Rapid diagnostics tests (RDTs) using anti-pan-Plasmodium lactate dehydrogenase (pLDH) antibody have been widely used along with peripheral blood smears (PBS) for the diagnosis of malaria. Malaria parasitemia, as determined by PBS, correlates with pLDH concentration in most cases [1]. Here, we report a case of malaria recurrence associated with false-negative RDT results, suggesting that caution must be exercised in the diagnosis of malaria when using anti-pLDH antibody.

A 22-year-old man was admitted to the hospital following 4 days of fever and chills. One month prior, the patient, with no remarkable medical history, was admitted to the Armed Forces Capital Hospital and was diagnosed with malaria. He was administered chloroquine for 3 days and primaquine for 11 days, and his symptoms subsided. However, the symptoms recurred 4 days before admission to our hospital. On admission, his leukocyte count was 6,400/µL; hemoglobin, 14.8 g/dL; platelet, 78,000/µL; and C-reactive protein was elevated, 3.61 mg/dL. No pathogens were isolated from sputum or throat cultures. PBS was performed and more than 300 fields were examined at high magnification (×400). At the time of admission, both PBS and RDT (Malaria Ag Pf/Pan test, Standard Diagnostics, Yongin, Korea) results were negative.

Due to negative RDT and PBS results and low clinical suspicion of malaria reinfection or recurrence, clinicians initiated a course of doxycycline for suspected epidemic hemorrhagic fever (EHF). On the 3rd day of hospitalization, PBS was repeated and yielded a positive result with a titer of 1,131/µL, rising to 1,360/µL on the 5th day. However, RDTs performed on both days remained negative. The patient was administered hydroxychloroquine and primaquine, and PBS results were negative after three days of treatment; he was discharged 10 days after admission (Fig. 1).

To evaluate the inconsistency between the RDT and PBS results on the 3rd and 5th days, the RDT was re-evaluated using internal quality control samples, but the results did not differ. A prozone phenomenon due to a high level of pLDH has been previously described [2], so to investigate this potential explanation, the specimens were diluted 2 and 4 times with normal saline; however, the results of this test were also negative.

Diagnosis of malaria based only on RDTs is cost-effective and operationally advantageous, since RDTs are easy to perform and provide results rapidly [3]. Most RDTs detect parasite histidine-rich protein 2 (pHRP-2) and pLDH from Plasmodium falciparum and Plasmodium spp., respectively. While the pHRP-2 antigen can be detected up to 40 days post-antibiotics treatment [4], the pLDH antigen has very short half-life of less than 2 days, and the median time on treatment for this test to become negative was reported as just 2 days [5]. Even though RDTs show good correlation with PBS results and can be used for evaluation of the treatment response and follow-up, low blood levels of pLDH can lead to false-negative results [16]. We were not able to evaluate the pLDH level in this patient's sample; the parasitemia was much higher than that reported previously with low detectable pLDH (<1,000/µL) [16]. Similarly, Jang et al. [1] reported 7 cases of high parasitemia with low pLDH levels, but did not provide additional detailed data regarding treatment history. pLDH levels reflect the level of viable parasitemia [7], and the World Health Organization guidelines also state that parasitemia is the most important criteria for confident diagnosis [8].

In this case, we suspect that doxycycline administration may have influenced the RDT results. Doxycycline exhibits antimicrobial activity via inhibition of bacterial cell wall synthesis and eliminates malarial parasites in hepatocytes. Due to its effectiveness, doxycycline is administered as a therapeutic and preventive drug for malaria [9]. Despite a lack of studies reporting a direct association between doxycycline treatment and RDT results, Pasricha et al. [10] showed that 50% of patients with malarial infection and negative PBS and RDT results at the time of admission, had a history of treatment with doxycycline. Even though anti-malarial drugs decrease the number of parasites in the blood [11], in our case, the malarial parasite titers were not low. These reports suggest that antibiotics administration lowered the pLDH concentration due to a decreased number of viable parasites, resulting in false-negative RDT results. Therefore, in patients receiving doxycycline, serial PBS testing should be performed to exclude false-negative malarial RDT results. Further studies are required to elucidate the effect of doxycycline administration on RDT results.