Bacillus amyloliquefaciens PMB05粉劑的種子處理於甘藍黑腐病之防治

Seed treatment of Bacillus amyloliquefaciens PMB05 powder for control black rot disease of cabbage

由Xanthomonas campestris pv. campestris（Xcc）所引起的甘藍黑腐病是一種種子傳播性病害，會導致甘藍產量的嚴重損失。因此，減少種子上初始接種原的數量是控制此病害的一種有效策略。由之前的研究結果表明，在汙染有Xcc的種子上處理Bacillus amyloliquefaciens PMB05的細菌懸浮液可以有效減少病害的發生。然而，這樣的處理仍然需要種子浸泡與後續乾燥的過程。在此限制下，本研究擬進一步探討是否利用發酵液結合噴霧乾燥的技術所製備完成的粉劑來處理種子後仍能有效減少黑腐病的發生。為了以麥芽糊精作為載體，本研究首先證明以麥芽糊精直接處理種子並不會抑制種子發芽。然後，將B. amyloliquefaciens PMB05發酵液和麥芽糊精的混合後進行噴霧乾燥，顯示在三種不同的入口溫度處理下，內生孢子數均達到10^9 CFU/g以上。進一步將粉劑處理於甘藍種子上，結果顯示入口溫度為140 °C與160 °C時，粉劑可以增加種子的發芽率。此外，本研究也推測來自於入口溫度120 °C的粉劑對種子發芽率的抑制可能是由於麥芽糊精在噴霧乾燥過程中所產生抑制物的影響。在後續以入口溫度160 °C處理之PMB05粉劑的分析結果顯示，PMB05在此狀態下於26 °C和4 °C下可以保持在90%以上存活率達16週以上。最重要的是，將受到Xcc污染的種子用1%的PMB05粉劑混合處理後，甘藍幼苗黑腐病的發病率與罹病度均顯著降低。此外，結果指出粉劑處理種子所獲得的葉片在利用細菌性誘引物誘導下，可觸發較強的植物免疫反應訊號，推測此種子處理可使甘藍得到更強的保護效果。綜上所述，本研究提供了噴霧乾燥之較佳條件並證明應用B. amyloliquefaciens PMB05粉劑來處理甘藍種子確實為防治甘藍黑腐病簡單又有效的方法。

Cabbage black rot disease (CBRD) caused by Xanthomonas campestris pv. campestris (Xcc) is a seedborne disease that causes severe yield losses. Therefore, reducing the pathogen on seed could be a possible strategy to control CBRD. Our previous study revealed that CBRD could be controlled by bacterial suspensions of Bacillus amyloliquefaciens PMB05 in Xcc-infested seeds. However, such treatments still require a process of seed soaking and subsequent drying. Under this limitation, this study intends to further explore whether using powder prepared by fermentation broths to treat infested seeds can still effectively reduce the occurrence of CBRD. In order to use maltodextrin as a carrier, it was first proved that seed germination was not inhibited by treatment with maltodextrin directly. Then, the mixture of B. amyloliquefaciens PMB05 fermentation broth and maltodextrin was spray-dried; the number of endospores reached more than 10^9 CFU/mL under the treatment with three distinct inlet temperatures. The 0.1% powder was further mixed with cabbage seeds, results showed that seed germination ratio was increased by the powders from the inlet temperature at 140 ℃ and 160 ℃. In addition, we also found that the inhibition of seed germination by the powder from the inlet temperature of 120 ℃ might be due to the compounds produced from maltodextrin during the spray drying process. Furthermore, the PMB05 powder produced at 160 ℃ of inlet temperature showed more than 90% of the survival rate for over 16 weeks at room temperature and 4 ℃ in this state. Most importantly, the disease incidence and disease severity of CBRD on seedlings were reduced significantly when the Xcc-infested seeds were coated with 0.1% of the PMB05 powder. In addition, we also found that the leaves obtained from the seed treatment can trigger stronger plant immune response signals upon the induction of a bacterial elicitor, suggesting that the seed treatment might give cabbage a more comprehensive protective effect. Combining the above results, we concluded that coating seeds with PMB05 powder is a convenient and effective way to control CBRD.