Isolation And Identification Of Leaf Brown Spot Caused Pathogens From Populus Euphractica Forest Of Tarim River And Identification Of A Novel Species

1 Populus euphratica Oliv. were the most original and oldest trees in the genus Populus. They had the quality of heat-resistant﹑anti-cold﹑anti-wind and sand﹑anti-saline﹑anti-droughty﹑anti-barren and so on. Thus, Populus euphratica played a very important role in maintaining ecosystem function in arid and semi-arid regions. Populus euphratica forests in Tarim River Valley were the largest natural Populus euphratica forests in the world. Their area was nearly 300,000 hm2, accounting for 90% of total Populus euphratica forests'area in China and more than 55% worldwide. They were extremely rare natural resource. The leaf brown spot disease was very prevalent in Populus euphratica forests. It began to appear light brown spot in August, the edge of spot was not regular, later the area of spot was gradually increscent and the spot colour turned into dark brown or dark. The leaf brown spot disease caused the premature and defoliation in Populus euphratica forests. Moreover, they would bring immense harm to Populus euphratica forests'natural growth and nutrient accumulation, finally pricking up P. euphratica forests to downfall.Now there are many researches on Populus pathogens, but lacking of researching on Populus euphratica forests. This paper studied on P.euphratica'leaf brown spot disease in Tarim River Valley and their isolation and screening of pathogens. Ascertain pathogens by inoculation test. The fungi were identified by morphological, 28S rDNA D1/D2 sequence homology, phylogenetic analysis and pathogenicity. The bacteria were characterized taxonomicolly by morphological, physiological and biochemical characteristics, 16S rDNA sequence homology, phylogenetic analysis and pathogenicity.The isolation of a total of 33 species, 32 different populations were identified and characterized, 24 isolates were identified as bacteria, including 16 isolates of genus Bacillus, 2 isolates of Kocuria, 2 isolates of Arthrobacter, 1 isolate of Pseudomonas, Micrococcus, Brevibacterium and Oceanobacillus, respectively. 8 isolates were identified as fungi, including 4 isolates of genus Penicillium, 2 isolates of Aspergillus, 1 isolate of Alternaria, 1 isolate of Quambalaria. The inoculation test indicated that strain X-9 was the main pathogen which caused the leaf brown spot disease of Populus euphratica forests. According to its morphological, 28S rDNA D1/D2 sequence homology, phylogenetic analysis, strain X-9 belonged to genus Quambalaria. Comparative 28S rDNA D1/D2 sequence analyses, the closest relative to strain X-9 was Quambalaria simpsonii GQ303321.1 (99% similarity), so strain X-9 was named as Quambalaria simpsonii X-9.2 A latency novel bacterium XH4 was characterized taxonomically by using a polyphasic approach, including morphological, physiological and biochemical characteristics, chemotaxonomic characteristics, G+C content of the genomic DNA, 16S rDNA sequence homology, phylogenetic analysis. Cells were Gram-negative, strictly aerobic, non-spore-forming, non-motile and rod-shaped (0.6–1.0μm wide and 1.0–1.5μm long). Colonies grown on LB were yellow, circular and convex with entire margins. Growth occurs between 20 and 45℃; the optimum temperature for growth is 30-37℃. The pH range for growth is 6.0-9.0, with an optimum between pH 6.0 and 8.0. Growth occurs in the presence of 0-3% (w/v) NaCl. Positive for catalase and oxidase. Comparative 16S rDNA sequence analyses, the closest relative to strain XH4 was Sphingobacterium mizutaii ATCC 33299T, the similarity was 96.02%. The GenBank number was FJ859899. MK-7 was the predominant menaquinone, and major fatty acids were iso-C15:0 (34.02%), summed feature 3 (C16:1ω7c and/or C16:1ω6c, 20.73%) and iso-C17:0 3-OH (14.74%). The G+C content of the DNA was 41.2mol%. On the basis of the data obtained in this polyphasic taxonomic study, strain XH4T should be placed in the genus Sphingobacterium as the type strain of a novel species, for which the name Sphingobacterium hotanense sp. nov. was proposed. The collection number in CCTCC and NRRL were CCTCC AB 209007 and NRRL B-59204, respectively.