Establishment of a production process for a novel vaccine candidate against Lawsonia intracellularis

Salazar, Santiago; Gutiérrez, Nicolás Antonio Sáez; Sánchez, Oliberto; Ramos, Eduardo Antônio Gonçalves; González, Alaín; Acosta, Jorge Berlanga; Ramos, Thelvia; Altamirano, Claudia; Alonso, Jorge R. Toledo; Montesino, Raquel

doi:10.1016/j.ejbt.2021.01.002

Cited by 1 publication

(4 citation statements)

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“…The production of individual antigens in E. coli under standard conditions allowed for the development of in vitro and in vivo experiments. As in a previous study, we obtained a 70-fold increase in protein yields when expressing the three antigens simultaneously by adjusting medium and culture conditions [30]. It is highly probable that a similar optimization applied to the individual antigen production also provides a significant improvement of antigen yields.…”

Section: Discussionsupporting

confidence: 71%

“…A favorable immune response pattern was observed with our new vaccine candidate formulated with even antigen amounts. These results were predictable due to outcomes with a similar vaccine candidate [ 29 , 30 ]. However, we obtained higher antibody titers and a superior cellular response in this study.…”

Section: Discussionmentioning

confidence: 99%

“…Inclusion bodies were extracted as previously described [ 30 ]. Briefly, the biomass was resuspended in PBS (137 mM NaCl (Sigma-Aldrich, Saint Louis, MO, USA), 2.7 mM KCl (Sigma-Aldrich, Saint Louis, MO, USA), 10 mM Na 2 HPO 4 (Sigma-Aldrich, Saint Louis, MO, USA), and 1.8 mM KH 2 PO 4 (Merck, Darmstadt, Germany), pH 7.4) containing 0.1% Triton X-100 (Sigma-Aldrich, Saint Louis, MO, USA) and then lysed by mechanical cell disruption in the bead mill Dyno-mill ML (Wab, Muttenz/Basel, Switzerland).…”

Section: Methodsmentioning

confidence: 99%

“…An in silico structural analysis allowed for protein modification in order to improve their immunological characteristics. This multi-antigenic and chimeric vaccine candidate showed promising results by inducing an effective immune response and exhibiting protecting signs on immunized pigs upon challenge [ 29 , 30 ]. To make the vaccine candidate production and the downstream process feasible, DNA sequences coding chimeric proteins were cloned into the same plasmid to express the three antigens simultaneously in E. coli as inclusion bodies.…”

Section: Introductionmentioning

confidence: 99%

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New Formulation of a Subunit Vaccine Candidate against Lawsonia intracellularis Increases Humoral and Cellular Immune Responses

Salazar,

Starck,

Villegas

et al. 2023

Vaccines

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Previously, we designed a subunit vaccine candidate based on three L. intracellularis antigens with promising results in pigs. In this study, antigens were produced individually to achieve an even antigen ratio in the formulation. The emulsion characterization included the drop size and the mechanical and thermal stability. Immune response was evaluated by indirect and sandwich ELISAs, qPCR, and flow cytometry. The vaccine candidate’s safety was assessed by histopathology and monitoring the clinical behavior of animals. The average production yielded for the chimeric antigen as inclusion bodies was around 75 mg/L. The formulation showed mechanical and thermal stability, with a ratio Hu/Ho > 0.85 and a drop size under 0.15 nm. Antigens formulated at a ratio of 1:1:1 induced a significant immune response in inoculated pigs that persisted until the end of the experiment (week 14). The dose of 200 μg significantly activated cellular response measured by transcriptional and translational levels of cytokines. The cell proliferation assay revealed an increment of lymphocytes T CD4+ at the same dose. Animals gained weight constantly and showed proper clinical behavior during immunization assays. This research demonstrated the immunological robustness of the new subunit vaccine candidate against Porcine Proliferative Enteropathy evenly formulated with three chimeric antigens of L. intracellularis.

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Section: Discussionsupporting

confidence: 71%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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