A novel antivirulent compound fluorothiazinone inhibits Klebsiella pneumoniae biofilm in vitro and suppresses model pneumonia.

The problematic treatment of infections caused by multiple-resistant Klebsiella, especially in ICU, is the leading cause of prolonged hospitalization and high mortality rates. The use of antibiotics for the prevention of infections is considered unreasonable as it may contribute to the selection of resistant bacteria. In this regard, the development of drugs that will be effective in preventing infection during various invasive procedures is extremely necessary. We have shown that the developed innovative antibacterial compound fluorothiazinone (FT) that suppresses the formation of biofilms is effective in the prevention of a model pneumonia caused by a multi-resistant clinical K. pneumoniae isolate. Prophylactic use followed by treatment with FT in mice with acute pneumonia modulates the local innate immune response without suppressing protective properties in the early stages of infection, while contributing to a decrease in the bacterial load in the organs and preventing lethal pathological changes in the lungs at later stages of K. pneumoniae infection. Further development of such antivirulence drugs and their use will reduce morbidity and mortality in nosocomial infections, as well as reduce the number of antibiotics used.

Fluorothiazinon inhibits the virulence factors of uropathogenic Escherichia coli involved in the development of urinary tract infection.

Uropathogenic Escherichia coli (UPEC) is the most common pathogenic bacterium associated with urinary tract infection. Due to the development of antibiotic resistance and MDR, UPEC infection has become a serious problem in the last decade. In order to combat resistance, it is necessary to develop innovative antimicrobial agents that act by different mechanisms than conventional antibiotics. Among the new therapeutic strategies, suppression of pathogen virulence has become a promising alternative, since it fundamentally reduces selective pressure and the development of resistance. In our study, we showed that the compound Fluorothiazinon suppressed UPEC's ability to form biofilms and to move using the flagellum, as well as to penetrate into cells. Prophylactic use with subsequent treatment of FT in rodent models led to an improvement in survival and significantly reduced the bacterial load in the organs of the urinary system, thereby inhibiting the development of ascending infection and preventing the development of pathological changes in prostate tissues. These results suggest that FT affects several UPEC virulence factors at once and if similar results can be found in clinical trials it can potentially be used as a new drug against UPEC.

[Single-domain adapted antibodies against Chlamydia trachomatis, preserving the development of chlamidic infection in vitro]. / Odnodomennye adaptirovannye antitela protiv Chlamydia trachomatis, podavliaiushchie razvitie khlamidiinoi infektsii v usloviiakh in vitro.

The technology for the generating of single-domain recombinant monoclonal antibodies (nanoantibodies) based on the immunization of a camel, cloning of induced sequences encoding single-domain antigen-recognizing fragments of non-canonical camel antibodies, as well as functional selection of clones of nanoantibodies by the phage display method, was used to obtain new effective tools for more efficient diagnostics of Chlamydia infection and to develop new approaches for effective therapy. Two promising nanoantibodies were obtained. They showed effective binding to extracellular and intracellular forms of C. trachomatis, and also had activity that inhibited the development of chlamydial infection in vitro.

[Cytokine gene polymorphism in type 2 diabetes mellitus in Russian women from eastern Europe].

AIM: To study the distribution of genotypes in the cytokine genes and their combinations with immunoregulatory activity in patients with type 2 diabetes mellitus (T2DM) and in healthy women. SUBJECTS AND METHODS: 586 Europeoid women from the eastern regions of Russia, including 374 healthy women aged 23-68 years and 212 women aged 28-69 years with T2DM complicated and uncomplicated by osteoporosis, were examined. Seven polymorphisms located in the promoter regions of the interleukin (IL) gene: TNF-alpha at positions C-863A, G-308A, G-238A, IL1B T-31C, IL6 G-174C, IL10 C-592A, VEGFA C-2578A were investigated. Restriction analysis of amplification products was applied. RESULTS: There were high associations of the predisposition and resistance to the development of T2DM with a number of polylocus cytokine genotype combinations having pro- and anti-inflammatory, angiogenic, and immunoregulatory activities. The association of the cytokine genes with T2DM was found to be mediated in nature through a relationship of the genotypes to the high or low production of regulatory cytokines and to different factors of regulation of lipid and carbohydrate metabolisms, inflammation, and bone remodeling. CONCLUSION: The high odds ratio and high specificity of the detected genetic combinations allow one to hope that they will be clinically used as predictors.

[Influence of Chlamydia trachomatis type III secretion system on regulation of cytokine response].

AIM: Develop in vitro model for studying production of cytokines by monocyte cells infected with Chlamydia trachomatis mediated by type III secretion system (TTSS). MATERIALS AND METHODS: Strain C. trachomatis L2/434/Bu was used in the experiments, culture of human monocytes U-937 was infected by this strain. Level of inflammatory cytokines was measured on flow analyzer Bio-Plex 200 (Bio-Rad Laboratories). Low molecular compound LHC-342 which belongs to the class of heterocyclic compounds was used as TTSS inhibitor. RESULTS: 24 hours after the infection with C. trachomatis culture 8 analyzed cytokines are induced in U-937 cells (IL-1beta, IL-4, IL-6, IL-8, IL-10, GM-CSF, IFN-gamma, TNFalpha). The most pronounced increase was observed for IL-8, GM-CSF and IFN-gamma. Introduction of TTSS inhibitor into the culture of infected cells suppressed chlamydia growth, but addition of FeSO4 restored the growth of chlamydiae. And activity associated with translocation of effector TTSS protein IncA to inclusion membrane was suppressed. Under the conditions of the obtained model of TTSS inhibition during intracellular development of C. trachomatis a significant decrease of 2 pro-inflammatory cytokines--IL-6 and IL-1beta--was observed. CONCLUSION: Cytokine response plays a key role in the protective immune response in chlamydia infection but at the same time induces immunopathologic conditions. The data obtained give reasons to assume role of C. trachomatis TTSS in the induction of this component of immune response that requires further detailed studies.

[Characteristic of early mucosal immune response in mice during intravaginal infection caused by Chlamydia muridarum].

AIM: Comparison of features of recruitment to infection focus of cells mediating early immune reactions in intravaginally infected mice that had previously received or not received covinan (progesterone analogue). MATERIALS AND METHODS: A/Sn and BALB/c line mice were used in the study. C. muridarum strain Nigg infection was carried out intravaginally or intraperitoneally. For synchronization of sexual cycle a group of mice received subcutaneously a synthetic analogue of progesterone--proligeston (covinan) at a single dose of 33 mg/kg. Acute urogenital infection was evaluated by culture method. Quantitative determination of C. muridarum DNA (including study of persistence) was carried out by real time PCR. Subpopulation structure of cell population of peritoneal and vaginal lavage was evaluated by flow cytofluorometry. RESULTS: Intravaginal infection of mice that had not received covinan resulted in a pronounced recruitment of cells into vaginal cavity at 24 hours after the infection. Influx of neutrophils, dendritic cells and T-lymphocytes was especially pronounced. Prior administration of covinan practically nullified cell recruitment to infection focus though partial preservation of subpopulations of activated dendritic cells and CD8+ T-cells was observed. CONCLUSION: In mice with artificially induced by progesterone sensitivity to chlamydias the ability of recruitment to the infection focus of cells that mediate early immune reactions is reduced, that gives evidence on the importance of these reactions for infection outcome.

[Development of vaccine preparation for urogenital Chlamydiosis prophylaxis].

Chlamydia trachomatis is one of the most widespread bacterial pathogens in the world that is transmitted sexually. Thereby a development of vaccine preparation for therapy and prophylaxis of infections caused by C. trachomatis is an actual topic of scientific research across the entire world. These vaccines could be used for both prophylaxis and therapy of already established chlamydia infection and, respectively, reduce the risk of chronic condition and prevent the spread of pathogen in the population. Features of chlamydia biology that are associated with obligate intracellular parasitism and immunopathologic state induced by this agent significantly complicate developments in this field. Currently a vaccine preparation that has reached clinical trials does not exist.

[The diurnal blood pressure and the EchoCG parameters of the left ventricle in patients with diabetic nephropathy]. / Sutochnyi profil' arterial'nogo davleniia i EkhoKG-parametry levogo zheludochka u bol'nykh diabeticheskoi nefropatiei.

The purpose of the study was to examine the geometry of the left ventricle (LV) in patients with type 1 diabetes mellitus (DM) in relation to the 24-hour profile of blood pressure (BP) and urinary albumin excretion. The study covered 60 patients with type 1 DM and normal creatinine clearance, including 20 patients with normal albuminuria, 23 with microalbuminuria, and 17 with proteinuria. 24-hour BP monitoring was performed oscillometrically; myocardial structural parameters were studied by echoCG. LV hypertrophy (LVH) was found in 1 patient with normal albuminuria and in 8 with proteinuria (5, 30.4, and 47%, respectively; chi2 = 9.3; p = 0.09). The frequency of concentric and eccentric types of LVH was equal. In patients with a lower nocturnal BP decrease, the LV myocardial mass index (LVMMI) and relative posterior wall thickness (RPWT) were higher than those in other patients (LVMMI, 120.8 +/- 24.6 and 95.0 +/- 23.1 g/m2, respectively; p < 0.001; RPWT, 0.35 +/- 0.06 and 0.31 +/- 0.06, respectively, p = 0.013). Multifactorial stepwise regression analysis has indicated that age, male sex, and proneinuria directly affected LVMMI (R2 = 0.70; p < 0.001). Diastolic BP, autonomic neuropathy, and hemoglobin levels were found to be independent predictors of RPWT (R2 = 0.70; p < 0.009). The findings suggest that there is a close relationship between diabetic neuropathy and LV remodeling in patients with type 1 DM. This relationship may be operative via factors, such as arterial hypertension, altered diurnal BP profile, autonomic neuropathy, and anemia.