臺灣褐根病菌無性繁殖系之判定與族群遺傳分析

Clone identification and population genetic analysis of Phellinus noxius collected from Taiwan

黃郁晴(Yu-Ching Huang)

指導教授 ： 劉瑞芬

共同指導教授 ： 張東柱(Tun-Tschu Chang)

國立臺灣大學/生物資源暨農學院/植物病理與微生物學研究所/碩士(2012年)

https://doi.org/10.6342/NTU.2012.00899

摘要

褐根病菌 (Phellinus noxius) 寄主範圍相當廣泛，可感染樟樹、榕樹、臺灣欒樹、鳳凰木等多種樹木，是近年來極受重視的病原菌。植物罹患褐根病後，初期不易看出明顯病徵，但感病後期常因根部木材逐漸腐朽，養分和水分的運輸功能也嚴重受損，導致罹病樹木萎凋、死亡或風倒，不僅破壞景觀，更可能造成公共危險。爲了擬定有效的褐根病防治策略，將需要瞭解更多其流行病學。本研究開發ISSR (Inter simple sequence repeats)和SSR (simple sequence repeats)分子標誌，分析自臺灣北部多個樣區所收集的褐根病菌菌株和擔孢子的基因型(genotype)，再根據分析結果判別其是否為相同無性繁殖系(clone)及親緣關係遠近。評估結果顯示，共計有9個ISSR引子和17個SSR分子標誌可以有效區分不同子實體來源的擔孢子菌株，同時亦可反映不同擔孢子之間的遺傳變異；SSR基因型分析亦顯示擔孢子菌株為單倍體。此外，在菌絲-斷生孢子的生長繼代過程中，菌株基因型容易發生變異。基因分型結果也顯示，自臺北市和平高中和螢橋國中約4公頃 (40000 m2) 校地的28棵罹病樹木，所分離的褐根病菌株分別屬於11個無性繁殖系，且除了部分栽植位置相近的罹病樹木之外，自其他樹木分離的菌株皆屬於不同的無性繁殖系。此外，褐根病菌分離株之菌絲大多為同質異核(heterokaryon)。本研究所開發的ISSR及SSR分子標誌可應用於探討褐根病菌的遺傳變異，不僅讓我們初步瞭解臺灣褐根病菌的遺傳特性，也以分子證據證實褐根病可藉由根接觸而傳染。但關於臺灣田間褐根病菌族群主要是以何種方式繁殖，以及是否存在高致病力之強勢小種，仍待分析更大量的樣本方能確定。

關鍵字

簡單重複性序列區間 ； 微衛星 ； 無性繁殖系 ； 褐根病 ； 簡單重複性序列 ； 族群遺傳

並列摘要

Since 1990s, brown root rot caused by Phellinus noxius has become a major tree disease in Taiwan. This fungal pathogen has a very wide host range, including Cinnamomum camphora、Ficus microcarpa、Koelreuteria henryi and Delonix regia. When infecting the trees, it can not only destroy bark and wood structure, but also seriously impair nutrient transportation and water uptake. In the early stage of infection, the infected trees may not show any obvious symptom, but as the disease develops, the tree may decline and eventually fall down or die within few months, thereby resulting in considerable economic loss and causing public danger. In order to investigate the disease dissemination pathway and epidemic dynamics of P. noxius, we have developed genotyping methods which were based on ISSR (inter-simple sequence repeat) or SSR (simple sequence repeat). These methods are useful not only for the distinction of basidiospores from different fruiting bodies, but also detection of genetic recombination in the process of sexual reproduction. The SSR prof n the process of sexual reproduction iles also verified the haploid nature of the basidiospore. Furthermore, single-arthrospore isolation might cause certain degree of genetic segregation. In the field survey, P. noxius isolates were obtained from 28 infected trees in two campuses (c.a. 4 hectares) of Taipei, and analyzed for their genotypes. The results revealed the presence of 11 clonal lineages, each occupying a distinct infection zone. As revealed by SSR profiles, mycelia of the field isolates were heterokaryon in most cases. Furthermore, for fungal isolates collected from infected trees nearby each other, most of them tend to display similar genotypes, indicating that root contact may serve as an important way for pathogen dissemination. However, to address important questions regarding epidemiology of P. noxius, including its major reproductive way and possible existence of a predominant virulent strain in the field, it is necessary to analyze more fungal isolates.

並列關鍵字

ISSR ； microsatellite ； clonality ； brown root rot ； simple sequence repeats ； population genetics ； Phellinus noxius

參考文獻

安寶貞、蔡志濃、張東柱、王姻婷。2005。台灣果樹及木本觀賞植物立枯型真菌性病害之調查與分布。植物病理學會刊 14:203-210。

蔡志濃。2008。褐根病菌之生物特性及分子診斷技術。國立中興大學植物病理學研究所博士論文。臺中。133頁。

Anderson, J. B., and Kohn, L. M. 1995. Clonality in soilborne, plant pathogenic fungi. Ann. Rev. Phyto. 33:369-391.

Ann, P. J., Chang, T. T., and Ko, W. H. 2002. Phellinus noxius brown root rot of fruit and ornamental trees in Taiwan . Plant Dis. 86:820-826.

Ann, P. J., Lee, H. L., and Huang, T. C. 1999. Brown root rot of 10 species of fruit trees caused by Phellinus noxius in Taiwan. Plant Dis. 83:746-750.