J. Hortl. Sci. Vol. 14(1) : 13-19, 2019

Original Research Paper

Evaluation of and cultivated varieties for bacterial wilt resistance T.H. Singh1*, D.C. Lakshmana Reddy2, C. Anand Reddy1, A.T. Sadashiva1 P. Pandyaraj1 and Y.B. Manoj1 Division of Vegetable crops, Indian Institute of Horticultural Research (IIHR), Hessaraghatta Lake Post, Bengaluru - 560 089, India 1Division of Vegetable crops, Indian Institute of Horticultural Research (ICAR-IIHR), Bengaluru, India 2Division of Biotechnology, Indian Institute of Horticultural Research (ICAR-IIHR), Bengaluru, India *Email : [email protected]

ABSTRACT Bacterial wilt caused by Ralstonia solanacearum is one of the major diseases in Solanum species including cultivated Eggplant (Solanum melongena L.). Bacterial wilt (BW) disease management in eggplant is difficult due to high survival rate of pathogen in soil and chemical application is not eco-friendly. The best way to avoid bacterial wilt in eggplant is using disease-resistant varieties. However, only a limited number of bacterial wilt resistant varieties are available and, there is a necessity to identify and/or develop new resistant varieties. In the current study, wild Solanum species, and eggplant cultivated varieties were evaluated against Ralstonia solanacearum, and disease incidence was recorded. The cultivated varieties IIHR-108, Pusa Purple Long and Rampur Local were identified as susceptible, whereas, IIHR-7 and CARI-1 were identified as resistant to bacterial wilt. These resistant wild and cultivated varieties can be used as a root-stock in bacterial wilt disease resistant breeding programmes.

Keywords: Wild Solanum species, Eggplant, Ralstonia solanacearum, Disease scoring, Grafting INTRODUCTION eggplant bacterial wilt is mainly caused by Race 1 and Eggplant (Solanum melongena L.) (2n=24) is one of Biovar 3 (Gopalakrishnan et al., 2005). The pathogen the most popular Solanaceous vegetable crop cultivated (Ralstonia solanacearum) can survive in soil upto globally. Asia is the major producer (93%) of eggplant ten years without any host and have the ability (FAO, 2016). It is originated from India and South to colonize in non-host including a vast range China (Daunay and Hazra, 2012). Eggplant is of symptomless weeds (Gopalakrishnan et.al., 2014). generally cultivated in open fields with hot and humid The pathogen enters into the plants through wounds conditions. Eggplant is susceptible to numerous or secondary root initiation points leading to pathogen diseases viz., bacterial wilt; fusarium wilt; verticillium colonization in the vascular parenchyma and cell wall wilt, early blight, spot, virus-Y (PVY), breakage (Ramesh, 2008). The initial wilt symptoms tobacco ring spot virus, spotted wilt virus are leaf drooping, followed by full-plant wilting and (TSWV), phytoplasma, and root-knot nematode. Duo vascular discoloration. When cut ends of wilted plant to these diseases, quality and quantity of eggplant placed in water, milky white ooze out can be observed. production is adversely affected. There are different methods of artificial inoculation/ Eggplant bacterial wilt (Ralstonia solanacearum) is screening of bacterial wilt like root cut inoculation, a major concern in India. The pathogen is race leaf or stem pricking and natural field infestation specific and has five different races. Ralstonia (Ramesh, 2008). These pathogenic spores remain solanacearum strains are divided into five different viable and active in the soil for several years making biovars based on the biochemical analysis. In India, the disease control almost impossible through any means of chemical treatments especially in the regions

13 Singh et al where repeated eggplant cultivation is taken up. Many inoculation, and these can be used as root-stock in of the commercial varieties/hybrids are highly grafting. susceptible to bacterial wilt. Chemical management MATERIALS AND METHODS often leads to the presence of chemical residues in the fruits, thus, raises the concern of food safety. The Plant Material and Phenotypic Characters most economical way of bacterial wilt control is to In this study, five eggplant cultivated varieties viz., develop bacterial wilt resistant varieties/hybrids. IIHR-7 (IC395334), IIHR-662 (CARI-1, IC0585684), Recently, grafting strategy using resistant root-stock IIHR-108 (Arka Kusumakar), IIHR-663 (Rampur has been proposed for soil-borne pathogens like Local), and seven wild species viz., Solanum gilo bacterial wilt, and it will help in bacterial wilt resistant (RS-3), Solanum indicum (RS-4), Solanum viarum varieties/hybrids development with the different genetic backgrounds. Hence, identification of a (RS-6), Solanum aethiopicum (RS-7), Solanum bacterial wilt root-stock will be of higher use and any mammosum (RS-8), and (RS-9 and preferable variety/hybrid can be grafted on it. Here, RS-9a) were used for bacterial wilt screening (Fig.1). we evaluated wild solanum species and cultivated These germplasm accessions were maintained at varieties for bacterial wilt resistance through artificial Indian Institute of Horticultural Research, Bengaluru.

Fig. 1. Eggplant cultivated varieties [IIHR-7 (IC395334), IIHR-662 (CARI-1, IC0585684), IIHR-108 (Arka Kusumakar), IIHR- 663 (Rampur Local)] and seven wild species [Solanum gilo, S. indicum, S. viarum, S. aethiopicum, S. mammosum, and S. torvum used for bacterial wilt screening. 14 J. Hortl. Sci. Vol. 14(1) : 13-19, 2019 Evaluation of eggplant for bacterial wilt resistance ., Plant height: incidence: high racters viz., viz., plantracters viz., Plant height: Plant viz., wilt wilt incidence: high t height: tall; growth tall; height: t otypic characters viz., characters otypic idge-less traits) cooking idge-less red and size, (pea purple shapeudder (turn cow’s te; fruit: small (medium- small fruit: te; g). (300-350 cm); fruit length length (18-20 fruit cm); height (80-90 cm); fruit cm); height (80-90 traits: stant).Quantitative stant).Quantitative traits: stant).Quantitative nd (look like tomato) and tomato) like (look nd 130 cm); fruit length (20- length fruit cm); 130 g). (5-8 ght lant height (130-150 cm); (130-150 lant height t length (15-18 cm); yield cm); lengtht (15-18 length (5.5-6.5 cm); fruit cm); length(5.5-6.5 small-round (white stripes (white small-round fruit cm); (125-135 height (3.0-3.5 cm); average fruit average cm); (3.0-3.5 reen at immature stage and stage at immature reen solitary); cooking qualities: cooking solitary); ht (100-115cm); fruit length fruit (100-115cm); ht ong-green; cooking qualities: cooking ong-green; Phenotypic Characters Phenotypic Table Table 1. Phenotypic traits of elite germplasm line Accession IIHR-108, is a pure line selection from IIHR-193 (A local collection from Karnataka). Phen Karnataka). from collection local (A IIHR-193 from selection line pure a is whi IIHR-108, color: Accession flower green; color: foliage and stem spreading; habit: growth medium/tall; height: Plant diameter long) fruit cm); green;(13-14 length fruit cooking cm); (85-100 qualities: height plant traits: good; Yield: 40t/ha; (susceptible).Quantitative crop duration: 140-150 days and g). bacterial (30-40 wilt weight cha Phenotypic and IIHR-322. ArkaKeshav, l fruit: between cross from purple; line derived breeding is Itan advanced color: flower green; color: foliage and stem spreading; habit: growth tall; height: planttraits: height (100-120 resistant).Quantitative (highly low incidence: wilt and bacterial good g). (40-45 weight fruit average cm); (3.2-3.5 diameter fruit cm); characters Phenotypic islands. Blair) (Port Nicobar & Andaman from collected was line germplasm This resi (highly low incidence: wilt bacterial round-green; fruit: spreading; habit: growth weight fruit medium/tall; average cm); (14-16 diameter fruit cm); (10-12 length fruit cm); (115-130 height plant Plan viz., characters Phenotypic India. UP, of village Rampur from collected was line germplasm This and in clusters borne are (fruits mixed bearing: fruit purple; long-violet fruit: spreading; habit: (115- height plant traits: susceptible).Quantitative (highly high incidence: wilt bacterial and good g). (70-75 weight fruit average cm); (4.0-4.5 diameter fruit cm); 24 This variety was throughdeveloped pure-lone selection by IARI, New Delhi. Phenotypic characters viz Medium; growth habit: spreading; andqualities: fruit:cooking good bacterial long-purple (glossy); traits: frui plant in cm); height Fruits (30-40 are born clusters.Quantitative susceptible). (highly 40-45tonnes/ha.about Plant height: ridge/r and both tall; tomato fruit:habit: like have small-growth erect; round (look heig plant traits: Quantitative resistant). (highly low incidence: wilt bacterial and bad g). qualities: (25-30 weight fruit average cm); (3.0-3.5 diameter fruit cm); (3.5-4.0 and small fruit: ribs); mid and stems on (thorns spreading habit: growth medium/tall, height: Plant traits: plant resistant).Quantitative (highly low incidence: wilt ripe) and when bacterial orange to g). (2-3 weight fruit average cm); (0.9-1.0 diameter fruit cm); (1.0-1.5 length fruit: ribs); mid stem the on present are thorns (dense resi spreading habit: (highly growth tall, height: Plant low incidence: wilt bacterial and ripening) after yellow turn and stage wei immature during fruit average cm); (2.0-3.0 diameter fruit cm); (2.5-3.0 length fruit cm); (100-120 height plant rou oblate small-red, fruit: tinge); purple (have spreading habit: growth medium/tall, height: Plant traits: fruit plantcm); height resistant).Quantitative (85-95 (highly low incidence: wilt bacterial g). (40-50 weight fruit average cm); (4.5-5.0 diameter Plant height: tall, habit: growth thornsspreading (dense are present on the fruit: stem mid ribs); plant traits: resistant).Quantitative (highly low incidence: wilt bacterial and ripening) on yellow g). (15-20 weight fruit average cm); (3.5-4.0 diameter fruit cm); (5.5-6.0 length g dark fruit: white; color: flower foliage); green (dark spreading habit: growth tall, height: Plant p traits: resistant).Quantitative (highly low incidence: wilt bacterial and ripening) on g). turn yellow (10-15 weight fruit average cm); (1.0-1.5 diameter fruit cm); (1.5-2.5 length fruit Solanum indicum Name of the line the of Name IIHR-108 Kusumakar) (Arka IIHR- (IC395334) 7 IIHR-662 IC0585684) (CARI-1, 663 IIHR- Local) (Rampur Pusa Purple Long Solanum gilo Solanum viarum Solanum aethiopicum Solanum torvum 1 2 3 4 5 6 7 8 9 10 11 S;. No. S;.

15 J. Hortl. Sci. Vol. 14(1) : 13-19, 2019 Singh et al

(Fig. 2). The plates were incubated at 28oC - 30oC Basic information of Solanum species and eggplant for 48 hours, and the TZC plates were checked for varieties were described in Table 1. development of avirulent and virulent colonies. The Artificial Screening of Bacterial Wilt separated virulent colonies were selected and suspended in sterile distilled water. The concentration R. solanacearum inoculum was prepared according of inoculum was recorded using a spectrophotometer to Urquhart et al. (1998) from bacterial wilted and stored at 4oC for further use. Soil drenching . The bacterium colonies were collected from method was used for inoculating the 25 days seedlings wilted plant using ooze out test, and streaked on with R. solanacearum suspension, with concentration Triphenyl Tetrozolium Chloride (TZC or TTC) plate

Figure 2. Ooze out test and R. Solanacearum virulence identification in TZC plate. of 1.0 x108 CFU / ml (OD 600 nm = 0.3) (Rashmi et (Markose, 1996). Controlling the bacterial wilt disease al., 2012). is practically not possible with the help of chemicals, and hence growing the resistant variety/hybrid is the A) Resistant B) Susceptible best approach. In the breeding or grafting, Observation and Bacterial Wilt Disease identification of best-durable resistant root-stock is the Screening first step. In various studies, high level of Ralstonia According to Hussain et al. (2005), bacterial wilt solanacearum resistance is reported in IIHR-7, symptoms and total number of wilted plants per CARI-1 and wild Solanum species Solanum torvum, germplasm/variety were recorded on a 0-5 scale, with LS 174, Solanum sisymbriifolium, Solanum viarum, minor modifications. Based on the percentage of Solanum mammosum, Solanum nigrum, Solanum wilted plants, accessions were categorized as highly maroniense and Solanum stramonifolium (Mochizuki resistant to highly susceptible. and Yamakawa, 1979, Gousset et al., 2005; Reddy et al., 2015; Bainsla et al., 2016). Eggplant cultivated Disease scoring scale varieties and wild Solanum species were artificially S.No. Percentage of disease Scale (0-5) inoculated with R. solanacearum inoculum Incidence (%) (Concentration ~1.0 x 108 cfu/ml). After 25-35 days 1 No wilt symptom (0%) Highly Resistant post-inoculation period, wilt incidence was observed (HR) (0) on IIHR-108, Rampur Local and Pusa Purple Long. 2 1 - 10% wilted plants Resistant (R) (1) Up to 7-8 weeks, no wilting symptoms were observed 3 11 -20% wilted plants Moderately Resistant on IIHR-7, CARI-1 and all wild Solanum species. (MR) (2) In the present study, susceptible check variety IIHR- 4 21-30% wilted plant Moderately Susceptible 108 has shown ~100% bacterial wilt incidence on (MS) (3) artificial inoculation. In the test accessions, bacterial 5 31- 40% wilted plants Susceptible (S) (4) wilt incidence ranged from 0 to 99.33 %. In general, 6 > 40% wilted plants Highly Susceptible the resistance level of eggplant cultivated varieties to (HS) (5) bacterial wilt in is limited (Bhavana and Singh, 2016). Resistant source from cultivated eggplant accessions RESULTS AND DISCUSSION will be highly useful in breeding due to their advantage Ralstonia solanacearum race 1 Biovar 3 is more in crossing, and no negative effects of fruits will hinder devastating and causes a severe problem in eggplant the breeding. Among the cultivable eggplant accessions, IIHR-7 (IC-395334) and IHR-662 cultivation mainly in hot & humid areas like India 16 J. Hortl. Sci. Vol. 14(1) : 13-19, 2019 Evaluation of eggplant for bacterial wilt resistance % 1.76 5.18 2.65 1.24 66.53 incidences 0.00 (0.00*) 0.00 (0.00*) 0.00 (0.00*) 0.00 (0.00*) 0.00 (0.00*) 0.00 4.38 (12.03*) 4.38 (12.19*) 4.50 Bacterial wilt Bacterial 99.33 (85.70*) 99.33 (27.64*) 21.54 (75.46*) 92.32 (30.93*) 26.45 6.22 4.52 4.56 0.79 1.80 8.22 4.39 0.82 0.80 1.14 3.45 9.97 Fruit yield 37.47 29.15 27.56 11.41 (t/ha) 0.29 0.19 2.69 3.80 7.52 fruit 42.67 42.67 36.72 32.84 25.67 19.84 33.17 30.83 50.60 11.54 291.67 wt. (g) wt. Average 6.79 4.93 6.97 9.64 plant 37.33 39.67 18.00 31.22 27.67 26.00 35.00 72.30 21.15 No. of No. 174.33 176.00 223.33 fruits / fruits 3.09 3.50 3.61 2.94 3.86 0.38 2.46 3.31 4.05 1.52 3.56 0.27 0.38 1.16 (cm) Fruit 10.48 10.72 diameter 5.13 1.61 3.42 5.57 5.65 2.12 9.83 0.67 0.94 2.85 9.57 (cm) Fruit 14.32 19.67 13.84 17.27 19.49 length 3.79 5.37 5.29 Plant 94.67 95.17 91.16 94.78 93.45 84.81 76.00 16.27 106.00 123.33 111.99 143.33 101.34 height 6.63 5.74 6.64 6.37 4.03 3.57 3.71 7.13 3.20 5.32 3.70 5.10 0.35 0.49 1.49 9.64 No. of No. primary branches Table Table 2. Mean performance of genotypes for bacterial wilt incidence and yield attributing traits Name of the genotype Name of IIHR-108 IIHR-7 (CARI-1) IIHR-662 IIHR-663( Long Purple Pusa Solanum gilo Solanum indicum Solanum viarum Solanum aethiopicum mammosum Solanum Solanum torvum 1 2 3 4 5 6 7 8 9 10 11 Sl. No. MEAN G. SEM SD CD (%) CV *Values in parenthesis are angular transformed values angular transformed areparenthesis in *Values 17 J. Hortl. Sci. Vol. 14(1) : 13-19, 2019 Singh et al

(CARI-1) were found to have high resistance with using Solanum torvum in grafting eggplant for no incidence of bacterial wilt. bacterial wilt and yield advantage has been proved. Among the wild species, bacterial wilt incidence The same can be used for tomato and other Solanum ranged from 0 (Solanum torvum, Solanum viarum, cultivation. Solanum torvum have graft advantages and Solanum indicum) to 26.45 % (Solanum gilo). and compatibilities with cultivated eggplant, hence, it Wild species are hardy in nature, and will have the can be effectively used in grafting programs of special advantage of strong root system for combating bacterial wilt resistant eggplant varieties/hybrids other abiotic stresses. These wild species cannot be (Ashok et al., 2017). Ashok et al. (2017) reported used in breeding programme easily because of Solanum torvum resistance with 5.7% susceptibility, negative fruit traits. However, these can be used as whereas, in our study it was found highly resistant with a root-stock for eggplant and tomato cultivation in no incidence. This variation in level of resistance may bacterial wilt prone areas. These wild species can be be due to the variability in the Solanum torvum effectively used for grafting the commercial varieties accessions or difference in Ralstonia solanacearum and hybrids of eggplant and tomato. Gopalakrishnan races. et al. (2005) identified high phenolic content, and ACKNOWLEDGEMENT some special root cortical cells in bacterial wilt resistant cultivar root system, and are thought to be Authors are thankful to the Director, Indian Institute responsible for controlling of R.solanacearum of Horticultural Research (ICAR-IIHR), Bengaluru, spreading and multiplication. Already, the success of India for providing research facilities.

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(MS Received 12 May 2018, Revised 14 May 2019, Accepted 24 June 2019)

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