Existence and characterization of allelic variants of Sao, a newly identified surface protein fromStreptococcus suis

Abstract

Surface antigen one (Sao) is a newly identified protein from the major zoonotic pathogen, Streptococcus suis. In search of functional proteins related to the pathogenesis of Chinese S. suis 2 (SS2), unexpectedly, a variant of Sao protein was obtained. To test its prevalence in S. suis, PCR assay was adopted to address the coding genes systematically. It was found that there are three allelic variants of sao gene, namely sao-S, sao-M, and sao-L based on the different lengths of the genes (approximately 1.5, approximately 1.7, and approximately 2.0 kb, respectively). These differences were determined to be caused by heterogeneity within the number of C-terminal repeat sequences (R), which had been seen as a pathogenicity-related domain in the plant pathogen, Xanthomonas oryzae. Two variants (sao-M and sao-L) were only found in SS2. All three variant proteins were prepared in vitro and their biochemical and biophysical properties were characterized. A soluble form of Sao-M protein was then used as a capture antigen to develop an enzyme-linked immunosorbent assay method to detect antibodies against SS2 in convalescent pig sera. Taken together, the results exhibit the properties of Sao proteins and provide an efficient Sao-M-based method for monitoring SS2 infection.

Functional Definition and Global Regulation of Zur, a Zinc Uptake Regulator in a Streptococcus suis Serotype 2 Strain Causing Streptococcal Toxic Shock Syndrome

“…The peak was collected, visualized by 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then stained with Coomassie brilliant blue R250 (Sigma, St. Louis, MO). The apparent molecular weights were estimated by comparison with standard protein markers (Sangon, Shanghai, China) run on the same gel (16). Chemical cross-linking assays.…”

“…A soluble protein encoded by gene 310 (named protein 310) was obtained as described previously (16) with some minor modifications. After sonication, the clarified bacterial supernatant was loaded onto affinity columns (for the His tag version, a nickel-ion [Ni ϩ ] affinity column [Qiagen], for the glutathione S-transferase [GST]-fused version, a glutathione Sepharose 4B column [Amersham]).…”

“…E. coli strains DH5␣ and BL21 were cultured in Luria-Bertani (LB) liquid medium or plated on LB agar. If necessary, either 50 g/ml of ampicillin (Sigma) or 50 g/ml of kanamycin (Sigma) was utilized to screen E. coli transformants (16).…”

“…Total bacterial protein was prepared by sonication from three strains of S. suis 2, i.e., the wild type (WT) (05ZYH33), the ⌬310 mutant, and the ⌬310 mutant complemented strain (C⌬310 mutant). They were then separated by 15% SDS-PAGE and transferred onto a polyvinylidene difluoride (PVDF) membrane for the Western blot-based detection of protein 310 as we described previously (16). Here, the first antibody is polyclonal anti-310 protein rabbit serum, which was obtained according to the customary immunization protocol for New Zealand White rabbits (33).…”

Diagnostik Antijen Olarak Rekombinant Sao-M Protein Kullanılarak Streptococcus suis’in Tanısı Amacıyla PPA-ELİSA Geliştirilmesi

“…Immunization with recombinant Sao protein was capable of provoking strong humoral antibody responses, diminish clinical signs and bacterial dissemination, improve survival rates and provide cross-serotype protection in pig and mouse vaccination protocols [24] , indicating rSao is a suitable antigen for subunit S. suis vaccine development [25] . Sao protein is encoded by three allelic variants of gene of difference lengths, Sao-S (1.5 kb), Sao-M (1.7 kb) and Sao-L (2.0 kb), and Sao-M is the most prevalent variant comprising about 80% [26] . We suspect that Sao-M has great potential to become a diagnostic antigen for detection of S. suis infection.…”