Clinical relevance of low-density Plasmodium falciparum parasitemia in untreated febrile children: A cohort study

Abstract

Background: Low-density (LD) Plasmodium infections are missed by standard malaria rapid diagnostic tests (standard mRDT) when the blood antigen concentration is below the detection threshold. The clinical impact of these LD infections is unknown. This study investigates the clinical presentation and outcome of untreated febrile children with LD infections attending primary care facilities in a moderately endemic area of Tanzania.

Methods/findings: This cohort study includes 2,801 febrile pediatric outpatients (median age 13.5 months [range 2-59], female:male ratio 0.8:1.0) recruited in Dar es Salaam, Tanzania between 01 December 2014 and 28 February 2016. Treatment decisions were guided by a clinical decision support algorithm run on a mobile app, which also collected clinical data. Only standard mRDT+ cases received antimalarials. Outcomes (clinical failure, secondary hospitalization, and death) were collected in follow-up visits or interviews on days 3, 7, and 28. After patient recruitment had ended, frozen blood from all 2,801 patients was tested for Plasmodium falciparum (Pf) by ultrasensitive-quantitative polymerase chain reaction (qPCR), standard mRDT, and "ultrasensitive" mRDT. As the latter did not improve sensitivity beyond standard mRDT, it is hereafter excluded. Clinical features and outcomes in LD patients (standard mRDT-/ultrasensitive-qPCR+, not given antimalarials) were compared with those with no detectable (ND) parasitemia (standard mRDT-/ultrasensitive-qPCR-) or high-density (HD) infections (standard mRDT+/ultrasensitive-qPCR+, antimalarial-treated). Pf positivity rate was 7.1% (n = 199/2,801) and 9.8% (n = 274/2,801) by standard mRDT and ultrasensitive qPCR, respectively. Thus, 28.0% (n = 76/274) of ultrasensitive qPCR+ cases were not detected by standard mRDT and labeled "LD". LD patients were, on average, 10.6 months younger than those with HD infections (95% CI 7.0-14.3 months, p < 0.001). Compared with ND, LD patients more frequently had the diagnosis of undifferentiated fever of presumed viral origin (risk ratio [RR] = 2.0, 95% CI 1.3-3.1, p = 0.003) and were more often suffering from severe malnutrition (RR = 3.2, 95% CI 1.1-7.5, p = 0.03). Despite not receiving antimalarials, outcomes for the LD group did not differ from ND regarding clinical failures (2.6% [n = 2/76] versus 4.0% [n = 101/2,527], RR = 0.7, 95% CI 0.2-3.5, p = 0.7) or secondary hospitalizations (2.6% [n = 2/76] versus 2.8% [n = 72/2,527], RR = 0.7,95% CI 0.2-3.2, p = 0.9), and no deaths were reported in any Pf-positive groups. HD patients experienced more secondary hospitalizations (10.1% [n = 20/198], RR = 0.3, 95% CI 0.1-1.0, p = 0.005) than LD patients. All the patients in this cohort were febrile children; thus, the association between parasitemia and fever cannot be investigated, nor can the conclusions be extrapolated to neonates and adults.

Conclusions: During a 28-day follow-up period, we did not find evidence of a difference in negative outcomes between febrile children with untreated LD Pf parasitemia and those without Pf parasitemia. These findings suggest LD parasitemia may either be a self-resolving fever or an incidental finding in children with other infections, including those of viral origin. These findings do not support a clinical benefit nor additional risk (e.g. because of missed bacterial infections) to using ultrasensitive malaria diagnostics at a primary care level.

Fig 1. Population flow chart.

After patient recruitment had ended, frozen blood from all 2,801 patients was tested for Pf by ultrasensitive qPCR, standard mRDT, and ultrasensitive mRDT. As the latter did not improve sensitivity beyond standard mRDT, it is hereafter excluded. ALMANACH, ALgorithms for the MANagement of Acute CHildhood illnesses; e-POCT, electronic point of care test; mRDT, malaria rapid diagnostic test; Pf, Plasmodium falciparum, qPCR, quantitative polymerase chain reaction; sd-mRDT,.

Fig 2. Temporal and age distribution of patients with LD, HD, and ND parasitemia.

(A) Predicted probability of being assigned to the HD or LD group according to age (fitted fractional polynomial plot). Shaded regions are 95% CI; (B) Predicted probability of parasite density by age (fitted fractional polynomial plot); (C) Number of patients with HD and LD parasitemia identified for each month of the 2016 recruitment period. HD, high-density Pf infection, LD, low-density Pf infection; RDT, rapid diagnostic test; ND, no detectable Pf parasitemia; sd-mRDT, standard malaria RDT; us-qPCR, ultrasensitive quantitative polymerase chain reaction.

Fig 3. Distribution of severe illness and anemia between patients with LD, HD, or ND parasitemia.

(A) Prevalence of severe illness amongst all patients (left) versus a subgroup excluding severely malnourished patients (right). (B) Log parasite densities in Pf-positive cases (qPCR-detectable: LD and HD), in the presence or absence of severe illness. (C) Prevalence of patients with moderate-to-severe anemia. (D) Log parasite densities in Pf-positive cases (qPCR-detectable: LD and HD), in the presence or absence of moderate-to-severe anemia. Severe malnutrition is defined as weight-for-age Z-score < −3 and/or MUAC <5–11 cm. Moderate-to-severe anemia is defined as memoglobin <9 g/dl. *p < 0.05. HD, high-density Pf infection, LD, low-density Pf infection; RDT, rapid diagnostic test; MUAC, mid-upper arm circumference; ND, no detectable Pf parasitemia; ns, nonsignificant; sd-mRDT, standard malaria RDT; us-qPCR, ultrasensitive quantitative polymerase chain reaction.

Fig 4. Distribution of inflammatory markers between patients with LD, HD, or ND parasitemia.

Violin plots showing the distribution in the concentrations of (A) PCT and (B) CRP found among ND, LD, and HD groups. Thick black bands: median. Dotted lines: quartiles. CRP was measured using a categorical quantitation test. Four categories exist: <10 mg/L, 10–40 mg/L, 40–80 mg/L, and >80 mg/L. The lower limit of this range is represented in the plot. *p < 0.05. CRP, C-reactive protein; HD, high-density Pf infection; LD, low-density Pf infection; ND, undetectable Pf parasitemia; ns, nonsignificant; PCT, procalcitonin; Sd-mRDT, standard malaria rapid diagnostic test; us-qPCR, ultrasensitive quantitative PCR.

Fig 5. Distribution of clinical outcomes between patients with LD, HD, or ND parasitemia.

(A) Prevalence of clinical failure (patients not cured) by day 7 postconsultation, (B) Prevalence of patients requiring secondary hospital admission. See S2 Table for the definition of severe illness. *p < 0.05. HD, high-density Pf infection; LD, low-density Pf infection; ND, undetectable Pf parasitemia; ns, nonsignificant; PCT, procalcitonin; sd-mRDT, standard malaria rapid diagnostic test; us-qPCR, ultrasensitive quantitative PCR.