In vitro antiviral activity and underlying molecular mechanisms of dipotassium glycyrrhetate against porcine reproductive and respiratory syndrome virus.
Porcine reproductive and respiratory syndrome virus (PRRSV) has caused large economic losses in the swine industry. Currently, there is no effective way to prevent PRRSV infection. In this study, we investigated the inhibitory effect of dipotassium glycyrrhetate (DG), a derivative of glycyrrhetinic acid, on PRRSV infection ability. The cytotoxicity of DG was measured by MTT assay, and the effects of DG on PRRSV N gene/protein were investigated using real-time PCR, western blot and immunofluorescence assay. In addition, the effect of DG on cell apoptosis was analysed by fluorescence staining. Our results indicated that DG could effectively inhibit virus replication and N gene expression in MARC-145 cells infected with PRRSV. When the infected cells received DG, the numbers of apoptotic cells were decreased, and the cleaved caspase-3 contents were decreased dramatically. Our study demonstrates that DG could effectively inhibit the PRRS virus via multiple pathways including inhibition of virus replication and N gene expression and reduction of apoptotic cells. DG can serve as a potential chemical for PRRSV prevention and control.


Natural nanogels crosslinked with S-benzyl-L-cysteine exhibit potent antibacterial activity.
Biofilm-forming bacteria E. coli and P. aeruginosa have both exhibited resistance against multiple antibiotics in clinical settings. To find a solution, researchers have turned to antibacterial structurally modified from natural materials that are harmless to the human body. Among these is DNA, a natural polymer composed of deoxyribose that when treated with HCl exposes its aldehyde groups and produces DNA-HCl. Here, we crosslinked these aldehyde groups with the primary amines in S-benzyl-L-cysteine (SBLC) using a Schiff reaction to obtain DNA-HCl-SBLC. We additionally treated alginate acid (AA) with EDAC, obtaining AA-EDAC, and substituting it with SBLC to produce AA-SBLC. We incorporated the above reactions with an emulsification process to produce nanogels (NGs) that were verified to be spherical and possessing benzene rings successfully grafted onto DNA-HCl and AA-EDAC. These natural NGs were proven to be negatively charged through zeta potential analysis and presented low cytotoxicity toward normal cells in cell organoid viability assays. These SBLC-modified polymers provided better inhibition of bacterial growth than those without modification. Moreover, after incubation with SBLC-modified NGs, bacteria expressed intracellular recA or pvdA in a dose-dependent manner, which was consistent with SEM data from damaged bacteria. Out of four tested NGs, DNA-HCl-SBLC NGs suppressed P. aeruginosa-induced sepsis most effectively and extended the lifespan of C. elegans. This study provides an alternative clinical solution to antibiotics-resistant biofilm strains.


Multidrug resistance mediated by P-glycoproteins.
Overexpression of P-glycoprotein genes is a well-established cause of one form of multidrug resistance. P-glycoproteins are plasma membrane proteins containing two ATP-binding sites and twelve putative transmembrane segments. P-glycoproteins are thought to act as ATP-dependent drug efflux pumps, actively extruding a range of structurally different, hydrophobic drugs from the cell. This simple model can account for the properties of multidrug resistant cells, even those that seem to require more complex explanations. The structure and function of P-glycoprotein genes has been studied in mammals and in several lower eukaryotes. These studies are helping to delineate the range of drugs that can be transported by P-glycoproteins; the genetic mechanisms that can lead to elevated cellular P-glycoproteins levels; and the evolution of the versatile and prolific P-glycoprotein gene family. The physiological function of the human P-glycoproteins encoded by the MDR1 and MDR3 (or MDR2) genes remains a matter of speculation.


Healon effect on the reactions of macrophages to a clear plastic surface.
Growth of peritoneal mouse macrophages in tissue culture on the bottom of clear plastic dishes is used as an experimental model to study the influence of Healon on the reactions of these cells. It is found that a layer of Healon on the plastic surface does not interfere with the adherence of the macrophages to the plastic, and allows for the formation of a continuous and well-developed separating membrane with phagocytizing epithelioid cells, fibroblast-like cells, and small giant cells.


De novo formation of a large cavernoma associated with a congenital torcular dural arteriovenous fistula: case report.
The authors report a case of a developmentally normal child with a congenital complex torcular dural arteriovenous fistula (DAVF) who later, in his teenage years, developed several vermian cavernomas within a large cerebellar developmental venous anomaly (DVA). The patient had initially presented with an abnormally large head circumference but no neurological deficits. He underwent several partial embolization procedures in an attempt to decrease the blood supply of the fistula over the course of 8 years. Nine years following initial presentation, he presented with a fourth ventricular hemorrhage, due to development of a new vermian cavernoma adjacent to a previously known vermian DVA and suffered subsequent mild left-sided hemiataxia from which he later recovered. CT angiographic images demonstrated that the vermian DVA drained into the left transverse sinus, which also drained the torcular arteriovenous fistula. A routine follow-up MRI examination 10 years following initial presentation demonstrated interval development of several large cavernomas in the cerebellum, all within the DVA. The patient had no new symptoms at that time and was neurologically intact. This case report highlights the de novo development of multiple cavernous malformations potentially secondary to DAVF-induced venous congestion in a preexisting DVA.


[Changes in the quantity of nucleic acids in the liver and kidneys of rats with experimental lead poisoning].
In the liver and kidneys of 125 control and test albino rats poisoned perorally daily with different doses of neutral lead acetate for 1, 3, 5, 10, 20, 30 and 45 days the RNA and DNA contest was measured. A statistically significant increase of nucleic acids was registered in both organs, earlier and stronger pronouced in the liver than in the kidneys. The amount of DNA changed quicker than that of DNA. In all the groups of animals the rise in the content was proportional to the introduced amount of lead (20, 40 and 100 mg/kg body weight). Following poisoning with 20 mg/kg of lead acetate the DNA and RNA accretion (statistically significant) remained after the 10th day roughly the same till the 45th day. The main reason for the DNA increment in the study organs seems to be an intensive synthesis, the rise of RNA is due to an inhibited activity of ribonuclease.


Exploring knowledge, attitudes, and practices related to alcohol in Mongolia: a national population-based survey.
The leading cause of mortality in Mongolia is Non-Communicable Disease. Alcohol is recognised by the World Health Organization as one of the four major disease drivers and so, in order to better understand and triangulate recent national burden-of-disease surveys and to inform policy responses to alcohol consumption in Mongolia, a national Knowledge, Attitudes and Practices survey was conducted. Focusing on Non-Communicable Diseases and their risk factors, this publication explores the alcohol-related findings of this national survey. A door-to-door, household-based questionnaire was conducted on 3450 people from across Mongolia. Participants were recruited using a multi-stage random cluster sampling technique, and eligibility was granted to permanent residents of households who were aged between 15 and 64 years. A nationally representative sample size was calculated, based on methodologies aligned with the WHO STEPwise approach to Surveillance. Approximately 50% of males and 30% of females were found to be current drinkers of alcohol. Moreover, nine in ten respondents agreed that heavy episodic drinking of alcohol is common among Mongolians, and the harms of daily alcohol consumption were generally perceived to be high. Indeed, 90% of respondents regarded daily alcohol consumption as either 'harmful' or 'very harmful'. Interestingly, morning drinking, suggestive of problematic drinking, was highest in rural men and was associated with lower-levels of education and unemployment. This research suggests that Mongolia faces an epidemiological challenge in addressing the burden of alcohol use and related problems. Males, rural populations and those aged 25-34 years exhibited the highest levels of risky drinking practices, while urban populations exhibit higher levels of general alcohol consumption. These findings suggest a focus and context for public health measures addressing alcohol-related harm in Mongolia.


A novel route to synthesize Freidinger lactams by micowave irradiation.
The incorporation of a Freidinger-like lactam structure into the backbone of peptides has been proven to be an useful strategy in the design of a variety of conformationally restricted targets. Several different strategies have been developed toward Freidinger lactams but no one resulted to be completely facile. Here, we report an efficient strategy that involves the iodo-derivatives in side chain of an appropriate amino acid used as electrophilic agent, and the standard solid phase peptide synthesis assisted by microwave irradiation. The methodology developed could be useful to perform Freidinger-like lactams with defined stereochemistry for routine use in solid phase peptide chemistry.


[Cytogenetic characteristics of patients with multiple myeloma in China: analysis of 100 case].
To summarize the cytogenetic characteristics of patients with multiple myeloma (MM) in China and clinical significance thereof. Specimens of bone marrow were collected from 100 patients with MM who visited the Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences in Tianjin, China. Chromosome banding analysis and fluorescence in situ hybridization (FISH) were conducted. (1) Clonal chromosome aberrations (CA) were detected in 16 of 93 patients (17.2%) by conventional cytogenetics (CC), and when subclonal CA was included CA were detected in 37.0% of the patients. Numerical aberration analysis showed that the most common trisomies were 11, 21, 3, 6, and 12, and the most common monosomies were 16, 13, 8, 22, and 11. Structural aberration analysis showed that the most frequently involved arms were 14q, 8q, 1q, 6q, 11q, 12p, and 1p, and the most frequently involved breakpoints were 14q32, 8q22 - 24, 11q13 - 14, 6p21, 1q10 - 11, and 12p12 - 13. Ploidy level was arranged in decreasing order as: hypodiploidy (46.1%), pseudodiploid (38.5%), hyperdiploidy (18.8%), and tri-/tetraploidy (15.4%). Chromosome 13 abnormality (C13A) was detected in 7 of the 100 patients (7%). The C13A prevalence rates by CC and interphase FISH were 6.0% and 33.3% respectively. Translocations involving chromosome 14 and/or 14q32 aberrations were detected in 6 patients (6.0%). (2) Univariate analysis showed that C13A, nonhyperdiploidy, and clonal CA were associated with significantly shorter overall survival (OS) and time to progression (TTP). Multivariate analysis showed that C13A was the only independent unfavorable factor. The concrete CA of the patients with MM in China are comparable to those from Western countries, among which C13A, nonhyperdiploidy, hyperdiploidy, and translocations involving immunoglobulin heavy chain locus (IgH) are recurrent. C13A, nonhyperdiploidy, and clonal CA are associated with shorter OS and time-to-progression, and C13A is the only independent adverse prognostic factor.


Performance improvement of N-doped carbon ORR catalyst via large through-hole structure.
N-doped carbon-based materials are crucial electrically conductive additives and non-metal electrocatalysts for the oxygen reduction reaction. At present, many pieces of research are focused on the effects of micropore, mesopore and hierarchical pore structure on the catalytic activity, however, there are few works concerning the role of large-dimension through-hole structure. Hence, in this work, we prepare two kinds of carbon materials with large through-hole structure, i.e. N-doped carbon hollow-spheres and hollow-tubes, as the oxygen reduction catalysts. The synthesis follows template-free morphology-controlled pyrolysis, which is more convenient than the preparation of conventional N-doped nanotubes and graphene. The resultant N-doped carbon hollow-spheres and hollow-tubes evidently enhanced their ORR catalytic activity, remarkable long-term stability and methanol resistance. The large-dimension through-hole structure is found to account for the increase in mass transfer.


Instructional design and assessment of an elective course on the use of drugs in sport.
The use and misuse of drugs in sport is becoming increasingly important globally, and the role of pharmacists is evolving in this regard. This paper describes the design and implementation of an elective course "Drugs in Sport" in an undergraduate pharmacy curriculum. The elective course was designed to introduce BSc pharmacy students to an evidence-based approach to safe, effective, and legal use of drugs in sport. The course covered why athletes take drugs, evolution of doping in sport, the international regulations regarding doping and anti-doping in sport including the World Anti-doping Agency Prohibited List, as well as testing and monitoring for drug use in sport. It also included the role of athlete support personnel (ASP) in preventing the use of prohibited substances by athletes. A web-based survey was conducted at the end of the course to assess the students' perspectives of the course. Students provided an evaluation of the course in terms of its content, methods of delivery, and assessment. Overall, the students demonstrated competence. They gained insight into international and national regulations regarding doping and anti-doping in sport and the potential role of pharmacists as ASP in providing support and advice for athletes and the public. Designing and implementing an evidence-based course on the use of drugs in sport customized for pharmacy students was achieved successfully. Such courses could provide an opportunity to advance the scope of pharmacy practice and possibly provide a new career path for future pharmacists.


Reticular telangiectatic erythema associated with an implantable cardioverter defibrillator.
Reticular telangiectatic erythema (RTE) is a skin reaction associated with implantable cardiac devices (ie, pacemakers and cardioverter defibrillators). We present a patient who developed an erythematous patch over the implantable cardioverter defibrillator site. We discuss the clinical features, histologic findings, and patch testing of this entity.


[Professor A.-C. Hollande at the School of Pharmacy of Nancy, 1912-1922].
André-Charles Hollande, born in Chambéry in 1881, obtained his pharmacy degree and science doctorate in Lyon and Grenoble. He came to the School of Pharmacy in Nancy in 1912 as director of studies in micrography and lecturer in natural history. During the war, while he was mobilized (1914-1917), he was head of the chemical and bacteriological laboratory of Chambéry and worked on clinical chemistry and bacteriology. Professor of natural history in Nancy in January 1920, professor of bacteriology some months after, he left Nancy for Montpellier in 1922. His research in Nancy was devoted to zoology (cytology of insects), parasitology, bacteriology (discovery of 5 bacterial species), clinical chemistry and technics (determination of simulations, histology), and finally cytology and colorations. He was also pharmacist inspector for the towns of Chaumont (Haute-Marne) and Epinal (Vosges).


Personalized Treatment of Periodontitis in a Patient with McArdle's Disease: The Benefits from Probiotics.
McArdle's disease is a severe glycogen storage disease characterized by intolerance to exercise; patients have a syndrome of muscle intolerance to stress, associated with myalgia, cramps, fatigue, and muscle weakness. Periodontal disease is a multifactorial pathology of the supporting tissues of the teeth: one of the main factors is the formation of bacterial biofilm; its control favors the prevention and the maintenance of good health of the oral cavity; and some systemic pathologies can worsen the periodontal disease and hinder its therapy. This case report concerns a woman with McArdle's disease diagnosed with periodontal disease. Material and Methods. A 54-year-old female patient with McArdle's disease has been diagnosed with Stage 3 generalized periodontitis, Grade B. At the baseline, the patient had 82 pockets with probing pocket depth (PPD) equal to or greater than 4 mm. The patient was instructed in the correct methods of oral hygiene and was advised toothpaste and mouthwash based on probiotics; subsequently, a debridement was performed to remove etiological factors using Dental-Biofilm Detection Topographic Technique (D-BioTECH). After 60 days, the number of pockets was reduced from 82 to 14 overall with PPD ≥ 4 mm and from 50 to 2 pockets with PPD ≥ 5 mm. Full mouth bleeding score (FMBS) increased from 48% to 15% and full mouth plaque score (FMPS) from 73% to 15%. In this case, the use of a correct brushing method combined with the D-BioTECH has reduced the disease state, with the use of probiotics at home to restore and maintain a healthy oral microbiome.


Pleistocene avifaunas and the overkill hypothesis.
At the end of the North American Pleistocene, birds and mammals suffered comparable degrees of generic extinction. Both the magnitude and pattern of avian extinction are incompatible with the hypothesis that humans played a major role in causing the demise of numerous North American mammalian genera at this time.


An empirical correction for moderate multiple scattering in super-heterodyne light scattering.
Frequency domain super-heterodyne laser light scattering is utilized in a low angle integral measurement configuration to determine flow and diffusion in charged sphere suspensions showing moderate to strong multiple scattering. We introduce an empirical correction to subtract the multiple scattering background and isolate the singly scattered light. We demonstrate the excellent feasibility of this simple approach for turbid suspensions of transmittance T ≥ 0.4. We study the particle concentration dependence of the electro-kinetic mobility in low salt aqueous suspension over an extended concentration regime and observe a maximum at intermediate concentrations. We further use our scheme for measurements of the self-diffusion coefficients in the fluid samples in the absence or presence of shear, as well as in polycrystalline samples during crystallization and coarsening. We discuss the scope and limits of our approach as well as possible future applications.


Prospective intervention study with a microarray-based, multiplexed, automated molecular diagnosis instrument (Verigene system) for the rapid diagnosis of bloodstream infections, and its impact on the clinical outcomes.
The Verigene Gram-positive blood culture test (BC-GP) and the Verigene Gram-negative blood culture test (BC-GN) identify representative Gram-positive bacteria, Gram-negative bacteria and their antimicrobial resistance by detecting resistance genes within 3 h. Significant benefits are anticipated due to their rapidity and accuracy, however, their clinical utility is unproven in clinical studies. We performed a clinical trial between July 2014 and December 2014 for hospitalized bacteremia patients. During the intervention period (N = 88), Verigene BC-GP and BC-GN was used along with conventional microbiological diagnostic methods, while comparing the clinical data and outcomes with those during the control period (N = 147) (UMIN registration ID: UMIN000014399). The median duration between the initiation of blood culture incubation and the reporting time of the Verigene system results was 21.7 h (IQR 18.2-26.8) and the results were found in 88% of the cases by the next day after blood cultures were obtained without discordance. The hospital-onset infection rate was higher in the control period (24% vs. 44%, p = 0.002), however, no differences were seen in co-morbidities and severity between the control and intervention periods. During the intervention period, the time of appropriate antimicrobial agents' initiation was significantly earlier than that in the control period (p = 0.001) and most cases (90%; 79/88) were treated with antimicrobial agents with in-vitro susceptibility for causative bacteria the day after the blood culture was obtained. The costs for antimicrobial agents were lower in the intervention period (3618 yen vs. 8505 yen, p = 0.001). The 30-day mortality was lower in the intervention period (3% vs. 13%, p = 0.019).


Interactions between shape-persistent macromolecules as probed by AFM.
Water-soluble shape-persistent cyclodextrin (CD) polymers with amino-functionalized end groups were prepared starting from diacetylene-modified cyclodextrin monomers by a combined Glaser coupling/click chemistry approach. Structural perfection of the neutral CD polymers and inclusion complex formation with ditopic and monotopic guest molecules were proven by MALDI-TOF and UV-vis measurements. Small-angle neutron and X-ray (SANS/SAXS) scattering experiments confirm the stiffness of the polymer chains with an apparent contour length of about 130 Å. Surface modification of planar silicon wafers as well as AFM tips was realized by covalent bound formation between the terminal amino groups of the CD polymer and a reactive isothiocyanate-silane monolayer. Atomic force measurements of CD polymer decorated surfaces show enhanced supramolecular interaction energies which can be attributed to multiple inclusion complexes based on the rigidity of the polymer backbone and the regular configuration of the CD moieties. Depending on the geometrical configuration of attachment anisotropic adhesion characteristics of the polymer system can be distinguished between a peeling and a shearing mechanism.


Heat of formation of the HOSO2 radical from accurate quantum chemical calculations.
The reaction HOSO(2)+O(2)-->HO(2)+SO(3) (2) is of crucial importance for sulfuric acid formation in the atmosphere, and reliable thermochemical data are required for an adequate modeling. The currently least well known thermochemical quantity of reaction (2) is the enthalpy of formation of the hydroxysulfonyl radical (HOSO(2)). We report on high-level quantum chemical calculations to predict the binding energy of the HO-SO(2) bond and deduce a value for the enthalpy of formation of HOSO(2) using the most reliable thermodynamic data of OH and SO(2). On the basis of anharmonic vibrational frequencies from calculations at the fc-CCSD(T)/cc-pV(T+d)Z level of theory, the enthalpy of reaction at 0 K for the reaction OH+SO(2)-->HOSO(2) (1) was computed to be Delta(R)H(0 K)(1)=-109.4+/-2.0 kJ/mol and the thermal corrections result in Delta(R)H(298 K)(1)=-114.7+/-3.0 kJ/mol. From these values, we obtain Delta(f)H(0 K)(HOSO(2))=-366.6+/-2.5 and Delta(f)H(298 K)(HOSO(2))=-374.1+/-3.0 kJ/mol, respectively. Accordingly, Delta(R)H(0 K)(2)=-8.5+/-3.0 and Delta(R)H(298 K)(2)=-9.5+/-3.0 kJ/mol.


Negative regulators and their mechanisms in NLRP3 inflammasome activation and signaling.
Inflammasomes are cytosolic multiprotein complexes that cause the release of biologically active interleukin-1β. The best-characterized inflammasome is the NLRP3 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 or Nod-like receptor protein 3) inflammasome. The NLRP3 inflammasome forms an assembly consisting of the ASC (apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain) adaptor protein and the effector, caspase-1 (cysteine-dependent aspartate-directed protease-1). Numerous agents and ligands derived from pathogens, modified self-cells and the environment induce NLRP3 inflammasome complex formation. NLRP3 inflammasome activation is tightly controlled at the transcriptional and post-translational levels to prevent unwanted excessive inflammation. Recent studies have highlighted the roles and mechanisms of several negative regulators that inhibit the assembly of NLRP3 inflammasome complexes and suppress inflammatory responses. The identification and characterization of new players in the regulation of NLRP3 inflammasome may lead to the development of inflammasome-targeting therapeutics against various inflammatory diseases related to NLRP3 inflammasome-associated pathogenesis.


The effect of icariin on autoimmune premature ovarian insufficiency via modulation of Nrf2/HO-1/Sirt1 pathway in mice.
Primary ovarian insufficiency (POI) is a common gynecological disease. Autoimmunity is a common cause of POI. Icariin (ICA) plays a therapeutic role in many autoimmune diseases. This study aims to investigate the effect of ICA on autoimmune POI mice and its effect on immune regulation. Sixty-three female BALB/c mice were randomized into three groups (control, POI, POI + ICA). POI and POI + ICA group were hypodermically injected with zona pellucida three peptides (pZP3) to induce autoimmune POI. Then the POI + ICA group was gavaged with ICA. A vaginal smear was to observe estrous cycles, hematoxylin-eosin staining was to count follicles. Enzyme-linked immunosorbent analysis determined serum FSH, LH, AMH, and anti-zona pellucida antibody (AZPAb) levels. In addition, flow cytometry detected the expression of Th1 cells and Treg cells, and Western blot was used to detect the expression of Nuclear factor E2 related factor 2(Nrf2), heme oxygenase-1 (HO-1), and Sirtuin-1 (Sirt1) proteins. pZP3 treatment decreased serum AMH levels and increased FSH, LH, and AZPAb levels. Additionally, decreases in the number of healthy follicles at all stages and an increase in the number of atretic follicles. Abnormal ovarian structure and an arrested estrous cycle were also noted. However, ICA rescued POI through up-regulating Nrf2, HO-1, and Sirt1 expressions and up-regulating Treg expressions. ICA treatment improved the structure of the injured ovarian and its function in autoimmune POI mice. The mechanism is achieved by increasing the expression of Nrf2/HO-1/Sirt1 pathway in the ovary and increasing Treg cells' expression.


Hepatic cyclooxygenase-2 expression protects against diet-induced steatosis, obesity, and insulin resistance.
Accumulation evidence links obesity-induced inflammation as an important contributor to the development of insulin resistance, which plays a key role in the pathophysiology of obesity-related diseases such as type 2 diabetes and nonalcoholic fatty liver disease. Cyclooxygenase (COX)-1 and -2 catalyze the first step in prostanoid biosynthesis. Because adult hepatocytes fail to induce COX-2 expression regardless of the proinflammatory stimuli used, we have evaluated whether this lack of expression under mild proinflammatory conditions might constitute a permissive condition for the onset of insulin resistance. Our results show that constitutive expression of human COX-2 (hCOX-2) in hepatocytes protects against adiposity, inflammation, and, hence, insulin resistance induced by a high-fat diet, as demonstrated by decreased hepatic steatosis, adiposity, plasmatic and hepatic triglycerides and free fatty acids, increased adiponectin-to-leptin ratio, and decreased levels of proinflammatory cytokines, together with an enhancement of insulin sensitivity and glucose tolerance. Furthermore, hCOX-2 transgenic mice exhibited increased whole-body energy expenditure due in part by induction of thermogenesis and fatty acid oxidation. The analysis of hepatic insulin signaling revealed an increase in insulin receptor-mediated Akt phosphorylation in hCOX-2 transgenic mice. In conclusion, our results point to COX-2 as a potential therapeutic target against obesity-associated metabolic dysfunction.


Effects of feeding drunken horse grass infected with Epichloë gansuensis endophyte on animal performance, clinical symptoms and physiological parameters in sheep.
Many reports showed that grass-endophyte symbiosis induced livestock poisoned. Yet, there is no study evaluating clinical symptoms and physiological parameters in sheep fed Epichloë gansuensis endophyte-infected grass. The objective of the present study was to investigate these indexes by feeding sheep with endophyte-infected A. inebrians (E+ Group) or endophyte-free A. inebrians (E- Group) drunken horse grass or alfalfa hay (Control Group). The Epichloë endophyte caused obvious toxicity symptoms in the sheep fed E+ A. inebrians, with 1 of the 5 sheep having died by the 35th day. The feed intake and body weight gain of the E+ Group were significantly less than the E- and control groups (P < 0.05). Serum concentrations of alanine aminotransferase (ALT, 45.5 mmol/L) and aspartate aminotransferase for the E+ group (AST, 139.3 mmol/L) were significantly (P < 0.05) greater than for the E- (ALT, 31.2 mmol/L; AST, 78.6 mmol/L) and control (ALT, 32.6 mmol/L; AST, 56.6 mmol/L) groups at the fifth week; serum concentration of creatinine for the E+ group (63.8 mmol/L) was also significantly (P < 0.05) greater than for E- (56.6 mmol/L) and control groups (58.5 mmol/L). Meanwhile, urine biochemical indices for the E+ group indicated that ketone and occult blood were significantly (P < 0.05) elevated compared to the other groups while urine pH values were significantly (P < 0.05) acidic. The relative weight of heart, brain, liver, lung and kidney for Group E+ were almost two fold more than the other groups, but uterus weight was about half that found for Group E- or Control. We conclude that the Epichloë endophyte infection is the cause of A. inebrians toxicity to sheep. Interestingly, none of the measured parameters differed significantly between E- and the control groups, which implied that drunken horse grass could be utilized efficiently by sheep when not infected by the Epichloë endophyte.


Cost-effectiveness and Net Monetary Benefit of Durvalumab Consolidation Therapy Versus No Consolidation Therapy After Chemoradiotherapy in Stage III Non-small Cell Lung Cancer in the Italian National Health Service.
The aim of this study was to evaluate the cost-effectiveness and net monetary benefit of durvalumab consolidation therapy compared with no consolidation therapy after chemoradiotherapy in patients with stage III non-small cell lung cancer with programmed cell death 1 ligand 1 expression ≥1% from the Italian National Health Service perspective. We developed a 12-month decision tree combined with a lifetime cohort Markov model in which patients were assigned to receive durvalumab consolidation therapy or active follow-up (Italian standard of care) after chemoradiotherapy to compare cost-effectiveness and net monetary benefit of the two strategies during a 40-year period. Clinical outcomes data were obtained from the respective clinical trials and extrapolated using survival analysis; cost data were derived from Italian official sources and relevant real-world studies. The incremental cost-effectiveness ratio, incremental cost-utility ratio, and incremental net monetary benefit were computed and compared against a 16,372 € per quality-adjusted life-year (QALY) willingness-to-pay threshold. We performed deterministic sensitivity analysis and probabilistic sensitivity analysis to assess how uncertainty affected results; we also performed scenario analyses to compare results under different pricing settings. In the base-case scenario, during a 40-year period, the total costs for patients treated with durvalumab consolidation therapy and active follow-up were €59,860 and €49,840 respectively; life-years gained were 3.47 and 3.31, respectively; and QALYs gained were 2.73 and 2.50, respectively, with an incremental cost-effectiveness ratio of €62,131 per life-year, an incremental cost-utility ratio of €42,322 per QALY, and an incremental net monetary benefit of €-6,144. We found that durvalumab was cost-effective (incremental net monetary benefit = 0) when a discount of 13% and 30% on its official price was applied, considering all other drugs priced according to official or maximum selling prices, respectively. Results were most sensitive to the progression-free survival rate for durvalumab and active follow-up, health utility in progression-free state, and price of subsequent treatments. Our analysis indicates that durvalumab consolidation is cost-effective when a discount is applied on its official price. These results suggest that durvalumab may deliver an incremental health benefit with a contained upfront cost during a 40-year period, from the Italian National Health Service perspective, providing added value in a potentially curative care setting.


Application of life cycle assessment for hospital solid waste management: A case study.
This study was meant to determine environmental aspects of hospital waste management scenarios using a life cycle analysis approach. The survey for this study was conducted at the largest hospital in a major city of Pakistan. The hospital was thoroughly analyzed from November 2014 to January 2015 to quantify its wastes by category. The functional unit of the study was selected as 1 tonne of disposable solid hospital waste. System boundaries included transportation of hospital solid waste and its treatment and disposal by landfilling, incineration, composting, and material recycling methods. These methods were evaluated based on their greenhouse gas emissions. Landfilling and incineration turned out to be the worst final disposal alternatives, whereas composting and material recovery displayed savings in emissions. An integrated system (composting, incineration, and material recycling) was found as the best solution among the evaluated scenarios. This study can be used by policymakers for the formulation of an integrated hospital waste management plan. This study deals with environmental aspects of hospital waste management scenarios. It is an increasing area of concern in many developing and resource-constrained countries of the world. The life cycle analysis (LCA) approach is a useful tool for estimation of greenhouse gas emissions from different waste management activities. There is a shortage of information in existing literature regarding LCA of hospital wastes. To the best knowledge of the authors this work is the first attempt at quantifying the environmental footprint of hospital waste in Pakistan.


Electroencephalography at the time of Covid-19 pandemic in Italy.
During the Covid-19 pandemic, government restrictions limited health care to urgent needs. Neurophysiology centers had to suddenly reschedule their activities, with a lack of specific recommendations about electroencephalography (EEG) execution. During the pandemic phase 1, we launched an online survey to understand the flaws and strengths of the EEG management in Italy at the time of Covid-19 pandemic. A 45-item online survey (published from April 16 to 30, 2020), endorsed by the Italian Society of Clinical Neurophysiology (SINC), the Italian League Against epilepsy (LICE), and the Italian Association of Neurophysiology technologists (AITN), collected EEG management data (EEG's number and type, indications, personnel and patients safety, devices' sanification) during the Covid-19 pandemic. We received responses from 206 centers. The number of EEGs performed was reduced by 76 ± 20%, and several types of specific EEG (video-EEG, ambulatory-EEG, LTM, polysomnography) were reduced at a minimum. Half of the centers performed inpatient EEGs only for urgencies. Repetitive seizures, encephalitis, and non-convulsive status epilepticus were the most common indications. Covid-19-positive patients received less EEG than negative ones (p < 0.0001). EEG requests came mainly not only from neurologists (n = 176) but also from general practitioners (n = 40), emergentists (n = 79), intensivists (n = 72), and other specialists (n = 53). Those centers which continued performing outpatient EEG examinations were instructed to perform the EEG after a Covid-19-related symptom screening for patients and using personal protective equipment (PPE) through all the procedure. Inpatient EEGs were performed using FFP2/FFP3 masks by neurophysiology technologists in only 50% of cases. Patients executed hyperventilation only for real clinical needs, but often (56%) with a mask. Italian neurophysiology centers strongly adhered to government restrictions of lockdown. Some issues emerged, ranging from the evaluation of a proper indication for EEG, technical procedures of EEG recording, and protection of neurophysiology technicians.


Molecular characterization of two insect nematode species (Oxyurida: Thelastomatidae) using small subunit (18S) ribosomal DNA sequence and secondary-structure analyses.
Nematodes of the family Thelastomatidae are parasitic in the alimentary tract of many arthropods, including Periplaneta americana L. In Meerut, Uttar Pradesh, India, two nematode species, namely Hammerschmidtiella indicus and Thelastoma icemi, belonging to this family have been reported. In the present study, the molecular phylogeny of these two nematode species was derived using small subunit (18S) sequence and secondary-structure analyses. The small subunit sequence analyses were carried out to explore the validation and systematics of these species. Phylogenetic analyses were performed for primary sequence data as well as using neighbour-joining and maximum-parsimony approaches. In contrast, the inferred secondary structures for the two species, using free-energy modelling, showed structural identities. As well as this, motif sequences were also found to be a promising tool for nematode species identification. The study provides molecular characterization based on primary sequence data of the 18S ribosomal DNA region of the nematodes along with secondary-structure data and motif sequences for inferences at higher taxonomic levels.


Endoscopic esophagogastric anastomosis with luminal apposition Axios stent (LAS) approach: a new concept for hybrid "Lewis Santy".
Esophagogastric anastomosis (EGA) has a high risk of leakage. Based upon our experience in endoscopic gastrojejunal anastomosis using LAS, the aim of this study was to verify the technical feasibility and the safety of performing an EGA using a hybrid approach (endoscopic and surgical). A pilot prospective study was performed on 8 survival pigs. The procedure was carried out in 2 stages: (i) surgical step consisting of an esogastrectomy by laparotomy with separated suture of the esophagus and stomach; (ii) endoscopic esophagogastric anastomosis using the LAS. The first 2 pigs allowed for the setting of the 2 steps procedure, and 6 were included in the study for assessing the efficacy and safety of the procedure with a 3-week survival course. The primary endpoint was morbidity and mortality. All procedures were successfull. The mean operative time was 98 minutes, with a mean endoscopic time of 46 minutes. Three early deaths occurred within the first weeks, unrelated to the LAS anastomosis. At 3 weeks, endoscopic assessment followed by necropsy demonstrated the right position and the endoscopic removability of the stent with good patency of the esophagogastric anastomosis, without leakage of the endoscopic suture. Pathological examination confirmed the patency of the anastomosis with fusion of mucosal and muscle layers. Endoscopic esophagogastric anastomosis with LAS is feasible and reproducible, without anastomotic leakage. It could be a new alternative to perform safe anastomoses, as part of a hybrid approach (surgical and endoscopic).


Bioinspired 2D Nanomaterials for Sustainable Applications.
The increasing demand for constructing ecological civilization and promoting socially sustainable development has encouraged scientists to develop bioinspired materials with required properties and functions. By bringing science and nature together, plenty of novel materials with extraordinary properties can be created by learning the best from natural species. In combination with the exceptional features of 2D nanomaterials, bioinspired 2D nanomaterials and technologies have delivered significant achievements. Here, the progress over the past decade in bioinspired 2D photonic structures, energy nanomaterials, and superwetting materials, is summarized, together with the challenges and opportunities in developing bioinspired materials for sustainable energy and environmental technologies.


Gestational diabetes mellitus (GDM) - do the number of fulfilled diagnostic criteria predict the perinatal outcome?
The aim of the study was to check whether the number of fulfilled diagnostic criteria of gestational diabetes mellitus (GDM) had any association with patients' characteristics and pregnancy outcomes. A total of 756 women with single pregnancies and GDM who gave birth at the 2nd Department of Obstetrics and Gynecology of the Medical University of Warsaw between 01.2013-12.2016 were included in a retrospective analysis. Patients were divided into 2 groups: A - 499 patients diagnosed with GDM on the basis of one diagnostic criterion, B - 257 patients diagnosed with GDM on the basis of more than one diagnostic criterion. Patients from group A had lower pre-pregnancy BMI than those from group B (median 24.9 kg/m2 vs. 26.5 kg/m2, p=0.0003). Women from group A were less frequently treated with insulin than women from group B (19.1% vs. 32.7%; p=0.00002). Group A had lower median OGTT levels than group B (85.9 mg/dL vs. 94.1 mg/dL, p=0,0001; 160.2 mg/dL vs. 197.6 mg/dL, p=0.0001; 144.8 mg/dL vs. 167.0 mg/dL,p=0.0001; respectively). Moreover, in group B the average week of labor was earlier than in group A (mean 38,1 and 38,5 weeks of gestation, p=0,0006). Patients who fulfilled more than one diagnostic criterion for GDM may have worse pregnancy outcome. We think that a number of fulfilled diagnostic criteria for GDM may be an important risk factor for insulin therapy during pregnancy and earlier gestational age at delivery.


Universal screening for intimate partner violence: a systematic review.
Intimate partner violence (IPV) is known to be prevalent among therapy-seeking populations. Yet, despite a growing understanding of the dynamics of IPV and of the acceptability of screening, universal screening practices have not been systematically adopted in family therapy settings. A rapidly growing body of research data-almost entirely conducted in medical settings-has investigated attitudes and practices regarding universal screening for IPV. This article is a systematic review of the IPV universal screening research literature. The review summarizes literature related to IPV screening rates and practices, factors associated with provider screening practice, the role of training and institutional support on screening practice, impact of screening on disclosure rates, client beliefs and preferences for screening, and key safety considerations and screening competencies. Implications for family therapy and recommendations for further inquiry and screening model development are provided.


Mechanical modulation of spontaneous emission of nearby nanostructured black phosphorus.
In this study, we investigate the spontaneous emission of a quantum emitter nearby black phosphorus (BP) sheet. The spontaneous emission can be modulated mechanically by rotating the BP sheet when the quantum emitter is placed parallel to the sheet. The spontaneous emission is dependent on the electron doping and rotation angle of BP with respect to the x-axis. The Purcell factor decreases with the increase in rotation angle under smaller electron doping. The Purcell factor increases with the increase in rotation angle under larger electron doping. The spontaneous emission of quantum emitter nearby two types of BP ribbon arrays tailored along armchair (type I) and zigzag (type II) directions is studied in detail. The spontaneous emission of quantum emitter parallel to type I is enhanced compared with that parallel to BP sheet. The spontaneous emission decreases remarkably for the quantum emitter parallel to type II compared with that parallel to BP sheet. The spontaneous emission can be flexibly modulated by rotating BP ribbon arrays mechanically in two types. The results obtained in this study provide a new method to actively modulate the spontaneous emission.


The Effect of Perforator Skeletonization on Pedicled Fasciocutaneous Flaps of the Lower Extremity: A Systematic Review.
Lower extremity defects often require free tissue transfer due to a paucity of local donor sites. Locoregional perforator-based flaps offer durable, single-stage reconstruction while avoiding the pitfalls of microsurgery. Multiple harvest techniques are described, yet few studies provide outcome comparisons. Specifically, no study has examined the impact of perforator flap pedicle skeletonization on reconstructive outcomes. This systematic review characterizes technique and impact of pedicle skeletonization on perforator-based fasciocutaneous flaps of the lower extremity. PubMed, Scopus, and the Cochrane Central Register of Controlled Trials were reviewed for literature examining perforator-based fasciocutaneous flaps from knee to ankle, from January 2000 through November 2018. The Preferred Reporting Items for Systematic Reviews-Individual Participant Data (PRISMA-IPD) structure was used. Thirty-six articles were included for quantitative analysis. Of 586 flaps, 365 were skeletonized (60.1%) with 58 major (9.9%) and 19 minor complications (3.2%). With skeletonization, overall reoperative rate was higher (odds ratio [OR]: 9.71, p = 0.004), specifically in propeller (OR: 12.50, p = 0.004) and rotational flaps (OR: 18.87, p = 0.004). The complication rate of rotational flaps also increased (OR: 2.60, p = 0.04). Notably, skeletonization reduced complications in flaps rotated 90 degrees or more (OR: 0.21, p = 0.02). Reoperative rate of distal third defects (OR: 14.08, p = 0.02), flaps over 48 cm2 (OR: 33.33, p = 0.01), and length to width ratios over 1.75 (OR: 7.52, p = 0.03) was increased with skeletonization. Skeletonization increased complications in traumatic defects (OR: 2.87, p = 0.04) and reduced complications in malignant defects (OR: 0.10, p = 0.01). Pedicled, perforator-based flaps can provide a reliable locoregional alternative to free tissue transfer for lower extremity defects. Though skeletonization increased the overall reoperative rate, the complication rate for flaps with 90 degrees or more of rotation was significantly reduced. This suggests skeletonization should be considered when large rotational movements are anticipated to reduce complications that can arise from pedicle compression and venous congestion.


Analytical description of a Fabry-Perot spectrometer. 7: TESS, a high-luminosity high-resolution twin-etalon scanning spectrometer.
The analytical description of the twin-etalon scanning spectrometer (TESS), i.e., an enhanced Fabry-Perot spectrometer, is presented in detail. Practical gains of >100 are attainable with this device, and it is also shown that this instrument is competitive with the spherical Fabry-Perot but with the ability to change the resolution at will. The spectral properties of the TESS are investigated for the determination of Doppler widths and shifts to emphasize the similarities and slight differences between this instrument and the single-etalon single-aperture Fabry-Perot. Practical considerations and tolerances required for operation of the TESS are discussed.


Coincidental Expression of Classic Hodgkin Lymphoma and Neurofibromatosis Type I and Literature Review.
Neurofibromatosis Type 1 (NF1) is a genetic disorder with an incidence of 1 in 2600 to 3000 individuals. It is a clinical diagnosis characterized by café-au-lait macules, neurofibromas, and axillary and/or groin freckling. Because of genetic mutations in the NF1 gene affecting the Ras/mitogen-activated protein kinase pathway, there is also risk of associated soft tissue sarcomas and hematologic malignancies. However, reports of classic Hodgkin lymphoma in patients with NF1 are sparse. We report an adolescent with NF1 who developed classic Hodgkin lymphoma. Although there is an unclear association between mutations in the NF1 gene and classic Hodgkin lymphoma, further studies are warranted.


Stability evaluation of 7 % chloral hydrate syrup contained in mono and multi-dose bottles under room and refrigeration conditions.
To evaluate the stability of an extemporaneously prepared 7% chloral hydrate syrup under different conditions of storage and dispensing. Three batches of 7% chloral hydrate syrup were prepared. Each batch was stored in 50 light-resistant glass containers of 60 mL with child-resistant caps and in two bottles of 1000 mL to simulate two forms of dispensing, mono and multi-dose, respectively. Twenty five mono-dose bottles and a multi-dose bottle of each batch were stored under room conditions (20 ± 1 °C) and the rest of the samples were stored in the fridge (5 ± 2 °C). The physical, chemical and microbiological stability was evaluated for 180 days. Stability was defined as retention of at least 95% of the initial concentration of chloral hydrate, the absence of both visible particulate matter, or color and/or odor changes and the compliance with microbiological attributes of non-sterile pharmaceutical products. At least 98% of the initial chloral hydrate concentration remained throughout the 180-day study period. There were no detectable changes in color, odor, specific gravity and pH and no visible microbial growth. These results were not affected by storage, room or refrigeration conditions or by the frequent opening or closing of the multi-dose containers. Extemporaneously compounded 7% chloral hydrate syrup was stable for at least 180 days when stored in mono or multi-dose light-resistant glass containers at room temperature and under refrigeration.


Terahertz vibration-rotation-tunneling spectroscopy of the ammonia dimer. II. A-E states of an out-of-plane vibration and an in-plane vibration.
Terahertz vibration-rotation-tunneling transitions have been measured between ca. 78.5 and 91.9 cm-1, and assigned to A-E (ortho-para) combinations of NH3 monomer states. The spectrum is complicated by inversion splittings that correlate to E symmetry monomer rovibronic states. Twenty progressions have been assigned to six excited states involving an out-of-plane vibration and an in-plane intermolecular vibration. The quality of the fit was affected by strong Coriolis interactions among these states and possibly an additional K = 2 state that was not explicitly observed in the data.


[A case of autoimmune pancreatitis associated with ulcerative colitis].
A 31-year-old man was admitted to our hospital because of upper abdominal pain. He had been diagnosed with ulcerative colitis (UC) at age 28, but the disease has been in remission since then. On admission, he had slight fever, abdominal pain, and bloody stools six times a day, and the serum levels of pancreatic enzyme and IgG4 were elevated. Diffuse enlargement of the pancreas was detected by abdominal computed tomography; furthermore, narrowing of the main pancreatic duct was revealed using endoscopic retrograde cholangiopancreatography. Based on these findings, he was given a diagnosis of autoimmune pancreatitis (AIP) associated with UC. Both diseases improved without using steroids. After discharge, he has not had any recurrence of AIP or UC despite not being on steroid treatment, although the serum IgG4 level has shown a slight tendency to elevate.


An electrochemical sensor for detection of trace-level endocrine disruptor bisphenol A using Mo2Ti2AlC3 MAX phase/MWCNT composite modified electrode.
Bisphenol A (BPA) is an industrially preferred material for the production of plastic and polycarbonate as well as a used material for the interior of food and beverage cans. In this study, synthesis and electrochemical sensor application of Mo2Ti2AlC3/MWCNT (multi-walled carbon nanotube) nanocomposite for BPA sensing was evaluated. Mo2Ti2AlC3 was used as MAX phase material in the design of the sensor, and MWCNT was preferred to increase conductivity and sensitivity. Cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were used to determine Mo2Ti2AlC3/MWCNT nanocomposite's electrochemical sensor performances which had LOD of 2.7 nM and LOQ of 8.91 nM in the linear working range of 0.01-8.50 μM calculated from DPV. The composite showed a single oxidation step against BPA which is diffusion-controlled and irreversible. The sensor was successfully applied for the determination of BPA in milk pack, plastic bottle, and can with recoveries ranging from 95.67% to 100.60%. In addition, sensor performance was examined through selectivity, repeatability, and reusability studies. HPLC as a standard determination method was carried out for accuracy of the voltammetric determination method in the real samples. The developed sensor could be applied to different areas from industry quality control to clinical analysis for the detection of BPA.


Parameters and symbols for use in nuclear magnetic resonance (IUPAC recommendations 1997).
NMR is now frequently the technique of choice for the determination of chemical structure in solution. Its uses also span structure in solids and mobility at the molecular level in all phases. The research literature in the subject is vast and ever-increasing. Unfortunately, many articles do not contain sufficient information for experiments to be repeated elsewhere, and there are many variations in the usage of symbols for the same physical quantity. It is the aim of the present recommendations to provide simple check-lists that will enable such problems to be minimised in a way that is consistent with general IUPAC formulation. The area of medical NMR and imaging is not specifically addressed in these recommendations, which are principally aimed at the mainstream use of NMR by chemists (of all sub-disciplines) and by many physicists, biologists, materials scientists and geologists etc. working with NMR. The document presents recommended notation for use in journal publications involving a significant contribution of nuclear magnetic resonance (NMR) spectroscopy. The recommendations are in two parts: (1) Experimental parameters which should be listed so that the work in question can be repeated elsewhere. (2) A list of symbols (using Roman or Greek characters) to be used for quantities relevant to NMR.


Surgical treatment of cervical kyphosis in Larsen syndrome: report of 3 cases and review of the literature.
A retrospective case series. To review the surgical results for midcervical kyphosis in 3 cases with Larsen syndrome, and to discuss the choice of surgical treatments. Cervical kyphosis is the most hazardous and serious manifestation of Larsen syndrome due to the risk of life-threatening paralysis, and thus usually requires surgical treatment. However, little information has been reported concerning surgical treatments for this challenging condition. Three patients with Larsen syndrome were surgically treated for midcervical kyphosis at our institution. An infant with mild cervical kyphosis was successfully treated with posterior arthrodesis using a halo immobilization, and anterior vertebral growth with a mature posterior fusion mass resulted in spontaneous correction of the kyphosis. In the remaining 2 infants with myelopathic symptoms due to severe and structural kyphosis, anterior decompression and fusion via a lateral approach followed by posterior fusion with segmental spinal instrumentation and halo immobilization resulted in improved neurologic symptoms and solid fusion. Posterior spinal fusion is only indicated for patients with mild and flexible cervical kyphosis, and anterior decompression and circumferential arthrodesis is required for patients with severe kyphotic deformity, who usually develop myelopathic symptoms. Anterior surgery for such a small patient with severe kyphosis involves much higher risk of spinal cord injury during decompression maneuvers and difficulty in stabilization of the reconstructed cervical spine. Therefore, all patients with Larsen syndrome should be screened with radiographs at the first visit to detect cervical kyphosis early so that posterior alone fusion is possible.


Empirical mode decomposition for self-mixing Doppler signals of hemodynamic optical probes.
A new type of optical probe based on laser Doppler self-mixing technology, for a truly non-contact measurement in a single location, and extraction of the temporal features of the distension wave in the arterial wall, was developed. The monitoring of temporal features allows the assessment of cardiovascular function when measurement is carried out at the carotid artery. An algorithm based on the short-time Fourier transform and empirical mode decomposition was applied to the test setup self-mixing signals for the determination of waveform features, with an accuracy of a few milliseconds and a root mean square error less than 3 ms. In vivo testing signals show great consistency in the measured pulse pressure waveform.


Lectin-magnetic separation (LMS) for isolation of Toxoplasma gondii oocysts from concentrated water samples prior to detection by microscopy or qPCR.
Although standard methods for analyzing water samples for the protozoan parasites Cryptosporidium spp. and Giardia duodenalis are available and widely used, equivalent methods for analyzing water samples for Toxoplasma gondii oocysts are lacking. This is partly due to the lack of a readily available, reliable immunomagnetic separation technique (IMS). Here we investigated the use of lectin-magnetic separation (LMS) for isolating T. gondii oocysts from water sample concentrates, with subsequent detection by microscopy or molecular methods. Four different types of magnetic beads coated with wheat germ agglutinin (WGA) were tested for capture of oocysts from clean or dirty water samples. Dynabeads (Myone T1 and M-280) consistently provided mean capture efficiencies from 1 ml clean water in excess of 97%. High recoveries were also found with Tamavidin beads (in excess of 90%) when LMS was used for capture from a small (1 ml) volume. Dissociation (required for detection by microscopy) using 0.1N hydrochloric acid (HCl), as standard in IMS, was not successful, but could be achieved using a combination of acidified pepsin (AP) and N-acetyl d-glucosamine. Although simple centrifugation was as effective as LMS when concentrating high numbers of oocysts from clean water, LMS provided superior results when oocysts numbers were low or the water sample was dirty. Application of LMS integrated with qPCR enabled detection of 10 oocysts per 10 ml dirty water sample concentrate. These findings indicate that LMS with WGA coupled to magnetic beads could be an efficient isolation step in the analysis of water sample concentrates for T. gondii oocysts, with detection either by microscopy or by qPCR.


Genome sequencing of Sporisorium scitamineum provides insights into the pathogenic mechanisms of sugarcane smut.
Sugarcane smut can cause losses in cane yield and sugar content that range from 30% to total crop failure. Losses tend to increase with the passage of years. Sporisorium scitamineum is the fungus that causes sugarcane smut. This fungus has the potential to infect all sugarcane species unless a species is resistant to biotrophic fungal pathogens. However, it remains unclear how the fungus breaks through the cell walls of sugarcane and causes the formation of black or gray whip-like structures on the sugarcane plants. Here, we report the first high-quality genome sequence of S. scitamineum assembled de novo with a contig N50 of 41 kb, a scaffold N50 of 884 kb and genome size 19.8 Mb, containing an estimated 6,636 genes. This phytopathogen can utilize a wide range of carbon and nitrogen sources. A reduced set of genes encoding plant cell wall hydrolytic enzymes leads to its biotrophic lifestyle, in which damage to the host should be minimized. As a bipolar mating fungus, a and b loci are linked and the mating-type locus segregates as a single locus. The S. scitamineum genome has only 6 G protein-coupled receptors (GPCRs) grouped into five classes, which are responsible for transducing extracellular signals into intracellular responses, however, the genome is without any PTH11-like GPCR. There are 192 virulence associated genes in the genome of S. scitamineum, among which 31 expressed in all the stages, which mainly encode for energy metabolism and redox of short-chain compound related enzymes. Sixty-eight candidates for secreted effector proteins (CSEPs) were found in the genome of S. scitamineum, and 32 of them expressed in the different stages of sugarcane infection, which are probably involved in infection and/or triggering defense responses. There are two non-ribosomal peptide synthetase (NRPS) gene clusters that are involved in the generation of ferrichrome and ferrichrome A, while the terpenes gene cluster is composed of three unknown function genes and seven biosynthesis related genes. As a destructive pathogen to sugar industry, the S. scitamineum genome will facilitate future research on the genomic basis and the pathogenic mechanisms of sugarcane smut.


Sound analysis of the magnetically levitated left ventricular assist device HeartMate 3™.
The HeartMate 3™ has shown lower rates of adverse events compared to previous devices due to the design and absence of mechanical bearings. For previous devices, sound analysis emerged as a way to assess pump function. The aims of this study were to determine if sound analysis can be applied to the HeartMate 3 in vivo and in vitro and to evaluate an electronic stethoscope. Sound recordings were performed with microphones and clinical accessible electronic stethoscope. The recordings were studied in both the time and the frequency domains. Recordings from four patients were performed to determine if in vivo and in vitro recordings are comparable. The results show that it is possible to detect sound from HeartMate 3 and the sound spectrum is clear. Pump frequency and frequency of the pulsatile mode are easily determined. Frequency spectra from in vitro and in vivo recordings have the same pattern, and the major proportion (96.7%) of signal power is located at the pump speed frequency ±40 Hz. The recordings from the patients show low inter-individual differences except from location of peaks originating from pump speed and harmonics. Electronic stethoscopes could be used for sound recordings, but the dedicated equipment showed a clearer sound spectrum. The results show that acoustic analysis can also be performed with the HeartMate 3 and that in vivo and in vitro sound spectrum is similar. The frequency spectra are different from previous devices, and methods for assessing pump function or thrombosis need further evaluation.


Generation of multiepitope cancer vaccines based on large combinatorial libraries of survivin-derived mutant epitopes.
Immune tolerance is the main challenge in the field of cancer vaccines, so modified peptide sequences or naturally occurring mutated versions of cancer-related wild-type (WT) antigens represent a promising pathway. However, the low immunogenicity of mutation-induced neoantigens and, particularly, their incapacity to activate CD8+ T cells are generating doubts on the success of neoantigen-based cancer vaccines in clinical trials. We developed a novel vaccine approach based on a new class of vaccine immunogens, called variable epitope libraries (VELs). We used three regions of survivin (SVN), composed of 40, 49 and 51 amino acids, along with the complete SVN protein to generate the VELs as multiepitope vaccines. BALB/c mice, challenged with the aggressive and highly metastatic 4T1 cell line, were vaccinated in a therapeutic setting. We showed significant tumor growth inhibition and, most importantly, strong suppression of lung metastasis after a single immunization using VEL vaccines. We demonstrated vaccine-induced broad cellular immune responses concomitant with extensive tumor infiltration of T cells, the activation of CD107a+ IFN-γ+ T cells in the spleen and a significant increase in the number of CD3+ CD8+ Ly6C+ effector T cells. In addition, we observed the presence of interferon-γ-, granzyme B- and perforin-producing lymphocytes along with modifications in the amount of CD11b+ Ly6Cint/low Ly6G+ granulocytic myeloid-derived suppressor cells and CD4+ CD25+ FoxP3+ regulatory T cells in the lungs and tumors of mice. In summary, we showed that the VELs represent a potent new class of cancer immunotherapy and propose the application of the VEL vaccine concept as a true alternative to currently available vaccine platforms.


Rotor modulations and recoupling strategies in 13C solid-state magic-angle-spinning NMR spectroscopy: probing molecular orientation and dynamics.
Recoupling strategies for anisotropic interactions enable the investigation of molecular structure, order and dynamics in a sensitive and site-specific fashion by solid-state NMR spectroscopy. Whereas magic-angle spinning (MAS) efficiently averages anisotropic interactions and enhances spectral resolution, recoupling pulse sequences selectively restore certain parts of rotor-modulated dipole-dipole couplings or chemical shift anisotropies (CSA). More specifically, it is possible to recouple either the omegaR- or the 2omegaR-modulated terms of an interaction Hamiltonian, which exhibit different orientation dependencies and, in this way, provide a means of distinguishing whether the observed NMR spectra are affected by molecular motion or by molecular orientation. Sideband patterns generated by reconversion rotor encoding allow for a precise and selective determination of coupling constants and anisotropies, which contain site-specific information on structure, orientation and/or dynamics of individual molecular segments. Corresponding recoupling schemes are presented in a common context, and the possibilities of exploiting these effects for the determination of order parameters of oriented materials, such as oriented polymer chains or extruded fibres of a discotic mesogen, are discussed. The obtained orientational order parameters are compared to results from two-dimensional wide angle X-ray scattering (WAXS).


Effect of 1,25-dihydroxyvitamin D3 on adenylate cyclase and protein kinase C in pig epidermis.
1,25-dihydroxyvitamin D3 (1,25(OH)2D3) is suggested to be involved in the regulation of keratinocyte proliferation and differentiation. Recent evidence also indicates its potential value for the treatment of psoriasis, where the alteration of various transmembrane signalling systems has been well documented. Using porcine epidermis, we investigated the effect of 1,25(OH)2D3 on adenylate cyclase and protein kinase C systems, both of which are markedly altered in the psoriatic hyperproliferative epidermis. The effect was compared with that of another anti-psoriatic agent, hydrocortisone. Neither 1,25(OH)2D3 nor hydrocortisone revealed any effect on cyclic AMP levels or adenylate cyclase responses of epidermis. Long-term (24 h) hydrocortisone treatment, however, resulted in increased beta-adrenergic-, and prostaglandin E-adenylate cyclase responses. 1,25(OH)2D3-treatment had no effect on the epidermal adenylate cyclase responses following 24 h of incubation. The addition of both 1,25(OH)2D3 and hydrocortisone to the incubation medium resulted in the attenuation of the hydrocortisone-induced beta-adrenergic-, and prostaglandin E-adenylate cyclase responses of the epidermis. Neither agent had any effect on the cholera toxin-induced and forskolin-induced cyclic AMP accumulations of the epidermis. Neither 1,25(OH)2D3 nor hydrocortisone had any effect on the epidermal protein kinase C activity. It has been suggested that various anti-psoriatic agents might reveal their effect through the modulation of the adenylate cyclase system. Since 1,25(OH)2D3 had no effect when it was added singly to the incubation medium and rather inhibited hydrocortisone-induced adenylate cyclase stimulation, it is suggested that 1,25(OH)2D3 reveals its therapeutic efficacy through the mechanism, probably independently of the adenylate cyclase system.


Bivoltine cocoon color sex-limited breeds of Bombyx mori (Lepidoptera: Bombycidae) show enhanced economic performance and fecundity following spermidine supplementation.
Sericulture has immense economic significance. Separating male and female silkworm pupae for egg production in grainage is a laborious task; hence, sex-limited breeds for cocoon color are advantageous for this process. The major constraint in sex-limited breeds is their low fecundity. Sex-limited female moths lay a lower number of eggs than nonsex-limited breeds. Polyamine, spermidine was shown to improve fecundity in several organisms, including the silkworm, Bombyx mori L. In the present study, cocoon color sex-limited breeds, HTO2SL, APS27SL, and SLFC27, were selected and fed with spermidine to improve fecundity and nutritional efficiency. The fifth-instar silkworm larvae of the selected breeds were subjected to standard rearing conditions with and without spermidine supplementation up to spinning. The spermidine-supplemented sex-limited breeds exhibited a significant increase in fecundity, nutritional indices, and economic parameters compared with the control sex-limited breeds. Among the 3 sex-limited breeds tested, the performance of APS27SL improved significantly.


RANKL is associated with persistent primary teeth and delayed permanent tooth emergence.
Tooth eruption is a process that is not fully understood. To evaluate whether genetic polymorphisms for RANK/RANKL/OPG are associated with delayed tooth emergence. To evaluate whether the relative expression of this genes is associated with persistent primary teeth. To evaluate whether genetic polymorphisms for RANK/RANKL/OPG could be involved in delayed tooth emergence, saliva samples from 160 children, aged 6-13 years old, were analysed. To test if there is correlation between gene expression of RANK/RANKL/OPG in children with delayed tooth emergence and persistent primary teeth, periapical tissue from 15 children with persistent primary teeth and from 15 control subjects were collected for qPCR analysis. Fifty-six children with delayed tooth emergence (35%) had at least one permanent tooth with delayed emergence. The T allele in RANKL (rs9594738) increased the risk of delayed tooth emergence (P = 0.02; OR = 1.71, 95%CI 1.09-2.75). The relative gene expression for RANKL and the ratio RANKL/OPG in children with delayed tooth emergence and persistent primary teeth were lower compared to controls (P = 0.02 and P = 0.005, respectively). Data suggest that the polymorphism rs9594738 in RANKL is associated with delayed permanent tooth emergence. Moreover, reduced relative gene expression of RANKL in periapical tissue is associated with persistent primary teeth.


B cells from the bench to the clinical practice.
B cells develop in bone marrow and undergo antigen-induced activation and terminal differentiation in germinal centres of secondary lymphoid organs. Each B cell is a clone which means that an individual B cell has a unique genetic code and produces only one type of antibody when stimulated by antigen being able to multiply itself and originate several B cells with the same antigen specificity (clonal selection theory). However their important role in adaptive immune responses is supported by the remarkable capacity of recognizing an unlimited array of antigens due to mechanisms of antibody diversity such as V(D)J recombination class switching and somatic hypermutation. B cells can also function as antigen presenting cells that can activate T cells improving the effectiveness of the immune response. Immune B cell tolerance surveillance through clonal deletion anergy and receptor editing is also necessary to avoid pathological conditions like autoimmune diseases. B cells can contribute to autoimmunity by autoantibody production cytokine synthesis antigen presentation T cell activation and ectopic lymphogenesis.


Current ultrafiltration techniques before, during and after pediatric cardiopulmonary bypass procedures.
Ultrafiltration, which is currently considered as a standard method to remove excess water administered during pediatric cardiopulmonary bypass (CPB), aims to minimize the adverse effects of hemodilution, such as tissue edema and blood transfusion. Three ultrafiltration techniques can be used before, during and after CPB procedures, including conventional ultrafiltration (CUF), modified ultrafiltration (MUF) and zero-balance ultrafiltration (Z-BUF). These methods are widely different, but they have common benefits on hemoconcentration, less requirement for blood products, and reduction of the systemic inflammatory responses (SIRS). The present review attempts to restate these ultrafiltration circuitries, application methods, end-points, and clinical impacts.


Hydrogen and Dihydrogen Bonds in the Reactions of Metal Hydrides.
The dihydrogen bond-an interaction between a transition-metal or main-group hydride (M-H) and a protic hydrogen moiety (H-X)-is arguably the most intriguing type of hydrogen bond. It was discovered in the mid-1990s and has been intensively explored since then. Herein, we collate up-to-date experimental and computational studies of the structural, energetic, and spectroscopic parameters and natures of dihydrogen-bonded complexes of the form M-H···H-X, as such species are now known for a wide variety of hydrido compounds. Being a weak interaction, dihydrogen bonding entails the lengthening of the participating bonds as well as their polarization (repolarization) as a result of electron density redistribution. Thus, the formation of a dihydrogen bond allows for the activation of both the MH and XH bonds in one step, facilitating proton transfer and preparing these bonds for further transformations. The implications of dihydrogen bonding in different stoichiometric and catalytic reactions, such as hydrogen exchange, alcoholysis and aminolysis, hydrogen evolution, hydrogenation, and dehydrogenation, are discussed.


AIDS incidence rates in Austria.
AIDS incidence rates were calculated for Austria based on cumulative data up to October 31st 1988. Geographical differences are obvious, provinces of the northwest showing higher incidence rates than south-east provinces. An association of high AIDS incidence rates with intravenous drug abuse is suggested by the data.


Regulation of pri-microRNA BIC transcription and processing in Burkitt lymphoma.
BIC is a primary microRNA (pri-miR-155) that can be processed to mature miR-155. In this study, we show the crucial involvement of protein kinase C (PKC) and nuclear factor-kappaB (NF-kappaB) in the regulation of BIC expression upon B-cell receptor triggering. Surprisingly, Northern blot analysis did not reveal any miR-155 expression upon induction of BIC expression in the Burkitt lymphoma-derived Ramos cell line, whereas other microRNAs were clearly detectable. Ectopic expression of BIC in Ramos and HEK293 cells resulted in miR-155 expression in HEK293, but not in Ramos cells, suggesting a specific block of BIC to miR-155 processing in Ramos. In line with the results obtained with Ramos, lack of miR-155 expression after induction of BIC expression was also observed in other Burkitt lymphoma cell lines, indicating a generic and specific blockade in the processing of BIC in Burkitt lymphoma. In contrast, induction of BIC expression in normal tonsillar B cells resulted in very high levels of miR-155 expression and induction of BIC expression in Hodgkin's lymphoma cell lines. It also resulted in elevated levels of miR-155. Our data provide evidence for two levels of regulation for mature miR-155 expression: one at the transcriptional level involving PKC and NF-kappaB, and one at the processing level. Burkitt lymphoma cells not only express low levels of BIC, but also prevent processing of BIC via an, as yet, unknown mechanism.


Transforming the fermented fish landscape: Microbiota enable novel, safe, flavorful, and healthy products for modern consumers.
Regular consumption of fish promotes sustainable health while reducing negative environmental impacts. Fermentation has long been used for preserving perishable foods, including fish. Fermented fish products are popular consumer foods of historical and cultural significance owing to their abundant essential nutrients and distinct flavor. This review discusses the recent scientific progress on fermented fish, especially the involved flavor formation processes, microbial metabolic activities, and interconnected biochemical pathways (e.g., enzymatic/non-enzymatic reactions associated with lipids, proteins, and their interactions). The multiple roles of fermentation in preservation of fish, development of desirable flavors, and production of health-promoting nutrients and bioactive substances are also discussed. Finally, prospects for further studies on fermented fish are proposed, including the need of monitoring microorganisms, along with the precise control of a fermentation process to transform the traditional fermented fish to novel, flavorful, healthy, and affordable products for modern consumers. Microbial-enabled innovative fermented fish products that consider both flavor and health benefits are expected to become a significant segment in global food markets. The integration of multi-omics technologies, biotechnology-based approaches (including synthetic biology and metabolic engineering) and sensory and consumer sciences, is crucial for technological innovations related to fermented fish. The findings of this review will provide guidance on future development of new or improved fermented fish products through regulating microbial metabolic processes and enzymatic activities.


[Malaria in the islands of the Comoro archipelago. Historical and geophysical aspects. Epidemiologic considerations].
Located on the maritime road to India, important harbour during slavery period, Comoro Archipelago was therefore free of malaria in the XVIIth century in spite of the presence of infected persons. In the XIXth century, malaria became endemic in Mayotte, Mohéli and Anjouan but only at the beginning of the XXth century in Grande Comore. The origin of vectors and main parasite, P. falciparum, are african. After quite a "natural" evolution, the situation is the same in these islands (continual high transmission) except Mayotte where a malaria programme is performed since 1976. But the historical differences, island by island, in the outbreak of malaria suggest various ecological balance which would be underlined by antimalarial activities.


Ephrin-B2 overexpression enhances integrin-mediated ECM-attachment and migration of B16 melanoma cells.
Eph-receptor tyrosine kinases (Eph-RTKs) and their membrane-bound receptor-like ligands, the ephrins, represent a cell-cell signaling system that directs cellular migration during development. Differential expression in cancer suggests similar roles in tumor progression. We have previously shown that ephrin-B2 mRNA is overexpressed in advanced malignant melanomas (MM). In this study, immunohistochemistry revealed a most prominent expression of ephrin-B2 in the invasive front of advanced MM. Therefore, we addressed the question of whether ephrin-B2 signaling modulates MM cell migration and matrix interaction. Using a wild-type ephrin-B2-negative B16 mouse MM subclone we show that overexpression of ephrin-B2 leads to the formation of multiple lamellipodia, enhanced polymerisation of actin fibers, and induction of focal adhesion complexes with constitutive activation of focal adhesion kinase. Consequently, ephrin-B2-overexpressing B16 cells display a significant increase of beta1-integrin-mediated attachment to matrix components, preferentially laminin and fibronectin. As a further effect of ephrin-B2 overexpression, we observed an accelerated migration in both Boyden chamber invasion experiments as well as in in vitro scratch-wound assays. We conclude that ephrin-B2 can act as a major modulator of cell migration and matrix interactions of MM cells, which possibly contributes to the expansion and metastatic spread of MM in vivo.


Acute blastomycosis thyroiditis.
Acute thyroiditis is very unusual, and fungal thyroiditis is even more rare. Cervical blastomycosis, on one occasion masquerading as a thyroid mass, has been reported. Here we report the first case of acute blastomycosis infection of the thyroid documented by biopsy and imaging studies. The patient was a 23-year-old woodcutter with no history or features of overt immunodeficiency. The initial response to Itraconazole therapy was satisfactory.


Trauma-Informed Care and Posttraumatic Growth Among Bereaved Youth: A Pilot Study.
Although a fair amount has been written about posttraumatic stress disorder among bereaved children and adolescents, less has been written about posttraumatic growth (PTG) and its predictors among this population. This study examines predictors of PTG and the impact of trauma-informed care on PTG among bereaved youth. A preexperimental, pretest-posttest design was applied to measure PTG among bereaved children (N = 32) before and after attending a healing camp that provides trauma-informed care. A regression model was applied to examine predictors of PTG. Results showed that children participating in the camp increased their PTG scores to a statistically significant degree. Circumstance of death (sudden or expected) was a predictor in this study. The results are discussed in relation to limitations, implications for future research, and practice.


Bioinformatics analysis and optimization of cell-free protein synthesis.
Cell-free protein synthesis offers substantial advantages over cell-based expression, allowing direct access to the protein synthetic reaction and meticulous control over the reaction conditions. Recently, we identified a number of statistically significant correlations between calculated and predicted properties of amino acid sequences and their amenability to heterologous cell-free expression. These correlations can be of practical use for predicting expression success and optimizing cell-free protein synthesis. In this chapter, we describe our approach and demonstrate how computational and predictive bioinformatics can be used to analyze and optimize cell-free protein expression.


Possibility of elevated parathyroid function in patients with calcium-containing nephrolithiasis as compared with normal controls.
109 patients with calcium-containing nephrolithiasis and 10 normal controls underwent oral calcium load test. After thorough examination, 6 of the 109 patients were diagnosed as having primary hyperparathyroidism (PHPT) and the remainder as having normocalcemic nephrolithiasis without PHPT. Following the oral calcium load test, the latter were operationally divided into 3 groups - normocalciuric nephrolithiasis (NN), n = 78; absorptive hypercalciuria (AH), n = 10, and renal hypercalciuria (RH), n = 15 - according to the criteria reported by Pak et al. Before the oral calcium load test, nephrogenous adenosine 3',5'-monophosphate (NcAMP), urinary adenosine 3'-5'-monophosphate (urinary cAMP), and plasma immunoreactive parathyroid hormone (iPTH) were determined to evaluate parathyroid function. This function, as assessed by mean basal NcAMP in the NN, AH and RH groups as well as the PHPT group, was significantly increased as compared with that in the normal controls. Within the NcAMP-elevated 4 groups, the mean basal NcAMP was highest in the PHPT group followed by the RH, AH and NN groups. In view of the mean basal NcAMP, disregarding the PHPT group, the NN and AH groups seemed to be intermediate types between the normal controls and the RH groups. Similar, but less distinctive results were obtained in the determination of urinary cAMP and plasma iPTH. On the other hand, when leaving the PHPT group out, the mean basal urinary calcium creatinine ratio (Ca/Cr) was highest in the RH group followed by the AH and NN groups, and lowest in normal controls, suggesting that the NN and AH groups were intermediate between normal controls and the RH group. The mean basal urinary Ca/Cr ratio in the PHPT group was moderately elevated but not remarkable. Almost similar tendencies were observed in 24-hour urinary calcium excretions on a calcium-restricted diet. A weakly positive correlation (r = 0.232, p less than 0.05) between basal NcAMP and basal urinary Ca/Cr ratio was observed in accumulated cases of the NN, AH and RH groups, whereas a negative correlation (r = -0.664, p less than 0.05) was obtained in normal controls. It is concluded that a possible abnormal calcium metabolism is suggested in stone formers without PHPT. Additionally, it is speculated that 'relative hypercalciuria' in NN and hypercalciuria in AH and RH might be accounted for in a single line of a primary renal leak of calcium.


Evidence of ssDNA in tubulofilamentous particles: their relationship to scrapie-associated fibrils.
Abnormal tubulofilamentous particles were identified by electron microscopy using a simple touch negative staining technique from brains of mice infected with four strains of the scrapie agent. Treatment by three proteolytic enzymes and subsequent treatment with DNase and mung bean nuclease of grids prepared from the infected animals confirmed previous observations that the tubulofilamentous particles observed in scrapie-effected brains are complex structures. The core of the tubulofilamentous particle scrapie-associated fibrils was revealed by treatment with SDS. Treatment with proteolytic enzymes and subsequent treatment with DNase or mung bean nuclease or S1 nuclease also revealed typical and transitional stages of scrapie-associated fibrils. However, treatment with RNase A had no effect. The data suggest that nucleic acid is a single-stranded DNA protected by a protein coat.


Regional and single glomerular blood flow in the rat kidney prepared for micropuncture. A methodological study.
Regional renal and single glomerular blood flow was investigated in rats using 141-Ce and 85-Sr labelled microspheres. The kidneys were prepared for micropuncture and also the artery was catheterized with a steel cannula in order to make injections of 133-Xenon and vasoactive substances selectively into one kidney. The microspheres were first separated by sedimentation into a narrow fraction and then injected as a plasma suspension into the carotid arter; the two batches were infused at half an hour intervals. Single glomeruli were sampled from kidney sections with the aid of silicon rubber casts allowing the identification of the glomeruli both with respect to their localisation and their postglomerular appearance. The results showed a relatively high blood flow in the superficial and juxtamedullary glomeruli of above 100 nl/min per 100 g rat whereas in the other glomeruli the blood flow was about 70 nl/min per 100 g rat. No differences could be observed between the right untouched kidney and the left prepared for micropuncture. Futhermore no discrepancies between the results from the two injections could be found. The data on the total renal blood flow obtained by the 133-Xenon wash-out method agreed well with the microsphere method.


Introducing Emerging Methods and Technologies.
With the current issue of The FEBS Journal, we are introducing a new category of invited review article contributions on Emerging Methods and Technologies. These articles provide an overview and discussion of recent, emerging methods that significantly advance and improve research efforts in the different fields of molecular and cellular research of our The FEBS Journal authors and readers. Deputy Editorial Manager Manuel Breuer and our Emerging Methods and Technologies Commissioning Editor Eric Chevet introduce the series.


Domain Shifts in Machine Learning Based Covid-19 Diagnosis From Blood Tests.
Many previous studies claim to have developed machine learning models that diagnose COVID-19 from blood tests. However, we hypothesize that changes in the underlying distribution of the data, so called domain shifts, affect the predictive performance and reliability and are a reason for the failure of such machine learning models in clinical application. Domain shifts can be caused, e.g., by changes in the disease prevalence (spreading or tested population), by refined RT-PCR testing procedures (way of taking samples, laboratory procedures), or by virus mutations. Therefore, machine learning models for diagnosing COVID-19 or other diseases may not be reliable and degrade in performance over time. We investigate whether domain shifts are present in COVID-19 datasets and how they affect machine learning methods. We further set out to estimate the mortality risk based on routinely acquired blood tests in a hospital setting throughout pandemics and under domain shifts. We reveal domain shifts by evaluating the models on a large-scale dataset with different assessment strategies, such as temporal validation. We present the novel finding that domain shifts strongly affect machine learning models for COVID-19 diagnosis and deteriorate their predictive performance and credibility. Therefore, frequent re-training and re-assessment are indispensable for robust models enabling clinical utility.


PHYSICAL AND BIOLOGICAL OBSERVATIONS ON HERPESVIRUS.
Smith, Kendall O. (Baylor University College of Medicine, Houston, Texas). Physical and biological observations on herpesvirus. J. Bacteriol. 86:999-1009. 1963.-The development of herpesvirus in human lung fibroblasts was studied by plaque assays and physical particle counts. Approximately 200 infectious units and 60,000 particles were produced by single cells during a single growth cycle. Production of physically recognizable particles preceded infectious virus particle production by about 5 to 6 hr, suggesting the occurrence of a maturation process during formation. Aggregation of particles in clusters and chains was observed in many cases. One of the mechanisms for this aggregation was the connection of particles by deoxyribonucleic acid strands. These strands appeared to connect some particles in a way that suggests a structural continuity between their cores.


Toward construction of a self-sustained clock-like expression system based on the mammalian circadian clock.
Despite recent advances in circadian biology, detailed understanding of how a biological pacemaker system is assembled, maintained, and regulated continues to be a significant challenge. We have assembled and characterized a first-generation, regulatable, self-sustained clock-like expression system based on key components of the mammalian circadian clock. The molecular setup of the clock-like oscillator was reduced to the core set of positive and negative elements common to all known circadian pacemakers. Sophisticated tetracycline-responsive multi-cistronic expression integrated with forefront lentiviral transduction tools enabled autoregulated reporter transgene expression in a human cell line. We characterized transgene expression kinetics of an artificial oscillator and showed that its expression profiles could be modulated by a serum shock and administration of regulating tetracycline antibiotics. Design of a generic mammalian clock-like expression system will offer novel opportunities to study circadian biology and may provide a unique tool for rhythmic expression of desired transgenes fostering advances in biopharmaceutical manufacturing, gene therapy, and tissue engineering.


Angiotensin converting enzyme gene polymorphism and ACE inhibition in diabetic nephropathy.
The antiproteinuric effect of angiotensin converting enzyme (ACE) inhibition in insulin-dependent diabetes mellitus (IDDM) patients with diabetic nephropathy varies considerably. Therefore, we tested the potential role of an insertion (I)/deletion (D) polymorphism of the ACE gene on this early antiproteinuric responsiveness in an observational follow-up study. Sixty (II, N = 13; ID, N = 26 and DD, N = 21) young hypertensive IDDM patients suffering from diabetic nephropathy were investigated during three months before and for the initial six month period during ACE inhibition [captopril 44 (SD 22) mg/24 hr, no differences in drug dose between groups]. Blood pressure (MABP) and albuminuria (ELISA) were measured three (1 to 6) times before and three (1 to 13) times during ACE inhibition. At baseline the groups (II/ID/DD) had comparable (1) mean arterial blood pressure (MABP mm Hg) of 113 +/- 10/108 +/- 9/114 +/- 8, (2) albuminuria (geometric mean with 95% CI) 1394 (747 to 2608)/1176 (844 to 1797) and 1261 (827 to 2017) mg/24 hr, and (3) serum creatinine (geometric mean with 95% CI), 80 (68 to 93)/85 (76 to 97)/103 (85 to 119) mumol/liter, respectively. Angiotensin converting enzyme inhibition induced a significant reduction in MABP, albuminuria and kidney function in all three groups (II/ID/DD; P < 0.05): (1) MABP (mean +/- SD) 12 +/- 7/5 +/- 7/8 +/- 9 mm Hg (ANOVA, P = 0.02); (2) albuminuria [mean (95% CI)] 61 (34 to 77)/22 (3 to 37)/31 (13 to 46) %, (ANOVA, P < 0.01); and (3) increasing serum creatinine [mean (95% CI)] 8 (4 to 12)/9 (3 to 16)/8 (0 to 16) % (ANOVA, NS), respectively. Adjusting for differences in reduction in MABP did not change the association between decrease in albuminuria and ACE/ID genotypes (P < 0.01). A multiple linear regression analysis revealed that the ACE/ID polymorphism, albuminuria and MABP at baseline independently influenced the decline in albuminuria after initiation of ACE inhibition (R2 = 0.21, P < 0.01). A significant association between changes in MABP and albuminuria was demonstrated (R2 = 0.16, P < 0.01). Our data show that hypertensive albuminuric IDDM patients with the II genotype are particularly susceptible to commonly advocated renoprotective treatment.


Notch down-regulation by endocytosis is essential for pigment cell determination and survival in the Drosophila retina.
The clathrin heavy chain is a fundamental element in endocytosis and therefore, in the internalization of several cell-surface receptors through which cells interact with their environment. Here we show that the only non-lethal mutant allele of the clathrin heavy chain identified to date in metazoans, the Drosophila Chc(4), involves the substitution of a residue at the knee region of the molecule that impairs clathrin-dependent endocytosis. We have investigated the consequences of this endocytic defect in Drosophila retinal development and found that it produces an inhibition of programmed cell death in the retinal lattice, followed by widespread death of interommatidial pigment cells once retinal development has been completed. Through genetic interactions and transgenic analyses, we show that Chc(4) phenotypes are caused by a Notch receptor gain-of-function, providing a dramatic example of the importance of Notch down-regulation by endocytosis. An increase in Notch signaling is also observed in Drosophila wings in response to the mutant clathrin, suggesting that Notch levels are controlled by clathrin-dependent endocytosis. We discuss the implications of these findings for current models on eye-development and for the role of endocytosis in Notch signaling.


[Polycythaemia as sole symptom of renal adenoma (author's transl)].
In two children, a 9 year-old boy and a 10 1/2 year-old girl, who presented with polycythaemia as the only symptom, the expected renal tumour was only found after exclusion of all other causes of polycythaemia. The delay in diagnosis was caused by technically inadequate intravenous urograms, which were erroneously passed as normal. In one child low kv X-ray exposition of the kidneys led to the diagnosis of a renal tumour. In the other child high-dose urography and tomography gave the indication for selective angiography. Normalization of the red blood count postoperatively verifies the connection between preoperative erythrocytosis and the renal tumour. Histologically both cases proved to be renal adenomas, which are extremely rare in childhood.


Imino proton exchange and base-pair kinetics in the AMP-RNA aptamer complex.
We report on the dynamics of base-pair opening in the ATP-binding asymmetric internal loop and flanking base-pairs of the AMP-RNA aptamer complex by monitoring the exchange characteristics of the extremely well resolved imino protons in the NMR spectrum of the complex. The kinetics of imino proton exchange as a function of basic pH or added ammonia catalyst are used to measure the apparent base-pair dissociation constants and lifetimes of Watson-Crick and mismatched base-pairs, as well as the solvent accessibility of the unpaired imino protons in the complex. The exchange characteristics of the imino protons identify the existence of four additional hydrogen bonds stabilizing the conformation of the asymmetric ATP-binding internal loop that were not detected by NOEs and coupling constants alone, but are readily accommodated in the previously reported solution structure of the AMP-RNA aptamer complex published from our laboratory. The hydrogen exchange kinetics of the non-Watson-Crick pairs in the asymmetric internal loop of the AMP-RNA aptamer complex have been characterized and yield apparent dissociation constants (alphaKd) that range from 10(-2) to 10(-7). Surprisingly, three of these alphaKd values are amongst the lowest measured for all base-pairs in the AMP-RNA aptamer complex. Comparative studies of hydrogen exchange of the imino protons in the free RNA aptamer and the AMP-RNA aptamer complex establish that complexation stabilizes not only the bases within the ATP-binding asymmetric internal loop, but also the flanking stem base-pairs (two pairs on either side) of the binding site. We also outline some preliminary results related to the exchange properties of a sugar 2'-hydroxyl proton of a guanosine residue involved in a novel hydrogen bond that has been shown to contribute to the immobilization of the bound AMP by the RNA aptamer, and whose resonance is narrow and downfield shifted in the spectrum.