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PMID: 1997416 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Changes in the surface of Leptospira interrogans serovar grippotyphosa during in vitro cultivation.

Infection and immunity ·Vol. 59 ·No. 3 ·1991-03-00 ·Pages 1131-40

Haake DA, Walker EM, Blanco DR, Bolin CA, Miller MN, Lovett MA

Abstract

Surface components of virulent and attenuated Leptospira interrogans serovar grippotyphosa were compared by using Triton X-114 solubilization and phase partitioning, immunoprecipitation of intact organisms, and freeze-fracture electron microscopy. Removal of the leptospiral outer membrane by using 0.1% Triton X-114 was demonstrated by whole-mount electron microscopy and by essentially complete solubilization of a lipopolysaccharidelike substance (LLS) from the outer membrane. Triton X-114 (0.1%) did not solubilize subsurface proteins, such as endoflagellar filaments or penicillin-binding proteins, which are markers for the periplasmic space and inner membrane, respectively. Triton X-114 solubilized material from both the virulent and attenuated strains, which partitioned into the hydrophobic, detergent phase, contained LLS and major proteins of 41 and 44 kDa, which were also immunoprecipitable from intact organisms. The virulent strain contained greater amounts of an LLS component with an apparent molecular mass of 30 kDa (R(f) = 0.57), whereas the attenuated strain contained larger amounts of an LLS component with an apparent molecular mass of 20 kDa (R(f) = 0.74). Differences in protein components between virulent and attenuated organisms were also detected; whereas the 41- and 44-kDa proteins were immunoprecipitated in equal amounts from both the virulent and attenuated strains, a 33-kDa protein was immunoprecipitated in significantly greater amounts from the attenuated strain. Quantitation of outer membrane particle density by freeze-fracture electron microscopy showed that both strains had a low transmembrane outer membrane protein content compared with that of typical gram-negative bacteria. The virulent and attenuated strains had 443 and 990 particles (P less than 0.000001) per micron, respectively, in the concave outer membrane fracture face. These findings suggest that in vitro cultivation of L. interrogans is accompanied by quantitative and qualitative changes in both LLS and outer membrane-associated proteins.

MeSH Terms
Animals Antibodies, Bacterial/immunology Antigens, Surface/analysis,immunology Bacterial Outer Membrane Proteins/analysis,immunology,ultrastructure Cricetinae Electrophoresis, Polyacrylamide Gel Freeze Fracturing Leptospira interrogans/analysis,immunology,pathogenicity,ultrastructure Molecular Weight Octoxynol Polyethylene Glycols Polysaccharides, Bacterial/analysis,immunology Precipitin Tests Rabbits Serial Passage Solubility Virulence
Chemicals
Antibodies, Bacterial Antigens, Surface Bacterial Outer Membrane Proteins Polysaccharides, Bacterial Polyethylene Glycols Octoxynol Nonidet P-40
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Haake D A
Department of Medicine, UCLA School of Medicine 90024.
Walker E M
Blanco D R
Bolin C A
Miller M N
Lovett M A
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Article Info
Journal
Infection and immunity
Abbr.
Infect Immun
ISSN
0019-9567
DOI
10.1128/iai.59.3.1131-1140.1991
Published
1991-03-00
Pages
1131-40
Language
English
Region
United States
NLM ID
0246127
PMCID
PMC258378
Subset
IM
Grants
NIAID NIH HHS · AI 12601 · United States
NIAID NIH HHS · AI 21352 · United States
NIAID NIH HHS · AI 29733 · United States
External Links
PubMed source DOI: 10.1128/iai.59.3.1131-1140.1991 PMC Full Text View journal details
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