Phytophthora kernoviae and P. ramorum: host susceptibility and sporulation potential on foliage of susceptible trees1

Phytophthora kernoviae and P. ramorum are introduced, invasive pathogens in the UK. Both species are adapted for aerial dispersion and have a wide host range, many of which are common to both pathogens. The diseases they cause are foliar necrosis and shoot tip dieback on both tree and ornamental hosts, and bleeding cankers on tree hosts. Inoculum is produced on infected foliage but not on bleeding cankers in both cases. Proactive measures to prevent disease spread and to evaluate the risks posed by these pathogens are being undertaken. Amongst others, these include using the detached leaf assay to get an indication of tree foliage susceptibility, and inoculating wounded stems of saplings to get an idea of under‐canopy sapling and nursery tree susceptibility. The sporulation potential on selected susceptible hosts was assessed, and finally, surveys which are still ongoing were carried out in south‐west England. Results of this work are presented and discussed.     

Susceptibility of Iberian trees to Phytophthora ramorum and P. cinnamomi

The capacity of Phytophthora ramorum to colonize the inner bark of 18 native and two exotic tree species from the Iberian Peninsula was tested. Living logs were wound-inoculated in a growth chamber with three isolates belonging to the EU1 and two to the NA1 clonal lineages of P. ramorum . Most of the Quercus species ranked as highly susceptible in experiments carried out in summer, with mean lesion areas over 100 cm² in Q. pubescens , Q. pyrenaica , Q. faginea and Q. suber and as large as 273 cm² in Q. canariensis , ca . 40 days after inoculation. Quercus ilex ranked as moderately susceptible to P. ramorum , forming lesions up to 133 cm² (average 17·2 cm²). Pinus halepensis and P. pinea were highly susceptible, exhibiting long, narrow lesions; but three other pine species, P. pinaster , P. nigra and P. sylvestris , were resistant to slightly susceptible. No significant difference in aggressiveness was found between the isolates of P. ramorum . In addition, there was evidence of genetic variation in susceptibility within host populations, and of significant seasonal variation in host susceptibility in some Quercus species. The results suggest a high risk of some Iberian oaks to P. ramorum , especially in forest ecosystems in southwestern Spain, where relict populations of Q. canariensis grow amongst susceptible understory species such as Rhododendron ponticum and Viburnum tinus . One isolate of P. cinnamomi used as positive control in all the inoculations was also highly aggressive to Iberian oaks and Eucalyptus dalrympleana .     

Evaluation of a rapid diagnostic field test kit for identification of Phytophthora species, including P. ramorum and P. kernoviae at the point of inspection

Plant health regulations to prevent the introduction and spread of Phytophthora ramorum and P. kernoviae require rapid, cost effective diagnostic methods for screening large numbers of plant samples at the time of inspection. Current on-site techniques require expensive equipment, considerable expertise and are not suited for plant health inspectors. Therefore, an extensive evaluation of a commercially available lateral flow device (LFD) for Phytophthora species was performed involving four separate trials and 634 samples. The assay proved simple to use, provided results in a few minutes and on every occasion a control line reacted positively confirming the validity of the test. LFD results were compared with those from testing a parallel sample, using laboratory methods (isolation and real-time PCR). The diagnostic sensitivity of the LFD (87·6%) compared favourably with the standard laboratory methods although the diagnostic specificity was not as stringent (82·9%). There were a small number ( n  = 28) of false negatives, but for statutory purposes where all positive samples must be identified to species level by laboratory testing, overall efficiency was 95·6% as compared with visual assessment of symptoms of between 20-30% for P. ramorum and P. kernoviae . This work demonstrates the value of the LFD for diagnosing Phytophthora species at the time of inspection and as a useful primary screen for selecting samples for laboratory testing to determine the species identification.     

Chlorophyll fluorescence for visualizing the spatial and temporal spread of Phytophthora alni subsp. alni in alder bark tissue

The impact of alder Phytophthora (Phytophthora alni subsp. alni) on corticular photosynthetic metabolism was explored via measurements of chlorophyll fluorescence. Stem inoculation induced a sharp reduction of maximum (F_v/F_m) and effective quantum yield of PSII (). Observations of the axial and radial spread of the pathogen revealed that near to the point of inoculation and in the whole centre of the stem lesion F_v/F_m and of the cortex chlorenchyma decreased to almost zero, indicating tissue necrosis. Low values of F_v/F_m and were also found in some presymptomatic regions beyond the visible stem lesion. In contrast, substantial photosynthetic activity was found in uninvaded parts of the inoculated trees and in the control. These stem parts showed a marked light‐adapted quantum efficiency of PSII as well as marked electron transport rates in their bark tissues. Thus, corticular photosynthesis stayed unaffected in these stem parts supporting stem carbon balance. Chlorophyll fluorescence measurements in the field illustrated that stem infection with P. alni subsp. alni and the effect on the bark tissues is not only highly heterogeneous but also underlies very quick temporal changes, due to a rapid destruction of the photosynthetic apparatus. The results show that chlorophyll fluorescence measurements and fluorescence imaging are useful indicators of tissue infection caused by Phytophthora infection of bark tissues. Chlorophyll fluorescence measurements can be used to map and visualize the spatial as well as temporal spread of bark pathogens and can give a first indication of invasion of the host tissue beyond the visible lesion.     

Pathogenicity and infectivity of Phytophthora ramorum vary depending on host species,infected plant part,inoculum potential,pathogen genotype,and temperature

A total of 25 ornamental plant species representing 10 families were inoculated using three genotypes, each representing one of the genetic lineages NA1, NA2, and EU1 of the pathogen Phytophthora ramorum. Leaves were inoculated using suspensions with two zoospore concentrations and exposure at three temperatures, while stems were inoculated using agar plugs colonized by mycelia. Susceptibility was determined by measuring either the success of pathogen reisolation or lesion length caused by the pathogen. Infectivity was determined by counting sporangia in washes of inoculated leaves or stems. Results from all three pathogen genotypes combined were used to rank each of the 25 plant species for susceptibility and infectivity, while pooled results per genotype from all 25 hosts combined were employed for a preliminary comparison of pathogenicity and infectivity among genotypes. Statistical analyses showed that leaf results were affected by the concentration of zoospores, temperature, plant host, pathogen genotype, and by the interaction between host and pathogen genotype. Stem results were mostly affected by host and by the interaction between host and pathogen genotype. Hosts ranked differently when looking at the various parameters, and differences in rankings were also significant when comparing stem and leaf results. Differences were identified among the 25 hosts and the three pathogen genotypes for all parameters: results can be used for decision-making regarding regulations or selection of plants to be grown where infestations by P. ramorum are an issue.     

基于去包络线和连续投影算法的枣园土壤电导率光谱检测研究

选取新疆阿拉尔市典型极端干旱区为研究对象,利用土壤高光谱特征对土壤电导率进行反演。为了准确快速检测土壤电导率,通过获取南疆阿拉尔市红枣种植区土壤电导率和高光谱信息,在去包络线处理基础上,分别采用相关性分析法和连续投影算法（SPA）筛选特征波长,并建立特征波长与土壤电导率的偏最小二乘回归模型,使用均方根误差(RMSE)、决定系数（R²）以及相对分析误差（RPD）对不同处理方法的模型效果进行评价。结果表明,基于原始光谱直接使用相关性分析法的预测精度RMSE=0.85566,R²=0.7479,RPD=2.7569;通过去包络线处理使用相关性分析筛选特征波长后,模型的预测精度RMSE=0.44490,R²=0.9500,RPD=6.4510;基于原始光谱使用SPA选择特征波长后,模型的预测精度RMSE=0.31178,R²=0.9707,RPD=8.4445;通过去包络线处理使用SPA选择特征波长后,模型的预测精度RMSE=0.30173,R²=0.9764,RPD=9.3215。综上,说明SPA方法具有较强的特征波长选择能力,基于去包络线处理+SPA的偏最小二乘回归反演模型的预测精度最好,可实现新疆阿拉尔地区土壤电导率的快速检测。     

胶东果园土壤电导率的时空分布特征及Kriging估值

应用传统统计学和地统计学方法,分析了2005年4~11月期间烟台农科院苹果园土壤电导率的空间变异性,并进行了Moran’s I系数分析和Kriging估值。结果表明:土壤电导率具有明显的空间变异性,半方差函数和Moran’s I系数分析说明了4月30日、6月29日和11月16日的空间自相关范围较大,相关性较强;4月20日、7月14日和8月16日的变程偏小,空间相关性较弱。土壤电导率的均值和变异系数随时间变化总体上近似呈现出先增加后减小的趋势,在空间分布上不同时期果园表层土壤电导率分布格局差异较大,土壤电导率的破碎化比较严重。     

宁夏不同类型盐渍化土壤水溶盐含量与其电导率的关系

根据宁夏盐渍化土壤的分布区域,采集了不同类型盐渍化土壤样品(0~20 cm)141个,测定了土壤浸提液电导率与水溶性盐含量及其盐分组成,系统研究了残渣烘干法土壤水溶盐含量与电导率之间的关系。结果如下:(1)土壤水溶性盐分离子阳离子以钠离子含量最高,镁、钙离子含量次之,阴离子以氯离子含量最高,硫酸根离子含量次之;(2)从阴离子组成来看,供试土壤以硫酸盐-氯化物型、氯化物-硫酸盐型和硫酸盐型为主,从阳离子组成来看,以镁钙盐型、钙镁盐型和钠盐型为主;(3)从5种函数模式中筛选出土壤浸提液电导率与其残渣烘干法水溶盐含量之间的最优回归关系,当不区分土壤盐分类型时,可采用二次式(通式)y=0.1609x2+2.9176x-0.0141求解土壤含盐量;当已知土壤盐分类型时,有针对性地选用不同盐分类型最优回归式计算;(4)提出了电导率法测定土壤水溶盐含量校正为残渣烘干法水溶盐含量的关系式。     

生物炭配施沼液对渗出液电导率、全氮含量及土壤理化性质的影响

为增加沼液中养分在土壤中的滞留量,提高沼液利用效率,采用室内土柱试验,设置不同沼液配比(沼液∶水(体积比)分别为1∶4、1∶6、1∶8)、生物炭混掺量(0.0%、0.5%、1.0%、2.0%(土壤质量分数))和生物炭混掺厚度(5、10、15、20 cm),探讨生物炭配施沼液对土柱渗出液电导率、土壤全氮、土壤容重、土壤水稳定性团聚体、土壤总孔隙度、pH值和土壤有机质的影响。结果表明:沼液配比为1∶8时,生物炭配施沼液对全氮淋失无明显作用,沼液配比为1∶6时,渗出液全氮淋失幅度为34.49%～66.79%,沼液配比为1∶4时,渗出液全氮淋失幅度为4.17%～71.67%,即生物炭配施沼液能有效地提高土壤中的全氮含量。随生物炭混掺量的增加,土壤>0.25 mm水稳定性团聚体含量、土壤总孔隙度和土壤有机质含量均有所增加,而随沼液配比的减小均逐渐降低,土壤>0.25 mm水稳定性团聚体较CK增加幅度为8.47%～31.99%,土壤总孔隙度增加幅度为0.76%～3.72%,土壤有机质增加幅度为4.83%～37.17%;土壤容重随土壤中生物炭混掺量和沼液配比的增大而逐渐减小,降低幅度为2...     

Usefulness of single copy genes containing introns in Phytophthora for the development of detection tools for the regulated species P. ramorum and P. fragariae

Introns are generally highly polymorphic regions within genes and were proven to be of great interest for discriminating among phylogenetically-close Phytophthora species. Phytophthora ramorum and P. fragariae are considered as quarantine pathogens by the European Union and accurate detection tools are therefore necessary for their monitoring. From introns located in different single copy genes (GPA1, RAS-like, and TRP1), we developed a series of PCR primers specific to P. ramorum and P. fragariae. The specificity of these primers was successfully checked with a wide collection of Phytophthora isolates and a protocol was developed to detect both pathogens directly in infected plant tissues. These genes should be of particular interest for the development of additional species-specific detection tools within the Phytophthora genus.     

Examination of latent infection in raspberry canes with Phytophthora rubi and P. idaei and transmission in micropropagation

Buds of raspberry canes from naturally and artificially infected root stocks were tested by nested PCR for the latent presence of Phytophthora rubi and P. idaei to establish the risk of introducing these pathogens into micropropagation. Fifteen out of 201 buds tested positive for P. rubi and only 1 out of 176 for P. idaei. These infections were not restricted to buds in the lower part of the cane and mostly do not appear to spread systemically up from the rootstock but seem to be caused by secondary infections of the canes. Buds from symptomless canes from infected root stocks were used for micropropagation and P. rubi could be detected in 5.8% of the obtained microplants after ten generations in one of the two trials. This indicates that micropropagation procedures are not absolutely infallible in eliminating infection and underlines the necessity of a zero tolerance towards diseases in the starting material.     

Epidemiological importance of Solanum sisymbriifolium, S. nigrum and S. dulcamara as alternative hosts for Phytophthora infestans

Lesions of Phytophthora infestans were found on woody nightshade ( Solanum dulcamara ), black nightshade ( S. nigrum ) and S. sisymbriifolium during a nationwide late blight survey in the Netherlands in 1999 and 2000. Pathogenicity and spore production of P. infestans isolates collected from potato ( S. tuberosum ), S. nigrum , S. dulcamara and S. sisymbriifolium were determined on several host plant species, and oospore formation in naturally infected and inoculated foliage of hosts was quantified. The present population of P. infestans in the Netherlands is pathogenic on S. nigrum , S. dulcamara and S. sisymbriifolium . Oospores were produced in leaves of S. nigrum , S. dulcamara and S. sisymbriifolium following infection with A1 and A2 isolates. Therefore these plant species should be regarded as alternative hosts for the late blight pathogen. In the case of S. nigrum and S. dulcamara infection was a relatively rare event, suggesting that diseased plants do not significantly contribute to the overall late blight disease pressure present in potato-production areas. Oospore production in ageing S. nigrum and S. dulcamara plants in autumn, however, may generate a considerable source of (auto) infections in following years. Considerable numbers of sporangia and oospores were produced on S. sisymbriifolium following infection with P. infestans . Additional field infection data are needed to evaluate the epidemiological consequences of a commercial introduction of S. sisymbriifolium as a potato cyst nematode trap crop.     

Genetic diversity,sensitivity to phenylamide fungicides and aggressiveness of Phytophthora ramorum on Camellia,Rhododendron and Viburnum plants in Spain

Phytophthora ramorum has been detected in official plant health surveys on Rhododendron, Viburnum and Camellia in ornamental nurseries in northern Spain since 2003. A collection of 94 isolates of P. ramorum was obtained from 2003 to 2008 from plants with symptoms at different geographical locations. Isolates were identified based on morphology and sequence of the rDNA ITS region. Mating type, genetic variation, sensitivity to phenylamide fungicides and aggressiveness of these isolates were determined. All isolates belonged to the A1 mating type, ruling out the possibility of genetic recombination. Seven microsatellite markers were used to study genetic diversity; three out of the seven microsatellite markers were polymorphic within the Spanish population of P. ramorum. This study confirms that all Spanish isolates of P. ramorum belonged to the EU1 lineage. Twelve intralineage genotypes were detected, five that are unique to Spain (EU1MG38, EU1MG41, EU1MG37, EU1MG39 and EU1MG40) and seven that are also present in at least one other European country (EU1MG1, EU1MG29, EU1MG22, EU1MG13, EU1MG2, EU1MG18 and EU1MG26). Genotypes EU1MG37, EU1MG39 and EU1MG40 were isolated from Rhododendron from one region; EU1MG38 and EU1MG41 were isolated from Camellia from two different regions. Isolates of genotype EU1MG38 were resistant to metalaxyl and mefenoxam. The level of genetic diversity within the Spanish population of P. ramorum is limited and indicates a relatively recent clonal expansion.     

Effect of burning and high temperature on survival of Xanthomonas translucens pv. pistaciae in infected pistachio branches and twigs

This paper reports the efficacy of burning and heat‐treating pistachio branches and twigs as a means of disposing of prunings from trees infected with Xanthomonas translucens pv. pistaciae (Xtp). Burning of pistachio wood, naturally infected with Xtp, was conducted twice under field conditions. Viable Xtp was detected in some non‐burned wood, but not in charcoal, ash or partially burned wood. Controlled laboratory experiments were conducted with pure cultures of Xtp and naturally and artificially infected pistachio wood. In liquid culture, 65°C was lethal to Xtp, whereas survival at 60°C or less varied with culture medium and duration of exposure. Xtp survived in infected wood exposed to 40–55°C for at least 60 min but was killed by exposure to 60°C for 15 min or more. Overall, the results of burning and heat treatment were consistent, and confirmed that burning was a reliable eradication technique to dispose of infected wood, such as prunings, providing the pathogen was exposed to a temperature of 60°C or greater for at least 15 min.     

Effect of cutting frequency on above‐ and belowground biomass production of Rumex alpinus,R. crispus,R. obtusifolius and the Rumex hybrid (R. patienta × R. tianschanicus) in the seeding year

Rumex species are important weeds in grasslands and on arable land. The Rumex hybrid (R. patienta × R. tianschanicus; cv. OK‐2, Uteusha) has been planted as a forage and energy crop since 2001 in the Czech Republic, but its ecological requirements and its potential to become a new weedy species have never been investigated. In 2010 and 2011, we performed a pot experiment to investigate the effect of none, one and two cuts per year on biomass production of Rumex OK‐2 and common broad‐leaved Rumex species (Rumex obtusifolius, R. crispus and R. alpinus). The higher cutting frequency can reduce the belowground biomass, but no effect on the aboveground biomass was detected. Flowering in the seeding year was recorded in only 50% of R. obtusifolius plants. Non‐flowering R. obtusifolius plants produced significantly more belowground biomass than flowering plants under no cutting or one cut treatments. The growth response of Rumex OK‐2 to different cutting treatments was very similar to R. crispus. These similarities indicate the weed potential of the hybrid to become a troublesome weedy species, similar to R. crispus.     

Aetiology and toxigenicity of Fusarium graminearum and F. pseudograminearum causing crown rot and head blight in Australia under natural and artificial infection

Globally fusarium head blight (FHB) of wheat is predominantly caused by Fusarium graminearum (FG) and crown rot (FCR) by F. pseudograminearum (FP). While both FG and FP can cause FHB in Australia, the reasons why the morphologically and culturally similar FG is not a major FCR pathogen has remained elusive. Using aetiology and toxigenicity, this study clarifies the contrasting roles of FG and FP in FCR and FHB in Australia. Naturally infected wheat from 42 sites during 2010 FHB epidemics, and wheat inoculated with either pathogen to induce FCR or FHB at three field plantings in 2011, were used to determine pathogen prevalence and deoxynivalenol (DON) content of the crown, stem base, stem top, rachis and grain. As the primary aetiological agent, FP prevalence in the crown correlated with FCR severity while FG in grain and/or the rachis correlated with FHB severity. FG was an effective colonizer of the crown and stem base but colonization was symptomless. DON content was linked to FG biomass in all tissues except the crown, where FP biomass was the main contributor. Of the 30 measures derived to analyse pathogen fitness in 2011, 10 described the superior fitness of FG for FHB; six defined FP fitness for FHB including inoculum dispersal; and eight defined FCR fitness of both FP and FG. FG had superior FHB fitness but weak saprophytic survival may have undermined its FCR fitness.     

Development of a real‐time PCR method for the detection of the dagger nematodes Xiphinema index,X. diversicaudatum,X. vuittenezi and X. italiae,and for the quantification of X. index numbers

The ectoparasitic dagger nematodes Xiphinema index and Xiphinema diversicaudatum, often at low numbers in the soil, are vectors of grapevine nepoviruses, which cause huge agronomical problems for the vineyard industry. This study reports a method, based on real‐time PCR, for the specific detection of these species and of the closely related non‐vector species Xiphinema vuittenezi and Xiphinema italiae. Specific primers and TaqMan probes were designed from the ribosomal DNA internal transcribed spacer 1 (ITS1), enabling the specific detection of single individuals of each of the X. index, X. diversicaudatum, X. italiae and X. vuittenezi species whatever the nematode population. The specificity of detection and absence of false positive reaction were confirmed in samples of each species mixed with the three other Xiphinema species or mixed with nematodes representative from other genera (non‐plant‐parasitic Dorylaimida, Longidorus sp., Meloidogyne spp., Globodera spp. and Pratylenchus sp.). The method was shown to be valid for the relative quantification of X. index numbers through its use, from crude nematode extracts of soil samples, in a greenhouse assay of grapevine accessions ranging from highly susceptible to resistant. As an alternative to time‐consuming microscopic identification and counting, this real‐time PCR method will provide a fast, sensitive and reliable diagnostic and relative quantification technique for X. index nematodes extracted from fields or controlled conditions.