PURIFICATION AND CHARACTERIZATION OF AN EXOPOLYSACCHARIDE OF BURKHOLDERIA (PSEUDOMONAS) PSEUDOMALLEI

被引:64
作者
STEINMETZ, I [1 ]
ROHDE, M [1 ]
BRENNEKE, B [1 ]
机构
[1] GESELL BIOTECHNOL FORSCH MBH, D-38124 BRAUNSCHWEIG, GERMANY
关键词
D O I
10.1128/IAI.63.10.3959-3965.1995
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
Burkholderia pseudomallei (basonym Pseudomonas pseudomallei) is the causative organism of melioidosis, a disease which is recognized as a major public health problem primarily in Southeast Asia and Northern Australia. In this paper, we report on the identification, purification, and characterization of a species-specific exopolysaccharide of B. pseudomallei. After immunization of mice with a B. pseudomallei strain exhibiting mucoid growth characteristics, we isolated an immunoglobulin G1 monoclonal antibody (MAb) (3015) with specificity for a carbohydrate structure as determined by immunoblotting following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Electron microscopy studies with MAb 3015 revealed reactivity with an exopolysaccharide with a capsule-like appearance in the immunizing strain. All of the mucoid and nonmucoid B, pseudomallei strains tested from geographically different tropical regions were recognized by MAb 3015 in an enzyme-linked immunosorbent assay or immunoblot, indicating that the exopolysaccharide is constitutively expressed among this species. Intensive testing for cross-reactivity including members of all the Pseudomonas rRNA groups showed no cross-reactivity except in the case of the closely related species Burkholderia mallei. A protocol for purification of the exopolysaccharide which is based principally on mechanical separation from the cell surface followed by repetitive ethanol precipitation steps and finally affinity chromatography using MAb 3015 was established. The exopolysaccharide yielded was of high purity. Gel permeation chromatography was performed, and the molecular mass was estimated to be >150 k)a. Sera from patients with melioidosis were strongly reactive with the purified exopolysaccharide, indicating its in vivo expression and immunogenicity in natural infection, The diagnostic value of the exopolysaccharide and its role in the pathogenesis of disease must still be determined.
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页码:3959 / 3965
页数:7
相关论文
共 25 条
[1]   INTERACTION BETWEEN LIMULUS AMEBOCYTE LYSATE AND SOLUBLE-ANTIGENS FROM PSEUDOMONAS-AERUGINOSA AND STAPHYLOCOCCUS-AUREUS STUDIED BY QUANTITATIVE IMMUNOELECTROPHORESIS [J].
BAEK, L ;
HOIBY, N ;
HERTZ, JB ;
ESPERSEN, F .
JOURNAL OF CLINICAL MICROBIOLOGY, 1985, 22 (02) :229-237
[2]   MELIOIDOSIS - A MAJOR CAUSE OF COMMUNITY-ACQUIRED SEPTICEMIA IN NORTHEASTERN THAILAND [J].
CHAOWAGUL, W ;
WHITE, NJ ;
DANCE, DAB ;
WATTANAGOON, Y ;
NAIGOWIT, P ;
DAVIS, TME ;
LOOAREESUWAN, S ;
PITAKWATCHARA, N .
JOURNAL OF INFECTIOUS DISEASES, 1989, 159 (05) :890-899
[3]   THE BACTERIAL GLYCOCALYX IN NATURE AND DISEASE [J].
COSTERTON, JW ;
IRVIN, RT ;
CHENG, KJ .
ANNUAL REVIEW OF MICROBIOLOGY, 1981, 35 :299-324
[4]   IMMUNOLOGIC STUDIES WITH MALLEOMYCES-MALLEI AND MALLEOMYCES-PSEUDOMALLEI .2. AGGLUTINATION AND COMPLEMENT FIXATION TESTS IN MAN AND LABORATORY ANIMALS [J].
CRAVITZ, L ;
MILLER, WR .
JOURNAL OF INFECTIOUS DISEASES, 1950, 86 (01) :52-62
[5]  
Dance D. A. B., 1990, Reviews in Medical Microbiology, V1, P143
[6]   IDENTIFICATION OF PSEUDOMONAS-PSEUDOMALLEI IN CLINICAL-PRACTICE - USE OF SIMPLE SCREENING-TESTS AND API20NE [J].
DANCE, DAB ;
WUTHIEKANUN, V ;
NAIGOWIT, P ;
WHITE, NJ .
JOURNAL OF CLINICAL PATHOLOGY, 1989, 42 (06) :645-648
[7]   MELIOIDOSIS - THE TIP OF THE ICEBERG [J].
DANCE, DAB .
CLINICAL MICROBIOLOGY REVIEWS, 1991, 4 (01) :52-60
[8]   PSEUDOMONAS-PSEUDOMALLEI - DANGER IN THE PADDY FIELDS [J].
DANCE, DAB .
TRANSACTIONS OF THE ROYAL SOCIETY OF TROPICAL MEDICINE AND HYGIENE, 1991, 85 (01) :1-3
[9]   COLORIMETRIC METHOD FOR DETERMINATION OF SUGARS AND RELATED SUBSTANCES [J].
DUBOIS, M ;
GILLES, KA ;
HAMILTON, JK ;
REBERS, PA ;
SMITH, F .
ANALYTICAL CHEMISTRY, 1956, 28 (03) :350-356
[10]  
HARLOW E, 1988, ANTIBODIES LAB MANUA, P522