Yeast Based Small Molecule Screen for Inhibitors of SARS-CoV

Severe acute respiratory coronavirus (SARS-CoV) emerged in 2002, resulting in roughly 8000 cases worldwide and 10% mortality. The animal reservoirs for SARS-CoV precursors still exist and the likelihood of future outbreaks in the human population is high. The SARS-CoV papain-like protease (PLP) is an attractive target for pharmaceutical development because it is essential for virus replication and is conserved among human coronaviruses. A yeast-based assay was established for PLP activity that relies on the ability of PLP to induce a pronounced slow-growth phenotype when expressed in S. cerevisiae. Induction of the slow-growth phenotype was shown to take place over a 60-hour time course, providing the basis for conducting a screen for small molecules that restore growth by inhibiting the function of PLP. Five chemical suppressors of the slow-growth phenotype were identified from the 2000 member NIH Diversity Set library. One of these, NSC158362, potently inhibited SARS-CoV replication in cell culture without toxic effects on cells, and it specifically inhibited SARS-CoV replication but not influenza virus replication. The effect of NSC158362 on PLP protease, deubiquitinase and anti-interferon activities was investigated but the compound did not alter these activities. Another suppressor, NSC158011, demonstrated the ability to inhibit PLP protease activity in a cell-based assay. The identification of these inhibitors demonstrated a strong functional connection between the PLP-based yeast assay, the inhibitory compounds, and SARS-CoV biology. Furthermore the data with NSC158362 suggest a novel mechanism for inhibition of SARS-CoV replication that may involve an unknown activity of PLP, or alternatively a direct effect on a cellular target that modifies or bypasses PLP function in yeast and mammalian cells

SPIKE1 is an upstream regulator of the WAVE-ARP2 /3 complexes and is required for epidermal morphogenesis

During cell morphogenesis the actin cytoskeleton controls the polarized distribution of proteins and organelles. ARP2/3 and its activator WAVE are conserved complexes that nucleate actin filaments. In the cell, ARP2/3 functions in plasma membrane recycling, organelle motility and vacuole biogenesis. The Rho-family GTPase, Rac1 activates WAVE complex by directly binding to SRA-1 subunit. Like other eukaryotes, plants have both the complexes. Mutation in the genes encoding subunits of WAVE and ARP2/3 complex in Arabidopsis causes stage-specific swelling and twisting of trichome and thus belongs to the \u27distorted (dis)\u27 class of mutants. Arabidopsis encodes 11 Rho-of-Plants (ROPs), and like their mammalian homologs, ROPs regulate endomembrane and cytoskeleton dynamics in a variety of cell types. However, to-date no GEF (Guanine-nucleotide-exchange-factor) has been identified as an upstream regulator of the ROP-WAVE-ARP2/3 pathway. To identify upstream players of ROP-WAVE-ARP2/3, we used trichome swelling as a mutant screening criterion where the SPIKE1 (SPK1) gene was identified. The C-terminal 300 amino acids of SPK1 encodes a conserved domain, DHR-2, that acts as a GEF for multiple ROPs. GEF activity is an essential component of SPK1 function because the spk1-3 protein that lacks an intact DHR2 domain fails to bind ROP and spk1-3 plants display a severe loss-of-function phenotype. This research will provide genetic and biochemical evidence for a SPK1-ROP-WAVE-ARP2/3 pathway. This is the first evidence of a well-defined pathway from GEF to actin filament nucleation in a multi-cellular organism

Characterization of a hemolytic and antibiotic-resistant Pseudomonas aeruginosa strain S3 pathogenic to fish isolated from Mahananda River in India.

Virulent strain Pseudomonas aeruginosa isolated from Mahananda River exhibited the highest hemolytic activity and virulence factors and was pathogenic to fish as clinical signs of hemorrhagic spots, loss of scales, and fin erosions were found. S3 was cytotoxic to the human liver cell line (WRL-68) in the trypan blue dye exclusion assay. Genotype characterization using whole genome analysis showed that S3 was similar to P. aeruginosa PAO1. The draft genome sequence had an estimated length of 62,69,783 bp, a GC content of 66.3%, and contained 5916 coding sequences. Eight genes across the genome were predicted to be related to hemolysin action. Antibiotic resistance genes such as class C and class D beta-lactamases, fosA, APH, and catB were detected, along with the strong presence of multiple efflux system genes. This study shows that river water is contaminated by pathogenic P. aeruginosa harboring an array of virulence and antibiotic resistance genes which warrants periodic monitoring to prevent disease outbreaks

Efficacy and safety of human papillomavirus vaccine for primary prevention of cervical cancer: A review of evidence from phase III trials and national programs

The Human Papillomavirus (HPV) vaccines have been widely introduced in the national immunization programs in most of the medium and high income countries following endorsement from national and international advisory bodies. HPV vaccine is unique and its introduction is challenging in many ways - it is the first vaccine developed to prevent any cancer, the vaccine is gender specific, it targets adolescent females who are difficult to reach by any health intervention programs. It is not unusual for such a vaccine to face scepticism and reservations not only from lay public but also from professionals in spite of the clinical trial results convincingly and consistently proving their efficacy and safety. Over the last few years millions of doses of the HPV vaccine have been administered round the world and the efficacy and safety data have started coming from the real life programs. A comprehensive cervical cancer control program involving HPV vaccination of the adolescent girls and screening of the adult women has been proved to be the most cost-effective approach to reduce the burden of cervical cancer. The present article discusses the justification of HPV vaccination in the backdrop of natural history of cervical cancer, the mechanism of action of the vaccines, efficacy and safety data from phase III randomized controlled trials as well as from the national immunization programs of various countries

Attitude towards adherence to long term therapy

Background: Chronic disease is a common public health problem worldwide. Adherence to long-term treatment is a key determinant of therapeutic success in patients with chronic diseases. Aim & Objective: The purpose of the study was to know the people’s attitude towards intake of drug, medication adherence and their relationship with socio demographic profile. Settings and Design: Data were collected from 729 individuals chronic disease sufferers chosen randomly from all 81 villages of Amdanga Block West Bengal through a household-based survey in a cross- sectional design. Methods and Material: A pre designed, pre tested, semi structured schedule containing socio-demographic profile and attitude among the respondents regarding adherence to long-term treatment. Statistical analysis used: To compare Attitude scores among different groups, median (IQR) attitude score was calculated and compared with Mann-Whitney U test and Kruskal-Wallis test to know the level of significance of variables. p–value < 0.05 considered statistically significant. Results: Attitude towards long-term adherence to treatment to chronic diseases were significantly associated to caste (p=0.043), education (p=0.001) and occupation (p=0.001) of the study subject. Conclusions: Attitude towards long-term adherence to treatment to chronic diseases were significantly associated to caste, education and occupation

Assessing vegetation fragmentation and plantation efficiency in an intertidal mudflat of Eastern India using Radar Forest Degradation Index and spatial metrics

Potentiality of Synthetic Aperture Radar (SAR) based Radar Forest Degradation Index (RFDI) combined with field observations for monitoring spatio-temporal dynamics of intertidal mudflat vegetation was assessed in this study. Five vegetation zones were delineated in the Junput mudflat of eastern India with very high classification accuracy (Kappa coefficient ≥ 0.79). Fragmentation and coalescence patterns of different vegetation zones under two plantation initiatives were also analysed by different spatial metrics. Results reveal gradual degradation of tree dominated vegetation zones and growth of shrub dominated and herbaceous ones from 2007 to 2019. Plantation of exotic species like Eucalyptus globulus and Casuarina equisetifolia along the shoreline had been found to be less effective against storms and sea surges. Conversely, native mangrove plantations and associated herbs had demonstrated remarkable growth in the intertidal areas. Based on the findings, the study pointed out that a zone-wise cum site-specific plantation strategy is needed towards developing effective bio-shields

Novel Influenza Virus NS1 Antagonists Block Replication and Restore Innate Immune Function▿ †

The innate immune system guards against virus infection through a variety of mechanisms including mobilization of the host interferon system, which attacks viral products mainly at a posttranscriptional level. The influenza virus NS1 protein is a multifunctional facilitator of virus replication, one of whose actions is to antagonize the interferon response. Since NS1 is required for efficient virus replication, it was reasoned that chemical inhibitors of this protein could be used to further understand virus-host interactions and also serve as potential new antiviral agents. A yeast-based assay was developed to identify compounds that phenotypically suppress NS1 function. Several such compounds exhibited significant activity specifically against influenza A virus in cell culture but had no effect on the replication of another RNA virus, respiratory syncytial virus. Interestingly, cells lacking an interferon response were drug resistant, suggesting that the compounds block interactions between NS1 and the interferon system. Accordingly, the compounds reversed the inhibition of beta interferon mRNA induction during infection, which is known to be caused by NS1. In addition, the compounds blocked the ability of NS1 protein to inhibit double-stranded RNA-dependent activation of a transfected beta interferon promoter construct. The effects of the compounds were specific to NS1, because they had no effect on the ability of the severe acute respiratory syndrome coronavirus papainlike protease protein to block beta interferon promoter activation. These data demonstrate that the function of NS1 can be modulated by chemical inhibitors and that such inhibitors will be useful as probes of biological function and as starting points for clinical drug development