Insight into the Antibacterial Action of Iodinated Imine, an Analogue of Rafoxanide: a Comprehensive Study of Its Antistaphylococcal Activity.

In this study, we have focused on a multiparametric microbiological analysis of the antistaphylococcal action of the iodinated imine BH77, designed as an analogue of rafoxanide. Its antibacterial activity against five reference strains and eight clinical isolates of Gram-positive cocci of the genera Staphylococcus and Enterococcus was evaluated. The most clinically significant multidrug-resistant strains, such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant S. aureus (VRSA), and vancomycin-resistant Enterococcus faecium, were also included. The bactericidal and bacteriostatic actions, the dynamics leading to a loss of bacterial viability, antibiofilm activity, BH77 activity in combination with selected conventional antibiotics, the mechanism of action, in vitro cytotoxicity, and in vivo toxicity in an alternative animal model, Galleria mellonella, were analyzed. The antistaphylococcal activity (MIC) ranged from 15.625 to 62.5 µM, and the antienterococcal activity ranged from 62.5 to 125 µM. Its bactericidal action; promising antibiofilm activity; interference with nucleic acid, protein, and peptidoglycan synthesis pathways; and nontoxicity/low toxicity in vitro and in vivo in the Galleria mellonella model were found to be activity attributes of this newly synthesized compound. In conclusion, BH77 could be rightfully minimally considered at least as the structural pattern for future adjuvants for selected antibiotic drugs. IMPORTANCE Antibiotic resistance is among the largest threats to global health, with a potentially serious socioeconomic impact. One of the strategies to deal with the predicted catastrophic future scenarios associated with the rapid emergence of resistant infectious agents lies in the discovery and research of new anti-infectives. In our study, we have introduced a rafoxanide analogue, a newly synthesized and described polyhalogenated 3,5-diiodosalicylaldehyde-based imine, that effectively acts against Gram-positive cocci of the genera Staphylococcus and Enterococcus. The inclusion of an extensive and comprehensive analysis for providing a detailed description of candidate compound-microbe interactions allows the valorization of the beneficial attributes linked to anti-infective action conclusively. In addition, this study can help with making rational decisions about the possible involvement of this molecule in advanced studies or may merit the support of studies focused on related or derived chemical structures to discover more effective new anti-infective drug candidates.

Improving the antimicrobial activity of old antibacterial drug mafenide: Schiff bases and their bioactivity targeting resistant pathogens.

Background: Increasing rates of acquired resistance have justified the critical need for novel antimicrobial drugs. One viable concept is the modification of known drugs. Methods & results: 21 mafenide-based compounds were prepared via condensation reactions and screened for antimicrobial efficacy, which demonstrated promising activity against both Gram-positive and Gram-negative pathogens, pathogenic fungi and mycobacterial strains (minimum inhibitory concentrations from 3.91 µM). Importantly, they retained activity against a panel of superbugs (methicillin- and vancomycin-resistant staphylococci, enterococci, multidrug-resistant Mycobacterium tuberculosis) without any cross-resistance. Unlike mafenide, most of its imines were bactericidal. Toxicity to HepG2 cells was also investigated. Conclusion: Schiff bases were significantly more active than the parent drug, with iodinated salicylidene and 5-nitrofuran/thiophene-methylidene scaffolds being preferred in identifying the most promising drug candidates.

Comprehensive insight into anti-staphylococcal and anti-enterococcal action of brominated and chlorinated pyrazine-based chalcones.

The greatest threat and medicinal impact within gram-positive pathogens are posed by two bacterial genera, Staphylococcus and Enterococcus. Chalcones have a wide range of biological activities and are recognized as effective templates in medicinal chemistry. This study provides comprehensive insight into the anti-staphylococcal and anti-enterococcal activities of two recently published brominated and chlorinated pyrazine-based chalcones, CH-0y and CH-0w. Their effects against 4 reference and 12 staphylococcal and enterococcal clinical isolates were evaluated. Bactericidal action, the activity in combination with selected conventional antibiotics, the study of post-antimicrobial effect (PAE, PAE/SME), and in vitro and in vivo toxicity, were included. In CH-0y, anti-staphylococcal activity ranging from MIC = 15.625 to 62.5 µM, and activity against E. faecium from 31.25 to 62.5 µM was determined. In CH-0w, anti-staphylococcal activity ranging from 31.25 to 125 µM, and activity against E. faecium and E. faecalis (62.5 µM) was revealed. Both CH-0y and CH-0w showed bactericidal action, beneficial impact on bacterial growth delay within PAE and PAE/SME studies, and non/low toxicity in vivo. Compared to CH-0w, CH-0y seems to have higher anti-staphylococcal and less toxic potential. In conclusion, chalcones CH-0y and CH-0w could be considered as structural pattern for future adjuvants to selected antibiotic drugs.

The Impact of Cultivation Media on the In Vitro Biofilm Biomass Production of Candida spp.

The yeasts of the genus Candida are among the most clinically significant fungal pathogenic agents. One of the unique features of the Candida species' pathogenicity is their ability to form biofilms. Generally, infections caused by biofilm-forming microorganisms tend to have chronic course and are difficult to treat. This fact highlights the need to search for drugs with anti-biofilm activities. At present, there are variety of protocols for performing antifungal anti-biofilm activity testing in which fundamental differences, especially in the choice of cultivation media for biofilm formation, can be noted. In our study, we focused on the effect of four different culture media on biofilm biomass formation in ten Candida spp. strains. With emphasis placed on clinical significance, strains of the C. albicans species were predominantly included in this study. Based on our results, we can conclude that the availability of other components in the culture media, such as amino acids or proteins, and not just the commonly mentioned glucose availability, helps promote the transition of Candida yeasts into a sessile form and leads to in vitro robust biofilm formation. We revealed that biofilm formation in C. albicans strains was enhanced, especially in media supplemented with fetal bovine serum (FBS). The nutritionally balanced cultivation medium with 10 g/L glucose and 10% (v/v) FBS evidently showed the most significant benefit for in vitro biofilm production in C. albicans strains.

A comparative analysis of protein virulence factors released via extracellular vesicles in two Candida albicans strains cultivated in a nutrient-limited medium.

One of the pathways for the delivery of virulence effector molecules into the extracellular environment of Candida albicans relies on the release of membrane-bound carriers which are called extracellular vesicles (EVs). Only a few studies aimed at investigating Candida albicans extracellular vesicles protein cargo and its potential contribution to the pathogenesis of C. albicans infections have been conducted to date. In this study, we mainly focused on a search for proteins with a demonstrated linkage to pathogenesis in EVs isolated from two C. albicans strains, the model strain ATCC 90028 and the clinical isolate from a woman suffering from vulvovaginal candidiasis. For the purpose of mimicking one of many hostile conditions during a host-pathogen interaction, C. albicans strains in a nutrient-limited medium were cultivated. We have hypothesized that this unfavourable, stressful condition could contribute to the induction of virulence effector molecules being released at a more extensive rate. In conclusion, 34 proteins with an undisputed linkage to C. albicans pathogenesis were detected in the extracellular vesicle cargoes of both strains. In case of the clinical isolate strain, no unique virulence-associated proteins were detected. In the C. albicans ATCC 90028 model strain, three unique proteins were detected, namely: agglutinin-like protein 3 (Als3), secreted aspartic protease 8 (Sap8) and cell surface superoxide dismutase [Cu-Zn] 6 (Sod6).

Synthesis and Antifungal Screening of 2-{[1-(5-Alkyl/arylalkylpyrazin-2-yl)ethylidene]hydrazono}-1,3-thiazolidin-4-ones.

Two novel thiosemicarbazones and eight novel 2-{[1-(5-alkyl/arylalkylpyrazin-2-yl)ethylidene]hydrazono}-1,3-thiazolidin-4-ones were prepared and tested against a panel of eight fungal strains-Candida albicans ATCC 44859, Candida tropicalis 156, Candida krusei E 28, Candida glabrata 20/I, Trichosporon asahii 1188, Aspergillus fumigatus 231, Lichtheimia corymbifera 272, and Trichophyton interdigitale 445. 1,3-Thiazolidin-4-ones exhibited activity against all strains, the most potent derivative was 2-{[1-(5-butylpyrazin-2-yl)ethylidene]hydrazono}e-1,3-thiazolidin-4-one. Susceptibility of C. glabrata to the studied 1,3-thiazolidin-4-ones (minimum inhibitory concentrations (MICs) were in the range 0.57 to 2.78 mg/L) is of great interest as this opportunistic pathogen is poorly susceptible to azoles and becomes resistant to echinocandins. Antifungal potency of thiosemicarbazones was slightly lower than that of 1,3-thiazolidin-4-ones.

Novel Halogenated Pyrazine-Based Chalcones as Potential Antimicrobial Drugs.

Chalcones, i.e., compounds with the chemical pattern of 1,3-diphenylprop-2-en-1-ones, exert a wide range of bio-activities, e.g., antioxidant, anti-inflammatory, anticancer, anti-infective etc. Our research group has been focused on pyrazine analogues of chalcones; several series have been synthesized and tested in vitro on antifungal and antimycobacterial activity. The highest potency was exhibited by derivatives with electron withdrawing groups (EWG) in positions 2 and 4 of the ring B. As halogens also have electron withdrawing properties, novel halogenated derivatives were prepared by Claisen-Schmidt condensation. All compounds were submitted for evaluation of their antifungal and antibacterial activity, including their antimycobacterial effect. In the antifungal assay against eight strains of selected fungi, growth inhibition of Candida glabrata and Trichophyton interdigitale (formerly T. mentagrophytes) was shown by non-alkylated derivatives with 2-bromo or 2-chloro substitution. In the panel of selected bacteria, 2-chloro derivatives showed the highest inhibitory effect on Staphylococcus sp. In addition, all products were also screened for their antimycobacterial activity against Mycobacterium tuberculosis H37RV My 331/88, M. kansasii My 235/80, M. avium 152/80 and M. smegmatis CCM 4622. Some of the examined compounds, inhibited growth of M. kansasii and M. smegmatis with minimum inhibitory concentrations (MICs) comparable with those of isoniazid.

Development of 3,5-Dinitrobenzylsulfanyl-1,3,4-oxadiazoles and Thiadiazoles as Selective Antitubercular Agents Active Against Replicating and Nonreplicating Mycobacterium tuberculosis.

Herein, we report the discovery and structure-activity relationships of 5-substituted-2-[(3,5-dinitrobenzyl)sulfanyl]-1,3,4-oxadiazoles and 1,3,4-thiadiazoles as a new class of antituberculosis agents. The majority of these compounds exhibited outstanding in vitro activity against Mycobacterium tuberculosis CNCTC My 331/88 and six multidrug-resistant clinically isolated strains of M. tuberculosis, with minimum inhibitory concentration values as low as 0.03 µM (0.011-0.026 µg/mL). The investigated compounds had a highly selective antimycobacterial effect because they showed no activity against the other bacteria or fungi tested in this study. Furthermore, the investigated compounds exhibited low in vitro toxicities in four proliferating mammalian cell lines and in isolated primary human hepatocytes. Several in vitro genotoxicity assays indicated that the selected compounds have no mutagenic activity. The oxadiazole and thiadiazole derivatives with the most favorable activity/toxicity profiles also showed potency comparable to that of rifampicin against the nonreplicating streptomycin-starved M. tuberculosis 18b-Lux strain, and therefore, these derivatives, are of particular interest.

Novel pyrazine analogs of chalcones: synthesis and evaluation of their antifungal and antimycobacterial activity.

Infectious diseases, such as tuberculosis and invasive mycoses, represent serious health problems. As a part of our long-term efforts to find new agents for the treatment of these diseases, a new series of pyrazine analogs of chalcones bearing an isopropyl group in position 5 of the pyrazine ring was prepared. The structures of the compounds were corroborated by IR and NMR spectroscopy and their purity confirmed by elemental analysis. The susceptibility of eight fungal strains to the studied compounds was tested. The results have been compared with the activity of some previously reported propyl derivatives. The only strain that was susceptible to the studied compounds was Trichophyton mentagrophytes. It was found that replacing a non-branched propyl with a branched isopropyl did not have a decisive and unequivocal influence on the in vitro antifungal activity against T. mentagrophytes. In vitro activity against Trichophyton mentagrophytes comparable with that of fluconazole was exhibited by nitro-substituted derivatives. Unfortunately, no compound exhibited efficacy comparable with that of terbinafine, which is the most widely used agent for treating mycoses caused by dermatophytes. Some of the prepared compounds were assayed for antimycobacterial activity against M. tuberculosis H37Rv. The highest potency was also displayed by nitro-substituted compounds. The results of the present study are in a good agreement with our previous findings and confirm the positive influence of electron-withdrawing groups on the B-ring of chalcones on the antifungal and antimycobacterial activity of these compounds.

Synthesis and antimycobacterial evaluation of 5-alkylamino-N-phenylpyrazine-2-carboxamides.

Substitution of chlorine in 5-chloro-N-phenylpyrazine-2-carboxamide (1) with simple n-alkylamines yielded a series of 5-alkylamino-N-phenylpyrazine-2-carboxamides (propylamino to octylamino derivatives), which possessed similar or increased activity against Mycobacterium tuberculosis H37Rv compared to parent 5-chloro derivative (1), with MIC ranging from 2.5 to 12.2 µM. 5-Butylamino to 5-heptylamino derivatives exerted similar activity also against Mycobacterium kansasii. Importantly, the substitution led also to significant decrease of in vitro cytotoxicity in HepG2 cell line. 5-Heptylamino-N-phenylpyrazine-2-carboxamide (1e) exerted MIC=2.5 µM (M.tbc) and IC50 >250 µM (HepG2). Further modification of alkylamino chain with terminal methoxy or hydroxy group lead to compounds with decreased or none activity, the decrease was proportional to the decrease of lipophilicity. 5-(2-Phenylethylamino) and 5-(3-phenylpropylamino) derivatives were also of decreased activity. On contrary to alkylamino derivatives derived from 1, alkylamino derivatives derived from 5-chloro-N-2-chlorophenylpyrazine-2-carboxamide (2) possessed substantially decreased or none activity. None of the prepared compounds was active against Mycobacterium avium.

6-Hydroxyquinolinium salts differing in the length of alkyl side-chain: synthesis and antimicrobial activity.

Quaternary ammonium salts substituted with a long alkyl chain exemplify a trustworthy group of medicinal compounds frequently employed as antifungal and antibacterial agents. A great asset of these surfactants underlying their widespread use is low local and system toxicity in humans. In this Letter, a series of novel quaternary 6-hydroxyquinolinium salts with varying length of N-alkyl chain (from C10 to C18) was synthesized and tested for in vitro activity against pathogenic bacterial and fungal strains. 6-Hydroxyquinolinium salt with C12 alkyl chain seems to be very interesting candidate due to a high antimicrobial efficacy and cytotoxic safety.

Cryptococcus neoformans meningoencephalitis in a patient with polyarteritis nodosa.

Case of 59-year-old male with chronic obstructive pulmonary disease and a number of comorbidities, who has developed meningoencephalitis caused by Cryptococcus neoformans var. grubii with polyarteritis nodosa diagnosed during hospitalization, was presented. Before evidence of meningoencephalitis, the patient was being treated with ketoconazole and low doses of fluconazole (200 mg/day) for alleged candidiasis. The dosage was increased (800 mg/day) following laboratory diagnosis of C. neoformans based on positive latex agglutination test and biochemical identification of encapsulated yeast isolated from the blood and CSF. Later, the yeast identification was confirmed by sequencing analysis. Owing to inadequate clinical response, fluconazole therapy was switched to voriconazole (400 mg/day) and later to intravenous amphotericin B (1.0 mg/kg per day). Despite of a temporary stabilization and improvement, which correlated with decline of cryptococcal antigen titers (from 1:1024 to 1:8), after 6 weeks, the patient's underlying condition deteriorated due to severe pancolitis and serious nosocomial bacterial infections. The patient died of multiorgan failure several days later. Our case demonstrates a possible connection between the development of life-threatening cryptococcosis and an autoimmune vasculitis disease and emphasizes that the outcome of the management of cryptococcal meningoencephalitis is highly dependent on early diagnosis, adequate treatment, including dosage, and last but not least control of underlying disease and risk factors.

1-Substituted-5-[(3,5-dinitrobenzyl)sulfanyl]-1H-tetrazoles and their isosteric analogs: A new class of selective antitubercular agents active against drug-susceptible and multidrug-resistant mycobacteria.

In this work, a new class of highly potent antituberculosis agents, 1-substituted-5-[(3,5-dinitrobenzyl)sulfanyl]-1H-tetrazoles and their oxa and selanyl analogs, is described. The minimal inhibitory concentration (MIC) values reached 1 µM (0.36-0.44 µg/mL) against Mycobacterium tuberculosis CNCTC My 331/88 and 0.25-1 µM against six multidrug-resistant clinically isolated strains of M. tuberculosis. The antimycobacterial effects of these compounds were highly specific because they were ineffective against all eight bacterial strains and eight fungal strains studied. Furthermore, these compounds exhibited low in vitro toxicity in four mammalian cell lines (IC50 > 30 µM). We also examined the structure-activity relationships of the compounds, particularly the effects on antimycobacterial activity of the number and position of the nitro groups, the linker between tetrazole and benzyl moieties, and the tetrazole itself. Relatively high variability of substituent R(1) on the tetrazole in the absence of negative effects on antimycobacterial activity allows further structural optimization with respect to toxicity and the ADME properties of the 1-substituted-5-[(3,5-dinitrobenzyl)sulfanyl]-1H-tetrazoles lead compounds.

Outbreak of fungal endophthalmitis due to Fusarium oxysporum following cataract surgery.

Outbreak of exogenous Fusarium endophthalmitis after cataract surgery was evaluated. Twenty patients developed postoperative endophthalmitis. In 19 eyes, pars plana vitrectomy (PPV) was performed, in 14 cases (74 %) with primary intraocular lens explantation. In one case, the PPV was not performed because of poor general condition of the patient. Symptoms of endophthalmitis (damaged vision, iritis, tyndallization in anterior chamber, hypopyon) occurred at intervals of 16-79 days (mean 31.3 days). Fungal etiology was documented in 12 eyes (60 %). Fusarium oxysporum was evidenced by culture and/or microscopy and confirmed by PCR and sequencing analysis. Eighteen (90 %) patients were treated with oral voriconazole (400 mg/day) for a period of 4-6 weeks. The final visual acuity was 6/15 in 1 case (5 %), 6/60 and worse in 17 eyes (85 %), and in 2 cases (10 %), enucleation had to be performed. Viscoelastic filling material was suggested the most likely source of infection. Endophthalmitis caused by Fusarium spp. are a potentially big threat for patients with serious impact on vision. Successful management of the infection is highly dependent on early diagnosis including species identification and antifungal susceptibility testing, and on aggressive and long-term treatment.

Investigation of the biological properties of (hetero)aromatic thiosemicarbazones.

Two series of thiosemicarbazone-based iron chelators (twenty-seven compounds) were designed and synthesized using a microwave-assisted approach. Quinoline and halogenated phenyl were selected as parent scaffolds on the basis of a similarity search. The lipophilicity of the synthesized compounds was measured using HPLC and then calculated. Primary in vitro screening of the synthesized compounds was performed against eight pathogenic fungal strains. Only a few compounds showed moderate activity against fungi, and (E)-2-(quinolin-2-ylvinyl)-N,N-dimethylhydrazine-carbothioamide appeared to be more effective than fluconazole against most of the fungal strains tested. Antiproliferative activity was measured using a human colon cancer cell line (HCT-116). Several of the tested compounds showed submicromolar antiproliferative activity. Compounds were also tested for their activity related to the inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. The structure-activity relationships are discussed for all of the compounds.

Anti-infective and herbicidal activity of N-substituted 2-aminobenzothiazoles.

In this study, a series of N-substituted 2-aminobenzothiazoles was prepared according to a recently developed method. Twelve compounds were tested for their activity related to the inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. Primary in vitro screening of the discussed compounds was also performed against fungal, bacterial and mycobacterial species. The biological activities of some compounds were comparable or higher than the standards phenoxymethylpenicillin or pyrazinamide. The most effective compounds demonstrated insignificant toxicity against the human monocytic leukemia THP-1 cell line. For all compounds, the structure-activity relationships are discussed.

Contribution to investigation of antimicrobial activity of styrylquinolines.

Series of new ring-substituted styrylquinolines and two oxorhenium complexes were prepared and characterized. The compounds were analyzed using RP-HPLC to determine lipophilicity. Primary in vitro screening of the synthesized compounds was performed against fungal and bacterial strains. Some compounds were active against bacteria at micromolar level and against fungi at submicromolar level. Compounds 5,7-dichloro-2-[2-(2-ethoxyphenyl)vinyl]quinolin-8-ol expressed excellent antifungal activity comparable with or higher than the standard fluconazole as well as antibacterial activity against Staphylococcus strains comparable with or higher than the standards bacitracin, penicillin and ciprofloxacin. The structure-activity relationships are discussed.

Investigating spectrum of biological activity of 4- and 5-chloro-2-hydroxy-N-[2-(arylamino)-1-alkyl-2-oxoethyl]benzamides.

In this study, a series of twenty-two 5-chloro-2-hydroxy-N-[2-(arylamino)-1-alkyl-2-oxoethyl]benzamides and ten 4-chloro-2-hydroxy-N-[2-(arylamino)-1-alkyl-2-oxoethyl]benzamides is described. The compounds were analyzed using RP-HPLC to determine lipophilicity. Primary in vitro screening of the synthesized compounds was performed against mycobacterial, bacterial and fungal strains. They were also evaluated for their activity related to the inhibition of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. The compounds showed biological activity comparable with or higher than the standards isoniazid, fluconazole, penicillin G or ciprofloxacin. For all the compounds, the relationships between the lipophilicity and the chemical structure of the studied compounds as well as their structure-activity relationships are discussed.

Synthesis and antimycobacterial properties of N-substituted 6-amino-5-cyanopyrazine-2-carboxamides.

A series of fifteen new compounds related to pyrazinamide (PZA) were synthesized, characterized with analytical data and screened for antimycobacterial, antifungal and antibacterial activity. The series consists of 6-chloro-5-cyanopyrazine-2-carboxamide and N-substituted 6-amino-5-cyanopyrazine-2-carboxamides, derived from the previous by nucleophilic substitution with various non-aromatic amines (alkylamines, cycloalkylamines, heterocyclic amines). Some of the compounds exerted antimycobacterial activity against Mycobacterium tuberculosis equal to pyrazinamide (12.5-25 µg/mL). More importantly, 6-chloro-5-cyanopyrazine-2-carboxamide and 5-cyano-6-(heptylamino)pyrazine-2-carboxamide were active against Mycobacterium kansasii and Mycobacterium avium, which are unsusceptible to PZA. Basic structure-activity relationships are presented. Only weak antifungal and no antibacterial activity was detected.

Synthesis, antimycobacterial, antifungal and photosynthesis-inhibiting activity of chlorinated N-phenylpyrazine-2-carboxamides.

A series of sixteen pyrazinamide analogues with the -CONH- linker connecting the pyrazine and benzene rings was synthesized by the condensation of chlorides of substituted pyrazinecarboxylic acids with ring-substituted (chlorine) anilines. The prepared compounds were characterized and evaluated for their antimycobacterial and antifungal activity, and for their ability to inhibit photosynthetic electron transport (PET). 6-Chloro-N-(4-chlorophenyl)pyrazine-2-carboxamide manifested the highest activity against Mycobacterium tuberculosis strain H37Rv (65% inhibition at 6.25 µg/mL). The highest antifungal effect against Trichophyton mentagrophytes, the most susceptible fungal strain tested, was found for 6-chloro-5-tert-butyl-N-(3,4-dichlorophenyl)pyrazine-2-carboxamide (MIC=62.5 µmol/L). 6-chloro-5-tert-butyl-N-(4-chlorophenyl)pyrazine-2-carboxamide showed the highest PET inhibition in spinach chloroplasts (Spinacia oleracea L.) chloroplasts (IC50=43.0 µmol/L). For all the compounds, the relationships between the lipophilicity and the chemical structure of the studied compounds as well as their structure-activity relationships are discussed.