A. Survey of leaf blight of litchi in Northern region of Bangladesh Survey was conducted from 13 October 2013 to 10 April, 2014 in the Northern region of Bangladesh. Altogether seven varieties of litchi viz., Bedana, Bombai, Haria Bombai, Madrajie, China-2, Chaina-3 and Kathali Lichu were surveyed. During the survey 30 saplings of litchi for each variety were selected randomly in each location. Number of saplings and number of healthy and diseased saplings were recorded from the selected nurseries. Each of the selected saplings was observed carefully and symptoms of leaf blight disease were recorded. The disease incidence and severity were evaluated following the formula of Rai and Mamatha, and Johnston, respectively.
B. Antibiotic sensitivity test of Pseudomonas syringae pv. syringae Diseased leaves were collected from the nurseries as shown in Fig. 1 and kept in sterile polythene bag and transported to the laboratory. Tissue planting (leaf cutting) method was used for collection of bacteria. The cut infected portions of the leaf were washed and cleaned in sterile distilled water, and plated on nutrient agar (NA). Plates were incubated at 280C for 2 days. Cream or off white colored colony of bacteria was appeared after incubation on NA medium. The cream color on the NA media was the colony of P. syringae pv. syringae. After isolation bacterial isolates were purified by streaking a single colony of each isolate by sub-culturing on nutrient agar medium as described. The isolates of bacteria were preserved in 10% skim milk, kept at -200C in refrigerator for antibiotic sensitivity test.
Sensitivity of P. syringae pv. syringae isolates to different antibiotics was determined in-vitro by employing Kirby-Bauer disc diffusion method. The procedure involved measuring the diameter of the zone of inhibition that results from diffusion of the agent into the medium surrounding the disc. Antimicrobial discs of 0.05% were used for the test. The used antibiotics were Gentamicin, Erythromycin and Doxycycline. With a sterile pipette a drop of test culture of bacteria was poured on NA plate. Sterile glass spreader was used to spread the culture homogenously on the medium. The plate was allowed to sit at room temperature at least 3 to 5 minutes, but no more than 15 minutes, for the surface of the agar plate to dry before proceeding to the next step. Three antibiotic discs were placed aseptically onto the surface of the inoculated plates applying appropriate special arrangement with the help of a sterile forceps. The plates were incubated at 35 0C ± 2 0C in the incubator. Following incubation, after 24 hrs the zone sizes were measured to the nearest millimeter using a ruler, include the diameter of the disc in the measurement. The plates were examined and the diameter of each zone of complete inhibition was measured in mm. At the time of measuring zone diameters, always rounded up to the next millimeter. All measurements were made with the unaided eye while viewing the back of illuminated with reflected light. The plate was viewed using a direct, vertical line of sight to avoid any parallax that may result in misreading. The zone size was recorded on the recording sheet. Growth up to the edge of the disc was reported as a zone of 0 mm. Distinct, discrete colonies within an obvious zone of inhibition were not considered. Data that were recorded depending on the areas of zone diameter were arranged according to maximum to minimum diameter in mm. For each antibiotic indicating on the recording sheet the zone size was reported as sensitive (S), intermediate (I), or resistant (R).
C. Management of bacterial leaf blight of litchi The experiment was carried out from July 2013 to June 2014 in the net house, Department of Plant Pathology, Bangladesh Agricultural University, Mymensingh, Bangladesh. Under this program, the saplings of three years old grown in pots were used for management of leaf blight of litchi. For the control of Bacterial Leaf Blight (BLB) disease of litchi (Variety: China-3) six different treatments were employed on litchi saplings. The treatments were: (i) T1 = Gentamicin applied as foliar spray @ 0.05%, (ii) T2 = Erithromycin applied as foliar spray @ 0.05%, (iii) T3 = Doxycycline applied as foliar spray @ 0.05%, (iv) T4 = Copper sulphate applied as foliar spray @ 0.05%, (v) T5 = BAU-Biofungicide applied as foliar spray @ 2% and (vi) T6 = Untreated control. The experiment was laid out in Completely Randomized Design (CRD) with three replications. The data were recorded on a) Height of the saplings (cm), b) Total number of branch/sapling, c) Total number of leaves/sapling, d) Number of diseased leaves/sapling and e) Percent leaf area diseased/sapling. The incidence and severity of bacterial leaf blight of litchi was assayed following the formula of Rai and Mamatha (2005), and Johnston (2000), respectively.