由Burkholderia gladioli引起的五種蘭花的細菌性葉斑病

Abstract

從台中、彰化、嘉義、高雄及屏東等地區的蝴蝶蘭、石斛蘭、文心蘭、萬代蘭及腎藥蘭等多處蘭園植株發現在葉片呈稍為軟化，但表皮仍硬的黑褐化且具黃暈的斑點，之後斑蔓延至全葉呈黑褐化，在花蕾上呈現小的斑點及消蕾的徵狀，經柯霍氏法則確認造成此徵狀的病原為細菌。該細菌經生理生化測定為革蘭氏陰性，好氣菌，在KB培養基上不會產生螢光色素，在NA上為白色菌落，培養後產生可溶性黃色色素，生理生化測試結果顯示該病原細菌與Burkholderia gladioli的特性相近，以Biolog鑑定該病原菌顯示為B. gladioli。再以對B.gladioli 具專一性的引子組 CMG-16-1/G-16-2進行PCR鑑定，供試菌株可產生468 bp的專一性條帶，應用另一組專一性引子組BGRPF3/BGRPR3鑑定，供試菌株可產生 350 bp的專一性條帶，顯示供試菌株為B. gladioli。再以f8-27/r1510引子組增幅該病原菌的16S rDNA片段，進行定序與鑑定，確認病原菌與GeneBank 中的 B. gladioli strain ATCC 10248 chromosome 1(accession number CP009323)及B. gladioli strain BgHL-01 16S ribosomal RNA gene (accession number JX566503)，相似度達99-100％，綜合各項鑑定試驗確認引起五種蘭花的細菌性葉斑病其病原菌為 Burkholderia gladioli。測試市售10種殺菌劑在一般推薦濃度下對供試病原菌生長的抑制效果，顯示除歐索林酸外，其餘的供試藥劑均無抑制效果。 Bacterial isolates were obtained from various orchid plants collected from Taichung, Changhua, Chiayi and Kaohsiung. Leaf spots appeared water-soaked, macerated, but had intact epidermal layers. In severe cases, the flower petals exhibited brown spots and failed to bloom. These symptoms were observed on Phalaenopsis, Dendrobium, Oncidium, Vanda, and Renanthera spp. and other hybrids. Bacterial isolates were identified based on physiological and biochemical tests and Biolog identification. PCR identification was also conducted using Burkholderia gladioli-specific primer pairs CMG-16-1/G-16-2 and BGRPF3/BGRPR3, which produced amplicons that are 468 bp and 350 bp in length, respectively. Further sequencing of 16S rDNA fragments using primer pair f8-27/r1510 confirmed that the bacterium was 99 to 100% identical with the 16S ribosomal RNA genes of Burkholderia gladioli ATCC 10248 chromosome 1 (accession number CP009323) and BgHL-01 (accession number JX566503). Among 10 fungicides screened in vitro, only oxolinic acid showed better control efficacy.