Early diagnosis of dengue depends on the detection of the virus by either nucleic acid amplification test (NAAT) [
1‐
4] or detection of dengue virus (DENV) nonstructural protein 1 (NS1) antigen especially that configured into the rapid detection test (RDT) format [
5‐
9]. The NS1 detection method is among the most widely used as it is rapid and simple to perform [
9]. The method, however, has its limitation, especially when utilized in dengue endemic regions where secondary dengue is common [
5,
9‐
11]. NS1 assay sensitivity in detection of secondary dengue infection is much lower, hence, may contribute to false negative results [
12,
13]. A complementary detection method is therefore needed [
14]. The NAAT has been suggested as the most suitable complementary test since the test allows for direct detection of DENV genome from samples of patients obtained during the viremic phase (< 5 days after fever onset). While there are a number of NAATs available, the most common NAAT method for detection of DENV has been the quantitative reverse-transcription polymerase chain reaction (RT-PCR) [
15‐
17]. The test is highly sensitive, specific, and can be easy to perform especially by trained personnel. Unfortunately, due to its requirement for highly specific equipment and reagents, usage of the test has been confined to the well-funded and well-equipped referral laboratories [
18,
19]. The use of NAAT in a resource-limited setting such as peripheral laboratories in many dengue endemic regions of the Southeast Asia is, therefore still limited [
20‐
22]. In recent years, extensive efforts have been undertaken to develop NAAT for the use in these resource-limited settings for various infectious diseases [
23‐
26]. Implementation of the NAAT as a preferred diagnostic test in this setting, however, remained challenging. The ideal diagnostic test should be a simple, rapid, sensitive, specific, and requires minimal laboratory infrastructure [
27]. A more cost-effective NAAT format that met all the aforementioned criteria hence, is needed.
The invention of the isothermal NAAT that requires no or minimum laboratory infrastructure has the potential to overcome the barrier to the use of NAAT in resource-limited setting [
28‐
31]. The isothermal NAAT usually has a simple protocol, easy to perform, does not require a sophisticated instrument, and straightforward result interpretation procedure [
32‐
34]. We recently reported a simple, rapid, sensitive, and specific single tube pan-dengue reverse-transcription recombinase polymerase amplification (RT-RPA) method for early detection of DENV [
1]. This method showed comparable sensitivity to the reference real-time RT-PCR test. The dengue RT-RPA assay was performed on an inexpensive portable fluorometer and took only approximately 20 min to perform with minimal reagent and equipment cost. This NAAT assay hence, has a potential for use in a resource-limited setting [
1]. However, a diagnostic assay with excellent performance in itself is insufficient. The feasibility of a diagnostic test for use in resource-limited setting relies heavily on the robustness of the assay and acceptability by the end users. With this in mind, in the present study, we assessed the operational utilities of the previously described dengue RT-RPA assay, which includes ease to use, the time required to perform the assay, and user acceptability.