Direct PCR with the CDC 2019 SARS-CoV-2 assay: optimization for limited-resource settings.

PCR-based diagnostics generally require nucleic acid extraction from patient specimens prior to amplification. As highlighted early in the COVID-19 pandemic, extraction steps may be difficult to scale during times of massive demand and limited reagent supply. Forgoing an extraction step, we previously reported that the N1 primer/probe-set of the widespread CDC COVID-19 assay maintains high categorical sensitivity (95%) and specificity (100%) with direct inoculation of viral transport media (VTM) into qRT-PCR reactions. In contrast, the N2 set demonstrated a prominent Ct delay and low sensitivity (33%) without extraction. In the current study, we have improved the performance of this modified CDC assay (in particular the N2 set) by incorporating N1/N2/RNase P multiplexing and dissecting the effects of annealing temperature, VTM interference, and inoculum volume. The latter two factors exerted a more prominent effect on the performance of N2 than N1, although these effects were largely overcome through elevated annealing temperature. This unextracted/multiplex protocol was evaluated with 41 SARS-CoV-2 positive and 43 negative clinical samples, demonstrating a categorical sensitivity of 92.7% and specificity of 100% versus the unmodified CDC methodology. Overall, this work offers a generalizable strategy to maximize testing capabilities for COVID-19 or other emerging pathogens when resources are constrained.

Powerful turbidity currents driven by dense basal layers.

Seafloor sediment flows (turbidity currents) are among the volumetrically most important yet least documented sediment transport processes on Earth. A scarcity of direct observations means that basic characteristics, such as whether flows are entirely dilute or driven by a dense basal layer, remain equivocal. Here we present the most detailed direct observations yet from oceanic turbidity currents. These powerful events in Monterey Canyon have frontal speeds of up to 7.2 m s-1, and carry heavy (800 kg) objects at speeds of ≥4 m s-1. We infer they consist of fast and dense near-bed layers, caused by remobilization of the seafloor, overlain by dilute clouds that outrun the dense layer. Seabed remobilization probably results from disturbance and liquefaction of loose-packed canyon-floor sand. Surprisingly, not all flows correlate with major perturbations such as storms, floods or earthquakes. We therefore provide a new view of sediment transport through submarine canyons into the deep-sea.

The role of biophysical cohesion on subaqueous bed form size.

Biologically active, fine-grained sediment forms abundant sedimentary deposits on Earth's surface, and mixed mud-sand dominates many coasts, deltas, and estuaries. Our predictions of sediment transport and bed roughness in these environments presently rely on empirically based bed form predictors that are based exclusively on biologically inactive cohesionless silt, sand, and gravel. This approach underpins many paleoenvironmental reconstructions of sedimentary successions, which rely on analysis of cross-stratification and bounding surfaces produced by migrating bed forms. Here we present controlled laboratory experiments that identify and quantify the influence of physical and biological cohesion on equilibrium bed form morphology. The results show the profound influence of biological cohesion on bed form size and identify how cohesive bonding mechanisms in different sediment mixtures govern the relationships. The findings highlight that existing bed form predictors require reformulation for combined biophysical cohesive effects in order to improve morphodynamic model predictions and to enhance the interpretations of these environments in the geological record.

Regulation of IL-8 promoter activity by verrucarin A in human monocytic THP-1 cells.

Macrocyclic trichothecenes have been frequently detected in fungi in water-damaged buildings and exhibited higher toxicity than the well-studied trichothecenes; however, the mechanism underlying their toxicity has been poorly understood. In this study, transcriptional regulation of the cytokine interleukin (IL)-8 by a macrocyclic trichothecene, verrucarin A (VA), in human monocytic THP-1 cells is reported. Consistent with previous findings, VA was 100-fold more cytotoxic than deoxynivalenol (DON), while ochratoxin A (OA) was not cytotoxic. In cells transduced with the wild-type IL-8 promoter luciferase construct, VA induced a biphasic dose response composed of an upregulation of luciferase expression at low concentrations of 0.01-1 ng/ml and a downregulation at high levels of 10 ng/ml and higher. In contrast, DON induced a sigmoid-shaped dose response with the EC50 of 11.6 ng/ml, while OA did not markedly affect the IL-8 expression. When cells were transduced with IL-8 promoter with a mutation of transcription factor nuclear factor-κB (NF-κB)-binding site, VA (1 ng/ml), DON (1000 ng/ml), and tumor necrosis factor (TNF) α (20 ng/ml)-induced luciferase expression were impaired. In addition, the NF-κB inhibitor caffeic acid phenethyl ester inhibited VA-, DON-, and TNFα-induced luciferase expression. Mutation of the CCAAT/enhancer-binding protein (CEBP) ß binding site of the IL-8 promoter affected only DON-, but not VA- and TNFα-induced luciferase expression. Taken together, these results suggested that VA activated IL-8 promoter via an NF-κB-dependent, but not CEBPß-dependent, pathway in human monocytes.

Effect of prior exercise on postprandial triglycerides in overweight young women after ingesting a high-carbohydrate meal.

To examine the effect of prior exercise on the postprandial lipid response to a high-carbohydrate meal in normal-weight (NW=BMI <25) and overweight (OW=BMI >or= 25) women (age 18-25), 10 NW and 10 OW participants completed 2 conditions separated by 1 month. In the morning, the day after control (CT=no exercise) or exercise conditions (EX=60 min cycling at 60% VO(2peak)), participants consumed a high-carbohydrate meal (80% CHO, 15% protein, 5% fat; 75 kJ/kg BM) followed by 6 hr of hourly blood sampling. Blood was analyzed for triglycerides (TG), blood glucose (BG), and insulin (IN). TG levels over the 6-hr period were lower in NW than OW (p= .021) and lower in EX than in CT (p= .006). Area under the curve (AUC) for TG was lower in NW than OW (p= .016) and EX than CT (p= .003). There were nonsignificant tendencies for reduced BG over time (p= .053) and AUC (p= .083), and IN AUC was lower in EX than in CT (p= .040) for both groups and lower in NW than in OW (p= .039). Prior exercise improved TG levels after a high-carbohydrate meal in both groups, and OW women demonstrated a greater postprandial lipemic response than NW regardless of condition. There were tendencies for improved glucose removal with prior exercise in NW vs. OW. Acute exercise can improve postprandial TG responses and might also improve postprandial BG and IN after a large meal in NW and OW young women.

Simultaneous surgeon and side-view video analysis comparing in situ fracture and stop-and-chop phacoemulsification.

A simultaneous surgeon and side-view camera video analysis demonstrated that in situ fracture requires more ultrasonic power behind the iris than the stop-and-chop technique, which requires a longer period of manipulation of larger fragments in the anterior chamber.

Best practice treatment algorithm for primary open-angle glaucoma: implications for U.S. ophthalmology practice.

The objective of this study was to develop a "best-practice" treatment algorithm for the management of primary open-angle glaucoma in patients receiving initial medical therapy, to serve as a consideration for future ophthalmology practice. For comparison, a baseline, "common-practice" treatment algorithm was also created that reflects current ophthalmology practice patterns. Survey instruments were developed based on a comprehensive review of relevant literature, along with input from a general ophthalmologist. A panel of eight ophthalmologists subspecialized in glaucoma management was surveyed. Consensus was achieved using a modified Delphi technique. A comparison of common- and best-practice treatment algorithms suggests that in contrast with expert opinion, nonselective beta blockers are currently used more often, and alpha-2 agonists less often, as first-line therapy for the treatment of primary open-angle glaucoma.