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TaqMan real-time PCR assays for single-nucleotide polymorphisms which identify Francisella tularensis and its subspecies and subpopulations
Get PDFFrancisella tularensis, the etiologic agent of tularemia and a Class A Select Agent, is divided into three subspecies and multiple subpopulations that differ in virulence and geographic distribution. Given these differences, there is a need to rapidly and accurately determine if a strain is F. tularensis and, if it is, assign it to subspecies and subpopulation. We designed TaqMan real-time PCR genotyping assays using eleven single nucleotide polymorphisms (SNPs) that were potentially specific to closely related groups within the genus Francisella, including numerous subpopulations within F. tularensis species. We performed extensive validation studies to test the specificity of these SNPs to particular populations by screening the assays across a set of 565 genetically and geographically diverse F. tularensis isolates and an additional 21 genetic near-neighbor (outgroup) isolates. All eleven assays correctly determined the genetic groups of all 565 F. tularensis isolates. One assay differentiates F. tularensis, F. novicida, and F. hispaniensis from the more genetically distant F. philomiragia and Francisella-like endosymbionts. Another assay differentiates F. tularensis isolates from near neighbors. The remaining nine assays classify F. tularensis-confirmed isolates into F. tularensis subspecies and subpopulations. The genotyping accuracy of these nine assays diminished when tested on outgroup isolates (i.e. non F. tularensis), therefore a hierarchical approach of assay usage is recommended wherein the F. tularensis-specific assay is used before the nine downstream assays. Among F. tularensis isolates, all eleven assays were highly sensitive, consistently amplifying very low concentrations of DNA. Altogether, these eleven TaqMan real-time PCR assays represent a highly accurate, rapid, and sensitive means of identifying the species, subspecies, and subpopulation of any F. tularensis isolate if used in a step-wise hierarchical scheme. These assays would be very useful in clinical, epidemiological, and/or forensic investigations involving F. tularensis
Landscape genetic connectivity in a riparian foundation tree is jointly driven by climatic gradients and river networks
Get PDFFremont cottonwood (Populus fremonti) is a foundation riparian tree species that drives community structure and ecosystem processes in southwestern U.S. ecosystems. Despite its ecological importance, little is known about the ecological and environmental processes that shape its genetic diversity, structure, and landscape connectivity. Here, we combined molecular analyses of 82 populations including 1312 individual trees dispersed over the species’ geographical distribution. We reduced the data set to 40 populations and 743 individuals to eliminate admixture with a sibling species, and used multivariate restricted optimization and reciprocal causal modeling to evaluate the effects of river network connectivity and climatic gradients on gene flow. Our results confirmed the following: First, gene flow of Fremont cottonwood is jointly controlled by the connectivity of the river network and gradients of seasonal precipitation. Second, gene flow is facilitated by mid-sized to large rivers, and is resisted by small streams and terrestrial uplands, with resistance to gene flow decreasing with river size. Third, genetic differentiation increases with cumulative differences in winter and spring precipitation. Our results suggest that ongoing fragmentation of riparian habitats will lead to a loss of landscape-level genetic connectivity, leading to increased inbreeding and the concomitant loss of genetic diversity in a foundation species. These genetic effects will cascade to a much larger community of organisms, some of which are threatened and endangered
Method of analysis for determining and correcting mirror deformation due to gravity
Get PDFThe Navy Precision Optical Interferometer, located near Flagstaff, Arizona, is a ground-based interferometer that collects, transports, and modulates stellar radiation from up to six primary flat collectors, known as siderostats, through a common vacuum relay system to a combiner. In the combiner, the modulated beams are superimposed, fringes obtained, and data recorded for further analysis to produce precise star positions or stellar details. The current number of observable stellar objects for the astrometric interferometer can increase from 6000 to at least 47,000 with the addition of full-aperture 20-deg down-tilting beam compressors in each optical train. Such an aperture increase, from the current 12.5 to 35 cm, opens the sky to many additional and fainter stars. Engineering analysis of our beam compressor primary mirror shows that the maximum allowable sag, 21 nm, occurs prematurely at 2.8-deg down-tilt angle. Furthermore, at the operational down-tilt angle of 20 deg, the wavefront deformation increases to 155 nm. We present a finite element analysis technique and design modification concept to reduce tilt-induced deformation on the mirror surface. This work is a first pass to determine the feasibility for a mechanical solution path forward. From this analysis, we found that four outwardly applied 17.8-N forces on the rear surface of the mirror could reduce sag from 155 to 32 nm at 20-deg down-tilt angle
Fact sheet: Long-term responses of Penstemon Clutei (Sunset Crater beardtongue) to root trenching and prescribed fire: Clues for population persistence
Get PDFPenstemon clutei (Sunset Crater beardtongue) is a rare plant species that grows on volcanic soils northeast of Flagstaff in northern Arizona. It is primarily restricted to tephra deposits from the Sunset Crater eruption at an elevation of approximately 7,000 feet and is typically found in open ponderosa pine forests and pinyon-juniper woodlands in areas containing a sparse understory. Its conservation status is ranked "imperiled" (G2) by NatureServe, a non-profit conservation organization, and it is on the U.S. Forest Service sensitive species list for Region 3 (Southwestern Region). Observations in the field suggest a positive link between disturbances (events that disrupt the ecosystem such as fire, logging, tornadoes, and death of trees by bark beetles) and reproduction and colonization. Published information on the ecology of this species is limited to about a half-dozen studies
Does distance impact willingness to pay for forested watershed restoration? A spatial probit analysis: Working paper series--14-06
Get PDFWhile the relationship between ecological restoration and forested watershed health is well established in the literature, funding for restoration remains a significant constraint. Thus, estimates of the benefits of restoration are essential for efficient decision-making. While much research exists estimating the non-market value of wildfire, less research exists estimating the value of forested watershed restoration, and no studies explicitly model spatial variables estimating Willingness to Pay (WTP) for forested watershed restoration. We estimate WTP for forested municipal watershed restoration in Flagstaff, AZ, located in an arid region of the southwestern United States. We find policy-relevant differences in estimated WTP when explicitly incorporating spatial information on the distance to restoration treatment area. Our results indicate that careful consideration of the distance to proposed environmental improvements may be necessary to ensure accurate WTP estimates from dichotomous choice Contingent Valuation models
Genotyping of burkholderia mallei from an outbreak of glanders in bahrain suggests multiple introduction events
Get PDFBackground: Glanders, caused by the gram-negative bacterium Burkholderia mallei, is a highly infectious zoonotic disease of solipeds causing severe disease in animals and men. Although eradicated from many Western countries, it recently emerged in Asia, the Middle-East, Africa, and South America. Due to its rareness, little is known about outbreak dynamics of the disease and its epidemiology. Methodology/Principal Findings: We investigated a recent outbreak of glanders in Bahrain by applying high resolution genotyping (multiple locus variable number of tandem repeats, MLVA) and comparative whole genome sequencing to B. mallei isolated from infected horses and a camel. These results were compared to samples obtained from an outbreak in the United Arab Emirates in 2004, and further placed into a broader phylogeographic context based on previously published B. mallei data. The samples from the outbreak in Bahrain separated into two distinct clusters, suggesting a complex epidemiological background and evidence for the involvement of multiple B. mallei strains. Additionally, the samples from Bahrain were more closely related to B. mallei isolated from horses in the United Arab Emirates in 2004 than other B. mallei which is suggestive of repeated importation to the region from similar geographic sources. Conclusion/Significance: High-resolution genotyping and comparative whole genome analysis revealed the same phylogenetic patterns among our samples. The close relationship of the Dubai/UAE B. mallei populations to each other may be indicative of a similar geographic origin that has yet to be identified for the infecting strains. The recent emergence of glanders in combination with worldwide horse trading might pose a new risk for human infections
Multiple mutations in the para-sodium channel gene are associated with pyrethroid resistance in Rhipicephalus microplus from the United States and Mexico.
Get PDFAcaricide resistant Rhipicephalus microplus populations have become a major problem for many cattle producing areas of the world. Pyrethroid resistance in arthropods is typically associated with mutations in domains I, II, III, and IV of voltage-gated sodium channel genes. In R. microplus, known resistance mutations include a domain II change (C190A) in populations from Australia, Africa, and South America and a domain III mutation (T2134A) that only occurs in Mexico and the U.S. We investigated pyrethroid resistance in cattle fever ticks from Texas and Mexico by estimating resistance levels in field-collected ticks using larval packet discriminating dose (DD) assays and identifying single nucleotide polymorphisms (SNPs) in the para-sodium channel gene that associated with resistance. We then developed qPCR assays for three SNPs and screened a larger set of 1,488 R. microplus ticks, representing 77 field collections and four laboratory strains, for SNP frequency. We detected resistance SNPs in 21 of 68 U.S. field collections and six of nine Mexico field collections. We expected to identify the domain III SNP (T2134A) at a high frequency; however, we only found it in three U.S. collections. A much more common SNP in the U.S. (detected in 19 of 21 field collections) was the C190A domain II mutation, which has never before been reported from North America. We also discovered a novel domain II SNP (T170C) in ten U.S. and two Mexico field collections. The T170C transition mutation has previously been associated with extreme levels of resistance (super-knockdown resistance) in insects. We found a significant correlation (r = 0.81) between the proportion of individuals in field collections that carried any two resistance SNPs and the percent survivorship of F1 larvae from these collections in DD assays. This relationship is accurately predicted by a simple linear regression model (R2 = 0.6635). These findings demonstrate that multiple mutations in the para-sodium channel gene independently associate with pyrethroid resistance in R. microplus ticks, which is likely a consequence of human-induced selection
ERI Spring-Summer 2014 Newsletter
Get PDFEcological Restoration Institute Spring-Summer 2014 Newslette
Final report on cooperative work plan FY2014
Get PDFThis report presents an integrated and coordinated series of actions for $2.4 million awarded to the ERI in Fiscal Years 2013 and 2014 under CFDA 10.694, Southwest Forest Health and Wildfire Prevention for the third and fourth years in a 5-year domestic grant.
The information provided herein reflects our annual progress as of 6/30/2014.
All of the activities (deliverables) summarized in this report respond to land manager and stakeholder requests and needs. The deliverables are informed by best available science and scientific evidence which is translated into the language and product appropriate for the target audience. The ERI actively delivers information using a variety of approaches that includes individual and group presentations and discussions, to printed and electronically accessible fact sheets, short technical reports, longer white papers and management reports, and peer reviewed archival literature
Kinesthetic investigations in the physics classroom
Get PDFCreating investigations that allow students to see physics in their everyday world and to be kinesthetically active outside of the traditional physics classroom can be incredibly engaging and effective. The investigations we developed were inquiry investigations in which students engaged in concrete experiences before we discussed the ab- stract concepts and derived the mathematical relationships. In this article, we describe the approach to inquiry used and an explanation of kinesthetic investigations in general. We then provide a description of several of the investigations and some examples of how students responded to them