Rapidly quantitative detection of Nosema ceranae in honeybees using ultra-rapid real-time quantitative PCR
- Author:
A-Tai TRUONG
1
;
Sedat SEVIN
;
Seonmi KIM
;
Mi-Sun YOO
;
Yun Sang CHO
;
Byoungsu YOON
Author Information
- Publication Type:Original Article
- From:Journal of Veterinary Science 2021;22(3):e40-
- CountryRepublic of Korea
- Language:English
-
Abstract:
Background:The microsporidian parasite Nosema ceranae is a global problem in honeybee populations and is known to cause winter mortality. A sensitive and rapid tool for stable quantitative detection is necessary to establish further research related to the diagnosis, prevention, and treatment of this pathogen.
Objectives:The present study aimed to develop a quantitative method that incorporates ultra-rapid real-time quantitative polymerase chain reaction (UR-qPCR) for the rapid enumeration of N. ceranae in infected bees.
Methods:A procedure for UR-qPCR detection of N. ceranae was developed, and the advantages of molecular detection were evaluated in comparison with microscopic enumeration.
Results:UR-qPCR was more sensitive than microscopic enumeration for detecting two copies of N. ceranae DNA and 24 spores per bee. Meanwhile, the limit of detection by microscopy was 2.40 × 104 spores/bee, and the stable detection level was ≥ 2.40 × 105 spores/ bee. The results of N. ceranae calculations from the infected honeybees and purified spores by UR-qPCR showed that the DNA copy number was approximately 8-fold higher than the spore count. Additionally, honeybees infected with N. ceranae with 2.74 × 104 copies of N. ceranae DNA were incapable of detection by microscopy. The results of quantitative analysis using UR-qPCR were accomplished within 20 min.
Conclusions:UR-qPCR is expected to be the most rapid molecular method for Nosema detection and has been developed for diagnosing nosemosis at low levels of infection.
