2023年9月,在克里特岛(Chania地区)的一个1.5公顷的果园中发现了30棵枯萎的30岁鳄梨(Perseaamericana)树(\'Hass\'嫁接在'Zutano\'幼苗上)。冠症状包括萎萎和叶萎黄,推进到落叶和广泛的死亡。在严重感染的树木上,Tap和食根腐烂,根组织呈棕色变色。这种疾病严重而广泛,导致300棵树的死亡率为5%。通过改良的土壤诱饵技术(Ferguson和Jeffers,1999).将表面消毒的鳄梨果实浸入含有土壤样品的水中。经过2到8天的时间,将来自果实表面上产生的坏死病变的组织碎片转移到ΡΑRP培养基上,然后在20°C下孵育(Ferguson和Jeffers,1999).通过将单个菌丝尖端转移到含有V8汁琼脂的新培养皿中,获得了三个分离株(AV2,AV12和AV11a)。它们生长在20℃,10天后用于鉴定。分离形成的珊瑚状菌落,具有丰富的簇状球形菌丝肿胀和末端或插入(比例为1:5)厚壁衣原体孢子,尺寸为20至36μm(平均29±0.8μm),具有特征性的厚壁(平均1.2±0.2μm)。Sporangia,在非无菌土壤提取物水中生产,卵形为卵形,持久性,无乳突,32至81μm(平均56±4.8μm)长,20至42μm(平均31±3.2μm)宽(n=100)。分离物是异系的,因为它们在单一培养物中不产生卵孢子。根据形态特征,将分离株鉴定为肉桂疫霉(Erwin和Ribeiro,1996年)。内部转录间隔区(ITS)包括ITS1,5.8SrDNA区,使用ITS1/ITS4和FM83/FM84引物扩增了ITS2以及三个代表性分离株的细胞色素c氧化酶亚基I(coxI)基因,分别(怀特等人。1990年;Martin和Tooley2003),并测序(GenBankacc。分别为PP506613到PP506615和PQ063867到PQ063869)。BLAST搜索显示与肉桂原同种型分离物(分别为KC478663和KU899315)的序列几乎100%同一性。按照土壤侵染方法进行了使用分离物AV2的致病性测试(Jung等人。1996)使用六岁的鳄梨“Zutano”幼苗。用蛭石-多种维生素汁混合物处理的六种未接种植物用作对照。植物(1m高)在温室条件下在盆中生长并定期浇水。接种后六周,所有接种的树木都显示出萎黄病,枯萎和根腐病,而对照植物保持无症状。症状与在该领域中观察到的症状相似,并且如前所述重新分离病原体并进行分子鉴定。这项研究首次记录了肉桂的发生,被广泛认为是全球最具破坏性的鳄梨病原体,关于希腊的鳄梨作物(Rodger等人。2019)。此外,这标志着这种病原体首次出现在克里特岛上,无论宿主物种如何。与鳄梨根腐病相关的疫霉物种的准确鉴定对于实施有效的疾病管理策略至关重要。特别是在选择合适的抗病砧木时。
In September 2023, thirty declining 30-year-old avocado (Persea americana) trees (\'Hass\' grafted on \'Zutano\' seedlings) were detected in a 1.5-ha orchard in the island of Crete (Chania region). Crown symptoms encompassed wilting and leaf chlorosis, advancing to defoliation and extensive dieback. Tap and feeder roots decayed and brown discoloration of root tissues was evident on heavily infected trees. The disease was severe and widespread, resulting in a 5% mortality rate among 300 trees. The pathogen was isolated with a modified soil baiting technique (Ferguson and Jeffers, 1999). Surface disinfected avocado fruits were immersed in water containing soil samples. Following a period of 2 to 8 days, tissue fragments from the resulting necrotic lesions on the fruit surface were transferred on ΡΑRP medium and subsequently incubated at 20°C (Ferguson and Jeffers, 1999). Three isolates (AV2, AV12 and AV11a) were obtained by transferring single hyphal tips to new Petri dishes containing V8 juice agar. They were grown at 20˚C and used for identification after 10 days. Isolates formed coralloid colonies with abundant clustered spherical hyphal swellings and terminal or intercalary (ratio 1:5) thick-walled chlamydospores measuring 20 to 36 μm (avg 29±0.8 μm) with characteristic thick walls (avg 1.2±0.2 μm). Sporangia, produced in non-sterile soil-extract water, were ovoid to obpyriform, persistent, non-papillate, 32 to 81 μm (avg 56±4.8 μm) long and 20 to 42 μm (avg 31±3.2 μm) wide (n=100). Isolates were heterothallic as they did not produce oospores in single cultures. Based on the morphological traits the isolates were identified as Phytophthora cinnamomi (Erwin and Ribeiro 1996). The internal transcribe spacer region (ITS) including ITS1, 5.8S rDNA region, and ITS2 as well as the cytochrome c oxidase subunit I (coxI) gene of the three representative isolates wereamplified with ITS1/ITS4 and FM83/FM84 primers, respectively (White et al. 1990; Martin and Tooley 2003), and sequenced (GenBank acc. PP506613 to PP506615 and PQ063867 to PQ063869, respectively). BLAST search revealed almost 100% identity with the sequences of P. cinnamomi ex-isotype isolate (KC478663 and KU899315 respectively). Pathogenicity tests using isolate AV2 were conducted following the soil infestation method (Jung et al. 1996) using six-year-old avocado \'Zutano\' seedlings. Six non-inoculated plants treated with vermiculite-multivitamin juice mixture were used as controls. Plants (1 m tall) were grown in pots under greenhouse conditions and watered regularly. Six weeks post inoculation, all inoculated trees showed chlorosis, wilting and root rot, while control plants remained symptomless. Symptoms were similar to those observed in the field and the pathogen was re-isolated and molecularly identified as previously described. This study presents the first documented occurrence of P. cinnamomi, widely regarded as the most destructive avocado pathogen globally, on avocado crops in Greece (Rodger et al. 2019). Additionally, this marks the first recorded presence of this pathogen on the island of Crete, regardless of the host species. The accurate identification of Phytophthora species associated with avocado root rot is essential for implementing an effective disease management strategy, particularly in the selection of appropriate disease-resistant rootstocks.