不同发酵条件对瓜类细菌性果斑病菌感染西瓜种子的影响

西(甜)瓜果斑病是西瓜嗜酸菌Acidovorax citrulli侵染瓜类作物引起的典型细菌病害,种子带菌是该病最重要的传播方式,防止种子感染是病害预防的首要环节。本研究分析了西瓜种子不同发酵条件对种子感染病菌的影响。研究通过人工接种西瓜发酵液,设置发酵温度为20℃和30℃,检测在不同发酵时间下发酵液pH、A.citrulli菌量、种子外部带菌量、种子内部带菌率,以及出苗发病率,系统地分析比较发酵条件对西瓜嗜酸菌侵染种子的影响。结果表明:从开始发酵到发酵24h,发酵液中A.citrulli菌量快速增长并达到峰值1011cfu/mL,随后逐渐减少;发酵24h的种子外部带菌量最高,达到4×109 cfu/mL;发酵过程中A.citrulli可侵染种皮内部(10%)及种仁(1%);发酵温度对种子感染A.citrulli及幼苗发病率的影响不显著;此外,发酵液菌量与pH相关性显著。以上研究结果表明:控制发酵温度并不能够减少A.citrulli菌的种子感染,减少发酵时间才是减少病菌增殖感染种子的防控关键。     

柑橘溃疡病菌检测方法及其在检疫中的优选

采用常规检测法、免疫斑点法、酶联免疫检测法和PCR检测法对柑橘溃疡病菌纯培养和柑橘叶片提取液进行平行测定。结果表明,对柑橘叶片提取液的检测,4种检测法检出的样品带菌率分别为11.12%、27.22%、18.69%、20.31%;对柑橘溃疡病菌纯培养的检测,4种方法检测结果一致。本实验同时比较了几种方法的灵敏度、特异性和稳定性,以及不同方法在各级检验检疫中的适应性。     

实时荧光定量PCR法检测十字花科细菌性黑斑病菌

为有效防控我国的检疫性有害生物十字花科细菌性黑斑病菌Pseudomonas syringae pv.maculicola在国内的传播与蔓延,通过设计1对特异性引物3539,利用132株靶标和非靶标菌为模板进行PCR扩增,建立了实时荧光定量PCR法,并进行了模拟种子带菌试验。结果显示,引物3539为只针对十字花科细菌性黑斑病菌扩增出的特异性产物;在模拟种子带菌检测中,常规PCR对菌悬液的检测限为10~5CFU/m L,实时荧光定量PCR的检测限为10~3CFU/m L,其中10~8CFU/m L菌液的Ct值最低,为22.90,10~3CFU/m L菌液的Ct值最高,为35.73,且不同浓度菌液间的Ct值均有显著差异;不同带菌率模拟种子的检测结果表明,常规PCR和实时荧光定量PCR能检测到的带菌率分别为0.5%和0.1%。研究表明,实时荧光定量PCR法不仅可用于病种的检测,也可用于病害的早期诊断。     

大白菜软腐细菌对幼苗根系侵入和潜伏侵染的研究

 灭菌土用利福霉素抗性细菌(Ecc RL4)接种后,播种大白菜,幼芽的成苗率降低,且表现矮化和叶色变深的症状。用含利福霉素的改良肉汁胨培养基分离,可以从病苗中回收到病原细菌。组织中病原细菌的含量为10²-10⁷CFU/克不等。病菌可从萌发过程中的芽和整个幼苗期的根部侵入,侵入后,从整个生育期的幼苗直到收获后的菜株内均可回收到病原菌。不同器官的含菌量有显著差别,通常以茎与叶柄的含菌量为高,叶片以半功能叶含菌量为高,根表现不稳定。幼苗带菌率50-100%不等,且随生育期的延长而增高,在播种后第17天均达100%。检测带菌率和腐烂率时,采用了厌气富集技术和含利福霉素的培养基,大大提高了组织内病原细菌的检出率和分离的选择性。     

西瓜细菌性果斑病菌快速免疫PCR检测

对种子携带的西瓜细菌性果斑病菌(Acidovorax avenae subsp. citrulli)进行简单抽提和纯化,不经过DNA提取而以病原菌为模板进行PCR检测.结果显示,对带菌种子提取液采用直接PCR法最低检出限为3600个细菌/mL,而免疫PCR最低检出限可达到600个细菌/mL.免疫PCR法可以有效富集病原后再扩增,方便快速,成本低,灵敏度高,适用于种子携带微量的西瓜细菌性果斑病菌的快速鉴定.     

黄龙病对柑橘叶际微生物组的影响

为明确柑橘叶际微生物组对黄龙病发生的响应规律，采用荧光定量PCR技术检测采集柑橘叶片感染黄龙病菌的情况，通过扩增子测序分析法对PCR检测结果为阴性和阳性叶片的微生物群落进行比较，并通过相关性分析和网络分析对黄龙病菌和叶际微生物的相关性进行解析。结果显示，夏季采集的柑橘叶片中黄龙病菌的检出率为7.5%，而到秋季黄龙病菌的检出率上升到32.3%。黄龙病的发生显著改变了柑橘叶际细菌群落的组成和结构，对于细菌类群的丰度也有一定影响；而对叶际真菌群落的组成和结构无显著影响。黄龙病菌与叶际细菌存在普遍的负相关关系，正相关关系相对较少但相关性更强。黄龙病菌可能通过与厌氧棍状菌Anaerotruncus sp.、梭菌Clostridiales sp.、假单胞菌Pseudomonas sp.、鞘脂单胞菌Sphingomonas sp.、毛螺菌Lachnospiraceae sp.和链球菌Streptococcus sp.的负向互作对柑橘叶际整个细菌群落产生负影响。表明黄龙病发生对柑橘叶际真菌和细菌群落的影响规律不同，同时黄龙病菌可能通过与几种主要细菌的负向互作来实现对叶际微生物组的影响。     

应用Nested PCR技术检测柑桔木虱及其寄主九里香的柑桔黄龙病带菌率

应用PCR及Nested PCR技术检测柑桔木虱及其寄主九里香的结果表明:PCR只可检测最低2头带菌木虱,Nested PCR可检测到单个带菌木虱。100头带菌木虱中,单虫检出率为96％。检测田间重、中等、轻病的柑桔园内的木虱,其带菌率依次为87％、53％和21％。在病芦柑上饲菌不同天数的木虱均能检测出带菌,其饲菌时间最短为1d。城市九里香叶片及在其叶上取食的木虱单虫,均能用Nested PCR检测出病原。饲菌木虱接种九里香及芦柑健苗,在植株尚未表现症状时,常规PCR难检测出病原,但用Nested PCR则能检测到病原,说明九里香不仅是木虱的寄主,而且是黄龙病病原的隐症寄主。     

大白菜根肿病发生特点及影响因子调查研究

通过对大白菜根肿病发生调查、种子带菌率测定、品种抗性鉴定、播期对发病影响等的研究,明确白菜品种、播种时间、前茬作物、白菜生育期、病菌致病性等是影响该病害发生的主要因子.     

大白菜软腐细菌从幼根侵染的组织病理学研究

 将大白菜种子置于培养皿内的滤纸上发芽。滤纸用不同浓度的洗涤与不洗涤的软腐欧氏杆菌(Erwinia carotovora pv.carotovoar Dye)悬浮液,离心上清液,培养基浸出液和灭菌生理盐水浸润。在25℃下培养26小时后,用细菌悬浮液处理的幼芽发病率为8-95%不等。病苗生长量为对照(用生理盐水处理)的17-80%。扫描电镜观察的结果指出,接种后病原细菌首先在根毛区发现,以后才在伸长区和成熟区发现大量病菌。成熟区表皮细胞与细胞交界处的细胞壁有被分解的迹象。在幼苗的导管及其附近的薄壁细胞内均发现了病原细菌,检出率分别为16%和3%。在病苗组织内和幼根表面的病菌可产生微纤丝。病苗幼茎经压气处理24小时后,对其断面进行扫描电镜观察,结果表明,病菌可将导管的外壁与纹孔膜分解而使导管解体仅剩螺蚊加厚部分或呈网孔状。     

薄壳山核桃疮痂病菌TaqMan荧光定量PCR检测方法的建立及应用

疮痂病是薄壳山核桃上最具毁灭性的病害,带菌植物材料是传播疮痂病的重要来源。准确、灵敏、快速的检测方法可为该病害流行规律调查和防控提供有力的依据。本文通过比较薄壳山核桃疮痂病菌Venturia effusa及其近似种之间的ITS序列差异,设计了特异性引物和TaqMan探针,建立了薄壳山核桃疮痂病菌的荧光定量PCR检测方法。特异性检测结果表明,该方法可以检测不同地区的薄壳山核桃疮痂病菌菌株,而对其近似种以及薄壳山核桃上的其他真菌均没有信号。本研究建立的检测方法对薄壳山核桃疮痂病菌DNA的最低检测限可达0.5 pg/μL。该方法用于田间样品检测时,检测时间仅需1 h,远快于常规的分离培养法。本研究建立的基于TaqMan探针的荧光定量PCR检测方法为薄壳山核桃疮痂病菌的快速检测和监测提供了有力工具。     

Dermestid beetles in ‘evolution Canyon’, lower Nahal Oren, Mt. Carmel, including new records for Israel

Fifteen species of dermestid beetles were recorded at ‘Evolution Canyon’ (EC), Lower Nahal Oren, Mt. Carmel, Israel. They represent ~35% of known Israeli dermestid species. The following three species were recorded for the first time in Israel:Trogoderma svriaca Dalla Torre, 1911;Ctesias svriaca Ganglbauer, 1904; andAnthrenus (s.str.) jordaniens Pic, 1934. Adults of 13 species were collected on the more solar radiated, warmer and climatically more fluctuating south-facing slope (SFS); ten species were collected on the opposite, north-facing slope (NFS), which was cooler and climatically more stable. The abundance of adult dermestid beetles was 1.9 times higher on the SFS than on the NFS (86 and 47, respectively). Species richness and abundance distribution at EC (three collecting stations on each slope and one at the valley bottom) were significantly negatively correlated with the plant cover that consisted of trees and bushes (Spearmanr _s ,P=0.007 and 0.039, respectively) and perennials (Spearmanr _s ,P=0.039 and 0.077, respectively), indicating that non-woody plants were preferred by adult dermestid beetles.     

Epidemiology of Toxigenic Fungi and their Associated Mycotoxins for Some Mediterranean Crops

Recent data on the epidemiology of the common mycotoxigenic species of Fusarium, Alternaria, Aspergillus and Penicillium in infected or colonized plants, and in stored or processed plant products from the Mediterranean area are reviewed. Emphasis is placed on the toxigenicity of the causal fungal species and the natural occurrence of well known mycotoxins (aflatoxins, ochratoxins, fumonisins, trichothecenes, zearalenone, patulin, Alternaria-toxins and moniliformin), as well as some more recently described compounds (fusaproliferin, beauvericin) whose toxigenic potential is not yet well understood. Several Fusarium species reported from throughout the Mediterranean area are responsible of the formation of mycotoxins in infected plants and in plant products, including: Fusarium graminearum, F. culmorum, F. cerealis, F. avenaceum, F. sporotrichioides and F. poae, which produce deoxynivalenol, nivalenol, fusarenone, zearalenone, moniliformin, and T-2 toxin derivatives in wheat and other small grains affected by head blight or scab, and in maize affected by red ear rot. Moreover, strains of F. verticillioides, F. proliferatum, and F. subglutinans, that form fumonisins, beauvericin, fusaproliferin, and moniliformin, are commonly associated with maize affected by ear rot. Fumonisins, were also associated with Fusarium crown and root rot of asparagus and Fusarium endosepsis of figs, caused primarily by F. proliferatum. Toxigenic A. alternata strains and associated tenuazonic acid and alternariols were commonly found in black mould of tomato, black rot of olive and citrus, black point of small cereals, and black mould of several vegetables. Toxigenic strains of A. carbonarius and ochratoxin A were often found associated with black rot of grapes, whereas toxigenic strains of A. flavus and/or P. verrucosum, forming aflatoxins and ochratoxin A, respectively, were found in moulded plant products from small cereals, peanuts, figs, pea, oilseed rape, sunflower seeds, sesame seeds, pistachios, and almonds. Finally, toxigenic strains of P. expansum and patulin were frequently found in apple, pear and other fresh fruits affected by blue mould rot, as well as in derived juices and jams.     

A survey of the genera of<Emphasis Type="Italic">Microgaster</Emphasis> latreille and<Emphasis Type="Italic">Hygroplitis</Emphasis> thomson (Hymenoptera: Braconidae: Microgastrinae) from China

The genera ofMicrogaster Latreille 1804 andHygroplitis Thomson 1895 from China are presented systematically in this paper. Thirty-two species ofMicrogaster and three species ofHygroplitis are known in China. Diagnosis, character variation, distribution and host of each species among the two genera are presented, including its host and distribution. Keys to the species ofMicrogaster andHygroplitis are given. http://www.phytoparasitica.org posting Dec. 19, 2006.     

Plant Viruses Transmitted by Whiteflies

One-hundred and fourteen virus species are transmitted by whiteflies (family Aleyrodidae). Bemisia tabaci transmits 111 of these species while Trialeurodes vaporariorum and T. abutilonia transmit three species each. B. tabaci and T. vaporariorum are present in the European–Mediterranean region, though the former is restricted in its distribution. Of the whitefly-transmitted virus species, 90% belong to the Begomovirus genus, 6% to the Crinivirus genus and the remaining 4% are in the Closterovirus, Ipomovirus or Carlavirus genera. Other named, whitefly-transmitted viruses that have not yet been ranked as species are also documented. The names, abbreviations and synonyms of the whitefly-transmitted viruses are presented in tabulated form together with details of their whitefly vectors, natural hosts and distribution. Entries are also annotated with references. Whitefly-transmitted viruses affecting plants in the European–Mediterranean region have been highlighted in the text.     

Broad bean mottle virus in Morocco; curculionid vectors,and natural occurrence in food legumes other than faba bean (Vicia faba)

Broad bean mottle virus (BBMV) was transmitted from infected to healthy faba-bean plants by the curculionid weevilsApion radiolus Kirby,Hypera variabilis Herbst,Pachytychius strumarius Gyll,Smicronyx cyaneus Gyll, andSitona lineatus L. The latter appeared to be an efficient vector: acquisition and inoculation occurred at the first bite, the rate of transmission was c. 41%, and virus retention lasted for at least seven days.S. lineatus transmitted the virus from faba bean to lentil and pea, but not to the three genotypes of chickpea tested. This is the first report on the generaHypera, Pachytychius, andSmicronyx as virus vectors, and onA. radiolus, H. variabilis, P. strumarius, andS. cyaneus as vectors of BBMV.Out of 351 samples of food legumes with symptoms suggestive of virus infection, 16, 11, 19, and 17% of the samples of chickpea, lentil, pea, and common bean, respectively, were found infected when tested for BBMV in DAS-ELISA. This is the first report on the natural occurrence of BBMV in chickpea, lentil, pea, and common bean. The virus should be regarded as a food-legume virus rather than a faba-bean virus solely, and is considered an actual threat to food legume improvement programmes.     

Characterization of potato potyvirus Y (PVY) isolates from seed potato batches. Situation of the NTN, Wilga and Z isolates

A collection of 38 PVY isolates from seed potato batches, originating from several Western European countries, was characterized by using current biological, serological and molecular tools differentiating PVY strains and groups. The correlation between the three kinds of tests was good but not absolute. No single serological or PCR method was able to discriminate among the five isolate groups found. Twenty-nine isolates belonged to the PVY^N strain and six to the PVY^O strain. No PVY^C was found. Two other isolates reacted serologically like PVY^O, but were unable to elicit a hypersensitive response from the Ny_tbr gene and probably represent the PVY^Z group. At the molecular level, these two isolates showed a combination of both PVY^O and PVY^N and could be recombinants of these strains. Another isolate reacted serologically like PVY^O, but induced vein necrosis in tobacco, like PVY^N-Wilga. Some PVY^N isolates caused tuber ring necrosis in glasshouse conditions. These might belong to the PVY^NTN group. The PVY^NTN, PVY^N-Wilga and PVY^Z groups probably represent pathotypes within strains PVY^N and PVY^O, respectively. The present study also confirms previous reports showing a high genetic variation at the 5 end within the PVYN strain.     

The autumn leafroller: Phenology,damage and parasitoids in a Dutch apple orchard

The phenology of the autumn leafroller,Syndemis musculana, a local pest of apple, was studied in order to forecast larval emergence. From 1983–1986, peak flight as determined with sexpheromone traps was always between 13–18 May. The duration of embryonic development was determined at various constant temperatures and used to estimate the periods of egg hatch in these four years. Each year, most eggs should have hatched in the second decade of June.Differences in attack rates between apple cultivars seem to be explained largely by the variation in picking time. Larvae are only half grown at the beginning of harvest (cv. James Grieve), and have gone into hibernation when the latest variety (cv. Golden Delicious) is picked. Moreover, the varieties Cox's Orange Pippin and Belle de Boskoop, picked about half time, are liable to receive additional damage by caterpillars brought with the picked fruits into storage.Various hymenopterous parasites were reared from caterpillars. As the only leafroller in the orchard which hibernates as mature larva,S. musculana may promote winter survival of some parasitoids, like the eulophidColpyclypeus florus.Samenvatting De fenologie van de herfstbladroller (Syndemis musculana Hübner), een incidentele plaag op appel, werd nader bepaald met het doel het uitkomen van de eieren te kunnen voorspellen. In 1983–1986 viel de piekvlucht, bepaald met behulp van feromoonvallen, steeds tussen 13 en 18 mei.De ontwikkelingsduur van de eieren bij verschillende constante temperaturen werd gebruikt om de periode van uitkomen te schatten. De meeste eieren zullen ieder jaar in de eerste helft van juni uitkomen.Geconstateerde verschillen in schade tussen appelrassen blijken goeddeels terug te voeren op verschillen in pluktijdstip. De rupsen van de herfstbladroller zijn pas half-was als de eerste appels eind augustus geplukt worden, terwijl tegen het einde van de oogst begin oktober de meeste al in winterslaap zijn. Met name tussentijdse rassen, als Cox's Orange Pippin and Schone van Boskoop, lopen extra schade op doordat grotere rupsen met de geplukte vruchten in de kist terecht komen.Uit de rupsen werden negen, al van andere boomgaardbladrollers bekende, sluipwespen gekweekt, Omdat deze bladrollersoort, als enige in de boomgaard, als volgroeide rups overwintert, lijkt zij bij uitstek geschikt als winterwaard.This study was carried out at the Experimental Orchard De Schuilenburg, Schuilenburg 3, 4041 BK Kesteren, the Netherlands, to which address correspondence should be addressed.     

Characterization of Sclerotinia minor populations in Texas peanut fields

In the summer of 2004 an epidemic of sclerotinia blight of peanut, a disease caused by Sclerotinia minor, occurred in Texas in fields where the disease was never previously detected. The disease was observed on many plants within one of the fields (>3000 disease foci), although most foci were <1 m. It is hypothesized that these observations were inconsistent with the recent introduction of a monocyclic pathogen, even if disease developed under conducive environmental conditions. The pattern of disease is most suggestive of the presence of foliar (ascospore) infections, although air temperature was above the known limits for apothecia development if the pathogen had arrived in the field in 2004 peanut seed. To further examine this epidemic, 232 isolates were collected, across a variety of spatial scales spanning this field and other Texas peanut fields, and evaluated for aggressiveness, fungicide sensitivity and genotypic diversity. There was wide variation among isolates for the phenotypic characteristics measured, but there was no evidence that a genotypically unique, highly aggressive, and fungicide resistant isolate had been introduced or evolved. The predominant genotype, TX1, which contained 154 isolates, was found in every county and field population.     

Eyespot of Cereals Revisited: ITS phylogeny Reveals New Species Relationships

Four species so far classified in Pseudocercosporella or Ramulispora (hyphomycetes) are associated with eyespot disease symptoms of cereals. Two of these have been linked to teleomorphs that were described in Tapesia. Sequence data derived from the Internal Transcribed Spacer region (ITS1, 5.8S and ITS2) of the rDNA operon showed, however, that the eyespot fungi associated with Tapesia are not congeneric with Ramulispora sorghi, the type of Ramulispora. The genus name Tapesia is now rejected in favour of the conserved name Mollisia, which appears to comprise heterogeneous fungi. Tapesia yallundae is not closely related to the type of Mollisia, M. cinerea, but clusters separately, being more closely allied to species with Cadophora anamorphs. A new holomorph genus, Oculimacula, is therefore proposed for teleomorphs of the eyespot fungi, while the anamorphs are accommodated in Helgardia gen. nov.     

Tumours of Begonia and some other ornamentals,induced by Corynebacterium fascians

In 1975 many tumours were observed in plants ofBegonia Schwabenland grown in Aalsmeer. Submersion of the roots ofNicotiana megalosiphon seedlings in a homogenate of tumorous tissue, induced tumours after two weeks. Short periods of submergence yielded results similar to those obtained after longer periods. Tumour homogenates lost their infectivity after ten min at 50°C. Aphids transmitted the infectious agent.Treatment with propylene oxide did not inhibit infectivity completely. Filtration through a 450 nm filter removed the infectious agent.Tobacco tumor virus or a viroid could not be isolated. Cultures ofCorynebacterium fascians, isolated from tumours ofN. megalosiphon were highly infectious and induced tumours in healthyN. megalosiphon andBegonia. Tumorous tissue homogenates ofPelargonium zonale, Dahlia sp.,Gladiolus sp., andLilium sp. also caused tumours inN. megalosiphon, from whichC. fascians was isolated. It was not possible to produce tumours inN. megalosiphon with homogenates from roses with symptoms of bud proliferation.Samenvatting In 1975 werden vele tumoren waargenomen inBegonia Schwabenland op Aalsmeerse bedrijven (Fig. 1). De infectiositeit van tumorweefsel kon goed en snel worden vastgesteld door de wortels van zaailingen vanNicotiana megalosiphon in een homogenaat van tumorweefsel te dompelen. Tumoren ontstonden na twee weken, de eindbeoordeling geschiedde na een maand (Fig. 2). Ook verschillende andereNicotiana spp.,Melilotus officinalis (Fig. 3) enPisum odoratum (Fig. 4) werden aangetast.Bij de infectiositeitstoets gaven zeer korte dompeltijden even goede resultaten als langere (Tabel 1). Infectieus sap verloor zijn infectievermogen na 10 min verhitting bij 50°C. Bladluizen brachten de smetstof over. Propyleenoxide verminderde de infectiositeit wel, doch onderdrukte deze niet totaal. Bij filtratie door een 450 nm filter bleef het infectieuse agens op het filter achter. Het tumor-inducerende agens was ook aanwezig in die delen van planten met tumoren welke gezond leken en het ging voor een gering deel over met zaad (Tabel 2).Uit tumoren konden wij geen tabakstumorvirus of een viroïde isoleren. Culturen vanCorynebacterium fascians, geïsoleerd uit tumoren vanN. megalosiphon bleken zeer infectieus en veroorzaakten tumoren inN. megalosiphon enBegonia. Homogenaten van tumorweefsel vanPelargonium zonale, dahlia (Fig. 5), gladiool (Fig. 6) enLilium Mid Century Hybrid Enchantment (Fig. 7) veroorzaakten ook tumoren opN. megalosiphon, waaruitC. fascians werd geïsoleerd. Met sap van kroeskopzieke rozen konden wijN. megalosiphon niet besmetten.