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Article

Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis

1
Research Center for Emerging Infections and Zoonoses, University of Veterinary Medicine Hannover, Foundation, 30559 Hannover, Germany
2
Lower Saxony State Office for Consumer Protection and Food Safety, 26203 Wardenburg, Germany
3
Field Station for Epidemiology in Bakum, University of Veterinary Medicine Hannover, Foundation, 30559 Hannover, Germany
4
Unit for Bioinformatics and Data Processing, Justus-Liebig-University Giessen, 35392 Giessen, Germany
5
Institute of Food Quality and Food Safety, University of Veterinary Medicine Hannover, Foundation, 30173 Hannover, Germany
6
Institute for Veterinary Food Science, Justus-Liebig-University Giessen, 35392 Giessen, Germany
*
Author to whom correspondence should be addressed.
Microorganisms 2021, 9(1), 41; https://doi.org/10.3390/microorganisms9010041
Received: 2 December 2020 / Revised: 19 December 2020 / Accepted: 21 December 2020 / Published: 25 December 2020
(This article belongs to the Special Issue Rapid Diagnosis of Microbial Pathogens)
Glaesserella parasuis is a fastidious pathogen that colonizes the respiratory tract of pigs and can lead to considerable economic losses in pig production. Therefore, a rapid detection assay for the pathogen, preferably applicable in the field, is important. In the current study, we developed a new and improved detection method using loop-mediated isothermal amplification (LAMP). This assay, which targets the infB gene, was tested on a collection of 60 field isolates of G. parasuis comprising 14 different serovars. In addition, 63 isolates from seven different closely related species of the family Pasteurellaceae, including A. indolicus, A. porcinus, and A. minor, and a species frequently found in the respiratory tract of pigs were used for exclusivity experiments. This assay showed an analytical specificity of 100% (both inclusivity and exclusivity) and an analytical sensitivity of 10 fg/µL. In further steps, 36 clinical samples were tested with the LAMP assay. An agreement of 77.1 (95% CI: 59.9, 89.6) and 91.4% (95% CI: 75.9, 98.2) to the culture-based and PCR results was achieved. The mean limit of detection for the spiked bronchoalveolar lavage fluid was 2.58 × 102 CFU/mL. A colorimetric assay with visual detection by the naked eye was tested to provide an alternative method in the field and showed the same sensitivity as the fluorescence-based LAMP assay. Overall, the optimized LAMP assay represents a fast and reliable method and is suitable for detecting G. parasuis in the laboratory environment or in the field. View Full-Text
Keywords: loop-mediated isothermal amplification (LAMP); Glaesserella parasuis; Haemophilus parasuis; pig; infB gene; rapid diagnostic; bronchoalveolar lavage fluid; BAL loop-mediated isothermal amplification (LAMP); Glaesserella parasuis; Haemophilus parasuis; pig; infB gene; rapid diagnostic; bronchoalveolar lavage fluid; BAL
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MDPI and ACS Style

Pilchová, V.; Seinige, D.; Hennig-Pauka, I.; Büttner, K.; Abdulmawjood, A.; Kehrenberg, C. Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis. Microorganisms 2021, 9, 41. https://doi.org/10.3390/microorganisms9010041

AMA Style

Pilchová V, Seinige D, Hennig-Pauka I, Büttner K, Abdulmawjood A, Kehrenberg C. Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis. Microorganisms. 2021; 9(1):41. https://doi.org/10.3390/microorganisms9010041

Chicago/Turabian Style

Pilchová, Veronika, Diana Seinige, Isabel Hennig-Pauka, Kathrin Büttner, Amir Abdulmawjood, and Corinna Kehrenberg. 2021. "Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis" Microorganisms 9, no. 1: 41. https://doi.org/10.3390/microorganisms9010041

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