Antibacterial and antibiofilm agents in the group of xanthone derivatives with piperazine moiety active against drug-resistant Helicobacter pylori strains.

Helicobacter pylori (H. pylori) cause chronic inflammation of the gastric mucosa which can lead to epithelial atrophy and metaplasia resulting in peptic ulcer disease and gastric cancer. The increasing resistance of H. pylori to antibiotics and chemotherapeutics used to treat the infection is a serious problem. However, it has been confirmed that the introduction of effective anti-H. pylori therapy can prevent the progression to cancerous changes. This problem calls for the search for new and effective therapies. Xanthones are a group of compounds with extensive biological activities, including antibacterial activity, also against H. pylori. Addressing this issue, the aim of the study was to evaluate the potential of a group of 13 xanthone derivatives against susceptible and resistant H. pylori strains. Moreover, our objective was to conduct tests aimed at determining their ability to inhibit biofilm formation. The antimicrobial evaluation revealed that benzylpiperazine coupled at the C-2 position to xanthone (compounds C11 and C12) had good selective bacteriostatic activity against reference and clinical H. pylori strains (MBC/MIC ratio >4) but with no activity against other bacteria such as Staphylococcus aureus, Escherichia coli, and Lactobacillus paracasei. Analysis of the activity of compounds C11 and C12 against the biofilm formed by H. pylori strain ATCC 700684, and the clinical strain showed that these compounds caused a significant reduction in the amount of biofilm produced (5-20×). Moreover, cell viability analysis confirmed a 3-4× reduction in the viability of cells forming biofilm after treatment with C11 and C12. Finally,both compounds did not impair human fibroblast viability at tested concentrations and were not mutagenic in the Ames test. Therefore, they could be promising leads as antibacterial candidates for multidrug-resistant strains of H. pylori.

Optimization of Helicobacter pylori Biofilm Formation in In Vitro Conditions Mimicking Stomach.

Helicobacter pylori is one of the most common bacterial pathogens worldwide and the main etiological agent of numerous gastric diseases. The frequency of multidrug resistance of H. pylori is growing and the leading factor related to this phenomenon is its ability to form biofilm. Therefore, the establishment of a proper model to study this structure is of critical need. In response to this, the aim of this original article is to validate conditions of the optimal biofilm development of H. pylori in monoculture and co-culture with a gastric cell line in media simulating human fluids. Using a set of culture-based and microscopic techniques, we proved that simulated transcellular fluid and simulated gastric fluid, when applied in appropriate concentrations, stimulate autoaggregation and biofilm formation of H. pylori. Additionally, using a co-culture system on semi-permeable membranes in media imitating the stomach environment, we were able to obtain a monolayer of a gastric cell line with H. pylori biofilm on its surface. We believe that the current model for H. pylori biofilm formation in monoculture and co-culture with gastric cells in media containing host-mimicking fluids will constitute a platform for the intensification of research on H. pylori biofilms in in vitro conditions that simulate the human body.

Vitamin B6 inhibits activity of Helicobacter pylori adenylosuccinate synthetase and growth of reference and clinical, antibiotic-resistant H. pylori strains.

The current therapies against gastric pathogen Helicobacter pylori are ineffective in over 20% of patients. Enzymes belonging to the purine salvage pathway are considered as novel drug targets in this pathogen. Therefore, the main aim of the current study was to determine the antibacterial activity of pyridoxal 5'-phosphate (PLP), an active form of vitamin B6, against reference and clinical strains of H. pylori. Using a broad set of microbiological, physicochemical (UV absorption, LC-MS, X-ray analysis) and in silico experiments, we were able to prove that PLP inhibits adenylosuccinate synthetase (AdSS) from H. pylori by the competition with GTP (IC50eq ∼30 nM). This behaviour was attributed to formation of a Schiff base with a lysine residue (a covalent bond with Lys322 in the GTP binding site of AdSS) and was potentiated by the presence of vitamin C. This antibacterial activity of PLP gives hope for its future use against H. pylori.

Latest Update on Outer Membrane Vesicles and Their Role in Horizontal Gene Transfer: A Mini-Review.

Outer membrane vesicles (OMVs) are spherical, lipid-based nano-structures, which are released by Gram-negative bacteria in both in vitro and in vivo conditions. The size and composition of OMVs depend on not only the producer bacterial species but also cells belonging to the same strain. The mechanism of vesicles' biogenesis has a key role in determining their cargo and the pattern of macromolecules exposed on their surface. Thus, the content of proteins, lipids, nucleic acids, and other biomolecules defines the properties of OMVs and their beneficial or harmful effects on human health. Many studies have provided evidence that OMVs can be involved in a plethora of biological processes, including cell-to-cell communication and bacteria-host interactions. Moreover, there is a growing body of literature supporting their role in horizontal gene transfer (HGT). During this process, OMVs can facilitate the spreading of genes involved in metabolic pathways, virulence, and antibiotic resistance, guaranteeing bacterial proliferation and survival. For this reason, a deeper understanding of this new mechanism of genetic transfer could improve the development of more efficient strategies to counteract infections sustained by Gram-negative bacteria. In line with this, the main aim of this mini-review is to summarize the latest evidence concerning the involvement of OMVs in HGT.

Ether Derivatives of Naringenin and Their Oximes as Factors Modulating Bacterial Adhesion.

Because of the close connection between adhesion and many vital cellular functions, the search for new compounds modulating the adhesion of bacteria belonging to the intestinal microbiota is a great challenge and a clinical need. Based on our previous studies, we discovered that O-lkyl naringenin derivatives and their oximes exhibit antimicrobial activity against antibiotic-resistant pathogens. The current study was aimed at determining the modulatory effect of these compounds on the adhesion of selected representatives of the intestinal microbiota: Escherichia coli, a commensal representative of the intestinal microbiota, and Enterococcus faecalis, a bacterium that naturally colonizes the intestines but has disease-promoting potential. To better reflect the variety of real-life scenarios, we performed these studies using two different intestinal cell lines: the physiologically functioning ("healthy") 3T3-L1 cell line and the disease-mimicking, cancerous HT-29 line. The study was performed in vitro under static and microfluidic conditions generated by the Bioflux system. We detected the modulatory effect of the tested O-alkyl naringenin derivatives on bacterial adhesion, which was dependent on the cell line studied and was more significant for E. coli than for E. faecalis. In addition, it was noticed that this activity was affected by the concentration of the tested compound and its structure (length of the carbon chain). In summary, O-alkyl naringenin derivatives and their oximes possess a promising modulatory effect on the adhesion of selected representatives of the intestinal microbiota.

Inhibitory activity of catecholic phosphonic and phosphinic acids against Helicobacter pylori ureolysis.

Catechols have been reported to be potent covalent inhibitors of ureases, and they exhibit activity by modifying cysteine residues at the entrance to enzymatic active sites. Following these principles, we designed and synthesized novel catecholic derivatives that contained carboxylate and phosphonic/phosphinic functionalities and assumed expanded specific interactions. When studying the chemical stability of the molecules, we found that their intrinsic acidity catalyzes spontaneous esterification/hydrolysis reactions in methanol or water solutions, respectively. Regarding biological activity, the most promising compound, 2-(3,4-dihydroxyphenyl)-3-phosphonopropionic acid (15), exhibited significant anti-urease potential (Ki = 2.36 µM, Sporosarcinia pasteurii urease), which was reflected in the antiureolytic effect in live Helicobacter pylori cells at a submicromolar concentration (IC50 = 0.75 µM). As illustrated by molecular modeling, this compound was bound in the active site of urease through a set of concerted electrostatic and hydrogen bond interactions. The antiureolytic activity of catecholic phosphonic acids could be specific because these compounds were chemically inert and not cytotoxic to eukaryotic cells.

Comparison of antibiofilm activity of low-concentrated hypochlorites vs polyhexanide-containing antiseptic.

Chronic wound infection is highly associated with morbidity and endangers the patient's life. Therefore, wound care products must have a potent antimicrobial and biofilm-eradicating effect. In this work, the antimicrobial/antibiofilm activity of two low-concentrated chlorine-based and releasing solutions was investigated on a total of 78 strains of methicillin-resistant Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans, using the cohesive spectrum of in vitro settings, including microtiter plate models, biofilm-oriented antiseptic test, cellulose-based biofilm model, biofilm bioreactors and Bioflux model. The antiseptic containing polyhexamethylene biguanide was used in the character of usability control of performed tests. The results obtained by static biofilm models indicate that low-concentrated chlorine-based and releasing solutions display none to moderate antibiofilm activity, while data obtained by means of the Bioflux model, providing flow conditions, indicate the moderate antibiofilm activity of substances compared with the polyhexanide antiseptic. Considering in vitro data presented in this manuscript, the earlier reported favorable clinical results of low-concentrated hypochlorites should be considered rather an effect of their rinsing activity combined with low cytotoxicity but not the antimicrobial effect per se. For the treatment of heavily biofilm-infected wounds, polyhexanide should be considered the agent of choice because of its higher efficacy against pathogenic biofilms.

Extracellular Vesicles of Probiotics: Shedding Light on the Biological Activity and Future Applications.

For many decades, the proper functioning of the human body has become a leading scientific topic. In the course of numerous experiments, a striking impact of probiotics on the human body has been documented, including maintaining the physiological balance of endogenous microorganisms, regulating the functioning of the immune system, enhancing the digestive properties of the host, and preventing or alleviating the course of many diseases. Recent research, especially from the last decade, shows that this health-benefiting activity of probiotics is largely conditioned by the production of extracellular vesicles. Although the importance of extracellular vesicles in the virulence of many live-threatening pathogens is widely described in the literature, much less is known with respect to the health-promoting effect of extracellular vesicles secreted by non-pathogenic microorganisms, including probiotics. Based on this, in the current review article, we decided to collect the latest literature data on the health-inducing properties of extracellular vesicles secreted by probiotics. The characteristics of probiotics' extracellular vesicles will be extended by the description of their physicochemical properties and the proteome in connection with the biological activities exhibited by these structures.

Activity of Novel Ultrashort Cyclic Lipopeptides against Biofilm of Candida albicans Isolated from VVC in the Ex Vivo Animal Vaginal Model and BioFlux Biofilm Model-A Pilot Study.

In recent years, clinicians and doctors have become increasingly interested in fungal infections, including those affecting the mucous membranes. Vulvovaginal candidiasis (VVC) is no exception. The etiology of this infection remains unexplained to this day, as well as the role and significance of asymptomatic vaginal Candida colonization. There are also indications that in the case of VVC, standard methods of determining drug susceptibility to antifungal drugs may not have a real impact on their clinical effectiveness-which would explain, among other things, treatment failures and relapse rates. The aim of the study was to verify the promising results obtained previously in vitro using standard methods, in a newly developed ex vivo model, using tissue fragments of the mouse vagina. The main goal of the study was to determine whether the selected ultrashort cyclic lipopeptides (USCLs) and their combinations with fluconazole at specific concentrations are equally effective against Candida forming a biofilm directly on the surface of the vaginal epithelium. In addition, the verification was also performed with the use of another model for the study of microorganisms (biofilms) in vitro-the BioFlux system, under microfluidic conditions. The obtained results indicate the ineffectiveness of the tested substances ex vivo at concentrations eradicating biofilm in vitro. Nevertheless, the relatively most favorable and promising results were still obtained in the case of combination therapy-a combination of low concentrations of lipopeptides (mainly linear analogs) with mycostatic fluconazole. Additionally, using BioFlux, it was not possible to confirm the previously obtained results. However, an inhibiting effect of the tested lipopeptides on the development of biofilm under microfluidic conditions was demonstrated. There is an incompatibility between the classic in vitro methods, the newer BioFlux method of biofilm testing, offering many advantages postulated elsewhere, and the ex vivo method. This incompatibility is another argument for the need, on the one hand, to intensify research on the pathomechanism of VVC, and, on the other hand, to verify and maybe modify the standard methods used in the determination of Candida susceptibility.