Genetic variation of St. Louis encephalitis virus

St. Louis encephalitis virus (SLEV) has been regularly isolated throughout the Americas since 1933. Previous phylogenetic studies involving 62 isolates have defined seven major lineages (I–VII), further divided into 14 clades. In this study, 28 strains isolated in Texas in 1991 and 2001–2003, and three older, previously unsequenced strains from Jamaica and California were sequenced over the envelope protein gene. The inclusion of these new sequences, and others published since 2001, has allowed better delineation of the previously published SLEV lineages, in particular the clades of lineage II. Phylogenetic analysis of 106 isolates identified 13 clades. All 1991 and 2001–2003 isolates from Nueces, Jefferson and Harris Counties (Texas Gulf Coast) group in clade IIB with other isolates from these counties isolated during the 1980s and 1990s. This lack of evidence for introduction of novel strains into the Texas Gulf Coast over a long period of time is consistent with overwintering of SLEV in this region. Two El Paso isolates, both from 2002, group in clade VA with recent Californian isolates from 1998–2001 and some South American strains with a broad temporal range. Overall, these data are consistent with multiple introductions of SLEV from South America into North America, and provide support for the hypothesis that in most situations, SLEV circulates within a locality, with occasional incursions from other areas. Finally, SLEV has much lower nucleotide (10.1 %) and amino acid variation (2.8 %) than other members of the Japanese encephalitis virus complex (maximum variation 24.6 % nucleotide and 11.8 % amino acid)

Study on Unsaturated Transport of Cement-Based Silane Sol Coating Materials

There are many types of concrete protection materials, but silane-based protective materials have excellent performance and durability. Experimental usage of silane sol-based waterproof materials is relatively mature and research studies on microscale mechanisms are relatively sparse. In this paper, molecular dynamics simulations are used to explain the microscopic transmission mechanism by analyzing the transport of water molecules and siloxane molecules in the gel pores, the local structure at the interface, and the molecular dynamics in the pores. Firstly, four models with different concentrations were constructed (0, 0.3, 0.6, and 0.9 mol/L). By comparison, it can be found that as the concentration increases, so does the effect of inhibiting the transport of water molecules in the pores. Based on the determination of the concentration, this paper corrects the arrangement. Next, the three commonly used silanes in the experiment were selected for simulation. It was found that octyltriethoxysilane has good stability and a waterproof effect. Among them, octyltriethoxysilane has a longer alkyl chain and is more stable at the interface, which destroys the original spatial correlation and weakens the capillary adsorption

Relative Contributions of Different Lifestyle Factors to Health-Related Quality of Life in the Elderly

Much of the previous literature has studied the relationship between individual lifestyle factors and the health-related quality of life (HRQOL). However, only a few studies combined them to explore their relative importance to the HRQOL in the elderly. This study assesses the HRQOL of the urban, rural, and institutionalized Chinese elderly and explores the relative contributions of different lifestyle factors to their HRQOL. The SF-36v2 Health Survey, the WHOQOL-OLD module, and the socio-demographic and lifestyle questionnaire were utilized in this study. Hierarchical regression was performed in order to analyze the results. The physical and mental component scores of the SF-36v2 survey were 47.05 ± 9.95 and 54.92 ± 9.92, respectively. The total score for the WHOQOL-OLD module was 73.01 ± 11.99, with institutionalized persons reporting lower scores. For the physical component of the elderly participants’ HRQOL, the R2 value changed the most (0.116) when exercise-and-labor-related factors were added in. For the mental component, sleep-related (0.054), and leisure-time-activity-related factors (0.053) caused the largest change of the R2 value. For the elderly-specific HRQOL, measured by the WHOQOL-OLD module, the leisure-time-activity-related factors caused the largest change in the R2 value (0.119), followed by exercise-and-labor-related factors (0.078). Heterogeneity was present among the three subgroups. In sum, compared with their community-dwelling counterparts, the HRQOL of institutionalized older people was relatively poor and different lifestyle factors contributed to the HRQOL differently

An in vitro CRISPR-Cas12a-mediated protocol for direct cloning of large DNA fragments

Summary: Large biosynthetic gene cluster (BGC) cloning is important for discovering natural product-based drugs and remains challenging in high GC content microorganisms (e.g., Actinobacteria). Here, we present an in vitro CRISPR-Cas12a-mediated protocol for direct cloning of large DNA fragments. We describe steps for crRNA design and preparation, genomic DNA isolation, and CRISPR-Cas12a cleavage and capture plasmid construction and linearization. We then detail target BGC and plasmid DNA ligation and transformation and screening for positive clones.For complete details on the use and execution of this protocol, please refer to Liang et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics

A Combination of Naturally Occurring Mutations in North American West Nile Virus Nonstructural Protein Genes and in the 3′ Untranslated Region Alters Virus Phenotype▿

We previously reported mutations in North American West Nile viruses (WNVs) with a small-plaque (sp), temperature-sensitive (ts), and/or mouse-attenuated (att) phenotype. Using an infectious clone, site-directed mutations and 3′ untranslated region (3′UTR) exchanges were introduced into the WNV NY99 genome. Characterization of mutants demonstrated that a combination of mutations involving the NS4B protein (E249G) together with either a mutation in the NS5 protein (A804V) or three mutations in the 3′UTR (A10596G, C10774U, A10799G) produced sp, ts, and/or att variants. These results suggested that the discovery of North American WNV-phenotypic variants is rare because of the apparent requirement of concurrent polygenic mutations

Envelope Protein Glycosylation Status Influences Mouse Neuroinvasion Phenotype of Genetic Lineage 1 West Nile Virus Strains

The introduction of West Nile virus (WNV) into North America has been associated with relatively high rates of neurological disease and death in humans, birds, horses, and some other animals. Previous studies identified strains in both genetic lineage 1 and genetic lineage 2, including North American isolates of lineage 1, that were highly virulent in a mouse neuroinvasion model, while other strains were avirulent or significantly attenuated (D. W. C. Beasley, L. Li, M. T. Suderman, and A. D. T. Barrett, Virology 296:17-23, 2002). To begin to elucidate the basis for these differences, we compared a highly virulent New York 1999 (NY99) isolate with a related Old World lineage 1 strain, An4766 (ETH76a), which is attenuated for mouse neuroinvasion. Genomic sequencing of ETH76a revealed a relatively small number of nucleotide (5.1%) and amino acid (0.6%) differences compared with NY99. These differences were located throughout the genome and included five amino acid differences in the envelope protein gene. Substitution of premembrane and envelope genes of ETH76a into a NY99 infectious clone backbone yielded a virus with altered in vitro growth characteristics and a mouse virulence phenotype comparable to ETH76a. Further site-specific mutagenesis studies revealed that the altered phenotype was primarily mediated via loss of envelope protein glycosylation and that this was associated with altered stability of the virion at mildly acidic pH. Therefore, the enhanced virulence of North American WNV strains compared with other Old World lineage 1 strains is at least partly mediated by envelope protein glycosylation