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1.
Microbiol Res ; 226: 1-9, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31284938

RESUMO

Citrus crops have great economic importance worldwide. However, citrus production faces many diseases caused by different pathogens, such as bacteria, oomycetes, fungi and viruses. To overcome important plant diseases in general, new technologies have been developed and applied to crop protection, including RNA interference (RNAi) and clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) systems. RNAi has been demonstrated to be a powerful tool for application in plant defence mechanisms against different pathogens as well as their respective vectors, and CRISPR/Cas system has become widely used in gene editing or reprogramming or knocking out any chosen DNA/RNA sequence. In this article, we provide an overview of the use of RNAi and CRISPR/Cas technologies in management strategies to control several plants diseases, and we discuss how these strategies can be potentially used against citrus pathogens.


Assuntos
Citrus/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Doenças das Plantas/genética , Doenças das Plantas/prevenção & controle , Interferência de RNA , Bactérias/genética , Bactérias/patogenicidade , Sistemas CRISPR-Cas , Citrus/microbiologia , Produtos Agrícolas , Fungos/genética , Fungos/patogenicidade , Edição de Genes/métodos , Engenharia Genética , Vírus/genética , Vírus/patogenicidade
2.
Microbiol Res ; 226: 27-33, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31284941

RESUMO

Postbloom fruit drop (PFD), caused mainly by Colletotrichum abscissum, is one of the most severe citrus diseases and can causes up to 80% fruit loss in favorable climatic conditions. According to the literature, other Colletotrichum species colonize hosts using distinct strategies: intracellular hemibiotrophic or subcuticular intramural necrotrophic colonization. However, so far, for C. abscissum only the necrotrophic stage has been described and some aspects remain unclear in PFD disease cycle. To better understand the disease cycle, microscopy studies could be applied. However, even using eGFP strains (expressing green fluorescent protein), the results are unclear due to the autofluorescence of citrus leaves. To eliminate this problem and to study the interaction between C. abscissum-citrus we used a destaining and staining methodologies, and we observed that in leaves, even applying injury before inoculation, C. abscissum does not colonize adjacent tissues. Apparently, in the leaves the fungus only uses the nutrients exposed in the artificial lesions for growth, and then produces large amount of spores. However, in flowers, C. abscissum penetrated and colonized the tissues of the petals 12 h after inoculation. In the early stages of infection, we observed the development of primary biotrophic hyphae, suggesting this species as a hemibiotrophic fungus, with a short biotrophic phase during flower colonization followed by dominant necrotrophic colonization. In conclusion, the use of an eGFP strain of C. abscissum and a different methodology of destaining and staining allowed a better understanding of the morphology and mechanisms used by this citrus pathogen to colonize the host.


Assuntos
Citrus/microbiologia , Colletotrichum/citologia , Colletotrichum/crescimento & desenvolvimento , Colletotrichum/patogenicidade , Doenças das Plantas/microbiologia , Flores/microbiologia , Frutas/microbiologia , Proteínas de Fluorescência Verde , Interações Hospedeiro-Patógeno , Hifas/citologia , Hifas/crescimento & desenvolvimento , Microscopia/métodos , Microscopia Confocal/métodos , Folhas de Planta , Esporos Fúngicos/citologia
3.
J Agric Food Chem ; 67(25): 6970-6977, 2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31150237

RESUMO

Huanglongbing (HLB), also known as citrus greening, is a bacterial disease that poses a devastating threat to the citrus industry worldwide. To manage this disease efficiently, we developed and characterized a ternary aqueous solution (TSOL) that contains zinc nitrate, urea, and hydrogen peroxide. We report that TSOL exhibits better antimicrobial activity than commercial bactericides for growers. X-ray fluorescence analysis demonstrates that zinc is delivered to citrus leaves, where the bacteria reside. FTIR and Raman spectroscopy, molecular dynamics simulations, and density functional theory calculations elucidate the solution structure of TSOL and reveal a water-mediated interaction between Zn2+ and H2O2, which may facilitate the generation of highly reactive hydroxyl radicals contributing to superior antimicrobial activity of TSOL. Our results not only suggest TSOL as a potent antimicrobial agent to suppress bacterial growth in HLB-infected trees, but also provide a structure-property relationship that explains the superior performance of TSOL.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Citrus/microbiologia , Nitratos/química , Nitratos/farmacologia , Doenças das Plantas/prevenção & controle , Rhizobiaceae/fisiologia , Compostos de Zinco/química , Compostos de Zinco/farmacologia , Peróxido de Hidrogênio/química , Peróxido de Hidrogênio/farmacologia , Doenças das Plantas/microbiologia , Folhas de Planta/microbiologia , Rhizobiaceae/efeitos dos fármacos , Espectroscopia de Infravermelho com Transformada de Fourier , Análise Espectral Raman , Relação Estrutura-Atividade , Ureia/química , Ureia/farmacologia
4.
Int J Food Microbiol ; 301: 9-18, 2019 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-31055161

RESUMO

A large amount of GRAS (generally recognized as safe) salts and concentrations were evaluated in in vitro tests (inhibition of mycelial growth on PDA dishes) against Lasiodiplodia theobromae, the causal agent of citrus Diplodia stem-end rot. Ammonium carbonate (AC, 0.2%), potassium sorbate (PS, 2.0%), potassium carbonate (PC, 0.2%), sodium methylparaben (SMP, 0.1%), sodium ethylparaben (SEP, 0.1%), sodium benzoate (SB, 2.0%), and potassium silicate (PSi, 2.0%) were selected as the most effective. Disease control ability of edible composite coatings formulated with hydroxypropyl methylcellulose (HPMC), beeswax (BW), and these selected antifungal GRAS salts was assessed in in vivo experiments with 'Ortanique' mandarins and 'Barnfield' oranges artificially inoculated with L. theobromae. Coatings containing 2% PS, 0.1% SEP, or 2% SB were the most effective reducing disease severity (up to 50% reduction) and were also applied to non-inoculated and cold-stored 'Barnfield' oranges to determine their effect on postharvest fruit quality. After periods of 21 and 42 d at 5 °C followed by 7 d of shelf life at 20 °C, coatings containing SEP and SB significantly reduced weight loss and did not adversely affect the physicochemical quality attributes (firmness, soluble solid content, titratable acidity, and ethanol and acetaldehyde content) and sensory flavor with respect to uncoated control fruit. Although the internal gas concentration (CO2 level) of coated fruit increased, the coatings did not induce off-flavors.


Assuntos
Ascomicetos/efeitos dos fármacos , Citrus/microbiologia , Microbiologia de Alimentos/métodos , Conservação de Alimentos/métodos , Derivados da Hipromelose/farmacologia , Sais/farmacologia , Antifúngicos/farmacologia , Frutas/microbiologia , Lipídeos/química , Lipídeos/farmacologia
5.
J Microbiol Biotechnol ; 29(6): 984-988, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31091865

RESUMO

Blue mold in citrus is caused by Penicillium italicum. In this study, the P. italicum-specific primers were developed for rapid detection based on the conserved genes RPB1 and RPB2 among Penicillium genomes. The two primer pairs RPB1-a and RPB1-b proved to be specific to detect P. italicum. The PCR assay among 39 fungal isolates and the colonial, pathogenic morphologies and molecular methods validated the specificity and reliability of these two primer pairs. This report provided a method and P. italicum-specific primers, which might greatly contribute to citrus postharvest industry.


Assuntos
Citrus/microbiologia , Primers do DNA/normas , Microbiologia de Alimentos/métodos , Técnicas de Tipagem Micológica/métodos , Penicillium/genética , Doenças das Plantas/microbiologia , Proteínas de Bactérias/genética , Primers do DNA/genética , Penicillium/classificação , Reação em Cadeia da Polimerase , RNA Polimerase II/genética , Reprodutibilidade dos Testes , Especificidade da Espécie
6.
J Food Sci ; 84(6): 1513-1521, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31120593

RESUMO

Geotrichum citri-aurantii causes sour rot in citrus fruits and is responsible for important economic losses during storage. However, the availability of chemical fungicides for the control of this pathogen is limited. Thus, the aim of this research was to evaluate the antifungal efficacy of thymol and carvacrol encapsulated in 2-hydroxylpropyl-beta-cyclodextrin (HP-ß-CD) (prepared by the microwave irradiation method [MW] and solubility method [S]) for inhibition of G. citri-aurantii using in vitro bioassays broth (micro and macrodilutions methods) and inoculated food testing. Both encapsulated thymol and carvacrol were shown to be effective for inhibiting G. citri-aurantii growth in in vitro assays. Thymol was more effective in inhibiting G. citri-aurantii, while better encapsulation was provided by MW. HP-ß-CD-thymol encapsulated by MW (HP-ß-CD-thymol-MW) showed the lowest 50% effective dose (ED50 = 1.16 mM), minimum inhibitory concentration (MIC = 5.06 mM), and minimum fungicide concentration (MFC = 52.6 mM). HP-ß-CD-thymol-MW was found highly effective in reducing the growth rate and mycelial growth inhibition. Finally, HP-ß-CD-thymol-MW and HP-ß-CD-carvacrol-MW showed a higher persistent effect than thymol and carvacrol in their natural form in inhibiting this fungus. Therefore, HP-ß-CD-thymol-MW could be a promising alternative to synthetic fungicides for controlling G. citri-aurantii, the causal agent of citrus sour rot. PRACTICAL APPLICATION: Encapsulated thymol and carvacrol in HP-ß-Cyclodextrins are effective for controlling G. citri-aurantii in in vitro experiments. Encapsulation of thymol and carvacrol by microwave irradiation method (MW) was more effective than the solubility (S) method. Thymol was more effective than carvacrol, and the best results on G. citri-auriantii inhibition were achieved using the HP-ß-CD-thymol-MW method (which gave the lowest ED50 , MIC, and MFC).


Assuntos
Citrus/microbiologia , Composição de Medicamentos/métodos , Fungicidas Industriais/farmacologia , Geotrichum/efeitos dos fármacos , Monoterpenos/farmacologia , Timol/farmacologia , beta-Ciclodextrinas/química , Fungicidas Industriais/química , Geotrichum/crescimento & desenvolvimento , Testes de Sensibilidade Microbiana , Monoterpenos/química , Doenças das Plantas/microbiologia , Timol/química
7.
J Agric Food Chem ; 67(20): 5720-5727, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31046262

RESUMO

The increasing drug resistance of phytopathogenic bacteria to conventional bactericides has driven the necessity for exploring new alternatives with a lower tendency to develop bacterial resistance. Here, we report a novel cationic symmetrical peptide P5VP5 (Ac- R+ LI R+ K+ V K+ R+ IL R+ -NH2 that enables self-assembly to form nanoparticles with excellent thermal stability. An in vitro assay showed that P5VP5 nanoparticles exhibited excellent antibacterial activity against Xanthomonas axonopodis pv citri with a MIC value of 20 µM. Meanwhile, under an in planta condition, treatment with peptide nanoparticles demonstrated the highest ability to reduce the development of citrus canker lesions in leaves. Moreover, the nanoparticles could destroy the biofilm formation, damage the cell membranes, and affect the cell membrane permeability, ultimately leading to the death of bacteria. Taken together, these nanoparticles are a promising antibacterial agent that can be used to control citrus canker and other plant diseases caused by bacteria.


Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Citrus/microbiologia , Nanopartículas/química , Xanthomonas/efeitos dos fármacos , Antibacterianos/química , Peptídeos Catiônicos Antimicrobianos/química , Biofilmes/efeitos dos fármacos , Doenças das Plantas/microbiologia , Folhas de Planta/microbiologia , Xanthomonas/fisiologia
8.
Anal Bioanal Chem ; 411(14): 3125-3133, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30989272

RESUMO

Huanglongbing (HLB) or citrus greening is a devastating disease of citrus trees that is caused by the gram-negative Candidatus Liberibacter spp. bacteria. The bacteria are phloem limited and transmitted by the Asian citrus psyllid, Diaphorina citri, and the African citrus psyllid, Trioza erytreae, which allows for a wider dissemination of HLB. Infected trees exhibit yellowing of leaves, premature leaf and fruit drop, and ultimately the death of the entire plant. Polymerase chain reaction (PCR) and antibody-based assays (ELISA and/or immunoblot) are commonly used methods for HLB diagnostics. However, they are costly, time-consuming, and destructive to the sample and often not sensitive enough to detect the pathogen very early in the infection stage. Raman spectroscopy (RS) is a noninvasive, nondestructive, analytical technique which provides insight into the chemical structures of a specimen. In this study, by using a handheld Raman system in combination with chemometric analyses, we can readily distinguish between healthy and HLB (early and late stage)-infected citrus trees, as well as plants suffering from nutrient deficits. The detection rate of Raman-based diagnostics of healthy vs HLB infected vs nutrient deficit is ~ 98% for grapefruit and ~ 87% for orange trees, whereas the accuracy of early- vs late-stage HLB infected is 100% for grapefruits and ~94% for oranges. This analysis is portable and sample agnostic, suggesting that it could be utilized for other crops and conducted autonomously. Graphical abstract.


Assuntos
Citrus/química , Citrus/microbiologia , Nutrientes/análise , Doenças das Plantas/microbiologia , Rhizobiaceae/isolamento & purificação , Análise Espectral Raman/instrumentação , Análise Espectral Raman/métodos , Citrus/genética , DNA Bacteriano/análise , DNA de Plantas/análise , Ensaio de Imunoadsorção Enzimática , Nutrientes/deficiência , Folhas de Planta/química , Folhas de Planta/genética , Reação em Cadeia da Polimerase , Reação em Cadeia da Polimerase em Tempo Real , Rhizobiaceae/genética
9.
BMC Plant Biol ; 19(1): 122, 2019 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-30940073

RESUMO

BACKGROUND: Citrus Huanglongbing (HLB) is a bacterial disease with high economic significance. The associated agent Candidatus Liberibacter asiaticus is a fastidious, phloem-limited, intracellular bacterium that is transmitted by an insect vector the Asian citrus psyllid (ACP). The genome of Ca. L. asiaticus contains protein secretion machinery that suggests host cell modulation capacity of this bacterium. RESULTS: A total of 28 candidate effectors, an important class of secreted proteins, were predicted from the Ca. L. asiaticus genome. Sequence specific primers were designed for reverse transcription (RT) and quantitative PCR (qPCR), and expression was validated for 20 of the effector candidates in infected citrus with multiple genetic background. Using detached leaf inoculation, the mRNA of effectors was detected from 6 h to 7 days post ACP exposure. It was observed that higher bacterial titers were associated with a larger number of effectors showing amplification across all samples. The effectors' expression were compared in citrus hosts with various levels of HLB tolerance, including susceptible Duncan grapefruit and Washington navel orange, tolerant citron and Cleopatra mandarin, and resistant Pomeroy trifoliate and Carrizo citrange. Across all genotypes relatively high expression was observed for CLIBASIA_03695, CLIBASIA_00460, CLIBASIA_00420, CLIBASIA_04580, CLIBASIA_05320, CLIBASIA_04425, CLIBASIA_00525 and CLIBASIA_05315 in either a host-specific or -nonspecific manners. The two genotypes in each HLB-response group also show effector-expression profiles that seem to be different. In a companion study, the expression of effectors was compared between leaves and roots of own-rooted citrus that had been Ca. L. asiaticus-infected for more than a year. Results indicated relatively high expression of CLIBASIA_03875, CLIBASIA_04800 and CLIBASIA_05640 in all leaf and some root tissues of citron, Duncan and Cleopatra. CONCLUSION: This temporal and spatial expression analysis of Ca. L. asiaticus effectors identified candidates possibly critical for early bacterial colonization, host tolerance suppression and long-term survival which are all worthy of further investigation.


Assuntos
Proteínas de Bactérias/genética , Citrus/microbiologia , Genoma Bacteriano/genética , Interações Hospedeiro-Patógeno , Doenças das Plantas/microbiologia , Rhizobiaceae/genética , Animais , Citrus/imunologia , Resistência à Doença , Genótipo , Hemípteros/microbiologia , Insetos Vetores/microbiologia , Floema/imunologia , Floema/microbiologia , Doenças das Plantas/imunologia , Folhas de Planta/imunologia , Folhas de Planta/microbiologia , RNA Bacteriano/genética , RNA Mensageiro/genética , Rhizobiaceae/fisiologia
10.
Plant Dis ; 103(6): 1156-1165, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30995151

RESUMO

Persian lime (Citrus latifolia Tan.) is an important and widely cultivated fruit crop in several regions of Mexico. In recent years, severe symptoms of gummosis, stem cankers, and dieback were detected in the Persian lime-producing region in the states of Veracruz and Puebla, Mexico. The aims of this study were to identify the species of Lasiodiplodia associated with these symptoms, determine the distribution of these species, and test their pathogenicity and virulence on Persian lime plants. In 2015, symptomatic samples were collected from 12 commercial Persian lime orchards, and 60 Lasiodiplodia isolates were obtained. Fungal identification of 32 representative isolates was performed using a phylogenetic analysis based on DNA sequence data of the internal transcribed spacer region and part of the translation elongation factor 1-α and ß-tubulin genes. Sequence analyses were carried out using the Maximum Likelihood and Bayesian Inference methods. Six Lasiodiplodia species were identified as Lasiodiplodia pseudotheobromae, Lasiodiplodia theobromae, Lasiodiplodia brasiliense, Lasiodiplodia subglobosa, Lasiodiplodia citricola, and Lasiodiplodia iraniensis. All Lasiodiplodia species of this study are reported for the first time in association with Persian lime in Mexico and worldwide. L. pseudotheobromae (46.9% of isolates) was the most frequently isolated species followed by L. theobromae (28.1%) and L. brasiliense (12.5%). Pathogenicity on Persian lime young plants using a mycelial plug inoculation method showed that all identified Lasiodiplodia species were able to cause necrotic lesions and gummosis, but L. subglobosa, L. iraniensis, and L. pseudotheobromae were the most virulent.


Assuntos
Ascomicetos , Citrus , Filogenia , Doenças das Plantas , Ascomicetos/classificação , Ascomicetos/genética , Teorema de Bayes , Citrus/microbiologia , DNA Fúngico , México , Doenças das Plantas/microbiologia , Análise de Sequência de DNA
11.
Phytopathology ; 109(8): 1344-1353, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30973310

RESUMO

Xylella fastidiosa releases outer membrane vesicles (OMVs) known to play a role in the systemic dissemination of this pathogen. OMVs inhibit bacterial attachment to xylem wall and traffic lipases/esterases that act on the degradation of plant cell wall. Here, we extended the characterization of X. fastidiosa OMVs by identifying proteins and metabolites potentially associated with OMVs produced by Temecula1, a Pierce's disease strain, and by 9a5c and Fb7, two citrus variegated chlorosis strains. These results strengthen that one of the OMVs multiple functions is to carry determinants of virulence, such as lipases/esterases, adhesins, proteases, porins, and a pectin lyase-like protein. For the first time, we show that the two citrus variegated chlorosis strains produce X. fastidiosa diffusible signaling factor 2 (DSF2) and citrus variegated chlorosis-DSF (likewise, Temecula1) and most importantly, that these compounds of the DSF (X. fastidiosa DSF) family are associated with OMV-enriched fractions. Altogether, our findings widen the potential functions of X. fastidiosa OMVs in intercellular signaling and host-pathogen interactions.


Assuntos
Citrus , Xylella , Citrus/microbiologia , Doenças das Plantas/microbiologia , Proteômica , Fatores de Virulência
12.
J Food Sci ; 84(4): 848-858, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30866044

RESUMO

This study assessed the compromised acceptance threshold (CAT) and rejection threshold (RT) of Citrus lemon (CLEO) and Citrus reticulata essential oil (CREO) in apple and orange juices. The efficacy of CLEO and CREO concentrations below the RT were evaluated alone and combined with mild heat treatment (MHT) (54 °C, up to 12 min) to inactivate the autochthonous spoilage bacteria Lactobacillus brevis, Lactobacillus plantarum, and Leuconostoc mesenteroides in apple and orange juices. The CAT of CLEO and CREO varied from 0.15 to 0.17 µL/mL in orange and apple juices. The RT of CLEO was approximately 0.58 µL/mL in apple and orange juices, and the RT of CREO was 0.68 µL/mL in both juices. When CLEO and CREO were assayed alone, the highest concentration (0.50 µL/mL) decreased counts of all strains approximately 2 log10 CFU/mL after 12 min of exposure to 54 °C. All concentrations of CLEO or CREO in combination with MHT acted synergistically against L. brevis, L. plantarum, and L. mesenteroides. Decreases in counts varied with the strain, CLEO and CREO concentrations, juice type, and exposure time to the combined treatment. CREO was more effective than CLEO in combination with MHT against the strains in apple and orange juices. Effective combinations of CLEO or CREO with MHT to control the autochthonous spoilage bacteria did not compromise the quality parameters (°Brix, pH, and titratable acidity) that characterize unsweetened juices. These results indicate CLEO or CREO at concentrations below the sensory RT in combination with MHT as a feasible technology to control autochthonous spoilage bacteria in fresh fruit juices. PRACTICAL APPLICATION: The present study provides novel information concerning the efficacy of sensorially accepted doses of CLEO and CREO combined with MHT against autochthonous spoilers in fruit juice. The valuable synergistic effects that can be observed when combining CLEO and CREO with MHT reveal a feasible preservation technology and alternative to traditional treatments that are successful because they help reduce treatment intensity, thereby avoiding adverse effects on the sensory, physicochemical, and nutritional properties of these products.


Assuntos
Citrus/microbiologia , Sucos de Frutas e Vegetais/microbiologia , Lactobacillales/classificação , Malus/microbiologia , Óleos Voláteis/farmacologia , Óleos Vegetais/farmacologia , Citrus/química , Conservação de Alimentos/métodos , Temperatura Alta , Lactobacillales/efeitos dos fármacos , Lactobacillales/fisiologia , Malus/química , Óleos Voláteis/química , Óleos Vegetais/química
13.
J Plant Physiol ; 236: 61-65, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30884323

RESUMO

The Lasbcp (CLIBASIA_RS00445) 1-Cys peroxiredoxin gene is conserved among all 13 sequenced strains of Candidatus Liberibacter asiaticus, the causal agent of Huanglongbing or "citrus greening" disease. LasBCP was previously characterized as a secreted peroxiredoxin with substrate specificity for organic peroxides, and as a potential pathogenicity effector. Agrobacterium-mediated transient expression of LasBCP in citrus leaves provided significant protection against peroxidation of free and membrane-bound lipids, thereby preserving the molecular integrity of the chlorophyll apparatus and reducing accumulation of lipid peroxidation products (oxylipins) following exposure to tert-butyl hydroperoxide (tBOOH, an organic peroxide). Oxylipins extracted from GUS-expressing citrus leaves reduced viability of L. crescens, the only Liberibacter species cultured to date. However, similar extracts obtained from LasBCP-expressing leaves were less inhibitory to L. crescens growth and viability in culture. Quantitative RT-PCR analyses showed coordinated transcriptional downregulation of oxylipin biosynthetic (CitFAD, CitLOX, CitAOS and CitAOC), and jasmonic acid (JA) (CitJAR1, CitCOI1 and CitJIN1) and salicylic acid (SA) (CitPAL, CitICS and CitPR1) signaling pathway genes in citrus leaves expressing LasBCP and treated with tBOOH. The negative response regulator of jasmonic acid CitJAZ1 was upregulated in LasBCP-expressing citrus leaves under similar conditions. These data clearly demonstrated a protective role of secreted LasBCP in favor of Las survival and colonization by alleviating ROS-induced lipid peroxidation in citrus host, preventing accumulation of antimicrobial oxylipins, and suppressing both localized and systemic immune responses in planta.


Assuntos
Citrus/microbiologia , Oxilipinas/metabolismo , Peroxirredoxinas/metabolismo , Doenças das Plantas/microbiologia , Imunidade Vegetal , Rhizobiaceae/metabolismo , Clorofila/metabolismo , Citrus/imunologia , Citrus/metabolismo , Ciclopentanos/metabolismo , Interações Hospedeiro-Patógeno , Peroxidação de Lipídeos , Doenças das Plantas/imunologia , Folhas de Planta/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Rhizobiaceae/enzimologia , Ácido Salicílico/metabolismo , Transdução de Sinais
14.
Syst Appl Microbiol ; 42(3): 348-359, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30910423

RESUMO

A collection of Pseudomonas strains was isolated in different regions of Tunisia in the period 2016-2017 from the fruits and leaves of Citrus sinensis cv. 'Valencia Late' and Citrus limon cv. 'Eureka' plants with symptoms of blast and black pit disease. A phylogenetic analysis of the housekeeping gene rpoD was used for strain identification at the species level. The results demonstrated the affiliation of these strains with the genus Pseudomonas and revealed the presence of 11 strains representing two putative new species in two monophyletic branches. These strains were analyzed morphologically and genotypically by multilocus sequence analyses of the rpoD, gyrB and 16S rRNA (rrs) gene sequences, and their phenotypic characteristics by API 20NE and Biolog GEN III. Plant pathogenic properties were confirmed on fruits and detached leaves of C. limon cv. 'Eureka'. Fatty acids and WC MALDI-TOF MS major protein profiles were determined. The genomes of both representatives were sequenced. The average nucleotide index and genome-to-genome distance from KC12T and E10BT are below the cut-off established for a described species. These results support the conclusion that the strains KC12T, KC17, KC20, KC22, KC24A, KC25 and KC26 represent a novel species of Pseudomonas, for which the name of Pseudomonas kairouanensis is proposed. The type strain is KC12T (=CECT9766 and CFBP 8662). The strains E10BT, E10AB, E10CB1 and Iy3BA represent another novel species of Pseudomonas for which the name of Pseudomonas nabeulensis is proposed; the type strain is E10BT (=CECT9765 and CFBP 8661).


Assuntos
Citrus/microbiologia , Filogenia , Doenças das Plantas/microbiologia , Pseudomonas/classificação , Antibacterianos/farmacologia , DNA Bacteriano/genética , Farmacorresistência Bacteriana , Ácidos Graxos/análise , Genes Bacterianos/genética , Genoma Bacteriano/genética , Tipagem de Sequências Multilocus , Hibridização de Ácido Nucleico , Pseudomonas/química , Pseudomonas/genética , Pseudomonas/patogenicidade , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Tunísia
15.
Ecotoxicol Environ Saf ; 173: 322-330, 2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-30784795

RESUMO

This study explores the possibility of using mycorrhization as a novel technique for diminishing the negative effects of boron (B) in the nutrient solution on seedlings of Carrizo citrange rootstock plants. For this, an experiment was planned for studying the physiological (gas exchange and chlorophyll fluorescence parameters), morphological (vegetative growth parameters), nutritional (organic solutes, carbohydrates) and oxidative stress responses of seedlings that were either mycorrhized (+AM, Rhizophagus irregularis; previously known as Glomus intraradices) or not mycorrhized (-AM), and irrigated with water containing different concentrations of B (0.5, 5 and 10 mg L-1). It was observed that an excess of B in the nutrient solution decreased the vegetative growth in both +AM and -AM plants, but this decrease was greater in -AM plants. Mycorrhized plants (+AM) under high B concentration accumulated less B in the leaves, and had a smaller reduction of net assimilation rate of CO2 and lower MDA concentration than non-mycorrhized plants. Thus, it can be concluded that mycorrhization increased the tolerance to high boron concentration in the irrigation water of citrange Carrizo seedlings by reducing both the B concentration in the plant tissue and the B toxicity in the physiological processes. The study of organic solutes and carbohydrates also pointed to a different response model between +AM and -AM plants that could be related to the different tolerance observed between these plants.


Assuntos
Boro/metabolismo , Citrus/metabolismo , Citrus/microbiologia , Glomeromycota/fisiologia , Micorrizas/fisiologia , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Simbiose
16.
Plant Cell Rep ; 38(5): 559-575, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30715581

RESUMO

KEY MESSAGE: Bacillus subtilis CLP extract activates defense gene expression and increases the unique protein production involving in pathways of ISR, SAR, ubiquitin-proteasome system, and glycolysis for stress responses in flavedo tissues. Cyclic lipopeptides (CLPs) of Bacillus subtilis ABS-S14 had ability to activate plant defensive pathways, increase resistance and control green mold rot caused by Penicillium digitatum in mandarin fruit. The current study investigated transcriptional and proteomic data to highlight the unique induction effect of CLPs produced by B. subtilis ABS-S14 on the defense mechanism of mandarins in response to P. digitatum attack, and their differences from those following the exogenous plant hormone application. The proteomic patterns of the flavedo tissues as affected by Bacillus CLP extract, salicylic acid (SA), methyl jasmonate (MeJA), and ethephon (Et) were explored. qPCR analysis revealed the great effects of CLP extract in enhancing the transcription of PAL, ACS1, GLU, POD, and PR1. Tryptic peptides by LC-MS analysis between treatments with and without fungal infection were compared. B. subtilis CLP extract empowered the plant's immune response to wound stress by the significant production of calmodulin-binding receptor-like cytoplasmic kinase 2, molybdenum cofactor sulfurase, and NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase. Ubiquitin carrier protein abundance was developed only in the treated flavedo with CLP extract coupled with P. digitatum infection. The gene expression and overall proteome findings involving pathways of ubiquitin proteasome system, ISR, SAR, and energy production provide a new insight into the molecular mechanisms of the antagonist B. subtilis ABS-S14 inducing resistance against green mold in mandarins.


Assuntos
Bacillus subtilis/patogenicidade , Proteínas de Bactérias/metabolismo , Citrus/metabolismo , Citrus/microbiologia , Lipopeptídeos/metabolismo , Peptídeos Cíclicos/metabolismo , Penicillium/patogenicidade , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Planta/metabolismo
17.
J Antibiot (Tokyo) ; 72(5): 306-310, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30792517

RESUMO

The isolation and structure elucidation of one new fungal metabolite, phenguignardic acid butyl ester (1a), and four previously reported metabolites (1b, 2a, 3-4) from the citrus phytopathogen Phyllosticta citricarpa LGMF06 are described. The new dioxolanone phenguignardic acid butyl ester (1a) had low phytotoxic activity in citrus leaves and fruits (at dose of 100 µg), and its importance as virulence factor in citrus black spot disease needs to be further addressed. Beside the phytotoxic analysis, we also evaluated the antibacterial (against methicillin sensitive and resistant Staphylococcus aureus) and cytotoxic (A549 non-small cell lung cancer, PC3 prostate cancer and HEL 299 normal epithelial lung) activities of the isolated compounds, which revealed that compounds 1a, 1b and 2a were responsible for the antibacterial activity of this strain.


Assuntos
Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Ascomicetos/química , Produtos Biológicos/isolamento & purificação , Produtos Biológicos/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Antibacterianos/química , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Produtos Biológicos/química , Produtos Biológicos/toxicidade , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citrus/microbiologia , Humanos , Estrutura Molecular , Doenças das Plantas/microbiologia
18.
Plant Dis ; 103(4): 619-628, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30789317

RESUMO

Phytophthora root rot, caused by several species of Phytophthora, is an important disease of citrus in California and other growing regions. For chemical management, mefenoxam and potassium phosphite have been available for many years, and resistance in Phytophthora spp. has been reported for both compounds. We evaluated the efficacy of the new Oomycota fungicides ethaboxam, fluopicolide, mandipropamid, and oxathiapiprolin, each with a different mode of action, against Phytophthora root rot of citrus in field and greenhouse studies. Root balls of navel orange trees on 'Carrizo citrange' rootstock were inoculated with P. nicotianae at planting in the field in fall 2013. Applications with 11 fungicide treatments were made 5 weeks after planting, in spring and fall 2014, and in spring 2015. Feeder roots and adjacent soil were collected before or after application. All of the new fungicides significantly reduced root rot incidence and Phytophthora soil populations to very low levels as compared with the control starting after the first application. Mefenoxam was only effective when a high label rate was used in the fourth application. Selected treatments also increased tree canopy size, trunk diameter, and fruit yield as compared with the control. A rate comparison with the four new fungicides was initiated in summer 2016 in another field trial using navel orange trees inoculated with P. citrophthora. Minimum effective rates to reduce Phytophthora root rot incidence and pathogen soil populations were determined after one and two applications in fall 2016 and summer 2017, respectively. Greenhouse studies confirmed the efficacy of the new fungicides. Based in part on our studies, fluopicolide recently received a federal and oxathiapiprolin a full registration for use on citrus, and registrations for ethaboxam and mandipropamid have been requested. These new compounds will provide highly effective treatment options and resistance management strategies using rotation and mixture programs for the control of Phytophthora root rot of citrus.


Assuntos
Citrus , Fungicidas Industriais , Phytophthora , Doenças das Plantas , California , Citrus/microbiologia , Fungicidas Industriais/farmacologia , Phytophthora/efeitos dos fármacos , Doenças das Plantas/prevenção & controle , Raízes de Plantas/microbiologia
19.
BMC Genomics ; 20(1): 55, 2019 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-30654743

RESUMO

BACKGROUND: The gram-negative Xanthomonas genus contains a large group of economically important plant pathogens, which cause severe diseases on many crops worldwide. The diffusible signal factor (DSF) - mediated quorum sensing (QS) system coordinates expression of virulence factors in plant pathogenic Xanthomonas spp. However, the regulatory effects of this system during the Xanthomonas- plant interactions remain unclear from both the pathogen and host aspects. RESULTS: In this study, we investigated the in planta DSF- mediated QS regulon of X. citri subsp. citri (Xac), the causal agent of citrus canker. We also characterized the transcriptional responses of citrus plants to DSF-mediated Xac infection via comparing the gene expression patterns of citrus trigged by wild type Xac strain 306 with those trigged by its DSF- deficient (∆rpfF) mutant using the dual RNA-seq approach. Comparative global transcript profiles of Xac strain 306 and the ∆rpfF mutant during host infection revealed that DSF- mediated QS specifically modulates bacterial adaptation, nutrition uptake and metabolisms, stress tolerance, virulence, and signal transduction to favor host infection. The transcriptional responses of citrus to DSF-mediated Xac infection are characterized by downregulation of photosynthesis genes and plant defense related genes, suggesting photosynthetically inactive reactions and repression of defense responses. Alterations of phytohormone metabolism and signaling pathways were also triggered by DSF-mediated Xac infection to benefit the pathogen. CONCLUSIONS: Collectively, our findings provide new insight into the DSF- mediated QS regulation during plant-pathogen interactions and advance the understanding of traits used by Xanthomonas to promote infection on host plants.


Assuntos
Proteínas de Bactérias/metabolismo , Citrus/microbiologia , Doenças das Plantas/microbiologia , Percepção de Quorum , Xanthomonas/metabolismo , Proteínas de Bactérias/genética , Parede Celular/metabolismo , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Genes Bacterianos , Interações Hospedeiro-Patógeno/genética , Ferro/metabolismo , Fotossíntese , Reguladores de Crescimento de Planta/metabolismo , Regulon/genética , Metabolismo Secundário/genética , Transcrição Genética , Transcriptoma/genética , Xanthomonas/genética
20.
Curr Microbiol ; 76(3): 279-289, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30689005

RESUMO

Endophytic fungi are characterized as microorganisms found within internal tissues of living plants without any immediate, overtly negative effects. The present study was carried out to isolate, taxonomically characterize and determine the spatiotemporal distribution of endophytic fungi associated with leaf, stem, trunk, and root of mandarin (Citrus reticulata cv. Siyahoo). To do so, the sampling program was done seasonally in four geographically isolated mandarin growing areas of Hormozgan province of Iran, including Siyahoo, Ahmadi, Sikhoran, and Roudan. In total, 702 fungal isolates were obtained from leaf, stem, trunk, and root of healthy mandarin trees divided into 26 distinct morphotypes based on morphological characteristics. The morphotypes were taxonomically characterized through phylogenetic analysis of the ITS1-5.8S-ITS4 rDNA region sequences. Accordingly, 10 different fungal orders from 5 fungal classes were identified, i.e., Saccharomycetes (Saccharomycetales), Eurotiomycetes (Eurotiales), Dothideomycetes (Capnodiales, Pleosporales, Dothideales), and Sordariomycetes (Diaporthales, Hypocreales, Microascales, Togniniales), all from Ascomycota, which represented 97.2% and Ustilaginomycetes (Ustilaginales) from Basidiomycota which represented 2.8% of the isolates. The Aureobasidium pullulans, Penicillium citrinum, and Dothideomycetes sp. were the most frequent isolates. The trunk and leaf showed the highest and lowest total colonization frequency and species richness of endophytic fungi, respectively, in all sampling periods. The results showed that the colonization frequency of endophytes in Hormozgan province was higher in autumn than that in spring, winter, and summer. The trunk showed the maximum diversity of endophytes over all seasons. The Shannon-Wiener (H') and Simpson indices had significant correlation with sampling cites and tissue type and the maximum value of Shannon and Simpson indices (H' = 3.05 and 1 - D = 0.94) was found in the specimens collected from Siyahoo. In conclusion, the three factors (season, location, and tissue type) all in together could determine fungal endophyte composition of C. reticulata.


Assuntos
Citrus/microbiologia , Fungos/classificação , Fungos/isolamento & purificação , Biodiversidade , DNA Ribossômico/genética , Endófitos/classificação , Endófitos/isolamento & purificação , Geografia , Irã (Geográfico) , Filogenia , Folhas de Planta/microbiologia , Raízes de Plantas/microbiologia , Caules de Planta/microbiologia , Estações do Ano , Especificidade da Espécie
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