Synthetic and Evolutionary Construction of a Chlorate-Reducing Shewanella oneidensis MR-1.

UnlabelledDespite evidence for the prevalence of horizontal gene transfer of respiratory genes, little is known about how pathways functionally integrate within new hosts. One example of a mobile respiratory metabolism is bacterial chlorate reduction, which is frequently encoded on composite transposons. This implies that the essential components of the metabolism are encoded on these mobile elements. To test this, we heterologously expressed genes for chlorate reduction from Shewanella algae ACDC in the non-chlorate-reducing Shewanella oneidensis MR-1. The construct that ultimately endowed robust growth on chlorate included cld, a cytochrome c gene, clrABDC, and two genes of unknown function. Although strain MR-1 was unable to grow on chlorate after initial insertion of these genes into the chromosome, 11 derived strains capable of chlorate respiration were obtained through adaptive evolution. Genome resequencing indicated that all of the evolved chlorate-reducing strains replicated a large genomic region containing chlorate reduction genes. Contraction in copy number and loss of the ability to reduce chlorate were also observed, indicating that this phenomenon was extremely dynamic. Although most strains contained more than six copies of the replicated region, a single strain with less duplication also grew rapidly. This strain contained three additional mutations that we hypothesized compensated for the low copy number. We remade the mutations combinatorially in the unevolved strain and determined that a single nucleotide polymorphism (SNP) upstream of cld enabled growth on chlorate and was epistatic to a second base pair change in the NarP binding sequence between narQP and nrfA that enhanced growth.ImportanceThe ability of chlorate reduction composite transposons to form functional metabolisms after transfer to a new host is an important part of their propagation. To study this phenomenon, we engineered Shewanella oneidensis MR-1 into a chlorate reducer. We defined a set of genes sufficient to endow growth on chlorate from a plasmid, but found that chromosomal insertion of these genes was nonfunctional. Evolution of this inoperative strain into a chlorate reducer showed that tandem duplication was a dominant mechanism of activation. While copy number changes are a relatively rapid way of increasing gene dosage, replicating almost 1 megabase of extra DNA is costly. Mutations that alleviate the need for high copy number are expected to arise and eventually predominate, and we identified a single nucleotide polymorphism (SNP) that relieved the copy number requirement. This study uses both rational and evolutionary approaches to gain insight into the evolution of a fascinating respiratory metabolism

A Francisella tularensis Live Vaccine Strain That Improves Stimulation of Antigen-Presenting Cells Does Not Enhance Vaccine Efficacy

Vaccination is a proven strategy to mitigate morbidity and mortality of infectious diseases. The methodology of identifying and testing new vaccine candidates could be improved with rational design and in vitro testing prior to animal experimentation. The tularemia vaccine, Francisella tularensis live vaccine strain (LVS), does not elicit complete protection against lethal challenge with a virulent type A Francisella strain. One factor that may contribute to this poor performance is limited stimulation of antigen-presenting cells. In this study, we examined whether the interaction of genetically modified LVS strains with human antigen-presenting cells correlated with effectiveness as tularemia vaccine candidates. Human dendritic cells infected with wild-type LVS secrete low levels of proinflammatory cytokines, fail to upregulate costimulatory molecules, and activate human T cells poorly in vitro. One LVS mutant, strain 13B47, stimulated higher levels of proinflammatory cytokines from dendritic cells and macrophages and increased costimulatory molecule expression on dendritic cells compared to wild type. Additionally, 13B47-infected dendritic cells activated T cells more efficiently than LVS-infected cells. A deletion allele of the same gene in LVS displayed similar in vitro characteristics, but vaccination with this strain did not improve survival after challenge with a virulent Francisella strain. In vivo, this mutant was attenuated for growth and did not stimulate T cell responses in the lung comparable to wild type. Therefore, stimulation of antigen-presenting cells in vitro was improved by genetic modification of LVS, but did not correlate with efficacy against challenge in vivo within this model system

Virtual embedded librarianship for information literacy teaching.

This paper, reports on the planning and preliminary results of an action research project undertaken for the redesign of an online distance learning information literacy (IL) module on the basis of virtual 'embedded librarianship'. The research project, which followed an action research design, brought together the IL module coordinator and an Academic Liaison Librarian, working at different institutions to collaboratively redesign the assessment and teaching of the module. Data were collected via a qualitative analysis of students' work and a series of open-ended questions addressed to students on the value of the approach followed. Students reacted positively to the embedded librarianship design and engaged constructively in situated learning. Challenges included time-zones differences, the contribution level of students and lack of confidence. The paper puts emphasis on educating future information professionals as embedded information literacy partners, promoting the development of transferable skills and a collaborative/sharing online working ethos

Pattern Transfer of Sub-10 nm Features via Tin-Containing Block Copolymers

Tin-containing block copolymers were investigated as materials for nanolithographic applications. Poly(4-trimethylstannylstyrene-block-styrene) (PSnS-PS) and poly(4-trimethylstannylstyrene-block-4-methoxystyrene) (PSnS-PMOST) synthesized by reversible addition–fragmentation chain transfer polymerization form lamellar domains with periodicities ranging from 18 to 34 nm. Thin film orientation control was achieved by thermal annealing between a neutral surface treatment and a top coat. Incorporation of tin into one block facilitates pattern transfer into SiO_2 via a two-step etch process utilizing oxidative and fluorine-based etch chemistries

Age and growth estimates for the nurse shark (Ginglymostoma cirratum) over 17 years in Bimini, The Bahamas

Age and growth estimates are essential for life history modeling in elasmobranchs and are used to inform accurate conservation and management decisions. The nurse shark (Ginglymostoma cirratum) is abundant in coastal waters of the Atlantic Ocean, yet many aspects of their life history remain relatively understudied, aside from their reproductive behavior. We used mark-recapture data of 91 individual G. cirratum from Bimini, The Bahamas, from 2003 to 2020, to calculate von Bertalanffy (vB) growth parameters, empirical growth rate, and age derived from the resulting length-at-age estimates. The Fabens method for estimating growth from mark-recapture methods was applied through a Bayesian framework using Markov chain Monte Carlo (MCMC) methods. This provided growth parameters with an asymptotic total length (L∞) of 303.28 cm and a growth coefficient (k) of 0.04 yr-1. The average growth rate for G. cirratum was approximately 8.68 ± 6.00 cm yr-1. This study also suggests that the previous maximum age for G. cirratum is likely underestimated, with the oldest individual predicted to be 43 years old. Our study is the first to present vB growth parameters and a growth curve for G. cirratum. It indicates that this species is slow-growing and long-lived, which improves our understanding of their life history

The role of host PrP in Transmissible Spongiform Encephalopathies

AbstractPrP has a central role in the Transmissible Spongiform Encephalopathies (TSEs), and mutations and polymorphisms in host PrP can profoundly alter the host's susceptibility to a TSE agent. However, precisely how host PrP influences the outcome of disease has not been established. To investigate this we have produced by gene targeting a series of inbred lines of transgenic mice expressing different PrP genes. This allows us to study directly the influence of the host PrP gene in TSEs. We have examined the role of glycosylation, point mutations, polymorphisms and PrP from different species on host susceptibility and the disease process both within the murine species and across species barriers

Interannual variability of primary production and dissolved organic nitrogen storage in the North Pacific Subtropical Gyre

Author Posting. © American Geophysical Union, 2012. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research 117 (2012): G03019, doi:10.1029/2011JG001830.The upper ocean primary production measurements from the Hawaii Ocean Time series (HOT) at Station ALOHA in the North Pacific Subtropical Gyre showed substantial variability over the last two decades. The annual average primary production varied within a limited range over 1991–1998, significantly increased in 1999–2000 and then gradually decreased afterwards. This variability was investigated using a one-dimensional ecosystem model. The long-term HOT observations were used to constrain the model by prescribing physical forcings and lower boundary conditions and optimizing the model parameters against data using data assimilation. The model reproduced the general interannual pattern in the observed primary production, and mesoscale variability in vertical velocity was identified as a major contributing factor to the interannual variability in the simulation. Several strong upwelling events occurred in 1999, which brought up nitrate at rates several times higher than other years and elevated the model primary production. Our model results suggested a hypothesis for the observed interannual variability pattern of primary production at Station ALOHA: Part of the upwelled nitrate input in 1999 was converted to and accumulated as semilabile dissolved organic nitrogen (DON), and subsequent recycling of this semilabile DON supported enhanced primary productivity for the next several years as the semilabile DON perturbation was gradually removed via export.This work was supported in part by the Center for Microbial Oceanography, Research and Education (C-MORE) (NSF EF-0424599), Hawaii Ocean Time series program (NSF OCE09–26766), the Gordon and Betty Moore Foundation, and the Marine Biological Laboratory.2013-03-1