ISW1a modulates cohesin distribution in centromeric and pericentromeric regions.

Cohesin is a highly conserved, multiprotein complex whose canonical function is to hold sister chromatids together to ensure accurate chromosome segregation. Cohesin association with chromatin relies on the Scc2-Scc4 cohesin loading complex that enables cohesin ring opening and topological entrapment of sister DNAs. To better understand how sister chromatid cohesion is regulated, we performed a proteomic screen in budding yeast that identified the Isw1 chromatin remodeler as a cohesin binding partner. In addition, we found that Isw1 also interacts with Scc2-Scc4. Lack of Isw1 protein, the Ioc3 subunit of ISW1a or Isw1 chromatin remodeling activity resulted in increased accumulation of cohesin at centromeres and pericentromeres, suggesting that ISW1a may promote efficient translocation of cohesin from the centromeric site of loading to neighboring regions. Consistent with the role of ISW1a in the chromatin organization of centromeric regions, Isw1 was found to be recruited to centromeres. In its absence we observed changes in the nucleosomal landscape at centromeres and pericentromeres. Finally, we discovered that upon loss of RSC functionality, ISW1a activity leads to reduced cohesin binding and cohesion defect. Taken together, our results support the notion of a key role of chromatin remodelers in the regulation of cohesin distribution on chromosomes.

The effects of age, size, and cage complexity on the behaviour of farmed female chinchillas (Chinchilla lanigera).

Even though chinchillas have been farmed for a century, there are not many studies concerning their behaviour in captivity or their optimal housing conditions, both of which are important factors in the assessment of their welfare. This study aimed to evaluate the effect of different cage types on chinchillas' behaviour and their reactions towards humans. Female chinchillas (n = 12) were kept in three types of cages: standard with a wire floor (S); standard with a deep litter floor of shavings (SR); and enlarged with a deep litter floor of shavings (LR). Animals spent 11 weeks in each type of cage. The chinchillas' reactions toward humans were observed via intruder test. Ethograms were prepared based on round-the-clock video recordings. The activity of the chinchillas was compared, taking into account the different cage types and the animals' varying responses to the hand test. The generalized ordered logistic regression model was used to ascertain whether cage type has an effect on a chinchilla's behaviour towards humans. To compare the time distribution of various activity between chinchillas, the non-parametric Scheirer-Ray-Hare test was used. Animals kept in LR cages presented significantly less timid reactions in comparison to those kept in S and SR cages. The chinchillas spent most of their time resting (68% of the day), in locomotion (23%), and eating or drinking (8%); they spent only 1% on grooming behaviour. Cage enrichment generally reduced the fear of humans. However, the average chinchilla response to the hand test was classified in each type of cage as "cautious". Analyses of the ethograms indicated that the chinchillas were active mostly during the dark stage of the day. In conclusion, the larger cage size and its enrichment (particularly litter) reduced the fearfulness and passivity of the animals, which could be evidence of better welfare conditions.

A Simple Approach to Connecting Pt100 by Utilizing an Electroacoustic Resonance Tube.

Temperature transducers are frequently employed to keep track of process variables with different kinds of industrial controllers. One of the widely used temperature sensors is Pt100. A novel approach of utilizing an electroacoustic transducer in signal conditioning for Pt100 is proposed in this paper. A "signal conditioner" is a resonance tube filled with air, which is operated in a free resonance mode. The Pt100 wires are connected to one of the leads of the speaker in the resonance tube where the temperature changes, which is related to Pt100 resistance. The resistance affects the amplitude of the standing wave that is detected by an electrolyte microphone. An algorithm for measuring the amplitude of the speaker signal is described, as well as the building and functioning of the electroacoustic resonance tube signal conditioner. The microphone signal is acquired as a voltage using LabVIEW software. A virtual instrument (VI) developed under LabVIEW provides a measure of the voltage using standard VIs. The findings of the experiments reveal a link between the measured amplitude of the standing wave within the tube and the change in Pt100 resistance as the ambient temperature changes. Additionally, the suggested method may interface with any computer system when a sound card is added to it without the need for any extra measuring tools. The maximum nonlinearity error at full-scale deflection (FSD) is estimated at roughly 3.77%, and the experimental results and a regression model are used to assess the relative inaccuracy of the developed signal conditioner. When comparing the proposed approach with well-known approaches for Pt100 signal conditioning, the proposed one has several advantages such as its simplicity of connecting Pt100 to a personal computer directly via the sound card of any personal computer. In addition, there is no need for a reference resistance to perform a temperature measurement using such a signal conditioner.

The Comparative Assessment of Effects on the Power System and Environment of Selected Electric Transport Means in Poland.

Currently, electric vehicles are a rapidly growing alternative to those with combustion engines and can contribute to reduction of CO2 emissions in the transport sector, especially when the energy to power electric motors is predominantly derived from renewable sources. Until now, the comparison of environmental impact and influence of electric transport means on the power systems was not fully addressed in the case of Poland. The purpose of the study is to describe, analyse and assess electric vehicles (EV) operation against performance indicators in Poland, especially the influence of electric transport means (ETM) (electric cars, trams, trolley buses and buses) on power system and environment. The influence on the power system was investigated for the Polish National Powers system using the simulation of different scenarios of loads generated by EV charging. The energy demand of the National Power System and daily load variability indices were determined. Based on the data of ETM powers consumption and emissions of energy production, the emissions of harmful gases per one km and per one person were calculated, as well as the financial outlays for energy necessary to drive 1 km per 1 passenger. To assess and compare the environmental impact of the selected ETM life cycle, the life cycle assessment method was used. The results of environmental impacts were determined for selected assessment methods: CML 2 and IPCC 2013 GWP 100. The functional unit in this study is one selected ETM with a service life of 100,000 km. Comparison of trams, trolley buses, buses and electric passenger cars indicates that most beneficial are electric buses which do not need rails or overhead lines, thus investment costs are lower.

The impact of chronic co-exposure to different heavy metals on small fibers of peripheral nerves. A study of metal industry workers.

BACKGROUND: Chronic exposure to heavy metals affects various organs, among them the brain and peripheral nerves. Polyneuropathy is mainly length-dependent with predominantly sensory symptoms. There have been few studies on small fiber neuropathy due to heavy metal intoxication. METHODS: We investigated 41 metal industry workers, mean age 51.3 ± 10.5 years, with at least 5 years' professional exposure to heavy metals, and 36 age- and sex-matched healthy controls. We performed neurological examinations, and assessed blood levels of cadmium, lead, and zinc protoporphyrin, urine levels of arsenic, standard, sensory and motor electrophysiological tests in the ulnar and peroneal nerves, sympathetic skin responses from the palm and foot, and quantitative sensation testing from dermatomes C8 and S1. DISCUSSION: The results of standard conduction tests of all nerves significantly differed between groups. The latency of sympathetic skin responses achieved from the foot was also statistically significantly prolonged in the study group. Significant differences were seen in both C8 and S1 regions for temperature and pain thresholds, and for vibratory threshold only in the S1 region, while the dispersions of low and high temperatures were important exclusively in the C8 region. CONCLUSIONS: We can conclude that co-exposure to many heavy metals results in explicit impairment of peripheral nerves. The lesion is more pronounced within small fibers and is predominantly connected with greater impairment of temperature-dependent pain thresholds. The evaluation of small fiber function should be considered in the early diagnosis of toxic polyneuropathy or in low-dose exposure to heavy metals.

Silver-decorated gel-shell nanobeads: physicochemical characterization and evaluation of antibacterial properties.

We report on the synthesis of composite nanobeads with antibacterial properties. The particles consist of polystyrene cores that are surrounded by sulfonic gel shells with embedded silver nanoparticles. The nanocomposite beads are prepared by sulfonation of polystyrene particles followed by accumulation of silver ions in the shell layer and subsequent reduction with sodium borohydride. The resulting material has been characterized by electron microscopy, vibrational and X-ray photoelectron spectroscopy and several other experimental techniques. It was shown that sodium borohydride reduces silver ions embedded in the gel layer producing silver nanoparticles but also transforms a fraction of sulfonic groups in the polymer to moieties with sulfur in a lower oxidation state, likely thiols. It is hypothesized that the generated thiol groups are anchoring the nanoparticles in the gel shell of the nanobeads stabilizing the whole structure. The silver-decorated nanobeads appear to be a promising material with considerable antimicrobial activity and were tested against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Staphylococcus epidermidis. The determined minimum inhibitory (MIC) and minimum biofilm inhibitory (MBIC) concentrations are comparable to those of non-incorporated silver nanoparticles.

Application of LCA Method for Assessment of Environmental Impacts of a Polylactide (PLA) Bottle Shaping.

In recent years, there has been a significant increase in the consumption of single-use packaging. Their material diversity is a significant barrier to recycling, causing overloading of landfills. Increasing negative environmental aspects have highlighted the need to develop solutions to achieve a relatively high efficiency of the bottle shaping process with the lowest possible energy consumption. The aim of the project is to try to describe the impact of this process on the state, transformation and development of the natural environment. The work concerns current issues of the impact of packaging on the natural environment. The main goal was to conduct a life cycle analysis (LCA) of beverage bottles made of polylactide. The functional unit comprised a total of 1,000 pieces of PLA bottles with a capacity of 1 L. The boundary of the adopted system included the steps from the delivery of the preforms to the production plant to their correct formation in the process of forming beverage bottles. Further stages of the production process were excluded from the system, such as beverage bottling, labeling, and storage and distribution. Processes related to transport and storage of raw material were also excluded. The LCA analysis was performed using the program of the Dutch company Pre Consultants called SimaPro 8.4.0. The ReCiPe 2016 method was chosen for the interpretation of the quantity of emitted substances into the natural environment. The test results were presented graphically on bar charts and subjected to verification and interpretation.

The Effect of Silver Nanoparticles on Listeria monocytogenes PCM2191 Peptidoglycan Metabolism and Cell Permeability.

Listeria monocytogenes is Gram-positive bacterial pathogen, a causative agent of food poisoning and systemic disease - listeriosis. This species is still susceptible to several conventionally used antibiotics but an increase in its resistance has been reported. For this reason the search for new, alternative therapies is an urgent task. Silver nanoparticles seem to be the promising antibacterial agent. Minimal inhibitory concentration of silver nanoparticles was determined. Sublethal concentrations were used in study of nanosilver effect on cells lysis by estimation of the number of cells surviving the treatment with 0.25 or 0.5 of minimal inhibitory concentrations of silver nanoparticles. Autolysis of isolated peptidoglycan was studied by measuring the absorbance of preparation subjected to nanosilver treatment. Silver nanoparticles effect on L. monocytogenes envelopes permeability was determined by measuring the efflux of cF, DNA and proteins. It was demonstrated that nanosilver enhanced the lysis of L. monocytogenes cells and, to the lesser extent, autolysis of isolated peptidoglycan. The increase in the efflux of carboxyfluoresceine, DNA and proteins was also noted. The obtained results allow to postulate that L. monocytogenes peptidoglycan, constituting the main component of cell wall, is the target of silver nanoparticles activity against this pathogen.

Transmembrane topology of the arsenite permease Acr3 from Saccharomyces cerevisiae.

Acr3 is a plasma membrane transporter, a member of the bile/arsenite/riboflavin transporter (BART) superfamily, which confers high-level resistance to arsenicals in the yeast Saccharomyces cerevisiae. We have previously shown that the yeast Acr3 acts as a low affinity As(III)/H+ and Sb(III)/H+ antiporter. We have also identified several amino acid residues that are localized in putative transmembrane helices (TM) and appeared to be critical for the Acr3 activity. In the present study, the topology of Acr3 was investigated by insertion of glycosylation and factor Xa protease cleavage sites at predicted hydrophilic regions. The analysis of the glycosylation pattern and factor Xa cleavage products of resulting Acr3 fusion constructs provide evidence supporting a topological model of Acr3 with 10 TM segments and cytoplasmically oriented N- and C-terminal domains. Next, we investigated the role of the hydrophilic loop connecting TM8 and TM9, the large size of which is unique to members of the yeast Acr3 family of metalloid transporters. We found that a 28 amino acid deletion in this region does not affect Acr3 folding, trafficking substrate binding, or transport activity. Finally, we constructed a homology-based structural model of Acr3 using the crystal structure of the Yersinia frederiksenii homologue of the human bile acid sodium symporter ASBT.

Genetic control of bacterial biofilms.

Nearly all bacterial species, including pathogens, have the ability to form biofilms. Biofilms are defined as structured ecosystems in which microbes are attached to surfaces and embedded in a matrix composed of polysaccharides, eDNA, and proteins, and their development is a multistep process. Bacterial biofilms constitute a large medical problem due to their extremely high resistance to various types of therapeutics, including conventional antibiotics. Several environmental and genetic signals control every step of biofilm development and dispersal. From among the latter, quorum sensing, cyclic diguanosine-5'-monophosphate, and small RNAs are considered as the main regulators. The present review describes the control role of these three regulators in the life cycles of biofilms built by Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella enterica serovar Typhimurium, and Vibrio cholerae. The interconnections between their activities are shown. Compounds and strategies which target the activity of these regulators, mainly quorum sensing inhibitors, and their potential role in therapy are also assessed.

Interactions of Escherichia coli molecular chaperone HtpG with DnaA replication initiator DNA.

The bacterial chaperone high-temperature protein G (HtpG), a member of the Hsp90 protein family, is involved in the protection of cells against a variety of environmental stresses. The ability of HtpG to form complexes with other bacterial proteins, especially those involved in fundamental functions, is indicative of its cellular role. An interaction between HtpG and DnaA, the main initiator of DNA replication, was studied both in vivo, using a bacterial two-hybrid system, and in vitro with a modified pull-down assay and by chemical cross-linking. In vivo, this interaction was demonstrated only when htpG was expressed from a high copy number plasmid. Both in vitro assays confirmed HtpG-DnaA interactions.

Identification of critical residues for transport activity of Acr3p, the Saccharomyces cerevisiae†As(III)/H+ antiporter.

Acr3p is an As(III)/H(+) antiporter from Saccharomyces cerevisiae belonging to the bile/arsenite/riboflavin transporter superfamily. We have previously found that Cys151 located in the middle of the fourth transmembrane segment (TM4) is critical for antiport activity, suggesting that As(III) might interact with a thiol group during the translocation process. In order to identify functionally important residues involved in As(III)/H(+) exchange, we performed a systematic alanine-replacement analysis of charged/polar and aromatic residues that are conserved in the Acr3 family and located in putative transmembrane segments. Nine residues (Asn117, Trp130, Arg150, Trp158, Asn176, Arg230, Tyr290, Phe345, Asn351) were found to be critical for proper folding and trafficking of Acr3p to the plasma membrane. In addition, we found that replacement of highly conserved Phe266 (TM7), Phe352 (TM9), Glu353 (TM9) and Glu380 (TM10) with Ala abolished transport activity of Acr3p, while mutation of Ser349 (TM9) to Ala significantly reduced the As(III)/H(+) exchange, suggesting an important role of these residues in the transport mechanism. Detailed mutational analysis of Glu353 and Glu380 revealed that the negatively charged residues located in the middle of transmembrane segments TM9 and TM10 are crucial for antiport activity. We also discuss a hypothetical model of the Acr3p transport mechanism.

The effect of oleanolic and ursolic acids on the hemolytic properties and biofilm formation of Listeria monocytogenes.

Oleanolic acid and ursolic acid are pentacyclic triterpenoids isolated from a variety of medicinal plants, which have antibacterial activity. Listeria monocytogenes is a Gram-positive facultative pathogen, being the causative agent of listeriosis. The present study was carried out to evaluate the in vitro effect of sub-inhibitory concentrations of both triterpene acids on the pathogenicity determinants of L. monocytogenes: their hemolytic activity and biofilm forming ability. Oleanolic and ursolic acids inhibited listeriolysin O activity without influencing toxin secretion. Biofilm formation, and the viability of L. monocytogenes cells in biofilms was diminished by both compounds. Thus, both acids affected L. monocytogenes virulence. It was also demonstrated that oleanolic acid bound to the peptidoglycan of L. monocytogenes and this interaction was influenced by teichoic acids.

Molecular analysis of vanA outbreak of Enterococcus faecium in two Warsaw hospitals: the importance of mobile genetic elements.

Vancomycin-resistant Enterococcus faecium represents a growing threat in hospital-acquired infections. Two outbreaks of this pathogen from neighboring Warsaw hospitals have been analyzed in this study. Pulsed-field gel electrophoresis (PFGE) of SmaI-digested DNA, multilocus VNTR analysis (MLVA), and multilocus sequence typing (MLST) revealed a clonal variability of isolates which belonged to three main lineages (17, 18, and 78) of nosocomial E. faecium. All isolates were multidrug resistant and carried several resistance, virulence, and plasmid-specific genes. Almost all isolates shared the same variant of Tn1546 transposon, characterized by the presence of insertion sequence ISEf1 and a point mutation in the vanA gene. In the majority of cases, this transposon was located on 50 kb or 100 kb pRUM-related plasmids, which lacked, however, the axe-txe toxin-antitoxin genes. 100 kb plasmid was easily transferred by conjugation and was found in various clonal backgrounds in both institutions, while 50 kb plasmid was not transferable and occurred solely in MT159/ST78 strains that disseminated clonally in one institution. Although molecular data indicated the spread of VRE between two institutions or a potential common source of this alert pathogen, epidemiological investigations did not reveal the possible route by which outbreak strains disseminated.

Modulation of antibiotic resistance and induction of a stress response in Pseudomonas aeruginosa by silver nanoparticles.

The objective of this study was to characterize the effects of silver nanoparticles on Pseudomonas aeruginosa. Their interactions with several conventional antibiotics and ability to induce a stress response were examined. Interactions between silver nanoparticles (AgNPs) and antibiotics against free-living cells and biofilm of P. aeruginosa were studied using the chequerboard method and time-kill assays. The ability of AgNPs to induce a stress response was determined by evaluation of cellular levels of the DnaK and HtpG chaperones using SDS-PAGE and Western blot analysis. Synergistic activity against free-living P. aeruginosa between AgNPs and ampicillin, streptomycin, rifampicin and tetracycline, but not oxacillin, ciprofloxacin, meropenem or ceftazidime, was demonstrated by the chequerboard method. No such interactions were observed against P. aeruginosa biofilm. The results of time-kill assays confirmed synergy only for the AgNPs-streptomycin combination. AgNPs induced the expression of chaperone DnaK. No induction of the HtpG chaperone was detected. In conclusion, AgNPs not only display potent bactericidal activity against P. aeruginosa, but also act synergistically with several conventional antibiotics to enhance their effect against free-living bacteria as determined by the chequerboard method. The time-kill assay proved synergy between AgNPs and streptomycin only. The ability of AgNPs to induce the major chaperone protein DnaK may influence bacterial resistance to antimicrobials.

Multiple cysteine residues are necessary for sorting and transport activity of the arsenite permease Acr3p from Saccharomyces cerevisiae.

The yeast transporter Acr3p is a low affinity As(III)/H(+) and Sb(III)/H(+) antiporter located in the plasma membrane. It has been shown for bacterial Acr3 proteins that just a single cysteine residue, which is located in the middle of the fourth transmembrane region and conserved in all members of the Acr3 family, is essential for As(III) transport activity. Here, we report a systematic mutational analysis of all nine cysteine residues present in the Saccharomyces cerevisiae Acr3p. We found that mutagenesis of highly conserved Cys151 resulted in a complete loss of metalloid transport function. In addition, lack of Cys90 and Cys169, which are conserved in eukaryotic members of Acr3 family, impaired Acr3p trafficking to the plasma membrane and greatly reduced As(III) efflux, respectively. Mutagenesis of five other cysteines in Acr3p resulted in moderate reduction of As(III) transport capacities and sorting perturbations. Our data suggest that interaction of As(III) with multiple thiol groups in the yeast Acr3p may facilitate As(III) translocation across the plasma membrane.

Silver nanoparticles as an alternative strategy against bacterial biofilms.

Biofilms are complex bacterial communities that resist the action of antibiotics and the human immune system. Bacteria within biofilms are the cause of numerous, almost impossible to eradicate, persistent infections. Biofilms can form on many medical devices and implants, and so have an enormous impact on medicine. Due to the lack of effective anti-biofilm antibiotics, novel alternative compounds or strategies are urgently required. This review describes some of the latest approaches in the field of biofilm treatment. New anti-biofilm technologies target different stages in the biofilm formation process. Some act to modify the colonized biomaterials to make them resistant to biofilm formation. One potentially important candidate treatment uses silver nanoparticles that show anti-bacterial and anti-biofilm activity. The biological action of nano-silver is complex and seems to involve a number of pathways. However, there have been few reports on the anti-biofilm activity of silver nanoparticles and the precise mechanism underlying their action remains unresolved. Here, we describe some anti-biofilm approaches employing AgNPs and consider the challenges and problems that need to be addressed in order to make silver nanoparticles a part of an effective anti-biofilm strategy.