The antibiofilm activity of selected substances used in oral health prophylaxis.

Oral health is a window to a patient's general well-being. Balance in oral microbiome functions is crucial for health maintenance. A state of oral dysbiosis may lead to a variety of local and systemic pathological conditions. The presence of dental plaque is related to the majority of oral infections. Proper oral hygiene is crucial and the most economic practice contributing to oral health prophylaxis. Aside from prophylactic treatments provided by dental practitioners, mouth rinses, containing antimicrobial agents, are one of the possible tools used for oral care. Our study was to determine whether available mouth rinses and selected products dedicated for professional use are efficient to eradicate biofilm formed by reference and clinical strains of Streptococcus mutans, Streptococcus sanguinis, Streptococcus oralis, Streptococcus mitis, Staphylococcus aureus, Enterococcus faecalis, Lactobacillus rhamnosus and Candida albicans on the surface of hydroxyapatite - major mineral component of a tooth. Therefore, such antimicrobials as chlorhexidine, cetylpyridine chloride, polyhexanide, silver nanoparticles, sulphonated phenolics, and natural antiplaque essential oils and coconut oil were analyzed. Applied experimental settings in in vitro models were designed to reflect accurately the recommended use of the tested substances, therefore four types of eradication procedure were conducted. Sialorrhea simulation was also performed to evaluate antibiofilm potential of diluted mouth rinses. Biofilm was investigated with quantitative method where absorbance values were measured. Statistical differences were assessed using the Kruskal-Wallis test with a post-hoc Dunnett's analysis. Results have shown that biofilms displayed a diversified sensitivity to the tested antimicrobials. The highest antibiofilm activity was detected for cetylpyridine chloride while the lowest for chlorhexidine. However the differences in E. faecalis biofilm reduction observed after the use of these two compounds were not statistically significant (p > 0.05), whereas all observed differences in S. aureus survival after exposure to the examined antimicrobial agents were statistically significant (p < 0.5). The PHMB, both in standard and in sialorrhea simulated conditions had the highest potential against streptococci. The coconut oil reduced C. albicans fungus biofilm by 65.48% but low eradication level was observed in case of bacterial biofilms. The dehydrating mechanism of action of sulfonated phenolics turned out to be ineffective against streptococcal biofilm which in turn was effectively eradicated by silver nanoparticles. The implementation of Antibiofilm Dressing's Activity Measurement method allowed to observe strain-related differences in terms of antimicrobial sensitivity. The obtained results may be introduced in everyday out-patient dental plaque prophylaxis as well as clinical environment.

Epidemiology of Staphylococcus pseudintermedius in cats in Poland.

Staphylococcus pseudintermedius is a well-known coagulase-positive staphylococcus that is mainly associated with the asymptomatic colonization of the skin of pets and mucous membranes. Little is still known about the occurrence of S. pseudintermedius in cats. The current study aimed to characterize the isolates of S. pseudintermedius from sick and healthy cats. This was achieved by examining their antibiotic resistance properties, biofilm formation, and genotype differences. Six hundred and seventy-six cats were swabbed (595 healthy and 81 sick cats). Thirty-five distinct S. pseudintermedius isolates from 27 cats were isolated. The prevalence of S. pseudintermedius in healthy and sick cats was 2.49% and 7.61%, respectively. In comparison, MRSP (methicillin-resistant Staphylococcus pseudintermedius) prevalence was 0.12% and 2.98%, respectively. Cats were more frequently colonized with S. pseudintermedius when kept with dogs, regardless of their health condition, with this result being statistically significant. Multidrug resistance was detected in 50%, and 38.46% of S. pseudintermedius isolates from healthy and sick cats, respectively. In contrast, genetic multidrug resistance was detected in 59% and 46.15% cases, respectively. Seven from eight isolated MRSPs were multidrug-resistant. Multi-locus sequence typing (MLST) assigned isolates to 19 types, of which 16 types submitted for the first time to the PubMLST database. The most frequently detected STs (sequence types) were 551 and 71. ST71 and ST551 were mainly isolated from cats with clinical signs of infection. All were MRSPs, regardless of cats' health. These isolates were characterized with the most frequent antibiotic resistance at the phenotypic and genotypic level.

Nasal carriage of various staphylococcal species in small ruminant lentivirus-infected asymptomatic goats.

The study was carried out in Polish goat population to estimate the prevalence of the nasal cavity infection with various staphylococcal species including methicillin-resistant Staphylococcus aureus(MRSA), investigate the potential permissive role of small ruminant lentivirus (SRLV) infection and determine the level of clonality of S. aureus nasal isolates. Nasal swabs and blood samples were collec-ted from 1300 clinically healthy adult goats from 21 Polish goat herds. Blood samples were serological-ly screened for SRLV. Staphylococci were isolated from nasal swabs and identified using classical microbiological methods, MALDI-TOF, multiplex-PCR, and their clonality was assessed using PFGE. Antimicrobial resistance was determined on the basis of minimum inhibitory concentration and by demonstration of the presence of the mecA gene encoding the multiplex-PCR PBP2a protein and of the five main types of staphylococcal cassette chromosome mec. The apparent prevalence of staphylococ-cal and S. aureus infection of the nasal cavity was 29.1% (CI 95%: 26.9%, 31.5%) and 7.3% (CI 95%: 6.1%, 8.8%), respectively. No relationship was found between the SRLV-infection and the presence of any staphylococcal species including S. aureus (p=0.143). Only 9.8% of S. aureus isolates were resistant to amoxicillin/clavulanic acid and 5.9% to chloramphenicol and ciprofloxacin. All tested isolates proved to be phenotypically and genotypically sensitive to methicillin, which yielded the appar-ent prevalence of MRSA of 0% (CI 95%: 0%, 7.0%). S. aureus isolates show high genetic similarity within goat herds, however vary considerably between herds. Goats do not appear to be an important source of S. aureus for humans in Poland.

Prevalence of Staphylococcus Species Colonization in Healthy and Sick Cats.

Staphylococcus is one of the most frequently isolated genera of opportunistic bacteria in animals and human beings. Staphylococci in mammals mostly inhabit the skin and mucous membranes. The objectives of the study were to investigate the distribution of staphylococcal species in healthy and sick cats in order to find diagnostic markers. The risk factors associated with colonization were also explored. Isolates from healthy (n=520) and sick cats (n=67) were identified at the species level using matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS). Swabs from conjunctival sacs, nares, skin, anus, and wounds were investigated using this technique. The diversity of the Staphylococcus species was high: 26 and 17 species in healthy and sick cats, respectively, and predominantly coagulase-negative staphylococci (CoNS) were isolated. The most frequently observed were S. felis and S. epidermidis in healthy cats, whereas S. felis and S. haemolyticus were most often found in sick animals. S. aureus strains were only isolated from healthy cats, whereas the only coagulase-positive Staphylococcus (CoPS) which occurred in the sick cats group was S. pseudintermedius. The sick, more frequently than the healthy animals, were colonized with S. pseudintermedius and S. haemolyticus and the relationship was statistically significant. Mostly, regardless of the state of their health, similar Staphylococcus species were isolated from cats; therefore, particular attention should be paid during the interpretation of diagnostic results.

Wasp venom immunotherapy in a patient with immune-mediated inflammatory central nervous system disease: is it safe?

No disponible

Genetic polymorphisms and expression of HLA-G and its receptors, KIR2DL4 and LILRB1, in non-small cell lung cancer.

Human leukocyte antigen-G (HLA-G) is a nonclassical HLA class I molecule absent from most normal tissues but detected in many malignant tumors. It is recognized by cells of the immune system using LILRB1, KIR2DL4 and LILRB2 receptors. We attempted to find out whether some polymorphisms of HLA-G, LILRB1 and KIR2DL4 genes are associated with susceptibility to nonsmall cell lung cancer (NSCLC). Four polymorphisms in HLA-G, i.e. -964A>G (rs1632947), -725C>G>T (rs1233334), -716T>G (rs2249863) in the promoter, and a 14 base pair insertion/deletion (14 bp indel) in the 3'-untranslated region (3'UTR), and five in LILRB1 - 5651G>A (rs41308748) in intron 14, 5717C>T L622L (rs1061684), 5724G>A E625K (rs16985478), 5774 C>A P641P (rs41548213) in exon 15, and 5806C>T (rs8101240) in 3'UTR - as well as 9620 9A/10A (rs11410751) polymorphism in exon 7 of KIR2DL4 were typed using different laboratory techniques. Only one single nucleotide polymorphism (SNP) in HLA-G (-964A>G) and one in LILRB1 (5724G>A) were found to influence the risk of NSCLC. In addition, 5724G>A was associated with protection from tumor cell infiltration of regional lymph nodes. Most importantly, we detected HLA-G and LILRB1 expression in tumor specimens, but no correlation with genetic polymorphisms was observed. HLA-G and LILRB1 protein expression levels in tumor tissue were significantly correlated with tumor stage.

Diffusion-weighted magnetic resonance imaging of breast lesions: the influence of different fat-suppression techniques on quantitative measurements and their reproducibility.

OBJECTIVE: The aim of this study was to evaluate the influence of different fat-suppression techniques on quantitative measurements and their reproducibility when applied to diffusion-weighted imaging (DWI) of breast lesions. METHODS: Twenty-five patients with different types of breast lesions were examined on a clinical 1.5-T magnetic resonance imaging (MRI) system. Two diffusion-weighted sequences with different fat-suppression methods were applied: one with spectral presaturation by inversion recovery (SPIR), and one with short-TI inversion recovery (STIR). The acquisition of both sequence variants was repeated with modified shim volume. Lesion-to-background contrast (LBC), apparent diffusion coefficients (ADC) ADC(0,1000) and ADC(50,1000), and their coefficients of variation (CV) were determined. RESULTS: In four patients, the image quality of DWI with SPIR was insufficient. In the other 21 patients, 46 regions of interest (ROI), including 11 malignant and 35 benign lesions, were analysed. The LBC, ADC(0,1000) and ADC(50,1000) values, which did not differ between initial and repeated measurements, were significantly higher for STIR than for SPIR. The mean CV improved from 10.8 % to 4.0 % (P = 0.0047) for LBC, from 6.3 % to 2.9 % (P = 0.0041) for ADC(0,1000), and from 6.3 % to 2.6 % (P = 0.0049) for ADC(50,1000). CONCLUSION: For STIR compared to SPIR fat suppression, improved lesion conspicuity, higher ADC values, and better measurement reproducibility were found in breast DWI. KEY POINTS: • Quality of fat suppression influences quantitative DWI breast lesion measurements. • In breast DWI, STIR fat suppression worked more reliably than SPIR. • Lesion-to-background contrast and its reproducibility were significantly higher with STIR fat suppression. • Lesional ADCs and their reproducibility were significantly higher with STIR fat suppression.

Ternary Pt/Rh/SnO2 electrocatalysts for oxidizing ethanol to CO2.

Ethanol, with its high energy density, likely production from renewable sources and ease of storage and transportation, is almost the ideal combustible for fuel cells wherein its chemical energy can be converted directly into electrical energy. However, commercialization of direct ethanol fuel cells has been impeded by ethanol's slow, inefficient oxidation even at the best electrocatalysts. We synthesized a ternary PtRhSnO(2)/C electrocatalyst by depositing platinum and rhodium atoms on carbon-supported tin dioxide nanoparticles that is capable of oxidizing ethanol with high efficiency and holds great promise for resolving the impediments to developing practical direct ethanol fuel cells. This electrocatalyst effectively splits the C-C bond in ethanol at room temperature in acid solutions, facilitating its oxidation at low potentials to CO(2), which has not been achieved with existing catalysts. Our experiments and density functional theory calculations indicate that the electrocatalyst's activity is due to the specific property of each of its constituents, induced by their interactions. These findings help explain the high activity of Pt-Ru for methanol oxidation and the lack of it for ethanol oxidation, and point to the way to accomplishing the C-C bond splitting in other catalytic processes.

Self-perpetuating changes in Sup35 protein conformation as a mechanism of heredity in yeast.

Recently, a novel mode of inheritance has been described in the yeast Saccharomyces cerevisiae. The mechanism is based on the prion hypothesis, which posits that self-perpetuating changes in the conformation of single protein, PrP, underlie the severe neurodegeneration associated with the transmissible spongiform enchephalopathies in mammals. In yeast, two prions, [URE3] and [PSI+], have been identified, but these factors confer unique phenotypes rather than disease to the organism. In each case, the prion-associated phenotype has been linked to alternative conformations of the Ure2 and Sup35 proteins. Remarkably, Ure2 and Sup35 proteins existing in the alternative conformations have the unique capacity to transmit this physical state to the newly synthesized protein in vivo. Thus, a mechanism exists to ensure replication of the conformational information that underlies protein-only inheritance. We have characterized the mechanism by which Sup35 conformational information is replicated in vitro. The assembly of amyloid fibres by a region of Sup35 encompassing the N-terminal 254 amino acids faithfully recapitulates the in vivo propagation of [PSI+]. Mutations that alter [PSI+] inheritance in vivo change the kinetics of amyloid assembly in vitro in a complementary fashion, and lysates from [PSI+] cells, but not [psi-] cells, accelerate assembly in vitro. Using this system we propose a mechanism by which the alternative conformation of Sup35 is adopted by an unstructured oilgomeric intermediate at the time of assembly.

[Molecular mimicry of bacterial polysaccharides and their role in etiology of infectious and autoimmune diseases]. / Mimikra czasteczkowa bakteryjnych antygenów polisacharydowych i jej rola w etiologii chorób infekcyjnych i autoimmunologicznych.

Molecular mimicry is one of the most important pathogenic factor of microorganism and is defined as a structural similarity of microbial molecules to host tissue contributing to the pathogenicity. Mimicry can be observed at the molecular, serological and functional level. In the review the infectious diseases have been discussed where the mimicry phenomenon may occur, and also autoimmune disease where due to the molecular mimicry bacterial structures are potent to induce adverse immune reactions. The cross-reacting molecules mimicking the host structures comprise colominic acid, sialic acid containing capsular polysaccharides of Streptococcus group B, phosphocholine containing antigen, lipopolysaccharides of Campylobacter jejuni contributing in induction of Guillain-Barré syndrome or Lewis antigen containing lipopolysaccharides of Helicobacter pylori inducing gut carcinoma. Knowledge on the phenomenon of molecular mimicry is important when new conjugate vaccine has to be constructed, because great care should be paid not to induce autoantibodies with synthetic immunogen. Investigation of microbial factors reveal that many autoimmune diseases are of infection etiology.

[Alcohol related hospitalization in the Department of Clinical Toxicology and Medicine Detoxification Ward of the Krakow Special Hospital of J. Babinski in Krakow]. / Alkohol jako przyczyna hospitalizacji w Klinice Toksykologii Collegium Medicum UJ i Oddziale Detoksykacji Szpitala Specjalistycznego im. J. Babinskiego w Krakowie.

The patients hospitalised because of acute ethanol intoxication or ethanol withdrawal syndrome at the Kraków Department of Clinical Toxicology and Detoxification Ward of J. Babinski Specialistic Hospital in the years 1997-2000 were subjected to the study. A significant increase in number of ethanol related hospitalisation was noted at the Department of Clinical Toxicology (from 1381 patients in year 1997 up to 1771 in year 2000), and at the Detoxification Ward of Babinski Hospital from 369 in 1997 and 849 patients in 2000 respectively. A significant increase in number of ethanol dependent patients admitted to the Department of Clinical Toxicology because of acute ethanol intoxication and the patients hospitalised because of ethanol withdrawal syndrome or signs of delirium tremens was noted. The same and even stronger trends in hospitalisation, particularly of those ethanol addicts presenting clinical symptoms of acute ethanol intoxication were observed at the Detoxification Ward of Babinski Hospital.

Bidirectional amyloid fiber growth for a yeast prion determinant.

The polymerization of many amyloids is a two-stage process initiated by the formation of a seeding nucleus or protofibril. Soluble protein then assembles with these nuclei to form amyloid fibers. Whether fiber growth is bidirectional or unidirectional has been determined for two amyloids. In these cases, bidirectional growth was established by time lapse atomic-force microscopy. Here, we investigated the growth of amyloid fibers formed by NM, the prion-determining region of the yeast protein Sup35p. The conformational changes in NM that lead to amyloid formation in vitro serve as a model for the self-perpetuating conformational changes in Sup35p that allow this protein to serve as an epigenetic element of inheritance in vivo. To assess the directionality of fiber growth, we genetically engineered a mutant of NM so that it contained an accessible cysteine residue that was easily labeled after fiber formation. The mutant protein assembled in vitro with kinetics indistinguishable from those of the wild-type protein and propagated the heritable genetic trait [PSI(+)] with the same fidelity. In reactions nucleated with prelabeled fibers, unlabeled protein assembled at both ends. Thus, NM fiber growth is bidirectional.

Twenty-first aminoacyl-tRNA synthetase-suppressor tRNA pairs for possible use in site-specific incorporation of amino acid analogues into proteins in eukaryotes and in eubacteria.

Two critical requirements for developing methods for the site-specific incorporation of amino acid analogues into proteins in vivo are (i) a suppressor tRNA that is not aminoacylated by any of the endogenous aminoacyl-tRNA synthetases (aaRSs) and (ii) an aminoacyl-tRNA synthetase that aminoacylates the suppressor tRNA but no other tRNA in the cell. Here we describe two such aaRS-suppressor tRNA pairs, one for use in the yeast Saccharomyces cerevisiae and another for use in Escherichia coli. The "21st synthetase-tRNA pairs" include E. coli glutaminyl-tRNA synthetase (GlnRS) along with an amber suppressor derived from human initiator tRNA, for use in yeast, and mutants of the yeast tyrosyl-tRNA synthetase (TyrRS) along with an amber suppressor derived from E. coli initiator tRNA, for use in E. coli. The suppressor tRNAs are aminoacylated in vivo only in the presence of the heterologous aaRSs, and the aminoacylated tRNAs function efficiently in suppression of amber codons. Plasmids carrying the E. coli GlnRS gene can be stably maintained in yeast. However, plasmids carrying the yeast TyrRS gene could not be stably maintained in E. coli. This lack of stability is most likely due to the fact that the wild-type yeast TyrRS misaminoacylates the E. coli proline tRNA. By using error-prone PCR, we have isolated and characterized three mutants of yeast TyrRS, which can be stably expressed in E. coli. These mutants still aminoacylate the suppressor tRNA essentially quantitatively in vivo but show increased discrimination in vitro for the suppressor tRNA over the E. coli proline tRNA by factors of 2.2- to 6.8-fold.

Subunit interactions influence the biochemical and biological properties of Hsp104.

Point mutations in either of the two nucleotide-binding domains (NBD) of Hsp104 (NBD1 and NBD2) eliminate its thermotolerance function in vivo. In vitro, NBD1 mutations virtually eliminate ATP hydrolysis with little effect on hexamerization; analogous NBD2 mutations reduce ATPase activity and severely impair hexamerization. We report that high protein concentrations overcome the assembly defects of NBD2 mutants and increase ATP hydrolysis severalfold, changing V(max) with little effect on K(m). In a complementary fashion, the detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate inhibits hexamerization of wild-type (WT) Hsp104, lowering V(max) with little effect on K(m). ATP hydrolysis exhibits a Hill coefficient between 1.5 and 2, indicating that it is influenced by cooperative subunit interactions. To further analyze the effects of subunit interactions on Hsp104, we assessed the effects of mutant Hsp104 proteins on WT Hsp104 activities. An NBD1 mutant that hexamerizes but does not hydrolyze ATP reduces the ATPase activity of WT Hsp104 in vitro. In vivo, this mutant is not toxic but specifically inhibits the thermotolerance function of WT Hsp104. Thus, interactions between subunits influence the ATPase activity of Hsp104, play a vital role in its biological functions, and provide a mechanism for conditionally inactivating Hsp104 function in vivo.

Nucleated conformational conversion and the replication of conformational information by a prion determinant.

Prion proteins can serve as genetic elements by adopting distinct physical and functional states that are self-perpetuating and heritable. The critical region of one prion protein, Sup35, is initially unstructured in solution and then forms self-seeded amyloid fibers. We examined in vitro the mechanism by which this state is attained and replicated. Structurally fluid oligomeric complexes appear to be crucial intermediates in de novo amyloid nucleus formation. Rapid assembly ensues when these complexes conformationally convert upon association with nuclei. This model for replicating protein-based genetic information, nucleated conformational conversion, may be applicable to other protein assembly processes.

Structure of the sialic acid-containing O-specific polysaccharide from Salmonella enterica serovar Toucra O48 lipopolysaccharide.

Lipopolysaccharide was extracted from cells of Salmonella enterica serovar Toucra O48 and, after mild acid hydrolysis (1% AcOH, 1 h, 100 degrees C or 0.1 M NaOH-AcOH, pH 4.5, 5 h, 100 degrees C), the O-specific polysaccharide was isolated and characterized. The core and an oligosaccharide containing a fragment of the repeating unit linked to the core region were also obtained, depending on hydrolysis conditions. On the basis of sugar and methylation analyses and NMR spectroscopy of the hydrolysis products, the biological repeating unit of the O-specific polysaccharide was shown to be the following trisaccharide: -->4)-alpha-Neup5Ac(2-->3)-L-alpha-FucpNAc(1-->3)-D-beta-Glc pNAc(1--> The polysaccharide O-chain was substituted with a single molar equivalent of O-acetyl group, distributed between the Neu5Ac O-9 and O-7 positions, in an approximate ratio of 7 : 3.

[Use of modern microscopic techniques for examining dialysis membrane properties]. / Zastosowanie nowoczesnych technik mikroskopowych w badaniach wlasnosci kapilar dializatorów.

The aim of the study was the microscopic evaluation of internal structure of cuprophane and polysulfone membrane and their surface analysis before and after reprocessing. The investigations were performed using an optical measurement system (Digital Instruments), a scanning electron microscope (SEM) and an atomic force microscope (AFM). We confirmed by SEM that reprocessing completely removed biofilm from both membranes surface. The analysis based on AFM visualized channels in the examined membrane. The diameter of the channels varied from 150 nm for cuprophane to 380 nm for polysulfone. The roughness expressed as root mean square (RMS) was higher for cuprophane than for polysulfone membrane. The physical differences between nanostrucure of the examined membranes might be responsible for lower biocompatibility of cuprophane.