Reply to Awandu et al

To the Editor—We thank Awandu and colleagues for their insightful comments on our recent article describing ultrasensitive diagnostic tests for detection of asymptomatic malaria in a highly endemic region of Gambella, Ethiopia [1]. The higher prevalence noted in Gambella based on ultrasensitive methods is most likely due to the improved limit of detection afforded by targeting highly abundant RNA. To test this hypothesis, we evaluated a subset (n = 48) of the P. falciparum-positive samples from the Gambella study using traditional nested polymerase chain reaction (PCR) [2]. Only 60.4% (29/48) was detected positive by nested PCR (Table 1). Therefore, using standard molecular methods, the prevalence is far lower and more in line with the meta-analysis quoted [3]. We summarize the various molecular methods and their limits of detection (LOD) in Table 2. Careful consideration has to be made when combining prevalence studies using different molecular methods due to the widely varying LOD. Nested Polymerase Chain Reaction (PCR) Results of Samples Positive for Plasmodium falciparum by Quantitative Reverse Transcriptase PCR (n = 48) From the Gambella Study Abbreviation: PCR, polymerase chain reaction. Nested Polymerase Chain Reaction (PCR) Results of Samples Positive for Plasmodium falciparum by Quantitative Reverse Transcriptase PCR (n = 48) From the Gambella Study Abbreviation: PCR, polymerase chain reaction. Variation in the Limit of Detections of the Different Molecular Tools for Diagnosing Plasmodium falciparum (modified from [12]) Abbreviations: LOD, limit of detection; NASBA, nucleic acid sequence-based amplification; PCR, polymerase chain reaction; qPCR, quantitative polymerase chain reaction; qRT-PCR, quantitative reverse transcriptase polymerase chain reaction; rRNA, ribosomal RNA; RT, reverse transcriptase; TARE-2, telomere-associated repetitive element 2; US-LAMP, ultrasensitive loop mediated amplification. Variation in the Limit of Detections of the Different Molecular Tools for Diagnosing Plasmodium falciparum (modified from [12]) Abbreviations: LOD, limit of detection; NASBA, nucleic acid sequence-based amplification; PCR, polymerase chain reaction; qPCR, quantitative polymerase chain reaction; qRT-PCR, quantitative reverse transcriptase polymerase chain reaction; rRNA, ribosomal RNA; RT, reverse transcriptase; TARE-2, telomere-associated repetitive element 2; US-LAMP, ultrasensitive loop mediated amplification. The role that very low level infections (<100 parasites per mL) detected only by ultrasensitive methods play in onward transmission remains unclear and necessitates further study. Despite no linear correlation between parasite count and gametocyte count, low parasitemia infections tend to produce a smaller number of infective gametocytes [4]. However, transmission depends not only on the number of gametocytes produced but also on the efficiency of the vectors, vector diversity, and the number of bites [5–7]. Furthermore, parasitemia can fluctuate in the asymptomatic individual and longitudinal studies suggest that gametocyte counts can also oscillate, implying these very low level infections may ultimately lead to onward transmission [8]. More studies like that of Hoffman et al on ultrasensitive rapid diagnostic tests and gametocyte carriage in asymptomatic individuals are needed, especially when coupled to membrane feeding assays [9]. Some studies have attempted to model the reservoir and the probability of onward transmission, but these models require validation [10]. Ultimately, public health programs need to evaluate the role of ultrasensitive diagnostics in reactive case detection, for example, in order to determine the value of such tools in accelerating elimination [11]. Potential conflicts of interest. All authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.

Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.