Determination Of Minimum Bactericidal Concentrations Research Articles

Determination of bactericidal activity against 3HC-2-Tre-labelled Mycobacterium abscessus (Mycobacteroides abscessus) by automated fluorescence microscopy

The minimum bactericidal concentration (MBC) of antibiotics is an important parameter for the potency of a drug in eradicating a bacterium as well as an important measure of the potential of a drug candidate in research and development. We have established a fluorescence-based microscopy method for the determination of MBCs against the non-tuberculous mycobacterium Mycobacterium abscessus (Mycobacteroides abscessus) to simplify and accelerate the performance of MBC determination compared to counting colony forming units on agar. Bacteria are labelled with the trehalose-coupled dye 3HC-2-Tre and analysed in a 96-well plate. The results of the new method are consistent with MBC determination by plating on agar. The method was used to evaluate the bactericidality of the antibiotics rifabutin, moxifloxacin, amikacin, clarithromycin and bedaquiline. Bactericidal effects against M. abscessus were observed, which are consistent with literature data.

Antibacterial and Antibiofilm Activity of Ethanol Extract of Batak Onion Bulbs (Allium chinense G.Don.) against Streptococcus mutans and Enterococcus faecalis.

Background: S.mutans is the main pathogen causing caries, while E.faecalis is the dominant microorganism in dental root canals. Batak onion is one of Indonesia's biological resources frequently used as condiments by Batak tribes and possesses antibacterial compounds such as alkaloids, saponins, and flavonoids. Aims: To analyze antibacterial activity test based on Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) and antibiofilm from 70% ethanol extract of Batak onion bulbs against S.mutans and E.faecalis. Material and Methods: This type of research is a laboratory experiment with a post-test only control group design. Determination of MIC by Kirby Baurer disc diffusion method, determination of MBC by streaking method, and colony tests are calculated using colony counters, determination of antibiofilm by Static Microtiter Plate Assays method, and checking the optical density at around 600nm. Data analysis of MIC, MBC, and antibiofilm using Oneway Anova and Post Hoc LSD tests. Results: MIC on S.mutans and E.faecalis was at a concentration of 0.78% with an average of one of inhibition of 9.00 ± 0.43 mm and 8.06 ± 0.20 mm; no MBC was found because no group was able to reduce bacteria 98-98%, Batak onion bulb extract has antibiofilm ability starting at a concentration of 0.78% with the ability to reduce S.mutans bacteria by 84.55% and 85.73% on E.faecalis. Conclusion: 70% ethanol extract of Batak onion bulbs can inhibit bacterial growth and biofilm of S.mutans and E.faecalis. The recommended dose for antibacterial and antibiofilm is a 6.25-50% concentration.

Photodynamic inactivation of Listeria monocytogenes using a natural aggregation-induced emission photosensitizer and its application in salmon preservation

Photodynamic sterilization has been regarded as a promising alternative to conventional antibacterial approaches. However, the photodynamic efficiency of conventional photosensitizer is often compromised by aggregation-induced quenching effects. Photosensitizers, especially natural products, with aggregation-induced emission (AIE) properties open a new avenue for photodynamic sterilization. Herein, we investigated the photodynamic antibacterial performance and mechanism of the AIE-type natural product of berberine (BBR) against Listeria monocytogenes (L. monocytogenes). The bactericidal effect was assessed through the determination of minimum bactericidal concentration and implementation of anti-biofilm experiments. The results showed that BBR can significantly suppress the proliferation of L. monocytogenes and can efficiently remove the L. monocytogenes-derived biofilm in a concentration-dependent trend. The mechanism of killing L. monocytogenes has been speculated based on bacterial morphology characterization, viability staining results, enzyme activity assays, DNA electrophoresis analysis, and molecular docking. The excellent anti-bacterial activity of BBR mediated photodynamic process may account for its abilities to the irreversibly disruption of bacterial membrane, inhibition of intracellular enzyme activity and decomposition of intracellular DNA molecules. We also demonstrated that BBR embedded sodium alginate/chitosan packaging film is feasible to maintain the freshness of salmon under light illumination by inhibiting the bacterial proliferation.

Antibacterial and Antibiofilm Potency of Menadione Against Multidrug-Resistant S. aureus.

Menadione is an analogue of 1,4-naphthoquinone (1,4-NQ) that possesses enormous pharmaceutical potential. The minimum inhibitory concentration (MIC) of menadione was determined against eighteen pathogens of the ESKAPE category, including thirteen multidrug-resistant (MDR) and five standard strains. From a total of eighteen pathogens, five strains of S. aureus (four: MDR and one: Standard strain) were considered further for detailed studies. This study included the determination of minimum bactericidal concentration (MBC), time-kill assay, scanning electron microscopic technique (SEM), and detection of reactive oxygen species (ROS). Additionally, the effect of menadione on biofilms of three strains of S. aureus was performed through crystal violet assay, SEM, and confocal laser scanning microscopy (CLSM). Menadione exerted substantial antibacterial activity against S. aureus (S8, S9, NCIM 5021) at a lower MIC (64µg/mL). Whereas, the MIC of 256µg/mL was displayed against J2 and J4 (MDR and biofilm-forming strains). The time-killing effect of menadione against S. aureus strains was observed after 9h at MBCs of 64µg/mL (NCIM 5021), 128µg/mL (S8, S9), and 512µg/mL (J2, J4). Enhanced levels of ROS in all five S. aureus were observed in presence of menadione (MICs and MBCs). The relation of enhanced ROS due to menadione activity invigorated us to explore its effect on S. aureus biofilms. We report menadione-mediated inhibition (> 90%) of biofilm formation (at respective MICs) and effect on preformed biofilms (> 85%) at 1024µg/mL. Menadione possessing antibacterial and antibiofilm potentials are imperative in the era of multidrug resistance developed by bacterial pathogens.

GC-MS and in vitro antibacterial potential of Cinnamomum camphora essential oil against some clinical antibiotic-resistant bacterial isolates.

Antibiotic resistance is increasing alarmingly in all parts of the world. Cinnamomum camphora (Linn.) Presl (C. camphora) is one of the earliest herbal remedies still in use today in traditional medicine. This study aimed to analyze the component of C. camphora grown widelyin Saudi Arabia (Qassim region) using GC-MS. Also, this study evaluates the in vitro antibacterial properties of C. camphora against certain clinical bacteria obtained from hospitals, including multi-drug resistant pathogens. Leaves of C. camphora tree were collected and essential oil was extracted for this study. The extract was subjected to GC-MS analysis. Eight clinical antibiotic-resistant pathogens were used in this study for the following assays: antibiotics susceptibility assay, determination of minimum inhibitory concentration (MIC), determination of minimum bactericidal concentration (MBC) and MIC index (MBC/MIC). The results show that the main components of the essential oil (EO) from the leaves ofC. camphorawere Eucalyptol. The EO had good antibacterial activity against eight clinical antibiotic-resistant pathogens, namely, Pseudomonas aeruginosa, Acinetobacter baumannii (two strains), Klebsiella pneumonia (two strains), Escherichia coli (one strain), Staphylococcus aureus (two strains). These findings may lead to a more complete knowledge of this aromatic plant's antibacterial action against antibiotic-resistant pathogens (in vitro).

Activity of Aegle marmelos (L) Correa (Buah Maja) Ethanol Extract to Multiresistant Eshecichia coli Clinical Isolates from Urinart Tract Infection Patients

The increase in resistance of Escherichia coli bacteria that caused urinary tract infections to antibiotics narrows the selection of effective treatment which encourages the development of treatment therapies using natural ingredients. Maja fruit [Aegle marmelos (L.) Correa] is a plant that has shown antibactericidal activity against gram-negative bacteria, one of which is Escherichia coli. This study aimed to determine the antibacterial activity of fruit Maja extract, minimum inhibition concentration (MIC) and the minimum bactericidal concentration (MBC) of fruit Maja extract and the comparative value of gentamycin against Escherichia coli resistance isolated from urinary tract infection patients. The methods are extraction of Maja fruit by using 96% ethanol, agar diffusion method with paper disc and determination of MIC and MBC using microdilution method. The results showed that the ethanol extract of Maja fruit had antibacterial activity with value of MIC50 as 1400-1500 μg/ml, the value of MIC90 was 1600 μg/ml and the value of MBC was in the range concentration of 1700-2000 μg/ml against E. coli isolate I. Against E. coli isolates II, value of MIC50 was in concentration of 2900 μg/ml, MIC90 was in concentration 3000 μg/ml and the value of the MBC is in range 3150-3500 μg/ml. The comparative values of the antibacterial activity of gentamycin on fruit maja extract are 1:5.871 x 103.

Study on the Minimum Bactericidal Concentration (MBC) of Maillard reaction products of Chitosan and Glucosamine prepared by Gamma-irradiation Method

The Maillard reaction products of chitosan and glucosamine (CTS-GA MRPs) were formed by gamma Co-60-irradiation method and determined their minimum bactericidal concentrations (MBC). The mixed solutions of chitosan - glucosamine were irradiated with a dose range of 0-100 kGy to induce the Maillard reaction products (MRPs). The formations of MRPs were determined by spectrophotometric analyses and the contents of remaining glucosamine were evaluated by high performance liquid chromatography (HPLC). The antibacterial activities of CTS-GA MRPs were tested by agar well diffusion test and MBC determination test against both gram negative (Escherichia coli) and positive bacteria (Bacillus subtilis) at acidic and alkaline pH was also carried out. By agar well diffusion test, irradiated CTS-GA solutions were able to form inhibition growth zones on E. coli plate whereas on B. subtilis plate, only CTS-GA MRPs irradiated at 25 kGy expressed this ability. The results of the MBC determination test indicated that CTS-GA MRPs formed at 25 kGy exhibited highly antibacterial activity in pH 5 and even pH 7. Therefore, this CTS-GA MRPs could be used as a promising preservative agent applied for meat and meat-product preservation.

ANTI-MICROBIAL EFFICACY OF AN ETHANOLIC EXTRACT OF MURRAYA KOENIGII AGAINST PERIODONTAL PATHOGENS LIKE PORPHYROMONAS GINGIVALIS AND AGGREGATIBACTER ACTINOMYCETEMCOMITANS: IN VITRO ANALYSIS

Periodontitis is primarily an infectious disease with Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans present in plaque biofilm as the key pathogens, causing destruction of supporting periodontal tissues. Herbal formulations are widely substituting synthetic anti-microbials due to their minimal adverse effects and cost effectiveness. Murraya koenigii (M. koenigii) is a well-known anti-microbial agent used in treating systemic infections. Hence its anti-microbial efficacy against key periodontal pathogens also needs to be tested. The aim of this study is to determine Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), and zone of inhibition of M. koenigii ethanolic extract against P. gingivalis and A. actinomycetemcomitans. Thioglycollate agar was used for culturing both P. gingivalis and A. actinomycetemcomitans. For determination of MIC and MBC broth dilution technique was used. For MIC, serial dilutions of extract were made and culture tubes were incubated in an anaerobic jar and observed for their turbidity. For MBC determination, dilution tubes sensitive to MIC were plated and incubated for the next 24 hours to monitor growth, and colony count was taken. MIC results showed A. actinomycetemcomitans was more sensitive to M. koenigii extract than P. gingivalis and got inhibited at 0.8 μg/ml. Also, MBC results showed extract has strong bactericidal activity towards A. actinomycetemcomitans. Disk diffusion test results showed bactericidal activity against both A. actinomycetemcomitans and P. gingivalis with a larger inhibition zone (15 mm) towards P. gingivalis at a concentration of 75 μl/ml. M. koenigii ethanolic extract is bactericidal against periodontal pathogens like A. actinomycetemcomitans and P. gingivalis and can be used as a safe and effective alternative for synthetic chemotherapeutic agents in the future.

In vivo Antibacterial Activity of Star Anise (Illicium verum Hook.) Extract Using Murine MRSA Skin Infection Model in Relation to Its Metabolite Profile.

IntroductionStar anise fruits (Illicium verum Hook.) have been used as an important treatment in traditional Chinese medicine. The previous studies reported the activity of the non-polar fractions as potential sources of antibacterial metabolites, and little was done concerning the polar fractions of star anise.MethodsThe antibacterial activity of the star anise aqueous methanolic (50%) extract against multidrug-resistant Acinetobacter baumannii AB5057 and methicillin-resistant Staphylococcus aureus (MRSA USA300) was investigated in vitro (disc diffusion assay, minimum bactericidal concentration determination, anti-biofilm activity and biofilm detachment activity). The antibacterial activity was further tested in vivo using a murine model of MRSA skin infection. Ultra-performance liquid chromatography coupled to high-resolution mass spectrometry (UPLC/HRMS) approach was applied for the identification of the metabolites responsible for the antibacterial activity. The antioxidant potential was evaluated using five in vitro assays: TAC (total antioxidant capacity), DPPH, ABTS, FRAP (ferric reducing antioxidant power) and iron-reducing power.ResultsIn vitro, star anise aqueous methanolic extract showed significant inhibition and detachment activity against biofilm formation by the multidrug-resistant and highly virulent Acinetobacter baumannii AB5057 and MRSA USA300. The topical application of the extract in vivo significantly reduced the bacterial load in MRSA-infected skin lesions. The extract showed strong antioxidant activity using five different complementary methods. More than seventy metabolites from different classes were identified: phenolic acids, phenylpropanoids, sesquiterpenes, tannins, lignans and flavonoids.ConclusionThis study proposes the potential use of star anise polar fraction in anti-virulence strategies against persistent infections and for the treatment of staphylococcal skin infections as a topical antimicrobial agent. To our knowledge, our research is the first to provide the complete polar metabolome list of star anise in an approach to understand the relationship between the chemistry of these metabolites and the proposed antibacterial activity.

Antibacterial Activity of Honey Samples from Ukraine.

The employment of natural substances such as beehive products with a preventive and therapeutic purpose has been a widespread custom since ancient times. In this investigation, the antibacterial activity of 41 honey samples from different Ukraine regions has been evaluated. For each honey, melissopalynological and physico-chemical analysis were performed in order to determine botanical origin, pH, glucose and fructose contents and free acidity. So, antibacterial activity against Staphylococcus aureus CCM 4223, Listeria monocytogenes ATCC 7644, Salmonella enterica serovar Typhimurium CCM 3807 and Escherichia coli ATCC 25922 was assessed through the determination of MIC (Minimum Inhibitory Concentration) and MBC (Minimum Bactericidal Concentration) values by the microdilutions method. The results show that the most susceptible bacterial strain was L. monocytogenes. Its growth was inhibited at a honey concentration ranging from 0.094 to 0.188 g/mL. The most resistant bacterial strain was S. aureus. As concerns MBC values, L. monocytogenes was the most susceptible bacteria, while S. aureus was the most resistant. Helianthus spp. honeys was the most effective against all tested bacterial strains, followed by Robinia spp. and multifloral honeys. Promising results for MIC tests have been found for Brassica spp.

Minimum bactericidal concentration of ciprofloxacin to Pseudomonas aeruginosa determined rapidly based on pyocyanin secretion

Infections due to Pseudomonas aeruginosa (P. aeruginosa) often exhibit broad-spectrum resistance and persistence to common antibiotics. Persistence is especially problematic with immune-compromised subjects who are unable to eliminate the inhibited bacteria. Hence, antibiotics must be used at the appropriate minimum bactericidal concentration (MBC) rather than at minimum inhibitory concentration (MIC) levels. However, MBC determination by conventional methods requires a 24 h culture step in the antibiotic media to confirm inhibition, followed by a 24 h sub-culture step in antibiotic-free media to confirm the lack of bacterial growth. We show that electrochemical detection of pyocyanin (PYO), which is a redox-active bacterial metabolite secreted by P. aeruginosa, can be used to rapidly assess the critical ciprofloxacin level required for bactericidal deactivation of P. aeruginosa within just 2 h in antibiotic-treated growth media. The detection sensitivity for PYO can be enhanced by using nanoporous gold that is modified with a self-assembled monolayer to lower interference from oxygen reduction, while maintaining a low charge transfer resistance level and preventing electrode fouling within biological sample matrices. In this manner, bactericidal efficacy of ciprofloxacin towards P. aeruginosa at the MBC level and bacterial persistence at the MIC level can be determined rapidly, as validated at later timepoints using bacterial subculture in antibiotic-free media.

Minimum Bactericidal Concentration Techniques in Mycobacterium tuberculosis: A Systematic Review.

Minimum bactericidal concentration (MBC) assay is an accepted parameter for evaluating new antimicrobial agents, and it is frequently used as a research tool to provide a prediction of bacterial eradication. To the best of our knowledge, there is no standardization among researchers regarding the technique used to detect a drug's MBC in Mycobacterium tuberculosis. Thus, the aim of this systematic review is to discuss the available literature in determining a drug's MBC in M. tuberculosis, to find the most commonly used technique and standardize the process. A broad and rigorous literature search of three electronic databases (PubMed, Web of Knowledge, and LILACS) was performed according to the PRISMA statement. We considered studies that were published from January 1, 1990 to February 19, 2019. Google Scholar was also searched to increase the number of publications. We searched for articles using the MeSH terms "microbiological techniques," "Mycobacterium," "antibacterial agents." In addition, free terms were used in the search. The search yielded 6,674 publications. After filter application, 5,348 publications remained. Of these, we evaluated the full text of 187 publications. By applying the inclusion criteria, 69 studies were included in the present systematic review. In the literature analyzed, a great variety in the techniques used to determine a drug's MBC in M. tuberculosis was observed. The most common variability is related to the culture media used, culture incubation time, and the percentage of bacterial death for the drug to be considered as bactericidal. The most commonly used technique for drug's MBC determination was carried out using the drug's minimum inhibitory concentration (MIC) assay. Aliquots from prior MIC values were subcultured in Middlebrook agar and incubated for 4 weeks at 35°C for determining the colony forming unit (CFU) with relevance to detect 99.9% bacilli killed or reduction in 3 log10 viable bacilli.

Determination of minimum bactericidal concentration, in single or combination drugs, against Mycobacterium tuberculosis.

Aim: To evaluate an assay to detect minimum bactericidal concentration (MBC) in Mycobacterium tuberculosis, using as single model rifampicin, isoniazid, levofloxacin (LVX) and linezolid (LNZ) and in combination. Material &methods: MBCs were carried out directly from resazurin microtiter assay plate and 3D checkerboard in M. tuberculosis H37Rv and five resistant clinical isolates. Results: The proposed MBC assay showed similar values to those determined by MGIT™, used as control. LVX and LNZ's MBC values were close to their MIC values. LNZ or LVX combined with isoniazid and rifampicin showed MBC value reduced in 63.7% of the assays. Conclusion: The proposed assay to determine MBCs of drugs can be applied to the study of new compounds with anti-M. tuberculosis activity to detect their bactericidal effect and also in laboratory routine for clinical dose adjustment of drugs according to the patient's profile.

Design, synthesis and antibacterial evaluation of novel 15-membered 11a-azahomoclarithromycin derivatives with the 1, 2, 3-triazole side chain

Macrolides are widely prescribed in clinic to treat various respiratory tract infections. However, due to their inappropriate use, the prevalence of macrolide-resistant strains among clinical isolates has become a concern for public health. Therefore, novel macrolides skeleton structures against resistant pathogens are badly needed. Thus, three series of novel 15-membered 11a-azahomoclarithromycin derivatives (series A-C) with the 1, 2, 3-triazole side chain were designed and synthesized through creatively opening the ring of clarithromycin (CAM), expanding the ring properly and introducing a suitable side chain of 1, 2, 3-triazole at the C12 and C13 positions, and evaluated for their antibacterial activity. The antibacterial results indicated that compounds 38b, 38l and 38v possessed strong antibacterial activity against Staphylococcus aureus ATCC25923 (0.25 μg/mL) and Bacillus subtilis ATCC9372 (0.25 μg/mL). Furthermore, compounds 9e and 38g were found to exhibit promising potent activity (8 μg/mL) against Streptococcus pneumonia AB11 expressing the ermB and mefA genes. In addition, the determination of minimum bactericidal concentration (MBC) indicated that the most promising compounds 38b, 38l, 38v, 9e and 38g were excellent bacteriostatic agents. The bactericidal curve showed that 9e exhibited antibacterial activity through bacteriostatic mechanism. Finally, 38b, 38l and 38v were confirmed to be non-toxic to MCF-7 breast cancer cells up to a concentration of 32 μg/mL in preliminary cytotoxicity assay. In summary, 38b, 38l, 38v, 9e and 38g can be served as lead compounds to provide a new perspective for further structural optimization.

Synergistic-antagonistic antibacterial potential of chitosan composites with <i>Moringa oleifera</i> leaf powder

Chitosan is very useful in everyday life in adsorption, cosmetics, pharmaceuticals, flocculants, anticancer and antimicrobial.In this study, chitosan was synthesized from chitin extracted from crayfish. The methods such as deproteinization, demineralization, and deacetylation respectively were used in the synthesis of chitosan from crayfish. Antimicrobial activity was studied and it was found that chitosan and Moringaleaf powder were good in inhibiting the growth of microorganisms; confirmed by the results obtained from the experiments. In evaluating the antimicrobial activity, the serial dilution method was used towards Escherichia coli, Staphylococcus aureus, Salmonella typhi, Proteus bulgaris and Streptococcus pneumonia. The antibacterial activity of chitosan composite with the leaf powder of Moringa oleifera Lam., was determined, using well diffusion, minimum inhibitory concentration (MIC) and minimum bactericidal concentration determination method. The composites show the synergistic effect at the higher chitosan to lower Moringa concentration and antagonistic effect at higher Moringa to lower chitosan concentrations in all the test organisms. The consequences of this research suggest that the chitosan, Moringa leaf powder, and their composites can be used to discover an antibacterial agent for developing new pharmaceuticals to control studied human pathogenic bacteria responsible for severe illness. Moringa oleiferais widely used in food and folk medicine; while chitosan is widely useful in food, detergents, textiles, leather, paper, pharmaceuticals, and cosmetics industries. Synergism/antagonism of Moringa-chitosan composites was based on concentrations on the tested organisms.Keywords: Antimicrobial, Biopolymer, Health, Synergism/antagonism

Fast-acting bactericidal activity of olanexidine gluconate against qacA/B-positive methicillin-resistant Staphylococcus aureus.

The qacA/B gene is one of the major determinants of resistance to antiseptics in methicillin-resistant Staphylococcus aureus (MRSA). Here, we compared the fast-acting bactericidal activity of skin antiseptics, including olanexidine gluconate (OLG), a new biguanide antiseptic agent introduced in Japan, against clinical qacA/B-positive MRSA strains by determination of minimum bactericidal concentration and time-kill assay. Our findings provide, for the first time, data indicating that the fast-acting bactericidal activity of OLG against qacA/B-positive MRSA is higher than that of chlorhexidine gluconate, even though both are biguanide antiseptics.

Assessments of Antibacterial Effects of Aqueous-Ethanolic Extracts of Sida rhombifolia's Aerial Part.

Infectious diseases are the critical problems of the world as a result of the emergence of different antimicrobial resistant microorganisms due to several reasons like misuses and repeating uses of antibiotics. Because of this, searching of new treatment method is important from natural substances to against those infectious diseases in both human and animals' aspects. Among those plants, Sida rhombifolia has various roles against those infectious diseases through its different phytochemical components. The objectives of this study were assessing the antibacterial activity of the aqueous-methanol extract of the plant's aerial part and knowing the phytochemical constituents of the plant. Preliminary phytochemical screening revealed that the extract of S. rhombifolia's aerial part possesses flavonoids, alkaloids, polyphenols, and quinines. In addition to this, the antibacterial activity of the plant extract was evaluated on five pathogenic bacteria species using agar well diffusion method at different concentrations of plant extracts. Minimum inhibition concentration and minimum bactericidal concentration determinations were done by tetrazolium chloride microtiter dilution assay. The inhibition zone of mean diameters ranging from 0.00 to 7.67mm against all test bacteria was significantly (p<0.05) much less than that of the positive control Chloroamphinicole (30μg/disc) with the range of 14.33mm-15mm of inhibition zone of diameters. The inhibition zones of the tested bacteria at the concentration of 62.5mg/ml were much less than the higher concentration (500mg/ml) and significantly different (p<0.05), whereas the MIC value ranges from 4.62 to 97.22mg/ml and the MBC value ranges from 4.62 to 125.00mg/ml. Even if the plant extract showed antibacterial activity, it was lower than that of other solvent extraction methods; so other solvent extraction methods and fractionates must be conducted to investigate the antibacterial activities of the plant extract on different bacterial strains and species that cause different diseases.

A new approach to determination of bactericidal activity of benzylpenycylin on Corynebacterium diphtheriae

Information on the bactericidal activity of benzylpenycylin is important when planning antibiotic therapy for a number of diseases caused by Corynebacterium diphtheriae (endocarditis, bacteremia, septicemia, etc.). However, methods for determining the minimum bactericidal concentration (MBC) of antibiotics are complicated. The aim of the study was to develop a method for testing of diphtheria causative agent susceptibility to the bactericidal action of penicillin without MBC determination. The minimum inhibitory concentrations (MICs) and MBCs in 80 strains of C. diphtheriae were determined using the standard broth dilution method (macromethod). The MICs were registered after 24 and 48 years of growth. After the first day of growth the MIC of penicillin was in the range of 0.017 to 0.5 mg / L, after second day — in the range of 0.035 to 0.5 mg / liter. The increase of the MICs for the second day of growth was observed in 47.5±5.6% of strains. MIC50 and MIC90 both after 24 hours and after 48 hours of growth were 0.13 and 0.25 mg/L, respectively. MBCs of penicillin was in the range of 0.5 to 32.0 mg/L, MBC50 was 4.0 mg/l, MBC90 – 8.0 mg/l. In 35,0±5,3% of the studied strains, high MBCs (8.0 mg / l or higher) was detected. It has been established that in C.diphtheriae the relationship between the MIC and the MBC of penicillin is clearly expressed, taking into account the two values of the MIC – for the first and second day of growth. Investigated strains are divided into three conditional groups: 1) with MIC 0.13 mg/L and basically with low MBCs; 2) with MIC 0.25 mg/L, in most cases are not susceptible to bactericidal action, and 3) strains with MIC 0.5 mg/L and high MBCs. The most pronounced were differences in the prevalence of corynebacteria with high MBCs in two groups of strains: MICs which did not exceed 0.13 mg/L after 24 and 48 hours, and those in whom the MIC was 0.25 mg/L or higher already on the first day of incubation (Student's coefficient t=4.13, p&lt;0.001). The obtained results can be used to improve the methods for determining of corynebacteria susceptibility to antimicrobials.

One-pot synthesis of biologically active 1,2,3-trisubstituted pyrrolo[2,3-b]quinoxalines through a palladium-catalyzed reaction with internal alkyne moieties.

Synthesis of 2,3-disubstituted 1-alkylpyrrolo[2,3-b]quinoxalines was accomplished through the reaction of 3-chloroquinoxalin-2-amines with internal alkynes in the presence of Pd(OAc)[Formula: see text], NaOAc, and KOtBu in DMSO. This method afforded desired pyrrolo[2,3-b]quinoxalines in 65-92% reaction yields. The minimum inhibition concentration and minimum bactericidal concentration determinations against Micrococcus luteus and Pseudomonas aeruginosa revealed that some of the synthesized compounds showed the same values compared to tetracycline. These compounds could be used in the future research for the development of new antibiotics.

Combining diagnostic methods for antimicrobial susceptibility testing – A comparative approach

BackgroundThe minimum inhibitory concentration (MIC) is a measure of antimicrobial susceptibility testing (AST) of a given antibiotic but provides insufficient information when bacterial killing is crucial, e.g., when treating immunocompromised patients. In these cases, the minimum bactericidal concentration (MBC) is a more reliable measure of antibiotic activity. Here, we aim to demonstrate and recommend combinations of methods for MIC and MBC measurements. We also aim to emphasize the importance of uniform protocols for these procedures including the time point for reading MIC results, which the authors suggest to be 20h. MethodsTo address the challenges with obtaining fast and reliable readouts on MIC as well as the kinetic and end-point effects of antibiotics, the broth micro dilution method, a calorimetric method and a microscopy-based screening system (MBSS) were evaluated in this study. For MBC determination, fluorophore staining with SYTO9 and propidium iodide was compared to the broth regrowth method. ResultsThree scenarios for combining the MIC and MBC methods depending on the investigators' primary concern (time, cost or sensitivity) are presented. Further, as the MBSS and the isothermal microcalorimetry method detected delayed bacterial growth up to 18h after initiation of experiments, the importance of reading MIC testing after a full 20h is emphasized. A one-fold change in MIC values can be observed when comparing data obtained at 16h and 20h of incubation. ConclusionThe authors suggest that combining MIC and MBC determinations will provide more detailed understanding of the bacteria susceptibility to antibiotic drugs and result in more clinically relevant data and optimized therapies. Furthermore, establishing 20h as a time point for reading MIC results will provide more uniform data across laboratories.