An extended panel of single nucleotide polymorphisms in Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis> L.) mitochondrial DNA

A total of 6,324 base pairs, distributed on eight different regions of the mitochondrial DNA (mtDNA) were re-sequenced for 32 specimens of Norwegian farmed and wild Atlantic salmon. A total of 16 new and three previously reported single nucleotide polymorphisms (SNPs) were detected. These SNPs were validated using 240 farm and 119 wild Norwegian salmon. Alignments of sequences obtained in this study and previously published sequences revealed another 15 previously unreported SNPs. The panel of mitochondrial SNPs detected in the present study will, in combination with previously identified SNPs, prove useful in designing efficient assays for analyzes of mtDNA variation.     

In‐depth characterization of Trichoderma reesei cellobiohydrolase TrCel7A produced in Nicotiana benthamiana reveals limitations of cellulase production in plants by host‐specific post‐translational modifications

Sustainable production of biofuels from lignocellulose feedstocks depends on cheap enzymes for degradation of such biomass. Plants offer a safe and cost‐effective production platform for biopharmaceuticals, vaccines and industrial enzymes boosting biomass conversion to biofuels. Production of intact and functional protein is a prerequisite for large‐scale protein production, and extensive host‐specific post‐translational modifications (PTMs) often affect the catalytic properties and stability of recombinant enzymes. Here we investigated the impact of plant PTMs on enzyme performance and stability of the major cellobiohydrolase TrCel7A from Trichoderma reesei, an industrially relevant enzyme. TrCel7A was produced in Nicotiana benthamiana using a vacuum‐based transient expression technology, and this recombinant enzyme (TrCel7A^rec) was compared with the native fungal enzyme (TrCel7A^nat) in terms of PTMs and catalytic activity on commercial and industrial substrates. We show that the N‐terminal glutamate of TrCel7A^rec was correctly processed by N. benthamiana to a pyroglutamate, critical for protein structure, while the linker region of TrCel7A^rec was vulnerable to proteolytic digestion during protein production due to the absence of O‐mannosylation in the plant host as compared with the native protein. In general, the purified full‐length TrCel7A^rec had 25% lower catalytic activity than TrCel7A^nat and impaired substrate‐binding properties, which can be attributed to larger N‐glycans and lack of O‐glycans in TrCel7A^rec. All in all, our study reveals that the glycosylation machinery of N. benthamiana needs tailoring to optimize the production of efficient cellulases.     

Nanoparticles of compacted DNA transfect postmitotic cells

Charge-neutral DNA nanoparticles have been developed in which single molecules of DNA are compacted to their minimal possible size. We speculated that the small size of these DNA nanoparticles may facilitate gene transfer in postmitotic cells, permitting nuclear uptake across the 25-nm nuclear membrane pore. To determine whether DNA nanoparticles can transfect nondividing cells, growth-arrested neuroblastoma and hepatoma cells were transfected with DNA/liposome mixtures encoding luciferase. In both models, growth-arrested cells were robustly transfected by compacted DNA (6,900-360-fold more than naked DNA). To evaluate mechanisms responsible for enhanced transfection, HuH-7 cells were microinjected with naked or compacted plasmids encoding enhanced green fluorescent protein. Cytoplasmic microinjection of DNA nanoparticles generated a approximately 10-fold improvement in transgene expression as compared with naked DNA; this enhancement was reversed by the nuclear pore inhibitor, wheat germ agglutinin. To determine the upper size limit for gene transfer, DNA nanoparticles of various sizes were microinjected into the cytoplasm. A marked decrease in transgene expression was observed as the minor ellipsoidal diameter approached 25 nm. In summary, suitably sized DNA nanoparticles productively transfect growth arrested cells by traversing the nuclear membrane pore.     

Ecology,sexual dimorphism,and jumping evolution in anurans

Sexual dimorphism (SD) is a common feature of animals, and selection for sexually dimorphic traits may affect both functional morphological traits and organismal performance. Trait evolution through natural selection can also vary across environments. However, whether the evolution of organismal performance is distinct between the sexes is rarely tested in a phylogenetic comparative context. Anurans commonly exhibit sexual size dimorphism, which may affect jumping performance given the effects of body size on locomotion. They also live in a wide variety of microhabitats. Yet the relationships among dimorphism, performance, and ecology remain underexamined in anurans. Here, we explore relationships between microhabitat use, body size, and jumping performance in males and females to determine the drivers of dimorphic patterns in jumping performance. Using methods for predicting jumping performance through anatomical measurements, we describe how fecundity selection and natural selection associated with body size and microhabitat have likely shaped female jumping performance. We found that the magnitude of sexual size dimorphism (where females are about 14% larger than males) was much lower than dimorphism in muscle volume, where females had 42% more muscle than males (after accounting for body size). Despite these sometimes-large averages, phylogenetic t-tests failed to show the statistical significance of SD for any variable, indicating sexually dimorphic species tend to be closely related. While SD of jumping performance did not vary among microhabitats, we found female jumping velocity and energy differed across microhabitats. Overall, our findings indicate that differences in sex-specific reproductive roles, size, jumping-related morphology, and performance are all important determinants in how selection has led to the incredible ecophenotypic diversity of anurans.     

Survival following renal transplantation in Saskatchewan, 1970--74: follow-up study using medical insurance records.

The patient history file of a medical care insurance plan can be used for statistical purposes in many ways. This is illustrated by the use of the records of the Saskatchewan Medical Care Insurance Commission to study the survival of patients receiving kidney transplants in the period 1970--74. During this period 48 males and 23 females received at least one renal transplant; these patients represented 34% of all males and 28% of all females undergoing regular renal dialysis. These period prevalence estimates are consistent with calculations based on incidence and point prevalence reported elsewhere. Life-table calculations showed the 4-year survival rate following first kidney transplantation in Saskatchewan to be 45% for all patients, 55% for those less than 45 years of age and 26% for those 45 years of age or older.     

Explorative multifactor approach for investigating global survival mechanisms of Campylobacter jejuni under environmental conditions

Explorative approaches such as DNA microarray experiments are becoming increasingly important in microbial research. Despite these major technical advancements, approaches to study multifactor experiments are still lacking. We have addressed this problem by using rotation testing and a novel multivariate analysis of variance (MANOVA) approach (50-50 MANOVA) to investigate interacting experimental factors in a complex experimental design. Furthermore, a new rotation testing based method was introduced to calculate false-discovery rates for each response. This novel analytical concept was used to investigate global survival mechanisms in the environment of the major food-borne pathogen C. jejuni. We simulated nongrowth environmental conditions by investigating combinations of the factors temperature (5 and 25 degrees C) and oxygen tension (anaerobic, microaerobic, and aerobic). Data were generated with DNA microarrays for information about gene expression patterns and Fourier transform infrared (FT-IR) spectroscopy to study global macromolecular changes in the cell. Microarray analyses showed that most genes were either unchanged or down regulated compared to the reference (day 0) for the conditions tested and that the 25 degrees C anaerobic condition gave the most distinct expression pattern with the fewest genes expressed. The few up-regulated genes were generally stress related and/or related to the cell envelope. We found, using FT-IR spectroscopy, that the amount of polysaccharides and oligosaccharides increased under the nongrowth survival conditions. Potential mechanisms for survival could be to down regulate most functions to save energy and to produce polysaccharides and oligosaccharides for protection against harsh environments. Basic knowledge about the survival mechanisms is of fundamental importance in preventing transmission of this bacterium through the food chain.     

The Lao Experience in Deploying Influenza A(H1N1)pdm09 Vaccine: Lessons Made Relevant in Preparing for Present Day Pandemic Threats

The Lao PDR, as did most countries of the Mekong Region, embarked on a pandemic vaccine initiative to counter the threat posed by influenza A(H1N1)pdm09. Overall, estimated vaccine coverage of the Lao population was 14%, with uptake in targeted health care workers and pregnant women 99% and 41%, respectively. Adverse Events Following Immunization accounted for only 6% of survey driven, reported vaccination experiences, with no severe consequences or deaths. Public acceptability of the vaccine campaign was high (98%). Challenges to vaccine deployment included: 1) no previous experience in fielding a seasonal influenza vaccine, 2) safety and efficacy concerns, and 3) late arrival of vaccine 10 months into the pandemic. The Lao success in surmounting these hurdles was in large measure attributed to the oversight assigned the National Immunization Program, and national sensitivities in responding to the avian influenza A(H5N1) crisis in the years leading up to the pandemic. The Lao “lessons learned” from pandemic vaccine deployment are made even more relevant four years on, given the many avian influenza strains circulating in the region, all with pandemic potential.