Studija znakova patogenosti kod Pasteurella multocida, izolirane iz životinja različitih vrsta

A significant number of microorganisms in natural and artificial environments exist in a structured formation – biofilm. This formation attaches to a certain surface, particularly the epithelium. The ability to form a similar structure has been observed in Pasteurella multocida, the causative agent of anthropozoonoses that affect domestic and wild animals, birds, companion animals and humans. The spectrum of pathogenetic action of P. multocida is wide and associated with the development of respiratory and multisystemic pathology, bacteraemia and other manifestations. Timely detection of P. multocida and treatment of the diseases it causes in farm and domestic animals is important to limit economic losses and improve social security. The main objective of this study was to determine the pathogenicity of P. multocida, its ability to form a biofilm, its resistance to antibiotics, and to identify the genes responsible for the formation of dermonecrotic toxin and biofilm formation. The paper presents the results of a study of 11 isolates of P. multocida: six isolates (54.5%) from rabbits, two isolates (18.2%) from dogs, two isolates (18.2%) from cats, and one isolate from pigs (9.2%). In all isolates, the gene ptfA was detected. This gene encodes the formation of type 4 fimbriae and participates in the formation of the biofilm, and the studied cultures in vitro formed a biofilm of different densities. The genome of eight isolates (72.7%) included the toxA gene (provides the formation of dermonecrotic toxin), while 45.4% of isolates had a complete set of the studied signs of pathogenicity, both in phenotypic (biofilm formation, mortality for laboratory animals) and genotypic (presence of toxA, ptfA) traits, and three isolates (27.3%) showed signs of multidrug resistance. The virulence of the toxA-negative isolates of P. multocida was lower than in toxA-positive isolates. The culture with the highest virulence (0.5x 101 CFU) and extreme resistance to antibiotics formed a biofilm of the highest density. The association of the gene in the biofilm-producing mechanism needs further evaluation, and further research is needed to identify the relationships between pathogens in Pasteurella multocida isolates from different species of animals and humans.Znatan broj mikroorganizama u prirodnim i umjetnim okruženjima postoji u obliku strukturirane formacije – biofilma i ta se formacija može vezati i na određenu površinu, posebice na epitel. Mogućnost formiranja slične strukture zamijećena je kod Pasteurella multocida, koja je uzročnik antropozoonoza, a pogađa i ljude, kućne ljubimce, ptice i domaće i divlje životinje. Spektar patogenih učinaka P. multocida prilično je širok i povezan je s razvojem respiratorne i multisistemske bolesti, bakterijemije i drugih manifestacija. Pravovremeno otkrivanje P. multocida i liječenje bolesti koje ova bakterija prouzroči u životinja na farmi i domaćih životinja važno je za ograničenje ekonomskih gubitaka i sigurnost društva. Glavni je cilj ove studije bio utvrditi patogenost P. multocida, sposobnost formiranja biofilma, otpornost na antibiotike, ali i identificirati gene odgovorne za formiranje dermonekrotičnog toksina, odnosno formiranje biofilma. Ovaj rad predstavlja rezultate studije 11 izolata P. multocida: 6 izolata (54,5 %) izoliranih iz zečeva, 2 (18,2 %) iz pasa, 2 (18,2 %) iz mačaka te 1 izolat izoliran iz svinja (9.2 %). U 100 % proučavanih izolata, otkriven je gen (ptfA) koji kodira formiranje fimbrija tipa 4 i sudjeluje u formiranju biofilma. Ispitane su kulture in vitro formirale biofilm različitih gustoća. U genomu 8 ispitanih izolata (72,7 %) otkrivena je prisutnost toxA gena (koji omogućuje formiranje dermonekrotičnog toksina). 45,4 % ispitanih izolata pokazalo je cijeli set proučavanih znakova patogenosti – fenotispkih (formiranje biofilma, smrtnost za laboratorijske životinje) karakteristika i genotipskih (prisutnost toxA, ptfA) karakteristika. Tri izolata (27,3 %) pokazala su otpornost na više lijekova. Otkriveno je da je kod toxA-negativnih izolata P. multocida virulencija bila niža u usporedbi s toxA-pozitivnim izolatima. Kultura s najvišom virulentnošću (0,5 x 101 CFU) i ekstremnom otpornošću na antibiotike formirala je biofilm najveće gustoće. Asocijacija gena u mehanizmu proizvodnje biofilma zahtijeva dodatnu procjenu, a potrebno je i dodatno istraživanje za identifikaciju odnosa između patogena među izolatima P. multocida izoliranima iz ljudi i različitih vrsta životinja

Structural Analysis of the Ancestral Haloalkane Dehalogenase AncLinB-DmbA

Haloalkane dehalogenases (EC 3.8.1.5) play an important role in hydrolytic degradation of halogenated compounds, resulting in a halide ion, a proton, and an alcohol. They are used in biocatalysis, bioremediation, and biosensing of environmental pollutants and also for molecular tagging in cell biology. The method of ancestral sequence reconstruction leads to prediction of sequences of ancestral enzymes allowing their experimental characterization. Based on the sequences of modern haloalkane dehalogenases from the subfamily II, the most common ancestor of thoroughly characterized enzymes LinB from Sphingobium japonicum UT26 and DmbA from Mycobacterium bovis 5033/66 was in silico predicted, recombinantly produced and structurally characterized. The ancestral enzyme AncLinB-DmbA was crystallized using the sitting-drop vapor-diffusion method, yielding rod-like crystals that diffracted X-rays to 1.5 & ANGS; resolution. Structural comparison of AncLinB-DmbA with their closely related descendants LinB and DmbA revealed some differences in overall structure and tunnel architecture. Newly prepared AncLinB-DmbA has the highest active site cavity volume and the biggest entrance radius on the main tunnel in comparison to descendant enzymes. Ancestral sequence reconstruction is a powerful technique to study molecular evolution and design robust proteins for enzyme technologies

Sex difference and intra-operative tidal volume: Insights from the LAS VEGAS study

BACKGROUND: One key element of lung-protective ventilation is the use of a low tidal volume (VT). A sex difference in use of low tidal volume ventilation (LTVV) has been described in critically ill ICU patients.OBJECTIVES: The aim of this study was to determine whether a sex difference in use of LTVV also exists in operating room patients, and if present what factors drive this difference.DESIGN, PATIENTS AND SETTING: This is a posthoc analysis of LAS VEGAS, a 1-week worldwide observational study in adults requiring intra-operative ventilation during general anaesthesia for surgery in 146 hospitals in 29 countries.MAIN OUTCOME MEASURES: Women and men were compared with respect to use of LTVV, defined as VT of 8 ml kg-1 or less predicted bodyweight (PBW). A VT was deemed 'default' if the set VT was a round number. A mediation analysis assessed which factors may explain the sex difference in use of LTVV during intra-operative ventilation.RESULTS: This analysis includes 9864 patients, of whom 5425 (55%) were women. A default VT was often set, both in women and men; mode VT was 500 ml. Median [IQR] VT was higher in women than in men (8.6 [7.7 to 9.6] vs. 7.6 [6.8 to 8.4] ml kg-1 PBW, P < 0.001). Compared with men, women were twice as likely not to receive LTVV [68.8 vs. 36.0%; relative risk ratio 2.1 (95% CI 1.9 to 2.1), P < 0.001]. In the mediation analysis, patients' height and actual body weight (ABW) explained 81 and 18% of the sex difference in use of LTVV, respectively; it was not explained by the use of a default VT.CONCLUSION: In this worldwide cohort of patients receiving intra-operative ventilation during general anaesthesia for surgery, women received a higher VT than men during intra-operative ventilation. The risk for a female not to receive LTVV during surgery was double that of males. Height and ABW were the two mediators of the sex difference in use of LTVV.TRIAL REGISTRATION: The study was registered at Clinicaltrials.gov, NCT01601223

Design and baseline characteristics of the finerenone in reducing cardiovascular mortality and morbidity in diabetic kidney disease trial

Background: Among people with diabetes, those with kidney disease have exceptionally high rates of cardiovascular (CV) morbidity and mortality and progression of their underlying kidney disease. Finerenone is a novel, nonsteroidal, selective mineralocorticoid receptor antagonist that has shown to reduce albuminuria in type 2 diabetes (T2D) patients with chronic kidney disease (CKD) while revealing only a low risk of hyperkalemia. However, the effect of finerenone on CV and renal outcomes has not yet been investigated in long-term trials. Patients and Methods: The Finerenone in Reducing CV Mortality and Morbidity in Diabetic Kidney Disease (FIGARO-DKD) trial aims to assess the efficacy and safety of finerenone compared to placebo at reducing clinically important CV and renal outcomes in T2D patients with CKD. FIGARO-DKD is a randomized, double-blind, placebo-controlled, parallel-group, event-driven trial running in 47 countries with an expected duration of approximately 6 years. FIGARO-DKD randomized 7,437 patients with an estimated glomerular filtration rate >= 25 mL/min/1.73 m(2) and albuminuria (urinary albumin-to-creatinine ratio >= 30 to <= 5,000 mg/g). The study has at least 90% power to detect a 20% reduction in the risk of the primary outcome (overall two-sided significance level alpha = 0.05), the composite of time to first occurrence of CV death, nonfatal myocardial infarction, nonfatal stroke, or hospitalization for heart failure. Conclusions: FIGARO-DKD will determine whether an optimally treated cohort of T2D patients with CKD at high risk of CV and renal events will experience cardiorenal benefits with the addition of finerenone to their treatment regimen. Trial Registration: EudraCT number: 2015-000950-39; ClinicalTrials.gov identifier: NCT02545049

Canagliflozin and renal outcomes in type 2 diabetes and nephropathy

BACKGROUND Type 2 diabetes mellitus is the leading cause of kidney failure worldwide, but few effective long-term treatments are available. In cardiovascular trials of inhibitors of sodium–glucose cotransporter 2 (SGLT2), exploratory results have suggested that such drugs may improve renal outcomes in patients with type 2 diabetes. METHODS In this double-blind, randomized trial, we assigned patients with type 2 diabetes and albuminuric chronic kidney disease to receive canagliflozin, an oral SGLT2 inhibitor, at a dose of 100 mg daily or placebo. All the patients had an estimated glomerular filtration rate (GFR) of 30 to &lt;90 ml per minute per 1.73 m2 of body-surface area and albuminuria (ratio of albumin [mg] to creatinine [g], &gt;300 to 5000) and were treated with renin–angiotensin system blockade. The primary outcome was a composite of end-stage kidney disease (dialysis, transplantation, or a sustained estimated GFR of &lt;15 ml per minute per 1.73 m2), a doubling of the serum creatinine level, or death from renal or cardiovascular causes. Prespecified secondary outcomes were tested hierarchically. RESULTS The trial was stopped early after a planned interim analysis on the recommendation of the data and safety monitoring committee. At that time, 4401 patients had undergone randomization, with a median follow-up of 2.62 years. The relative risk of the primary outcome was 30% lower in the canagliflozin group than in the placebo group, with event rates of 43.2 and 61.2 per 1000 patient-years, respectively (hazard ratio, 0.70; 95% confidence interval [CI], 0.59 to 0.82; P=0.00001). The relative risk of the renal-specific composite of end-stage kidney disease, a doubling of the creatinine level, or death from renal causes was lower by 34% (hazard ratio, 0.66; 95% CI, 0.53 to 0.81; P&lt;0.001), and the relative risk of end-stage kidney disease was lower by 32% (hazard ratio, 0.68; 95% CI, 0.54 to 0.86; P=0.002). The canagliflozin group also had a lower risk of cardiovascular death, myocardial infarction, or stroke (hazard ratio, 0.80; 95% CI, 0.67 to 0.95; P=0.01) and hospitalization for heart failure (hazard ratio, 0.61; 95% CI, 0.47 to 0.80; P&lt;0.001). There were no significant differences in rates of amputation or fracture. CONCLUSIONS In patients with type 2 diabetes and kidney disease, the risk of kidney failure and cardiovascular events was lower in the canagliflozin group than in the placebo group at a median follow-up of 2.62 years

NMR Spectral Characteristics of Ultrahigh Pressure High Temperature Impact Glasses of the Giant Kara Crater (Pay-Khoy, Russia)

In this study, we carried out the analysis of the impact melt vein glasses from the Kara impact crater (Russia) in comparison to low-pressure impact melt glasses (tektites) of the Zhamanshin crater (Kazakhstan). 27Al, 23Na, and 29Si MAS NMR spectra of the samples of these glasses were analyzed. The samples of the natural glass contained inclusions of crystalline phases, paramagnetic elements that greatly complicate and distort the NMR signals from the glass phase itself. Taking into account the Mossbauer distribution of Fe in these glasses, the analysis of the spectra of MAS NMR of glass network-former (Si, Al) and potential network-modifiers (Na) of nuclei leads to the conclusion that the Kara impact melt vein glasses are characterized by complete polymerization of (Si,Al)O4 tetrahedral structural units. The NMR features of the glasses are consistent with the vein hypothesis of their formation under conditions of high pressures and temperatures resulting in their fluidity, relatively slow solidification with partial melt differentiation, polymerization, and precipitation of mineral phases as the impact melt cools. The 70 Ma stability of the Kara impact vein glass can be explained by the stabilization of the glass network with primary fine-dispersed pyroxene and coesite precipitates and by the high polymerization level of the impact glass

Problems of Professional Self-Development Among Undergraduates in the Digital Space, Identified During the COVID-19 Pandemic

The article examines the problems of professional self- development among undergraduates in the digital space, identified during the COVID-19 pandemic. The results of the study showed that students have characteristics that are necessary for further personal growth. They want to know themselves and ready to change and learn new things, improve themselves. Conducted during a pandemic and distance learning in self- isolation showed that 50% of students expressed satisfaction with this form of education, explaining their opinion with a high level of independent search for the necessary information, opportunities for self-development, self-realization, and self-improvement. Among the difficulties of distance learning, all surveyed identified: search and critical analysis of digital information; communicative dissatisfaction, the need for approval. The surveyed students in their self-development need a micro- and macro- environment that would create a secure creative educational space. It is necessary to search for new solutions to overcome psychological barriers associated with the need for self-development as future highly qualified specialists in the context of modern trends in the development of digital education

Crystallization and Crystallographic Analysis of a Bradyrhizobium Elkanii USDA94 Haloalkane Dehalogenase Variant with an Eliminated Halide-Binding Site

Haloalkane dehalogenases are a very important class of microbial enzymes for environmental detoxification of halogenated pollutants, for biocatalysis, biosensing and molecular tagging. The double mutant (Ile44Leu + Gln102His) of the haloalkane dehalogenase DbeA from Bradyrhizobium elkanii USDA94 (DbeA&Delta;Cl) was constructed to study the role of the second halide-binding site previously discovered in the wild-type structure. The variant is less active, less stable in the presence of chloride ions and exhibits significantly altered substrate specificity when compared with the DbeAwt. DbeA&Delta;Cl was crystallized using the sitting-drop vapour-diffusion procedure with further optimization by the random microseeding technique. The crystal structure of the DbeA&Delta;Cl has been determined and refined to the 1.4 &Aring; resolution. The DbeA&Delta;Cl crystals belong to monoclinic space group C121. The DbeA&Delta;Cl molecular structure was characterized and compared with five known haloalkane dehalogenases selected from the Protein Data Bank

Improved Biopharmaceutical Properties of Oral Formulations of 1,2,4-Thiadiazole Derivative with Cyclodextrins: in Vitro and in Vivo Evaluation

The synthesized 1,2,4-thiadiazole derivative displaying biological activity has low aqueous solubility and dissolution rate. Novel oral formulations of thiadiazole with β- and hydroxypropyl-β-cyclodextrins were obtained by grinding and freeze-drying methods with the purpose to improve the aqueous solubility. Complex formation of 1,2,4-thiadiazole derivative with cyclodextrins was confirmed by means of solid-state ¹³C MAS CP/TOSS NMR. Solubility, dissolution rate and permeability of the solid inclusion complexes were evaluated in different biorelevant media (SGF, FaSSGF, FaSSIF) simulating the conditions in the gastrointestinal tract. It was demonstrated that the content of biorelevant media affects the properties of the inclusion complexes. In particular, solubilizing effect of cyclodextrins became less pronounced when the micelles of taurocholic acid and lecithin are formed in the dissolution media. The inclusion of thiadiazole into cyclodextrin cavity is in competition with its partitioning into the micelles and this should be taken into account when the in vivo behavior is predicted. The results of in vitro and in vivo experiments were found to be in agreement and showed the highest solubility, dissolution rate and bioavailability of the freeze-dried complexes of thiadiazole with hydroxypropyl-β-cyclodextrin. These complexes can be proposed as more effective dosage forms for oral administration