The role of fat dormouse (Glis glis L.) as reservoir host for spirochete Borrelia burgdorferi sensu lato in the region of Gorski Kotar, Croatia

To determine whether some of the B. burgdorferi sensu lato genospecies associate with fat dormouse as a reservoir host, we investigated the prevalence of infection in questing animals. A total of 45 adult fat dormice (30 female and 15 male) were captured by hunters during their hunting season in the region of Gorski Kotar, Croatia. Dead animals were aseptically dissected, and the urinary bladder tissue was used for isolation attempt and for deoxyribonucleic acid (DNA) extraction. Out of 45 DNA samples extracted from urine bladder tissue, we found four (8.88%) to be polymerase chain reaction (PCR) positive. The RFLP analysis of the PCR product after cleavage with DraI and MseI distinguished between the three major genospecies: B. burgdorferi sensu stricto, B. garinii and B. afzelii. All positive samples were typed as B. afzelii with a unique DraI or MseI pattern. The results of the analysis of urinary bladder tissue samples culture for the presence of Borrelia were negative. Results showed that a prevalence of the Borrelia infection among population of fat dormice indicated their epizootiological involvement as a reservoir of Borrelia spirochetes. Furthermore, this work is an initial step in the investigation of the molecular epidemiology/epizootiology of Lyme borreliosis in Croatia.     

Kinetic Constant Variability in Bacterial Oxidation of Elemental Sulfur

Wide ranges of growth yields on sulfur (from 2.4 × 10¹⁰ to 8.1 × 10¹¹ cells g⁻¹) and maximum sulfur oxidation rates (from 0.068 to 1.30 mmol liter⁻¹ h⁻¹) of an Acidithiobacillus ferrooxidans strain (CCM 4253) were observed in 73 batch cultures. No significant correlation between the constants was observed. Changes of the Michaelis constant for sulfur (from 0.46 to 15.5 mM) in resting cells were also noted.     

Apoptose et ségrégation méiotique dans les spermatozoïdes d'hommes porteurs de translocations

Men with a chromosomal translocation produce a significant percentage of unbalanced spermatozoa. In order to determine a correlation between chromosomal anomalies and apoptosis in human sperm, we analysed DNA fragmentation and meiotic segregation in sperm from men with a (13;14) Robertsonian translocation. We studied sperm from 12 (13;14) translocation carriers and 9 proven fertile men with a normal karyotype. Meiotic segregation of chromosomes 13 and 14 was analysed using dual-colour fluorescencein situ hybridization with locus-specific probes for chromosomes 13 and 14. Apoptosis in spermatozoa was measured byin situ TUNEL assay. The meiotic segregation study showed a significantly increased frequency of unbalanced spermatozoa for chromosomes 13 and 14 in (13;14) carriers (15.9%) compared to the control population (1.3%) (p=0.00016). The study of apoptosis showed an increase of DNA fragmentation in (13;14) carriers (34.9%) compared to the control population (13.8%) (p=0.0036). This increased apoptosis was observed in spermatozoa presenting an increase of unbalanced chromosomal anomalies concerning chromosomes 13 and 14, but with a predominance of balanced spermatozoa compared to the theoretical risk of meiotic segregation. These results suggest that apoptosis could be involved as a regulatory mechanism to eliminate unbalanced chromosomal spermatozoa in men with a (13;14) Robertsonian translocation.     

The secretion of N-acetylglucosaminidase during germ-tube formation in Candida albicans

N-Acetylglucosaminidase was induced by either N-acetylglucosamine or N-acetylmannosamine in several strains of Candida albicans. Enzyme activity was not induced in a N-acetylglucosamine non-utilizing mutant which is unable to express the first three steps in the N-acetylglucosamine catabolic pathway. The enzyme, purified 500-fold, had a specific activity of 36.8 units (mg protein)-1 and catalysed the hydrolysis of p-nitrophenyl-beta-n-acetylglucosamine, N,N'-diacetylchitobiose and N,N',N"-triacetylchitotriose. No activity was observed toward colloidal chitin, hyaluronic acid or mucin. The cellular distribution of N-acetylglucosaminidase was determined by measuring in situ enzyme activity before and after acid treatment of intact cells. N-Acetylglucosaminidase (80-88% of the total cellular activity) was rapidly secreted to the periplasm when the enzyme was induced either during yeast growth at 28 degrees C or germ-tube formation at 37 degrees C. Export of the enzyme from the periplasm into the medium was fourfold greater during germ-tube formation, and after 6 h incubation the amount of enzyme released into the medium represented 70% of cell-associated enzyme activity.     

Final thermal conditions override the effects of temperature history and dispersal in experimental communities

Predicting the effect of climate change on biodiversity is a multifactorial problem that is complicated by potentially interactive effects with habitat properties and altered species interactions. In a microcosm experiment with communities of microalgae, we analysed whether the effect of rising temperature on diversity depended on the initial or the final temperature of the habitat, on the rate of change, on dispersal and on landscape heterogeneity. We also tested whether the response of species to temperature measured in monoculture allowed prediction of the composition of communities under rising temperature. We found that the final temperature of the habitat was the primary driver of diversity in our experimental communities. Species richness declined faster at higher temperatures. The negative effect of warming was not alleviated by a slower rate of warming or by dispersal among habitats and did not depend on the initial temperature. The response of evenness, however, did depend on the rate of change and on the initial temperature. Community composition was not predictable from monoculture assays, but higher fitness inequality (as seen by larger variance in growth rate among species in monoculture at higher temperatures) explained the faster loss of biodiversity with rising temperature.     

Multiresistance of Staphylococcus xylosus and Staphylococcus equorum from Slovak Bryndza cheese

Staphylococcus xylosus, Staphylococcus equorum, and Staphylococcus epidermidis strains were isolated from Bryndza cheese and identified using PCR method. The antimicrobial susceptibility of these strains was assessed using disc diffusion method and broth microdilution method. The highest percentage of resistance was detected for ampicillin and oxacillin, and in contrary, isolates were susceptible or intermediate resistant to ciprofloxacin and chloramphenicol. Fourteen of the S. xylosus isolates (45 %) and eleven of the S. equorum isolates (41 %) exhibited multidrug resistance. None of the S. epidermidis isolate was multiresistant. The phenotypic resistance to oxacillin was verified by PCR amplification of the gene mecA.     

A virus-like particle-based Epstein-Barr virus vaccine

Epstein-Barr Virus (EBV) is an ubiquitous human herpesvirus which can lead to infectious mononucleosis and different cancers. In immunocompromised individuals, this virus is a major cause for morbidity and mortality. Transplant patients who did not encounter EBV prior to immunosuppression frequently develop EBV-associated malignancies, but a prophylactic EBV vaccination might reduce this risk considerably. Virus-like particles (VLPs) mimic the structure of the parental virus but lack the viral genome. Therefore, VLPs are considered safe and efficient vaccine candidates. We engineered a dedicated producer cell line for EBV-derived VLPs. This cell line contains a genetically modified EBV genome which is devoid of all potential viral oncogenes but provides viral proteins essential for the assembly and release of VLPs via the endosomal sorting complex required for transport (ESCRT). Human B cells readily take up EBV-based VLPs and present viral epitopes in association with HLA molecules to T cells. Consequently, EBV-based VLPs are highly immunogenic and elicit humoral and strong CD8+ and CD4+ T cell responses in vitro and in a preclinical murine model in vivo. Our findings suggest that VLP formulations might be attractive candidates to develop a safe and effective polyvalent vaccine against EBV.     

Mitochondrial Complex I: structure, function, and implications in neurodegeneration

Mitochondrial Complex I (NADH Coenzyme Q oxidoreductase) is the least understood of respiratory complexes. In this review we emphasize some novel findings on this enzyme that are of relevance to the pathogenesis of neurodegenerative diseases. Besides Coenzyme Q (CoQ), also oxygen may be an electron acceptor from the enzyme, with generation of superoxide radical in the mitochondrial matrix. The site of superoxide generation is debated: we present evidence based on the rational use of several inhibitors that the one-electron donor to oxygen is an iron-sulphur cluster, presumably N2. On this assumption we present a novel mechanism of electron transfer to the acceptor, CoQ. Strong evidence is accumulating that electron transfer from Complex I to Complex III via CoQ is not performed by operation of the CoQ pool but by direct channelling within a super-complex including Complex I, Complex III and bound CoQ. Besides structural evidence of a Complex I -Complex III aggregate obtained by native electrophoresis, we have obtained kinetic evidence based on metabolic flux analysis, demonstrating that Complexes I and III behave as an individual enzyme. Quantitative and qualitative changes of phospholipids, including peroxidation, may affect the supercomplex formation. Complex I is deeply involved in pathological changes, including neurodegeneration. Maternally inherited mutations in mitochondrial DNA genes encoding for Complex I subunits are at the basis of Leber's Hereditary Optic Neuropathy; a decrease of electron transfer in the complex, due to the mutations, is not sufficient per se to explain the clinical phenotype, and other factors including proton translocation and oxygen radical generation have been considered of importance. Complex I changes are also involved in more common neurological diseases of the adult and old ages. In this review we discuss Parkinson's disease, where the pathogenic involvement of Complex I is better understood; the accumulated evidence on the mode of action of Complex I inhibitors and their effect on oxygen radical generation is discussed in terms of the aetiology and pathogenesis of the disease.     

Role of Genetic Variants of Autophagy Genes in Susceptibility for Non-Medullary Thyroid Cancer and Patients Outcome

Autophagy is a central process in regulation of cell survival, cell death and proliferation and plays an important role in carcinogenesis, including thyroid carcinoma. Genetic variation in autophagy components has been demonstrated to influence the capacity to execute autophagy and is associated with disease susceptibility, progression and outcome. In the present study, we assessed whether genetic variation in autophagy genes contributes to susceptibility to develop thyroid carcinoma, disease progression and/or patient outcome. The results indicate that patients carrying the ATG5 single nucleotide polymorphisms rs2245214 have a higher probability to develop thyroid carcinoma (OR 1.85 (95% CI 1.04–3.23), P = 0.042). In contrast, no significant differences could be observed for the other genetic variants studied in terms of thyroid carcinoma susceptibility. Furthermore, none of the selected genetic variants were associated with clinical parameters of disease progression and outcome. In conclusion, genetic variation in ATG5, a central player in the autophagy process, is found to be associated with increased susceptibility for thyroid carcinoma, indicating a role for autophagy in thyroid carcinogenesis.