Inactivated enterotoxigenic Escherichia coli (ETEC) vaccine with a natural-based nanoadjuvant enhances specific IgG serum levels in mice

CECCHINI ME; MONTIRONI ID; ARSAUTE S; RAVIOLO J; CAMACHO N; VANCOLLI ML; PALMA S; MAÑAS F; BELLINGERI R; CARIDDI LN

Congreso; Reunión conjunta SAIC SAI&FAIC SAFIS; 2022

The immunoadjuvant potential of Minthostachys verticillata essential oil (EO) has been previously demonstrated. Therefore, this study aimed to evaluate the adjuvant effect of the EO-based nanoemulsion (NEO) incorporated in an experimental inactivated vaccine against enterotoxigenic Escherichia coli (ETEC). The NEO was synthetized with 20% v/v of EO, 1% v/v of a surfactant mixture (0.75% Tween 80 and 0.25% Span 60) and 79% v/v of distillated water; the nanodroplet size was 105.6 nm (PDI=0,301). Binding between the NEO and ETEC was assessed by SEM. Experimental vaccines were prepared under aseptic conditions with formaldehyde-inactivated ETEC at 1x109 UFC/mL with either safe concentrations of NEO (1; 0.75 and 0.5 mg/mL), incomplete Freund's adjuvant (IFA) (50%), EO (1 mg/mL), surfactant mixture (Tween80/Span60 1 mg/mL) or saline solution. Saline-treated animals were included as a negative control. Mice were divided into eight groups of four animals each and vaccines were administrated subcutaneously on days 1, 14, 28 and 42. Animals were kept under controlled temperature, provided ad libitum access to food and water and were sacrificed 7 days after the last inoculation. For each group, a pool of sera was obtained by centrifugation and anti-ETEC IgG titers were determined through the indirect ELISA method. Assays were performed by triplicate. Results showed that IgG titers of the groups that received the vaccines formulated with NEO as adjuvant were significantly increased (p˂0.05) compared to control groups without adjuvant, reaching similar values of those in IFA group. Moreover, IgG titers produced by the NEO at concentrations of 1 and 0.5 mg/mL were significantly higher (p˂0.05) than those that received the EO-adjuvanted formulation, suggesting that the process of nanoencapsulation enhanced the immunomodulating activity of the EO.