Current Trends in the Management of Helicobacter pylori Infection in Serbia: Preliminary Results from the European Registry on H. pylori Management.

BACKGROUND: Helicobacter pylori (H. pylori) is the most common chronic bacterial infection. Treatment effectiveness remains a subject of debate considering bacterial antimicrobial resistance. Our aim was to analyze the diagnostic methods and eradication treatments for H. pylori infection in Serbia. METHODS: An observational multicenter prospective study was conducted in Serbia, as part of the European Registry on H. pylori Management (Hp-EuReg). Demographics, treatment indication, diagnostic methods, previous eradication attempts, and treatment were collected at AEG-REDCap e-CRF. Modified intention-to-treat (mITT) and per-protocol (PP) effectiveness analyses were performed. Safety, compliance, and bacterial antimicrobial resistance rates were reported. Data were quality checked. RESULTS: Overall, 283 patients were included, with a mean age of 55 ± 15 years. Dyspepsia (n = 214, 77%) was the most frequent treatment indication, and histology (n = 144, 51%) was the most used diagnostic method. Overall eradication rate was 95% (PP) and 94% (mITT). Most prevalent first-line therapy was quadruple PPI + clarithromycin + amoxicillin + metronidazole, with a 96% effectiveness (p < 0.001). Second-line main treatment choice was triple amoxicillin + levofloxacin, with a 95% effectiveness (p < 0.05). Single-capsule Pylera® was the most prescribed third-line therapy, with 100% effectiveness (p < 0.05). Longer treatment duration was associated with a higher eradication rate in first-line therapy (p < 0.05). Clarithromycin and quinolone resistance rates in first-line were 24% and 8.3%, respectively. The overall adverse events' incidence rate was 13.4%, and therapy compliance was 97%. CONCLUSIONS: Considering the high eradication rate, 14-day non-bismuth quadruple concomitant therapy is a reasonable first-line choice, while quinolone-based therapy and single-capsule Pylera® should be considered as rescue therapy options.

Dissemination of Metallo-ß-Lactamase-Producing Pseudomonas aeruginosa in Serbian Hospital Settings: Expansion of ST235 and ST654 Clones.

This nationwide study aimed to investigate the molecular characteristics of metallo-ß-lactamase (MBL)-producing Pseudomonas aeruginosa in Serbia, underlying resistance mechanisms, the genetic context of detected MBL genes, and the clonal relationship between isolates harboring genes-encoding MBL. Overall, 320/5334 isolates collected from 2018 to 2021 were identified as P. aeruginosa. Carbapenem-resistant P. aeruginosa (CRPA) were screened for the presence of blaVIM, blaIMP, and blaNDM, genes whereas MBL-positive isolates were tested for the presence of the blaCTX-M-2, blaPER, blaTEM, blaSHV, blaVEB, and blaGES. Multilocus sequence typing and phylogenomic analysis were performed for P. aeruginosa-producing MBL. The majority of the P. aeruginosa isolates were recovered from the lower respiratory tract (n = 120; 37.5%) and wound specimens (n = 108; 33.75%). CRPA isolates accounted for 43.1% (n = 138) of the tested isolates, 31 out of them being blaNDM-1-positive (22.5%). The colistin resistance rate was 0.3%. MLST analysis revealed the occurrence of ST235 (n = 25) and ST654 (n = 6), mostly confined to Serbia. The distribution of beta-lactamase-encoding genes in these isolates suggested clonal dissemination and possible recombination: ST235/blaNDM-1, ST235/blaNDM-1/blaPER-1, ST654/blaNDM-1, ST654/blaNDM-1/blaPER-1, and ST654/blaNDM-1/blaGES-5. High-risk clones ST235 and ST654 identified for the first time in Serbia, are important vectors of acquired MBL and ESBL and their associated multidrug resistance phenotypes represent a cause for considerable concern.

Tigecycline susceptibility of multidrug-resistant Acinetobacter baumannii from intensive care units in the western Balkans.

Tigecycline can be effective to treat infections of carbapenem-resistant Acinetobacter baumannii (CRAB) however, no interpretive criteria have been approved so far. The objectives of this study were to evaluate the proportion of CRAB isolates and to compare gradient test with a broth microdilution (BMD) method for tigecycline susceptibility testing of A. baumannii.This study included 349 multidrug-resistant (MDR) Acinetobacter spp. collected from Serbia, Montenegro, Bosnia and Herzegovina in 2016 and 2017. Antibiotic susceptibility testing was performed by disk diffusion, VITEK2, gradient, ComASP Colistin. Tigecycline susceptibilities were interpreted according to breakpoints of European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Food and Drug Administration (FDA).Majority of the tested isolates were CRAB (92.8%). Tigecycline MIC50/MIC90 values were 4/8 µg/mL by BMD and 0.5/4 µg/mL by gradient test. Essential agreement for BMD and gradient test amounted to 65.1%. With EUCAST breakpoints, categorical agreement (CA) was achieved in 38% isolates. Major discordance (MD-false susceptibility/resistance) and minor discordance (mD-false categorization involving intermediate results) were observed in 10% and 57% A. baumannii, respectively. With FDA breakpoints, CA, MD and mD were observed in 44%, 16% and 47% isolates, respectively. Colistin resistance was 2.1%.The study highlights a high proportion of CRAB and several discordances between BMD and gradient test which may lead to inappropriate therapy.

Molecular epidemiology of invasive and non-invasive group B Streptococcus circulating in Serbia.

Streptococcus agalactiae (group B Streptococcus, GBS) remains the leading cause of invasive diseases in neonates and an important cause of infections in the elderly. The aim of this study was to access the prevalence of GBS genito-rectal colonisation of pregnant women and to evaluate the genetic characteristics of invasive and non-invasive GBS isolates recovered throughout Serbia. A total of 432 GBS isolates were tested for antimicrobial susceptibility, capsular polysaccharide (CPS) types and the presence of the hvgA gene. One hundred one randomly selected isolates were further characterized by clustered regularly interspaced short palindromic repeats (CRISPRs) analysis and/or multilocus sequence typing (MLST). The prevalence of GBS colonization in pregnant women was 15%. Overall, six capsular types (Ia, Ib, II to V) were identified, the most common being III (32.2%) and V (25.2%). The hiper-virulent clone type III/ST17 was present in 43.1% and 6.3% (p < 0.05) of paediatric and adults isolates, respectively. Comparative sequence analysis of the CRISPR1 spacers content indicated that a few clones comprised the vast majority of the tested GBS isolates. Thus, it was estimated that dominant clones recovered from infants were CPS III/ST17 in late-onset infections (19/23; 82.6%), and Ia/ST23 in early-onset disease (44.4%). Conversely, genotype CPS V/ST1 was the most prevalent in adults (4/9; 25.4%). All isolates were susceptible to penicillin. Macrolide resistance (23.1%) was strongly associated with the ermB gene and constitutive resistance to clindamycin (63.9%). The majority of strains was resistant to tetracycline (86.6%), mostly mediated by the tetM gene (87.7%). GBS isolates of CPS V/ST1 and CPS III/ST23 were significantly associated with macrolide and tetracycline resistance, respectively. In conclusion, hyper-virulent CPS III/ST17 and V/ST1 were recognized as dominant GBS clones in this study.

Papillary Thyroid Carcinoma: A Malignant Tumor with Increased Antioxidant Defense Capacity.

Papillary thyroid carcinoma (PTC) is the commonest thyroid malignancy worldwide for which the radiation exposure is the most influential risk factor. The levels of oxidative stress in PTC are not well characterized on the tissue level. The objective of this study was to evaluate total oxidant status (TOS) and total antioxidant status (TAS) in PTC and benign goiter (BG) tissues and to examine their association with clinicopathological characteristics. Tumor and normal thyroid tissue samples were collected from 59 PTC patients, and goiter tissues were collected from 50 BG patients. TOS and TAS were quantified in the tissue homogenates by spectrophotometric assays. TOS values in tumor tissues did not differ significantly from normal and goiter tissues; however, PTC tissues have significantly higher TAS values than normal and goiter tissues. TOS values correlated with retrosternal growth in BG patients. The significant correlations were found between TOS and TAS values and thyroid function parameters. In 17 PTC patients with multiple tumor foci (multicentric phenotype), TAS values were significantly lower, compared to 42 patients with unicentric PTC. TAS and TOS are the most useful predictors of thyroid capsular invasion by PTC. The age, sex, body mass index, smoking, familial history of thyroid disease and nodule size did not influence TOS and TAS in PTC or BG patients. In conclusion, we show the profiles of TOS and TAS in PTC and BG tissues. Importantly, PTC tissues possess increased antioxidant capacity. The redox status influences the parameters of the thyroid function and tumor's biological behavior.

Effect of Childhood Pneumococcal Conjugate Vaccination on Invasive Disease Serotypes in Serbia.

In Serbia, PCV10 was introduced into the routine immunization for children under 2 in 2018 and replaced by PCV13 in 2022. We evaluated their impact on the distribution of invasive pneumococcal disease (IPD) serotypes across all age groups. Overall, 756 isolates were obtained from patients with IPD between 2010 and 2023 through laboratory surveillance. In the post-vaccination period, serotypes 14, 19F, 23F, and 6A significantly declined, while 3 and 19A considerably increased. This was especially evident in the ≤2 years group, making these serotypes the most prevalent among them. Serotype 3 dominated, representing 19.1% of all invasive isolates prior to 2018 and 33.1% thereafter. While serotype coverage of PCV10 has significantly decreased in the ≤2 years group (from 74.2% before 2018 to 29.5% after 2018), PCV13 coverage was 63.9% after 2018. In the post-PCV period, non-PCV13 serotypes, such as 9N, 10A, 15A, 15B, 15C, 22F, 6C, 6D, and 7C, increased across all isolates. Antibiotic non-susceptibility considerably decreased after 2018. MLST analysis showed shifts in sequence type prevalence, with pre-PCV lineages replaced and ongoing serotype 3 persistence, alongside potential capsule-switching events. These findings emphasize a noticeable shift in the distribution of serotypes and adaptability of pneumococcal populations, highlighting the importance of ongoing surveillance and the requirement for the urgent introduction of higher valent vaccines into the National Immunization Program.

Potential influence of antimicrobial resistance gene content in probiotic bacteria on the gut resistome ecosystems.

Antimicrobial resistance (AMR) poses a substantial threat to human health. The commensal bacteria of the gut microbiome were shown to serve as a reservoir of antibiotic resistance genes (ARGs), termed the gut resistome, which has the potential to transfer horizontally to pathogens and contribute to the emergence of drug-resistant bacteria. Namely, AMR traits are generally linked with mobile genetic elements (MGEs), which apart from disseminating vertically to the progeny, may cross horizontally to the distantly related microbial species. On the other hand, while probiotics are generally considered beneficiary to human health, and are therefore widely consumed in recent years most commonly in conjunction with antibiotics, the complexities and extent of their impact on the gut microbiome and resistome have not been elucidated. By reviewing the latest studies on ARG containing commercial probiotic products and common probiotic supplement species with their actual effects on the human gut resistome, this study aims to demonstrate that their contribution to the spread of ARGs along the GI tract merits additional attention, but also indicates the changes in sampling and profiling of the gut microbiome which may allow for the more comprehensive studying of the effects of probiotics in this part of the resistome.

Comparative genomics and molecular epidemiology of colistin-resistant Acinetobacter baumannii.

This study aimed to investigate the prevalence and resistance mechanisms of colistin-resistant Acinetobacter baumannii (ColRAB) isolates in Serbia, assess their genetic relatedness to other circulating A. baumannii isolates in the neighbouring European countries, and analyse the global genomic epidemiology of ColRAB isolates. A total of 784 isolates of A. baumannii were recovered from hospitalised patients in Serbia between 2018 and 2021. The antimicrobial susceptibility testing was performed using disk diffusion and broth microdilution. All ColRAB isolates were subjected to DNA isolation and whole-genome sequencing (WGS). Overall, 3.94 % (n = 30) isolates were confirmed as ColRAB. Results of mutational and transcriptional analysis of genes associated with colistin resistance indicate the central role of the two-component regulating system, PmrAB, and increased expression of the pmrC gene in ColRAB. Most of the isolates (n = 29, 96.6 %) belonged to international clone II, with the most common sequence type being STPas2 (n = 23, 76.6 %). Based on the WGS analysis, ColRAB isolates belonging to the same ST isolated in various countries were grouped into the same clusters, indicating the global dissemination of several high-risk clonal lineages. Phylogenomic analysis of ColRAB isolates, together with all previously published A. baumannii genomes from South-Eastern European countries, showed that colistin resistance arose independently in several clonal lineages. Comparative genomic analysis revealed multiple genes with various roles (transcriptional regulation, transmembrane transport, outer membrane assembly, etc.), which might be associated with colistin resistance in A. baumannii. The obtained findings serve as the basis for further studies, contributing to a better understanding of colistin resistance mechanisms in A. baumannii.

Serotype distribution, antimicrobial susceptibility and molecular epidemiology of invasive Streptococcus pneumoniae in the nine-year period in Serbia.

Streptococcus pneumoniae is one of the leading bacterial pathogens that can cause severe invasive diseases. The aim of the study was to characterize invasive isolates of S. pneumoniae obtained during the nine-year period in Serbia before the introduction of the pneumococcal conjugate vaccines (PCVs) into routine vaccination programs by determining: serotype distribution, the prevalence and genetic basis of antimicrobial resistance, and genetic relatedness of the circulating pneumococcal clones. A total of 490 invasive S. pneumoniae isolates were included in this study. The serotype, antimicrobial susceptibility, and ST of the strains were determined by the Quellung reaction, disk- and gradient-diffusion methods, and multilocus sequence typing (MLST), respectively. The most common serotypes in this study were 3, 19F, 14, 6B, 6A, 19A, and 23F. The serotype coverages of PCV10 and PCV13 in children less than 2 years were 71.3 and 86.1%, respectively, while PPV23 coverage in adults was in the range of 85-96%, depending on the age group. Penicillin and ceftriaxone-non-susceptible isolates account for 47.6 and 16.5% of all isolates, respectively. Macrolide non-susceptibility was detected in 40.4% of isolates, while the rate of multidrug- and extensive-drug resistance was 20.0 and 16.9%, respectively. The MLST analysis of 158 pneumococci identified 60 different STs belonging to the 16 Clonal Complexes (CCs) (consisting of 42 STs) and 18 singletons. The most common CC/ST were ST1377, CC320, CC15, CC273, CC156, CC473, CC81, and CC180. Results obtained in this study indicate that the pre-vaccine pneumococcal population in Serbia is characterized by high penicillin and macrolides non-susceptibility, worrisome rates of MDR and XDR, as well as a high degree of genetic diversity. These findings provide a basis for further investigation of the changes in serotypes and genotypes that can be expected after the routine introduction of PCVs.

Carbapenem-Resistant Acinetobacter baumannii: Biofilm-Associated Genes, Biofilm-Eradication Potential of Disinfectants, and Biofilm-Inhibitory Effects of Selenium Nanoparticles.

This study aimed to investigate the biofilm-production ability of carbapenem-resistant Acinetobacter baumannii (CRAB), the biofilm-eradication potential of 70% ethanol and 0.5% sodium hypochlorite, the effects of selenium nanoparticles (SeNPs) against planktonic and biofilm-embedded CRAB, and the relationship between biofilm production and bacterial genotypes. A total of 111 CRAB isolates were tested for antimicrobial susceptibility, biofilm formation, presence of the genes encoding carbapenemases, and biofilm-associated virulence factors. The antibiofilm effects of disinfectants and SeNPs against CRAB isolates were also tested. The vast majority of the tested isolates were biofilm producers (91.9%). The bap, ompA, and csuE genes were found in 57%, 70%, and 76% of the CRAB isolates, with the csuE being significantly more common among biofilm producers (78.6%) compared to non-biofilm-producing CRAB (25%). The tested disinfectants showed a better antibiofilm effect on moderate and strong biofilm producers than on weak producers (p < 0.01). The SeNPs showed an inhibitory effect against all tested planktonic (MIC range: 0.00015 to >1.25 mg/mL) and biofilm-embedded CRAB, with a minimum biofilm inhibitory concentration of less than 0.15 mg/mL for 90% of biofilm producers. In conclusion, SeNPs might be used as promising therapeutic and medical device coating agents, thus serving as an alternative approach for the prevention of biofilm-related infections.

Isolation, Characterization, Genome Analysis and Host Resistance Development of Two Novel Lastavirus Phages Active against Pandrug-Resistant Klebsiella pneumoniae.

Klebsiella pneumoniae is a global health threat and bacteriophages are a potential solution in combating pandrug-resistant K. pneumoniae infections. Two lytic phages, LASTA and SJM3, active against several pandrug-resistant, nosocomial strains of K. pneumoniae were isolated and characterized. Their host range is narrow and latent period is particularly long; however, their lysogenic nature was refuted using both bioinformatic and experimental approaches. Genome sequence analysis clustered them with only two other phages into the new genus Lastavirus. Genomes of LASTA and SJM3 differ in only 13 base pairs, mainly located in tail fiber genes. Individual phages, as well as their cocktail, demonstrated significant bacterial reduction capacity in a time-dependent manner, yielding up to 4 log reduction against planktonic, and up to 2.59 log on biofilm-embedded, cells. Bacteria emerging from the contact with the phages developed resistance and achieved numbers comparable to the growth control after 24 h. The resistance to the phage seems to be of a transient nature and varies significantly between the two phages, as resistance to LASTA remained constant while resensitization to SJM3 was more prominent. Albeit with very few differences, SJM3 performed better than LASTA overall; however, more investigation is needed in order to consider them for therapeutic application.

In Search for Reasons behind Helicobacter pylori Eradication Failure-Assessment of the Antibiotics Resistance Rate and Co-Existence of Helicobacter pylori with Candida Species.

Helicobacter pylori eradication is characterized by decreasing successful eradication rates. Although treatment failure is primarily associated with resistance to antibiotics, other unknown factors may influence the eradication outcome. This study aimed to assess the presence of the antibiotics resistance genes in H. pylori and the presence of Candida spp., which are proposed to be endosymbiotic hosts of H. pylori, in gastric biopsies of H. pylori-positive patients while simultaneously assessing their relationship. The detection and identification of Candida yeasts and the detection of mutations specific for clarithromycin and fluoroquinolones were performed by using the real-time PCR (RT-PCR) method on DNA extracted from 110 gastric biopsy samples of H. pylori-positive participants. Resistance rate to clarithromycin and fluoroquinolone was 52% and 47%, respectively. Antibiotic resistance was associated with more eradication attempts (p < 0.05). Candida species were detected in nine (8.18%) patients. Candida presence was associated with older age (p < 0.05). A high rate of antibiotic resistance was observed, while Candida presence was scarce, suggesting that endosymbiosis between H. pylori and Candida may not be a major contributing factor to the eradication failure. However, the older age favored Candida gastric mucosa colonization, which could contribute to gastric pathologies and microbiome dysbiosis.

Current Susceptibility Surveillance and Distribution of Antimicrobial Resistance in N. gonorrheae within WHO Regions.

Neisseria gonorrhoeae (N. gonorrhoeae) is the etiological agent of the second most common sexually transmitted disease in the world, gonorrhoea. Currently recommended and last available first-line therapy is extended-spectrum cephalosporins most often combined with azitromycin. However, misuse of antibiotics and the abilities of N. gonorrhoeae to acquire new genetic and plasmid-borne resistance determinants has gradually led to the situation where this bacterium has become resistant to all major classes of antibiotics. Together with a generally slow update of treatment guidelines globally, as well as with the high capacity of gonococci to develop and retain AMR, this may lead to the global worsening of gonococcal AMR. Since effective vaccines are unavailable, the management of gonorrhoea relies mostly on prevention and accurate diagnosis, together with antimicrobial treatment. The study overviews the latest results of mostly WHO-initiated studies, primarily focusing on the data regarding the molecular basis of the resistance to the current and novel most promising antibacterial agents, which could serve to establish or reinforce the continual, quality-assured and comparable AMR surveillance, including systematic monitoring and treatment with the use of molecular AMR prediction methods.

Antimicrobial Susceptibility Testing: A Comprehensive Review of Currently Used Methods.

Antimicrobial resistance (AMR) has emerged as a major threat to public health globally. Accurate and rapid detection of resistance to antimicrobial drugs, and subsequent appropriate antimicrobial treatment, combined with antimicrobial stewardship, are essential for controlling the emergence and spread of AMR. This article reviews common antimicrobial susceptibility testing (AST) methods and relevant issues concerning the advantages and disadvantages of each method. Although accurate, classic technologies used in clinical microbiology to profile antimicrobial susceptibility are time-consuming and relatively expensive. As a result, physicians often prescribe empirical antimicrobial therapies and broad-spectrum antibiotics. Although recently developed AST systems have shown advantages over traditional methods in terms of testing speed and the potential for providing a deeper insight into resistance mechanisms, extensive validation is required to translate these methodologies to clinical practice. With a continuous increase in antimicrobial resistance, additional efforts are needed to develop innovative, rapid, accurate, and portable diagnostic tools for AST. The wide implementation of novel devices would enable the identification of the optimal treatment approaches and the surveillance of antibiotic resistance in health, agriculture, and the environment, allowing monitoring and better tackling the emergence of AMR.

Redox metabolism correlates with cellular turnover and clinical phenotype of papillary thyroid carcinoma and colloid goiter.

Introduction: Papillary thyroid carcinoma (PTC) and colloid goiter (CG) represent the most common thyroid malignant and benign diseases, respectively. Oxidative stress is considered to have an important role in the pathogenesis of both diseases, but without sufficient and comprehensive data. The aim was to evaluate the redox profile, its influence on cell survival of PTC, comparing it with CG as a control and its relation with demographic, pathological and clinical parameters. Material and methods: We evaluated for the first time the PTC and CG tissue profile of advanced oxidation protein products (AOPP) and total thiols as parameters of redox metabolism and deoxyribonuclease I (DNase I) and deoxyribonuclease II (DNase II) activity as biomarkers of cell turnover and apoptosis. Tissue levels of biochemical parameters were quantified in PTC and CG tissue using spectrophotometric methods. Study parameters were evaluated in light of different demographic, clinical and pathological features of PTC and CG. Results: Papillary thyroid carcinoma tissue is characterized by increased antioxidant activity and a normal prooxidation level. Biochemical parameters show numerous correlations with demographic and clinical characteristics of PTC and CG patients. DNase I and II activities are dependent upon the AOPP concentration in PTC tissue. The size of CG can be predicted with combined use of AOPP, DNase I and DNase II. AOPP is the most powerful predictor of PTC capsular invasion, multicentric intrathyroid dissemination and lymph node metastasis phenotype. Conclusions: Evaluated parameters can be used for assessment of tumor redox and survival status and the clinical course of PTC and CG.

Cinnamon essential oil and its emulsion as efficient antibiofilm agents to combat Acinetobacter baumannii.

Acinetobacter baumannii is an emerging nosocomial pathogen resistant to a wide spectrum of antibiotics, with great potential to form a biofilm, which further aggravates treatment of infections caused by it. Therefore, searching for new potent agents that are efficient against A. baumannii seems to be a necessity. One of them, which has already been proven to possess a wide spectrum of biological activities, including antimicrobial effect, is cinnamon essential oil. Still, further increase of antibacterial efficacy and improvement of bioavailability of cinnamon oil is possible by emulsification process. The aim of this study was comparative analysis of cinnamon essential oil and its emulsion against biofilm forming A. baumannii clinical isolates. Furthermore, the investigation of toxicological aspects of possible applications of essential oil and emulsion was done as well. Gas chromatography-mass spectrometry of essential oil indicated trans-cinnamaldehyde as the most abundant component. The cinnamon emulsion was synthesized from cinnamon essential oil by combining modified low- and high- energy methods. Synthesized emulsion was characterized with Fourier-transform infrared spectroscopy and photon correlation spectroscopy. Both substances exhibited significant antibacterial (minimal inhibitory concentrations in the range 0.125-0.5 mg/ml) and antibiofilm effects (inhibitions of formation and reduction of pre-formed biofilm were 47-81 and 30-62%, respectively). Compared to essential oil, the efficacy of emulsion was even stronger considering the small share of pure oil (20%) in the emulsion. The result of biofilm eradication assay was confirmed by scanning electron microscopy. Even though the cytotoxicity was high especially for the emulsion, genotoxicity was not determined. In conclusion, strong antibacterial/antibiofilm effect against A. baumannii of the cinnamon essential oil and the fact that emulsification even potentiated the activity, seems to be of great significance. Observed cytotoxicity implicated that further analysis is needed in order to clearly determine active principles being responsible for obtained antibacterial/antibiofilm and cytotoxic properties.

Matrix metalloproteinases and membrane damage markers in sera of patients with acute myocardial infarction.

Coronary artery disease is a multifunctional disease and represents one of the leading causes of death worldwide. Oxidative stress appears as an etiological factor for myocardial damage during acute myocardial infarction. Some data suggest that acute coronary syndromes may also be influenced by matrix metalloproteinases through degradation of the fibrous cap of vulnerable atherosclerotic lesions. It has been indicated that gelatinases A and B play a key role in acute myocardial infarction and deoxyribonuclease I has been postulated to be a novel early phase marker of disease. The aim was to study activity of gelatinases A and B in acute myocardial infarction and its association with some membrane damage markers. Seventy-five patients with disease and seventy-five healthy controls were enrolled. Activities of lactate dehydrogenase, malate dehydrogenase, and deoxyribonuclease I were estimated using standard spectrophotometric assay and isoforms of lactate and malate dehydrogenases were determined using direct zymography. Activity of dehydrogenases was significantly higher in patients, while deoxyribonuclease I was lower. Isoform 2 of lactate dehydrogenase was significantly higher in the patient group. Gelatinases A and B were detected only in patients group. The results suggest determination of serum malate dehydrogenase activity to be used as an additional parameter for acute myocardial infarction diagnosis. Those findings suggest important role of gelatinases A and B as biomarkers of early stage of acute myocardial infarction together with membrane damage parameters.

Study of the venom proteome of Vipera ammodytes ammodytes (Linnaeus, 1758): A qualitative overview, biochemical and biological profiling.

Vipera ammodytes (Va), is the European venomous snake of the greatest medical importance. We analyzed whole venom proteome of the subspecies V. ammodytes ammodytes (Vaa) from Serbia for the first time using the shotgun proteomics approach and identified 99 proteins belonging to four enzymatic families: serine protease (SVSPs), L-amino acid oxidase (LAAOs), metalloproteinases (SVMPs), group II phospholipase (PLA2s), and five nonenzymatic families: cysteine-rich secretory proteins (CRISPs), C-type lectins (snaclecs), growth factors -nerve (NGFs) and vascular endothelium (VEGFs), and Kunitz-type protease inhibitors (SPIs). Considerable enzymatic activity of LAAO, SVSPs, and SVMPs and a high acidic PLA2 activity was measured implying potential of Vaa to produce haemotoxic, myotoxic, neuro and cardiotoxic effects. Moreover, significant antimicrobial activity of Vaa venom against Gram-negative (Klebsiella pneumoniae, Pseudomonas aeruginosa) and Gram-positive bacteria (Staphylococcus aureus) was found. The crude venom shows considerable potential cytotoxic activity on the C6 and HL60 and a moderate level of potency on B16 cell lines. HeLa cells showed the same sensitivity, while DU 145 and PC-3 are less sensitive than as normal cell line. Our data demonstrated a high complexity of Vaa and considerable enzymatic, antibacterial and cytotoxic activity, implying a great medical potential of Vaa venom as a promising source for new antibacterial and cytostatic agents.

Trends in molecular characteristics and antimicrobial resistance of group B streptococci: a multicenter study in Serbia, 2015-2020.

Group B Streptococcus (GBS) is a major cause of neonatal morbidity and mortality. Serbia has not fully implemented preventive measures against GBS neonatal diseases. Therefore, we aimed to assess the maternal GBS colonisation and invasive neonatal disease rate, to reveal the trends of antimicrobial resistance and serotype distribution of GBS from various patient groups. Randomly selected non-invasive (n = 991) and all invasive GBS (n = 80) collected throughout Serbia from 2015 to 2020 were tested for antimicrobial susceptibility, capsular typing, and hvgA detection. Overall, 877/5621 (15.6%) pregnant women were colonised with GBS. Invasive GBS infections incidence in infants (0.18/1000 live births) showed a decreasing trend (0.3 to 0.1/1000 live births). Type III was overrepresented in infants with invasive infections (n = 35, 58.3%), whereas type V predominated among colonised adults (n = 224, 25.5%) and those with noninvasive (n = 37, 32.5%) and invasive infections (n = 8, 40%). The hypervirulent clone III/ST17 was highly associated with invasive infections (n = 28, 35%), particularly late-onset disease (n = 9, 47.4%), showing an increase from 12.3 to 14.8%. The GBS resistance to erythromycin and clindamycin was 26.7% and 22.1%, respectively, with an upward trend. The emergence of the hypervirulent clone III/ST17 and the escalation in GBS resistance highlight an urgent need for continuous monitoring of GBS infections.

A Laboratory-Based Surveillance Study of Invasive Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae Diseases in a Serbian Pediatric Population-Implications for Vaccination.

The aim of this study was to present the epidemiology of invasive diseases caused by Neisseria meningitidis and Streptococcus pneumoniae in the pre-vaccine period, and Haemophilus influenzae in the post-vaccine period in a pediatric population from Serbia. Among the meningococci, serogroup B dominated (83%), followed by serogroup C (11.3%). High antigenic diversity was found, with fine type P1.5-1,10-4 being the most frequent. Moderate susceptibility to penicillin was common (55%). Within pneumococci, serotypes 19F, 14, 6B, 6A, 18C, 23F, 3, and 7F prevailed, while 19A was rare (3.6%). The coverages of PCV10 and PCV13 were 68% and 84%, respectively. Major sequence types were ST320, ST15, ST273, ST271, and ST81. Non-susceptibility to penicillin (66.7%), cefotaxime (37%), and macrolides (55%) was predominantly detected in vaccine-related serotypes. Among the 11 invasive H. influenzae isolates collected, there were six Hib, three non-type b, and two non-typeable strains (ntHi) that were antibiotic susceptible. These results imply a potential benefit of future Men-B vaccine implementations. For pneumococci, as PCV10 was recently introduced, a significant reduction of morbidity and antibiotic resistance might be expected. The efficiency of Hib vaccination is evident, but a shift towards non-type b and ntHi strains may be anticipated.