Detection of honeybee bacterial pathogens with
upconversion-linked immunosorbent assay

a 2021

Detection of honeybee bacterial pathogens with upconversion-linked immunosorbent assay

PASTUCHA, Matěj, Eliška ODSTRČILÍKOVÁ, Veronika POLÁCHOVÁ, Julian BRANDMEIER, Antonín HLAVÁČEK et. al.

Basic information

Original name

Detection of honeybee bacterial pathogens with upconversion-linked immunosorbent assay

Authors

PASTUCHA, Matěj, Eliška ODSTRČILÍKOVÁ, Veronika POLÁCHOVÁ, Julian BRANDMEIER, Antonín HLAVÁČEK, Hans-Heiner GORRIS, Zdeněk FARKA and Petr SKLÁDAL

Edition

UPCONline 2021, 2021

Other information

Language

English

Type of outcome

Konferenční abstrakt

Field of Study

10406 Analytical chemistry

Country of publisher

France

Confidentiality degree

není předmětem státního či obchodního tajemství

Organization unit

Faculty of Science

Abstract

V originále

European and American Foulbroods (EFB and AFB) are the most serious honeybee diseases caused by bacteria. In the case of an outbreak of either disease, an effective diagnosis method is necessary, sensitive enough to detect the pathogen before the manifestation of clinical symptoms. Bacterial pathogens are traditionally detected by cultivation on selective media. Cultivations are lengthy but very sensitive and have low requirements on the laboratory equipment. However, their limits become apparent when the bacterial strain is difficult to cultivate or when the results are needed quickly. Enzyme-linked immunosorbent assay (ELISA) is a classic method for the detection of various analytes. Modern nanoparticle labels provide improved sensitivity and enhanced properties compared to conventional labels. Luminescent nanoparticles can substitute traditional fluorophores and particularly the photon-upconversion nanoparticles (UCNP) bring the intriguing possibility of background-free luminescence measurement. We developed novel polyclonal antibodies detecting Melissococcus plutonius (EFB) and Paenibacillus larvae (AFB). In both cases, a “traditional” ELISA was not sensitive enough to reveal the early stages of the diseases, and we improved it by applying streptavidin-labeled UCNPs (BSA-coated UCNPs for EFB and PEGylated for AFB). With the upconversion readout, we achieved an LOD of 3.4×10^2 CFU mL−1 for M. plutonius and 2.9×10^3 CFU/mL for P. larvae. This represents a 400- and 22-fold improvement over the ELISA. Furthermore, we have successfully detected bacteria in spiked samples of bees, larvae, and hive debris.

Links

LQ1601, research and development project

Name: CEITEC 2020 (Acronym: CEITEC2020)

Investor: Ministry of Education, Youth and Sports of the CR

LTAB19011, research and development project

Name: Luminiscenční imunostanovení jako citlivý nástroj pro diagnostiku nemocí včel

Investor: Ministry of Education, Youth and Sports of the CR, Sensitive bioanalytical tools for the surveillance of honey bee diseases, INTER-ACTION

Citovat

PASTUCHA, Matěj, Eliška ODSTRČILÍKOVÁ, Veronika POLÁCHOVÁ, Julian BRANDMEIER, Antonín HLAVÁČEK, Hans-Heiner GORRIS, Zdeněk FARKA and Petr SKLÁDAL. Detection of honeybee bacterial pathogens with upconversion-linked immunosorbent assay. In UPCONline 2021. 2021.

@proceedings{1757238,
   author = {Pastucha, Matěj and Odstrčilíková, Eliška and Poláchová, Veronika and Brandmeier, Julian and Hlaváček, Antonín and Gorris, HansandHeiner and Farka, Zdeněk and Skládal, Petr},
   booktitle = {UPCONline 2021},
   language = {eng},
   title = {Detection of honeybee bacterial pathogens with upconversion-linked immunosorbent assay},
   year = {2021}
}

TY  - CONF
ID  - 1757238
AU  - Pastucha, Matěj - Odstrčilíková, Eliška - Poláchová, Veronika - Brandmeier, Julian - Hlaváček, Antonín - Gorris, Hans-Heiner - Farka, Zdeněk - Skládal, Petr
PY  - 2021
TI  - Detection of honeybee bacterial pathogens with upconversion-linked immunosorbent assay
N2  - European and American Foulbroods (EFB and AFB) are the most serious honeybee diseases caused by bacteria. In the case of an outbreak of either disease, an effective diagnosis method is necessary, sensitive enough to detect the pathogen before the manifestation of clinical symptoms. Bacterial pathogens are traditionally detected by cultivation on selective media. Cultivations are lengthy but very sensitive and have low requirements on the laboratory equipment. However, their limits become apparent when the bacterial strain is difficult to cultivate or when the results are needed quickly. Enzyme-linked immunosorbent assay (ELISA) is a classic method for the detection of various analytes. Modern nanoparticle labels provide improved sensitivity and enhanced properties compared to conventional labels. Luminescent nanoparticles can substitute traditional fluorophores and particularly the photon-upconversion nanoparticles (UCNP) bring the intriguing possibility of background-free luminescence measurement. We developed novel polyclonal antibodies detecting Melissococcus plutonius (EFB) and Paenibacillus larvae (AFB). In both cases, a “traditional” ELISA was not sensitive enough to reveal the early stages of the diseases, and we improved it by applying streptavidin-labeled UCNPs (BSA-coated UCNPs for EFB and PEGylated for AFB). With the upconversion readout, we achieved an LOD of 3.4×10^2 CFU mL−1 for M. plutonius and 2.9×10^3 CFU/mL for P. larvae. This represents a 400- and 22-fold improvement over the ELISA. Furthermore, we have successfully detected bacteria in spiked samples of bees, larvae, and hive debris.
ER  -

Detailed Information on Publication Record