Protease inhibitors targeting coronavirus and filovirus entry.

In order to gain entry into cells, diverse viruses, including Ebola virus, SARS-coronavirus and the emerging MERS-coronavirus, depend on activation of their envelope glycoproteins by host cell proteases. The respective enzymes are thus excellent targets for antiviral intervention. In cell culture, activation of Ebola virus, as well as SARS- and MERS-coronavirus can be accomplished by the endosomal cysteine proteases, cathepsin L (CTSL) and cathepsin B (CTSB). In addition, SARS- and MERS-coronavirus can use serine proteases localized at the cell surface, for their activation. However, it is currently unclear which protease(s) facilitate viral spread in the infected host. We report here that the cysteine protease inhibitor K11777, ((2S)-N-[(1E,3S)-1-(benzenesulfonyl)-5-phenylpent-1-en-3-yl]-2-{[(E)-4-methylpiperazine-1-carbonyl]amino}-3-phenylpropanamide) and closely-related vinylsulfones act as broad-spectrum antivirals by targeting cathepsin-mediated cell entry. K11777 is already in advanced stages of development for a number of parasitic diseases, such as Chagas disease, and has proven to be safe and effective in a range of animal models. K11777 inhibition of SARS-CoV and Ebola virus entry was observed in the sub-nanomolar range. In order to assess whether cysteine or serine proteases promote viral spread in the host, we compared the antiviral activity of an optimized K11777-derivative with that of camostat, an inhibitor of TMPRSS2 and related serine proteases. Employing a pathogenic animal model of SARS-CoV infection, we demonstrated that viral spread and pathogenesis of SARS-CoV is driven by serine rather than cysteine proteases and can be effectively prevented by camostat. Camostat has been clinically used to treat chronic pancreatitis, and thus represents an exciting potential therapeutic for respiratory coronavirus infections. Our results indicate that camostat, or similar serine protease inhibitors, might be an effective option for treatment of SARS and potentially MERS, while vinyl sulfone-based inhibitors are excellent lead candidates for Ebola virus therapeutics

Inhibitors of SARS-CoV entry--identification using an internally-controlled dual envelope pseudovirion assay.

Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) emerged as the causal agent of an endemic atypical pneumonia, infecting thousands of people worldwide. Although a number of promising potential vaccines and therapeutic agents for SARS-CoV have been described, no effective antiviral drug against SARS-CoV is currently available. The intricate, sequential nature of the viral entry process provides multiple valid targets for drug development. Here, we describe a rapid and safe cell-based high-throughput screening system, dual envelope pseudovirion (DEP) assay, for specifically screening inhibitors of viral entry. The assay system employs a novel dual envelope strategy, using lentiviral pseudovirions as targets whose entry is driven by the SARS-CoV Spike glycoprotein. A second, unrelated viral envelope is used as an internal control to reduce the number of false positives. As an example of the power of this assay a class of inhibitors is reported with the potential to inhibit SARS-CoV at two steps of the replication cycle, viral entry and particle assembly. This assay system can be easily adapted to screen entry inhibitors against other viruses with the careful selection of matching partner virus envelopes

No Evidence for XMRV Nucleic Acids, Infectious Virus or Anti-XMRV Antibodies in Canadian Patients with Chronic Fatigue Syndrome

The gammaretroviruses xenotropic murine leukemia virus (MLV)-related virus (XMRV) and MLV have been reported to be more prevalent in plasma and peripheral blood mononuclear cells of chronic fatigue syndrome (CFS) patients than in healthy controls. Here, we report the complex analysis of whole blood and plasma samples from 58 CFS patients and 57 controls from Canada for the presence of XMRV/MLV nucleic acids, infectious virus, and XMRV/MLV-specific antibodies. Multiple techniques were employed, including nested and qRT-PCR, cell culture, and immunoblotting. We found no evidence of XMRV or MLV in humans and conclude that CFS is not associated with these gammaretroviruses

No Evidence of Murine Leukemia Virus-Related Viruses in Live Attenuated Human Vaccines

The association of xenotropic murine leukemia virus (MLV)-related virus (XMRV) in prostate cancer and chronic fatigue syndrome reported in previous studies remains controversial as these results have been questioned by recent data. Nonetheless, concerns have been raised regarding contamination of human vaccines as a possible source of introduction of XMRV and MLV into human populations. To address this possibility, we tested eight live attenuated human vaccines using generic PCR for XMRV and MLV sequences. Viral metagenomics using deep sequencing was also done to identify the possibility of other adventitious agents.All eight live attenuated vaccines, including Japanese encephalitis virus (JEV) (SA-14-14-2), varicella (Varivax), measles, mumps, and rubella (MMR-II), measles (Attenuvax), rubella (Meruvax-II), rotavirus (Rotateq and Rotarix), and yellow fever virus were negative for XMRV and highly related MLV sequences. However, residual hamster DNA, but not RNA, containing novel endogenous gammaretrovirus sequences was detected in the JEV vaccine using PCR. Metagenomics analysis did not detect any adventitious viral sequences of public health concern. Intracisternal A particle sequences closest to those present in Syrian hamsters and not mice were also detected in the JEV SA-14-14-2 vaccine. Combined, these results are consistent with the production of the JEV vaccine in Syrian hamster cells.We found no evidence of XMRV and MLV in eight live attenuated human vaccines further supporting the safety of these vaccines. Our findings suggest that vaccines are an unlikely source of XMRV and MLV exposure in humans and are consistent with the mounting evidence on the absence of these viruses in humans

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Impact of Gamification on Consumers’ Favorability in Cause-Related Marketing Programs: Between-Subjects Experiments

BackgroundSuccessful cause-related marketing (CRM) campaigns can help companies stand out from their competitors; however, CRM may not have pleasant outcomes, even if it receives substantial investment. ObjectiveThis research aimed to investigate how gamified CRM projects influence consumers’ favorability. MethodsWe introduced 3 different CRM projects in 3 different studies. Every project had 2 versions according to the level of gamification, and participants were randomly assigned into these 2 groups. Additionally, we used a 2 (gamification: lower, higher) 2 (rules presentation: without visual cues, with visual cues) between-subjects design to test the moderation role of rules presentation in gamified CRM projects. ResultsIn Study 1, we identified that the highly gamified CRM program induces more enjoyment (F1,139=21.11, P<.001) and higher favorability (F1,139=14.57, P<.001). Moreover, we found that enjoyment played a mediation role between gamification and favorability (P<.001) in Study 2. In addition, the results of Study 3 indicated rules presentation in a gamified CRM program can moderate the indirect effect of gamification on favorability via enjoyment (index of the moderated mediation: 95% CI –1.12 to –0.10; for rules presentation with visual cues: 95% CI 0.69 to 1.40; for rules presentation without visual cues: 95% CI 0.08 to 0.83). ConclusionsOverall, this research contributes to the CRM literature and suggests gamification is an effective way of managing CRM campaigns

Rubber Identification Based on Blended High Spatio-Temporal Resolution Optical Remote Sensing Data: A Case Study in Xishuangbanna

As an important economic resource, rubber has rapidly grown in Xishuangbanna of Yunnan Province, China, since the 1990s. Tropical rainforests have been replaced by extensive rubber plantations, which has resulted in ecological problems such as the loss of biodiversity and local water shortages. It is vitally important to accurately map the rubber plantations in this region. Although several rubber mapping methods have been proposed, few studies have investigated methods based on optical remote sensing time series data with high spatio-temporal resolution due to the cloudy and foggy weather conditions in this area. This study presented a rubber plantation identification method that used spatio-temporal optical remote sensing data fusion technology to obtain vegetation index data at high spatio-temporal resolution within the optical remote sensing window in Xishuangbanna. The analysis of the proposed method shows that (1) fused optical remote sensing data with high spatio-temporal resolution could map the rubber distribution with high accuracy (overall accuracy of up to 89.51% and kappa of 0.86). (2) Fused indices have high R2 (R2 greater than 0.8, where R is the correlation coefficient) with the indices that were derived from the Landsat observed data, which indicates that fusion results are dependable. However, the fusion accuracy is affected by terrain factors including elevation, slope, and slope aspects. These factors have obvious negative effects on the fusion accuracy of high spatio-temporal resolution optical remote sensing data: the highest fusion accuracy occurred in areas with elevations between 1201 and 1400 m.a.s.l., and the lowest accuracy occurred in areas with elevations less than 600 m.a.s.l. For the 5 fused time series indices (normalized difference vegetation index (NDVI), enhanced vegetation index (EVI), normalized difference moisture index (NDMI), normalized burn ratio (NBR), and tasseled cap angle (TCA)), the fusion accuracy decreased with increasing slope, and increasing slope had the least impact on the EVI, but the greatest negative impact on the NDVI; the slope aspect had a limited influence on the fusion accuracies of the 5 time series indices, but fusion accuracy was lowest on the northwest slope. (3) EVI had the highest accuracy of rubber plantation classification among the 5 time series indices, and the overall classification accuracies of the time series EVI for the four different years (2000, 2005, 2010, and 2015) reached 87.20% (kappa 0.82), 86.91% (kappa 0.81), 88.85% (kappa 0.84), and 89.51% (kappa 0.86), respectively. The results indicate that the method is a promising approach for rubber plantation mapping and the detection of changes in rubber plantations in this tropical area

2D Shape Deformation Using Nonlinear Least Squares Optimization

Abstract This paper presents a novel 2D shape deformation algorithm based on nonlinear least squares optimization. The algorithm aims to preserve two local shape properties: Laplacian coordinates of the boundary curve and local area of the shape interior, which are together represented in a non-quadratic energy function. An iterative Gauss-Newton method is used to minimize this nonlinear energy function. The result is an interactive shape deformation system that can achieve physically plausible results that are hard to achieve with previous linear least squares methods. Besides preserving local shape properties, we also introduce a scheme to preserve the global area of the shape which is useful for deforming incompressible objects