Blister Spot Research Articles

WITHDRAWN: Categorising Apple Fruit Diseases Employing Hybrid Neural Clustering Classifier

Agricultural field plays a major role in development of India. The farmers are supplemented with additional profit by exporting fruits from India. The devastating problem in economic losses and production in agricultural industry is caused by diseases in fruits. Manual identification of defected fruit is time consuming and a difficult approach. Based on the pictures representing the symptoms of the fruit, Web-based system helps non experts also in the identification of the fruit disease. In this paper ten types of apple fruit diseases are classified by using the proposed Hybrid Neural Clustering (HNC) Classifier. The results of the feature extraction vector is supplied as the input to the classifier. There are two phases in this proposed method .The first phase is used to train the images by using the K Means Clustering to cluster the vector points and followed up with the second phase which is used to test by applying the Feed Forward Back propagation neural network (FFBP). The output received is the whole set of images that are classified into the various apple fruit diseases. The various diseases classified are Alternia Rot, Black Rot, Scab, Grey Mould, Bitter Rot, Blister Spot, Brown Rot, White Rot, Blue Mould, Powdery Mildew. The proposed method is evaluated by precision, recall and F Measure and it scored higher values than existing methods. Proposed method provides 98% of accuracy than other existing method.

Exoenzymes as a pathogenicity factor for Colletotrichum gloeosporioides associated with coffee plants

ABSTRACT Phytopathogenic fungi during the penetration and colonization process are capable of secreting several enzymes, which enable infection of the host live tissue, acting on the degradation of wax, cuticle and cell walls. The ability of a pathogenic agent to produce enzymes or not can determine the severity degree of a disease. In this study, 33 isolates of Colletotrichum gloeosporioides related to anthracnose and blister spot on coffee trees were evaluated for their ability to produce hydrolytic enzymes (amylase, lipase, protease, laccase, pectinase and cellulase) and specific cell wall degrading enzymes “CWDEs” (polygalacturonase, polymethylgalacturonase and pectin-lyase), as well as their relationship with the pathogenicity/aggressiveness of isolates. For all isolates of C. gloeosporioides, extracellular enzymes could be detected, except cellulases. Isolates I-9 and I-24 produced the highest levels of extracellular enzymes, as well as CWDEs. They also had the highest disease intensity indexes, suggesting a relationship between enzymes and aggressiveness of the isolates.

Investigating variability and behaviour of Colletotrichum gloeosporioides strains from lesions of coffee blister spot

AbstractCoffee blister spot has been associated with species from the Colletotrichum genus, but there is no information on the variability of isolates present on leaf lesions. This study evaluated a population of Colletotrichum gloeosporioides strains from blister spot lesions in Coffea arabica. Colletotrichum spp. isolates were collected from blister spot lesions on leaves of coffee trees from Catuaí and Topázio cultivars (Coffea arabica). Monosporic cultures were obtained from colonies with sporulation. A pathogenicity test was carried out by inoculation of pathogens on the leaves of young coffee plants. C. gloeosporioides strains were characterized by morphologial, cytological and physiological analyses. The molecular analysis was carried out using Inter‐Retrotransposon Amplified Polymorphism (IRAP) markers. C. gloeosporioides strains showed no pathogenicity on coffee plants and presented a wide variability in all traits evaluated. The presence of sexual strains, formation of CATs (conidial anastomosis tubes) among conidial strains and high mycelial compatibility among strains observed suggest the occurrence of sexual and asexual recombination. The role of these C. gloeosporioides strains on the lesions of coffee plant leaves is unclear.

Phytotoxic Metabolites Produced by Diaporthella cryptica, the Causal Agent of Hazelnut Branch Canker.

From the culture filtrates of Diaporthella cryptica, an emerging hazelnut pathogen, 2-hydroxy-3-phenylpropanoate methyl ester and its 3-(4-hydroxyphenyl) and 3-(1 H-indol-3-yl) analogues, named crypticins A-C, were isolated together with the well-known tyrosol. Crypticins A-C were identified by spectroscopic (essentially nuclear magnetic resonance and high-resolution electrospray ionization mass spectrometry) methods. The R absolute configuration (AC) of crypticin A was determined by comparing its optical rotation and electronic circular dichroism (ECD) spectrum with those of papuline, the methyl ester of (-)( S)-phenyllactic acid isolated as the main phytotoxin of Pseudomonas syringae pv. papulans, responsible for apple blister spot. The ACs of crypticins B and C were determined by time-dependent density functional theory calculations of their ECD spectra. Papuline and the new metabolites herein isolated, except tyrosol, were tested at 1 mg/mL on cork oak, grapevine, hazelnut, and holm oak leaves using the leaf puncture assay. They were also tested on tomato cuttings at 0.5 and 0.05 mg/mL. In the leaf puncture assay, none of the compounds was found to be active. Crypticin C and papuline were active in the tomato cutting assay. Additionally, crypticin C displayed moderate inhibitory effect against Phytophthora cambivora.

Colletotrichum on coffee trees in south of minas gerais -Brazil: physiological, molecular and pathogenic description

Fungi of the genus Colletotrichum spp. are cosmopolitan and are responsible for disease in many plants of agronomic importance. In coffee crops, three species have been described: Colletotrichum acutatum, Colletotrichum kahawae and Colletotrichum gloeosporioides. In Brazil were only reported the species C. acutatum and C. gloeosporioides, which symptoms are attributed as antracnoses, dieback and blister spot. This study evaluates the population of Colletotrichum spp. through morpho-physiological, genetic and pathogenic characteristics, in several coffee plantations in the southern of Minas Gerais. All isolates showed similar morphological and physiological characteristics, being identified as C. gloeosporioides. The best medium for mycelial growth was agar-malt extract with pH 5.5 and for spore production oat-agar medium with pH 6.5. When inoculated on berries, all isolates were pathogenic with variable levels of aggressiveness, but only 57% of these were pathogenic for hypocotyls. Three isolates, I-9, I-24 and I-26, were identified featuring the highest rates of aggressiveness in berries and hypocotyls.

Viability and susceptibility of propagation material from coffee plants to Colletotrichum sp.

This study aimed to verify the viability of propagation material from coffee plants descended from germplasm susceptible to blister spot disease as well as its susceptibility to Colletotrichum sp. relative to commercial coffee cultivars. In the first experiment, fruits were harvested from plants with and without symptoms of blister spot and sowed in trays containing a commercial sterilized substrate. The percentages of germinated seeds, viable plantlets and seedlings were evaluated. In diseased tissues, pathogens were isolated and identified though a pathogenicity test. In the second experiment, ten commercial cultivars and one cultivar originating from plants with blister spot were inoculated with the pathogens to assess the severity of anthracnose. Significant differences were not observed with respect to seed germination. However, the viability of plantlets and seedlings was reduced in the cultivar originating from plants with blister spot (Genotype Originated from Diseased Plants-GODP). These plants showed characteristic symptoms of blister spot, including necrosis in the leaves and hypocotyls, wilting and death. In the necrotic lesions, we observed characteristic sporulation of Colletotrichum sp. The cultivar most susceptible to anthracnose in cotyledonary leaves was Catuaí Vermelho (GODP), which presented the highest area under the disease progress curve (AUDPC). In conclusion, the viability of propagation material from coffee plants that had descended from plants with symptoms of blister spot (GODP) was reduced compared with plants from other genotypes, although seed germination was not affected. Moreover, GODP species are more susceptible to Anthracnose on the cotyledonary leaves relative to the other analyzed commercial cultivars. This work is the first to report on different symptoms exhibited by seedlings originating from the seeds of plants with symptoms of blister spot.

GFP-fluorescent protein on the study of blister spot in coffee plants

In Brazil, Colletotrichum gloeosporioides is associated with a complex of symptoms in coffee culture. Although this pathogen had its pathogenesis observed and identified, its importance has still been questioned due to its several endophytic forms, raising doubts as to the real importance of the pathosystem. The aim of this study was to demonstrate, by using an isolate transformed with the gene gfp, the infection and colonization capability of C. gloeosporioides in coffee seedlings. After the fourth day of inoculation, manifestation of symptoms as punctual necrosis could be observed, which progressed during the evaluation period, culminating in the death of seedlings. Epifluorescence microscopy confirmed the presence of the pathogen in the seedlings, as well as the visualization of internal colonization of tissues, acervulus formation and conidium production, confirming that it was responsible for the observed symptoms.

English

Blister Spot is a disease transmitted through seeds. This characteristic of the disease reveals the studies of severity can offer more replies than the studies of incidence. Due to the inexistence of standardized for visual quantification of the Blister Spot’s severity, the aim of this work was to build up and validate a diagrammatic scale for evaluating the severity of Blister Spot in coffee tree. Leaves in the field were collected with different intensities of symptoms disease, and electronically determined the real severity. Based on the frequency distribution of the severity values, and according to the law of visual stimulus of Weber-Fechner, the maximum and minimum limits, and the intermediate levels of the scale were defined. The validation was realized by eight evaluators that estimated the severity in 50 leaves with different intensities of symptoms. An evaluation without the aid of the diagrammatic scale was realized, and two others with its use, having intervals of seven days. The accuracy, precision, repeatability and reproducibility of the estimate were evaluated. The developed scale shows seven 0 (0%), 1 (0.1-3%), 2 (3.1-6%), 3 (6.1-12%), 4 (12.1-25%) and5 (≥25.1%). Using the scale proposed, the evaluators presented better levels of accurancy, precision, reproducibility and repeatability in the estimate, once compared to the evaluators who did not use the diagrammatic scale. The diagrammatic scale was adjusted to aid in the visual estimate of the severity of the Blister Spot in coffee leaves   Key words: Coffea arabica, Colletotrichum gloeosporioides, pathometry.

Nutrients distribution in diseased coffee leaf tissue

Knowing the structure and distribution of nutrients in plant tissues can clarify some mechanisms of pathogen attack in plants and plant defense against infection, thus helping management strategies. The aim of this study was verify differences in distribution of mineral nutrients in coffee leaf tissues around foliar lesions of bacterial blight of coffee, blister spot, cercospora leaf, phoma leaf spot and coffee leaf rust. Fragments of leaf tissue surrounding the lesions were dehydrated in silica gel, carbon covered and subjected to X-ray microanalysis (MAX). Thirty-three chemical elements were detected in leaf tissue; however, there was variation in potassium and calcium contents surrounding the lesions. The highest potassium content was found in asymptomatic tissues surrounding the lesions, decreasing toward the transition zone and reaching minimum content in symptomatic tissues. The highest calcium content was found in symptomatic tissues, decreasing toward the transition zone and reaching minimum content in asymptomatic tissues. Therefore, MAX can be used to analyze the composition and distribution of nutrients in plant tissues and, if associated with mineral nutrition, it may help understand host-pathogen relationships and plant disease management.

Genetic transformation with the gfp gene of Colletotrichum gloeosporioides isolates from coffee with blister spot

Genetic transformation with the gfp gene of Colletotrichum gloeosporioides isolates from coffee with blister spot

Efeito de Colletotrichum gloeosporioides Penz, agente etiológico da mancha manteigosa, na germinação e viabilidade de sementes de cafeeiro

Foi avaliada a influência de Colletotrichum. gloeosporioides na sanidade, germinação, viabilidade e vigor de sementes de cafeeiro. Sementes das cultivares Catuaí Vermelho, provenientes de plantas com sintomas de mancha manteigosa, e de lavouras sadias de Catuaí Vermelho IAC 44 e Mundo Novo 379-19 foram submetidas a inoculação por C. gloeosporioides, pelo contato direto com a colônia do fungo, por 0; 24; 48; 72; 96 e 120 horas. As variáveis analisadas foram sanidade, germinação, índice de velocidade de emergência e viabilidade pelo teste de tetrazólio. Verificou-se aumento da incidência de C. gloeosporioides com o do tempo de exposição ao fungo, sendo esta de 26% em Catuaí Vermelho, 21,5% em Catuaí Vermelho IAC-44 (21,5%) e de 11,0% em Mundo Novo IAC 379-19. O fungo afetou a germinação das sementes e o índice de velocidade de emergência das sementes da cultivar Catuaí Vermelho e Catuaí Vermelho IAC 44 a partir de 96 horas de exposição. O efeito de C. gloeosporioides na germinação e no estabelecimento de plântulas de cafeeiro é variável, dependendo da cultivar utilizada, da presença do pergaminho e do tempo de exposição ao fungo, mas a viabilidade das sementes decresce com o aumento do tempo de exposição ao fungo.

Genetic transformation with the gfp gene of Colletotrichum gloeosporioides isolates from coffee with blister spot.

Blister spot (Colletotrichum gloeosporioides) is now widespread in most coffee producing states of Brazil, becoming a limiting factor for production. The lack of data relating to the reproduction of typical symptoms (light green, oily patches) leaves a gap within the pathosystem, forcing the search for new methodologies for monitoring the disease. Monitoring of genetically modified organisms has proven to be an effective tool in understanding the host × pathogen interactions. Thus, the present study was carried out to evaluate the effectiveness of two systems of genetic transformation in obtaining mutants using the gfp reporter gene. Using the two transformation systems (PEG and electroporation) revealed the efficiency of both, confirmed by fluorescence microscopy and resistance to the antibiotic hygromycin-B, when incorporated into the culture medium. The fungus maintained its cultural and morphological characteristics when compared to wild strains. When inoculated on coffee seedlings, it was found that the pathogenicity of the processed isolates had not changed.

Eventos do processo de infecção de Colletotrichum gloeosporioides inoculados em folhas de Coffea arabica L

O presente trabalho teve por objetivo obter informações sobre os eventos de pré-penetração, penetração e colonização de isolados de C. gloeosporioides, obtidos de mangueira e cafeeiro, quando inoculados em folhas de cafeeiros (Coffea arabica L.) da cultivar Catucaí Vermelho. As folhas foram selecionadas, padronizadas e lavadas, demarcando-se áreas circulares de 0,5 cm de diâmetro na face abaxial, inoculando-se uma alíquota de 20 µL da suspensão de conídios. Utilizou-se um isolado obtido de mangueira e dois isolados obtidos de cafeeiro com mancha manteigosa. Realizaram-se avaliações com 3, 6, 8, 12, 24, 36, 48, 72, 96, 144 e 240 horas após a inoculação (hai). Todos os materiais foram processados e observados em microscópico eletrônico de varredura. Os conídios de todos os isolados aderiram freqüentemente nas depressões das células da epiderme e células-guarda dos estômatos, formando septo antes da germinação. A penetração, na maior parte, se deu por via direta e algumas vezes por estômatos. Isolados de cafeeiro germinaram em folhas de 6 a 8 hai, produzindo apressórios 12 hai e acérvulos de 96 a 144 hai. O isolado de mangueira germinou de 6 a 8 hai com formação de apressório de 8 a 12 hai e produziu novos conídios diretamente em hifas conidiogênicas. Não foi observada a formação acérvulos para este isolado.

Sensibilidade de Colletotrichum gloeosporioides (mancha manteigosa do cafeeiro) a diferentes concentrações de fungicidas

Com o objetivo de avaliar a eficiência de alguns fungicidas sobre Colletotrichum gloeosporioides, agente etiológico da mancha manteigosa do cafeeiro (Coffea arabica L.), testes in vitro foram conduzidos no Laboratório de Diagnose e Controle/UFLA. Utilizou-se o método de incorporação de fungicidas ao meio de cultura MEA 2% para a avaliação da inibição do crescimento micelial e em lâmina escavada contendo água com fungicida para a germinação de conídios. Os fungicidas, tetraconazol, triadimenol, chlorotalonil e mancozeb foram testados quanto à inibição do crescimento do micelial (nas concentrações de 1, 5, 10, 25, 50, 100, 500 e 1.000 mg L-1) e quanto à inibição da germinação de conídios (nas concentrações de 1, 5, 10, 25, 50 e 100 mg L-1). Os fungicidas tetraconazol e triadimenol apresentaram alta eficiência na inibição do crescimento micelial. Os fungicidas chlorotalonil e mancozeb mostraram baixa eficiência e ineficiência, respectivamente. Quanto à germinação dos conídios, os fungicidas que demonstraram maior eficiência em baixas concentrações foram o chlorotalonil e o tetraconazol.

Estudos histoplásticos da interação Colletotrichum gloeosporioides: cafeeiro

Nos estudos envolvendo a interação Colletotrichum gloeosporioides-cafeeiro (Coffea arabica L.), poucas são as informações a respeito do modo de penetração e colonização deste patógeno. Estudou-se por meio da microscopia eletrônica de varredura (MEV), os eventos de pré-penetração e penetração de C. gloeosporioides em hipocótilos de cafeeiros e a colonização natural de ramos e frutos provenientes de plantas com mancha manteigosa. Realizaram-se três ensaios. No primeiro, fez-se a observação dos eventos de pré-penetração de C. gloeosporioides inoculados em hipocótilos de plântulas da cv. Acaiá Cerrado com e sem ferimentos; no segundo, observou-se a colonização de ramos enfermos em que havia murcha drástica e necrose local e, no terceiro, a colonização de frutos enfermos. A germinação dos conídios nos hipocótilos feridos ocorreu 6 horas após a inoculação, com a formação de um ou dois tubos germinativos terminais e a adesão dos conídios nas depressões dos hipocótilos. Apressórios globosos a subglobosos e de contorno regular surgiram 12 horas após a inoculação. Até 72 horas, não foi possível observar a formação de acérvulos sobre os tecidos submetidos à inoculação. Nos ramos observava-se colonização nos vasos floema e parênquima cortical. Nos frutos em diferentes estádios de maturação foi observada a colonização do tecido próximo à epiderme e colapso dos estômatos na área lesionada.

Evaluation of the antifungal activity by plant extracts against Colletotrichum gloeosporioides Penz

Aiming to develop more efficient and environmental friendly methods than those available to control Colletotrichum gloeosporioides Penz, which causes blister spot in coffee trees, a search for plants able to produce substances active against such pathogen was carried out. Thus, extracts of 48 plant species, collected at Alto Rio Grande region, in Minas Gerais, were prepared and submitted to in vitro assays with that fungus. The best results were obtained with the extracts prepared from Digitalis lanata Ehrh, Origanum manjorona L., Plantago lanceolata Hook. and Stevia rebaudiana (Bertoni) Bertoni, which inhibited C. gloeosporioides spores germination. After dilution of some active extracts with aqueous 1 % Tween 80 solution in a 1:2 or 1:3 ratio (extract:aqueous solution), their antifungal activity vanished. Some of the active extracts were also submitted to freeze drying and none of them presented any alteration in their antifungal activity. Concluding, several plants presented potential to be used in the search for new bioactive substances to control C. gloeosporioides, especially O. manjorona L., which inhibited 96 % of the fungus spores germination.

Efeito da temperatura no crescimento micelial, produção e germinação de conídios de Colletotrichum spp. isolados de Coffea arabica L

Espécies do gênero Colletotrichum são agentes etiológicos de importantes doenças do cafeeiro, porém, ainda pouco estudadas no Brasil. Diante desse fato, objetivou-se com este trabalho foi estudar o efeito da temperatura no crescimento, produção e germinação de conídios de diferentes isolados de Colletotrichum spp. Foram avaliados 8 isolados, obtidos de folhas com sintoma de mancha manteigosa, seca de ponteiros e assintomático. Os isolados apresentaram comportamento diferenciado na velocidade de crescimento micelial, capacidade de esporulação e na formação de estruturas reprodutivas, com alta capacidade adaptativa em diferentes temperaturas.

Rapid Diagnosis of Pseudomonas syringae pv. papulans, the Causal Agent of Blister Spot of Apple, by Polymerase Chain Reaction Using Specifically Designed hrpL Gene Primers

ABSTRACT The identification and detection of Pseudomonas syringae pv. papulans, the causal agent of blister spot of apple, on apple leaves and fruit was achieved by polymerase chain reaction amplification of a specific DNA fragment of the hrpL sequence. The consensus primers hrpL(1) and hrpL(2) were designed based on the alignment of pseudomonad hrpL gene sequences available in nucleic acid data banks. This primer set produced a 631-bp amplicon from 37 of the 57 pseudomonads strains tested. These strains belonged to genomospecies 1 and 2, as described by Gardan et al. (8). The amplicon obtained from 30 of these strains was digested with eight restriction enzymes. Three different restriction patterns were produced from strains belonging to genomospecies 1, resulting in A1 and A2 patterns, while strains belonging to genomospecies 2 were characterized by a B pattern. Patterns A1 and A2 differed at only two sites, a Bsp 143I site located at nucleotide 360 and a MseI site located at nucleotides 22-24. Group A2 consisted solely of P. syringae pv. papulans strains. The hrpL gene in P. syringae pv. papulans strain CFBP3323 was sequenced. Two primer sets, Pap1/Pap2 and Pap1/Pap3, were designed and tested for specificity to P. syringae pv. papulans. These primers amplified expected fragments of 242 and 303 bp, respectively. Pap1/Pap2 amplified a fragment only with P. syringae pv. papulans DNA, while Pap1/Pap3 amplified all tested strains belonging to genomospecies 1. A diagnostic procedure using the Pap1/Pap2 primer set was successful for the detection of P. syringae pv. papulans in diseased fruit and artificially inoculated leaves.

Epiphytic occurrence of Pseudomonas syringae pv. papulans (Rose) in France, where blister spot has never been seen

Pseudomonas syringae pv. papulans (PSP) the causal agent of blister spot, on the apple cultivar Mutsu in the USA, Canada and Italy, has not been described in France. A study on epiphytic populations of P. syringae isolated from French apple orchards revealed two isolates called KA54 and E121, whose biochemical characterisation showed high similarities with PSP strains. Identical symptoms were obtained with KA54, E121 and PSP strains, after vacuum inoculation of detached immature fruits of the cultivar Fuji, and young leaves of the cultivars Fuji, Mutsu, Gala and Golden Delicious. Koch's postulate was verified. These results indicate the presence of PSP in France. Differential characterisation criteria including serological, molecular and pathogenicity tests are proposed.

Effectiveness of Fosetyl-Aluminum and Streptomycin Alone and In Combination for Control of Blister Spot on ‘Mutsu’ Apples in Ohio and New York

The efficacies of fosetyl-Al (Aliette 80 WDG) and streptomycin (Agri-mycin 17 WP) were evaluated alone and in combination for control of blister spot of ‘Mutsu’ apple in Ohio and New York. In both test orchards, streptomycinresistant strains of Pseudomonas syringae pv. papulans were present. The results of field tests conducted in two consecutive years in Ohio were very similar. Fosetyl-Al and streptomycin applied alone resulted in significantly fewer infected fruit and significantly lower disease severity than the nontreated control. In 1997 only, the combination treatment of fosetyl-Al and streptomycin resulted in significantly less infected fruit and significantly less disease severity than the other treatments. For field experiments conducted in two consecutive years in New York, streptomycin and fosetyl-Al plus streptomycin resulted in significantly fewer infected fruit and significantly less disease severity than the untreated control, with no significant differences between treatments. Fosetyl-Al decreased disease severity and increased the percentage of marketable fruit in 1995, but it did not in 1996. The results suggest that fosetyl-Al and streptomycin usually provide a significant level of blister spot control when applied alone. When applied in combination, they may have an additive effect that increases the level of disease control. However, the level of control under the heavy disease pressure that occurred in both years of testing in Ohio was still less than 50% as compared to the nontreated control. The level of control obtained in New York tests was also unsatisfactory. Results indicate that fosetyl-Al and streptomycin alone or in combination are not highly efficacious in orchards where streptomycin-resistant strains of P. syringae pv. papulans are present, and their use in such orchards is not economically feasible. The lack of effective chemical controls could increase the economic importance of blister spot in the U.S. Accepted for publication 28 November 2000. Published 4 December 2000.