ALBERT

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The 〈em〉Streptomyces〈/em〉 sp. strain AV05 isolated from an organic amendment was found to impact both growth and fumonisin production of 〈em〉Fusarium verticillioides〈/em〉 during in vitro direct confrontation. In order to investigate the interactions between the 〈em〉Streptomyces〈/em〉 sp. strain AV05 and 〈em〉F〈/em〉. 〈em〉verticillioides〈/em〉, a metabolomic approach was used. The study of the endometabolomes of the microorganisms was carried out in two different conditions: the microorganisms were cultivated alone or in confrontation. The aim of this study was to examine the modifications of the endometabolome of 〈em〉F〈/em〉. 〈em〉verticillioides〈/em〉 in confrontation with the 〈em〉Streptomyces〈/em〉 strain. The metabolites involved in these modifications were identified using 2D NMR. Many metabolites were found to be overproduced in confrontation assays with the 〈em〉Streptomyces〈/em〉 strain, notably 16 proteinogenic amino acids, inosine, and uridine. This suggested that fungal metabolic pathways such as protein synthesis have been affected due to interaction. Thus, metabolomic studies, as well as proteomics or transcriptomics, are useful for deciphering the mechanisms of interactions between biological control agents and mycotoxigenic fungi. This comprehension is one of the key elements of the improvement of the selection and use of antagonistic agents.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Mastitis in dairy cows is generally considered to be the most expensive disease for dairy farmers worldwide. The overuse of antibiotics is a major problem in the treatment of bovine mastitis, and bacteriophage therapy is expected to provide an alternative treatment. The primary aim of this study was to evaluate the efficacy of a phage cocktail against mastitis in a mouse model. First, a 〈em〉Staphylococcus aureus〈/em〉 strain was isolated from milk samples taken from mastitis cows from dairy farms in Xinjiang, China, and it was designated as Sau-XJ-21. Next, two phages (designated as vBSM-A1 and vBSP-A2) with strong lytic activity against Sau-XJ-21 were isolated from mixed sewage samples collected from three cattle farms in Xinjiang. Phages vBSM-A1 and vBSP-A2 were identified as members of the Myoviridae and Podoviridae families, respectively. The two phages exhibited a wide range of hosts, especially phage vBSM-A1. To evaluate the effectiveness of the two phages in the treatment against mastitis, female lactating mice were used 10–14 days after giving births. The mice were divided into six groups; one group was kept as healthy control, while the remaining five groups were inoculated with the isolated 〈em〉S. aureus〈/em〉 strain to induce mastitis. Four hours after bacterial inoculation, mice in these groups were injected with 25 μL phosphate buffer saline (negative control), ceftiofur sodium (positive control), or phage, either individually or as a cocktail. The mice were sacrificed 20 h later, and the mammary glands were removed and subjected to further analysis, including the quantitation of colony-forming units (CFU), plaque-forming units (PFU), and gross macroscopic as well as histopathology observation. Mice with induced mastitis exhibited significantly improved mastitic pathology and decreased bacterial counts after they had been given phage treatments, with the phage cocktail being more superior than either phage alone. Furthermore, the cocktail treatment also maintained the highest intramammary phage titer without spreading systemically. The effectiveness of the phage cocktail was comparable to that produced by ceftiofur sodium〈em〉.〈/em〉 According to the data obtained for the mouse model of mastitis, phage therapy could be considered as an innovative alternative to antibiotics for the treatment of bovine mastitis.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Lower respiratory tract infection due to 〈em〉Pseudomonas aeruginosa〈/em〉 has become increasingly challenging, resulting in a worse morbidity and mortality. Airway remodeling is a common phenomenon in this process, to which epithelial-mesenchymal transition (EMT) may contribute as an important promoter. Previous studies showed that epithelium-specific integrin αvβ6–mediated EMT was involved in pulmonary fibrosis via transforming growth factor-β1 (TGF-β1) signaling, but whether integrin αvβ6 plays a role in the 〈em〉P. aeruginosa〈/em〉–associated airway remodeling remains unknown. BEAS-2B cells were incubated with lipopolysaccharide (LPS) from 〈em〉P. aeruginosa〈/em〉 in the presence or the absence of integrin αvβ6–blocking antibodies. Morphologic changes were observed by an inverted microscopy. The EMT markers were detected using Western blotting and immunofluorescence. The activation of TGF-β1-Smad2/3 signaling pathway was assessed. Furthermore, matrix metalloproteinase (MMP)-2 and -9 in the medium were measured using ELISA. 〈em〉P. aeruginosa〈/em〉’s LPS decreased the expression of the epithelial marker E-cadherin and promoted the mesenchymal markers, vimentin and α-smooth muscle actin in BEAS-2B cells. The expression of integrin αvβ6 was significantly increased during EMT process. Blocking integrin αvβ6 could attenuate 〈em〉P. aeruginosa〈/em〉’s LPS-induced EMT markers’ expression via TGF-β1-Smad2/3 signaling pathway. Furthermore, blocking integrin αvβ6 could prevent morphologic changes and oversecretion of MMP-2 and -9. Integrin αvβ6 mediates epithelial-mesenchymal transition in human bronchial epithelial cells induced by lipopolysaccharides of 〈em〉P. aeruginosa〈/em〉 via TGF-β1-Smad2/3 signaling pathway and might be a promising therapeutic target for 〈em〉P. aeruginosa〈/em〉–associated airway remodeling.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Excessive use of antibiotics contributes to the selection of resistant bacteria and intestinal colonization with multiresistant pathogens poses a risk factor for subsequent infections. The present study assessed vancomycin-resistant enterococci (VRE) carriage rates in patients admitted to our tertiary care hospital. Stool samples sent for routine culturing were screened with vancomycin containing solid or broth enrichment media. VRE isolates were identified with matrix-assisted laser desorption/ionization-time of flight mass spectrometry and antibiotic susceptibilities were tested by E-test. Vancomycin resistance genes were detected by polymerase chain reaction. Medical records of carriers were examined for suspected risk factors for colonization. Altogether 3025 stool specimens were analyzed. Solid media identified a VRE carriage rate of 2.2% while broth enrichment detected 5.8%. Seventy percent of the isolates were 〈em〉Enterococcus faecium. VanB〈/em〉 genotype was detected in 38.2%, 〈em〉VanA〈/em〉 in 37.3%, 〈em〉VanC1〈/em〉 in 22.6%, and 〈em〉VanC2〈/em〉 in 1.9%. All VRE were sensitive to linezolid, daptomycin, and tigecycline. Collective risk factors for carriage were diabetes, normal flora absence, 〈em〉Clostridioides difficile〈/em〉 positivity, longer hospital stay, and advanced age. 78.5% of the carriers received antibiotic therapy which was metronidazole in most cases (47.3%)〈em〉.〈/em〉 We recommend regular screening of risk groups such as patients with diabetes, history of recent hospitalization, or former 〈em〉C. difficile〈/em〉 infection as an imperative step for preventing VRE dissemination.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The objectives of this study were (i) to isolate and characterize of cultivable denitrifying bacteria using classic microbiological and molecular methods, (ii) to compare of 16S rRNA and 〈em〉nosZ〈/em〉 genes as molecular markers, (iii) to determine bacterial community structure and diversity in soil samples using single-strand conformation polymorphism (SSCP) analysis. In this study, 49 bacterial isolates were cultivated and phylogenetic analyses grouped them into two phyla: 〈em〉Proteobacteria〈/em〉 (37 species) and 〈em〉Firmicutes〈/em〉 (12 species). Our study showed that the 〈em〉nosZ〈/em〉 functional gen could be used to identify denitrifying bacteria abundance in environment but could not be used to identify pure bacterial cultures. In addition, the bacterial community structure showed significant differences among the various soil types. Phylogenetic analysis of community structure indicated that 51 clones could be divided into 2 phylotypes. Uncultured bacteria (80.4%) and 〈em〉Gammaproteobacteria〈/em〉 (19.6%) were the dominant components of the soil bacterial community. For 16S rRNA, PCR products of 49 bacteria were obtained with 27F-1492R primer pairs. For 〈em〉nosZ〈/em〉, PCR products were obtained with primers 1F-1R (259 bp), 2F-2R (267 bp), and F-1622R (453 bp) of 39 bacteria that the single 〈em〉nosZ〈/em〉 band provided on the agarose gel. The bacterial 16S rRNA gene clone library was dominated by 〈em〉Gammaproteobacteria〈/em〉 and 〈em〉Bacilli〈/em〉. The 〈em〉nosZ〈/em〉 clone sequences did not represent the bacteria from which they were obtained but were found to be closer to the environmental clones. Our study showed that the 〈em〉nosZ〈/em〉 functional gene could be used to identify denitrification abundance in environment but could not be used to identify pure bacterial cultures. It was also found that the 〈em〉nosZ〈/em〉 sequences showed uncultured denitrifier species.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Vancomycin is often used in orthopedic surgery as a local prophylaxis of bacterial infection. The aim of this work was to compare the release of vancomycin and its biologically inactive crystalline degradation products (CDP-1) during in vitro experiments from different types of local antibiotic delivery systems (bone grafts and bone cements). The concentrations of vancomycin and its crystalline degradation products were determined by high-performance liquid chromatography. Each experiment was performed in a phosphate buffer solution over 21 days. Morselized bone grafts, synthetic bone cements Palacos and Copal, and synthetic bone grafts were tested as local carriers of vancomycin. The highest concentration approximately 670 mg/L of vancomycin was released from synthetic bone grafts Actifuse. Even after 21 days, the concentration of vancomycin was still above the minimum inhibitory concentration (MIC). The maximum concentration of vancomycin released in two experiments with human bone grafts exceeded 600 mg/L during the first day and was still above MIC level 21 days later when the experiment was concluded. By comparing the synthetic bone cements Palacos and Copal, Copal had the average maximum concentration of only 32.4 mg/L and Palacos 35.7 mg/L. The concentration of vancomycin fell below the MIC for vancomycin-resistant 〈em〉Staphylococcus aureus〈/em〉 (VRSA) on the seventh day with Palacos and the ninth day with Copal. This study showed the insufficient concentration of released vancomycin from synthetic bone cements at the end of the experiment. For improvement of local prophylaxis, it would be beneficial to increase the amount of vancomycin in bone cements.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The present study was designed to characterize six 〈em〉Trueperella〈/em〉 (〈em〉T.〈/em〉) 〈em〉abortisuis〈/em〉 strains, cultured over a period of 5 months from fetus and abortion material of six pigs of a single farm in Mecklenburg-West Pomerania federal state, Germany. It was of interest to investigate the epidemiological relationships of the six strains among each other and whether a single bacterial clone was responsible for the abortion situation of the single farm. All six strains were identified phenotypically, by MALDI-TOF MS analysis and by phylogenetic analysis based on 16S rRNA gene and 〈em〉gap〈/em〉 (encoding the glyceraldehyde 3-phosphate dehydrogenase) and 〈em〉tuf〈/em〉 (encoding elongation factor tu) gene sequencing. Further genotypic comparison was performed using different genomic DNA fingerprint methods including BOX-PCR, (GTG)〈sub〉5〈/sub〉-PCR, and three RAPD-PCRs. The sequence analysis of the genes 〈em〉gap〈/em〉 and 〈em〉tuf〈/em〉 and the genomic DNA fingerprinting results revealed, as noval findings, that the six 〈em〉T. abortisuis〈/em〉 strains cultured from a single farm represent six different bacterial clones showing a genetic variability of this bacterial species in the pig population. All six 〈em〉T. abortisuis〈/em〉 strains were isolated in mixed culture with several other bacterial species. However, the 〈em〉T. abortisuis〈/em〉 strain, generally found in high numbers, seemed to be responsible for the abortion situation in the farm.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Getting into a weakened organism, non-tuberculosis mycobacteria (NTMB) contact not only with the cells of the microorganism but also with the microflora of the human body; however, these interactions are poorly understood. The purpose of this work was to study the effect of NTMB supernatants on the properties of conditionally pathogenic microorganisms in their interaction with red blood cells. We used strains of non-tuberculous mycobacteria 〈em〉Mycobacterium iranicum〈/em〉 and 〈em〉M. rutilum〈/em〉, as well as strains of 〈em〉Staphylococcus epidermidis〈/em〉 and 〈em〉Escherichia coli〈/em〉. Using the fluorescence in situ hybridisation (FISH) method, the processes of adhesion to the surface of erythrocytes and the intra-erythrocyte penetration of cells of 〈em〉S. epidermidis〈/em〉 and 〈em〉E. coli〈/em〉 under the influence of NTMB supernatants were studied. To study changes in the haemoglobin molecule under the action of the supernatants of NTMB, spectral analysis was performed. Statistical processing was performed using STATISTIKA 6.0. The results showed that the supernatants of 〈em〉M. iranicum〈/em〉 and 〈em〉M. rutilum〈/em〉 increased the adhesion of conditionally pathogenic bacteria with a low level of AntiHbA to the surface of erythrocytes by 3–4 times. It also increased the intra-erythrocyte penetration of cells of 〈em〉S. epidermidis〈/em〉 and 〈em〉E. coli〈/em〉 relative to the control values. As a result of studying the haemoglobin spectrum of erythrocytes under the influence of 〈em〉M. iranicum〈/em〉, a decrease in the optical density values of oxyhaemoglobin by a factor of 2 relative to the values in the control sample was noted. Thus, the supernatants of NTMB have a multidirectional effect on the interaction of opportunistic microorganisms with erythrocytes, increasing the adhesive activity and the penetration of cells into the erythrocytes, as well as reducing the optical density of oxyhaemoglobin.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim of the present study was to develop an ion-selective electrode method for the continuous determination of the intracellular pH in 〈em〉Lactobacillus plantarum〈/em〉 using a small-scale bioreactor. This method employed a salicylate-selective electrode basing on the distribution of salicylic acid across the cytoplasmic membrane. This developed electrode responded to salicylate concentrations above 20 μmol/L with a Nernstian sensitivity. The energized and concentrated cells were added into a thermostated small-scale bioreactor that contained the salicylate anions dissolved in a 100 mmol/L potassium phosphate buffer at different pH values. The changes in salicylate concentration that occurred in the medium containing bacterial suspension were measured as a voltage change. The cells of 〈em〉Lactobacillus plantarum〈/em〉 showed maintenance of pH homeostasis at the studied pH ranging from 4.0 to 7.0, and they kept a neutral intracellular pH up to 5.8. The simplicity of the measuring preparation and the relatively low cellular concentration, as well as the advantages of the small-scale bioreactor, lead us to believe that the described method can facilitate the study of the physicochemical factors on the intracellular pH of lactic acid bacteria using a single pH probe in one method.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Microbial contamination poses a great threat to aviation system security through mechanisms such as microbiologically influenced corrosion (MIC), fuel filter clogging, and fuel deterioration. In this study, a survey of microbial contamination in aviation fuel obtained from aircraft fuel tanks was performed to test the relationship between microbial contamination and aircraft service life. The contaminating microorganisms were counted, isolated, identified, and subjected to preliminary characterization. A low risk of microbial contamination in the selected samples was confirmed, and there was no significant difference in the counts between culturable bacteria and fungi (〈em〉p〈/em〉 〉 0.05). Phylogenetic analysis tree indicated that the diversity of culturable microorganisms was rather low, with 17 bacterial isolates belonging to 13 genera and 12 fungal isolates belonging to 5 genera. No yeast was isolated. The growth characteristics of these isolates indicated that the aircraft fuel tanks harbored various microorganisms that were able to utilize the aviation fuel as a source of carbon and energy. Meanwhile, some isolates caused emulsification and produced acid. The conclusions of this study were that various hazardous microorganisms can root in aircraft aviation fuel tanks. There was no relationship between microbial contamination and aircraft service life (〈em〉p〈/em〉 〉 0.05), and continuous good maintenance suppressed microbial proliferation.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The antifouling, antimicrobial, elution behavior, skin irritant, and cytotoxicity properties of water-soluble phosphate glass on stainless steel were evaluated. Water-soluble phosphate glass samples with 35% Cu (mol/mol) were prepared by altering the network modifier (Na〈sub〉2〈/sub〉O, K〈sub〉2〈/sub〉O) and network former (P〈sub〉2〈/sub〉O〈sub〉5〈/sub〉, B〈sub〉2〈/sub〉O〈sub〉3〈/sub〉) compositions. The materials were melted at temperatures within the range of 850–950 °C. The melt was then quenched and ground into fine particles using a twin roll mill. The resulting water-soluble glasses were prepared as glass frit (size 〈 100 μm) using a sieve. The amorphous phase was determined by X-ray diffraction and differential thermal analysis. Water-soluble glasses with a reduced Cu ion elution rate of 1.2 ppm per week were formed because the chemical resistances of the formulated glasses improved as the P〈sub〉2〈/sub〉O〈sub〉5〈/sub〉 content decreased and the B〈sub〉2〈/sub〉O〈sub〉3〈/sub〉 content increased. To test its antifouling properties, the glass frit was mixed with paint and coated onto a STS316L sheet. The surface roughness was increased markedly from 1.4 to 19.2 nm, increasing the specific surface area for antimicrobial activity. It was demonstrated that the proposed method was able to form noncytotoxic, nonirritant, water-soluble glasses with 99.9% antimicrobial activity against 〈em〉Staphylococcus aureus.〈/em〉 These results suggest that water-soluble phosphate glass on STS316L sheets could be useful in filtration plants.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Nosocomial infections are an important cause of morbi-mortality worldwide. The increase in the rate of resistance to conventional drugs in these microorganisms has stimulated the search for new therapeutic options. The nitro moiety (NO〈sub〉2〈/sub〉) is an important pharmacophore of molecules with high anti-infective activity. We aimed to synthesize new nitro-derivates and to evaluate their antibacterial and anti-〈em〉Candida〈/em〉 potential in vitro〈em〉.〈/em〉 Five compounds [3-nitro-2-phenylchroman-4-ol (〈strong〉3〈/strong〉); 3-nitro-2-phenyl-2H-chromene (〈strong〉4a〈/strong〉); 3-nitro-2-(4-chlorophenyl)-2H-chromene (〈strong〉4b〈/strong〉); 3-nitro-2-(4-fluorophenyl)-2H-chromene (〈strong〉4c〈/strong〉), and 3-Nitro-2-(2,3-dichlorophenyl)-2H-chromene (〈strong〉4d〈/strong〉)] were efficiently synthesized by Michael-aldol reaction of 2-hydroxybenzaldehyde with nitrostyrene, resulting in one β-nitro-alcohol (〈strong〉3〈/strong〉) and four nitro-olefins (〈strong〉4a〈/strong〉–〈strong〉4d〈/strong〉). The antibacterial and anti-〈em〉Candida〈/em〉 potentials were evaluated by assaying minimal inhibitory concentration (MIC), minimum fungicidal concentration (MFC), and minimum bactericidal concentration (MBC). Mono-halogenated nitro-compounds (〈strong〉4b〈/strong〉 and 〈strong〉4c〈/strong〉) showed anti-staphylococcal activity with MIC values of 15.6–62.5 μg/mL and MBC of 62.5 μg/mL. However, the activity against Gram-negative strains was showed to be considerably lower and our data suggests that this effect was associated with the outer membrane. Furthermore, nitro-compounds 〈strong〉4c〈/strong〉 and 〈strong〉4d〈/strong〉 presented activity against C〈em〉andida〈/em〉 spp. with MIC values ranging from 7.8–31.25 μg/mL and MFC of 15.6–500 μg/mL. In addition, these compounds were able to induce damage in fungal cells increasing the release of intracellular material, which was associated with actions on the cell wall independent of quantitative changes in chitin and β-glucan. Together, these findings show that nitro-compounds can be exploited as anti-staphylococcal and anti-〈em〉Candida〈/em〉 prototypes.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Flap endonuclease is a structure-specific nuclease which cleaves 5′-flap of bifurcated DNA substrates. Genome sequence of 〈em〉Thermococcus kodakarensis〈/em〉 harbors an open reading frame, Tk1281, exhibiting high homology with archaeal flap endonucleases 1. The corresponding gene was cloned and expressed in 〈em〉Escherichia coli〈/em〉, and the gene product was purified to apparent homogeneity. Tk1281 was a monomer of 38 kDa and catalyzed the cleavage of 5′-flap from double-stranded DNA substrate containing single-stranded DNA flap. The highest cleavage activity was observed at 80 °C and pH 7.5. Under optimal conditions, Tk1281 exhibited apparent 〈em〉V〈/em〉〈sub〉max〈/sub〉 and 〈em〉K〈/em〉〈sub〉m〈/sub〉 values of 278 nmol/min/mg and 37 μM, respectively, against a 54-nucleotide double-stranded substrate containing a single-stranded 5′-flap of 27 nucleotides. A unique feature of Tk1281 is its highest activation in the presence of Co〈sup〉2+〈/sup〉 and no activation with Mn〈sup〉2+〈/sup〉. To the best of our knowledge, this is the first cloning and characterization of a flap endonuclease from the genus 〈em〉Thermococcus〈/em〉.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Chromera velia〈/em〉 is a marine photosynthetic relative of human apicomplexan parasites. It has been isolated from coral reefs and is indicted for being involved in symbioses with hermatypic corals. 〈em〉C. velia〈/em〉 has been subject to intensive research, but still very little is known of its response to light quality and quantity. Here, we have studied the growth and compositional responses of 〈em〉C. velia〈/em〉 to culture under monochromatic light (blue, green or red), at two photon flux densities (PFD, 20 and 100 μmol photons m〈sup〉−2〈/sup〉 s〈sup〉−1〈/sup〉). Our results show that 〈em〉C. velia〈/em〉 growth rate is unaffected by the quality of light, whereas it responds to PFD. However, light quality influenced cell size, which was smaller for cells exposed to blue monochromatic light, regardless of PFD. PFD strongly influenced carbon allocation: at 20 μmol photons m〈sup〉−2〈/sup〉 s〈sup〉−1〈/sup〉, carbon was mainly allocated into proteins while at 100 μmol photons m〈sup〉−2〈/sup〉 s〈sup〉−1〈/sup〉, carbon was allocated mainly into carbohydrate and lipid pools. The blue light treatment caused a decrease in the lipids and carbohydrates to proteins and thus suggested to affect nitrogen metabolism in acclimated cells. Whole-cell absorption spectra revealed the existence of red-shifted chlorophyll 〈em〉a〈/em〉 antenna not only under red light but in all low PFD treatments. These findings show the ability of 〈em〉C. velia〈/em〉 to successfully adapt and thrive in spectrally very different environments of coral reefs.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The following series of articles form a special issue organized by the Algatech Center of the Institute of Microbiology CAS dedicated to the memory of Dr. Ivan Šetlík.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The rapid emergence of resistance in pathogenic bacteria together with a steep decline in economic incentives has rendered a new wave in the drug development by the pharmaceutical industry and researchers. Since cyanobacteria are recognized as wide producers of pharmaceutically important compounds, we investigated thirty-four cyanobacterial extracts prepared by solvents of different polarities for their antimicrobial potential. Almost all tested cyanobacterial strains exhibited some degree of antimicrobial bioactivity, with more general effect on fungal strains compared with bacteria. Surprisingly ~50% of cyanobacterial extracts exhibited specific activity against one or few bacterial indicator strains with Gram-positive bacteria being more affected. Extracts of two most promising strains were subjected to activity-guided fractionation and determination of the minimum inhibitory concentration (MIC) against selected bacterial and fungal isolates. Multiple fractions were responsible for their antimicrobial effect with MIC reaching low-micromolar concentrations and in some of them high level of specificity was recorded. Twenty-six bioactive fractions analyzed on LC-HRMS/MS and Global Natural Product Social Molecular Networking (GNPS) online workflow using dereplication resulted in identification of only forty-nine peptide spectrum matches (PSMs) with eleven unique metabolites spectrum matches (MSMs). Interestingly, only three fractions from 〈em〉Nostoc calcicola〈/em〉 Lukešová 3/97 and four fractions from 〈em〉Desmonostoc〈/em〉 sp. Cc2 showed the presence of unique MSMs suggesting the presence of unknown antimicrobial metabolites among majority of bioactive fractions from both the strains. Our results highlight potential for isolation and discovery of potential antimicrobial bioactive lead molecules from cyanobacterial extracts.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Citrus black spot (CBS) and post-bloom fruit drop (PFD), caused by 〈em〉Phyllosticta citricarpa〈/em〉 and 〈em〉Colletotrichum abscissum〈/em〉, respectively, are two important citrus diseases worldwide. CBS depreciates the market value and prevents exportation of citrus fruits to Europe. PFD under favorable climatic conditions can cause the abscission of flowers, thereby reducing citrus production by 80%. An ecofriendly alternative to control plant diseases is the use of endophytic microorganisms, or secondary metabolites produced by them. Strain LGMF1631, close related to 〈em〉Diaporthe〈/em〉 cf. 〈em〉heveae〈/em〉 1, was isolated from the medicinal plant 〈em〉Stryphnodendron adstringens〈/em〉 and showed significant antimicrobial activity〈em〉,〈/em〉 in a previous study. In view of the potential presented by strain LGMF1631, and the absence of chemical data for secondary metabolites produced by 〈em〉D.〈/em〉 cf. 〈em〉heveae〈/em〉, we decided to characterize the compounds produced by strain LGMF1631. Based on ITS, TEF1, and TUB phylogenetic analysis, strain LGMF1631 was confirmed to belong to 〈em〉D.〈/em〉 cf. 〈em〉heveae〈/em〉 1. Chemical assessment of the fungal strain LGMF1631 revealed one new 〈em〉seco〈/em〉-dihydroisocoumarin [cladosporin B (〈strong〉1〈/strong〉)] along with six other related, already known dihydroisocoumarin derivatives and one monoterpene [(−)-(1〈em〉S〈/em〉,2〈em〉R〈/em〉,3〈em〉S〈/em〉,4〈em〉R〈/em〉)-〈em〉p〈/em〉-menthane-1,2,3-triol (〈strong〉8〈/strong〉)]. Among the isolated metabolites, compound 〈strong〉5〈/strong〉 drastically reduced the growth of both phytopathogens in vitro and completely inhibited the development of CBS and PFD in citrus fruits and flowers. In addition, compound 〈strong〉5〈/strong〉 did not show toxicity against human cancer cell lines or citrus leaves, at concentrations higher than used for the inhibition of the phytopathogens, suggesting the potential use of (−)-(3〈em〉R〈/em〉,4〈em〉R〈/em〉)-〈em〉cis〈/em〉-4-hydroxy-5-methylmellein (〈strong〉5〈/strong〉) to control citrus diseases.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The biogenesis of the cyanobacterial photosystem II (PSII) complex requires a number of auxiliary assembly factors that improve efficiency of the process but their precise function is not well understood. To assess a possible synergic action of the Ycf48 and Ycf39 factors acting in early steps of the biogenesis via interaction with the nascent D1 subunit of PSII, we constructed and characterised a double mutant of the cyanobacterium 〈em〉Synechocystis〈/em〉 PCC 6803 lacking both these proteins. In addition, we also deleted the 〈em〉ycf39〈/em〉 gene in the double mutant lacking Ycf48 and Pam68, the latter being a ribosomal factor promoting insertion of chlorophyll (Chl) into the CP47 subunit of PSII. The resulting double ΔYcf48/ΔYcf39 and triple ΔYcf48/ΔPam68/ΔYcf39 mutants were deficient in PSII and total Chl, and in contrast to the source mutants, they lost the capacity for autotrophy. Interestingly, autotrophic growth was restored in both of the new multiple mutants by enhancing Chl biosynthesis using a specific ferrochelatase inhibitor. Taking together with the weak radioactive labelling of the D1 protein, these findings can be explained by inhibition of the D1 synthesis caused by the lack and/or incorrect binding of Chl molecules. The results emphasise the key importance of the sufficient Chl supply for the PSII biogenesis and also support the existence of a so far enigmatic regulatory mechanism leading to the reduced overall Chl biosynthesis/accumulation when the PSII assembly is impaired.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉When it comes to women’s health, treating vaginal infections makes up a high proportion of the gynecological services. Among the forms of vaginitis, vulvovaginal candidiasis (VVC) is considered the second most common. Demand for new treatment alternatives is increasingly relevant, especially for therapies with fewer side effects, better tolerability, and lower cost, while still offering improved quality of life in terms of disease prevention. This study intended to investigate the alternative therapies described for the adjuvant treatment of vulvovaginitis caused by 〈em〉Candida〈/em〉 species, including alternative and complementary treatment methods used by women. This literature review is based on articles written in English and Portuguese in the PubMed, Google Scholar, and SciELO databases. This study was conducted for the most part using the Brazilian Government’s Capes Periodicals Portal, which directs to Google Scholar and PubMed. Since the 1980s, there has been growing interest in alternative therapies in Brazil, a trend which also began in other Western countries in the second half of the twentieth century. Some alternative treatments include substances with antifungal activity, some substances help restore the balance of the vaginal microbiota, while others have an inhibitory activity on microbial virulence factors. The proper use of therapeutic alternatives can effectively contribute to the treatment of VVC, but it should be remembered that some chemical products, such as boric acid or vinegar, and even natural products such as propolis, garlic, and tea tree may have undesirable side effects, having not been tested by well-designed clinical studies. Even so, alternative therapies in the treatment of VVC do have support in the scientific literature.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Asymptomatic uropathogenic 〈em〉Escherichia coli〈/em〉 (UPECs) are the leading cause of asymptomatic bacteriuria (ABU) in humans. So this study aimed to identify and characterize ABU UPECs from hospitalized patients of Kolkata, India, with respect to their antibiogram profile, phylogeny, pathogenicity islands, and virulence factor gene acquisition and FimH mutations in comparison to symptomatic UPECs. 〈em〉E. coli〈/em〉 was detected biochemically in 44.44% (20/45) and 32.26% (20/62) of urine culture-positive asymptomatic and symptomatic hospitalized individuals respectively. Ninety-five percent of the asymptomatic isolates were multidrug resistant (MDR) compared to the symptomatic isolates (100%). Significant predominance of unknown phylogroup, pathogenicity island markers (PAI IV536, PAI I CFT073), and distribution patterns of different virulence factor genes respectively was evident among both groups. A significant correlation was observed between both groups of isolates with respect to their antibiotic resistances (except imipenem, amikacin, and nitrofurantoin), prevalence of phylogenetic groups and PAIs, and virulence factor gene (〈em〉fimH〈/em〉, 〈em〉papC〈/em〉, 〈em〉papEF〈/em〉, 〈em〉papGII〈/em〉, 〈em〉iucD〈/em〉, and 〈em〉cnf1〈/em〉) acquisition. Pathoadaptive FimH adhesin mutations, especially hot spot mutation V27A, were detected in 80% asymptomatic isolates mostly reported in symptomatic ones worldwide. Moreover, this is the first study from India that reported incidence of “Unknown” phylogroup, pathogenicity island markers, and potentially pathoadaptive FimH mutations in asymptomatic UPECs isolated from hospitalized patients which further indicated that these ABU 〈em〉E. coli〈/em〉 might have originated from their symptomatic counterparts due to unbridled use of unprescribed antibiotics. Therefore, this study demands antibiotic de-escalation along with regular and intricate monitoring at the molecular level for efficient management of ABU that addresses a major public health concern.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Mutations occurring in viral polymerase gene of hepatitis B virus (HBV) due to the use of nucleos(t)id analogs reduce the activity of the drugs by causing antiviral resistance. In this study, it was aimed to evaluate mutations responsible for drug resistance and drug resistance mutation rates in patients followed up by the diagnosis of chronic hepatitis B (CHB). A total of 318 CHB patients were included in the study. HBV mutations were detected using the INNO-LiPA commercial kit based on the reverse hybridization principle. Drug resistance mutation was detected in 46.86% (149/318) of the patients. The rates of drug resistance were found 36.79% (117/318) for lamivudine resistance, 12.58% (40/318) for entecavir (ETV), and 7.86% (25/318) for adefovir. In 10 patients, the possible tenofovir (TDF) resistance (3.14%) was found. Single-drug and double-drug resistances were detected in 34.59% and in 11.01% of the patients, respectively. Triple drug resistance was detected in only 1.26% of the patients. Unlike various studies in Turkey and in other countries, remarkable resistance to ETV and TDF were found in this study. The high rate of the probable TDF resistance was striking, with 3.14%.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Some strains of the genus 〈em〉Enterococcus〈/em〉 are effective probiotic bacteria if they meet safety and probiotic criteria. In our study, 17 canine enterococci previously selected from a group of 160 isolates based on safety criteria were screened for some functional properties relevant to their use as probiotics. The results of antimicrobial resistance testing showed sensitivity of eleven strains to EFSA recommended antimicrobials. In contrast, the most frequent resistance was observed for cefotaxim (15/17) and oxacillin (13/17). PCR detection of resistance genes (〈em〉vanA〈/em〉, 〈em〉vanB〈/em〉, 〈em〉vanC〈/em〉, 〈em〉tetM〈/em〉, 〈em〉tetL〈/em〉, 〈em〉ermB〈/em〉, and 〈em〉mefA〈/em〉) revealed the presence of 〈em〉mefA〈/em〉 gene in five 〈em〉Enterococcus faecium〈/em〉 strains and 〈em〉vanA〈/em〉 gene in one strain. The production of enzymes commonly associated with intestinal diseases was in general rare (β-glucosidase 2/17, α-chymotrypsin 1/17, 〈em〉N〈/em〉-acetyl-β-glucosaminidase 0/17, and β-glucuronidase 0/17). The measurement of strain survival rate (%) under the conditions simulating gastric (pH 2.5) and bile juices (0.3% bile) showed considerable differences between strains (〈 0.01 to 4.7% after 90 min for gastric juices, 48.0 to 254.0% after 180 min for bile). The concentration of produced 〈span〉l〈/span〉-lactic acid ranged between 83.1 to 119.3 mmol/L after 48 h cultivation depending on the strain. All strains fermented 16 out of 49 different carbohydrates (range from 17 to 23/49). Antimicrobial activity was recorded for two strains against some species of 〈em〉Listeria〈/em〉 sp. and 〈em〉Enterococcus〈/em〉 sp. Finally, two 〈em〉E. faecium〈/em〉 candidates (IK25 and D7) were selected for testing in dogs, and hereafter they could possibly extend the currently limited range of beneficial bacteria of canine origin used as a dietary supplement for dogs.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Paclobutrazol, (2RS, 3RS)-1-(4-chlorophenyl)-4, 4-dimethyl-2-(1H-1,2,4-triazol-1-yl) pentan-3-ol, is a plant growth retardant that mainly inhibits gibberellins (GAs) biosynthesis. In agricultural practice, paclobutrazol is applied to arrest vegetative growth so as to increase the reproductive growth of many orchard fruit, as well as grain crops. However, due to its over-application and chemical stability, paclobutrazol accumulates in soil and inhibits the growth of subsequent crops, especially those grown for vegetative purposes. The present study focused mainly on the changes in the soil bacterial community following application of paclobutrazol. Mung bean (〈em〉Vigna radiata〈/em〉) plants were treated with paclobutrazol and cultivated for three consecutive seasons. Soil samples were collected and analyzed by denaturing gradient gel electrophoresis (DGGE) using 16S rDNA gene fragments and clone library analyses. The results obtained through clustering and clonal sequencing analysis showed that the bacterial community was affected by paclobutrazol, and in addition, was more diverse in the third stage of mung bean plant cultivation. The results of the study showed that paclobutrazol affected bacterial composition, and the population of bacteria varied greatly across time.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim of this study was to evaluate the contribution of plasmid-mediated genes and efflux to fluoroquinolone resistance in collection of 〈em〉Achromobacter〈/em〉 spp. gathered during a 3-year period. Susceptibility to ciprofloxacin and levofloxacin was tested by disk diffusion and microdilution tests for a collection of 98 〈em〉Achromobacter〈/em〉 spp. clinical isolates. Identification of fluoroquinolone-resistant isolates was performed by sequencing and phylogenetic analyses of the 〈em〉nrdA〈/em〉 gene. Genetic relatedness among resistant isolates was determined by pulsed-field gel electrophoresis (PFGE) analysis. The influence of an H〈sup〉+〈/sup〉 conductor cyanide 〈em〉m〈/em〉-chlorophenyl hydrazone (CCCP) and a resistance-nodulation-division-type efflux pump inhibitor phenylalanine-arginine beta-naphthylamide (PAβN) on minimal inhibitory concentration (MIC) value was evaluated by broth microdilution. The presence of the plasmid-mediated 〈em〉qnrA〈/em〉, 〈em〉qnrB〈/em〉, 〈em〉qnrC〈/em〉, 〈em〉qnrS〈/em〉, and 〈em〉aac-(6′)-Ib-cr〈/em〉 genes was investigated by PCR and sequencing. 〈em〉Achromobacter〈/em〉 spp. isolates that were resistant or intermediately resistant to fluoroquinolones in disk diffusion tests (44/98) were subjected to microdilution. As a result, 20/98 isolates were confirmed to be resistant to ciprofloxacin while 10/98 was resistant to levofloxacin. CCCP decreased twofold MIC value for ciprofloxacin in six isolates and more than 16 times in one isolate, while MIC value for levofloxacin was decreased in all isolates (twofold to more than eightfold). Fluoroquinolone-resistant isolates were identified as 〈em〉A. xylosoxidans〈/em〉 with the 〈em〉nrdA〈/em〉 gene sequencing. PFGE revealed that resistant isolates belonged to seven different genotypes. Ten isolates belonging to four genotypes were positive for the 〈em〉aac-(6′)-Ib-cr〈/em〉 gene. Although resistance to fluoroquinolones was not widespread among analyzed isolates, detected contribution of efflux pumps and the presence of the 〈em〉aac-(6′)-Ib-cr〈/em〉 gene present a platform for emergence of more resistant strains.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim of this study was to determine the presence and prevalence of 〈em〉Wolbachia〈/em〉 bacteria in natural population of fleas (Insecta: Siphonaptera) in Turkey, and to exhibit the molecular characterization and the phylogenetic reconstruction at the positive isolates with other species in GenBank, based on 16S rDNA sequences. One hundred twenty-four flea samples belonging to the species 〈em〉Ctenocephalides canis〈/em〉, 〈em〉C. felis〈/em〉, and 〈em〉Pulex irritans〈/em〉 were collected from animal shelters in Kayseri between January and August 2017. All flea species were individually screened for the presence of 〈em〉Wolbachia〈/em〉 spp. by polymerase chain reaction (PCR) targeting the 16S ribosomal RNA gene. According to PCR analyses, 〈em〉Wolbachia〈/em〉 spp. were found prevalent in 〈em〉C. canis〈/em〉 and 〈em〉P. irritans〈/em〉 fleas, while it was not detected in the 〈em〉C. felis〈/em〉 species. Totally, 20 isolates were purified from agarose gel and sequenced with the same primers for molecular characterization and phylogenetic analyses. The sequence analyses revealed 17 polymorphic sites and 2 genetically different 〈em〉Wolbachia〈/em〉 isolates, representing two different haplotypes in two flea species. The distribution patterns, molecular characterization, and phylogenetic status of 〈em〉Wolbachia〈/em〉 spp. of fleas in Turkey are presented for the first time with this study. Understanding of the role of 〈em〉Wolbachia〈/em〉 in vector biology may provide information for developing 〈em〉Wolbachia〈/em〉-based biological control tools.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Continuous increasing incidence of allergic diseases is calling for identifying early prognostic markers pointing to increased risk of allergy development and establishing protocols for preventive strategies limiting allergy development in predisposed individuals. It is important to better understand the critical events occurring in early postnatal life, especially the interaction of a newborn with microbial compounds important for the maturation of the neonatal immune system and setting immunoregulatory responses as well. Dendritic cells (DC) together with the cytokine microenvironment play an important role in priming of immune responses. The capacity of monocyte-derived DC (moDC) from cord blood of children of healthy and allergic mothers to respond to microbial antigens (〈em〉Escherichia coli〈/em〉 O86 (EcO86) and delipidated 〈em〉Bacillus firmus〈/em〉 (DBF)) was tested by flow cytometry and quantitative real-time PCR. Both EcO86 and DBF were able to promote maturation of moDC, but moDC of children of allergic mothers expressed higher levels of activation markers CD80 and CD83. Increased gene expression of IL-6 and lower expression of indol-amine 2,3 dioxygenase were observed in moDC of neonates of allergic mothers, in comparison to healthy ones. A higher gene expression and an increased presence of activation markers on moDC of newborns of allergic mothers indicate a generally higher reactivity of these cells, possibly enabling easier development of inappropriate immune response after an allergen encounter.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Neoscytalidium〈/em〉 (or 〈em〉N.〈/em〉) 〈em〉dimidiatum〈/em〉 and 〈em〉N. novaehollandiae〈/em〉 are two aggressive plant pathogenic species that affect several agricultural crops. Early detection and identification of these fungi are of critical importance to bring about the effective minimization to the threat they pose to the infected plants. Herein, two species of 〈em〉Neoscytalidium〈/em〉 were rapidly discriminated by utilizing the rRNA internal transcribed (ITS4-5.8S-ITS5) PCR primers. A total of 100 isolates of 〈em〉Neoscytalidium〈/em〉 species, which were isolated from Iraqi canker-infected fig trees, were included in this study. Two discrete electrophoretic PCR bands were observed in 〈em〉Neoscytalidium〈/em〉 isolates—A-variants were about 546 bp, while B-variants were about 993 bp in length. The comprehensive phylogenetic analysis of both DNA variants revealed that A-variants resided between 〈em〉N. novaehollandiae〈/em〉 and 〈em〉N. hyalinum〈/em〉, while B-variants were closely related to 〈em〉N. dimidiatum〈/em〉. Furthermore, the highly specific re-constructed tree of both electrophoretic variants demonstrated that B-variants share a high similarity with 〈em〉N. novaehollandiae〈/em〉. Additionally, the secondary structures for both variants were predicted computationally to reveal the structural patterns that each variant follows. In conclusion, a small rRNA locus comprising 22 nucleotides that differs in the two variants is potentially responsible for this species-specific classification. The main divergence in the amplified loci led to the classification of these fungal variants into two main species, namely 〈em〉N. dimidiatum〈/em〉 and 〈em〉N. novaehollandiae〈/em〉, demonstrating that the amplification by ITS4–ITS5 rRNA fragment is a beneficial strategy that can be employed for the assessment of 〈em〉Neoscytalidium〈/em〉 diversity in the natural ecosystems.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Turkeys and broilers have been identified as important reservoirs for 〈em〉Campylobacter jejuni〈/em〉 which is of public health significance. The evaluation of the genotypes among 〈em〉C〈/em〉. 〈em〉jejuni〈/em〉 strains within different reservoirs is critical for our understanding of the epidemiology of this infectious agent. The present study aimed to compare the genetic diversity and differences of 〈em〉C〈/em〉. 〈em〉jejuni〈/em〉 isolates from turkeys and broilers using flagellin PCR-RFLP typing (〈em〉flaA〈/em〉 typing) technique, in terms of the ease of use and discriminatory power. Sixty 〈em〉C〈/em〉. 〈em〉jejuni〈/em〉 isolates were detected biochemically and confirmed by duplex-PCR from turkeys and broilers (30 strains from each bird species). Then, a 〈em〉flaA〈/em〉 gene fragment (1725 bp) of 〈em〉C〈/em〉. 〈em〉jejuni〈/em〉 isolates was amplified and amplicons were digested with HpyF3I enzyme. Restriction analysis by HpyF3I gave four different 〈em〉flaA〈/em〉 patterns (H1, H2, H3, H4) among all tested 〈em〉C〈/em〉. 〈em〉jejuni〈/em〉 isolates. In broiler isolates, all four patterns were observed but in turkey isolates, only H2 and H4 patterns were present. The results clearly demonstrated that distribution of the 〈em〉flaA〈/em〉 typing patterns differed depending on the host species (broiler/turkey). H1 and H3 〈em〉flaA〈/em〉 types are more prevalent in broiler than turkey isolates, while H2 type is significantly more prevalent within isolates from turkey (〈em〉p〈/em〉 〈 0.05). The 〈em〉flaA〈/em〉 typing technique by digestion with HpyF3I enzyme can almost give us a clue to the source of infection in local outbreaks.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Mixed infections and heteroresistance of 〈em〉Helicobacter pylori〈/em〉 contribute to decreased efficacy of treatments. This study aimed to investigate frequency of clarithromycin heteroresistance and its link with mixed infections, medication history, and disease severity. A total of 40 pairs of 〈em〉H〈/em〉. 〈em〉pylori〈/em〉 strains were isolated from the antrum and corpus of 97 patients. Susceptibility of the strains to clarithromycin was measured by agar dilution method. Site-specific mutations of 〈em〉23S rRNA〈/em〉 at A2143G, A2142G, and A2142C positions were analyzed by PCR and genomic relatedness of pairs of the strains was determined by random amplified polymorphic DNA (RAPD)-PCR. The results showed a prevalence of 35% (14/40) clarithromycin resistance. Diversity of the antrum and corpus isolates in resistance to clarithromycin was detected among 17.5% (7/40) of the patients. Similarly, diversity in MIC value was also detected in two patients infected with the sensitive strains. Significant difference in frequency of resistance was detected among patients with peptic ulcer disease (PUD) (MIC90 32 μg/mL) and severe gastritis (MIC90 16 μg/mL), compared with those who suffered from non-ulcer dyspepsia (NUD) (MIC90 8 μg/mL) and chronic gastritis (MIC90 0.25 μg/mL). MIC values showed 8–32 folds increased levels in the corpus. A2142G, A2143G, and A2142C mutations were detected in three, two, and two patients, respectively, but not observed in 46% of the resistant strains. RAPD-PCR fingerprints showed identical molecular patterns for the isolates of the corpus and antrum in each patient. In conclusion, microevolution of 〈em〉H〈/em〉. 〈em〉pylori〈/em〉 strains during chronic infection, rather than mixed infection, and inappropriate medication appear to be main reasons of treatment failure in adults.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Pediococcus pentosaceus〈/em〉 GS4 (MTCC 12683), a probiotic lactic acid bacterium (LAB), was found to produce bacteriocin in spent culture. Antibacterial and antagonistic potential of this bacteriocin against reference strains of 〈em〉Staphylococcus aureus〈/em〉 (ATCC 25923), 〈em〉Escherichia coli〈/em〉 (ATCC 25922), 〈em〉Pseudomonas aeruginosa〈/em〉 (ATCC 25619), and 〈em〉Listeria monocytogenes〈/em〉 (ATCC 15313) was proven by double-layer and well diffusion methods wherein nisin and ampicillin were used as positive controls. Bacteriocin in supernatant was purified and analyzed by SDS-PAGE, RP-HPLC, and circular dichroism (CD). The physico-chemical properties of purified bacteriocin were characterized being treated at different temperatures (30 to 110 °C), pH (3.0 to 12.0), with different enzymes (α-amylase, pepsin, and lysozyme), and organic solvents (hexane, ethanol, methanol, and acetone) respectively. The molar mass of bacteriocin (named pediocin GS4) was determined as 9.57 kDa. The single peak appears at the retention time of 2.403 with area amounting to 25.02% with nisin as positive control in RP-HPLC. CD analysis reveals that the compound appears to have the helix ratio of 40.2% with no beta sheet. The antibacterial activity of pediocin GS4 was optimum at 50 °C and at pH 5.0 and 7.0. The pediocin GS4 was not denatured by the treatment of amylase and lysozyme but was not active in the presence of organic solvents. This novel bacteriocin thus m ay be useful in food and health care industry.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Upsurge in the instances of antibiotic-resistant uropathogenic 〈em〉Escherichia .coli〈/em〉 (UPECs) strains has repositioned the attention of researchers towards a century old antimicrobial approach popularly known as phage therapy. Rise of extended spectrum beta lactamase (ESBL) and biofilm producing strains has added another step of hurdle in treatment of uropathogens with conventional antibiotics, thus providing a further impetus for search for exploring new therapeutic measures. In this direction, bacteriophages, commonly called phages, are recently being considered as potential alternatives for treatment of UPECs. Phages are the tiniest form of viruses which are ubiquitous in nature and highly specific for their host. This review discusses the possible ways of using natural phages, genetically engineered phages, and phage lytic enzymes (PLEs) as an alternative antimicrobial treatment for urinary tract infections. The review also sheds light on the synergistic use of conventional antibiotics with phages or PLEs for treatment of uropathogens. These methods of using phages and their derivatives, alone or in combination with antibiotics, have proved fruitful so far in in vitro studies. However, in vivo studies are required to make them accessible for human use. The present review is a concerted effort towards putting together all the information available on the subject.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Homeostatic mechanisms preventing the toxicity of heavy metal ions in cells involve, among others, compartmentalization and binding with peptidaceous ligands, particularly the cysteinyl-rich metallothioneins (MTs). We have previously shown that in natural conditions Zn-overaccumulating ectomycorrhizal (EM) fungus 〈em〉Russula bresadolae〈/em〉 stores nearly 40% of Zn bound with cysteinyl- and hystidyl-containing RaZBP peptides, which resemble MTs, while the detoxification of Zn and Cd in EM 〈em〉Hebeloma mesophaeum〈/em〉 relies upon compartmentalization in small vesicles and vacuoles, respectively. Here, we examined the performance of Ra〈em〉ZBP1〈/em〉 gene expressed in 〈em〉H. mesophaeum〈/em〉 mycelium with respect to handling of Zn and Cd. Expression of Ra〈em〉ZBP1〈/em〉 impaired growth of the mycelium on low-Zn medium by 60%, the growth was partly ameliorated upon the addition of Zn and remained considerable up to 2 mmol/L Zn, while the growth of the wild-type and control mycelia transformed with empty T-DNA was severely reduced in the presence of 0.5 mmol/L Zn; furthermore, Ra〈em〉ZBP1〈/em〉 slightly added to Cd tolerance in the range of Cd concentrations of 0.625 to 8 μmol/L. Staining of Zn- or Cd-exposed hyphal cells with Zn- or Cd-specific fluorescent tracers did not indicate that the expression of Ra〈em〉ZBP1〈/em〉 would redirect the flow of the metals away from their innate sinks. Size exclusion chromatography of extracted metal species revealed that the complexes corresponding to Zn/Cd-RaZBP1 are present only in minute levels. Considering that RaZBP1 inhibited growth at low Zn, and despite the benefit that it provided to 〈em〉H. mesophaeum〈/em〉 in the presence of high Zn and moderate Cd, these data indicate that the binding of excess Zn and Cd with RaZBP1 is not a trait that would be outright transmitted to 〈em〉H. mesophaeum〈/em〉.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉We have worked out a rapid 1-day test based on photosynthesis measurements to estimate suitable growth temperature of microalgae cultures. To verify the proposed procedure, several microalgae—〈em〉Chlorella〈/em〉, 〈em〉Nostoc〈/em〉, 〈em〉Synechocystis〈/em〉, 〈em〉Scenedesmus〈/em〉, and 〈em〉Cylindrospermum—〈/em〉were cultured under controlled laboratory conditions (irradiance, temperature, mixing, CO〈sub〉2〈/sub〉, and nutrient supply) to find the optima of photosynthetic activity using the range between 15 and 35 °C. These activities were recorded at each temperature step after 2 h of acclimation which should be sufficient as oxygen production and the PQ cycle are regulated by fast processes. Photosynthetic activity was measured using three techniques—oxygen production/respiration, saturating pulse analysis of fluorescence quenching, and fast fluorescence induction kinetics—to estimate the temperature optima which should correspond to high growth rate. We measured all variables that might have been directly related to growth—photosynthetic oxygen evolution, maximum photochemical yield of PSII, 〈em〉F〈/em〉〈sub〉v〈/sub〉/〈em〉F〈/em〉〈sub〉m〈/sub〉, relative electron transport rate rETR〈sub〉max〈/sub〉, and the transients 〈em〉V〈/em〉〈sub〉j〈/sub〉 and 〈em〉V〈/em〉〈sub〉i〈/sub〉 determined by fast fluorescence induction curves. When the temperature optima for photosynthetic activity were verified in growth tests, we found good correlation. For most of tested microalgae strains, temperature around 30 °C was found to be the most suitable at this setting. We concluded that the developed test can be used as a rapid 1-day pre-screening to estimate a suitable growth temperature of microalgae strains before they are cultured in a pilot scale.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉In this work, the key moments of the development of the so-called thin-layer cascades (TLC) for microalgae production are described. Development started at the end of the 1950s when the first generation of TLCs was set-up in former Czechoslovakia. Since, similar units for microalgae culturing, which are relatively simple, low-cost and highly productive, have been installed in a number of other countries worldwide. The TLCs are characterized by microalgae growth at a low depth (〈 50 mm) and fast flow (0.4–0.5 m/s) of culture compared to mixed ponds or raceways. It guarantees a high ratio of exposed surface to total culture volume (〉 100 1/m) and rapid light/dark cycling frequencies of cells which result in high biomass productivity (〉 30 g/m〈sup〉2〈/sup〉/day) and operating at high biomass density, 〉 10 g/L of dry mass (DW). In TLCs, microalgae culture is grown in the system of inclined platforms that combine the advantages of open systems—direct sun irradiance, easy heat derivation, simple cleaning and maintenance, and efficient degassing—with positive features of closed systems—operation at high biomass densities achieving high volumetric productivity. Among significant advantages of thin layer cascades compared to raceway ponds are the operation at much higher cell densities, very high daylight productivities, and the possibility to store the culture in retention tanks at night, or in unfavourable weather conditions. Concerning the limitations of TLCs, one has to consider contaminations by other microalgae that limit cultivation to robust, fast-growing strains, or those cultured in selective environments.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Due to limitations in commercial diagnostic methods, this study aimed to develop a reliable real-time polymerase chain reaction (Rt-PCR) assay for early diagnosis of brucellosis. Optimization of the Rt-PCR method was performed on serum samples spiked by 〈em〉Brucella melitensis〈/em〉 with different densities ranging from 10〈sup〉1〈/sup〉 to 10〈sup〉8〈/sup〉 colony-forming units (cfu)/mL; each density was prepared in ten samples. The limit of detection was investigated by using Thermo DNA extraction kit with Maxima SYBR Green Rt-PCR and two TaqMan probe–based Rt-PCR protocols performed by QuantiTect and TEMPase multiplex PCR master mixes in two thermal cyclers, which were Rotor-Gene and Bio-Rad. The validation of the optimized protocol was carried on 20 brucellosis-negative samples and 20 samples spiked with 〈em〉B. melitensis〈/em〉 by using a combination of Thermo DNA extraction kit with TEMPase PCR master mix. SYBR Green Rt-PCR yielded positive results on all samples having ≥ 10〈sup〉4〈/sup〉 cfu/mL of 〈em〉B. melitensis〈/em〉 in both thermal cyclers. Its limit of detection was 112 DNA copies per reaction. The positivity of both probe-based Rt-PCR protocols was 100% and 80% on the samples having 10〈sup〉3〈/sup〉 cfu/mL and 10〈sup〉2〈/sup〉 cfu/mL of 〈em〉B. melitensis〈/em〉, respectively. The limit of detection of probe-based protocols was defined as 4 DNA copies per reaction. The optimized Rt-PCR protocol showed high-level accuracy, precision, specificity, and sensitivity, each having a rate of 100%. The current study indicated that the TaqMan probe–based Rt-PCR protocol optimized and validated with serum samples can be reliably used for early diagnosis of brucellosis.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Enteroviruses have been associated with a host of clinical presentations including acute flaccid paralysis (AFP). The site of primary replication for most enteroviruses is the gastrointestinal tract (GIT) and lactic acid bacteria (LAB) may confer protection in the GIT against them. This study therefore investigates the antiviral potential of some selected lactic acid bacteria against enterovirus isolates recovered from AFP cases. The antiviral activities of 〈em〉Lactobacillus plantarum〈/em〉, 〈em〉Lactobacillus amylovorus〈/em〉, and 〈em〉Enterococcus hirae〈/em〉 in broth culture, their cell-free supernatant (CFS), and bacterial cell pellets were assayed against 〈em〉Echovirus〈/em〉 7 (E7), E13, and E19 in a pre- and post-treatment approach using cytopathic effect (CPE) and cell viability (MTT) assay. The tested 〈em〉Lactobacillus plantarum〈/em〉, 〈em〉Lactobacillus amylovorus〈/em〉, and 〈em〉Enterococcus hirae〈/em〉 strains have good antiviral properties against E7 and E19 but not against E13. 〈em〉Lactobacillus amylovorus〈/em〉 AA099 shows the highest activity against E19. The pre-treatment approach displays better antiviral activities compared to post-treatment approach. The LAB in broth suspension have better antiviral activities than their corresponding CFS and bacterial pellet. Lactic acid bacteria used in this study have the potential as antiviral agents.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The 〈em〉Candida haemulonii〈/em〉 complex (〈em〉Candida haemulonii〈/em〉, 〈em〉Candida haemulonii〈/em〉 var. 〈em〉vulnera〈/em〉, and 〈em〉Candida duobushaemulonii〈/em〉) comprises emerging opportunistic human fungal pathogens with recognized multidrug-resistance profiles. Little is known about the virulence markers produced by this fungal complex. However, it is recognized that 〈em〉Candida〈/em〉 spp. express a large array of peptidases, which play multiple roles in different aspects of fungal-host interactions. In the present study, we have identified proteolytic enzymes in clinical isolates of the 〈em〉C〈/em〉. 〈em〉haemulonii〈/em〉 complex using zymographic assays. Peptidases able to hydrolyze gelatin, casein, albumin, hemoglobin, and immunoglobulin G were detected in cell-free supernatants and cellular extracts taken from the three species forming the 〈em〉C〈/em〉. 〈em〉haemulonii〈/em〉 complex. Overall, peptidases were preferentially evidenced at physiological pH and temperatures of 37–42 °C, with molar masses between 35 and 85 kDa. Peptidase profiles of 〈em〉C〈/em〉. 〈em〉haemulonii〈/em〉 and 〈em〉C〈/em〉. 〈em〉haemulonii〈/em〉 var. 〈em〉vulnera〈/em〉 isolates were quite similar, contrasting to the peptidases produced by 〈em〉C〈/em〉. 〈em〉duobushaemulonii〈/em〉. Almost all peptidases were inhibited by phenylmethanesulfonyl fluoride (PMSF), thus classifying them as serine-type peptidases. Additionally, proteolytic cleavage of soluble azoalbumin was blocked by PMSF (65–95% inhibition depending on the fungal isolate). These unprecedented results have demonstrated the capability of the 〈em〉C〈/em〉. 〈em〉haemulonii〈/em〉 complex to produce serine-type peptidases with an ability to cleave a broad spectrum of proteins, including key host components.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉A group of 59 putative strains of 〈em〉Staphylococcus intermedius〈/em〉/〈em〉Staphylococcus pseudintermedius〈/em〉 deposited in the Czech National Collection of Type Cultures (CNCTC, National Institute for Public Health, Prague, Czech Republic) and the National Reference Laboratory for Staphylococci (NRL for Staphylococci, National Institute for Public Health, Prague, Czech Republic) was reclassified using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). There the biggest human collection of 〈em〉S. pseudintermedius〈/em〉 in Europe was analysed; 44 samples (75%) were of human origin. Twenty-two percent (〈em〉n〈/em〉 = 13) of the strains were isolated from animals, and two staphylococci were of unknown origin. This study revealed the prevalence of 〈em〉Staphylococcus pseudintermedius〈/em〉 (94%, 〈em〉n〈/em〉 = 53) vs. 〈em〉Staphylococcus intermedius〈/em〉 (6%, 〈em〉n〈/em〉 = 6) in the collection of human and veterinary staphylococci after reclassification. Results of PCR-RFLP analysis were verified by comparison with a repetitive element sequence-based polymerase chain reaction (Rep-PCR) analysis on 26 (44%) randomly selected strains. Due to a low〈strong〉-〈/strong〉resolution ability of PCR-RFLP to separate 〈em〉Staphylococcus intermedius〈/em〉 from 〈em〉Staphylococcus delphini〈/em〉, 〈strong〉four〈/strong〉 isolates of 〈em〉Staphylococcus intermedius〈/em〉 were biochemically verified further to exclude the presence of 〈em〉Staphylococcus delphini〈/em〉 in the collection. Our results indicate that 〈em〉S. intermedius〈/em〉 and 〈em〉S. pseudintermedius〈/em〉 have occurred independently over an age-long period of their co-evolution.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Here, we report a case of neonatal calf meningitis due to 〈em〉Streptococcus gallolyticus〈/em〉 subsp. 〈em〉gallolyticus〈/em〉 (SGG). Clinical, pathological and microbiological findings were evaluated. API Strep, 16S rRNA gene sequencing, 〈em〉rpoB〈/em〉 gene sequencing and 〈em〉sodA〈/em〉 gene sequencing were used for the complete identification of SGG. This is the first documented report of neonatal calf meningitis due to SGG in veterinary medicine.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Inorganic polyphosphate is involved in architecture and functioning of yeast cell wall. The strain of 〈em〉Saccharomyces cerevisiae〈/em〉 constitutively overexpressing acid phosphatase Pho5 was constructed for studying the Pho5 properties and its possible participation in polyphosphate metabolism. The parent strain was transformed by the vector carrying the 〈em〉PHO5〈/em〉 gene under a strong constitutive promoter of glyceraldehyde-3-phosphate dehydrogenase of 〈em〉S. cerevisiae〈/em〉. The culture liquid and biomass of transformant strain contained approximately equal total acid phosphatase activity. The levels of acid phosphatase activity associated with the cell wall and culture liquid increased in the transformant strain compared to the parent strain ~ 10- and 20-fold, respectively. The Pho5 preparation (specific activity of 46 U/mg protein and yield of 95 U/L) was obtained from culture liquid of overproducing strain. The overproducing strain had no changes in polyphosphate level. The activity of Pho5 with long-chained polyP was negligible. We concluded that Pho5 is not involved in polyphosphate metabolism. Purified Pho5 showed a similar activity with p-nitrophenylphosphate, ATP, ADP, glycerophosphate, and glucose-6-phosphate. The substrate specificity of Pho5 and its extracellular localization suggest its function: the hydrolysis of organic compounds with phosphoester bonds at phosphate limitation.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Bacillus circulans〈/em〉 528 produces a restriction endonuclease, Bci528I which is an isoschizomer of EcoRI. We purified the enzyme, using Sephadex G-150, Phospho-cellulose, DEAE-cellulose, Hepharin-Sepharose CL-6B chromatography. The specific activity of Bci528I was 29,400 U/mg·protein. Bci528I recognizes 5′-GAATTC-3′ in dsDNA and cleaves between G and A of the recognition sequence, producing a symmetric four base 5′overhang.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim of the study was to screen 〈em〉Yarrowia lipolytica〈/em〉 strains for keto acid production and determine optimal conditions for pyruvic acid biosynthesis from glycerol by the best producer. The analyzed parameters were thiamine concentration, medium pH, stirring speed, and substrate concentration. The screening was performed in flask cultures, whereas pyruvic acid production was carried out in 5-L stirred-tank reactor with 2 L of working volume. In total, 24 〈em〉Y. lipolytica〈/em〉 strains were compared for their abilities to produce pyruvic and α-ketoglutaric acids. The total concentration of both acids ranged from 0.1 to 15.03 g/L. Ten strains were selected for keto acid biosynthesis in bioreactor. The 〈em〉Y. lipolytica〈/em〉 SKO 6 strain was identified as the best producer of pyruvic acid. In the selected conditions (thiamine concentration 1.5 μg/L, pH 4.0, stirring speed 800 rpm, 150 g/L of glycerol), the strain 〈em〉Y. lipolytica〈/em〉 SKO 6 produced 99.3 g/L of pyruvic acid, with process yield of 0.63 g/g and volumetric production rate of 1.18 g/L/h. Higher titer of pyruvic acid was obtained during fed-batch culture with 200 g/L of glycerol, reaching 125.8 g/L from pure glycerol (yield 0.68 g/g) and 124.4 g/L from crude glycerol (yield 0.62 g/g). Results obtained for the strain 〈em〉Y. lipolytica〈/em〉 SKO 6 proved the suitability of microbial production of pyruvic acid at industrial scale.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉An agar well diffusion assay (AWDA) was used to isolate a high bacteriocin-producing strain with a broad spectrum of antibacterial activity, strain MXG-68, from Inner Mongolia traditional fermented koumiss. 〈em〉Lactobacillus plantarum〈/em〉 MXG-68 was identified by morphological, biochemical, and physiological characteristics and 〈em〉16S rDNA〈/em〉 analysis. The production of antibacterial substance followed a growth-interrelated model, starting at the late lag phase of 4 h and arriving at a maximum value in the middle of the stationary phase at 24 h. Antibacterial activity was abolished or decreased in the presence of pepsin, chymotrypsin, trypsin, proteinase, and papain K. The results showed that antibacterial substances produced by 〈em〉L. plantarum〈/em〉 MXG-68 were proteinaceous and could thus be classified as the bacteriocin, named plantaricin MXG-68. The molar mass of plantaricin MXG-68 was estimated to be 6.5 kDa, and the amino acid sequence of its N-terminal was determined to be VYGPAGIFNT. The mode of plantaricin MXG-68 action was determined to be bactericidal. Bacteriocin in cell-free supernatant (CFS) at pH 7 was stable at different temperatures (60 °C, 80 °C, 100 °C, 121 °C for 30 min; 4 °C and − 20 °C for 30 days), as well as at pH 2.0–10.0. Antibacterial activity maintained stable after treatment with organic solvents, surfactants, and detergents but increased in response to EDTA. Response surface methodology (RSM) revealed the optimum conditions of bacteriocin production in 〈em〉L. plantarum〈/em〉 MXG-68, and the bacteriocin production in medium optimized by RSM was 26.10% higher than that in the basal MRS medium.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Staphylococcus aureus〈/em〉 (〈em〉S. aureus〈/em〉) is an important causative agent of contagious intermammary infections in dairy cattle. 〈em〉S. aureus〈/em〉 is also considered as an important foodborne pathogen and cause of food poisoning cases and outbreaks worldwide. In order to understand the molecular ecology of 〈em〉S. aureus〈/em〉, the present study compared phenotypic and genotypic characteristics of 70 〈em〉S. aureus〈/em〉 isolates from bovine mastitis milk samples collected during the period from August 2001 to March 2014 in different regions of Northern Germany. The 〈em〉S. aureus〈/em〉 isolates were characterised phenotypically, as well as genotypically for their genetic diversity using multi-locus sequence typing (MLST), 〈em〉spa〈/em〉 typing and the presence of virulence genes encoding 16 staphylococcal enterotoxins (〈em〉sea-selu〈/em〉), toxic shock syndrome toxin (〈em〉tst〈/em〉), thermonuclease (〈em〉nuc〈/em〉), clumping factor (〈em〉clfA〈/em〉 and 〈em〉clfB〈/em〉), coagulase (〈em〉coa〈/em〉) and the methicillin resistance gene 〈em〉mecA〈/em〉. A total of 16 sequence types were grouped into eight clonal complexes (CCs), and 17 〈em〉spa〈/em〉 types were identified. These included six novel sequence types and one novel 〈em〉spa〈/em〉 type. The majority of bovine mastitis milk-associated sequence types belonged to the clonal complex CC5, CC97, CC133, and CC151 and showed closely related genotypes or lineages with sequence types of human origin. The genotype CC133 (ST133-t1403) was predominant, constituting 27.1% of the isolates. In addition, the 〈em〉S. aureus〈/em〉 isolates displayed nine different enterotoxigenic profiles. All 〈em〉S. aureus〈/em〉 were methicillin-susceptible (MSSA). The current study provides new information on phenotypic and genotypic traits of 〈em〉S. aureus〈/em〉 isolates from bovine mastitis〈em〉.〈/em〉 The comparison of characteristics of isolates from the present study originating from mastitis milk showed similarities with human isolates. This might help to better understand the distribution of 〈em〉S. aureus〈/em〉 in the one health context.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Endophytic fungi live inside vegetal tissues without causing damage to the host plant and may provide lead compounds for drug discovery. The co-culture of two or more endophytic fungi can trigger silent gene clusters, which could lead to the isolation of bioactive compounds. In this study, two endophytic strains isolated from 〈em〉Handroanthus impetiginosus〈/em〉 leaves, identified as 〈em〉Talaromyces purpurogenus〈/em〉 H4 and 〈em〉Phanerochaete〈/em〉 sp. H2, were grown in mixed and axenic cultures. The meroterpenoid austin was detected only in the extracts from the mixed culture. Once isolated, austin displayed very interesting trypanocidal activity, with an IC〈sub〉50〈/sub〉 value of 36.6 ± 1.2 μg/mL against 〈em〉Trypanosoma cruzi〈/em〉 in the epimastigote form. The results obtained highlight the importance of the co-culturing of endophytic fungi to obtain natural bioactive products. The findings also enhance our understanding of the ecological relationships between endophytic fungi.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Anaerobic ammonium-oxidizing (anammox) bacteria play an essential part in nitrogen removal in constructed wetlands. The objective of the present study was to explore the spatiotemporal dynamics of anammox bacterial communities and the associated factors in a full-scale constructed wetland for the treatment of polluted surface water. The abundance and diversity of anammox bacterial communities were characterized using quantitative polymerase chain reaction (PCR) and clone library analysis, respectively. Anammox bacterial diversity, richness, and abundance in the treatment wetland differed considerably among sampling sites and seasons, whereas anammox bacterial community structure tended to change slightly with site and time. Anammox abundance was likely influenced by temperature and the contents of nitrate and nitrite nitrogen. The increase of carbon and nitrogen contents could lower wetland anammox bacterial diversity and richness. Moreover, anammox bacterial diversity, richness, and abundance were also affected by wetland vegetation type. 〈em〉Candidatus Brocadia〈/em〉 dominated in the treatment wetland, whereas 〈em〉Candidatus Kuenenia〈/em〉 and a novel anammox phylotype were also detected. This work could provide some new insights towards anaerobic ammonium oxidization in surface water treatment wetland.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Today, many microbial amylases are available commercially and they have almost completely replaced chemical hydrolysis in several industry processes. Amylases from microorganisms have a broad spectrum of industrial applications as they are more stable than amylases obtained from plants and animals. The objective of this work was to use potato baits in an Atlantic Forest remnant located in Ribeirão Preto, São Paulo, Brazil, in order to obtain amylase-producing fungi with potential for biotechnological application. In addition, the culture conditions for the fungal strain that presented higher production of glucoamylase were standardized using industrial wastes. For this, 6 PET bottles containing potatoes as baits were scattered at different points in an Atlantic forest remnant. After 6 days, the samples were collected, and the filamentous fungi were isolated in Petri dishes. Fungi screening was carried out in Khanna liquid medium with 1% starch Reagen®, at 30 °C, pH 6.0, under static conditions for 4 days. Proteins and glucoamylase activity were determined by Bradford and 3,5-dinitrosalicylic acid (DNS), respectively. Among all isolated fungi, 〈em〉A. carbonarius〈/em〉 showed the highest glucoamylase production. Its best cultivation conditions were observed in Khanna medium, 4 days, at 30 °C, pH 6.0, under static condition with 0.1% yeast extract and 1% starch Reagen®. Wheat and brewing residues were also used as inducers for large quantities of glucoamylase production. 〈em〉A. carbonarius〈/em〉 showed to be a good alternative for the wheat and brewing waste destinations in order to obtain high added value products.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Biofilm-associated bacterial infections represent one of the major threats to modern medical treatments. Bacteria encased in biofilm matrix are more resistant towards antimicrobials and thus the capability of microbes to persist and nurture in a biofilm seems to be the foremost aspect of pathogenesis and therapeutic failure. Therefore, there is a pressing demand for new drugs active against microbial biofilms. In the current study, anti-biofilm potential of 〈em〉Lactobacillus〈/em〉 spp. cell-free supernatants (CFSs) against 〈em〉Cronobacter sakazakii〈/em〉 and 〈em〉Listeria monocytogenes〈/em〉 was characterized using crystal violet staining and MTT assay. CFSs of goat milk origin lactobacilli not only prevented biofilm formation but also disrupted preformed biofilms. Neutralized and heat-treated preparations of 〈em〉Lactobacillus〈/em〉 CFSs also inhibited biofilm formation by test pathogens. The results were quantitatively confirmed by light and fluorescent microscopy observations. Biofilms developed under static conditions displayed typical compact microcolonies with uniform distribution over the surface, while upon CFS challenge, biofilms were disrupted with presence of dead cells. These findings highlight the anti-biofilm potency of 〈em〉Lactobacillus〈/em〉 spp. strains of goat milk origin and their potential application in food industries.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉To understand the role of phospholipids on Cdr1p (drug exporter)-mediated drug resistance in yeast, the phospholipids biosynthesis genes 〈em〉PSD1〈/em〉, 〈em〉PSD2〈/em〉, 〈em〉CHO2〈/em〉, and 〈em〉OPI3〈/em〉 were deleted in a strain of 〈em〉Saccharomyces cerevisiae〈/em〉 already overexpressing Cdr1-GFP of 〈em〉Candida albicans〈/em〉 as a heterologous system. The effect of phospholipids biosynthesis gene deletion was analyzed on Cdr1p-GFP-mediated drug resistance as well as its localization. The results indicate that phospholipids biosynthesis disruption makes the cell sensitive to several drugs including fluconazole (FLC), with Δ〈em〉psd1〈/em〉/Cdr1-GFP being worst affected. Interestingly, unlike sterols and sphingolipids, the localization of Cdr1p was unaffected by phospholipid biosynthesis gene disruption. Concomitantly, phospholipids mutants also showed an increase in reactive oxygen species (ROS) generation, as verified by fluorescence probe 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) method. In addition, the sensitivity of phospholipid mutants with FLC was found to be synergistic to ROS generation, resulting in further reduction of growth. Thus, this study proposes phospholipid biosynthesis as a novel target for antifungal therapy.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉In Slovakia, dairy products made from ewes’ milk have a long tradition. These products include the lactic acid product called “žinčica” which is a by-product occurring during the preparation of ewes’ lump cheese. There is no information in the literature regarding the special properties of the microbiota, especially lactic acid Firmicutes, which can survive in “žinčica.” From the safety aspect, enterococci are a controversial group of bacteria, and those from “žinčica” have never been tested for their properties. The “žinčica” used in our study was supplied by several different agrofarms producing ewes’ lump cheese in central Slovakia. The species 〈em〉Enterococcus faecium〈/em〉 (strains EF30E1, EF32E1, EF34E1, EF34E5) and 〈em〉Enterococcus faecalis〈/em〉 (strains EE30E4, EE35E1, E31E2, altogether 7) were detected in samples from “žinčica” identified using MALDI-TOF spectrometry with secure genus identification/probable species identification and then confirmed by means of PCR. Enterococci were hemolysis-negative and the genes of the typical enterococcal virulence factors were mostly absent; the g〈em〉elE〈/em〉 gene was found in two 〈em〉E. faecium〈/em〉 strains (EF30E1 and EF32E1), the 〈em〉agg〈/em〉 gene was detected in 〈em〉E. faecalis〈/em〉 EE35E1, and the 〈em〉esp〈/em〉 gene was found in two 〈em〉E. faecalis〈/em〉 strains (EE30E4 and EE31E2). No strains harbored the 〈em〉cytolysin A〈/em〉 gene. Biofilm formation was detected in four strains (EF30E1, EF32E1, EF34E1, and EF34E5), indicating highly positive and low-grade positive biofilm formation. Enterococci were mostly susceptible to antibiotics tested for their phenotype. This is the first study to analyze enterococci in “žinčica.”〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Ascochyta blight of chickpea is caused by 〈em〉Ascochyta rabiei〈/em〉 (Pass.) Labr. which is primarily seedborne. For rapid detection and precise identification of 〈em〉A. rabiei〈/em〉, a sequence-characterized amplified region (SCAR) marker was developed for detection of genomic DNA and infected plant DNA. An SSR primer amplified monomorphic band was cloned in pGEM®-T easy vector and sequenced. The best primer pair was selected and validated on 〈em〉A. rabiei〈/em〉. The specificity and sensitivity of the SCAR-based marker designated as MBAR was evaluated using conventional PCR and real-time PCR. The marker produced consistently an amplicon size of 196 bp in all 〈em〉A. rabiei〈/em〉 isolates tested. The sensitivity of the marker was 0.1 ng of genomic fungal DNA and 0.5 ng of plant DNA by conventional PCR and 0.5 pg of 〈em〉A. rabiei〈/em〉 DNA and 1.0 pg of plant DNA by real-time PCR. This is the first SCAR marker having high specificity and sensitivity towards 〈em〉A. rabiei〈/em〉. The marker may be useful in detecting the pathogen before the disease appearance and in plant quarantine program to detect the pathogen in seed lots.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Klebsiella pneumoniae〈/em〉 infections have always been an important problem in public health, but today, the increasing resistance of these bacteria to antibiotics due to β-lactamases production has renewed interest in 〈em〉K. pneumoniae〈/em〉 infections. The aim of the study was to present a case of a neurosurgical patient with multidrug-resistant 〈em〉K. pneumoniae〈/em〉 ST11 infection after craniectomy. Four 〈em〉K. pneumoniae〈/em〉 isolates from various clinical materials of the patient undergone identification and susceptibility testing with the Vitek2 system. Tests for β-lactamases production were performed according to EUCAST guidelines. Strains were analyzed for 〈em〉bla〈/em〉 genes responsible for β-lactamase production (〈em〉bla〈/em〉〈sub〉TEM〈/sub〉, 〈em〉bla〈/em〉〈sub〉SHV〈/sub〉, 〈em〉bla〈/em〉〈sub〉CTX-M〈/sub〉, 〈em〉bla〈/em〉〈sub〉VIM〈/sub〉, 〈em〉bla〈/em〉〈sub〉IMP〈/sub〉, 〈em〉bla〈/em〉〈sub〉NDM〈/sub〉, 〈em〉bla〈/em〉〈sub〉KPC〈/sub〉, 〈em〉bla〈/em〉〈sub〉OXA-48〈/sub〉) using PCR. Moreover, the genetic relatedness of these isolates was determined by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). All tested strain presented multidrug resistance. The highest susceptibility was observed for imipenem, meropenem, and ertapenem. The strain isolated from the nervous system was ESBL-positive with 〈em〉bla〈/em〉〈sub〉SHV-11〈/sub〉, 〈em〉bla〈/em〉〈sub〉TEM-1〈/sub〉, and 〈em〉bla〈/em〉〈sub〉CTX-M-15〈/sub〉 genes. Additionally, the strain from urine was 〈em〉bla〈/em〉〈sub〉KPC-3〈/sub〉-positive. Molecular typing revealed that all strains belonged to the same clone and identified two PFGE profiles. The analysis of MLST allelic profile showed that tested 〈em〉K. pneumoniae〈/em〉 strains belonged to ST11. Identification of ST11 〈em〉K. pneumoniae〈/em〉 as etiological factor of infection unfavorably impacts on prognosis among neurosurgical patient after craniectomy.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Peptidyl-prolyl 〈em〉cis-trans〈/em〉 isomerases (PPIase) exhibit chaperone activity and assist in protein folding by increasing the rate of 〈em〉cis-trans〈/em〉 transition on proline-peptide bonds. The current study aimed to identify and characterize three genes, 〈em〉ppiA〈/em〉, 〈em〉ppiB〈/em〉, and 〈em〉ppiC〈/em〉, which encode proteins of the PPIase family in the bacterium 〈em〉Salmonella enterica〈/em〉 serovar Typhimurium. 〈em〉Salmonella〈/em〉 Typhimurium is a facultative intracellular zoonotic pathogen that causes food- and water-borne gastroenteritis in humans (leading to bacteremia in immune-compromised subjects). Recombinant clones for the three genes were constructed and sequenced and the sequences submitted to NCBI GenBank. Three-dimensional structures for the corresponding proteins were predicted by comparative modeling. A maximum-likelihood phylogenetic gene tree constructed for the three genes showed a low evolutionary mean diversity, indicating strong evolutionary conservation. Further, single-gene deletion mutant strains, generated for the respective genes, were observed to be more susceptible to the stationary phase of growth and heat stress conditions and showed reduced survival within macrophage cells line. The present study thus indicates that 〈em〉ppiA〈/em〉, 〈em〉ppiB〈/em〉, and 〈em〉ppiC〈/em〉 genes are conserved among 〈em〉Salmonella〈/em〉 genome, are critical for the growth of 〈em〉Salmonella〈/em〉 Typhimurium in the examined stress conditions, and may play a role in its responses and virulence.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The presence of a diversity of virulence factors such as Chaperone-Usher (CU) fibers in uropathogenic 〈em〉Escherichia coli〈/em〉 (UPEC) pathotypes virulome has offered these bacteria a unique opportunity to select the right factors in suitable condition. Understanding the structures and mechanisms of these infectious weapons enables us to prevent the occurrence of infections and/or to treat the urinary tract infections (UTIs) with effective methodologies. This review summarizes the current knowledge on the mechanisms of CU fimbriae (CUF) formation in UPEC. Several CU fibers including Auf, Dr, F1C, S, Type 9, Type 3, Type 1, and P fimbriae have been recognized in UPEC pathotypes. These fimbrial organelles may have cross-talk with each other in the presence of different environmental factors. In other words, the expression or the silence of their genes are associated with a wide range of items comprising environmental factors, genetical factors, physiological factors, etc. Recognition, detection, and identification of virulome and the related characteristics, properties, and mechanisms in UPEC enable us to create new methodologies to have a definite prevention and treatment for UTIs in this regard.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Biofilm formation is regarded as an important factor in the establishment of infections caused by 〈em〉Staphylococcus aureus〈/em〉. In the present study, phenotypic and molecular assays were used to evaluate antibiofilm potential of thiosemicarbazide (Tsc) conjugated with silver nanoparticles (Ag NPs) and functionalized by glutamic acid (Ag@Glu/Tsc NPs) against methicillin-resistant 〈em〉S. aureus〈/em〉 (MRSA). Ag NPs were synthesized using precipitation method and conjugated to Tsc using glutamic acid. The NPs were characterized using SEM and FTIR spectroscopy analyses. Then, antibiofilm potential of the prepared NPs against MRSA strains was evaluated using phenotypic method and their effects on the expression of biofilm-associated genes 〈em〉icaA〈/em〉 and 〈em〉icaD〈/em〉. Finally, the genes involved with the synthesis of intercellular adhesion molecules were determined. According to the results, Ag@Glu/Tsc NPs inhibited biofilm formation of MRSA strains up to 76.7% compared with the control. In addition, expression of the biofilm-associated genes 〈em〉icaA〈/em〉 and 〈em〉icaD〈/em〉 reduced by 66.7% and 60.3%, respectively in the presence of sub-inhibitory concentration of Ag@Glu/Tsc NPs. In conclusion, Ag@Glu/Tsc NPs could be considered as a potent antibacterial agent to inhibit bacterial biofilms.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉This study aimed to analyze the proinflammatory cytokine mRNA expression in the urinary tract of BALB/c mice infected with bacterial strains with uropathogenic potential. Groups of four 6-week-old female BALB/c mice were intraurethrally inoculated with 5 × 10〈sup〉7〈/sup〉 colony-forming units (CFU) of 〈em〉P. mirabilis〈/em〉 ATCC29906, EAEC O42, 〈em〉P. mirabilis〈/em〉 RTX339, or sterile saline (control group) and then sacrificed at 0, 2, 4, 7, or 10 days post-infection (p.i.). Samples were cultured to determine the CFU/mL in urine or CFU/g in the bladders and kidneys. Cytokine expression (tumor necrosis factor (TNF)-α and interleukin (IL)-1β, -6, and -8) was evaluated in the target organs using real-time PCR and immunohistochemistry; histology was examined with hematoxylin and eosin staining. The results are presented as the means and standard deviations and were compared using one-way ANOVA, with 〈em〉p〈/em〉 〈 0.05 indicating significant differences. Bacteriuria was not detected in the infected groups; bacterial colonization occurred in the target organs at all time points, but was higher in mice infected with EAEC O42 or 〈em〉P. mirabilis〈/em〉 RTX339 at 7 days p.i. The expression of all cytokine mRNAs was seen, but only the levels of the IL-8 protein increased in situ at 7 days p.i. in the 〈em〉P. mirabilis〈/em〉 RTX339 and EAEC O42 groups in both organs. Morphological alterations, observed in all of the infected groups, were more prominent in the EAEC O42 and 〈em〉P. mirabilis〈/em〉 RTX339 groups. The findings provide insights into the uropathogenicity and inflammatory cytokine expression in the urinary tract of mice infected with three previously untested bacterial strains.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Cerebral abscesses caused by dark-pigmented 〈em〉Fonsecaea〈/em〉 fungi are rare, especially in otherwise healthy individuals. In this case report, we present a 61-year-old man from Moldova, living in the Czech Republic, who had worked as a locksmith on oil platforms in Turkmenistan, Kazakhstan, Sudan, and Iraq since 1999, and was admitted to a neurology ward for a sudden motion disorder of the right leg, dysarthria, and hypomimia. Imaging revealed presence of expansive focus around the left lateral ventricle of the brain and a pronounced peripheral edema. The intracranial infectious focus was excised under intraoperative SonoWand guidance. Tissue samples were histologically positive for dark-pigmented hyphae, suggesting dematiaceous fungi. Therefore, liposomal amphotericin B therapy was initiated immediately. 〈em〉Fonsecaea monophora〈/em〉 was provisionally identified using ITS rDNA region sequencing directly from brain tissue. The identification was subsequently confirmed by cultivation and DNA sequencing from culture. The strain exhibited in vitro sensitive to voriconazole (MIC = 0.016 μg/mL) and resistance to amphotericin B (MIC = 4 μg/mL); therefore, the amphotericin B was replaced with voriconazole. Postoperatively, a significant clinical improvement was observed and no additional surgery was required. Based on the literature review, this is the third documented case of cerebral infection due to this pathogen in patients without underlying conditions and the first such case in Europe.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The 〈em〉luxS〈/em〉 gene is responsible for the synthesis of AI-2 (autoinducer-2), a signaling molecule that participates in quorum sensing regulation in a large number of bacteria. In this work, we investigated which phenotypes are regulated by 〈em〉luxS〈/em〉 gene in 〈em〉Serratia proteamaculans〈/em〉 94, psychrotrophic strain isolated from spoiled refrigerated meat. AI-2 was identified in 〈em〉S. proteamaculans〈/em〉 94, and the 〈em〉luxS〈/em〉 gene involved in its synthesis was cloned and sequenced. A mutant with the inactivated 〈em〉luxS〈/em〉 gene was constructed. Inactivation of the 〈em〉luxS〈/em〉 gene was shown to lead to the absence of AI-2 synthesis, chitinolytic activity, swimming motility, suppression of the growth of fungal plant pathogens 〈em〉Rhizoctonia solani〈/em〉 and 〈em〉Helminthosporium sativum〈/em〉 by volatile compounds emitted by 〈em〉S. proteamaculans〈/em〉 94 strain, and to a decrease of extracellular proteolytic activity. The knockout of the 〈em〉luxS〈/em〉 gene did not affect synthesis of 〈em〉N〈/em〉-acyl-homoserine lactones, lipolytic, and hemolytic activities of 〈em〉S. proteamaculans〈/em〉 94.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Enzymes of microbial origin are of immense importance for organic material decomposition leading to bioremediation of organic waste, bioenergy generation, large-scale industrial bioprocesses, etc. The market demand for microbial cellulase enzyme is growing more rapidly which ultimately becomes the driving force towards research on this biocatalyst, widely used in various industrial activities. The use of novel cellulase genes obtained from various thermophiles through metagenomics and genetic engineering as well as following metabolic engineering pathways would be able to enhance the production of thermophilic cellulase at industrial scale. The present review is mainly focused on thermophilic cellulolytic bacteria, discoveries on cellulase gene, genetically modified cellulase, metabolic engineering, and their various industrial applications. A lot of lacunae are yet to overcome for thermophiles such as metagenome analysis, metabolic pathway modification study, search of heterologous hosts in gene expression system, and improved recombinant strain for better cellulase yield as well as value-added product formation.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Diseases of the central nervous system (CNS) mean for the human organism a potentially dangerous situation. An investigation of cerebrospinal fluid (CSF) provides important information about a character of CNS impairment in the decision-making diagnostic and therapeutic algorithm. The authors present a brief overview of available cerebrospinal fluid assays, shortened indication criteria, a recommended algorithm of CSF assessment in different suspected diseases, and a view of the external quality system. The whole portfolio of obtainable CSF methodology is further subdivided according to the adequate choice into the first and inevitable basic routine panel, and following complicated analyses of highly specialized character. The basic panel is considered for standard laboratories, the complete specialized assessment should be provided by a super-consulting laboratory.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉〈em〉Beauveria bassiana〈/em〉 is widely studied as an alternative to chemical acaricides in controlling the cattle tick 〈em〉Rhipicephalus microplus.〈/em〉 Although its biocontrol efficiency has been proved in laboratory and field scales, there is a need to a better understanding of host interaction process at molecular level related to biocontrol activity. In this work, applying a proteomic technique multidimensional protein identification technology (MudPIT), the differential secretome of 〈em〉B. bassiana〈/em〉 induced by the host 〈em〉R. microplus〈/em〉 cuticle was evaluated. The use of the host cuticle in a culture medium, mimicking an infection condition, is an established experimental model that triggers the secretion of inducible enzymes. From a total of 236 proteins, 50 proteins were identified exclusively in infection condition, assigned to different aspects of infection like host adhesion, cuticle penetration and fungal defense, and stress. Other 32 proteins were considered up- or down-regulated. In order to get a meaningful global view of the secretome, several bioinformatic analyses were performed. Regarding molecular function classification, the highest number of proteins in the differential secretome was assigned in to hydrolase activity, enzyme class of all cuticle-degrading enzymes like lipases and proteases. These activities were also further validated through enzymatic assays. The results presented here reveal dozens of specific proteins and different processes potentially implicated in cattle tick infection improving the understanding of molecular basis of biocontrol of 〈em〉B. bassiana〈/em〉 against 〈em〉R. microplus.〈/em〉〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Mucormycosis is a rare fungal infection in immunocompetent patients, whereas in immunocompromised, it may be systemic and disseminated infection associated with high mortality. Mucormycosis is one of the most rapidly progressing and fulminant forms of fungal infections; 〈em〉Mucor circinelloides〈/em〉 is rarely isolated species, also from immunocompromised patients. The reported case of mucormycosis after a traffic accident indicates that it may be the result of a contamination of wound by 〈em〉M. circinelloides〈/em〉 coming from the environment. The fungal strain was identified by phenotypic methods and confirmed by molecular methods. Etest method was used for susceptibility testing of the fungal strain. No mycotoxins were detected in the analyzed sample. The infection was successfully treated with amphotericin B, but amputation of the lower limb was necessary.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉This is the first exhaustive report on the fungal community biodiversity in hypersaline water in România. A total of 27 fungal strains (19 molds and eight yeast) have been isolated from Lopătari hypersaline water, Buzau County. Based on classical investigation, these strains have been identified as belonging to the genera 〈em〉Aureobasidium〈/em〉, 〈em〉Alternaria〈/em〉, 〈em〉Aspergillus〈/em〉, 〈em〉Penicillium〈/em〉, and 〈em〉Fusarium〈/em〉. The molecular characterization of fungal isolates at species level was performed using PCR-RFLP analysis of the 5.8S-ITS region. PCR products were digested with different combinations of endonucleases. The most frequently isolated species were 〈em〉Aspergillus niger〈/em〉 (14.81% of all isolates), 〈em〉A. versicolor〈/em〉, (14.81%) and 〈em〉Penicillium crustosum〈/em〉 (14.81%). In addition, ribosomal restriction patterns which exhibited profiles specific to 〈em〉Aureobasidium pullulans〈/em〉 were derived, and to discriminate between 〈em〉Aureobasidium〈/em〉 isolates, the elongase-encoding gene (〈em〉ELO〈/em〉) was chosen as a genetic marker followed by digestion with endonuclease 〈em〉Hha〈/em〉I. Five yeast isolates displayed restriction patterns corresponding to 〈em〉Aureobasidium melanogenum〈/em〉 (18.52%) and three isolates to 〈em〉Aureobasidium pullulans〈/em〉 (11.11%). In addition, the RFLP types of 〈em〉Aureobasidium pullulans〈/em〉 varieties with 〈em〉Hha〈/em〉I are clearly distinguished and could be applied to assess the intraspecific variability.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Rapid diagnostics of fungal pneumonia and initiation of appropriate therapy are still challenging. In this study, we used two panfungal assays to test bronchoalveolar lavage fluid (BALF) samples to prove their ability to confirm invasive fungal disease diagnosis and identify causative agents. Two methods targeting different fungal rDNA regions were used, and the obtained PCR products were sequenced directly or after cloning. In total, 106 BALF samples from 104 patients were tested. After sequencing, we obtained 578 sequences. Four hundred thirty-seven sequences were excluded from further analysis due to duplication (〈em〉n〈/em〉 = 335) or similarity with sequences detected in the extraction control sample (〈em〉n〈/em〉 = 102); 141 unique sequences were analyzed. Altogether, 23/141 (16%) of the fungi detected belonged to pathogenic species, and 63/141 (45%) were identified as various yeasts; a variety of environmental or very rare fungal human pathogens represented 29/141 (21%) of the total and 26/141 (18%) were described as uncultured fungus. Panfungal PCR detected fungal species that would be missed by specific methods in only one case (probable cryptococcosis). Panfungal PCR followed by sequencing has limited use for testing BALF samples due to frequent commensal or environmental fungal species pickup.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The ability of 〈em〉Diplodia pinea〈/em〉 to inhibit 〈em〉Armillaria〈/em〉 sp., 〈em〉Bjerkandera adusta〈/em〉, 〈em〉Botrytis cinerea〈/em〉, and 〈em〉Rhizoctonia〈/em〉 sp. mycelium growth was analyzed using the double-culture method. Wild-type fungal strains were incubated in a biochemical oxygen demand incubator using potato agar dextrose medium at 24 ± 2 °C for 35 days in darkness. 〈em〉D. pinea〈/em〉 significantly inhibited the growth of all fungi species tested (30.75 to 98.37% inhibition) and showed moderate antagonistic activity (antagonistic index, 14.5). Chemical analysis of 〈em〉D. pinea〈/em〉 culture broth extracts revealed steroids, triterpenes, and phenolic compounds. Alkaloids were qualitatively detected in the mycelium crude extract. The presence of these compounds may be related to the antagonistic activity observed. The inhibition ability of 〈em〉D. pinea〈/em〉 is due to competition with the tested fungi for substrate and space.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Endo-glucanase (cellulase) and xylanase have high industrial demand due to their vast application in industrial processes. This study reports statistical based experimental optimization for co-production of endo-glucanase and xylanase from 〈em〉Bacillus sonorensis〈/em〉 BD92. Response surface methodology (RSM) involving central composite design (CCD) with full factorial experiments (2〈sup〉3〈/sup〉) was applied to elucidate the components that significantly affect co-production of endo-glucanase and xylanase. The optimum co-production conditions for endo-glucanase and xylanase were as follows: carboxymethyl cellulose (CMC) 20 g/L, yeast extract 15 g/L, and time 72 h. The maximum endo-glucanase and xylanase production obtained was 1.46 and 5.69 U/mL, respectively, while the minimum endo-glucanase and xylanase production obtained was 0.66 and 0.25 U/mL, respectively. This statistical model was efficient because only 20 experimental runs were necessary to assess the highest production conditions, and the model accuracy was very satisfactory as coefficient of determination (〈em〉R〈/em〉〈sup〉2〈/sup〉) was 0.95 and 0.89 for endo-glucanase and xylanase, respectively. Further, potential application of these enzymes for saccharification of lignocellulosic biomass (wheat bran, wheat straw, rice straw, and cotton stalk) was also investigated. The results revealed that the biomass was susceptible to enzymatic saccharification and the amount of reducing sugars (glucose and xylose) increased with increase in incubation time. In conclusion, 〈em〉Bacillus sonorensis〈/em〉 BD92 reveals a promise as a source of potential endo-glucanase and xylanase producer that could be useful for degrading plant biomass into value-added products of economic importance using precise statistically optimized conditions.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉A quarantine organism, “〈em〉Candidatus〈/em〉 Phytoplasma mali,” is the causal agent of apple proliferation, one of the most important apple diseases in Europe. The genetic diversity of this pathogen in Central and Southern Europe has already been reported; however, almost no data exists from Eastern Europe. In this study, “〈em〉Ca〈/em〉. P. mali” strains, which were identified in 14 apple trees from the Bulgarian germplasm collection, were characterized by restriction fragment length polymorphism (RFLP) and sequence analysis of four genomic loci. In total, nine distinct genetic lineages were recognized based on the combination of the following detected RFLP profiles: two profiles for the 16S-23S rDNA region (16SrX-A2, -A3), four profiles for the 〈em〉secY〈/em〉 gene (one previously known: secY(X)-A, and three new: secY-C, secY-D, secY-E), three profiles for the 〈em〉rpl22-rps3〈/em〉 genes (rpX-A, rpX-B, rpX-F), and one profile for the nitroreductase- and rhodanese-like gene (AT-1). Phylogenetic analysis of the Bulgarian and other European “〈em〉Ca〈/em〉. P. mali” strains based on 16S-23S rRNA gene sequences confirmed RFLP grouping, regardless of the phytoplasma origin. In a phylogenetic tree based on the 〈em〉secY〈/em〉 data, only German strains formed separate clade from the other strains. The tree based on rp genes did not correspond to RFLP profiles. Unexpectedly, when using nitroreductase and rhodanese-like gene sequences, the Bulgarian strains clustered separately from the other European strains. Apart from the identification of different “〈em〉Ca〈/em〉. P. mali” strains, the paper also recommends the unification of the rpX-subgroup nomenclature to avoid future confusions. Both aims of this paper provide valuable tools to understand the epidemiology of this quarantine pathogen.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Due to the increasing number of 〈em〉Candida albicans〈/em〉’ infections and the resistance of this pathogenic fungus to drugs, new therapeutic strategies are sought. One of such strategies may be the use of static magnetic field (SMF). 〈em〉C. albicans〈/em〉 cultures were subjected to static magnetic field of the induction 0.5 T in the presence of fluconazole and amphotericin B. We identified a reduction of 〈em〉C. albicans〈/em〉 hyphal length. Also, a statistically significant additional effect on the viability of 〈em〉C. albicans〈/em〉 was revealed when SMF was combined with the antimycotic drug amphotericin B. The synergistic effect of this antimycotic and SMF may be due to the fact that amphotericin B binds to ergosterol in plasma membrane and SMF similarly to MF could influence domain orientation in plasma membrane (PM).〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim was to evaluate in vitro possible interactions, gene expression, and biofilm formation in species of 〈em〉Candida albicans〈/em〉, 〈em〉Streptococcus mitis〈/em〉, and 〈em〉Streptococcus sanguinis〈/em〉 and their in vivo pathogenicity. The in vitro analysis evaluated the effects of 〈em〉S. mitis〈/em〉 and 〈em〉S. sanguinis〈/em〉 on 〈em〉C. albicans〈/em〉’s biofilm formation by CFU count, filamentation capacity, and adhesion (〈em〉ALS1〈/em〉, 〈em〉ALS3〈/em〉, 〈em〉HWP1〈/em〉) and transcriptional regulatory gene (〈em〉BCR1〈/em〉, 〈em〉CPH1〈/em〉, 〈em〉EFG1〈/em〉) expression. In vivo studies evaluated the pathogenicity of the interaction of the microorganisms on 〈em〉Galleria mellonella〈/em〉, with analyses of the CFU per milliliter count and filamentation. In vitro results indicated that there was an observed decrease in CFU (79.4–71.5%) in multi-species biofilms. The interaction with 〈em〉S. mitis〈/em〉 inhibited filamentation, which seems to increase its virulence factor with over-expression of genes 〈em〉ALS1〈/em〉, 〈em〉ALS3〈/em〉, and 〈em〉HWP1〈/em〉 as well the interaction with 〈em〉S. sanguinis〈/em〉 as 〈em〉ALS3〈/em〉 and 〈em〉HWP1〈/em〉. 〈em〉S. mitis〈/em〉 upregulated 〈em〉BRC1〈/em〉, 〈em〉CPH1〈/em〉, and 〈em〉EFG1〈/em〉. The histological images of in vivo study indicate an increase in the filamentation of 〈em〉C. albicans〈/em〉 when in interaction with the other species. It was concluded that 〈em〉S. mitis〈/em〉 interaction suggests increased virulence factors of 〈em〉C. albicans〈/em〉, with periods of lower virulence and proto-cooperation in the interaction with 〈em〉S. sanguinis〈/em〉.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The moving volutin (polyphosphate) granules known as “dancing bodies” can be observed in the vacuoles of the yeast cells. The aim of work was to study the effects of cultivation conditions and influences of physico-chemical factors on the motion of vacuolar volutin granules in 〈em〉Saccharomyces cerevisiae〈/em〉 cells. The motion of granules is a non-Markovian process. It does not depend on the cell cycle phase, but depends on the growth stage. The maximal number of cells with “dancing bodies” was observed under cultivation of yeast at 25–28 °C and pH 5.4–5.8. Irradiation by non-ionizing electromagnetic radiation (EMR) of extremely high frequency (61.22 GHz, 100 μW, 30 min) had no effect on granule motion. After irradiation by non-ionizing EMR of very high frequency (40.68 MHz, 30 W, 30 min) the number of cells with “dancing bodies” decreased significantly and in 2 h restored almost to the control value. The possible nature of the moving volutin granules phenomenon due to metabolic processes is discussed.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉Actinomycete strain YIM PH20520, isolated from the rhizosphere soil sample of 〈em〉Panax notoginseng〈/em〉 collected in Wenshang, Yunnan Province, China, exhibited antifungal activity against root-rot pathogens of the 〈em〉Panax notoginseng〈/em〉. The structures of bioactive molecules, isolated from the ethyl acetate extract of the fermentation broth of the strain, were identified as echinosporin (1) and 7-deoxyechinosporin (2) based on extensive spectroscopic analyses. 1 exhibited antifungal activity against four tested root-rot pathogens of 〈em〉Panax notoginseng〈/em〉 include 〈em〉Fusarium oxysporum〈/em〉, 〈em〉Fusarium solani〈/em〉, 〈em〉Alternaria panax〈/em〉, and 〈em〉Phoma herbarum〈/em〉 with the MIC value at 64, 64, 32, and 64 μg/mL, respectively. 2 exhibited antifungal activities against 〈em〉F〈/em〉. 〈em〉oxysporum〈/em〉, 〈em〉F〈/em〉. 〈em〉solani〈/em〉, 〈em〉A〈/em〉. 〈em〉panax〈/em〉, and 〈em〉P〈/em〉. 〈em〉herbarum〈/em〉 with the MIC value at 128, 128, 64, and 128 μg/mL, respectively. Based on the phylogenetic analyses, the closest phylogenetic relative of strain YIM PH20520 is 〈em〉Amycolatopsis speibonae〈/em〉 JS72〈sup〉T〈/sup〉 (97.69%), so strain YIM PH20520 was identified as 〈em〉Amycolatopsis〈/em〉 strain. To the best of our knowledge, this is the first report of echinosporin antibiotics isolated from 〈em〉Amycolatopsis〈/em〉 strain besides 〈em〉Streptomyces〈/em〉 strain and their antifungal activity against four tested root-rot pathogens of the 〈em〉Panax notoginseng〈/em〉. The results provide a reliable evidence for the following related biosynthetic investigations on 〈em〉Amycolatopsis〈/em〉 strain YIM PH20520 due to echinosporins antibiotics’ unique tricyclic acetal-lactone structures.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The aim of this work was to compare production of endotoxin and to determine susceptibility to antibiotics in two groups of specimens—wild-type strains 〈em〉Ochrobactrum anthropi〈/em〉 isolated from the environment and the strains isolated from patients with cystic fibrosis. The determination of the endotoxin produced by the test strains was carried on by using a limulus amebocyte lysate test (LAL test). Determination of ATB sensitivity was accomplished by means of a broth dilution method in a microtiter plate (MIC). No significant difference was found between the group of ochrobacters isolated from the environment and the group of ochrobacters isolated from cystic fibrosis patients. Antibiotic sensitivity testing has indicated that the resistance to tigecycline, trimethoprim/sulfamethoxazole, and gentamicin was slightly higher in strains isolated from cystic fibrosis patients in comparison with strains isolated from the environment. In general, most of the test strains were sensitive to most of the antibiotics tested. Significant resistance has been demonstrated for cefotaxime. Resistance was also found for gentamicin in strains number 4 and 7. The MIC was equal to the breakpoint for this antibiotic (8000 mg/L).〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The present study aims to evaluate the diagnostic yield of bronchoalveolar lavage (BAL) fluid in patients with hematological malignancies and describe the most common pathogens detected in BAL fluid (BALF.) An analysis of 480 BALF samples was performed in patients with hematological malignancies over a period of 7 years. The results of culture methods, PCR, and immunoenzymatic sandwich microplate assays for 〈em〉Aspergillus〈/em〉 galactomannan (GM) in BALF were analyzed. Further, the diagnostic thresholds for 〈em〉Aspergillus〈/em〉 GM and 〈em〉Pneumocystis jiroveci〈/em〉 were also calculated. Microbiological findings were present in 87% of BALF samples. Possible infectious pathogens were detected in 55% of cases; 32% were classified as colonizing. No significant difference in diagnostic yield or pathogen spectrum was found between non-neutropenic and neutropenic patients. There was one significant difference in BALF findings among intensive care units (ICU) versus non-ICU patients for 〈em〉Aspergillus〈/em〉 spp. (22% versus 9%, 〈em〉p〈/em〉 = 0.03). The most common pathogens were 〈em〉Aspergillus〈/em〉 spp. (〈em〉n〈/em〉 = 86, 33% of BAL with causative pathogens) and 〈em〉Streptococcus pneumoniae〈/em〉 (〈em〉n〈/em〉 = 46, 18%); polymicrobial etiology was documented in 20% of cases. A quantitative PCR value of 〉 1860 cp/mL for 〈em〉Pneumocystis jirovecii〈/em〉 was set as a diagnostic threshold for pneumocystis pneumonia. The absorbance index of GM in BALF of 0.5 was set as a diagnostic threshold for aspergillosis. The examination of BAL fluid revealed the presence of pathogen in more than 50% of cases and is, therefore, highly useful in this regard when concerning pulmonary infiltrates.〈/p〉

Beschreibung: 〈h3〉Abstract〈/h3〉 〈p〉The family 〈em〉Bifidobacteriaceae〈/em〉 constitutes an important phylogenetic group that particularly includes bifidobacterial taxa demonstrating proven or debated positive effects on host health. The increasingly widespread application of probiotic cultures in the twenty-first century requires detailed classification to the level of particular strains. This study aimed to apply the glutamine synthetase class I (〈em〉gln〈/em〉AI) gene region (717 bp representing approximately 50% of the entire gene sequence) using specific PCR primers for the classification, typing, and phylogenetic analysis of bifidobacteria and closely related scardovial genera. In the family 〈em〉Bifidobacteriaceae〈/em〉, this is the first report on the use of this gene for such purposes. To achieve high-value results, almost all valid 〈em〉Bifidobacteriaceae〈/em〉 type strains (75) and 15 strains isolated from various environments were evaluated. The threshold value of the 〈em〉gln〈/em〉AI gene identity among 〈em〉Bifidobacterium〈/em〉 species (86.9%) was comparable to that of other phylogenetic/identification markers proposed for bifidobacteria and was much lower compared to the 16S rRNA gene. Further statistical and phylogenetic analyses suggest that the 〈em〉gln〈/em〉AI gene can be applied as a novel genetic marker in the classification, genotyping, and phylogenetic analysis of isolates belonging to the family 〈em〉Bifidobacteriaceae.〈/em〉〈/p〉