Comparison of the biological responses induced by lipopolysaccharide and endotoxin of Treponema hyodysenteriae and Treponema innocens

Abstract

The chemical composition and classical biologic activities of lipopolysaccharide (LPS; phenol-water) and endotoxin (butanol-water) preparations from virulent Treponema hyodysenterinae and avirulent Treponema innocens were examined. The LPS and endotoxin preparations from T. hyodysenteriae B204 contained approximately 80.9 and 35.2% hexose, 0.12 and 0.45% thiobarbituric acid-reactive compound, and <1 and 11.3% protein, respectively. The LPS and endotoxin preparations of T. innocens B1555a contained approximately 56.3 and 37.8% hexose, 0.45 and 0.4% thiobarbituric acid-reactive compound, and <1 and 26% protein, respectively. A silver-stained 7.5 to 15% sodium dodecyl sulfate-polyacrylamide gel showed four bands for the T. hyodysenteriae preparations, while the T. innocens preparations failed to resolve into discrete bands on electrophoresis. We determined by the Limulus amebocyte lysate assay that the treponemal preparations had comparable amounts of endotoxin activity when Eschenichia coli LPS was used as a standard. The 50% lethal doses of LPS and endotoxin from T. hyodysenteriae for BALB/cByJ mice were 380 and 80 ,ug, respectively. The treponemal preparations were poor adjuvants, failed to induce a dermal Shwartzman reaction, and were not pyrogenic. The treponemal LPS preparations, unlike the endotoxin preparations, were not mitogenic for murine spleen cells. Differences in virulence between the two treponemal species could not be associated with the biologic activities of the respective LPS or endotoxin moieties, but the endotoxin preparations were consistently more active than the purified LPS preparations. Lipopolysaccharide (LPS) is a molecule that is found in the outer membrane of gram-negative bacteria and is associated with numerous biologic effects on the mammalian immune system (25). These responses include B-cell mitogenicity, polyclonal antibody induction, adjuvanticity, macrophage activation, immunogenicity, pyrogenicity, lethality, induction of tolerance, and inflammatory reactions (25, 33, 34, 39). Different extraction methods for LPS have been reported (5, 8, 24, 46). LPS, when free of contaminating protein, is extracted by the hot phenol-water method of Westphal and Jann (46). Endotoxin preparations, which consist of LPS as well as lipid A-associated protein(s), is extracted by either the trichloroacetic acid methods of Boivin and Mesrobeanu (5) and Staub (36) or the butanol-water method of Morrison and Leive (24). Endotoxin possesses all of the aforementioned biologic activities and the ability to stimulate lymphoreticular cells from LPS-hyporesponsive C3H/HeJ mice (10, 45). This stimulation has been shown to reside with the protein moieties that are associated with endotoxin (10, 23, 25). In comparison with the LPS from Escherichia coli, there are numerous gram-negative organisms which have LPS with various chemical and biologic characteristics (12, 31, 35). These differences include the inability to detect the presence of 2-keto-3-deoxyoctonate (KDO) (9), differences in fatty acid composition ...

Induction of interleukin (IL)-1beta and IL-8 mRNA expression in porcine macrophages by lipopolysaccharide from Serpulina hyodysenteriae

“…Preparations of endotoxin or LPS were dissolved in pyrogen-free saline (2 mg/ml on a dry weight basis), sterilized by being heated at 100ЊC for 10 min, and stored at 4ЊC until use. The serpulinal LPS and endotoxin preparations had comparable amounts of endotoxin activity compared with E. coli LPS in the Limulus amebocyte lysate assay, as has been previously reported (7).…”

“…Serpulina hyodysenteriae, a gram-negative anaerobic spirochete, induces a mucohemorrhagic diarrheal disease of growing swine (9). Putative virulence determinants of S. hyodysenteriae include a ␤-hemolysin(s) (11,17,18) and a lipopolysaccharide (LPS) or LPS-like molecule (7,15,16). This LPS-like molecule isolated by phenol-water extraction possesses various in vitro biologic activities, including toxicity for murine peritoneal macrophages, enhancement of C3 and Fc␥ receptor-mediated phagocytosis, splenocyte mitogenesis, and stimulation of leukocyte chemotaxis (14).…”

“…LPS from E. coli K235 and from Salmonella typhimurium was obtained from Sigma Chemical Co., St. Louis, Mo. LPS (phenol-water extract) and endotoxin (butanol-water extract) from S. hyodysenteriae B204 and B234 were prepared as previously described (7). It has been shown that the LPS and endotoxin preparations differ in the percentage of protein, hexose, and 2-keto-3-deoxyoctanate (7).…”

Purification and immunological characterization of a major low-molecular-weight lipoprotein from Borrelia burgdorferi

“…Reports differ as to whether spirochetes contain lipopolysaccharide (LPS) in their outer membrane. Leptospira interrogans, Treponema phagedenis (biotype Reiter), and Serpula (Treponema) hyodysenteriae apparently do contain an LPSlike substance (3,21,23,28,35,36,41,63,66,76,77), while Treponema pallidum apparently does not (3,63). There is conflicting evidence regarding the presence of LPS in Borrelia spp.…”

[beta]-hemolysin and the pathogenesis of Serpulina hyodysenteriae infection

“…In vitro properties of LPS and endotoxin include polyclonal B cell activation and activation of macrophages resulting in interleukin-1 (IL-1) and tumor necrosis factor (TNF) release. LPS and endotoxin preparations from S. hyodysenteriae and S. innocens do possess these properties, but in order for them to be observed, 5 to 50 times as much LPS or endotoxin is required when compared to E. coli or Salmonella spp (51,141). Furthermore, the dermal Shwartzman reaction was not elicited in rabbits treated with serpulinal endotoxin.…”