Low Cost Tuberculosis Vaccine Antigens in Capsules: Expression in Chloroplasts, Bio-Encapsulation, Stability and Functional Evaluation In Vitro

Tuberculosis (TB) caused by Mycobacterium tuberculosis is one of the leading fatal infectious diseases. The development of TB vaccines has been recognized as a major public health priority by the World Health Organization. In this study, three candidate antigens, ESAT-6 (6kDa early secretory antigenic target) and Mtb72F (a fusion polyprotein from two TB antigens, Mtb32 and Mtb39) fused with cholera toxin B-subunit (CTB) and LipY (a cell wall protein) were expressed in tobacco and/or lettuce chloroplasts to facilitate bioencapsulation/oral delivery. Site-specific transgene integration into the chloroplast genome was confirmed by Southern blot analysis. In transplastomic leaves, CTB fusion proteins existed in soluble monomeric or multimeric forms of expected sizes and their expression levels varied depending upon the developmental stage and time of leaf harvest, with the highest-level of accumulation in mature leaves harvested at 6PM. The CTB-ESAT6 and CTB-Mtb72F expression levels reached up to 7.5% and 1.2% of total soluble protein respectively in mature tobacco leaves. Transplastomic CTB-ESAT6 lettuce plants accumulated up to 0.75% of total leaf protein. Western blot analysis of lyophilized lettuce leaves stored at room temperature for up to six months showed that the CTB-ESAT6 fusion protein was stable and preserved proper folding, disulfide bonds and assembly into pentamers for prolonged periods. Also, antigen concentration per gram of leaf tissue was increased 22 fold after lyophilization. Hemolysis assay with purified CTB-ESAT6 protein showed partial hemolysis of red blood cells and confirmed functionality of the ESAT-6 antigen. GM1-binding assay demonstrated that the CTB-ESAT6 fusion protein formed pentamers to bind with the GM1-ganglioside receptor. The expression of functional Mycobacterium tuberculosis antigens in transplastomic plants should facilitate development of a cost-effective and orally deliverable TB booster vaccine with potential for long-term storage at room temperature. To our knowledge, this is the first report of expression of TB vaccine antigens in chloroplasts

Measurement of the Bottom-Strange Meson Mixing Phase in the Full CDF Data Set

We report a measurement of the bottom-strange meson mixing phase \beta_s using the time evolution of B0_s -> J/\psi (->\mu+\mu-) \phi (-> K+ K-) decays in which the quark-flavor content of the bottom-strange meson is identified at production. This measurement uses the full data set of proton-antiproton collisions at sqrt(s)= 1.96 TeV collected by the Collider Detector experiment at the Fermilab Tevatron, corresponding to 9.6 fb-1 of integrated luminosity. We report confidence regions in the two-dimensional space of \beta_s and the B0_s decay-width difference \Delta\Gamma_s, and measure \beta_s in [-\pi/2, -1.51] U [-0.06, 0.30] U [1.26, \pi/2] at the 68% confidence level, in agreement with the standard model expectation. Assuming the standard model value of \beta_s, we also determine \Delta\Gamma_s = 0.068 +- 0.026 (stat) +- 0.009 (syst) ps-1 and the mean B0_s lifetime, \tau_s = 1.528 +- 0.019 (stat) +- 0.009 (syst) ps, which are consistent and competitive with determinations by other experiments.Comment: 8 pages, 2 figures, Phys. Rev. Lett 109, 171802 (2012

Quorum sensing:Implications on rhamnolipid biosurfactant production

Quorum sensing (QS) has received significant attention in the past few decades. QS describes population density dependent cell to cell communication in bacteria using diffusible signal molecules. These signal molecules produced by bacterial cells, regulate various physiological processes important for social behavior and pathogenesis. One such process regulated by quorum sensing molecules is the production of a biosurfactant, rhamnolipid. Rhamnolipids are important microbially derived surface active agents produced by Pseudomonas spp. under the control of two interrelated quorum sensing systems; namely las and rhl. Rhamnolipids possess antibacterial, antifungal and antiviral properties. They are important in motility, cell to cell interactions, cellular differentiation and formation of water channels that Currently, biosurfactants are unable to compete economically with chemically synthesized compounds in the market due to high production costs. Once the genes required for biosurfactant production have been identified, they can be placed under the regulation of strong promoters in nonpathogenic, heterologous hosts to enhance production. The production of rhamnolipids could be increased by cloning both the rhlAB rhamnosyltransferase genes and the rhlRI quorum sensing system into a suitable bacterium such as E. coli or P. putida and facilitate rhamnolipid production. Biosurfactants can also be genetically engineered for different industrial applications assuming there is a strong understanding of both the genetics and the structure-function relationships of each component of the molecule. Genetic engineering of surfactin has already been reported, with recent papers describing the creation of novel peptide structures from the genetic recombination of several peptide synthetases. Recent application of dynamic metabolic engineering strategies for controlled gene expression could lower the cost of fermentation processes by increasing the product formation. Therefore, by integrating a genetic circuit into applications of metabolic engineering the biochemical production can be optimized. Furthermore, novel strategies could be designed on the basis of information obtained from the studies of quorum sensing and biosurfactants produced suggesting enormous practical applications.</p

Observation of the Y(4140)Y(4140) structure in the J/ψ ϕJ/\psi\,\phi Mass Spectrum in B±→J/ψ ϕKB^\pm\to J/\psi\,\phi K cays

The observation of the $Y(4140)$ structure in $B^\pm\rightarrow J/\psi\,\phi K^\pm$ decays produced in $\bar{p} p$ collisions at \sqrt{s}=1.96~\TeV is reported with a statistical significance greater than 5 standard deviations. A fit to the $J/\psi\,\phi$ mass spectrum is performed assuming the presence of a Breit-Wigner resonance. The fit yields a signal of $19^{+6}_{-5}$ resonance events, and resonance mass and width of 4143.4^{+2.9}_{-3.0}(\mathrm{stat})\pm0.6(\mathrm{syst})~\MeVcc and 15.3^{+10.4}_{-6.1}(\mathrm{stat})\pm2.5(\mathrm{syst})~\MeVcc respectively. The parameters of this resonance-like structure are consistent with values reported from an earlier CDF analysis.Comment: 7 pages, 2 figures, submited to Phys. Rev. Let

Search for Higgs bosons produced in association with b quarks

We present a search for neutral Higgs bosons decaying into bb̄, produced in association with b quarks in pp̄ collisions. This process could be observable in supersymmetric models with high values of tanβ. The event sample corresponds to 2.6fb -1 of integrated luminosity collected with the CDF II detector at the Fermilab Tevatron collider. We search for an enhancement in the mass of the two leading jets in events with three jets identified as coming from b quarks using a displaced vertex algorithm. A data-driven procedure is used to estimate the dijet mass spectrum of the nonresonant multijet background. The contributions of backgrounds and a possible Higgs boson signal are determined by a two-dimensional fit of the data, using the dijet mass together with an additional variable which is sensitive to the flavor composition of the three tagged jets. We set mass-dependent limits on σ(pp̄→)×B(→bb̄) which are applicable for a narrow scalar particle produced in association with b quarks. We also set limits on tanβ in supersymmetric Higgs models including the effects of the Higgs boson width. © 2012 American Physical Society

Bi-allelic loss-of-function CACNA1B mutations in progressive epilepsy-dyskinesia

The occurrence of non-epileptic hyperkinetic movements in the context of developmental epileptic encephalopathies is an increasingly recognized phenomenon. Identification of causative mutations provides an important insight into common pathogenic mechanisms that cause both seizures and abnormal motor control. We report bi-allelic loss-of-function CACNA1B variants in six children from three unrelated families whose affected members present with a complex and progressive neurological syndrome. All affected individuals presented with epileptic encephalopathy, severe neurodevelopmental delay (often with regression), and a hyperkinetic movement disorder. Additional neurological features included postnatal microcephaly and hypotonia. Five children died in childhood or adolescence (mean age of death: 9 years), mainly as a result of secondary respiratory complications. CACNA1B encodes the pore-forming subunit of the pre-synaptic neuronal voltage-gated calcium channel Cav2.2/N-type, crucial for SNARE-mediated neurotransmission, particularly in the early postnatal period. Bi-allelic loss-of-function variants in CACNA1B are predicted to cause disruption of Ca2+ influx, leading to impaired synaptic neurotransmission. The resultant effect on neuronal function is likely to be important in the development of involuntary movements and epilepsy. Overall, our findings provide further evidence for the key role of Cav2.2 in normal human neurodevelopment