Academic literature on the topic 'Agar diffusion method'

The microbial factor is essential in the etiology of mastitis. In this regard, bacteriological diagnostics is one of the decisive moments in the recognition and differentiation of pathological conditions of the mammary gland. However, it takes a lot of time to make a diagnosis using general methods, special culture media and reagents.

Antibacterial activity testing can be done using the agar diffusion method, including agar well difussion and disk diffusion agar methods. This study aims to compare two antibacterial testing methods to analyze the anti-bacterial activity of a yogurt starter against the bacteria Eschericia coli and Staphilococcus aureus. The study was conducted experimentally with 5 concentrations of yogurt starter, namely 2%, 4%, 6%, 8%, and 10%. Testing antibacterial activity using two methods ,disk diffusion agar and well difussion agar methods. The research showed that agar well diffusion method obtained antibacterial activity greater than the disk diffusion method for E. coli and S. aureus.

Gramicidin, an antimicrobial peptide active against Gram positive bacteria, is commonly used in pharmaceutical preparations for topical use. Considering that only the turbidimetric method has been described in the literature, the present study sought to develop and validate an agar diffusion method for the dosage of gramicidin. The method was developed and validated using the Kocuria rhizophila ATCC 9341 as a test microorganism. Two designs were used: a 3x3 parallel-line model, and a 5x1 standard curve. The validation demonstrated that the method follows the linear model (r²= 0.994), presenting a significant regression between the zone diameter of growth inhibition and the logarithm of the concentration within the range of 5 to 25.3 µg/mL. The results obtained for both designs were precise, having a relative standard deviation (R.S.D.) for intra-day precision of 0.81 for the 3x3 assay and 1.90 for the 5x1 assay. For the inter-day precision, the R.S.D. was 1.35 for the 3x3 and 2.64 for the 5x1. The accuracy was verified and results confirmed to be accurate, having a tolerance interval of 95%, which lay within permitted limits and appropriate trueness. In addition, the method was considered selective, with limit of detection and upper and lower limits of quantification of 2.00, 5.00 and 25.3 µg/mL, respectively. No difference in precision between the designs used in the agar diffusion method was evident (p>0.05). The method proved to be appropriate for the microbiological dosage of the raw material gramicidin.

The improved agar diffusion method for determination of residual antimicrobial agents was investigated, and the sensitivities of various combinations of test organisms and assay media were determined using 7 organisms, 5 media, and 31 antimicrobial agents. Bacillus stearothermophilus and synthetic assay medium (SAM) showed the greatest sensitivity for screening penicillins (penicillin G and ampicillin). The combination of Bacillus subtilis and minimum medium (MM) was the most sensitive for tetracyclines (oxytetracycline and chlortetracycline), B. stearothermophilus and SAM or Micrococcus luteus and Mueller-Hinton agar (MHA) for detecting tylosin and erythromycin, B. subtilis and MHA for aminoglycosides (streptomycin, kanamycin, gentamicin, and dihydrostreptomycin), B. stearothermophilus and SAM for polyethers (salinomycin and lasalocid), and B. subtilis and MM or Clostridium perfringens and GAM for polypeptides (thiopeptin, enramycin, virginiamycin, and bacitracin). However, gram-negative bacterium Escherichia coli ATCC 27166 and MM were better for screening for colistin and polymixin-B. For detecting the synthetic drugs tested, the best combination was B. subtilis and MM for sulfonamides, E. coli 27166 and MM for quinolones (oxolinic acid and nalidixic acid), B. subtilis and MM for furans (furazolidone), and the bioluminescent bacterium Photobacterium phosphoreum and luminescence assay medium for chloramphenicol and oxolinic acid. The results showed that the use of four assay plates, B. stearothermophilus and SAM, B. subtilis and MM, M. luteus and MHA, and E. coli 27166 and MM, was superior to the currently available techniques for screening for residual antimicrobial agents in edible animal tissues.

Elizabethkingia anophelis has recently emerged as a cause of life-threatening infections. This study compared the results of antimicrobial susceptibility testing (AST) conducted for E. anophelis through different methods. E. anophelis isolates collected between January 2005 and June 2019 were examined for their susceptibility to 14 antimicrobial agents by using disk diffusion, gradient diffusion (Etest; bioMérieux S.A., Marcy l’Etoile, France), and agar dilution methods. The agar dilution method was the reference assay. According to the agar dilution method, the isolates exhibited the highest susceptibility to minocycline (100%), doxycycline (97.6%), rifampin (95.2%), and levofloxacin (78.6%). A very major error rate of >1.5% was observed for nine antibiotics tested using the disk diffusion method. The overall categorical agreement rate between the disk diffusion and agar dilution methods was 74.8%, and ceftazidime, minocycline, levofloxacin, and rifampin met the minimum requirements for discrepancy and agreement rates. The Etest method tended to produce lower log2 minimum inhibitory concentrations for the antibiotics, except for trimethoprim–sulfamethoxazole and rifampin; the method resulted in very major errors for nine antibiotics. The overall essential and categorical agreement rates between the Etest and agar dilution methods were 67.3% and 76.1%, respectively. The Etest method demonstrated acceptable discrepancy and agreement rates for ceftazidime, minocycline, doxycycline, levofloxacin, and rifampin. AST results obtained through the disk diffusion and Etest methods for multiple antibiotics differed significantly from those obtained using the agar dilution method. These two assays should not be a routine alternative for AST for E. anophelis.

Objectives. To develop a new method for determining total antioxidants in serum and to evaluate the total antioxidant capacity of organisms.Design and Methods. Sodium hyposulfite (Na2S2O3) and serum were used to evaluate the linearity and precision of the potassium permanganate agar method. The area of serum diffusion in samples from 30 intensive care unit (ICU) patients compared with 44 healthy subjects was determined by the potassium permanganate agar method.Results. The linearity (R2in the linear experiment of Na2S2O3was 0.994;R2in the linear experiment of serum was 0.987) and precision (coefficient of variation of area of high level serum diffusion within-run, between-run, and between-day and coefficient of variation of area of low serum diffusion within-run, between-run, and between-day were all less than 10%) were acceptable using the potassium permanganate agar method. Total antioxidants of serum between the ICU group and the healthy group were different (p=0.002, two tailed).Conclusions. Total antioxidants in serum can be determined by the potassium permanganate agar method. The total antioxidant capacity of an organism can be evaluated by the amount of total antioxidants in serum.

Background:- Methicillin resistant Staphylococcus aureus (MRSA) are responsible for hospital and community acquired infections. There are many laboratory methods for detection of MRSA. Chromogenic media have been used for the last few years for the quick detection of MRSA. Objective:- Aim of this study was to compare the performance of conventional methods and chromogenic media for the detection of MRSA in a tertiary care hospital. Material and method: - 200 consecutive isolates of S. aureus confirmed by conventional methods, collected in a tertiary care hospital were used for this study. Cefoxitin and oxacillin disc diffusion test used as conventional methods and Chromogenic media i.e. oxacillin resistant screen agar base (ORSAB) was used for detection of methicillin resistant Staphylococcus aureus. All confirmed MRSA were checked by gold standard mecA base PCR method. Result: - Out of 200 isolates of Staphylococcus aureus, 50,52 and 47 strains were MRSA by Cefoxitin disc diffusion method, oxacillin disc diffusion method and oxacillin resistant screen agar base (ORSAB) method respectively. Specificity was 100%, 98.66%, 98.66% by Cefoxitin disc diffusion, oxacillin disc diffusion and ORSAB method respectively. Conclusion: - In conclusion, cefoxitin disc diffusion was the best for the phenotypic detection of MRSA because their sensitivity and specificity were better than oxacillin and ORSAB.

Propolis exhibits antimicrobial, anti-inflammatory and other biological effects. The aim of this study was to evaluate the activity of 30 % ethanolic extract of Bulgarian propolis against 94 Helicobacter pylori strains by three methods. By the agar-well diffusion method, only 13.8 % of the strains exhibited no inhibition by 30 μl propolis extract (containing 9 mg propolis) and all isolates were inhibited to some extent by 90 μl of the extract (27 mg propolis) per well. The mean diameters of growth inhibition by 30, 60 or 90 μl propolis extract or 30 μl 96 % ethanol per well were 16.8, 19.2, 27.5 and 8.3 mm, respectively. The propolis extract was more active than the ethanol (P < 0.001). With 90 μl propolis extract per well, 69.4 % of the strains exhibited large diameters of growth inhibition (⩾20 mm) versus 26.6 % with 30 μl per well (P < 0.001). With moist propolis discs, inhibition was detected in more strains (92.1 %) than with dried discs (78.2 %, P < 0.05), with mean inhibitory diameters of 18.7 and 13.8 mm, respectively. By the agar dilution method, 100 and 300 μg propolis ml−1 inhibited the growth of 57.1 % and 76.2 %, respectively, of the 21 strains tested. In conclusion, Bulgarian propolis had a strong and dose-dependent activity against most of the H. pylori strains tested. Although the effect of propolis on H. pylori in vitro is promising, further microbiological, pharmacological and clinical trials are required.

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This study describes the therapeutic potential of extracts and standardized fractions of Arrabidaea chica leaves. A. chica is a Bignoniaceae popularly known as “crajiru”. The genus Arrabidaea occurs in Tropical America, from the Southern Mexico to the Southern Brazil. The red color of its dried leaves is attributed to two flavonoidal pigments: carajurina (main pigment) and carajurona. Chemical studies described the isolation of saponins and flavonoids from the plant leaves; purified 3- desoxyanthocyanidins were reported as anti-inflammatory. The infusion or decoction of the plant leaves is used in the folk medicine to treat anemia, inflammation and in skin wound-healing. In this work, the antimicrobial activity of the standardized extracts and fractions of A. chica cultivated at Embrapa Amazônia Ocidental, Manaus, were evaluated against fungal and bacterial microorganisms grown either from local domestic dogs and cats, or from human samples supplied by the Microorganism Collection of FIOCRUZ, in Manaus, Brazil. The plant dried leaves were extracted with increasing polarity solvents and progressively purified in preparative thin layer chromatography (TLC) and silica-gel column; the semi-purified extracts were standardized in TLC. The agar diffusion method; bioautography; minimum inhibitory concentrations tested against Staphylococcus epidermidis (CBAM 293), Staphylococcus aureus (CBAM 324), Pseudomonas aeruginosa (CBAM 232), Escherichia coli (CBAM 002), Trichophyton mentagrophytes (CFAM 1288), Microsporum canis (CFAM 1289), Malassezia pachydermatis (CFAM 1290) e Candida albicans (CFAM 1285). The standardized fractions were effective against all these microorganisms, but more intensively against Microsporum canis and Staphylococcus epidermidis. The results might favour the use of the standardized sub-fractions of A. chica as topic phytotherapic agent to treat canine external otitis. So far oleanolic and ursolic acids were identified as the main compounds in the active semi-purified fraction but other compounds of the leaves extract were not discarded. Later studies will consider the veterinarian use of the standardized extract, of the active pure entities and the convenience of the natural active antibiotic mix.
Este estudo analisou o potencial terapêutico de extratos e frações purificadas da planta amazônica Arrabidaea chica visando seu uso tópico como medicamento e eficácia comprovada em doenças cutâneas. A. chica Verl., é uma Bignoniaceae conhecida popularmente como crajiru. O gênero Arrabidaea ocorre na América tropical, do sul do México ao sul do Brasil. A cor avermelhada da folha seca e sua propriedade tintorial são devidas a dois pigmentos flavonoídicos: a carajurina, que é o pigmento principal e a carajurona. Dela foram isolados saponinas e flavonóides; As 3-desoxiantocianidinas, descritas na planta parecem possuir atividade antiinflamatória. A medicina popular utiliza o decocto ou a infusão das folhas para tratar anemia, inflamações e na cicatrização da pele. Neste trabalho, a atividade antimicrobiana de extratos e frações padronizadas da A. chica cultivada na Embrapa Amazônia Ocidental, em Manaus/AM, foi avaliada contra fungos e bactérias de amostras clínicas coletadas de animais domésticos e contra amostras humanas depositadas na coleção de Microrganimos da FIOCRUZ, Manaus/AM. Para isso, as folhas secas da planta foram extraídas com solventes de polaridade crescente, as frações foram progressivamente purificadas em cromatografia de placa ou coluna de sílica-gel, os extratos semi-purificados foram padronizados em cromatografia líquida de alta eficiência acoplada à espectrometria de massas. Os testes de difusão em ágar, bioautografia e concentração inibitória mínima foram usados para avaliar a atividade antimicrobiana das subfrações padronizadas frente aos microrganismos Staphylococcus epidermidis (CBAM 293), Staphylococcus aureus (CBAM 324), Pseudomonas aeruginosa (CBAM 232), Escherichia coli (CBAM 002), Trichophyton mentagrophytes (CFAM 1288), Microsporum canis (CFAM 1289), Malassezia pachydermatis (CFAM 1290) e Candida albicans (CFAM 1285). As frações padronizadas foram ativas contra todos esses microrganismos, com melhores resultados contra M. pachydermatis e S. epidermidis. Os resultados foram favoráveis à utilização das subfrações padronizadas na formulação de um produto fitoterápico para uso tópico em otite canina. Nas frações ativas foram identificados os ácidos oleanólico e ursólico. Estudos posteriores deverão avaliar a possibilidade de uso humano das frações purificadas ou dos compostos identificados.

Microorganisms are considered to be crucial selective factor affecting reproductive success of birds. It is hypothesized that egg-white antimicrobial proteins and incubation behavior are the most important defense mechanisms that eliminates the risk of microbial trans-shell infection. The latest studies supposed that incubation temperatures may significantly affect the antimicrobial activity of egg white proteins. The concentration of egg white antimicrobial proteins as well as incubation patterns differ among altricial and precocial species of birds. However, experimental study testing the effect of incubation temperature on the antimicrobial potential of antimicrobial egg white proteins in altricial and precocial birds is missing. In this study we tested in manipulative experiment the effect of partial and full incubation, the concentration of lysozyme and ovotransferrin and their interactions on the antimicrobial activity of egg whites of two model species - Japanese quail (Coturnix japonica) and home pigeon (Columba livia). Antimicrobial protein activity was determined by the agar well diffusion method against two Gram-positive bacteria - Bacillus subtilis and Micrococcus luteus that have been documented as pathogenic egg-white invaders of several birds. Moreover, we analyzed the effect of...

The utilization of in vitro tests with a tiered testing strategy for detection of mild ocular irritants can reduce the use of animals for testing, provide mechanistic data on toxic effects, and reduce the uncertainty associated with dose selection for clinical trials. The first section of this thesis describes how in vitro methods can be used to improve the prediction of the toxicity of chemicals and ophthalmic products. The proper utilization of in vitro methods can accurately predict toxic threshold levels and reduce animal use in product development. Sections two, three and four describe the development of new sensitive in vitro methods for predicting ocular toxicity. Maintaining the barrier function of the cornea is critical for the prevention of the penetration of infections microorganisms and irritating chemicals into the eye. Chapter 2 describes the development of a method for assessing the effects of chemicals on tight junctions using a human corneal epithelial and canine kidney epithelial cell line. In Chapter 3 a method that uses a primary organ culture for assessing single instillation and multiple instillation toxic effects is described. The ScanTox system was shown to be an ideal system to monitor the toxic effects over time as multiple readings can be taken of treated bovine lenses using the nondestructive method of assessing for the lens optical quality. Confirmations of toxic effects were made with the utilization of the viability dye alamarBlue. Chapter 4 describes the development of sensitive in vitro assays for detecting ocular toxicity by measuring the effects of chemicals on the mitochondrial integrity of bovine cornea, bovine lens epithelium and corneal epithelial cells, using fluorescent dyes. The goal of this research was to develop an in vitro test battery that can be used to accurately predict the ocular toxicity of new chemicals and ophthalmic formulations. By comparing the toxicity seen in vivo animals and humans with the toxicity response in these new in vitro methods, it was demonstrated that these in vitro methods can be utilized in a tiered testing strategy in the development of new chemicals and ophthalmic formulations.

Urinary tract infection is one of the most common bacterial infectious diseases encountered in clinical practice. The development and spread of multidrug resistant isolates are of great global health burden; among them, extended spectrum beta-lactamase (ESBL) producing Enterobacteriaceae has been a prime concern. This topic describes the resistance patterns of eighty three (83) Gram negative uropathogens to different classes of antibiotics. Bacteria isolates were obtained from patients of all age groups who sought medical attention at a secondary and tertiary hospital in Northern Ghana. Culture and isolation methods employed were the quantitative urine culture on Cysteine Lysine Electrolyte Deficient (CLED) agar and standard biochemical tests. ESBL production was detected using the CLSI recommended phenotypic confirmatory test along with routine antibiotic susceptibility test, adopting the Kirby-Bauer disk diffusion method. Out of 83 isolates, seven (7) Gram negative uropathogens were characterized and ESBLs were detected in 32 of the isolates. Escherichia coli was the pathogen with most ESBL positive strains. Generally high and multiple drug resistance were recorded in both ESBL and non-ESBL strains to the empirical drugs, however, ESBL positive strains significantly (p = 0.000) showed greater resistance. A notable finding was the appreciable resistance exhibited by ESBL strains to last line treatment drugs that include aminoglycosides and imipenem.

● To optimize antimicrobial therapy for the management of individual patient’s infection. ● For surveillance purposes, which in turn inform local/national/international clinical guidelines. ● For the management of infection control and prevention. Broadly speaking, resistance is detected by observing its phenotypic expression (activity of the candidate drug(s) against the target bacterium) or detecting the underlying genotypic determinant (resistance genes). Commonly used methods in clinical diagnostic laboratories generally fall under the ‘phenotypic’ category. These share similar traits— ease of use, reproducibility, scalability, quick turnaround of results and relative low cost of materials/reagents required. Moreover, decades of experience and fine-tuning have seen them established as methods of choice in most microbiology laboratories. Most phenotypic test methods are reliant on the use of clinical breakpoints set by national and international bodies (e.g. EUCAST and CLSI) to determine susceptibility/resistance. These guidelines are regularly subject to updates with input from leading experts and latest research findings. It is important for clinical diagnostic laboratories to adhere to best practice guidance set out by these bodies and keep up-to-date with the latest guidelines. Growth characteristics (on artificial media) of the bacterium of interest are extremely important in conventional phenotypic methods. As this presents a big obstacle for slow growers and ‘unculturable’ pathogens (e.g. Mycobacterium tuberculosis, Mycoplasma spp.) it has led to the introduction of genotypic methods of resistance detection in the clinical diagnostic laboratory. meteoric rise in the world of microbiology. Compared with conventional phenotypic methods, molecular genotypic-based tests are better suited for automation and reduce dependence on skilled workers for result interpretation. They therefore deliver the rapid turnaround demanded by modern medicine. Antimicrobial susceptibility tests (ASTs) is a term used to describe a range of phenotypic methods that employ direct observation of the action of antimicrobials against a target microorganism. This is the most commonly used method in clinical diagnostic laboratories for detecting resistance in bacteria. A. Disc diffusion Growth medium: Standardized agar plates (usually unsupplemented, but addition(s) may be necessary for bacteria with specific growth requirements). Antibacterial component: Fixed dose in standard size circular paper discs or tablets.

ABSTRACT Background: Trichophyton rubrum is an infectious dermatophyte fungus which is the most common cause of dermatophytosis. Fungal resistance and the side effects of therapy are problems of antifungal agents. Phytochemical test of brotowali stem extract (Tinospora crispa) consist of flavonoids, phenols and triterpenoids which have antifungal effects. This study aimed to examine the effectiveness of brotowali stem extract (Tinospora crispa) as antifungal agent towards the growth of Trichophyton rubrum in vitro using agar well diffusion method. Subjects and Method: This was an experimental study using brotowali stem extract with concentration of 10%, 12,5%, 25%, 30%, 40%, 50%, 60%, 75%, and 100%. The dependent variable was Trichophyton rubrum growth. The independent variable was brotowali stem extract (Tinospora crispa). The data were obtained from the inhibition zone showed in agar well diffusion method in Sabouraud Dextrose Agar media. The data were analyzed using Kruskal-Wallis test. Results: The average diameter of inhibition zone of each variance (10%, 12,5%, 25%, 30%, 40%, 50%, 60%, 75%, and 100%) were 2.167 mm, 6.367 mm, 7.0 mm, 10.67 mm, 119 mm, 13.07 mm, 15.8 mm, 17.96 mm dan 17.13 mm, respectively, and they were statistically significant (p= 0.001). Conclusion: Brotowali stem extract has weak antifungal effectiveness at concentration 10%, 12,5%, 25%, intermediate antifungal effectiveness at concentration 30% and strong antifungal effectiveness at concentration 40%, 50%, 60%, 75% and 100%. Keywords: antifungal, brotowali stem, well diffusion, Trichophyton rubrum Correspondence: Fajriati Zulfa. Faculty of Medicine, Universitas Pembangunan Nasional ‘Veteran’ Jakarta. Jl. RS Fatmawati, Pondok Labu, Jakarta Selatan 12450, Telp. (021) 7656971. Email: nurulnerza@gmail.com DOI: https://doi.org/10.26911/the7thicph.05.02

ABSTRACT Background: Pityriasis versicolor or Tinea versicolor is a skin disease caused by the Malassezia furfur which is often found in Indonesia. People can use anti-fungal drugs to treat this disease. However, long-term use of anti-fungal drugs is relatively more expensive and can have side effects for its users. Cocoa bean husk contains flavonoids, saponins, and alkaloids which have anti-fungal effects. This study aimed to determine the antifungal effectiveness of the cocoa bean husk extract on the growth of M. furfur. Subjects and Methods: This was an experimental study using cocoa bean husk extract with a concentration variance of 25%, 50%, 75%, 100%, with a positive control for ketoconazole 2% and a negative control using distilled water. The test was carried out by the well diffusion method using Sabouraud Dextrose Agar media. The inhibition of fungal growth was calculated by looking at the clear zone formed after 48 hours. Data were analyzed using Kruskal-Wallis and Post hoc Mann Whitney statistical tests. Results: The mean diameter of the inhibition zone at a concentration of 25%, 50%, 75% and 100% was 3.42 mm, 4.07 mm, 4.9 mm, and 7.3 mm, respectively, and it was statistically significant (p = 0.001). Conclusion: Cocoa bean husk extract has weak anti-fungal effectiveness at concentrations of 25%, 50%, and 75%, while at 100% it has moderate effectiveness. Keywords: antifungal, Pityriasis versicolor, cocoa bean shell, well diffusion, Malassezia furfur Correspondence: Yuni Setyaningsih. Department of Parasitology, Faculty of Medicine, Universitas Pembangunan Nasional “Veteran” Jakarta. DOI: https://doi.org/10.26911/the7thicph.05.01