Antimicrobial resistance patterns and integron carriage of Escherichia coli isolates causing community-acquired infections in Turkey

We aimed to observe antimiCrobial resistance patterns and integron carriage of Escherichia coli isolates causing community-acquired infections. Two hundred sixty-eight E. coli strains were obtained from outpatients with various infections at different polyclinics at the 82nd Year of State Hospital in Rize, Turkey. Susceptibility to antimicrobials was tested using a disk diffusion method. The presence of integrons was examined using PCR with specific primers. Positive PCR results were confirmed by sequencing. A broth mating method was used for conjugation assays. Extragenic palindromic-FOR was performed using the oligonucleotide primer BOXA1R. Resistance frequency for ampicillin,trimethoprim/sulfamethoxazole, and tetracycline was determined as 50.6%, 33.5%, and 36.8% respectively. No strains were resistant to amikacin. Seventy isolates were positive for the intll gene, of which 49 carried gene cassettes. Eleven isolates were positive for the int12 gene, eight of swhich carried gene cassettes. Seven gene cassettes (dfrAl, dfrA5, dfrA7, dfrA17, aadAl, aadA5, and sat2) were predominantly harbored in integrons. We detected conjugative plasmids harboring integrons in two E. coli strains. Four strain clusters were yielded by BOX-FOR fingerprints showing that they were clonally related. No apparent relationship occurred among class 1 and 2 integron-carrying strains. We conclude that integrons are widespread in genetically variable E. coli strains and will continue to mediate dissemination of resistance genes in the community.We aimed to observe antimiCrobial resistance patterns and integron carriage of Escherichia coli isolates causing community-acquired infections. Two hundred sixty-eight E. coli strains were obtained from outpatients with various infections at different polyclinics at the 82nd Year of State Hospital in Rize, Turkey. Susceptibility to antimicrobials was tested using a disk diffusion method. The presence of integrons was examined using PCR with specific primers. Positive PCR results were confirmed by sequencing. A broth mating method was used for conjugation assays. Extragenic palindromic-FOR was performed using the oligonucleotide primer BOXA1R. Resistance frequency for ampicillin, trimethoprim/sulfamethoxazole, and tetracycline was determined as 50.6%, 33.5%, and 36.8% respectively. No strains were resistant to amikacin. Seventy isolates were positive for the intll gene, of which 49 carried gene cassettes. Eleven isolates were positive for the int12 gene, eight of swhich carried gene cassettes. Seven gene cassettes (dfrAl, dfrA5, dfrA7, dfrA17, aadAl, aadA5, and sat2) were predominantly harbored in integrons. We detected conjugative plasmids harboring integrons in two E. coli strains. Four strain clusters were yielded by BOX-FOR fingerprints showing that they were clonally related. No apparent relationship occurred among class 1 and 2 integron-carrying strains. We conclude that integrons are widespread in genetically variable E. coli strains and will continue to mediate dissemination of resistance genes in the community.This work was supported by Recep Tayyip Erdogan University Research Fund grants BAP-2012.106.01.11 and BAP-2011. 102.03.3

Investigation of class 1 integrons with antibiotic resistance genes in multidrug-resistant acinetobacter baumannii strains and determination of plant extract effects on multidrug-resistant isolates

This study aimed to investigate the presence of resistance genes in multidrug-resistant A. baumannii isolates as well as to determine the antibacterial activity of selected plant extracts against isolates. 41 strains were isolated from various clinical samples. PCR tests were performed using the primers. Methanol was used as solvent for the preparation of the plant extracts. MIC values of the plant extracts were determined by the broth microdilution method. The bla(OXA-)(23), bla(CTX-M-1), bla(CTX-M-2), bla(GES) genes and Class 1 integrons were detected in five isolated strains. The lowest MIC value (2.25 mg/mL) was determined for the Echinacea putpurea extract, while the highest MIC value (50 mg/mL) was determined for the Morus alba extract. Determination of the antibacterial effect of plants extracts used in the study against A. baumannii isolates shows the importance of screening the antibacterial activity of plants in the fight against antibiotic resistance.Studiul a avut ca scop investigarea genelor de rezistență ale izolatelor de A. baumannii multirezistente, precum și determinarea activității antibacteriene a unor extracte vegetale supra a 41 specii microbiene izolate din diferite probe clinice. Metanolul a fost folosit ca solvent de extracție. Valorile concentrației minime inhibitorii ale extractelor obținute au fost determinate prin metoda microdiluției. Genele blaOXA-23, blaCTX-M-1 , blaCTX-M2 și blaGES, alături de integroni din clasa 1 au fost decelate pentru 5 specii izolate. Valoarea MIC cea mai scăzută (2,25 mg/mL) a fost obținută pentru extractul din Echinaceea purpurea, în timp ce cea mai mare valoare MIC (50 mg/mL) a fost determinată pentru extractul de Morus alba. Determinarea efectului antibacterian ale extractelor vegetale împotriva izolatelor de A. baumannii arată importanța screeningului activității antibacteriene a plantelor în lupta împotriva rezistenței la antibiotice

Cloning, expression, and characterization of a novel CTP synthase gene from Anoxybacillus gonensis G2

The cytidine-5'-triphosphate (CTP) synthase (EC 6.4.3.2) gene (pyrG) was cloned and sequenced from the thermophilic bacterium Anoxybacillus gonensis G2 (Ago). The gene is 1590 bp in length and encodes a protein of 530 amino acids, with a molecular mass of 59.5 kDa. The amino acid sequence of CTP synthase shares approximately 90%–94% similarity to Bacillus sp., and it belongs to the triad glutamine amidotransferases, which utilize a Cys–His–Glu triad for activity. Multiple sequence alignments revealed that the enzyme includes conserved amino acids responsible for catalytic activity and the binding of a divalent metal ion (Mg+2). AgoCTP synthase (AgoG2CTPs) was overproduced in Escherichia coli BL21 (DE3) pLysS as recombinant and purified by nickel affinity chromatography. Its biochemical characterization showed that the enzyme had maximal activity at pH 9.0–10.0 and 65 ºC. Km, Vmax, and kcat were found to be approximately 12.415 mM, 0.381 U/L, and 0.762 s–1 at 65 ºC, respectively. CTP synthase promotes the formation of CTP in dividing cells and is a recognized target for anticancer and antibacterial drugs. The results obtained from this study can be improved upon with the use of different species and substrates

COVID-19 vaccine immunity in oncology patients

BACKGROUND: To investigate the effect of vaccine types applied in our country against 2019 coronavirus disease on the formation of protective antibodies in oncology patients. MATERIALS AND METHODS: The data of 81 cancer patients who received at least one dose of vaccine for COVID-19 and radiotherapy were analyzed retrospectively. At any time after the vaccination, blood samples were taken and the antibody titers against the vaccine were measured. RESULTS: There were 28 (34.6 %) patients who received two doses of vaccine and 48 patients (59.3 %) who received 3 doses of vaccine (Sinovac only), while 26 patients (32.1 %) were given both vaccines. The mean time for antibody measurement was 62 days after the last vaccination. IgG levels were significantly higher in patients who received Biontech vaccine than in those who received Sinovac (r = 0.525; p < 0.001). While chemotherapy was the factor that decreased the mean IgM level (p = 0.044), advanced disease (stages 3 and 4) was a significant factor that increased the mean IgG level (p = 0.047). A statistically significant negative correlation was found between IgM antibody level and WBC count after first vaccination (r = ‒0.251; p = 0.024). For every WBC count unit increase in the first vaccination period, there was a 1.333-fold increase in the risk of IgM negativity. CONCLUSION: The Biontech vaccine produced higher antibody levels in advanced oncology patients. While the application of radiotherapy in cancer patients was not found to be an effective factor in the vaccination status, it was determined that the application of chemotherapy significantly reduced IgM levels (Tab. 5, Ref. 28). Text in PDF www.elis.skRecep Tayyip Erdogan University ; Coordination Unit for Projects of Scientific Researc

OXA- and GES-type beta-lactamases predominate in extensively drug-resistant Acinetobacter baumannii isolates from a Turkish University Hospital

This study was supported by grants from Recep Tayyip Erdogan University (BAP-2012.106.01.11 and BAP-2011.102.03.3). AYP was supported by the Australian National Health and Medical Research Council (APP1047916 and APP1010114).We determined the antibiotic susceptibility and genetic mechanisms of resistance in clinical strains of Acinetobacter baumannii from Istanbul, Turkey. A total of 101 clinical strains were collected between November 2011 and July 2012. Antimicrobial susceptibility was performed using the Vitek 2 Compact system and E-test. Multiplex PCR was used for detecting bla(OXA-51-like), bla(OXA-23-like), bla(OXA-40-like) and bla(OXA-58-like) genes. ISAba1, bla(IMP-like), bla(VIM-like), bla(GES), bla(VEB), bla(PER-2), aac-3-Ia and aac-6'-Ib and NDM-1 genes were detected by PCR and sequencing. By multiplex PCR, all strains were positive for bla(OXA-51), 79 strains carried bla(OXA-23) and one strain carried bla(OXA-40). bla(OXA-51) and bla(OXA-23) were found together in 79 strains. ISAba1 element was detected in 81 strains, and in all cases it was found upstream of bla(OXA-51). GES-type carbapenemases were found in 24 strains (GES-11 in 16 strains and GES-22 in 8 strains) while bla(PER-2), bla(VEB-1), bla(NDM-1), bla(IMP)- and bla(VIM)-type carbapenemases were not observed. Aminoglycoside modifying enzyme (aac-3-Ia and aac-6'Ib) genes were detected in 13 and 15 strains, respectively. Ninety-seven (96%) A. baumannii strains were defined as MDR and of these, 98% were extensively drug resistant (sensitive only to colistin). Colistin remains the only active compound against all clinical strains. As seen in other regions, OXA-type carbapenemases, with or without an upstream ISAba1, predominate but GES-type carbapenemases also appear to have a significant presence. REP-PCR analysis was performed for molecular typing and all strains were collected into 12 different groups. To our knowledge, this is the first report of GES-11 and OXA-40 in A. baumannii from Turkey

Evaluating molecular epidemiology of carbapenem non-susceptible Klebsiella pneumoniae isolates with MLST, MALDI-TOF MS, PFGE

Background: This study aimed to evaluate antibiotic resistance genes and virulence genes and the clonal relationship of the carbapenem-nonsusceptible Klebsiella pneumoniae strains by molecular methods which are isolated from various clinical specimens from patients treated in tertiary care hospital in Turkey. Methods: Identification of 32 carbapenem non-susceptible K. pneumoniae were determined by VITEK-2 (BioMérieux, France) automated system. Thirteen colistin-resistant strains were tested with the broth microdilution method. Various antibiotic resistance genes and virulence genes frequently seen in carbapenem-resistant strains were screened by PCR. Immunochromatographic tests used in the rapid diagnosis of carbapenemases were compared with PCR results. In addition, PFGE, MLST and MALDI-TOF MS methods were used to determine the clonal relationship among these strains. Results: PCR demonstrated that 31 of the strains carried at least one of the carbapenemase genes. In one strain, the coexistence of bla OXA−48+NDM was shown. The most common resistance genes were determined as bla SHV (84.3%), bla CTX−M−1 (46.8%), bla OXA−48 (40.6%), bla KPC (40.6%), bla TEM (31.2%), bla NDM (18.8%) respectively. Among the virulence genes; magA (68.7%) was the most common, followed by kpn (59.3%) and K2 (9.3%). Immunochromatographic tests were found to be 100% compatible with PCR results. All colistin-resistant isolates were also found to be resistant by colistin broth microdilution. In PFGE analysis, 25 different genotypes were determined and clustering isolates were collected in 5 different clusters and the clustering rate was 35.4%. In MLST analysis, ST101 type was determined as the most common ST type with a rate of 29%. ST101 is followed by ST16, ST307, ST14, ST147, ST309, ST377, ST395 and ST2096, respectively. The compatibility rate between MALDI-TOF MS and VITEK-2 was found 94.3%, in bacterial identification. In MALDI-TOF MS typing, the maximum similarity between the strains was less than 70% and clustering not shown. Conclusion: In addition to OXA-48, which is endemic in our country, it has been determined that KPC, which is more common in the world, is becoming increasingly common in our region. ST101 type was determined as the most common type between the strains. To the best of our knowledge, this is the first study that compares these three methods in our country. There may be differences between bacterial identifications made with VITEK-2 and MALDI-TOF MS. In this study, it was observed that MALDI-TOF MS analyses were not compatible with the typing of strains according to PFGE and MLST analysis results.Recep Tayyip Erdogan University Scientific Research Projects Uni

Distribution of Intestinal Parasites in Patients Admitted to the Government and University Hospitals in Rize Province

AimThe aim of this study was to retrospectively determine the prevalence of intestinal parasites in patients, whose examination material was sent to parasitology laboratory of two different hospitals in Rize province during the period of January 2012 to June 2013.Materials and MethodsStool samples were examined by direct macroscopy, native-lugol and trichrome staining methods for protozoan (cyst or trophozoites) and helminths (eggs or larvae). Cellophane preparations were examined for Enterobius vermicularis eggs.ResultsA total of 9.994 samples were investigated and parasites were identified from 240 samples (2.4%). Entamoeba coli (59.6%) was determined the most common parasite among the identified parasites and it was followed by E. vermicularis (12.5%), G. intestinalis (12.1%) and Entamoeba histolytica/dispar/dispar (9.6 %).ConclusionThere is less studies showing the prevalence of intestinal parasites in our region. It is compared to the work done in different places and times, parasites ration in our province was found very low (2.4%). The acceptance of the appropriate sample and the use of high sensitivity methods of investigation should be expanded for the effective recognition of parasitosis and their successful treatment. There is a slight improvement for socio-economic and environmental conditions in our country, but the parasitic infections are still a current and continous health problem in our society

Comparison of verona integron-borne metallo-beta-lactamase (VIM) variants reveals differences in stability and inhibition profiles

DUZGUN, AZER OZAD/0000-0002-6301-611X; Abboud, Martine I./0000-0003-2141-5988; Brem, Jurgen/0000-0002-0137-3226; McDonough, Michael A/0000-0003-4664-6942; Rydzik, Anna/0000-0003-3158-0493; DUZGUN, AZER OZAD/0000-0002-6301-611X; McDonough, Michael/0000-0003-4664-6942; Schofield, Christopher/0000-0002-0290-6565; SANDALLI, Cemal/0000-0002-1298-3687WOS: 000376490800025PubMed: 26666919Metallo-beta-lactamases (MBLs) are of increasing clinical significance; the development of clinically useful MBL inhibitors is challenged by the rapid evolution of variant MBLs. the Verona integron-borne metallo-beta-lactamase (VIM) enzymes are among the most widely distributed MBLs, with > 40 VIM variants having been reported. We report on the crystallographic analysis of VIM-5 and comparison of biochemical and biophysical properties of VIM-1, VIM-2, VIM-4, VIM-5, and VIM-38. Recombinant VIM variants were produced and purified, and their secondary structure and thermal stabilities were investigated by circular dichroism analyses. Steady-state kinetic analyses with a representative panel of beta-lactam substrates were carried out to compare the catalytic efficiencies of the VIM variants. Furthermore, a set of metalloenzyme inhibitors were screened to compare their effects on the different VIM variants. the results reveal only small variations in the kinetic parameters of the VIM variants but substantial differences in their thermal stabilities and inhibition profiles. Overall, these results support the proposal that protein stability may be a factor in MBL evolution and highlight the importance of screening MBL variants during inhibitor development programs.Rhodes Trust; Scientific and Technology Council of Turkey; Recep Tayyip Erdogan Universitesi Research FundRecep Tayyip Erdogan University [BAP-2013.102.03.13]; Medical Research CouncilMedical Research Council UK (MRC) [MR/L007665/1]; Medical Research Council/Canadian Grant [G1100135]; Biochemical Society Krebs Memorial Award; Medical Research CouncilMedical Research Council UK (MRC) [G1100135, MR/N002679/1] Funding Source: researchfishThe Rhodes Trust provided funding to Anne Makena. Scientific and Technology Council of Turkey provided funding to Cemal Sandalli. Recep Tayyip Erdogan Universitesi Research Fund provided funding to Aysegul Saral, Aysegul C. Cicek, and Cemal Sandalli under grant number BAP-2013.102.03.13. Medical Research Council provided funding to Jurgen Brem, Michael A. McDonough, Anna M. Rydzik, and Christopher J. Schofield under grant number MR/L007665/1. Medical Research Council/Canadian Grant provided funding to Jurgen Brem, Michael A. McDonough, Anna M. Rydzik, and Christopher J. Schofield under grant number G1100135. Biochemical Society Krebs Memorial Award provided funding to Martine I. Abboud

Molecular epidemiological analysis of integron gene cassettes and tetA/tetB/tetD gene associations in Escherichia coli strains producing extended-spectrum beta-lactamase (ESBL) in urine cultures

Background. Epidemiological studies of tetracycline (TE) resistance genes and integron gene cassettes, particularly in urine samples, are limited in Turkey. Objectives. To investigate antibiotic susceptibility profiles, extended-spectrum beta-lactamase (ESBL) positivity, tet gene types, class-I/-II integron gene cassettes, and clonal relationships among tet-resistant isolates of Escherichia coli from urine cultures of outpatients. Materials and methods. Isolates were identified using conventional methods and the automated Vitek (R) 2 Compact system. Antimicrobial susceptibility was performed for 19 antibiotics. The ESBL production was performed using the Kirby-Bauer disk diffusion test. The double disk synergy test was used for confirmatory testing. Polymerase chain reaction (PCR) was used to determine the presence of class-I/-II integron gene cassettes and tetA, tetB and tetD resistance genes. The pulsed-field gel electrophoresis typing was performed to identify clonal relations. Results. A total of 121 isolates were obtained and found to be resistant or sensitive to ampicillin and amikacin/imipenem. Resistance to ceftazidime, cefotaxime and ceftriaxone was determined to be 31.3%, 77.6% and 83.1%, respectively. Tetracycline resistance was detected in 82 isolates, mostly caused by the tetB gene. No tet gene was detected in the remaining 39 isolates. Although 64 out of 82 isolates carried a class-I integron, only 4 had a class-II integron (with sizes of 800-2900 base pairs). Furthermore, tet genes were identified with different size class-I integron gene cassettes. However, tet genes were not detected in any isolate identified with integron gene cassette II. Clonally, the isolates were found to be related in subgroups because they were community-acquired. Conclusions. This study showed thatthe tetB gene is most commonly found in E. coli isolates grown in urine samples from the Turkish population

High prevalence of NDM metallo-β-lactamase among ESBL-producing Escherichia coli İsolates

Resistance to β-lactams in Enterobacteriaceae has been increasing worldwide. This study aimed to determine the frequency of β-lactamase genes and antibiotic resistance rates of 140 extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates obtained from urinary tract infection in Ordu Province, Turkey. Isolates were identified by classic methods and by automated system. ESBL production was confirmed by double disk synergy test and antimicrobial susceptibility was investigated by disk diffusion method. All isolates were screened for β-lactamase coding genes from three groups (A, B, and D) by polymerase chain reaction. The highest rate of susceptible isolates was observed for imipenem (IPM, 99.3%) and ertapenem (ETP, 97.9%), and the highest rate of resistant isolates was observed for cefuroxime (97.9%), ceftriaxone (97.2%), and cefazolin (90.7%). In our study, blaCTX-M1-like group was the most prevalent β-lactamase (n = 109), followed by blaTEM (n = 68), blaCTX-M2 (n = 22), and blaSHV (n = 2). By contrast to low resistance rate to IPM and ETP, we determined blaNDM in 31 isolates (22.1%). In co-prevalence of blaNDM-1 and ESBL-coding genes, a low carbapenem resistance was determined. We can confirm that blaCTX-M1-types are still the most frequent β-lactamase coding gene in Turkey. Our study showed the highest prevalence of blaNDM-1 metallo-β-lactamase coding gene in ESBL-producing E. coli