Long-read direct RNA sequencing of the mitochondrial transcriptome of Saccharomyces cerevisiae reveals condition-dependent intron abundance.

Mitochondria fulfil many essential roles and have their own genome, which is expressed as polycistronic transcripts that undergo co- or posttranscriptional processing and splicing. Due to the inherent complexity and limited technical accessibility of the mitochondrial transcriptome, fundamental questions regarding mitochondrial gene expression and splicing remain unresolved, even in the model eukaryote Saccharomyces cerevisiae. Long-read sequencing could address these fundamental questions. Therefore, a method for the enrichment of mitochondrial RNA and sequencing using Nanopore technology was developed, enabling the resolution of splicing of polycistronic genes and the quantification of spliced RNA. This method successfully captured the full mitochondrial transcriptome and resolved RNA splicing patterns with single-base resolution and was applied to explore the transcriptome of S. cerevisiae grown with glucose or ethanol as the sole carbon source, revealing the impact of growth conditions on mitochondrial RNA expression and splicing. This study uncovered a remarkable difference in the turnover of Group II introns between yeast grown in either mostly fermentative or fully respiratory conditions. Whether this accumulation of introns in glucose medium has an impact on mitochondrial functions remains to be explored. Combined with the high tractability of the model yeast S. cerevisiae, the developed method enables to monitor mitochondrial transcriptome responses in a broad range of relevant contexts, including oxidative stress, apoptosis and mitochondrial diseases.

Continuous synthesis of E. coli genome sections and Mb-scale human DNA assembly.

Whole-genome synthesis provides a powerful approach for understanding and expanding organism function1-3. To build large genomes rapidly, scalably and in parallel, we need (1) methods for assembling megabases of DNA from shorter precursors and (2) strategies for rapidly and scalably replacing the genomic DNA of organisms with synthetic DNA. Here we develop bacterial artificial chromosome (BAC) stepwise insertion synthesis (BASIS)-a method for megabase-scale assembly of DNA in Escherichia coli episomes. We used BASIS to assemble 1.1 Mb of human DNA containing numerous exons, introns, repetitive sequences, G-quadruplexes, and long and short interspersed nuclear elements (LINEs and SINEs). BASIS provides a powerful platform for building synthetic genomes for diverse organisms. We also developed continuous genome synthesis (CGS)-a method for continuously replacing sequential 100 kb stretches of the E. coli genome with synthetic DNA; CGS minimizes crossovers1,4 between the synthetic DNA and the genome such that the output for each 100 kb replacement provides, without sequencing, the input for the next 100 kb replacement. Using CGS, we synthesized a 0.5 Mb section of the E. coli genome-a key intermediate in its total synthesis1-from five episomes in 10 days. By parallelizing CGS and combining it with rapid oligonucleotide synthesis and episome assembly5,6, along with rapid methods for compiling a single genome from strains bearing distinct synthetic genome sections1,7,8, we anticipate that it will be possible to synthesize entire E. coli genomes from functional designs in less than 2 months.

Development of unconventional treatments for mastitis in dairy cattle.

Background: The increase in the intensity of livestock industries, by improving the technology of animal product manufacturing, largely depends on the correct organization of herd reproduction and the intensity of using the biological capabilities of the animal organism. Various diseases, including such common diseases as mastitis, complicate the successful reproduction and growth of enterprise productivity. The widespread use of antibiotic-containing drugs for the treatment of mastitis creates a number of inevitable consequences for the body. The relevance of the study is due to the fact that the residual content of antibiotics in the collected milk after the course of treatment poses a serious danger to human health and reduces the quality of dairy products obtained from such milk. Aim: The authors set the task of developing a new, antibiotic-free method of treating bovine mastitis. This paper is devoted to the problem of improving methods of treating the subclinical mastitis in dairy cattle by alternative methods during the interlactation period. Methods: The leading method for the study of this problem is an experimental method that allows to develop and test a veterinary homeopathic substance for the treatment of subclinical mastitis in cows during the interlactation period. Results: This paper presents materials on the typification of microflora in the milk of cows with subclinical mastitis and the effectiveness of a veterinary homeopathic substance developed by the authors. The use of the veterinary homeopathic substance provided a high therapeutic effect, and did not cause side effects and complications in cows. Conclusion: The resulting veterinary substance was tested and introduced into the "Izhevskiy" natural complex of Akmola region as a new alternative method of treating subclinical mastitis in cows. Based on this substance, a drug for the treatment of mastitis will be developed and proposed for production.

Using in silico modelling and FRET-based assays in the discovery of novel FDA-approved drugs as inhibitors of MERS-CoV helicase.

A Förster resonance energy transfer (FRET)-based assay was used to screen the FDA-approved compound library against the MERS-CoV helicase, an essential enzyme for virus replication within the host cell. Five compounds inhibited the helicase activity with submicromolar potencies (IC50, 0.73-1.65 µM) and ten compounds inhibited the enzyme with micromolar potencies (IC50, 19.6-502 µM). The molecular operating environment (MOE) was used to dock the identified inhibitors on the MERS-CoV helicase nucleotide binding. Strong inhibitors docked well in the nucleotide-binding site and established interactions with some of the essential residues. There was a reasonable correlation between the observed IC50 values and the MOE docking scores of the strong inhibitors (r 2 = 0.74), indicating the ability of the in silico docking model to predict the binding of strong inhibitors. In silico docking could be a useful complementary tool used with the FRET-based assay to predict new MERS-CoV helicase inhibitors. The identified inhibitors could potentially be used in the clinical development of new antiviral treatment for MERS-CoV and other coronavirus related diseases, including coronavirus disease 2019 (COVID-19).

Multiscale structural complexity of natural patterns.

Complexity of patterns is key information for human brain to differ objects of about the same size and shape. Like other innate human senses, the complexity perception cannot be easily quantified. We propose a transparent and universal machine method for estimating structural (effective) complexity of two-dimensional and three-dimensional patterns that can be straightforwardly generalized onto other classes of objects. It is based on multistep renormalization of the pattern of interest and computing the overlap between neighboring renormalized layers. This way, we can define a single number characterizing the structural complexity of an object. We apply this definition to quantify complexity of various magnetic patterns and demonstrate that not only does it reflect the intuitive feeling of what is "complex" and what is "simple" but also, can be used to accurately detect different phase transitions and gain information about dynamics of nonequilibrium systems. When employed for that, the proposed scheme is much simpler and numerically cheaper than the standard methods based on computing correlation functions or using machine learning techniques.

PMMA Thin Film with Embedded Carbon Quantum Dots for Post-Fabrication Improvement of Light Harvesting in Perovskite Solar Cells.

Perovskite solar cells (PSCs) with a standard sandwich structure suffer from optical transmission losses due to the substrate and its active layers. Developing strategies for compensating for the losses in light harvesting is of significant importance to achieving a further enhancement in device efficiencies. In this work, the down-conversion effect of carbon quantum dots (CQDs) was employed to convert the UV fraction of the incident light into visible light. For this, thin films of poly(methyl methacrylate) with embedded carbon quantum dots (CQD@PMMA) were deposited on the illumination side of PSCs. Analysis of the device performances before and after application of CQD@PMMA photoactive functional film on PSCs revealed that the devices with the coating showed an improved photocurrent and fill factor, resulting in higher device efficiency.

The functional landscape of the human phosphoproteome.

Protein phosphorylation is a key post-translational modification regulating protein function in almost all cellular processes. Although tens of thousands of phosphorylation sites have been identified in human cells, approaches to determine the functional importance of each phosphosite are lacking. Here, we manually curated 112 datasets of phospho-enriched proteins, generated from 104 different human cell types or tissues. We re-analyzed the 6,801 proteomics experiments that passed our quality control criteria, creating a reference phosphoproteome containing 119,809 human phosphosites. To prioritize functional sites, we used machine learning to identify 59 features indicative of proteomic, structural, regulatory or evolutionary relevance and integrate them into a single functional score. Our approach identifies regulatory phosphosites across different molecular mechanisms, processes and diseases, and reveals genetic susceptibilities at a genomic scale. Several regulatory phosphosites were experimentally validated, including identifying a role in neuronal differentiation for phosphosites in SMARCC2, a member of the SWI/SNF chromatin-remodeling complex.

100-fold improvement in carrier drift mobilities in alkanephosphonate-passivated monocrystalline TiO2 nanowire arrays.

Single crystal rutile titania nanowires grown by solvothermal synthesis are actively being researched for use as electron transporting scaffolds in perovskite solar cells, in low detection limit ultraviolet photodetectors, in photoelectrochemical water-splitting, and in chemiresistive and electrochemical sensing. The electron drift mobility (µ n ) in solution-grown TiO2 nanowires is very low due to a high density of deep traps, and reduces performance in optoelectronic devices. In this study, the effects of molecular passivation of the nanowire surface by octadecylphosphonic acid (ODPA), on carrier transport in TiO2 nanowire ensembles, were investigated using transient space charge limited current measurements. Infrared spectroscopy indicated the formation of a highly ordered phosphonate monolayer with a high likelihood of bidentate binding of ODPA to the rutile surface. We report the hole drift mobility (µ p ) for the first time in unpassivated solvothermal rutile nanowires to be 8.2 × 10-5 cm2 V-1 s-1 and the use of ODPA passivation resulted in µ p improving by nearly two orders of magnitude to 7.1 × 10-3 cm2 V-1 s-1. Likewise, ODPA passivation produced between a 2 and 3 order of magnitude improvement in µ n from ∼10-5-10-6 cm2 V-1 s-1 to ∼10-3 cm2 V-1 s-1. The bias dependence of the post-transit photocurrent decays in ODPA-passivated nanowires indicated that minority carriers were lost to trapping and/or monomolecular recombination for small values of bias (<5 V). Bimolecular recombination was indicated to be the dominant recombination mechanism at higher bias values.

Rare case of spot diagnosis of nephrotic syndrome in a Saudi male.

A 26- year- old Saudi male school teacher presented with clinical features of nephrotic syndrome. Physical examination revealed loss of subcutaneous fat over the face and upper arms (partial lipodystrophy) and drusen in both eyes. The serum level of complement C3 was low, with normal C4. Renal biopsy revealed features consistent with membranoproliferative glomerulo- nephritis (MPGN) type II [Dense Deposit Disease (DDD)]. He was treated with prednisolone and mycophenolate mofetil with partial remission. He also received lipitor and lisinopril. The occurrence of partial lipo- dystrophy and drusen in a patient with nephrotic syndrome provides a rare clinical opportunity of making a bedside diagnosis of type II MPGN even before kidney biopsy.

Collateral blood vessels in acute ischaemic stroke: a potential therapeutic target.

Ischaemic stroke results from acute arterial occlusion leading to focal hypoperfusion. Thrombolysis is the only proven treatment. Advanced neuroimaging techniques allow a detailed assessment of the cerebral circulation in patients with acute stroke, and provide information about the status of collateral vessels and collateral blood flow, which could attenuate the effects of arterial occlusion. Imaging of the brain and vessels has shown that collateral flow can sustain brain tissue for hours after the occlusion of major arteries to the brain, and the augmentation or maintenance of collateral flow is therefore a potential therapeutic target. Several interventions that might augment collateral blood flow are being investigated.

Invited review: feeding behavior of goats.

Factors influencing the feeding behavior of goats include grazing management practices, type of vegetation and season, breed and stage of production, group size, and properties of diets fed in confinement. Considerable information has been gathered from visual observation during daylight. However, tools are now available to characterize the feeding behavior of goats while grazing and while in confinement throughout the day. Global positioning system collars can be used to assess horizontal and vertical distances traveled, up or down position of the head, and movement within pasture or rangeland areas. A commercially available leg activity monitor allows estimation of the number of steps and time spent standing, lying, and moving rapidly without grazing. However, these measurements do not directly determine grazing. Therefore, prediction equations based on visual observation must be developed. Classification tree analysis is a robust method in developing these equations because the decision tree can be pruned or expanded to provide the best fit. Another equipment system determines time spent eating, ruminating, and remaining idle from the pattern of jaw movement. In addition to use of n-alkanes as internal markers to estimate digestibility, their profile can provide an indication of the botanical composition of the selected diet. Automated feeding systems for confined goats permit determinations such as number of feeder visits and meals, eating time, and rate and pattern of feed intake. Heart rate measured while goats are in normal production settings can be used to predict total energy expenditure through multiplication by energy expenditure per heartbeat of individual animals. To partition the activity energy cost, an estimate of ME intake or measures of changes in body energy status and milk energy yield are needed to determine other sources of heat to be subtracted from total energy expenditure. These methods create the opportunity to gain a fuller understanding of factors influencing the feeding behavior of goats and the relationships with levels and efficiencies of production.

Homocysteine reduces endothelial progenitor cells in stroke patients through apoptosis.

Homocysteine (Hcy) is a risk factor for vascular dysfunction. High levels of Hcy may result in vascular injury accelerating atherosclerosis leading to ischemia. After ischemia, endothelial progenitor cells (EPCs) migrate from bone marrow to repair damaged sites either through direct incorporation of EPCs or by repopulating mature endothelial cells. This study looks into the relationship between increased Hcy in patients with cerebrovascular disease (CVD) and EPCs. Some patients with hyperhomocysteinemia were treated with B vitamins to evaluate if the treatment reverses the elevated Hcy and its impact on their EPC levels. EPCs were treated with Hcy to determine the in vitro effects of Hcy. Our clinical findings show that elevated Hcy levels have an inverse relationship with EPC levels and B vitamin intervention can reverse this effect. Our in vitro work shows that Hcy-mediated EPC toxicity is due to apoptosis involving caspase-8, cytochrome c release, and caspase-3 activation. Vitamin B(6), and B(9) significantly impair Hcy-mediated EPC caspase-3 activation in vitro. Our clinical and in vitro data together indicate that increased Hcy results in a decrease in EPC numbers. This decrease in EPC by Hcy may be occurring through increased apoptosis and B vitamins (B(6), B(9)) intervention can attenuate such effects.

Serum cystatin C as a marker of renal function in patients with acute renal failure.

Serum creatinine level is the traditionally used tool to detect changes in renal function. Serum cystatin C (CC) has been suggested to be an equally effective marker of renal function. We measured the serum levels of creatinine and CC in 73 patients with acute renal failure (ARF) and 300 age and sex matched healthy controls. The serum CC was measured by particle enhanced nephlometric immunoassay (PENIA). As expected, the serum creatinine and CC levels were significantly higher in ARF patients than the healthy controls and serum CC levels correlated significantly with serum creatinine (r = 0.47, p< 0.0001). This correlation further increased for multiple measurements, (r = 0.51, p< 0.0001. No gender difference was noted. Serum CC also correlated significantly with calculated GFR. Correlation of serum CC with serum creatinine and calculated GFR was much greater in patients with deteriorating renal function, compared to patients with improving renal function (p< 0.0001). Our study further suggests that the serum CC is a good marker of renal function in ARF patients, especially those with worsening renal function. Further larger studies are needed to evaluate its role in detecting early ARF and institute possible intervention.

Serum cystatin C in pregnant women: reference values, reliable and superior diagnostic accuracy.

BACKGROUND: A simple, endogenous, accurate and minimally invasive marker of glomerular filtration rate (GFR) is much desired in clinical nephrology. Cystatin C fulfills all criteria to be a marker for GFR. For early detection of renal impairment in pregnant women, it is necessary to determine serum cystatin C reference values and the correlation with GFR. The present study was therefore undertaken. METHOD: Healthy pregnant women were followed during pregnancy and the postnatal period. Patient demographics included age, height, weight, BMI, parity, total blood count, LFT, urea, creatinine, Na, K, and blood sugar. Serum cystatin C was estimated using particle enhanced nephlo-immunoassay method. All the parameters were recorded at the start of pregnancy and then in each trimester and the postnatal period. Regression analysis correlation coefficient, ANOVA and the Student's t-test were used for analysis using the SPSS statistical package. RESULTS: A total of 197 pregnant women were included. Mean serum cystatin C for all the women was 0.82 +/- 0.184 mg/l. Serum cystatin C levels were high -0.89 +/- 0.12 mg/l in the first trimester, decreased significantly to 0.651 +/- 0.14 mg/l during the second trimester (p = 0.0000 compared to first trimester), and increased again to 0.82 +/- 0.191 mg/l in the third trimester. After delivery the level rose to 0.94 +/- 0.12 mg/l. A strong correlation was found between serum cystatin C and serum creatinine. A strong negative correlation was found between GFR and cystatin C values in the women (r = -0.546, p = 0.000). A linear relationship was found between GFR and cystatin C levels. A significant increase in the GFR was noted with the progression of pregnancy from 128.06 +/- 29.7 ml/min in the first trimester to 155.2 +/- 29.59 ml/min during second trimester (p = 0.006). A decline in the level of cystatin C exactly parallel to the increase in the GFR was noted with the progression of pregnancy. Interestingly cystatin C was found to have a strong negative correlation with gestational age (r = -0.663, p = 0.000). CONCLUSION: Our results indicate that the mean serum cystatin C levels reflect changes in the GFR during the entire pregnancy and also in the postnatal period. Moreover, serum cystatin C levels are independent of age, height, weight, or blood sugar level. Cystatin C can be used for close supervision and early diagnosis of renal impairment in pregnant patients. Cystatin C is a reliable, useful and promising marker of GFR in pregnant women.

Solution of the quantum fluid dynamical equations with radial basis function interpolation

The paper proposes a numerical technique within the Lagrangian description for propagating the quantum fluid dynamical (QFD) equations in terms of the Madelung field variables R and S, which are connected to the wave function via the transformation psi = exp(R + iS)/[symbol: see text]. The technique rests on the QFD equations depending only on the form, not the magnitude, of the probability density rho = magnitude of psi 2 and on the structure of R = [symbol: see text]/2 ln rho generally being simpler and smoother than rho. The spatially smooth functions R and S are especially suitable for multivariate radial basis function interpolation to enable the implementation of a robust numerical scheme. Examples of two-dimensional model systems show that the method rivals, in both efficiency and accuracy, the split-operator and Chebychev expansion methods. The results on a three-dimensional model system indicates that the present method is superior to the existing ones, especially, for its low storage requirement and its uniform accuracy. The advantage of the new algorithm is expected to increase for higher dimensional systems to provide a practical computational tool.

Effect of intravenous cyclophosphamide pulse therapy on renal functions and histopathology in patients with severe lupus nephritis.

Despite the wide use of intravenous cyclophosphamide (IC) in lupus nephritis (LN), there are few published studies showing the effect of this treatment on renal histology. In this prospective study, we report the effect of IC on the evolution of histopathologic features in successive renal biopsies in patients with LN. Thirty patients with class IV or V LN were started on IC (10-15 mg/kg) administered once every month for six months followed by three monthly for another six doses making a total of two years of therapy. The clinical course of the disease, serum creatinine and 24 hours urinary protein and creatinine clearance were tested at entry and subsequently during each follow-up visit. Repeat renal biopsy was performed after completion of two years of therapy. The mean serum creatinine of the study patients was 166.3 + 42 tmol/L at entry which decreased to 104 + 46.4 tmol/L at two years (P + 2.4 g at entry to 1.39 + 1.54 g at two years (P + 31 ml/min at the start of treatment to 64 + 32 ml/min at two years of therapy (P 200 tmol/L, of whom six progressed to variable degrees of chronic renal failure. Repeat renal biopsy was performed in 21 patients. The original biopsy of these patients showed class IV in 17 and class V in four patients. On repeat biopsy, five of class IV disease had progressed to advanced sclerosis, four to class V, and five remained unchanged. The remaining three patients with class IV LN changed to one each of class I, II and III. Of the four patients with class V, one progressed to advanced sclerosis, one changed to class III and two remained the same. There was a significant decrease (P < 0.05) in the activity index although there was a significant increase in the chronicity index (P < 0.001). Multivariat analysis for possible risk factors for progression to chronic renal failure showed initial high serum creatinine to be a powerful predictor of renal failure. In conclusion, IC pulse therapy is effective in improving or stabilizing renal function in patients with class IV or V LN. The only poor prognostic determinant observed was higher initial serum creatinine value.

Prevalence of glomerular diseases: king khalid university hospital, saudi arabia.

To obtain a more recent and comprehensive insight into the prevalence of glomerular diseases in our patient population, medical records of 200 patients with biopsy proven glomerulonephritis (GN), between January 1994 and June 1999, at the King Khalid University Hospital, Riyadh, Saudi Arabia were analyzed. Primary glomerular disease was found to be the most prevalent, accounting for 63.5% of all glomerular diseases. Among primary glomerular diseases, focal and segmental glomerulosclerosis (FSGS) was the most common histological lesion (34.6%) and was associated with a high prevalence of hypertension (86.4%), nephrotic syndrome (68.18%), hematuria (63.6%) and renal functional impairment (27.3%). Mesangioproliferative GN was the second most common lesion (25.1%) followed by mesangiocapillary GN (15.7%), IgA nephropathy (10.2%), and minimal change disease (8.5%). Amongst secondary glomerular diseases, lupus nephritis was the most prevalent (24.5%). In conclusion, primary glomerular diseases constituted the commonest group encountered and the prevalence of FSGS was quite high with male sex and young adults predominating. FSGS was also associated with a high prevalence of end-stage renal disease. Further collaborative studies are necessary to explore the predisposing factors and associations of glomerular disease, especially FSGS.

Capillary gas chromatography determination of aliphatic hydrocarbons in fish and water from Oman.

Aliphatic hydrocarbon residues were monitored in two fish species, Indian mackerel (Rastrelliger Kanaguria) and Indian scad (Decapterus russelli) collected from local markets in three different cities in Oman. Residues were also monitored in water samples collected from two sites of the Gulf of Oman. Sampling was conducted twice, in July and again in February to have a thorough understanding of the dynamics of aliphatic hydrocarbon residues. Results indicated the presence of petrogenic aliphatic hydrocarbons ranging from C12-C28 in fish and water samples alike. Fish and water chromatograms were characterized by an unresolved complex mixture (UCM), a confirmatory sign of petrogenic origin of monitored hydrocarbons. Residue levels showed some variations related to sampling sites and time of sampling.

The pattern of diabetic nephropathy among Saudi patients with noninsulin-dependent diabetes mellitus.

There were no studies on the different stages of diabetic nephropathy in Saudi Arabia, particularly the earliest stages. We have therefore investigated the frequency of occurrence of varying degrees of proteinuria including microalbuminuria in noninsulin-dependent diabetes mellitus (NIDDM) Saudi patients as well as the correlation of varying degrees of proteinuria with other diabetic complications and risk factors. One hundred and twenty-five NIDDM patients were studied. Fifty-seven were males and 68 were females. Their mean age was 49.8 +/- 10 years with a mean duration of diabetes of 9.48 +/- 6 years. The mean of HbA1c was 10.3 +/- 2.6%, serum creatinine was 76.7 +/- 23 mmol/L, creatinine clearance 94.3 mL/min, glomerular filtration rate 129.7 +/- 44 and effective renal plasma flow was 496.5 +/- 153. The pattern of proteinuria group was as follows: nephrotic range proteinuria 5.6%, clinical proteinuria 30.4%, microalbuminuria 16.8%. Hypertension and retinopathy were present in 36.8% and 37% of the patients respectively. A significant correlation was found between the presence of hypertension, duration of diabetes and development of diabetic nephropathy. Similarly, a significant correlation was found between retinopathy and the degree of proteinuria. In conclusion, the pattern of diabetic nephropathy in the Saudi NIDDM patients is similar to that in the Western world. Hypertension and duration of diabetes mellitus are important risk factors in the development of diabetic nephropathy. There is a good correlation between retinopathy and the degree of proteinuria.