Abstract
Burkholderia pseudomallei is the causative agent of melioidosis, a severe infection endemic to many tropical regions. Lipopolysaccharide (LPS) is recognized as an important virulence factor used by B. pseudomallei. Isolates of B. pseudomallei have been shown to express one of four different types of LPS (typical LPS, atypical LPS types B and B2, and rough LPS) and in vitro studies have demonstrated that LPS types may impact disease severity. The association between LPS types and clinical manifestations, however, is still unknown, in part because an effective method for LPS type identification is not available. Thus, we developed antigen capture immunoassays capable of distinguishing between the LPS types. Mice were injected with B or B2 LPS for atypical LPS-specific monoclonal antibody (mAb) isolation; only two mAbs (3A2 and 5B4) were isolated from mice immunized with B2 LPS. Immunoblot analysis and surface plasmon resonance demonstrated that 3A2 and 5B4 are reactive with both B2 and B LPS where 3A2 was shown to possess higher affinity. Assays were then developed using capsular polysaccharide-specific mAb 4C4 for bacterial capture and 4C7 (previously shown to bind typical LPS) or 3A2 mAbs for typical or atypical LPS strain detection, respectively. The evaluations performed with 197 strains of Burkholderia and non-Burkholderia species showed that the assays are reactive to B. pseudomallei and Burkholderia mallei strains and have an accuracy of 98.8% (zero false positives and two false negatives) for LPS typing. The results suggest that the assays are effective and applicable for B. pseudomallei LPS typing.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 358-367 |
| Number of pages | 10 |
| Journal | American Journal of Tropical Medicine and Hygiene |
| Volume | 96 |
| Issue number | 2 |
| DOIs | |
| State | Published - 2017 |
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ASJC Scopus subject areas
- Parasitology
- Infectious Diseases
- Virology
Cite this
Development of immunoassays for burkholderia pseudomallei typical and atypical lipopolysaccharide strain typing. / Nualnoi, Teerapat; Norris, Michael H.; Tuanyok, Apichai; Brett, Paul J.; Burtnick, Mary N.; Keim, Paul S; Settles, Erik W.; Allender, Christopher J.; AuCoin, David P.
In: American Journal of Tropical Medicine and Hygiene, Vol. 96, No. 2, 2017, p. 358-367.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Development of immunoassays for burkholderia pseudomallei typical and atypical lipopolysaccharide strain typing
AU - Nualnoi, Teerapat
AU - Norris, Michael H.
AU - Tuanyok, Apichai
AU - Brett, Paul J.
AU - Burtnick, Mary N.
AU - Keim, Paul S
AU - Settles, Erik W.
AU - Allender, Christopher J.
AU - AuCoin, David P.
PY - 2017
Y1 - 2017
N2 - Burkholderia pseudomallei is the causative agent of melioidosis, a severe infection endemic to many tropical regions. Lipopolysaccharide (LPS) is recognized as an important virulence factor used by B. pseudomallei. Isolates of B. pseudomallei have been shown to express one of four different types of LPS (typical LPS, atypical LPS types B and B2, and rough LPS) and in vitro studies have demonstrated that LPS types may impact disease severity. The association between LPS types and clinical manifestations, however, is still unknown, in part because an effective method for LPS type identification is not available. Thus, we developed antigen capture immunoassays capable of distinguishing between the LPS types. Mice were injected with B or B2 LPS for atypical LPS-specific monoclonal antibody (mAb) isolation; only two mAbs (3A2 and 5B4) were isolated from mice immunized with B2 LPS. Immunoblot analysis and surface plasmon resonance demonstrated that 3A2 and 5B4 are reactive with both B2 and B LPS where 3A2 was shown to possess higher affinity. Assays were then developed using capsular polysaccharide-specific mAb 4C4 for bacterial capture and 4C7 (previously shown to bind typical LPS) or 3A2 mAbs for typical or atypical LPS strain detection, respectively. The evaluations performed with 197 strains of Burkholderia and non-Burkholderia species showed that the assays are reactive to B. pseudomallei and Burkholderia mallei strains and have an accuracy of 98.8% (zero false positives and two false negatives) for LPS typing. The results suggest that the assays are effective and applicable for B. pseudomallei LPS typing.
AB - Burkholderia pseudomallei is the causative agent of melioidosis, a severe infection endemic to many tropical regions. Lipopolysaccharide (LPS) is recognized as an important virulence factor used by B. pseudomallei. Isolates of B. pseudomallei have been shown to express one of four different types of LPS (typical LPS, atypical LPS types B and B2, and rough LPS) and in vitro studies have demonstrated that LPS types may impact disease severity. The association between LPS types and clinical manifestations, however, is still unknown, in part because an effective method for LPS type identification is not available. Thus, we developed antigen capture immunoassays capable of distinguishing between the LPS types. Mice were injected with B or B2 LPS for atypical LPS-specific monoclonal antibody (mAb) isolation; only two mAbs (3A2 and 5B4) were isolated from mice immunized with B2 LPS. Immunoblot analysis and surface plasmon resonance demonstrated that 3A2 and 5B4 are reactive with both B2 and B LPS where 3A2 was shown to possess higher affinity. Assays were then developed using capsular polysaccharide-specific mAb 4C4 for bacterial capture and 4C7 (previously shown to bind typical LPS) or 3A2 mAbs for typical or atypical LPS strain detection, respectively. The evaluations performed with 197 strains of Burkholderia and non-Burkholderia species showed that the assays are reactive to B. pseudomallei and Burkholderia mallei strains and have an accuracy of 98.8% (zero false positives and two false negatives) for LPS typing. The results suggest that the assays are effective and applicable for B. pseudomallei LPS typing.
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U2 - 10.4269/ajtmh.16-0308
DO - 10.4269/ajtmh.16-0308
M3 - Article
VL - 96
SP - 358
EP - 367
JO - American Journal of Tropical Medicine and Hygiene
JF - American Journal of Tropical Medicine and Hygiene
SN - 0002-9637
IS - 2
ER -