scholarly journals Amplificação isotérmica mediada por loop para detecção de patógenos de plantas

2022 ◽  
pp. 7-34
Author(s):  
Carolina Garcia Neves ◽  
Ismail Teodoro de Souza Júnior ◽  
Danielle Ribeiro de Barros
Keyword(s):  
Plant Pathogens ◽  
Bacillus Stearothermophilus ◽  
Research Area ◽  
Molecular Technique ◽  
Naked Eye ◽  
Molecular Tests ◽  
Amplification Method ◽  
Target Dna ◽  
Sophisticated Equipment ◽  
Bacterium Bacillus

Disease control is crucial to minimize potential losses in agriculture and thereby maintain high crop yield. However, for its effectiveness, the pathogen must be detected early and correctly in the production fields. Different methods of diagnosis can be used, from those based on symptoms to molecular tests. Loop-mediated isothermal amplification (LAMP) is a molecular technique that has been widely used in several biological fields, due to the ease with which it can be applied. The reaction can be carried out in a single thermal condition, due to the use of Bst DNA polymerase, isolated from the bacterium Bacillus stearothermophilus, which has high displacement activity. LAMP is a highly exponential amplification method that produces the target DNA in amounts 109 -1010 times between 45 and 60 minutes at 60-65°C. Its advantages are the visualization of results directly with the naked eye and the fact that it does not need sophisticated equipment for its application. In phytopathology, the technique has been gaining prominence in the detection of fungi, viruses, bacteria, nematodes and phytoplasmas, as well as in the monitoring of fungicide-resistant fungi. LAMP can benefit agriculture so that early, accurate and sensitive diagnostics can be carried out in the fields of cultivation and minimize losses caused by diseases. In this review, we present and discuss LAMP tests, developed for plant pathogens detection, which can be useful for researchers who wish to use the technique in their research area

scholarly journals A Novel Rhizospheric Bacterium, Bacillus Velezensis NKMV-3 as a Biocontrol Agent against Alternaria Leaf Blight in Tomato

2021 ◽  
Author(s):  
Vignesh Murthy ◽  
VedhaHari BodethalaNarayanan ◽  
MubarakAli Davoodbasha ◽  
MadhanShankar ShankarRamakrishanan
Keyword(s):  
Early Blight ◽  
Plant Pathogens ◽  
Alternaria Solani ◽  
Inhibitory Effect ◽  
Rrna Gene ◽  
Bacillus Velezensis ◽  
Bacterium Bacillus ◽  
Blight Disease ◽  
Molecular Sequencing

Abstract A novel strain of Bacillus isolated from rhizosphere has shown to be excellent biocontrol agents against various plant pathogens. In this study, a first report of a Bacillus strain NKMV-3 which effectively controlling Alternaria solani, which cause the Early Blight disease in tomato. Based on the cultural and molecular sequencing of 16S rRNA gene sequence, the identity of the strain was confirmed as Bacillus velezensis NKMV-3. The presence of the lipopeptide which are antibiotic synthesis genes namely Iturin C, Surfactin A, Fengycin B and D were confirmed through gene amplification. In addition, lipopetides was also confirmed through liquid chromatography. The extract showed inhibitory effect against A.solani in-vitro and detached tomato leaf assays. Bacillus velezensis strain NKMV-3 based formulations may provide an effective solution in controlling early blight disease in tomato and other crops.

scholarly journals Analysis of microbial populations in plastic–soil systems after exposure to high poly(butylene succinate-co-adipate) load using high-resolution molecular technique

2021 ◽  
Vol 33 (1) ◽  
Author(s):  
Benjawan Tanunchai ◽  
Kantida Juncheed ◽  
Sara Fareed Mohamed Wahdan ◽  
Vusal Guliyev ◽  
Maria Udovenko ◽  
...  
Keyword(s):  
High Resolution ◽  
Community Composition ◽  
Plant Pathogens ◽  
Microbial Control ◽  
High Load ◽  
Soil Microbiome ◽  
Molecular Technique ◽  
Human Pathogens ◽  
Soil Microbial Diversity ◽  
Surrounding Soil

Abstract Background Bio-based and biodegradable plastics are considered as plastics of the future owing to their ability to decompose under various environmental conditions. However, their effects on the soil microbiome are poorly characterised. In this study, we aimed to investigate the effects of an important bio-based and biodegradable plastic, polybutylene succinate-co-adipate (PBSA), on soil microbial diversity and community composition using high-resolution molecular technique (Illumina sequencing) targeting all three microbial domains: archaea, bacteria, and fungi. Results Adding high load of PBSA to soil (6% (w/w)) caused a significant decline in archaeal (13%) and fungal (45%) richness and substantial changes in both bacterial (Proteobacteria, Actinobacteria, and Acidobacteria) and fungal (Eurotiomycetes, Sordariomycetes, Leotiomycetes, and Dothideomycetes) community composition compared with no PBSA addition to soil. The combined effects of PBSA and (NH4)2SO4 fertilisation on the soil microbiome were much greater than the effects of PBSA alone. We only detected opportunistic human pathogens in low abundance on PBSA and in the surrounding soil. However, some plant pathogenic fungi were detected and/or enriched on the PBSA films and in surrounding soil. Apart from plant pathogens, many potential microbial control agents and plant growth-promoting microorganisms were also detected/enriched owing to PBSA addition. Adding high load of PBSA together with (NH4)2SO4 fertilisation can either eliminate some plant pathogens or enrich specific pathogens, especially Fusarium solani, which is economically important. Conclusions We conclude that high load of bio-based and biodegradable PBSA plastic may negatively affect soil microbiome.

A Review on the Efficiency of NASBA Molecular Technique for Diagnosis of Acute Toxoplasmosis in Pregnant Mothers

2021 ◽  
Author(s):  
Mohammad Shabanpour ◽  
Yahya Ehteshaminia ◽  
Shayan Poyandeh ◽  
Seif Ali Mahdavi
Keyword(s):  
Nucleic Acid ◽  
Detection Methods ◽  
Molecular Technique ◽  
Acid Extraction ◽  
Rna Amplification ◽  
Isothermal Method ◽  
Amplification Method ◽  
Acute Form ◽  
Acute Toxoplasmosis ◽  
Pregnant Mothers

Introduction: Toxoplasmosis is a worldwide disease caused by an intracellular protozoan called Toxoplasma gondii and in mothers who become infected during pregnancy, it can cause serious damage to the fetus by passing through the placenta. The aim of this study was to review the effectiveness of the NASBA molecular technique in diagnosing the acute form of Toxoplasmosis in pregnant mothers.  Material and Methods: In this study, the websites of PubMed, Google Scholar, SID, Magiran, Web of Science, IranDoc were searched and articles related to the title have been reviewed from 1990 to 2020.    Results: Nucleic Acid Sequence-Based Amplification (NASBA) is an isothermal method that has the processes of nucleic acid extraction, amplification, and identification of amplified products. This technique is based on transcription and is specifically used for RNA amplification, so it is highly specific in identifying living and active microorganisms. All steps in this amplification method are performed at 41 °C and the amplified products can be identified by appropriate detection methods such as electrochemical luminescence (ECL). Conclusion: Since all steps of amplification are performed by NASBA at the same temperature of 41°C, unlike molecular PCR technique, a thermocycler is not required, so setting it up will not cost much for laboratories and it can be useful in providing a suitable solution for toxoplasmosis screening in pregnant mothers.

scholarly journals Functional proteomics and correlated signaling pathway of the thermophilic bacterium Bacillus stearothermophilus TLS33 under cold-shock stress

PROTEOMICS ◽  
2005 ◽  
Vol 5 (17) ◽  
pp. 4456-4471 ◽  
Author(s):  
Supachai Topanurak ◽  
Supachok Sinchaikul ◽  
Boonyaras Sookkheo ◽  
Suree Phutrakul ◽  
Shui-Tein Chen
Keyword(s):  
Signaling Pathway ◽  
Bacillus Stearothermophilus ◽  
Cold Shock ◽  
Thermophilic Bacterium ◽  
Functional Proteomics ◽  
Bacterium Bacillus ◽  
Shock Stress

scholarly journals A Novel Rapid Visual Detection of Toxoplasma Gondii by Combining Recombinase Polymerase Amplification and Lateral Flow Dipstick Coupled With CRISPR-Cas13a Fluorescence Assay

2021 ◽  
Author(s):  
Jinhong Zhao ◽  
Yuanyuan Li ◽  
Qiqi Xue ◽  
Zhiwei Zhu ◽  
Minghui Zou ◽  
...  
Keyword(s):  
Visual Detection ◽  
High Specificity ◽  
Lateral Flow ◽  
Parasitic Disease ◽  
Blood Samples ◽  
Naked Eye ◽  
Detection Assay ◽  
Lateral Flow Dipstick ◽  
Sophisticated Equipment ◽  
Positive Rate

Abstract Background: Toxoplasmosis caused by infecting with Toxplasma gondii is a kind of parasitic disease that prevalent all over the world and does great harm to pregnant women and newborns. Effective, rapid and accurate diagnosis T. gondii is urgently needed to prevent and treatment the toxoplasmosis. The purpose of this study was to develop a rapid visual detection assay using recombinase aided amplification (RAA) and lateral flow dipstick (LFD) coupled with CRISPR-Cas13a fluorescence, henceforth RAA-Cas13a-LFD, for detection of T. gondii.Methods: Targeting 529bp gene of T. gondii, the primers and probes for RAA-Cas13a-LFD assay were designed and screened. The reaction time of RAA-LFD-Cas13a assay was optimized, as well as the sensitivity and specificity was further validated. Finally, the diagnostic performance of T. gondii was evaluated using the RAA-Cas13a-LFD assay for clinical blood samples.Results: The RAA-Cas13a-LFD assay was performed in an incubator block at 37℃ within 2h, and the amplicons were visible through LFD for naked eye visualization. The detection limit of the developed RAA-Cas13a-LFD assay was 1×10-6 ng/μL with high specificity for T. gondii. Compared with qPCR assay, there was a consistent positive rate among the clinical blood samples. Conclusion: In this study, A rapid and visual RAA-Cas13a-LFD assay was developed. It requires no sophisticated equipment and shows promise for on-site surveillance of T. gondii.

scholarly journals Phytopathogenic species of fungi and fungal-like organisms identified in plant samples delivered to the Plant Disease Clinic in 2018–2020

2021 ◽  
Vol 61 (3) ◽  
Keyword(s):  
Host Plants ◽  
Plant Disease ◽  
Plant Pathogens ◽  
Ornamental Plants ◽  
Single Species ◽  
Plant Diseases ◽  
Effective Management ◽  
Fusarium Spp ◽  
Molecular Tests ◽  
Disease Clinic

One of the conditions for effective management of farm is an access to quick diagnostics of plant pathogens in order to reduce the occurrence of plant diseases. The Plant Diseases Clinic receives samples of infected plants supplied by growers and gardeners from all over Poland. In the years 2018–2020, a total of 274 samples were tested at the Clinic for the presence of fungi and fungal-like organisms pathogenic for plants. The tests were carried out using the microscopic method, and in case of doubt, the result was confirmed by molecular tests. The most frequently studied plant was tomato (26%), followed by strawberry (9%), cucumber (5%) and tobacco, sugar beet, onion, blueberry, raspberry, lettuce, cauliflower and potato. Conifers were also a large group, such as: thujas, cypresses and pines; a total of 17 host plants. Single species of ornamental plants were very numerous, e.g. gerbera, anthurium, aster, geranium, phlox, chrysanthemum and others. The fungi of the genus Fusarium spp. constituted about 38% of infections. This was followed by Alternaria spp. (26%), Botrytis cinerea (11%) and Cladosporium sp. (10%). The remaining diseases were caused by Pythium sp., Rhizoctonia sp., Colletotrichum sp., Ulocladium sp., Pestalotia sp. and Phytophthora sp. In recent years, the greatest threat to tomatoes and strawberries has been the fungi of the Fusarium genus, and the pathogens of the Pythium genus to cucumbers.

Loop-Mediated Isothermal Amplification Method for Rapid Detection of Shigella dysenteriae

2011 ◽  
Vol 140 ◽  
pp. 369-373 ◽  
Author(s):  
Qing Ping Zhong ◽  
Li Wang ◽  
Bin Wang ◽  
Hong Yuan Chen
Keyword(s):  
Isothermal Condition ◽  
Lamp Assay ◽  
High Specificity ◽  
Isothermal Amplification ◽  
Shigella Dysenteriae ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification ◽  
Amplification Method ◽  
Target Dna ◽  
Pcr Method

The study was aimed to develop a loop-mediated isothermal amplification (LAMP) method which amplifies DNA with high specificity and rapidity for the detection of Shigella dysenteriae. A set of four primers was designed for recognizing six distinct sequences on the target ipaH of S. dysenteriae. By the method, the target DNA was amplified within 1h under isothermal condition at 65 °C. The sensitivities of the LAMP for detecting pure culture and genomic DNA were 1.04 CFU/ml and 1.06 fg/μl, while the sensitivities of PCR method were 1.04×102 CFU/ml and 1.06 pg/μl. Furthermore, the LAMP assay was examined for its ability to detect S. dysenteriae in artificially contaminated lettuce sample, the detection limits of this LAMP assay and the PCR method were 4.60 CFU/g and 4.60×102 CFU/g, respectively.

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