ABSTRACT
Phytophthora parasitica is an important oomycete that causes disease in a variety of plants, dimethomorph fungicides being specific for oomycetes. The aim of this study was to use RNA-seq to rapidly discover the mechanism by which dimethomorph acts in the treatment of P. parasitica. We found that the expression of 832 genes changed significantly after the dimethomorph treatment, including 365 up-regulated genes and 467 down-regulated genes. According to the Gene Ontology (GO) enrichment analysis, pathway enrichment and verification test results, the following conclusions are obtained: (i) the treatment of P. parasitica with dimethomorph causes changes in the expression levels of genes associated with the cell wall and cell wall synthesis; (ii) dimethomorph treatment results in reduced permeability of the cell membrane and changes in the expression of certain transport-related proteins; (iii) dimethomorph treatment increased reactive oxygen species and reduced the expression of genes related to the control of oxidative stress.
Phytophthora parasitica es un importante oomiceto que origina enfermedades en una variedad de plantas; el fungicida dimetomorf es específico contra oomicetos. El objetivo de este estudio fue utilizar la tecnología de RNA-seq para descubrir rápidamente el mecanismo por el que el dimetomorf actúa en el tratamiento de P. parasitica. Descubrimos que la expresión de 832 genes se modificaba significativamente tras el tratamiento con dimetomorf, incluyendo 365 genes que son sobrerregulados y 467 genes que son subrregulados. El análisis de enriquecimiento de ontología de genes (GO), análisis de enriquecimiento de las vías y pruebas de verificación permitieron extraer las conclusiones siguientes: 1) el tratamiento de P. parasitica con dimetomorf origina cambios en los niveles de expresión de los genes relacionados con la pared celular y su síntesis; 2) el tratamiento con dimetomorf origina una reducción de la permeabilidad de la membrana celular, así como cambios en la expresión de ciertas proteínas relacionadas con el transporte, y 3) el tratamiento con dimetomorf incrementó las especies reactivas del oxígeno y redujo la expresión de los genes relacionados con el control del estrés oxidativo.
Subject(s)
Phytophthora/drug effects , RNA, Messenger/biosynthesis , Morpholines/pharmacology , Fungicides, Industrial/pharmacology , RNA-Seq , Phytophthora/genetics , Plant Diseases/parasitology , RNA, Messenger/genetics , Carrier Proteins/biosynthesis , Carrier Proteins/genetics , Cell Membrane Permeability/drug effects , Cell Membrane Permeability/genetics , Cell Wall/metabolism , Gene Expression Regulation/drug effects , Sequence Alignment , Reactive Oxygen Species , Oxidative Stress/genetics , beta-Glucans/analysis , Real-Time Polymerase Chain Reaction , Gene OntologyABSTRACT
Phytophtora capsici es un patógeno que incide sobre cultivos de la familia de las solanáceas causando pérdidas económicas en cultivos de pimientos, tomates, berenjenas y cur-cubitáceas. En este trabajo evaluamos el efecto del quitosano de bajo grado de polimerización (QBP) sobre el crecimiento de P. capsici y sobre la regulación génica de este fitopatógeno a nivel transcripcional. A una concentración de 0,4mg/l de QBP se obtuvo un 88% de inhibición en el crecimiento; concentraciones superiores a 1,6 mg/l inhibieron el crecimiento en un 100%. Mediante ensayos de cambio en la movilidad electroforética de ácidos nucleicos se comprobó que el quitosano interactúa con el ADN y el ARN del hongo frente a concentraciones entre 2 y 4 mg/l de ADN y entre 0,5 y 3 mg/l de ARN. Además, se efectuó un análisis de despliegue diferencial de los productos de amplificación por RT-PCR de los ARN mensajeros de P. capsici obtenidos en presencia o ausencia de QBP; este mostró cambios en el perfil de expresión inducidos por el tratamiento con quitosano. El análisis bioinformático de las secuencias de los transcritos expresados diferencialmente sugiere que el QBP afectó la regulación génica de elementos involucrados en la síntesis de quitina y de proteínas de unión a hidratos de carbono.
Phytophthora blight of peppers, caused by oomycete Phytophthora capsici, currently causes economic losses in crops such as peppers, tomatoes, eggplant and cucurbits. In this work, we evaluated the effect of chitosan with low degree of polymerization (LDP) on growth and gene expression of P. capsici cultures. LDP chitosan inhibited 88% of P. capsici mycelial growth at concentrations up to 0,4 mg/l, whereas at concentrations higher than 1,6 mg/l it completely inhibit growth. Gel mobility shift assays demonstrated that chitosan interacts with DNA and RNA of the fungus at concentrations ranging from 2 to 4 mg/l for DNA and 0,5 to 3 mg/l for RNA. The differential display analysis of RT-PCR-amplification products of P. capsici messenger RNA revealed changes in gene expression profiles after the chitosan treatment. Bioinformatic analysis of sequences from selected differentially-expressed bands showed the gene regulation of elements involved in chitin synthesis and carbohydrate-binding proteins.
Subject(s)
Phytophthora/genetics , Gene Expression/drug effects , Gene Expression Regulation/drug effects , Chitosan/administration & dosage , Phytophthora/drug effects , Electrophoretic Mobility Shift Assay/methods , Chitosan/therapeutic use , PolymerizationABSTRACT
The objective of this work was to conduct a temporal evaluation of incidence of rot base and sanitary severity, and to relate the impact on the seed pathology of common bean cultivars. In the 2015-2016 harvest, in the city of Ipameri, Goiás, ten cultivars of common bean were evaluated (BRS Estilo©, BRS Pérola©, IPR Tangará©, IPR Tuiuiú©, IPR Uirapuru©, IAC Milênio©, Imperador©, IAC F3 R2©, IAC OTG© and IPR Campos Gerais©) and distributed into five blocks, totaling 40 experimental units. The incidence of wilt and base rot and sanitary severity were analyzed by taking ten random samples per block at 21, 28, 56, 63 and 69 days after planting. At the end of 120 days, a total of 20 plants were harvested per cultivar, and 250 seeds were harvested for application of the Blotter Test method. From 21 to 69 days after planting, the cultivars BRS Pérola© and IPR Campos Gerais© deserve to be highlighted for presenting the lowest incidence of wilt in the reproductive and vegetative cycles. The cultivar BRS Pérola© showed the lowest incidence of wilt and sanitary severity. In the analysis of harvested seeds, the cultivar BRS Pérola© presented high physiological quality for all evaluated parameters. On the other hand, Cramberry (OTG)© showed low physiological potential in germination and vigor tests.(AU)
O objetivo deste trabalho foi realizar uma avaliação temporal da incidência de podridão-do-colo e severidade fitossanitária, e relacionar o impacto na patologia de sementes de cultivares comerciais de feijoeiro-comum. Na safra 2015-2016, no município de Ipameri, Goiás, foram avaliados dez cultivares de feijoeiro (BRS Estilo©, BRS Pérola©, IPR Tangará©, IPR Tuiuiú©, IPR Uirapuru©, IAC Milênio©, Imperador©, IAC F3 R2©, IAC OTG© e IPR Campos Gerais©), distribuídos em cinco blocos, totalizando 40 unidades experimentais. Analisou-se temporalmente a incidência da murcha e podridão-do-colo e a severidade fitossanitária tomando dez amostragens aleatórias por bloco aos 21, 28, 56, 63 e 69 dias após o plantio. Ao final dos 120 dias, colheu-se um total de 20 plantas por cultivar, sendo extraídas 250 sementes de cada para aplicação do método "Blotter Test". Dos 21 aos 69 dias após o plantio, merecem destaque os cultivares BRS Pérola© e IPR Campos Gerais©, por apresentarem as menores incidências de murcha nos ciclos reprodutivo e vegetativo. O cultivar BRS Pérola© apresentou as menores incidências de murchas e severidade fitossanitária. Na análise de sementes colhidas, o cultivar BRS Pérola© apresentou elevada qualidade fisiológica para todos os parâmetros avaliados. Em contrapartida, o cultivar Cramberry (OTG)© mostrou baixo potencial fisiológico nos testes de germinação e vigor.(AU)
Subject(s)
Phytophthora/growth & development , Seeds/microbiology , Incidence , PhaseolusABSTRACT
Papaya (Carica papaya) is one of the most cultivated and consumed tropical fruit worldwide. Its production might be limited by preharvest and postharvest diseases. The fruit rot caused by Phytophthora palmivora is one of the most important postharvest diseases of papaya in Brazil. The control of these diseases is usually made with fungicide applications. Therefore, studies concerning biocontrol of postharvest diseases might generate data that may reduce the environmental impacts caused by pesticides. Thus, the biological control by Trichoderma in postharvest diseases is an alternative to the use of fungicides for the postharvest control of P. palmivora in the papaya fruit. Four antagonists [T. asperellum (SF04), T. virens (255C1), T. harzianum (THP) and T. longibrachiatum (4088)] were tested, as follow: 1) Trichoderma spp. applied 1 hour after inoculation of P. palmivora and; 2) Trichoderma spp. applied 24 hours after inoculation of P. palmivora; 3) Trichoderma spp. applied 1 hour before inoculation of P. palmivora, and 4) Trichoderma spp. applied 24 hours before inoculation of P. palmivora. All Trichoderma significantly (P£0,05) reduced the incidence and severity of disease. The 4088 (T. longibrachiatum) isolate was the best controller agent of P. palmivora in postharvest.
O mamão é uma fruta muito cultivada e consumida nas regiões tropicais e subtropicais do mundo e apresenta diversos problemas fitossanitários. Sendo assim, estudos de doenças pós-colheita com biocontroladores viabilizam a diminuição de impactos causados pelo uso de fungicidas. A podridão-dos-frutos (Phytophthora palmivora) é uma importante doença pós-colheita em mamão no Brasil. Neste contexto, o controle biológico desta doença na póscolheita com Trichoderma é uma alternativa viável ao uso de fungicidas e foi aplicado neste estudo para avaliar a eficácia de Trichoderma spp. para o biocontrole de P. palmivora em mamão na pós-colheita. Foram utilizados quatro potenciais antagonistas: T. asperellum (SF04), T. virens (255C1), T. harzianum (THP) e T. longibrachiatum (4088). E as frutas foram submetidas aos seguintes tratamentos: Inoculação de P. palmivora e 1 hora depois inoculação do Trichoderma spp.; Inoculação de P. palmivora e 24 horas depois inoculação do Trichoderma spp.; Inoculação de Trichoderma spp. e 1 hora depois inoculação do P. palmivora e; Inoculação de Trichoderma spp. e 24 horas depois inoculação do P. palmivora. Todos os isolados de Trichoderma reduziram significativamente tanto na incidência como na severidade da doença. O isolado 4088 (T. longibrachiatum) foi o melhor no controle da podridão.
Subject(s)
Phytophthora , Trichoderma , Pest Control, Biological , Crops, Agricultural , Carica , FruitABSTRACT
The papaya fruit rot (Phytophthora palmivora) is responsible for significant losses. To reduce diseases, especially in areas with climate and humidity favorable to pathogens, are adopted chemical methods, which sometimes increase the cost of production and cause severe environmental impacts. Alternatively, there are products, such as, phosphites of potassium and acibenzolar-S-methyl (ASM) that might be efficient on disease control and less aggressive to environment. Phosphites of K and ASM were evaluated in this study on the control effectiveness of papaya fruit rot at different dosages in preharvest and postharvest. The severity and percentage of disease control were evaluated for each treatment. For the pre-harvest treatments (applied six days before harvest), the phosphite of K [240 g L-1 K2O, 340 g L-1 P2O5 and 50 g L-1 (Reforce® + Salicylic Acid)] at 3 or 6 mL L-1significantly reduced disease severity, and, reduced fruit ripening. On postharvest application, ASM reduced disease severity.
A podridão dos frutos do mamoeiro (Phytophthora palmivora) é responsável por perdas significativas e para minimizar a doença, principalmente em locais com clima favorável ao patógeno, medidas de controle químico são adotadas. Porém, estas medidas podem elevar o custo da produção e causar severos impactos ambientais. Alternativamente, existem produtos como os fosfitos de potássio (K) e acibenzolar-S-metil (ASM) que podem ser eficientes no controle de doenças e menos agressivos ao ambiente. Fosfitos de K e ASM foram avaliados neste trabalho quanto à eficácia do controle da podridão de frutos de mamão 'Sunrise Solo' tipo exportação, cultivados no Sul da Bahia, em diferentes doses e formulações comerciais na pré e pós-colheita. Para cada tratamento foi avaliada a severidade da doença e o percentual de controle da doença, sendo também verificada a influência nas características fisioquímicas na fruta. Para os tratamentos pré-colheita, aplicados aos seis dias antes da colheita, o fosfito de K [240 g L-1 K2O, 340 g L-1 P2O5 e 50 g L-1 C7H6O3 (Reforce® + Ácido Salicílico)] a 3 ou 6 mL L-1 reduziu significativamente a doença, e influenciou diretamente no atraso da maturação da fruta. Na pós-colheita, ASM incitou redução na severidade da doença. Todos os tratamentos não alterarão as características fisioquímicas da fruta.
Subject(s)
Phytophthora , Phosphites , Crops, Agricultural , Carica , Failure to ThriveABSTRACT
Abstract A total of 276 endophytic bacteria were isolated from the root nodules of soybean (Glycine max L.) grown in 14 sites in Henan Province, China. The inhibitory activity of these bacteria against pathogenic fungus Phytophthora sojae 01 was screened in vitro. Six strains with more than 63% inhibitory activities were further characterized through optical epifluorescence microscopic observation, sequencing, and phylogenetic analysis of 16S rRNA gene, potential plant growth-promoting properties analysis, and plant inoculation assay. On the basis of the phylogeny of 16S rRNA genes, the six endophytic antagonists were identified as belonging to five genera: Enterobacter, Acinetobacter, Pseudomonas, Ochrobactrum, and Bacillus. The strain Acinetobacter calcoaceticus DD161 had the strongest inhibitory activity (71.14%) against the P. sojae 01, which caused morphological abnormal changes of fungal mycelia; such changes include fracture, lysis, formation of a protoplast ball at the end of hyphae, and split ends. Except for Ochrobactrum haematophilum DD234, other antagonistic strains showed the capacity to produce siderophore, indole acetic acid, and nitrogen fixation activity. Regression analysis suggested a significant positive correlation between siderophore production and inhibition ratio against P. sojae 01. This study demonstrated that nodule endophytic bacteria are important resources for searching for inhibitors specific to the fungi and for promoting effects for soybean seedlings.
Subject(s)
Plant Growth Regulators/metabolism , Soybeans/growth & development , Soybeans/microbiology , Bacteria/isolation & purification , Root Nodules, Plant/microbiology , Endophytes/isolation & purification , Antibiosis , Phylogeny , Phytophthora/cytology , Phytophthora/growth & development , Phytophthora/drug effects , Bacteria/classification , Bacteria/metabolism , DNA, Ribosomal/genetics , DNA, Ribosomal/chemistry , RNA, Ribosomal, 16S/genetics , Cluster Analysis , China , Sequence Analysis, DNA , Endophytes/classification , Endophytes/metabolismABSTRACT
Black rot disease in orchids is caused by the water mold Phytophthora palmivora. To gain better biocontrol performance, several factors affecting growth and antifungal substance production by Pseudomonas aeruginosa RS1 were verified. These factors include type and pH of media, temperature, and time for antifungal production. The results showed that the best conditions for P. aeruginosa RS1 to produce the active compounds was cultivating the bacteria in Luria-Bertani medium at pH 7.0 for 21 h at 37 °C. The culture filtrate was subjected to stepwise ammonium sulfate precipitation. The precipitated proteins from the 40% to 80% fraction showed antifungal activity and were further purified by column chromatography. The eluted proteins from fractions 9–10 and 33–34 had the highest antifungal activity at about 75% and 82% inhibition, respectively. SDS-PAGE revealed that the 9–10 fraction contained mixed proteins with molecular weights of 54 kDa, 32 kDa, and 20 kDa, while the 33–34 fraction contained mixed proteins with molecular weights of 40 kDa, 32 kDa, and 29 kDa. Each band of the proteins was analyzed by LC/MS to identify the protein. The result from Spectrum Modeler indicated that these proteins were closed similarly to three groups of the following proteins; catalase, chitin binding protein, and protease. Morphological study under scanning electron microscopy demonstrated that the partially purified proteins from P. aeruginosa RS1 caused abnormal growth and hypha elongation in P. palmivora. The bacteria and/or these proteins may be useful for controlling black rot disease caused by P. palmivora in orchid orchards.
Subject(s)
Ammonium Sulfate , Bacteria , Carrier Proteins , Catalase , Chitin , Chromatography , Electrophoresis, Polyacrylamide Gel , Fungi , Hydrogen-Ion Concentration , Hyphae , Microscopy, Electron, Scanning , Molecular Weight , Phytophthora , Pseudomonas aeruginosa , Pseudomonas , WaterABSTRACT
Dual biocontrol of both insects and plant pathogens has been reported for certain fungal entomopathogens, including Beauveria bassiana and Lecanicillum spp. In this study, we demonstrate, for the first time, the dual biocontrol potential of two fungal isolates identified by morphological and phylogenetic analyses as Isaria javanica. Both these isolates caused mortality in the greater wax moth, and hence can be considered entomopathogens. Spores of the isolates were also pathogenic to nymphs of the green peach aphid (Myzus persicae), with an LC₅₀ value of 10⁷ spores/mL 4 days after inoculation and an LT₅₀ of 4.2 days with a dose of 10⁸ spores/mL. In vitro antifungal assays also demonstrated a strong inhibitory effect on the growth of two fungi that are pathogenic to peppers, Colletotrichum gloeosporioides and Phytophthora capsici. These results indicate that I. javanica isolates could be used as novel biocontrol agents for the simultaneous control of aphids and fungal diseases, such as anthracnose and Phytophthora blight, in an integrated pest management framework for red pepper.
Subject(s)
Aphids , Beauveria , Capsicum , Colletotrichum , Fungi , Hemiptera , In Vitro Techniques , Insecta , Mortality , Moths , Nymph , Pest Control , Phytophthora , Plants , Prunus persica , SporesABSTRACT
The most effective disease management method for yield reducing diseases affecting tomatoes is the use of fungicides. This study evaluated the efficacy of chemical control on three Phytophthora sp. isolates, pathogenic to tomatoes. The effect of fungicides on mycelial growth of Phytophthora sp. and on tomato wilt was evaluated in vitro and in vivo. Two tests were done in Petri plates, and one on seedlings, in completely randomized design as a 4x3 factorial, with 5 replications. In vitro tests were done in growth chamber, at 25°C. The experimental unit consisted of a 5-mm diameter fungal mycelial plug placed 30 mm away from a filter paper disk, of similar size, soaked in fungicide, over cornmeal agar. The first test evaluated four commercial products registered for the control of potato blight: chlorothalonil+metalaxyl (Folio Gold® 742.5 WP); propamocarb chloridrate (Infinito® 687.5 CS), metalaxyl-m+mancozeb (Ridomil Gold® 68 WP), cymoxanil + manconzeb (Curzate® MZ 72 WG), at the recommended doses. The other assays evaluated three doses of Infinito (0.125%, 0.150% or 0.175%) and Ridomil. In vivo test was done in the greenhouse, and the experimental unit consisted of one pot, containing one tomato seedling, cultivar Alambra F1. Fungicide was drenched on the seedling soil one day prior to inoculation with 50,000 zoospores per pot. Data of mycelia growth inhibition by fungicide were submitted to analysis of variance and the averages compared by the Tukey test at 5% significance; efficacy was determined as a function of Ridomil®, the standard fungicide. In the first test, regardless of isolate, Infinito® presented performance similar to Ridomil® with efficacy of 98.5%, while Folio Gold® presented efficacy of 57.3% and Curzate® had no fungicide effect. Growth of isolate PP3 was smaller in all fungicides. In the second in vitro test, all three doses of Infinito® had efficacy above 82%. The best control was observed on isolates PP3 and PP4. In the third test, in vivo, no significant differences were observed in root matter among the standard fungicide and the doses of Infinito®; however, efficacy of Infinito® at 0.175% was 14% greater than that obtained with Ridomil®. It can be concluded that Infinito® is one more option for the control of tomato wilt.
O método mais efetivo no manejo das doenças fúngicas que afetam o tomateiro e reduzem a produtividade significativamente é o uso de fungicidas. Este trabalho avaliou a eficiência do controle químico sobre três isolados de Phytophthora sp. patogênicos ao tomateiro. O efeito de fungicidas no crescimento micelial de Phytophthora sp. e na murcha de tomateiro foi avaliado in vitro e in vivo, respectivamente. Foram realizados dois testes em placa de Petri e um teste em mudas, em delineamento experimental inteiramente casualizado (DIC), como fatorial 4x3, com 5 repetições. Os testes in vitro ocorreram em câmara de crescimento, a 25°C. A parcela experimental correspondeu a um disco de micélio fúngico disposto a 30 mm de um disco de papel embebido com solução de fungicida, ambos com 5 mm de diâmetro e dispostos sobre meio sólido a base de milho. O primeiro teste avaliou quatro produtos comerciais indicadas para o controle da mela ou requeima no tomateiro: clorotalonil+metalaxil (Folio Gold® 742,5 PM); propamocarbe (Infinito® 687,5 SC), metalaxil-M+mancozeb (Ridomil Gold® 68 PM), cimoxanil + manconzeb (Curzate® MZ 72 WG), nas dosagens recomendadas. Os demais ensaios avaliaram três doses de Infinito (0,125%, 0,150% e 0,175%) e Ridomil. O teste in vivo foi realizado em casa de vegetação, sendo a parcela experimental representada por um vaso, contendo uma muda de tomate, variedade Alambra F1. Fez-se "drench" para aplicação do fungicida no dia anterior à inoculação de 50.000 zoósporos por vaso. Dados de inibição do crescimento micelial por fungicidas foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey a 5% de significância; e a eficácia foi calculada em função do Ridomil®, fungicida padrão. No primeiro ensaio, independentemente do isolado, Infinito® apresentou desempenho semelhante ao Ridomil® com uma eficácia de 98,5%, enquanto Folio Gold® apresentou eficácia de 57,3% e Curzate não apresentou efeito fungicida. O crescimento do isolado PP3 foi menor em todos os produtos. No segundo teste in vitro as três doses de Infinito® tiveram eficácia superior a 82%. O melhor controle foi observado sobre os isolados PP3 e PP4. No terceiro teste, in vivo, não houve diferença significativa na massa de raízes, entre o padrão e as doses de Infinito®, no entanto, a eficácia de Infinito® a 0,175% foi 14% maior que a obtida para o Ridomil®. Conclui-se que Infinito® é mais uma opção para controle da mela em tomateiro
Subject(s)
Phytophthora , Solanum lycopersicum , MycosesABSTRACT
El objetivo principal fue la identificación de microorganismos del género Phytophthora que afectan a bosques naturales mixtos y viveros de Pinus sp. y Quercus sp., con importancia socioeconómica, en los departamentos de Guatemala y Sacatepéquez. Para el aislamiento de Phytophthora se procesaron muestras de suelo y tejido vegetal. En el departamento de Guatemala, del bosque natural se obtuvieron 10 muestras de Quercus sp. y 45 de Pinus sp., de los viveros se obtuvieron 11 muestras de Quercus sp. y 88 de Pinus sp. En el departamento de Sacatepéquez se obtuvieron del bosque natural 15 muestras de Quercus sp. y 48 de Pinus sp.; y en los viveros, 58 muestras de Pinus sp. y 25 de Quercus sp. Un total de 13 muestras procedentes de los viveros fueron positivas a la presencia de Phytophthora sp. en Pinus maximinoi, 10 procedentes del departamento de Guatemala, y tres de Sacatepéquez. Referente al tipo de crecimiento de la colonia en medio PDA, se obtuvieron cinco de tipo estolonífero, cinco tipo semipetaloide, una colonia de tipo estelado y dos colonias sin ningún tipo de patrón de crecimiento. Las pruebas de patogenicidad realizadas con la cepa VP16 mostraron alta incidencia y severidad para las especies de Pinus caribaea, P. oocarpa, P. pseudostrobus, P. maximinoi y en menor grado en Pinus tecunumanii.
The mean purpose of this research was to identify microorganism belonging to Phytophthora genera which are affecting mixed natural forests and nurseries of Pinus sp. and Quercus sp. These species have economic and social impact in provinces such as Guatemala and Sacatepéquez. Soil and vegetal tissue were used to isolate Phytophthora from natural forest of Guatemala, 45 Pinus sp. and 10 Quercus sp. were sampled and from nurseries 88 Pinus sp. and 11 Quercus sp. From Sacatepéquez province, from natural forest system were sampled 48 Pinus sp. and 15 Quercus sp. From nurseries were sampled 58 Pinus sp. and 25 Quercus sp. After processing the samples from soil and roots 13 were found positive to Phytophthora sp. in Pinus maximinoi, 10 from Guatemala and three from Sacatepéquez Provinces. The culture of Phytophthora sp. on PDA produced two colonies without define form and five stoloniferous, five semipetaloid, one stelade type colonies. VP16 isolate was inoculated in five species of pine for pathogenicity test, causing high percentages of incidence and severity on Pinus caribaea, P. oocarpa, P. pseudostrobus and P. maximinoi and low rates of incidence and severity on Pinus tecunumanii.
Subject(s)
Humans , Animals , Male , Forests , Phytophthora , Pinus , Forests , Trees/microbiologyABSTRACT
Phytophthora diseases have become a major impediment in the citrus production in Thailand. In this study, an isolate of Phytophthora denominated as PHY02 was proven to be causal pathogen of root rot of Pomelo (Citrus maxima) in Thailand. The isolate PHY02 was morphologically characterized and identified as Phytophthora palmivora based on molecular analysis of an internal transcribed spacer rDNA sequence. This work also presents in vitro evaluations of the capacities of Chaetomium spp. to control the P. palmivora PHY02. As antagonists, Chaetomium globosum CG05, Chaetomium cupreum CC3003, Chaetomium lucknowense CL01 inhibited 50~61% mycelial growth, degraded mycelia and reduced 92~99% sporangial production of P. palmivora PHY02 in bi-culture test after 30 days. Fungal metabolites from Chaetomium spp. were tested against PHY02. Results showed that, methanol extract of C. globosum CG05 expressed strongest inhibitory effects on mycelial growth and sporangium formation of P. palmivora PHY02 with effective dose ED50 values of 26.5 microg/mL and 2.3 microg/mL, respectively. It is interesting that C. lucknowense is reported for the first time as an effective antagonist against a species of Phytophthora.
Subject(s)
Chaetomium , Citrus , DNA, Ribosomal , Methanol , Phytophthora , Sporangia , ThailandABSTRACT
Thailand is one of the largest citrus producers in Southeast Asia. Pathogenic infection by Phytophthora, however, has become one of major impediments to production. This study identified a pathogenic oomycete isolated from rotted roots of pomelo (Citrus maxima) in Thailand as Phytophthora nicotianae by the internal transcribed spacer ribosomal DNA sequence analysis. Then, we examined the in vitro and in vivo effects of Chaetomium globosum, Chaetomium lucknowense, Chaetomium cupreum and their crude extracts as biological control agents in controlling this P. nicotianae strain. Represent as antagonists in biculture test, the tested Chaetomium species inhibited mycelial growth by 50~56% and parasitized the hyphae, resulting in degradation of P. nicotianae mycelia after 30 days. The crude extracts of these Chaetomium species exhibited antifungal activities against mycelial growth of P. nicotianae, with effective doses of 2.6~101.4 microg/mL. Under greenhouse conditions, application of spores and methanol extracts of these Chaetomium species to pomelo seedlings inoculated with P. nicotianae reduced root rot by 66~71% and increased plant weight by 72~85% compared to that in the control. The method of application of antagonistic spores to control the disease was simple and economical, and it may thus be applicable for large-scale, highly effective biological control of this pathogen.
Subject(s)
Asia, Southeastern , Biological Control Agents , Chaetomium , Citrus , Complex Mixtures , DNA, Ribosomal , Hyphae , Methanol , Oomycetes , Phytophthora , Plants , Seedlings , Sequence Analysis , Spores , Thailand , TobaccoABSTRACT
Laccases are blue copper oxidases (E.C. 1.10.3.2) that catalyze the one-electron oxidation of phenolics, aromatic amines, and other electron-rich substrates with the concomitant reduction of O2 to H2O. A novel laccase gene pclac2 and its corresponding full-length cDNA were cloned and characterized from Phytophthora capsici for the first time. The 1683 bp full-length cDNA of pclac2 encoded a mature laccase protein containing 560 amino acids preceded by a signal peptide of 23 amino acids. The deduced protein sequence of PCLAC2 showed high similarity with other known fungal laccases and contained four copper-binding conserved domains of typical laccase protein. In order to achieve a high level secretion and full activity expression of PCLAC2, expression vector pPIC9K with the Pichia pastoris expression system was used. The recombinant PCLAC2 protein was purified and showed on SDS-PAGE as a single band with an apparent molecular weight ca. 68 kDa. The high activity of purified PCLAC2, 84 U/mL, at the seventh day induced with methanol, was observed with 2,2'-azino-di-(3-ethylbenzothialozin-6-sulfonic acid) (ABTS) as substrate. The optimum pH and temperature for ABTS were 4.0 and 30 ºC, respectively . The reported data add a new piece to the knowledge about P. Capsici laccase multigene family and shed light on potential function about biotechnological and industrial applications of the individual laccase isoforms in oomycetes.
Subject(s)
Laccase/genetics , Laccase/metabolism , Phytophthora/enzymology , Cloning, Molecular , Conserved Sequence , Enzyme Stability , Gene Expression , Hydrogen-Ion Concentration , Laccase/chemistry , Laccase/isolation & purification , Molecular Weight , Open Reading Frames , Protein Structure, Tertiary , Phytophthora/genetics , Pichia/genetics , Protein Sorting Signals/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , TemperatureABSTRACT
Based on previous results of 16S rDNA sequence homologuous and results of physic-biochemical indexes and morphological characteristics in the present work, bacterial strain ge15 isolated from roots of ginseng plants was identified as Stenotrophomonas rhizophila. Confronting incubation results showed that, strain ge15 inhibited the growth of Alternaria panax, Phytophthora cactorum, and Cylindrocapon destructans significantly, and the width of inhibition zone was 13.3, 24.0, 12.0 mm, respectively. Further results showed that the emergence rate and seedling survive rate of ge15 treatment was significantly higher than those of the control, and which was similar to pesticide carbendazol treatment. The ge15 strain has good application potential in ginseng diseases control without contamination.
Subject(s)
Alternaria , Physiology , Antibiosis , Biological Control Agents , Hypocreales , Physiology , Panax , Microbiology , Phytophthora , Physiology , Plant Diseases , Microbiology , Plant Roots , Microbiology , Seedlings , Microbiology , Stenotrophomonas , Classification , Cell Biology , PhysiologyABSTRACT
Sequence characterized amplified region (SCAR) markers are one of the most effective and accurate tools for microbial identification. In this study, we applied SCAR markers for the rapid and accurate detection of Phytophthora katsurae, the casual agent of chestnut ink disease in Korea. In this study, we developed seven SCAR markers specific to P. katsurae using random amplified polymorphic DNA (RAPD), and assessed the potential of the SCAR markers to serve as tools for identifying P. katsurae. Seven primer pairs (SOPC 1F/SOPC 1R, SOPC 1-1F/SOPC 1-1R, SOPC 3F/SOPC 3R, SOPC 4F/SOPC 4R, SOPC 4F/SOPC 4-1R, SOPD 9F/SOPD 9R, and SOPD 10F/SOPD 10R) from a sequence derived from RAPD fragments were designed for the analysis of the SCAR markers. To evaluate the specificity and sensitivity of the SCAR markers, the genomic DNA of P. katsurae was serially diluted 10-fold to final concentrations from 1 mg/mL to 1 pg/mL. The limit of detection using the SCAR markers ranged from 100 microg/mL to 100 ng/mL. To identify the limit for detecting P. katsurae zoospores, each suspension of zoospores was serially diluted 10-fold to final concentrations from 10 x 10(5) to 10 x 10(1) zoospores/mL, and then extracted. The limit of detection by SCAR markers was approximately 10 x 10(1) zoospores/mL. PCR detection with SCAR markers was specific for P. katsurae, and did not produce any P. katsurae-specific PCR amplicons from 16 other Phytophthora species used as controls. This study shows that SCAR markers are a useful tool for the rapid and effective detection of P. katsurae.
Subject(s)
Cicatrix , DNA , Ink , Korea , Limit of Detection , Phosphatidylcholines , Phytophthora , Polymerase Chain Reaction , Sensitivity and SpecificityABSTRACT
The purpose of this study was to evaluate the effectiveness of a commercial biofungicide such as Trichomix-HV in controlling damping-off disease in cucumber seedlings of greenhouses. In this regard, 504 fungal isolates were collected from greenhouses at 31 districts in city of Jiroft in Iran. Pythium aphanidermatum, P.ultimum, P. irregulare, Phytophthora drechsleri, and Ph. melonis accounted for 9.9%, 8.3%, 4.5%, 4.9%, and 21% of total isolates collected, respectively. Isolates of P. aphanidermatum obtained from commercial cucumber in greenhouses were tested in vitro and under greenhouse conditions for sensitivity to chemical and biological treatments. To this aim, Trichomix-HV a commercial formulation of Trichoderma harzianum strain T969 and the fungicides Metalaxyl and Metalaxyl MZ were amended into the culture medium as well as into sterilized or non-sterilized greenhouse soils inoculated by the pathogen and containing plants at the seedling stage. Trichomix-HV significantly [P < 0.001] reduced seedling infection at a rate of 82% when applied into soil medium at a concentration of 107 conidia ml/L and reduced vegetative growth of Pythium aphanidermatum in vitro. The result from this study shows that Trichomix-HV can be effectively used as a biocontrol agent for controlling damping-off cucumber seedlings and having the potential to replace chemical fungicides as a mean of disease control
Subject(s)
Fungicides, Industrial , Greenhouse Effect , Cucumis , Pythium , Phytophthora , TrichodermaABSTRACT
<p><b>OBJECTIVE</b>To observe the effects of a biotic elicitor fungal hyphae extract, an abiotic elicitor methyl jasmonate and their synergistic action on the accumulation of phenolic acids and tanshinones in Salvia miltiorrhiza hairy root.</p><p><b>METHOD</b>Different elicitors were added to S. miltiorrhiza hairy root, which was subcultured for 21 days, the dry weight and contents of phenolic acids and tanshinones were determined at different harvest-time.</p><p><b>RESULT</b>S. miltiorrhiza hairy root growth was significantly inhibited by all three treatments and the accumulation of cryptotanshinone and dihydrotanshinone were promoted by each elicition. As for the accumulation of phenolic acids, there were differences between fungal elicitor and methyl jasmonate treatments, they were promoted by methyl jasmonate while inhibited in a certain extent by fungal hyphae extract.</p><p><b>CONCLUSION</b>Fungal elicitor, methyl jasmonate and their synergistic action have significant influence on accumulation of components in S. miltiorrhiza hairy root, and the effect varies between phenolic acids and tanshinones. There is no correlation between production of water-soluble ingredients and fat-soluble components on the whole under three different treatments.</p>
Subject(s)
Acetates , Pharmacology , Cyclopentanes , Pharmacology , Abietanes , Metabolism , Fungal Proteins , Metabolism , Pharmacology , Hydroxybenzoates , Metabolism , Membrane Glycoproteins , Metabolism , Pharmacology , Oxylipins , Pharmacology , Phytophthora , Chemistry , Metabolism , Plant Extracts , Metabolism , Plant Roots , Metabolism , Microbiology , Salvia miltiorrhiza , Metabolism , MicrobiologyABSTRACT
Antagonistic mechanisms of Trichoderma viride M3, Tv04-2, and T. harzianum ThB, were studied against Phytophthora nicotianae, the pathogen of stem blight disease on Schizonepeta tenuifolia by dual-culture, hydrolase activity, volatile and nonvolatile substances. Results indicated that competitive, mycoparasitism and antagonism were the antagonistic mechanisms of three Trichoderma spp. against P. nicotianae. Hydrolase activity showed that M3 was the highest for beta-1, 3-glucanases activity while ThB was the highest for proteases activity among the three T. strains, and they could produce volatile and non-volatile substances, also.
Subject(s)
Fungal Proteins , Metabolism , Hydrolases , Metabolism , Lamiaceae , Parasitology , Peptide Hydrolases , Metabolism , Pest Control, Biological , Phytophthora , Microbiology , Physiology , Plant Diseases , Parasitology , Trichoderma , PhysiologyABSTRACT
Phytophthora parasitica causes serious widespread, and difficult-to-control root rots in warmer regions. This oomycete is one of the most important pathogen of citrus. This paper reports the biological control of the pathogen by a strain of Serratia marcescens R-35, isolated from citrus rhizosphere. In greenhouse trials, the bacterium suppressed more than 50 percent of the disease and promoted the plant growth.
Phytophthora parasitica é um oomiceto que causa sérios problemas fitossanitários em diferentes espécies de plantas em regiões tropicais e o controle tem sido difícil. Este patógeno é um dos mais importante à citricultura. Este trabalho relata o controle biológico do patógeno por uma linhagem de Serratia marcescens R-35, isolada da rizosfera de citros. Em condições de casa-de-vegetação, a bactéria reduziu em mais de 50 por cento a incidência da doença, ao mesmo tempo que promoveu o crescimento de plantas.
Subject(s)
In Vitro Techniques , Pest Control, Biological , Phytophthora , Plants , Serratia marcescens , Methods , VirulenceABSTRACT
RQRT-PCR technique was evaluated for its validity as an alternative to Northern blotting for quantification of plant gene expression in diseased tissues of Hevea. Reliable RT-PCR results could be obtained by co-amplification of housekeeping actin gene as the internal control along with the gene of interest. The product of interest was quantified relative to that of the internal control by measuring net intensity of bands. Expression levels of defense-related beta-1,3-glucanase gene was studied in the pathogen infected tissues of rubber. The beta-1,3-glucanase gene was found to be induced in infected leaf tissues and reached a peak at 48 h after inoculation. The beta-1,3-glucanase gene expression during pathogen infection was determined through Northern blot hybridization also, using 18S RNA as the internal control. RQRT-PCR and Northern hybridization showed almost similar results, thereby validating the use of this technique to study the gene expression in rubber.