Phytoplasma transmission by in vitro graft inoculation as a basis for a preliminary screening method for resistance in fruit trees

1999 ◽  
Vol 48 (2) ◽  
pp. 283-287 ◽  
Author(s):  
Jarausch ◽  
Lansac ◽  
Bliot ◽  
Dosba
Keyword(s):  
Screening Method ◽  
Fruit Trees ◽  
Preliminary Screening

A Modified Technique for Preliminary Screening of Compounds for Anthelmintic Activity against Eggs and Larvae of Bursate Nematodes of Sheep

1964 ◽  
Vol 38 (1-2) ◽  
pp. 47-56 ◽  
Author(s):  
I. W. Parnell
Keyword(s):  
Screening Method ◽  
Anthelmintic Activity ◽  
Modified Technique ◽  
Free Living ◽  
Preliminary Screening ◽  
Small Glass ◽  
Glass Tubes ◽  
Eggs And Larvae ◽  
Made In

A technique is described for measuring the larvicidal activity of compounds against the free-living stages of bursate nematodes of sheep. Cultures were made in small glass tubes in 2 oz. “pomade” jars, and the larvicidal effects of the compounds were measured by comparing the number of infective larvae which were recovered from treated cultures with the numbers from the control cultures.This technique provides another in vitro screening method for compounds which may be effective as anthelmintics against bursate nematodes.

An integrated screening method for evaluating the pulmonary toxicity of inhaled particulates: Role of microscopic techniques in assessing pulmonary macrophage clearance functions

1990 ◽  
Vol 48 (3) ◽  
pp. 826-827
Author(s):  
David B. Warheit ◽  
Lena Achinko ◽  
Mark A. Hartsky
Keyword(s):  
Bronchoalveolar Lavage ◽  
Pulmonary Toxicity ◽  
Screening Method ◽  
Pulmonary Response ◽  
Inhaled Particles ◽  
Cytochemical Analysis ◽  
Pulmonary Macrophages ◽  
Integrated Screening

There is a great need for the development of a rapid and reliable bioassay to evaluate the pulmonary toxicity of inhaled particles. A number of methods have been proposed, including lung clearance studies, bronchoalveolar lavage analysis, and in vitro cytotoxicity tests. These methods are often limited in scope inasmuch as they measure only one dimension of the pulmonary response to inhaled, instilled or incubated dusts. Accordingly, a comprehensive approach to lung toxicity studies has been developed.To validate the method, rats were exposed for 6 hours or 3 days to various concentrations of either aerosolized alpha quartz silica (Si) or carbonyl iron (CI) particles. Cells and fluids from groups of sham and dust-exposed animals were recovered by bronchoalveolar lavage (BAL). Alkaline phosphatase, LDH and protein values were measured in BAL fluids at several time points postexposure. Cells were counted and evaluated for viability, as well as differential and cytochemical analysis. In addition, pulmonary macrophages (PM) were cultured and studied for morphology, chemotaxis, and phagocytosis by scanning electron microscopy.

scholarly journals A high-throughput screening method for evolving a demethylase enzyme with improved and new functionalities

2020 ◽  
Author(s):  
Yuru Wang ◽  
Christopher D Katanski ◽  
Christopher Watkins ◽  
Jessica N Pan ◽  
Qing Dai ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Screening Method ◽  
Targeted Mutagenesis ◽  
Rna Repair ◽  
Dna And Rna ◽  
Modified Dna ◽  
Dna Substrates ◽  
Trna Sequencing

Abstract AlkB is a DNA/RNA repair enzyme that removes base alkylations such as N1-methyladenosine (m1A) or N3-methylcytosine (m3C) from DNA and RNA. The AlkB enzyme has been used as a critical tool to facilitate tRNA sequencing and identification of mRNA modifications. As a tool, AlkB mutants with better reactivity and new functionalities are highly desired; however, previous identification of such AlkB mutants was based on the classical approach of targeted mutagenesis. Here, we introduce a high-throughput screening method to evaluate libraries of AlkB variants for demethylation activity on RNA and DNA substrates. This method is based on a fluorogenic RNA aptamer with an internal modified RNA/DNA residue which can block reverse transcription or introduce mutations leading to loss of fluorescence inherent in the cDNA product. Demethylation by an AlkB variant eliminates the blockage or mutation thereby restores the fluorescence signals. We applied our screening method to sites D135 and R210 in the Escherichia coli AlkB protein and identified a variant with improved activity beyond a previously known hyperactive mutant toward N1-methylguanosine (m1G) in RNA. We also applied our method to O6-methylguanosine (O6mG) modified DNA substrates and identified candidate AlkB variants with demethylating activity. Our study provides a high-throughput screening method for in vitro evolution of any demethylase enzyme.

scholarly journals Identification of Potential Kinase Inhibitors within the PI3K/AKT Pathway of Leishmania Species

Biomolecules ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1037
Author(s):  
Rodrigo Ochoa ◽  
Amaya Ortega-Pajares ◽  
Florencia A. Castello ◽  
Federico Serral ◽  
Darío Fernández Do Porto ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Screening Method ◽  
Molecular Targets ◽  
Leishmania Species ◽  
Akt Pathway ◽  
Protein Protein Interaction ◽  
Starting Point ◽  
Binding Pockets ◽  
Protein Protein Interaction Networks

Leishmaniasis is a public health disease that requires the development of more effective treatments and the identification of novel molecular targets. Since blocking the PI3K/AKT pathway has been successfully studied as an effective anticancer strategy for decades, we examined whether the same approach would also be feasible in Leishmania due to their high amount and diverse set of annotated proteins. Here, we used a best reciprocal hits protocol to identify potential protein kinase homologues in an annotated human PI3K/AKT pathway. We calculated their ligandibility based on available bioactivity data of the reported homologues and modelled their 3D structures to estimate the druggability of their binding pockets. The models were used to run a virtual screening method with molecular docking. We found and studied five protein kinases in five different Leishmania species, which are AKT, CDK, AMPK, mTOR and GSK3 homologues from the studied pathways. The compounds found for different enzymes and species were analysed and suggested as starting point scaffolds for the design of inhibitors. We studied the kinases’ participation in protein–protein interaction networks, and the potential deleterious effects, if inhibited, were supported with the literature. In the case of Leishmania GSK3, an inhibitor of its human counterpart, prioritized by our method, was validated in vitro to test its anti-Leishmania activity and indirectly infer the presence of the enzyme in the parasite. The analysis contributes to improving the knowledge about the presence of similar signalling pathways in Leishmania, as well as the discovery of compounds acting against any of these kinases as potential molecular targets in the parasite.

scholarly journals Yeast-based automated high-throughput screens to identify anti-parasitic lead compounds

Open Biology ◽  
2013 ◽  
Vol 3 (2) ◽  
pp. 120158 ◽  
Author(s):  
Elizabeth Bilsland ◽  
Andrew Sparkes ◽  
Kevin Williams ◽  
Harry J. Moss ◽  
Michaela de Clare ◽  
...  
Keyword(s):  
Side Effects ◽  
Trypanosoma Brucei ◽  
Fluorescent Proteins ◽  
Screening Method ◽  
Lead Compounds ◽  
Dihydrofolate Reductases ◽  
Parasitic Activity ◽  
High Throughput Screens ◽  
Specific Inhibitors

We have developed a robust, fully automated anti-parasitic drug-screening method that selects compounds specifically targeting parasite enzymes and not their host counterparts, thus allowing the early elimination of compounds with potential side effects. Our yeast system permits multiple parasite targets to be assayed in parallel owing to the strains’ expression of different fluorescent proteins. A strain expressing the human target is included in the multiplexed screen to exclude compounds that do not discriminate between host and parasite enzymes. This form of assay has the advantages of using known targets and not requiring the in vitro culture of parasites. We performed automated screens for inhibitors of parasite dihydrofolate reductases, N -myristoyltransferases and phosphoglycerate kinases, finding specific inhibitors of parasite targets. We found that our ‘hits’ have significant structural similarities to compounds with in vitro anti-parasitic activity, validating our screens and suggesting targets for hits identified in parasite-based assays. Finally, we demonstrate a 60 per cent success rate for our hit compounds in killing or severely inhibiting the growth of Trypanosoma brucei , the causative agent of African sleeping sickness.

scholarly journals Formulation and in-vitro preliminary screening of polyphyto antilithiatic combination

2013 ◽  
Vol 2 (3) ◽  
pp. 214-217
Author(s):  
Abhishek Bharadwaj ◽  
Kumud Upadhayaya ◽  
N.V. Satheesh Madhav
Keyword(s):  
Preliminary Screening

IN VITRO SELECTION OF DISEASE RESISTANCE IN FRUIT TREES

1995 ◽  
pp. 19-26 ◽  
Author(s):  
F. Hammerschlag ◽  
D. Ritchie ◽  
D. Werner ◽  
G. Hashmil ◽  
L. Krusberg ◽  
...  
Keyword(s):  
Disease Resistance ◽  
In Vitro Selection ◽  
Fruit Trees ◽  
Selection Of

scholarly journals In Vitro Evaluation of Five Antimicrobial Peptides against the Plant Pathogen Erwinia amylovora

Biomolecules ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 554
Author(s):  
Rafael J. Mendes ◽  
Laura Regalado ◽  
João P. Luz ◽  
Natália Tassi ◽  
Cátia Teixeira ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Antimicrobial Peptides ◽  
Erwinia Amylovora ◽  
Growth Curves ◽  
Fruit Trees ◽  
Membrane Permeabilization ◽  
Control Measures ◽  
Minimal Bactericidal Concentration ◽  
Pome Fruit

Fire blight is a major pome fruit trees disease that is caused by the quarantine phytopathogenic Erwinia amylovora, leading to major losses, namely, in pear and apple productions. Nevertheless, no effective sustainable control treatments and measures have yet been disclosed. In that regard, antimicrobial peptides (AMPs) have been proposed as an alternative biomolecule against pathogens but some of those AMPs have yet to be tested against E. amylovora. In this study, the potential of five AMPs (RW-BP100, CA-M, 3.1, D4E1, and Dhvar-5) together with BP100, were assessed to control E. amylovora. Antibiograms, minimal inhibitory, and bactericidal concentrations (minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC), growth and IC50 were determined and membrane permeabilization capacity was evaluated by flow cytometry analysis and colony-forming units (CFUs) plate counting. For the tested AMPs, the higher inhibitory and bactericidal capacity was observed for RW-BP100 and CA-M (5 and 5–8 µM, respectively for both MIC and MBC), whilst for IC50 RW-BP100 presented higher efficiency (2.8 to 3.5 µM). Growth curves for the first concentrations bellow MIC showed that these AMPs delayed E. amylovora growth. Flow cytometry disclosed faster membrane permeabilization for CA-M. These results highlight the potential of RW-BP100 and CA-M AMPs as sustainable control measures against E. amylovora.

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