The 2021 World Health Statistics Report released by the World Health Organization (WHO) notes that pathogens are still one of the important causes of harm to human health and life, especially for children under 6 years of age [1]. Respiratory infections, diarrhea and malaria caused by pathogens are important causes of death [2]. Among the various types of pathogens, pathogenic microorganism transmission is more difficult to detect, and outbreaks are more sudden, difficult to control, and often more destructive. Taking severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as an example, since December 2019, coronavirus disease (COVID-19) caused by the virus has continued to rapidly spread worldwide and is by far the largest public health event. According to the WHO, as of July 25, 2022, nearly 570 million people around the world had been infected with SARS-CoV-2, and more than 6.35 million people had died, seriously threatening human life and safety [3]. With the emergence of mutant strains such as Delta and Omicron and asymptomatic infections, the early diagnosis of the disease and the prevention and control of the epidemic have presented huge challenges [4].

Early strategies for pathogenic microorganism detection were mainly based on the isolation and culture of pathogens, which is very time consuming and labor intensive, and it is difficult to meet the needs for the prevention and control of sudden epidemic situations using sus methods. Although the detection of pathogenic microorganisms based on immunological methods, which are now commonly used, is simple and rapid, the application of this approach is limited due to the existence of a detection window [5]. The development of polymerase chain reaction (PCR)-based methods has led to revolutionary changes in the detection of pathogenic microorganisms [6]. By amplifying the nucleic acid sequences of pathogenic microorganisms by PCR, it has become possible to detect a single pathogenic microorganism with high sensitivity and high specificity [7]. Quantitative PCR and digital PCR technology can even achieve accurate quantification of pathogenic microorganisms [8,9]. However, the reliance of this methodology on thermal cyclers makes it difficult to expand the PCR-based nucleic acid detection of pathogenic microorganisms to primary medical units or remote areas with limited resources. In the early 1990s, some scholars began to apply nucleic acid isothermal amplification technology and combined it with other technologies, leading to the widespread use of nucleic acid isothermal amplification technology in many fields. Compared with PCR, nucleic acid isothermal amplification technology does not require a long reaction time, and the requirements for the reaction instrument are not overly strict, allowing the rapid screening and detection of pathogenic microorganisms to be realized [10].

According to the characteristics of nucleic acid amplification at a specific temperature, isothermal amplification techniques have received much attention. The main categories of this technology are strand displacement amplification (SDA), loop-mediated isothermal amplification (LAMP), helicase-dependent isothermal DNA amplification (HDA), recombinase polymerase amplification (RPA) and rolling circle amplification (RCA) [11], [12], [13]. Compared with traditional methods and PCR technology, isothermal amplification technology can achieve accurate and rapid detection in relatively imperfect laboratory facilities.

RPA is a nucleic acid isothermal amplification technology developed by the British company TwistDx Inc. in 2006 [14]. In RPA technology, a complex formed by recombinase combined with primers identifies homologous sequences in the template, which a triggers strand exchange reaction, initiates DNA synthesis after localization, and exponentially amplifies the target region of the template (Fig. 1). RPA technology can perform nucleic acid amplification rapidly at a constant temperature of 25°C to 42°C, and the products can be monitored in real time by fluorescence quantification with probe-based methods. This technology can also be combined with the use of side-flow chromatography strips, biochips, gel electrophoresis and other methods of detection [15,16]. In this paper, the principle, development process, advantages and disadvantages of RPA technology as well as its application for the detection of viruses, bacteria, fungi and other pathogens are summarized, and the direction of the future development of this technology is considered, together with domestic and foreign research progress.