液化澱粉芽孢桿菌Ba01防治馬鈴薯青枯病之探討

茄科青枯病為馬鈴薯和番茄重要細菌性病害，由青枯病菌Ralstonia solanacearum引起，除健康種苗、水旱輪作及合理化施肥等防治方法外，尚推薦液化澱粉芽孢桿菌Bacillus amyloliquefaciens PMB01。本實驗室曾發現液化澱粉芽孢桿菌Ba01對馬鈴薯瘡痂病具防治效果，但目前Ba01或PMB01對馬鈴薯青枯病之防治仍未有相關研究。此研究發現，Ba01可產生蛋白酶、澱粉酶、纖維素酶、脂酶等胞外水解酶，並具溶磷能力。與已上市之PMB01相比，Ba01具有較佳的澱粉、蛋白質、纖維素分解能力及溶磷效果。此外，Ba01在泳動、表面移行與生物膜生成的能力亦較佳。對峙培養試驗發現，Ba01與PMB01皆可以抑制馬鈴薯青枯病菌 race 1菌株RSN371與race 3菌株RSN245、RSN373、RSN439之生長。然而，混合Ba01與PMB01並未較單獨施用Ba01或PMB01獲得更佳拮抗青枯病菌之效果。此4株青枯病菌以RSN439具較強毒力，故作為後續試驗菌株。於盆栽試驗中，單獨接種RSN439罹病度達100%，而在分別澆灌Ba01或PMB01後，其罹病度顯著降至20% (P= 0.0009)與40% (P = 0.0135)。於植物的防禦反應，發現同時接種RSN439與Ba01或PMB01能增加活化氧(reactive oxygen species, ROS)於馬鈴薯葉片的累積。總結，液化澱粉芽孢桿菌Ba01可有效防治馬鈴薯青枯病，其作用機制可能與抑制病原菌、胞外分解酶的分泌、溶磷作用及誘導ROS累積有關。

關鍵字

馬鈴薯青枯病；青枯病菌；液化澱粉芽孢桿菌；生物防治

並列摘要

Solanaceae bacterial wilt caused by Ralstonia solanacearum has been an important disease of potato and tomato. Though there were no recommended bactericides, current ways to control the bacterial wilt included using bacteria-free tubers, rotation with cereals or rice, and applying Bacillus amyloliquefaciens PMB01 as a biocontrol agent. B. amyloliquefaciens Ba01 was isolated from healthy potato tuber, and demonstrated the antibacterial effect against potato common scab. In this study, we found that B. amyloliquefaciens Ba01 effectively inhibited the growth of potato bacterial wilt pathogen R. solanacearum, and it can enhance phosphate solubilization and secrete multiple extracellular enzymes, such as protease, amylase, cellulase and lipase, which could facilitate the growth of plants. Meanwhile, Ba01 exerted better swimming/swarming ability compared to PMB01, indicating it might colonize well in potato plants. We also found that Ba01 had better ability in biofilm formation than PMB01. In terms of antibacterial activity, both Ba01 and PMB01 can inhibit R. solanacearum race 1 strain RSN371, and race 3 strains RSN245, RSN373 and RSN439, but there were no synergistic effect between these two biocontrol agents. Furthermore, RSN439 has stronger virulence to potato among four R. solanacearum strains, hence we determined the disease severity of potato inoculated with R. solanacearum RSN439, and found that the disease severity significantly reduced to 20% (P= 0.0009) or 40% (P= 0.0135) after treatment with Ba01 or PMB01, respectively. As for plant defensive responses, leaves obtained from potato plants inoculated with R. solanacearum RSN439 and Ba01 or PMB01 simultaneously increased the accumulation of reactive oxygen species (ROS). In conclusion, potato bacterial wilt can be controlled by B. amyloliquefaciens Ba01 effectively due to the inhibition of R. solanacearum, the secreting of extracellular enzymes, phosphate solubilization and ROS accumulation in potato plants.