Journal of Applied Botany and Food Quality 86, 138 - 142 (2013), DOI:10.5073/JABFQ.2013.086.019 Agricultural Biotechnology and Phytopathology Laboratory, Department of Botany, University of Karachi, Karachi, Pakistan Management of root diseases of eggplant and watermelon with the application of asafoetida and seaweeds Ghulam Nabi Baloch, Samrah Tariq, Syed Ehteshamul-Haque, Mohammad Athar, Viqar Sultana, Jehan Ara (Received October 10, 2012) Summary Eggplant (Solanum melongena L.) and watermelon (Citrullus la- natus (Thunb.) Matsum. & Nakai) are highly susceptible to root rotting fungi Fusarium solani, F. oxysporum, Macrophomina phaseolina and root knot nematode (Meloidogyne spp.) causing huge losses each year in Pakistan. In field experiments, application of asafoetida, a medicinal gum from Ferula assafoetida and sea- weeds Spatoglossum variabile, Stokeyia indica and Melanothamnus afaqhusainii showed significant suppressive effect on root rotting fungi Fusarium solani, Macrophomina phaseolina and root knot nematode Meloidogyne incognita) attacking watermelon and eggplant and improved plant growth in soil naturally infested with root rotting fungi and artificially infested with root knot nematode. Length of vine of watermelon, shoot length of eggplant and fresh shoot weight were higher in seaweed and asafoetida treated plants as compared to control or Topsin-M, a fungicide, treated plants. Seaweed and asafoetida treated plants also showed earlier fruiting than control or fungicide treated plants. At farmer’s field seaweed showed similar suppressive effect on F. solani and M. phaseolina and root knot nematode on watermelon in soil naturally infested by these pathogens. Application of seaweed produced healthy plants and number of fruits and weight were significantly higher in seaweed and asafoetida treated plants. Asafoetida and seaweeds offer a non- chemical means of disease management. Introduction Cucurbits and vegetable crops are highly susceptible to a number of root and soilborne diseases causing great losses in yield and quality (Sharma et al., 2004; Chehri et al., 2010). Watermelon (Citrullus lanatus (Thunb.) Matsum & Nakai) is a cucurbit fruit, grown throughout the world and consumed as fresh fruit. Fusarium wilt of watermelon caused by Fusarium oxysporum f. sp. niveum is among the most important diseases of watermelon (Zhou and evertS, 2004). The disease is well known as mature watermelon vine decline (sudden wilt of watermelon). Infected plants also showed the presence of Fusarium solani, Macrophomina phaseolina, Rhizoctonia solani, Monosporascus cannonballus and Pythium spp. besides F. oxysporum (BoughalleB and el mahjouB, 2006). The disease is best controlled through the use of wilt resistant cultivars and crop rotations for a minimum of 5 to 7 years (martyn, 1996). However, resistance in commercial cultivars often no longer is effective due to the presence of the highly aggressive race of F. oxysporum (martyn, 1987). Similarly, among the various vegetables, eggplant (Solanum melongena) is one of the most common and extensively grown all over the world. Eggplant wilt complex caused by a number of fungal genera such as Fusarium, Verticilium, Rhizoctonia, Sclerotium and Phytophthora take a considerable portion of produce annually (najar et al., 2011). Moreover, eggplant is highly susceptible to toot knot nematodes, species of Meloidogyne (Zarina and Shahina, 2010). Several nonchemical methods including addition of organic amend- ments are an effective method for controlling soilborne pathogens and diseases in various field crops (Chellemi, 2002; huBer, 1980; Zhou and evertS, 2004). Marine bioactive substances extracted from seaweeds have been used for several decades to enhance plant growth and productivity. (rathore et al., 2009; Stirk and van Staden, 1997). The application of seaweeds as organic soil amendment, has been increased in the recent years due to raising awareness about the adverse effect of chemical pesticides (Sultana et al., 2007; 2008; 2009; 2011). Similarly, asafoetida or hing, a dry latex or resinous gum from Ferula assafoetida has been widely used in various indigenous systems of medicine in India and Pakistan. It is regarded as an effective remedy for worms and other intestinal parasites. From old ages, farmers in Malir, Karachi area of Sindh, are applying asafoetida for preventing the plants from root diseases particularly nematode attacks and increasing yield. However, scientific reasons are still unclear. In this study, efficacy of some seaweeds and asafoetida were evaluated in field plots on watermelon and eggplants in suppressing the soilborne diseases and their effect on plant growth. The experiment was also conducted at farmer’s field on watermelon. Materials and methods Asafoetitda (Ferula assafoetida) was purchased from local market whereas seaweeds Melanothamnus afaqhusainii Shameel, Spatoglossum variabile Fig. et De Notar [= S. lubricum Fig. et De Notar] and Stokeyia indica Thivy et Doshi [= Cystoseira indica (Thivy et Doshi) Mairh] were collected at the coastal area of Karachi at low tide. The seaweeds were washed under tap water, dried under shade, ground to powder and stored in polyethylene bags at room temperature until used. Field plot experiments The experiments were conducted at the Crop Diseases Research Institute, Pakistan Agricultural Research Council, Karachi Univer- sity Campus, Karachi. Dry powder of seaweeds Melanothamnus afaqhusainii, Spatoglossum variabile and Stokeyia indica were mixed in sandy loam soil at 70 g per two meter rows and watered 2-3 days interval to allow the organic matters to decompose, where- as aqueous suspension of asafoetida (200 pp) was applied at 400 ml per 2 meter row. The soil had a natural infestation of 5-16 sclerotia/g of soil of Macrophomina phaseolina (Sheikh and ghaffar, 1975), 5-12% colonization of Rhizoctonia solani on sorghum seeds used as baits (Wilhelm, 1955) and 2500cfu/gm of soil of mixed population of Fusarium oxysporum and F. solani as determined by soil dilution (naSh and Snyder, 1962). After two weeks of seaweed decomposition, 12 seeds of watermelon were sown in each row. Whereas in the case of eggplant, three-week-old seedlings of equal size, raised in steam sterilized soil, were transplanted in each row at 12 seedlings per row. After one week of seed germination or seedling transplantation, each row was inoculated with aqueous suspension of Meloidogyne incognita eggs/juveniles at 2000 / two meter row. Topsin-M (200 ppm) at 400 ml / 2 meter row and carbofuran (1 g / Management of root diseases of eggplant and watermelon 139 2 m row) served as positive control against fungi and nematode respectively. Each treatment was replicated four times and plants were watered 2-3 days intervals depending upon requirement of plants. To determine the efficacy of seaweeds and asafoetida on the root pathogens and plant growth, plants were uprooted after six weeks of nematode inoculation. Observations were made on yield, plant height, fresh shoot weight, root length and root weight. Nematode infection was determined by counting the numbers of galls per root system (tayler and SaSSer, 1978). To determine nematode penetration and infection by root-infecting fungi, roots from each plant were cut into 1 cm long pieces and five pieces of tap roots from each plant were used for assessment of fungal infection. The remaining roots were mixed thoroughly, and 1 gram sub-sample was wrapped in muslin cloth and dipped in boiling 0.25% acid fuchsin stain for 3-5 minutes. Roots were left in the stain to cool, and then washed under tap water to remove excess stain. Roots were transferred to vials containing glycerol and water (1:1 v:v) with a few drops of lactic acid. Roots were macerated in an electric blender for 45 seconds and the resulting suspension was suspended in 50 ml water. Numbers of juveniles and females in five 5 ml sub samples were counted with the aid of dissecting microscope and numbers of nematode/g root were calculated (Siddiqui and ehteShamul- haque, 2001). To determine the incidence of fungal infection, 1 cm long root pieces from tap roots (five pieces from each plant) were surface disinfested with 1% Ca (OCl)2 solution and plated onto potato dextrose agar amended with penicillin (100,000 units/l) and streptomycin (0.2 g/l). After incubation for 5 days at 28°C, colonies of Macrophomina phaseolina, Rhizoctonia solani and species of Fusarium were recorded. The experiment was repeated twice. Data were subjected to analysis of variance (ANOVA) and means were separated using the least significant difference (LSD) according to gomeZ and GomeZ (1984). Farmer’s field experiment Efficacy of asafoetida and seaweeds in protecting the watermelon from root rotting fungi and root knot nematode were also evaluated at farmer’s field in Malir, Karachi. Watermelon field about 6 acres was selected before the sowing of seeds. Fifteen meter rows were selected for each treatment at different locations in the field and dry powder of seaweeds Melanothamnus afaqhusainii, Spatoglossum variabile and Stokeyia indica were mixed in soil at 35 g per meter and watered 2-3 days interval to allow the organic matters to de- compose, whereas aqueous suspension of asafoetida (200 pp) was applied at 200 ml per meter. The soil had variable natural infestation of root rotting fungi and root knot nematode. After two weeks of amendment, seeds of watermelon were sown in each row. Observations were recorded after 80 days of seed germination. Results Field plot experiment Eggplant Fusarium solani and Macrophomina phaseolina were found in higher frequencies in control plants than R. solani or F. oxysporum. Plants grown in seaweed or asafoetida amended soil showed less infection of M. phaseolina than control plants. Spatoglossum variabile was also found effective against F. solani (Tab. 1). Seaweed and asafoetida also showed significant suppressive effect on root knot nematode by reducing gall formation on roots and nematode’s penetration in roots as compared to untreated control (Tab. 2). Efficacy of asafoetida and seaweed against root knot nematode is comparable with carbofuran, a nematicide. Taller plants and greater fresh shoot weight was produced by S. indica (Tab. 2). Watermelon Application of seaweeds Spatoglossum variabile, Melanothamnus afaqhusainii and asafoetida caused a suppressive effect on root rotting fungi by reducing the infection of M. phaseolina, R. solani and F. oxysporum on watermelon roots. Stokeyia indica was effective against F. solani (Tab. 3). Seaweeds and asafoetida also showed suppressive effect on root knot nematode by reducing the nematode’s penetration in roots (Tab. 4). Vine length and fresh weight were significantly higher in watermelon grown in Spatoglossum variabile amended soil than other treatments (Tab. 4). Plants grown in sea- weed or asafoetida amended soils also showed earlier emergence of fruits than control of pesticides treatments. Farmer’s field experiment Watermelon Infection of Fusarium solani was found in higher frequencies in control or treated watermelon plants. However, plants grown in S. variabile amended soil showed less infection of F. solani than other treatments, whereas topsin-M treated plants showed no in- fection of F. solani (Tab. 3). Macrophomina phaseolina infection was found in control plants only. Nematode penetration in water- melon roots were found less in S. indica S. variablie, asafoetida amended soil or in topsin-m treated plants (Tab. 4). Number of fruits and fruit weight were higher in plants grown in asafoetida or seaweed Tab. 1: Effect of asafoetida and seaweed on the infection of root infecting fungi on eggplant in field plot experiment. No. Treatments M. phaseolina R. solani F. solani F. oxysporum Infection % 1. Control 68.7 6.2 31.2 6.2 2. Topsin-M 12.5 0 12.5 0 3. Carbofuran 18.7 31.2 43.7 6.2 4. Asafoetida 6.2 50 43.7 0.0 5. Stokeyia indica 0.0 0.0 50 0.0 6. Spatoglossum variabile 0.0 6.2 18.7 0.0 7. Melanothamnus afaqhusainii 12.5 12.5 25 0.0 LSD0.05 Treatments= 10.31 Pathogens= 7.82 1Mean values in column showing differences greater than LSD values are significantly different at p< 0.05. 2Mean values in rows showing differences greater than LSD values are significantly different at p< 0.05. 140 G.N. Baloch, S. Tariq, S. Ehteshamul-Haque, M. Athar, V. Sultana, J. Ara Tab. 2: Effect of asafoetida and seaweed on the infection of root knot nematode and growth of eggplant in field plot experiment. No. Treatments No. of knots J2/females Plant Fresh Root Fresh /g roots height shoot length root (cm) wt. (g) (cm) wt. (g) 1. Control 92.5 192 14.2 20.2 7.7 11.7 2. Topsin-M 23.2 124 21.2 29.7 5.2 4.1 3. Carbofuran 16 80 26.5 33.7 11 10.2 4. Asafoetida 16.5 80 18.7 32.3 9.7 12.7 5. Stokeyia indica 7 72.7 38.5 40.1 10.5 11.7 6. Spatoglossum variabile 12 79.2 24.5 29.5 8.2 4.5 7. Melanothamnus afaqhusainii 24.2 102.5 17 27.6 6.7 2.6 LSD0.05 35.11 15.31 9.61 14.01 3.11 3.61 1Mean values in column showing differences greater than LSD values are significantly different at p< 0.05. Tab. 3: Effect of asafoetida and seaweeds on the infection of root infecting fungi on watermelon in field plot experiment (after 45 days of nematode inocu- lation) and at farmer’s field (after 80 days of sowing). No. Treatments M. phaseolina R. solani F. solani F. oxysporum Infection % Field plot experiment 1. Control 31.2 25 37.5 18.7 2. Topsin-M 43.7 25 50 12.5 3. Carbofuran 12.5 6.2 43.7 18.7 4. Asafoetida 6.2 0 18.7 0 5. Stokeyia indica 50 12.5 18.7 0 6. Spatoglossum variabile 6.2 6.2 31.2 6.2 7. Melanothamnus afaqhusainii 0 0 50 0 Farmer’s field experiment 1. Control 18.7 6.2 37.5 0 2. Topsin-M 0 0 0 0 3. Asafoetida 0 0 12.5 0 4. Stokeyia indica 0 6.2 37.5 0 5. Spatoglossum variabile 0 0 18.7 0 6. Melanothamnus afaqhusainii 0 0 31.2 0 LSD0.05 Field plot experiment Treatments= 9.071 Pathogens= 6.862 Experiment at farmer’s field Treatments= 7.31 Pathogens= 5.22 1Mean values in column showing differences greater than LSD values are significantly different at p< 0.05. 2Mean values in rows showing differences greater than LSD values are significantly different at p< 0.05. amended soil than control plants. Vine length and fresh weight was found significantly higher in plants grown in M. afaqhusainii amended soil (Tab. 4). Discussion Protecting the plant roots from soilborne plant diseases with or- ganic amendments and improving plant growth and yield is practiced since long by the farmers throughout the world. The suppressive effect of organic amendments on diseases is primarily associated with a reduction in pathogen inoculums density in amended soil (Chun and loCkWood, 1985; Sun and huang, 1985; SuBBa- rao et al., 1999). In this study, soil amendment with seaweed or asafoetida in general showed suppressive effect on soilborne pathogens and have a positive effect on growth of watermelon both in field plots and at farmer’s field. Plant grown in seaweed or asafoetida amended soil showed earlier fruiting, greater number of fruits and weight than control plants. It was reported that seaweed extracts increased plant resistant to pests and diseases, improve plant growth, yield and quality (rathore et al., 2009; Stirk and van Staden, 1997; verkleij, 1992). Application of seaweed to plants can result in decreased levels of nematode attack (ara et al., 1997; Wu et al., 1997; 1998), due to presence of betaines (Wu et al., 1997) and root rotting fungi (Sultana et al., 2007; 2008; 2009; 2011) due to 1-aminocyclopropane-1-carboxylic acid (ACC), which has antimicrobial activity (NELSON and van Standen, 1985) or due to volatile compounds and essential oils (kajiWara et al., 2006). Seaweed could also affect cell metabolism through the induction of the synthesis of antioxidant molecules which could favor plant growth and plant resistance to stress (ZHANG and SCHMIDT, 2000). TARIQ et al. (2011) has already reported the presence of polyphenols and antioxidant activity of seaweeds from the Karachi Management of root diseases of eggplant and watermelon 141 coast. In this study, the potential of asafoetida in suppressing the soilborne diseases has been confirmed. Asafoetida or hing, is regarded as an effective remedy for worms and other intestinal parasites and pos- seses antimicrobial and antioxidant activities (iranShahy and iranShahi, 2011). Biological activity of sesquiterpene coumarins from Ferula species has also been reported (naZari and iranShahi, 2011). Similarly lee et al. (2009) reported antiviral and cytotoxic agents from Ferula assafoetida. Growth inhibition of plant patho- genic fungus Fusarium by the essential oils extracted from asafetida has also been reported (Sitara et al., 2008). Due to the damaging influence on crop yields, the root knot nematodes Meloidogyne incognita, M. javanica, M. arenaria and M. hapla are considered economically important pests worldwide (roBertSon et al., 2006). The damages are much higher in tropical and subtropical countries where environmental factors favor their survival and dispersal (Sikora and fernandeZ, 2005). In this study, the application of asafoetida and seaweeds besides, suppressing the same root rotting fungi also showed significant reduction in gall formation on roots and nematode’s penetration into roots of eggplant and improve plant growth. Eggplant is susceptible to several insects and pest, particularly Fusarium wilts (F. oxysporum f. sp. melongenae) and root knot nematodes (Meloidogyne spp.) which reduced the yield (PhaP et al., 2010). Due to high susceptibility, eggplant is widely used for maintaining the pure culture of root knot nematode (Siddiqui et al., 2001; Sikander et al., 2009). In this study, efficacy of asafoetida and seaweed is comparable to carbofuran, a commercial nematicide. Nonchemical management of soilborne pests has been practiced for centuries. Only in the last 40 years agricultural producers have come to rely on synthetic chemicals for the control of soilborne pest and diseases (Chellemi, 2002). However, due to raising awareness about the adverse effect of chemicals, organic forming is gaining popularity (Sultana et al., 2011). The ability of seaweeds and asafoetida in reducing the root knot nematode on a highly susceptible crop showed its great potential in future nematode pest management. Acknowledgements Financial support provided the Dean, Faculty of Science, University of Karachi is sincerely acknowledged. 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Address of the author: Mr. Ghulam Nabi Baloch, Agricultural Biotechnology and Phytopathology Laboratory, Department of Botany, University of Karachi, Karachi-75270, Pakistan. Miss Samrah Tariq, Agricultural Biotechnology and Phytopathology Laboratory, Department of Botany, University of Karachi, Karachi-75270, Pakistan. Prof. Dr. Syed Ehteshamul-Haque, Agricultural Biotechnology and Phytopathology Laboratory, Department of Botany, University of Karachi, Karachi-75270, Pakistan. Dr. Mohammad Athar, Pest Detection and Emergency Projects, California Department of Food and Agriculture, 3288 Meadowview Road, Sacramento, CA 95832, USA. E-mail: atariq@cdfa.ca.gov Prof. Dr. Viqar Sultana, Biotechnology and Drug Development Laboratory, Department of Biochemistry, University of Karachi, Karachi-75270, Pakistan. Prof. Dr. Jehan Ara, Post Harvest Technology and Food Biochemistry Laboratory, Department of Food Science and Technology, University of Karachi, Karachi-75270, Pakistan.