Virulence and antibiotic resistance genes in Campylobacter spp. in the Czech Republic.

OBJECTIVE: Thermotolerant species of the genus Campy-lobacter are the important agents causing human foodborne infections throughout the world. The aims of this study were to evaluate the presence of nine putative virulence genes in Campylobacter spp. isolated from patients and from foods (poultry meat, pork liver), to determine the resistance of Campylobacter isolates to eight antibiotic agents and to detect four resistance genes.Matherial and methods: The presence of the virulence genes cdtA, cdtB, cdtC, virB11, ciaB, wlaN, iam, dnaJ and racR was detected by polymerase chain reaction (PCR) in 94 Campylobacter spp. isolates from humans and 123 campylobacters from foods. The phenotypic resistance to selected antimicrobial agents was tested with microdilution method in 82 human isolates and 91 food isolates. The isolates with antibiograms were tested for the presence of blaOXA-61, tet(O), aph-3-1 and cmeB genes by PCR with specific primers. RESULTS: In both human and food C. jejuni isolates the preva-lence of the studied virulence genes, especially dnaJ, racR, ciaB genes and the toxigenic genes cdtA, cdtB, cdtC, was considerably higher than in C. coli isolates. The only exception was the iam gene identified in only C. coli. The tested isolates of both C. jejuni and C. coli were highly resistant to quinolone antibiotics. Additionally, C. coli was also more resistant to erythromycin, streptomycin and, in case of isolates from pork liver, to tetracycline. High prevalence rates of genes encoding antibiotic resistance was noted for the blaOXA-61 and tet(O) genes in both Campylobacter species. CONCLUSIONS: The presented study is the first to assess the presence of genes for virulence and resistance to antibiotics in thermotolerant Campylobacter spp. isolated from humans and foods in the Czech Republic. The resistance of Campylobacter isolates to eight antibiotic agents was also assessed. The prevalence of genes responsible for virulence and resistance is rather varied in thermotolerant Campylobacter spp.

[Prevalence and characteristics of thermotolerant Campylobacter spp. in the human food chain]. / Výskyt a charakteristika termotolerantních kampylobakteru v potravinovém retezci cloveka.

OBJECTIVES: To monitor the prevalence of Campylobacter spp. in poultry in slaughterhouses, poultry and pork liver at retail, and cows milk in Moravia. To determine the resistance of animal isolates to selected antibiotics; and to compare it with an antibiogram of human strains. MATERIAL AND METHODS: Throughout the year 2013, the following samples were collected in the South Moravian and Olomouc Regions: mixed samples of broiler cecal contents in slaughterhouses, fresh and frozen chickens and pork liver at retail, and raw cows milk from vending machines. The samples were both qualitatively and quantitatively analyzed for the presence of Campylobacter spp. The isolates recovered were tested for resistance to antibiotics. For comparison, antimicrobial resistance was also studied in human isolates from the same regions. RESULTS: A total of 41.8% of the tested food samples were found to contain Campylobacter spp.. The most contaminated (73.2%) were fresh chickens. Campylobacter spp. were not detected in raw cows milk samples. The isolates showed high levels of resistance to quinolone antibiotics and, in the case of C. coli, also to tetracycline and streptomycin. CONCLUSION: The studied commodities were frequently contaminated with Campylobacter spp. The levels of contamination (in CFU/g) varied between commodities and so, evidently, did the real risk for human infections. When antibiotic therapy is needed, quinolone antibiotics cannot be used. Adherence to high standards of consumer safe food handling is crucial for the prevention of diseases.

[Zoonotic potential of raw cow's milk in the Czech Republic]. / Zoonotický potenciál syrového kravského mléka v CR.

BACKGROUND: To assess current microbiological risks of raw cow's milk. The presented work reports the prevalence of selected bacteria on Czech dairy farms in 2010 and compares the results with a similar study in 2002. MATERIAL AND METHODS: The prevalence of bacteria was studied by culture methods using milk filters made from non-woven fabric collected from dairy farms throughout the year 2010. Together 260 filters from 65 dairy farms were tested. The prevalence of Campylobacter spp., Listeria monocytogenes, Staphylococcus aureus, Escherichia coli O 157 and Salmonella spp. in filters was tested. In staphylococci, genes encoding enterotoxin-production were studied. In Campylobacter spp., resistance to antibiotics was ascertained. RESULTS: In 2010, the prevalence rates of selected bacteria on farms were as follows: Campylobacter spp. 3%, Escherichia coli O 157 0.4%, Salmonella spp. 0.8%, Listeria monocytogenes 10% and Staphylococcus aureus 31%. In 12% of the tested filters, S. aureus with the genetic make-up for enterotoxin production was isolated. Antibiotic resistance in Campylobacter spp. isolated from dairy farms is different from that in human or poultry isolates. CONCLUSION: Raw cow's milk contains bacteria capable of causing human alimentary tract diseases. The above microbiological risks for consumers are eliminated by proper storage conditions (below 10 degrees C) and heat treatment (boiling) of raw milk.

[Detection of ESBL-positive strains of Escherichia coli in pigs in the Czech Republic]. / Záchyt ESBL-pozitivních kmenu Escherichia coli u prasat v Ceské republice.

BACKGROUND: The goals of the presented regional study were monitoring the prevalence of Enterobacteriaceae producing broad-spectrum beta-lactamases in slaughter pigs and their basic molecular biology analysis. MATERIAL AND METHODS: In the presented study, rectal swabs from 118 slaughter pigs from 5 farms in the Olomouc Region were analyzed. Bacteriological tests aimed at detection and identification of bacteria from the Enterobacteriaceae family producing broad-spectrum beta-lactamases. Suspected isolates were further analyzed using both phenotypic and genotypic methods. RESULTS: From the group of 118 analyzed samples, seven Escherichia coli strains with the presence of the bla genes encoding CTX-M-1 beta-lactamases were isolated. Genes for TEM and SHV enzymes were not detected. This is the first report of ESBL-positive isolates of Escherichia coli in pigs in the Czech Republic.

[Brucella suis--a little-known zoonotic agent]. / Brucella suis--málo známé zoonotické agens.

Brucellosis is a rare but serious bacterial zoonosis. Officially, the Czech Republic is among countries that are free from brucellosis in livestock. In the country, sporadic imported human infections may occur, caused by e.g. Brucella melitensis. In wild hare populations, however, rare cases of infection caused by Brucella suis are still observed, potentially threatening humans. The short communication reports two cases of hare brucellosis in Moravia, Czech Republic, and experiences with isolate identification by the MALDI-TOF method.

Probing the reaction mechanism between a laser welded polyamide thin film and titanium with XPS and ToF-SIMS.

The biomedical industry uses more and more polymer/metal hybrid assemblies because of the ability to combine the advantages and lower the inconveniences of both materials. The key is to assemble them. Among the high variety of existing assembling techniques, laser welding appears as an excellent option. It is a quick process allowing a great design flexibility, high reproducibility without intermediate material needed to create the adhesion, which is advantageous for biomedical applications. The laser welding process creates strong adhesion between dissimilar materials, but the root cause for adhesion is still unclear. The analytical challenge is to gain an information at the molecular level from an interface that is deeply buried between the two materials. Such a study requires extremely surface sensitive analytical methods, such as ToF-SIMS or XPS in order to detect chemical bonds, but also a method to expose the interface to the X-ray or ion beam. In order to investigate the chemical bonding at the interface between polyamide-6.6 and titanium, mirror polished titanium surfaces were prepared, on which a thin polyamide-6.6 film was spin-coated. The samples were laser welded, and after dissolving the polymer thin film, XPS and ToF-SIMS measurement were performed. The ToF-SIMS data interpretation was assisted by a principal component analysis. This multivariate analysis is rather common for ToF-SIMS data but is more rarely used to solve adhesion problems. This allowed to show the nature of the chemical bond at the interface and to propose a reaction mechanism.

Extended-spectrum beta-lactamase-producing Escherichia coli in turkey meat production farms in the Czech Republic: national survey reveals widespread isolates with bla(SHV-12) genes on IncFII plasmids.

AIM: The occurrence and epidemiology of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in the environment of turkey farms in the Czech Republic were studied. METHODS AND RESULTS: Extended-spectrum beta-lactamase-producing E. coli isolates were found on 8 (20%) of 40 turkey farms surveyed. A total of 200 environmental smears were examined, and a total of 25 ESBL-producing E. coli were isolated. These isolates were analysed using XbaI pulsed-field gel electrophoresis and divided into nine pulsotypes. Most of the isolates harboured the gene bla(SHV-12) on a 40-kb plasmid of the IncFII group with an identical EcoRV restriction profile. Indistinguishable or clonally related SHV-12-producing isolates belonging to the same pulsotypes were found at some unrelated farms. CONCLUSIONS: Widespread occurrence of ESBL-producing E. coli isolates with bla(SHV-12) carried on IncFII plasmids in meat production flocks in the Czech Republic was demonstrated. SIGNIFICANCE AND IMPACT OF THE STUDY: Results indicate vertical transmission of ESBL-producing E. coli within the turkey production pyramid. The study shows the risk of multiresistant ESBL-producing bacteria and antibiotic-resistance genes being transmitted to humans via the food chain.

Epidemiology and characterization of Staphylococcus epidermidis isolates from humans, raw bovine milk and a dairy plant.

Geographically related Staphylococcus epidermidis isolates from human patients (n=30), dairy farms (farmers and individual raw milk from cattle, n=36) and a dairy plant (n=55) were examined for epidemiological relatedness by pulsed-field gel electrophoresis and, using in vitro methods, for the ability to produce biofilm and antimicrobial resistance. Methicillin-resistant isolates (MRSE) were also identified and characterized. Isolates from farmers and dairy cattle were found to be genetically related, while isolates from human patients were highly diverse. Some dairy plant isolates (18.2%) were closely related to those from dairy farms. Biofilm production and resistance to antimicrobial agents were most typical for isolates from human patients, of which 76.7% were MRSE. Methicillin resistance was also widespread in farm-related isolates (61.1%). This study indicates the possible transmission of S. epidermidis between cattle and farmers. Dairy products were not proven to be an important source of either human infections or methicillin-resistant strains.

Hybrid layers deposited by an atmospheric pressure plasma process for corrosion protection of galvanized steel.

Finding alternative treatments to reproduce anticorrosion properties of chromated coatings is challenging since both physical barrier and self-healing effects are needed. Siloxane based treatments are known to be a promising way to achieve physical barrier coatings, mainly plasma polymerized hexamethyldisiloxane (ppHMDSO). In addition, it is known that cerium-based coatings can also provide corrosion protection of metals by means of self-healing effect. In this frame, innovative nanoAlCeO3/ppHMDSO layers have thus been deposited and studied. These combinations allow to afford a good physical barrier effect and active properties. Liquid siloxane and cerium-based particles mixture is atomized and introduced as precursors into a carrier gas. Gas mixture is then injected into an atmospheric pressure dielectric barrier discharge (DBD) where plasma polymerization of the siloxane precursor occurs. The influence of cerium concentration on the coating properties is investigated: coating structure and topography have been studied by scanning electron microscopy (SEM) and interferometry, and corrosion resistance of these different coatings is compared by electrochemistry techniques: polarization curves and electrochemical impedance spectroscopy (EIS). Potential self-healing property afforded by cerium in the layer was studied by associating EIS measurements and nanoscratch controlled damaging. Among the different combinations investigated, mixing of plasma polymerized HMDSO and AICeO3 nanoparticles seems to give promising results with a good physical barrier and interesting electroactive properties. Indeed, corrosion currents measured on such coatings are almost as low as those measured with the chromated film. Combination of nanoscratch damaging of layers with EIS experiments to investigate self-healing also allow to measure the active protection property of such layers.

Isolation of Leptospira interrogans from suburban rats in Tandil, Buenos Aires, Argentina.

The main objective of this study was to investigate the role of wild rodents as Leptospira spp. reservoirs in a suburban area of Tandil city, Buenos Aires province (Argentina), where a person had died due to pulmonary leptospirosis. The specific objectives were: to estimate the rodent density near the patient's home, to determine the serological prevalence and isolation of leptospirosis from wild rodents, and to identify the isolated strains. The area examined was a suburban neighbourhood in Tandil near the Langueyú stream, where the patient's house is located. Rattus norvegicus were trapped on the stream banks during two nights and a high capture rate (70%), was obtained. All rats (42) were examined serologically by the microscopic agglutination test (MAT), and 22 of them (52.3%) reacted with Leptospira serovars castellonis, canicola, grippotyphosa, icterohaemorrhagiae and hebdomadis at a titer of 1:50. The kidneys from 25 animals were cultured, and 24 isolates of L. interrogans (96%) were obtained. The isolated strains were identified as Icterohaemorrhagiae serogroup by MAT performed with rabbit hyperimmune reference sera. These findings showed a high density of suburban rodents highly infected with pathogenic leptospira, sharing environment in close contact with humans with evidence of leptospiral disease.

Chronic wasting disease.

Chronic wasting disease (CWD) is the only known prion disease affecting free-ranging animals and has become a serious epidemic in North America. Although any case was reported from Europe, the spread of the disease to other continents and regions cannot be excluded, because the transmission of CWD is the most efficient among prion diseases. This article reviews the host range of CWD including experimentally infected animals, models for potential transmissibility to humans, clinical signs of the disease, pathogenesis, and methods for CWD detection.

Sepsis induces long-term metabolic and mitochondrial muscle stem cell dysfunction amenable by mesenchymal stem cell therapy.

Sepsis, or systemic inflammatory response syndrome, is the major cause of critical illness resulting in admission to intensive care units. Sepsis is caused by severe infection and is associated with mortality in 60% of cases. Morbidity due to sepsis is complicated by neuromyopathy, and patients face long-term disability due to muscle weakness, energetic dysfunction, proteolysis and muscle wasting. These processes are triggered by pro-inflammatory cytokines and metabolic imbalances and are aggravated by malnutrition and drugs. Skeletal muscle regeneration depends on stem (satellite) cells. Herein we show that mitochondrial and metabolic alterations underlie the sepsis-induced long-term impairment of satellite cells and lead to inefficient muscle regeneration. Engrafting mesenchymal stem cells improves the septic status by decreasing cytokine levels, restoring mitochondrial and metabolic function in satellite cells, and improving muscle strength. These findings indicate that sepsis affects quiescent muscle stem cells and that mesenchymal stem cells might act as a preventive therapeutic approach for sepsis-related morbidity.

Conducting, semiconducting and insulating objects observed by low-energy electron holography.

To demonstrate the potential of low energy in line projection holography, we study the reconstruction of the experimental holograms of three electrically different objects: a conducting, a semiconducting and an insulating object. The reconstructions of these holograms provide meaningful results for a large range of magnification of the object. The comparison between the reconstructed images and the scanning electron microscopy (SEM) micrographs of the same objects shows that the shapes and the dimensions of the reconstructed objects are identical to those obtained by conventional SEM. So, the simple assumptions needed to the reconstruction are justified. The reconstructions show a 2 nm resolution and appear superior than the best obtained SEM micrographs, when available. We also show some limitations of the reconstruction process. We point out that both numerical artifacts and experimental conditions are responsible for these limitations.

Imaging charged objects using low-energy-electron coherent beams.

The microscopic shapes, unobservable by conventional electron optics, of objects surrounded by electrostatic fields are observed. The method directly uses electron-coherent-beam imaging concepts and is successfully applied to recently developed low-energy-electron projection holography. Experimental demonstrations on objects charged in a controlled way like a biased metallic tip as well as on a self-charging insulator object are shown.

Improved low energy electron projection in-line holograms reconstruction: application to the holograms of a tungsten tip.

A modification of the classical calculation algorithm is presented which allows to minimize the effect of the long range oscillations created by the twin object in the reconstruction process of in-line holograms obtained by low energy electron projection microscopy. The reconstructed shapes of a tungsten tip, observed with different magnifications are shown.

Plesiomonas shigelloides and its serovars in animals in the Czech Republic--region Moravia.

In the period of 1994-1997, 4,552 samples of sectional, clinical material and environment were examined for the presence of Plesiomonas shigelloides. In 55 cases (1,21%) Plesiomonas shigelloides was isolated. Within 55 isolated strains, 26 serovars have been successfully identified. Among other secured cases was Plesiomonas shigelloides isolated in pig (Sus scrofa domestica), mouse Mus musculus), binturong (Arctictis binturong) and raccoon (Pocyon lotor). In mandarin duck (Aix galericulata) the strain of Plesiomonas shigelloides with a new 0 antigen-097 was isolated. In regard of the particular biological classes, the isolation of Plesiomonas shigelloides in fish (2.63%) predominate, followed by reptiles (1.66%), mammals (0.94%) and birds (0.79%). Plesiomonas shigelloides in amphibians was not demonstrated. This work surveys the incidence of Plesiomonas shigelloides and its serovars in animals in the region Moravia in the Czech Republic.

[Group A bovine rotavirus: diagnosis and antigenic characterization of strains circulating in the Argentine Republic, 1994-1999]. / Rotavirus bovino grupo A: diagnóstico y caracterización antigénica de cepas circulantes en la República Argentina, 1994-1999.

Group A Bovine Rotavirus (BRV) has been identified as a major cause of neonatal diarrhea in cattle. The study was aimed to determine the prevalence of BRV and to antigenically characterize the G-types of circulating strains in dairy and beef herds in Argentina. A total of 1129 stool samples from diarrheic calves was analyzed from 1994 to 1999. The samples were initially screened for RV by ELISA and PAGE, and then G-typed using monoclonal antibodies (Mab) directed against G1, G2, G3, G6 and G10-specific epitopes. Forty percent (452/1129) of the samples were positive for RV by ELISA, while 24.7% (279/1129) were also positive for PAGE. VP7 was detected in the 70.5% (319/452) of the positive samples using a broadly reactive Mab (C60); 32.6% (104/319) were G6, 15.4% (49/319) were G10, and 6% (19/319) were G1. However, 46.1% (147/319) of the samples remained untypable. Rotavirus diarrhea prevalences were comparable in beef and dairy herds (87.3% and a 74.4%, respectively). Finally, G6 was the most prevalent G-type circulating in beef herds while G10 prevailed in dairy herds. A better understanding of RV epidemiology will contribute to the optimization of current vaccines and prevention programs of RV diarrhea in calves.

Humoral immune response to infectious agents in aborted bovine fetuses in Argentina.

The purpose of this study was to characterize the exposure of bovine aborted fetuses from beef and dairy herds of the humid pampas of Argentina to different infectious agents by the evaluation of fetal fluid antibodies. Presence of fetal antibodies to bovine viral diarrhea virus genotype 1 (BVDV-1), bovine herpes virus type 1 (BHV-1), Leptospira interrogans, Brucella abortus, and Neospora caninum was determined. Of the 95 fetuses processed, 66 came from 49 beef herds and 29 from 12 dairy herds. The average gestational age of the aborted fetuses was 7.1 months. Antibodies to the mentioned agents were detected in 65 of the 95 fetal fluids (68.4%). In addition, antibodies to more than one infectious agent were detected in 32 fetuses (33.7%), suggesting fetal exposure to multiple antigens during gestation. There were antibodies to BVDV-1, BHV-1, N. caninum and Leptospira interrogans in 43 (45.2%), 29 (30.5%), 26 (27.4%) and 5 (5.2%) specimens, respectively. Antibodies to B. abortus were not detected in any of the fetal fluids. The results of this study provide information on the determination of antibodies in fluids from bovine aborted fetuses exposed to different infectious agents in the region.

Prevalence and characteristics of Escherichia coli strains producing extended-spectrum ß -lactamases in slaughtered animals in the Czech Republic.

Resistance of bacteria to antibiotics is a global medical problem requiring close cooperation between veterinary and human physicians. Raw materials and foods of animal origin may be not only a source of pathogenic bacteria causing alimentary tract infections but also a source of bacteria with a dangerous extent of resistance to antibiotics, potentially entering the human food chain. This article presents results of the first study in the Czech Republic detecting the presence of Enterobacteriaceae-producing extended-spectrum b -lactamases (ESBLs) in swabs collected in slaughterhouses from surfaces of healthy animal carcasses. In 2012, swabs taken from pig (n = 166) and cattle (n = 140) carcass surfaces were analyzed. In 17 % of 53 studied slaughterhouses, ESBL-producing Escherichia coli strains were isolated. ESBLs were found in 11 and 4 % of porcine and bovine samples, respectively. Swabs collected from pigs yielded 18 ESBL-producing E. coli strains. The bla genes were found to encode production of CTX-M-1 group enzymes in 16 strains, SHV in one case, and both CTX-M-1-like and TEM in another case. In swabs taken from cattle, five ESBL-producing E. coli strains were isolated. In three cases, the bla genes for CTX-M-1-like production were identified; in two cases, genes for both CTX-M-1-like and TEM production were found. The similarity/identity of ESBL-positive isolates was compared by pulsed-field gel electrophoresis. This is the first report and characterization of the presence and nature of ESBL-producing E. coli in swabs collected from surfaces of healthy pig and cattle carcasses in slaughterhouses in the Czech Republic.