Biology subjects

Gintsburg, A. L.

Publications and source records attributed to Gintsburg, A. L..

10 recordsLinked to original sources

bioRxiv2026

Susceptibility of Klebsiella pneumoniae Clinical Isolates in Biofilms to Antibiotics and Assessment of Secondary Drug Effects

Biofilms pose a significant challenge to antimicrobial therapy. Bacteria in biofilms differ from planktonic counterpart in their altered metabolism, collective behavior, protective role of extracellular matrix and diversified microbial subpopulations. These attributions significantly influence bioavailability and activity of antibiotics. The presence of bacterial aggregates during acute infections expands the problem to many other conditions previously not discussed in the biofilm context. Klebsiella pneumoniae is a leading cause of life-threatening hospital-acquired infections and is included in the WHO Bacterial Priority Pathogens List due to increasing antimicrobial resistance. The combination of antimicrobial resistance and the ability to form biofilms severely limits the efficacy of antibiotic treatments. In this study, we investigated the in vitro susceptibility of mature biofilms to 13 antimicrobials of K. pneumoniae clinical isolates from a single hospital. The resistance profiles of the local clinical isolates were consistent with the global epidemiology of K. pneumoniae. Minimal biofilm eradication concentrations (MBEC) for mature biofilms were defined with two assays (biomass and metabolic activity measurements) and brought into relation with susceptibility breakpoints and plasma (Cmax). Colistin sulfate, tigecycline, cephalosporins and combination of imipenem with cilastatin were the most potent biomass eradicators, while suppression of metabolic activity was barely reachable. Moreover, we observed a notable increase in metabolic activity upon exposure to sub-MBEC concentrations of antibiotics. Finally, our data broach a subject of antibiotic prioritization with respect to biofilm tolerance. IMPORTANCEThis study addresses the critical gap between standard antibiotic susceptibility testing and the tolerance of biofilm and microbial aggregates during infections caused by K. pneumoniae. By systematically evaluating mature biofilms from a significant number of clinical isolates, we demonstrate that colistin and tigecycline show potent activity against both biofilm biomass and metabolic activity, whereas cephalosporins primarily reduce biomass without effectively suppressing bacterial metabolism, and other drugs have only weak effects on biofilms at clinically achievable concentrations. Furthermore, the alarming observation that sub-inhibitory biofilm eradication concentration (sub-MBEC) of antibiotic can paradoxically increase the metabolic activity of biofilms highlights a potential risk factor for therapy failure and resistance development. Our findings contribute to the necessary evidence base for prioritizing existing antibiotics in the limited armamentarium against biofilm-forming K. pneumoniae.

microbiology↗

bioRxiv2025

Circular mRNA against Clean Cap linear mRNA vectors comprehensive comparison, expression, active and passive immunization

The mRNA platform has revolutionised vaccine technology by providing a universal, fast and easily scalable production process. There are two main types of mRNA - linear (cap-dependent) and circular (cap-independent), each with its own advantages. Although both vector types are continuously being improved, there has been no comprehensive comparative analysis of the most efficient existing vectors of each type. This paper provides a comparison of expression efficiency, protective and therapeutic activities of circular and linear mRNA vectors. We examined different combinations of linear vectors: containing either N1-methylpseudouridines or uridine and capped with either ARCA (m7G(5')ppp(5')G) or CleanCap (m7G(5')ppp(5')m2G). Circular vectors contained both commonly used IRES of coxsackievirus B3 and new a IRES of human rhinoviruses B6. Preliminary luciferase assay showed that modified linear vectors exhibited significantly higher expression levels both in vitro and in vivo. A similar but less dramatic difference was observed in expression of the target protein. At the same time, immunogenicity and protective activity of linear and circular vectors were equal.

immunology↗

bioRxiv2025

Comparison candidate Tick-borne encephalitis virus vaccines based on mRNA, adenovirus serotype 25 and chimera of YFV vaccine strain

BackgroundDespite the availability of several licensed inactivated vaccines, the development of new vaccines against the Tick-borne encephalitis virus (TBEV) remains an important task, especially in countries endemic to this pathogen. The risk of infection with TBEV increases every year because of increase in the number of ticks, the emergence of tick carriers into new territories, and active human activity in areas of TBEV natural foci. Annual reports of vaccination failures have prompted us to search for approaches to creating a new vaccine. ObjectivesIn our study, we used the same PrM and E antigens to produce three vaccine candidates against tick-borne encephalitis (TBE): 1) a live attenuated YFV 17DD-UN vaccine strain, 2) recombinant adenovirus (rAd) vectors, and 3) mRNA encapsulated in lipid nanoparticles (LNP). MethodsWe generated and assessed the immunogenicity and protective efficacy of three candidate TBEV vaccines based on mRNA, simian adenovirus type 25, and a chimera of the YFV 17DD-UN attenuated strain. ResultsAnalysis of the virus-neutralizing titers in the blood sera of immunized mice revealed a statistically significant difference among the three candidate vaccines. The immunogenicity and protective efficacy of the candidate mRNA-LNP vaccine were found to be higher than those of the other two vaccines. ConclusionsBased on the results of our study, it can be concluded that the mRNA-based platform is more promising for the creation of a vaccine against TBEV.

immunology↗

bioRxiv2024

Seroprevalence and cross-reactivity of neutralizing antibodies to six types of adenoviruses

BackgroundThe use of adenoviral vector vaccines is an effective way to induce an antiviral immune response. Most vaccines used worldwide are based on human adenovirus type 5 (Ad5). Pre-existing natural immunity to Ad5 is a limitation to the frequent use of Ad5-based vectors in gene therapy and vaccination. In the present study, we assessed the seroprevalence and cross-reactivity of neutralizing antibodies to six adenovirus types: Ad1, Ad2, Ad5, Ad19, Ad26, and Ad37. MethodsThe study used immunological and virological research methods. Adenovirus vectors encoding the enhanced green fluorescent protein (EGFP) were developed by the N.F. Gamaleya National Research Center for Epidemiology and Microbiology. For seroprevalence and cross-neutralation study we used a panel of sera from 101 volunteers. ResultsThe conducted correlation analysis showed a strong relationship between seropositivity and the geometric mean titer of neutralizing antibodies to adenovirus. An increase in antibody titers indicates the formation of a humoral immune response and population immunity. We determined that there is no cross-reactivity between NtAbs to Ad1, Ad2, Ad5 and Ad26. ConclusionsBased on the presented data, we assume that Ad1 or Ad2 should be considered the most promising alternative candidates for the development of vaccine vectors.

immunology↗

bioRxiv2024

Major role of S-glycoprotein in providing immunogenicity and protective immunity in mRNA lipid nanoparticle vaccines based on SARS-CoV-2 structural proteins

Recently we have developed an mRNA lipid nanoparticle (mRNA-LNP) platform providing efficient long-term expression of an encoded gene in vivo after both intramuscular and intravenous application. Based on this platform, we have generated mRNA-LNP coding SARS-CoV-2 structural proteins M, N, S from different virus variants and studied their immunogenicity separately or in combinations in vivo. As a result, all candidate vaccine compositions coding S and N proteins induced excellent anti-RBD and N titers of binding antibodies. T cell responses mainly represented specific CD4+ T cell lymphocyte producing IL-2 and TNF-. mRNA-LNP coding M protein did not show high immunogenicity. High neutralizing activity was detected in sera of mice vaccinated with mRNA-LNP coding S protein (alone or in combinations) against closely related strains but was not detectable or significantly lower against an evolutionarily distant variant. Our data showed that the addition of mRNAs encoding S and M antigens to the mRNA-N in the vaccine composition enhanced immunogenicity of mRNA-N inducing more robust immune response to the N protein. Based on our results, we suggested that the S protein plays a key role in enhancement of immune response to the N protein in the mRNA-LNP vaccine.

immunology↗

bioRxiv2023

Trivalent mRNA vaccine-candidate against seasonal flu with cross-specific humoral immune response

ABSTRACTSeasonal influenza remains a serious global health problem, leading to high mortality rates among the elderly and individuals with comorbidities. It also imposes a substantial economic burden through increased absenteeism during periods of active pathogen circulation. Vaccination is generally accepted as the most effective strategy for influenza prevention. As both influenza A and B viruses circulate and cause seasonal epidemics, vaccines need to include multiple antigens derived from different viral subtypes. While current influenza vaccines are effective, they still have limitations, including narrow specificity for certain serological variants, which may result in a mismatch between vaccine antigens and circulating strains. Additionally, the rapid variability of the virus poses challenges in providing extended protection beyond a single season. Therefore, mRNA technology is particularly promising for influenza prevention, as it enables the rapid development of multivalent vaccines and allows for quick updates of their antigenic composition. mRNA vaccines have already proven successful in preventing COVID-19 by eliciting rapid cellular and humoral immune responses. In this study, we present the development of a trivalent mRNA vaccine candidates, evaluate its immunogenicity using the hemagglutination inhibition assay, and assess its efficacy in animals. We demonstrate the higher immunogenicity of the mRNA vaccine candidates compared to the inactivated split influenza vaccine and its enhanced ability to generate a cross-specific humoral immune response. These findings highlight the potential mRNA technology in overcoming current limitations of influenza vaccines and hold promise for ensuring greater efficacy in preventing seasonal influenza outbreaks.

immunology↗

bioRxiv2022

Single-domain antibody delivery using an mRNA platform protects against lethal doses of botulinum neurotoxin A

Single-domain antibodies (sdAbs, VHHs, or nanobodies) are a promising tool for the treatment of both infectious and somatic diseases. Their small size greatly simplifies any genetic engineering manipulations. Such antibodies have the ability to bind hard-to-reach antigenic epitopes through long parts of the variable chains, the third complementarity-determining regions (CDR3s). VHH fusion with the canonical immunoglobulin Fc fragment allows the Fc-fusion single-domain antibodies (VHH-Fc) to significantly increase their neutralizing activity and serum half-life. Previously we have developed and characterized VHH-Fc specific to botulinum neurotoxin A (BoNT/A), that showed a 1000-fold higher protective activity than monomeric form when challenged with five times the lethal dose (5 LD50) of BoNT/A. During the COVID-19 pandemic, mRNA vaccines based on lipid nanoparticles (LNP) as a delivery system have become an important translational technology that has significantly accelerated the clinical introduction of mRNA platforms. We have developed an mRNA platform that provides long-term expression after both intramuscular and intravenous application. The platform has been extensively characterized using firefly luciferase (Fluc) as a reporter. An intramuscular administration of LNP-mRNA encoding VHH-Fc antibody made it possible to achieve its rapid expression in mice and resulted in 100% protection when challenged with up to 100 LD50 of BoNT/A. The presented approach for the delivery of sdAbs using mRNA technology greatly simplifies drug development for antibody therapy and can be used for emergency prophylaxis.

molecular biology↗

bioRxiv2022

Cross-reactive single-domain antibodies to hemagglutinin stem region protect mice from group 1 influenza A virus infection

The continued evolution of influenza viruses reduces the effectiveness of vaccination and antiviral drugs. The identification of novel and universal agents for influenza prophylaxis and treatment is an urgent need. We have previously described two potent single-domain antibodies (VHH), G2.3 and H1.2, which efficiently neutralize H1N1 and H5N2 influenza viruses in vivo. In this study, we modified these VHHs with Fc-fragment to enhance their antiviral activity. Reformatting of G2.3 into bivalent Fc-fusion molecule increased its in vitro neutralizing activity against H1N1 and H2N3 viruses up to 20-fold and, moreover, resulted in obtaining the ability to neutralize H5N2 and H9N2 subtypes. We demonstrated that a dose as low as 0.6 mg/kg of G2.3-Fc or H1.2-Fc administered systemically or locally before infection could protect mice from lethal challenges with both H1N1 and H5N2 viruses. Furthermore, G2.3-Fc reduced the lung viral load to an undetectable level. Both VHH-Fc showed in vivo therapeutic efficacy when delivered via systemic or local route. The findings support G2.3-Fc as a potential therapeutic agent for both prophylaxis and therapy of Group 1 influenza A infection.

immunology↗

bioRxiv2021

Biofilm matrix proteome of clinical strain of P. aeruginosa

Extracellular matrix plays a pivotal role in biofilm biology and proposed as a potential target for therapeutics development. As matrix is responsible for some extracellular functions and influence bacterial cytotoxicity against eukaryotic cells, it must have unique protein composition. P. aeruginosa is one of the most important pathogens with emerging antibiotic resistance, but only a few studies were devoted to matrix proteomes and there are no studies describing matrix proteome for any clinical isolates. Here we report the first biofilm matrix proteome of P. aeruginosa isolated from bronchoalveolar lavage of patient in intensive care unit. We have identified the largest number of proteins in the matrix among all published studies devoted to P. aeruginosa biofilms. Comparison of matrix proteome with proteome from embedded cells let us to identify several enriched bioprocess groups. Bioprocess groups with the largest number of overrepresented in matrix proteins were oxidation-reduction processes, proteolysis, and transmembrane transport. The top three represented in matrix bioprocesses concerning the size of the GO annotated database were cell redox homeostasis, nucleoside metabolism, and fatty acid synthesis. Finally, we discuss the obtained data in a prism of antibiofilm therapeutics development.

microbiology↗

bioRxiv2021

Single-domain antibodies efficiently neutralize SARS-CoV-2 variants of concern

Virus-neutralizing antibodies are one of the few treatment options for COVID-19. The evolution of SARS-CoV-2 virus has led to the emergence of virus variants with reduced sensitivity to some antibody-based therapies. The development of potent antibodies with a broad spectrum of neutralizing activity is urgently needed. Here we isolated a panel of single-domain antibodies that specifically bind to the receptor-binding domain of SARS-CoV-2 S glycoprotein. Three of the selected antibodies exhibiting most robust neutralization potency were used to generate dimeric molecules. We observed that these modifications resulted in up to a 200-fold increase in neutralizing activity. The most potent heterodimeric molecule efficiently neutralized each of SARS-CoV-2 variant of concern, including Alpha, Beta, Gamma, Delta and Omicron variants. This heterodimeric molecule could be a promising drug candidate for a treatment for COVID-19 caused by virus variants of concern.

immunology↗