Capsular polysaccharide determines the serotyping of Riemerella anatipestifer

Abstract

Riemerella anatipestifer ( R. anatipestifer ) is a Gram-negative pathogen that causes significant economic losses to the farming industry. The polysaccharides that determine serotype are well understood in other bacteria; they have not yet been fully characterized in R. anatipestifer . In this study, we unexpectedly discovered a strain, RCAD0392, that is capable of cross-agglutination. Whole genome sequencing revealed base mutations disrupting a gene on its capsular polysaccharide synthesis gene cluster. By constructing homologous gene deletion mutants in CH-2, we demonstrated that deletion of this gene leads to altered serological phenotypes. India ink and transmission electron microscopy experiments revealed the disappearance of capsule on the surface of the bacteria, indicating the association of the gene with capsule synthesis. In addition, agar-gel precipitin tests showed that lipopolysaccharide binds to antisera of multiple serotypes, while the capsular polysaccharide only binds to the corresponding antisera. This suggests that capsular polysaccharide is the specific antigen that determines the serotype of R. anatipestifer . IMPORTANCE Riemerella anatipestifer ( R. anatipestifer ) is one of the most important veterinary pathogens with at least 21 serotypes. However, the exact polysaccharide(s) that determine R. anatipestifer serotype is still unknown. This study has provided a preliminary exploration of the relationship between capsular polysaccharides and serotyping in R. anatipestifer and suggests possible directions for further investigation of the genetic basis of serotypes in this bacterium.

The GE296_RS03820 and GE296_RS03830 genes are involved in capsular polysaccharide biosynthesis in Riemerella anatipestifer

“…These genes include a wza-like gene (the B739_1124 gene) of RA-CH-1 (corresponding to the GE296_RS03840 gene of RA-LZ01), the M949_RS00050 gene of the CH3 strain and the G148_RS04320 gene (corresponding to the GE296_ RS03845 gene) of CH-2. [6][7][8] The CPS synthesis in R. anatipestifer involves multiple genes, as predicted in Figure 1. Hence, we aimed to identify additional key genes that have significant effects on the CPSs of R. anatipestifer.…”

“…Only three references have been identified, indicating the involvement of specific genes in CPS biosynthesis in R. anatipestifer . These genes include a wza ‐like gene (the B739_1124 gene) of RA‐CH‐1 (corresponding to the GE296_RS03840 gene of RA‐LZ01), the M949_RS00050 gene of the CH3 strain and the G148_RS04320 gene (corresponding to the GE296_RS03845 gene) of CH‐2 6–8 . The CPS synthesis in R. anatipestifer involves multiple genes, as predicted in Figure 1.…”

“…Previous studies have reported the presence of certain genes involved in capsule biosynthesis, such as wza-like, M949_RS00050, and G148_RS04320 genes. [6][7][8] However, the complete mechanism of capsular biosynthesis in R. anatipestifer remains to be fully understood, as it can contribute to understanding the pathogenicity of the bacterium and provide targets for prevention and treatment. Therefore, it is necessary to identify the key genes related to the biosynthesis of CPS in R. anatipestifer.…”

“…R. anatipestifer is known to produce capsules, but the understanding of its capsular biosynthesis is limited. Previous studies have reported the presence of certain genes involved in capsule biosynthesis, such as wza ‐like, M949_RS00050 , and G148_RS04320 genes 6–8 . However, the complete mechanism of capsular biosynthesis in R. anatipestifer remains to be fully understood, as it can contribute to understanding the pathogenicity of the bacterium and provide targets for prevention and treatment.…”

“…FIGURE6 The bacterial burdens in the blood and brain tissue of ducklings at 24 h, 48 h and 72 h post infection. The loads in blood are presented as log 10 CFU/mL , and those in brain tissue are presented as log 10 CFU/g (***p < .001 was considered to indicate statistical significance).…”