Secreted Extracellular Products of Flavobacterium covae as Potential Immunogenic Factors for Protection against Columnaris Disease in Channel Catfish (Ictalurus punctatus)

Abstract

Columnaris disease caused by Flavobacterium covae leads to substantial economic losses in commercially important fish species worldwide. The US channel catfish (Ictalurus punctatus) industry is particularly vulnerable to this disease. Therefore, there is an urgent need to develop a vaccine to reduce the economic losses caused by this disease. Secreted extracellular products (SEPs) are considered to be essential bacterial virulence factors that often provide immunogenicity and protection. The current study sought to identify the main SEPs of F. covae and to evaluate their potential to provide protection in channel catfish against columnaris disease. SDS-PAGE analysis of SEPs revealed five protein bands with molecular weights ranging from 13 to 99 kDa. Mass spectrometry analysis showed that these SEPs were hypothetical protein (AWN65_11950), zinc-dependent metalloprotease (AWN65_10205), DNA/RNA endonuclease G (AWN65_02330), outer membrane protein beta-barrel domain (AWN65_12620), and chondroitin-sulfate-ABC endolyase/exolyase (AWN65_08505). Catfish fingerlings were vaccinated with SEPs, SEPs emulsified with mineral oil adjuvant, or heat-inactivated SEPs, or they were sham-immunized through intraperitoneal (IP) injection. After 21 days, an F. covae challenge showed 58.77% and 46.17% survival in the catfish vaccinated with the SEPs and the SEPs emulsified with adjuvant compared to the sham-vaccinated control (100% mortality within 120 h post-infection). However, the heat-inactivated SEPs failed to provide significant protection (23.15% survival). In conclusion, although SEPs contain potentially important immunogenic proteins, further work is needed to optimize their use for long-lasting protection against columnaris disease in fish. These results are significant given the economic impact of columnaris disease on fish farming worldwide.

Keywords: Flavobacterium covae; protection; secreted extracellular proteins.

Figure 1

SDS-PAGE with Coomassie blue stain showing purified SEPs of F. covae 94-081 grown in FCGM. Molecular weights in kilodaltons (kDa) are shown for the standard protein marker in the right column.

Figure 2

Percent survival of channel catfish fingerlings immunized with the SEPs, heat-inactivated SEPs, or SEPs emulsified with adjuvant (Adj + SEPs) and challenged with F. covae 94-081 strain at 21 days post-exposure. Data are presented as mean ± SE of four replicate tanks.