Mitochondrial Physiology and Gene Expression Analyses Reveal Metabolic and Translational Dysregulation in Oocyte-Induced Somatic Nuclear Reprogramming

While reprogramming a foreign nucleus after somatic cell nuclear transfer (SCNT), the enucleated oocyte (ooplasm) must signal that biomass and cellular requirements changed compared to the nucleus donor cell. Using cells expressing nuclear-encoded but mitochondria-targeted EGFP, a strategy was developed to directly distinguish maternal and embryonic products, testing ooplasm demands on transcriptional and post-transcriptional activity during reprogramming. Specifically, we compared transcript and protein levels for EGFP and other products in pre-implantation SCNT embryos, side-by-side to fertilized controls (embryos produced from the same oocyte pool, by intracytoplasmic injection of sperm containing the EGFP transgene). We observed that while EGFP transcript abundance is not different, protein levels are significantly lower in SCNT compared to fertilized blastocysts. This was not observed for Gapdh and Actb, whose protein reflected mRNA. This transcript-protein relationship indicates that the somatic nucleus can keep up with ooplasm transcript demands, whilst transcription and translation mismatch occurs after SCNT for certain mRNAs. We further detected metabolic disturbances after SCNT, suggesting a place among forces regulating post-transcriptional changes during reprogramming. Our observations ascribe oocyte-induced reprogramming with previously unsuspected regulatory dimensions, in that presence of functional proteins may no longer be inferred from mRNA, but rather depend on post-transcriptional regulation possibly modulated through metabolism

Prognostic model to predict postoperative acute kidney injury in patients undergoing major gastrointestinal surgery based on a national prospective observational cohort study.

Background: Acute illness, existing co-morbidities and surgical stress response can all contribute to postoperative acute kidney injury (AKI) in patients undergoing major gastrointestinal surgery. The aim of this study was prospectively to develop a pragmatic prognostic model to stratify patients according to risk of developing AKI after major gastrointestinal surgery. Methods: This prospective multicentre cohort study included consecutive adults undergoing elective or emergency gastrointestinal resection, liver resection or stoma reversal in 2-week blocks over a continuous 3-month period. The primary outcome was the rate of AKI within 7 days of surgery. Bootstrap stability was used to select clinically plausible risk factors into the model. Internal model validation was carried out by bootstrap validation. Results: A total of 4544 patients were included across 173 centres in the UK and Ireland. The overall rate of AKI was 14·2 per cent (646 of 4544) and the 30-day mortality rate was 1·8 per cent (84 of 4544). Stage 1 AKI was significantly associated with 30-day mortality (unadjusted odds ratio 7·61, 95 per cent c.i. 4·49 to 12·90; P < 0·001), with increasing odds of death with each AKI stage. Six variables were selected for inclusion in the prognostic model: age, sex, ASA grade, preoperative estimated glomerular filtration rate, planned open surgery and preoperative use of either an angiotensin-converting enzyme inhibitor or an angiotensin receptor blocker. Internal validation demonstrated good model discrimination (c-statistic 0·65). Discussion: Following major gastrointestinal surgery, AKI occurred in one in seven patients. This preoperative prognostic model identified patients at high risk of postoperative AKI. Validation in an independent data set is required to ensure generalizability

Review of clinical and laboratory features of human Brucellosis

Infection with Brucella spp. continues to pose a human health risk globally despite strides in eradicating the disease from domestic animals. Brucellosis has been an emerging disease since the discovery of Brucella melitensis by Sir David Bruce in 1887. Although many countries have eradicated B. abortus from cattle, in some areas B. melitensis and B. suis have emerged as causes of this infection in cattle, leading to human infections. Currently B. melitensis remains the principal cause of human brucellosis worldwide including India. The recent isolation of distinct strains of Brucella from marine mammals as well as humans is an indicator of an emerging zoonotic disease. Brucellosis in endemic and non-endemic regions remains a diagnostic puzzle due to misleading non-specific manifestations and increasing unusual presentations. Fewer than 10% of human cases of brucellosis may be clinically recognized and treated or reported. Routine serological surveillance is not practiced even in Brucella - endemic countries and we suggest that this should be a part of laboratory testing coupled with a high index of clinical suspicion to improve the level of case detection. The screening of family members of index cases of acute brucellosis in an endemic area should be undertaken to pick up additional unrecognised cases. Rapid and reliable, sensitive and specific, easy to perform and automated detection systems for Brucella spp. are urgently needed to allow early diagnosis and adequate antibiotic therapy in time to decrease morbidity / mortality. The history of travel to endemic countries along with exposure to animals and exotic foods are usually critical to making the clinical diagnosis. Laboratory testing is indispensable for diagnosis. Therefore alertness of clinician and close collaboration with microbiologist are essential even in endemic areas to correctly diagnose and treat this protean human infection. Existing treatment options, largely based on experience gained > 30 years ago, are adequate but not optimal. In our experience, an initial combination therapy with a three drug-regimen followed by a two-drug regimen for at least six weeks and a combination of two drugs with a minimum of six weeks seems warranted to improve outcome in children and adult patients respectively with laboratory monitoring. A safe and effective vaccine in humans is not yet available. Prevention is dependent upon the control of the disease in animal hosts, effective heat treatment of dairy produce and hygienic precautions to prevent occupational exposure. This review compiles the experiences and diagnostic and treatment paradigms currently employed in fighting this disease

Finding Potent Inhibitors Against SARS-CoV-2 Main Protease Through Virtual Screening, ADMET, and Molecular Dynamic Simulation Studies

We have performed vitual screening of 1000 ligands and identified 9 compounds that can potentially inhibit SARS-CoV-2 3CLpro. This docking study is further complemented by the MD/MMGBSA study.<br /

Computational Investigation of Structural Dynamics of SARS-CoV-2 Methyltransferase-Stimulatory Factor Heterodimer Nsp16/nsp10 Bound to the Cofactor SAM

Recently, a highly contagious novel coronavirus (COVID-19 or SARS-CoV-2) has emerged as a global threat in people\u27s health and global economies. Identification of the potential targets and development of a vaccine or antiviral drugs is an urgent demand. The 5’-capping mechanism of eukaryotic mRNA and some viruses such as coronaviruses (CoVs) are essential for maintaining the RNA stability, protein translation, and for viral immune escape. SARSCoV encodes S-adenosyl-L-methionine dependent (SAM) methyltransferase (MTase) enzyme characterized by nsp16 (2’-O-MTase) for generating the capped structure. The present article highlights the binding mechanisms of nsp16 and nsp10 to identify the role of nsp10 in MTase activity. Furthermore, the conformational dynamics and energetic behind the SAM binding to nsp16 in its monomer and dimer form was analyzed by using an extensive molecular dynamics simulation along with the Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PBSA). Our results show that the presence of nsp10 increases the favorable van der Waal and electrostatic interactions between the SAM and nsp16, thus nsp10 acts as a stimulator for its strong binding. The interaction profile suggests that hydrophobic interactions were predominately identified for protein-protein interactions. Also, the stable hydrogen bond between Ala83 (nsp16) and Tyr96 (nsp10), and between Gln87 (nsp16) and Leu45 (nsp10) plays a vital role in the nsp16-nsp10 interface. Further, Computational Alanine Scanning (CAS) mutagenesis was performed, which revealed hotspot mutants, namely I40A, V104A, and R86A for the dimer association. Therefore, the dimer interface of nsp16/nsp10 could also be a potential target to suppress the 2’-O-MTase activity of SARS-CoV-2. Overall, our study provides a comprehensive understanding of the dynamic and thermodynamic process of binding of nsp16 and nsp10 that will contribute to the novel design of peptide inhibitors based on nsp16

Elucidating Biophysical Basis of Binding of Inhibitors to SARS-CoV-2 Main Protease by Using Molecular Dynamics Simulations and Free Energy Calculations

The recent outbreak of novel “coronavirus disease 2019” (COVID-19) has spread rapidlyworldwide, causing a global pandemic. In the absence of a vaccine or a suitablechemotherapeutic intervention, it is an urgent need to develop a new antiviral drug to fight thisdeadly respiratory disease. In the present work, we have elucidated the mechanism of bindingof two inhibitors, namely α-ketoamide and Z31792168 to SARS-CoV-2 main protease (Mproor 3CLpro) by using all-atom molecular dynamics simulations and free energy calculations. Wecalculated the total binding free energy (ΔGbind) of both inhibitors and further decomposedΔGbind into various forces governing the complex formation using the MolecularMechanics/Poisson-Boltzmann Surface Area (MM/PBSA) method. Our calculations revealthat α-ketoamide is more potent (ΔGbind= - 9.05 kcal/mol) compared to Z31792168 (ΔGbind= -3.25 kcal/mol) against COVID-19 3CLpro. The increase in ΔGbind for α-ketoamide relative toZ31792168 arises due to an increase in the favorable electrostatic and van der Waalsinteractions between the inhibitor and 3CLpro. Further, we have identified important residuescontrolling the 3CLpro-ligand binding from per-residue based decomposition of the binding freeenergy. Finally, we have compared ΔGbind of these two inhibitors with the anti-HIV retroviraldrugs, such as lopinavir and darunavir. It is observed that α-ketoamide is more potent comparedto both lopinavir and darunavir. In the case of lopinavir, a decrease in the size of the van derWaals interactions is responsible for the lower binding affinity compared to α-ketoamide. Onthe other hand, in the case of darunavir, a decrease in the favorable intermolecular electrostaticand van der Waals interactions contributes to lower affinity compared to α-ketoamide. Ourstudy might help in designing rational anticoronaviral drugs targeting the SARS-CoV-2 mainprotease. </div

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Not AvailableThe moribund shrimps were exhibiting yellow discoloration of the cephalothorax, blackening of gills and ulcers in the uropod and telson region. PCR for OIE listed viral pathogens ruled out known viral infections. No bacteria was present in haemolymph however, low level of Vibrios i.e. Vibrio harveyi, V. parahaemolyticus and V. alginolyticus was found both in the water collected from the broodstock tanks and affected tissue region. Histopathological examination of affected tissues revealed presence of highly invasive fungal hyphae both by routine and specific fungal stain. PCR amplification of the ITS region (approximately 900 bp) and sequencing confirmed presence of Pythium insidiosum. Phylogenetic analysis of this isolate placed it among the environmental isolates.NFD