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1.
Apple proliferation (AP), caused by ‘Candidatus Phytoplasma mali’, is an economically important disease affecting many apple-growing areas in Europe. A new TaqMan real-time PCR assay was established for absolute quantification of ‘Ca. P. mali’ by using a single-copy gene of the host plant as a reference, which is amplified with the pathogen DNA in a single-tube reaction. Normalised estimates of phytoplasma concentration are ultimately expressed as the number of phytoplasma cells per host plant cell. The assay was used to monitor the ‘Ca. P. mali’ titre over the course of two growing seasons in roots and branches of symptomatic and asymptomatic but AP-positive apple trees. All 252 root samples from symptomatic and asymptomatic trees tested positive, with an average number of 59.8 ± 5.68 (standard error) and 55.1 ± 9.83 ‘Ca. P. mali’ per host cell, respectively. From the 378 shoot samples analysed, 81% of the symptomatic and only 20% of the asymptomatic samples were AP-positive with an average number of 9.4 ± 1.04 and 0.7 ± 0.13 ‘Ca. P. mali’ per host cell, respectively. This strengthens evidence that not the pathogen occurrence alone but the presence of a certain quantity of ‘Ca. P. mali’ in the aerial tree sections is involved in symptom expression. In addition, pronounced seasonality of the phytoplasma concentration was found, not only in branches, but also for the first time in roots of symptomatic and asymptomatic apple trees. Highest phytoplasma levels in roots were detected from December to May.  相似文献   

2.
In the present study, the pathogenicity of 36 isolates of Guignardia species isolated from asymptomatic ‘Tahiti’ acid lime fruit peels and leaves, ‘Pêra-Rio’ sweet orange leaves and fruit peel lesions, and a banana leaf were characterized. For pathogenicity testing, discs of citrus leaves colonized by Phyllosticta citricarpa under controlled laboratory conditions were kept in contact with the peels of fruit that were in susceptible states. In addition, pathogenicity was related to morphological characteristics of colonies on oatmeal (OA) and potato dextrose agar (PDA). This allowed the morphological differentiation between G. citricarpa and G. mangiferae. Polymerase chain reactions (PCRs) were also used to identify non-pathogenic isolates based on primers specific to G. citricarpa. A total of 14 pathogenic isolates were detected during pathogenicity tests. Five of these were obtained from leaf and fruit tissues of the ‘Tahiti’, which until this time had been considered resistant to the pathogen. Given that the G. citricarpa obtained from this host was pathogenic, it would be more appropriate to use the term insensitive rather than resistant to categorize G. citricarpa. A non-pathogenic isolate was obtained from lesions characteristic of citrus black spot (CBS), indicating that isolation of Guignardia spp. under these conditions does not necessarily imply isolation of pathogenic strains. This also applied to Guignardia spp. isolates from asymptomatic citrus tissues. Using fluorescent amplified fragment length polymorphism (fAFLP) markers, typically pathogenic isolates were shown to be more closely related to one another than to the non-pathogenic forms, indicating that the non-pathogenic isolates display higher levels of genetic diversity.  相似文献   

3.
‘Candidatus Liberibacter solanacearum’ is associated with the Zebra Chip (ZC) disorder of potatoes. A panel of eight simple sequence repeat (SSR) markers was developed and used to genetically characterize ‘Ca. L. solanacearum’ strains obtained from ZC-affected potato plants in the United States and Mexico. The multilocus SSR markers in this study effectively differentiated genotypes and estimated genetic diversity of ‘Ca. L. solanacearum’ strains. Genotype assignment analyses identified two major lineages of ‘Ca. L. solanacearum’ in the North American populations while only one lineage type was identified in Mexican population. No clear genetic structure was found among haplotypes based on geographical proximity or host. The high resolution power of the SSR marker system developed in this study provides a useful tool for genotyping closely related strains and tracking sources of the pathogen. Genotype information combined with epidemiological data will advance knowledge of ZC disease and will facilitate development of effective disease management.  相似文献   

4.
Zebra chip, an emerging disease of potatoes, has recently been associated with ‘Candidatus Liberibacter solanacearum’ in New Zealand. The phloem-limited bacterium is known to be vectored by the tomato potato psyllid (Bactericera cockerelli). In this study, the role of tuber transmission in the spread of Ca. L. solanacearum was investigated by re-planting potato tubers infected with Ca. L. solanacearum in the absence of the psyllid. Nested PCR demonstrated that Ca. L. solanacearum could be transmitted from the mother tubers both to the foliage of growing plants and to progeny tubers, resulting in symptomatic and asymptomatic plants. Of 62 Ca. L. solanacearum-infected tubers four did not sprout symptomatic of zebra chip. A further two plants developed foliar symptoms associated with zebra chip during the growing season and died prematurely. Fifty-six of the infected tubers produced asymptomatic plants, although Ca. L. solanacearum was detected in the foliage of 39 of them indicative of transmission into asymptomatic progeny plants. At harvest, Ca. L. solanacearum was found in the daughter tubers of only five of the 39 asymptomatic plants, and only one of these plants was found to have zebra chip symptoms in the daughter tubers. Our results show that tuber transmission of Ca. L. solanacearum could play a role in the life cycle of this pathogen, providing a source for acquisition by Bactericera cockerelli and for movement of the pathogen to other regions of New Zealand via transport of seed tubers.  相似文献   

5.
‘Candidatus Liberibacter asiaticus’ is the most widespread of the three species of ‘Ca. Liberibacter’ that cause citrus greening disease (huanglongbing). To ascertain the phylogenetic relationships among Indian isolates that have higher diversity in the 16S rDNA than Asian isolates of this species, we collected symptomatic leaves from Northeast India, Papua New Guinea and Timor-Leste (East Timor) and detected ‘Ca. L. asiaticus’ by PCR using primers specific for nusG–rplK genes and 16S rDNA. Phylogenetic analysis with 16S rDNA sequences and single nucleotide polymorphisms of the omp gene region revealed that the Northeast Indian isolates were genetically closer to Asian-common isolates from Japan, Taiwan, and Vietnam than to Indian isolates reported previously. Thus, the Asian-common strains of ‘Ca. L. asiaticus’ are apparently also present in Northeast India.  相似文献   

6.
Epidemiology ofVerticillium dahliae on olive trees (cvs. ‘Barnea’ and ‘Souri’) irrigated with saline water was studied over a period of 3 years. Disease incidence in cv. Barnea increased from 18.9% to 40.5% in the first year, while disease severity decreased from 4.2 to 2.4 (on a scale of 2–10). In the first month after pruning of diseased branches (June 1999), disease incidence was 26.9% and it increased to 60.6% by May 2000. No changes in disease severity were observed from this time until the end of the study. In cv. Souri, disease incidence doubled during the 3 years of the study (from 22.1% to 44.8%), and disease severity increased from 2.4 to 2.9. The disease caused significant reduction of yield in cv. Barnea in 1999 and 2000 (48.3% and 12.0%, respectively). In cv. Souri yield was measured only in 2001, where a reduction of 66.8% was observed.V. dahliae was isolated from both diseased and symptomless trees. A significant decrease in the isolation rate ofV. dahliae from diseased cv. Barnea trees occurred between the first year (47.9%) and the third year (5.3%); positive isolations were obtained in all seasons, without differences between seasons. In cv. Souri, the fungus was isolated in significantly higher rates during the winter and spring (55% and 50%, respectively). Our findings strongly support the assumption that natural recovery occurs in cv. Barnea 4–5 years after planting, but not in cv. Souri. http://www.phytoparasitica.org posting July 10, 2003.  相似文献   

7.
Phoma macdonaldii is one of the most important pathogens of sunflower (Heliantus annuus) in France. In order to determine the inheritance of resistance to the disease, five sunflower genotypes with wide genetic variability for resistance to two ‘collar’ and two ‘root’ Phoma isolates were crossed in a diallel programme. Four separate experiments were undertaken under controlled conditions. In each one, the response of parental genotypes and their F1 hybrids were evaluated with one of the four Phoma isolates. Analysis of variance was performed to determine the effects of genotype on disease severity score when inoculated with ‘collar’ or ‘root’ Phoma isolates and showed significant variability among parents and F1 hybrids for disease severity score. Diallel analysis showed that general combining ability (GCA) and specific combining ability (SCA) effects for resistance to ‘collar’ and ‘root’ Phoma isolates were highly significant for each of the four isolates indicating that both kinds of gene effects were important in controlling the resistance. The GCA/SCA ratios were more than one for three out of four isolates showing that additive genetic effects were more important than non-additive effects for resistance to three of the studied Phoma isolates. Hence, conventional breeding methods could be recommended to achieve genetic improvement to such ‘collar’ and ‘root’ Phoma isolates.  相似文献   

8.
Citrus Huanglongbing (HLB) is a devastating disease of citrus known to be associated with a fastidious, phloem-limited Gram-negative, yet to be cultured bacterium in the genus Candidatus Liberibacter. In the present study we have developed a method to quantify viable Candidatus Liberibacter asiaticus (Las) with the aid of ethidium monoazide (EMA) which can differentiate live from dead cells. First, calibration curves were developed with the aid of quantitative real-time PCR (QPCR) by using a plasmid template consisting of a 703 bp DNA fragment of rplKAJL-rpoBC (β-operon) region. Standard equations were then developed to quantify Las genome equivalents in citrus, periwinkle, and Asian citrus psyllid, Diaphorina citri. To overcome the limitation of quantitative PCR in discriminating between live and dead bacterial cells, EMA was used to inhibit the amplification of DNA from the dead cells of Las in plant samples. By using the standard equations and EMA-QPCR methods developed in this study, we found that the proportion of viable cells in citrus and periwinkle ranged from 17–31% and 16–28%, respectively. It was determined that a minimum bacterial concentration is required for HLB symptom development by quantifying the population of Las in symptomatic and asymptomatic leaves. The EMA-QPCR methodology developed in the present study should provide an accurate assessment of viable HLB pathogen, providing a tool to investigate disease epidemiology and thus act as a crucial component for disease assessment and management. The authors P. Trivedi and U. S. Sagaram contributed equally to this work.  相似文献   

9.
Citrus huanglongbing (HLB), associated with Candidatus Liberibacter asiaticus and Ca. L. americanus and transmitted by the Asian psyllid Diaphorina citri, is the most serious disease of citrus worldwide because of crop devastation and difficulty to control. Since 2004, approximately 3 million trees were eliminated in attempts to limit its spread in Brazil. Where HLB becomes endemic, the disease progression in the orchard and the increasing symptom severity throughout the tree canopy can be relatively fast, greatly reducing the economic life of affected orchards because of tree decline and yield reduction. Although the majority of the fruit from symptomatic branches drop before harvest, a significant amount of affected fruit remain attached, are available for harvest, and can affect juice quality. To quantify and compare the effects of HLB on fruit quality of the most important sweet orange cultivars grown in São Paulo State, 4–6 year-old sweet orange trees from 26 blocks (two of ‘Valencia Americana’, eight of ‘Hamlin’, four of ‘Westin’, seven of ‘Pera’, and five of ‘Valencia’) were selected prior to harvest. In each block, 14–21 HLB-symptomatic trees were chosen. In each tree, the quality of 20 fruit normal in appearance from asymptomatic branches and 20 symptomatic fruit from symptomatic branches were assessed. In general, compared to normal fruit, the symptomatic fruit were small, light, more acidic, and had lower juice percentage, Brix, total soluble solids per box, total soluble solids per fruit, and Brix/acidity ratio. These effects of fruit quality were less pronounced on early and mid season sweet orange cultivars than on late season cv. Valencia.  相似文献   

10.
The objectives of this work were (i) to determine the influence of temperature on infection of citrus by ‘Candidatus Liberibacter asiaticus’ and ‘Candidatus Liberibacter americanus’, the two bacterial species associated with citrus huanglongbing (HLB) in Brazil, and (ii) to determine the influence of temperature on citrus colonization by ‘Ca. L. asiaticus’, which has taken over from ‘Ca. L. americanus’ as the predominant species in Brazil since 2008. Two experiments were carried out with graft‐inoculated Valencia oranges on Rangpur lime rootstocks. Immediately after inoculation the plants were maintained for 423 days in growth chambers under the following night/day temperature conditions: 17/22, 22/27 or 27/32°C, with a dark/light photoperiod of 8/16 h. Infection and colonization of plants were determined using quantitative PCR (qPCR). ‘Candidatus Liberibacter americanus’ did not infect the plants maintained at 27/32°C; however, infection by ‘Ca. L. asiaticus’ occurred at all studied temperatures. Two months after inoculation, ‘Ca. L. asiaticus’ was distributed throughout the inoculated plants, with mean Ct values in the range of 30–31 for leaves and 25–28 for roots. Over time, ‘Ca. L. asiaticus’ reached the highest titres in mature leaves (mean Ct value = 26·7) of citrus plants maintained at 22/27°C. ‘Candidatus Liberibacter asiaticus’ colonization of citrus plants was negatively affected by the daily temperature regime of 27/32°C (mean Ct value in mature leaves = 33·6).  相似文献   

11.
Two surveys (2005/2006 and 2009) were conducted in the state of São Paulo, Brazil, to investigate the incidence of ‘Candidatus Liberibacter asiaticus’ and ‘Ca. L. americanus’, two liberibacters associated with citrus huanglongbing (HLB) disease and both transmitted by Diaphorina citri, in orange jasmine (Murraya exotica), a widespread ornamental tree in cities and villages. The graft‐transmissibility of the two species, and their DNA relatedness to citrus‐associated liberibacters, were also investigated. Quantitative PCR was applied to PCR‐positive orange jasmine and HLB‐positive citrus growing in backyards and orchards to assess their inoculum source potentials. Liberibacters were detected in 91 of 786 sampled orange jasmine plants in 10 of 76 sampled locations. PCR‐positive trees exhibited yellow shoots and/or dieback symptoms indistinguishable from those on PCR‐negative trees. ‘Candidatus Liberibacter americanus’ was more common in 2005/2006 (96·6%) and ‘Ca. L. asiaticus’ in 2009 (84·8%). rplJ nucleotide sequences were identical within all populations of either species. Graft transmission succeeded only in homologous host combinations, including ‘Ca. L. americanus’ (2/10) from/to orange jasmine and ‘Ca. L. americanus’ (5/18) and ‘Ca. L. asiaticus’ (5/9) from/to citrus. Symptoms were mild and developed less rapidly in orange jasmine than in citrus, probably as a result of lower liberibacter multiplication rates. Respective titres of ‘Ca. L. americanus’ and ‘Ca. L. asiaticus’ in orange jasmine averaged 4·3 and 3·0 log cells g?1 tissue, compared with 5·5 and 7·3 in citrus. The results indicate that orange jasmine does not favour liberibacter multiplication as much as citrus. However, its importance in HLB epidemics should not be underestimated as it is a preferred host of D. citri and is not under any strict tree‐eradication programme or measures for insect control.  相似文献   

12.
Candidatus Phytoplasma prunorum was detected for the first time in almond (Prunus dulcis Mill.) cv. ‘Abiod’ in Tunisia. Infected trees showed emergence of new growth during dormancy and leafed out before flowers opened in addition to early defoliation in summer. Phytoplasma was detected by nested polymerase chain reaction (PCR) using universal phytoplasma primer pairs P1/P7 and F2n/R2. A band with expected size was observed in samples collected from five symptomatic, but not symptomless almond trees. PCR products (1.2 kbp) were used for restriction fragment length polymorphism (RFLP) analysis after digestion with endonucleases RsaI and SspI. RFLP patterns obtained were similar to those reported previously for the European stone fruit yellows (ESFY, 16SrX-B). Identification has been further confirmed by PCR using ESFY specific primer pairs (ECA1/ECA2). This is the first report of Ca. Phytoplasma prunorum infecting almonds in Tunisia.  相似文献   

13.
The spider mites Tetranychus urticae Koch and Panonychus ulmi (Koch) (Tetranychidae) cause severe economic losses to vegetable farms and deciduous fruit orchards in Turkey. One of their predators, the ladybird beetle Stethorus gilvifrons (Muls.) (Col., Coccinellidae), aggregates on mite-infested patches of plants. The present study assessed whether there is a role for herbivore-induced plant volatiles (HIPVs) and/or odors emitted directly from these two mite species in the aggregative response of ladybird beetles. The olfactory responses of the predator females to volatiles from T. urticae- and/or P. ulmi-infested sweet pepper (four cultivars, viz. ‘Demre’, ‘Yalova Carliston’, ‘Kandil Dolma’ and ‘Yag Biberi’), kidney bean (cv. ‘Barbunya’) and apple (M9 rootstock) were investigated using a two-choice olfactometer. Our results showed that HIPVs emitted from both T. urticae- and P. ulmi-infested plants significantly attracted S. gilvifrons adults for all plants except the sweet pepper cv. Yag Biberi. In addition, it was found that volatiles from apple plants infested by T. urticae and, especially, P. ulmi are more attractive for S. gilvifrons females than those emitted by other infested plants. The results also suggest that the odors of T. urticae adults and their products might influence the attraction of S. gilvifrons females.  相似文献   

14.
In order to accelerate breeding and selection for disease resistance to Fusarium wilt, it is important to develop bioassays which can differentiate between resistant and susceptible cultivars efficiently. Currently, the most commonly used early bioassay for screening Musa genotypes against Fusarium oxysporum f. sp. cubense (Foc) is a pot system, followed by a hydroponic system. This paper investigated the utility of in vitro inoculation of rooted banana plantlets grown on modified medium as a reliable and rapid bioassay for resistance to Foc. Using a scale of 0 to 6 for disease severity measurement, the mean final disease severities of cultivars expressing different levels of disease reaction were significantly different (P ≤ 0.05). Twenty-four days after inoculation with Foc tropical race 4 at 106 conidia ml−1, the plantlets of two susceptible cultivars had higher final disease severities than that of four resistant cultivars. Compared with ‘Guangfen No.1’, ‘Brazil Xiangjiao’ is highly susceptible to tropical race 4 and its mean final disease severity was the highest (5.27). The plantlets of moderately resistant cultivar ‘Formosana’ had a mean final disease severity (3.53) lower than that of ‘Guangfen No.1’ (4.33) but higher than that of resistant cultivars: ‘Nongke No.1’, GCTCV-119, and ‘Dongguan Dajiao’ (1.87, 1.73, and1.53, respectively). Promising resistant clones acquired through non-conventional breeding techniques such as in vitro selection, genetic transformation, and protoplast fusion could be screened by the in vitro bioassay directly. Since there is no acclimatization stage for plantlets used in the bioassay, it helps to improve banana breeding efficiency.  相似文献   

15.
In a study of date fruit damage caused byOligonychus spp., we investigated whether the cultivar affects phenology, and on what hosts the mites over-winter. Samples were taken from ‘Deglet Noor’, ‘Barhi’ and ‘Medjool’ trees from mid-April through mid-September during the years 1999–2002. In the ground-cover mites were monitored by collecting Bermuda grass (Cynodon dactylon) under each sampling tree. Over 99% of the mites collected on Deglet Noor and Barhi fruit were identified asO. afrasiaticus. Mean population levels ofO. afrasiaticus reached ten mites or more (initiation of infestation) on Medjool in the second half of May, whereas on Deglet Noor this did not occur before the first week of July. On Barhi the initiation of infestation varied between plots and years, ranging from the second half of May to the beginning of July, but always occurred earlier than Deglet Noor. Mite populations on the pinnae remained low from June through October, not exceeding seven mites per pinna, whereas on fruit strands they reached peak populations of approximately 4000 mites per strand. The sex ratio (proportion of females) ofO. afrasiaticus on fruit of all three cultivars was highly female-biased, usually above 0.85. During winter,O. afrasiaticus was found on Bermuda grass in the orchard ground-cover as well as on fronds of all three cultivars. http://www.phytoparasitica.org posting Jan. 30, 2003.  相似文献   

16.
Eleven soybean cultivars with different levels of susceptibility to virulent isolate SG915 ofCalonectria ilicicola were examined for reaction to metabolites produced by the isolate. When the culture filtrate from isolate SG915 was applied to trifoliates from 11 cultivars, cvs. ‘Cajun’ and ‘Asgrow 7986’ exhibited reduced wilting severity. However, there was no correlation between sensitivity to culture filtrate and susceptibility to the fungal isolate. Wilting severity on cv. ‘Riverside 699’ was greatest when trifoliates were treated with culture filtrates from isolates SG915 (highly virulent) and C31 (less virulent). The dilution end-point for culture filtrates of virulent isolate SG915 was determined to be 1:8. Nonautoclaved culture filtrates caused complete wilt of soybean trifoliates after 36 h, but autoclaved culture filtrates demonstrated a reduced ability to wilt leaves. Electrolyte leakage from treated leaf tissues increased over time regardless of the concentrations of culture filtrate tested. The greatest electrolyte losses were observed during the initial 30 min incubation of leaf tissues. The highest concentration of culture filtrate (50%, v/v) induced more electrolyte loss than the low concentration (10%, v/v) or control. These results suggest that toxic metabolites ofC. ilicicola may be involved in disease development with leaf symptom expression.  相似文献   

17.
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18.
The occurrence of Asian grey weevil Myllocerus undatus (Curculionidae: Coleoptera) damaging coconut (Cocos nucifera L.) seedlings is reported for the first time from Kerala, India. A mild to moderate level of infestation damaging 5–10% leaf lamina of un-split leaves with a typical notching-like symptom along the leaf margins is noticed on the majority of the coconut seedlings belonging to different coconut varieties, viz., ‘Chowghat Orange Dwarf’, ‘Chowghat Green Dwarf’, ‘Malayan Green Dwarf’, ‘West Coast Tall’ and Dwarf x Tall hybrids. M. undatus has not been reported as a destructive pest on any of the crops from India and is considered to be a pest of quarantine significance. Adult weevils have a characteristic tri-spined hind femur and elytra strongly angled broader than the prothorax.  相似文献   

19.
The phylogeny of ‘Candidatus Phlomobacter fragariae’(Ca. P. fragariae), the agent of the strawberry marginal chlorosis (SMC), and the proteobacterium associated with the low sugar content syndrome of sugar beet (SBRp) is not well understood. The spoT-spoU-recG genetic locus initially characterised by genome walking from a ‘Ca. P. fragariae’ partial spoT sequence was used to determine relatedness of ‘Ca. P. fragariae’ and SBRp with bacteria detected in hemipteran insects. Both plant pathogenic bacteria belong to the same phylogenetic group as bacteria of the arsenophonus clade detected in hemipteran insects. The SBRp is closely related to arsenophonus-like proteobacteria from cixiids and more distantly related to psyllid and delphacid secondary endosymbionts, whereas the relatives of ‘Ca. P. fragariae’ remain to be discovered. No genetic variability was found among isolates of ‘Ca. P. fragariae’ or SBRp. Implications for explaining the emergence of both ‘Ca. P. fragariae’ and SBRp as epidemic plant pathogens are discussed.  相似文献   

20.
Apple proliferation caused by ‘Candidatus Phytoplasma mali’ is a disease of apple trees gaining increasing importance in Europe. The present study describes a high-throughput method for simultaneous typing of ‘Ca. P. mali’ at two genetic loci. This novel approach combines terminal restriction fragment length polymorphism (T-RFLP) analysis of a putative rhodanese-like protein gene and the analysis of the variable number of tandem repeats (VNTR) of the ribosomal protein L22 gene. The typing approach was applied to analyse a collection of DNA isolates from 310 apple trees tested positive for ‘Ca. P. mali’. Samples were taken between 2002 and 2010 in South Tyrol (Northern Italy). In addition, 15 samples of Cacopsylla melanoneura and 19 of C. picta were typed. Seven combined genetic profiles were found in the samples of infected apple trees: AT-2/rpX-A (81.0%), AT-1/rpX-D (8.4%), AT-1/rpX-E (4.2%), AT-1/rpX-A (3.2%), AT-1/rpX-B (1.6%), AT-1/rpX-C (1.0%) and AP/rpX-A (0.3%), and one mixed infection AP + AT-1/rpX-A + rpX-D (0.3%). Subtype rpX-E was discovered for the first time. In C. melanoneura samples the most frequent subtype was AT-1/rpX-E, followed by AT-1/rpX-D and AT-1/rpX-C. All C. picta samples displayed subtype AT-2/rpX-A. Analysis of the temporal distribution of subtype frequencies in apple trees revealed that exclusively subtype AT-1 in combination with four rpX subtypes was present in South Tyrol in the period from 2002 to 2004. From 2006 onwards subtype AT-2/rpX-A became dominant with an average frequency of 90%. The data obtained suggest that there may be a co-adaptation of particular ‘Ca. P. mali’ subtypes with different insect vector species.  相似文献   

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