Prevalence of carbapenemases in Enterobacterales from urine specimens in an university hospital in Istanbul, Turkey.

We aimed to investigate the prevalence of carbapenemases in Enterobacterales strains isolated from urine specimens between July 2019 and July 2020.CIM and modified CIM tests were applied as well as detection of blaOXA-48, blaNDM, blaVIM, blaKPC and blaIMP genes was performed by multiplex PCR.One hundred fifty of 3,242 Enterobacterales strains were found to be carbapenem resistant and 46 were included in the study. Forty five (98%) of the 46 strains included in the study were Klebsiella spp. and one (2%) of them was Escherichia coli. Susceptibility to ceftazidime-avibactam, amikacin and gentamicin was 97%, 11% and 9%, respectively. Forty three (94%) isolates were found positive at 2 and 4 h with CIM test. Forty four (97%) strains were found positive at 4 h and 43 (94%) strains were found positive at 2 h with modified CIM test.While blaOXA-48, blaNDM and blaOXA-48 with blaNDM association were found in Klebsiella spp. isolates in 55%, 27% and 11%, respectively, blaVIM, blaKPC, blaIMP were not found. Only blaOXA-48 and blaNDM-1 were detected in the E. coli strain.None of the investigated genes were detected in three Klebsiella strains but with whole genome analysis the combination of blaOXA-534, blaCMY-99 and blaKPC-3 was found in the first strain, blaOXA-370 in the second strain and no resistance gene was found in the third strain.Ceftazidime-avibactam was found to be active against 97% of strains, and the most common resistance genes were blaOXA-48 and blaNDM-1. Previously undetected resistance genes have been identified in our country.

The Relationship between Gastroduodenal Pathologies and Helicobacter pylori cagL (Cytotoxin-Associated Gene L) Polymorphism.

BACKGROUND: The polymorphisms in the region between 58 and 62 amino acids of the 194-amino acid CagL protein (CagL hypervariable motif) affect the binding affinity of CagL to integrin α5ß1 (ITGA5B1) receptor in host epithelial cells and have an effect on the development of various gastrointestinal diseases. We aimed to evaluate the associations of gastroduodenal pathologies, with the polymorphisms of cagL gene of Helicobacter pylori (H. pylori) and also associations between vacA genotypes and cagL polymorphisms. METHODS: A total of 19 gastric cancer, 16 duodenal ulcer, and 26 non-ulcer dyspepsia patients were included in this case-control study. All cases had H. pylori. A fragment of 651 bp from gene cagL (hp0539) and cagA, vacA genes was amplified by polymerase chain reaction. Purified polymerase chain reaction products were sequenced by Sanger sequencing, and nucleotide sequences were translated into amino acid sequences. RESULTS: All of the H. pylori strains had cagL and cagA genes. In the 16 (84%) gastric cancer cases, the D58 amino acid polymorphism was significant than the 4 (15.4%) duodenal ulcer cases (P = .029), and the D58/K59 amino acid polymorphism was significant in 12 (63.1%) of the gastric cancer cases than 1 (3.85%) duodenal ulcer case (P = .008). D58/K59 and DKIGQ (n = 10; 52.63%) were the most common polymorphisms in the gastric cancer and were associated with the vacA genotype s1/m2, respectively (P = .022 and P = .008). The D58/K59 amino acid polymorphism was found to have a significant Odds Ratio (OR) value of 8.9 (P = .0017) in multivariate logistic regression analysis. CONCLUSIONS: The risk of gastric cancer development is 8.9 times higher with D58/K59 polymorphism.

Epstein-Barr Virus and Helicobacter pylori co-infection in patients with gastric cancer and duodenale ulcer.

We aimed to detect EBV/Hp (Epstein-Barr Virus/Helicobacter pylori) co-infection by determining the number of copies of EBV/EBER-1 in the gastric biopsy samples of the Hp (+) GC, peptic ulcer (PU), and non-ulcer dyspepsia (NUD) cases. The patient group (PG), with 34 patients (34 GC and 30 PU patients) and a control group with 40 NUD cases were included. All patients and controls were Hp positive. EBV/EBNA-1 IgG were measured by the Anti-EBNA-1 ELISA IgG kit. Determination and quantification of EBV/EBER-1 gene region was performed by qPCR. EBV/EBER-1 positivity was 35.29% (12/34), 6.6% (2/30) and 2.5% (1/40) in GC, PU and 40 NUD cases, respectively. A significant difference was found between the GC and NUD cases (p=0.001). A significant difference was found between the groups for mean EBV/EBER-1 copy numbers (p=0.019). No significant difference was found between GC and the NUD cases (p=0.1455) for EBV/EBNA-1 IgG antibody positivity. EBV/EBER-1 positivity (OR=3.319), and age ≥55 years old (OR=2.331) were found to be a significant in multivariate logistic regression. In conclusion, our data suggest that the GC risk by EBVand Hp co-infection increased 3.3 times.

Point Mutations at gyrA and gyrB Genes of Levofloxacin Resistant Helicobacter pylori Strains and Dual Resistance with Clarithromycin.

BACKGROUND: Spontaneous point mutations in genes encoding gyrA/B subunits of DNA gyrase are responsible for fluoroquinolone resistance. We aimed to determine the clarithromycin and levofloxacin resistance phenotypically in H. pylori strains and to investigate the mutations responsible for levofloxacin resistance and the effects of these mutations on dual antibiotic resistance. METHODS: A total of 65 H. pylori isolates were included. The E-test method was used for the clarithromycin and le-vofloxacin antimicrobial susceptibility test. Real-time PCR was used to detect the point mutations. RESULTS: Twenty-four (36.9%) of 65 H. pylori strains were phenotypically resistant to clarithromycin and 14 (21.5%) to levofloxacin. The phenotypic levofloxacin resistance rate of strains with Asn87Lys and Asp91Asn mutations were significantly higher (gyrA gene) (p < 0.05). The phenotypic levofloxacin resistance rate of strains with Arg484Lys and Asp481Glu mutations were significantly higher (gyrB gene) (p < 0.05). The Asn87Lys mutation increased the risk of phenotypes being resistant to levofloxacin 70.156 times and Asp91Asn mutation increased 125,427 times higher. Seven (10.8%) of 65 H. pylori strains showed dual resistance to both levofloxacin and clarithromycin. The rate of being dual resistant with A2143G mutation (clarithromycin resistance) was found to be significantly higher (p < 0.05). CONCLUSIONS: The Asn87Lys and Asp91Asn mutations in the gyrA gene had a phenotypically enhancing effect on levofloxacin resistance, while the presence of Asp481Glu and Arg484Lys mutations in the gyrB gene did not. The existence of dual resistance was developed with the increase in clarithromycin and levofloxacin resistance rates.

Association between human leukocyte antigen gene polymorphisms and multiple EPIYA-C repeats in gastrointestinal disorders.

BACKGROUND: Polymorphisms of human leukocyte antigen (HLA) genes are suggested to increase the risk of gastric cancer (GC). AIM: To investigate the HLA allele frequencies of patients with GC relative to a control group in terms of CagA+ multiple (≥ 2) EPIYA-C repeats. METHODS: The patient group comprised 94 patients [44 GC and 50 duodenal ulcer (DU) patients], and the control group comprised 86 individuals [(50 non-ulcer dyspepsia patients and 36 people with asymptomatic Helicobacter pylori (H. pylori)]. Polymerase chain reaction was performed for the amplification of the H. pylori cagA gene and typing of EPIYA motifs. HLA sequence-specific oligonucleotide (SSO) typing was performed using Lifecodes SSO typing kits (HLA-A, HLA-B HLA-C, HLA-DRB1, and HLA-DQA1-B1 kits). RESULTS: The comparison of GC cases in terms of CagA+ multiple (≥ 2) EPIYA-C repeats showed that only the HLA-DQB1*06 allele [odds ratio (OR): 0.37, P = 0.036] was significantly lower, but significance was lost after correction (Pc = 0.1845). The HLA-DQA1*01 allele had a high ratio in GC cases with multiple EPIYA-C repeats, but this was not significant in the univariate analysis. We compared allele frequencies in the DU cases alone and in GC and DU cases together using the same criterion, and none of the HLA alleles were significantly associated with GC or DU. Also, none of the alleles were detected as independent risk factors after the multivariate analysis. On the other hand, in a multivariate logistic regression with no discriminative criterion, HLA-DQA1*01 (OR = 1.848), HLA-DQB1*06 (OR = 1.821) and HLA-A*02 (OR = 1.579) alleles were detected as independent risk factors for GC and DU. CONCLUSION: None of the HLA alleles were detected as independent risk factors in terms of CagA+ multiple EPIYA-C repeats. However, HLA-DQA1*01, HLA-DQB1*0601, and HLA-A*2 were independent risk factors with no criterion in the multivariate analysis. We suggest that the association of these alleles with gastric malignancies is not specifically related to cagA and multiple EPIYA C repeats.

Evaluating the effectiveness of anti-tuberculosis treatment by detecting Mycobacterium tuberculosis 85B messenger RNA expression in sputum.

BACKGROUND: The antigen 85 complex (85B) is secreted in large quantities from growing mycobacteria and the presence of bacterial mRNA is an indicator of cell viability. The quantitative detection of 85B mRNA expression levels can be used to assess the success of anti-tuberculosis treatment outcomes to detect viable mycobacteria cells. Therefore, we evaluated the levels of 85B mRNA of Mycobacterium tuberculosis strains in patients with pulmonary tuberculosis. METHODS: Thirty patients with primary tuberculosis were included in this study. The sputum specimens of patients were collected on days 0, 15, and 30 days and were cultured and evaluated by 85B mRNA-based RT-qPCR. RESULTS: Overall, 23 of the studied tuberculosis strains were susceptible to the primary anti-tuberculosis antibiotics used in this study, 7 were resistant. By the 30th day of treatment, 85B mRNA was detected in only one of the susceptible strains, but in all 7 of the resistant strains, though the relative gene expression varied between the strains. This difference between the susceptible and resistant strains at day 30 was statistically significant (p < 0.05). CONCLUSION: 85B mRNA expression levels could be used to follow up on primary tuberculosis cases. 85B mRNA seems to be a good diagnostic marker for monitoring anti-tuberculosis treatment outcomes.

The Prevalence of Periodontal Pathogenic Bacteria in Atherosclerotic Cardiovascular Disease.

BACKGROUND: A possible link between periodontal pathogenic bacteria and atherosclerosis may exist based on the inflammatory mechanisms initiated by bacteria found in periodontal lesions. Our aim was to investigate the presence of DNA originating from T. denticola, C. rectus, T. forsythia, and P. gingivalis in the vascular tissue specimens obtained from patients who underwent surgery for arteriosclerotic vascular disease in this study. METHODS: A total of 96 patients diagnosed with valvular heart disease due to atherosclerosis and 85 patients with advanced aortic valve stenosis due to rheumatic fever and had undergone aortic valve replacement were included as the study (PG) and the control groups (CG), respectively. Atheroma plaques and vascular tissue specimens were collected from PG and CG during cardiovascular surgical procedures. Revitalization of the lyophilized T. denticola, ATCC 35405; C. rectus, ATCC 33238; P. gingivalis, ATCC 33277 and T. forsythia, ATCC 43037 strains was performed according to the manufacturer's instructions. C. rectus, T. forsythia, and T. denticola DNA samples were analyzed using the one-step in-house PCR method. RESULTS: In one (1.04%) and three (3.13%) out of 96 atherosclerotic PG tissue specimens, P. gingivalis and T. for-sythia DNA were detected, respectively. No T. denticola or C. rectus DNA was found in the study specimens. Periodontal pathogenic bacteria were not observed in 85 CG tissue specimens. There was no statistically significant difference between PG and CG for the presence of P. gingivalis and T. forsythia DNA using Fischer's Exact test (p > 0.05). CONCLUSIONS: In conclusion, with the case-control studies on a small scale such as in our study, it is not possible to determine a causality relationship between periodontal pathogenic bacteria and formation of atherosclerosis. Periodontal pathogenic bacteria may not be the only factor that causes inflammatory diseases associated with atherosclerosis. Host response and inflammatory mechanisms may be affected by other factors such as ethnicity, dietary habits, nutritional availability, and lifestyle. Taken together, it is difficult to conclude a causal link between periodontal pathogenic bacteria and formation of atherosclerosis.

Association between polymorphisms in HLA-A, HLA-B, HLA-DR, and DQ genes from gastric cancer and duodenal ulcer patients and cagL among cagA-positive Helicobacter pylori strains: The first study in a Turkish population.

Colonization of the human gastric mucosa by H. pylori may cause peptic and duodenal ulcers (DUs), gastric lymphomas, and gastric cancers. The cagL gene is a component of cag T4SS and is involved in cagA translocation into host. An association between the risk of gastric cancer and the type of HLA class II (DR and/or DQ) was suggested in different populations. The aim of this study was to investigate, the clinical association of the cagL gene with host HLA alleles in H. pylori strains that were isolated from patients with gastric cancer, DU, and non-ulcer dyspepsia (NUD) and to determine the HLA allele that confers susceptibility or resistance for the risk of gastric cancer and DU development in Turkish patients. A total of 94 patients (44 gastric cancer and 50 DU patients; 58 male, 36 female; mean age, 49.6 years), and 86 individuals (50 NUD patients and 36 persons with normal gastrointestinal system [NGIS]; 30 male, 56 female; mean age, 47.3 years) were included as the patient and the control groups, respectively. CagA and cagL were determined by PCR method. DNA from peripheral blood samples was obtained by EZ-DNA extraction kit. For HLA SSO typing, LIFECODES SSO Typing kits (HLA-A, HLA-B HLA-C, HLA-DRB1 and HLA-DQA1/B1 kits) were used. The CagL/CagA positivity distribution in the groups were as follows: 42 (95.4%) gastric cancer, 46 (92%) DU and, 34 (68%) NUD and no NGIS cases. The HLA-DQA1*01 (OR: 3.82) allele was significantly different, suggesting that these individuals with H. pylori strains harbouring the CagL/CagA positivity are susceptible to the risk of gastric cancer and DU, and the HLA-DQA1*05 (OR, 0.318) allele was suggested as a protective allele for the risk of gastric cancer and DU using univariate analyses. HLA-DQA1*01 (OR, 2.21), HLA-DQB1*06 (OR, 2.67), sex (male, OR, 2.27), and CagL/CagA/(<2) EPIYA C repeats (OR, 5.72) were detected independent risk factors that increased the risk of gastric cancer and DU using multivariate analyses. However, the HLA-DRB1*04 (OR, 0.28) allele was shown to be a protective allele, which decreased the risk of gastric cancer and DU. Gastric pathologies result from an interaction between bacterial virulence factors, host epigenetic and environmental factors, and H. pylori strain heterogeneity, such as genotypic variation among strains and variations in H. pylori populations within an individual host.