Genetic diversity and aggressiveness of Calonectria pteridis in Eucalyptus spp.

Calonectria leaf blight, caused by Calonectria pteridis, is currently one of the main foliar diseases in eucalypt plantations in Brazil. In warm and high rainfall regions, the disease can be a limiting factor for eucalypt production when planting susceptible genotypes. The most effective method for controlling this disease in the field is the use of resistant genotypes, which requires knowledge of the genetic variability and aggressiveness of the pathogen population for effective deployment of plant resistance. This work evaluated the genetic diversity and aggressiveness of C. pteridis populations obtained from infected eucalypt plants in Monte Dourado (Pará state) and Imperatriz (Maranhão state), Brazil. To study the genetic diversity, 16 ISSR primers were tested, five of which amplified polymorphic, reproducible and informative bands. Thirty-one closely related genotypes were identified from 84 isolates studied, indicating that the population has a low genetic diversity. The aggressiveness of seven isolates, selected according to geographic origin and their clustering in the ISSR-based dendogram, was determined by inoculation of a hybrid Eucalyptus grandis × E. urophylla clone under controlled conditions. Disease severity was assessed by both measuring the percentage of plant defoliation and assigning a score according to a diagrammatic scale of symptoms. A high correlation between the two evaluation methods was observed, which revealed significant differences in aggressiveness among the isolates. The diagrammatic scale is recommended for disease evaluation because results are obtained much faster, before the occurrence of severe defoliation. No correlation between clustering in the ISSR-based phylogenetic analysis and aggressiveness was observed.     

First report of Calonectria cerciana causing leaf blight of Eucalyptus in northern India

Eucalyptus spp. and their hybrids are frequently cloned and mass planted across farmland tracts and commercial plantations in northern India. It is a viable feeder species to the paper and pulp industries in this region. In 2018 and 2019, during field surveys conducted in northern India, a serious leaf blight disease was frequently observed in E. tereticornis plantations. Isolation from the blighted leaf samples consistently yielded fungal isolates having Calonectria‐like morphology. Morphological features coupled with sequence analysis of partial β‐tubulin (TUB2) and partial translation elongation factor‐alpha (TEF1) gene regions of two fungal isolates confirmed the species as Ca. cerciana. In detached leaf assays and glasshouse inoculation experiments, both isolates produced symptoms similar to those observed on the naturally infected leaves. Koch's postulates were fulfilled by re‐isolating Ca. cerciana from the inoculated leaves. This work is the first to confirm that Ca. cerciana is associated with a serious leaf blight disease of Eucalyptus in northern India and is an important addition to the taxonomy of Calonectria fungi in India.     

浙江省大豆红冠腐病的病原菌鉴定

大豆红冠腐病(red crown rot of soybean,RCR)是世界范围内的重要的大豆病害之一,病害症状为罹病植株茎基部变红,表面聚集了大量橙红色球状子囊壳。对采自浙江省桐庐县的疑似大豆红冠腐病的病原进行分离和培养,从分离获得的菌株中选择1个代表菌株ZJHZ01进行后续试验和分析,为浙江省大豆产区的红冠腐病的防控提供依据。致病性测定结果表明:菌株ZJHZ01接种大豆茎基部可引起发病,回接症状与田间自然发病症状较为相似。病原菌落初为白色,后变为鹅黄色,菌落中可产生大量红褐色微菌核和厚垣孢子,在PDA、V8和康乃馨叶片培养基中极易产生子囊孢子,与已报道的大豆红冠腐病菌的菌落形态特征相似,初步认为ZJHZ01为冬青丽赤壳(Calonectria ilicicola)。进一步在分子生物学水平上,对其TUB、HIS和EF-1α序列进行PCR扩增和测序,对比Gen Bank中已经录入的丽赤壳属相应序列。基于以上3种基因联合序列,通过采用邻接法构建了的系统发育树。结果表明:菌株ZJHZ01与Gen Bank中C.ilicicola菌株GDBL01存在着极高的同源性,序列相似度达到了99%以上。通过对浙江省桐庐地区大豆产区的大豆病株的病原菌进行致病性、形态学和分子生物学分析,明确了浙江省桐庐大豆产区的红冠腐病是由C.ilicicola侵染所致,这是该病害在浙江省的首次报道。     

EuPOD基因克隆与焦枯病菌胁迫表达分析

植物过氧化物酶是植物体内活性氧清除过程中的关键酶之一,在植物的抗逆性中发挥着重要作用。以对焦枯病菌高抗品系尾细桉为材料,克隆得到一个 POD 基因序列,并命名为 EuPOD,通过实时荧光定量聚合酶链式反应( qRT￣PCR)检验该基因在尾细桉叶片中的表达。结果表明该基因cDNA长999 bp,其编码蛋白由332个氨基酸组成;基于氨基酸序列的系统进化树分析表明, EuPOD在进化上与蓖麻、可可、苹果的同源基因亲缘关系较近,相似性达到了75％,与拟南芥的相似性达到65％,与水稻的相似性较低,只有48％。 EuPOD在桉树焦枯病菌侵染后不同时间所受诱导表达量不同,在12 h的表达量达到高峰。     

Cylindrocladium brown leaf spot on Howea belmoreana caused by Calonectria ilicicola (anamorph: Cylindrocladium parasiticum) in Japan

Brown leaf spot disease caused by Cylindrocladium was found on Howea belmoreana on Hachijojima Island, Tokyo, Japan, in December 2001. Typical symptoms were incited after artificial inoculation. A culture of white mycelia, isolated from leaf spot symptoms, produced reddish perithecia of a nectriaceous fungus. Based on morphological and molecular analyses, this fungus was identified as Calonectria ilicicola (anamorph: Cylindrocladium parasiticum). Pathogenicity of this fungus on five plants cultivated on Hachijojima Island was confirmed by artificial inoculation. This report is the first on Cylindrocladium brown leaf spot of H. belmoreana caused by C. ilicicola (anamorph: Cy. parasiticum).     

中国丽赤壳属 Calonectria 真菌物种及遗传多样性

丽赤壳属(Calonectria)真菌中含有一些在世界范围内分布广且危害大的病原菌。通过基于多基因序列的系统发育分析并结合形态学特征,我国共发现并鉴定了28个丽赤壳属物种,其中27个种是在桉树人工林叶片、桉树林下土壤或者桉树苗上发现。本文对我国丽赤壳属真菌的物种多样性以及遗传多样性进行了概述,对各个物种的形态特征、菌落特点、寄主范围、分布地域和危害特点等进行了详细的描述,并对未来我国丽赤壳属真菌的物种分类以及引起病害的防控研究提供了思路。     

12种桉树焦枯病病原菌对10种桉树无性系致病性初步分析

为了明确在中国发现的桉树叶片焦枯病病原菌Calonectria spp.对桉树的致病力大小,并测定不同桉树无性系对Calonectria病原菌抗病性的强弱,本试验采用在中国发现的12种Calonectria属（Ca. cerciana、Ca. chinensis、Ca. hongkongensis、Ca. microconidialis、Ca. papillata、Ca.parakyotensis、Ca. pauciramosa、Ca. pentaseptata、Ca. pseudoreteaudii、Ca. seminaria、Ca. terrestris、Ca. tetraramosa）共29株致病菌对10个桉树无性系（DH32-22、DH32-29、EC152、EC153、EC155、G1、K31、OC14、U6、W5）进行了室内离体叶片致病性测定。结果显示：12种被测试Calonectria病原菌均能在不同程度上使10个桉树无性系离体叶片产生病斑;不同种Calnectria病原菌对同一桉树无性系的致病性差异显著,不同桉树无性系对同种Calonectria致病菌的抗病能力也存在显著差异。     

Effect of initial darkness duration on the pathogenicity of Calonectria pseudonaviculata on boxwood

Light is an important environmental stimulus that regulates many physiological activities within plants and pathogenic fungi. Boxwood blight (causal agent: Calonectria pseudonaviculata) is a newly emergent disease in the United States and a significant threat to the boxwood industry. A disease‐forecasting model has been developed to aid boxwood growers in their management decisions, but light is not included as a variable within the programme. Growth chamber experiments were conducted to evaluate the effect of an initial dark period on disease severity, mycelial growth and conidial germination of C. pseudonaviculata. Treatments varied by darkness duration (0, 6, 12 or 24 h dark), and were followed by a 14 h day length at a 25/21 °C day/night temperature. Neither boxwood blight severity nor fungal colony development was enhanced by an initial dark period. However, darkness duration was associated with a significant increase in conidial germination in vitro. This study suggests the predictive performance of the current boxwood blight disease‐forecasting model would not be improved by inclusion of photoperiod.     

Root and Collar Rot of Milkwort Caused by Cylindrocladium pauciramosum, a New Record for Europe

A new disease of milkwort (Polygala myrtifolia) was observed on several commercial nurseries in southern Italy. Diseased plants showed wilting, stunting, chlorosis or loss of foliage, and rotting of the basal stem as well as the crown and roots. A Cylindrocladium species was consistently found associated with crown, basal stem, and root lesions. The etiology of this disease was proved on milkwort, by fulfilling Koch's postulates. Two hundred Cylindrocladium isolates were collected from the most important Sicilian and Calabrian ornamental production areas from different host plants. Isolates were identified as Cylindrocladium pauciramosum (teleomorph Calonectria pauciramosa) on the basis of their obpyriform to broadly ellipsoidal terminal vesicles, conidiophore branching pattern, conidium morphology, as well as mating type studies with tester strains of C. pauciramosum for selected isolates. This is the first record of this pathogen from Europe and it is the first report of C. pauciramosum on milkwort.     

桉树焦枯病菌CpSit1基因的鉴定与表达

利用Blast及TCDB数据库对桉树焦枯病菌(Calonectria pseudoreteaudii)的Cpsit1基因进行鉴定;再利用SMART数据库和Prot Param、TMHMM、PHD、Pro Comp 9.0在线分析工具对Cpsit1基因编码蛋白的理化性质、跨膜螺旋、蛋白质二级结构和亚细胞定位进行预测分析;最后采用qRT-PCR方法对CpSit1基因在焦枯病菌侵染桉树过程中的表达情况进行分析.结果表明:CpSit1基因长度为1 780 bp,其编码的蛋白序列共有592个氨基酸残基组成,并将其鉴定为铁载体-铁:H+同向转运蛋白.其保守结构域为MFS_1;共含有14个跨膜螺旋;在二级结构中α螺旋占45.10%,延伸链占26.01%,无规则卷曲占28.89%.通过qRT-PCR相对定量的方法分析焦枯病菌侵染桉树24、48和72 h后CpSit1基因的表达情况,结果显示,CpSit1基因在这3个时段均发生上调表达,但24 h的表达量明显大于48和72 h.说明桉树焦枯病菌CpSit1基因在病原菌侵染寄主的过程中通过调控铁载体-铁化合物的转运来完成铁元素的摄入,协助其在寄主中的定植.