scholarly journals The Tobacco Trichome Exudate Z-abienol and Its Relationship With Plant Resistance to Phytophthora nicotianae

Plant Disease ◽  
2017 ◽  
Vol 101 (7) ◽  
pp. 1214-1221 ◽  
Author(s):  
William T. Steede ◽  
Justin M. Ma ◽  
David P. Eickholt ◽  
Katherine E. Drake-Stowe ◽  
Sheri P. Kernodle ◽  
...  
Keyword(s):  
Close Association ◽  
Hyphal Growth ◽  
Field Resistance ◽  
Phytophthora Nicotianae ◽  
In Vitro Assays ◽  
Black Shank ◽  
Plant Parts ◽  
Tobacco Cultivar ◽  
High Level

In previous research, we discovered a favorable quantitative trait locus (QTL) in cigar tobacco cultivar ‘Beinhart 1000’ designated as Phn15.1, which provides a high level of partial resistance to the black shank disease caused by Phytophthora nicotianae. A very close genetic association was also found between Phn15.1 and the ability to biosynthesize Z-abienol, a labdanoid diterpene exuded by the trichomes onto above-ground plant parts, and that imparts flavor and aroma characteristics to Oriental and some cigar tobacco types. Because accumulation of Z-abienol is considered to be undesirable for cultivars of other tobacco types, we herein describe a series of experiments to gain insight on whether this close association is due to genetic linkage or pleiotropy. First, in an in vitro bioassay, we observed Z-abienol and related diterpenes to inhibit hyphal growth of P. nicotianae at concentrations between 0.01 and 100 ppm. Secondly, we field-tested transgenic versions of Beinhart 1000 carrying RNAi constructs for downregulating NtCPS2 or NtABS, two genes involved in the biosynthesis of Z-abienol. Thirdly, we also field tested a recombinant inbred line population segregating for a truncation mutation in NtCPS2 leading to an interrupted Z-abienol pathway. We observed no correlation between field resistance to P. nicotianae and the ability to accumulate Z-abienol in either the transgenic materials or the mapping population. Results suggest that, although Z-abienol may affect P. nicotianae when applied at high concentrations in in vitro assays, the compound has little effect on black shank disease development under natural field conditions. Thus, it should be possible to disassociate Phn15.1-mediated black shank resistance identified in cigar tobacco cultivar Beinhart 1000 from the ability to accumulate Z-abienol, an undesirable secondary metabolite for burley and flue-cured tobacco cultivars.

scholarly journals In vitro evaluation of fungal antagonists of Phytophthora nicotianae

2017 ◽  
Vol 38 (SI 2 - 6th Conf EFPP 2002) ◽  
pp. 634-637
Author(s):  
R. Nicoletti ◽  
F. Raimo ◽  
E. Cozzolino
Keyword(s):  
Mycelial Growth ◽  
Central Italy ◽  
Phytophthora Nicotianae ◽  
Aspergillus Sydowii ◽  
Black Shank ◽  
Culture Filtrates ◽  
Dual Cultures ◽  
Penicillium Brevicompactum ◽  
Fusarium Chlamydosporum

As tobacco black shank epidemics caused by Phytophthora nicotianae occurred in central Italy in the late 1990s, fungal antagonists of the pathogen were searched in the rhizosphere of tobacco plants. Isolates of Aspergillus sydowii, Fusarium chlamydosporum, Gliocladium roseum, Penicillium brevicompactum, P. chrysogenum, Scopulariopsis candida and Trichoderma harzianum were recovered. Antagonism of these isolates toward P. nicotianae was evaluated in vitro: even if no hyphal interactions were observed in dual cultures, aberration in mycelial growth and morphology of sporangia occurred in most cases. Unlike those of T. harzianum, concentrated culture filtrates of A. sydowii, F. chlamydosporum, G. roseum, P. brevicompactum, P. chrysogenum, inhibited growth of all P. nicotianae isolates tested, while culture filtrates of S. candida caused aberrant mycelial growth.

scholarly journals Sensitivity of Phytophthora nicotianae From Tobacco to Fluopicolide, Mandipropamid, and Oxathiapiprolin

Plant Disease ◽  
2016 ◽  
Vol 100 (10) ◽  
pp. 2119-2125 ◽  
Author(s):  
Tianli Qu ◽  
Yuanyuan Shao ◽  
Alexander S. Csinos ◽  
Pingsheng Ji
Keyword(s):  
Mycelial Growth ◽  
Agar Medium ◽  
Phytophthora Nicotianae ◽  
Limited Information ◽  
Growth Reduction ◽  
Pathogen Population ◽  
Baseline Sensitivity ◽  
Resistance Development ◽  
Black Shank

Black shank incited by Phytophthora nicotianae is a devastating disease in the production of tobacco. Fungicides have been commonly used for managing the disease; however, there is only a narrow pool of effective fungicides. A few new fungicides became available in recent years, including fluopicolide, mandipropamid, and oxathiapiprolin, which reduced diseases incited by oomycetes under field conditions. Limited information is available regarding sensitivity of P. nicotianae isolates to these new fungicides. Research was conducted to determine effects of the three new fungicides on P. nicotianae isolates from tobacco in Georgia. Studies with 106 isolates indicated that they did not grow when agar medium was amended with the fungicides at the rate of 1 μg/ml. Twenty isolates were used for in vitro studies to determine sensitivity to the fungicides. Fluopicolide, mandipropamid, and oxathiapiprolin inhibited mycelial growth of the isolates with mean EC50 values (effective concentrations that provide 50% growth reduction) of 0.09, 0.04, and 0.001 μg/ml, respectively. EC50 values of fluopicolide, mandipropamid, and oxathiapiprolin for inhibiting sporangial formation were 0.15, 0.03, and 0.0002 μg/ml, respectively. EC50 values for suppressing zoospore germination averaged 0.16, 0.04, and 0.002 μg/ml for fluopicolide, mandipropamid, and oxathiapiprolin, respectively. Results from the study indicated that P. nicotianae isolates from tobacco in Georgia were sensitive to the fungicides, with lower EC50 for oxathiapiprolin than for fluopicolide and mandipropamid. The information on effectiveness and baseline sensitivity of fungicides on P. nicotianae will facilitate monitoring of resistance development in the pathogen population.

scholarly journals Expression of a Novel Small Antimicrobial Protein from the Seeds of Motherwort (Leonurus japonicus) Confers Disease Resistance in Tobacco

2006 ◽  
Vol 73 (3) ◽  
pp. 939-946 ◽  
Author(s):  
Xingyong Yang ◽  
Yuehua Xiao ◽  
Xiaowen Wang ◽  
Yan Pei
Keyword(s):  
Storage Proteins ◽  
Bacterial Pathogen ◽  
Hyphal Growth ◽  
Antimicrobial Activities ◽  
In Vitro Assays ◽  
Antimicrobial Protein ◽  
Heat Stable ◽  
Antimicrobial Materials ◽  
Rapid Amplification

ABSTRACT Medicinal plants are valuable resources of natural antimicrobial materials. A novel small protein with antimicrobial activities, designated LJAMP1, was purified from the seeds of a medicinal herb, motherwort (Leonurus japonicus Houtt). LJAMP1 is a heat-stable protein with a molecular mass of 7.8 kDa and a determined isoelectric point of 8.2. In vitro assays showed that LJAMP1 inhibits the growth of an array of fungi and bacteria. The hyphal growth inhibition by LJAMP1 was more evident against hyphomycete fungi, such as Alternaria alternata, Cercospora personata, and Aspergillus niger. The N-terminal amino acid sequence of LJAMP1 was determined, and its coding gene was consequently cloned by the rapid amplification of cDNA ends. The gene LJAMP1 has no intron and encodes a polypeptide of 95 amino acids, in which the first 27 residues was deduced as a signal peptide. The mature LJAMP1 shows relatively low identity to plant napin-like storage proteins. Northern blot assays revealed that LJAMP1 is expressed preferentially in seeds. Bioassays in transgenic tobacco demonstrated that that overexpression of LJAMP1 significantly enhanced the resistance of tobacco against not only the fungal pathogen A. alternata but also the bacterial pathogen Ralstonia solanacearum, while no visible alteration in plant growth and development was observed.

Virulent bacteriophage infection of Rhizobium leguminosarum

1972 ◽  
Vol 18 (4) ◽  
pp. 375-384 ◽  
Author(s):  
A. N. Ley ◽  
H. R. Warner ◽  
Phyllis L. Kahn
Keyword(s):  
Molecular Weight ◽  
Rna Synthesis ◽  
Rhizobium Leguminosarum ◽  
Deoxyribonucleic Acid ◽  
In Vitro Assays ◽  
Tail Fiber ◽  
Bacteriophage Infection ◽  
Cell Extracts ◽  
High Level

Bacteriophage 317, which virulently infects Rhizobium leguminosarum, has an eclipse period of 60–70 min and a latent period of 100 min at 30°. Electron micrographs of the phage indicated head, tail, and tail-fiber structural components.Base analysis of phage 317 deoxyribonucleic acid (DNA) indicated the presence of equimolar amounts of adenine and thymine and of guanine and cytosine, which suggests that the DNA is double stranded. The DNA has a molecular weight of 41 × 106 daltons as determined from electron micrographs.The results of 14C-uracil incorporation studies showed that net ribonucleic acid (RNA) synthesis was markedly inhibited after infection. There was a slight stimulation in DNA synthesis after infection as indicated by 14C-thymidine incorporation.The results of in vitro assays of enzymes involved in the biosynthesis of DNA showed that deoxyuridine 5′-triphosphate nucleotidohydrolase (dUTPase) and deoxythymidine 5′-monophosphate kinase (dTMP kinase) increased 50- and 30-fold respectively, after infection. The reason for the increased dUTPase activity is not readily apparent. Phage 317 DNA contains only the standard bases, unlike the DNA of other phages that induce an increase in this enzyme after infection. A high level of deoxythymidine 5′-monophosphatase (dTMPase) was observed in both uninfected and infected crude cell extracts. Further work is necessary to see if similar changes occur in Rhizobium during the establishment of symbiosis with legumes.

scholarly journals Antagonism of Trichoderma harzianum ETS 323 on Botrytis cinerea Mycelium in Culture Conditions

2012 ◽  
Vol 102 (11) ◽  
pp. 1054-1063 ◽  
Author(s):  
Chi-Hua Cheng ◽  
Chia-Ann Yang ◽  
Kou-Cheng Peng
Keyword(s):  
Amino Acid ◽  
Botrytis Cinerea ◽  
Trichoderma Harzianum ◽  
Hyphal Growth ◽  
Extracellular Proteins ◽  
In Vitro Assays ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
Trichoderma Spp

Previous studies have shown that the extracellular proteins of Trichoderma harzianum ETS 323 grown in the presence of deactivated Botrytis cinerea in culture include a putative l-amino acid oxidase and have suggested the involvement of this enzyme in the antagonistic mechanism. Here, we hypothesized that the mycoparasitic process of Trichoderma spp. against B. cinerea involves two steps; that is, an initial hyphal coiling stage and a subsequent hyphal coiling stage, with different coiling rates. The two-step antagonism of T. harzianum ETS 323 against B. cinerea during the mycoparasitic process in culture was evaluated using a biexponential equation. In addition, an l-amino acid oxidase (Th-l-AAO) was identified from T. harzianum ETS 323. The secretion of Th-l-AAO was increased when T. harzianum ETS 323 was grown with deactivated hyphae of B. cinerea. Moreover, in vitro assays indicated that Th-l-AAO effectively inhibited B. cinerea hyphal growth, caused cytosolic vacuolization in the hyphae, and led to hyphal lysis. Th-l-AAO also showed disease control against the development of B. cinerea on postharvest apple fruit and tobacco leaves. Furthermore, an apoptosis-like response, including the generation of reactive oxygen species, was observed in B. cinerea after treatment with Th-l-AAO, suggesting that Th-l-AAO triggers programmed cell death in B. cinerea. This may be associated with the two-step antagonism of T. harzianum ETS 323 against B. cinerea.

scholarly journals Characterization of the Black Shank Pathogen, Phytophthora nicotianae, Across North Carolina Tobacco Production Areas

Plant Disease ◽  
2018 ◽  
Vol 102 (6) ◽  
pp. 1108-1114 ◽  
Author(s):  
Courtney A. Gallup ◽  
Kestrel L. McCorkle ◽  
Kelly L. Ivors ◽  
David Shew
Keyword(s):  
North Carolina ◽  
Natural Populations ◽  
Hyphal Growth ◽  
The United States ◽  
The State ◽  
Phytophthora Nicotianae ◽  
Mating Types ◽  
Black Shank ◽  
Nuclear Regions

Black shank disease of tobacco, caused by the oomycete Phytophthora nicotianae, is a major threat to production in the United States and tobacco-producing areas worldwide. In a statewide survey of North Carolina, the rapid shift from race 0 to race 1 was documented. Collected pathogen isolates were characterized phenotypically for mating type and mefenoxam sensitivity, and genotypically by comparing sequences from three cytoplasmic and two nuclear regions. Both the A1 and A2 mating types were found throughout the state. When both mating types were recovered from the same field, pairings of isolates yielded viable oospores, indicating for the first time the potential for sexual sporulation by P. nicotianae in natural populations. Because the loss of complete resistance required a renewed use of the fungicide mefenoxam, a subset of the survey isolates was screened for sensitivity to the fungicide. All isolates were sensitive, with a mean effective concentration to inhibit 50% of hyphal growth of 0.4 μg/ml that was similar across mating types and races. Molecular characterization of 226 isolates revealed that the pathogen exists as multiple clonal types within the state. Genetic diversity among the pathogen population and the potential for sexual recombination may help explain the ability of the pathogen to rapidly adapt to host resistance genes.

scholarly journals Biocontrol of PGPR strain Bacillus amyloliquefaciens Ba168 against Phytophthora nicotianae on tobacco

2019 ◽  
Author(s):  
Dongsheng Guo ◽  
Chenhong Yuan ◽  
Yunyan Luo ◽  
YaHan Chen ◽  
Meihuan Lu ◽  
...  
Keyword(s):  
Crude Extract ◽  
Dry Weight ◽  
Plant Growth Promoting Rhizobacteria ◽  
Sem Analysis ◽  
Phytophthora Nicotianae ◽  
Black Shank ◽  
Biocontrol Efficacy ◽  
Plant Growth Promoting ◽  
Pgpr Strain

AbstractTobacco black shank (TBS) caused by Phytophthora nicotianae is destructive to almost all kinds of tobacco cultivars and is widespread in many tobacco-planted countries. Here, an isolated plant growth-promoting rhizobacteria (PGPR) strain Ba168 is promise in biocontrol of TBS. In vitro assays disclosed a strong P. nicotianae suppression activity and the field utilization potential (FUP) by characterized the crude extract of culture filtrates of Ba168. P. nicotianae’s growth was inhibited by the crude extract at minimum inhibitory concentration (MIC) of 5μl/mL. Extracellular conductivity, pH and the wet, dry weight of P. nicotianae’s mycelia, were significantly different after treated with different concentrations of the crude extract and the deformity and perforation of treated P. nicotianae’s hyphae can be observed in scanning electron microscope (SEM) analysis. Proteome characterizations of the crude extract were used as supplementary proofs that further evaluated FUP of Ba168. We then identified strain Ba168 as B. amyloliquefaciens by its genetic and phenotypic characteristics. Field assays comparatively evaluated TBS control efficacy of these PGPRs and agrochemicals. Pooling analysis of the results showed that the biocontrol efficacy of Ba168 preparation is only lower than Mixture of Propamocarb hydrochloride and Azoxystrobin (MPA) but better than other tested subjects. Although the existence of differences in biocontrol efficacy, PGPR preparations effectively reduced the disease index of tobacco.ImportanceThis work demonstrates the promising biocontrol potential of B. amyloliquefaciens Ba168 and highlights the positive roles of PGPR in suppression of this soil-borne disease.

Production of Mycobacterium bovis Antigens Included in Recombinant Occlusion Bodies of Baculovirus

2019 ◽  
Vol 29 (1-6) ◽  
pp. 83-90
Author(s):  
Luciana Villafañe ◽  
Marina Andrea Forrellad ◽  
María Gabriela López ◽  
Sergio Garbaccio ◽  
Carlos Garro ◽  
...  
Keyword(s):  
Mycobacterium Bovis ◽  
Intradermal Injection ◽  
In Vitro Assays ◽  
Chimeric Proteins ◽  
Purified Protein Derivative ◽  
E Coli ◽  
Occlusion Bodies ◽  
Ifn Γ ◽  
High Level

Bovine tuberculosis (bTB) is a disease produced by <i>Mycobacterium bovis</i> that affects livestock, wild animals, and humans. The classical diagnostic method to detect bTB is measuring the response induced with the intradermal injection of purified protein derivative of <i>M. bovis</i> (PPDb). Another ancillary bTB test detects IFN-γ produced in whole blood upon stimulation with PPDb, protein/peptide cocktails, or individual antigens. Among the most used <i>M. bovis</i> antigens in IFN-γ assays are the secreted proteins ESAT-6 and CFP-10, which together with antigen Rv3615c improve the sensitivity of the test in comparison to PPDb. Protein reagents for immune stimulation are generally obtained from <i>Escherichia coli</i>, because this bacterium produces a high level of recombinant proteins. However, <i>E. coli</i> recombinant antigens are in general contaminated with lipopolysaccharides and other components that produce non-specific IFN-γ secretion in in vitro assays. In this work, we produced the relevant ESAT-6, CFP-10, and Rv3615c <i>M. bovis</i> antigens as fusions to the polyhedrin protein from the baculovirus AcMNPV. We obtained chimeric proteins effectively incorporated to the occlusion bodies and easily purified the recombinant polyhedra with no reactive contaminants. In an IFN-γ assay, these fusion proteins showed equivalent sensibility but better specificity than the same <i>M. bovis</i> proteins produced in <i>E. coli</i>.

scholarly journals Feedback Regulation of N Fixation in Frankia-Alnus Symbiosis Through Amino Acids Profiling in Field and Greenhouse Nodules

2020 ◽  
Vol 33 (3) ◽  
pp. 499-508
Author(s):  
Anne-Emmanuelle Hay ◽  
Aude Herrera-Belaroussi ◽  
Marjolaine Rey ◽  
Pascale Fournier ◽  
Philippe Normand ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nitrogen Fixation ◽  
Amino Acid ◽  
Feedback Regulation ◽  
Metabolic Model ◽  
In Vitro Assays ◽  
N Fixation ◽  
Abundance Pattern ◽  
High Level

Symbiosis established between actinorhizal plants and Frankia spp., which are nitrogen-fixing actinobacteria, promotes nodule organogenesis, the site of metabolic exchange. The present study aimed to identify amino acid markers involved in Frankia-Alnus interactions by comparing nodules and associated roots from field and greenhouse samples. Our results revealed a high level of citrulline in all samples, followed by arginine (Arg), aspartate (Asp), glutamate (Glu), γ-amino-n-butyric acid (GABA), and alanine (Ala). Interestingly, the field metabolome approach highlighted more contrasted amino acid patterns between nodules and roots compared with greenhouse samples. Indeed, 12 amino acids had a mean relative abundance significantly different between field nodule and root samples, against only four amino acids in greenhouse samples, underlining the importance of developing “ecometabolome” approaches. In order to monitor the effects on Frankia cells (respiration and nitrogen fixation activities) of amino acid with an abundance pattern evocative of a role in symbiosis, in-vitro assays were performed by supplementing them in nitrogen-free cultures. Amino acids had three types of effects: i) those used by Frankia as nitrogen source (Glu, Gln, Asp), ii) amino acids stimulating both nitrogen fixation and respiration (e.g., Cit, GABA, Ala, valine, Asn), and iii) amino acids triggering a toxic effect (Arg, histidine). In this paper, a N-metabolic model was proposed to discuss how the host plant and bacteria modulate amino acids contents in nodules, leading to a fine regulation sustaining high bacterial nitrogen fixation.

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