Development ofClonostachys rosea and interactions withBotrytis cinerea in rose leaves and residues

The effect of microclimate variables on development ofClonostachys rosea and biocontrol ofBotrytis cinerea was investigated on rose leaves and crop residues. C.rosea established and sporulated abundantly on inoculated leaflets incubated for 7–35 days at 10°, 20° and 30°C and then placed on paraquat—chloramphenical agar (PCA) for 15 days at 20°C. On leaflets kept at 10°C, the sporulation after incubation on PCA increased from 60% to 93% on samples taken 7 to 21 days after inoculation, but decreased to 45% on material sampled after 35 days. A similar pattern was observed on leaves incubated at either 20° or 30°C. The sporulation ofC. rosea on leaf disks on PCA was not affected when the onset of high humidity occurred 0, 4, 8, 12 or 16 h after inoculation. However, sporulation was reduced to 54–58% on leaflets kept for 20–24 h under dry conditions after inoculation and before being placed on PCA. The fungus sporulated on 68–74% of the surface of leaf disks kept for up to 24 h at high humidity after inoculation, but decreased to 40–51% if the high humidity period before transferral to PCA was prolonged to 36–48 h. The growth ofC. rosea on leaflets was reduced at low inoculum concentrations (10³ and 10⁴ conidia/ml) because of competition with indigenous microorganisms, but at higher concentrations (10⁵ and 10⁶ conidia/ml) the indigenous fungi were inhibited. Regardless of the time of application ofC. rosea in relation toB. cinerea, the pathogen’s sporulation was reduced by more than 99%. The antagonist was able to parasitize hyphae and conidiophores ofB. cinerea in the leaf residues. AsC. rosea exhibited flexibility in association with rose leaves under a wide range of microclimatic conditions, and in reducingB. cinerea sporulation on rose leaves and residues, it can be expected to suppress the pathogen effectively in rose production systems.     

The involvement of ethylene and calcium in gray mold of pelargonium,ruscus, and rose plants

Leaves ofPelargonium domesticum andRuscus hypoglossum infected byBotrytis cinerea Pers., produced 12 and 1.5 nl ethylene/h/g, respectively, 34 days after inoculation; wounded or healthy leaves and phyloclades of them produced much lower amounts. When the fungus was grown on dead leaves it produced negligible amounts of the gas even when supplemented with methionine. Exogenous ethylene enhanced gray mold development in both hosts. Silver thiosulfate, aminooxyacetic acid (AOA) and aminoethoxyvinylglycine significantly inhibited disease development in pelargonium, and the latter two compounds inhibited ethylene production. AOA inhibited disease development and ethylene production in cut rose flowers; calcium ions inhibited disease development whereas the chelator EGTA [ethylene glycol bis-(β-aminoethyl ether)N,N,N′,N′-tetraacetic acid] enhanced it. Disease suppression by an excess of Ca²⁺ was correlated with repression of ethylene production by the flowers, whereas deficiency in Ca²⁺ increased disease severity. Publication of the Agricultural Research Organization. No. 2413-E, 1988 series.     

Trichoderma Biocontrol of Colletotrichum acutatum and Botrytis cinerea and Survival in Strawberry

Trichoderma isolates are known for their ability to control plant pathogens. It has been shown that various isolates of Trichoderma, including T. harzianum isolate T-39 from the commercial biological control product TRICHODEX, were effective in controlling anthracnose (Colletotrichum acutatum) and grey mould (Botrytis cinerea) in strawberry, under controlled and greenhouse conditions. Three selected Trichoderma strains, namely T-39, T-161 and T-166, were evaluated in large-scale experiments using different timing application and dosage rates for reduction of strawberry anthracnose and grey mould. All possible combinations of single, double or triple mixtures of Trichoderma strains, applied at 0.4% and 0.8% concentrations, and at 7 or 10 day intervals, resulted in reduction of anthracnose severity; the higher concentration (0.8%) was superior in control whether used with single isolates or as a result of combined application of two isolates, each at 0.4%. Only a few treatments resulted in significant control of grey mould. Isolates T-39 applied at 0.4% at 2 day intervals, T-166 at 0.4%, or T-161 combined with T-39 at 0.4% were as effective as the chemical fungicide fenhexamide. The survival dynamics of populations of the Trichoderma isolates (T-39, T-105, T-161 and T-166) applied separately was determined by dilution plating and isolates in the mixtures calculated according to the polymerase chain reaction (PCR) using repeat motif primers. The biocontrol isolates were identified to the respective species T. harzianum (T-39), T. hamatum (T-105), T. atroviride (T-161) and T. longibrachiatum (T-166), according to internal transcribed spacer sequence analysis.     

Characterization of Laboratory Mutants of Botrytis cinerea Resistant to QoI Fungicides

Mutants of Botrytis cinerea with moderate and high resistance to pyraclostrobin, a Qo inhibitor of mitochondrial electron transport at the cytochrome bc ₁ complex, were isolated at a high mutation frequency, after nitrosoguanidine mutagenesis and selection on medium containing pyraclostrobin and salicylhydroxamate (SHAM), a specific inhibitor of cyanide-resistant (alternative) respiration. Oxygen uptake in whole cells was strongly inhibited in the wild-type strain by pyraclostrobin and SHAM, but not in the mutant isolates. Cross-resistance studies with other Qo and Qi inhibitors (QoIs and QiIs) of cytochrome bc ₁ complex of mitochondrial respiration showed that the mutation(s) for resistance to pyraclostrobin also reduced the sensitivity of mutant strains to other QoIs as azoxystrobin, fluoxastrobin, trifloxystrobin and picoxystrobin, but not to famoxadone and to the QiIs cyazofamid and antimycin-A. An increased sensitivity of pyraclostrobin-resistant strains to the carboxamide boscalid, an inhibitor of complex II, and to the anilinopyrimidine cyprodinil, a methionine biosynthesis inhibitor, was observed. Moreover, no effect of pyraclostrobin resistance mutation(s) on fungitoxicity of the hydroxyanilide fenhexamid, the phenylpyrrole fludioxonil, the benzimidazole benomyl, and to the phenylpyridinamine fluazinam, which affect other cellular pathways, was observed. Study of fitness parameters in the wild-type and pyraclostrobin-resistant mutants of B. cinerea showed that most mutants had a significant reduction in the sporulation, conidial germination and sclerotia production. Experiments on the stability of the pyraclostrobin-resistant phenotype showed a reduction of resistance, mainly in moderate resistant strains, when the mutants were grown on inhibitor-free medium. However, a rapid recovery of the resistance level was observed after the mutants were returned to a selective medium. Studies on the competitive ability of mutant isolates against the wild-type parent strain, by applications of a mixed conidial population, showed that, in vitro, all mutants were less competitive than the wild-type strain. However, the competitive ability of high resistant mutants was higher than the moderate ones. Pathogenicity tests on cucumber seedlings showed that all mutant strains tested exhibited an infection ability similar with the wild-type parent strain. Preventive applications of the commercial product of F-500 25EC (pyraclostrobin) were effective against lesion development on cotyledons by the wild-type, but ineffective, even at high concentrations, against disease caused by the pyraclostrobin-resistant isolates. Boscalid (F-510 50WG) was found equally effective against the disease caused by the wild-type or pyraclostrobin-resistant mutants. This is the first report indicating the appearance of B. cinerea strains resistant to QoI fungicides by the biochemical mechanism of site modification and the risk for field resistance.     

Studies on the Inherent Resistance Risk to Fenhexamid in Botrytis cinerea

After chemical mutagenesis with N-methyl-N-nitrosoguanidine (MNNG) two phenotypes that were highly or moderately resistant to fenhexamid, were isolated from a wild-type strain of Botrytis cinerea, at a mutation frequency of 0.9 × 10^–5. Resistance factors, based on EC⁵⁰ values, were 460–570 and 10–15, respectively. The mutation(s) for resistance to fenhexamid did not affect the sensitivity of mutant strains to the benzimidazole benomyl, the phenylpyridinamine fluazinam, the anilinopyrimidine cyprodinil, the guanidine iminoctadine or to the sterol-biosynthesis-inhibiting fungicides fenarimol, fenpropimorph and tridemorph. On the contrary, an increased sensitivity (EC⁵⁰ ratios of 0.2–0.6) of fenhexamid-resistant strains to the phenylpyrrole fludioxonil and the dicarboximide iprodione was observed. Study of fitness parameters of fenhexamid-resistant isolates of both phenotypic classes showed that these mutation(s) had no effect on mycelial growth and sensitivity to high osmolarity, but they did affect one or more of some other characteristics, such as sporulation, conidial germination and sclerotia production. In tests on cucumber seedlings under greenhouse conditions, all highly fenhexamid-resistant isolates tested presented decreased infection ability compared with the wild-type. Preventive applications of a commercial formulation of fenhexamid, Teldor 50 WP, were effective against lesion development on cotyledons by the wild-type, but ineffective, even in high concentrations, against disease caused by the fenhexamid-resistant isolates. The risk of resistance problems arising during commercial use of fenhexamid is discussed.     

Phenotypic Differences Between vacuma and transposa subpopulations of Botrytis cinerea

One hundred and twenty-one single-spore strains of Botrytis cinerea isolated from Bordeaux vineyards were molecularly characterized as either transposa or vacuma, two subpopulations of B. cinerea distinguished by the presence of transposable elements. Forty-three vacuma and 68 transposa strains were distributed into two main classes (mycelial or sclerotial) by morphological phenotype according to the organ of origin. Strains isolated from overwintering sclerotia produced exclusively sclerotial colonies. The mycelial growth rate of 21 transposa and 13 vacuma strains was significantly influenced by agar-medium and temperature. The mycelial growth rate was significantly strain-dependent at favourable temperatures (15, 20 and 25 °C), but not at limiting ones (5 and 28 °C): vacuma strains showed the fastest growth rates. The strains of the two subpopulations were similar in virulence on both host species tested (Vitis vinifera and Nicotiana clevelandii). The grapevine leaves were significantly more susceptible to B. cinerea than those of tobacco. A significant negative correlation was established between virulence and mycelial growth rate. The epidemiological consequences concerning population structure of B. cinerea in vineyards are discussed.     

Ultrastructural study on the interaction between Physcomitrella patens and Botrytis cinerea

Little is known about the interactions of bryophytes with their pathogens compared with those of flowering plants. This study used the moss Physcomitrella patens and fungal pathogen Botrytis cinerea to investigate their interaction at the ultrastructural level. Infection behaviour of B. cinerea in moss tissues was observed for 1–7 days after inoculation. Some appressoria directly penetrated P. patens cytoplasm through leaf cell walls, and others entered the intercellular spaces of leaves and infected cytoplasm. Infection hyphae were observed in sclerenchyma cells of P. patens stems for the first time. The results demonstrated that the ratio of hyphae‐inoculated leaf cells to leaf numbers could be used to quantify the disease development process at 24 h after inoculation. The ultrastructural studies revealed two responses of P. patens to B. cinerea inoculation: reinforcement of cell walls, including papillae formation; and cell death. This was the first observation of papillae for P. patens–B. cinerea interaction at the ultrastructural level. Papillae in P. patens had a similar ultrastructure to those of higher plants, and may be involved in the defence response to B. cinerea infection. Thus papillae formation is probably an evolutionarily conserved defence mechanism from the early land colonization by plants. Cell death during the P. patens–B. cinerea interaction had some features of programmed cell death, with hydrogen peroxide produced in cytoplasm membranes, suggesting something like a hypersensitive response. These ultrastructural studies suggest that P. patens could be a useful system for studying phytopathogens and could provide theoretical bases for defence mechanisms in evolutionary development.     

Biological control ofBotrytis cinerea in residues and flowers of Rose (Rosa hybrida)

Microbial isolates from living petals, petal residues and leaf residues of rose, and from laboratory collections, were evaluated for control ofBotrytis cinerea in rose. In leaf residues artificially infested withB. cinerea, isolates of the filamentous fungiGliocladium roseum, FR136 (unidentified) andTrichoderma inhamatum reduced sporulation of the pathogen by >90%, other filamentous fungi were 25–90% effective, and those of yeasts and bacteria were <50% effective. In artificially inoculated petal residues, no microbe reduced sporulation ofB. cinerea by >75%, but isolates ofCladosporium oxysporum and four yeasts were 51–75% effective, and three filamentous fungi, eight yeasts andBacillus subtilis isolates were 26–50% effective. Isolates ofT. inhamatum, C. oxysporum andG. roseum performed best againstB. cinerea among isolates evaluated in leaf residues naturally infested with the pathogen and indigenous microorganisms. Totals of ten isolates of filamentous fungi (includingC. oxysporum andC. cladosporioides), two of yeasts and five ofBacillus subtilis completely prevented lesion production byB. cinerea in detached petals, and a further six isolates of filamentous fungi (includingG. roseum) and six yeasts were 90–99% effective. Isolates ofC. oxysporum, C. cladosporioides andB. subtilis, the most effective microorganisms againstB. cinerea in flower buds, reduced number of lesions in the range of 42–65% compared with 59–89% for à standard fungicide (vinclozolin). It is suggested that application of leading antagonists Jo living rose leaves and flowers should optimize control of inoculum production byB. cinerea when the tissues die. Optimal biocontrol of lesion production in flower buds requires a better understanding of the microenvironment of petals.     

Effects of Host and Microbial Factors on Development of Clonostachys rosea and Control of Botrytis cinerea in Rose

Development of Clonostachys rosea in rose leaves and petals and control of Botrytis cinerea by the agent were investigated. C. rosea germinated, established endophytic growth, and sporulated abundantly whether the tissues were mature, senescent or dead when inoculated. Germination incidence was moderate on mature and senescent leaves (47% and 35%) and petals (31% and 43%), and high (>98%) on dead tissues. Sporulation of C. rosea in tissues inoculated when mature, senescent or dead averaged 41%, 61%, and 75% in leaves, and 48%, 87% and 53% in petals. When leaves were wounded with needles before inoculation, germination of C. rosea increased from 45–56% to 90–92%, but sporulation became high (>75%) regardless of wounds. When leaves were inoculated with C. rosea at 0–24h after wounding and subsequently with B. cinerea, germination of the pathogen was reduced by 25–41% and sporulation by 99%. A humid period prior to inoculation of senescent or dead leaves promoted communities of indigenous fungi, reduced sporulation of C. rosea and B. cinerea, and, in dead leaves, increased control of the pathogen associated with C. rosea. Applied at high density, isolates of indigenous Penicillium sp. and Alternaria alternata from rose interacted with C. rosea and reduced control of the pathogen by 16% and 21%, respectively. In conclusion, C. rosea markedly suppressed sporulation of B. cinerea in rose leaves and petals regardless of developmental stage, minor wounds, and natural densities of microflora. This versatility should allow C. rosea to effectively control inoculum production of B. cinerea in rose production systems.     

Relationships of aphid and mite infestations to control ofBotrytis cinerea byClonostachys rosea in rose(Rosa hybrida) leaves

Infestations of aphids(Macrosiphum rosae L.) and of twospotted spider mites(Tetranychus urticae Koch) were examined in relation to growth and sporulation ofClonostachys rosea andBotrytis cinerea, and to suppression of the pathogen by the agent, in green rose leaves. Leaves were infested artificially with 10 aphids/leaflet for 3 h, or naturally with 15-30 aphids/leaflet for 7-12 days or with undetermined numbers of mites for 10-12 days. Leaves that had or had not been infested were inoculated withC. rosea, withB. cinerea, or withC. rosea plusB. cinerea. Germination incidence and germ tube growth ofC. rosea andB. cinerea on the phylloplane in most instances were much greater in leaves previously infested with aphids or mites compared with noninfested leaves. After combined inoculation,C. rosea suppressed germination ofB. cinerea from 47% to 19% in noninfested leaves, but in leaves that had been infested the agent was ineffective and germination incidence of the pathogen increased to 75-93%. Previous infestation with naturally introduced aphids or mites, but not brief infestations of artificially introduced aphids, markedly increased sporulation ofC. rosea after the leaves died during an initial 7-15 days of incubation on a paraquat agar medium, regardless of whether or notB. cinerea was present. Sporulation ofB. cinerea was similarly increased when inoculated alone. After 15-20 days, however, conidiophores of the agent or pathogen covered most of the leaf surface in these treatments. In leaves inoculated withC. rosea plusB. cinerea, the agent suppressed sporulation of the pathogen almost completely in both previously infested and noninfested leaves. Thus, aphid and mite infestations did not compromise the ability ofC. rosea to suppress inoculum production byB. cinerea in the leaves. Increased nutrient availability on the phylloplane through exudation or as honeydew or frass is proposed as a basis to explain effects of the pest infestations onC. rosea andB. cinerea.     

Honey bee dispersal of the biocontrol agent Trichoderma harzianum T39: effectiveness in suppressing Botrytis cinerea on strawberry under field conditions

Botrytis cinerea, which causes grey mould, is a major pathogen of many crops. On strawberry, isolates of Trichoderma spp. can effectively control B. cinerea, but frequent application is necessary. Bees can be used to disseminate biological control agents to the target crop. We tested the ability of honey bees to disseminate Trichoderma harzianum T39 to control B. cinerea in strawberry in the field during the winter in Israel over two consecutive seasons. We used the recently developed ‘Triwaks’ dispenser for loading the bees with the T. harzianum inoculum. During both years, grey mould developed in late January in untreated control plots; at low to medium disease levels it was partially controlled by fungicide treatment, and was best controlled in bee-visited plots. At high disease levels neither chemical nor biological control was effective. To assess the spatial distribution of inoculum by bees, we sampled flowers up to 200 m from the hives and found effective levels of T. harzianum even at 200 m. The approach used in this study provides an effective control of grey mould in strawberry in conditions of low to medium grey mould incidence.     

Survey of mutations of a histidine kinase gene BcOS1 in dicarboximide-resistant field isolates of Botrytis cinerea

Previously, we cloned a putative osmosensing histidine kinase gene (BcOS1) and revealed that a single amino acid substitution, isoleucine to serine at codon 365, conferred dicarboximide resistance in field isolates of Botrytis cinerea. This point mutation (type I) occurred within the restriction enzyme TaqI site of the wild-type BcOS1 gene. Thus, a procedure was developed for detecting the type I mutation of the BcOS1 gene using a polymerase chain reaction (PCR) in combination with restriction fragment-length polymorphism (RFLP). Diagnosis by PCR-RFLP was conducted on the 105 isolates isolated from 26 fields in Japan. All dicarboximide-sensitive isolates (49 isolates) had the wild-type BcOS1 gene, and the 43 isolates with the type I mutation were resistant to dicarboximides without exception. These data indicate that dicarboximide-resistant isolates with type I mutation are widespread throughout Japan. However, other types of dicarboximide resistance were detected among isolates from Osaka; among the 24 resistant isolates from Osaka, 12 had the BcOS1 gene without the type I mutation. BcOS1 gene sequencing of these resistant isolates classified them into two groups, type II and type III. The type II isolates have three amino acid substitutions within BcOS1p (³⁶⁸Val to Phe, ³⁶⁹Gln to His, and ⁴⁴⁷Thr to Ser). The type III isolates have two amino acid substitutions within BcOS1p (³⁶⁹Gln to Pro and ³⁷³Asn to Ser). These amino acid changes are located on the amino acid repeat domain in BcOS1p. The three types of resistant isolates were all moderately resistant to dicarboximides without significant osmotic sensitivity, and their pathogenicity on cucumber leaves was also very similar to that of the wild-type isolate.     

Morphological characterization of the interaction between Diplocarpon rosae and various rose species

Blackspot, caused by Diplocarpon rosae , is the most severe and ubiquitous disease of garden roses, but information is lacking about genotype-specific forms of resistance and susceptibility of the host. Macro- and microscopic analyses of 34 rose genotypes with a defined monoconidial culture black spot inoculum identified susceptible and resistant rose genotypes and further genotype-specific subdivisions, indicating the presence of partial forms of resistance and different resistance mechanisms. In total, eight interaction types were characterized, five representing compatible (types 1–5) and three representing incompatible interactions (types 6–8). The incompatible interactions were characterized by the lack of any visible fungal structures beneath the cuticle (type 8), single-cell necroses (type 7) or necroses of larger cell clusters (type 6), the latter two types with penetration hyphae and haustoria in epidermal cells.     

Relationship between Concentrations of Botrytis Cinerea Conidia in Air, Environmental Conditions, and the Incidence of Grey Mould in Strawberry Flowers and Fruits

Atmospheric concentrations of Botrytis cinerea conidia were monitored for two seasons in a strawberry crop in Moguer (Huelva, southwestern Spain). Concentrations of conidia were estimated using a Burkard volumetric spore sampler. A diurnal pattern of conidial release was observed. Airborne conidial concentration was significantly and positively correlated with the average solar radiation and mean temperature, and negatively with rainfall and relative humidity. Among the weather variables considered, solar radiation showed the most consistent results in the regression analysis, explaining over 40% of airborne conidial concentration variability. Correlation between Botrytis fruit rot incidence and accumulated number of conidia over seven days was significant and positive. Two regression models containing three variables explained over 62 and 52% of the fruit rot incidence variability. A positive but non-significant correlation was established between B. cinerea incidence in flowers and airborne conidial concentration. It was not possible to fit a consistent regression model to relate flower infection incidence to conidial concentration or weather variables.     

Comparative effects of temperature and humidity on the activity of three potential antagonists of rose powdery mildew

Three reported antagonists of cucumber powdery mildew,Stephanoascus flocculosus, Stephanoascus rugulosus, andTilletiopsis washingtonensis, were tested and compared under different environmental conditions for their potential for controlling rose powdery mildew, caused bySphaerotheca pannosa var.rosae. Under controlled conditions in vitro, all three fungi induced a rapid collapse of conidia, conidiophores and hyphae ofS. pannosa var.rosae on detached leaflets of miniature roses within 48 h following their application, as observed under a SEM. Both temperature and relative humidity (r.h.) affected the activity of the antagonists differently. The colonization of powdery mildew was maximal at 26 °C, especially forSt. rugulosus andT. washingtonensis. Maximal colonization was achieved at the highest r.h. tested (90%) for all three antagonists but onlySt. flocculosus maintained a colonization of 80% or better under lower r.h. These observations stress the importance of considering environmental conditions when assessing the activity of antagonistic microorganisms.     

Studies on red light-induced resistance of broad bean to Botrytis cinerea: I. Possible production of suppressor and elicitor by germinating spores of pathogen

When detached broad bean leaves were preinoculated with virulent strain B304 of Botrytis cinerea 24 h before a challenge inoculation with strain B304, lesion formation by B304 was significantly inhibited in red light but not in the dark. In leaves that were preinoculated with avirulent strain 021 and then challenged by B304, however, lesion formation was not inhibited even under red light. Such differences in lesion formation after the challenge inoculation with an avirulent strain were also observed with lesions caused by Alternaria alternata, a nonpathogen of broad bean and by avirulent strain 021R in the presence of germination fluid from spores of strains B304 and 021R. These results suggest the possibility that virulent B. cinerea produced a suppressor involved in induced susceptibility and an elicitor involved in resistance induced by red light during spore germination.     

Role of strawberry volatile organic compounds in the development of Botrytis cinerea infection

Botrytis cinerea, the main pathogen of strawberry, has the ability to remain quiescent in unripe tissue and develop disease symptoms in ripe fruit. As strawberry ripening is characterized by an increase of aroma compounds, the role of volatile emission in the development of infection was investigated. Thirty‐five strawberry volatile organic compounds (VOCs) were tested on B. cinerea in vitro and volatile emission was analysed in strawberry harvested at four ripening stages by headspace solid‐phase microextraction/gas chromatography–mass spectrometry and proton transfer reaction–time of flight–mass spectrometry. The coupling of such data sets made it possible to conclude that key strawberry aroma compounds stimulate B. cinerea conidial germination and some typical wound‐volatiles stimulate pathogen conidial germination or mycelial growth. This study is the first report of fungal stimulation by some VOCs naturally occurring in strawberry: the esters ethyl butanoate, cis‐3‐hexenyl acetate, trans‐2‐hexenyl acetate, methyl butanoate and hexyl butanoate, the furanones furaneol and mesifurane, and the alcohol trans‐2‐hexenol. The results of this work provide advances in understanding the functional role of fruit VOCs and suggest, for the first time, that fruit VOCs may influence the development of B. cinerea from the latent phase and that they could favour the invasive growth of B. cinerea after wounding. In particular, ethyl butanoate and furaneol could signal strawberry ripening, and the green leaf volatiles trans‐2‐hexenol, trans‐2‐hexenyl acetate and cis‐3‐hexenyl acetate could signal the presence of damaged tissues that are easier sites for penetration by B. cinerea.     

Microscopical studies of the effect of metalaxyl on the interaction between sunflower,Helianthus annuus L. and downy mildew,Plasmopara halstedii

The mode of action of the fungicide metalaxyl againstPlasmopara halstedii, the causal agent of sunflower downy mildew, was studied following its application before, during and after artificial contamination of seedlings. The development of the fungus within the treated seedlings was examined microscopically and compared to that occuring in untreated genetically susceptible or resistant genotypes. Hypersensitive-like reactions and necrotic zones leading to the inhibition of fungus growth within the hypocotyl were observed for the three modes of application. This suggests that sunflower defence mechanisms activated by genetical resistance are also involved in the control of downy mildew by metalaxyl.     

LncRNA在柑橘木虱与黄龙病病原菌互作中的差异表达分析及验证

为明确柑橘黄龙病的唯一自然传播媒介——柑橘木虱Diaphorina citri的长链非编码RNA(long non-coding RNA,lncRNA)是否参与调控黄龙病病原菌Candidatus Liberibacter asiaticus(CLas)的侵染及复制,采用生物信息学预测及PCR扩增方法进行lncRNA的预测、特征分析、验证及差异表达分析。结果显示,柑橘木虱的13个转录组RNA-Seq数据中共有10 192个lncRNA基因,对应15 747条lncRNA转录本;与蛋白质编码基因相比,柑橘木虱lncRNA具有更少的外显子数量和更短的转录本长度;随机选取的10条lncRNA基因中,有7条lncRNA基因在无菌柑橘木虱广州品系或赣州品系中有表达,其中1条lncRNA基因TCONS_00034665在无菌广州品系和无菌赣州品系中存在差异表达;带菌和无菌柑橘木虱成虫中预测获得2个差异表达的lncRNA基因TCONS_00096118和TCONS_00234564,实时荧光定量PCR验证发现TCONS_00234564与预测结果一致,在带菌柑橘木虱成虫中高表达。表明lncRNA参与了黄龙病病原菌与寄主柑橘木虱的互作。