The present study was carried out to find out the resistance against bacterial wilt of the 15 cross combinations/ hybrids and six parents and screened them to find out suitable cross combination of inbred lines for developing hybrid variety with high yield and wilting resistant. The materials required and methodology are described below under the following sub-heading.
Experimental site: The present experiment was conducted at the Plant Pathology lab and glass house of Plant Pathology Section of Horticulture Research Centre (HRC) of Bangladesh Agricultural Research Institute (BAIR), Gazipur, Bangladesh during the period from September 2016-17.
Materials: Six parents and 15 cross combinations/ hybrids total 21 eggplant genotypes were used in this experiment. The parents were selected not only on the basis of high yield and other yield contributing characters and quality but also on the basis bacterial wilt resistant. Therefore, the selected parents exhibited some degrees of resistant and the main objective of the experiment was to find out how the intensity of resistance turns to the hybrids form the parents.
Seeding production: Seeds of the parents and cross combinations were sown in steel tray containing sterilized soils mixture (soil: sand: compost) in the ratio of 3: 2: 1 on 1st September 2016. Before sowing, the seeds were treated with GA3 solution (100 ppm) for 24 hours at room temperature for quick and uniform germination. Seedlings were germinated on trays with sterilized soils mixture for a period of ten days. Ten days after seed germination young seedlings were transferred to the individual plastic pot (9cm diameter) containing sterilized soil mixed with 50% organic matter (v/v).
Bacterial cultures, Preparation of inoculum and Stock cultures: Pure culture of R. solanearum strain used in this study was isolated form wilted eggplant showing typical symptoms of bacterial wilt form the research field of Plant Pathology Section, HRC, BARI. The pathogen on isolation on triphenyl tetrazolium chloride (TZC) agar medium (Kelman, 1954) yielded grayish white, fluidal colonies with light pink centers. After purification, the isolate was characterized as race1 according to pathogenicity and host range (Buddenhagen et al., 1962), and biochemical test (Hayward, 1994). The identification of the isolates was also confirmed by Cock’s Postulate. All isolates were kept as stock culture in lyophilized form and in 5 ml sterilized tap water kept at 150C. Before preparation of inoculum, the bacteria was streaked on TZC from the stock culture. The inoculum was prepared by culturing the virulent and fluidal colony of R. solanacearum on casamino acid-peptone-glucose (CPG) agar medium.
Experimental design: The plants of 15 cross combinations and six parents were arranged in the glass house following Completely Randomized Design (CRD) with three replications and ten plants were inoculated in each replication. Line to line and plant to plant distance were maintained 30 cm and 25 cm respectively. The plants were inoculated when they were approximately 35 days old (15 days after transplanting in plastic pot).
Inoculation: Suspension of R. solacearum was prepared form the stock culture which was grown on CPG for 48 hours at 300C. A sterile toothpick was used to inoculate the bacterial suspension containing approximately 2.0 ´ 108 cfu/ml (colony forming unit per milliliter) was inserted at the axil of 3rd fully expanded leaf of the plants. Plants were observed regularly for the initiation of symptoms of wilt, incubation period, 25% and 50% plant wilt (days after inoculation). After recording the 50% wilted plant data regularly, data recorded at seven days interval and final data recorded at 42 days after inoculation. The initiation of the first wilt-symptoms after inoculation and the number of wilted plants for each accession was recorded and graded on a 0-5 rating scale of Winstead and Kelnan (1952) with some modifications. The modified rating scale is;
0 = Plants did not show any symptoms of wilt; highly resistant (HR)
1 = 1 - 20% plants wilted; resistant (R)
2 = 21 - 40% plants wilted; moderately resistant (MR)
3 = 41 - 60% plants wilted; moderately susceptible (MS)
4 = 61 - 80% plants wilted; susceptible (S)
5 = more than 80% plants wilted; highly susceptible (HS)
The reaction of the hybrids and parents were categorized as highly resistant, resistant, moderately resistant, moderately susceptible to highly susceptible, depending on the percentage of wilt of the plants. The incubation period, time taken to produce 25% and 50% wilt at days after inoculation (DAI) were also recorded. Final data on wilting (recorded at 42 DAI) were converted into percent wilt and the reaction of the genotypes was graded on a 0-5 rating scale according to Winstead and Kelman (1952).