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題 名 | 種帶植物病原細菌之偵測=Detection of Seedborne Plant Pathogenic Bacteria |
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作 者 | 黃㯖昌; | 書刊名 | 植物保護學會會刊 |
卷 期 | 39:1 1997.03[民86.03] |
頁 次 | 頁13-21 |
分類號 | 433.8 |
關鍵詞 | 種帶植物病原細菌; 偵測; 種子檢疫; Seedborne plant pathogenic bacteria; Detection; Seed quarantine; |
語 文 | 中文(Chinese) |
中文摘要 | 超過45種植物病原細菌證實可經種子傳播,一般細菌最常污染種子表面,引起維束管系統性病害者,則可侵入種皮或種子內部組織。附著於種子表面的細菌存活期限較短,由數月至1-2年;存在於種子組織內者則可以殘存甚久,例如Curtobacterium flaccumfaciens pv. flaccumfaciens(syn. Coryebacterium flaccumfaciens)可在菜豆種子組織內存活達24年。種帶細菌是重要的病害第一次感染源,因此,先進國家對於種子的健康極為重視,以十字花科蔬菜黑腐病(Xanthomonas campestris pv. campestris)及菜豆黃暈葉枯病(Pseudomonas syringae pv. phaseolicola)為例, 其用於生產種苗的種子,帶菌容許率均為0。美國並特別成立一委員會名為NC-135,專門統籌十字花科蔬菜種帶病菌的防治研究與技術推廣。種子檢疫是確保種子健康的首要措施,精確、有效率的病菌偵測與鑑定技術則是種子檢疫的基石,優良偵測方法須符合專一、敏感、簡便、速效及低成本等條件。最早被用於偵測種帶細菌者為試種法(growing-on),該法可顯示種子傳病的比率,但其結果容易受環境及生物因子干擾,而且曠日廢時,所需空間大。較可靠而實用的偵測技術,通常先以浸清、洗滌或研磨自種子萃取病菌,而後將萃取液展佈於選擇性培養基,俟菌落長出後,依菌落形態配合寄主接種或噬菌體、抗體、基因探針測試,以確認病菌。近幾年來,聚合脢鏈反應(PCR)廣泛被應用於基因的偵測與鑑定,當引子(primers)選定得當時,該方法之靈敏度與專一性均頗高,符合種子病菌偵測技術的需求。Schaad氏先以一般性培養基回收菜豆種帶細菌,而後以選自植物毒質(phaseolotoxin)基因的二對引子,直接就自培養基上洗出的細菌進行二段聚合脢鏈反應,由放大所得的特定基因片段判定 P. syringae pv. phaseolicola是否存在,可測出每毫升種子浸出液含2-3個細胞的菌量。 Audy等人則直接快速萃取種帶病菌的核酸後,以二組特定引子進行聚合脢鏈反應,同步偵測P.syringae pv. phaseolicola及菜豆葉枯病菌(Xanthomonas campesrtris pv. phaseoli),可測出0.01%之帶菌種子。 |
英文摘要 | Over 45 plant pathogenic bacteria are known to be seed-transmitted. The bacteria are often carried on the surface of the seed, whereas those which cause vascular or systemic infections might invade seed coat or other tissue of the seed. Bacteria borne on the surface of the seed may survive for a limited period of time ranging from several months to 1-2 years, whereas bacteria existing within seed tissues usually show a surprisingly prolonged longevity.For instance, Curtobacterium flaccumfaciens pv. flaccumfaciens (Syn. Corynebacterium flaccumfaciens) was found to have survived for 24 years in Phaseolus bean seed kept under laboratory conditions. Since seedborne bacteria serve as primary inocula of diseases, seed sanitation is a great concern in many advanced countries. For example, the tolerance levels for seedborne Xanthomonas campestris pv. campestris, the causal agent of black rot of cruicf ers, and Pseudomonas syringae pv. phaseolicola, the causal agent of halo blight of bean, have been both set at zero for the seeds for see or transplant production. In addition, a regional Cooperative States Research Service Committee NC-135 was organized in the U.S to entirely devote to the research of seedborne bacteria and the extension of seed sanitation.Seed assays based on precise and efficient techniques for detecting seedborne bacteria are of great importance in terms of the production of healthy seeds. The requirements for a good detection technique are high specificity and sensitivity, simplicity, speed, and low cost. Growing-on is the method which was first adopted for this purpose. The major advantage of growing-on is that it indicates the probability of seed-transimitted diseases. However, the results obtained from this method are often interfered by other biotic and environmental factors. In addition, it is time-comsuming and requires a lot of space.Direct isolation that includes extracting the bacterium by washing, soaking, or grinding and then plating the extract onto solid selective medium is one of the most reliable and practical methods. The resultant colonies grown on the medium can be further identified by host inoculations, phage plaques, serological methods, or DNA hybridization probes. Polymerase chain reaction(PCR), a newly developed novel technique, has been successfully applied to detect and identify specific nucleic acid in recent years. When both primers and the procedure are designed properly, it demonstrates great sensitivity and specificity, that are in agreement with the requirement of a good detection technique, in detecting seedborne bacteria. Schaad et al. designed two pairs of primers based on the phaseolotoxin gene of P. syringae pv. phaseolicola and conducted a two consecutive rounds of PCR to amplify a specific DNA fragment in bacterial washings from general agar media on which seedborne bacteria were grown. They successfully detected 2-3 cfu/ml of P. syringae pv. phaseolicola in original washings of bean seeds. Audy et al. designed 2 pairs of primers specific to P. syringae pv. phaseolicola and Xanthomonas campesrtris pv. phaseoli (common blight of bean)respectivel;y, and used to carry out a PCR assay following a quick extraction of bacterial genomic DNA. Both of the pathogens were concurrently detected in batches containing as few as 1 infected seed in 10,000 seeds. |
本系統中英文摘要資訊取自各篇刊載內容。