A non-destructive sugar-feeding assay for parasite detection and estimating the extrinsic incubation period of Plasmodium falciparum in individual mosquito vectors

AbstractDespite its epidemiological importance, the time Plasmodium parasites take to achieve development in the vector mosquito (the extrinsic incubation period, EIP) remains poorly characterized. A novel non-destructive assay designed to estimate EIP in single mosquitoes, and more broadly to study Plasmodium – Anopheles vectors interactions, is presented. The assay uses small pieces of cotton wool soaked in sugar solution to collect malaria sporozoites from individual mosquitoes during sugar feeding to monitor infection status over time. This technique has been tested across four natural malaria mosquito species of Africa and Asia, six parasite isolates of Plasmodium falciparum, and across a range of temperatures relevant to malaria transmission in field conditions. We find that monitoring individual infectious mosquitoes is feasible, although due to the frequency of mosquito sugar feeding and inter-individual variation in infection intensity, there is inherent risk that this technique will result in some false negatives. The sensitivity rate ranged from 0.27 to 0.81 depending on mosquito species and on infection intensity in mosquitoes used to collect saliva. Using this non-destructive technique, the estimated median extrinsic incubation period of P. falciparum at 27°C was 11 to 14 days depending on mosquito species and parasite isolate. Long-term individual tracking also revealed that sporozoite transfer onto cotton wool can occur at least until day 40 post-infection. In addition to contributing to a better understanding of EIP and mosquito to human transmission with implications for improving epidemiological models, this technique also allows to link different transmission traits at the mosquito individual level. As one example, we found a significant relationship between EIP and mosquito lifespan, with short individual EIP associated with short mosquito lifespan. Correlations between mosquito/parasite traits often reveal trade-offs and constraints and have important implications for understanding the evolution of parasite transmission strategies.

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Microclimate variables of the ambient environment deliver the actual estimates of the extrinsic incubation period of Plasmodium vivax and Plasmodium falciparum: a study from a malaria-endemic urban setting, Chennai in India

Malaria Journal ◽

10.1186/s12936-018-2342-1 ◽

2018 ◽

Vol 17 (1) ◽

Cited By ~ 7

Author(s):

Shalu Thomas ◽

Sangamithra Ravishankaran ◽

N. A. Johnson Amala Justin ◽

Aswin Asokan ◽

T. Maria Jusler Kalsingh ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Plasmodium Vivax ◽

Incubation Period ◽

Urban Setting ◽

Ambient Environment ◽

Extrinsic Incubation Period ◽

Microclimate Variables

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Multiple blood feeding in mosquitoes shortens the Plasmodium falciparum incubation period and increases malaria transmission potential

PLoS Pathogens ◽

10.1371/journal.ppat.1009131 ◽

2020 ◽

Vol 16 (12) ◽

pp. e1009131

Author(s):

W. Robert Shaw ◽

Inga E. Holmdahl ◽

Maurice A. Itoe ◽

Kristine Werling ◽

Meghan Marquette ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Incubation Period ◽

Time Course ◽

Mosquito Control ◽

Mosquito Species ◽

Blood Feeding ◽

Egg Laying ◽

Sub Saharan Africa ◽

Reproductive Capacity ◽

Transmission Potential

Many mosquito species, including the major malaria vector Anopheles gambiae, naturally undergo multiple reproductive cycles of blood feeding, egg development and egg laying in their lifespan. Such complex mosquito behavior is regularly overlooked when mosquitoes are experimentally infected with malaria parasites, limiting our ability to accurately describe potential effects on transmission. Here, we examine how Plasmodium falciparum development and transmission potential is impacted when infected mosquitoes feed an additional time. We measured P. falciparum oocyst size and performed sporozoite time course analyses to determine the parasite’s extrinsic incubation period (EIP), i.e. the time required by parasites to reach infectious sporozoite stages, in An. gambiae females blood fed either once or twice. An additional blood feed at 3 days post infection drastically accelerates oocyst growth rates, causing earlier sporozoite accumulation in the salivary glands, thereby shortening the EIP (reduction of 2.3 ± 0.4 days). Moreover, parasite growth is further accelerated in transgenic mosquitoes with reduced reproductive capacity, which mimic genetic modifications currently proposed in population suppression gene drives. We incorporate our shortened EIP values into a measure of transmission potential, the basic reproduction number R0, and find the average R0 is higher (range: 10.1%–12.1% increase) across sub-Saharan Africa than when using traditional EIP measurements. These data suggest that malaria elimination may be substantially more challenging and that younger mosquitoes or those with reduced reproductive ability may provide a larger contribution to infection than currently believed. Our findings have profound implications for current and future mosquito control interventions.

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Vector survival and parasite infection: the effect of Wuchereria bancrofti on its vector Culex quinquefasciatus

Parasitology ◽

10.1017/s0031182004005153 ◽

2004 ◽

Vol 129 (1) ◽

pp. 43-50 ◽

Cited By ~ 11

Author(s):

K. KRISHNAMOORTHY ◽

S. SUBRAMANIAN ◽

G. J. VAN OORTMARSSEN ◽

J. D. F. HABBEMA ◽

P. K. DAS

Keyword(s):

Incubation Period ◽

Culex Quinquefasciatus ◽

Geometric Mean ◽

Wuchereria Bancrofti ◽

Parasite Load ◽

Infected Mosquito ◽

Parasite Development ◽

Mosquito Vector ◽

Extrinsic Incubation Period ◽

Parasite Loss

This paper investigates a cohort of 2187 laboratory reared Culex quinquefasciatus fed on 69 human volunteers, including 59 persons with different levels of Wuchereria bancrofti microfilariae and 10 without microfilaria. Mosquitoes were followed until death. Mosquito survival was analysed in relation to the level of microfilaria in the human and larval count in the dead mosquito. Vector mortality during the extrinsic incubation period (12 days post-engorgement) was significantly higher in mosquitoes fed on microfilaraemic volunteers (50%) than in those fed on amicrofilaraemics (29%). Both the percentage infected and the geometric mean parasite density was significantly higher among mosquitoes which died before 13 days (45% infected and 10 larvae per infected mosquito) than those surviving beyond 13 days (39% and 2·2), suggesting a parasite loss of more than 80% during the extrinsic incubation period. A large proportion (62%) of the mosquitoes that died during the early of phase of parasite development were infected (36% in low, 26% in medium and 90% in high human Mf-density). Survival analysis showed that the parasite load in mosquitoes and the human Mf-density for a given parasite load are independent risk factors of vector survival. Overall, the hazard of dying was found to be 11–15 times higher among mosquitoes fed on microfilaraemic volunteers than those fed on amicrofilaraemics. The hazard doubles for every increase of about 60–70 parasites in the vector. As a consequence of the parasite-induced reduction in vector survival, the transmission success of the parasite is reduced. The implication of the results on control/elimination of lymphatic filariasis using mass-drug administration is discussed.

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THE EFFECT OF VARIOUS TEMPERATURES IN MODIFYING THE EXTRINSIC INCUBATION PERIOD OF THE YELLOW FEVER VIRUS IN AËDES AEGYPTI*

American Journal of Epidemiology ◽

10.1093/oxfordjournals.aje.a117853 ◽

1932 ◽

Vol 16 (1) ◽

pp. 163-176 ◽

Cited By ~ 18

Author(s):

NELSON C. DAVIS

Keyword(s):

Aedes Aegypti ◽

Incubation Period ◽

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus ◽

Extrinsic Incubation Period

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STUDIES ON PLASMODIUM VIVAX. 2. THE INFLUENCE OF TEMPERATURE ON THE DURATION OF THE EXTRINSIC INCUBATION PERIOD*

American Journal of Epidemiology ◽

10.1093/oxfordjournals.aje.a117892 ◽

1932 ◽

Vol 16 (3) ◽

pp. 851-853 ◽

Cited By ~ 2

Author(s):

MARK F. BOYD

Keyword(s):

Plasmodium Vivax ◽

Incubation Period ◽

Influence Of Temperature ◽

Extrinsic Incubation Period ◽

Influence Of Temperature On

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Extrinsic Incubation Period (EIP)

Encyclopedia of Parasitology ◽

10.1007/978-3-662-43978-4_1145 ◽

2016 ◽

pp. 974-974

Author(s):

Heinz Mehlhorn

Keyword(s):

Incubation Period ◽

Extrinsic Incubation Period

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Extrapair Fertilization in the Black-Browed Reed Warbler (Acrocephalus Bistrigiceps): Effects on Mating Status and Nesting Cycle of Cuckolded and Cuckolder Males

The Auk ◽

10.1093/auk/122.4.1086 ◽

2005 ◽

Vol 122 (4) ◽

pp. 1086-1096

Author(s):

Shoji Hamao ◽

Daichi S. Saito

Keyword(s):

Incubation Period ◽

Mate Guarding ◽

Reed Warbler ◽

Fertile Period ◽

Mating Status ◽

Trade Offs ◽

Socially Monogamous ◽

The Relationship ◽

Extrapair Copulation

Abstract We studied the relationship between the occurrence of extrapair fertilizations (EPFs) and mating status and nesting cycle of mates, respectively, in a population of Black-browed Reed Warblers (Acrocephalus bistrigiceps). Overall, 6.4% (9 of 140) of the chicks were sired by extrapair males, and 13.5% (5 of 37) of the broods contained at least one extrapair chick. Socially polygynous males were cuckolded more frequently than socially monogamous males (40.0% and 3.7%, respectively). In three of four cases where cuckolded males were polygynous, the fertile periods of their mates overlapped. Except for two floating males that obtained EPFs, five out of seven cuckolder males were paired. Males succeeded in EPFs when the fertile period of the extrapair females largely overlapped the incubation period of their own mates. These findings are consistent with the hypothesis that there are trade-offs between extrapair copulation activity and both mate guarding and rearing of nestlings. Fertilisation Extraconjugale chez Acrocephalus bistrigiceps: Effets des Mâles Infidèles et Cocus sur le Statut de Couple et le Cycle de Nidification

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Exploring the lower thermal limits for development of the human malaria parasite, Plasmodium falciparum

Biology Letters ◽

10.1098/rsbl.2019.0275 ◽

2019 ◽

Vol 15 (6) ◽

pp. 20190275 ◽

Cited By ~ 4

Author(s):

Jessica L. Waite ◽

Eunho Suh ◽

Penelope A. Lynch ◽

Matthew B. Thomas

Keyword(s):

Plasmodium Falciparum ◽

Future Climate Change ◽

Parasite Development ◽

Temperature Dependent ◽

Human Malaria Parasite ◽

Thermal Limits ◽

Degree Day ◽

Extrinsic Incubation Period ◽

Parasite Plasmodium ◽

Parasite Plasmodium Falciparum

The rate of malaria transmission is strongly determined by parasite development time in the mosquito, known as the extrinsic incubation period (EIP), since the quicker parasites develop, the greater the chance that the vector will survive long enough for the parasite to complete development and be transmitted. EIP is known to be temperature-dependent but this relationship is surprisingly poorly characterized. There is a single degree-day model for EIP of Plasmodium falciparum that derives from a limited number of poorly controlled studies conducted almost a century ago. Here, we show that the established degree-day model greatly underestimates the rate of development of P. falciparum in both Anopheles stephensi and An. gambiae mosquitoes at temperatures in the range of 17–20°C. We also show that realistic daily temperature fluctuation further speeds parasite development. These novel results challenge one of the longest standing models in malaria biology and have potentially important implications for understanding the impacts of future climate change.

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