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Background: Shigellosis is the leading cause of diarrheal deaths worldwide and is particularly dangerous in children under 5 years of age in low- and middle-income countries. Additionally, the rise in antibiotic resistance has highlighted the need for an effective Shigella vaccine. Previously, we have used the Multiple Antigen-Presenting System (MAPS) technology to generate monovalent and quadrivalent Salmonella MAPS vaccines that induce functional antibodies against Salmonella components. Methods: In this work, we detail the development of several monovalent vaccines using O-specific polysaccharides (OSPs) from four dominant serotypes, S. flexneri 2a, 3a, and 6, and S. sonnei. We tested several rhizavidin (rhavi) fusion proteins and selected a Shigella-specific protein IpaB. Quadrivalent MAPS were made with Rhavi-IpaB protein and tested in rabbits for immunogenicity. Results: Individual MAPS vaccines generated robust, functional antibody responses against both IpaB and the individual OSP component. Antibodies to IpaB were effective across Shigella serotypes. We also demonstrate that the OSP antibodies generated are specific to each homologous Shigella O type by performing ELISA and bactericidal assays. We combined the components of each MAPS vaccine to formulate a quadrivalent MAPS vaccine which elicited similar antibody and bactericidal responses compared to their monovalent counterparts. Finally, we show that the quadrivalent MAPS immune sera are functional against several clinical isolates of the serotypes used in the vaccine. Conclusions: This quadrivalent MAPS Shigella vaccine is immunogenicity and warrants further study.

Bloodstream infections in low- and middle-income countries (LMICs) are most frequently attributed to invasive Salmonella disease caused by four primary serovars of Salmonella enterica: Typhi, Paratyphi A, Typhimurium, and Enteritidis. We showed previously that a bivalent vaccine targeting S. Typhi and S. Paratyphi A using a Multiple Antigen-Presenting System (MAPS) induced functional antibodies against S. Typhi and S. Paratyphi. In the current study, we describe the preclinical development of a first candidate quadrivalent combination Salmonella vaccine with the potential to cover all four leading invasive Salmonella serotypes. We showed that the quadrivalent Salmonella MAPS vaccine, containing Vi from S. Typhi, O-specific Polysaccharide (OSP) from S. Paratyphi A, S. Enteritidis and S. Typhimurium, combined with the Salmonella-specific protein SseB, elicits robust and functional antibody responses to each of the components of the vaccine. Our data indicates that the application of MAPS technology to the development of vaccines targeting invasive forms of Salmonella is practical and merits additional consideration.

Infections by Salmonella Typhi and Paratyphi A strain are still a major cause of morbidity and mortality in developing countries. Generation of antibodies against the Vi capsular polysaccharide of S. Typhi via either pure polysaccharide or protein–polysaccharide conjugate is a very effective way to protect against S. Typhi. To date, there is no commercially available vaccine against S. Paratyphi A. The O-specific polysaccharide (OSP) has been generally considered a good vaccine target for Paratyphi A. Here, a bivalent vaccine against Vi and OSP was generated using the Multiple Antigen Presenting System (MAPS). Three different protein constructs, including CRM197, rEPA of Pseudomonas, and a pneumococcal fusion protein SP1500-SP0785, were fused to Rhizavidin (Rhavi) and evaluated their impact on immunogenicity when incorporated as fusion proteins affinity-bound to the two polysaccharides. We compared the antibody responses, antibody avidity, and cidal activity of sera post-immunization with monovalent vs. combination vaccines. We also wished to evaluate the generation of Vi-specific memory B cells in mice. We found little interference when combination vaccine was compared to monovalent vaccines with respect to antibody concentration and cidal activity of sera. Significant affinity maturation was noted for both Vi and OSP antigens. Thus, our preclinical results with a combination Vi- and OSP-MAPS vaccine strongly support the feasibility of this approach and its application of this approach to other important salmonella and Shigella species.

The authors would like to update the XPS spectrum in Figure 3c [...]

We investigate the possibilities to realize light extraction from single crystal diamond (SCD) nanopillars. This was achieved by dedicated 519 nm laser-induced spin-state initiation of negatively charged nitrogen vacancies (NV−). We focus on the naturally-generated by chemical vapor deposition (CVD) growth of NV−. Applied diamond was neither implanted with 14N+, nor was the CVD synthesized SCD annealed. To investigate the possibility of light extraction by the utilization of NV−’s bright photoluminescence at room temperature and ambient conditions with the waveguiding effect, we have performed a top-down nanofabrication of SCD by electron beam lithography (EBL) and dry inductively-coupled plasma/reactive ion etching (ICP-RIE) to generate light focusing nanopillars. In addition, we have fluorinated the diamond’s surface by dedicated 0 V SF6 ICP plasma. Light extraction and spin manipulations were performed with photoluminescence (PL) spectroscopy and optically detected magnetic resonance (ODMR) at room temperature. We have observed a remarkable effect based on the selective 0 V SF6 plasma etching and surprisingly, in contrast to literature findings, deactivation of NV− centers. We discuss the possible deactivation mechanism in detail.

A simple and inexpensive method using planar electrodes was proposed for the measurement of the imaginary part of the Clausius-Mossotti factor, K i , of particle/cell for electrorotation (ER) and travelling wave dielectrophoresis (twDEP). It is based on the balance between the dielectrophoretic and viscous torques on a particle undergoing ER subject to dual frequency operation in an ER chamber. A four-phase ac voltage signal with a given frequency is applied for generating ER for measurement, and another two-phase signal is applied at a selected frequency for generating a negative dielectrophoretic force for confining the particle motion, instead of using laser tweezer or three-dimensional electrodes in the literature. Both frequencies can be applied to the same electrodes in a four-electrode ER system and to alternative different electrodes in an eight-electrode ER system, and both systems are capable for providing accurate measurement. The measurements were validated by comparing with the theoretical result using sephadex particles in KCl solution, and with the existing experimental results for various human cancer cells in medium with conductivity from 0.01−1.2 S/m, using ER with optical tweezer and dual frequency twDEP. Contrast between the ER and the twDEP methods (the current two available methods) was discussed and commented. The present method could provide measurement for wider frequency range and more accurate result near K i = 0, in comparison with the results using the twDEP method. However, the twDEP method could perform much more rapid measurement. Detailed forces and torque were calculated inside the ER chamber for understanding the physics and assessing the characteristics of the dual frequency ER method. This study is of academic interest as the torque in ER and the force in twDEP can be calculated only when K i is known. It also finds biomedical applications as the K i -spectra can be served as physical phenotypes for different cells, and can be applied for deriving dielectric properties of cells.

In contrast to the delicate 3D electrodes in the literature, a simple flow-through device is proposed here for continuous and massive lysis of cells using electricity. The device is essentially a rectangular microchannel with a planar electrode array built on its bottom wall, actuated by alternating current (AC) voltages between neighboring electrodes, and can be incorporated easily into other biomedical systems. Human whole blood diluted 10 times with phosphate-buffered saline (about 6 × 108 cells per mL) was pumped through the device, and the cells were completely lysed within 7 s after the application of a 20 V peak-to-peak voltage at 1 MHz, up to 400 μL/hr. Electric field and Maxwell stress were calculated for assessing electrical lysis. Only the lower half-channel was exposed to an electric field exceeding the irreversible threshold value of cell electroporation (Eth2), suggesting that a cross flow, proposed here primarily as the electro-thermally induced flow, was responsible for bringing the cells in the upper half-channel downward to the lower half-channel. The Maxwell shear stress associated with Eth2 was one order of magnitude less than the threshold mechanical stresses for lysis, implying that an applied moderate mechanical stress could aid electrical lysis.

A new coumarin-based fluorescent probe, containing an allylic esters group, has been designed and synthesized for sensing cysteine in physiological pH. In this fluorescent probe, the coumarin was applied as the fluorophore and an allylic esters group was combined as both a fluorescence quencher and a recognition unit. The probe can selectively and sensitively detect cysteine (Cys) over homocysteine, glutathione, and other amino acids, and has a rapid response time of 30 min and a low detection limit of 47.7 nM. In addition, the probe could be applied for cell imaging with low cytotoxicity.