An evaluation of fusion partner proteins for paratransgenesis in Asaia bogorensis.

Mosquitoes transmit many pathogens responsible for human diseases, such as malaria which is caused by parasites in the genus Plasmodium. Current strategies to control vector-transmitted diseases are increasingly undermined by mosquito and pathogen resistance, so additional methods of control are required. Paratransgenesis is a method whereby symbiotic bacteria are genetically modified to affect the mosquito's phenotype by engineering them to deliver effector molecules into the midgut to kill parasites.

An inducible transposon mutagenesis approach for the intracellular human pathogen .

is a prolific human pathogen that can cause serious long-term conditions if left untreated. Recent developments in genetics have opened the door to conducting targeted and random mutagenesis experiments to identify gene function. In the present study, an inducible transposon mutagenesis approach was developed for using a self-replicating vector to deliver the transposon-transposase cassette - a significant step towards our ultimate aim of achieving saturation mutagenesis of the genome.

Progress towards an inducible, replication-proficient transposon delivery vector for .

Genetic systems have been developed for but the extremely low transformation frequency remains a significant bottleneck.  Our goal is to develop a self-replicating transposon delivery vector for which can be expanded prior to transposase induction. We made / shuttle vectors bearing the C9  transposase under control of the promoter and a novel rearrangement of the transposon with the β-lactamase gene.

Blood meal-induced inhibition of vector-borne disease by transgenic microbiota.

Vector-borne diseases are a substantial portion of the global disease burden; one of the deadliest of these is malaria. Vector control strategies have been hindered by mosquito and pathogen resistances, and population alteration approaches using transgenic mosquitos still have many hurdles to overcome before they can be implemented in the field. Here we report a paratransgenic control strategy in which the microbiota of Anopheles stephensi was engineered to produce an antiplasmodial effector causing the mosquito to become refractory to Plasmodium berghei.

Charles Darwin Synthetic Interview: A 19th Century Scientist Speaks in the 21 Century.

Charles Darwin is largely unknown and poorly understood as a historical figure. Similarly the fundamental principles of evolution are often misstated, misunderstood, or entirely rejected by large numbers of Americans. Simply trying to communicate more facts about Darwin, or facts supporting the principles of evaluation, is inadequate; neither students nor members of the public will care or retain the information.

Inhibition of Plasmodium berghei Development in Mosquitoes by Effector Proteins Secreted from Asaia sp. Bacteria Using a Novel Native Secretion Signal.

Novel interventions are needed to prevent the transmission of the Plasmodium parasites that cause malaria. One possible method is to supply mosquitoes with antiplasmodial effector proteins from bacteria by paratransgenesis. Mosquitoes have a diverse complement of midgut microbiota including the Gram-negative bacteria Asaia bogorensis.

Current perspectives on unconventional shale gas extraction in the Appalachian Basin.

The Appalachian Basin is home to three major shales, the Upper Devonian, Marcellus, and Utica. Together, they contain significant quantities of tight oil, gas, and mixed hydrocarbons. The Marcellus alone is estimated to contain upwards of 500 trillion cubic feet of natural gas.

Draft Genome Sequence of Asaia sp. Strain SF2.1, an Important Member of the Microbiome of Anopheles Mosquitoes.

Asaia spp. are abundant members of the microbiota of Anopheles mosquitoes, the principle vectors of malaria. Here, we report the draft genome sequence of Asaia sp.

Fighting malaria with engineered symbiotic bacteria from vector mosquitoes.

The most vulnerable stages of Plasmodium development occur in the lumen of the mosquito midgut, a compartment shared with symbiotic bacteria. Here, we describe a strategy that uses symbiotic bacteria to deliver antimalaria effector molecules to the midgut lumen, thus rendering host mosquitoes refractory to malaria infection. The Escherichia coli hemolysin A secretion system was used to promote the secretion of a variety of anti-Plasmodium effector proteins by Pantoea agglomerans, a common mosquito symbiotic bacterium.

Ribosome display of combinatorial antibody libraries derived from mice immunized with heat-killed Xylella fastidiosa and the selection of MopB-specific single-chain antibodies.

Pierce's disease is a devastating lethal disease of Vitus vinifera grapevines caused by the bacterium Xylella fastidiosa. There is no cure for Pierce's disease, and control is achieved predominantly by suppressing transmission of the glassy-winged sharpshooter insect vector. We present a simple robust approach for the generation of panels of recombinant single-chain antibodies against the surface-exposed elements of X.

Secretion of anti-Plasmodium effector proteins from a natural Pantoea agglomerans isolate by using PelB and HlyA secretion signals.

The insect-vectored disease malaria is a major world health problem. New control strategies are needed to supplement the current use of insecticides and medications. A genetic approach can be used to inhibit development of malaria parasites (Plasmodium spp.

Bacterial genetic methods to explore the biology of mariner transposons.

Mariners are small DNA mediated transposons of eukaryotes that fortuitously function in bacteria. Using bacterial genetics, it is possible to study a variety of properties of mariners, including transpositional ability, dominant-negative regulation, overexpresson inhibition, and the function of cis-acting sequences like the inverted terminal repeats. In conjunction with biochemical techniques, the structure of the transposase can be elucidated and the activity of the elements can be improved for genetic tool use.

Technological advances to enhance agricultural pest management.

Biotechnology offers new solutions to existing and future pest problems in agriculture including, for the first time, possible tools to use against insect transmitted pathogens causing plant diseases. Here, we describe the strategy first described as Autocidal Biological Control applied for the development of conditional lethal pink bollworm strains. When these strains are mass-reared, the lethal gene expression is suppressed by a tetracycline repressor element, which is activated by the presence of chlorotetracycline, a normal component of the mass-rearing diet.

Hyperactive Himar1 transposase mediates transposition in cell culture and enhances gene expression in vivo.

The use of nonviral delivery systems results in transient gene expression, in part because of the low efficiency of DNA integration. Previously, vectors based on transposon systems such as Sleeping Beauty have been shown to be able to increase stable transfection efficiencies in cell culture and in animal models. Himar1, a reconstructed active transposon belonging to the Tc1/mariner superfamily, also has been used as a vector for stable gene delivery, but the rate of transposition after transfection is low.

The N-terminus of Himar1 mariner transposase mediates multiple activities during transposition.

Mariner family transposons are perhaps the most widespread transposable elements of eukaryotes. While we are beginning to understand the precise mechanism of transposition of these elements, the structure of their transposases are still poorly understood. We undertook an extensive mutagenesis of the N-terminal third of the transposase of the Himar1 mariner transposon to begin the process of determining the structure and evolution of mariner transposases.

Early intermediates of mariner transposition: catalysis without synapsis of the transposon ends suggests a novel architecture of the synaptic complex.

The mariner family is probably the most widely distributed family of transposons in nature. Although these transposons are related to the well-studied bacterial insertion elements, there is evidence for major differences in their reaction mechanisms. We report the identification and characterization of complexes that contain the Himar1 transposase bound to a single transposon end.

A bacterial genetic screen identifies functional coding sequences of the insect mariner transposable element Famar1 amplified from the genome of the earwig, Forficula auricularia.

Transposons of the mariner family are widespread in animal genomes and have apparently infected them by horizontal transfer. Most species carry only old defective copies of particular mariner transposons that have diverged greatly from their active horizontally transferred ancestor, while a few contain young, very similar, and active copies. We report here the use of a whole-genome screen in bacteria to isolate somewhat diverged Famar1 copies from the European earwig, Forficula auricularia, that encode functional transposases.

Recent horizontal transfer of mellifera subfamily mariner transposons into insect lineages representing four different orders shows that selection acts only during horizontal transfer.

We report the isolation and sequencing of genomic copies of mariner transposons involved in recent horizontal transfers into the genomes of the European earwig, Forficula auricularia; the European honey bee, Apis mellifera; the Mediterranean fruit fly, Ceratitis capitata; and a blister beetle, Epicauta funebris, insects from four different orders. These elements are in the mellifera subfamily and are the second documented example of full-length mariner elements involved in this kind of phenomenon. We applied maximum likelihood methods to the coding sequences and determined that the copies in each genome were evolving neutrally, whereas reconstructed ancestral coding sequences appeared to be under selection, which strengthens our previous hypothesis that the primary selective constraint on mariner sequence evolution is the act of horizontal transfer between genomes.