Leptospirosis, caused by pathogenic Leptospira, is one of the most important zoonoses in the world. Several molecular techniques have been developed for detection and differentiation between pathogenic and saprophytic Leptospira spp. The aim of this study was to develop a rapid and simple assay for specific detection and differentiation of pathogenic Leptospira spp. by multiplex real-time PCR (TaqMan) assay using primers and probes targeting Leptospira genus specific 16S ribosomal RNA gene, the pathogen specific lig A/B genes and nonpathogen Leptospira biflexa specific 23S ribosomal RNA gene. Sixteen reference strains of Leptospira spp. including pathogenic and nonpathogenic and ten other negative control bacterial strains were used in the study. While the 16S primers amplified target from both pathogenic and non-pathogenic leptospires, the ligA/B and the 23S primers amplified target DNA from pathogenic and non-pathogenic leptospires, respectively. The multiplex real-time PCR (TaqMan) assay detection limit, that is, the sensitivity was found approximately 1 x 10(2) cells/ml for ligA/B gene and 23S ribosomal RNA gene, and 10 cells/ml 16S ribosomal RNA. The reaction efficiencies were 83-105% with decision coefficients of more than 0.99 in all multiplex assays. The multiplex real-time PCR (TaqMan) assay yielded negative results with the ten other control bacteria. In conclusion, the developed multiplex real-time PCR (TaqMan) assay is highly useful for early diagnosis and differentiation between pathogenic and non-pathogenic leptospires in a reaction tube as having high sensitivity and specificity.
Our results suggest that highly detectable PSA mRNA expression levels in preoperative samples seem to be a significant predictable factor for prostate cancer recurrence.
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.
Introduction: This study aimed to differentiate among the species belonging to the Candida parapsilosis complex, which is one of the leading causes of systemic mycoses through molecular characterisation and to determine their antifungal resistance pattern. Material and Methods: Ninety-five C. parapsilosis complex isolates identified using two yeast identification systems, BioMérieux ID 32C and Vitek 2™ with YST card, were included in this study. Molecular characterisation was achieved by polymerase chain reaction-restriction fragment length polymorphism analysis of the secondary alcohol dehydrogenase gene. Isolates that could not be identified using this technique were identified by internal transcribed spacer (ITS) sequencing. Antifungal susceptibility was evaluated using the Vitek 2™ AST-YS06 card. Results: Out of the 95 isolates, 94 were identified as C. parapsilosis sensu stricto, and only one isolate was identified as C. orthopsilosis; the latter result was achieved using ITS sequencing. C. orthopsilosis was susceptible against all tested antifungal agents. Among the C. parapsilosis sensu stricto isolates, three were found to be resistant: one to amphotericin B, two to fluconazole of which one showed intermediate resistance to voriconazole. Discussion: Isolation rates of C. orthopsilosis and C. metapsilosis in this study show that they are rare species. It was not possible to compare resistance among these three species due to very low isolation rates of C. orthopsilosis and C. metapsilosis. The use of molecular identification tests for these species in routine laboratory settings remains controversial. Nevertheless, their isolation rates and antifungal resistance patterns are important from the epidemiological viewpoint.
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