Figures and data in Evidence from a natural experiment that malaria parasitemia is pathogenic in retinopathy-negative cerebral malaria

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Figure 1

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Potential pathways to clinically-defined cerebral malaria and genetic bottle necks.

There are three potential pathogenetic routes to WHO-defined cerebral malaria (CM). The first, shown in red, is the classical pathway: a malaria infection evolves into retinopathy-positive (Ret+) CM. The second and third possibilities produce retinopathy-negative (Ret-) CM. In (a) the coma is entirely the result of another etiology and the malaria parasitemia is incidental. In (b), the coma is a product of the interaction between the malaria parasitemia and an additional cause (or causes) of coma. Sickle cell trait is underrepresented in patients with Ret+ and Ret- cerebral malaria (CM) because of the bottleneck at the transition between 'malaria infection' (asymptomatic malaria) and 'malaria disease' (uncomplicated malaria). Blood group O is underrepresented in patients with Ret+ CM, but not in those with Ret- CM. Taken together, the results for sickle cell trait and blood group O suggest that some Ret- CM cases occur through pathway (b) (because sickle cell trait is underrepresented in Ret- CM) and that malaria parasites contribute to the pathogenesis of these cases, and that sickle cell trait reduces the pathogenetic potential of malaria infection for Ret- CM but do not provide evidence that blood group O reduces the pathogenetic potential of malaria infection for Ret- CM.

https://doi.org/10.7554/eLife.23699.004

Tables

Table 1

Characteristics of study participants at admission, Means ± SD for continuous variables. The proportions of missing data are shown in Appendix 1. There are 3704 community controls, but their characteristics are not shown because only their genotypes and not their clinical characteristics were collected. Bold denotes p-value less than 0.05.

https://doi.org/10.7554/eLife.23699.003

	Retinopathy Positive CM	Retinopathy Negative CM	Non-Malaria Hospital Controls	p-value, Ret + vs. Ret -	p-value, Ret + vs. Controls	p-value, Ret – vs. Controls
Number of participants	438	288	204
Female	50%	52%	43%	0.54	0.11	0.05
Age (months)	40 ± 26	44 ± 30	46 ± 30	0.10	0.05	0.53
Mid-upper arm circumference (cm)	14.9 ± 1.6	15.0 ± 1.7	14.8 ± 1.8	0.72	0.53	0.39
Weight (kg)	12 ± 4	13 ± 5	13 ± 6	0.37	0.29	0.74
Height (cm)	90 ± 16	91 ± 17	91 ± 20	0.30	0.40	1.00
Temperature (°C)	38.6 ± 1.2	38.4 ± 1.4	37.7 ± 1.5	0.03	<0.001	<0.001
Febrile (Temperature≥ 37.5°C)	81%	77%	56%	0.23	<0.001	<0.001
Pulse rate – beats/minute	152 ± 26	148 ± 24	139 ± 28	0.06	<0.001	<0.001
Respiratory rate – breaths/minute	47 ± 15	45 ± 13	45 ± 15	0.12	0.17	0.93
Liver size – cm below costal margin	2.0 ± 1.9	1.5 ± 1.9	1.1 ± 1.7	<0.001	<0.001	0.04
Spleen size – cm below costal margin	1.7 ± 2.1	1.6 ± 2.1	0.9 ± 1.6	0.56	<0.001	<0.001
Deep breathing	33%	25%	30%	0.03	0.59	0.18
Blantyre Coma Score: 0 1 2 3 4 5	14% 35% 49% 1% 0% 0%	19% 38% 43% 0% 0% 0%	28% 40% 23% 3% 0% 5%	0.01	0.08	0.90
CSF opening pressure – mm of water	176 ± 75	152 ± 82	176 ± 99	0.001	0.96	0.07
Hematocrit -- %	19.8 ± 6.9	28.2 ± 7.5	28.1 ± 9.6	<0.001	<0.001	0.86
Platelets	81,220± 67,219	161,600± 124,747	248,400± 162,287	<0.001	<0.001	0.86
Malaria parasitemia – parasites/mm³	230,500± 321,924	180,500± 280,676	3,619± 28,917	0.03	<0.001	<0.001
White blood cells	13,040± 9163	13,020± 8923	13,930± 9544	0.97	0.29	0.31
Lactate – mmol/liter	8.6 ± 5.0	7.3 ± 4.4	5.5 ± 3.9	0.05	<0.001	0.007
Blood glucose – mmol/liter	6.1 ± 3.9	6.8 ± 4.4	7.6 ± 5.3	0.03	<0.001	0.05
CSF white cell count – % ≥ 5	16%	20%	24%	0.31	0.06	0.37
Blood culture positive for pathogen	4%	2%	14%	0.51	<0.001	<0.001
HIV positive	18%	17%	15%	0.91	0.59	0.66
Outcomes
Discharge outcome: Full recovery Neurological Sequalae Died	69% 10% 21%	78% 10% 12%	57% 15% 28%	0.003	0.01	<0.001

Table 2

The top panel displays sickle cell trait (HbAS) proportions in retinopathy-positive (Ret+) cerebral malaria (CM), retinopathy-negative (Ret-) CM and control groups. The bottom panel displays ABO blood group gene proportions in Ret+ CM, Ret- CM and control groups. The last two rows of each panel display the odds ratios comparing controls to true Ret+ and true Ret- CM groups, which account for the fact that there is measurement error in observed retinopathy status (false discovery rate = 0.07 and false omission rate = 0.05).

https://doi.org/10.7554/eLife.23699.005

	Ret+ CM	Ret- CM	Non-malaria hospital controls	Community controls
Sample size	438	287	192	3657
HbAS^*	0	1	8	175
HbAA	437	286	184	3482
Proportion of HbAS	0	.003	.042	.048

			Odds ratio (95% CI)
Non-malaria hospital controls vs. community controls			0.87 (0.36, 1.78)
Controls vs. true Ret- CM			14.33 (3.21, 257.24)
Controls vs. true Ret+ CM			1223.22 (9.87, $\infty$ )

	Ret+ CM	Ret- CM	Non-malaria hospital controls	Community controls
Sample size	433	286	199	3543
Blood Group O	175	135	96	1739
Blood Group A, B or AB	258	151	103	1804
Proportion of Blood Group O	.404	.472	.482	.491

			Odds ratio (95% CI)
Non-malaria hospital controls vs. community controls			0.97 (0.72, 1.30)
Controls vs. true Ret- CM			1.03 (0.83, 1.29)
Controls vs. true Ret+ CM			1.23 (1.01, 1.50)

* HbAS (sickle cell trait) means that that the person has one normal and one abnormal copy of the hemoglobin beta gene. HbAA means the person has two normal copies of the hemoglobin beta gene.

Table 3

Inferences for lower bound on malaria parasitemia attributable fraction of Ret- CM (fraction of Ret- CM cases that would be prevented if malaria parasitemia were to be eliminated) under the sufficient-component cause model based on Figure 1 presented in Materials and methods. Inferences under the main model and sensitivity analyses that vary the effect of HbAS on malaria parasitemia incidence rate, the false discovery rate ( $F D R$ ) and the false omission rate ( $F O R$ ) for malarial retionopathy.

https://doi.org/10.7554/eLife.23699.006

Effect of HbAS on malaria parasitemia incidence rate	$F D R$	$F O R$	Lower bound on malaria parasitemia attributable fraction of Ret- CM Estimate (95% CI)
Main Model
No Effect	.07	.05	.93 (.68, 1)
Sensitivity Analyses
Reduce 10%	.07	.05	.92 (.64, 1)
Reduce 41%	.07	.05	.88 (.46, 1)
No Effect	.30	.11	.94 (.75, 1)
Reduce 10%	.30	.11	.94 (.72, 1)
Reduce 41%	.30	.11	.91 (.58, 1)
No Effect	0	.11	.92 (.62, 1)
Reduce 10%	0	.11	.91 (.58, 1)
Reduce 41%	0	.11	.86 (.37, 1)
No Effect	.30	0	.95 (.77, 1)
Reduce 10%	.30	0	.94 (.74, 1)
Reduce 41%	.30	0	.91 (.61, 1)
No Effect	0	0	.92 (.66, 1)
Reduce 10%	0	0	.92 (.63, 1)
Reduce 41%	0	0	.87 (.44, 1)

Appendix 1—table 1

Sample sizes.

https://doi.org/10.7554/eLife.23699.008

	Retinopathy-positive CM	Retinopathy-negative CM	Non-malaria hospital controls	Community controls
Sample Size	438	288	204	3704

Appendix 1—table 2

Missing data proportions for genetic traits.

https://doi.org/10.7554/eLife.23699.009

	Retinopathy-positive CM	Retinopathy-negative CM	Non-malaria hospital controls	Community controls
Sickle Cell Trait	.002	0	.054	.010
Blood Group	.011	.003	.025	.043

Appendix 1—table 3

Missing data proportions for demographic and clinical variables.

https://doi.org/10.7554/eLife.23699.010

	Retinopathy-positive CM	Retinopathy-negative CM	Non-malaria hospital controls
Female	.018	.021	.005
Age (months)	0	0	.005
Mid-upper arm circumference (cm)	.016	.014	.054
Weight (kg)	0	0	0
Height (cm)	.009	.024	.034
Temperature (°C)	0	0	0
Pulse rate – beats/minute	.002	0	.010
Respiratory rate – breaths/minute	0	0	.005
Liver size – cm below costal margin	.009	.024	.010
Spleen size – cm below costal margin	.005	.014	.010
Deep breathing	.007	.021	0
Blantyre Coma Score:	0	0	0
CSF opening pressure – mm of water	.420	.330	.623
Hematocrit -- %	.009	.024	.034
Platelets	.153	.160	.132
Malaria parasitemia – parasites/mm³	.039	.042	.025
White blood cells	.082	.097	.118
Lactate – mmol/liter	.653	.753	.564
Blood glucose – mmol/liter	.014	.003	0
CSF white cell count – % ≥ 5	.277	.170	.275
Blood culture positive for pathogen	.039	.024	.059
HIV positive	.144	.153	.353
Discharge outcome	0	.007	0

Appendix 1—table 4

Odds ratio comparing community controls to true Ret+ CM and Ret- CM groups, which account for the fact that there is measurement error in observed retinopathy status (false discovery rate = 0.07 and false omission rate = 0.05). The p-values are two-sided p-values for testing that the odds ratio equals 1.

https://doi.org/10.7554/eLife.23699.011

	Odds ratio (95% CI)	p-value
HbAS
Controls vs. true Ret- CM	14.43 (3.23, 258.94)	<0.0001
Controls vs. true Ret+ CM	1234.96 (9.93, $\infty$ )	<0.0001
BGO
Controls vs. true Ret- CM	1.03 (0.83, 1.29)	0.79
Controls vs. true Ret+ CM	1.23 (1.01, 1.51)	0.04

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Source code 1: https://doi.org/10.7554/eLife.23699.007
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Dylan S Small
Terrie E Taylor
Douglas G Postels
Nicholas AV Beare
Jing Cheng
Ian JC MacCormick
Karl B Seydel

(2017)

Evidence from a natural experiment that malaria parasitemia is pathogenic in retinopathy-negative cerebral malaria

eLife 6:e23699.

https://doi.org/10.7554/eLife.23699

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Potential pathways to clinically-defined cerebral malaria and genetic bottle necks.

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