Bloodstream infections (BSI) result in significant morbidity and mortality rates, and delayed administration of appropriate antimicrobial treatment is a major predictor of poor outcomes. T2 magnetic resonance (T2MR®) (T2 Biosystems®, Lexington, MA, USA) is an innovative technology that can rapidly identify pathogens from a sample of whole blood in a remarkably short time frame of 3-5 h. We are evaluating if the T2Bacteria Panel (T2BP) contributes to the etiological diagnosis of bloodstream infections when combined with standard blood cultures (BC). The study was performed between December 2018 and March 2019, and a total of 28 patients with suspected BSI were included. The most notable finding of our study was that the addition of T2BP to BC in a diagnostic workflow led to a statistically significant higher rate of T2BP-targeted bacteria identification in patients with suspected BSI (46.4% versus 7.1%, P = 0.001) when compared to BC alone. Considering the measures of diagnostic accuracy, T2BP showed 100.00% sensitivity, 88.24% specificity, 100% negative predictive value (NPV), and 84.62% positive predictive value (PPV). Our findings give valuable insights for microbiologists and clinicians into this molecular method and its advantages in routine diagnostics of BSI.

Clostridioides difficile infection (CDI) is one of the most significant causes of diarrhea in hospitalized patients. The aim of this study was to investigate the incidence and epidemiology of CDI after the COVID-19 pandemic in hospitalized patients in a rehabilitation center in Thessaloniki, Greece. Α retrospective observational cohort study was performed in inpatients diagnosed with diarrhea of all ages (January 2023 - December 2023) who were initially screened for CDI. From the total cohort of patients with proven CDI, some patients were randomly selected based on their monthly isolation incidence throughout the study period, to investigate their epidemiological data and clinical characteristics. Laboratory diagnosis of CDI was performed by enzyme immunoassay, followed by specific anaerobic culture and molecular testing for detection of toxigenic C. difficile. The isolated C. difficile strains were further characterized by PCR ribotyping. The annual incidence of CDI during the study period was 27.1% (130/480). The linear trend of CDI incidence decreased from 32.5% to 18.2% (P = 0.024). The all-cause mortality rate was 5.0% (3/60). A positive correlation was observed between the length of hospital stay and the number of recurrences (r = 0.546, P < 0.001), while 28 patients (46.7%) experienced recurrence of the infection. Seven different PCR ribotypes were identified in this study. C. difficile tcdA+, tcdB+, cdtA+, cdtB+ PCR ribotype 181 (RT181) was the predominant (76.6%, 46/60), followed by toxin A-negative PCR RT017 (11.6%, 7/60). The annual incidence of CDI decreased after the COVID-19 pandemic. Our study demonstrates predominance of C. difficile RT181 with tcdA+, tcdB+, cdtA+, cdtB+ toxin gene profile after COVID-19 pandemic in Northern Greece.

Hepatitis C virus (HCV) is a leading cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma worldwide. HCV has 8 genotypes (GT) and 86 subtypes and distribution of GTs varies based on geographical regions, transmission routes and even in cultural groups. The determination of viral genotype is crucial in choosing antiviral treatment, determining the duration of therapy, and monitoring treatment respose. Since 2014, with the usage of direct-acting antiviral agents (DAAs) in the treatment of HCV infections, a cure rate over 95% could be possible. Epidemiological data are important to combat a chronic HCV infections. Due to its geographical location, Turkey is like a bridge connecting Asia and Europe. Istanbul is the biggest and most crowded city of Turkey and has received immigration from many different countries, especially from Syria, in recent years and immigration still goes on. In addition, the COVID-19 pandemic has had devastating effects in our country. In this study, we determined the HCV genotypes in Health Sciences University Ümraniye Training and Research Hospital, in Istanbul between 2016 and 2022. Of the 322 patients analyzed during this 7-year period, HCV GT1b was the most prevalent GT in 65.2%, followed by GT3 in 15.5%, GT1a in 10.6%. Our data serve as a great mirror for HCV epidemiology in Turkey and contribute to global data.

Urinary tract infections are becoming difficult to treat every year due to antibiotic resistance. Uropathogenic Escherichia coli (UPEC) isolates pose a threat with a combined expression of multidrug-resistance and biofilm formation. ST131 clone is a high-risk pandemic clone due to its strong association with antimicrobial resistance, which has been reported frequently in recent years. This study aims to define risk factors, clinical outcomes, and bacterial genetics associated with ST131/O25b UPEC. In this study, antibiotic susceptibility and species-level identification of 61 clinical E. coli strains were determined by automated systems. Detection of extended-spectrum beta-lactamases was assessed by double-disk synergy test. Biofilm formation was quantified by spectrophotometric method. Virulence genes (iutA, sfa cnf-1, iroN, afa, papA, fimA), antibiotic resistance genes (blaCTX-M, blaTEM, blaSHV, blaOXA, qnrA, qnrB, qnrS, ant(2')-Ia, ant(3)-Ia, aac(3)-IIa, mcr-1, mcr-2, mcr-3, mcr-4) were investigated by PCR. The following beta-lactamase genes were identified, blaTEM (n = 53, 86.8%), blaCTX-M (n = 59, 96.7%), blaSHV (n = 47, 77.0%), and blaOXA-1 (n = 27, 44.2%). Our data revealed that 93.4% of (57/61) E. coli isolates were biofilm-producers. O25pabBspe and trpA2 were investigated for the presence of ST131/O25b clone. Among multidrug resistant isolates, co-existence of O25pabBspe and trpA2 was detected in 29 isolates (47.5%). The fimH30 and H30Rx subclones were detected in four isolates that are strong biofilm-producers. These results suggest that clinical E. coli strains may become reservoirs of virulence and antibiotic resistance genes. This study demonstrates a significant difference in biofilm formation between E. coli ST131 and non-ST131 isolates. Moreover, 86.21% (n = 25) of ST131 isolates produced strong to moderate biofilms, while only 43.75% (n = 14) of non-ST131 isolates showed the ability to form strong biofilms. Presence of iutA and fimA genes in the majority of ST131 strains showed an important role in biofilm formation. These findings suggest application of iutA and fimA gene suppressors in treatment of infections caused by biofilm-producing drug-resistant ST131 strains.

Klebsiella pneumoniae is an opportunistic pathogen and it can cause human mucosal lesions through the intestine, leading to bacteremia and abscess formation in liver and spleen. Previous studies have shown that K. pneumoniae can enter or cross cells through the intestinal epithelium, but the mechanism is unknown. In this study, we treated the intestinal epithelial cell line Caco-2 with KP1195, a clinically isolated strain with high adhesion and invasion of intestinal epithelial cells. The results showed that the treatment of K. pneumoniae could increase the expression of integrin gene and further disrupt the changes of cytoskeleton. Treating Caco-2 with cytoskeletal inhibitor cytorelaxin D can significantly increase the efficiency of K. pneumoniae invading Caco-2 cells. These data suggest that disruption of the cytoskeleton through integrins may be one of the mechanisms by which K. pneumoniae increases intracellular invasion. This study provides a theoretical basis for further understanding of the mechanism of K. pneumoniae entering intestinal epithelial cells.

The aim of this study is to evaluate the antimicrobial susceptibility of invasive isolates of Serratia marcescens, associated with blood stream infections (BSIs) in patients hospitalized in Varna University Hospital, Bulgaria, as well as to identify the genetic mechanisms responsible for 3rd generation cephalosporin and carbapenem-resistance among these isolates. A total of 45 consecutive S. marcescens isolates, obtained from blood cultures of 45 patients with BSIs, hospitalized during an 8-year period (2016-2023) were included. Species identification and antimicrobial susceptibility testing were done by Phoenix (BD, USA) and Vitek 2 (BioMerieux, France) systems and the results were interpreted according to EUCAST guidelines. The genetic mechanisms of beta-lactam resistance were studied by PCR. During the study period, a total of 45 patients were diagnosed with S. marcescens-associated BSIs. All infections were defined as nosocomial, predominantly intensive care unit-acquired (42.2%) and 28.8% were central venous catheter-associated. The following antimicrobial resistance rates were found: ceftriaxone, piperacillin/tazobactam, 57.8%; ceftazidime, 55.6%; cefepime, trimethoprime/sulfamethoxazole, 53.3%; gentamicin, 48.8%; ciprofloxacin, 44.5%; amikacin, 15.6%; carbapenems, 2.2%. The blaCTX-M was identified in 88.9% of the tested 3rd generation cephalosporin resistant isolates. Among these, 50% were also blaTEM positive. The single carbapenem-resistant isolate harboured blaKPC, blaCTX-M1/9, blaCMY-2 and blaTEM. This study demonstrates S. marcescens as a problematic nosocomial pathogen and we report a KPC-producing S. marcescens clinical isolate from a BSI in Bulgaria.

Delftia acidovorans is an aerobic, non-fermenting Gram-negative bacterium (NFGNB), found in soil, water and hospital environments. It is rarely clinically significant, most commonly affecting hospitalized or immunocompromised patients. The present study aimed to explore the genomic characteristics of a Bulgarian clinical D. acidovorans isolate (designated Dac759) in comparison to all strains of this species with available genomes in the NCBI Genome database (n = 34). Dac759 was obtained in 2021 from the sputum of a 65-year-old female immunocompetent outpatient with bronchitis. Species identification using MALDI-TOF mass spectrometry, antimicrobial susceptibility testing, whole-genome sequencing (WGS), and phylogenomic analysis were performed. The isolate demonstrated high-level resistance to colistin (16 mg L-1); resistance to gentamicin; reduced susceptibility to piperacillin, piperacillin-tazobactam, ceftazidime, cefepime, ciprofloxacin, and levofloxacin; and susceptibility to imipenem, meropenem, amikacin, and tobramycin. The observed genome size (6.43 Mb) and GC content (66.76%) were comparable with the accessible data from sequenced D. acidovorans genomes. A limited number of resistance determinants were identified in the assembled genome as follows: blaOXA-459, emrE, oqxB, and mexCD-oprJ. The phylogenomic analysis indicated a high heterogenicity of the included D. acidovorans genomes. In conclusion, to the best of our knowledge, this is the first documented case of a clinically relevant D. acidovorans isolate in Bulgaria. Unlike the majority of reports in the literature, Dac759 affected a patient with no malignancies or other preexisting comorbidities. With this in mind, its genome sequence is a valuable resource for the fundamental study of uncommon bacterial pathogens of public health importance.

The Acidaminococcus genus is a part of the normal flora in gastrointestinal tract. It is a strictly anaerob Gram-negative coccus that is rarely pathogenic. We report the case of a 58-year-old man, who presented to surgery department A of the Charles Nicolle hospital, complaining of a wide inflammatory lesion in the anterior abdominal wall evolving for two weeks. Patient's anamnestic data included smoking, hypertension, and diabetes mellitus with poor compliance. The patient underwent flattening with excision of necrotic tissues and surgical drainage using a DELBET blade. Empirical antibiotic therapy with imipenem 1gx3/d, teicoplanin 400 mg 1 inj x2/d and gentamicin 400 mg 1 inj/d was administered pending bacteriological results. The bacteriological examination of a sample of necrotic tissue, after 72 h of incubation at 37 °C in anaerobic atmosphere, was able to detect a Gram-negative coccus, that the VITEK2 ANC system identified as Actinomyces canis with an accuracy of 80%. Whole genome sequencing was subsequently performed, that identified Acidaminococcus sp. AM33-14BH and demonstrated the following resistance genes: cfxa, tet(X) and tet(Q). An antibiogram for anaerobes was performed showing that the strain was resistant to amoxicillin but sensitive to amoxicillin-clavulanic acid, piperacillin-tazobactam, ertapenem, imipenem, meropenem and rifampin. Patient's condition improved after treatment with imipenem for 2 weeks, followed by oral amoxicillin-clavulanic acid for 16 days.This work highlights the role of molecular biology in the diagnosis of infections caused by anaerobes. Although the Vitek 2 ANC card provides rapid and acceptable identification of the most common anaerobic bacteria, improvements are needed for the identification of bacteria in the genera Acidaminococcus and Actinomyces.

Hospital acquired infections greatly affect recovery and survival in pediatric surgical patients. We evaluated prevalence and antimicrobial resistance of ESKAPE pathogens in neonates and infants subjected to cardiac surgery in a tertiary hospital in Central Kazakhstan between 2019 and 2023 (2,278 patients) using routine methods of microbiological detection. ESKAPE pathogens were found in 1,899 out of 2,957 samples (Staphylococcus aureus - 35.3%, Klebsiella pneumoniae - 27.8%, Acinetobacter baumannii - 14.5%, Pseudomonas aeruginosa - 12.4%, Enterobacter sp. - 8.8%, Enterococcus faecium - 1.2%). The total prevalence of ESKAPE increased significantly from 45.1 to 76.9% (P = 0.005) during the study period. The resistance significantly increased in methicillin-resistant S. aureus (MRSA, from 13.7 to 41.9%, P = 0.041) but decreased in carbapenem-resistant P. aeruginosa (from 64.3 to 37.7%, P = 0.037) and carbapenem-resistant A. baumannii (from 48.5 to 19.1%, P = 0.039). Gradual but non-significant changes were shown in third-generation cephalosporin resistant K. pneumoniae (from 63.6 to 45.2%) and carbapenem-resistant K. pneumoniae (from 0 to 8.3%). The relative prevalence of ESKAPE pathogens steadily increased in our pediatric cardiac surgery patients in 2019-2023. The most frequent were S. aureus, K. pneumoniae, and A. baumannii, with dramatically increasing tendencies for MRSA. Our results highlight the necessity for a well-designed infection control strategy and constant microbiological monitoring in pediatric cardiac surgery departments.

We report a nosocomial outbreak caused by a multidrug-resistant carbapenemase-producing Klebsiella pneumoniae (MDRCPKp), that was detected in six patients admitted to the medical intensive care unit between 20th of December 2023 and 15th of January 2024 in Ankara, Turkey. The investigation of this outbreak was started on 29th of December 2023. During the outbreak 11 samples were collected from the six patients with MDRCPKp. Pulsed-field gel electrophoresis (PFGE) was performed to determine the genetic relatedness and clonality of MDRCPKp strains. MDRCPKp was isolated in the tracheal aspiration culture, blood, urine, and screening samples. Five patients with MDRCPKp colonization developed healthcare-associated infection. In one patient MDRCPKp was isolated from tracheal aspirate and the screening cultures were considered as colonization not infection. PFGE analysis revealed that all isolates belonged to the same K. pneumoniae clone. MDRCPKp strain of this outbreak exhibited multidrug resistance and co-produced OXA-48 and NDM-1. This outbreak ended after application of strict infection control measures. An outbreak of MDRCPKp can occur in hospitals, especially in the intensive care units; thus, it should be detected early by infection control teams. A strong collaboration between infection control team and microbiology laboratory is essential to cope with MDR bacterial outbreaks in hospitals.

This study examined the prevalence and antibiotic resistance pattern of blaCTX-M extended-spectrum β-lactamase positive Salmonella species isolated from a hospital in Weifang. Salmonella strains were isolated from hospitalized patients from January 2018 to April 2023. Whole-genome sequencing was performed by Illumina platform. CTX-M-producing Salmonella were identified by Comprehensive Antibiotic Research Database (CARD). Strain susceptibility to six antimicrobial agents was assessed by BD Phoenix™ M50 System. MLST analysis confirmed sequence types and additionally, serotypes were determined by SeqSero2. Genetic environments of blaCTX-M genes were analyzed by Isfinder and BLASTn. Single nucleotide polymorphisms were used to construct a phylogenetic tree to analyze homology. A total of 34 CTX-M-producing Salmonella were detected. The most prevalent serotype was Salmonella enterica subsp. enterica 1,4,[5],12:i:- (14/34, 41.18%), belonging to ST34, followed by Salmonella Enteritidis (10/34, 29.41%), belonging to ST11. The highest resistance rate was detected to ampicillin (97.06%), followed by ceftriaxone (94.12%) and ceftazidime (58.83%). In CTX-M-producing Salmonella five types of blaCTX-M genes were identified, the most prevalent was blaCTX-M-55 (47.06%, 16/34), followed by blaCTX-M-14, blaCTX-M-65, blaCTX-M-125, and blaCTX-M-27 at 26.47% (9/34), 11.77% (4/34), 8.82% (3/34), and 5.88% (2/34), respectively. Apart from blaCTX-M, 40 antibiotic resistance genes were also detected, conveying resistance to multiple drugs and the most frequent genes were namely, mcr-1.1, aph(6)-Id, aph(3″)-Ib, oqxAB, qnrB6, qnrS1. According to genetic environment analysis, the insertion sequence ISEcp1 was prevalent upstream of the blaCTX-M gene. Our study demonstrates that multiple resistance genes are carried by clinical isolates of Salmonella spp. however, the dominant ESBL genotype is CTX-M-55, that is associated with ISEcp1.

Antibiotic resistance constitutes a significant public health challenge, with diverse reservoirs of resistant bacteria playing pivotal roles in their dissemination. Among these reservoirs, pets are carrying antibiotic-resistant strains. The objective of this study was to assess the resistance profiles of Escherichia coli, and the prevalence of extended-spectrum β-lactamase (ESBL) producing E. coli strains in dogs and cats from Tamaulipas, Mexico. A total of 300 stool samples (150 dogs and 150 cats) from healthy pets were subjected to analysis. Antibiotic susceptibility testing and the identification of ESBLs were carried out by disc diffusion method. The presence of resistance genes, class 1, 2, and 3 integrons (intI1, intI2, and intI3) and phylogroups was determined by PCR analysis. The findings reveal that 42.6% (128/300) of the strains exhibited resistance to at least one of the eight antibiotics assessed, and 18.6% (56/300) demonstrated multidrug resistance (MDR), that distributed across 69 distinct resistance patterns. Altogether 2.6% of E. coli strains (8/300) were confirmed as TEM and CTX-M type ESBL producers. These outcomes underscore the roles of dogs and cats in Tamaulipas as reservoirs for the dissemination of MDR and/or ESBL strains. The results underscore the necessity for conducting prevalence studies on ESBL-producing E. coli, forming a foundation for comprehending the present scenario and formulating strategies for the control and mitigation of this issue.

Escherichia coli A382 was isolated in July 2024 from a positive blood culture obtained from the central venous catheter of a male patient undergoing chemotherapy at the Hospital of Volos, Thessaly, Greece. Whole-genome sequencing analysis revealed that the isolate A382 is E. coli belonging to the ST410 high-risk clone, which co-harbors the blaKPC-3 and blaSHV-182 genes on an IncX3 plasmid. It also harbors blaTEM-1 and has five replicons, as follows: IncX3, IncQ1, CoIRNAI, IncF1A, and IncFIB. Complete genome analysis revealed that E. coli A382 isolate carries a range of virulence factors (iutA, iucC, fimH, fdeC, yehA, yehD, yehC, yehB, cgs, ahha, ccI, hlyE, papC, irp2, fyuA, lpfA, and nlpl) and many other non-beta-lactam resistance determinants, including dfrA14 and sul2, but it is susceptible to aminoglycosides, nitrofurantoin, tigecycline, colistin and ceftazidime-avibactam. In conclusion in this study, we describe the phenotypic and genome characteristics of an extensively drug-resistant E. coli ST410.

Oral Squamous cell carcinoma (OSCC) is the 14th most frequent cancer with 300,000 new cases and 100,000 deaths reported annually. Even with advanced therapy, the treatment outcomes are poor at advanced stages of the disease. The diagnosis of early OSCC is of paramount clinical value given the high mortality rate associated with the late stages of the disease. Recently, the role of microbiome in the disease manifestation, including oral cancer, has garnered considerable attention. But, to establish the role of bacteria in oral cancer, it is important to determine the differences in the colonization pattern in non-tumour and tumour tissues. In this study, 16S rRNA based metagenomic analyses of 13 tumorous and contralateral anatomically matched normal tissue biopsies, obtained from patients with advanced stage of OSCC were evaluated to understand the correlation between OSCC and oral microbiome. In this study we identified Fusobacterium, Prevotella, Capnocytophaga, Leptotrichia, Peptostreptococcus, Parvimonas and Bacteroidetes as the most significantly enriched taxa in OSCC lesions compared to the non-cancerous tissues. Further, PICRUSt2 analysis unveiled enhanced expression of metabolic pathways associated with L-lysine fermentation, pyruvate fermentation, and isoleucine biosynthesis in those microbes associated with OSCC tissues. These findings provide valuable insights into the distinctive microbial signatures associated with OSCC, offering potential biomarkers and metabolic pathways underlying OSCC pathogenesis. While our focus has primarily centred on microbial signatures, it is essential to recognize the pivotal role of host factors such as immune responses, genetic predisposition, and the oral microenvironment in shaping OSCC development and microbiome composition.

Although the relationship between vaginal microorganisms and fertility has been well established, only few studies have investigated vaginal microorganisms in women undergoing in vitro fertilization (IVF). Our aim was to study the differences in vaginal microbiota between infertile women with repeated implantation failure (RIF) and those who achieved clinical pregnancy in their first frozen embryo transfer cycle. We compared the vaginal microbiota of patients with a history of RIF (n = 37) with that of the control group (n = 43). Following DNA extraction, metagenomic sequencing was employed for the analysis of alpha and beta diversities, distinctions in bacterial species, and the functional annotation of microbial genes. Furthermore, disparities between the two groups were revealed. Alpha diversity analysis revealed that the Shannon index was higher in the RIF group (P < 0.05). There were differences in the beta diversity between groups (P = 0.16). At the bacterial family level, the relative abundance of Actinomycetaceae (P = 0.013) and Ruminococcaceae (P = 0.013) were significantly higher in the RIF group. At the genus level, the abundances of Actinomyces (P = 0.028) and Subdoligranulum (P = 0.013) were significantly higher in the RIF group. At the species level, the abundances of Prevotella timonensis (P = 0.028), Lactobacillus jensenii (P = 0.049), and Subdoligranulum (P = 0.013) were significantly higher in the RIF group. Significant differences in family, genus, species, alpha and beta diversity were observed in the vaginal microbiota between groups. Notably, among these findings, the Subdoligranulum genus emerged as the most prominent correlating factor.

Klebsiella pneumoniae is one of the most prevalent bacteria causing urinary tract infections (UTIs). Its increasing resistance to a wide array of antibiotics limits available treatment options. This study investigated the characteristics and trends of antimicrobial resistance in K. pneumoniae isolated from UTIs in Crete, Greece, during 2017 and 2022. Among the 11,946 Enterobacteriaceae isolated from urine specimens, a total of 1,771 K. pneumoniae isolates were identified (14.8%), with an isolation frequency secondary to Escherichia coli (66.3%). K. pneumoniae isolates increased over the years, with a peak in the year 2022. Higher resistance rates were detected in ciprofloxacin (41%), trimethoprim/sulfamethoxazole (TMP/SMX) (38.1%) and nitrofurantoin (33.9%). Resistance to ciprofloxacin, amoxicillin/clavulanic acid, tigecycline, and TMP/SMX significantly increased from 33.7%, 24%, 6%, and 33.1%, respectively, over the years 2017-2019, to 47.8%, 34.2%, 14.3% and 42.8%, respectively, over the period 2020-2022. ESBL production and carbapenem resistance were decreased by 2.2% and 3.7%, respectively, over the two three-year periods (2017-2019 and 2020-2022). Among the 278 carbapenem-resistant K. pneumoniae (CRKP) isolates, 164 (59%), 66 (23.7%), 18 (6.5%) and 16 (5.8%) were positive for KPC, NDM, VIM and OXA-48 enzymes, respectively. Only 14 (5%) isolates harboured two carbapenemase genes, namely 10 (3.6%) both blaNDM and blaVIM, and 4 (1.4%) both blaKPC and blaNDM. Females, inpatients and the elderly were more frequently affected by CRKP. The frequency of multidrug-resistant (MDR) and extensively drug-resistant (XDR) isolates were 32.6% and 7.7%, respectively. Continuous surveillance of local microbial prevalence and monitoring of antimicrobial resistance patterns provide critical information to guide the empiric therapy for UTIs and control the spread of MDR bacteria.

Carbapenem-resistant Enterobacterales (CRE) have become a major public health problem worldwide. The aim of this study was to investigate efficacy of ceftazidime/avibactam and plazomicin on carbapenem-resistant Klebsiella pneumoniae and Escherichia coli isolates. Susceptibility of imipenem, meropenem, ertapenem, ceftazidime/avibactam and plazomicin was investigated by broth-microdilution method. Major carbapenemases NDM, VIM, IMP, KPC, OXA-48 as well as other β-lactamases namely, TEM, SHV, OXA-1-like, CTX-M, ACC, FOX, MOX, DHA, CIT, EBC, VEB, GES, PER were investigated by PCR. A total of 120 carbapenem-resistant isolates (60 E. coli and 60 K. pneumoniae) were included in this study and blaOXA-48-like was found in 78.33%, blaNDM in 26.66%, blaKPC in 7.5%, blaIMP in 5.83%, and blaVIM in 5%. Among 94 isolates with the blaOXA-48-like gene, 22.3% were resistant to ceftazidime/avibactam and 51.1% were resistant to plazomicin. Of 32 isolates with blaNDM, 31 (96.9%) were resistant to ceftazidime/avibactam and 30 (93.75%) were resistant to plazomicin, and both antibiotics had limited effects against blaNDM carriers (P < 0.001). Of the 12 isolates with blaNDM+OXA-48 combination, 11 (91.7%) were resistant to ceftazidime/avibactam and plazomicin. The effect of both antibiotics was significantly lower in strains with blaNDM+OXA-48 combination (P < 0.005).The most common carbapenemase genes in this study were blaOXA-48-like and blaNDM. Ceftazidime/avibactam demonstrated a good efficacy among OXA-48 producing K. pneumoniae and E. coli, however, plazomicin had a significantly lower antibacterial effect in our study. Both antimicrobial agents should be considered as an option by evaluating combined susceptibility results and gene patterns obtained by regional and global molecular data in the treatment of CRE infections.

The rate of pandrug-resistant Acinetobacter baumannii strains is on the rise in all continents. This bacterium can acquire resistance to all antibiotics, even to colistin. Alterations in the lipid A or/and the two-component pmrAB were earlier detected in colistin resistance. We investigated and analyzed two strains of A. baumannii (ABRC1 and ABRC2) isolated from two patients admitted to intensive care unit with a septic shock. Both strains were resistant to all tested antibiotics including colistin with a MIC >256 mg L-1. Colistin resistance genes (pmrA, pmrB, lpxA, lpxC, lpxD, and lpsB) of two strains (ABRC1 and ABRC2) were investigated by PCR and sequencing. Obtained nucleic acid sequences were aligned with reference sequences of ATCC 19606 and 17987. In this study two amino acid mutations, N287D in the lpxC gene and E117K in the lpxD gene, were detected in both ABRC1 and ABRC2 strains. ABRC1 had an additional H200L mutation in the pmrA gene. Both colistin resistant strains harbored the same A138T mutation in the pmrB gene. The ABRC2 strain also had an alteration in the kinase domain, specifically an R263S substitution of the histidine kinase domain. Three identical mutations were found in the lpsB gene of both A. baumannii strains: Q216K + H218G + S219E. As a result, a newly deduced protein sequence in both ABRC1 and ABRC2 strains differed from those described in ATCC 17978 and 19606 strains was determined. Colistin resistance is multifactorial in A. baumannii. In our study we detected novel mutations in colistin resistant A. baumannii clinical isolates.

The present study aimed to explore the genomic characteristics of eight New Delhi metallo-β-lactamase-1 (NDM-1)-producing carbapenem-resistant Pseudomonas aeruginosa (CRPA) isolates from a Bulgarian tertiary hospital (2021-2023) in comparison to blaNDM-1-positive strains originating from the Balkans. Antimicrobial susceptibility testing, phenotypic assays for carbapenemase activity, PCR screening, whole-genome sequencing (WGS), and phylogenomic analysis were performed. Seven of the CRPA isolates investigated (Minimum inhibitory concentration values of imipenem and meropenem >32 mg L-1) were also resistant to piperacillin-tazobactam, ceftazidime, ceftazidime-avibactam, cefepime, ceftolozane-tazobactam, amikacin, tobramycin, ciprofloxacin, and levofloxacin, but were susceptible to colistin (0.5-2 mg L-1) and cefiderocol (0.25-1 mg L-1). The P. aeruginosa Pae57 isolate (designated Pae57) remained susceptible to aminoglycosides as well. WGS uncovered the co-existence of blaNDM-1 and blaGES-1. The isolates belonged to the ST654 high-risk clone, except for Pae57 (ST611). Alignment against reference sequences revealed the presence of a Tn21 transposon harboring bleMBL-blaNDM-1-ISAba125. It was similar to that found in the P. aeruginosa ST654 NDM1_1 strain (GCA_020404785.1) from Serbia. Phylogenomic analysis of our isolates indicated that seven of them (ST654) differed from each other in no more than 44 single-nucleotide polymorphisms (SNPs). Pae57 (ST611) was strikingly different (>21,700 SNPs) compared to all Balkan strains. In conclusion, to our knowledge this is the first report of blaNDM-1-positive P. aeruginosa ST611 isolation, which indicates the transmission dynamics of this determinant between high-risk and potentially high-risk P. aeruginosa clones. Obtained results unveil the dissemination of clonally related NDM-1-producing P. aeruginosa strains in the monitored hospital for approximately a 2-year period.

Pseudomonas aeruginosa has been in the center of attention for several years as an opportunistic human pathogen implicated in many severe acute and chronic infections particularly in immunocompromised patients. Its high persistence and resistance against many antimicrobial agents are mostly attributed to biofilm formation. Biofilms are microbial communities mainly consisting of extracellular polymeric substances that encapsulate bacteria together and protect them from extracellular stresses. This cell aggregation is a stress response that P. aeruginosa employes as a survival strategy during growth with the toxic detergents. This process has shown to involve several operons such as psl, pel, and alg. Here we used P. aeruginosa strain PAO1 in control group, 40 P. aeruginosa strains from sink and 40 strains from surface of public places. Biofilm formation and gene expression were measured before and after exposure to sub minimum inhibitory concentration (sub-MIC) of biocides chlorhexidine diacetate and benzalkonium chloride. The qRT-PCR and biofilm formation results demonstrated an increase in biofilm formation ability and gene expression of pslA/B and pelA/B in two groups collected from sink and surface in contrast to the control group. A remarkable increase was observed in the biofilm formation and expression of pslA in the bacterial strain collected from the sink after exposure to biocides chlorhexidine diacetate. Both Pel and Psl appeared to have redundant functions as structural scaffolds in biofilms. Sub-MIC levels of detergents can improve biofilm formation ability of P. aeruginosa and therefore trigger resistance.

The aim of this prospective pilot study was to compare culture and microbiome results of the removed tonsils of patients with assumed distant focal disease (11 patients) and those who underwent a tonsillectomy, due to other reasons, such as recurrent tonsillitis, tonsil stones or snoring (nine patients). Aerobic culture was carried out for samples taken from the surface of the tonsils by swabs before tonsillectomy for all 20 patients. The squeezed detritus and the tissue samples of removed tonsils, taken separately for the right and left tonsils, were incubated aerobically and anaerobically. The microbiome composition of tissue samples of removed tonsils was also evaluated. Based on the culture results of the deep samples Staphylococcus aureus was the dominating pathogen, besides a great variety of anaerobic and facultative anaerobic bacteria present in the oral microbiota in those patients who underwent tonsillectomy due to distant focal diseases. Microbiome study of the core tissue samples showed a great diversity on genus and species level among patients of the two groups however, S. aureus and Prevotella nigrescens were present in higher proportion in those, whose tonsils were removed due to distant focal diseases. Our results may support previous findings about the possible triggering role of S. aureus and P. nigrescens leading to distant focal diseases. Samples taken by squeezing the tonsils could give more information about the possible pathogenic/triggering bacteria than the surface samples cultured only aerobically.