Marcela Pejchalová

Prevalence and diversity of Arcobacter spp. in the Czech Republic.

The aim of this study was to examine 634 samples of chicken, lamb, pork, beef, fish, samples from the intensive animal industry and from poultry for slaughter, as well as from the domestic breeding of poultry, horses, pigs, and lambs, from surface water, and from clinical samples for the presence of Arcobacter. All the samples were examined with a cultivation method, followed by confirmation by multiplex PCR. The method of multiplex PCR applied directly to a liquid medium after enrichment was applied only to the samples with the highest probability of the presence of arcobacters. Arcobacter spp. were detected in 11.8% of the samples, of which A. butzleri, A. cryaerophilus, and A. skirrowii were found in 6.6, 5.1, and 0.2% of the samples, respectively. The sources of the arcobacters were chicken meat from the retail market, intensive animal production facilities, domestic chicken breeding facilities, lamb raising environments, surface water and wastewater, and beef swabs taken in a meat processing factory. No occurrence of arcobacters was identified in the swabs from slaughter turkeys, ducks, and wild poultry. No arcobacters were found in horse and pig breeding environments, on pork, or on the swabs of fish. Forty-two rectal swabs taken from humans were also free of Arcobacter. Seventeen isolates of Arcobacter were further identified by sequencing the 16S rRNA gene. Varied genotypes were observed among A. butzleri from chicken meat and chicken breeds, and A. cryaerophilus from wastewater and chicken breeds. They were similar to the genotypes present in wastewater, porcine feces, human stool, and human blood obtained from databases. Our results revealed that the chicken meat from the retail market is an important source of arcobacters. Cross-contamination during handling of chicken carcass practices could play a key role in the spread of Arcobacter.

Isolation ofArcobacter butzleri andA. cryaerophilus in samples of meats and from meat-processing plants by a culture technique and detection by PCR

A pilot survey of sources of contamination with arcobacters (representing a potential risk for humans) was done in a wide range of samples involved various kinds of meat (beef, pork, meat products, chilled chickens,etc.) from a retail level and domestic farming. Sanitary practices in slaughterhouses and production lines were checked in two different plants (a beef and pork production and a chicken processing plant). The method is based on a selective enrichment to isolate suspect strains, in combination with a PCR technique specific for arcobacters. The choice of a suitable enrichment broth and a plating agar was made with the use of pure bacterial strains and by means of real meat samples seeded withArcobacter butzleri. The PCR technique was optimized to allow differentiation of a 1223 bp product, typical of the genusArcobacter, and a product of 686 bp, specific forA. butzleri a total number of 198 samples were tested, of that 33 (17 %) were found to be positive for the genusArcobacter but only 22 (11 %) forA. butzleri.

Inhibitory Effects of Some Spice and Herb Extracts Against Arcobacter butzleri , A. cryaerophilus , and A. skirrowii

Seventeen spice and medicinal plant extracts (methanol and chloroform) were assayed for their antimicrobial activity against Arcobacter butzleri, A. cryaerophilus, and A. skirrowii. In general, all of the tested extracts were able, to a different extent, to inhibit the growth of the selected Arcobacter species. Cinnamon, bearberry, chamomile, sage and rosemary extracts showed strong antimicrobial activity toward arcobacter strains tested. Overall, the methanol extracts showed better activity than the chloroform extracts (P < 0.05); however, enhanced antibacterial activity of chloroform extracts of cinnamon and rosemary has been observed in comparison with their methanol counterparts. The inhibitory dose of the most active extracts (the diameter of zone of inhibition ≥ 20 mm) was determined using the disc-diffusion method as well.

Detection of aflatoxigenic fungi in feeds using the PCR method.

The possibility of using the polymerase chain reaction (PCR) to speed up and specify the detection of aflatoxigenic fungi isolated from feed was investigated. The method, applied to 2 genes encoding the biosynthesis of aflatoxins (apa-2 andver-1), was optimized on two collection cultures (Aspergillus flavus CCM F-108 andA. parasiticus CCM F-550). The specificity of the optimized PCR method was proved on collection cultures of different kinds of fungi. Fifty feed samples out of which 18 showed positive findings of aflatoxigenic fungi on anAspergillus Flavus and Parasiticus Agar (AFPA) medium were tested. Isolated strains ofAspergillus strains were verified using the PCR method; its reaction products were detected in 1% agarose gel by electrophoresis. The results almost exclusively matched those gained from the AFPA medium.

Synthesis, structural characterization, docking, lipophilicity and cytotoxicity of 1-[(1R)-1-(6-fluoro-1,3-benzothiazol-2-yl)ethyl]-3-alkyl carbamates, novel acetylcholinesterase and butyrylcholinesterase pseudo-irreversible inhibitors

In the current study, sixteen novel derivatives of (R)-1-(6-fluorobenzo[d]thiazol-2-yl)ethanamine were synthesized as acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitors. Chemical structures together with purity of the synthesized compounds were substantiated by IR, (1)H, (13)C, (19)F NMR, high resolution mass spectrometry and elemental analysis. The optical activities were confirmed by optical rotation measurements. The synthesized compounds were evaluated for their AChE and BChE inhibitory activities. In addition, the cytotoxicity of the most active compounds was investigated against human cell lines employing XTT tetrazolium salt reduction assay and xCELLigence system allowing a label-free assessment of the cells proliferation. Our results demonstrated that the inhibitory mechanism was confirmed to be pseudo-irreversible, in line with previous studies on carbamates. Compounds indicated as 3b, 3d, 3l and 3n showed the best AChE inhibitory activity of all the evaluated compounds and were up to tenfold more potent than standard drug rivastigmine. The binding mode was determined using state-of-the-art covalent docking and scoring methodology. The obtained data clearly demonstrated that 3b, 3d, 3l and 3n benzothiazole carbamates possess high inhibitory activity against AChE and BChE and concurrently negligible cytotoxicity. In conclusion, our results indicate, that these derivatives could be promising in an effective therapeutic intervention for Alzheimers disease.

Antimicrobial effect of oxidized cellulose salts

Antimicrobial properties of oxidized cellulose and its salts in linters (-L) and microsphere (-M) form (OKCEL® H-L, OKCEL® Zn-M, OKCEL® ZnNa-L, OKCEL® ZnNa-M and OKCEL® Ag-L) were tested by a dilution method against a spectrum of microbial strains: Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Bacillus licheniformis, Aspergillus niger, Penicillium chrysogenum, Rhizopus oryzae, Scopulariopsis brevicaulis, Candida albicans and Candida tropicalis. OKCEL® Ag-L exhibited antimicrobial activity in the range 0.1–3.5% w/v against all the bacteria and fungi involved in this study. Strong inhibition by OKCEL® ZnNa-M was observed for Staphylococcus epidermidis, Bacillus licheniformis, Rhizopus oryzae, Candida albicans and Candida tropicalis in the range 0.5–2.0% w/v. Antimicrobial effects of oxidized cellulose and its salts in textile form were investigated by a diffusion and dilution method against the spectrum of above-cited microbial strains extended by Clostridium perfringens. Generally, OKCEL® Ag-T, OKCEL® Zn-T and OKCEL® H-T showed high antimicrobial activity against populations of Pseudomonas aeruginosa, Bacillus licheniformis and Staphylococcus epidermidis. OKCEL® Zn-T was the only sample suppressing the growth of species.

Persistence of Arcobacter butzleri CCUG 30484 on plastic, stainless steel and glass surfaces

The persistence of A. butzleri CCUG 30484 on various surfaces under 32% and 64% relative humidity suspended in physiological saline or nutrient broth to simulate relatively clean or soiled conditions was studied using various isolation techniques. Our study revealed that A. butzleri CCUG 30484 cells were able to survive for a considerable period of time, even after the droplet of suspending medium has been visibly dried. An extended survival on polypropylene coupons at both humidity levels was observed, particularly at soiled conditions.

Inhibition of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii by plant oil aromatics.

The inhibitory effect of some plant oil aromatics against three strains of Arcobacter butzleri, two strains of Arcobacter cryaerophilus, and one strain of Arcobacter skirrowii was evaluated. When MICs were determined using the broth macrodilution method, cinnamaldehyde was most inhibitory followed by thymol, carvacrol, caffeic acid, tannic acid, and eugenol (P < 0.001). Sublethal concentrations of the three most potent plant oil aromatics also were examined. Overall, cinnamaldehyde was the most bacteriostatic against all arcobacters tested except A. butzleri when these strains were exposed to the MIC25 of this aromatic aldehyde. The bacteriostatic activities of thymol and carvacrol were concentration and species dependent.

Synthesis, structural characterization, antimicrobial and antifungal activity of substituted 6-fluorobenzo[d]thiazole amides

A series of novel 1-[(1R)-1-(6-fluoro-1,3-benzothiazol-2-yl)ethyl]-3-substituted phenyl amides was synthesized by the condensation reaction of (1R)-1-(6-fluoro-1,3-benzothiazol-2-yl)ethanamine with substituted benzoyl chlorides under mild conditions. Their structures were confirmed by 1H NMR, 13C NMR and 19F NMR spectra, elemental analyses and in three cases also by single-crystal X-ray diffraction techniques. The optical activities were confirmed by optical rotation measurements. All the synthesized compounds were screened for antibacterial and antifungal activity against a variety of bacterial and fungal strains. Some of the compounds reveal antibacterial and antifungal activity comparable or slightly better to that of chloramphenicol, cefoperazone and amphotericin B used as medicinal standards.

Susceptibility to 18 drugs and multidrug resistance of Arcobacter isolates from different sources within the Czech Republic

OBJECTIVES Arcobacter spp. are considered to be potential foodborne pathogens, and consumption of contaminated food containing these bacteria could endanger human and animal health. Arcobacter butzleri and Arcobacter cryaerophilus are the species most frequently isolated from food of animal origin and from other samples. The aim of this study was to evaluate the susceptibility of arcobacters isolated in the Czech Republic. No information about antibiotic susceptibility and multidrug resistance of arcobacters isolated in the Czech Republic is available in the literature before now. METHODS The antimicrobial resistance of A. butzleri (n=80) and A. cryaerophilus (n=20) isolated from meat of animal origin, water sources and clinical samples was examined by the disk diffusion method. RESULTS Arcobacters were resistant to one or more antimicrobial agents in 99% (99/100) of tested isolates. Most of the Arcobacter isolates were resistant to β-lactam antibiotics, i.e. ampicillin (81.0%), amoxicillin/clavulanic acid (28.0%), cefalotin (73.0%) and aztreonam (93.0%). Arcobacters were also frequently resistant to lincosamides, i.e. clindamycin (98.0%). Of the aminoglycosides, amikacin, gentamicin and tobramycin were evaluated to be the most effective antibiotics among those tested against arcobacters. CONCLUSIONS These results demonstrate substantial resistance in Arcobacter isolates to 18 antimicrobial agents commonly used in medical and veterinary medicine. Multidrug resistance was found in 93.8% (75/80) of A. butzleri isolates and 70.0% (14/20) of A. cryaerophilus isolates.