Novel autochthonous strains from Cyprinus carpio as candidates for probiotic use and microplastic-degrading properties

In the modern era, identifying and characterizing novel bacterial strains with possible probiotic potential and environmental bioremediation capabilities is an emerging focus in microbiology and biotechnology. This study analysed the cultivable gut microbiota of the freshwater fish, Cyprinus carpio, and identified six different bacterial genera, including Citrobacter, Serratia, Bacillus, Enterococcus, and Kocuria. Among these, two novel autochthonous strains—Hafnia alvei UUNT_MP41 and Hafnia paralvei UUNT_MP29—were isolated and selected for further investigation due to their promising probiotic traits and potential to degrade microplastics in aquatic ecosystems. Both strains were evaluated for antibacterial activity against pathogens and susceptibility to a broad spectrum of antibiotics. Whole-genome analysis using next-generation sequencing (NGS) revealed the presence of genes potentially associated with probiotic properties, such as ClpB, as well as genes potentially involved in the biodegradation of common microplastics, including the tesA gene, a homolog of the PpEst gene from the genome of Pseudomonas pseudoalcaligenes, and the lipR gene, a homolog of the EstC9 gene from the genome of Acidocella sp. Here, we performed a more in-depth analysis of the similarity between the genes/proteins we identified as potentially involved in plastic biodegradation and previously described ones. Notably, the identified strains' potential to degrade microplastics under conditions relevant to the human gastrointestinal system positions them as candidates for a new generation of dual-function probiotics, supporting both human health and microplastic detoxification. These findings lay the groundwork for future development of multifunctional probiotic formulations with environmental and therapeutic benefits

Increased sensivity to infection due to memantine-induced blocking of inlammatory cell trafficking

Memantine is a selective N-methyl-d-aspartate receptor antagonist, used clinically for treating Alzheimer’s disease. The proposed mechanism of action is the blocking of inflammatory cells infiltration into the brain and spinal cord. Previously, we uncovered the potential of memantine to ameliorate symptoms of experimental autoimmune encephalomyelitis (EAE) in old rats [previously published in: Bufan B et al (2024), Biomedicines 12 (4):717]. Here, we have treated female dark agouti rats (3 and 22 months old) with memantine (60 mg/kg body weight) for 7 days (1st–7th day post-immunization for EAE). On the 7th day, we assayed peritoneal and blood leukocytes for their phenotype and functionality. Memantine treatment reduced the expression of the surface major histocompatibility complex class II (MHCII) on peritoneal macrophages. Peritoneal macrophages were weakly responsive to in vitro LPS stimulation. Macrophages from old rats produced more nitric oxide (NO) ex vivo compared to macrophages from young rats. Cellularity of peritoneal cavity was inversely correlated with NO production, however memantine treatment seemed to reverse that correlation in old rats. Peritoneal cells from memantine-treated rats responded differently to phorbol 12-myristate 13-acetate (PMA) – there was an increased dihidrorhodamine fluorescence in all leucokytes, which was abrogated with memantine treatment, and in the old rat group there was a stronger PMA stimulation but reduced inhibitory effect of memantine. Among blood lymphocytes, there was a decreased expression of CD49d (CD43+CD62L+CD49d+), as well as an increased percentage of CD62L+ cells. Finally, while our results show a more pronounced effect of memantine in young rats, we have noticed that old rats under memantine treatment showed signs of nonEAE distress (rough fur, lethargy, respiratory issues). These results show the fast effect of memantine on a systemic level, as well as the potential downsides of its mechanism of action. *The authors marked with an asterisk equally contributed to the work

Mpox Cases in Serbia, 2022

Background: On 23 July 2022, the World Health Organization (WHO) declared the mpox multi-country outbreak as a Public Health Emergency of International Concern. This study aimed to identify the epidemiological and clinical characteristics of confirmed mpox cases reported in Serbia in 2022. Methods: The mpox WHO case definition was used. Incidence rates (IRs) and incidence rate ratios (IRRs) by age groups and nomenclature of territorial units for statistics level 3 (NUTS-3) with 95% confidence intervals (CIs) were calculated. Results: Between June and October 2022, 43 laboratory-confirmed cases were reported. All were unvaccinated males, with the mean age of 34 (±7.4) years. Out of the total, 72.1% cases were men who have sex with men (MSM), who reported sexual intercourse either with multiple or unknown partners (p < 0.01). Fifteen cases (34.9%) lived with HIV, mostly in the 30–39 age group (p = 0.023). People living in Belgrade City NUTS-3 were six times more likely to become infected compared to South Backa citizens (IRR: 6.03, 95% CI: 1.47–25.53). Conclusions: In Serbia, mpox mainly affected MSM aged 30–39 and living in urban areas. Health promotion and vaccine implementation should be prioritized in populations with a higher risk

Regulation of Erythropoietin Activity in Clear Renal Cell Carcinoma

Abstract: Clear-cell renal cell carcinoma (ccRCC) is associated with the mutated von Hippel–Lindau (VHL) gene leading to the activation of hypoxia-inducible factor 1A (HIF1A) and subsequent overexpression of erythropoietin (EPO). We analyzed tumor and healthy tissues from 43 ccRCC patients after radical nephrectomy and cultured 786-O (biallelic VHL inactivation) and Caki-1 (wild-type VHL) cells in normal (21% O2) and low oxygen (3% O2) with 10% and 2% fetal bovine serum (FBS). DNA sequencing, including Sanger sequencing, MLPA and LOH, revealed 27 somatic mutations of VHL in ccRCC. HIF1A protein showed decreased or no expression in tumors compared to healthy tissue, independent of VHL alteration. The 786-O cells showed increased HIF1A protein expression after 48 h under low oxygen and 10% FBS. EPO and erythropoietin receptor (EPOR) were significantly decreased in ccRCC without HIF1A expression. EPO mRNA increased in the 786-O cells at 3% O2 after 48 h, while the Caki-1 cells had low or no EPO expression. Hypoxia increased EPOR mRNA in the Caki-1 cells at 10% FBS, but decreased in the 786-O cells at 2% FBS after 48 h. JAK2/STAT5A activity was increased only in HIF1A-positive tumors. These results suggest that EPO/EPOR activation in ccRCC is mainly driven by low oxygen, not VHL regulation of hypoxia-related responses

Herd Immunity To The Measles, Mumps And Rubella Viruses Among The Belgradian Population In May, 2024

Background/Objectives: In the Republic of Serbia, anti-measles vaccination was first introduced in 1971, while combined vaccination (measles, mumps, rubella) was made mandatory in 1996 as part of the national vaccination program. Reported prevalence values for 2023 were: < 0.75 cases per 100K population for measles; 0.09 cases per 100K for mumps; and no cases of rubella. Methods: This cross-sectional study was performed in May, 2024 as part of the project "Herd Immunity to Vaccine-Preventable and Other Relevant Infections in the Belgradian Population." It focused on assessing herd immunity to measles, mumps and rubella (MMR) among residents, insofar as these remain a public concern despite the availability of vaccines. A total of 2,533 subjects were distributed across nine age groups, covering those aged 1-70+ years and various professional groups, residing in Belgrade. Participants were stratified by age and activity. Upon obtaining individual information by online questionnaire, and receiving a signed statement of in-formed consent, blood samples were obtained for IgG antibody testing (ELISA) to de-termine MMR serological status. The results were compared to national and interna-tional immunization standards to evaluate herd immunity levels. Results: Our results indicate varying levels of immunity for each virus, with specific demographic groups showing different immunity levels. Total measles seroprevalence during this study was 74.7%, with significant variation across all age groups. While high seropositivity was observed in both children (90.7%) and elder age groups (98.4%), middle-aged individuals in the age group 30-49 years showed significantly lower IgG levels. Between 2021 and 2023, there were no registered cases of rubella detected in Serbia, which indicates a high level of immunity. This was confirmed here with consistently high IgG levels across all age groups, with an average seropositivity of 94.8%. Average mumps seropositivity across all age groups was 85.1%. The lowest value was in the young child (1-5 years) age group (76.1%); the highest was in the elderly group (92.6%). Conclusions: The current findings suggest that the Belgradian population has strong overall immunity to MMR, yet with some concerns regarding measles immunity in middle-aged adults, suggesting a potential need for catch-up vaccinations. While rubella status indicates strong herd immunity and minimal risk of outbreaks, mumps immunity in some groups (children, middle-aged adults) is below the protective threshold. While it is still sufficient to pre-vent widespread transmission, it should be closely observed. To our knowledge, this study is the first of its kind to provide data about MMR seroprevalence in Belgrade. Findings indicate the need for constant surveillance and revaccination of vulnera-ble/seronegative groups

Investigation of novel clusters of bacterial head-to-tail cyclized peptides from the Bacillus pumilus genome

Bacterial headtotail cyclized peptides are a group of ribosomally synthesized and post -translationally modified peptides (RiPPs) of relatively large size. During maturation, the peptideundergoes cyclization, forming a peptide bond between its Cterminus and Nterminus. Comparedto other cyclic RiPPs bacterial cyclized peptides are distinguished by their larger cyclic structureand likely by their cyclization mechanism. The sequences of these peptides are highly diverse, andare not similar to each other. However, they share key properties: remarkable resistance to hightemperatures, proteases, and pH fluctuations. These features make them promising candidates forapplications in food preservation and medicine, for example, as antimicrobial agents. All thesepeptides are hydrophobic, and their mechanism of action is thought to involve disrupting the targetorganism’s membrane. It is believed that they form pores in the membrane, altering ionpermeability and ultimately leading to cell death. However, information on their mode of actionremains scarce, and the mechanisms for specific representat ives of this group are not fullyunderstood. Recently, our research team identified two novel clusters of bacterial headtotailcyclized peptides in the genome of Bacillus pumilus. The clusters differ from each other in boththe primary sequence of the precursor peptide and the number of genes within the cluster.Moreover, one of the clusters contains two precursor peptides, which is highly unusual for a clusterof this type. We tested the antibacterial activity of this strain against a range of Gram positive andGram negative bacteria, observing zones of growth inhibition likely linked to the production ofcyclized peptides. To further investigate their antibacterial activity, functions of genes, andcyclization mechanism, we constructed a recombinant shuttle vector based on the pHT01 plasmidfor heterologous expression of these clusters in another Bacillus strain.BeCELS 2025: Belgrade Conference for Early-Career Life Scientists, taking place on Friday, September 5, 2025, at the Institute of Molecular Genetics and Genetic Engineering (IMGGE) in Belgrad

Supplementary information for the article: Miljković, R.; Marinković, E.; Prodić, I.; Kovačević, A.; Protić-Rosić, I.; Vasić, M.; Lukić, I.; Gavrović-Jankulović, M.; Stojanović, M. Ameliorative Effect of Banana Lectin in TNBS-Induced Colitis in C57BL/6 Mice Relies on the Promotion of Antioxidative Mechanisms in the Colon. Biomolecules 2025, 15 (4), 476. https://doi.org/10.3390/biom15040476.

Supplementary Table S1. Histology scoring for estimation of disease severitySupplementary material for: [https://intor.torlakinstitut.com/handle/123456789/987] Related to the published version: [https://doi.org/10.3390/biom15040476]Related to the published version: [https://doi.org/10.3390/biom15040476

Cathepsin L inhibition enhances microglial CX3CR1 expression in EAE

Cathepsin L (CatL), a lysosomal cysteine protease, is highly expressed in antigen-presenting cells and plays a role in the processing of autoantigens. It is widely expressed throughout the CNS and is associated with microglia-driven neuroinflammatory responses. Previous studies suggest that inhibition of catL activity may offer therapeutic potential for the treatment of multiple sclerosis. The aim of this study was to investigate the effects of a selective catL inhibitor [CC(=O)c1cccc(NC(=O)c2cnc3sccn3c2=O) c1] on microglia in mice immunized for experimental autoimmune encephalomyelitis (EAE). The catL inhibitor was administered intraperitoneally for five consecutive days from the onset of clinical signs of disease. Mononuclear cells were isolated from the spinal cord (SC) of EAE rats and analysed by flow cytometry. Given that this was the first in vivo application of this inhibitor, hepatotoxicity and nephrotoxicity were also investigated. The administered catL inhibitor reduced the infiltration of the SC by CD4+ T lymphocytes. On the other hand, the proportion of CD11b+ cells among the mononuclear cells isolated from the SC was higher in the treated EAE mice. CatL inhibitor decreased the proportion of CD45high cells among CD11b+ cells, most likely monocyte-derived macrophages that infiltrated the SC and increased the proportion of CD45low/int cells among CD11b+ cells, which correspond to microglia. The proportion of neuroprotective CX3CR1+ cells among microglial cells was significantly higher in treated mice than in untreated EAE mice. These findings reinforce the hypothesis that catL could serve as a potential therapeutic target for treating multiple sclerosis

Expression of terminal galactose and sialic acid on serum IgA in IgA multiple myeloma

Introduction/Objective. IgA multiple myeloma has a poor prognosis, and altered glycosylation of myeloma IgA may be one of contributing factors. This study examined the expression of terminal galactose and sialic acid (SA) on serum IgA oligosaccharides in patients with IgA myeloma, compared to healthy control sera. Methods. Serum samples from 15 IgA myeloma patients and pooled serum from 100 healthy donors were analyzed. IgA was purified using peptide M affinity chromatography. Terminal galactose and SA expression on isolated IgA was analyzed by Ricinus communis agglutinin I and Sambucus nigra agglutinin lectin blotting. Results. IgA-heavy chains from both healthy individuals and all myeloma patients expressed galactose. SA was present in healthy control and in 14 out of 15 myeloma patients. Compared to controls, myeloma IgA showed 12–63% a reduction in galactose and a 67–97% reduction in SA expression on heavy chains. Notable galactosylation of IgA-light chains was observed in only three, while weak SA expression was seen in 14 myeloma cases. Healthy IgA was predominantly monomeric and expressed both galactose and SA. Myeloma IgA existed in both and polymeric forms expressing detectable galactose level, though with different expression levels among individuals. At the same time, SA was undetectable. Conclusion. The results of this study showed altered glycosylation of myeloma IgA. Compared to healthy control, myeloma IgA-heavy chains expressed reduced terminal galactose and SA. Notable galactosylation of light chains was observed in three cases. Unlike SA, galactose was detectable on intact monomeric and polymeric multiple myeloma IgA

Modulation of T-Cell-Dependent Humoral Immune Response to Influenza Vaccine by Multiple Antioxidant/Immunomodulatory Micronutrient Supplementation

Notwithstanding prevalence gaps in micronutrients supporting immune functions, the significance of their deficits/supplementation for the efficacy of vaccines is underinvestigated. Thus, the influence of supplementation combining vitamins C and D, zinc, selenium, manganese, and N-acetyl cysteine on immune correlates/surrogates of protection conferred by a quadrivalent influenza vaccine (QIV) in mice was investigated. The supplementation starting 5 days before the first of two QIV injections given 28 days apart increased the serum titres of total and neutralizing IgG against each of four influenza strains from QIV. Accordingly, the frequencies of germinal center B cells, follicular CD4+ T helper (Th) cells, and IL-21-producing Th cells increased in secondary lymphoid organs (SLOs). Additionally, the supplementation improved already increased IgG response to the second QIV injection by augmenting not only neutralizing antibody production, but also IgG2a response, which is important for virus clearance, through favoring Th1 differentiation as indicated by Th1 (IFN-γ)/Th2 (IL-4) signature cytokine level ratio upon QIV restimulation in SLO cell cultures. This most likely partly reflected antioxidant action of the supplement as indicated by splenic redox status analyses. Thus, the study provides a solid scientific background for further research aimed at repurposing the use of this safe and inexpensive micronutrient combination to improve response to the influenza vaccine