苹果轮纹病室内快速评价体系的建立

为探索高效、稳定且操作简便的苹果轮纹病室内评价方法,选用分离菌株Slg08O3-1-1-1菌饼分别对离体富士苹果两年生枝条以及当年生嫩梢、叶片和成熟果实4种材料进行接种,并对试验结果和接种方法进行比较分析.结果显示,在有伤和无伤条件下各接种材料接种无菌PDA块均不发病,接种病菌仅在有伤情况下致病;枝条烫伤接种发病最慢且病斑最小,嫩梢针刺1针接种较叶痕接种发病明显;1针及10针刺伤接种叶片正、反面,正面1针刺伤接种发病较快且一致;果实去除果皮接种较1针、10针刺伤后接种发病显著.用4个致病力不同的菌株验证,发现4种离体材料均能验证不同菌株间的致病力差异,且结果与田间无伤接种当年生新梢一致.说明4种离体材料均可准确、快速评价苹果轮纹病,其中正面针刺1针接种叶片的方法最优.     

苹果树腐烂病菌拮抗放线菌JPD-1的筛选及鉴定

为筛选对苹果树腐烂病菌Valsa mali具有拮抗作用的放线菌,从苹果树根际土壤中分离获得放线菌,利用平板对峙法和生长速率法筛选拮抗活性较高的菌株,通过菌株培养特征、形态特征、生理生化特性及16S rDNA序列分析确定其分类地位,并采用离体枝条烫伤接种法测定所筛选获得的拮抗菌株对苹果树腐烂病的防效。结果表明,从苹果树根际土壤中共分离获得28株放线菌,其中菌株JPD-1对苹果树腐烂病菌的拮抗作用最强,抑制率为72.50%,该菌在高氏合成1号培养基上生长最好,培养7 d时菌落生长旺盛,气生菌丝白色,基生菌丝深粉红色,无可溶性色素产生;经形态特征观察、生理生化试验及16S rDNA序列分析,将菌株JPD-1鉴定为公牛链霉菌Streptomyces tau‐ricus。菌株JPD-1发酵滤液对苹果树腐烂病菌的抑制率及对离体枝条上苹果树腐烂病的防效均随稀释倍数的增大而减弱,发酵滤液原液使苹果树腐烂病菌菌丝膨大、末端畸形,抑制率为78.14%,对离体枝条上苹果树腐烂病的防效为76.37%。表明分离到的放线菌菌株JPD-1可以用作苹果树腐烂病的生防材料,具有很好的开发及应用前景。     

引起乙烯褪绿蜜橘果实腐烂的主要致病真菌分离及鉴定

为明确贮藏过程中乙烯褪绿蜜橘果实腐烂的主要致病真菌,以重庆市北碚产区无核蜜橘为研究材料,利用乙烯利对其果实进行褪绿处理,观察贮藏期间褪绿蜜橘果实的发病症状,确定病害种类,统计发病率;并从发病果实中分离主要致病真菌,采用形态学特征观察和ITS序列分子生物学分析对其进行鉴定。结果表明,乙烯褪绿加剧贮藏蜜橘果实病害的发生。从发病果实中共分离出5株主要致病真菌菌株,分别编号为A、B、C、D、E;结合形态学特征和5株致病真菌的ITS序列分析结果,A、B菌株均被鉴定为胶孢炭疽菌Colletotrichum gloeosporioides,但这2株菌株具有不同的菌落形态及致病特性;C菌株被鉴定为焦腐病菌Lasiodiplodia theobromae;D菌株被鉴定为交链格孢菌Alternaria alternata;E菌株被鉴定为棘孢曲霉Aspergillus aculeatus。A、B菌株引起的果实腐烂占乙烯褪绿蜜橘总腐烂果实的81.96%,是引起贮藏过程中乙烯褪绿蜜橘果实腐烂的主要致病真菌,其次为C菌株,D、E菌株为次要致病真菌。     

苹果树腐烂病内生拮抗放线菌A-2的鉴定及其活性评价

以苹果树腐烂病菌Valsa mali为靶标菌,从健康苹果树枝条中获得一株高效内生放线菌A-2。采用生长速率法、对峙培养法和玻片法测定了菌株A-2的抑菌谱及对苹果树腐烂病菌菌丝生长和孢子萌发的影响;利用离体枝条烫伤接种法测定了菌株A-2对腐烂病的防治效果。结果表明:A-2发酵滤液对腐烂病菌的抑制率达90%以上,对其他9种常见的植物病原真菌也具有良好的拮抗作用;菌株A-2可导致腐烂病菌菌丝畸形、分支增多和局部膨大;不同稀释倍数的A-2发酵滤液对苹果离体枝条腐烂病均有明显的防效,接种病原菌前后24 h和同时施用500倍发酵滤液的防效均达70%以上。根据培养特征、生理生化特性和16S rDNA 序列分析,将菌株A-2鉴定为卡伍尔链霉菌Streptomyces cavourensis。     

葡萄溃疡病菌转化子致病力快速评价体系的建立

为探索快速、稳定、高效的葡萄溃疡病菌转化子致病力评价方法,分别采用无伤接种、昆虫针针刺接种、打孔器打孔接种等3种接种方法,将随机选取的来自葡萄溃疡病菌REMI转化子库的50株转化子分别接种‘富士’苹果果实和‘夏黑’葡萄一年生绿枝条,测量病斑大小.结果表明,用葡萄绿枝条接种能区分不同转化子的致病力,而苹果果实则仅能将致病力减弱的转化子筛选出来.在葡萄绿枝条上致病力减弱的转化子有96.15％在‘富士’苹果果实上的致病力也是减弱的,而用苹果果实接种未筛选到致病力增强转化子.上述结果为获得致病力突变体奠定了基础.     

离体水稻叶片划伤接种鉴定稻瘟菌的致病型

抗瘟品种的培育和抗瘟基因布局需要快速、准确、大规模地定性水稻抗源及其后代的抗瘟基因型和稻瘟菌致病型。为此, 本研究建立了水稻离体叶片划伤接种方法。该方法依据主效抗瘟基因抵抗稻瘟菌在寄主体内扩展的特点, 通过针刺在水稻叶片上造成伤口, 避免了寄主侵入抗性的干扰, 从而有利于抗扩展性的定性鉴定。作者利用活体喷雾接种、叶片无划伤接种和本研究建立的离体叶片划伤接种等3种接种方法, 在秧苗4~6叶龄期, 对菌株12-DG-68在24个水稻抗瘟单基因系上的致病反应进行了测定, 结果显示：叶片划伤接种的检测结果稳定、一致; 而叶片无划伤接种和活体喷雾接种的检测结果假抗性比例分别为12.5%和4.2%, 不同叶龄期的叶片间反应型不一致率达7%。此外, 离体叶片划伤接种还可利用菌丝块接种, 以鉴定分生孢子产量低的菌株的致病型。因此, 水稻叶片划伤接种是一种准确、稳定和方便的稻瘟菌接种方法, 可用于大规模定性测定水稻抗源及其后代的抗瘟基因型和稻瘟菌的致病型。     

湘西地区猕猴桃细菌性溃疡病抗性资源筛选及其抗性机理研究

由Pseudomonas syringae pv.actinidiae(Psa)引起的溃疡病是猕猴桃生产中威胁最大的细菌性病害。种植抗病品种是防治猕猴桃溃疡病最有效途径,猕猴桃抗源是抗病育种的物质基础。本试验通过离体接种,在室内评价了4个不同种的7个猕猴桃资源及品种对溃疡病的抗性。结果表明:供试材料离体叶片、枝条接种溃疡病菌后,其发病时间、病斑大小、发病率差异明显。按病斑大小排序,其抗性强弱依次为毛花猕猴桃‘M9808’对萼猕猴桃‘S9801’美味猕猴桃‘M-06’美味猕猴桃‘米良1号’中华猕猴桃‘Z-12’中华猕猴桃‘东红’中华猕猴桃‘红阳’(CK)。其中,毛花猕猴桃‘M9808’抗病性最强,表现为发病最晚,离体叶片、枝条分别于接种后10 d、20 d发病,比对照推迟了8 d、10 d;病斑最小,为对照的1/25、1/11;发病率最低,为对照的1/28、1/10。抗病性相关酶活性比对发现,不同材料的PAL、CAT、POD酶活性差异较大,抗性材料均高于感病材料,且峰值出现时间早于感病材料。其中,毛花猕猴桃‘M9808’的PAL峰值最高,接种后5 d出现,比对照早10 d;CAT、POD峰值接种后10 d出现,比对照早5 d。说明毛花猕猴桃‘M9808’对溃疡病抗性最强,可作为今后猕猴桃抗病育种或抗性砧木的理想材料。这为猕猴桃抗病育种提供了理论依据,为猕猴桃溃疡病的防控提供了参考。     

剑麻抗斑马纹病鉴定技术研究

为了促进形成一套剑麻抗斑马纹病的鉴定技术,本研究对剑麻斑马纹病最适宜接种条件进行了探讨,并利用活体叶片接种和离体叶片接种两种方式,以抗性不同的8个剑麻种质为试验材料,在人工气候箱和恒温室中进行剑麻抗斑马纹病接种试验,并与大田抗病性进行比较.结果表明:针刺法接种,25～30℃保湿培养是该病最适宜的发病条件;两种接种方法都能比较准确地鉴定剑麻种质对斑马纹病的抗病性,而活体叶片接种较离体叶片接种鉴定抗病性更加准确.     

马尾松梢枯病的研究

由松壳色单隔孢(Diplodia pinea(Desm.)Kickx)引起的马尾松梢枯病在我国广东省首次发现。马尾松梢枯病的主要症状有梢枯、嫩梢弯曲、溃疡、流脂、叶枯,甚至全株死亡。病原菌的特征是分生孢子双孢,褐色,大小为28.5～39.9×14～17微米。松壳色单隔孢能侵染马尾松(Pinus massoniana)、湿地松(P.elliotii)、加勒比松(P.caribaea)和火炬松(P.taeda)。病原菌在受病的针叶、枝条和球果上越冬。每年3～5月第一次发病,7～8月第二次发病。当年生嫩梢未老化之前,病原菌能从伤口侵入或从嫩叶、嫩梢直接侵入。嫩梢老化以后,只能从伤口侵入。在自然条件下,病害主要发生在嫩梢伸长的后期,很少发生在嫩梢伸长的前期。在病树枯枝、枯叶上,松壳色单隔孢和多毛孢菌(Pestalotia sp.)之间存在生存竞争的关系,经接种试验证明后者对马尾松没有侵染能力,可利用它进行生物防治的研究。     

荔枝炭疽病室内抗病评价体系的建立

由炭疽菌(Colletotrichum gloeosporioides)引起的荔枝炭疽病是产业中最严重的病害之一，其周年发生，可以侵染叶片、花穗和果实。为评估荔枝资源的炭疽病抗病性、筛选抗性种质，本研究利用离体叶片，分析了接种方法、叶片叶龄、接种位置、发病时间和病斑测量方法对抗病性评价的影响。结果表明，选用叶片完全舒展、达到最大叶面积、刚完成转色的叶片，在叶片背面针刺接种炭疽菌菌饼(直径5 mm),接种后72 h观察发病情况，并测量病斑面积，能够较好的评估荔枝的抗病性。利用此方法分析了55份荔枝资源接种炭疽菌后的叶片病斑面积，利用欧式平方距离法将病情指数分为6级，根据发病情况将抗性等级归类为免疫、高抗、中抗、中感和高感。本研究建立了利用荔枝离体叶片评价抗炭疽病的技术方法，明确了评价标准，为荔枝抗病品种的选育、培育和抗病基因的筛选提供了技术支持。     

Factors affecting the severity of bacterial canker of pear caused by Pseudomonas syringae pv. syringae

Several factors affecting the severity of bacterial canker of pear were studied. In the orchard, infection of shoots by Pseudomonas syringae pv. syringae occurred only when the inoculum dose exceeded 10⁶ colony-forming units/shoot. However, under favourable conditions in a growth chamber, cankers formed on detached shoots inoculated with 5 cfu/shoot. A second-order polynomial relationship was established between log₁₀ transformed canker length and log₁₀ transformed inoculum dose. In orchard and growth chamber experiments, shoots were susceptible from the time of bud swell until after fruit harvest. The severity of Pseudomonas canker of detached shoots increased if they were frozen at – 10°C for 24 h before inoculation. Shoots were most susceptible when inoculated immediately after wounding, and no cankers developed in the orchard when 3-day-old wounds were inoculated. Additionally, no cankers resulted from inoculation of leaf scars at leaf drop. Actively growing, current-season shoots were more susceptible than shoots that had set a terminal bud. The practical implications of these results are discussed as a basis for control of bacterial canker of pear.     

新疆部分地区苹果、核桃和杨树腐烂病的病原鉴定

 树木腐烂病在新疆主林果种植区均有不同程度的发生,近年呈加重趋势。本研究在调查了新疆阿克苏、和田、伊犁、喀什和库尔勒等地区苹果树、核桃树和杨树3种林果腐烂病田间发病情况的基础上,重点对14个样本点的样品进行病原物的分离与鉴定。结果表明,各样本点3种林果均有不同程度的腐烂病发生,分离鉴定结果为引起苹果树腐烂病病原菌主要为Valsa mali var. mali(分离频率为64.5%),其次是V. mali var. pyri(25.8%)、V. malicola(3.2%)和V. nivea(6.5%);引起核桃树和杨树腐烂病病原菌为V. sordida(分离频率为83.3%和96%)和V. nivea(分离频率为16.7%和4%)。离体枝条及苹果果实上的致病性测定结果表明,3种林果上分离得到的病原菌均能感染原寄主。引起新疆部分地区苹果树腐烂病的主要病原菌是Valsa mali var. mali,核桃树和杨树腐烂病的主要病原菌是V. sordida。同时,基于实际生产中存在感病枯死杨树枝干作为苹果园、核桃园树体支撑木的现象,推测3种林果腐烂病菌间可能存在交叉感染的潜在风险。     

Population structure of Valsa ceratosperma, causal fungus of Valsa canker, in apple and pear orchards

On the basis of mycelial compatibility, the population structure of Valsa ceratosperma, the causal fungus of Valsa canker on broadleaf trees, was investigated in apple and pear orchards in Japan. Field strains of V. ceratosperma from a single canker on trunks or scaffold limbs belonged to different mycelial compatibility groups. Thus, the population structure of this fungus was complex in most orchards. Because mycelia of strains originating from different conidia from the same pycnidium were compatible, infection by this fungus is thought to be ascospores.     

苹果树腐烂病斑季节扩展动态

本研究通过田间人工接种苹果树腐烂病病菌并在发病后对病斑进行连续观察,明确苹果树腐烂病在不同季节发生的难易程度和病斑扩展动态。结果表明,不同生长季节,苹果树腐烂病从人工接种到扩展为可见病斑(病斑径向扩展5mm)所需的时间不同,其中春季所需时间最短,平均为7.7d;其次是夏季,平均为10.2d;秋季所需时间最长,平均为12.5d。不同季节接种后形成的腐烂病斑在发病后2个月内都会出现一个快速扩展过程,随后病斑停止或缓慢扩展,经2~5个月后,再次出现一个快速扩展过程。无论哪个季节接种形成的病斑,第二个快速扩展期都出现在春季或秋季。     

7种杀菌剂对3种林木腐烂病菌的毒力测定及活性评价

林木腐烂病是苹果树、梨树和杨树等林木枝干的重要真菌性病害。为了筛选出对苹果树腐烂病菌Valsa mali var. mali、梨树腐烂病菌V. mali var. pyri和杨树腐烂病菌V. sordida等3种不同寄主腐烂病菌都能有效防控的杀菌剂,本研究开展室内毒力试验比较了7种杀菌剂对3种腐烂病病原菌菌丝生长和分生孢子萌发的抑制效果,并进一步通过田间活性测定试验比较7种杀菌剂对梨树腐烂病病斑扩展和分生孢子发生的防治效果,同时测定了增效剂8.6%聚乙二醇(PEG)对7种杀菌剂的增效作用。毒力测定结果表明,苯醚甲环唑、戊唑醇、吡唑醚菌酯和丙唑·多菌灵对3种腐烂病病原菌菌丝生长和分生孢子萌发的抑制作用较强,其中EC50平均值最低的是苯醚甲环唑,而戊唑醇的MIC平均值最低,在0.33 mg/L浓度下对3种腐烂病病原菌的菌丝生长和分生孢子萌发抑制率均达到100%。田间试验结果表明,45%苯醚甲环唑SC、43%戊唑醇SC和35%丙唑·多菌灵SE对梨树腐烂病病斑扩展和分生孢子萌发的防治效果突出,其中45%苯醚甲环唑SC 30.00 mg/L对病斑扩展防治效果达到82.23%,孢子萌发抑制效果达到85.96%,田间防治效果最好。10%丙硫唑SC+8.6% PEG处理组对病斑扩展防治效果提高了15.39百分点,达到73.46%,分生孢子萌发抑制率提高了23.75百分点,达到83.06%,增效作用显著。本研究为苹果树、梨树和杨树等3种寄主腐烂病的化学防控提供了科学依据。     

梨和苹果腐烂病菌不同培养表型菌株的致病性分析

The pathogenicity of three strains (F-SD-8, F-BJ-2c-2 and F-HN-2a-1) of Valsa mali var. pyri causing pear canker and one strain (F-SX-A6) of V. mali var. mali causing apple canker in China were comparatively tested by wound inoculation on in vitro twigs of pear, apple and some other woody plants, and in vivo twigs of pear. Significant pathogenicity differentiation was detected in V. mali var. pyri. Generally strains F-SD-8 and F-BJ-2c-2 were highly pathogenic on pear although their culturing characteristics differed greatly. The strain F-SX-A6 was more aggressive on apple than on pear, and the strain F-HN-2a-1 showed significant lower pathogenicity on ten pear cultivars and other seven species of woody plants. Our results confirmed that two variants of V. mali had host preference and were also aggressive to crabapple, apricot, and peach besides apple and pear. Meanwhile, strains F-SD-8 and F-BJ-2c-2 could induce the formation of pycnidia on in vivo twigs of pear, which was not observed on in vivo twigs inoculated with F-HN-2a-1 and F-SX-A6.     

Mycoparasitic Trichoderma isolates as a biocontrol agent against Valsa ceratosperma,the causal agent of apple valsa canker

European Journal of Plant Pathology - Apple valsa canker, caused by the fungus Valsa ceratosperma, is one of the most destructive diseases of this crop. Conventional fungicide treatments are not...     

Phenotypic,molecular and pathogenic characterization of Phlyctema vagabunda,causal agent of olive leprosy

Olive leprosy, caused by the fungus Phlyctema vagabunda, is a classic fruit rot disease widespread in the Mediterranean basin. From 2009 to 2013, new disease symptoms consisting of small circular necrotic leaf lesions, coin branch canker and shoot dieback were observed in Spanish and Portuguese olive orchards showing intense defoliation. Phlyctema‐like anamorphs were consistently isolated from leaves and shoots with symptoms. Representative isolates from affected leaves, shoots and fruits were characterized based on morphology of colonies and conidia, optimum growth temperature and comparison of DNA sequence data from four regions: ITS, tub2, MIT and rpb2. In addition, pathogenicity tests were performed on apple and olive fruits, and on branches and leaves of olive trees. Maximum mycelial growth rate ranged between 0.54 and 0.73 mm per day. Conidia produced on inoculated apple fruits showed slight differences in morphology among the representative fungal isolates evaluated. Phylogenetic analysis clustered all of the Phlyctema‐like isolates in the same clade, identifying them as Phlyctema vagabunda. On fruits, influence of wounding, ripening and cultivar resistance was studied, with cv. Blanqueta being the most susceptible cultivar. On branches, a mycelial‐plug inoculation method reproduced olive leprosy symptoms and caused shoot dieback. On leaves, Koch's postulates were fulfilled and the pathogen caused characteristic necrotic spots and plant defoliation. This is the first time that the pathogenicity of P. vagabunda in olive leaves has been demonstrated.     

Incidence-density relationship of pear scab (Venturia nashicola) on fruits and leaves

Lesion-count data on fruits/leaves from two regions of China and on leaves from controlled-environment studies were used to investigate incidence-density [incidence of leaves/fruits with lesion(s) and average number of lesions per leaf/fruit] and incidence-incidence [incidences of leaves and shoots with lesion(s)] relationships. Few of the datasets for the number of lesions per fruit/leaf could be fitted satisfactorily by a Poisson distribution. Three two-parameter distributions (negative binominal, Neyman type A and Polya-Aeppli) provided significantly better fit than the Poisson distribution, indicating a degree of aggregation in the number of lesions on a single leaf/fruit. However, many datasets could still not satisfactorily be fitted by these distributions. The dynamics of aggregation of lesions on leaves/fruits was well described by Taylor's power-law model. Regression models provided accurate predictions of the average number of lesions per leaf/fruit from the incidence of leaves or fruits with lesion(s). Nevertheless, the incidence-density relationship varied considerably between regions and between leaf and fruit scab. Field data also indicated that the number of scabbed leaves per shoot showed some degree of aggregation. The incidence of leaves with scab could be predicted accurately from the incidence of shoots with scab. The incidence-density relationships developed in this study could be used in making practical disease-management decisions when incidence of leaves with scab is less than 35%.