Blood-Stage Parasitaemia and Age Determine Plasmodium falciparum and P. vivax Gametocytaemia in Papua New Guinea

Abstract

A better understanding of human-to-mosquito transmission is crucial to control malaria. In order to assess factors associated with gametocyte carriage, 2083 samples were collected in a cross-sectional survey in Papua New Guinea. Plasmodium species were detected by light microscopy and qPCR and gametocytes by detection of pfs25 and pvs25 mRNA transcripts by reverse-transcriptase PCR (qRT-PCR). The parasite prevalence by PCR was 18.5% for Plasmodium falciparum and 13.0% for P. vivax. 52.5% of all infections were submicroscopic. Gametocytes were detected in 60% of P. falciparum-positive and 51% of P. vivax-positive samples. Each 10-fold increase in parasite density led to a 1.8-fold and 3.3-fold increase in the odds of carrying P. falciparum and P. vivax gametocytes. Thus the proportion of gametocyte positive and gametocyte densities was highest in young children carrying high asexual parasite densities and in symptomatic individuals. Dilution series of gametocytes allowed absolute quantification of gametocyte densities by qRT-PCR and showed that pvs25 expression is 10-20 fold lower than pfs25 expression. Between 2006 and 2010 parasite prevalence in the study site has decreased by half. 90% of the remaining infections were asymptomatic and likely constitute an important reservoir of transmission. However, mean gametocyte densities were low (approx. 1-2 gametocyte/μL) and it remains to be determined to what extent low-density gametocyte positive individuals are infective to mosquitos.

Fig 1. Age trends in P. falciparum and P. vivax blood-stage and gametocyte prevalence and density.

Error bars represent CI₉₅.

Fig 2. pvs25 transcript numbers versus light microscopically determined P. vivax gametocyte counts.

Dilution series were made before storing samples in RNAprotect (5 dilution steps) or after RNA extraction of samples stored in TRIzol (3 dilution steps). Dots represent means of technical triplicates (TRIzol) or quintuplicates (RNAprotect) of dilutions except for the highest concentration (no replicates). Shaded grey shows the CI₉₅ of the intercept of the regression of compiled sample sets.

Fig 3. Relationship between parasite density and probability that a sample is gametocyte positive.

Left: Probability (with CI₉₅) that a sample is gametocyte positive vs. parasite density, calculated using a general additive model. Right: correlation between gametocyte density (measured as pfs25 or pvs25 transcripts/uL) and parasite density by qPCR.