Enhanced expression of codon optimized Mycobacterium avium subsp. paratuberculosis antigens in Lactobacillus salivarius

It is well documented that open reading frames containing high GC content show poor expression in A+T rich hosts. Specifically, G+C-rich codon usage is a limiting factor in heterologous expression of Mycobacterium avium subsp. paratuberculosis (MAP) proteins using Lactobacillus salivarius. However, re-engineering opening reading frames through synonymous substitutions can offset codon bias and greatly enhance MAP protein production in this host. In this report, we demonstrate that codon-usage manipulation of MAP2121c can enhance the heterologous expression of the major membrane protein (MMP), analogous to the form in which it is produced natively by MAP bacilli. When heterologously over-expressed, antigenic determinants were preserved in synthetic MMP proteins as shown by monoclonal antibody mediated ELISA. Moreover, MMP is a membrane protein in MAP, which is also targeted to the cellular surface of recombinant L. salivarius at levels comparable to MAP. Additionally, we previously engineered MAP3733c (encoding MptD) and show herein that MptD displays the tendency to associate with the cytoplasmic membrane boundary under confocal microscopy and the intracellularly accumulated protein selectively adheres to the MptD-specific bacteriophage fMptD. This work demonstrates there is potential for L. salivarius as a viable antigen delivery vehicle for MAP, which may provide an effective mucosal vaccine against Johne’s disease

In vivo evaluation of an Ultra-fast Disintegrating Wafer matrix : a molecular simulation approach to the ora-mucoadhesivity

The purpose of this study was to design and evaluate the performance of an Ultra-fast Disintegrating Wafer (U-D-WAF) loaded with highly water soluble diphenhydramine hydrochloride (DPH) through the oramucosa of the Large White Pig model. For the first time this work explored the oramucosivity of the U-D-WAF by detailed molecular modeling of the matrix on buccal tissue in order to mechanistically deduce the mucodhesivity. The U-D-WAF was formulated using a blend of hydroxypropylcellulose, poly(acrylic) acid, sodium starch glycolate and β –cyclodextrin in accordance with a Box-Benkhen experimental design for optimization prior to ex vivo permeation and in vivo release studies in the Large White Pig. Molecular simulation studies assess the mucoadhesivity of the U-D-WAF to the oramucosa. A mean Drug Entrapment Efficiency of 72.96 ± 4.32%, disintegration time of 29.33 ± 15.91 s and drug release after 60 s of 86.32 ± 20.37% was recorded. Ex vivo permeation studies revealed cumulative drug permeation of 86.32 ± 20.34% 60 s after onset. In vivo evaluation of the optimized U-D-WAF had a Cmax = 59 μgL−1 approximately 300 s after administration. The ultrafast disintegration of the U-D-WAF matrix with desirable mucoadhesivity in in vitro and in vivo studies makes it suitable for effective and rapid oramucosal drug delivery.The National Research Foundation (NRF) and the Technology Innovation Agency (TIA) of South Africa.http://www.elsevier.com/locate/jddst2018-02-20hj2018Paraclinical Science

Synthesis and Biological Evaluation of 2-Hydroxy-3-[(2-aryloxyethyl)amino]propyl 4-[(Alkoxycarbonyl)amino]benzoates

A series of twenty substituted 2-hydroxy-3-[(2-aryloxyethyl)amino]propyl 4-[(alkoxycarbonyl)amino]benzoates were prepared and characterized. As similar compounds have been described as potential antimycobacterials, primary in vitro screening of the synthesized carbamates was also performed against two mycobacterial species. 2-Hydroxy-3-[2-(2,6-dimethoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, 2-hydroxy-3-[2-(4-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride, and 2-hydroxy-3-[2-(2-methoxyphenoxy)ethylamino]-propyl 4-(butoxycarbonylamino)benzoate hydrochloride showed higher activity against M. avium subsp. paratuberculosis and M. intracellulare than the standards ciprofloxacin, isoniazid, or pyrazinamide. Cytotoxicity assay of effective compounds was performed using the human monocytic leukaemia THP-1 cell line. Compounds with predicted amphiphilic properties were also tested for their effects on the rate of photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. All butyl derivatives significantly stimulated the rate of PET, indicating that the compounds can induce conformational changes in thylakoid membranes resulting in an increase of their permeability and so causing uncoupling of phosphorylation from electron transport

AIDS-related mycoses: the way forward.

The contribution of fungal infections to the morbidity and mortality of HIV-infected individuals is largely unrecognized. A recent meeting highlighted several priorities that need to be urgently addressed, including improved epidemiological surveillance, increased availability of existing diagnostics and drugs, more training in the field of medical mycology, and better funding for research and provision of treatment, particularly in developing countries

Search for dark matter produced in association with bottom or top quarks in √s = 13 TeV pp collisions with the ATLAS detector

A search for weakly interacting massive particle dark matter produced in association with bottom or top quarks is presented. Final states containing third-generation quarks and miss- ing transverse momentum are considered. The analysis uses 36.1 fb−1 of proton–proton collision data recorded by the ATLAS experiment at √s = 13 TeV in 2015 and 2016. No significant excess of events above the estimated backgrounds is observed. The results are in- terpreted in the framework of simplified models of spin-0 dark-matter mediators. For colour- neutral spin-0 mediators produced in association with top quarks and decaying into a pair of dark-matter particles, mediator masses below 50 GeV are excluded assuming a dark-matter candidate mass of 1 GeV and unitary couplings. For scalar and pseudoscalar mediators produced in association with bottom quarks, the search sets limits on the production cross- section of 300 times the predicted rate for mediators with masses between 10 and 50 GeV and assuming a dark-matter mass of 1 GeV and unitary coupling. Constraints on colour- charged scalar simplified models are also presented. Assuming a dark-matter particle mass of 35 GeV, mediator particles with mass below 1.1 TeV are excluded for couplings yielding a dark-matter relic density consistent with measurements