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Quinones-Garcia, S.

Publications and source records attributed to Quinones-Garcia, S..

2 recordsLinked to original sources

A recombinant SARS-CoV-2 RBD antigen expressed in insect cells elicits immunogenicity and confirms safety in animal models

COVID-19 pandemic has accelerated the development of vaccines against its etiologic agent, SARS-CoV-2. However, the emergence of new variants of the virus requires new immunization strategies in addition to the current vaccines approved for human administration. In the present report, the immunological and safety evaluation in mice and hamsters of a subunit vaccine based on the RBD sub-domain with two adjuvants of oil origin is described. The RBD protein was expressed in insect cells and purified by chromatography until >95% purity. The protein was shown to have the appropriate folding as determined by ELISA and flow cytometry binding assays to its receptor, as well as by its detection by hamster immune anti-S1 sera under non-reducing conditions. In immunization assays in mice and hamsters, the purified RBD formulated with adjuvants based on oil-water emulsifications and squalene was able to stimulate specific neutralizing antibodies and confirm the secretion of IFN-{gamma} after stimulating spleen cells with the purified RBD. The vaccine candidate was shown to be safe, as demonstrated by the histopathological analysis in lungs, liver and kidney. These results demonstrate the potential of the purified RBD administered with adjuvants through an intramuscular route, to be evaluated in a challenge against SARS-CoV-2 and determine its ability to confer protection against infection.

immunology↗

Silencing of an efflux pump coding gene decreases the efflux rate of pyrazinoic acid in Mycobacterium smegmatis

BackgroundTuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (MTB). The recommended treatment for TB is based on the use of first-line drugs, including pyrazinamide (PZA). PZA is also a drug used in the treatment of multidrug-resistant TB (MDR-TB) because of its main effect against the latent stage. The main cause of resistance to PZA is mutations in the pncA gene, which compromise the activity of the encoded enzyme pyrazinamidase (PZAse), which hydrolyzes PZA into POA, the active antituberculosis molecule. The mechanism of action of PZA requires that POA is expelled from the bacterium by an efflux mechanism. After that, if the extracellular medium is sufficiently acidic, POA is protonated and returns to the cytosol, releasing the proton and repeating the cycle, resulting lethal to the bacteria. The efflux pump responsible for extruding the POA to the extracellular environment is not yet known. Mycobacterium smegmatis is naturally resistant to PZA and has a 900-fold faster POA efflux rate than MTB, and has the advantage to be a faster growing mycobacterium. MethodsIn the present study we have silenced the transcription of several genes encoding efflux pumps in M. smegmatis by CRISPRi (CRISPR interference). These genes (MSMEG_0250, MSMEG_3815, MSMEG_0241, MSMEG_5046 and MSMEG_0410) were homologous to efflux pump genes in MTB. POA efflux rate was measured, and a quantitative Waynes test was performed after silencing each gene. ResultsSilencing of MSMEG_0250, resulted in approximately 5-fold decrease in the POA efflux rate in M. smegmatis (P<0.0001). None of the other silenced genes showed a notable decrease in POA efflux rate.

molecular biology↗