1. Epidemiology and Global Health
  2. Immunology and Inflammation
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The complex relationship of exposure to new Plasmodium infections and incidence of clinical malaria in Papua New Guinea

  1. Natalie E Hofmann
  2. Stephan Karl
  3. Rahel Wampfler
  4. Benson Kiniboro
  5. Albina Teliki
  6. Jonah Iga
  7. Andreea Waltmann
  8. Inoni Betuela
  9. Ingrid Felger
  10. Leanne J Robinson  Is a corresponding author
  11. Ivo Mueller  Is a corresponding author
  1. Swiss Tropical and Public Health Institute, Switzerland
  2. University of Basel, Switzerland
  3. Walter and Eliza Hall Institute of Medical Research, Australia
  4. Papua New Guinea Institute of Medical Research, Papua New Guinea
  5. University of Melbourne, Australia
  6. Burnet Institute, Australia
  7. ISGlobal, Barcelona Centre for International Health Research, Hospital Clínic-University of Barcelona, Spain
  8. Institut Pasteur, France
Research Article
Cite this article as: eLife 2017;6:e23708 doi: 10.7554/eLife.23708
4 figures, 6 tables, 2 data sets and 5 additional files

Figures

Figure 1 with 2 supplements
P. falciparum and P. vivax molFOB (A), prevalence by qPCR (B) and LM (C) by week of follow-up.

Blue lines, P. falciparum; red lines, P. vivax; solid lines, placebo arm; dashed lines, PQ arm. Open circles in (B) mark enrolment qPCR prevalence for each species.

https://doi.org/10.7554/eLife.23708.005
Figure 1—figure supplement 1
Definition of new infections for calculating molFOB.

Definition of P. falciparum new infections in two exemplary children is shown. The study design and timelines of follow-up are shown in upper panel: enrolment visit (‘E’), followed by radical treatment (black bar ‘T’) and 235 days of follow-up. The presence of P. falciparum clones by sampling visit is visualized below. Columns represent sampling visits, rows represent P. falciparum msp2 alleles, that is distinct P. falciparum clones. Grey solid circles, P. falciparum negative sample, grey open circle, missing sample due to missed follow-up visit; red circle: sample positive for respective Pf-msp2 allele. New infections were defined as a positive sample preceded by two samples negative for this allele (black rectangles), excluding missed samples (see Child 2, allele F, days 120–200). The time point of new infections is marked by arrows for the two children.

https://doi.org/10.7554/eLife.23708.006
Figure 1—figure supplement 2
P. ovale and P. malariae prevalence by qPCR during follow-up.

Purple lines, P. ovale; green lines, P. malariae; solid lines, Placebo arm; dashed lines, PQ arm. Open circles mark enrolment qPCR prevalence for each species.

https://doi.org/10.7554/eLife.23708.007
Distribution of P.falciparum molFOB (A) and P. vivax molFOB by treatment arm (B).

Relative frequencies among the 466 children are shown.

https://doi.org/10.7554/eLife.23708.009
Heterogeneity in molFOB (A, B) and clinical episode risk (C, D) of P.falciparum (A, C) and P. vivax (B, D).

Upper panels show the kriging fit of model predictions of molFOB and clinical episode risk of children in both treatment arms. Lower panels show the standard error relative to the kriging estimate. Dots represent study participants’ houses and are color-coded according to village. Black lines: vehicle-accessible road; dark grey lines: vehicle-inaccessible road; light grey lines: river; red/white cross: health center or aid post; grey square: school or enrolment location. Maps were prepared using ArcGIS 10.2 (Esri, USA).

https://doi.org/10.7554/eLife.23708.010
The incidence of P.falciparum (A) and P. vivax (B) clinical episodes relative to molFOB.

Mean clinical episode incidence is shown as bars (left axis) and proportion of clinical episode incidence divided by molFOB as connected dots (right axis). Error bars represent 95% CIs. p-values refer to the differences between groups in the proportion of clinical episodes and new infections, assessed by Chi2 or Fisher’s exact test.

https://doi.org/10.7554/eLife.23708.014

Tables

Table 1
Characteristics of study participants by village.
https://doi.org/10.7554/eLife.23708.003
VillageN% femaleMean age (±SD)Mean weight (±SD)% LLIN use at enrolment*Mean LLIN use during follow-up
(%, range)
Mean Hb
(±SD)
Amahup119537.6 (±1.5)19.8 (±3.3)9999 (50–100)11.1 (±1.0)
Albinama99437.7 (±1.5)20.0 (±3.3)9597 (78–100)11.7 (±1.8)
Balanga54597.8 (±1.6)19.8 (±4.3)9699 (83–100)11.3 (±1.1)
Balif93517.8 (±1.5)20.3 (±3.3)9199 (69–100)11.7 (±1.2)
Bolumita70507.4 (±1.7)19.3 (±2.9)7792 (56–100)10.7 (±1.0)
Numangu31557.4 (±1.6)19.2 (±4.6)100100 (92–100)12.1 (±1.4)
Total466517.6 (±1.5)19.8 (±3.5)93100 (50–100)11.4 (±1.4)
  1. * LLIN use in the night preceding enrolment.

    † Information on LLIN use in the previous night was collected at each follow-up visit and averaged across follow-up per participant. Mean LLIN use by village was calculated from the averaged individual LLIN use.

  2. Hb: Haemoglobin.

Table 2
Plasmodium infection status at enrolment by village.
https://doi.org/10.7554/eLife.23708.004
VillageP. falciparumP. vivaxP. malariaeP. ovale
N pos.Prevalence by qPCR (CI95)% mixed*Mean density (IQR)Mean MOI (range)N pos.Prevalence by qPCR
(CI95)
% mixed*Mean density (IQR)Mean MOI (range)N pos.Prevalence by qPCR
(CI95)
% mixed*N pos.Prevalence by qPCR
(CI95)
% mixed*
Albinama1818 (11–27)72131 (38–189)1.4 (1–4)5455 (44–65)243 (1–17)1.8 (1–7)99 (5–17)6755 (2–12)100
Amahup1412 (7–19)5756 (14–105)1.6 (1–5)4639 (30–48)243 (1–29)2.2 (1–7)1210 (6–17)8300
Balanga1528 (17–42)6779 (30–848)1.7 (1–5)2343 (30–57)432 (1–28)2.0 (1–7)917 (8–30)5623 (0–14)50
Balif89 (4–17)6364 (10–325)2.0 (1–4)3538 (30–48)142 (1–14)1.9 (1–6)78 (3–15)5700
Bolumita5071 (59–81)80331 (62–1988)2.2 (1–8)4767 (55–78)813 (2–27)2.9 (1–10)2840 (29–52)89811 (5–22)100
Numangu826 (13–45)75192 (30–848)1.1 (1–2)1858 (39–75)283 (1–25)1.6 (1–5)413 (4–31)5000
Overall11324 (20–28)73163 (20–1103)1.9 (1–8)22348 (43–52)373 (1–23)2.2 (1–10)6915 (12–18)75153 (2–5)93
p-value§<0.0010.0340.0860.047<0.001<0.0010.9470.020<0.0010.086<0.0010.133
  1. * % of infections by qPCR that are mixed-species infections.

    † Geometric mean of species-specific 18S rRNA copy numbers per µl blood.

  2. ‡ MOI, multiplicity of infection: number of Pf-msp2 and Pv-msp1F3 alleles per infection.

    § Differences between villages were tested for using Chi2 and Fisher’s exact test (prevalence, proportion mixed) or Kruskal-Wallis test (MOI, log10-transformed parasite density).

Table 3
Multivariable predictors for time to recurrent blood-stage infection with Plasmodium species by qPCR
https://doi.org/10.7554/eLife.23708.008
VariableP. vivaxP. falciparumP. malariaeP. ovale
AHR*CI95p-valueAHR*CI95p-valueAHR*CI95p-valueAHR*CI95p-value
PQ treatment0.180.13–0.25<0.0010.730.52–1.020.0640.510.22–1.190.1210.310.12–0.750.010
Age0.950.87–1.040.2471.050.94–1.170.3610.980.75–1.290.9050.960.74–1.260.793
LLIN use at enrolment0.620.39–0.980.0430.840.49–1.440.5311.330.33–6.090.7150.950.26–3.430.936
Hb at enrolment (g/dl)0.880.80–0.980.0190.900.80–1.020.0990.830.61–1.120.2240.920.66–1.280.634
Village
 Albinama (ref)1111
 Amahup0.450.29–0.710.0010.580.31–1.110.1010.340.07–1.790.2052.830.29–27.480.370
 Balanga2.151.40–3.31<0.0011.810.99–3.300.0540.920.24–3.600.9107.740.85–70.450.070
 Balif1.000.66–1.540.9830.600.30–1.190.1450.240.03–2.070.1934.600.51–41.410.173
 Bolumita3.342.09–5.33<0.0014.732.69–8.30<0.0011.210.34–4.310.77019.432.19–172.370.008
 Numangu0.830.44–1.590.5832.291.17–4.500.0150.820.15–4.530.8233.170.19–52.410.420
Infection status at enrolment (by qPCR)
 Uninfected (ref)1111
P. vivax1.270.91–1.780.1651.370.86–2.200.1860.920.20–4.180.9132.170.68–6.970.192
P. falciparum1.360.84–2.190.2051.560.86–2.820.1453.540.85–14.720.0831.250.26–5.900.779
P. malariae0.830.38–1.850.6550.990.38–2.560.9776.351.31–30.810.0221.580.17–14.300.676
 Mixed P.f. or P.v.1.741.14–2.650.0102.081.25–3.480.0053.370.88–12.900.0762.030.55–7.530.287
  1. * AHRs were modeled using Cox proportional hazard regression.

    † Mixed infection including P. falciparum or P. vivax infection in conjunction with one or more other Plasmodium spp.

  2. PQ: Primaquine; LLIN: long-lasting insecticide-treated net; Hb: haemoglobin.

Table 4
Multivariable predictors of Pv- and Pf-molFOB per follow-up interval.

Model predictions from this model were used for mapping molFOB in Figure 3A.

https://doi.org/10.7554/eLife.23708.011
VariableP. vivaxP. falciparum
PQ armPlacebo armCombined arms
IRR*CI95p-valueIRR*CI95p-valueIRR*CI95p-value
PQ treatmentn.a.n.a.n.a.n.a.n.a.n.a.0.890.65–1.220.474
New P. falc. infections in interval1.320.92–1.890.1341.100.85–1.420.466n.a.n.a.n.a.
New P. vivax infections in intervaln.a.n.a.n.a.n.a.n.a.n.a.1.150.97–1.360.100
Age0.860.74–1.010.0590.950.87–1.040.3051.030.92–1.140.640
LLIN use at enrolment0.960.51–1.790.8970.620.43–0.910.0131.070.7–1.620.755
Hb at enrolment (g/dl)0.850.72–1.010.0630.910.85–0.990.0250.850.75–0.970.013
Village
 Albinama (ref)111
 Amahup0.020–0.11<0.0010.560.34–0.910.0200.520.25–1.070.074
 Balif0.850.4–1.80.6641.741.16–2.610.0071.810.98–3.350.059
 Balanga0.280.1–0.820.0201.130.73–1.730.5900.750.37–1.520.423
 Bolumita1.520.73–3.170.2682.671.83–3.9<0.0016.053.32–11.05<0.001
 Numangu0.50.15–1.680.2640.760.4–1.430.3942.81.39–5.640.004
Study Day
 Day 0–35 (ref)111
 Day 36–801.370.54–3.480.5091.991.39–2.84<0.0012.421.44–4.070.001
 Day 81–1751.340.57–3.120.5030.890.61–1.30.5381.130.7–1.840.616
 Day > 1750.650.25–1.690.3740.560.38–0.830.0040.870.48–1.560.643
  1. *IRRs were modeled per sampling interval using negative binomial generalized estimating equations allowing for repeated visits with log-link and an exchangeable correlation structure.

    † n.a., not applicable.

  2. molFOB in the follow-up interval (time-varying covariate).

    PQ: Primaquine; LLIN: long-lasting insecticide-treated net; Hb: haemoglobin.

Table 5
Multivariable predictors for time to P. vivax and P. falciparum clinical episodes.

Model predictions from this model were used for mapping the relative risk of clinical malaria episodes in Figure 3C and D.

https://doi.org/10.7554/eLife.23708.013
VariableP. vivaxP. falciparum
AHR*CI95p-valueAHR*CI95p-value
PQ treatment0.760.34–1.680.4971.791.05–3.030.031
P. vivax molFOB1.071.04–1.09<0.001n.a.n.a.n.a.
P. falciparum molFOBn.a.n.a.n.a.1.151.11–1.21<0.001
Age0.620.46–0.840.0020.980.85–1.130. 799
LLIN use at enrolment0.840.24–2.880.7780.440.22–0.870.018
Hb at enrolment (g/dl)0.950.74–0.670.6680.850.71–1.010.070
Village
 Albinama (ref)11
 Amahup0.890.23–3.460.8710.650.20–2.080.465
 Balif1.480.45–4.860.5181.260.50–3.140.626
 Balanga0.850.21–3.530.8271.390.59–3.300.455
 Bolumita0.990.24–4.030.9871.320.58–3.030.508
 Numangu1.000.23–4.310.9974.292.06–8.97<0.001
Infection status at enrolment (by qPCR)
 Uninfected (ref)11
P. vivax0.770.29–2.070.6081.640.91–2.950.101
P. falciparum1.740.59–5.110.3160.970.34–2.770.954
 Mixed P.f. or P.v.1.590.56–4.500.3811.240.57–2.680.582
  1. * AHRs were modeled using multiple failure Cox proportional hazard regression.

    † n.a., not applicable

  2. ‡ Average molFOB until the time of failure (time-varying covariate).

    PQ: Primaquine; LLIN: long-lasting insecticide-treated net; Hb: haemoglobin.

Table 6
Multivariable predictors for odds of P. falciparum clinical episodes
https://doi.org/10.7554/eLife.23708.015
P. falciparum episode
VariableOR*CI95p-value
PQ treatment1.420.80–2.520.226
P. vivax qPCR positive0.350.15–0.780.011
P. falciparum molFOB1.211.10–1.34<0.001
Age0.930.80–1.090.370
LLIN at enrolment0.370.16–0.830.016
Hb (g/dl) at enrolment0.880.70–1.110.292
Village
 Albinama (ref)1
 Amahup0.410.12–1.390.154
 Balif0.90.26–3.080.870
 Balanga1.190.42–3.390.747
 Bolumita1.480.42–5.170.540
 Numangu4.171.64–10.580.003
Study Day
 Day 0–801
 Day 81–1750.990.51–1.910.972
 Day > 1750.830.39–1.750.629
  1. * ORs were modeled using a binomial generalized estimating equation with logit link function using an exchangeable correlation structure.

    † Determined as P. vivax positive at the same or previous sampling visit.

  2. molFOB in the follow-up interval (time-varying covariate).

    PQ: Primaquine; LLIN: long-lasting insecticide-treated net; Hb: haemoglobin.

Data availability

The following data sets were generated
  1. 1
The following previously published data sets were used
  1. 1

Additional files

Supplementary file 1

Univariate factors.

Table 1. Univariate/PQ-adjusted predictors for time to recurrent blood-stage infection with Plasmodium species by qPCR. Table 2. Univariate (P. vivax) and univariate/PQ-treatment-adjusted (P. falciparum) predictors for Pv- and Pf-molFOB by follow-up interval. Table 3. Univariate/PQ-treatment-adjusted predictors for time to P. vivax and P. falciparum episodes. Table 4. Univariate/PQ-treatment-adjusted predictors for odds of P. falciparum clinical episodes.

https://doi.org/10.7554/eLife.23708.012
Supplementary file 2

Multivariable predictors for time to recurrent blood-stage infection with Plasmodium species by LM.

https://doi.org/10.7554/eLife.23708.016
Supplementary file 3

Multivariable predictors for P. falciparum and P. vivax density by qPCR during follow-up.

https://doi.org/10.7554/eLife.23708.017
Supplementary file 4

Multivariable predictors of Pv-molFOB (combining primaquine and placebo arms) per follow-up interval.

This model is similar to that presented in Table 4 in the main text but combines the treatment arms for P. vivax. Model predictions from this model were used for mapping molFOI in Figure 3B.

https://doi.org/10.7554/eLife.23708.018
Transparent reporting form
https://doi.org/10.7554/eLife.23708.019

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