Malaria Parasites in the Asymptomatic: Looking for the Hay in the Haystack

doi:10.1016/j.pt.2015.11.015

Trends in Parasitology

Volume 32, Issue 4, April 2016, Pages 296-308

https://doi.org/10.1016/j.pt.2015.11.015 Get rights and content

Trends

Understanding the determinants of asymptomatic malaria infections, as a silent driver of transmission, has become essential for the success of elimination campaigns.

Transmission intensity, parasite virulence, immune resistance and tolerance to infection, host genetic factors, pregnancy, or comorbidities can potentially affect the transition between symptomatic and asymptomatic infection and therefore constitute candidate areas for further research.

Research is needed to understand how the risk of asymptomatic malaria changes when transmission is abruptly interrupted, as well as the timeframes needed to lose host defenses that maintain infections at subclinical levels.

Elimination strategies could be guided by the surveillance of molecular and immunological biomarkers that mirror the dynamics of asymptomatic infections and allow prediction of the risk of asymptomatic infections in target populations.

With malaria elimination back on the international agenda, programs face the challenge of targeting all Plasmodium infections, not only symptomatic cases. As asymptomatic individuals are unlikely to seek treatment, they are missed by passive surveillance while remaining infectious to mosquitoes, thus acting as silent reservoirs of transmission. To estimate the risk of asymptomatic infections in various phases of malaria elimination, we need a deeper understanding of the underlying mechanisms favoring carriage over disease, which may involve both pathogen and host factors. Here we review our current knowledge on the determinants leading to Plasmodium falciparum symptomless infections. Understanding the host–pathogen interactions that are most likely to affect transitions between malaria disease states could guide the development of tools to tackle asymptomatic carriers in elimination settings.

Section snippets

Being Asymptomatic

Peaceful coexistence with the infected host, rarely causing clinical symptoms, is a common but poorly understood phenomenon that occurs for many human pathogens [1]. Because such asymptomatic cases of infection occur without eventual overt symptoms, they do not come to clinical attention, thus representing a large hidden reservoir of active infection that permits their persistence and eventual spread to other human hosts. This is the case for many malaria infections in semi-immune individuals

Determinants of Asymptomatic Malaria

Why some individuals develop clinical manifestations during malaria infection while others remain asymptomatic is poorly understood. The general state of health and physiological condition of the host, particularly subtle immunity variations, together with host genetic predisposition and parasite factors involved in the virulence of the infection might influence the progression of malaria infection toward an asymptomatic outcome (Figure 1).

Disease Patterns During Changes of Malaria Transmission

Understanding the determinants and clinical consequences of malaria declines and resurgences, as well as the timescales for changing disease presentations, has become a priority in the context of the current goal to eradicate malaria. Several studies have shown a shift in the peak of acute malaria incidence toward older ages as malaria exposure wanes 14, 63, which may precede changes in the number of episodes and disease spectrum in an area with declining transmission intensities [64]. These

Concluding Remarks

Malaria elimination programs must include tactics to target asymptomatic reservoirs that can sustain transmission. As the relative contribution of asymptomatic infections to the overall pool of infections may change during the course of elimination activities, surveillance and response strategies should be adapted to the different epidemiological scenarios [71]. However, our understanding of how malaria transmission, host defenses, and parasite factors interact to trigger the clinical spectrum

Acknowledgments

This work was supported by the Instituto de Salud Carlos III (PI13/01478, CES10/021-I3SNS to A.M. and CP11/00269 from the Miguel Servet program to Q.B.) and the Departament d’Universitats i Recerca de la Generalitat de Catalunya (AGAUR; grant 2014SGR263 to A.M.). The authors thank Eduard Rovira-Vallbona, Silvie Huijben, and Azucena Bardají for helpful comments on an earlier version of the manuscript.

Glossary

Active case detection: detection by health workers of malaria cases at community and household level in population groups that are considered to be at high risk. Active case detection can be conducted as fever screening followed by parasitological examination of all febrile patients or as parasitological examination of the target population without prior fever screening.
Adaptive immunity: also known as acquired immunity; one of the two main immune strategies in vertebrates (the other being the

References (105)

H.A. Babiker
The role of asymptomatic P. falciparum parasitaemia in the evolution of antimalarial drug resistance in areas of seasonal transmission
Drug Resist. Updat.
(2013)
M.S. Oakley
Clinical and molecular aspects of malaria fever
Trends Parasitol.
(2011)
M.T. Duraisingh
Heterochromatin silencing and locus repositioning linked to regulation of virulence genes in Plasmodium falciparum
Cell
(2005)
J.D. Smith
Decoding the language of var genes and Plasmodium falciparum sequestration
Trends Parasitol.
(2001)
O.C. Finney
Predicting antidisease immunity using proteome arrays and sera from children naturally exposed to malaria
Mol. Cell. Proteomics
(2014)
P.E. Duffy
Variant proteins on the surface of malaria-infected erythrocytes – developing vaccines
Trends Parasitol.
(2001)
M.J. Miller
Observations on the natural history of malaria in the semi-resistant West African
Trans. R. Soc. Trop. Med. Hyg.
(1958)
W.P. O’Meara
Effect of a fall in malaria transmission on morbidity and mortality in Kilifi, Kenya
Lancet
(2008)
R.W. Snow et al.
The consequences of reducing transmission of Plasmodium falciparum in Africa
Adv. Parasitol.
(2002)
Q. de Mast
Is asymptomatic malaria really asymptomatic? Hematological, vascular and inflammatory effects of asymptomatic malaria parasitemia
J. Infect.
(2015)

P.E. Szmitko

Plasmodium falciparum malaria occurring 8 years after leaving an endemic area

Diagn. Microbiol. Infect. Dis.

(2009)

M.T. Bretscher

The distribution of Plasmodium falciparum infection durations

Epidemics

(2011)

S.E. Clarke

Risk of malaria attacks in Gambian children is greater away from malaria vector breeding sites

Trans. R. Soc. Trop. Med. Hyg.

(2002)

V. Jeney

Control of disease tolerance to malaria by nitric oxide and carbon monoxide

Cell Rep.

(2014)

A. Casadevall et al.

Host–pathogen interactions: basic concepts of microbial commensalism, colonization, infection, and disease

Infect. Immun.

(2000)

K.A. Lindblade

The silent threat: asymptomatic parasitemia and malaria transmission

Expert Rev. Anti Infect. Ther.

(2013)

F.P. Alves

Asymptomatic carriers of Plasmodium spp. as infection source for malaria vector mosquitoes in the Brazilian Amazon

J. Med. Entomol.

(2005)

T. Bousema

Asymptomatic malaria infections: detectability, transmissibility and public health relevance

Nat. Rev. Microbiol.

(2014)

M. Tanner

Malaria eradication and elimination: views on how to translate a vision into reality

BMC Med.

(2015)

Disease Surveillance for Malaria Elimination

(2012)

M.L. Gatton et al.

Investigating antigenic variation and other parasite–host interactions in Plasmodium falciparum infections in naive hosts

Parasitology

(2004)

M.L. Gatton et al.

Evaluation of the pyrogenic threshold for Plasmodium falciparum malaria in naive individuals

Am. J. Trop. Med. Hyg.

(2002)

J.A. Simpson

Population dynamics of untreated Plasmodium falciparum malaria within the adult human host during the expansion phase of the infection

Parasitology

(2002)

L.H. Miller

Malaria biology and disease pathogenesis: insights for new treatments

Nat. Med.

(2013)

T. Smith

An epidemiologic model of the incidence of acute illness in Plasmodium falciparum malaria

Am. J. Trop. Med. Hyg.

(2006)

T. Smith

Relationships between the outcome of Plasmodium falciparum infection and the intensity of transmission in Africa

Am. J. Trop. Med. Hyg.

(2004)

P. Boisier

Relationship between parasite density and fever risk in a community exposed to a low level of malaria transmission in Madagascar highlands

Am. J. Trop. Med. Hyg.

(2002)

K. Chotivanich

Parasite multiplication potential and the severity of falciparum malaria

J. Infect. Dis.

(2000)

M. Pinkevych

Decreased growth rate of P. falciparum blood stage parasitemia with age in a holoendemic population

J. Infect. Dis.

(2014)

L. Mancio-Silva

Sir2a regulates rDNA transcription and multiplication rate in the human malaria parasite Plasmodium falciparum

Nat. Commun.

(2013)

C.J. Merrick

Epigenetic dysregulation of virulence gene expression in severe Plasmodium falciparum malaria

J. Infect. Dis.

(2012)

S. Mok

Drug resistance Population transcriptomics of human malaria parasites reveals the mechanism of artemisinin resistance

Science

(2015)

L. Turner

Severe malaria is associated with parasite binding to endothelial protein C receptor

Nature

(2013)

G.M. Warimwe

Plasmodium falciparum var gene expression is modified by host immunity

Proc. Natl. Acad. Sci.U.S.A.

(2009)

N. Falk

Analysis of Plasmodium falciparum var genes expressed in children from Papua New Guinea

J. Infect. Dis.

(2009)

M. Rottmann

Differential expression of var gene groups is associated with morbidity caused by Plasmodium falciparum infection in Tanzanian children

Infect. Immun.

(2006)

A. Claessens

Generation of antigenic diversity in Plasmodium falciparum by structured rearrangement of var genes during mitosis

PLoS Genet.

(2014)

S. Tukwasibwe

Differential prevalence of transporter polymorphisms in symptomatic and asymptomatic falciparum malaria infections in Uganda

J. Infect. Dis.

(2014)

R. Ord

Seasonal carriage of pfcrt and pfmdr1 alleles in Gambian Plasmodium falciparum imply reduced fitness of chloroquine-resistant parasites

J. Infect. Dis.

(2007)

K. Sonden

Asymptomatic multiclonal Plasmodium falciparum infections carried through the dry season predict protection against subsequent clinical Malaria

J. Infect. Dis.

(2015)

A.R. Wargo

Transmission stage investment of malaria parasites in response to in-host competition

Proc. Biol. Sci.

(2007)

R. Medzhitov

Disease tolerance as a defense strategy

Science

(2012)

P.D. Crompton

Malaria immunity in man and mosquito: insights into unsolved mysteries of a deadly infectious disease

Annu. Rev. Immunol.

(2014)

M.M. Stevenson et al.

Innate immunity to malaria

Nat. Rev. Immunol.

(2004)

G.M. Warimwe

Plasmodium falciparum var gene expression homogeneity as a marker of the host–parasite relationship under different levels of naturally acquired immunity to malaria

PLoS ONE

(2013)

M.M. Medeiros

Natural antibody response to Plasmodium falciparum merozoite antigens MSP5 MSP9 and EBA175 is associated to clinical protection in the Brazilian Amazon

BMC Infect. Dis.

(2013)

K.J. Torres

Genome-level determination of Plasmodium falciparum blood-stage targets of malarial clinical immunity in the Peruvian Amazon

J. Infect. Dis.

(2015)

D.I. Stanisic

Immunoglobulin G subclass-specific responses against Plasmodium falciparum merozoite antigens are associated with control of parasitemia and protection from symptomatic illness

Infect. Immun.

(2009)

P. Bejon

Analysis of immunity to febrile malaria in children that distinguishes immunity from lack of exposure

Infect. Immun.

(2009)

D.D. Laishram

The complexities of malaria disease manifestations with a focus on asymptomatic malaria

Malar. J.

(2012)

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The existence of asymptomatic infectious humans complicates disease control efforts. Specifically, since asymptomatic infectious humans do not exhibit clinical symptoms of malaria and hence, cannot be detected through fever-based methods (Galatas et al., 2016; Ramaswamy et al., 2020), they serve as hidden reservoirs for malaria parasites that can be picked up my mosquitoes and conveyed to other humans. Hence, they are critical in the spread and persistence of malaria (Alves et al., 2002; Bousema et al., 2004; Eke et al., 2006; Lindblade et al., 2013; Dal-Bianco et al., 2007; Laishram et al., 2012).
The mosquito-borne disease, malaria, continues to impose a devastating health and economic burden worldwide. In malaria-endemic areas, insecticide-treated nets (ITNs) have been useful in curtailing the burden of the disease. However, mosquito resistance to insecticides, decay in ITN efficacy, net attrition, etc., undermine the effectiveness of ITNs in combatting malaria. In this study, mathematical models that account for asymptomatic infectious humans (through a partially immune class or a separate asymptomatic infectious class), insecticide resistance, and decay in ITN efficacy are proposed and analyzed. Analytical and numerical results of the models when ITN efficacy is constant show that there are parameter regimes for which a backward bifurcation occurs. Local and global sensitivity analyses are performed to identify parameters (some of which are potential targets for disease control) with the most significant influence on the control reproduction ( $R_{c}$ ) and disease prevalence. These influential parameters include the maximum biting rate of resistant mosquitoes, ITN coverage, initial ITN efficacy against sensitive mosquitoes, the probability that an infectious mosquito (human) infects a susceptible human (mosquito), and the rate at which adult mosquitoes develop (lose) resistance to insecticides. Simulations of the models show that accounting for asymptomatic infectious humans through a separate class, or not accounting for the decay in ITN efficacy leads to an underestimation of disease burden. In particular, if the initial efficacy of ITNs against sensitive and resistance mosquitoes is 96%, the minimum ITN coverage required to reduce $R_{c}$ below one (and hence, contain malaria) is approximately 11% (27%) lower when ITN efficacy is averaged (constant) for a model with a separate asymptomatic class. For the model with a partially immune class and decaying ITN efficacy, reducing $R_{c}$ below one is impossible even if the entire populace uses ITNs. The study shows that replacing ITNs before their prescribed lifespans, or designing ITNs with longer lifespans is important for malaria control. Furthermore, the study shows that piperonyl butoxide (PBO) ITNs (which inhibit or reverse insecticide resistance) outperform regular ITNs in malaria control. Hence, prospects for effectively controlling malaria are enhanced by widespread use of high quality ITNs (e.g. PBO ITNs), especially if the useful lifespans of the ITNs are long enough and the ITNs are replaced before the end of their useful lifespans.

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Trends in Parasitology

ReviewMalaria Parasites in the Asymptomatic: Looking for the Hay in the Haystack

Trends

Section snippets

Being Asymptomatic

Determinants of Asymptomatic Malaria

Disease Patterns During Changes of Malaria Transmission

Concluding Remarks

Acknowledgments

Glossary

Drug Resist. Updat.

Trends Parasitol.

Cell

Trends Parasitol.

Mol. Cell. Proteomics

Trends Parasitol.

Trans. R. Soc. Trop. Med. Hyg.

Lancet

Adv. Parasitol.

J. Infect.

Diagn. Microbiol. Infect. Dis.

Epidemics

Trans. R. Soc. Trop. Med. Hyg.

Cell Rep.

Host–pathogen interactions: basic concepts of microbial commensalism, colonization, infection, and disease

Infect. Immun.

The silent threat: asymptomatic parasitemia and malaria transmission

Expert Rev. Anti Infect. Ther.

Asymptomatic carriers of Plasmodium spp. as infection source for malaria vector mosquitoes in the Brazilian Amazon

J. Med. Entomol.

Asymptomatic malaria infections: detectability, transmissibility and public health relevance

Nat. Rev. Microbiol.

Malaria eradication and elimination: views on how to translate a vision into reality

BMC Med.

Disease Surveillance for Malaria Elimination

Investigating antigenic variation and other parasite–host interactions in Plasmodium falciparum infections in naive hosts

Parasitology

Evaluation of the pyrogenic threshold for Plasmodium falciparum malaria in naive individuals

Am. J. Trop. Med. Hyg.

Population dynamics of untreated Plasmodium falciparum malaria within the adult human host during the expansion phase of the infection

Parasitology

Malaria biology and disease pathogenesis: insights for new treatments

Nat. Med.

An epidemiologic model of the incidence of acute illness in Plasmodium falciparum malaria

Am. J. Trop. Med. Hyg.

Relationships between the outcome of Plasmodium falciparum infection and the intensity of transmission in Africa

Am. J. Trop. Med. Hyg.

Relationship between parasite density and fever risk in a community exposed to a low level of malaria transmission in Madagascar highlands

Am. J. Trop. Med. Hyg.

Parasite multiplication potential and the severity of falciparum malaria

J. Infect. Dis.

Decreased growth rate of P. falciparum blood stage parasitemia with age in a holoendemic population

J. Infect. Dis.

Sir2a regulates rDNA transcription and multiplication rate in the human malaria parasite Plasmodium falciparum

Nat. Commun.

Epigenetic dysregulation of virulence gene expression in severe Plasmodium falciparum malaria

J. Infect. Dis.

Drug resistance Population transcriptomics of human malaria parasites reveals the mechanism of artemisinin resistance

Science

Severe malaria is associated with parasite binding to endothelial protein C receptor

Nature

Plasmodium falciparum var gene expression is modified by host immunity

Proc. Natl. Acad. Sci.U.S.A.

Analysis of Plasmodium falciparum var genes expressed in children from Papua New Guinea

J. Infect. Dis.

Differential expression of var gene groups is associated with morbidity caused by Plasmodium falciparum infection in Tanzanian children

Infect. Immun.

Generation of antigenic diversity in Plasmodium falciparum by structured rearrangement of var genes during mitosis

PLoS Genet.

Differential prevalence of transporter polymorphisms in symptomatic and asymptomatic falciparum malaria infections in Uganda

J. Infect. Dis.

Seasonal carriage of pfcrt and pfmdr1 alleles in Gambian Plasmodium falciparum imply reduced fitness of chloroquine-resistant parasites

J. Infect. Dis.

Asymptomatic multiclonal Plasmodium falciparum infections carried through the dry season predict protection against subsequent clinical Malaria

J. Infect. Dis.

Transmission stage investment of malaria parasites in response to in-host competition

Proc. Biol. Sci.

Disease tolerance as a defense strategy

Science

Malaria immunity in man and mosquito: insights into unsolved mysteries of a deadly infectious disease

Review
Malaria Parasites in the Asymptomatic: Looking for the Hay in the Haystack