苹果树腐烂病室内快速评价方法的研究

 腐烂病是我国苹果树上一种严重的树干皮层腐烂病害,本研究拟探索快速、可靠、稳定及操作简便的病害室内评价方法。在"富士"苹果离体叶片、嫩梢、果实和枝条上造成不同伤口,强致病菌株03-8接种,25℃保湿培养后调查发现,不接菌对照有、无伤口均不发病;各种伤口接种病原菌均能发病,但伤口类型对病害影响很大。叶片正面较反面更有利于发病,叶片正面1针和10针的刺伤发病无明显差异;嫩梢叶痕接种较刺伤接种发病轻;果实表面针刺1针和10针及去除果皮造成伤口对发病影响大,差异显著;枝条烫伤接种后10d发病明显而其它材料在接种后1.5~2d即可发病。进一步用4个致病力不同的菌株验证评价方法的可靠性和稳定性,发现接种完全展开叶、嫩梢和枝条均可以充分显示不同菌株之间的致病力差异,但前两者重复性好、试验周期短、鉴定效率高,且操作简便、材料易得,因此建议用离体叶片或嫩梢作为材料,针刺1针接种,25℃保湿培养2d后调查,作为室内准确、快速评价苹果树腐烂病的方法,该方法可用于筛选抗病材料、评价分离株的致病性和药剂的防病效果等。     

83份新疆野苹果种质资源对梨火疫病菌的抗病性评价

2022年4—5月,以国家果树种质新疆野苹果资源圃83份新疆野苹果种质资源为试材,采用室内离体叶片和离体枝条接种方法鉴定评价83份新疆野苹果对梨火疫病的抗性。结果表明,梨火疫病菌对83份不同类型新疆野苹果均有致病力。离体叶片和离体枝条接种结果存在一定差异,离体叶片接种的抗性资源多于离体枝条接种。抗病的新疆野苹果种质资源来自新源分布区较多。筛选出高抗新疆野苹果种质资源1份,抗病种质资源4份,感病资源7份,高感资源13份。5份新疆野苹果种质资源可作为抗病砧木选育的基础材料。     

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.     

胶东地区套袋苹果果面斑点病病原鉴定与防治

2014年套袋苹果果面斑点病在烟台严重发生。室内分离鉴定结果表明,其病原菌为链格孢(Alternaria mali)和点枝顶孢(Acremonium stictum),在苹果不同品种上侵染的种类也不同。经室内离体果实接种病菌测定其致病性,针刺有伤接种处理均发病,无伤接种处理均不发病。表明该病原菌是弱寄生菌,只通过伤口侵染。田间防治试验结果表明,10%多抗霉素可湿性粉剂1000倍液喷雾对该病害有较好的防治效果。     

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。     

河南和广西地区紫云英匍柄霉叶斑病的病原菌鉴定

2019年-2020年,在我国河南省信阳市和广西壮族自治区南宁市种植紫云英的田间发现一种紫云英叶斑病。通过常规组织分离法分离得到7株真菌（菌株号为XZYB1~XZYB6、GX1）,经单孢纯化、形态学观察和多基因系统发育学分析（ITS-GAPDH-CAL）,结果显示7株真菌均为黄芪匍柄霉Stemphylium astragali。依据柯赫氏法则,选取菌株XZYB1和XZYB4无伤接种紫云英离体叶片,选取菌株GX1活体接种紫云英叶片进行致病性测定。接种5 d后离体叶片和活体植株叶片均发病,发病症状与田间症状一致,且再分离得到的菌株与接种菌株相同。据此,确定黄芪匍柄霉为该叶斑病的病原菌,并将病害定名为紫云英匍柄霉叶斑病。这是该病害在河南和广西发生的首次报道。本研究也为匍柄霉叶斑病的诊断与后续的防治策略提供了科学依据和理论指导。     

立枯丝核菌AG-1IA对水稻致病力及粗毒素活性的测定方法研究

水稻纹枯病是严重影响水稻生产的真菌病害,水稻相对病斑高度法作为评估立枯丝核菌AG-1 IA致病力的传统方法,需要将水稻培养至分蘖末期或抽穗期,再进行活体接种,耗时耗力,且试验条件不易控制。本研究以水稻相对病斑高度法(活体接种)为对照,利用水稻离体叶鞘法和离体叶片法测定了水稻纹枯病菌致病力和粗毒素活性,比较活体接种法与离体接种法的相关性。结果表明水稻离体叶片法、水稻离体叶鞘法和水稻相对病斑法测定致病力和粗毒素活性结果均呈显著正相关,水稻离体叶片法和离体叶鞘法测定结果能够准确反映立枯丝核菌AG-1 IA的致病力和粗毒素活性,离体评估方法操作方便,试验条件易于控制。因此,水稻离体叶片法、叶鞘法可以替代活体接种法作为测定立枯丝核菌AG-1 IA致病力和粗毒素活性的方法。水稻离体叶片法、叶鞘法与活体接种法结合起来可作为测定立枯丝核菌AG-1 IA毒素活性测定的新方法。本研究的结果还表明,水稻不同组织对水稻纹枯病菌毒素敏感性不同,水稻的叶鞘组织比叶片组织对纹枯病菌敏感。由于粗毒素的致病力与活体菌株接种的致病力存在显著差异,因此纹枯病菌毒素以外的致病因子,也是不容忽视的。在定量分析立枯丝核菌AG-1 IA与水稻的互作中,需要将不同水稻的组织进行区分,才能更加精准地了解互作机制。     

不同龄期枝条对苹果轮纹病菌的抗病性研究

本研究采用轮纹病菌分生孢子和菌丝接种不同龄期的苹果枝条,检测枝条皮孔发病率,以明确不同龄期苹果枝条对轮纹病菌的抗病性差异.结果表明,当年生‘富士’苹果枝条的幼嫩部分接种后发病最重,皮孔的发病率达80％以上,随枝条的生长发育,抗病性逐渐增强,枝条发育成熟后,皮孔发病率降至40％左右.在7月份接种的1～5年生‘富士’苹果成熟枝条中,皮孔的发病率没有显著差异.在8月份接种的‘富士’苹果枝条中,1年生枝条的皮孔发病率显著高于5年生枝条的发病率.未发育成熟的苹果枝条对轮纹病菌的抗侵入能力差,枝条发育成熟后抗病能力明显增强.1～5年生枝条上发育完整的皮孔对轮纹病的抗侵染能力没有显著差异,但在枝条的迅速加粗生长期,低龄枝条上皮孔结构变化更大,对轮纹病菌的侵入也更敏感.     

重庆市烟草赤星病菌致病力测定

利用离体叶片悬滴接种法,将2005年在重庆市各烟区采集分离的烟草赤星病24个代表菌株进行了致病力测定。结果显示,所有测试菌株均具有致病力,但存在明显的致病力差异,其中强致病力菌株5个,较强菌株8个,中等致病力菌株10个,弱致病力菌株1个,分别占供试菌株的20.8%、33.3%、41.7%和4.2%。这表明试验分离获得的大多数菌株致病力居中或偏强。对来源性不同的菌株的致病力进行分析比较,发现重庆各烟区菌株的致病力有所不同,而且同一烟区的菌株致病力也存在一定的差异。离体叶片试验表明,赤星病菌不同病原菌株对烟草叶片的穿透生长能力与致病力呈明显的正相关。     

广西杧果炭疽病病菌致病力差异测定

采用组织分离法从广西不同地区不同生境分离获得19株杧果炭疽病病菌,通过离体针刺接种测定了菌株的致病力差异。结果表明,以接种病菌后7 d的病情指数进行聚类分析,将可19株菌株划分为3个致病类型:强致病型、中等致病型、弱致病型。各类型菌株所占比例大小为:弱致病型中等致病型强致病型。表明广西杧果炭疽病病菌不同菌株的致病力存在明显差异。其中,弱致病型菌株占优势。     

Seasonal dynamics of <Emphasis Type="Italic">Botryosphaeria dothidea</Emphasis> infections and symptom development on apple fruits and shoots in China

Botryosphaeria dothidea is one of the most important pathogens of apple trees in China. It causes warts and cankers on shoots and fruit rot. In this study, multi-season experiments were conducted to investigate the seasonal dynamics of B. dothidea infection and symptom development on twigs and fruits of cv. Fuji through spraying and rain-splash inoculations. Fruit were most susceptible to infection from late May to mid-August. The length of the incubation period, i.e. from inoculation to the first lesion was observed, generally decreased with the age of fruit at the time of inoculation. Fruit inoculated in May–June developed visible symptoms from early August onwards, with a peak in early September. However, fruit inoculated in September developed visible symptoms within one week after inoculation. Twigs remained susceptible to infection by conidia from the initial extension stage in early May to late September. New extension shoots were most susceptible from June to mid-August and then became gradually less susceptible over time. When inoculated before the end of August, twigs developed visible lesions within the same season. In contrast, when inoculated late in the season, most lesions became visible during the subsequent season. The incubation period on twigs was shorter for inoculations conducted in June–August, with the shortest being 25 days. These results may contribute to the improvement of the management of this disease.     

绛红褐链霉菌YSSPG3的发酵条件及其发酵滤液对杂交竹梢枯病的防治作用

绛红褐链霉菌YSSPG3是从四川碧峰峡撑×绿杂交竹健康植株根际分离到的一株对杂交竹梢枯病菌有抗菌活性的放线菌。为进一步明确该菌的生防潜力,采用正交设计进行发酵培养基优化,并通过分生孢子萌发试验、菌丝抑制试验、离体枝叶接种试验及田间防治试验,研究了优化发酵培养滤液对杂交竹梢枯病的防治效果。结果显示,最优培养基组成为葡萄糖5 g、甘油5 g、黄豆饼粉3 g、蛋白5 g、NaCl 2.5 g、CaCO32 g、蒸馏水1 000 mL。优化发酵培养滤液对病原菌分生孢子萌发及菌丝生长均有明显的抑制作用,可导致芽管、菌丝畸形。去菌发酵原液及其10倍稀释液处理离体枝叶能有效抑制病斑扩展,接种后4天的防治效果在70%以上。田间喷施去菌发酵原液及其5倍稀释液防治效果均达80%以上,极显著高于浓度为30 mg/mL的化学药剂50%多菌灵和70%甲基硫菌灵(P<0.01)。     

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.     

Survival and dispersal of a marked strain of Pseudomonas syringae in a maple nursery

A pathogenic Pseudomonas syringae strain resistant to rifampicin and nalidixic acid was sprayed upon and colonized maple twigs and perennial ryegrass. The inoculated twigs were sampled at intervals of 2-3 weeks from July 1985 to September 1986, and epiphytic populations of the marked strain recovered during this time ranged from undetectable to 10⁴ colony-forming units/g. The results showed that this strain of P. syringae could overwinter on maple twigs and potentially serve as a source of inoculum in the spring. Aerial dispersal was also investigated. The marked strain inoculated onto grass growing in pots was detected on medium in inverted petri plates, on maple leaves and with an Andersen sampler positioned from 12 to 100 cm above the grass. The number of detectable cells that dispersed vertically upwards was low even in the presence of wind, rain, or irrigation water. The marked strain inoculated onto maple branches was isolated from grasses under the inoculated trees, showing that P. syringae was dispersed downward. Lateral dispersal of the marked strain from inoculated to uninoculated trees was not detected. The identity of the antibiotic-resistant strain isolated from the trees and grasses was confirmed by DNA restriction-fragment profile analysis.     

防治苹果树腐烂病杀菌剂的室内筛选

 The inhibitory effects of 10 fungicides on apple tree valsa canker were compared on dishes and twigs. The results showed that conidia germination and mycelial growth were inhibited by all fungicedes tested. The EC₅₀ value of Difenoconazole was 6. 1×10^-3μg·mL^-1, the lowest among tested fungicide in inhibiting conidia germination. Tebuconazole and Imazalil also showed obvious inhibition effect. Thiophanatemethyl was the least efficient with the EC₅₀ value 2.6×10¹ μg·mL^-1. Difenoconazole also showed the highest activity for inhibiting mycelial growth with ECho value of 2.3×10^-2 μg·mL^-1. Tebuconazole was better than other fungicide. Thiophanate-methyl and Propineb were the least efficient with quite high EC₅₀ value. Furthermore, the size of the lesion after inoculation on excised twigs also revealed that Difenoconazole was most efficient because it showed the smallest lesion of 394 mm² compared with other tested fungicides. So Difenoconazole and Tebuconazole could be used to control apple tree valsa canker in field to subsititute forbided fungicides such as asomate.     

Wound‐healing in cocoa (theobroma cacao l.) stems and its effect on canker caused by phytophthora palmivora (butl.) butler

Investigations on canker development in six cocoa genotypes showed IMC 67 to be resistant, ICS 1, TSH 1188 moderately resistant and TSH 1076, SCA 6 and P 18 as susceptible. Plant wound‐healing components (lignin, callose and suberin) were detected in healthy, wounded and wound‐inoculated (Phytophthora palmivora) tissues of all the genotypes, although at different rates. Lignin concentration was negatively correlated with canker lesion size and positively correlated with time of complete healing. A strong positive correlation was also observed between time of healing and lesion size. The rate of healing of uninoculated wounds was fastest in IMC 67, moderate in ICS 1 and TSH 1188 and slowest in TSH 1076, SCA 6 and P 18 while cessation of lesion expansion was observed at 6 weeks in IMC 67, 8 in ICS 1, TSH 1188 and 10 in TSH 1076, SCA 6 and P 18. This study revealed that lignification plays a role in wound‐healing and consequently a role in resistance of coca stems to P. palmivora infection.     

Association of citrus psorosis B symptoms with a sequence variant of the Citrus psorosis virus RNA 2

Citrus psorosis virus (CPsV), the type species of genus Ophiovirus, is the presumed causal agent of a bark scaling disease in citrus plants. CPsV virions are kinked filaments composed of three negative‐strand RNA molecules and a ～48‐kDa coat protein. The virus induces two different syndromes: psorosis A (PsA), characterized by limited bark scaling lesions in the trunk and main limbs, and a more aggressive form of the disease called psorosis B (PsB) with rampant bark lesions affecting even thin branches and chlorotic blotches in old leaves. In the greenhouse, the PsA and PsB syndromes can be induced by graft inoculating healthy citrus seedlings with non‐lesion or with lesion bark inoculum from PsA‐affected field trees. PsA‐ and PsB‐inducing CPsV sub‐isolates obtained by this procedure from the same tree showed identical single‐strand conformation polymorphism (SSCP) profiles in homologous segments of the RNAs 1 and 3, whereas segments of the RNA 2 enabled discrimination between PsA‐ and PsB‐associated sequence variants. SSCP analysis of the RNA 2 population present in different tissues of psorosis‐infected plants showed that: (i) PsA‐inducing isolates contain PsB‐associated sequence variants at low frequency, (ii) the PsB‐associated sequence variant is predominant in blistered twigs and gummy pustules affecting old leaves, characteristic of PsB isolates, and (iii) the PsB‐associated sequence variant accumulates preferentially in bark lesions of the trunk and limbs. SSCP analysis of the RNA 2 population also enabled monitoring of interference between PsA‐ and PsB‐associated variants in plants co‐inoculated with both psorosis types.     

Use of phages for identifying the citrus canker bacterium Xanthomonas campestris pv. citri in Taiwan

Phages CP115 and CP122, which were isolated from canker lesions on grapefruit and Liucheng sweet orange, respectively, showed a high degree of specificity with respect to lysis of test bacterial strains. When used jointly, they lysed 135 (97·8%) out of 138 Xanthomonas campestris pv. citri strains isolated from the canker lesions on leaves, twigs, and fruits of various citrus species, cultivars, and hybrids grown throughout Taiwan, but they did not lyse other X. campestris pathovars and other phytopathogenic bacteria, nor other bacteria isolated from soil, clinical or environmental samples. Of 252 CP115/CP122-sensitive and 78 CP115/CP122-resistant bacterial strains with colony characteristics typical of or similar to those of X. campestris pv. citri , isolated from canker lesions of various citrus plants in diverse growing regions in Taiwan, 250 (99·2%) and 76 (97·4%) strains were pathogenic and non-pathogenic, respectively, when inoculated into Liucheng sweet orange or Mexican lime. Thus, phages CP115 and CP122, when used jointly, appear to be applicable for identifying X. campestris pv. citri in Taiwan.     

Pathogenicity of Marssonina betulae and other fungi on birch

Marssonina betulae , Discula betulina , Melanconium bicolor and Fusarium avenaceum were inoculated onto shoots of 1- and 2-year-old seedlings of Betula pendula and B. pubescens and symptom development monitored over several seasons. Marssonina betulae caused disease on B. pendula , but not on B. pubescens . On B. pendula symptoms included discrete lesions, which often girdled, causing dieback of inoculated leading shoots, and the development of secondary sunken cankers on the main stems, which were usually centred around a dead sideshoot. Cankers on the main stems expanded during subsequent growing and dormant seasons, and often coalesced, girdling stems and causing the death of some seedlings. All isolates of M. betulae caused disease on B. pendula and conidia were able to infect young shoots in early flush without requiring a wound. Discula betulina caused lesions and dieback on B. pendula and B. pubescens within 3 months of inoculation, but disease did not progress thereafter and all inoculated seedlings recovered. Melanconium bicolor and F. avenaceum caused very little disease on either birch species. This study showed that M. betulae is an aggressive pathogen on B. pendula , causing sunken stem cankers and progressive crown dieback, which are symptoms commonly observed on young, planted birch at field sites across Scotland.