Abstract
Lactococcus garvieae (L. garvieae) is an important pathogen that causes enormous economic losses in both marine and freshwater aquaculture. At present, antibiotics are the only option for farmers to reduce the losses caused by L. garvieae. However, the usage of antibiotics leads to environmental pollution and the production of drug-resistant strains of bacteria. Therefore, vaccination is preferred as an alternative method to prevent infectious diseases. In this study, we describe an effective approach to the production of an oral biofilm vaccine, using bacteria grown on chitosan particles to form biofilms, and thus providing an inactive pathogen that enhances the immune response in fish. We observed the formation of a biofilm on chitosan particles and administered the novel oral biofilm vaccine to fish. We analyzed the immune responses, including antibody production, phagocytic ability, albumin/globulin ratio and immune-related genes, of vaccinated and control groups of black mullet. Our results show that the phagocytic ability of the biofilm vaccine group was 84%, which is significantly higher than that of the control group, and the antibody production in this group was significantly higher compared with the other group. The mRNA expression levels of immune-related genes (TLR2, IL-1β, TNF-α) were significantly upregulated in the spleen after vaccination. In challenge experiments, the relative percent survival (RPS) was 77% in the biofilm vaccine group, 18% in the whole-cell vaccine group, and 0% in the chitosan particle group at 32 days post-vaccination. In addition, we also found that the relative percent survival (RPS) at 1 day post-vaccination was 74% in the biofilm vaccine group, 42% in the whole-cell vaccine group, and 26% in the chitosan particle group. In both long-term and short-term challenge experiments, the viability of the biofilm vaccine group was significantly higher than that of the whole-cell, chitosan particle and PBS groups. We conclude that based on its protective effect, the L. garvieae biofilm vaccine is better than the whole-cell vaccine when challenged several weeks after vaccination. In addition, the biofilm vaccine also has a greater protective effect than the whole-cell vaccine when challenged immediately after vaccination. Therefore, the biofilm vaccine might represent a novel method for the prevention and treatment of L. garvieae infection.
Highlights
Lactococcus garvieae is a facultative anaerobic, non-motile and Gram-positive bacteria that infects many fish species and causes enormous economic losses in both marine and freshwater aquacultures [1]
The results show that bacteria attached to the chitosan and began to form a biofilm at 3 h, reaching a peak at 48 h (Figure 2A,C)
The results indicate that the phagocytic ability of the biofilm vaccine group (84%) was significantly higher than that of the whole-cell vaccine, chitosan particle and PBS
Summary
Lactococcus garvieae is a facultative anaerobic, non-motile and Gram-positive bacteria that infects many fish species and causes enormous economic losses in both marine and freshwater aquacultures [1]. 80% of persistent bacterial infections may be related to biofilm formation [4]. The composition of the biofilm matrix enables bacteria to deceive the host immune system and escape the immune response. After Streptococcus pneumonia infection, the bacteria trigger an acute response, as bacteraemia stimulates the immune system of the host and induces humoral immunity against planktonic bacterial antigens. The biofilm antigenicity and immunogenicity are considerably different from those of the planktonic bacteria, and the biofilm elicits a different immune response in the host [7]. In a mouse model study, a biofilm LytB protein vaccine was immunogenic, and enhanced complement-mediated immunity and the phagocytosis of different serotypes of S. pneumoniae [10]
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