Truncated Surface Protective Antigen (SpaA) of Erysipelothrix rhusiopathiae Serotype 1a Elicits Protection against Challenge with Serotypes 1a and 2b in Pigs

Abstract

Erysipelothrix rhusiopathiae is a causal agent of swine erysipelas, which is of economic importance in the swine industry by virtue of causing acute septicemia, chronic arthritis, and endocarditis. However, little is known about the genetic properties of its protective antigens. Recently, a surface protective antigen (SpaA) gene was identified from serotype 2 in a mouse model. We clonedspaA from virulent strain Fujisawa (serotype 1a) and determined that the N-terminal 342 amino acids without C-terminal repeats of 20 amino acids have the ability to elicit protection in mice. Fusions of 342 amino acids of Fujisawa SpaA and histidine hexamer (HisSpa1.0) protected pigs against challenge with both serotype 1 and serotype 2, the most important serotypes in the swine industry. Pigs immunized with HisSpa1.0 reacted well with both HisSpa1.0 and intact SpaA by enzyme-linked immunosorbent assay and immunoblotting. Serum collected at the time of challenge from a pig immunized with HisSpa1.0 markedly enhanced the in vitro phagocytic and killing activity of pig neutrophils against the bacteria. DNA sequences of protective regions of spaA genes from five strains of serotypes 1 and 2 were almost identical. The full DNA sequences also seemed to be conserved among strains of all 12 serotype reference strains harboring thespaA gene by restriction fragment length polymorphism analysis of PCR products. These results indicates that SpaA is a common protective antigen of serotypes 1 and 2 of E. rhusiopathiae in swine and will be a useful tool for development of new types of vaccines and diagnostic tools for effective control of the disease.

Identification of surface protective antigen (spa) types in Erysipelothrix reference strains and diagnostic samples by spa multiplex real-time and conventional PCR assays

“…The Spa types of the Erysipelothrix reference strains with serotypes 1-26 and N were tested using the spa multiplex real-time and conventional PCR assays. Consistent with previous results identified by SDS-PAGE and Western blotting (Makino et al 1998;Imada et al 1999;Shimoji et al 1999;To and Nagai 2007), we found spaA in E. rhusiopathiae serotypes 1a, 1b, 2, 5, 9, 12, 15, 16, 17 and N; spaB (including spaB1 and spaB2) in E. rhusiopathiae serotypes 4, 6, 11, 19 and 21; and spaC in E. sp. strain 2 serotype 18.…”

“…However, to our knowledge, a quick method such as we describe to determine the spa type of the Erysipelothrix spp. has not been previously reported, in spite of the fact that techniques utilizing SDS-PAGE and Western blotting (Makino et al 1998;Imada et al 1999;Shimoji et al 1999;To and Nagai 2007) and conventional PCR assays for spaA, spaB and spaC (Ingebritson et al 2010) have been recently described. For the spaB gene detection in the multiplex real-time PCR, we first designed a spaB1 probe but found it was not able to recognize serotype 11.…”

“…Previously, the SpaA protein was found to be highly immunogenic and has been regarded as the superior protective antigen for developing new vaccines (Timoney and Groschup 1993;Makino et al 1998;Imada et al 1999;Shimoji et al 1999Shimoji et al , 2002Cheun et al 2004). However, it also has been shown that specific antibodies against each Spa type react strongly with the homologous Spa protein but weakly with heterologous Spa proteins (To and Nagai 2007).…”

“…Because the Spa antigen is one of the most clearly defined surface proteins and is a protective antigen, the detection and identification of the Spa type may be important for evaluating the antigenicity and pathogenicity of field Erysipelothrix isolates. However, the full deter-mination of Spa protein type usually requires sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting with appropriate antisera (Makino et al 1998;Imada et al 1999;Shimoji et al 1999;To and Nagai 2007), which is time-consuming. In this study, we describe the development of a spa multiplex real-time and a conventional PCR assay for the detection and differentiation of Spa-related genes in Erysipelothrix spp.…”

Characterization of spaC‐type Erysipelothrix sp. isolates causing systemic disease in ornamental fish

“…Virulence factor profiles of isolates from terrestrial animals are fairly uniform (Janßen et al, 2015) and include an immunologically significant surface-protective antigen (Spa) for which three isoforms, SpaA, SpaB and SpaC, have been identified (To & Nagai, 2007). SpaA is a common protective antigen of E. rhusiopathiae (Imada, Goji, Ishikawa, Kishima, & Sekizaki, 1999) and virulence determinant that plays a role in cell adhesion, as well as resistance to serum killing and phagocytosis by macrophages (Borrathybay, Gong, Zhang, & Nazierbieke, 2015).…”