Linaldeddu B T, Franceschini A, Luque J, Phillips A J L
Dipartimento di Protezione delle Piante, Sezione di Patologia Vegetale, Università degli Studi di Sassari, Via E. De Nicola 9, I-07100 Sassari, Italy.
Departamento de Protecció Vegetal, Institut de Recerca i Tecnologia Agroalimentàries, Centre de Cabrils, Ctra. de Cabrils s.n., E-08348 Cabrils, Barcelona, Spain.
Plant Dis. 2007 Mar;91(3):324. doi: 10.1094/PDIS-91-3-0324A.
A survey was carried out in the spring of 2003 to study the fungi associated with declining trees in a cork oak (Quercus suber L.) forest located in Sassari Province, Sardinia, Italy (40°52'N, 9°01'E) at an altitude of 150 m (above sea level). Several isolates obtained from live twigs and branches showing sunken necrotic bark lesions were identified as Fusicoccum parvum Pennycook & Samuels (teleomorph Botryosphaeria parva Pennycook & Samuels). Neither pycnidia nor ascomata were observed on the symptomatic samples collected. On potato dextrose agar (PDA) at 25°C, the isolates developed an aerial and compact mycelium, initially white but becoming gray after 4 to 6 days, and produced pycnidia after 1 month on sterile cork oak twigs placed on the surface of PDA. Conidia from culture were hyaline, ellipsoidal to fusiform, externally smooth, thin walled, nonseptate, 12 to 19 (15.5) × 5.5 to 8.5 (6.5) μm, with length/width ratio of 2.4 ± 0.1 (mean ± standard error). Identity was confirmed by analysis of the nucleotide sequences of the internal transcribed spacer (ITS) from the rRNA repeat and the translation elongation factor 1-alpha (EF1-α), as done elsewhere (1,4). BLAST searches at GenBank showed a high identity with reference sequences (ITS: >99%; EF1-α: 100%). Representative sequences of both regions were deposited at GenBank (ITS: Accession No. DQ487157; EF1-α: Accession No. DQ487158). Pathogenicity tests were carried out on seven 2-year-old cork oak seedlings maintained in a greenhouse at 14 to 26°C with the B. parva strain CBS 119937 obtained in this study. A mycelial plug (3 to 4 mm) taken from the margin of an actively growing colony on PDA was put in a shallow wound made by a scalpel on the basal part of the stem of each seedling. Sterile PDA plugs were placed into similar wounds on three control seedlings. The inoculation points were wrapped in Parafilm to retain moisture for 1 week. After 4 weeks, all seedlings inoculated with B. parva died and showed a collapse of the stem cortical tissues associated with dark brown discolorations and vascular necrosis measuring 10.9 ± 0.4 cm. No symptoms were visible in the control seedlings. The pathogen was reisolated from all the inoculated seedlings, thus fulfilling Koch's postulates. The results confirm the virulence of this fungus and point to its possible involvement in the aetiology of cork oak decline. B. parva is a cosmopolitan, plurivorous pathogen causing disease in several hosts of economic importance, such as grapevine (3), kiwi (2), and Eucalyptus spp. trees (1). To our knowledge, this is the first report of B. parva causing canker disease on cork oak trees. References: (1) A. Gezahgne et al. S. Afr. J. Bot. 70:241, 2004. (2) S. R. Pennycook and G. J. Samuels. Mycotaxon 24:445, 1985. (3) A. J. L. Phillips. Phytopathol. Mediterr. 41:3, 2002. (4) B. Slippers et al. Mycologia 96:83, 2004.
2003年春季，在意大利撒丁岛萨萨里省（北纬40°52′，东经9°01′）海拔150米（高于海平面）的一片栓皮栎（Quercus suber L.）林中开展了一项调查，以研究与树木衰退相关的真菌。从呈现树皮凹陷坏死病斑的活嫩枝和枝条上分离得到的多个菌株被鉴定为细小色二孢（Fusicoccum parvum Pennycook & Samuels，有性型为小葡萄座腔菌Botryosphaeria parva Pennycook & Samuels）。在采集的有症状样本上未观察到分生孢子器和子囊盘。在25°C的马铃薯葡萄糖琼脂（PDA）培养基上，这些菌株形成了气生且紧密的菌丝体，最初为白色，但4至6天后变为灰色，并在置于PDA表面的无菌栓皮栎嫩枝上培养1个月后产生了分生孢子器。培养产生的分生孢子无色，椭圆形至梭形，表面光滑，薄壁，无隔膜，大小为12至19（15.5）×5.5至8.5（6.5）μm，长宽比为2.4±0.1（平均值±标准误差）。如在其他地方所做的那样（1,4），通过分析核糖体RNA重复序列的内部转录间隔区（ITS）和翻译延伸因子1-α（EF1-α）的核苷酸序列来确认其身份。在GenBank上进行的BLAST搜索显示与参考序列具有高度同源性（ITS：>99%；EF1-α：100%）。这两个区域的代表性序列已存入GenBank（ITS：登录号DQ487157；EF1-α：登录号DQ487158）。用本研究中获得的小葡萄座腔菌菌株CBS 119937对7株2年生栓皮栎幼苗进行了致病性测试，这些幼苗在14至26°C的温室中培养。从PDA上一个活跃生长的菌落边缘取下一个菌丝块（3至4毫米），放入每株幼苗茎基部用手术刀造成的浅伤口中。将无菌PDA块放入3株对照幼苗的类似伤口中。接种点用Parafilm包裹以保持一周的湿度。4周后，所有接种小葡萄座腔菌的幼苗均死亡，茎皮层组织出现萎蔫，伴有深褐色变色和10.9±0.4厘米的维管束坏死。对照幼苗未出现症状。从所有接种的幼苗中重新分离出了病原菌，从而满足了柯赫氏法则。结果证实了这种真菌的致病性，并表明其可能与栓皮栎衰退的病因有关。小葡萄座腔菌是一种世界性的、多食性病原菌，可在几种具有经济重要性的寄主上引起病害，如葡萄（3）、猕猴桃（2）和桉属树木（1）。据我们所知，这是关于小葡萄座腔菌引起栓皮栎溃疡病的首次报道。参考文献：（1）A. Gezahgne等人，《南非植物学杂志》70:241，2004年。（2）S. R. Pennycook和G. J. Samuels，《真菌分类学》24:445，1985年。（3）A. J. L. Phillips，《地中海植物病理学》41:3，2002年。（4）B. Slippers等人，《真菌学》96:83，2004年。
