High rate of colistin and fosfomycin resistance among carbapenemase-producing Enterobacteriaceae in Turkey.

When the problem with carbapenem-resistant Enterobacteriaceae (CRE) increases, the older antimicrobial agents such as colistin and fosfomycin are used for the treatment of these infections. In this study, the broth microdilution method for colistin and the agar dilution method for fosfomycin were used for a total of 147 multidrug-resistant (MDR) or extensively drug-resistant (XDR) strains of CRE. The study included Klebsiella pneumoniae (91.16%), Escherichia coli (7.48%), Enterobacter cloacae (0.68%), and Serratia marcescens (0.68%). All these strains produce various types of carbapenemase, including OXA-48, NDM, and KPC. Some of these strains also have three different carbapenemase mechanisms, including OXA-48 (78.23%), NDM (2.04%), and KPC (0.68%) or OXA-48 and NDM (10.88%), or OXA-48 and KPC (0.68%). About 76.19% of the strains and 67.35% of the strains were resistant for colistin and fosfomycin, respectively. A total of 21 out of 35 colistin-susceptible strains were found to be susceptible to fosfomycin. This study showed that the resistance rates of colistin and fosfomycin are high. The MDR and XDR strains of CRE are spreading in our region and thus a monitoring system for CRE should be followed. Moreover, the applicability of antimicrobial stewardship programs should be increased in all inpatient and outpatient settings.

Evaluation of the Abbott Real Time HCV genotype II assay for Hepatitis C virus genotyping.

OBJECTIVE: The determination of HCV genotypes and subtypes is very important for the selection of antiviral therapy and epidemiological studies. The aim of this study was to evaluate the performance of Abbott Real Time HCV Genotype II assay in HCV genotyping of HCV infected patients in Kayseri, Turkey. METHODS: One hundred patients with chronic hepatitis C admitted to our hospital were evaluated between June 2012 and December 2012, HCV RNA levels were determined by the COBAS® AmpliPrep/COBAS® TaqMan® 48 HCV test. HCV genotyping was investigated by the Abbott Real Time HCV Genotype II assay. With the exception of genotype 1, subtypes of HCV genotypes could not be determined by Abbott assay. Sequencing analysis was used as the reference method. RESULTS: Genotypes 1, 2, 3 and 4 were observed in 70, 4, 2 and 24 of the 100 patients, respectively, by two methods. The concordance between the two systems to determine HCV major genotypes was 100%. Of 70 patients with genotype 1, 66 showed infection with subtype 1b and 4 with subtype 1a by Abbott Real Time HCV Genotype II assay. Using sequence analysis, 61 showed infection with subtype 1b and 9 with subtype 1a. In determining of HCV genotype 1 subtypes, the difference between the two methods was not statistically significant (P>0.05). HCV genotype 4 and 3 samples were found to be subtype 4d and 3a, respectively, by sequence analysis. There were four patients with genotype 2. Sequence analysis revealed that two of these patients had type 2a and the other two had type 2b. CONCLUSION: The Abbott Real Time HCV Genotype II assay yielded results consistent with sequence analysis. However, further optimization of the Abbott Real Time HCV Genotype II assay for subtype identification of HCV is required.

Is rapid antibacterial susceptibility testing medium reliable for routine laboratory practices?

OBJECTIVE: Early detection of antibiotic susceptibility profile of the isolates has critical importance in terms of immediate beginning of the appropriate treatment and increasing of treatment success, such as meningitis, bacteriemia and sepsis. In the present study, it was aimed to compare the antibiotic susceptibility results of Quicolor (Salubris Inc., Massachusetts, USA) and standard disk diffusion method. METHODS: One hundred twenty three isolates were included in this study (80 Enterobacteriaceae, 15 Staphylococci and 28 nonfermentative Gram-negative bacteria). Antibiotic susceptibility in clinical isolates was evaluated using Mueller-Hinton (MH) agar and Quicolor (ES and NF) agar plates. RESULTS: For Enterobacteriaceae, frequency of total concordance, major error, and minor error between the tests were found as 96.8%, 0.8%, and 2.4%, respectively. For Staphylococci, frequency of total concordance, major error, and minor error among the tests were found as 95.7%, 3.5%, and 0.8%, respectively. For non fermentative bacteria, frequency of total concordance, major error, and minor error among the tests were found as 83.9%, 9.6%, and 6.4%, respectively. CONCLUSIONS: Quicolor media provided reliable susceptibility results in enteric bacteria and Staphylococci. However, further studies including higher number of nonfermentative bacteria are required to determine whether the chromogenic media are appropriate for this group of bacteria.

Retrospective evaluation of "Rods and Rings" pattern detected in the anti-nuclear antibody (ANA) indirect immunofluorescence (IIF) test.

Introduction: Anti-rods and rings (anti-RR) antibodies have recently been described as a cytoplasmic pattern in IIF-based screening of autoantibodies on HEp-2 cells and ICAP has named it as AC-23. It is most frequently related to drug-induced antibody generation. This study aimed to investigate the clinical significance of AC-23 positivity and its relevance to the diagnosis and/or follow-up of the associated diseases and/or drug use. Methods: A multicenter retrospective study was conducted among 10 hospitals from six different provinces in Türkiye from January 2017 to December 2021. The laboratory data and clinical information of 600 patients with positive anti-RR antibodies out of 547.558 HEp-2 IIF ANA samples were analyzed. Results: The distribution of AC-23 positive patients by year indicated a steady increase between 2017-2021. Anti-RR prevalence in post-COVID-19 period was significantly higher than that of pre-COVID-19 period (p=0.00). Concomitant ANA positivity was detected in 56.5% of patients, the most common patterns being AC-4 and AC-5 (41.1%). The most frequent pathology among the anti-RR positive patients was an autoimmune disease (19.83%); 28.57% of which had rheumatoid arthritis and 17.65% autoimmune liver disease. Among the 600 patients, 65 (10.83%) were diagnosed as hepatitis C virus (HCV) infection. Available data for 38 of the HCV patients revealed that 71.05% of them had a history of interferon alfa+ribavirin and 28.95% of them had a history of NS3/4/5A/5B polymerase inhibitor or protease inhibitor drug use. Significant increase in the rate of anti-RR positivity was observed in the post-COVID-19 period when compared to pre-COVID-19 period (p:0.00). Discussion: This is the first multicenter study in Türkiye about the clinical association of anti-RR antibodies which may be ignored during routine HEp-2 IIF testing. Pathologies other than HCV should be taken into consideration in terms of the possible role of anti-RR in autoimmune diseases and other pathologies. The preliminary data obtained in this study suggest that anti-RR antibody development might also be associated to COVID-19, supporting the several previous data related to the potential of viruses triggering the formation of autoantibodies. Large-scale prospective studies should elucidate the clinical significance of RR pattern and determine its role in patient diagnosis and follow-up.

Distribution of hepatitis C virus genotypes in Kayseri region, in Turkey: unexpected rate of genotype 4.

BACKGROUND: Hepatitis C virus (HCV) is an important cause of chronic liver disease. There are six genotypes and more than 80 subtypes of HCV. The aim of this study was to investigate the distribution of HCV genotypes in Middle Anatolia in Turkey, and the association of HCV genotypes with pre-treatment HCV RNA viral load, serum transaminase levels, and histopathological grade of liver fibrosis. METHODS: A total of 160 patients (103 female, 57 male) with chronic hepatitis C were retrospectively evaluated. HCV RNA level was determined by commercial real time PCR method. HCV RNA positive sera were genotyped by the Abbott Real Time HCV Genotype II assay and sequenced by the ABI Prism 310 Genetic Analyzer. Gender, age, serum ALT, AST, HCV RNA viral load, and fibrosis staging of liver were determined in all patients. RESULTS: Genotype 1b was the most frequent (64.7%) followed by genotype 4d (28.3%), 2 (4.4%), and la (2.5%). The HCV genotype results were found consistent with both methods. The gender distribution of the 160 HCV infected patients was 57 male/103 female. Log HCVRNA was significantly higher in genotype 1b compared to genotype 4 and 1a. Stage of liver fibrosis, histology activity index, serum ALT and AST levels did not differ between groups depending on genotypes. Advanced liver fibrosis (Group 2) was found in 36 (76.6%) patients with genotype 1b and in 10 (21.3%) patients with genotype 4 and only in 1 patient (2.1%) with genotype 1a. CONCLUSIONS: HCV genotype 1b is the most frequent type (64.7%) in this region. Prevalence of genotype 4 in this region is higher than the national HCV genotype distribution. Serum transaminase levels and liver fibrosis scores are not associated with HCV genotypes.

Molecular epidemiology, virulence factors, and antifungal susceptibility of Candida inconspicua strains isolated from clinical samples in Turkey.

In this study, it was aimed to evaluate the molecular epidemiology, virulence factors, and antifungal susceptibility of clinical Candida inconspicua isolates. All isolates were identified by phenotypic methods and sequence analysis of ITS 1-2, D1/D2, EF-1 alpha. Proteinase, phospholipase, and esterase activities, biofilm formation, and antifungal susceptibilities were determined. All thirty isolates identified as Candida norvegensis by phenotypic methods were reidentified as C. inconspicua by sequence analysis, demonstrating the inadequacy of phenotypic methods to differentiate these 2 species. The gene regions examined in terms of determining evolutionary relatedness did not show intraspecies nucleotide variations. Therefore, different molecular approaches are needed to evaluate molecular epidemiology. Esterase, phospholipase, and biofilm formation were found to be positive in 100%, 100%, and 36.6% of the strains, respectively. The MIC50/MIC90 values for fluconazole and flucytosine were found to be higher than the other tested antifungals, which should be taken into account in the treatment.

Identification, molecular characterization, and antifungal susceptibility of Cyberlindnera fabianii strains isolated from urinary tract.

OBJECTIVES: Cyberlindnera fabianii is an opportunistic pathogen isolated from clinical specimens. It can be incorrectly identified as Candida utulis by phenotypic methods. This study aimed to accurately identify Cy.fabianii strains isolated from the urinary tract, and to determine their molecular characterization and antifungal susceptibilities as well. METHODS: Twenty-nine yeast strains isolated from urinary tract samples were studied. Strains were identified by phenotypically, sequence analysis and MALDI-TOF MS. Sequence analysis using different gene regions (ITS1-2,D1/D2,EF-1-alpha) in ribosomal DNA was performed for the molecular analysis. Phylogenetic analysis was done by the neighbor-joining method. Antifungal susceptibilities of strains were determined for nine antifungals by reference broth microdilution and the Sensititre YeastOne broth microdilution method (SensititreTMYeastOneTMAST Plate, Thermo Fisher Scientific™,USA) according to CLSI M60-Ed2 recommendations. RESULTS: All strains were identified as C.utulis phenotypically by conventional methods, however all strains were identified as Cy.fabianii by sequence analysis and MALDI-TOF MS. It was observed that the gene regions examined in terms of determining evolutionary relatedness did not show intraspecies nucleotide variations. In all strains, the MIC50/MIC90 values for fluconazole were higher than the other antifungals tested. CONCLUSION: Cy.fabianii should be considered in fluconazole-resistant urinary tract yeast infections. Although conventional phenotypical methods were insufficient to identify Cy.fabianii, it could be correctly identified with sequence analysis using different gene regions (ITS1-2,D1/D2,EF-1-alpha) in ribosomal DNA and MALDI-TOF MS.

Molecular epidemiology and antifungal susceptibilities of Aspergillus species isolated from patients with invasive aspergillosis.

OBJECTIVE: The aim of this study was to evaluate the demographic data, molecular epidemiology, and in vitro antifungal susceptibility results of patients with Aspergillus isolated from various clinical specimens. METHODS: A total of 44 Aspergillus strains were studied. The definition of invasive aspergillosis in patients was made according to European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria. Strains were phenotypically and molecularly identified. Demographic characteristics of patients and genotypes of strains were evaluated. Phylogenetic analysis was done by the The Unweighted Pair-Group Method with Arithmetic Mean (UPGMA). Antifungal susceptibility of strains was determined according to The Clinical and Laboratory Standards Institute (CLSI)-M61-Ed2 and The European Committee on Antimicrobial Susceptibility Testing (EUCAST). RESULTS: A total of 11 patients were classified as proven and 33 as probable invasive aspergillosis. There was a statistically significant difference in age groups, subdisease, neutropenic, and receiving chemotherapy between groups. A total of 23 strains were identified as Aspergillus fumigatus, 12 as Aspergillus niger, 6 as Aspergillus flavus, and 3 as Aspergillus terreus. Phylogenetic analysis revealed five different genotypes. No statistical difference was found in the comparisons between patients groups and genotype groups. There was a statistically significant difference between genotype groups and voriconazole, posaconazole, and itraconazole Minimum Inhibition Concentration (MIC). CONCLUSION: Accurate identification of strains and antifungal susceptibility studies should be performed due to azole and amphotericin B resistance. Genotyping studies are important in infection control due to identifying sources of infection and transmission routes.

Molecular epidemiology and antifungal susceptibilities of Aspergillus species isolated from patients with invasive aspergillosis

SUMMARY OBJECTIVE: The aim of this study was to evaluate the demographic data, molecular epidemiology, and in vitro antifungal susceptibility results of patients with Aspergillus isolated from various clinical specimens. METHODS: A total of 44 Aspergillus strains were studied. The definition of invasive aspergillosis in patients was made according to European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria. Strains were phenotypically and molecularly identified. Demographic characteristics of patients and genotypes of strains were evaluated. Phylogenetic analysis was done by the The Unweighted Pair-Group Method with Arithmetic Mean (UPGMA). Antifungal susceptibility of strains was determined according to The Clinical and Laboratory Standards Institute (CLSI)-M61-Ed2 and The European Committee on Antimicrobial Susceptibility Testing (EUCAST). RESULTS: A total of 11 patients were classified as proven and 33 as probable invasive aspergillosis. There was a statistically significant difference in age groups, subdisease, neutropenic, and receiving chemotherapy between groups. A total of 23 strains were identified as Aspergillus fumigatus, 12 as Aspergillus niger, 6 as Aspergillus flavus, and 3 as Aspergillus terreus. Phylogenetic analysis revealed five different genotypes. No statistical difference was found in the comparisons between patients groups and genotype groups. There was a statistically significant difference between genotype groups and voriconazole, posaconazole, and itraconazole Minimum Inhibition Concentration (MIC). CONCLUSION: Accurate identification of strains and antifungal susceptibility studies should be performed due to azole and amphotericin B resistance. Genotyping studies are important in infection control due to identifying sources of infection and transmission routes.

The Investigation of HCV RNA in Tear Fluid and Aqueous Humor in Patients with Anti-HCV Antibody Positive Who Underwent Cataract Surgery.

PURPOSE: To obtain aqueous humor and tear fluid samples during cataract surgery of the hepatitis C virus (HCV)-antibody-positive patients in order to analyze them for HCV RNA and compare these measurements with serum HCV RNA levels. METHODS: Twenty-nine anti-HCV-positive patients were included this study. HCV RNA viral load levels were determined by commercial real-time polymerase chain reaction system. Antibodies to HCV were screened with the enzyme-linked immunosorbent assay (ELISA) using anti-HCV test kit. RESULTS: Log10 HCV RNA levels were found to be 6.00 ± 1.06 IU/mL in serum, 2.76 ± 0.36 IU/mL in the aqueous humor, and 1.91 ± 0.93 IU/mL in tear fluid. No correlation was detected between samples for HCV RNA positivity (p = .390, κ = .102). We have observed that, viral RNA was detected in the aqueous humor, while not in serum in one case, whereas viral RNA was detected in tear fluid but not in serum in another case. CONCLUSIONS: Although viral load detected in aqueous humor and tear fluid samples was considerably lower compared to the serum samples, it can still be important in terms of infectivity.

Diagnostic Accuracy of a New Urinalysis System, DongJiu, for Diagnosis of Urinary Tract Infection.

OBJECTIVE: The microscopic and chemical analysis of urine is essential for the diagnosis of patients with urinary tract infections (UTI). Quantitative urine culture is the 'gold standard' method for the diagnosis of UTI, but it is labour-intensive and time consuming. The DongJiu 8602 is a new automated urinalysis system with dipstick and microscopy testing. The aim of this study was to evaluate the analytical and diagnostic performance of DongJiu 8602 in comparison to urine culture as the reference method. METHODS: This retrospective study included 3970 urine samples, with a preliminary diagnosis of UTI and for which both urine analysis and urine culture were requested. Cut-off values for bacteria and white blood cells (WBCs) were determined by comparing the results with urine cultures. The cut-off values by the receiver operating characteristic (ROC) curve technique, sensitivity, and specificity were calculated for microscopy and dipstick parameters (nitrite and leukocyte esterase). RESULTS: Among the 3970 urine specimens submitted for culture, 3275 cultures (82.5%) were negative, and 695 were (17.5%) positive. The best cut-off values obtained from ROC analysis were 13/µL for bacteriuria (sensitivity: 31.1%, specificity: 91.8%), and 31/µL for WBCs (sensitivity: 48.6%, specificity: 85.9 %). We observed no significant difference between the area under the curves (AUC) of microscopy and dipstick parameters (p>0.05). CONCLUSION: According to our data, performances of microscopic and dipstick parameters of DongJiu 8602 were not satisfactory. Although, analytical sensitivity was improved slightly with combined assessment of microscopic and dipstick results for the lack of UTI, it did not achieve expected levels.

Evaluation of chromogenic agar, [corrected] VITEK2 YST and VITEK® MS for identification of Candida strains isolated from blood cultures.

The aim of this study is to compare conventional methods, chromogenic agar, [corrected] VITEK2 YST card and VITEK®MS system for the identification of Candida strains isolated from blood cultures. Fifty-four strains were identified according to conventional methods, chromogenic agar, [corrected] VITEK2 YST card and VITEK®MS. Sequencing was used as the reference method. The 54 strains included 32 Candida parapsilosis, 19 Candida albicans, 1 Candida glabrata and 2 Candida tropicalis according to the reference method. One C. albicans and one C. glabrata isolate were misidentified as C. parapsilosis by chromogenic agar. [corrected]. Two C. parapsilosis and three C. albicans isolates were misidentified by VITEK2 YST card. Chromogenic agar, [corrected] VITEK2 YST card and VITEK®MS identified correctly 96.2%, 90.7% and 100% of all strains, respectively. We found that the chromogenic agar, [corrected] VITEK2 YST card and VITEK®MS system are easy, rapid and accurate alternative methods for the identification of yeast species in the clinical microbiology laboratory.

Interdigital foot infections: Corynebacterium minutissimum and agents of superficial mycoses.

Interdigital foot infections are mostly caused initially by dermatophytes, yeasts and less frequently by bacteria. Erythrasma caused by Corynebacterium minutissimum can be confused with superficial mycoses. The aim of the study was to determine the prevalence of the etiologic agents of superficial mycoses and the frequency of Corynebacterium minutissimum in interdigital foot infections. All the samples obtained from the 121 patients with interdigital foot infections were examined directly with the use of 20% potassium hydroxide mounts and Gram stain under the microscope and cultured on Sabouraud's dextrose agar plates. In identification of superficial mycoses, the rate was found to be 14% with the cultural method and 14% with direct microscopic examination. Using a combination of direct microscopic examination and culture, a 33.8% ratio was achieved. In the culture of these samples, the most isolated factor was Trichophyton rubrum (33.7%). In 24 of the patients (19.8%) Corynebacterium minutissimum was detected by Gram staining, in 6 of these patients Trichophyton rubrum was found, Trichophyton mentagrophytes was found in 2 and Trichosporon spp. was found in 1. The examination of interdigital foot lesions in the laboratory, the coexistence of erythrasma with dermatophytes and yeast should be considered.

Interdigital foot infections: Corynebacterium minutissimum and agents of superficial mycoses

Interdigital foot infections are mostly caused initially by dermatophytes, yeasts and less frequently by bacteria. Erythrasma caused by Corynebacterium minutissimum can be confused with superficial mycoses. The aim of the study was to determine the prevalence of the etiologic agents of superficial mycoses and the frequency of Corynebacterium minutissimum in interdigital foot infections. All the samples obtained from the 121 patients with interdigital foot infections were examined directly with the use of 20% potassium hydroxide mounts and Gram stain under the microscope and cultured on Sabouraud's dextrose agar plates. In identification of superficial mycoses, the rate was found to be 14% with the cultural method and 14% with direct microscopic examination. Using a combination of direct microscopic examination and culture, a 33.8% ratio was achieved. In the culture of these samples, the most isolated factor was Trichophyton rubrum (33.7%). In 24 of the patients (19.8%) Corynebacterium minutissimum was detected by Gram staining, in 6 of these patients Trichophyton rubrum was found, Trichophyton mentagrophytes was found in 2 and Trichosporon spp. was found in 1. The examination of interdigital foot lesions in the laboratory, the coexistence of erythrasma with dermatophytes and yeast should be considered.