Assessment of therapeutic responses to gametocytocidal drugs in Plasmodium falciparum malaria

Abstract

Indirect clinical measures assessing anti-malarial drug transmission-blocking activity in falciparum malaria include measurement of the duration of gametocytaemia, the rate of gametocyte clearance or the area under the gametocytaemia-time curve (AUC). These may provide useful comparative information, but they underestimate dose-response relationships for transmission-blocking activity. Following 8-aminoquinoline administration P. falciparum gametocytes are sterilized within hours, whereas clearance from blood takes days. Gametocytaemia AUC and clearance times are determined predominantly by the more numerous female gametocytes, which are generally less drug sensitive than the minority male gametocytes, whereas transmission-blocking activity and thus infectivity is determined by the more sensitive male forms. In choosing doses of transmission-blocking drugs there is no substitute yet for mosquito-feeding studies.

Figure 1

Dose–response relationships for single-dose primaquine in reducing the infectivity of Plasmodium falciparum- infected subjects to anopheline mosquitoes. Pooled data from all studies conducted [13]. Vertical axes shows the proportions of fed anopheline mosquitoes which were infected. Upper (a): Oocyst formation assessed from blood sampled 48 hours after primaquine dose; Lower (b): Sporozoite formation assessed from blood sampled 48 hours after primaquine dose. Primaquine given with an artemisinin derivative is shown in green circles, and with no anti-malarial or a non-artemisinin derivative is shown in red circles. In these studies 29 patients received no primaquine. The size of the circle is proportional to the number of subjects in each group (shown within). Corresponding adult primaquine doses are indicated in square boxes.

Figure 2

Gametocyte clearance following different drug regimens in falciparum malaria. Times from appearance of gametocytaemia (= 0 if gametocytaemia present on admission) to disappearance of gametocytaemia after different seven-day drug regimens in adults with acute falciparum malaria studied in Thailand [4]. Primaquine was given daily in a dose of 0.25 mg base/kg or *0.5 mg/kg.

Figure 3

Durations of gametocytaemia following different drug regimens in falciparum malaria. The comparison of durations of Plasmodium falciparum gametocytaemia following quinine alone compared with quinine plus 0.25 mg base/kg from Pukrittayakamee et al. [4]. These are the same patients as in Figure 2.

Figure 4

Dose–response relationship for primaquine in reducing the duration of (female) gametocyte carriage assessed by Pfs25 transcripts. Mean (95%CI) duration of female gametocyte carriage in Ugandan children with falciparum malaria treated with artemether-lumefantrine (AL) and different doses of primaquine as reported by Eziefula et al.[16]. Duration was estimated by fitting of a deterministic compartmental mathematical model to repeated Pfs25 quantitative real time nucleic acid sequence- based analysis gametocyte prevalence estimates. The vertical dashed line indicates the set threshold for non- inferiority compared with the primaquine 0.75 mg/kg reference group (non-inferiority margin of 2.5 days). The currently recommended 0.25 mg/kg dose is indicated by the horizontal line.

Figure 5

Male and female gametocytaemia in falciparum malaria. Artificial infection with Plasmodium falciparum for malariatherapy showing typical delay between the peaks of asexual and sexual parasitaemia, and the respective densities of male and female gametocytes as reported by Ciuca et al.[22].

Figure 6

Proposed relationship between male and female gametocyte clearance and transmission blocking effects in falciparum malaria. If male gametocytes are more sensitive to transmission-blocking drugs than female gametocytes, and female gametocytes predominate, then gametocyte clearance times are determined by the female gametocytaemia, and transmission-blocking effects are determined mainly by male gametocytaemia. In this illustration of gametocytaemia responses to drug treatment clearance half-lives are one day for male (blue) and two days for female gametocytes (pink). The limits of gametocyte detection by microscopy and Pfs25mRNA (which detects predominantly females) are shown by the dotted lines. If a density of >1,000/mL was required for mosquito infection, then in this illustration the maximum duration of possible infectivity is three days compared to a clearance time measured by Pfs25mRNA of seven days. In fact, drugs such as primaquine sterilize rapidly (Figure 7) suggesting that loss of infectivity precedes gametocyte clearance so the post-treatment duration of infectivity in this illustration is likely to be < one day.

Figure 7

The temporal dissociation in falciparum malaria between changes in gametocytaemia and effects on infectivity following primaquine administration. Individual data from studies of mosquito infectivity following anti-malarial treatments of falciparum malaria with plasmoquine or primaquine in which oocyst assessments were made in mosquitoes which fed ~24 hours after drug administration [10]. Oocysts were assessed typically in 10-20 mosquitoes 6-7 days after feeding. Each pair of circles or diamonds represents a studied patient.

Figure 8

The effects of primaquine alone on asexual and sexual parasitaemia in falciparum malaria. Volunteer study reported by Rieckmann et al. [52] in which primaquine (45 mg base) only was given. The volunteers were infected with the Malayan Camp strain of P. falciparum. Anopheles stephensi was used as the vector. Volunteer 3 data are shown. Gametocytaemia was quantitated by microscopy, oocysts and sporozoites were assessed by microscopy after dissection (arrows are the days when mosquitoes were dissected, the numerator is the number positive, the denominator is the number of mosquitoes dissected). The subsequent infectivity of these mosquitoes (i.e., from the same batches) to other volunteers was also assessed; a different, healthy, non-immune volunteer was bitten by 75 mosquitoes before, and 12, 24 and 48 hours after primaquine had been given to volunteer 3; + (red) denotes that the volunteer became infected with falciparum malaria, 0 denotes the volunteer did not become infected.

Figure 9

Effects of very low dose primaquine on transmissibility of falciparum malaria. Malariatherapy with Plasmodium falciparum (Panama strain) in two patients who were treated with primaquine 3 mg daily (starting at vertical yellow arrow) as reported by Young et al[53]. Daily direct mosquito feeding was performed. Black line is asexual parasitaemia, vertical lines are temperature (fever), vertical boxes represent mosquito feeds and dark hatching shows proportion infected. Mosquitoes (10 each) were dissected for oocysts and sporozoites; red + = infected (≥8 of 10), grey 0 = none infected. Note that infectivity persisted until the fourth day of treatment in patient 1316 (upper panel), and the second day in patient 1321 (lower panel) and also that the young circulating gametocytes in patient 1316 were initially not infectious. There was a moderate and delayed effect on gametocytaemia (red dashed line).

Figure 10

Proposed approximate dose-response relationships for primaquine in falciparum malaria. Approximate dose-response relationships in falciparum malaria for primaquine inhibition of infectiousness (blue dashed curve) presumably reflecting susceptibility of male gametocytes, and effects on gametocyte clearance (pink dashed curve) reflecting susceptibility of female gametocytes. Arrow shows currently recommended transmission-blocking primaquine dose of 0.25 mg/kg. Studies are in progress to characterize these relationships more accurately.