Virulence determinants and antibiotic resistance in staphylococci isolated from the skin of captive bred reptiles.

Knowledge of the composition and properties of skin microbiota in healthy reptiles is essential for preservation strategies and thus the prevention of skin dysbiosis leading to dermatological diseases. Despite the greatly increasing popularity of reptiles as pets, only a few studies have dealt with this topic. Therefore, the aim of this work was to analyse species composition of bacteria isolated from skin swabs of 40 reptiles (17 species) using MALDI-TOF spectrometry and to characterise the virulence properties of identified staphylococci (n = 51). The most common species were Staphylococcus xylosus and S. sciuri. Bacilli, enterococci, Escherichia coli, Salmonella sp. and Acinetobacter sp. were also common. The most frequent antimicrobial resistance in staphylococcal isolates was observed for ampicillin (100.0%) and cefoxitin (98.0%) with the blaZ gene being most prevalent (58.8%). In contrast, all staphylococci were susceptible to gentamicin, kanamycin and imipenem. Slime and biofilm production was observed in 86.3% and 76.5% of isolates, respectively. Gelatinase, DNase, protease and lipase activity was found more rarely (41.2%; 25.5%; 27.5% and 21.6%). Since reptiles are a reservoir of bacteria for their owners, common multi-drug resistance (84.3%, MAR index average 0.29 ± 0.09) and biofilm formation must be kept in mind, especially in the case of injury when handling reptiles.

High Prevalence of Clostridioides difficile Ribotype 176 in the University Hospital in Kosice.

Dysbiosis of the gut microbiota, caused by antibiotics, plays a key role in the establishment of Clostridioides difficile CD). Toxin-producing strains are involved in the pathogenesis of Clostridioides difficile infection (CDI), one of the most common hospital-acquired infections. We cultured a total of 84 C. difficile isolates from stool samples of patients hospitalized at Louis Pasteur University Hospital in Kosice, Slovakia, that were suspected of CDI and further characterized by molecular methods. The presence of genes encoding toxin A, toxin B, and binary toxin was assessed by toxin-specific PCR. CD ribotypes were detected using capillary-based electrophoresis ribotyping. A total of 96.4% of CD isolates carried genes encoding toxins A and B, and 54.8% of them were positive for the binary toxin. PCR ribotyping showed the presence of three major ribotypes: RT 176 (n = 40, 47.6%); RT 001 (n = 23, 27.4%); and RT 014 (n = 7, 8.3%). Ribotype 176 predominated among clinical CD isolates in our hospital. The proportion of RT 176 and RT 001 in four hospital departments with the highest incidence of CDI cases was very specific, pointing to local CDI outbreaks. Based on our data, previous use of antibiotics represents a significant risk factor for the development of CDI in patients over 65 years of age.

Antibiotic resistance in the invasive bacteria Escherichia coli.

OBJECTIVES: The beta-lactamases with extended spectrum of activity (ESBL) are medically one of the most important group of enzymes. Another group of beta-lactamases representing of Enterobacteriaceae is group of the AmpC-type cephalosporinases. The presented study provides identification and determination of the spectrum of resistance against different and clinically used antimicrobial drugs in the clinical isolates of Escherichia coli. METHODS: These isolates had origin in different departments of the L. Pasteur University Hospital in Kosice. The goal was the detection of beta-lactamase production with extended-spectrum effect and testing of AmpC-type cephalosporinases by several phenotypic tests in clinical isolates. MALDI-TOF MS analysis was performed on a Microflex MALDI Biotyper. Samples were positively tested for ESBL with the use of the disc diffusion method. PCR were performed with a series of primers designed for the detection of Ambler class A, B and C beta-lactamase genes. RESULTS: For all 485 isolates, we determined the production of ESBL, which we detected in 166 E. coli isolates, which represents a 34.2% prevalence of ESBL production. It is clear from the results that the prevalence of ESBL-producing E. coli out of the total number of E. coli investigated reached 34.2%. In the monitored period, we confirmed at least one resistance gene from 485 E. coli in 188 positive isolates. CONCLUSIONS: We describe a complex ESBL epidemiology. The study revealed a high rate of ESBL-producing E. coli isolates; blaTEM and blaSHV enzymes dominated in ESBL-positive E. coli isolates in the L. Pasteur University Hospital in Kosice.

Virulence, phylogenetic background and antimicrobial resistance in Escherichia coli associated with extraintestinal infections.

BACKGROUND: Escherichia coli (E. coli) is a major cause of urinary tract infections and bloodstream infections and an important agent in the resistance to antibiotics. The present study sought to determine associations between virulence, phylogenetic background and antimicrobial resistance of E. coli strains isolated from patients with extraintestinal infections. METHODS: A total of three hundred ten E. coli strains were isolated from blood, skin and soft tissue and urine. PCR methods were used to detect four main phylogenetic groups (A, B1, B2 and D) and 11 virulence genes (3 toxins, 3 adhesins, 1 siderophore, 4 capsule synthesis proteins and protectins). Standard broth microdilution test was used to determine sensitivity to 12 antimicrobial drugs. RESULTS: The most common and the most virulent phylogenetic group B2 was found in 193 (62.3%) isolates. The lowest virulence was observed among the group A. Analysis of virulence factors revealed the kpsMTII gene in 212 (68.4%), aer in 194 (62.6%) and tra in 184 (59.4%) of isolates, respectively. Multi-drug resistant (MDR) phenotype was noticed in 165 (53.2%) isolates. Lower representation of the MDR phenotype was detected in E. coli containing all groups of virulence genes and in the avirulent E. coli. CONCLUSIONS: Our study documented that E. coli associated with 3 different extraintestinal infections contain various virulence factors. Genes afa, pap, aer, neuC show significant differences among the 3 groups of the strains tested and might be the prerequisite virulence factors in bloodstream infections. Isolates containing all groups of virulence genes predominantly originate in the blood and belong to the B2 phylogenetic group. Overall, we identified significantly higher incidence of all the groups of virulence genes examined among the B2 group. Prevalence of the MDR phenotype and high levels of resistance to ampicillin, ciprofloxacin and trimetoprim/sulfamethoxazole reflect the trend observed worldwide in recent years.

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.

Hepameta-- prevalence of hepatitis B/C and metabolic syndrome in population living in separated and segregated Roma settlements: a methodology for a cross-sectional population-based study using community-based approach.

BACKGROUND: Roma represent one of the largest and oldest minorities in Europe. Health of many of them, particularly those living in settlements, is heavily compromised by poor dwelling, low educational level, unemployment, and poverty rooted in generational poverty, segregation and discrimination. The cross-sectional population-based study using community based approach aimed to map the prevalence of viral hepatitis B/C and metabolic syndrome in the population living in separated and segregated Roma settlements and to compare it with the occurrence of the same health indicators in the majority population, considering selected risk and protective factors of these health indicators. METHODS: The sample consisted of 452 Roma (mean age = 34.7; 35.2% men) and 403 non-Roma (mean age = 33.5; 45.9% men) respondents. Data were collected in 2011 via questionnaire, anthropometric measures and analysed blood and urine samples. A methodology used in the study as well as in the following scientific papers is described in the Methods section (i.e. study design, procedures, samples, methods including questionnaire, anthropometric measurements, physical measurements, blood and urine measurements). CONCLUSIONS: There are regions of declining prosperity due to high unemployment, long-term problems with poverty and depleted resources. Populations living in these areas, i.e. in Central and Eastern Europe in Roma settlements, are at risk of poverty, social exclusion and other factors affecting health. Therefore, we should look for successful long-term strategies and tools (e.g. Roma mediators, terrain work) in order to improve the future prospects of these minorities.

Association between metabolic syndrome and hepatitis B virus infection in the Roma population in eastern Slovakia: a population-based study.

BACKGROUND: The simultaneous presence of chronic hepatitis B (CHB) and metabolic syndrome (MS) in the high-risk Roma community constitutes a high risk for liver cirrhosis and potentially hepatocellular carcinoma. This study aims to explore the relationship between MS and CHB. METHODS: Data from the cross-sectional HepaMeta Study conducted in Slovakia in 2011 among Roma living in rural communities were used. Participants were tested for the presence of MS, and lipid levels--total cholesterol, high density lipoproteins (HDL), low density lipoproteins (LDL), triglycerides (TG), apolipoprotein B100, and CHB HBsAg and anti-HBc IgG were also monitored. Viral load was measured in HBsAg-positive patients. RESULTS: A total of 452 patients were screened; MS was diagnosed in 29.6% of patients, and 12.5% had CHB. Anti-HBc IgG antibodies were present in 52.8% of patients. CHB patients had lower levels of total cholesterol (5.45 +/-1.21 vs. 4.71 +/- 1.23 mmol/l; p = 0.035), LDL cholesterol (median 2.2 mmol/l, interquartile range 0.88 mmol/l vs. 2.5 mmol/l, interquartile range 0.9 mmol/l; p = 0.01) and apolipoprotein B100 (median 0.66 mmol/l, interquartile range 0.26 mmol/l vs. 0.74 mmol/l, interquartile range 0.29 mmol/l; p = 0.025). Patients diagnosed with MS had a higher HBV DNA load than patients without MS (1,728.2 +/- 14.33 IU/ml vs. 12,779.1 +/- 20.9 IU/ml; p = 0.037). CHB patients with TC and apolipoprotein B100 within the reference range had a lower hepatitis B DNA (HBV DNA) load than patients with high or low values of TC or apolipoprotein B100. CONCLUSION: The prevalence of chronic hepatitis B and simultaneous presence of MS was high among Roma. HBsAg-positive patients had lower levels of total and LDL cholesterol along with decreased apolipoprotein B100. The viral load of chronic hepatitis B patients with MS was higher than in patients without MS.

Gamma-glutamyl transpeptidase level associated with metabolic syndrome and proinflammatory parameters in the young Roma population in eastern Slovakia: a population-based study.

BACKGROUND: Elevated gamma-glutamyl transpeptidase (GGT) is present approximately in half of all patients with non-alcoholic fatty liver disease (NAFLD). NAFLD is the liver manifestation of metabolic syndrome (MS). This study aimed to explore the relationship between GGT and MS or proinflammatory parameters. METHODS: Data from the cross-sectional HepaMeta study conducted in Slovakia in 2011 among Roma living in rural communities were used. Participants (n = 446) were divided into 2 groups; those with elevated GGT and those with normal GGT levels. MS was diagnosed according to the International Diabetes Federation criteria; presence of central obesity and low density lipoproteins (LDL) or high density lipoproteins (HDL), high triglycerides, hypertension, glucose intolerance or type 2 diabetes. Participants were tested for the presence of MS and its components, and biochemical tests for lipid levels (total cholesterol, HDL, LDL, TG) and inflammatory parameters (high sensitivity C-reactive protein--hs-CRP and ferritin) were performed. RESULTS: Of 446 Roma participants, only 29 (6.5%) had GGT levels above the normal value. After exclusion of patients with viral hepatitis and alcohol abuse, patients with elevated GGT suffered from MS more often (p < 0.001), and patients with more MS components had a higher risk of elevated GGT. We found a significant association between GGT and the individual MS components, except HDL (waist circumference > or = 94 cm in men or 80 cm in women: p < 0.01; BMI > 30: p < 0.001; fasting glucose > or = 5.6 mmol/l: p < 0.001; arterial hypertension: p < 0.05, and TAG > or = 1.7 mmol/l: p < 0.001). Patients with elevated GGT levels had also significantly higher hs-CRP (hs-CRP > 2 mg/l: p < 0.001; hs-CRP > 3 mg/l: p < 0.001) and ferritin (ferritin > 300 mg/l: p < 0.01) levels. CONCLUSION: Patients with MS have more significantly elevated levels of GGT. There is a significant association of GGT with individual MS components, except HDL and inflammatory parameters (hs-CRP, ferritin).

Prevalence of cardiovascular risk factors in relation to metabolic syndrome in the Roma population compared with the non-Roma population in the eastern part of Slovakia.

BACKGROUND: Obesity-induced metabolic syndrome is a multiple risk factor for cardiovascular (CV) risk factors and type 2 diabetes, and ethnic minorities seem to have unfavourable medical risk factors in general more frequently than majority populations. OBJECTIVE: The aim of this study was to evaluate the prevalence of cardiovascular risk factors in relation to metabolic syndrome in the Roma population compared with the non-Roma population residing in the eastern part of Slovakia. RESULTS: 123 Roma and 79 non-Roma patients with metabolic syndrome were evaluated. Men between 40-55 years of age had 4.76-times higher odds and women 5.26-times higher odds for metabolic sydrome compared with the younger population. We found statistically significant higher waist circumference in the Roma subpopulation and higher body mass index as well, although in selected population with metabolic syndrome. HDL cholesterol was significantly lower in both Roma men and women, and LDL cholesterol was not significant in men and women with metabolic syndrome. Triglycerides levels were significantly higher in non-Roma women only. High-sensitivity C-reactive protein (hsCRP) values were not in correlation with age but were associated with the increasing number of fulfilled criteria for metabolic syndrome in both subgroups (Roma, non-Roma), independently of gender. CONCLUSION: Our study confirmed higher prevalence of obesity, metabolic syndrome and other CV risk factors associated with metabolic syndrome among younger Roma population, which may be associated with increased cardiovascular disease (CVD) morbidity and mortality among elderly Roma compared with non-Roma.

High hepatitis B and low hepatitis C prevalence in Roma population in eastern Slovakia.

BACKGROUND: Viral hepatitis B and C prevalence in the Roma population of eastern Slovakia is largely unknown. This study aimed to explore the prevalence and associated risk factors of chronic viral hepatitis B and C among Roma living in segregated communities in eastern Slovakia. METHODS: Data from the cross-sectional HepaMeta study conducted in Slovakia in 2011 among Roma living in rural communities were used. Participants were tested for the presence of HBsAg, anti-HBc IgG and anti-HCV. The risk factors were assessed mainly via a structured questionnaire/interview. RESULTS: Altogether 452 Roma were screened, and 11 were excluded due to missing data. A total of 441 patients were included (mean age 34.7 +/- 9.14 years; 35.2% men). 12.5% of participants were HBsAg positive, 40.4% anti-HBc IgG positive while negative for HBsAg and 47.2% of participants were negative for all serological markers of hepatitis B. Hepatitis C prevalence was very low (0.7%), while 2 out of 3 anti-HCV positive participants were coinfected with hepatitis B. Risk factors for hepatitis B infection were male sex, higher age, tattoo, and previous imprisonment. No difference was found in intravenous drug use, blood transfusions and sexual behaviour. CONCLUSION: More than half of the Roma residing in eastern Slovakia have been infected at one point in life with the hepatitis B virus, and 12.5% are HBsAg positive. Hepatitis C prevalence is very low, which is probably due to very low intravenous drug use.

Clinical and biochemical determinants of metabolic syndrome among Roma and non-Roma subjects in the eastern part of Slovakia.

BACKGROUND: The metabolic syndrome (MS) is a clustering of cardiovascular risk. The high prevalence of metabolic syndrome among populations of lower socioeconomic status is a cause of concern and calls for an effective public health response. OBJECTIVES: The aim of this study was to determine the prevalence of metabolic syndrome in the Roma population compared with the non-Roma population in the eastern part of Slovakia and to determine the parameter which has the strongest association with metabolic syndrome. RESULTS: 123 Roma and 79 non-Roma patients with metabolic syndrome were evaluated. In the subgroup of Roma men, we found that waist circumference conferred the highest chance of MS (more than 12-times), followed by triglycerides (TG) (3.670-times). In the subgroup of non-Roma men, we found that waist circumference conferred the highest chance of MS (more than 16-times), followed by high-density lipoprotein (HDL) (4.348-times increased risk per one unit decrease in HDL). In the subgroup of Roma women as well as non-Roma women, we found that serum TG conferred the highest chance of MS, followed by waist circumference for Roma women. Comparing non-classical risk factors for MS we found that only age (with OR 1.977) and high-sensitivity C-reactive protein (hsCRP) (OR 1.887) were significant and independent predictors of MS in Roma men. Among Roma women apolipoprotein B100 was also found to be an independent predictor of MS, besides age and hsCRP. CONCLUSION: Our study confirmed that the prevalence of metabolic syndrome is strongly associated with hypertriglyceridemic waist, besides other risk factors, a marker of the atherogenic metabolic triad among younger Roma population, which may be the reason for the increased cardiovascular (CV) morbidity and mortality in elderly Roma compared with non-Roma. In light of these results, better prevention of CV events for Roma minority settlements in Slovakia should be provided.

Hepatitis B virus infection in patients with metabolic syndrome: a complicated relationship. Results of a population based study.

BACKGROUND: The presence of hepatitis B infection (HBI) and metabolic syndrome (MS) at the same time constitutes a high risk for liver cirrhosis and potentially hepatocellular carcinoma. AIM: In this study we aim to explore the relationship between MS and HBI. METHODS: We used data from the cross-sectional HepaMeta study conducted in 2011 in Slovakia. Patients were tested for presence of MS, while lipid levels (total cholesterol, HDL, LDL, TG, apolipoprotein B100 and HBI (HBsAg and antiHBcIgG)) were also monitored. Viral load was measured in HBsAg positive patients. RESULTS: Altogether 855 patients were screened, MS was diagnosed in 25.1% of patients and 7.9% of patients presented with HBI. AntiHBcIgG antibodies were present in 34.6% patients. HBI patients had lower levels of total and LDL cholesterol along with a decreased apolipoprotein B100 (4.54 ± 0.84 vs. 5.0 ± 0.99 mmol/l, P=0.001; 2.29 ± 0.58 vs. 2.6 ± 0.68 mmol/l, P=0.001 and 0.71 ± 0.21 vs. 0.77 ± 0.23 mmol/l, P=0.013 respectively). Patients diagnosed with MS had higher HBV DNA load than patients without MS - 1300.2 (95% CI 506.06-3440.41) vs. 7661.3 (95% CI 2008.17-29,228.06) IU/ml; P=0.011. HBI patients with TC and apolipoprotein B100 in the reference range had lower HBV DNA load than patients with high or low values of TC or apolipoprotein B100. CONCLUSION: Hepatitis B patients had lower levels of total and LDL cholesterol along with a decreased apolipoprotein B100. Viral load of chronic hepatitis B patients with MS was higher than that in patients without MS.

[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.

Involvement of virulence properties and antibiotic resistance in Escherichia coli strains causing pyelonephritis in children.

A total of 145 Escherichia coli strains causing pyelonephritis in children were investigated for the prevalence of genes encoding the following virulence factors (VFs): P fimbria (67.6 %), S fimbria (53.8 %), AFA adhesins (2.8 %), cytotoxic necrotizing factor 1 (37.9 %), α-hemolysin (41.4 %), and aerobactin (71.7 %). One hundred and thirty-six (93.8 %) isolates harbored at least one of the virulence genes detected in the present study. Statistically significant co-occurrent presence of two VF genes was found for α-hly-cnf1, α-hly-sfa, cnf1-sfa (p<0.001), and α-hly-pap (p=0.001). Twenty-six profiles of VF genes were detected in this study. The combinations of aer-pap and aer-pap-sfa-α-hly-cnf1 were presented with the highest frequency-both of them in 28 isolates (19.3 %). All E. coli strains included in the study were susceptible to meropenem, amikacin, and tobramycin; the highest frequency resistance was found toward ampicillin (43.4 %), piperacillin (31.7 %), tetracycline (15.9 %), and trimethoprim/sulfamethoxazole (11.7 %). The resistance to the other tested antimicrobial drugs did not exceed 3 % incidence. Overall, 55.9 % strains were susceptible to all tested anti-infective agents. Antimicrobial resistance of E. coli strains toward trimethoprim/sulfamethoxazole statistically significantly correlated with the presence of α-hly (p<0.001), sfa (p<0.01), and cnf1 (p<0.05).

Prevalence, epidemiological aspects and clinical importance of TT virus infection in Slovakia.

The aim of the study was to assess the prevalence, clinical impact and importance of different risk factors of transmission of TTV infection in Slovakia using two PCR methods. Sera of 426 adult persons were examined. TTV DNA was identified by PCR using primers from N22 and untranslated region (UTR) respectively. The established prevalence of TTV tested with N22 and UTR primers according to patients groups was: acute hepatitis of unknown etiology 4 resp. 28 of 37, acute hepatitis B 3 resp. 29 of 38, chronic hepatitis B 11 resp. 41 of 44, chronic hepatitis C 10 resp. 93 of 102, hemodialysis patients 13 resp. 72 of 72, health care workers 0 resp. 27 of 33, control group 8 resp. 83 of 100. Using N22 primers, TTV infection occurred more frequently in chronic hepatitis B group compared with health care workers, if UTR primers were used the group of hemodialysis patients differed significantly from both acute hepatitis groups, health care workers and controls (p < 0.05). From possible risk factors hemodialysis and transfusion count showed notable differences. Bilirubin and aminotransferase levels did not differ between TTV positive and negative groups. No pathogenetic role of TT virus in liver injury was confirmed.

[Selected mechanisms of resistance to antimicrobial drugs in Pseudomonas aeruginosa and their detection]. / Vybrané mechanizmy rezistencie na antimikrobiálne látky u Pseudomonas aeruginosa a ich detekcia.

By now most clinically significant bacterial species have resistance mechanisms against almost all accessible antibiotics. One of the most significant nosocomial pathogens is Pseudomonas aeruginosa. Its antibiotic resistance directly correlates with higher morbidity, mortality and longer hospital stay. An essential condition for preserving the antibiotic efficacy against this pathogen is the exploration of its resistance mechanisms. To protect itself against the action of antibacterials, P. aeruginosa uses the cellular wall, which prevents the drug from entering the bacterium, as also an active efflux from the cell. It is also capable of modifying the molecule of the antimicrobial. The bacterium can effectively combine all these mechanisms and the result are multiresistant bacterial clones. The paper describes some diagnostic methods that reveal the resistance mechanisms of the species Pseudomonas aeruginosa.