Microplate Alamar Blue Assay (MABA) and Low Oxygen Recovery Assay (LORA) for Mycobacterium tuberculosis

2015 ◽  
pp. 281-292 ◽  
Author(s):  
Sanghyun Cho ◽  
Hyung Sup Lee ◽  
Scott Franzblau
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Alamar Blue Assay ◽  
Alamar Blue ◽  
Low Oxygen ◽  
Recovery Assay

scholarly journals Rapid, Low-Technology MIC Determination with Clinical Mycobacterium tuberculosis Isolates by Using the Microplate Alamar Blue Assay

1998 ◽  
Vol 36 (2) ◽  
pp. 362-366 ◽  
Author(s):  
Scott G. Franzblau ◽  
Richard S. Witzig ◽  
James C. McLaughlin ◽  
Patricia Torres ◽  
Guillermo Madico ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Low Cost ◽  
Multidrug Resistant ◽  
Appropriate Technology ◽  
Alamar Blue Assay ◽  
Alamar Blue ◽  
Solid Media ◽  
Resistant Strains ◽  
Stable Reagent ◽  
Initial Results

A colorimetric, microplate-based Alamar Blue assay (MABA) method was used to determine the MICs of isoniazid (INH), rifampin, streptomycin (SM), and ethambutol (EMB) for 34 PeruvianMycobacterium tuberculosis isolates (including both pansensitive and multidrug-resistant strains) and the H37Rv strain by using bacterial suspensions prepared directly from solid media. Results for all isolates were available within 8 days. Discordant results were observed on initial tests for 3 of 16 INH-susceptible isolates, 5 of 31 EMB-susceptible isolates, and 2 of 4 SM-resistant isolates (by the BACTEC 460 system). The overall agreements between the MICs obtained by MABA and the results obtained with the BACTEC 460 system were 87.9% for initial results and 93.6% after retesting 12 of 17 samples with discrepant results. Interpretation of MABA endpoints improved with technical experience. The MABA is a simple, rapid, low-cost, appropriate technology which does not require expensive instrumentation and which makes use of a nontoxic, temperature-stable reagent.

scholarly journals Villarinol, a new Alkenoyloxyalkenol Derivative from the Endemic Philippine Rubiaceae species Villaria odorata

2012 ◽  
Vol 7 (6) ◽  
pp. 1934578X1200700 ◽  
Author(s):  
Allan Patrick G. Macabeo ◽  
Jalil A. Avila ◽  
Grecebio Jonathan D. Alejandro ◽  
Scott G. Franzblau ◽  
Simeon F. Kouam ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Alamar Blue Assay ◽  
Alamar Blue

Villarinol (1), a new alkenoyloxy alkenol metabolite, has been isolated from the dichloromethane extract of Villaria odorata, an endemic Rubiaceae Philippine plant, along with the known compounds stigmasterol and 4-hydroxybenzaldehyde. The structure of 1 was elucidated on the basis of spectroscopic and spectrometric studies. The extracts of V. odorata exhibited moderate inhibition against Mycobacterium tuberculosis H37Rv, based on the colorimetric microplate alamar blue assay.

scholarly journals Antituberculosis Activity of Brotowali (Tinospora crispa) Extract and Fractions against Mycobacterium tuberculosis using Microplate Alamar Blue Assay Method

2017 ◽  
Vol 22 (2) ◽  
pp. 124
Author(s):  
Retno Wahyuningrum ◽  
Ritmaleni Ritmaleni ◽  
Tatang Irianti ◽  
Subagus Wahyuono ◽  
Takushi Kaneko ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Pathogenic Bacteria ◽  
Ethanolic Extract ◽  
Multidrug Resistant ◽  
Alamar Blue Assay ◽  
Assay Method ◽  
Drug Candidate ◽  
Antituberculosis Activity ◽  
Alamar Blue ◽  
Resistant Tuberculosis

Tuberculosis (TB), in which caused by pathogenic bacteria, Mycobacterium tuberculosis, has become the major causes of death among all of infectious diseases. The increasing incidence of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) has created a need to discover a new antituberculosis drug candidate. The aim of this study was to screen extract and fractions of Tinospora crispa for activity against Mycobacterium tuberculosis H37Rv using the Microplate Alamar Blue Assay (MABA) method. T. crispa extract was prepared by maceration in ethanol (96%) and antituberculosis activity was carried out using MABA method. The result of this study showed that ethanolic extract of T. crispa exhibit antituberculosis activity with minimum inhibition concentration of 12.5 mg/ml.

scholarly journals Evaluation of Accuracy of Microplate Alamar Blue Assay and Proportion Method for Prompt Detection of Mycobacterium tuberculosis and Clinical Isolates of Multidrug-resistant M. tuberculosis

2021 ◽  
Vol 14 (3) ◽  
Author(s):  
Alireza Jafari ◽  
Raj Goswami ◽  
Hesamaddin Shirzad Aski ◽  
Nasser Behnampour ◽  
Masoomeh Taziki ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Positive Predictive Value ◽  
Negative Predictive Value ◽  
Predictive Value ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Alamar Blue Assay ◽  
Reference Laboratory ◽  
Alamar Blue ◽  
Resistant Tuberculosis

Background: Tuberculosis is appraised to cause the deaths of more than a billion people in the last decades. Objectives: The current study compares the performance of microplate Alamar blue assay for clinical isolates of Mycobacterium tuberculosis and multidrug-resistant tuberculosis. Microplate Alamar blue assay was performed in a central tuberculosis laboratory at Golestan University of Medical Sciences in Gorgan, Iran. Methods: In the first step, the microplate Alamar blue assay was used for the detection of 78 clinical isolates in the Golestan Regional Tuberculosis Reference Laboratory, and the results were compared with those of the proportion assay. In the second step, the microplate Alamar blue assay and the proportion assay were used for the drug susceptibility of 35 isolates. Results: In the microplate Alamar blue assay, the sensitivity was 100 (90.97 - 100), with a specificity of 74.36 (57.87 - 86.96), positive predictive value of 79.59 (65.66 - 89.76), and negative predictive value of 100 (88.06 - 100). For the microplate Alamar blue assay with rifampin, the sensitivity was 100 (89.11 - 100), specificity was 100 (29.24 - 100), positive predictive value was 100 (89.11 - 100), and negative predictive value was 100 (29.24 - 100). For the microplate Alamar blue assay with isoniazid, the sensitivity was 84.38 (67.21 - 94.72), specificity was 66.67 (9.43 - 99.16), positive predictive value was 96.43 (81.65 - 99.91), and negative predictive value was 28.57 (3.67 - 70.96). Conclusions: We found high accuracy between the microplate Alamar blue assay with rifampin and the proportion assay. The rapid and low-cost microplate Alamar blue assay is an inexpensive and appropriate assay for the detection of rifampin-resistant tuberculosis in low-income countries.

scholarly journals Low-Oxygen-Recovery Assay for High-Throughput Screening of Compounds against Nonreplicating Mycobacterium tuberculosis

2007 ◽  
Vol 51 (4) ◽  
pp. 1380-1385 ◽  
Author(s):  
Sang Hyun Cho ◽  
Saradee Warit ◽  
Baojie Wan ◽  
Chang Hwa Hwang ◽  
Guido F. Pauli ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
High Throughput ◽  
High Throughput Screening ◽  
Bacterial Infections ◽  
Antimicrobial Agents ◽  
Physiological State ◽  
Recovery Phase ◽  
Low Oxygen ◽  
Anaerobic Conditions ◽  
Recovery Assay

ABSTRACT Screening for new antimicrobial agents is routinely conducted only against actively replicating bacteria. However, it is now widely accepted that a physiological state of nonreplicating persistence (NRP) is responsible for antimicrobial tolerance in many bacterial infections. In tuberculosis, the key to shortening the 6-month regimen lies in targeting this NRP subpopulation. Therefore, a high-throughput, luminescence-based low-oxygen-recovery assay (LORA) was developed to screen antimicrobial agents against NRP Mycobacterium tuberculosis. M. tuberculosis H37Rv containing a plasmid with an acetamidase promoter driving a bacterial luciferase gene was adapted to low oxygen conditions by extended culture in a fermentor with a 0.5 headspace ratio. The MICs of 31 established antimicrobial agents were determined in microplate cultures maintained under anaerobic conditions for 10 days and, for comparative purposes, under aerobic conditions for 7 days. Cultures exposed to drugs under anaerobic conditions followed by 28 h of “recovery” under ambient oxygen produced a luminescent signal that was, for most compounds, proportional to the number of CFU determined prior to the recovery phase. No agents targeting the cell wall were active against NRP M. tuberculosis, whereas drugs hitting other cellular targets had a range of activities. The calculated Z′ factor was in the range of 0.58 to 0.84, indicating the suitability of the use of LORA for high-throughput assays. This LORA is sufficiently robust for use for primary high-throughput screening of compounds against NRP M. tuberculosis.

scholarly journals Microplate alamar blue assay versus BACTEC 460 system for high-throughput screening of compounds against Mycobacterium tuberculosis and Mycobacterium avium.

1997 ◽  
Vol 41 (5) ◽  
pp. 1004-1009 ◽  
Author(s):  
L Collins ◽  
S G Franzblau
Keyword(s):  
Mycobacterium Tuberculosis ◽  
High Throughput ◽  
Mycobacterium Avium ◽  
High Throughput Screening ◽  
Antimicrobial Agents ◽  
Alamar Blue Assay ◽  
Alamar Blue ◽  
Mycobacterium Tuberculosis H37rv ◽  
Large Numbers ◽  
Highly Correlated

In response to the need for rapid, inexpensive, high-throughput assays for antimycobacterial drug screening, a microplate-based assay which uses Alamar blue reagent for determination of growth was evaluated. MICs of 30 antimicrobial agents against Mycobacterium tuberculosis H37Rv, M. tuberculosis H37Ra, and Mycobacterium avium were determined in the microplate Alamar blue assay (MABA) with both visual and fluorometric readings and compared to MICs determined in the BACTEC 460 system. For all three mycobacterial strains, there was < or = 1 dilution difference between MABA and BACTEC median MICs in four replicate experiments for 25 to 27 of the 30 antimicrobics. Significant differences between MABA and BACTEC MICs were observed with 0, 2, and 5 of 30 antimicrobial agents against H37Rv, H37Ra, and M. avium, respectively. Overall, MICs determined either visually or fluorometrically in MABA were highly correlated with those determined in the BACTEC 460 system, and visual MABA and fluorometric MABA MICs were highly correlated. MICs of rifampin, rifabutin, minocycline, and clarithromycin were consistently lower for H37Ra compared to H37Rv in all assays but were similar for most other drugs. M. tuberculosis H37Ra may be a suitable surrogate for the more virulent H37Rv strain in primary screening of compounds for antituberculosis activity. MABA is sensitive, rapid, inexpensive, and nonradiometric and offers the potential for screening, with or without analytical instrumentation, large numbers of antimicrobial compounds against slow-growing mycobacteria.

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