scholarly journals Baseline Sensitivity of Fusarium virguliforme to Fluopyram Fungicide

Plant Disease ◽  
2017 ◽  
Vol 101 (4) ◽  
pp. 576-582 ◽  
Author(s):  
Jie Wang ◽  
Carl A. Bradley ◽  
Olivia Stenzel ◽  
Dianne K. Pedersen ◽  
Ursula Reuter-Carlson ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mycelial Growth ◽  
Soybean Seed ◽  
Inhibition Assay ◽  
Baseline Sensitivity ◽  
Growth Inhibition Assay ◽  
Fusarium Virguliforme ◽  
Fungicide Sensitivity ◽  
Conidia Germination

Fluopyram, a succinate dehydrogenase inhibitor (SDHI) fungicide, was recently registered for use as a soybean seed treatment for management of sudden death syndrome (SDS) caused by Fusarium virguliforme. Although registered and now used commercially, in vitro baseline fungicide sensitivity of F. virguliforme to fluopyram has not yet been established. In this study, the baseline sensitivity of F. virguliforme to fluopyram was determined using in vitro growth of mycelium and germination of conidia assays with two collections of F. virguliforme isolates. A total of 130 and 75 F. virguliforme isolates were tested using the mycelial growth and conidia germination assays, respectively, including a core set of isolates that were tested with both assays. In the mycelial growth inhibition assay, 113 out of 130 isolates (86.9%) were inhibited 50% by effective concentrations (EC50) less than 5 µg/ml with a mean EC50 of 3.35 µg/ml. For the conidia germination assay, 73 out of 75 isolates (97%) were determined to have an estimated EC50 of less than 5 µg/ml with a mean EC50 value of 2.28 µg/ml. In a subset of 20 common isolates that were phenotyped with both assays, conidia germination of F. virguliforme was determined to be more sensitive to fluopyram (mean EC50 = 2.28 µg/ml) than mycelial growth (mean EC50 = 3.35 µg/ml). Hormetic effects were observed in the mycelial growth inhibition assay as 22% of the isolates demonstrated more growth on medium amended with the lowest fluopyram concentration (1 µg/ml), as compared with the nonfluopyram amended control. It was not possible to determine EC50 values for nine out of 185 isolates (4.8%), as those isolates were not inhibited by 50% even at the highest fluopyram concentrations of 100 µg/ml for mycelial growth and 20 µg/ml for conidia germination inhibition assays. On the whole, the F. virguliforme population appears to be sensitive to fluopyram, and this study enables future monitoring of fungicide sensitivity.

scholarly journals Sensitivity distribution of Venturia inaequalis to fenarimol in Québec apple orchards

2005 ◽  
Vol 75 (1) ◽  
pp. 35-43 ◽  
Author(s):  
O. Carisse ◽  
J.R. Pelletier
Keyword(s):  
Growth Inhibition ◽  
Frequency Distribution ◽  
Radial Growth ◽  
Venturia Inaequalis ◽  
Apple Scab ◽  
Inhibition Assay ◽  
Baseline Sensitivity ◽  
Growth Inhibition Assay ◽  
Sensitivity Distribution ◽  
Ergosterol Synthesis

This study was initiated to quantify the baseline sensitivity of apple scab (Venturia inaequalis) to fenarimol, an ergosterol synthesis-inhibiting fungicide. In 1988, 576 monoconidial isolates of Venturia inaequalis were collected from 26 commercial orchards throughout Quebec. Sensitivity to fenarimol was assessed by radial growth inhibition assay. The ED50 values for the 26 orchards ranged from 0.024 to 5.212 (μ g mL-1 with a mean ED50 of 0.156 μg ml-1. Reduced sensitivity, expressed as ED50, was found in three orchards for an overall frequency of 4.51% of isolates. Sensitive isolates had a mean ED50 of 0.079 μg ml-1, whereas isolates with reduced sensitivity had a mean ED50 of 1.714 μ g mL-1, yielding a resistance factor of about 22. Four populations were identified based on the frequency distribution of ED50 values.

scholarly journals Development of a Murine Mycobacterial Growth Inhibition Assay for Evaluating Vaccines against Mycobacterium tuberculosis

2009 ◽  
Vol 16 (7) ◽  
pp. 1025-1032 ◽  
Author(s):  
Marcela Parra ◽  
Amy L. Yang ◽  
JaeHyun Lim ◽  
Kristopher Kolibab ◽  
Steven Derrick ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Immune Responses ◽  
Growth Inhibition ◽  
In Vitro Assays ◽  
Functional Assay ◽  
Inhibition Assay ◽  
Growth Inhibition Assay ◽  
Vitro Assay ◽  
Mycobacterial Growth

ABSTRACT The development and characterization of new tuberculosis (TB) vaccines has been impeded by the lack of reproducible and reliable in vitro assays for measuring vaccine activity. In this study, we developed a murine in vitro mycobacterial growth inhibition assay for evaluating TB vaccines that directly assesses the capacity of immune splenocytes to control the growth of Mycobacterium tuberculosis within infected macrophages. Using this in vitro assay, protective immune responses induced by immunization with five different types of TB vaccine preparations (Mycobacterium bovis BCG, an attenuated M. tuberculosis mutant strain, a DNA vaccine, a modified vaccinia virus strain Ankara [MVA] construct expressing four TB antigens, and a TB fusion protein formulated in adjuvant) can be detected. Importantly, the levels of vaccine-induced mycobacterial growth-inhibitory responses seen in vitro after 1 week of coculture correlated with the protective immune responses detected in vivo at 28 days postchallenge in a mouse model of pulmonary tuberculosis. In addition, similar patterns of cytokine expression were evoked at day 7 of the in vitro culture by immune splenocytes taken from animals immunized with the different TB vaccines. Among the consistently upregulated cytokines detected in the immune cocultures are gamma interferon, growth differentiation factor 15, interleukin-21 (IL-21), IL-27, and tumor necrosis factor alpha. Overall, we have developed an in vitro functional assay that may be useful for screening and comparing new TB vaccine preparations, investigating vaccine-induced protective mechanisms, and assessing manufacturing issues, including product potency and stability.

scholarly journals The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells

F1000Research ◽  
2021 ◽  
Vol 10 ◽  
pp. 257
Author(s):  
Rachel Tanner ◽  
Emily Hoogkamer ◽  
Julia Bitencourt ◽  
Andrew White ◽  
Charelle Boot ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Protective Immunity ◽  
Vaccine Development ◽  
Autologous Serum ◽  
Inhibition Assay ◽  
Growth Inhibition Assay ◽  
Vaccine Candidates ◽  
Mycobacterial Growth

The only currently available approach to early efficacy testing of tuberculosis (TB) vaccine candidates is in vivo preclinical challenge models. These typically include mice, guinea pigs and non-human primates (NHPs), which must be exposed to virulent M.tb in a ‘challenge’ experiment following vaccination in order to evaluate protective efficacy. This procedure results in disease development and is classified as ‘Moderate’ in severity under EU legislation and UK ASPA licensure. Furthermore, experiments are relatively long and animals must be maintained in high containment level facilities, making them relatively costly. We describe an in vitro protocol for the direct mycobacterial growth inhibition assay (MGIA) for use in the macaque model of TB vaccine development with the aim of overcoming some of these limitations. Importantly, using an in vitro assay in place of in vivo M.tb challenge represents a significant refinement to the existing procedure for early vaccine efficacy testing. Peripheral blood mononuclear cell and autologous serum samples collected from vaccinated and unvaccinated control animals are co-cultured with mycobacteria in a 48-well plate format for 96 hours. Adherent monocytes are then lysed to release intracellular mycobacteria which is quantified using the BACTEC MGIT system and colony-forming units determined relative to an inoculum control and stock standard curve. We discuss related optimisation and characterisation experiments, and review evidence that the direct NHP MGIA provides a biologically relevant model of vaccine-induced protection. The potential end-users of the NHP MGIA are academic and industry organisations that conduct the assessment of TB vaccine candidates and associated protective immunity using the NHP model. This approach aims to provide a method for high-throughput down-selection of vaccine candidates going forward to in vivo efficacy testing, thus expediting the development of a more efficacious TB vaccine and offering potential refinement and reduction to the use of NHPs for this purpose.

A practical in vitro growth inhibition assay for the evaluation of TB vaccines

Vaccine ◽  
2009 ◽  
Vol 28 (2) ◽  
pp. 317-322 ◽  
Author(s):  
Kristopher Kolibab ◽  
Marcela Parra ◽  
Amy L. Yang ◽  
Liyanage P. Perera ◽  
Steven C. Derrick ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
In Vitro Growth ◽  
Inhibition Assay ◽  
Growth Inhibition Assay

scholarly journals Inhibition of Mycobacterial GrowthIn Vitrofollowing Primary but Not Secondary Vaccination with Mycobacterium bovis BCG

2013 ◽  
Vol 20 (11) ◽  
pp. 1683-1689 ◽  
Author(s):  
Helen A. Fletcher ◽  
Rachel Tanner ◽  
Robert S. Wallis ◽  
Joel Meyer ◽  
Zita-Rose Manjaly ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mycobacterium Bovis ◽  
Primary Vaccination ◽  
Inhibition Assay ◽  
Peripheral Blood Mononuclear ◽  
Growth Inhibition Assay ◽  
Ifn Γ ◽  
Mycobacterial Growth

ABSTRACTDespite the widespread use of theMycobacterium bovisBCG vaccine, there are more than 9 million new cases of tuberculosis (TB) every year, and there is an urgent need for better TB vaccines. TB vaccine candidates are selected for evaluation based in part on the detection of an antigen-specific gamma interferon (IFN-γ) response. The measurement of mycobacterial growth in blood specimens obtained from subjects immunized with investigational TB vaccines may be a betterin vitrocorrelate ofin vivovaccine efficacy. We performed a clinical study with 30 United Kingdom adults who were followed for 6 months to evaluate the abilities of both a whole-blood- and a novel peripheral blood mononuclear cell (PBMC)-based mycobacterial growth inhibition assay to measure a response to primary vaccination and revaccination with BCG. Using cryopreserved PBMCs, we observed a significant improvement in mycobacterial growth inhibition following primary vaccination but no improvement in growth inhibition following revaccination with BCG (P< 0.05). Mycobacterial growth inhibition following primary BCG vaccination was not correlated with purified protein derivative (PPD) antigen-specific IFN-γ enzyme-linked immunospot (ELISPOT) responses. We demonstrate that a mycobacterial growth inhibition assay can detect improved capacity to control growth following primary immunization, but not revaccination, with BCG. This is the first study to demonstrate that anin vitrogrowth inhibition assay can identify a difference in vaccine responses by comparing both primary and secondary BCG vaccinations, suggesting thatin vitrogrowth inhibition assays may serve as better surrogates of clinical efficacy than the assays currently used for the assessment of candidate TB vaccines.

scholarly journals The in vitro direct mycobacterial growth inhibition assay (MGIA) for the early evaluation of TB vaccine candidates and assessment of protective immunity: a protocol for non-human primate cells

F1000Research ◽  
2021 ◽  
Vol 10 ◽  
pp. 257
Author(s):  
Rachel Tanner ◽  
Emily Hoogkamer ◽  
Julia Bitencourt ◽  
Andrew White ◽  
Charelle Boot ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Protective Immunity ◽  
Vaccine Development ◽  
Autologous Serum ◽  
Inhibition Assay ◽  
Growth Inhibition Assay ◽  
Vaccine Candidates ◽  
Mycobacterial Growth

The only currently available approach to early efficacy testing of tuberculosis (TB) vaccine candidates is in vivo preclinical challenge models. These typically include mice, guinea pigs and non-human primates (NHPs), which must be exposed to virulent M.tb in a ‘challenge’ experiment following vaccination in order to evaluate protective efficacy. This procedure results in disease development and is classified as ‘Moderate’ in severity under EU legislation and UK ASPA licensure. Furthermore, experiments are relatively long and animals must be maintained in high containment level facilities, making them relatively costly. We describe an in vitro protocol for the direct mycobacterial growth inhibition assay (MGIA) for use in the macaque model of TB vaccine development with the aim of overcoming some of these limitations. Importantly, using an in vitro assay in place of in vivo M.tb challenge represents a significant refinement to the existing procedure for early vaccine efficacy testing. Peripheral blood mononuclear cell and autologous serum samples collected from vaccinated and unvaccinated control animals are co-cultured with mycobacteria in a 48-well plate format for 96 hours. Adherent monocytes are then lysed to release intracellular mycobacteria which is quantified using the BACTEC MGIT system and colony-forming units determined relative to an inoculum control and stock standard curve. We discuss related optimisation and characterisation experiments, and review evidence that the direct NHP MGIA provides a biologically relevant model of vaccine-induced protection. The potential end-users of the NHP MGIA are academic and industry organisations that conduct the assessment of TB vaccine candidates and associated protective immunity using the NHP model. This approach aims to provide a method for high-throughput down-selection of vaccine candidates going forward to in vivo efficacy testing, thus expediting the development of a more efficacious TB vaccine and offering potential refinement and reduction to the use of NHPs for this purpose.

Management of collar rot disease in chickpea by Trichoderma species

2020 ◽  
Vol 7 (03) ◽  
Author(s):  
PREM PANDEY ◽  
G. C. SAGAR ◽  
SUNDARMAN SHRESTHA2 ◽  
HIRAKAJI MANANDHAR ◽  
RITESH K. YADAV ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mycelial Growth ◽  
Sclerotium Rolfsii ◽  
Maximum Growth ◽  
Pot Experiment ◽  
Trichoderma Spp ◽  
Trichoderma Species ◽  
Collar Rot ◽  
Rot Disease

Nine isolates of Trichoderma spp. were isolated from different agro- ecological regions of Nepal viz; Jumla, Palpa, Chitwan, Tarahara, Banke, Illam and Salyan and screened against Sclerotium rolfsii Sacc. Adreded soil borne phytopathogen causing collar rot of chickpea in chickpea; In-vitro efficacy of nine fungal antagonist (Trichoderma spp.) against Sclerotium rolfsii were screened. Pot experiment was done to find out the effective management of S. rolfsi through Tricoderma using different methods i.e. Seed treatment, soil drenching and soil application. All the tested isolates of Trichoderma spp. were found effective on mycelial growth inhibition and sclerotial parasitization of S. rolfsii. Trichoderma isolated from Palpa district showed maximum growth inhibition (%) of pathogen periodically after 48(93.78%), 72(96.00%), 96(97.96%) and 120(100.00%) hours of inoculation. Parasitized sclerotium showed minimum sclerotial germination on agar plates. Moreover, Trichoderma species isolated from Palpa districts showed second best percent mycelial growth inhibition periodically at 72(25.00%), 120(29.16%), 168(29.16%) and 216(29.16%).In pot experiment at 40 days after sowing, Seedling height was maximum in soil drenching with 30g per 100ml of water (22.27cm) and Mortality percentage of seedlings was least or highest disease control was observed in seed treated with 109cfu/ml (0.000%).

scholarly journals A non-human primate in vitro functional assay for the early evaluation of TB vaccine candidates

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Rachel Tanner ◽  
Andrew D. White ◽  
Charelle Boot ◽  
Claudia C. Sombroek ◽  
Matthew K. O’Shea ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mononuclear Cells ◽  
Functional Assay ◽  
Intermediate Precision ◽  
Peripheral Blood Mononuclear ◽  
Growth Inhibition Assay ◽  
Early Evaluation ◽  
Human Primate

AbstractWe present a non-human primate mycobacterial growth inhibition assay (MGIA) using in vitro blood or cell co-culture with the aim of refining and expediting early tuberculosis vaccine testing. We have taken steps to optimise the assay using cryopreserved peripheral blood mononuclear cells, transfer it to end-user institutes, and assess technical and biological validity. Increasing cell concentration or mycobacterial input and co-culturing in static 48-well plates compared with rotating tubes improved intra-assay repeatability and sensitivity. Standardisation and harmonisation efforts resulted in high consistency agreements, with repeatability and intermediate precision <10% coefficient of variation (CV) and inter-site reproducibility <20% CV; although some systematic differences were observed. As proof-of-concept, we demonstrated ability to detect a BCG vaccine-induced improvement in growth inhibition in macaque samples, and a correlation between MGIA outcome and measures of protection from in vivo disease development following challenge with either intradermal BCG or aerosol/endobronchial Mycobacterium tuberculosis (M.tb) at a group and individual animal level.

scholarly journals Fungicide Sensitivity of Pythium spp. Associated with Cavity Spot of Carrot in California and Michigan

Plant Disease ◽  
2012 ◽  
Vol 96 (3) ◽  
pp. 384-388 ◽  
Author(s):  
Xiao Hong Lu ◽  
R. Michael Davis ◽  
S. Livingston ◽  
J. Nunez ◽  
Jianjun J. Hao
Keyword(s):  
United States ◽  
Growth Inhibition ◽  
Active Ingredient ◽  
Mycelial Growth ◽  
The United States ◽  
Fungicide Sensitivity ◽  
First Report ◽  
Cavity Spot ◽  
Pythium Violae

The identity of 172 isolates of Pythium spp. from cavity spot lesions on carrot produced in California and Michigan was determined, and their sensitivity to three fungicides was examined. Pythium violae accounted for 85% of California isolates, with P. irregulare, P. dissotocum (the first report as a carrot pathogen in the United States), P. ultimum, and P. sulcatum making the balance. P. sulcatum, P. sylvaticum, and P. intermedium were the most commonly recovered (85%) species in Michigan; others from Michigan included P. intermedium, P. irregulare, and an unclassified strain, M2-05. On fungicide-amended media, 93% of isolates were sensitive to mefenoxam (inhibition of mycelial growth was >60% at 10 μg active ingredient [a.i.]/ml); however, two of five isolates of P. irregulare from California were highly resistant (≤60% inhibition at 100 μg a.i./ml); about half of the isolates of P. intermedium and P. sylvaticum and a single isolate of P. violae were highly or intermediately resistant to mefenoxam (>60% inhibition at 100 μg a.i./ml, or ≤60% inhibition at 10 μg a.i./ml). P. dissotocum, P. irregulare, P. sulcatum, M2-05, and three of seven isolates of P. intermedium were insensitive to fluopicolide (effective concentrations for 50% growth inhibition [EC50] were >50 μg a.i./ml), while P. sylvaticum, P. ultimum, P. violae, and some isolates in P. intermedium were sensitive (EC50 < 1 μg a.i./ml). All isolates were sensitive to zoxamide (EC50 < 1 μg a.i./ml). Sensitivity baselines of P. violae to zoxamide and fluopicolide were established.

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