering the plant as endophytic bacteria and are able to create nodules, whereas extracellular PGPR are found in the rhizosphere, rhizoplane or within the apoplast of the root cortex, but not inside the cells (Dimkpa et al., 2009; Rajkumar et al., 2009). Since endophytic bacteria live within the plant, they could be better protected from biotic and

Plant-associated bacteria can promote plant growth as well as reduce and/or control of environmental stresses which together affect phytoremediation efficiency through several approaches directly and indirectly, within the plant and/or in the rhizosphere (Dimkpa et al., 2009; Glick, 2004, 2010; Kang et al., 2010; Rajkumar et al., 2009; Weyens et al., 2009; Yang et al., 2009). Furthermore, in the case of organic pollutants, there are a number of soil microorganisms that are capable of degrading xenobiotic compounds and consequently

Regarding to plant-bacteria symbiosis, there are several mechanisms which induce abiotic stress tolerance within the plant or in the rhizosphere which are mentioned in the following.

1. Production of phytohormones (e.g. auxins, cytokinins, gibberellins) which can change root morphology is an adaptation mechanism of plant species exposed to environmental stresses (Dimkpa et al., 2009; Weyens et al., 2009). Indole acetic acid as a sub-group of auxins together with nitric oxide are produced in plant shoot transported

phytoremediation efficiency.

**4.1 Plant-bacteria symbiosis** 

They are as follows:

**4. Enhancing phytoremediation efficiency** 

al., 2005). It could also be considered that NaCl might increase the occurance of CdCl+ which may be absorbed by the roots and translocated to the shoots (Lefevre et al., 2009).

Abiotic stresses such as salinity and organic and inorganic pollutants could adversely affect seed germination of plants (Soleimani et al., 2010b; Besalatpour et al., 2008). However, some plants such as *Frankenia* species have been reported to germinate successfully even though in response to abiotic stresses which demonstrate their uses in remediation and revegetation projects in areas affected by salinity (Easton & Kleindorfer, 2009).

Another response of plants upon exposure to heavy metals is oxidative stress which leads to cellular damage. In addition, metal accumulation by plant tissues disturbs cellular ionic homeostasis (Yadav, 2010). Salts and heavy metals could induce oxidative stress in plant which generate active oxygen species and consequently damage plant photosynthetic apparatus resulting in a loss of chlorophyll content and decline in photosynthetic rate and biomass production as well (Qureshi et al., 2005). Total antioxidant activity may increase with increasing environmental pollutants suggesting the capacity of plant to enhance antioxidant defense in response to pollutant stress. Antioxidant enzymes (e.g. dehydroascorbate reductase, glutathione peroxidase, glutathione-S-transferase and superoxide dismutases) may play an important role in plant cell against environmental abiotic stressors (Babar Ali et al., 2005). Reduced forms of phytophenolics act as antioxidant in plant facing to heavy metal stress, while oxidized form (i.e. phenoxyl radicals) can exhibit prooxidant activities under conditions that prolong the radical life time (Dimkpa et al., 2009; Sakihama et al., 2002). Hence, Johnstone et al. (2005) suggested that the test of total antioxidant activity could be mentioned as a new approach to identify putative algal phytoremediator as well as to monitor the effects of water quality on the biological components of polluted aquatic ecosystems.

Generally, the main mechanisms of higher plants in the presence of a metal stress include: stimulation of antioxidant systems in plants, complexation or co-precipitation, immobilization of toxic metal ions in growth media, uptake processes and compartmentation of metal ions within plants (Pilon-Smits, 2005; Liang et al., 2007; Jahangir et al., 2008). To minimize the detrimental effects of heavy metal stress, plants use detoxification mechanisms which are mainly based on chelation and subcellular compartmentalization (Mejáre & Bülow, 2001; Yadav, 2010). A principal class of heavy metal chelator known in plants is phytochelatins (PCs), a family of Cys-rich peptides. PCs are synthesized non-translationally from reduced glutathione in a transpeptidation reaction catalyzed by the enzyme phytochelatin synthase. Therefore, availability of glutathione is very essential for PCs synthesis in plants at least during their exposure to heavy metals (Yadav, 2010). One strategy of plants against xenobiotic stress such as phytotoxic chlorophenols is increasing of extracellular peroxidases enzymes capable of catalyzing their oxidative dechlorination which could be a protection approach of some aquatic plants (e.g. *Spirodela punctata*) against pollution stress (Jansen et al., 2004).

In the case of hyperaccumulators which are extensively used to remediate soil contaminated with heavy metals, the major involved processes in response to excess amounts of metals are i) bioactivation of metals in the rhizosphere through root–microbe interaction, ii) enhanced uptake by metal transporters in the plasma membranes, iii) detoxification of metals by chelation with phytochelatins, metallothioneins, metal-binding proteins in the cytoplasm and/or cell wall, and iv) sequestration of metals into the vacuole by tonoplast-located transporter proteins (Yang et al., 2005).

Understanding the plant response to abiotic stresses, mainly due to excess environmental pollutants, can be important in selecting a suitable approach to prevent decreasing phytoremediation efficiency.

### **4. Enhancing phytoremediation efficiency**

Due to limitations of phytoremediation such as low biomass of hyperaccumulator species, plant sensitivity to high concentrations of environmental pollutants as well as other abiotic stresses and less efficiency of ions and compounds which have low bioavailability to uptake by plants, several approaches have been mentioned in recent decays to boost the efficiency of this technology. Although there are some chemicals (e.g. surfactants and ligands) which may increase phytoextraction, phytodegradation or phytostimulation of pollutants through the enhancement of bioavailability of organic and in-organic compounds in media, naturebased methods like using plant-microorganisms symbiosis not only seem to be more acceptable due to having less side-effects by protection of food chain but could also be efficient in remediation process by increasing plant biomass (Weyens et al., 2009). In the following, we mainly discuss several approaches including plant symbiosis with fungi and bacteria as well as plant genetic engineering which have revealed improvement of phytoremediation efficiency of various environmental pollutants consequently.

### **4.1 Plant-bacteria symbiosis**

4 Plants and Environment

al., 2005). It could also be considered that NaCl might increase the occurance of CdCl+ which

Abiotic stresses such as salinity and organic and inorganic pollutants could adversely affect seed germination of plants (Soleimani et al., 2010b; Besalatpour et al., 2008). However, some plants such as *Frankenia* species have been reported to germinate successfully even though in response to abiotic stresses which demonstrate their uses in remediation and revegetation

Another response of plants upon exposure to heavy metals is oxidative stress which leads to cellular damage. In addition, metal accumulation by plant tissues disturbs cellular ionic homeostasis (Yadav, 2010). Salts and heavy metals could induce oxidative stress in plant which generate active oxygen species and consequently damage plant photosynthetic apparatus resulting in a loss of chlorophyll content and decline in photosynthetic rate and biomass production as well (Qureshi et al., 2005). Total antioxidant activity may increase with increasing environmental pollutants suggesting the capacity of plant to enhance antioxidant defense in response to pollutant stress. Antioxidant enzymes (e.g. dehydroascorbate reductase, glutathione peroxidase, glutathione-S-transferase and superoxide dismutases) may play an important role in plant cell against environmental abiotic stressors (Babar Ali et al., 2005). Reduced forms of phytophenolics act as antioxidant in plant facing to heavy metal stress, while oxidized form (i.e. phenoxyl radicals) can exhibit prooxidant activities under conditions that prolong the radical life time (Dimkpa et al., 2009; Sakihama et al., 2002). Hence, Johnstone et al. (2005) suggested that the test of total antioxidant activity could be mentioned as a new approach to identify putative algal phytoremediator as well as to monitor the effects of water quality on the biological

Generally, the main mechanisms of higher plants in the presence of a metal stress include: stimulation of antioxidant systems in plants, complexation or co-precipitation, immobilization of toxic metal ions in growth media, uptake processes and compartmentation of metal ions within plants (Pilon-Smits, 2005; Liang et al., 2007; Jahangir et al., 2008). To minimize the detrimental effects of heavy metal stress, plants use detoxification mechanisms which are mainly based on chelation and subcellular compartmentalization (Mejáre & Bülow, 2001; Yadav, 2010). A principal class of heavy metal chelator known in plants is phytochelatins (PCs), a family of Cys-rich peptides. PCs are synthesized non-translationally from reduced glutathione in a transpeptidation reaction catalyzed by the enzyme phytochelatin synthase. Therefore, availability of glutathione is very essential for PCs synthesis in plants at least during their exposure to heavy metals (Yadav, 2010). One strategy of plants against xenobiotic stress such as phytotoxic chlorophenols is increasing of extracellular peroxidases enzymes capable of catalyzing their oxidative dechlorination which could be a protection approach of some aquatic plants (e.g.

In the case of hyperaccumulators which are extensively used to remediate soil contaminated with heavy metals, the major involved processes in response to excess amounts of metals are i) bioactivation of metals in the rhizosphere through root–microbe interaction, ii) enhanced uptake by metal transporters in the plasma membranes, iii) detoxification of metals by chelation with phytochelatins, metallothioneins, metal-binding proteins in the cytoplasm and/or cell wall, and iv) sequestration of metals into the vacuole by tonoplast-located

may be absorbed by the roots and translocated to the shoots (Lefevre et al., 2009).

projects in areas affected by salinity (Easton & Kleindorfer, 2009).

components of polluted aquatic ecosystems.

transporter proteins (Yang et al., 2005).

*Spirodela punctata*) against pollution stress (Jansen et al., 2004).

Generally there are several bacterial species in the rhizosphere called rhizobacteria. Root zone bacteria which have shown beneficial effects on various plants are named plant growth-promoting rhizobacteria (PGPR) and categorized into 2 main parts; extracellular and intracellular PGPR (Dimkpa et al., 2009). The latter group includes bacteria which are capable of entering the plant as endophytic bacteria and are able to create nodules, whereas extracellular PGPR are found in the rhizosphere, rhizoplane or within the apoplast of the root cortex, but not inside the cells (Dimkpa et al., 2009; Rajkumar et al., 2009). Since endophytic bacteria live within the plant, they could be better protected from biotic and abiotic stresses in comparison to rhizospheric bacteria (Rajkumar et al., 2009).

Plant-associated bacteria can promote plant growth as well as reduce and/or control of environmental stresses which together affect phytoremediation efficiency through several approaches directly and indirectly, within the plant and/or in the rhizosphere (Dimkpa et al., 2009; Glick, 2004, 2010; Kang et al., 2010; Rajkumar et al., 2009; Weyens et al., 2009; Yang et al., 2009). Furthermore, in the case of organic pollutants, there are a number of soil microorganisms that are capable of degrading xenobiotic compounds and consequently reduce their related stress to plants in contaminated soils (Glick, 2010).

Regarding to plant-bacteria symbiosis, there are several mechanisms which induce abiotic stress tolerance within the plant or in the rhizosphere which are mentioned in the following.

### **4.1.1 Mechanisms underlying abiotic stress tolerance within the plant**

They are as follows:

1. Production of phytohormones (e.g. auxins, cytokinins, gibberellins) which can change root morphology is an adaptation mechanism of plant species exposed to environmental stresses (Dimkpa et al., 2009; Weyens et al., 2009). Indole acetic acid as a sub-group of auxins together with nitric oxide are produced in plant shoot transported

Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress 7

One of the approaches to enhance phytoremediation efficiency is using of plant-fungi association. In this regard using of arbuscular mycorrhizal fungi (AMF) that are naturally present in the roots of most plant species where they form a mutualistic association, as well as endophytic fungi which live systemically within the aerial portion of many grass species, can improve plant tolerance to biotic and abiotic stresses (Hildebrandt et al., 2007; Kuldau & Bacon 2008; Lingua et al., 2008; Soleimani et al., 2010a). The role of these groups of fungi on

Environmental stresses such as organic and inorganic pollutants trigger oxidative stress commonly showed by increasing content of malondialdehyde in plant. Mycorrhizal fungi are important factors which have the ability to regulate oxidative stress (i.e. reducing the amount of malondialdehyde), as a general strategy, to protect plants from abiotic and biotic stresses (Bressano et al., 2010). Several researches have revealed using AMF as a useful method showing more beneficial effects of phytoremediation, especially in metal contaminated soils (Bressano et al., 2010; Jiang et al., 2008; Lingua et al., 2008; Schützendübe & Polle, 2002). The enhancement of phytoextraction efficiency of *Brassica juncea* L. inoculated with Acacia-associated fungi reported by Jiang et al. (2008). It has been confirmed that mycorrhizal fungi in association with poplars are suitable for phytoremediation purposes underscore the importance of appropriate combinations of plant genotypes and fungal symbionts (Lingua et al., 2008). Furthermore, some fungi have the potential to degrade organic pollutants via extracellular or intracellular oxidation using various enzymes such as laccase, peroxidase, nitroreductase and transferases (Harm et al., 2011), and thereafter

AMF can reduce metal stress in host plants or improve plant growth and development via several ways. Production and excretion of organic acids (e.g. citrate and oxalate) may increase dissolution of primary minerals containing phosphate which is one of the main nutrients for plant (Harm et al., 2011). Furthermore, release of siderophores can enhance iron uptake by plant and boost the growth. In the other hand, increasing the metal solubility or metal-complexing through acidification of the mycosphere could enhance metal uptake by plants which is important in phytoextraction. Extra-hyphal immobilization may occur through the complexation of metals by glomalin (i.e. metal-sorbing glycoproteins excreted by AMF) and biosorption to cell wall constituents such as chitin and chitosan (Harm et al., 2011). This process could be important in phytostabilization of heavy metals in contaminated soils considering that fungal mycelia and glomain could only increase soil aggregate stability against wind and water erosion (Harm et al., 2011). Methalothionin is another protein excreted by some mycorrhizal fungi which can also be important to reduce heavy metal stress in plants (Schutzendubel & Polle, 2002). It would be possible for heavy metals to storage in vacuoles or complex by cytoplasmic metallothioneins in fungi cells or volatilize via metal transformation (Harm et al., 2011). To alleviate heavy metal stress in plants associated with AMF, several genes encoding proteins (e.g. metallothionein, 90 kD heat shock protein, Glutathione-S-transferase) potentially involved in metal tolerance are expressed which are varied in their response to different heavy metals (Hildebrandt et al., 2007). However, improvement of plant mineral nutrition and health and also detoxification of metals in plants associated with AMF could be important to use them in phytoremediation of soil and water contaminated with heavy metals and/or organic

**4.2 Plant-fungi symbiosis** 

**4.2.1 AMF and plant abiotic stress** 

reduce stress of organic compounds in soil.

pollutants (Lingua et al., 2008).

reducing abiotic stresses is mentioned in the following.

to root tips and consequently enhance cell elongation, root growth, root surface area and development of lateral roots (Dimkpa et al., 2009).


### **4.1.2 Mechanisms underlying abiotic stress tolerance in the rhizosphere**

They are as follows:


### **4.2 Plant-fungi symbiosis**

6 Plants and Environment

2. Inoculation with non-pathogenic rhizobacteria can induce signaling cascades and plant systemic resistance, alter the selectivity for Na, K and Ca ions resulting in higher K/Na ratios and change in membrane phospholipid content as well as the saturation pattern

3. Bacteria may produce osmolytes, such as glycine betaine, act synergistically with plant

4. PGPR containing 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity reduces ethylene level within the plant and consequently facilitates plant growth under stress conditions (Dimkpa et al., 2009; Glick, 2004; Kang et al., 2010). The possible mechanisms described by Kang et al. (2010) accordingly. "ACC synthesized in plant tissues by ACC synthase is thought be exuded from plant roots and be taken up by neighboring bacteria. Subsequently, the bacteria hydrolyze ACC to ammonia and 2 oxobutanoate. This ACC hydrolysis maintains ACC concentrations low in bacteria and permits continuous ACC transfer from plant roots to bacteria. Otherwise, ethylene can be produced from ACC and then cause stress responses including growth inhibition."

1. Rhizobacterial with the capability of nitrogen fixation can positively influence on host plant growth by increasing nitrogen availability (Dimkpa et al., 2009; Kang et al., 2010; Rajkumar et al., 2009). Therefore they can act as a biofertilizer which affect plant growth

2. The mobility of heavy metals in contaminated soils can be significantly reduced through root zone bacteria which finally cause precipitation of metals as insoluble compounds in soil and sorption to cell components or intracellular sequestration

3. Bacterial migration from the rhizoplane to the rhizosphere plays a role in reducing plant uptake of some metals (e.g. Cd) in biologically unavailable complex forms

4. Iron–chelating siderophores complexes can be taken up by the host plant, resulting in a higher fitness (Dimkpa et al., 2009). They can also form complexes with other nonsoluble metals (e.g. Pb) and enhancing their ability to uptake by hyperaccumulators

5. Bacterial exopolysaccharides lead to the development of soil sheaths around the plant

6. Root zone bacteria can influence pH and redox potential in the rhizosphere, for instance, through the release of organic acids. This can have positive effects on the availability of nutrients (e.g. phosphorous) for the plant (Dimkpa et al., 2009; Weyens et

7. Under abiotic stress such as high concentration of environmental pollutants causing plant less tolerant in response to biotic stress (e.g. disease, pathogens), PGPR can act as biocontrol agents which mitigate the effect of pathogenic organisms (Gerhardt et al.,

root, which reduces the flow of sodium into the stele (Dimkpa et al., 2009).

and development of lateral roots (Dimkpa et al., 2009).

osmolytes, accelerating osmotic adjustment (Dimkpa et al., 2009).

**4.1.2 Mechanisms underlying abiotic stress tolerance in the rhizosphere** 

of lipids (Dimkpa et al., 2009).

(Kang et al., 2010).

(Gerhardt et al., 2009).

(Dimkpa et al., 2009).

(Dimkpa et al., 2009).

such as *Brassica napus* (Rajkumar et al., 2009).

al., 2009; Rajkumar et al., 2009).

2009)

They are as follows:

to root tips and consequently enhance cell elongation, root growth, root surface area

One of the approaches to enhance phytoremediation efficiency is using of plant-fungi association. In this regard using of arbuscular mycorrhizal fungi (AMF) that are naturally present in the roots of most plant species where they form a mutualistic association, as well as endophytic fungi which live systemically within the aerial portion of many grass species, can improve plant tolerance to biotic and abiotic stresses (Hildebrandt et al., 2007; Kuldau & Bacon 2008; Lingua et al., 2008; Soleimani et al., 2010a). The role of these groups of fungi on reducing abiotic stresses is mentioned in the following.

### **4.2.1 AMF and plant abiotic stress**

Environmental stresses such as organic and inorganic pollutants trigger oxidative stress commonly showed by increasing content of malondialdehyde in plant. Mycorrhizal fungi are important factors which have the ability to regulate oxidative stress (i.e. reducing the amount of malondialdehyde), as a general strategy, to protect plants from abiotic and biotic stresses (Bressano et al., 2010). Several researches have revealed using AMF as a useful method showing more beneficial effects of phytoremediation, especially in metal contaminated soils (Bressano et al., 2010; Jiang et al., 2008; Lingua et al., 2008; Schützendübe & Polle, 2002). The enhancement of phytoextraction efficiency of *Brassica juncea* L. inoculated with Acacia-associated fungi reported by Jiang et al. (2008). It has been confirmed that mycorrhizal fungi in association with poplars are suitable for phytoremediation purposes underscore the importance of appropriate combinations of plant genotypes and fungal symbionts (Lingua et al., 2008). Furthermore, some fungi have the potential to degrade organic pollutants via extracellular or intracellular oxidation using various enzymes such as laccase, peroxidase, nitroreductase and transferases (Harm et al., 2011), and thereafter reduce stress of organic compounds in soil.

AMF can reduce metal stress in host plants or improve plant growth and development via several ways. Production and excretion of organic acids (e.g. citrate and oxalate) may increase dissolution of primary minerals containing phosphate which is one of the main nutrients for plant (Harm et al., 2011). Furthermore, release of siderophores can enhance iron uptake by plant and boost the growth. In the other hand, increasing the metal solubility or metal-complexing through acidification of the mycosphere could enhance metal uptake by plants which is important in phytoextraction. Extra-hyphal immobilization may occur through the complexation of metals by glomalin (i.e. metal-sorbing glycoproteins excreted by AMF) and biosorption to cell wall constituents such as chitin and chitosan (Harm et al., 2011). This process could be important in phytostabilization of heavy metals in contaminated soils considering that fungal mycelia and glomain could only increase soil aggregate stability against wind and water erosion (Harm et al., 2011). Methalothionin is another protein excreted by some mycorrhizal fungi which can also be important to reduce heavy metal stress in plants (Schutzendubel & Polle, 2002). It would be possible for heavy metals to storage in vacuoles or complex by cytoplasmic metallothioneins in fungi cells or volatilize via metal transformation (Harm et al., 2011). To alleviate heavy metal stress in plants associated with AMF, several genes encoding proteins (e.g. metallothionein, 90 kD heat shock protein, Glutathione-S-transferase) potentially involved in metal tolerance are expressed which are varied in their response to different heavy metals (Hildebrandt et al., 2007). However, improvement of plant mineral nutrition and health and also detoxification of metals in plants associated with AMF could be important to use them in phytoremediation of soil and water contaminated with heavy metals and/or organic pollutants (Lingua et al., 2008).

Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress 9

hyperaccumulation or phytovolatization resulting in plants being more tolerant to heavy metals. Alteration of enzymes which are involved in oxidative stress may also produce an altered metal tolerance in plants. Since having highly branched root systems with large surface area is important for efficient uptake of toxic metals, introduction of genes affecting root biomass can improve rhizofiltration of heavy metals in some hyperaccumulator plants (Eapen & D'Souza, 2005). Besides, the phytoremediation potential of most hyperaccumulating plants is limited because of their low biomass and slow growth and close association with a special habitant. Therefore, biomass of hyperaccumulator plants can be changed by introduction of genes affecting phytohormone synthesis resulting in enhanced biomass (Eapen & D'Souza, 2005; Kotrba et al., 2009). Transgenic plants expressing bacterial ACC deaminase genes can decrease ethylene level which is a major problem that reduces phytoremediation efficiency in plants exposed to abiotic stresses

Genetically engineered plants can offer new characteristics that may not be met in normal plants (Table 1). Transgenic plants for phytoremediation presenting new or improved characteristics are engineered by the introduction and/or overexpression of genes taken from other organisms, such as bacteria or mammals. Bacteria and mammals are heterotrophs and have the enzymes necessary for achieving complete mineralization of organic pollutants; therefore bacterial and mammalian degradative enzymes can complete

Tolerance to toxic elements is a key factor in bioremediation. Plants which are more persistent in a harsh environment tend to maintain a high biomass and fast growth rate in regions unfavorable for growth and have more time for accumulating metals from the soil. (Eapen & D'Souza, 2005). Metal tolerance can significantly be increased by over-expression of proteins involved in intracellular metal sequestration but may not be utilized for metal accumulation. Tolerance and accumulation are highly independent traits, therefore they should both be manipulated to obtain a suitable plant for phytoremediation. (Eapen &

Increasing the plant's ability to convert a toxic element into a less toxic form can improve its tolerance to excess amount of that toxic trace element. Typically, such a plant could be able to accumulate higher amounts of the detoxified form. In order to create a model system for phytoremediation of heavy metals a MerP protein was expressed in transgenic *Arabidopsis*. The transgenic *Arabidpsis* showed higher tolerance and accumulation capacity for mercury, cadium and lead when compared with the control plant (Hsieh et al., 2009). Organomercury can be converted to metallic Hg volatized from the leaf surface in plants with capability to produce bacterial mercuric reductase and organomercurial lyase (Kotrba, et al., 2009). Besides, volatiziation of selenium compounds could be promoted via overexpressing genes encoding enzymes involved in production of gas methylselenide species (Kotrba, et al., 2009). Some of the genetically engineered plants and sources of the genes involved in the

Ferredoxin, a stress-sensitive protein, was replaced in tobacco chloroplasts by an isofunctional protein, a cyanobacterial flavodoxin that usually exist in photosynthetic microorganisms such as algae and bacteria and is missing in plants. The resulting transgenic plants showed tolerance to some abiotic stresses such as drought, chilling, oxidants, heat

There are specific genes called "pollutant-responsive elements" (PRE) that can be induced by the presence of particular toxic chemicals in the environment. They should be identified

(Kawahigashi, 2009).

the metabolic efficiencies of plants (Van Aken, 2008).

D'Souza, 2005; Karenlampi et al., 2000).

process are mentioned in Table 1.

and iron starvation (Zurbriggen et al., 2008).

### **4.2.2 Endophytic fungi and plant abiotic stress**

Endophytic fungi are a group of fungi that live their entire life cycle within the aerial portion of many grass species, forming nonpathogenic, systemic and usually intercellular associations (Soleimani et al., 2010a). Endophytes induce mechanisms of drought avoidance (morphological adaptations), drought tolerance (physiological and biochemical adaptations), and drought recovery in infected grasses (Malinowski and Belesky, 2000). In response to phosphorous deficiency, root morphology of host plant is altered or exudation of phenolic-like compounds may modify the rhizosphere conditions (Malinowski and Belesky, 2000). Aluminium toxicity mainly in acidic soils can be reduced on root surface of endophyte-infected plants through Al sequestration which appears to be related to exudation of phenolic-like compounds with Al-chelating activity (Malinowski and Belesky, 2000). Besides, drought and light stress as well as salt stress could be reduced in endophyteinfected plants via release of some proteins (e.g. dehydrins) and phenolic-like compounds in the rhizosphere (Kuldau & Bacon, 2008; Malinowski and Belesky, 2000). Several researches have also demonstrated the positive effect of endophytic fungi on phytoremediation of heavy metals as well as organic pollutants such as petroleum hydrocarbons (Soleimani et al., 2010a, 2010b). However, there is a lack of information regarding the effect of endophytic fungi on plant tolerance in response to stress of pollutants, especially organic pollutants, in both laboratory and field conditions.

#### **4.3 Transgenic plants**

Plants absorb toxic elements by the same pathways they take up essential elements. There should be a vast investigation on the processes involved in metal uptake, transport and storage by hyperaccumulating plants. Improving the number of absorption sites, changing specificity of uptake system to decrease competition by unwanted cations and enhancing intracellular binding sites should be considered to improve heavy metal accumulation in plants (Eapen & D'Souza, 2005). Modification of plant characteristics through the genetic engineering to enhance metal uptake, transport and accumulation as well as plant tolerance to abiotic stresses is a new approach for phytoremediation (Karenlampi,et al., 2000).

The first transgenic plants for phytoremediation were developed to enhance heavy metal tolerance including tobacco plants (*Nicotiana tabacum*) with a yeast metallothionein gene that gives tolerance to cadmium, and *Arabidopsis thaliana* that overexpressed a mercuric ion reductase gene for higher tolerance to mercury. Since each heavy metal may have a specific mechanism for uptake, therefore it is important to design suitable strategies for developing transgenic plants specific for each metal (Eapen & D'Souza, 2005).

There are different possible areas for genetic manipulation to create a suitable transgenic plant for phytoremediation. Generally, genes can be transferred from any living source to develop efficient transgenic plants for phytoremediation. Storage and detoxification of metals in some metal accumulating plants is due to metal storage in epidermal cells. Hence, genes can be inserted and/or overexpressed to produce metallothioneins, phytochelatins and metal chelators to improve plant tolerance and metal accumulation, thus play a role in detoxification of metals in plants (Eapen & D'Souza, 2005; Hassan et al., 2011). Genetic manipulation of metal transporters can be effective in modification of metal tolerance/accumulation in plants. One of the key essential features of metal hyperaccumulators is root-to-shoot translocation of ions. A strong metal sink in the shoots and improved xylem loading and repressed metal sequestration in root vacuoles are possible ways to enhance the root-to-shoot translocation (Hassan et al., 2011). Different metabolic pathways from various organisms can be presented into plants for

Endophytic fungi are a group of fungi that live their entire life cycle within the aerial portion of many grass species, forming nonpathogenic, systemic and usually intercellular associations (Soleimani et al., 2010a). Endophytes induce mechanisms of drought avoidance (morphological adaptations), drought tolerance (physiological and biochemical adaptations), and drought recovery in infected grasses (Malinowski and Belesky, 2000). In response to phosphorous deficiency, root morphology of host plant is altered or exudation of phenolic-like compounds may modify the rhizosphere conditions (Malinowski and Belesky, 2000). Aluminium toxicity mainly in acidic soils can be reduced on root surface of endophyte-infected plants through Al sequestration which appears to be related to exudation of phenolic-like compounds with Al-chelating activity (Malinowski and Belesky, 2000). Besides, drought and light stress as well as salt stress could be reduced in endophyteinfected plants via release of some proteins (e.g. dehydrins) and phenolic-like compounds in the rhizosphere (Kuldau & Bacon, 2008; Malinowski and Belesky, 2000). Several researches have also demonstrated the positive effect of endophytic fungi on phytoremediation of heavy metals as well as organic pollutants such as petroleum hydrocarbons (Soleimani et al., 2010a, 2010b). However, there is a lack of information regarding the effect of endophytic fungi on plant tolerance in response to stress of pollutants, especially organic pollutants, in

Plants absorb toxic elements by the same pathways they take up essential elements. There should be a vast investigation on the processes involved in metal uptake, transport and storage by hyperaccumulating plants. Improving the number of absorption sites, changing specificity of uptake system to decrease competition by unwanted cations and enhancing intracellular binding sites should be considered to improve heavy metal accumulation in plants (Eapen & D'Souza, 2005). Modification of plant characteristics through the genetic engineering to enhance metal uptake, transport and accumulation as well as plant tolerance

The first transgenic plants for phytoremediation were developed to enhance heavy metal tolerance including tobacco plants (*Nicotiana tabacum*) with a yeast metallothionein gene that gives tolerance to cadmium, and *Arabidopsis thaliana* that overexpressed a mercuric ion reductase gene for higher tolerance to mercury. Since each heavy metal may have a specific mechanism for uptake, therefore it is important to design suitable strategies for developing

There are different possible areas for genetic manipulation to create a suitable transgenic plant for phytoremediation. Generally, genes can be transferred from any living source to develop efficient transgenic plants for phytoremediation. Storage and detoxification of metals in some metal accumulating plants is due to metal storage in epidermal cells. Hence, genes can be inserted and/or overexpressed to produce metallothioneins, phytochelatins and metal chelators to improve plant tolerance and metal accumulation, thus play a role in detoxification of metals in plants (Eapen & D'Souza, 2005; Hassan et al., 2011). Genetic manipulation of metal transporters can be effective in modification of metal tolerance/accumulation in plants. One of the key essential features of metal hyperaccumulators is root-to-shoot translocation of ions. A strong metal sink in the shoots and improved xylem loading and repressed metal sequestration in root vacuoles are possible ways to enhance the root-to-shoot translocation (Hassan et al., 2011). Different metabolic pathways from various organisms can be presented into plants for

to abiotic stresses is a new approach for phytoremediation (Karenlampi,et al., 2000).

transgenic plants specific for each metal (Eapen & D'Souza, 2005).

**4.2.2 Endophytic fungi and plant abiotic stress** 

both laboratory and field conditions.

**4.3 Transgenic plants** 

hyperaccumulation or phytovolatization resulting in plants being more tolerant to heavy metals. Alteration of enzymes which are involved in oxidative stress may also produce an altered metal tolerance in plants. Since having highly branched root systems with large surface area is important for efficient uptake of toxic metals, introduction of genes affecting root biomass can improve rhizofiltration of heavy metals in some hyperaccumulator plants (Eapen & D'Souza, 2005). Besides, the phytoremediation potential of most hyperaccumulating plants is limited because of their low biomass and slow growth and close association with a special habitant. Therefore, biomass of hyperaccumulator plants can be changed by introduction of genes affecting phytohormone synthesis resulting in enhanced biomass (Eapen & D'Souza, 2005; Kotrba et al., 2009). Transgenic plants expressing bacterial ACC deaminase genes can decrease ethylene level which is a major problem that reduces phytoremediation efficiency in plants exposed to abiotic stresses (Kawahigashi, 2009).

Genetically engineered plants can offer new characteristics that may not be met in normal plants (Table 1). Transgenic plants for phytoremediation presenting new or improved characteristics are engineered by the introduction and/or overexpression of genes taken from other organisms, such as bacteria or mammals. Bacteria and mammals are heterotrophs and have the enzymes necessary for achieving complete mineralization of organic pollutants; therefore bacterial and mammalian degradative enzymes can complete the metabolic efficiencies of plants (Van Aken, 2008).

Tolerance to toxic elements is a key factor in bioremediation. Plants which are more persistent in a harsh environment tend to maintain a high biomass and fast growth rate in regions unfavorable for growth and have more time for accumulating metals from the soil. (Eapen & D'Souza, 2005). Metal tolerance can significantly be increased by over-expression of proteins involved in intracellular metal sequestration but may not be utilized for metal accumulation. Tolerance and accumulation are highly independent traits, therefore they should both be manipulated to obtain a suitable plant for phytoremediation. (Eapen & D'Souza, 2005; Karenlampi et al., 2000).

Increasing the plant's ability to convert a toxic element into a less toxic form can improve its tolerance to excess amount of that toxic trace element. Typically, such a plant could be able to accumulate higher amounts of the detoxified form. In order to create a model system for phytoremediation of heavy metals a MerP protein was expressed in transgenic *Arabidopsis*. The transgenic *Arabidpsis* showed higher tolerance and accumulation capacity for mercury, cadium and lead when compared with the control plant (Hsieh et al., 2009). Organomercury can be converted to metallic Hg volatized from the leaf surface in plants with capability to produce bacterial mercuric reductase and organomercurial lyase (Kotrba, et al., 2009). Besides, volatiziation of selenium compounds could be promoted via overexpressing genes encoding enzymes involved in production of gas methylselenide species (Kotrba, et al., 2009). Some of the genetically engineered plants and sources of the genes involved in the process are mentioned in Table 1.

Ferredoxin, a stress-sensitive protein, was replaced in tobacco chloroplasts by an isofunctional protein, a cyanobacterial flavodoxin that usually exist in photosynthetic microorganisms such as algae and bacteria and is missing in plants. The resulting transgenic plants showed tolerance to some abiotic stresses such as drought, chilling, oxidants, heat and iron starvation (Zurbriggen et al., 2008).

There are specific genes called "pollutant-responsive elements" (PRE) that can be induced by the presence of particular toxic chemicals in the environment. They should be identified

Enhancing Phytoremediation Efficiency in Response to Environmental Pollution Stress 11

many types of toxic chemicals (Kawahigashi, 2009). Special bacterial genes which encode enzymes involved in the breakdown of explosives, such as cytochrome P450 and nitroreductase have been also used in manipulating higher plants to enhance plant tolerance, uptake, and detoxification of contaminated environments (Van Aken, 2009). Therefore, using transgenic plants or combined use of them with microorganisms in the rhizosphere could be mentioned as a promising technique to reduce abiotic stresses in plants which are used in phytoremediation. Future researches, especially in the field

Abiotic stresses, especially due to high level of organic and inorganic pollutants, are major limiting factors which could adversely affect phytoremediation. To reduce the effects of these stresses in plants which are used in phytoremediation, using bacteria- or fungi symbiosis as well as plant genetic engineering could be a promising way to enhance remediation efficiency. In practical point of view, it should be considered that combined enhanced-phytoremediation approaches are possible to use too. Finally, understanding the involved mechanisms in the mentioned enhancing methods would be a useful tool to extend use of phytoremediation based on these approaches. Since most of researches have been carried out in laboratory

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conditions, can distinguish the efficiency of this approach.

conditions, field trials are needed to perform in contaminated sites.

**5. Conclusion** 

**6. Acknowledgment** 

15320383

review.

**7. References** 

and characterized so that they can be fused with reporter genes and be introduced into plants. For example the promoter of the barley gene HvhsplT which is expressed in the presence of some heavy metals had been fused to the reporter gene. This new gene combination was used to make a transformed tobacco plant which could be used as a bioindicator for monitoring heavy metal pollution (Mociardini, et al., 1998).

A combined use of transgenic plants and bacteria in the rhizosphere could improve phytoremediation of contaminated environments and may overcome the current limitations of phytoremediation such as low detoxification and absorption efficiency. The combination of plants for removing or degrading toxic pollutants and rhizospheric microorganisms for improving the availability of hydrophobic compounds can be effective in breaking down


Table 1. Genes involved in plant genetic engineering for phytoremediation of heavy metals (Hsieh et al., 2009; Karenlampi et al., 2000; Kotrba et al., 2009; Kolodyazhnaya et al., 2009).

many types of toxic chemicals (Kawahigashi, 2009). Special bacterial genes which encode enzymes involved in the breakdown of explosives, such as cytochrome P450 and nitroreductase have been also used in manipulating higher plants to enhance plant tolerance, uptake, and detoxification of contaminated environments (Van Aken, 2009).

Therefore, using transgenic plants or combined use of them with microorganisms in the rhizosphere could be mentioned as a promising technique to reduce abiotic stresses in plants which are used in phytoremediation. Future researches, especially in the field conditions, can distinguish the efficiency of this approach.

### **5. Conclusion**

10 Plants and Environment

and characterized so that they can be fused with reporter genes and be introduced into plants. For example the promoter of the barley gene HvhsplT which is expressed in the presence of some heavy metals had been fused to the reporter gene. This new gene combination was used to make a transformed tobacco plant which could be used as a

A combined use of transgenic plants and bacteria in the rhizosphere could improve phytoremediation of contaminated environments and may overcome the current limitations of phytoremediation such as low detoxification and absorption efficiency. The combination of plants for removing or degrading toxic pollutants and rhizospheric microorganisms for improving the availability of hydrophobic compounds can be effective in breaking down

gshI *Brassica juncea* Cd tolerance , Cd, Zn, Cu and Pb

AsPCS1 *Arabidopsis thaliana* As and Cd tolerance and accumulation

APS1 *Brassica juncea* Se tolerance and accumulation ,Cd

merP *Arabidopsis thaliana* Hg accumulation and tolerance ADC *Oryza sativa* Heavy metal tolerance

*Nicotiana tabacum* Cd tolerance , Cu accumulation

*L.tulipifera* Hg and Au resistance

*Brassica juncea* Cd accumulation, Cd and As tolerance

*and Zea mays* Heavy metal tolerance

*Nicotiana tabacum* Fe accumulation

OAS-TL *N. tabacum* Cd and Ni tolerance

SMT *Brassica juncea* Se accumulation SMT and APS1 *Brassica juncea* Se accumulation

TaPCS1 *Nicotiana. glauca* Pb and Cd tolerance

HisCUP1 *Nicotiana tabacum* Cd accumulation , Cd tolerance Ferritin *Oryza sativa* Fe accumulation

SOS1 *Oryza sativa* Heavy metal tolerance

SAMDC *Oryza sativa* Heavy metal tolerance Table 1. Genes involved in plant genetic engineering for phytoremediation of heavy metals (Hsieh et al., 2009; Karenlampi et al., 2000; Kotrba et al., 2009; Kolodyazhnaya et al., 2009).

MT-I and MT-II *Nicotiana tabacum* Cd tolerance ZAT *Arabidopsis thaliana* Zn tolerance

NtCBP4 *Arabidopsis thaliana* Ni tolerance, Pb and Ni accumulation

accumulation

As and Cd tolerance and accumulation

accumulation

Pb, Cd, Zn, Cu and Ni accumulation

bioindicator for monitoring heavy metal pollution (Mociardini, et al., 1998).

**Gene Target plant Effect**

*Brassica juncea*

*Arabidopsis thaliana* 

gshI and gshII *Arabidopsis thaliana and* 

CUP1 *Brassica oleracea*

FRE1 and FRE2 *Arabidopsis thaliana*

AtPCS1 *Nicotiana tabacum*

SOD *Nicotiana tabacum*

GSH1 and

merApe9 and merA18

Abiotic stresses, especially due to high level of organic and inorganic pollutants, are major limiting factors which could adversely affect phytoremediation. To reduce the effects of these stresses in plants which are used in phytoremediation, using bacteria- or fungi symbiosis as well as plant genetic engineering could be a promising way to enhance remediation efficiency. In practical point of view, it should be considered that combined enhanced-phytoremediation approaches are possible to use too. Finally, understanding the involved mechanisms in the mentioned enhancing methods would be a useful tool to extend use of phytoremediation based on these approaches. Since most of researches have been carried out in laboratory conditions, field trials are needed to perform in contaminated sites.

### **6. Acknowledgment**

The authors are very grateful to the researchers whose their results have been used in this review.

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**2** 

*India* 

**Morphophysiological Investigations in Some Dominant Alien Invasive Weeds** 

Allelopathy generally refers to any direct or indirect, harmful or beneficial effect of one plant on other plants, animals including microorganisms, through the production of chemical compounds that are released into the environment (Rice 1984). These donor plants affect the germination, growth and development of the recipient plant species (Einhellig 1987). The science of allelopathy has a very crucial role in maintaining the phytodiversity / biodiversity of a particular region. In fact, the phenomenon of biodiversity is the reflection of allelopathic interactions in that area. The losses in phytodiversity which are taking place at an alarming rate throughout the world is mainly ascribed to introduction of invasive / alien species which substitute the native ones. Invaded plant species and their success as

"The invasive plants are also known as alien, exotic or introduced ones, which are new to a specific area, become dominant, replacing / substituting the native plant species". These wide-spreading, non-indigenous species adversely affect the habitats they are invading in. The most important aspect of the alien plant is their rapid growth, establishment over new and large areas. Introduced species often find no natural enemies in their new habitat and therefore spread easily and quickly, especially in open disturbed areas. Invasive plants reproduce fast, either vegetatively or by seed. Their phenomenal growth allows them to

The general ecological impacts of invasive plants on natives and their surrounding

Competition with (and/or replacement of) native plants along with rare and

Loss of habitat and food sources of native insects, birds, wildlife and plants including

Acute competition for space, water, sunlight and nutrients due to its reduction

well as secret have always threatened the world's biodiversity.

overwhelm and displace existing vegetation and form dense Monothickets.

**1.1 Ecological impacts of invasive plants on the environment** 

environment are given in nutshell:-

 Disruption of native plant-animal associations Elimination of native plant communities Prevention of establishment of native plants

Change of the soil structure and chemistry

endangered once.

microorganisms.

**1. Introduction** 

*1Department of Botany, Abasaheb Garware College, Pune, M.S.* 

Nivedita Ghayal1 and Kondiram Dhumal2

*2Department of Botany, University of Pune, Pune, M.S.* 


## **Morphophysiological Investigations in Some Dominant Alien Invasive Weeds**

Nivedita Ghayal1 and Kondiram Dhumal2 *1Department of Botany, Abasaheb Garware College, Pune, M.S. 2Department of Botany, University of Pune, Pune, M.S.* 

*India* 

### **1. Introduction**

14 Plants and Environment

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Allelopathy generally refers to any direct or indirect, harmful or beneficial effect of one plant on other plants, animals including microorganisms, through the production of chemical compounds that are released into the environment (Rice 1984). These donor plants affect the germination, growth and development of the recipient plant species (Einhellig 1987). The science of allelopathy has a very crucial role in maintaining the phytodiversity / biodiversity of a particular region. In fact, the phenomenon of biodiversity is the reflection of allelopathic interactions in that area. The losses in phytodiversity which are taking place at an alarming rate throughout the world is mainly ascribed to introduction of invasive / alien species which substitute the native ones. Invaded plant species and their success as well as secret have always threatened the world's biodiversity.

"The invasive plants are also known as alien, exotic or introduced ones, which are new to a specific area, become dominant, replacing / substituting the native plant species". These wide-spreading, non-indigenous species adversely affect the habitats they are invading in. The most important aspect of the alien plant is their rapid growth, establishment over new and large areas. Introduced species often find no natural enemies in their new habitat and therefore spread easily and quickly, especially in open disturbed areas. Invasive plants reproduce fast, either vegetatively or by seed. Their phenomenal growth allows them to overwhelm and displace existing vegetation and form dense Monothickets.

### **1.1 Ecological impacts of invasive plants on the environment**

The general ecological impacts of invasive plants on natives and their surrounding environment are given in nutshell:-


Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 17

Weeds have many ill / negative characters, which cannot be neglected at all. Many of the weeds cause damages to agroecosystems and also disturb/ reduce natural phytodiversity. Weeds cause great harm to the crops in various ways as they cause 30 – 40% yield losses, increase the expenditure of various cultural practices, reduce the efficiency of agricultural implements. Perennial weeds reduce quality of fertile lands, cause obstacles for water flowing in canals. Weeds reduce crop yield and its quality as they compete with crops for resources like soil, water, nutrients and light. Weeds are alternative hosts for many pests and pathogens. Many weeds like *Prosopis, Calotropis* etc. reduce the germination capacity of crops' seeds due to the phytotoxins/ allelochemicals/ ecochemicals, many a times which are

Aquatic weeds like *Eichhornia* and different types of algae produce toxins, which are harmful to aquatic flora and fauna. Weeds harbour organisms like mosquitoes, which cause or transmit diseases. Some weeds are poisonous to humans and produce pollens, which cause allergies. These studies will be more helpful, if emphasis on interactions among the plants is highly focused by the researchers. Studies on allelopathic potential and the biochemical characterization of native and invasive weeds has become the top priority to get

the secondary metabolites, secreted by them in the soil.

rid of the ill effects of native and invasive weeds.

Diagram 1. Mechanism of action and physiological effects

Allelopathic interactions are primarily based on the synthesis and release of secondary metabolites by higher plants that initiate a wide array of biochemical reactions, which induce several biological changes, however, many of these are yet to be understood. In

**2.1 Allelopathic interactions between plants** 

 Morphophysiological interactions with native flora and fauna through release of allelochemicals / ecochemicals

### **1.2 Scope of allelopathy**

At present it is well established that allelopathic phenomenon exist in different ecosystems including forest ecosystem and it can be exploited for increasing the productivity of crops as well as forest plant species in sustainable manner. According to Reigosa et al. (1999), allelopathy can affect distribution pattern of plants and biodiversity. They further explained that in a climax forest, germination and growth of understorey species must cope with allelochemicals released by the dominant trees. Those trees could release different chemicals, producing differences in the species composition. Similarly, Carballeira and Reigosa (1999) also indicated that monocultures (pure stands) allow the accumulation of particular allelochemicals affecting species composition. The occurrence of some weeds growing better than others within a monoculture could be a result of accumulation of allelochemicals. The allelopathic effects can affect small-scale vegetation patterns, by strengthening the associations between plants or not allowing them to grow in their vicinity. The research in allelopathy has increased greatly from 1960 onwards (Putnam 1985).

Inderjit *et al.* (2005a, b) has discussed in detail the challenges, achievements and opportunities in allelopathy research. They further highlighted that the novel research findings of allelopathy relevant to enzymes and genes involved in production of putative allelochemicals, allelochemical persistence in the rhizosphere, the molecular target sites of allelochemicals in sensitive plant species and the influence of allelochemicals upon other organisms will lead to enhanced utilization of natural products for pest management or as pharmaceuticals and nutraceuticals. The research and development in allelopathy is of extreme urgency for improvement of agriculture, forestry and global environment (Reigosa and Pedrol, 2002), because it deals majorly with invasive and native plant species.

### **2. Allelobiogenesis - concept and mechanism**

Allelobiogenesis is nothing but the stress created by allelopathic effects of donor plants on recipient plants. In other words it is biotic stress and at the same time it is abiotic stress also. Allelobiogenesis is a typical stress combination of biotic and abiotic factors. Plant – plant, plant – animal and plant – micro-organism interactions can be considered as biotic stress.

The influence / stress of one plant on the other plant is mainly through the phytochemicals / ecochemicals / allelochemicals released by these donor plants. Hence the stress is obviously of abiotic nature. These allelochemicals are mainly secondary metabolites like alkaloids, glycosides, tannins, flavonoids, phenols etc. and the stress created by such allelochemicals is abiotic stress. The stress created by such allelopathic interaction or allelochemicals is allelobiogenesis. A weed exhausting nutrients from the soil voraciously and producing nutrient stress on associated crop as it shows dominance on associated crop by its faster growth and encroachment over crop species is biotic allelobiogenesis, e.g. *Parthenium* the invasive weed growing in association with *Sorghum.* 

The exotic weed *Parthenium* is releasing large no. of allelochemicals through root exudation, leaching and volatilization in the surroundings and these allelochemicals cause very adverse effects on seed germination and growth and all the metabolic processes such as photosynthesis, respiration, absorption of water and minerals. This stress can be well explained as "abiotic allelobiogenesis".

Morphophysiological interactions with native flora and fauna through release of

At present it is well established that allelopathic phenomenon exist in different ecosystems including forest ecosystem and it can be exploited for increasing the productivity of crops as well as forest plant species in sustainable manner. According to Reigosa et al. (1999), allelopathy can affect distribution pattern of plants and biodiversity. They further explained that in a climax forest, germination and growth of understorey species must cope with allelochemicals released by the dominant trees. Those trees could release different chemicals, producing differences in the species composition. Similarly, Carballeira and Reigosa (1999) also indicated that monocultures (pure stands) allow the accumulation of particular allelochemicals affecting species composition. The occurrence of some weeds growing better than others within a monoculture could be a result of accumulation of allelochemicals. The allelopathic effects can affect small-scale vegetation patterns, by strengthening the associations between plants or not allowing them to grow in their vicinity.

The research in allelopathy has increased greatly from 1960 onwards (Putnam 1985).

and Pedrol, 2002), because it deals majorly with invasive and native plant species.

**2. Allelobiogenesis - concept and mechanism** 

*Parthenium* the invasive weed growing in association with *Sorghum.* 

explained as "abiotic allelobiogenesis".

Inderjit *et al.* (2005a, b) has discussed in detail the challenges, achievements and opportunities in allelopathy research. They further highlighted that the novel research findings of allelopathy relevant to enzymes and genes involved in production of putative allelochemicals, allelochemical persistence in the rhizosphere, the molecular target sites of allelochemicals in sensitive plant species and the influence of allelochemicals upon other organisms will lead to enhanced utilization of natural products for pest management or as pharmaceuticals and nutraceuticals. The research and development in allelopathy is of extreme urgency for improvement of agriculture, forestry and global environment (Reigosa

Allelobiogenesis is nothing but the stress created by allelopathic effects of donor plants on recipient plants. In other words it is biotic stress and at the same time it is abiotic stress also. Allelobiogenesis is a typical stress combination of biotic and abiotic factors. Plant – plant, plant – animal and plant – micro-organism interactions can be considered as biotic stress. The influence / stress of one plant on the other plant is mainly through the phytochemicals / ecochemicals / allelochemicals released by these donor plants. Hence the stress is obviously of abiotic nature. These allelochemicals are mainly secondary metabolites like alkaloids, glycosides, tannins, flavonoids, phenols etc. and the stress created by such allelochemicals is abiotic stress. The stress created by such allelopathic interaction or allelochemicals is allelobiogenesis. A weed exhausting nutrients from the soil voraciously and producing nutrient stress on associated crop as it shows dominance on associated crop by its faster growth and encroachment over crop species is biotic allelobiogenesis, e.g.

The exotic weed *Parthenium* is releasing large no. of allelochemicals through root exudation, leaching and volatilization in the surroundings and these allelochemicals cause very adverse effects on seed germination and growth and all the metabolic processes such as photosynthesis, respiration, absorption of water and minerals. This stress can be well

allelochemicals / ecochemicals

**1.2 Scope of allelopathy** 

Weeds have many ill / negative characters, which cannot be neglected at all. Many of the weeds cause damages to agroecosystems and also disturb/ reduce natural phytodiversity. Weeds cause great harm to the crops in various ways as they cause 30 – 40% yield losses, increase the expenditure of various cultural practices, reduce the efficiency of agricultural implements. Perennial weeds reduce quality of fertile lands, cause obstacles for water flowing in canals. Weeds reduce crop yield and its quality as they compete with crops for resources like soil, water, nutrients and light. Weeds are alternative hosts for many pests and pathogens. Many weeds like *Prosopis, Calotropis* etc. reduce the germination capacity of crops' seeds due to the phytotoxins/ allelochemicals/ ecochemicals, many a times which are the secondary metabolites, secreted by them in the soil.

Aquatic weeds like *Eichhornia* and different types of algae produce toxins, which are harmful to aquatic flora and fauna. Weeds harbour organisms like mosquitoes, which cause or transmit diseases. Some weeds are poisonous to humans and produce pollens, which cause allergies. These studies will be more helpful, if emphasis on interactions among the plants is highly focused by the researchers. Studies on allelopathic potential and the biochemical characterization of native and invasive weeds has become the top priority to get rid of the ill effects of native and invasive weeds.

Diagram 1. Mechanism of action and physiological effects

### **2.1 Allelopathic interactions between plants**

Allelopathic interactions are primarily based on the synthesis and release of secondary metabolites by higher plants that initiate a wide array of biochemical reactions, which induce several biological changes, however, many of these are yet to be understood. In

Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 19

Biological species invasions alter ecological systems in a multitude of ways. Worldwide an estimated 80% of endangered species could suffer losses due to competition with or

To have the information about the previous work done on allelopathy in general, its role in different fields of agriculture and botany, different types of interactions such as weed – weed, weed – crop, the impact of leachates, extracts and residues on recipient plants, allelochemicals existing in different donor plants, their chemical structures, mode of release of these ecochemicals in the environment, their accumulation, mechanism of action, their effect on seed germination, seedling growth, mineral nutrition, microbial activity in the soil

Studies on allelopathy were made thousands of years before the term was coined by Molisch (1937). The term allelopathy is derived from two Latin words *Allelon* means each other and *pathos* means to suffer. He, for the first time studied the effect of numerous plant species and their plant parts viz.- roots, shoots, leaves, flowers, fruits, leachates, extracts and residues on seed germination, seedling growth and maturity of crops. Later on many scientists at different corners of the world, contributed to this field by carrying out the research on various aspects of allelopathy. At present the research on allelopathy is being carried out in more than 85 countries. In India, the research in this field took a great speed after 1950. Vilai-Santisopasri (2003) studied the allelopathic effects of *Eupatorium adenophorum* Spreng. on growth of some crops and weeds. Hierro and Callaway (2003) had investigated in detail the invasion of exotic plants and their role in allelopathy. Many workers like Rice (1979), Gill and Sandhu (1996), Pawar and Chavan (1999), Chou (1999), Wang et al. (2001), Cheema et al. (2002) had great contribution in allelopathy through their basic research. Recently, many researchers like Narwal et al. (2003a, b), Podolska et al. (2003), Navaz et al. (2003), Batish et al. (2002), Singh and Singh (2003) and Azania et al*.* (2003) have introduced

According to Fujii et al. (2002) allelopathy now refers to any process involving secondary metabolites produced by plants, microorganisms, viruses and fungi, that influence the growth and development of agricultural and biological systems. The allelopathy workers like Bhatt and Chauhan (2000), Singh and NarsingRao (2003) and Leather and Einhellig (2005) also claimed that secondary metabolites produced by donor plants, when released into environment, play a key role in ecology and physiology of recipient plants. They further advocated that the released allelochemicals as well as the phytochemicals present in the leachates **/** extracts have stimulatory or inhibitory influence on seed germination,

The allelopathic impact of invasive weeds on seed germination, seedling growth, growth parameters like plant height, number of leaves per plant, leaf area, yield contributing parameters like number of flowers and fruits per plant, weight of fruit and grains etc in different crops had been studied in detail by Rice (1979), Patil and Hegde (1988), Devi et al*.* (1997), Kulvinder et al*.* (1999), Bhalerao et al*.* (2000a, b), Wang et al. (2001), Kong and Hu (2001), Lin et al. (2002), Bhalerao (2003), Jadhav (2006), Hase (2008) and Vaidya (2009). Presently the allelopathy research work is mainly focused on identification of

According to many researchers allelopathy now refers to any process involving secondary metabolites produced by plants, microorganisms, viruses and fungi that influence the

predation by invasive species.

**3. Compilation of updated work** 

etc. a review of literature is given in nutshell.

multidisciplinary approach in allelopathy.

seedling growth and yield of recipient plants.

allelochemicals, their mode of action and ecological significance.

nature, many plant species grow together and interact with each other by inhibiting or stimulating the growth and development through allelopathic interactions. In any ecosystem, the dominant plants growing within it are exhibited in the form of pure stands or monothickets. Such ecosystems always show the zones of inhibition around them (Nilsen 2002, Chase and Leibold 2003). The ecosystems infested by dominant weeds show drastic alterations in their structure and function.

All the weed species, which are the part of dynamic ecosystems, originate in natural environment and become hurdle to the crops (Aldrich 1984). These weeds have some diagnostic features, such as short seed dormancy period, high rate of seed germination, rapid seedling growth, high reproductive ability, life cycle of a short span, very high environmental plasticity, self-compatibility, effective and efficient methods of dispersal of propagules, production of different types of novel ecochemicals or allelochemicals and tolerance to biotic and abiotic stresses (Baker 1965), which enable them to grow and survive in varied habitats and inhospitable ecological conditions.

As a result of this these weeds are becoming dominant throughout the world (Colautti and MacIsaac 2004, Lee and Klasing 2004, Jeschke and Strayer, 2005). As stated by Li et al. (2009), the invasion of exotic weeds is mainly due to their easier establishment and faster growth under diverse environmental conditions. Lonsdale (1999) claimed that the propagules' pressure, adaptive characters and susceptible environment favour the invasibility to which Carlton (2001) called biological invasion.

### **2.2 Plant invasions and encroachments**

The whole biosphere is facing the problem of invasion of different weed species, hence studies on plant invasions and allelopathy will help in understanding the mechanism of invasions, and consequences of them on global biodiversity and ecosystem functioning. These invasions pose many ecological, economic and social problems. A team approach to solve these complicated problems is necessary.

According to MacDougall and Turkington (2005), the alien species highly out compete the native species or escape from adverse environmental conditions and dominate the community. According to vacant niche hypothesis (Elton 1958) the empty places such as barren lands, roadsides, open grounds etc. are generally invaded by such weeds. There are different hypotheses explaining the invasion mechanisms (Inderjit et al. 2005a, b). The diversity of these weeds is governed by population, ecosystem dynamics, disturbances, nutrient supply and climatic factors. The biotic restrictions on them, force to skip from their previous habitat and start surviving in new habitats, helping in the process of invasion. The enemy release hypothesis advocated by Mack et al. (2000) also supports the above view. If the invader is resistant enough and tolerant to herbivory, then its competitive ability increases and it becomes very aggressive due to production of some defensive chemicals (Carpenter and Cappuccino 2005).

The disturbances by some plant species, grazing pressure, fluctuation in resource availability (Davis et al. 2000), soil moisture, available light (Meekins and Mc Carthy 2001), phenotypic plasticity and hybridization (Daehler 2003) results in to successful invasion. The novel weapon hypothesis (Callaway and Ridenour 2004), biotic resistance hypothesis (Maron and Vilà 2001), and the genetic shift hypothesis (DeWalt et al. 2004) also explain the mechanism of invasion. To understand the distribution of invasive weeds and their associates in a natural community, the eco-distribution mapping is of paramount importance.

Biological species invasions alter ecological systems in a multitude of ways. Worldwide an estimated 80% of endangered species could suffer losses due to competition with or predation by invasive species.

### **3. Compilation of updated work**

18 Plants and Environment

nature, many plant species grow together and interact with each other by inhibiting or stimulating the growth and development through allelopathic interactions. In any ecosystem, the dominant plants growing within it are exhibited in the form of pure stands or monothickets. Such ecosystems always show the zones of inhibition around them (Nilsen 2002, Chase and Leibold 2003). The ecosystems infested by dominant weeds show drastic

All the weed species, which are the part of dynamic ecosystems, originate in natural environment and become hurdle to the crops (Aldrich 1984). These weeds have some diagnostic features, such as short seed dormancy period, high rate of seed germination, rapid seedling growth, high reproductive ability, life cycle of a short span, very high environmental plasticity, self-compatibility, effective and efficient methods of dispersal of propagules, production of different types of novel ecochemicals or allelochemicals and tolerance to biotic and abiotic stresses (Baker 1965), which enable them to grow and survive

As a result of this these weeds are becoming dominant throughout the world (Colautti and MacIsaac 2004, Lee and Klasing 2004, Jeschke and Strayer, 2005). As stated by Li et al. (2009), the invasion of exotic weeds is mainly due to their easier establishment and faster growth under diverse environmental conditions. Lonsdale (1999) claimed that the propagules' pressure, adaptive characters and susceptible environment favour the invasibility to which

The whole biosphere is facing the problem of invasion of different weed species, hence studies on plant invasions and allelopathy will help in understanding the mechanism of invasions, and consequences of them on global biodiversity and ecosystem functioning. These invasions pose many ecological, economic and social problems. A team approach to

According to MacDougall and Turkington (2005), the alien species highly out compete the native species or escape from adverse environmental conditions and dominate the community. According to vacant niche hypothesis (Elton 1958) the empty places such as barren lands, roadsides, open grounds etc. are generally invaded by such weeds. There are different hypotheses explaining the invasion mechanisms (Inderjit et al. 2005a, b). The diversity of these weeds is governed by population, ecosystem dynamics, disturbances, nutrient supply and climatic factors. The biotic restrictions on them, force to skip from their previous habitat and start surviving in new habitats, helping in the process of invasion. The enemy release hypothesis advocated by Mack et al. (2000) also supports the above view. If the invader is resistant enough and tolerant to herbivory, then its competitive ability increases and it becomes very aggressive due to production of some defensive chemicals

The disturbances by some plant species, grazing pressure, fluctuation in resource availability (Davis et al. 2000), soil moisture, available light (Meekins and Mc Carthy 2001), phenotypic plasticity and hybridization (Daehler 2003) results in to successful invasion. The novel weapon hypothesis (Callaway and Ridenour 2004), biotic resistance hypothesis (Maron and Vilà 2001), and the genetic shift hypothesis (DeWalt et al. 2004) also explain the mechanism of invasion. To understand the distribution of invasive weeds and their associates in a natural community, the eco-distribution mapping is of paramount

alterations in their structure and function.

Carlton (2001) called biological invasion.

**2.2 Plant invasions and encroachments** 

solve these complicated problems is necessary.

(Carpenter and Cappuccino 2005).

importance.

in varied habitats and inhospitable ecological conditions.

To have the information about the previous work done on allelopathy in general, its role in different fields of agriculture and botany, different types of interactions such as weed – weed, weed – crop, the impact of leachates, extracts and residues on recipient plants, allelochemicals existing in different donor plants, their chemical structures, mode of release of these ecochemicals in the environment, their accumulation, mechanism of action, their effect on seed germination, seedling growth, mineral nutrition, microbial activity in the soil etc. a review of literature is given in nutshell.

Studies on allelopathy were made thousands of years before the term was coined by Molisch (1937). The term allelopathy is derived from two Latin words *Allelon* means each other and *pathos* means to suffer. He, for the first time studied the effect of numerous plant species and their plant parts viz.- roots, shoots, leaves, flowers, fruits, leachates, extracts and residues on seed germination, seedling growth and maturity of crops. Later on many scientists at different corners of the world, contributed to this field by carrying out the research on various aspects of allelopathy. At present the research on allelopathy is being carried out in more than 85 countries. In India, the research in this field took a great speed after 1950.

Vilai-Santisopasri (2003) studied the allelopathic effects of *Eupatorium adenophorum* Spreng. on growth of some crops and weeds. Hierro and Callaway (2003) had investigated in detail the invasion of exotic plants and their role in allelopathy. Many workers like Rice (1979), Gill and Sandhu (1996), Pawar and Chavan (1999), Chou (1999), Wang et al. (2001), Cheema et al. (2002) had great contribution in allelopathy through their basic research. Recently, many researchers like Narwal et al. (2003a, b), Podolska et al. (2003), Navaz et al. (2003), Batish et al. (2002), Singh and Singh (2003) and Azania et al*.* (2003) have introduced multidisciplinary approach in allelopathy.

According to Fujii et al. (2002) allelopathy now refers to any process involving secondary metabolites produced by plants, microorganisms, viruses and fungi, that influence the growth and development of agricultural and biological systems. The allelopathy workers like Bhatt and Chauhan (2000), Singh and NarsingRao (2003) and Leather and Einhellig (2005) also claimed that secondary metabolites produced by donor plants, when released into environment, play a key role in ecology and physiology of recipient plants. They further advocated that the released allelochemicals as well as the phytochemicals present in the leachates **/** extracts have stimulatory or inhibitory influence on seed germination, seedling growth and yield of recipient plants.

The allelopathic impact of invasive weeds on seed germination, seedling growth, growth parameters like plant height, number of leaves per plant, leaf area, yield contributing parameters like number of flowers and fruits per plant, weight of fruit and grains etc in different crops had been studied in detail by Rice (1979), Patil and Hegde (1988), Devi et al*.* (1997), Kulvinder et al*.* (1999), Bhalerao et al*.* (2000a, b), Wang et al. (2001), Kong and Hu (2001), Lin et al. (2002), Bhalerao (2003), Jadhav (2006), Hase (2008) and Vaidya (2009). Presently the allelopathy research work is mainly focused on identification of allelochemicals, their mode of action and ecological significance.

According to many researchers allelopathy now refers to any process involving secondary metabolites produced by plants, microorganisms, viruses and fungi that influence the

Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 21

growth and development of agricultural and biological systems. The allelopathy workers also claimed that secondary metabolites produced by donor plants, when released into environment, play a key role in ecology and physiology of recipient plants. They further advocated that the released allelochemicals as well as the phytochemicals present in the leachates **/** extracts/ residues have stimulatory or inhibitory influence on seed germination,

Today this subject has come into lime-light because of its multidisciplinary nature, which covers agriculture, biological sciences, biochemistry, physiology, biotechnology and even

The light has been thrown on the success of invasive alien weeds outside their native boundary and probable causes of this. The biological processes and specific characteristics of invasive weeds are important factors in their introduction, spread, and establishment that threatens the ecosystems, habitats, or species with economic/ environmental harm. Therefore the detailed investigations on the ecological, physiological and molecular aspects of invasive weeds' allelopathy should be conducted in order to understand community

Isolation, identification and characterization of allelochemicals present in roots, stems, leaves, flowers, fruits, seeds, bark, residues, litter, dried leaves (trash) and their leachates, extracts and residues have a pivotal role in allelopathy research, without which any predictions, possibilities, hypothesis and explanations are not possible. Asteraceae plants with their leachates, extracts and residues of different plant parts are well known for their allelopathic activity because of their allelochemicals like phenolic acids and terpenoids

Many researchers like Ghayal et al*.* (2007a, b, c) and Li et al. (2009) have given prime importance for identification of allelochemicals, ecochemicals, novel bioactive compounds which are the secondary metabolites existing in their leachates, extracts and residues. They have characterized diverse groups of allelochemicals like terpenoids, flavonoids, phenolic

The big campus of University of Pune established in 1949, at Ganeshkhind occupies an area of 164.8 hectares, which is situated about seven km north-west of Pune city proper and lies between 18034' North latitude and 73053' East longitude at an elevation of about 1880 m. At

Ganeshkhind stands on pediment surface of amygdaloidal basalt. These rocks are traversed by many veins and veinlets of silica and chalcedony. The poor soil of study area is reddish brown on higher grounds and deeper dark brown (black cotton soil) on flat areas. The soils are alkaline and are of pedocal type (Varadpande 1972). The average rainfall, climate and other environmental conditions of the campus are more or less similar to that of Pune city. The railfall is restricted to couple of months in monsoon and the maximum annual rainfall is 31.78cm. The temperature during hot season goes up to 40 – 420C but normally it is cool as

seedling growth and yield of recipient plants.

genetic engineering.

(Chon et al. 2003).

**4. About the study area** 

compared to Pune city.

**3.1 Invasion success of weeds** 

structure and declining phytodiversity.

**3.2 Allelochemicals in invasive and native weed species** 

compounds and essential oils existing in the invasive and native weeds.

present 1/4 th area is occupied by roads, buildings and gardens.

Diagram 2. Different pathways of synthesis of allelochemicals

growth and development of agricultural and biological systems. The allelopathy workers also claimed that secondary metabolites produced by donor plants, when released into environment, play a key role in ecology and physiology of recipient plants. They further advocated that the released allelochemicals as well as the phytochemicals present in the leachates **/** extracts/ residues have stimulatory or inhibitory influence on seed germination, seedling growth and yield of recipient plants.

Today this subject has come into lime-light because of its multidisciplinary nature, which covers agriculture, biological sciences, biochemistry, physiology, biotechnology and even genetic engineering.

### **3.1 Invasion success of weeds**

20 Plants and Environment

Diagram 2. Different pathways of synthesis of allelochemicals

The light has been thrown on the success of invasive alien weeds outside their native boundary and probable causes of this. The biological processes and specific characteristics of invasive weeds are important factors in their introduction, spread, and establishment that threatens the ecosystems, habitats, or species with economic/ environmental harm. Therefore the detailed investigations on the ecological, physiological and molecular aspects of invasive weeds' allelopathy should be conducted in order to understand community structure and declining phytodiversity.

### **3.2 Allelochemicals in invasive and native weed species**

Isolation, identification and characterization of allelochemicals present in roots, stems, leaves, flowers, fruits, seeds, bark, residues, litter, dried leaves (trash) and their leachates, extracts and residues have a pivotal role in allelopathy research, without which any predictions, possibilities, hypothesis and explanations are not possible. Asteraceae plants with their leachates, extracts and residues of different plant parts are well known for their allelopathic activity because of their allelochemicals like phenolic acids and terpenoids (Chon et al. 2003).

Many researchers like Ghayal et al*.* (2007a, b, c) and Li et al. (2009) have given prime importance for identification of allelochemicals, ecochemicals, novel bioactive compounds which are the secondary metabolites existing in their leachates, extracts and residues. They have characterized diverse groups of allelochemicals like terpenoids, flavonoids, phenolic compounds and essential oils existing in the invasive and native weeds.

### **4. About the study area**

The big campus of University of Pune established in 1949, at Ganeshkhind occupies an area of 164.8 hectares, which is situated about seven km north-west of Pune city proper and lies between 18034' North latitude and 73053' East longitude at an elevation of about 1880 m. At present 1/4 th area is occupied by roads, buildings and gardens.

Ganeshkhind stands on pediment surface of amygdaloidal basalt. These rocks are traversed by many veins and veinlets of silica and chalcedony. The poor soil of study area is reddish brown on higher grounds and deeper dark brown (black cotton soil) on flat areas. The soils are alkaline and are of pedocal type (Varadpande 1972). The average rainfall, climate and other environmental conditions of the campus are more or less similar to that of Pune city. The railfall is restricted to couple of months in monsoon and the maximum annual rainfall is 31.78cm. The temperature during hot season goes up to 40 – 420C but normally it is cool as compared to Pune city.

Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 23

*Bidens pilosa Cassia uniflora* Very common *Cassia uniflora Achyranthes aspera* Very common *Triumfetta rhomboidea Cassia obtusifolia* Common *Alternanthera tenella Boerhaavia erecta* Common *Cassia absus Achyranthes aspera* Common *Cassia uniflora Synedrella nodiflora* Occasional *Cassia uniflora Parthenium hysterophorus* Rare

species

species

species

*Synedrella nodiflora Cassia uniflora* Very common *Synedrella nodiflora Achyranthes aspera* Common *Cassia uniflora Tithonia tagetiflora* Common *Cassia uniflora Achyranthes aspera* Common *Rauwolfia tetraphylla Euphorbia geniculata* Common

*Cassia uniflora Achyranthes aspera* Common *Cassia uniflora Alternanthera tenella* Common *Lantana camara Cassia uniflora* Occasional *Acalypha ciliata Cassia uniflora* Common

Mishra et al. (1997), Chapin et al. (2000), Kumar et al. (2004), Jadhav (2006), Saswade (2007) and Thakur and Khare (2009) have also carried out the phytosociological studies on various invasive and native weeds. The phytosociological studies on *Potentilla recta* and other species*,* the invasive, noxious weed from Eurasia were carried out by Werner and Soule (1976). Zouhar (2003), Endress and Parks (2004) had also conducted phytosociological studies on these invasive weeds from U.S. The dominance of *Cassia uniflora* and *Synedrella* in the study area may be attributed to their aggressive nature, allelolpathic potential, adaptations in morphological and reproductive features along with specific type of physiological, biochemical and enzymological mechanisms allowing their faster growth and

Along with the two dominant weed species like Cassia uniflora and Synedrella nodiflora at all the four sites, the major co-occuring species recorded were Acalypha ciliata, Boerhaavia erecta, Cassia obtusifolia, Lagasca mollis, Peristrophe bicalyculata, Parthenium hysterophorus and Triumfetta rhomboidea. The fact worth to mention was establishment of

Nature of association

Nature of association

> Nature of association

No. Dominant species Associated

Table 1. (b) Weed-weed interactions at site II in Pune university campus

Table 1. (c) Weed-weed interactions at site III in Pune university campus

No. Dominant species Associated

Table 1. (d) Weed-weed interactions at site IV in Pune university campus

tolerance to biotic and abiotic stress conditions.

**4.2 Invasion components** 

No. Dominant species Associated

### **4.1 Weed floristic of study area**

The phytosociological studies were conducted at all the four selected sites in Pune University campus. The different plant species were recorded with respect to their distribution, frequency, density and abundance. All four study sites (I to IV) showed the vegetation composition of various types of herbs, shrubs, trees, climbers and twinners.

The results on phytosociological studies at site I reported in Table 1 (a) indicated that this site was having highest weed diversity. The most abundant weeds at this site with maximum frequencies were *Cassia uniflora* (93.33%), *Achyranthes* (90.00%), *Synedrella nodiflora*  (90.00%) and *Oplismenus* (83.33%). These were followed by *Acalypha*, *Bidens, Boerhaavia* and *Euphorbia geniculata,* all these were at par having frequency (80.00%). *Alternanthera tenella* was also showing better population with frequency 76.67%. Remaining native plants like *Cynotis* and *Calotropis* and invasive plants have shown less frequency, density and abundance.

The results of site II recorded Table 1(b) revealed that at this site, 43 different weed genera were recorded including about 19 genera of invasive weeds. The most abundant weeds with maximum frequencies were *Synedrella nodiflora* (96.67%), *Cassia uniflora* (93.33%), and *Cassia absus* (86.67%). These were succeeded by *Acalypha*, *Bidens* and *Euphorbia geniculata* which were at par having frequency (83.33%). Remaining plants showed discrete occurrence. At sites I and II *Cassia uniflora* was showing very thick population density, as these sites were exposed to sunlight for longer period.

The results on phytosociological studies of site III reported in Table 1 (c) illustrated that at this site, only 24 genera were reported including about 12 genera of invasive weeds. The plants recorded at this site in order of highest frequency were *Synedrella nodiflora* (93.33%), *Cassia uniflora* (80.00%) and *Rauwolfia* (73.33%)*.* As opposite to sites I and II, at site III, there was complete dominance of *Synedrella nodiflora* only, which was virtually forming monothickets / pure stands. *Synedrella nodiflora* being shade lover was showing luxuriant growth and very high dominance at this site. The shady conditions along with high soil moisture favoured the luxuriant growth of *Synedrella nodiflora* and *Rauwolfia.* 

The results listed in Table 1 (d) regarding phytosociological studies carried out at site IV revealed that, there were about 32 genera of weed species including 14 genera of invasive weeds. The weeds with highest frequency were *Cassia uniflora* (96.67%), *Synedrella nodiflora* (93.33%) and *Euphorbia geniculata* (76.67%)*,* which were followed by *Acalypha, Achyranthes* and *Alternanthera*. At this site there was highest human interference as compared to the remaining three sites. The fast growing invasive weed *Lantana camara* showed successful invasion only at this site with frequency 63.33%.


Table 1. (a) Weed-weed interactions at site I in Pune university campus

The phytosociological studies were conducted at all the four selected sites in Pune University campus. The different plant species were recorded with respect to their distribution, frequency, density and abundance. All four study sites (I to IV) showed the vegetation composition of various types of herbs, shrubs, trees, climbers and twinners. The results on phytosociological studies at site I reported in Table 1 (a) indicated that this site was having highest weed diversity. The most abundant weeds at this site with maximum frequencies were *Cassia uniflora* (93.33%), *Achyranthes* (90.00%), *Synedrella nodiflora*  (90.00%) and *Oplismenus* (83.33%). These were followed by *Acalypha*, *Bidens, Boerhaavia* and *Euphorbia geniculata,* all these were at par having frequency (80.00%). *Alternanthera tenella* was also showing better population with frequency 76.67%. Remaining native plants like *Cynotis* and *Calotropis* and invasive plants have shown less frequency, density and

The results of site II recorded Table 1(b) revealed that at this site, 43 different weed genera were recorded including about 19 genera of invasive weeds. The most abundant weeds with maximum frequencies were *Synedrella nodiflora* (96.67%), *Cassia uniflora* (93.33%), and *Cassia absus* (86.67%). These were succeeded by *Acalypha*, *Bidens* and *Euphorbia geniculata* which were at par having frequency (83.33%). Remaining plants showed discrete occurrence. At sites I and II *Cassia uniflora* was showing very thick population density, as these sites were

The results on phytosociological studies of site III reported in Table 1 (c) illustrated that at this site, only 24 genera were reported including about 12 genera of invasive weeds. The plants recorded at this site in order of highest frequency were *Synedrella nodiflora* (93.33%), *Cassia uniflora* (80.00%) and *Rauwolfia* (73.33%)*.* As opposite to sites I and II, at site III, there was complete dominance of *Synedrella nodiflora* only, which was virtually forming monothickets / pure stands. *Synedrella nodiflora* being shade lover was showing luxuriant growth and very high dominance at this site. The shady conditions along with high soil

The results listed in Table 1 (d) regarding phytosociological studies carried out at site IV revealed that, there were about 32 genera of weed species including 14 genera of invasive weeds. The weeds with highest frequency were *Cassia uniflora* (96.67%), *Synedrella nodiflora* (93.33%) and *Euphorbia geniculata* (76.67%)*,* which were followed by *Acalypha, Achyranthes* and *Alternanthera*. At this site there was highest human interference as compared to the remaining three sites. The fast growing invasive weed *Lantana camara* showed successful

> *Cassia uniflora Achyranthes aspera* Very common *Cassia uniflora Blainvillea acmella* Occasional *Acalypha ciliata Cassia uniflora* Rare *Alternanthera tenella Cassia uniflora* Common *Synedrella nodiflora Cassia uniflora* Common *Oplismenus compositus ---* Very common *Euphorbia geniculata Cassia uniflora* Common

Associated species

Nature of association

moisture favoured the luxuriant growth of *Synedrella nodiflora* and *Rauwolfia.* 

**4.1 Weed floristic of study area** 

exposed to sunlight for longer period.

invasion only at this site with frequency 63.33%.

species

Table 1. (a) Weed-weed interactions at site I in Pune university campus

No. Dominant

abundance.


Table 1. (b) Weed-weed interactions at site II in Pune university campus


Table 1. (c) Weed-weed interactions at site III in Pune university campus


Table 1. (d) Weed-weed interactions at site IV in Pune university campus

Mishra et al. (1997), Chapin et al. (2000), Kumar et al. (2004), Jadhav (2006), Saswade (2007) and Thakur and Khare (2009) have also carried out the phytosociological studies on various invasive and native weeds. The phytosociological studies on *Potentilla recta* and other species*,* the invasive, noxious weed from Eurasia were carried out by Werner and Soule (1976). Zouhar (2003), Endress and Parks (2004) had also conducted phytosociological studies on these invasive weeds from U.S. The dominance of *Cassia uniflora* and *Synedrella* in the study area may be attributed to their aggressive nature, allelolpathic potential, adaptations in morphological and reproductive features along with specific type of physiological, biochemical and enzymological mechanisms allowing their faster growth and tolerance to biotic and abiotic stress conditions.

### **4.2 Invasion components**

Along with the two dominant weed species like Cassia uniflora and Synedrella nodiflora at all the four sites, the major co-occuring species recorded were Acalypha ciliata, Boerhaavia erecta, Cassia obtusifolia, Lagasca mollis, Peristrophe bicalyculata, Parthenium hysterophorus and Triumfetta rhomboidea. The fact worth to mention was establishment of

Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 25

Allelochemicals always affect many aspects of plants' ecology e.g. distribution, growth, succession, structure of communities, dominance, diversity and productivity (Takeuchi et al. 2001). The population of *Cassia uniflora* had always shown its shifting nature, i.e. area with monothickets during first year will show very less population next year on the same spot. It may be due to the resource exhaust, autotoxicity and heavy accumulation of allelochemicals making the area inhospitable for its own growth. However, it requires further confirmation

An attempt was made to study the morphological and reproductive features of invasive and native weeds from Pune university campus (Table 2). The maximum plant height was recorded for *Cassia uniflora,* (104.66cm), which was followed by *Triumfetta* (103.66cm) and

The root length indicates easy and proper establishment of the plants. The maximum root

The highest number of branches per plant was observed in *Achyranthes* and followed by *Triumfetta,* while the weed species like *Acalypha* and *Oplismenus* were without any branch.

*Cassia obtusifolia* had maximum third leaf area and it was followed by *Rauwolfia* and *Triumfetta.* The third leaf area in remaining weeds was very less as compared to them. The results on fresh biomass per plant indicated that *Cassia obtusifolia* was having highest biomass and *Triumfetta* was next to it. The weight of fresh biomass in the remaining weeds

The dry biomass per plant showed wide variations ranging from 0.89g to 17.43g. The *Cassia*  species e.g. *C. obtusifolia* and *C. uniflora* were having highest dry biomass 17.43g and 12.4g

The highest fresh biomass per m2 area was recorded in *Triumfetta, Cassia uniflora* and *Blainvillea.* The remaining weeds recorded comparatively less biomass per m2 area in the

Many research workers like Sen (1977), Weaver and McWilliams (1980), Wilson (1988) had given due importance to morphological studies of native and invasive weeds. Ehrenfeld (2003) proposed that the invasive plants share many physical characteristics and tend to alter habitats. They have very high productivity and above ground biomass. They grow earlier in the season and show faster growth rate than native species. All such features might be applicable to *Cassia* and *Synedrella,* because of which they are dominant in the

Unless we know the morphological features of weeds, the attempts for their effective management are difficult. With this view, Sutherland (2004) has described all the morphological details of different terrestrial and aquatic weeds of India. Similarly Monaco et al. (2002) had also investigated the various morphological characteristics of different weeds from USA. The life cycle of a plant can be understood well by its morphological structures, developmental processes and whole plant activities that occur during each phase of its life cycle. He further stated that selected phenotypes dominate their neighbours, because the timing of their life history optimizes their relative fitness and minimizes mortality. Same explanation may be true for the dominance of different invasive weeds

including *Cassia* and *Synedrella* over the natives of Pune University campus.

length was recorded for *Alternanthera* followed by *Rauwolfia* and *Achyranthes*.

The number of branches in remaining weeds was next to the above weed species.

**4.4 Morphological specifications of invasive and native weeds** 

and experimentation in details.

*Achyranthes* (96.33cm)*.* 

was comparatively very less.

campus of Pune University.

campus, showing luxuriant growth.

per plant respectively.

monothickets (Bhakat et al. 2006) of Cassia uniflora at sites I, II and IV and that of Synedrella at site III.

The results of GPS mapping of weeds in Pune University campus strongly support the phytosociological observations recorded through quadrat studies. The GPS mapping had given the exact latitude, longitude and altitude of each plant species. More than 200 waypoints were recorded to confirm the dominance of selected weed species like *Cassia uniflora, Synedrella nodiflora, Alternanthera tenella, Bidens pilosa, Blainvillea acmella, Acalypha ciliata, Euphorbia geniculata, Triumfetta rhomboidea, Cassia obtusifolia* etc. It has also indicated the dominance of *Cassia uniflora* and *Synedrella nodiflora* at all the four selected sites. These selected weeds were located at 18055' north latitude and 73082'east longitude and an altitude of 568.63m and 571.48m respectively.

The Millenium Ecosystem Assessment (2005) have claimed that invasive species are the most important drivers of ecosystem change, which can very well alter the vegetational set up of a particular area. Such phytosociological studies on various weeds occurring in crop ecosystems have been carried out by researchers like Bartariya et al. (2005) and Seal et al. (2009) have also reported the dominance of invasive weeds in different ecosystems.

### **4.3 Secrets of invasion / encroachment and aggressiveness**

The invasive weeds can exploit many niches left available and keep changing the phytodiversity of these niches or ecosystems. Unless the phytosociological studies of such areas are carried out, it is difficult to know the extent of encroachment over natives and invasion by the invasive plants. Phytosociology will help to understand the growth characteristics, dominance, distribution and adaptations which enable these plants to sustain the changes in the environment. These studies help to determine the distribution, prevalence, competing ability, behaviour and survival of weeds (Rao, 2000). The results recorded in Tables 1(a, b, c and d) clearly showed the higher dominance of *Cassia uniflora*  and *Synedrella nodiflora*, at different sites in the university campus amongst the co-occuring species. These two invasive weed species have caused the reduction in the native phytodiversity of Pune University campus. As suggested by Rizvi and Rizvi (1992), allelopathic interactions of these weeds might be playing a crucial role in existing vegetation pattern of Pune University campus.

The dominance of *Cassia uniflora* and *Synedrella* in the study area may be attributed to their aggressive nature, allelolpathic potential, adaptations in morphological and reproductive features along with specific type of physiological, biochemical and enzymological mechanisms allowing their faster growth and tolerance to biotic and abiotic stress conditions.

Plants are chemically well defended in their environments, because their exposure to any stress leads to the qualitative and quantitative changes in the plant biochemicals and enzymes as a part of defense mechanism. These defensive chemicals are nothing but allelochemicals only, which act as feeding deterrents or alter the physiology and development of the attacking organisms (Pathipati UshaRani 2008). Even the different organic compounds often have role in ecological development which mediates interactions between the donor plants and the recipient organisms. The defensive allelochemicals and organic compounds have crucial role in the weed–weed associations formed in the campus of Pune University. The allelopathic potential of invasive weeds like *Cassia* and *Synedrella* can be ascribed to the above mentioned factors.

monothickets (Bhakat et al. 2006) of Cassia uniflora at sites I, II and IV and that of Synedrella

The results of GPS mapping of weeds in Pune University campus strongly support the phytosociological observations recorded through quadrat studies. The GPS mapping had given the exact latitude, longitude and altitude of each plant species. More than 200 waypoints were recorded to confirm the dominance of selected weed species like *Cassia uniflora, Synedrella nodiflora, Alternanthera tenella, Bidens pilosa, Blainvillea acmella, Acalypha ciliata, Euphorbia geniculata, Triumfetta rhomboidea, Cassia obtusifolia* etc. It has also indicated the dominance of *Cassia uniflora* and *Synedrella nodiflora* at all the four selected sites. These selected weeds were located at 18055' north latitude and 73082'east longitude and an altitude

The Millenium Ecosystem Assessment (2005) have claimed that invasive species are the most important drivers of ecosystem change, which can very well alter the vegetational set up of a particular area. Such phytosociological studies on various weeds occurring in crop ecosystems have been carried out by researchers like Bartariya et al. (2005) and Seal et al.

The invasive weeds can exploit many niches left available and keep changing the phytodiversity of these niches or ecosystems. Unless the phytosociological studies of such areas are carried out, it is difficult to know the extent of encroachment over natives and invasion by the invasive plants. Phytosociology will help to understand the growth characteristics, dominance, distribution and adaptations which enable these plants to sustain the changes in the environment. These studies help to determine the distribution, prevalence, competing ability, behaviour and survival of weeds (Rao, 2000). The results recorded in Tables 1(a, b, c and d) clearly showed the higher dominance of *Cassia uniflora*  and *Synedrella nodiflora*, at different sites in the university campus amongst the co-occuring species. These two invasive weed species have caused the reduction in the native phytodiversity of Pune University campus. As suggested by Rizvi and Rizvi (1992), allelopathic interactions of these weeds might be playing a crucial role in existing vegetation

The dominance of *Cassia uniflora* and *Synedrella* in the study area may be attributed to their aggressive nature, allelolpathic potential, adaptations in morphological and reproductive features along with specific type of physiological, biochemical and enzymological mechanisms allowing their faster growth and tolerance to biotic and abiotic stress

Plants are chemically well defended in their environments, because their exposure to any stress leads to the qualitative and quantitative changes in the plant biochemicals and enzymes as a part of defense mechanism. These defensive chemicals are nothing but allelochemicals only, which act as feeding deterrents or alter the physiology and development of the attacking organisms (Pathipati UshaRani 2008). Even the different organic compounds often have role in ecological development which mediates interactions between the donor plants and the recipient organisms. The defensive allelochemicals and organic compounds have crucial role in the weed–weed associations formed in the campus of Pune University. The allelopathic potential of invasive weeds like *Cassia* and *Synedrella*

(2009) have also reported the dominance of invasive weeds in different ecosystems.

**4.3 Secrets of invasion / encroachment and aggressiveness** 

at site III.

of 568.63m and 571.48m respectively.

pattern of Pune University campus.

can be ascribed to the above mentioned factors.

conditions.

Allelochemicals always affect many aspects of plants' ecology e.g. distribution, growth, succession, structure of communities, dominance, diversity and productivity (Takeuchi et al. 2001). The population of *Cassia uniflora* had always shown its shifting nature, i.e. area with monothickets during first year will show very less population next year on the same spot. It may be due to the resource exhaust, autotoxicity and heavy accumulation of allelochemicals making the area inhospitable for its own growth. However, it requires further confirmation and experimentation in details.

### **4.4 Morphological specifications of invasive and native weeds**

An attempt was made to study the morphological and reproductive features of invasive and native weeds from Pune university campus (Table 2). The maximum plant height was recorded for *Cassia uniflora,* (104.66cm), which was followed by *Triumfetta* (103.66cm) and *Achyranthes* (96.33cm)*.* 

The root length indicates easy and proper establishment of the plants. The maximum root length was recorded for *Alternanthera* followed by *Rauwolfia* and *Achyranthes*.

The highest number of branches per plant was observed in *Achyranthes* and followed by *Triumfetta,* while the weed species like *Acalypha* and *Oplismenus* were without any branch. The number of branches in remaining weeds was next to the above weed species.

*Cassia obtusifolia* had maximum third leaf area and it was followed by *Rauwolfia* and *Triumfetta.* The third leaf area in remaining weeds was very less as compared to them.

The results on fresh biomass per plant indicated that *Cassia obtusifolia* was having highest biomass and *Triumfetta* was next to it. The weight of fresh biomass in the remaining weeds was comparatively very less.

The dry biomass per plant showed wide variations ranging from 0.89g to 17.43g. The *Cassia*  species e.g. *C. obtusifolia* and *C. uniflora* were having highest dry biomass 17.43g and 12.4g per plant respectively.

The highest fresh biomass per m2 area was recorded in *Triumfetta, Cassia uniflora* and *Blainvillea.* The remaining weeds recorded comparatively less biomass per m2 area in the campus of Pune University.

Many research workers like Sen (1977), Weaver and McWilliams (1980), Wilson (1988) had given due importance to morphological studies of native and invasive weeds. Ehrenfeld (2003) proposed that the invasive plants share many physical characteristics and tend to alter habitats. They have very high productivity and above ground biomass. They grow earlier in the season and show faster growth rate than native species. All such features might be applicable to *Cassia* and *Synedrella,* because of which they are dominant in the campus, showing luxuriant growth.

Unless we know the morphological features of weeds, the attempts for their effective management are difficult. With this view, Sutherland (2004) has described all the morphological details of different terrestrial and aquatic weeds of India. Similarly Monaco et al. (2002) had also investigated the various morphological characteristics of different weeds from USA. The life cycle of a plant can be understood well by its morphological structures, developmental processes and whole plant activities that occur during each phase of its life cycle. He further stated that selected phenotypes dominate their neighbours, because the timing of their life history optimizes their relative fitness and minimizes mortality. Same explanation may be true for the dominance of different invasive weeds including *Cassia* and *Synedrella* over the natives of Pune University campus.

Morphophysiological Investigations in Some Dominant Alien Invasive Weeds 27

spike or cyme. Only in *Euphorbia,* it was cyathium, and in *Oplismenus* it was panicle. In almost all the weeds the inflorescence was axillary, but only in *Achyranthes* and *Cassia absus*,

For the number of inflorescences per plant *Triumfetta,* was at first rank, which was followed sequentially by *Alternanthera, Synedrella* and *Cassia uniflora.* The weed species like *Blainvillea*, *Acalypha* and *Achyranthes* were at medium position, while the least number was noted in

The number of flowers, florets and flowerbuds per inflorescence was also a diagnostic feature for all the weeds. *Acalypha* was having highest number of male and female flowers together, whilst *Achyranthes* was following it (Table 3a). *Oplismenus* was in third position and it was followed by *Synedrella*. Remaining weeds had very few numbers of flowers**/**

The number of fruits per inflorescence is very important reproductive character which was

Snl > Ach > Bdn > Alt > Tum > Bln > Aca > Cul > Cfl > Cab > Opl > Eug > Raw (Table 3b). The number of seeds per plant was not showing the same order as that of number of fruits (Table 3b). This was due to the number of seeds per fruit, which is again a specific and variable character for a particular plant. The descending sequence for number of seeds per

Cfl > Snl > Ach > Cul > Aca > Bdn > Tum > Cab > Alt > Bod > Bln > Eug > Opl > Raw.

Snl Axillary heads 60.0 c ± 3.00 21.0 cd ± 1.05 Alt Axillary clusters 75.0 b ± 5.25 9.0 fgh ± 0.63 Eug Cyathium 9.0 i ± 0.27 8.0 fgh ± 0.24

Ach Terminal spikes 21.0 g ± 1.26 36.0 b ± 2.16 Bln Heads in erect terminal cymes 25.33 f ± 1.01 13.0 efg ± 0.52 Aca Androgynous spikes 23.0 fg ± 1.61 178.3 a ± 12.48

Cab Terminal raceme 8.66 i ± 0.43 14.0 ef ± 0.7 Cfl Axillary subsessile pairs of flowers 9.33 i ± 0.27 14.33 ef ± 0.42 Bdn Corymbose panicled heads 15.66 h ± 0.93 16.66 de ± 0.99 Raw Umbelliform cyme 9.66 i ± 0.67 7.33 gh ± 0.51 Opl Panicle 2.0 j ± 0.08 24.0 c ± 0.96 p**=** <0.001 <0.001

plant

panicles 30.0 e ± 1.8 13.0 efg ± 0.78

38.0 d ± 1.52 8.0 fgh ± 0.32

90.0 a ± 2.7 4.0 h ± 0.12

No. of flowers, florets or floral buds per inflo.

florets**/** flowerbuds and the lowest number of flowers was in *Triumfetta.*

species Type of Inflorescence No. of Inflo. /

Crowded upwards

cymes

Table 3. (a) Reproductive features of invasive and native weeds

Cul Axillary raceme/ subsessile pairs,

Bod Umbels in terminal corymbose

Tum Dense terminal and leaf opposed

it was terminal.

*Oplismenus.* 

in the order of –

plant was:

Weed

