Efficient gene inactivation in Bacillus anthracis

A procedure for high-efficiency gene inactivation in Bacillus anthracis has been developed. It is based on a highly temperature-sensitive plasmid vector carrying kanamycin resistance cassette surrounded by DNA fragments flanking the desired insertion site. The approach was tested by constructing glutamate racemase E1 (racE1), glutamate racemase E2 (racE2) and comEC knock-out mutants of B. anthracis strain DeltaANR. Allelic replacements were observed at high frequencies, ranging from approximately 0.5% for racE2 up to 50% for racE1 and comEC. The system can be used for genetic validation of potential drug targets.     

Levodopa-induced dyskinesia is associated with increased thyrotropin releasing hormone in the dorsal striatum of hemi-parkinsonian rats

Background

Dyskinesias associated with involuntary movements and painful muscle contractions are a common and severe complication of standard levodopa (L-DOPA, L-3,4-dihydroxyphenylalanine) therapy for Parkinson''s disease. Pathologic neuroplasticity leading to hyper-responsive dopamine receptor signaling in the sensorimotor striatum is thought to underlie this currently untreatable condition.

Methodology/Principal Findings

Quantitative real-time polymerase chain reaction (PCR) was employed to evaluate the molecular changes associated with L-DOPA-induced dyskinesias in Parkinson''s disease. With this technique, we determined that thyrotropin releasing hormone (TRH) was greatly increased in the dopamine-depleted striatum of hemi-parkinsonian rats that developed abnormal movements in response to L-DOPA therapy, relative to the levels measured in the contralateral non-dopamine-depleted striatum, and in the striatum of non-dyskinetic control rats. ProTRH immunostaining suggested that TRH peptide levels were almost absent in the dopamine-depleted striatum of control rats that did not develop dyskinesias, but in the dyskinetic rats, proTRH immunostaining was dramatically up-regulated in the striatum, particularly in the sensorimotor striatum. This up-regulation of TRH peptide affected striatal medium spiny neurons of both the direct and indirect pathways, as well as neurons in striosomes.

Conclusions/Significance

TRH is not known to be a key striatal neuromodulator, but intrastriatal injection of TRH in experimental animals can induce abnormal movements, apparently through increasing dopamine release. Our finding of a dramatic and selective up-regulation of TRH expression in the sensorimotor striatum of dyskinetic rat models suggests a TRH-mediated regulatory mechanism that may underlie the pathologic neuroplasticity driving dopamine hyper-responsivity in Parkinson''s disease.     

Molecular Evidence for High Frequency of Multiple Paternity in a Freshwater Shrimp Species Caridina ensifera

Background

Molecular genetic analyses of parentage provide insights into mating systems. Although there are 22,000 members in Malacostraca, not much has been known about mating systems in Malacostraca. The freshwater shrimp Caridina ensifera blue, is a new species belonging to Malacostraca which was discovered recently in Sulawesi, Indonesia. Due to its small body size and low fecundity, this species is an ideal species to study the occurrence and frequency of multiple paternity and to understand of how the low fecundity species persist and evolve.

Methodology/Principal Findings

In this study, we developed four polymorphic microsatellites from C. ensifera and applied them to investigate the occurrence and frequency of multiple paternity in 20 C. ensifera broods caught from Lake Matano, Sulawesi. By genotyping the mother and all offspring from each brood we discovered multiple paternity in all 20 broods. In most of the 20 broods, fathers contributed skewed numbers of offspring and there was an apparent inverse correlation between reproductive success of sires and their relatedness to mothers.

Conclusions/Significance

Our results in combination with recent reports on multiple paternity in crayfish, crab and lobster species suggests that multiple paternity is common in Malacostraca. Skewed contribution of fathers to the numbers of offspring and inverse correlation between reproductive success of sires and their relatedness to mothers suggest that sperm competition occurred and/or pre- and postcopulatory female choice happen, which may be important for avoiding the occurrence of inbreeding and optimize genetic variation in offspring and for persistence and evolution of low fecundity species.     

A biomimetic robotic jellyfish (Robojelly) actuated by shape memory alloy composite actuators

An analysis is conducted on the design, fabrication and performance of an underwater vehicle mimicking the propulsion mechanism and physical appearance of a medusa (jellyfish). The robotic jellyfish called Robojelly mimics the morphology and kinematics of the Aurelia aurita species. Robojelly actuates using bio-inspired shape memory alloy composite actuators. A systematic fabrication technique was developed to replicate the essential structural features of A. aurita. Robojelly's body was fabricated from RTV silicone having a total mass of 242 g and bell diameter of 164 mm. Robojelly was able to generate enough thrust in static water conditions to propel itself and achieve a proficiency of 0.19 s(-1) while the A. aurita achieves a proficiency of around 0.25 s(-1). A thrust analysis based on empirical measurements for a natural jellyfish was used to compare the performance of the different robotic configurations. The configuration with best performance was a Robojelly with segmented bell and a passive flap structure. Robojelly was found to consume an average power on the order of 17 W with the actuators not having fully reached a thermal steady state.     

Mixed Cation FAxPEA1–xPbI3 with Enhanced Phase and Ambient Stability toward High‐Performance Perovskite Solar Cells

In this work, different from the commonly explored strategy of incorporating a smaller cation, MA⁺ and Cs⁺ into FAPbI₃ lattice to improve efficiency and stability, it is revealed that the introduction of phenylethylammonium iodide (PEAI) into FAPbI₃ perovksite to form mixed cation FA_xPEA_1–xPbI₃ can effectively enhance both phase and ambient stability of FAPbI₃ as well as the resulting performance of the derived devices. From our experimental and theoretical calculation results, it is proposed that the larger PEA cation is capable of assembling on both the lattice surface and grain boundaries to form quais‐3D perovskite structures. The surrounding of PEA⁺ ions at the crystal grain boundaries not only can serve as molecular locks to tighten FAPbI₃ domains but also passivate the surface defects to improve both phase and moisture stablity. Consequently, a high‐performance (PCE:17.7%) and ambient stable FAPbI₃ solar cell could be developed.     

Structure-selected RBM immunogens prime polyclonal memory responses that neutralize SARS-CoV-2 variants of concern

Successful control of the COVID-19 pandemic depends on vaccines that prevent transmission. The full-length Spike protein is highly immunogenic but the majority of antibodies do not target the virus: ACE2 interface. In an effort to affect the quality of the antibody response focusing it to the receptor-binding motif (RBM) we generated a series of conformationally-constrained immunogens by inserting solvent-exposed RBM amino acid residues into hypervariable loops of an immunoglobulin molecule. Priming C57BL/6 mice with plasmid (p)DNA encoding these constructs yielded a rapid memory response to booster immunization with recombinant Spike protein. Immune sera antibodies bound strongly to the purified receptor-binding domain (RBD) and Spike proteins. pDNA primed for a consistent response with antibodies efficient at neutralizing authentic WA1 virus and three variants of concern (VOC), B.1.351, B.1.617.2, and BA.1. We demonstrate that immunogens built on structure selection can be used to influence the quality of the antibody response by focusing it to a conserved site of vulnerability shared between wildtype virus and VOCs, resulting in neutralizing antibodies across variants.