Development of an Acid-Resistant Salmonella Typhi Ty21a Attenuated Vector For Improved Oral Vaccine Delivery.
The licensed oral, live-attenuated bacterial vaccine for typhoid fever, Salmonella enterica serovar Typhi strain Ty21a, has also been utilized as a vaccine delivery platform for expression of diverse foreign antigens that stimulate protection against shigellosis, anthrax, plague, or human papilloma virus. However, Ty21a is acid-labile and, for effective oral immunization, stomach acidity has to be either neutralized with buffer or by-passed with Ty21a in an enteric-coated capsule (ECC). Several studies have shown that efficacy is reduced when Ty21a is administered in an ECC versus as a buffered liquid formulation, the former limiting exposure to GI tract lymphoid tissues. However, the ECC was selected as a more practical delivery format for both packaging/shipping and vaccine administration ease. We have sought to increase Ty21a acid-resistance to allow for removal from the ECC and immune enhancement. To improve Ty21a acid-resistance, glutamate-dependent acid resistance genes (GAD; responsible for Shigella spp. survival at very low pH) were cloned on a multi-copy plasmid (pGad) under a controllable arabinose-inducible promoter. pGad enhanced acid survival of Ty21a by 5 logs after 3 hours at pH 2.5, when cells were pre-grown in arabinose and under conditions that promote an acid-tolerance response (ATR). For genetically 100% stable expression, we inserted the gad genes into the Ty21a chromosome, using a method that allowed for subsequent removal of a selectable antibiotic resistance marker. Further, both bacterial growth curves and survival assays in cultured human monocytes/macrophages suggest that neither the genetic methods employed nor the resulting acid-resistance conferred by expression of the Gad proteins in Ty21a had any effect on the existing attenuation of this vaccine strain.
Laboratory of Mucosal Pathogens and Cellular Immunology, Food and Drug Administration-Center for Biologics Evaluation and Research, New Hampshire Avenue, Silver Spring, Maryland, United States of America.
Madushini N
http://orcid.org/0000-0003-4954-2487
MN
Dharmasena
Laboratory of Mucosal Pathogens and Cellular Immunology, Food and Drug Administration-Center for Biologics Evaluation and Research, New Hampshire Avenue, Silver Spring, Maryland, United States of America.
Catherine M
CM
Feuille
Laboratory of Mucosal Pathogens and Cellular Immunology, Food and Drug Administration-Center for Biologics Evaluation and Research, New Hampshire Avenue, Silver Spring, Maryland, United States of America.
Carly Elizabeth C
CE
Starke
Environmental Microbial and Food Safety Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland, United States of America.
Arvind A
AA
Bhagwat
Laboratory of Mucosal Pathogens and Cellular Immunology, Food and Drug Administration-Center for Biologics Evaluation and Research, New Hampshire Avenue, Silver Spring, Maryland, United States of America.
Scott
S
Stibitz
Laboratory of Mucosal Pathogens and Cellular Immunology, Food and Drug Administration-Center for Biologics Evaluation and Research, New Hampshire Avenue, Silver Spring, Maryland, United States of America.
Dennis J
DJ
Kopecko
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