Background: Candida glabrata is the third leading cause of candidaemia in Turkey; however, the data regarding antifungal resistance mechanisms and genotypic diversity in association with their clinical implication are limited.Objectives: To assess genotypic diversity, antifungal susceptibility and mechanisms of drug resistance of C glabrata blood isolates and their association with patients' outcome in a retrospective multicentre study.Patients/Methods: Isolates from 107 patients were identified by ITS sequencing and analysed by multilocus microsatellite typing, antifungal susceptibility testing, and sequencing of PDR1 and FKS1/2 hotspots (HSs).Results: Candida glabrata prevalence in Ege University Hospital was twofold higher in 2014‐2019 than in 2005‐2014. Six of the analysed isolates had fluconazole MICs ≥ 32 µg/mL; of them, five harboured unique PDR1 mutations. Although echinocandin resistance was not detected, three isolates had mutations in HS1‐Fks1 (S629T, n = 1) and HS1‐Fks2 (S663P, n = 2); one of the latter was also fluconazole‐resistant. All patients infected with isolates carrying HS‐FKS mutations and/or demonstrating fluconazole MIC ≥ 32 µg/mL (except one without clinical data) showed therapeutic failure (TF) with echinocandin and fluconazole; seven such isolates were collected in Ege (n = 4) and Gulhane (n = 3) hospitals and six detected recently. Among 34 identified genotypes, none were associated with mortality or enriched for fluconazole‐resistant isolates.Conclusion: Antifungal susceptibility testing should be supplemented with HS‐FKS sequencing to predict TF for echinocandins, whereas fluconazole MIC ≥ 32 µg/mL may predict TF. Recent emergence of C glabrata isolates associated with antifungal TF warrants future comprehensive prospective studies in Turkey.

Infections caused by multi drug-resistant gram-negative bacilli are increasingly reported worldwide. Colistin, tigecycline and aminoglycosides are almost the only and last choice antibiotics in the treatment of infections caused by carbapenem-resistant Enterobacterales members. Ceftazidime-avibactam is a novel antibiotic combination consisting of a broad-spectrum cephalosporin and avibactam with good antimicrobial activity against carbapenem-resistant Enterobacterales members. The aim of this study was to assess the in vitro activity of ceftazidime-avibactam and colistin against carbapenem-resistant Klebsiella pneumoniae isolates and to obtain local antimicrobial surveillance data. A total of 150 carbapenem-resistant K.pneumoniae isolates obtained from various clinical samples of the patients hospitalized in our hospital between 2018-2021 were included in the study. Duplicate isolates were excluded from the study. The isolates were recovered from blood (n= 72), tracheal aspirate (n= 40), wound (n= 20), biopsy and abscess (n= 10), steril body fluid (n= 5), and peripheral venous catheter (n= 3) samples. Isolates were identified by matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS, Bruker Daltonics, Germany). The minimum inhibitory concentration (MIC) values of the isolates for meropenem, colistin, ceftazidime, and ceftazidime-avibactam were determined by broth microdilution method. Susceptibility of the isolates to the tested antibiotics was evaluated by the European Committee of Antimicrobial Susceptibility Testing (EUCAST) criteria. The presence of carbapenemases (VIM, IMP, NDM, KPC, and OXA-48) was investigated by polymerase chain reaction (PCR) using specific primers. The mcr-1, mcr-2, mcr-3, mcr-4, and mcr-5 genes were evaluated by PCR for plasmid-mediated colistin resistance. All K.pneumoniae isolates were found to be positive for at least one of the carbapenemase genes evaluated in the study. The blaOXA-48 gene was detected in 107 (71.3%), blaKPC gene in 25 (16.7%); blaNDM gene in 7 (4.7%), co-production of blaOXA-48 and blaKPC genes in 10 (6.7%), co-production of blaOXA-48 and blaNDM genes in 1 (0.6%) isolate. None of the isolates harbored the blaVIM and blaIMP genes. None of the mcr genes screened in the study were detected among the isolates. The susceptibility of the isolates to ceftazidime-avibactam and colistin was 92.7% (139/150) and 48% (72/150), respectively. The MIC50 and MIC90 values for meropenem, ceftazidime, ceftazidime-avibactam, and colistin of the isolates were determined as 32/256, > 128/> 128, 1/8, and 4/16 µg/ml, respectively. Of the ceftazidimeavibactam resistant isolates, seven were positive for blaNDM, three for blaKPC, and one for both blaOXA-48 and blaNDM genes. High ceftazidime-avibactam MIC levels (> 128 µg/ml) were detected in metallo-betalactamase producing isolates. Consequently, our data suggested that ceftazidime-avibactam exhibited as a good alternative therapeutic choice for carbapenem-resistant K.pneumoniae isolates. It is noteworthy that high rate of colistin resistance was detected in K.pneumoniae isolates. Another notable finding of this study is the increase in K.pneumoniae isolates producing blaKPC for our country. To prevent the development of resistance which is observed even in last-choice therapeutic antibiotics, the principles of rational antibiotic use should be followed. The appropriate antimicrobial susceptibility testing should be routinely performed for surveillance of ceftazidime-avibactam and colistin.

Colistin, one of the last-line treatment options for carbapenem-resistant microorganisms, presents challenges in determining its antimicrobial susceptibility. This study aims to compare colistin broth disk elution (CBDE), modified CBDE (mCBDE), the colistin agar test (CAT), and rapid polymyxin NP (RPNP) tests with the reference broth microdilution (BMD) in determining colistin susceptibility of 116 multidrug-resistant (MDR) Klebsiella pneumoniae isolates. Isolates were collected from blood and respiratory tract samples of different patients between 2022 and 2023 in Türkiye. For each isolate, a single 0.5 McFarland inoculum was prepared to inoculate CBDE, mCBDE, and CAT in parallel, then the suspension was adjusted to a density of 3-3.5 for RPNP. All tests were conducted simultaneously under the same incubation conditions. In the comparative analysis of the tests, bias, essential agreement (EA), and sensitivity were assessed according to ISO 20776-2:2021. Additionally, EA, categorical agreement (CA), major error (ME), and very major error (VME) were evaluated by the CLSI guidelines. The minimum inhibitory concentration (MIC) values for MIC50 and MIC90 were 2 µg/ml and 16 µg/ml, respectively. All methods achieved the CA above 90%, with no VMEs (0%). The ME rates for CBDE, mCBDE, CAT, and RPNP were 3.4%, 8.6%, 12%, and 10.3%, respectively. The EA rates of CBDE and CAT were 95.6% and 91.3%, respectively, while bias rates were 10.8% and 24.7%. The sensitivity of both mCBDE and RPNP was 100%. Consistent with other studies in this field, a comprehensive evaluation of all methods revealed that 58.3% (7/12) of the isolates responsible for MEs had an MIC value of 1 µg/ml, 25% (3/12) had an MIC of 2 µg/ml, and 16.6% (2/12) had an MIC of 0.25 µg/ml. Most errors occurred in isolates with MIC values near the breakpoint. The CBDE demonstrated the best performance in all criteria assessed. This study highlights the benefits of alternative phenotypic methods compared to reference BMD for assessing colistin susceptibility, including ease of use, cost-effectiveness, and rapid results. Our findings emphasize the need for more comprehensive multicenter studies to validate and improve these methods, especially those with more isolates with MIC values near the breakpoint.

Introduction: Proteus species are frequently isolated from urinary tract infections (UTIs) and diabetic foot infections (DFIs). We aimed to evaluate the presence of virulence genes in P. mirabilis and P. vulgaris strains isolated from DFI and UTI. Methodology: A total of 78 Proteus isolates (57 P. mirabilis and 21 P. vulgaris) collected from patients were studied. The isolates were identified using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometer. The presence of virulence-associated genes (hlyA, mrpA, atfA, pmfA, hmpA, ptaA, ureA, ureC, zapA, ireA, rsbA, flaA, and ucaA) was evaluated by polymerase chain reaction (PCR). Results: 46 isolates were obtained from wound cultures, and 32 were obtained from the midstream urine cultures. All virulence genes, except hlyA, were detected in the study. ureA was the most detected gene in both UTI (100%) and DFI isolates (84.8%). The distributions of ureC, flaA, hpmA, ireA, rsbA, pmfA, zapA, ucaA, ptaA, atfA, and mrpA genes in DFI and UTI isolates were as follows: 82.6% and 96.9%, 71.7% and 93.8%, 69.6% and 93.8%, 69.6% and 96.9%, 69.6% and 96.9, 67.4% and 81.3%, 65.2% and 43.8%, 54.3 and 71.9%, 34.8% and 96.9%, 26.1 and 93.8%, and 23.9% and 46.9%, respectively. Conclusions: We demonstrated that P. mirabilis and P. vulgaris express a variety of virulence genes related to pathogenicity. All virulence genes were found to be more frequent in UTI isolates except zapA. There is limited data on the virulence factors of Proteus species in DFIs. Further studies are needed to investigate virulence genes in wound isolates.

Increasing resistance to first-line antibiotics used in the treatment of infections caused by Salmonella and Shigella species is emerging. Azithromycin presents a good alternative treatment option for Salmonella and Shigella infections. However, there are limited data regarding the susceptibility of azithromycin in Turkey. In this study, we aimed to evaluate the susceptibility of Salmonella and Shigella species to azithromycin, to determine and compare the minimum inhibitory concentration (MIC) values and disk diffusion zone diameters. In addition, susceptibility to meropenem and first-line antibiotic options in isolates was also investigated. A total of 170 Salmonella, 76 Shigella clinical isolates collected between 2014 and 2018 in our hospital were tested for their susceptibility to azithromycin, meropenem, ampicillin, pefloxacin, trimetoprim-sulfamethoxazole, ceftazidime, and cefotaxime. Isolates were identified by matrix-assisted laser desorption ionization time of flight mass spectrometry. The isolates were confirmed and serotyped by the reference laboratory using the conventional slide agglutination method. Susceptibility of the isolates to azithromycin and other antibiotics was evaluated by Kirby-Bauer disk diffusion method. MIC values of azithromycin were determined by the reference broth microdilution method. Combined disk diffusion test was used for the detection of extended spectrum beta-lactamase (ESBL) production. Polymerase chain reaction was performed for macrolide and carbapenem resistance genes and the detected resistance genes were confirmed by sequencing. Of the 76 Shigella isolates tested, 64 (84.2%) were identified as Shigella sonnei, 10 (13.2%) as Shigella flexneri, one (1.3%) as Shigella boydii, and one (1.3%) as Shigella dysenteriae. Among the 170 Salmonella isolates, 131 (77%) were identified as Salmonella enteritidis, 11(6.5%) as Salmonella Typhimurium, 8 (4.7%) as Salmonella Kentucky, 5 (2.9%) as Salmonella Paratyphi B, 4 (2.4%) as Salmonella Infantis, 3 (1.8%) as Salmonella Cholerasuis, and 8 (4.7%) as other serovars (Salmonella Agona, Salmonella Dabou, Salmonella Gallinarum, Salmonella Hadar, Salmonella Muenchen, Salmonella Newport, Salmonella Paratyphi C, Salmonella Senftenberg), respectively. ESBL production was determined as 7.9% (6/76) in Shigella isolates and 2.9% (5/170) in Salmonella isolates. A carbapenem resistant S.Senftenberg isolate positive for the bla OXA-48 resistance gene was detected in our study. Meropenem MIC value of the isolate was detected as > 32 mu g/ml with gradient diffusion test. Among all isolates, only one S.boydii isolate was detected as resistant to azithromycin with a MIC value of 128 mu g/ml. The isolate was positive for the existence of mphA gene by PCR. In the disk diffusion test, azithromycin inhibition zone diameters were >= 12 mm in all of the tested isolates, except for the azithromycin-resistant isolate, and the azithromycin MICs were determined as <= 16 mu g/ml by broth microdilution. Increasing resistance to commonly used antibiotics in Salmonella and Shigella species is emerging. The detection of a carbapenem-resistant Salmonella isolate in our study indicates that the spread of carbapenem resistance to other Enterobacterales species may cause global problems. Antimicrobial susceptibility testing of azithromycin for Salmonella and Shigella species has been difficult to establish due to the lack of approval in vitro breakpoints for all species. Consequently, our data shows that azithromycin exhibits as a good alternative therapeutic choice for the treatment of gastrointestinal diseases caused by Salmonella and Shigella species. Further studies are needed to provide appropriate in vitro breakpoints supported by clinical data.