Aortic Arch Thrombus and Pulmonary Embolism in a COVID-19 Patient

© 2020 Elsevier Inc. Background: Coronavirus disease 2019 (COVID-19) is associated with endothelial inflammation and a hypercoagulable state resulting in both venous and arterial thromboembolic complications. We present a case of COVID-19-associated aortic thrombus in an otherwise healthy patient. Case Report: A 53-year-old woman with no past medical history presented with a 10-day history of dyspnea, fever, and cough. Her pulse oximetry on room air was 84%. She tested positive for severe acute respiratory syndrome coronavirus 2 infection, and chest radiography revealed moderate patchy bilateral airspace opacities. Serology markers for cytokine storm were significantly elevated, with a serum D-dimer level of 8180 ng/mL (normal \u3c 230 ng/mL). Computed tomography of the chest with i.v. contrast was positive for bilateral ground-glass opacities, scattered filling defects within the bilateral segmental and subsegmental pulmonary arteries, and a large thrombus was present at the aortic arch. The patient was admitted to the intensive care unit and successfully treated with unfractionated heparin, alteplase 50 mg, and argatroban 2 μg/kg/min. Why Should an Emergency Physician Be Aware of This?: Mural aortic thrombus is a rare but serious cause of distal embolism and is typically discovered during an evaluation of cryptogenic arterial embolization to the viscera or extremities. Patients with suspected hypercoagulable states, such as that encountered with COVID-19, should be screened for thromboembolism, and when identified, aggressively anticoagulated

Clinical and molecular characterization of a novel PLIN1 frameshift mutation identified in patients with familial partial lipodystrophy.

Perilipin 1 is a lipid droplet coat protein predominantly expressed in adipocytes, where it inhibits basal and facilitates stimulated lipolysis. Loss-of-function mutations in the PLIN1 gene were recently reported in patients with a novel subtype of familial partial lipodystrophy, designated as FPLD4. We now report the identification and characterization of a novel heterozygous frameshift mutation affecting the carboxy-terminus (439fs) of perilipin 1 in two unrelated families. The mutation cosegregated with a similar phenotype including partial lipodystrophy, severe insulin resistance and type 2 diabetes, extreme hypertriglyceridemia, and nonalcoholic fatty liver disease in both families. Poor metabolic control despite maximal medical therapy prompted two patients to undergo bariatric surgery, with remarkably beneficial consequences. Functional studies indicated that expression levels of the mutant protein were lower than wild-type protein, and in stably transfected preadipocytes the mutant protein was associated with smaller lipid droplets. Interestingly, unlike the previously reported 398 and 404 frameshift mutants, this variant binds and stabilizes ABHD5 expression but still fails to inhibit basal lipolysis as effectively as wild-type perilipin 1. Collectively, these findings highlight the physiological need for exquisite regulation of neutral lipid storage within adipocyte lipid droplets, as well as the possible metabolic benefits of bariatric surgery in this serious disease.Wellcome TrustThis is the author accepted manuscript. The final version is available from the American Diabetes Association via http://dx.doi.org/10.2337/db14-010

Assessment and comparison of four lab made tris-base extenders for preservation of Labrador retriever dog semen at 4ºC

131-139The present study was aimed to develop a most effective extender for long term storage of Labrador dog semen at 4oC. Tris-citric acid-fructose was supplemented with egg yolk (TEY), egg yolk plasma (TEYP), low density lipoproteins (TLDL) and coconut water (TCW) @10, 15 and 20; 10, 15 and 20; 11-15; and 25, 50 and 75 percent, respectively. Extended semen was stored at 4oC and analyzed for sperm attributes and lipid peroxidation. Values for motility, viability and plasma membrane integrity remained significantly (p < 0.05) higher in 15% TEY, 15% TEYP, 13% TLDL and 25%/50% TCW from 0 hr to 72 hrs of storage. It indicated superiority of 15% TEY, 15% TEYP, 13% TLDL and 25% /50% TCW over other concentrations for storage of Labrador dog semen at 4oC. A significant (p < 0.05) decline in motility, viability and membrane integrity was observed from 0-72 hrs of preservation in all extenders, but sperm attributes were still >50% at 72 hrs of preservation. Decline was comparatively less in 15% TEY, 15% TEYP, 25%/50% TCW and 13% TLDL compared to other concentrations. Values for acrosome integrity also differ significantly (p > 0.05) among different concentrations of extenders except TEY. Lipid peroxidation did not vary among different extenders except TEY. In conclusion, both 15% TEY and 15% TEYP were better than 13% TLDL and 25% TCW. TEY extender may be substituted with TEYP for preservation of dog semen at 4°C and further interventions may improve TLDL and TCW extenders

Prediction of buffalo bull fertility on the basis of sperm motion traits, viability, membrane integrity, heat shock protein (HSP70) expression and fertility associated antigen (FAA)

The present study was conducted on 20 buffalo bulls to predict the fertility on the basis of: (a) Computer assisted semen analysis (CASA) based sperm motion traits, (b) viability, (c) membrane integrity, (d) expression for HSP70, and (e) assessment of fertility associated antigen (FAA). Six frozen semen straws from each buffalo bulls were analyzed for sperm motion traits, viz. individual motility, progressive motility, average path velocity (VAP), straight line velocity (VSL), curvilinear velocity (VCL), amplitude of lateral head deviation (ALH), beat cross frequency (BCF), straightness (STR), linearity (LIN) and sperm size. Viability and hypoosmotic swelling tests (HOST) were also conducted. Heat shock protein 70 (HSP70) expression and quantification was conducted using a real time PCR. Fertility associated antigen (FAA) was assessed in fresh semen from buffalo bulls using test. Fertility trial was conducted on 250 normal cycling buffaloes following estrus synchronization with GnRH-PGF- GnRH protocol. It was concluded that buffalo bull fertility could be best predicted on the basis of sperm motion traits (VCL, VAP, VSL, ALH, LIN), motility, viability, HOST and HSP70 expression. However, FAA assessment in fresh semen did not indicate fertility

A Metabolomic Endotype of Bioenergetic Dysfunction Predicts Mortality in Critically Ill Patients with Acute Respiratory Failure

Acute respiratory failure (ARF) requiring mechanical ventilation, a complicating factor in sepsis and other disorders, is associated with high morbidity and mortality. Despite its severity and prevalence, treatment options are limited. In light of accumulating evidence that mitochondrial abnormalities are common in ARF, here we applied broad spectrum quantitative and semiquantitative metabolomic analyses of serum from ARF patients to detect bioenergetic dysfunction and determine its association with survival. Plasma samples from surviving and non-surviving patients (N = 15/group) were taken at day 1 and day 3 after admission to the medical intensive care unit and, in survivors, at hospital discharge. Significant differences between survivors and non-survivors (ANOVA, 5% FDR) include bioenergetically relevant intermediates of redox cofactors nicotinamide adenine dinucleotide (NAD) and NAD phosphate (NADP), increased acyl-carnitines, bile acids, and decreased acyl-glycerophosphocholines. Many metabolites associated with poor outcomes are substrates of NAD(P)-dependent enzymatic processes, while alterations in NAD cofactors rely on bioavailability of dietary B-vitamins thiamine, riboflavin and pyridoxine. Changes in the efficiency of the nicotinamide-derived cofactors\u27 biosynthetic pathways also associate with alterations in glutathione-dependent drug metabolism characterized by substantial differences observed in the acetaminophen metabolome. Based on these findings, a four-feature model developed with semi-quantitative and quantitative metabolomic results predicted patient outcomes with high accuracy (AUROC = 0.91). Collectively, this metabolomic endotype points to a close association between mitochondrial and bioenergetic dysfunction and mortality in human ARF, thus pointing to new pharmacologic targets to reduce mortality in this condition

Simultaneous Analysis of Multiple Mycobacterium tuberculosis Knockdown Mutants In Vitro and In Vivo

Mycobacterium tuberculosis (Mtb) represents one of the most persistent bacterial threats to human health and new drugs are needed to limit its impact. Conditional knockdown mutants can help validate new drug targets, but the analysis of individual mutants is laborious and time consuming. Here, we describe quantitative DNA tags (qTags) and their use to simultaneously analyze conditional Mtb knockdown mutants that allowed silencing the glyoxylate and methylcitrate cycles (via depletion of isocitrate lyase, ICL), the serine protease Rv3671c, and the core subunits of the mycobacterial proteasome, PrcB and PrcA. The impact of gene silencing in multi-strain cultures was determined by measuring the relative abundance of mutant-specific qTags with real-time PCR. This achieved accurate quantification over a broad range of qTag abundances and depletion of ICL, Rv3671c, or PrcBA resulted in the expected impairment of growth of Mtb with butyrate as the primary carbon source, survival during oxidative stress, acid stress and starvation. The impact of depleting ICL, Rv3671c, or PrcBA in multi-strain mouse infections was analyzed with two approaches. We first measured the relative abundance of mutant-specific qTags in total chromosomal DNA isolated from bacteria that were recovered from infected lungs on agar plates. We then developed a two-step amplification procedure, which allowed us to measure the abundances of individual mutants directly in infected lung tissue. Both strategies confirmed that inactivation of Rv3671c and PrcBA severely reduced persistence of Mtb in mice. The multi-strain infections furthermore suggested that silencing ICL not only prevented growth of Mtb during acute infections but also prevented survival of Mtb during chronic infections. Analyses of the ICL knockdown mutant in single-strain infections confirmed this and demonstrated that silencing of ICL during chronic infections impaired persistence of Mtb to the extent that the pathogen was cleared from the lungs of most mice

Evaluating the Sensitivity of Mycobacterium tuberculosis to Biotin Deprivation Using Regulated Gene Expression

In the search for new drug targets, we evaluated the biotin synthetic pathway of Mycobacterium tuberculosis (Mtb) and constructed an Mtb mutant lacking the biotin biosynthetic enzyme 7,8-diaminopelargonic acid synthase, BioA. In biotin-free synthetic media, ΔbioA did not produce wild-type levels of biotinylated proteins, and therefore did not grow and lost viability. ΔbioA was also unable to establish infection in mice. Conditionally-regulated knockdown strains of Mtb similarly exhibited impaired bacterial growth and viability in vitro and in mice, irrespective of the timing of transcriptional silencing. Biochemical studies further showed that BioA activity has to be reduced by approximately 99% to prevent growth. These studies thus establish that de novo biotin synthesis is essential for Mtb to establish and maintain a chronic infection in a murine model of TB. Moreover, these studies provide an experimental strategy to systematically rank the in vivo value of potential drug targets in Mtb and other pathogens

ESCAPADE: Encryption-type-ransomeware: system call based pattern detection

Encryption-type ransomware has risen in prominence lately as the go-to malware for threat actors aiming to compromise Android devices. In this paper, we present a ransomware detection technique based on behaviours observed in the system calls performed by the malware. We identify and present some common high-level system call behavioural patterns targeted at encryption-type ransomware and evaluate these patterns. We further present our repeatable and extensible methodology for extracting the system call log and patterns

Protein inactivation in mycobacteria by controlled proteolysis and its application to deplete the beta subunit of RNA polymerase

Using a component of the Escherichia coli protein degradation machinery, we have established a system to regulate protein stability in mycobacteria. A protein tag derived from the E. coli SsrA degradation signal did not affect several reporter proteins in wild-type Mycobacterium smegmatis or Mycobacterium tuberculosis. Expression of the adaptor protein SspB, which recognizes this modified tag and helps deliver tagged proteins to the protease ClpXP, strongly decreased the activities and protein levels of different reporters. This inactivation did not occur when the function of ClpX was inhibited. Using this system, we constructed a conditional M. smegmatis knockdown mutant in which addition of anhydrotetracycline (atc) caused depletion of the beta subunit of RNA polymerase, RpoB. The impact of atc on this mutant was dose-dependent. Very low amounts of atc did not prevent growth but increased sensitivity to an antibiotic that inactivates RpoB. Intermediate amounts of RpoB knockdown resulted in bacteriostasis and a more substantial depletion led to a decrease in viability by up to 99%. These studies identify SspB-mediated proteolysis as an efficient approach to conditionally inactivate essential proteins in mycobacteria. They further demonstrate that depletion of RpoB by ∼93% is sufficient to cause death of M. smegmatis