1. INTRODUCTION. oldest commercial crops, plays a key role in the economic and social affairs of the world

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1 1. INTRODUCTION Cotton (Gossypium spp.) regarded as white gold is one of the important and oldest commercial crops, plays a key role in the economic and social affairs of the world (Shah et al., 2011; Akhtar et al., 2013). It is grown chiefly for its fibre, providing basic input to the textile industry. In India, cotton occupies five per cent of the total cultivated area and contributes about 85 per cent of raw material in textile manufacturing of the world (AICCIP, 2014). China ranks first in the world cotton production with 33 million bales, followed by India (25 million bales), and the United States (16.7 million bales) (USDA, 2014). On a worldwide basis, cotton is cultivated in an area of million ha during with a production of million bales of lint accounting for a productivity (lint) of 766 kg ha -1 (USDA, 2014). In India, cotton is cultivated in an area of million ha during with an average annual production of million bales of lint and productivity (lint) of kg ha -1. In Tamil Nadu, cotton is cultivated in an area of 1.17 million ha during with production of 2.80 million bales of lint and productivity (lint) of 726 kg ha -1 (Cotcorp.gov.in., 2014). In India, the productivity of cotton is very low due to many constraints including diseases. Diseases are inherent compounds of agro ecosystem that must be dealt continuously and on knowledge basis. Cotton is affected by various diseases caused by fungi, bacteria and viruses. Of these, wilt of cotton is a vascular disease caused by Fusarium oxysporum f. sp. vasinfectum (Atk.) Snyder and Hans is an important soil borne pathogen, distributed worldwide. The disease is now widespread and causes substantial crop losses in most of the major cotton-producing areas of the

2 2 world (Colyer, 2001). In India, it was first reported in Nagpur and the loss due to wilt is estimated to be 5-47% and in Tamil Nadu about 10-15% loss was reported in Palladam areas (Hemalatha, 2008). Since, the first report of Fusarium wilt of cotton, in Alabama (Atkinson, 1892), the disease has increased in importance (Davis et al., 2006) and is responsible for losses averaging $20 million each year across the cotton belt of the United States of America (Blasingame et al., 2008). The loss in terms of production potential attributable to Fusairum wilt in the U.S.A has been reported as 13.6% (Glass et al., 2013). Under conducive environmental conditions, high losses occur when susceptible cultivars are grown on heavily infested soil. Losses are greatest on sandy soils that are infested with M. incognita (DeVay et al., 1997). Fusarium wilt in cotton is one of the first reports of an interaction between a nematode and a fungus, and is now one of the most recognized disease complexes in the world (Colyer et al., 1997). Both pathogens are common in most cotton producing areas and often inhabit the same fields. These two pathogens often infect cotton simultaneously, forming a complex that increases the incidence and severity of Fusarium wilt (Colyer and Vernon, 2003). Currently, six distinct races, restricted to defined geographic areas, have been described for this wilt pathogen. Races 1 and 2 were described in the United States and Tanzania, race 3 in Egypt, Sudan, and Israel, race 4 in India, race 5 in Sudan, and race 6 in Brazil and Paraguay (Hillocks, 1992; Kim et al., 2005). Recent studies have found that races 1 and 3 are mildly virulent and cause wilt symptoms in the presence of M. incognita. However, race 4 of Fov, is capable of causing severe wilt symptoms and economic loss in the absence of nematodes (Ulloa et al., 2006). Fov invades the host through the tap roots behind the root tip. The combined effect of fungal

3 3 metabolites and the production of lipoidal substances by the host in response to infection may lead to the occlusion of the vascular tissues resulting in wilt symptoms (Shi et al., 1992; Hillocks, 1992). Initial symptoms of Fusarium wilt include chlorosis and necrosis of the leaf margins. Severely infected plants may be killed or if they survive, may often remain stunted throughout the season (Chawla et al., 2012). In most severely affected plants, leaves wilt, drop off and the plants may die (Nelson, 1981; Colyer, 2001). Internal symptoms of Fov infection include brown discoloration in the vasculature. Plants infected at a later stage of crop growth may be wilted and stunted, and leaves may display chlorosis and necrosis. The characteristic brown vascular discoloration is most evident in the roots and lower stem, but may be apparent in upper branches as well (Hao et al., 2009). Plants that develop symptoms early usually die before producing any bolls, whereas plants that develop symptoms after the onset of flowering often survive but produce fewer bolls. Once a field is infested with Fov, the fungus usually persists indefinitely in the decaying plant tissues and soil in the form of chlamydospores (Nelson, 1981; Mai and Abawi, 1987). The fungus also survives in association with non-hosts, a particularly challenging trait since the ability to colonize roots of weeds and other crops allows Fov to persist in soil for years or decades (Smith and Snyder, 1975). The fungus is capable of surviving for over 10 years in the soil not planted with cotton (Smith et al., 2001). Fov also persists in seed, which can occur by contamination of cotton bolls in the field (Bennett et al., 2008). Thus, the ability of the pathogen to survive in soils for long periods has important consequences on disease management.

4 4 Management of F. oxysporum f. sp. vasinfectum using chemical fungicides has been the prevailing control method for over fifty years. Though effective fungicides are available to mange the soil borne diseases, they will not be reliable as a long term solution because of the concerns about exposure risks, health and environment hazards. Moreover, the frequent applications of fungicides may lead to the development of tolerance in the target organisms. As a result, in recent years, the biological control especially using fungal and bacterial antagonists against fungal plant pathogen has gained considerable attention and appears to be promising as a viable alternative to chemical control (Hafez et al., 2013; Sivasakthi et al., 2014). Management of soil borne diseases through biological control by addition of antagonistic microorganisms is known to be ecofriendly and effective method (Cook and Baker, 1983). Members of the genus Pseudomonas, Bacillus and Trichoderma have been known for their potential antifungal, plant growth promoting and plant defense inducing activities (Zaidi et al., 2004). Though a huge number of strains are commercially available in the market, the native isolates of certain biocontrol agents showed superiority over other isolates for the management of soil borne diseases (Dubey and Patel, 2001). Hence, the possibility of exploiting the native antagonistic potential was thought off for the management of Fusarium wilt disease, which may represent a potential eco-friendly strategy. However, application of single antagonistic strain in a variety of eco systems often results in inconsistent management of the diseases. One of the strategies for overcoming such inconsistent performance is to combine two or more beneficial microbes in a biocontrol preparation (Raupach and Kloepper, 1998). Such combinatory

5 5 approach has the potential to overcome problems that occur with individual biocontrol agents (Meyer and Roberts, 2002) as combinations of biocontrol agents have the potential for extensive colonization of the rhizosphere, broad spectrum of activity, and consistency of biocontrol resulting in enhanced efficacy and reliability of biological control. Several researchers have also tested different biocontrol strains with different modes of actions in combination (Droby, 2001) to increase the efficacy and the consistency of disease control. Another possible reason for the inconsistent performance of the antagonists might be due to the failure of the organisms to get established in the new environment due to lack of proper nutritional base in the soil which resulted in limited commercial use of biocontrol approaches for the suppression of plant pathogens (Akkim et al., 2013). Hence, it was thought to use a suitable organic amendment so that it could serve as a nutrient base and also help in maintaining vigour of the plants. Organic amendments of different kinds used as nutrient sources for crop production have been found more effective in control of root diseases of plants (Nagaraju et al., 2010). Thus, use of such potential organic amendment along with native biocontrol agents in the integrated disease management system might provide an alternative, sustainable and inexpensive means of managing diseases like cotton wilt caused by Fov and nematodes. Also, an integrated approach would always ensure the maximum suppression of the disease and higher yield without any deleterious effect on the ecosystem. In this direction the present investigation was carried out to study the combination effect of using an effective biocontrol agent along with an organic amendment for the sustainable eco-friendly management of wilt disease of cotton with the following objectives:

6 6 To survey for the incidence of Fusarium wilt disease of cotton in Cuddalore, Villupuram, Perambalur and Salem districts of Tamil Nadu and to assess the cultural and pathogenic variations among the isolates of F. oxysporum f. sp. vasinfectum. To identify the effective native antagonist(s) for the management of the Fusarium wilt disease and to assess the effect due to application of individual and combination of antagonists on the biometrics and wilt incidence of cotton. To develop an integrated management practice for the effective management of cotton Fusarium wilt disease and To study the induction of defense related enzymes in cotton plants pretreated with biocontrol agents, organic amendment and challenge inoculated with the pathogen.