Related Experiment Video
Updated: May 15, 2026
Rapid and Specific Detection of Acinetobacter baumannii Infections Using a Recombinase Polymerase Amplification/Cas12a-based System
Published on: April 25, 2025
Rapid visual detection of Mycoplasma gallisepticum by combining recombinase-aided amplification with lateral-flow
Huiyong Xuan1, Shu Wang1, XinYi Ren1
1Key Laboratory of Animal Disease Prevention and Control of Xinjiang Production and Construction Corps, College of Animal Science and Technology, Shihezi University, Shihezi city, Xinjiang province, 832000, China.
Abstract:
Mycoplasma gallisepticum (MG) is a significant avian pathogen that adversely affects chickens and turkeys globally and is classified as a notifiable disease agent by the World Organisation for Animal Health. Therefore, rapid and accurate diagnosis is a critical factor in controlling MG infections. This study established a simple, rapid, and visual detection method for MG using recombinase-aided amplification (RAA) combined with a lateral flow dipstick (LFD). Following primer and probe screening, the RAA-LFD method was optimized for primer concentration, probe concentration, reaction temperature, and time, and its specificity and sensitivity were analyzed. The results demonstrated that under isothermal conditions at 39°C, RAA could amplify the target gene within 20 minutes, and the products could be visualized via LFD within 5 minutes. The method showed no cross-reactivity with Mycoplasma synoviae, Mycoplasma bovis, Pasteurella multocida, Mannheimia haemolytica, Staphylococcus aureus, Salmonella spp., Escherichia coli, Listeria monocytogenes, Enterococcus faecium, or Enterococcus faecalis. Furthermore, the RAA-LFD method exhibited high sensitivity and good reproducibility, with a detection limit of 10 copies/μL. A total of 100 clinical samples suspected of MG infection were tested using RAA, PCR, and real-time fluorescence quantitative PCR (qPCR). The concordance rates between the RAA detection method and conventional PCR or qPCR were 96% and 98%, respectively. These results indicate that the established RAA-LFD method is easy to operate, highly specific, and sensitive, making it suitable for grassroots-level field diagnosis of MG infections.
Insights
A new recombinase-aided amplification (RAA) combined with lateral flow dipstick (LFD) method offers rapid, visual detection of Mycoplasma gallisepticum (MG) in avian species. This highly specific and sensitive test is ideal for field diagnosis of this notifiable avian pathogen.
Area of Science:
- Veterinary Microbiology
- Molecular Diagnostics
- Avian Pathology
Background:
- Mycoplasma gallisepticum (MG) is a significant global avian pathogen affecting chickens and turkeys.
- Accurate and rapid diagnosis of MG is crucial for disease control and is a priority for the World Organisation for Animal Health.
- Current diagnostic methods may lack the speed and accessibility required for effective field deployment.
Purpose of the Study:
- To develop a simple, rapid, and visual diagnostic method for Mycoplasma gallisepticum.
- To combine recombinase-aided amplification (RAA) with lateral flow dipstick (LFD) technology for MG detection.
- To optimize and validate the performance characteristics of the novel RAA-LFD assay.
Main Methods:
- Primer and probe screening for target gene amplification via RAA.
- Optimization of RAA reaction conditions (temperature, time, concentrations) and LFD visualization.
- Specificity testing against a panel of avian pathogens and sensitivity analysis using defined DNA concentrations.
Main Results:
- The optimized RAA-LFD assay detected MG within 20 minutes under isothermal conditions (39°C), with results visible on LFD in 5 minutes.
- The method demonstrated high specificity, showing no cross-reactivity with other Mycoplasma species or common bacterial pathogens.
- Sensitivity was determined to be 10 copies/μL, with high concordance (96-98%) compared to conventional PCR and qPCR in clinical samples.
Conclusions:
- The developed RAA-LFD assay is a highly specific, sensitive, and rapid method for Mycoplasma gallisepticum detection.
- Its ease of operation and visual output make it suitable for field applications and grassroots-level diagnostics.
- This innovative diagnostic tool can significantly aid in the control of MG infections in poultry populations.
