Immune Response of Nile Tilapia (Oreochromis niloticus) Vaccinated With Diatom-Based Oral Vaccines Against Piscine Francisellosis

Abstract

Piscine francisellosis is a highly infectious and economically significant disease caused by Francisella orientalis in tilapia (Oreochromis spp.). There are currently no approved treatments or commercial vaccines for this disease in cultured fish. Injectable vaccines using diatoms as antigen expression vectors have demonstrated efficacy in tilapia models; however, no oral vaccine trials have been performed. We hypothesised that fusion proteins consisting of F. orientalis IglC and flagellin expressed in Thalassiosira pseudonana diatoms will act as self-adjuvanting antigen delivery systems to confer a protective immune response against F. orientalis in tilapia when administered as top-coated feed. Different treatments were immunised and subsequently provided with one or two boosters prior to challenge. Fish were challenged with virulent F. orientalis via immersion thirty days post initial immunisation. Tilapia immune response was assessed 24 h post-challenge by quantifying il-12, il-10, ifn-γ and tgf-β gene expression in gills and internal organs. Morbidity and mortality were monitored for 21 days after challenge and bacterial load was assessed in survivors. Findings indicate significant changes in the expression of ifn-γ and tgf-β in immunised fish, but similar mortality rates and bacterial load across all exposed groups.

Keywords: Francisella orientalis; cytokine; diatom; immersion challenge; oral vaccine; tilapia.

FIGURE 1

Kaplan–Meier survival curves of tilapia fingerlings from day 0 to day 21 post immersion challenge with Francisella orientalis. Each curve represents the average survival of 4 parallel tanks holding n = 16 fish. Groups that do not share letters are significantly different as determined by Log‐rank (Mantel Cox) test with Bonferroni correction (α = 0.05). (A) Wild‐type diatoms, (B) flagellin‐expressing diatoms, (C) IglC‐expressing diatoms and (D) flagellin‐IglC‐expressing diatoms, including both challenged and unchallenged groups.

FIGURE 2

Cumulative mortality rate of tilapia fingerlings 21 days post challenge with 1.6 × 10⁵ CFU Francisella orientalis/mL tank water for 5 h at 25°C. Each column represents the mean with standard error of 4 replicate tanks (n = 16 fish each). Columns with asterisks (*) were unchallenged controls. Groups that do not share letters are significantly different (p < 0.05) as determined by one‐way ANOVA with post hoc Tukey Test.

FIGURE 3

Francisella orientalis genome equivalent (GE) quantification in internal organs of tilapia fingerlings by qPCR. Treatments (n = 10/group) sampled from survivors at 21 days post challenge with 1.6 × 10⁵ CFU F. orientalis/mL tank water for 5 h at 25°C. Mortalities (n = 20) across groups were sampled at the time of death. Columns represent mean with standard error. Different letters are significantly different (p < 0.05) as determined by one‐way ANOVA with post hoc Tukey Test.

FIGURE 4

Relative gene expression of interleukin 12, interferon gamma, interleukin 10 and transforming growth factor beta in (A) gills and (B) internal organs of n = 8/group tilapia fingerlings 24 h post challenge with 1.6 × 10⁵ CFU Francisella orientalis/mL tank water for 5 h at 25°C. Columns represent mean with standard error. Gene expression was normalised against the expression of the average of actb and ubce reference genes. Different letters within each gene are significantly different (p < 0.05) as determined by one‐way ANOVA with post hoc Tukey Test.