Introducing Bacillus licheniformis as the causal agent of pistachio dieback in Iran

During 2004 and 2006 growing seasons some pistachio trees in Kerman province of Iran showed dieback symptoms. Initial symptoms were observed at the beginning of the growing season and they developed during two weeks after green tip stage, as shoot tips turned black and dieback occurred. During the growing season, the symptoms developed and vessel destruction was observed. If these stems were not pruned during winter, dieback developed in the spring. During the growing season affected pistachio samples were collected and surface disinfected with 0.01% mercury chloride. Pieces of affected vessels were grown on nutrient agar (NA) and incubated at 25°C for three-to-four days. Bacterial colonies with Bacillus characteristics were isolated and 15 representative strains were selected for further characterisation. The pathogenicity of selected strains was verified on 2–3 year-old pistachio seedlings using injection of bacterial suspension (10⁷ cfu p/ml) and control plants inoculated with distilled water; vessel destruction developed after 20 days, and bacterial causal agent was isolated from seedlings. No symptoms were observed in control plants. The strains were Gram positive, motile, with central spores and caused a hypersensitivity reaction (HR) on tobacco and geranium; they were positive for anaerobic growth, nitrate reduction, utilisation of citrate, VP test, urease, catalase, growth at pH 5.7, 7% NaCl and 45°C, acid production from arabinose, xylose, glucose and mannitol, and anaerobic fermentation of glucose. They could hydrolyze starch, aesculin, Tween 80 and gelatin but indole production was negative. Based on the characteristics of the isolated strains, they were identified as Bacillus licheniformis. This is the first report of Bacillus licheniformis as a causal agent of pistachio dieback.