Abstract
The spirochetes Brachyspira hyodysenteriae and B. pilosicoli are pig intestinal pathogens that are the causative agents of swine dysentery (SD) and porcine intestinal spirochaetosis (PIS), respectively. Although some inactivated bacterin and recombinant vaccines have been explored as prophylactic treatments against these species, no effective vaccine is yet available. Immunoproteomics approaches hold the potential for the identification of new, suitable candidates for subunit vaccines against SD and PIS. These strategies take into account the gene products actually expressed and present in the cells, and thus susceptible of being targets of immune recognition. In this context, we have analyzed the immunogenic pattern of two B. pilosicoli porcine isolates (the Spanish farm isolate OLA9 and the commercial P43/6/78 strain) and one B. hyodysenteriae isolate (the Spanish farm V1). The proteins from the Brachyspira lysates were fractionated by preparative isoelectric focusing, and the fractions were analyzed by Western blot with hyperimmune sera from challenged pigs. Of the 28 challenge-specific immunoreactive bands detected, 21 were identified as single proteins by MS, while the other 7 were shown to contain several major proteins. None of these proteins were detected in the control immunoreactive bands. The proteins identified included 11 from B. hyodysenteriae and 28 from the two B. pilosicoli strains. Eight proteins were common to the B. pilosicoli strains (i.e., elongation factor G, aspartyl-tRNA synthase, biotin lipoyl, TmpB outer membrane protein, flagellar protein FlaA, enolase, PEPCK, and VspD), and enolase and PEPCK were common to both species. Many of the identified proteins were flagellar proteins or predicted to be located on the cell surface and some of them had been previously described as antigenic or as bacterial virulence factors. Here we report on the identification and semiquantitative data of these immunoreactive proteins which constitute a unique antigen collection from these bacteria.
Highlights
Bacteria in the Brachyspiraceae family are gram-negative and spiral-shaped Spirochaetes
Note that for all strains, an immunoblot analysis with control serum from non-challenged pigs revealed some reactive bands (Supplementary Figure S3). This cross-reactivity was consistent with previous reports that showed that healthy pig serum detects B. hyodysenteriae surface antigens (Wannemuehler et al, 1988) or some of the recombinant proteins tested for a vaccine against swine dysentery (SD) (Song et al, 2009)
The immunoproteomics approach applied in this study has been demonstrated to be very effective for the characterization of new Brachyspira antigens
Summary
Bacteria in the Brachyspiraceae family are gram-negative and spiral-shaped Spirochaetes. Members of this phylum are characterized by their unique motility and a loosely coiled morphology caused by the existence of periplasmic flagella (Charon and Goldstein, 2002). Species of the Brachyspiraceae family are anaerobic, host-associated intestinal bacteria in pigs, humans and other species, and can cause gastrointestinal pathologies and mortality (Stanton, 2006). SD mainly affects pigs in the growing-finishing periods. This contributes to the high cost of the disease, which is associated with mortality, which is relatively low when animals are treated (Hamdy, 1974), and with high morbidity, growth retardation and the need for continual in-feed medication
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