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Home / Equine Research Bank / PLoS Negl Trop Dis / Development of a sensitive competitive enzyme-linked immunos...
PLoS neglected tropical diseases2021; 15(12); e0010007; doi: 10.1371/journal.pntd.0010007

Development of a sensitive competitive enzyme-linked immunosorbent assay for serodiagnosis of Burkholderia mallei, a Tier 1 select agent.

Abstract: Glanders is a highly contagious and potentially serious disease caused by Burkholderia mallei, a Tier 1 select agent. In this study, we raised a monoclonal antibody (mAb) against the lipopolysaccharide (LPS) of B. mallei and developed a competitive enzyme-linked immunosorbent assay (cELISA) for B. mallei infection. Using the titrated optimal conditions of B. mallei-LPS (2 ng) for microtiter plate coating, sample serum dilution at 1:20 and 3.5 ng/μL anti-LPS mAb B5, the cutoff value of the cELISA was determined using serum samples from 136 glanders-free seronegative horses in Hong Kong. All calculated percentage inhibition (PI) values from these seronegative samples were below 39.6% inhibition (1.5 standard deviations above mean PI) and was used as the cutoff value. The diagnostic sensitivity of the developed LPS-based cELISA was first evaluated using sera from donkeys and mice inoculated with B. mallei. An increasing trend of PI values above the defined cELISA cutoff observed in the donkey and mouse sera suggested positive detection of anti-LPS antibodies. The sensitivity and specificity of the LPS-based cELISA was further evaluated using 31 serologically positive horse sera from glanders outbreaks in Bahrain and Kuwait, of which 30 were tested positive by the cELISA; and 21 seronegative horse sera and 20 seronegative donkey sera from Dubai, of which all were tested negative by the cELISA. A cELISA with high sensitivity (97.2%) and specificity (100%) for the detection of B. mallei antibodies in different animals was developed.
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Publication Date: 2021-12-21 PubMed ID: 34932554PubMed Central: PMC8691619DOI: 10.1371/journal.pntd.0010007Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research outlines the development of a highly sensitive competitive enzyme-linked immunosorbent assay (cELISA) for detecting Burkholderia mallei, the bacteria behind a potentially serious contagious disease known as Glanders. The method is based on identifying the presence of a specific monoclonal antibody against lipopolysaccharide (LPS) of B. mallei, which was found to have a high sensitivity and specificity in different animals.

Creation of the cELISA

  • The researchers created a monoclonal antibody (mAb) against the lipopolysaccharide (LPS) of B. mallei, a bacteria that causes the highly contagious disease known as Glanders.
  • The optimal conditions for the cELISA were determined and involved B. mallei-LPS (2 ng) for microtiter plate coating and sample serum dilution at 1:20 and 3.5 ng/μL anti-LPS mAb B5.
  • The cELISA’s cutoff value was determined using serum samples from 136 glanders-free seronegative horses in Hong Kong. The calculated percentage inhibition (PI) values from these samples served to establish the threshold of the cELISA, below which a negative result is presumed.

Diagnostic Applications and Evaluations

  • Initially, the researchers checked the sensitivity of the LPS-based cELISA using sera from donkeys and mice that had been inoculated with B. mallei.
  • These results showed an increasing trend of the PI values, suggesting that the anti-LPS antibodies had been detected. This indicated that the cELISA was effective in identifying the presence of B. mallei.

Further Testing on Real-Life Samples

  • Upon initial success, the team went on to evaluate the sensitivity and specificity of the cELISA by testing 31 positive horse sera from glanders outbreaks in Bahrain and Kuwait. Of these, 30 tested positive by the cELISA.
  • Moreover, 21 seronegative horse sera and 20 seronegative donkey sera from Dubai were all also tested, and each was shown to be negative by the cELISA.
  • The sensitivity (97.2%) and specificity (100%) of the developed cELISA was thus established, meaning that the assay is highly efficient in distinguishing true positives and true negatives in regards to the presence of B. mallei antibodies in different animals.

Cite This Article

APA
Wernery U, Chan E, Raghavan R, Teng JLL, Syriac G, Siu SY, Joseph M, Yeung ML, Jia L, Cai JP, Chiu TH, Lau SKP, Woo PCY. (2021). Development of a sensitive competitive enzyme-linked immunosorbent assay for serodiagnosis of Burkholderia mallei, a Tier 1 select agent. PLoS Negl Trop Dis, 15(12), e0010007. https://doi.org/10.1371/journal.pntd.0010007

Publication

PLoS neglected tropical diseases
ISSN: 1935-2735
NlmUniqueID: 101291488
Country: United States
Language: English
Volume: 15
Issue: 12
Pages: e0010007

Researcher Affiliations

Wernery, Ulrich
  • Central Veterinary Research Laboratory, Dubai, United Arab Emirates.
Chan, Elaine
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
Raghavan, Rekha
  • Central Veterinary Research Laboratory, Dubai, United Arab Emirates.
Teng, Jade L L
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
  • State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong.
  • Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong.
Syriac, Ginu
  • Central Veterinary Research Laboratory, Dubai, United Arab Emirates.
Siu, Sing-Yung
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
Joseph, Marina
  • Central Veterinary Research Laboratory, Dubai, United Arab Emirates.
Yeung, Man-Lung
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
  • State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong.
  • Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong.
Jia, Lilong
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
Cai, Jian-Piao
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
Chiu, Tsz-Ho
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
Lau, Susanna K P
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
  • State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong.
  • Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong.
Woo, Patrick C Y
  • Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
  • State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong.
  • Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong.

MeSH Terms

  • Animals
  • Antibodies, Bacterial / blood
  • Burkholderia mallei / immunology
  • Burkholderia mallei / isolation & purification
  • Enzyme-Linked Immunosorbent Assay / methods
  • Equidae
  • Glanders / blood
  • Glanders / diagnosis
  • Glanders / microbiology
  • Horse Diseases / blood
  • Horse Diseases / diagnosis
  • Horse Diseases / microbiology
  • Horses
  • Mice
  • Sensitivity and Specificity
  • Serologic Tests / methods

Conflict of Interest Statement

The authors have declared that no competing interests exist.

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