Analyses were based on 8897 patients, 89.6% treated with an ACT and 10.4% with non-ACTs (Additional file 2: Table S2, Additional file 3: Fig. S1), providing 38,864 Hb measurements (93% and 7% for ACT and non-ACT, respectively).
Study characteristics are in Additional file 4: Table S3: 98% (n = 8728) of subjects had at least one other recorded value post-D0 (58% at D3, 80% at D7, 73% at D28); 18% (n = 1645) children <5 years-old enrolled in Liberia, Mozambique, Rwanda and Uganda had daily measurements on D0 through D3; an additional 11% (n = 1000) had daily measurements on D0 through D2. All but two studies (Uganda and Rwanda) had at least three measurements: D0, D28 and one in-between: D7 (11%, in four countries), D14 (20%, two countries), and D3, D7, and D14 (17%, two countries).
Patients’ baseline characteristics by study are in Table 1 and Fig. 1: 77% were children <5 years of age, 24.1% were 5–11 years-old, 4.5% were 12–14 years-old, and 11.0% ≥15. The geometric mean D0 parasite density was 13,790 overall, ranging from 594 (Liberia [19]) to 39,023 parasites/μl (Mozambique [13]).
Anaemia and haemoglobin levels on presentation with acute falciparum malaria
Sixty-seven percent (67.3%, n = 5978/8897) of subjects had anaemia on D0, 63% mild or moderate. The mean D0 Hb was 10.2 g/dl (standard deviation 2.05) and the median 10.1 g/dl (range 3.4–18.3).
Hb increased with age (from a median of 9.6 g/dl in < 5 years-old to 14.1 g/dl in males ≥15 years), corresponding to a decreasing prevalence of anaemia (from 77.6% to 22.1%, respectively, Fig. 2).
A multivariate logistic regression model with mixed effects found that the risk of D0 anaemia decreased significantly with increasing age (p = 0.001 for all comparisons vs. under 5’s). Parasitological recurrence during follow-up was more frequent in subjects with D0 anaemia (AOR 1.45, 95% CI 1.23–1.73, p = 0.001).
Overall, higher D0 parasite density correlated with lower Hb by linear regression, mixed effect model (Fig. 3) except for <5 year-olds who had higher D0 geometric mean parasite densities (43,938 μl) than older subjects (2784 μl) (Additional file 5: Fig. S2).
Anaemia during treatment and follow-up
Occurrence of anaemia
From D0 to last observation, 82.7% of the patients had at least one Hb measurement fulfilling the anaemia criteria (any grade of severity), corresponding to 26,408 (67.9%) of the total 38,864 Hb measurements; of these, 20,430 (77.4%) occurred after treatment start (D1-D28). Moderate or more severe anaemia (≥grade 2) occurred in 47.6% of all measurements (18,486/38,864), in 76.5% of episodes (14,148/18,486) after treatment start (D1-D28). Post-D0 anaemia was less frequent on ACT: all-grade (77.3% vs. 80.3%, p = 0.036); ≥grade 2 anaemia (76.4% vs. 81.1%, p = 0.007).
Severity of anaemia
Five patients had very severe anaemia (grade 4) at D0. Overall, 19 patients had 27 records of transient very severe anaemia, corresponding to 2/1000 patients (19/8897). All episodes of very severe anaemia occurred by D14 in ACT groups, corresponding to 1 per 1000 (27/26,408) of all anaemia episodes, with no difference between treatments. Only one patient had very severe anaemia on D28 concomitant with recurrent infection, while all other patients recovered after D14. No case of very severe anaemia was observed in patients treated with non-ACT.
Changes in anaemia status
Of the 8720 patients with at least one additional post-D0 Hb measurement, 67.9% were anaemic on presentation and 47.1% when last seen – the latter was composed of 41.9% who remained anaemic plus an additional 5.2% who became anaemic post-D0. Anaemia grade worsened in 31.9% patients.
Mixed-effect multivariate logistic regression model showed an increased risk of anaemia worsening in patients with higher D0 parasitaemia (AOR 1.65, 95% CI 1.52–1.79, p = 0.001) and Hb (AOR 1.57, 95% CI 1.51–1.63, p = 0.001), younger age (AOR 0.95, 95% CI 0.94–0.97, p = 0.001, per year of age). It detected no significant association with parasitological failure (AOR 0.94, 95% CI 0.81–1.12, p = 0.452) and no difference between treatments, except a lower risk with AQ + SP (AOR 0.51, 95% CI 0.36–0.72, p = 0.001) compared to ASAQ.
Haemoglobin levels during treatment and follow-up
Haemoglobin levels changes (D0 – D3 – D28)
Ten countries and 15 sites (Burkina Faso, Gabon, Liberia, Mali, Mozambique, Nigeria, Rwanda, Uganda, Zambia, Zanzibar) had Hb measurements on D0, D3 and then weekly from D7-D28 (n = 4886 inclusive of the 1645 above; median age 2 years, range 6 months to 14 years). Between D0 and D28, Hb increased by +13.8% (from 9.4 to 10.7 g/dl).
Haemoglobin levels changes between D0 and D3
On D3, Hb concentrations decreased −4.6% (95% CI -4.3% to −4.9%), from 9.4 to 9.0 g/dl; such decrease was of −4.5% (95% CI −.2% to −4.8%), from 9.3 to 8.9 g/dl, in patients treated with ACT and −7% (95% CI -8.5%-5.6%) on AS. The D3 drop in Hb was greater for subjects with normal D0 Hb (from 11.9 to 10.8 g/dl, −9.3%, 95% CI -9.9% to −8.6%) than for those who were anaemic (from 8.9 to 8.6 g/dl, −3.4%, 95% CI -3.7% to −3.0%, p = 0.001).
A multivariate logistic regression model with mixed-effect showed that the risk was higher for patients with higher D0 parasitaemia (AOR 2.51, 95% CI 2.23–2.83, p = 0.001) and Hb (AOR 1.19, 95% CI 1.14–1.24, p = 0.001) and lower for older subjects (AOR 0.87, 95% CI 0.83–0.92, p = 0.001), while no significant association was found with parasitological failure (AOR 1.13, 95% CI 0.95–1.34, p = 0.177) and treatment (all comparisons p > 0.1).
Time to haemoglobin nadir, children <5 years-old
Two studies enrolled 1645 children <5 years-old (median age 2.3 years, range 0.5–4.9) in Liberia [19], Mozambique, Rwanda, and Uganda [13] and recorded Hb on D0-D3, D7, D14, D21, D28. D0 parasite density was 23,908 /μl overall but varied across sites (p = 0.001). Median time-to-nadir (Tn) was D2 Table 2), and was independent of treatment (all comparisons p > 0.1). The predicted median Tn calculated by linear regression was 35 h, ranging from 29 h with ASAQ in Uganda to 48 h with DP in Rwanda (Additional file 6: Fig. S3).
The nadir occurred in 78% between D1–3, with the highest proportion on D2 (34%). The highest rate of Hb decrease (−12.6%) and slope (−1.25) were at D1. The PRRd1 was highest (98.4%) for the patients whose Hb increased post-D0 (HDRd1 + 8.3%) and lowest (83.8%) in those with nadir at D3 (HDRd1–1.8%). Excluding the 11% subjects with no decrease, overall Hb dropped from 9.8 g/dl to 8.7 g/dl (−1.18 g/dl or - 11.4%, 95% CI -11.0% to −11.8%, p = 0.001) (Table 2). The patterns of evolution of Hb levels are shown in Additional file 7: Fig. S4.
Subjects whose Hb increased after baseline (nadir on D0) had significantly lower D0 parasitaemia (14,410 μl) and Hb (9.0 g/dl) than those whose Hb decreased post-D0 (parasite density = 25,403/μl; Hb = 9.9 g/dl, p = 0.001). Subjects with higher D0 parasitaemia had longer Tn (r = 0.281, p = 0.001, Spearman test); the correlation between parasitaemia and Tn occurring between D1 and D7 was also significant (r = 0.060, p = 0.021, Spearman test) (Fig. 4). Tn was longer, and the drop in Hb greater, in patients without D0 anaemia (mean Tn 3.0 days; Hb change −14.7) than those presenting with anaemia (mean Tn 2.1 days; Hb change −10.4%, p = 0.001). The PCT (but not PRRd1) of patients whose Hb increased right after treatment start was significantly faster (p = 0.001) than that of patients with a decrease in Hb (whatever the time to nadir); no difference was apparent for either parameters between the groups with shorter and longer time to nadir.
Different approaches using linear regression mixed-effect multivariate analyses produced consistent results, whether the model includes the PRRd1, PCT or HDRd1. The Hb percentage decrease at Tn was greater in subject with higher D0 Hb and parasitaemia (p = 0.001) and those with longer PCT (p = 0.001), and smaller in those with higher PRRd1. A slight difference between models was that the Hb percentage decrease was independent of age when PRRd1 is used in the model (p = 0.088) but lower in older patients (p = 0.010) when using PCT. It was lower in patients treated with AL or DP than ASAQ (p = 0.001 in all comparison except DP when using PCT, p = 0.070).
Special attention was paid to the interaction between Hb and parasitaemia at D1 (when on average ~ 93% of the parasites were cleared – see Table 2). The rate at which Hb fell (measured as Hb decrease rate from D0 to D1, HDRd1) was slower in patients with longer Tn and faster in patients with higher D0 Hb (p = 0.001). Concerning parasitaemia, HDRd1 was higher in patients with higher D0 parasitaemia (p = 0.001), and those with longer PCT (p = 0.001), but was unrelated to PRRd1 (p = 0.112) and age (p = 0.296 and p = 0.124 in either models). HDRd1 was less in patients treated with AL or DP than with ASAQ (p = 0.001).
Haemoglobin recovery post-treatment
After an initial decrease in Hb concentrations, a + 19.4% increase from D3 to D28 (from 8.9 to 10.7 g/dl) was observed with a steady significant linear increment in Hb of +0.065 g/dl/day throughout D28, i.e. +0.6% daily (Fig. 5). Patients with normal D0 Hb levels experienced a significant slower recovery (+0.047 vs. +0.073 g/dl/day, p = 0.001; corresponding to a daily Hb increase of +0.4% and +0.7%, respectively) than those with pre-treatment anaemia.
Linear regression mixed-effect model found that the recovery in under 5’s was not significantly different from the 5–15 years-old (p = 0.059), but that it was significantly faster in patients with anaemia on D0 than those without anaemia (p = 0.001). No between-treatment difference was detected.