# Time-of-day of infection: impact on liver stage malaria parasites in untreated and drug-treated hosts

## TL;DR

This study investigates how the time of day affects the ability of malaria parasites to infect mice, finding no significant impact from the host's circadian rhythm.

## Contribution

The study provides new insights into the lack of influence of host circadian rhythms on malaria parasite liver-stage establishment.

## Key findings

- Pyrimethamine treatment reduced parasite liver burdens but not significantly based on host time-of-day.
- Host time-of-day did not affect parasite liver burdens in untreated mice.
- Circadian rhythms in hosts do not significantly impact vector-to-host malaria transmission.

## Abstract

Circadian clocks are thought to have evolved owing to the benefits of anticipating daily environmental rhythms. Daily environmental rhythms that impact on fitness include interactions between organisms, such as host–pathogen interactions. For example, host susceptibility to infection for taxonomically diverse hosts and pathogens varies across the circadian cycle. We previously revealed that mosquito vectors are less susceptible to malaria (Plasmodium) infection during their active phase (night time), and here we test whether a similar pattern occurs for infection of the mammalian host.

We used Plasmodium berghei-infected Anopheles stephensi mosquitoes to infect mice during their rest or active phase, both in untreated and pyrimethamine-treated mice. We assessed the parasites’ success in establishing at the first site of replication (in the liver) by quantifying parasite burdens using quantitative PCR (qPCR), adjusted for sporozoite inocula. By independently manipulating the photoschedules of vectors and hosts, we standardise the time-of-day for parasites and mosquitoes used to initiate infections, and thus, directly test the impact of host time-of-day on the parasites’ ability to establish an infection.

The three experiments we conducted showed that pyrimethamine treatment reduced parasite liver burdens, but not in a biologically significant manner dependent on host time-of-day (active/rest phase). Furthermore, host time-of-day did not affect parasite liver burdens in untreated hosts.

Understanding the roles of host, parasite, and vector rhythms on malaria transmission is important given that mosquitoes are altering the time of day that they bite. That rhythms, per se, do not affect vector-to-host transmission suggests that the impacts of time-of-day on components of vectorial capacity are more epidemiologically influential than host rhythms.

The online version contains supplementary material available at 10.1186/s13071-025-06986-7.