黃龍病菌於柑橘寄主體內之移動與影響亞洲柑橘木蝨 (半翅目：扁木蝨科) 獲菌因子之探討

Abstract

由韌皮部侷限細菌所引起的柑橘黃龍病 (Citrus huanglongbing, HLB) 是影響柑橘生產的重要病害，危害世界各大柑橘產區，而年產量高達50萬公噸以上的臺灣亦受嚴重波及。根據2017年田間調查的結果顯示臺灣約有31.9%之罹病率，且各品種皆會染病，造成果實的品質及產量大打折扣。目前已知病害的感染途徑主要藉由嫁接及媒介昆蟲進行傳播，而自1973年起積極推廣健康柑橘種苗概念之下，柑橘園中的亞洲柑橘木蝨 (Diaphorina citri Kuwayama) 似乎成為病害擴散的一大隱憂，對於媒介木蝨傳病生態學的相關研究，則可提供黃龍病防治策略的重要參考。本研究藉由分析不同生長期的柑橘品種其新梢帶菌的情形，以推估黃龍病菌自病徵明顯的老熟組織移動至幼嫩葉部所需的時間，結果發現柳橙在新梢生長一週後即可測得黃龍病菌，往後隨週數的進展菌量呈微幅增長；椪柑及桶柑則在兩週過後病原菌便開始快速增殖，到第四週時菌量達所有品種中的最高，而黃龍病菌在白金柚中的移動最為緩慢，至三、四週後才可在新梢中檢測到少量存在。病原菌移動至新梢後，因木蝨對嫩葉處具有強烈的偏好，進而提供受吸引前來的木蝨獲菌的機會，文中亦針對可能影響木蝨獲菌的因子進行探討，在調查不同齡期若蟲的獲菌情形中，發現自二齡起木蝨即具獲菌的能力，又以四齡及五齡若蟲的帶菌量最高，五齡若蟲的帶菌量約與初獲菌的二齡若蟲相差近130倍；取食病株的時間長短亦會造成獲菌差異，在14天時初有30%的帶菌蟲比例，到28天時則升高至80%，說明獲菌情形可能隨時間的延長而提升；而不同的柑橘品種及其罹病情形也是影響木蝨獲菌的可能因子，以罹病三個月後的柳橙最早有木蝨獲菌的記錄，椪柑及文旦亦在四、五個月時相繼出現木蝨獲菌的情形，而任一品種皆發生在完整病徵出現以前。進入木蝨體內的黃龍病菌隨開放系統進行循環，以持續取食月橘屬 (Murraya spp.) 植株的試驗中觀察病原菌於媒介昆蟲體內的增殖效率，而長至四週後仍能檢測到帶菌情形，證實病原於蟲體內的保存效力，一旦經有效獲菌後，則可能終生成為潛在感染媒介，在田間造成黃龍病傳播的威脅。

Citrus huanglongbing (HLB) is a destructive disease among citrus industry caused by a phloem-limited bacteria (Candidatus Liberibacter asiaticus, CLas), and it happens in citrus production areas worldwide. With over than 500,000 tons of annual production, Taiwan also suffers from this serious disease. According to the HLB field survey in 2017, there was 31.9% infection in Taiwan. Because all citrus cultivars are susceptible to HLB, it caused great damage on fruit quality and production. As known, the spread of HLB in the field is mainly due to the grafting of infectious scions and insect vector transmission. However, with the help of citrus healthy budwood program which was firstly promoted in 1973, vector psyllids (Diaphorina citri Kuwayama) seems to play an important role in spreading the disease. Researches on transmission ecology of HLB by the psyllid may contribute to the good solution for HLB management. By analyzing new shoot infection status in different weeks, we attempted to find out the movement of CLas from symptomatic old tissues to young leaves among different cultivars, and the results showed that movement in Liucheng cultivar requiring the least time. CLas reached new shoots in only 1 week, and the copy numbers increased slightly. Same movement tendency could be observed in Mandarins (Ponkan and Tankan cultivars), CLas copy numbers rapidly increased after 2 weeks and reach amount of 10,000 copies in week 4, which is also the highest amount in the study. The movement of CLas was much slower in Oroblanco, high ratio of CLas-infected new shoots could only be observed after 3 weeks. Because of the high preference of psyllids to immature foliar tissues, CLas moving to new shoots may offer a good chance for pathogen acquisition. In this study, we also investigated the factors that might influence CLas acquisition. Among different nymphal stages, CLas was firstly detected in the 2nd nymphal stage, and got the highest average copy numbers per psyllid in the 4th and 5th nymphal stages, which had about 130 times in difference. Also, acquisition access period on HLB-infected plant seemed to be another influence factors, percentages of CLas-infected psyllid raised from 30% to 80% when prolonging the period from 14 to 28 days. Moreover, other factors like plant infection status among different cultivars could lead to variation in CLas acquisition. After CLas inoculation by grafting, pathogen acquisition by D. citri firstly occurred after 3 months on infectious Liucheng, 4 and 5 months on Ponkan and Wentan, respectively; while they all happened during asymptomatic phase of infection. As CLas acquired by the psyllids, they followed the open system and circulated in insect bodies. Continuous feeding tests on Murraya spp. made a good way to observe CLas proliferation in D. citri. The results showed that CLas could be still detected until 4 weeks, and it proved the ability of CLas retention in insect vectors. Indicating that once D. citri acquired CLas via feeding, they could probably become a persistent infectious vector, and post the threats to HLB spreading in the fields.