Analysis of Whole-Genome Sequences of Pathogenic Gram-Positive and Gram-Negative Isolates from the Same Hospital Environment to Investigate Common Evolutionary Trends Associated with Horizontal Gene Exchange, Mutations and DNA Methylation Patterning.

Hospital-acquired infections are a generally recognized problem for healthcare professionals. Clinical variants of Gram-negative and Gram-positive pathogens are characterized with enhanced antibiotic resistance and virulence due to mutations and the horizontal acquisition of respective genetic determinants. In this study, two Escherichia coli, two Klebsiella pneumoniae, three Pseudomonas aeruginosa, two Staphylococcus aureus, one Staphylococcus epidermidis and one Streptococcus pneumoniae showing broad spectra of antibiotic resistance were isolated from patients suffering from nosocomial infections in a local hospital in Almaty, Kazakhstan. The aim of the study was to compare general and species-specific pathways of the development of virulence and antibiotic resistance through opportunistic pathogens causing hospital-acquired infections. The whole-genome PacBio sequencing of the isolates allowed for the genotyping and identification of antibiotic resistance and virulence genetic determinants located in the chromosomes, plasmids and genomic islands. It was concluded that long-read sequencing is a useful tool for monitoring the epidemiological situation in hospitals. Marker antibiotic resistance mutations common for different microorganisms were identified, which were acquired due to antibiotic-selective pressure in the same clinical environment. The genotyping and identification of strain-specific DNA methylation motifs were found to be promising in estimating the risks associated with hospital infection outbreaks and monitoring the distribution and evolution of nosocomial pathogens.

Effects of Traumatic Brain Injury on the Gut Microbiota Composition and Serum Amino Acid Profile in Rats.

Traumatic brain injury (TBI) heavily impacts the body: it damages the brain tissue and the peripheral nervous system and shifts homeostasis in many types of tissue. An acute brain injury compromises the "brain-gut-microbiome axis", a well-balanced network formed by the brain, gastrointestinal tract, and gut microbiome, which has a complex effect: damage to the brain alters the composition of the microbiome; the altered microbiome affects TBI severity, neuroplasticity, and metabolic pathways through various bacterial metabolites. We modeled TBI in rats. Using a bioinformatics approach, we sought to identify correlations between the gut microbiome composition, TBI severity, the rate of neurological function recovery, and blood metabolome. We found that the TBI caused changes in the abundance of 26 bacterial genera. The most dramatic change was observed in the abundance of Agathobacter species. The TBI also altered concentrations of several metabolites, specifically citrulline and tryptophan. We found no significant correlations between TBI severity and the pre-existing gut microbiota composition or blood metabolites. However, we discovered some differences between the two groups of subjects that showed high and low rates of neurological function recovery, respectively. The present study highlights the role of the brain-gut-microbiome axis in TBI.

Insights into herpesvirus assembly from the structure of the pUL7:pUL51 complex.

Herpesviruses acquire their membrane envelopes in the cytoplasm of infected cells via a molecular mechanism that remains unclear. Herpes simplex virus (HSV)-1 proteins pUL7 and pUL51 form a complex required for efficient virus envelopment. We show that interaction between homologues of pUL7 and pUL51 is conserved across human herpesviruses, as is their association with trans-Golgi membranes. We characterized the HSV-1 pUL7:pUL51 complex by solution scattering and chemical crosslinking, revealing a 1:2 complex that can form higher-order oligomers in solution, and we solved the crystal structure of the core pUL7:pUL51 heterodimer. While pUL7 adopts a previously-unseen compact fold, the helix-turn-helix conformation of pUL51 resembles the cellular endosomal complex required for transport (ESCRT)-III component CHMP4B and pUL51 forms ESCRT-III-like filaments, suggesting a direct role for pUL51 in promoting membrane scission during virus assembly. Our results provide a structural framework for understanding the role of the conserved pUL7:pUL51 complex in herpesvirus assembly.

Most people suffer from occasional cold sores, which are caused by the herpes simplex virus. This virus causes infections that last your entire life, but for the most part it lies dormant in your cells and reactivates only at times of stress. When it reactivates, the virus manipulates host cells to make new virus particles that may spread the infection to other people. Like many other viruses, herpes simplex viruses also steal jelly-like structures known as membranes from their host cells to form protective coats around new virus particles. In cells from humans and other animals, proteins belonging to a molecular machine known as ESCRT form filaments that bend and break membranes as the cells require. Many viruses hijack the ESCRT machinery to wrap membranes around new virus particles. However, herpes simplex viruses do not follow the usual rules for activating this machine. Instead, they rely on two viral proteins called pUL7 and pUL51 to hot-wire the ESCRT machinery. Previous studies have shown that these two proteins bind to each other, but it remained unclear how they work. Butt et al. used a combination of biochemical and biophysical techniques to solve the three-dimensional structures of pUL7 and pUL51 when bound to each other. The experiments determined that the structure of pUL51 resembles the structures of different components in the ESCRT machinery. Like the ESCRT proteins, pUL51 formed filaments, suggesting that pUL51 bends membranes in cells and that pUL7 blocks it from doing so until the time is right. Further experiments showed that the equivalents of pUL7 and pUL51 in other members of the herpes virus family also bind to each other in a similar way. These findings reveal that herpes simplex viruses and their close relatives have evolved a different strategy than many other viruses to steal membranes from host cells. Interfering with this mechanism may provide new avenues for designing drugs or improving vaccines against these viruses. The pUL7 and pUL51 proteins may also inspire new tools in biotechnology that could precisely control the shapes of biological membranes.

Inner tegument proteins of Herpes Simplex Virus are sufficient for intracellular capsid motility in neurons but not for axonal targeting.

Upon reactivation from latency and during lytic infections in neurons, alphaherpesviruses assemble cytosolic capsids, capsids associated with enveloping membranes, and transport vesicles harboring fully enveloped capsids. It is debated whether capsid envelopment of herpes simplex virus (HSV) is completed in the soma prior to axonal targeting or later, and whether the mechanisms are the same in neurons derived from embryos or from adult hosts. We used HSV mutants impaired in capsid envelopment to test whether the inner tegument proteins pUL36 or pUL37 necessary for microtubule-mediated capsid transport were sufficient for axonal capsid targeting in neurons derived from the dorsal root ganglia of adult mice. Such neurons were infected with HSV1-ΔUL20 whose capsids recruited pUL36 and pUL37, with HSV1-ΔUL37 whose capsids associate only with pUL36, or with HSV1-ΔUL36 that assembles capsids lacking both proteins. While capsids of HSV1-ΔUL20 were actively transported along microtubules in epithelial cells and in the somata of neurons, those of HSV1-ΔUL36 and -ΔUL37 could only diffuse in the cytoplasm. Employing a novel image analysis algorithm to quantify capsid targeting to axons, we show that only a few capsids of HSV1-ΔUL20 entered axons, while vesicles transporting gD utilized axonal transport efficiently and independently of pUL36, pUL37, or pUL20. Our data indicate that capsid motility in the somata of neurons mediated by pUL36 and pUL37 does not suffice for targeting capsids to axons, and suggest that capsid envelopment needs to be completed in the soma prior to targeting of herpes simplex virus to the axons, and to spreading from neurons to neighboring cells.

Dual Function of the pUL7-pUL51 Tegument Protein Complex in Herpes Simplex Virus 1 Infection.

The tegument of herpesviruses is a highly complex structural layer between the nucleocapsid and the envelope of virions. Tegument proteins play both structural and regulatory functions during replication and spread, but the interactions and functions of many of these proteins are poorly understood. Here we focus on two tegument proteins from herpes simplex virus 1 (HSV-1), pUL7 and pUL51, which have homologues in all other herpesviruses. We have now identified that HSV-1 pUL7 and pUL51 form a stable and direct protein-protein interaction, their expression levels rely on the presence of each other, and they function as a complex in infected cells. We demonstrate that expression of the pUL7-pUL51 complex is important for efficient HSV-1 assembly and plaque formation. Furthermore, we also discovered that the pUL7-pUL51 complex localizes to focal adhesions at the plasma membrane in both infected cells and in the absence of other viral proteins. The expression of pUL7-pUL51 is important to stabilize focal adhesions and maintain cell morphology in infected cells and cells infected with viruses lacking pUL7 and/or pUL51 round up more rapidly than cells infected with wild-type HSV-1. Our data suggest that, in addition to the previously reported functions in virus assembly and spread for pUL51, the pUL7-pUL51 complex is important for maintaining the attachment of infected cells to their surroundings through modulating the activity of focal adhesion complexes. IMPORTANCE: Herpesviridae is a large family of highly successful human and animal pathogens. Virions of these viruses are composed of many different proteins, most of which are contained within the tegument, a complex structural layer between the nucleocapsid and the envelope within virus particles. Tegument proteins have important roles in assembling virus particles as well as modifying host cells to promote virus replication and spread. However, little is known about the function of many tegument proteins during virus replication. Our study focuses on two tegument proteins from herpes simplex virus 1 that are conserved in all herpesviruses: pUL7 and pUL51. We demonstrate that these proteins directly interact and form a functional complex that is important for both virus assembly and modulation of host cell morphology. Further, we identify for the first time that these conserved herpesvirus tegument proteins localize to focal adhesions in addition to cytoplasmic juxtanuclear membranes within infected cells.

Conserved Tryptophan Motifs in the Large Tegument Protein pUL36 Are Required for Efficient Secondary Envelopment of Herpes Simplex Virus Capsids.

UNLABELLED: Herpes simplex virus (HSV) replicates in the skin and mucous membranes, and initiates lytic or latent infections in sensory neurons. Assembly of progeny virions depends on the essential large tegument protein pUL36 of 3,164 amino acid residues that links the capsids to the tegument proteins pUL37 and VP16. Of the 32 tryptophans of HSV-1-pUL36, the tryptophan-acidic motifs (1766)WD(1767) and (1862)WE(1863) are conserved in all HSV-1 and HSV-2 isolates. Here, we characterized the role of these motifs in the HSV life cycle since the rare tryptophans often have unique roles in protein function due to their large hydrophobic surface. The infectivity of the mutants HSV-1(17(+))Lox-pUL36-WD/AA-WE/AA and HSV-1(17(+))Lox-CheVP26-pUL36-WD/AA-WE/AA, in which the capsid has been tagged with the fluorescent protein Cherry, was significantly reduced. Quantitative electron microscopy shows that there were a larger number of cytosolic capsids and fewer enveloped virions compared to their respective parental strains, indicating a severe impairment in secondary capsid envelopment. The capsids of the mutant viruses accumulated in the perinuclear region around the microtubule-organizing center and were not dispersed to the cell periphery but still acquired the inner tegument proteins pUL36 and pUL37. Furthermore, cytoplasmic capsids colocalized with tegument protein VP16 and, to some extent, with tegument protein VP22 but not with the envelope glycoprotein gD. These results indicate that the unique conserved tryptophan-acidic motifs in the central region of pUL36 are required for efficient targeting of progeny capsids to the membranes of secondary capsid envelopment and for efficient virion assembly. IMPORTANCE: Herpesvirus infections give rise to severe animal and human diseases, especially in young, immunocompromised, and elderly individuals. The structural hallmark of herpesvirus virions is the tegument, which contains evolutionarily conserved proteins that are essential for several stages of the herpesvirus life cycle. Here we characterized two conserved tryptophan-acidic motifs in the central region of the large tegument protein pUL36 of herpes simplex virus. When we mutated these motifs, secondary envelopment of cytosolic capsids and the production of infectious particles were severely impaired. Our data suggest that pUL36 and its homologs in other herpesviruses, and in particular such tryptophan-acidic motifs, could provide attractive targets for the development of novel drugs to prevent herpesvirus assembly and spread.

Effect of vasopressin on the expression of genes for key enzymes of hyaluronan turnover in Wistar Albino Glaxo and Brattleboro rat kidneys.

Hyaluronan (HA), the major glycosaminoglycan of the interstitial matrix, is heterogeneously distributed within the kidney. Using real-time RT-PCR, we tested the assumption that renal HA may be involved in the long-term effect of vasopressin on water reabsorption. The expression of the genes encoding hyaluronan synthase-2 (Has2), hyaluronidase-1 and hyaluronidase-2 (Hyal1 and Hyal2) was studied in the kidneys of Wistar Albino Glaxo (WAG) and homozygous vasopressin-deficient Brattleboro rats treated with the V2 receptor-selective vasopressin analogue dDAVP (100 µg (kg body wt)(-1), i.p., twice a day for 2 days). The Has2 mRNA content was the highest in the kidney papilla of the hydrated WAG and control Brattleboro rats, devoid of vasopressin. In WAG rats, dDAVP induced a considerable decrease in Has2 mRNA content in the papilla, with less pronounced changes in the cortex. The changes elicited by dDAVP in Brattleboro rats tended to be the same as in WAG rats, but weaker. In contrast to Has2, dDAVP treatment caused a significant increase in the Hyal1 and Hyal2 mRNA content in the renal papilla of WAG and Brattleboro rats. In rats of both strains, there was a good fit between Hyal1 and Hyal2 transcriptional levels and changes in hyaluronidase activity in the renal tissue. It is suggested that vasopressin is able to inhibit the synthesis of HA and concomitantly promote its degradation in the interstitium of the renal papilla, thereby facilitating water flow between elements of the renal countercurrent system. The implications for this effect are discussed in the context of the data in the literature.

Comparison of propagation of atrial excitation with the cardiopotential distribution on the body surface of hypertensive rats.

OBJECTIVE: Arterial hypertension is associated with the risk of developing atrial arrhythmia. This research was aimed at studying the sequence of depolarization along the atrial epicardium and formation of the cardioelectric field on the body surface of hypertensive rats. METHODS: The study was carried out on eleven ISIAH rats (with hereditary stress-induced hypertension). We analyzed spatial-temporal characteristics of body surface potential map (BSPM), time characteristics of electrocardiogram during atrial depolarization, sequence of atrial epicardial depolarization of rats. Statistical analysis was performed using independent samples t-test. RESULTS: The results indicated that in 27% of hypertensive rats in the pulmonary vein (PV) sleeves, early excitation areas are formed 2.0 ± 0.5 ms after the beginning of depolarization of the sino-atrial node area, in 73% of animals, the area of the PV sleeves is excited 6.5 ± 0.4 ms after the beginning of depolarization. In experimental animals, the beginning of inversion of areas of positive and negative cardioelectric potentials on BSPM does not differ. In 73% of rats, duration of inversion on BSPM was 7.7 ± 1.9 ms, and in 27% - 3.9 ± 0.1 ms (p=0.011). The formation of early activation zones in PV of rats with arterial hypertension testifies to possible wandering focus in the myocardium of PV sleeves. Stress-induced hypertension results in actual risk of atrial arrhythmias, which originate at the base of the PV. CONCLUSION: In rats with arterial hypertension, two early depolarization zones are revealed in the sinus node area and in the PV return to the left atrium, projected on BSPM by mutual positions of negative and positive potential zones.

Biochemical and kinetic analysis of the GH3 family beta-xylosidase from Aspergillus awamori X-100.

The beta-xylosidase from Aspergillus awamori X-100 belonging to the family 3 glycoside hydrolase revealed a distinctive transglycosylating ability to produce xylooligosaccharides with degree of polymerization more than 7. In order to explain this fact, the enzyme has been subjected to the detailed biochemical study. The enzymatic hydrolysis of p-nitrophenyl beta-D-xylopyranoside was found to occur with overall retention of substrate anomeric configuration suggesting cleavage of xylosidic bonds through a double-displacement mechanism. Kinetic study with aryl beta-xylopyranosides substrates, in which leaving group pK(a)s were in the range of 3.96-10.32, revealed monotonic function of log(k(cat)) and no correlation of log(k(cat)/Km) versus pKa values indicating deglycosylation as a rate-limiting step for the enzymatic hydrolysis. The classical bell-shaped pH dependence of k(cat)/Km indicated two ionizable groups in the beta-xylosidase active site with apparent pKa values of 2.2 and 6.4. The kinetic parameters of hydrolysis, Km and k(cat), of p-nitrophenyl beta-D-1,4-xylooligosaccharides were very close to those for hydrolysis of p-nitrophenyl-beta-D-xylopyranoside. Increase of p-nitrophenyl-beta-D-xylopyranoside concentration up to 80 mM led to increasing of the reaction velocity resulting in k(cat)(app)=81 s(-1). Addition of alpha-methyl D-xylopyranoside to the reaction mixture at high concentration of p-nitrophenyl-beta-D-xylopyranoside (50 mM) caused an acceleration of the beta-xylosidase-catalyzed reactions and appearance of a new transglycosylation product, alpha-methyl D-xylopyranosyl-1,4-beta-D-xylopyranoside, that was identified by 1H NMR spectroscopy. The kinetic model suggested for the enzymatic reaction was consistent with the results obtained.

From management to mediation: local forestry management and the forestry crisis in post-socialist Russia.

The local branches of the Russian Forestry Service, the leskhozy, were known for their efficiency and management skills in the Soviet era and were one of the very few community-based "Soviet-type" institutions to survive the transition. This article examines the role of the leskhozy in the new market economy. Our analysis is based on data from interviews with informants attached to the forestry sector in the Murmansk area. In some cases their knowledge of the leskhozy stretches back to the emergence of the system in 1947. Our principal finding is that the struggle to survive as a federal body in the current legal and economic climate is forcing the leskhozy to relegate sustainable forestry management, presumably their primary raison d'être, to the lower portions of their list of priorities. Several consequences result. There is a heightened incidence of illegal logging, and corruption informs the allocation of forest areas to private interests. Stumpage prices have plummeted as timber from subsidized commercial cutting (ostensibly sanitary cutting or thinning) has flooded the markets. The root cause of these tribulations lies with the market-based harvesting permit system. Its introduction in the 1990s did little to eliminate the self-seeking practices of the old Soviet forestry management hierarchies. In the free market, local forestry managers can turn their dual responsibilities to their own advantage inasmuch as they control the allocation of harvesting permits while at the same time controlling logging practices.

Dermatitis de contacto por Zovirax@ crema / Contact dermatitis from Zovirax cream

No disponible

Acné necrótico (varioliforme) / Acne necrotica (varioliformis)

Se presenta un paciente varón de 44 años que consultó por papulopústulas pruriginosas localizadas en cuero cabelludo, cara y cuello con tendencia a formar costras hemorrágicas y cicatrices deprimidas. Histológicamente se apreció un infiltrado de predominio linfocitario con tendencia a la necrosis del folículo y con signos de exocitosis folicular linfocitaria. Por los datos clínicos e histológicos se realizó el diagnóstico de acné necrótico. Fue tratado con diversos antibióticos, mostrando remisiones y recidivas frecuentes. Tras 6 meses de tratamiento con isotretinoína oral se obtuvo una remisión de casi 2 años. El acné necrótico es una foliculitis necrosante crónica en la que se distinguen dos formas clínicas, una superficial o miliar y otra profunda con tendencia a la formación de cicatrices o varioliforme (AU)