Design, construction and evaluation of multi-epitope antigens for diagnosis of Lyme disease.

INTRODUCTION: Introduction and objective. Lyme disease (LD) is the most common vector-borne disease in the temperate zone of the Northern Hemisphere. Diagnosis of LD is mainly based on clinical symptoms supported with serology (detection of anti-Borrelia antibodies) and is often misdiagnosed in areas of endemicity. MATERIAL AND METHODS: In this study, the chimeric proteins (A/C-2, A/C-4 and A/C-7.1) consisting of B-cell epitopes of outer surface proteins OspA and OspC from Borrelia genospecies prevalent in Eastern Slovakia, were designed, over-expressed in E. coli, and used to detect specific anti-Borrelia antibodies in serologically characterized sera from patients with Lyme-like symptoms to evaluate their diagnostic potential. RESULTS: Results showed that chimeras vary in their immuno-reactivity when tested with human sera. Compared with the results obtained from a two-tier test, the application of recombinant multi-epitope chimeric proteins as diagnosis antigens, produced fair agreement in the case of A/C-2 (0.20<κ<0.40) and good agreement (0.60<κ<0.80) when A/C-7.1 was used as capture antigen. Chimera A/C-4 were excluded from further study due to loss of reactivity with OspA-specific antibodies. CONCLUSIONS: The combination of specific B-cell epitopes from OspA and OspC proteins may improve the diagnostic accuracy of serologic assays, but further studies are required to address this hypothesis.

Distribution and antifungal susceptibility of yeasts isolates from intensive care unit patients.

Yeasts frequently colonize non-sterile sites in the body. The aim of the study was to determine distribution in clinical samples and antifungal susceptibility to five antifungals. From January 2013 through June 2015, 800 isolates were obtained from intensive care unit patients. Candida albicans (58.9%), Candida glabrata (20.4%), Candida krusei (8.6%), and Candida parapsilosis (3.6%) were the leading species. Majority of the C. albicans isolates were susceptible to the fluconazole. Elevated voriconazole minimal inhibitory concentrations (MICs) were observed in isolates exhibiting high fluconazole MICs, most frequently in C. glabrata. Isolates with echinocandins MICs suggesting reduced susceptibility were only sporadic cases with the exception of Trichosporon spp. The amphotericin B MICs were slightly higher for some C. krusei.

Identification of B-cell epitopes of Borrelia burgdorferi outer surface protein C by screening a phage-displayed gene fragment library.

Outer surface protein C (OspC) of Borrelia stimulates remarkable immune responses during early infection and is therefore currently considered a leading diagnostic and vaccine candidate. The sensitivity and specificity of serological tests based on whole protein OspC for diagnosis of Lyme disease are still unsatisfactory. Minimal B-cell epitopes are key in the development of reliable immunodiagnostic tools. Using OspC fragments displayed on phage particles (phage library) and anti-OspC antibodies isolated from sera of naturally infected patients, six OspC epitopes capable of distinguishing between LD patient and healthy control sera were identified. Three of these epitopes are located at the N-terminus (OspC E1 aa19-27, OspC E2 aa38-53, OspC E3 aa62-66) and three at the C-terminal end (OspC E4 aa155-163, OspC E5 aa184-190 and OspC E6 aa201-207). OspC E1, E4 and E6 were highly conserved among LD related Borreliae. To our knowledge, epitopes OspC E2, E3 and E5 were identified for the first time in this study. Minimal B-cell epitopes may provide fundamental data for the development of multi-epitope-based diagnostic tools for Lyme disease.

[Methods of phenotypic determination of New Delhi metallo-beta-lactamase (NDM-1) in Klebsiella penumoniae isolated in Slovakia].

OBJECTIVES: Reported is the first isolation and phenotypic determination of Klebsiella pneumoniae producing New Delhi metallo-beta-lactamase (NDM-1) isolated from patients in the Slovak Republic. MATERIAL AND METHODS: Between 27 October 2012 and 22 January 2013, twenty-five isolates of Klebsiella pneumoniae collected from 5 patients were identified with MIC of meropenem ≥ 32 mg/L and MIC of ertapenem ≥ 4 mg/L in screening tests. Next, all isolates were assessed with the modified Hodge test, combined disk test with EDTA, double disk synergy test with EDTA and MBL E-test. To confirm production of MBL in isolated strains of Klebsiella pneumoniae, all strains were sent to the National Reference Center for Antibiotic Resistance in Bratislava. RESULTS: All strains were positive in all phenotypic tests. In the first carbapenem-resistant isolate, NDM-1 production was confirmed by PCR amplification, sequencing and comparison with the GenBank. CONCLUSIONS: To the best of our knowledge, this is the first case of isolation of NDM-1 from Klebsiella pneumoniae in the Slovak Republic. As of 31 January 2014, with well-established and strict epidemiological and preventive measures, there was no further spread or another outbreak of NDM-1 producing Enterobacteriaceae in Louis Pasteur University Hospital in Kosice.