Vol 58, No 6 (2024)

Antibiotic and, more broadly, antimicrobial resistance is a naturally occurring biological phenomenon and a major public health problem. The first cases of mass emergence of drug-resistant strains of bacteria were observed in the mid-20th century; Since then, cases of resistance have been reported worldwide, and in the last two decades, multiple bacterial resistance has been increasingly reported. Factors contributing to the development of bacterial resistance include the overuse of antibacterial agents in humans or livestock and the release of antibacterial agents into the environment. Unfortunately, the development of new effective antibiotics is declining, which requires strengthening this work, as well as the search for alternative methods of treating infectious diseases.

Despite nearly a century of therapy for gonococcal infection with a variety of antimicrobials, more than 80 million cases of this disease are reported annually worldwide. The gonorrhea pathogen, Neisseria gonorrhoeae, exhibits an exceptional ability to develop resistance to antibiotics due to its high genetic flexibility. As an obligate pathogen, the gonococcus has evolved mechanisms to evade host defenses, engaging with the innate and adaptive immune responses in both men and women. The bacterium can establish residence within epithelial cells, macrophages, and neutrophils. Through genetic variability and horizontal gene transfer, strains resistant to each of the drugs used in gonorrhea therapy have emerged. The type IV secretion system plays a critical role in horizontal gene transfer, driving the development of antimicrobial resistance. This review explores the pathogenesis and immune evasion mechanisms, antibiotic resistance formation, genetic variability, laboratory analysis methods for the pathogen, and emerging trends in the diagnosis and treatment of gonococcal infections.

The review considers issues related to the spread of antibiotic resistance genes in environmental microbial communities. “Hotspots” of adaptive evolution, accumulation and spread of antibiotic-resistant bacteria and genetic material of antibiotic resistance are highlighted. Such “hotspots” include anthropogenic ecosystems, such as municipal wastewater treatment plants, municipal solid waste landfills, livestock enterprises, and agrocenoses. The influence of various types of pollutants and biotic factors on enhancement of mutagenesis and horizontal transfer of antibiotic resistance genes is considered. The role of mobile genetic elements in mobilization and accelerated spread of resistance determinants is shown. Special attention is paid to the role of oxidative stress and stress regulons, which are activated for realization and control of molecular genetic mechanisms of adaptive evolution of bacteria and horizontal distribution of genetic material in bacterial populations. Oxidative stress is identified as one of the main activators of genome destabilization and adaptive evolution of bacteria.

The acute and subchronic toxicity of the pharmacological pair based on encapsulated Citrobacter freundii C115H methionine γ-lyase enzyme/prodrug (methiin) was studied in female ICR mice. The drug showed a weak/moderate dose-dependent hepatotoxic effect. Most of the identified changes in liver morphology were insignificant or mild deviations from the norm. Long-term use of a single therapeutic dose per mouse of 1.5 U C. freundii C115H methionine γ-lyase @ (PEG−P(Asp)₇₀/PLL₇₀)-PICsome / 2 mg methiin led to a slight decrease in the weight of animals without obvious signs of intoxication. A quarter of the animals in this group had no deviations from the norm in liver morphology. No nephrotoxic effect in all study groups was found.

Cystathionine γ-lyase (CSE) is a key enzyme for the H₂S generation in such pathogenic bacteria as Staphylococcus aureus, Pseudomonas aeruginosa, etc. Suppression of CSE activity significantly increases the sensitivity of bacteria to the action of antibiotics. Here, we present a method for the synthesis of a novel indole-based CSE inhibitor, 3-amino-5-[(6-bromo-1H-indol-1-yl)methyl]thiophene, named MNS1. The synthesis of this compound is based on the modification of substituted thiophene as the main structural fragment, which is involved the alkylation of 6-bromoindole at the final stages. The dissociation constant of the MNS1 complex with bacterial CSE (from S. aureus, SaCSE) was 0.5 μM, which was an order of magnitude lower than that for human CSE (hCSE). The MNS1 compound was shown to effectively enhance the antibacterial effect of gentamicin against Bacillus subtilis, allowing it to be used as an antibiotic potentiator, to inhibit the growth of CSE-expressing bacterial cells.

Bacillus cereus sensu lato s.l. comprises genetically, morphologically and physiologically similar gram-positive spore-forming bacterial species with high pathogenic potential, such as B. anthracis, B. cereus and B. thuringiensis. Toxin-producing strains of B. cereus s.l. pose a major threat to human health. The high degree of similarity between these species makes it very difficult to identify them and to take adequate measures to treat the diseases they cause. Previously, we characterized a clinical isolate CCGC 19/16 belonging to B. cereus s.l. that exhibited features of both B. cereus and B. cytotoxicus. In the present work, CCGC 19/16 was identified as B. cytotoxicus using multilocus sequence typing (MLST) and mass spectrometric analysis. It was also shown that, unlike other representatives of the B. cytotoxicus species, strain CCGC 19/16 is not thermotolerant. Unlike B. cereus, strain CCGC 19/16 is sensitive to most antibiotics and shows increased motility. Like B. cereus strain CCGC 19/16 forms β-hemolysis zones in blood agar. In addition, it has been shown that prolonged storage of samples prior to analysis can lead to misidentification of the isolate. Our results indicate that “rapid methods” of analysis using single genes have insufficient resolving power in the identification of B. cereus s.l. species. The combination of MLST analysis with MALDI-TOF MS provides sufficient resolution.

Prescribing adequate empiric therapy is not possible without local monitoring of the spread of antibiotic-resistant bacteria in each hospital. The aim of the study was to compare the frequency of β-lactamase-producing Enterobacterales in patients of therapeutic and surgical units. Antibiotic susceptibility was evaluated by disk-diffusion method. The production of extended-spectrum β-lactamases (ESBL) was detected by the double disk method, and carbapenemases were determined by a modified method of carbapenem inactivation. Carbapenemase genes and the expression were quantified by real-time PCR and immunochromatography assay. Among the isolated Enterobacterales, more than one third produced ESBL in both the therapeutic (35.51%) and surgical (39.85%) units. The proportion of carbapenemase producers was comparable in both groups, and amounted to 8.41% and 9.77%, respectively. Metallo-β-lactamases were predominant in surgical units, whereas serine lactamases were predominated in therapeutic ones. Among the out-of-hospital Enterobacterales, β-lactamase producers were isolated less frequently than among the nosocomial ones in both the therapeutic (31.48% and 56.6%) and surgical (45.45% and 51%) wards, but without statistically significant differences. During the three years of the study, we did not detect an increase in the proportion of β-lactamase producers in surgical and therapeutic units, but local monitoring should certainly be continued in order to develop a local strategy for the adequate use of antibacterial drugs.

Currently, empirical therapy regimens are often used in the treatment of infectious diseases that are not based on data on pathogen resistance. One of the main reasons for the unjustified prescription of antibacterial drugs is the lack of rapid and at the same time universal methods of determining antibiotic resistance of the pathogen. The most widely used culture techniques, such as the microdilution method, require a long time to generate the necessary number of bacterial cells. Less time-consuming methods of resistance assessment (genomic or proteomic) are based on the determination of specific markers (resistance genes, overexpression of certain proteins, etc.); in this case, the specific protocol is most often applicable to a narrow number of both microorganism strains and antibiotics. Previously, we have demonstrated the possibility of using Raman spectroscopy (RS) technology for quantitative determination of the product of bacterial cell activity in the MTT аssay, formazan, directly in the cell suspension. The absence of the formazan isolation step simplifies the assay and increases its accuracy. The analysis time did not exceed 2 h while maintaining the versatility of the MTT аssay itself. Limitations of the developed protocol for RS detection of MTT аssay results include a high sensitivity threshold of 10⁷ CFU/mL for bacterial cell concentration, so a preliminary stage of cultivation is necessary for samples with low bacterial content. Here, we propose a method to increase the sensitivity of formazan determination by utilizing the effect of surface-enhanced Raman scattering (SERS) on gold nanoparticles. As a result of the study, the optimal conditions for SERS analysis of formazan in both solution and suspension of Escherichia coli cells are selected. Formazan signal amplification due to the use of SERS on gold nanoparticles instead of RS allowed us to reduce the sensitivity threshold for the number of bacterial cells in the sample at least 30 times, up to 3 × 10⁵ CFU/mL. This sensitivity is not the limit of the SERS technology capabilities because of the introduction of other types of nanoparticles (more optimal in shape, size, concentration, etc.) into the experiment will allow to achieve even higher signal amplification.

The extracellular matrix (ECM) provides structural support and regulates cellular activity. Its disruption during metabolic pathologies or aging can lead to disease development. Developing ECM protectors is crucial for the etiological prevention and treatment of pathologies associated with ECM alterations. Key mechanisms of pathological changes in the ECM include non-enzymatic reactions such as glycation and glycoxidation. The potential of agents as ECM protectors can be assessed by their ability to inhibit these processes. In this study, compounds based on heterocyclic scaffolds, including partially hydrogenated isoindole fragments, were investigated for their ability to slow down the formation of advanced glycation end-products (AGEs). The study employed a combination of in silico and in vitro approaches. In the in silico study, the energies of the frontier molecular orbitals of the compounds were determined using the ab initio method with the 6-311G(d,p) basis set. Their antiglycation activity was then investigated in the glycation reaction of bovine serum albumin (BSA) with glucose, using albumin as a model protein. Pyridoxamine served as a reference compound. The antiglycation activity of the compounds was evaluated spectrofluorometrically by measuring the fluorescent products at excitation/emission wavelengths of 440/520 nm, which are not typically used for assessing antiglycation properties. At these wavelengths, glycation and oxidation products in human skin can be detected, which correlate with chronological age, unlike some other glycation products. Experimentally, it was found that the energies of the frontier molecular orbitals of the compounds can serve as predictors of their ability to slow down the formation of fluorescent products detected at 440/520 nm. Inhibiting the formation of such products may be significant for the treatment and prevention of diseases, including metabolic, fibrotic, or age-associated conditions. It was also established that at a concentration of 100 µM, the antiglycation properties are most pronounced in the series of hydrogenated 3a,6-epoxyisoindole-7-carboxylic acids (compounds of type XIII) and cyclopenta[b]furo[2,3-c]pyrrole-3-carboxylic acids (structures of type XIX).

Proteins of the TRIM family are involved in both innate immunity and the nervous system processes and may play an important role in the development of neurodegenerative diseases. In this work, we analyzed the expression of 35 genes of the TRIM family in neural progenitors (NPs), terminally differentiated neurons (TDNs) and glial derivatives (NGs) obtained from induced pluripotent stem cells (iPSCs) of healthy donors (HD) and patients with Parkinson’s disease (PD), in the absence of inflammatory stimuli and upon the induction of a nonspecific inflammatory response under the influence of TNFα. In NPs and TDNs of PD patients, compared with HD cells, differences in expression were observed for only a small number of TRIM genes. Under the influence of TNFα in TDNs, the expression of individual TRIM genes was activated, which was more significant in the cells of patients with PD compared to cells of HDs. In NGs of PD patients, the expression of many TRIM genes was initially reduced compared to HD cells and remained low or further decreased after exposure to TNFα. The data obtained demonstrate differences in the network of the TRIM family members in PD neurons and glia compared to control, and also show the multidirectional influence of the inflammatory stimulus on the expression of a number of TRIM genes in these types of cells. Considering the important role of many TRIM genes in the functioning of the innate immune system, it can be assumed that, in PD, more significant disturbances in the functioning of genes of this family occur in glia compared to neurons.