一株疫病拮抗青霉P.st10菌株的抗菌活性及其对辣椒疫病的盆栽防效

筛选得到一株疫霉菌的拮抗青霉Penicillium striatisporumP.st10。该菌株强烈抑制疫霉菌及核盘菌的菌丝生长。无菌滤液抑制辣椒疫霉孢子囊的形成及孢子囊、游动孢子的萌发,且能杀死菌丝细胞。该菌株在辣椒根际有很强的定殖能力,有机肥对其定殖有增强作用,接种后40d在根际的青霉菌数量,青霉处理为0.76×104cfu/g,而青霉菌肥处理为2.55×104cfu/g。盆栽试验显示:处理7d后,土壤只接种辣椒疫霉对照发病率为93%;同时接种青霉培养滤液和辣椒疫霉处理为16%;接种青霉培养滤液、新鲜菌丝和辣椒疫霉处理为15%。     

百合疫病病原菌的鉴定及培养基的筛选

从具典型症状的新鲜百合疫病植株茎基部病组织中分离到百合疫霉菌,根据其病原菌菌丝的形态、菌落特征,厚垣孢子、游动孢子囊和卵孢子的形态和大小,以及病原菌致病性测定,该病原菌鉴定为烟草疫霉Phytophthora nicotianae van Brede de Haan.供试的16种培养基中,病原菌在胡萝卜琼脂培养基(CaA)和辣椒琼脂培养基(PeA)上生长最好,生长速率分别为1.771和1.770mm/h.在常规培养条件下,病原菌不易产生厚垣孢子、游动孢子囊和卵孢子,在低温、皮氏溶液和土壤浸出液中分别诱导产生出大量厚垣孢子、游动孢子囊和卵孢子.     

非寄主和寄主种子分泌物对大豆疫霉活性的影响

为探寻非寄主和寄主种子分泌物中抗病信号分子,通过显微观察,采用菌丝生长速率法和离体接种法对不同种子分泌物处理后大豆疫霉Phytophthora sojae的游动孢子数、孢子囊数、游动孢子释放后残留的空囊数、成囊和未成囊的游动孢子数、萌发和未萌发的胞囊数、菌落直径、卵孢子数进行测量,并计算抑制率,明确非寄主菜豆和寄主大豆抗病品种、感病品种种子分泌物对大豆疫霉游动孢子趋化性、生长发育和侵袭力的影响。结果显示,非寄主菜豆种子分泌物不吸引大豆疫霉游动孢子,显著抑制大豆疫霉孢子囊形成、胞囊萌发和卵孢子产生,抑制率依次为97.3%、73.0%和17.5%,然后溶解胞囊,最终导致游动孢子对下胚轴侵袭力降低,抑制率为67.1%。寄主大豆种子分泌物能吸引大豆疫霉游动孢子,感病品种种子分泌物吸引力高于抗病品种。感病品种种子分泌物对大豆疫霉生长发育无显著影响,但促进大豆疫霉游动孢子侵袭力;抗病品种种子分泌物显著抑制大豆疫霉孢子囊形成、胞囊萌发和卵孢子产生,抑制率依次为86.6%、34.3%和12.8%,然后溶解胞囊,但作用强度小于非寄主菜豆种子分泌物,最终导致游动孢子对下胚轴的侵袭力降低,抑制率为24.2%。表明非寄主菜豆和寄主大豆抗病品种的种子分泌物对大豆疫霉有抑菌活性,大豆疫霉的非寄主和寄主抗病性与种子分泌物有关。     

抗甲霜灵辣椒疫霉菌株的生物学特性

为了明确抗甲霜灵辣椒疫霉菌株的适生性,以田间自然产生及室内诱变获得的抗甲霜灵辣椒疫霉菌株为材料,测定了抗甲霜灵辣椒疫霉菌株的主要生物学性状。各菌株的游动孢子囊产生量为9.6×103～13.0×103个/mL,孢子囊释放率为61.3%～68.4%,游动孢子萌发率为35.3%～42.1%,适温条件下的菌丝平均生长速率约为10.28～11.55 mm/天;各菌株在含100μg/mL孔雀石绿的CA培养基上均不能生长,淀粉水解指数为0.18～0.95;接种辣椒、番茄、南瓜、西葫芦和茄子等果实所致病斑平均直径分别为34.11、18.47、30.19、37.78和32.73 mm。经方差分析,抗甲霜灵辣椒疫霉菌株的生物学性状与敏感菌株无显著差异,表明甲霜灵并未改变其生物学性状,辣椒疫霉对甲霜灵具有较高的抗性风险。     

链霉菌Sa-21对烟草黑胫病的防效及抑菌作用研究

烟草疫霉引起的烟草黑胫病,是烟草生产上的重要病害之一。为了明确链霉菌Sa-21防治烟草黑胫病的应用潜力,用盆栽试验测定了链霉菌Sa-21的生防效果,并用生长速率法等手段探讨了链霉菌Sa-21对烟草疫霉的抑菌作用。盆栽试验表明,链霉菌Sa-21菌株发酵液对苗期和成株期烟草黑胫病有较好的控制效果,最高防效可达98.39%。菌株Sa-21粗浸膏对烟草疫霉菌丝生长的抑菌中浓度为56.1 μg/mL,50 μg/mL的粗浸膏处理后可以降低50%的菌丝生物量。菌株Sa-21粗浸膏对烟草疫霉孢子囊的产生、游动孢子释放、休止孢萌发以及细胞膜透性均有抑制作用,100 μg/mL的粗浸膏基本能够完全抑制孢子囊的产生,并且对游动孢子释放的抑制率达50%以上;粗浸膏浓度为150 μg/mL时,对休止孢萌发的抑制率达94.35%;随着粗浸膏浓度增加,菌丝细胞膜的透性也会加大。研究结果表明,链霉菌Sa-21代谢物对烟草疫霉的生长繁殖产生抑制作用,并为链霉菌Sa-21生防菌剂的开发及其对烟草黑胫病的防治提供了理论依据。     

核桃内生细菌HT-6对致病疫霉的持续抑制作用及其抑菌特性研究

为进一步了解核桃内生细菌HT-6菌株在防治马铃薯晚疫病中的利用潜力,采取延长对峙培养时间以及致病疫霉预先培养后再接种HT-6的方法,对其抑菌特性及持续抑菌能力进行了评价。结果显示,延长对峙培养时间到第15天时,致病疫霉菌落半径为11.0 mm,与对照(40.0 mm)仍有极显著差异;以可溶性淀粉和葡萄糖代替黑麦培养基中的蔗糖,对于抑菌作用无明显影响;HT-6菌液浓度与抑菌效果关系密切,最适浓度分别为OD值1.088和1.928的菌液,其抑菌率分别为95.33%和91.78%,且随OD值升高其抑菌率降低;预先培养致病疫霉10 d后再接种HT-6菌液继续对峙培养21 d,仍可强效抑制致病疫霉生长;镜检发现受抑制的致病疫霉孢子囊数量显著低于对照,菌丝体和孢子囊的畸形率显著高于对照。     

恶疫霉有性杂交后代的生物学研究

 将2个带有不同抗药性标记的恶疫霉菌株配对,培养36 d后诱导其卵孢子萌发,从3760个单卵孢株中,获得50株带有双亲标记的杂交个体。对其中32株杂交个体的生长速率、有性和无性繁殖能力、卵孢子和游动孢子萌发率、致病力及对高温的耐受力等生物学性状进行测定。结果显示,测试的32株杂交个体在LBA培养基上均能较好地生长,有24株的杂交个体的生长速率介于2亲本之间;在LBA和SL培养基上,大多数杂交个体均能产生较大数量的卵孢子,有37%的杂交个体在SL培养基上产卵孢子能力显著大于双亲,仅有1个杂交个体不产生卵孢子;在人工培养条件下,大多数供试杂交个体可以产生较大数量的孢子囊,其中15株杂交个体产孢子囊能力处于双亲之间;被测的杂交个体产生的卵孢子或游动孢子均可以萌发形成有效的单孢株,有22株的杂交个体的卵孢子萌发率大于50%;被测的杂交个体接种在苹果上都有较强的致病力,有16株杂交个体的致病力显著大于双亲。表明在群体水平上恶疫霉有性杂交后代具有较强的生活能力,提示同宗配合恶疫霉不同菌株间的有性重组,对该种的群体遗传多样性可能具重要作用。     

“霜霉威(Propamocarb)”“甲霜灵(Metalaxyl)”对我国辣椒疫霉菌作用方式比较及交互抗性的研究

 通过霜霉威(Propamocarb商品名普力克)、甲霜灵(Metalaxyl商品名瑞毒霉)对辣椒疫霉菌(Phytophthora capsici)6个菌株作用方式的体外测定结果表明,霜霉威浓度在2000ppm时对P. capsici孢子囊和卵孢子的形成,游动孢子的释放、游动及休止孢萌发、菌丝体的生长没有明显的抑制作用。而甲霜灵对P. capsici的各种孢子的产生及菌丝体的生长有较强的抑制作用,对菌丝体生长抑制的Ec₅₀和Ec₉₅分别为0.5596ppm和2.1511ppm;对孢子囊形成抑制的Ec₅₀和Ec₉₅分别为0.1520ppm和15.0032ppm。0.1ppm的甲霜灵可显著抑制卵孢子的生成。但500ppm的甲霜灵对于游动孢子的释放和休止孢萌发没有明显的抑制作用。对霜霉威和甲霜灵的体内活性试验结果表明,800ppm的霜霉威对辣椒疫霉菌的内吸保护防效达94%,1.0ppm的甲霜灵对该病的防效达100%。室内初步试验结果表明,甲霜灵与霜霉威对P. capsici没有交互抗性。     

福建省大豆疫病病原鉴定及其核糖体DNA-ITS序列分析

 从福建省龙海大豆根腐病株上分离的疫霉菌株中,选取6个代表菌株,对病原菌进行了形态特征、致病性、寄主范围鉴定及核糖体DNA-ITS序列分析,结果表明,该菌为疫霉属真菌,在黑麦琼脂培养基上生长缓慢,菌丝致密、无隔,形成菌丝膨大体,近直角分支,分支处稍缢缩。水培后产生大量椭圆形孢子囊,不形成乳突,通过内层出方式产生新孢子囊,游动孢子在孢子囊内形成,同宗配合,藏卵器球形,雄器侧生;接种后可出现典型的大豆疫病症状;人工接种只侵染大豆、豇豆和菜豆等少数豆科植物。其核糖体DNA-ITS序列分析表明,分离菌株与GenBank中大豆疫霉的ITS序列的同源性均为99.8%,仅有2个碱基的差异,结合形态特征和致病性测定,将这些病原菌鉴定为Phytophthora sojae. 这是首次报道大豆疫霉菌在福建省存在。     

“霜霉威(Propamocarb)”“甲霜灵(Metalaxyl)”对我国辣椒疫霉菌作用方式比较及交互抗性的研究

通过霜霉威(Propamocarb商品名普力克)、甲霜灵(Metalaxyl商品名瑞毒霉)对辣椒疫霉菌(Phytophthora capsici)6个菌株作用方式的体外测定结果表明,霜霉威浓度在2000ppm时对P. capsici孢子囊和卵孢子的形成,游动孢子的释放、游动及休止孢萌发、菌丝体的生长没有明显的抑制作用。而甲霜灵对P. capsici的各种孢子的产生及菌丝体的生长有较强的抑制作用,对菌丝体生长抑制的Ec_(50)和Ec_(95)分别为0.5596ppm和2.1511ppm;对孢子囊形成抑制的Ec_(50)和Ec_(95)分别为0.1520ppm和15.0032ppm。0.1ppm的甲霜灵可显著抑制卵孢子的生成。但500ppm的甲霜灵对于游动孢子的释放和休止孢萌发没有明显的抑制作用。对霜霉威和甲霜灵的体内活性试验结果表明,800ppm的霜霉威对辣椒疫霉菌的内吸保护防效达94％,1.0ppm的甲霜灵对该病的防效达100％。室内初步试验结果表明,甲霜灵与霜霉威对P. capsici没有交互抗性。     

Modified agar-based media for culturing Phytophthora infestans

Studies were undertaken to evaluate locally available subtrates for use in a culture medium for Phytophthora infestans (Mont.) de Bary employing a protocol similar to that used for the preparation of rye A agar. Test media preparations were assessed for growth, sporulation, oospore formation, and long-term storage of P. infestans. Media prepared from grains and fresh produce available in Thailand and Asian countries such as black bean (BB), red kidney bean (RKB), black sesame (BSS), sunflower (SFW) and sweet corn supported growth and sporulation of representative isolates compared with rye A, V8 and oat meal media. Oospores were successfully formed on BB and RKB media supplemented with ??-sitosterol. The BB, RKB, BSS and SFW media maintained viable fungal cultures with sporulation ability for 8?months, similar to the rye A medium. Three percent and 33% of 135 isolates failed to grow on V8 and SFW media, respectively.     

Riboflavin activates defense responses in tobacco and induces resistance against Phytophthora parasitica and Ralstonia solanacearum

The effects of riboflavin on defense responses and secondary metabolism in tobacco (Nicotiana tabacum cv. NC89) cell suspensions and the effects of protecting tobacco seedlings against Phytophthora parasitica var. nicotianae and Ralstonia solanacearum were investigated. Defense responses elicited by riboflavin in tobacco cells included an oxidative burst, alkalinization of the extracellular medium, expression of 4 defense-related genes with different kinetics and intensities, and accumulation of 2 total phenolic compounds, scopoletin and lignin. When applied to tobacco plants challenged by P. parasitica and R. solanacearum, riboflavin treatment resulted in 47.9% and 48.0% protection, respectively. These results suggest that riboflavin can both induce a series of defense responses and secondary metabolism in cell suspensions and protect tobacco against P. parasitica and R. solanacearum.     

Response of transgenic cucumber expressing a rice class I chitinase gene to two fungal pathogens with different infectivities

A transgenic cucumber line (CR32) over-expressing the rice class I chitinase gene exhibited resistance to Phytophthora rot (Phytophthora nicotianae var. parasitica) but not to Fusarium wilt (Fusarium oxysporum f. sp. cucumerinum). The infection behavior of these fungi on CR32 and nontransgenic plants was examined with an optical microscope. In zoosporangia of P. nicotianae var. parasitica, the rates of germination and penetration on leaves of both CR32 and nontransgenic plants were almost equal. After infection, however, the growth of infection hyphae was markedly suppressed in CR32 compared with their growth in the nontransgenic plants. In F. oxysporum f. sp. cucumerinum, the infection hyphae localized in petiole vessels of both CR32 and nontransgenic plants, and growth did not differ in the two plants. We investigated the antifungal activity of a high-molecular-weight fraction (HF) and a low-molecular-weight fraction (LF) of crude leaf extracts from CR32 and from the nontransgenic line. CR32 HF, which included the rice chitinase, had antifungal activity only against F. oxysporum f. sp. cucumerinum. In contrast, CR32 LF, which did not have the rice chitinase, had strong antifungal activity against the two fungi. These results suggested that a low-molecular-weight antifungal substance(s) was induced in CR32 and might function as a factor of resistance to P. nicotianae var. parasitica, which has cell walls that almost never contain chitin. Because rice chitinase has already been demonstrated not to localize in vessels of CR32, the infection localization of F. oxysporum f. sp. cucumerinum in vessels might enable the fungus to avoid antifungal substance(s), resulting in Fusarium wilt in transgenic cucumber.     

烟草黑胫病菌培养特性的研究

研究了云南烟草黑胫病菌在不同培养基、不同温度的生长情况及不同菌龄与不同诱导时间孢子囊的产生数量。结果表明,菌株在8种供试培养基中,以燕麦培养基和选择性培养基上菌落生长最好,其次为胡萝卜培养基,在番茄汁和马铃薯培养基上生长最差。同一菌株在不同培养基上产孢量不同、产生的孢子囊大小亦有所不同。05P226菌株在玉米培养基上产孢量最高;05P55菌株在燕麦培养基上产孢量最高,在番茄汁培养基上产生的孢子囊最大,而05P62菌株在燕麦培养基上产孢量最高,在烟叶汁培养基上产生的孢子囊最大。在30℃菌丝生长速率最高,其次为28℃,以35℃生长最差。供试3个菌株间生长差异显著,以05P226生长最好,05P55菌株次之,05P62生长最差。05P226、05P55菌株皆以培养14d、诱导72h产孢量最高,最适宜试验中产孢培养。     

Characteristics and pathogenicity of six Phytophthora isolates from pot plants

From several greenhouse plants showing foot and root rot symptomsPhytophthora isolates were gathered. Isolates fromPeperomia, Saintpaulia andSinningia were identified asP. nicotianae van Breda de Haan var.nicotianae and an isolate fromBegonia asP. cryptogea Pethybr. & Laff. Cardinal temperatures for the various isolates were determined. The specific and non-specific pathogenicity of the isolates was studied by inoculating the different crops with the different isolates, including aP. cryptogea isolate fromGerbera and aP. nicotianae var.nicotianae isolate from carnation. TheP. nicotianae var.nicotianae isolates appeared to be morphologically identical. Some of the isolates were similar in host range, but others exhibited differences in host specificity at 20°C as well as 25, 30 or 35°C. The same applies for the twoP. cryptogea isolates.     

Phosphorous acid treatments control Phytophthora diseases in Australia1

Treatments with a partially neutralized formulation of phosphorous acid containing potassium phosphite were assessed for control of Phytophthora diseases in subtropical and temperate crops in Australia. In Queensland, trunk injections of phosphite (10% solution) controlled severe root rot (Phytophthora cinnamomi) of avocado trees and resulted in the recovery of trees. Single pre-harvest sprays (2.5 kg ha^-1) of phosphite controlled root and heart rot (P. cinnamomi) of pineapples. Foliar sprays of phosphite (64 g per tree) controlled root rot (P. nicotianae var. parasitica) and trunk canker (P. citrophthora) of mandarin trees. In Victoria, a foliar spray of phosphite (300 g ha^-1) reduced root rot (P. clandestina) of subterranean clover and increased dry matter by 1.96 to 5.11 t ha^-1. Trunk injections of phosphite (10% solution) controlled trunk rot (P. cactorum) of peach trees and foliar sprays (10 kg ha^-1) reduced severity of root rot (P. nicotianae var. nicotianae) of tomatoes.     

Control of phytophthora gummosis of citrus with systemic fungicides in Brazil1

Phytophthora gummosis and phytophthora root rot are the most important fungal diseases of citrus in the State of São Paulo, Brazil. Phytophthora citrophthora and P. nicotianae var. parasitica are the main species that incite both diseases in all the citrus-growing areas of the State. Studies on the control of gummosis and root rot with systemic fungicides have been under way in the State since 1980. The efficacy of fosetyl-A1 and metataxyl to control gummosis incited by both fungi in lemon and sweet orange trees was confirmed. The best control was achieved when foliar sprays of fosetyl-A1 were combined with soil applications of metataxyl. The best timing was determined for application of these systemic products to control gummosis in the state.     

Acquired resistance triggered by elicitins in tobacco and other plants

Elicitins are a family of proteins excreted byPhytophthora spp. They exhibit high sequence homology but large net charge differences. They induce necrosis in tobacco plants which then become resistant to the tobacco pathogenPhytophthora parasitica var.nicotianae. In stem-treated plants, resistance was not restricted to the site of elicitin application, but could be demonstrated by petiole inoculation at all levels on the stem. Resistance was already maximum after two days and lasted for at least two weeks. It was effective not only towardsP. p. var.nicotianae infection, but also against the unrelated pathogenSclerotinia sclerotiorum. In contrast to dichloroisonicotinic acid, an artificial inducer of systemic acquired resistance, which was increasingly effective with doses ranging from 0.25 to 5mole per plant, the basic elicitin cryptogein exhibited a threshold effect, inducing near total resistance and extensive leaf necrosis above 0.1 nmole per plant. Between 1 and 5 nmole, acidic elicitins (capsicein and parasiticein) protected tobacco plants with hardly any necrotic symptom. Elicitins exhibited similar effects in various tobacco cultivars andNicotiana species, although with quantitative differences, but induced neither necrosis nor protection in other SolanaceÆ (tomato, petunia and pepper). Among 24 additional species tested belonging to 18 botanical families, only some BrassicaceÆ, noticeably rape, exhibited symptoms in response to elicitins, in a cultivar-specific manner. Elicitins appear to be natural specific triggers for systemic acquired resistance and provide a tool for unraveling the mechanisms leading to its establishment.Abbreviations AR acquired resistance - HR hypersensitive response - INA 2,6-dichloroisonicotinic acid - Ppn Phytophthora parasitica var.nicotianae - SAR systemic acquired resistance     

Effect of high-power monochromatic (pulsed UV laser) and low-power broadband UV radiation on Phytophthora spp. in irrigation water

Cysts or zoospores of Phytophthora capsici, P. citrophthora and P. nicotianae were suspended in distilled water or in recycled irrigation water collected from commercial nurseries. Propagules, suspended in shallow layers of still water in 60 mm-diameter plastic Petri dishes with lids off, were exposed to various UV doses emitted from either an excimer laser or a mercury vapor lamp. The laser emitted a monochromatic (248 nm, 5 eV/pulse), pulsed UV beam with an intensity of 1 to 2 mJ/cm²/pulse with 20-ns pulse durations and a high peak power of 50 to 100 kW/cm²/pulse. The UV lamp was from a commercial water purifier that emitted broadband (continuous) UV radiation at an intensity of ～8 mW/cm². Survival was assessed by culturing aliquots of the treated or non-treated suspensions onto corn meal agar amended with ampicillin (250 ppm) and rifampin (10 ppm), and then counting numbers of developing colonies. The UV dose (as energy per unit area) required to kill propagules was smaller when they were suspended in distilled water than when suspended in recycled nursery water. The reduced kill effectiveness in recycled water appears to be related to UV-absorbing soluble chemicals. Cysts and zoospores were equally susceptible to UV, although the high-peak power pulsed-UV laser source with ultra-short exposure times appeared to have greater kill effectiveness than the conventional Hg-vapor UV lamp. Phytophthora capsici and P. nicotianae isolates were somewhat less sensitive to UV than isolates of P. citrophthora obtained from various hosts and geographical regions. Furthermore, hyphae of Phytophthora spp. were less susceptible to UV than were cysts or zoospores.