Characterization, modes of interactions with DNA/BSA biomolecules and anti-tumor activity of newly synthesized dinuclear platinum(II) complexes with pyridazine bridging ligand.

Platinum-based drugs are widely recognized efficient anti-tumor agents, but faced with multiple undesirable effects. Here, four dinuclear platinum(II) complexes, [{Pt(1,2-pn)Cl}2(µ-pydz)]Cl2 (C1), [{Pt(ibn)Cl}2(µ-pydz)]Cl2 (C2), [{Pt(1,3-pn)Cl}2(µ-pydz)]Cl2 (C3) and [{Pt(1,3-pnd)Cl}2(µ-pydz)]Cl2 (C4), were designed (pydz is pyridazine, 1,2-pn is ( ±)-1,2-propylenediamine, ibn is 1,2-diamino-2-methylpropane, 1,3-pn is 1,3-propylenediamine, and 1,3-pnd is 1,3-pentanediamine). Interactions and binding ability of C1-C4 complexes with calf thymus DNA (CT-DNA) has been monitored by viscosity measurements, UV-Vis, fluorescence emission spectroscopy and molecular docking. Binding affinities of C1-C4 complexes to the bovine serum albumin (BSA) has been monitored by fluorescence emission spectroscopy. The tested complexes exhibit variable cytotoxicity toward different mouse and human tumor cell lines. C2 shows the most potent cytotoxicity, especially against mouse (4T1) and human (MDA-MD468) breast cancer cells in the dose- and time-dependent manner. C2 induces 4T1 and MDA-MD468 cells apoptosis, further documented by the accumulation of cells at sub-G1 phase of cell cycle and increase of executive caspase 3 and caspase 9 levels in 4T1 cells. C2 exhibits anti-proliferative effect through the reduction of cyclin D3 and cyclin E expression and elevation of inhibitor p27 level. Also, C2 downregulates c-Myc and phosphorylated AKT, oncogenes involved in the control of tumor cell proliferation and death. In order to measure the amount of platinum(II) complexes taken up by the cells, the cellular platinum content were quantified. However, C2 failed to inhibit mouse breast cancer growth in vivo. Chemical modifications of tested platinum(II) complexes might be a valuable approach for the improvement of their anti-tumor activity, especially effects in vivo.

In vitro activity of imipenem/relebactam and ceftazidime/avibactam against carbapenem-resistant Klebsiella pneumoniae from blood cultures in a University hospital in Serbia.

The study aimed to investigate prevalence of carbapenem-resistant Klebsiella pneumoniae (CRKP) blood culture isolates and their susceptibility to two new antibiotics, imipenem/relebactam and ceftazidime/avibactam. Out of 765 isolates recovered from blood cultures in a tertiary care hospital in Serbia between 2020 and 2023, 143 non-repetitive K. pneumoniae strains were included in this study. Minimum inhibitory concentration (MIC) values of the examined antimicrobial drugs was determined by VITEK 2 system, MIC test strip (imipenem/relebactam and ceftazidime/avibactam), and broth microdilution method (tigecycline and colistin). Carbapenemase-encoding genes (blaKPC, blaOXA-48-like, blaNDM, blaVIM, blaIMP) were detected using a multiplex-PCR assay, the BioFire-Blood Culture Identification 2-panel. This closed molecular assay is designed for the BioFire® FilmArray® system, enabling automated sample preparation, amplification, detection, and analysis (bioMérieux, France). Results revealed that K. pneumoniae was the most common isolate from blood cultures in 2022. The prevalence of K. pneumoniae was about 11.6% in 2020 and 2021, while in 2022 it raised to over 30%. Also, the frequency of CRKP increased from 11.76% in 2020, through 15.29% in 2021 to 72.94% in 2022. The majority of CRKP carried blaOXA-48-like (60.0%), followed by blaKPC (16.47%), and blaNDM (8.24%) genes, while 14.12% harboured both blaOXA-48-like and blaNDM genes. Only 25.88% of CRKP isolates were resistant to ceftazidime/avibactam, while 51.76% were resistant to imipenem/relebactam and colistin. The rapid spread of CRKP is particularly concerning because therapeutic options are limited to a few antibiotics. While imipenem/relebactam and colistin showed similar antimicrobial activity against CRKP clinical isolates, ceftazidime/avibactam proved to be the most effective antibiotic.

Prevalence of multidrug-resistant Gram-negative bacteria from blood cultures and rapid detection of beta-lactamase-encoding genes by multiplex PCR assay.

Introduction: This study aimed to determine the prevalence of multidrug-resistant Gram-negative bacteria (GNB) from blood cultures in a tertiary-care hospital and the multiplex PCR assay's ability to detect resistance genes. Methods: A total of 388 GNB isolates obtained from hospitalized patients between November 2019 and November 2021 were included in the study. Antimicrobial susceptibility testing was done by VITEK 2 system and broth microdilution method. Beta-lactamase-encoding genes were detected by multiplex PCR assays, BioFire-Blood Culture Identification 2 (BCID2) panel (bioMérieux, France). Extended-spectrum beta-lactamases (ESBLs) were detected phenotypically with VITEK AST-GN71 card (bioMérieux, France). The isolates of GNB were classified into multidrug-resistant, extensively-drug-resistant, and pandrug-resistant categories, and their prevalence and distribution in different wards, including coronavirus diseases 2019 (COVID-19) intensive care units (ICU), were calculated. Results: Results revealed that all isolates of Acinetobacter baumannii and Pseudomonas aeruginosa were multidrug-resistant as well as 91.6% of Enterobacter cloacae, 80.6% of Proteus mirabilis, and 76.1% of Klebsiella pneumoniae, respectively. In fermentative bacteria, blaOXA-48-like (58.1%), blaNDM (16.1%), blaKPC (9.7%) and blaVIM (6.5%) genes were detected. More than half of Enterobacter cloacae (58.3%) and Klebsiella pneumoniae (53.7%) produced ESBLs. Among non-fermenters, the blaNDM gene was carried by 55% of Pseudomonas aeruginosa and 19.5% of Acinetobacter baumannii. In the COVID-19 ICU, Acinetobacter baumannii was the most common isolate (86.1%). Conclusions: This study revealed high proportions of multidrug-resistant blood isolates and various underlying resistance genes in Gram-negative strains. The BCID2 panel seems to be helpful for the detection of the most prevalent resistance genes of fermentative bacteria.

An outbreak of Legionnaires' disease in newborns in Serbia.

Legionnaires' disease is an atypical pneumonia caused by inhaling small droplets of water containing the bacterium Legionella spp. In newborns, it is a rare event, usually associated with water births and the use of air conditioning systems or air humidifiers. A nosocomial outbreak of Legionnaires' disease in the maternity ward of a secondary-care hospital in Arandjelovac, Serbia is described.Two male newborns were found to be infected with Legionnella pneumophila. On Days 7 and 6 of life, respectively, they were transferred to a tertiary-care hospital with signs of severe pneumonia which was radiologically confirmed. L. pneumophila was detected in tracheal secretions from both infants by RT-PCR, and its antigens were also positive in urine samples. The source of infection in the secondary-care hospital was the internal hot water heating system, and the main contributory factor to the emergence of the infection was the low temperature of the hot water which did not kill the bacteria during the available exposure time.These two cases highlight the importance of being cautious about possible Legionnaires' disease in maternity wards with outdated or irregularly maintained internal water supply systems. The adoption of official guidelines for the control and regular maintenance of water supply systems, including the multidisciplinary cooperation of all relevant parties, forms the basis for direct monitoring for Legionella and the prevention of new outbreaks.Abbreviations: BCYE: buffered charcoal yeast extract agar; GVPC: glycine vancomycin polymyxin cycloheximide agar; LD - Legionnaires' disease; TR-PCR: Reverse transcription polymerase chain reaction.

The first nationwide multicenter study of Acinetobacter baumannii recovered in Serbia: emergence of OXA-72, OXA-23 and NDM-1-producing isolates.

BACKGROUND: The worldwide emergence and clonal spread of carbapenem-resistant Acinetobacter baumannii (CRAB) is of great concern. The aim of this nationwide study was to investigate the prevalence of CRAB isolates in Serbia and to characterize underlying resistance mechanisms and their genetic relatedness. METHODS: Non-redundant clinical samples obtained from hospitalized patients throughout Serbia were included in the prospective, observational, multicenter study conducted from January to June 2018. Samples were initially screened for the presence of Acinetobacter baumannii-calcoaceticus (Acb) complex using conventional bacteriological techniques. Acb complexes recovered from clinical samples obtained from inpatients with confirmed bacterial infections were further evaluated for the presence of A. baumannii. Identification to the species level was done by the detection of the blaOXA-51 gene and rpoB gene sequence analysis. Susceptibility testing was done by disk diffusion and broth microdilution method. CRAB isolates were tested for the presence of acquired carbapenemases (blaOXA-24-like, blaOXA-23-like,blaOXA-58-like, blaOXA-143-like, blaIMP, blaVIM, blaGIM, blaSPM, blaSIM, blaNDM) by PCR. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS: Acb complex was isolated in 280 out of 2401 clinical samples (11.6%). Overall, A. baumannii was identified in 237 out of 280 Acb complex (84.6%). CRAB prevalence was found to be 93.7% (237/222). The MIC50/MIC90 for imipenem and meropenem were 8/> 32 µg/mL and 16/> 32 µg/mL, respectively. Although susceptibility was high for colistin (95.7%; n = 227) and tigecycline (75.1%; n = 178), ten isolates (4.3%) were classified as pandrug-resistant. The following carbapenemases-encoding genes were found: 98 (44.2%) blaOXA-24-like, 76 (34.5%) blaOXA-23-like, and 7 (3.2%) blaNDM-1. PFGE analysis revealed six different clusters. MLST analysis identified three STs: ST2 (n = 13), ST492 (n = 14), and ST636 (n = 10). Obtained results evaluated that circulating CRAB clones in Serbia were as follows: blaOXA66/blaOXA23/ST2 (32.4%), blaOXA66/blaOXA23/blaOXA72/ST2 (2.7%), blaOXA66/blaOXA72/ST492 (37.8%), and blaOXA66/blaOXA72/ST636 (27.1%). CONCLUSION: This study revealed extremely high proportions of carbapenem resistance among A. baumannii clinical isolates due to the emergence of blaOXA-72, blaOXA-23, and blaNDM-1 genes among CRAB isolates in Serbia and their clonal propagation.