**3. The effects of infection on behavior and physiology of mosquito vectors**

Since vectorial capacity suffers major influence of vector behavior, studying the degree of modulation that arbovirus exerts on *A. aegypti*'s behavior is a key factor for understanding infection dynamics and host pathogenesis. In a recent work, Gaburro et al. have shown that infection by Zika virus leads to neuro-excitation in *A. aegypti*'s brain, inducing changes in the mosquito's behavior. The increase in neuronal spikes in infected versus non-infected females reflected on an increase in flight activity when females were studied in pools [57]. The authors found replicating virus in ZIKV-infected female brains, characterizing the tropism for the central nervous system, as well as in sensory organs like antennas and eyes, potentially affecting neuronal communication. Likewise, dengue virus was also found to be neurotropic in mosquitoes [58].

A consequence of the neurotropic characteristic of these arboviruses is the alteration in the patterns of locomotor activity and feeding behavior. For instance, *A. triseriatus* becomes more avid for refeeding when infected by La Crosse virus [59, 60], while *Aedes aegypti* becomes more active when infected with serotype 2 of dengue virus [61] and with Zika virus when females are monitored in groups in cages (**Figure 4**) [57]. However, the assumption that virus infection would modulate behavior in a way to increase virus transmission and vectorial capacity is not always met. The example of West Nile virus indicates a possible decrease in virus transmission, where the mosquito vector *Culex pipiens* becomes less avid for host-seeking when infected with the virus [63]. Likewise, for individually monitored females of *A. aegypti*, Zika virus infection reduces flight activity, suggesting that infected mosquitoes may remain associated with closely distributed human hosts (**Figure 4**) [62].

#### **Figure 4.**

*Locomotor activity of virus-infected and not infected females. (A) Females infected with serotype 2 of dengue virus (modified from [61]). (B) Females infected with Zika virus (modified from [62]). The Zeitgeber time means the time passed, in hours, after light is turned on.*

#### **Figure 5.**

*Effect of Zika virus infection on the fecundity (A) and fertility (B) of Aedes aegypti females, on the third gonotrophic cycles. The lack of significance is represented by p values >0.05 obtained by using the nonparametric Mann-Whitney tests. Error bars represent mean ± s.d of three independent experiments (modified from [62]).*

Arbovirus infection is also responsible for changes in physiological traits implicated in the estimate of the vectorial capacity. The number of female mosquitoes per host is one of the most important parameters of the vectorial capacity and is directly influenced by life history traits like the number of eggs laid by females (fecundity) and the number of viable offspring (fertility). These traits have been reported altered by arbovirus infection, although the effect varies depending on the virus. Dengue-infected females of *A. aegypti* produce a significantly lower number of eggs with a lower hatching rate [45], while ZIKV-infected and non-infected females do not show significant differences in fecundity and fertility (**Figure 5**) [62].

### **4. Human environmental impact and the effects on vector-host interaction and the risk of disease transmission**

Human occupation may lead to profound alterations in the environment, such as global warming and light pollution. Some of these changes impose new selective pressures to all organisms involved with the infection, say pathogens, vectors, and hosts, but also their predators and the vegetation used as nutrition or habitat. The modeling of the effects of global warming on disease transmission indicates a shift in the global distribution of *Aedes*-borne virus with mild to severe effects on the risk of transmission [64, 65]. Concerning light pollution, the increase in domiciliary and peridomiciliary lighting may extend the phase of activity of *A. aegypti* by a couple

**75**

circadian clock.

*The Effects of Infection on Mosquito Rhythmic Behavior DOI: http://dx.doi.org/10.5772/intechopen.89409*

of environmental factors on synchronizing these rhythms.

**5. Molecular control of the behavior**

with tissue-specific functions [70].

transmission [46].

of hours, which may raise the biting rate and the chance of arbovirus transmission. A recent work showed that artificial lighting at night make house sparrows, the reservoirs of West Nile virus, to become infectious for a period of 2 days longer than house sparrows that get dark nights [66]. This leads to an increase of 41% in the potential of disease outbreak. Comparatively, in light-night areas, nocturnal mosquitoes like *Anopheles* species will begin their phase of activity when human hosts are still active and not under a bed net, leading to a higher chance of malaria

Altogether, both vectors and hosts undergo behavioral and physiological changes triggered by the virus infection, and in turn, the influence of environmental variation is behind all facets of this interaction. The next section will discuss the endogenous mechanisms regulating rhythmic behavior and physiology, as well as the role

The different behaviors exhibited by mosquitoes are, in general, driven by internal biological clocks that generate circadian rhythms. These rhythms present a period of nearly, but not exactly, 24 h and are responsible for responses such as host-seeking, breeding site seeking, activity, and rest, among others [67].

These rhythms are directly influenced by natural cues from the environment, and the most important ones are the light/dark and the temperature cycles. These stimuli are received by specific receptors, like photoreceptors (in the eyes and head) and thermoreceptors (along the whole body) and are transmitted to the internal pacemaker or the biological clock itself. Thus, a rhythm or a physiological response is generated from the interaction of the stimuli with the pacemaker neurons [68]. The pacemaker neurons are so-called because they express the clock genes, which are the components of the circadian clock. These genes interact with each other and recruit kinases, phosphatases, and transcription factors to generate oscillating expression in a 24-h cycle [69]. They are also responsible for the regulation of many other genes, the clock-controlled genes (CCGs), that are directly associated

*Drosophila melanogaster* is the insect model for studying circadian rhythm, but it is already known that many features of the circadian clock of other insects differ considerably from the fly clock. The *Drosophila* clock is formed by three interconnected autoregulatory loops, in which the proteins coded by *Clock* (*Clk*) and *cycle* (*cyc*) genes play a central role. In the first loop, the heterodimer CLK-CYC binds to an E-box sequence in the promoter region of *period* (*per*) and *timeless* (*tim*) genes, activating their transcription. Once in the cytoplasm, the transcripts are translated into proteins that accumulate during the early night and later enter the nucleus to repress their own transcription. This cycle lasts 24 h due to the posttranslational modifications controlled by the activity of kinases such as DOUBLETIME (DBT), CASEIN KINASE 2A (CK2A), and SHAGGY (SGG), which together with phosphatases such as PP2A stabilize PER and TIM [71, 72]. In the second loop, two transcription factors, VRI and PDP1e, are involved, respectively, in the repression and activation of *Clk* and *cyc* genes. Finally, a third interconnected loop involves the activation of *clockwork orange* (*cwo*) gene and the repression exerted by its product, CWO, in PER targets [73]. **Figure 6** summarizes the *D. melanogaster* molecular

An interesting feature of this clock is its property of environmental synchronization, which adjusts the period to exactly 24 h. One of the most important synchronizers (or *Zeitgebers*) is light. The light-induced resetting mechanism is

*The Effects of Infection on Mosquito Rhythmic Behavior DOI: http://dx.doi.org/10.5772/intechopen.89409*

*Vector-Borne Diseases - Recent Developments in Epidemiology and Control*

*means the time passed, in hours, after light is turned on.*

Arbovirus infection is also responsible for changes in physiological traits implicated in the estimate of the vectorial capacity. The number of female mosquitoes per host is one of the most important parameters of the vectorial capacity and is directly influenced by life history traits like the number of eggs laid by females (fecundity) and the number of viable offspring (fertility). These traits have been reported altered by arbovirus infection, although the effect varies depending on the virus. Dengue-infected females of *A. aegypti* produce a significantly lower number of eggs with a lower hatching rate [45], while ZIKV-infected and non-infected females do

*Effect of Zika virus infection on the fecundity (A) and fertility (B) of Aedes aegypti females, on the third gonotrophic cycles. The lack of significance is represented by p values >0.05 obtained by using the nonparametric Mann-Whitney tests. Error bars represent mean ± s.d of three independent experiments (modified from [62]).*

*Locomotor activity of virus-infected and not infected females. (A) Females infected with serotype 2 of dengue virus (modified from [61]). (B) Females infected with Zika virus (modified from [62]). The Zeitgeber time* 

not show significant differences in fecundity and fertility (**Figure 5**) [62].

**4. Human environmental impact and the effects on vector-host** 

Human occupation may lead to profound alterations in the environment, such as global warming and light pollution. Some of these changes impose new selective pressures to all organisms involved with the infection, say pathogens, vectors, and hosts, but also their predators and the vegetation used as nutrition or habitat. The modeling of the effects of global warming on disease transmission indicates a shift in the global distribution of *Aedes*-borne virus with mild to severe effects on the risk of transmission [64, 65]. Concerning light pollution, the increase in domiciliary and peridomiciliary lighting may extend the phase of activity of *A. aegypti* by a couple

**interaction and the risk of disease transmission**

**74**

**Figure 4.**

**Figure 5.**

of hours, which may raise the biting rate and the chance of arbovirus transmission. A recent work showed that artificial lighting at night make house sparrows, the reservoirs of West Nile virus, to become infectious for a period of 2 days longer than house sparrows that get dark nights [66]. This leads to an increase of 41% in the potential of disease outbreak. Comparatively, in light-night areas, nocturnal mosquitoes like *Anopheles* species will begin their phase of activity when human hosts are still active and not under a bed net, leading to a higher chance of malaria transmission [46].

Altogether, both vectors and hosts undergo behavioral and physiological changes triggered by the virus infection, and in turn, the influence of environmental variation is behind all facets of this interaction. The next section will discuss the endogenous mechanisms regulating rhythmic behavior and physiology, as well as the role of environmental factors on synchronizing these rhythms.

### **5. Molecular control of the behavior**

The different behaviors exhibited by mosquitoes are, in general, driven by internal biological clocks that generate circadian rhythms. These rhythms present a period of nearly, but not exactly, 24 h and are responsible for responses such as host-seeking, breeding site seeking, activity, and rest, among others [67].

These rhythms are directly influenced by natural cues from the environment, and the most important ones are the light/dark and the temperature cycles. These stimuli are received by specific receptors, like photoreceptors (in the eyes and head) and thermoreceptors (along the whole body) and are transmitted to the internal pacemaker or the biological clock itself. Thus, a rhythm or a physiological response is generated from the interaction of the stimuli with the pacemaker neurons [68].

The pacemaker neurons are so-called because they express the clock genes, which are the components of the circadian clock. These genes interact with each other and recruit kinases, phosphatases, and transcription factors to generate oscillating expression in a 24-h cycle [69]. They are also responsible for the regulation of many other genes, the clock-controlled genes (CCGs), that are directly associated with tissue-specific functions [70].

*Drosophila melanogaster* is the insect model for studying circadian rhythm, but it is already known that many features of the circadian clock of other insects differ considerably from the fly clock. The *Drosophila* clock is formed by three interconnected autoregulatory loops, in which the proteins coded by *Clock* (*Clk*) and *cycle* (*cyc*) genes play a central role. In the first loop, the heterodimer CLK-CYC binds to an E-box sequence in the promoter region of *period* (*per*) and *timeless* (*tim*) genes, activating their transcription. Once in the cytoplasm, the transcripts are translated into proteins that accumulate during the early night and later enter the nucleus to repress their own transcription. This cycle lasts 24 h due to the posttranslational modifications controlled by the activity of kinases such as DOUBLETIME (DBT), CASEIN KINASE 2A (CK2A), and SHAGGY (SGG), which together with phosphatases such as PP2A stabilize PER and TIM [71, 72]. In the second loop, two transcription factors, VRI and PDP1e, are involved, respectively, in the repression and activation of *Clk* and *cyc* genes. Finally, a third interconnected loop involves the activation of *clockwork orange* (*cwo*) gene and the repression exerted by its product, CWO, in PER targets [73]. **Figure 6** summarizes the *D. melanogaster* molecular circadian clock.

An interesting feature of this clock is its property of environmental synchronization, which adjusts the period to exactly 24 h. One of the most important synchronizers (or *Zeitgebers*) is light. The light-induced resetting mechanism is

dependent upon CRYPTOCHROME (CRY), which is a photoreceptor that induces TIM phosphorylation and leads it to degradation via proteossome [74, 75]. Other stimuli act as *Zeitgebers*, such as temperature and food, and their importance varies from species to species. In *A. aegypti*, temperature cycles are a very strong environmental cue, although the molecular mechanisms for entrainment are still unknown [76].

Molecular studies regarding the circadian clock in *A. aegypti* have been purely descriptive, because of the lack of genetic tools to unravel the function of clock genes in this species. Moreover, the phylogenetic distance between *Aedes aegypti* and *Drosophila melanogaster* implies significant differences between the two species in several biological aspects, beyond the circadian expression pattern. One significant difference is the presence, in *A. aegypti*, of a second cryptochrome gene, called *cryptochrome 2* (*cry2*), which does not exist in *D. melanogaster* [77]. This orthologue in mammals is a transcriptional repressor [78], and some studies done in *Danaus plexippus* confirmed this function in monarch butterflies [79]. Therefore, it is reasonable that this gene also plays this role in mosquitoes as well.

In a general manner, the circadian expression pattern of the main clock genes in *A. aegypti* under light dark cycles (LD12:12, which means 12 h of light followed by 12 h of dark) and constant temperature presents some similarities to what is observed in the *D. melanogaster* model. Genes *per* and *tim* present a cyclic mRNA expression with a peak in ZT 17, in the middle of the dark phase, and *vri* mRNA peak expression occurs some hours earlier than *Pdp1* mRNA peak expression (ZT 11 × ZT 17, respectively). However, two striking differences in relation to the *Drosophila* clock can be seen. The first one is related to the expression of the genes that encode the transcriptional activators, *Clk* and *cyc.* In Drosophila, *Clk* is the

#### **Figure 6.**

*The molecular circadian clock of Drosophila melanogaster. The heterodimer CLK-CYC plays a crucial role in the maintenance of the 24-h cycle, integrating the three regulatory feedback loops. In the first loop, CLK-CYC binds to the E-box regions in per and tim promoters and activates their transcription. Once in the cytoplasm, they are translated into proteins and suffer posttranslational modifications, caused by kinases and phosphatases. Once they can accumulate, the heterodimer PER-TIM enters in the nucleus and represses its own transcription. In the second loop, CLK-CYC heterodimer binds to E-boxes and activates* vri *and* Pdp*1ε transcription. VRI binds to V/P boxes and inhibits* Clk *transcription, while PDP1ε accumulates a few hours later and shifts VRI from V/P box and activates Clk transcription. Finally, the third loop is related to the activation of* clockwork orange *gene and the inhibition of CLK-CYC by this gene product. The light synchronization is fired by a conformational change of CRY, which allows it to bind to TIM and leads to its degradation via proteasome, reinitiating the cycle. Straight arrows indicate following steps in the feedback loops; curved arrows indicate activation interactions; blocked line indicates inhibitory interaction. Colored forms represent proteins and wavy lines represent the mRNA. Based on [71, 72].*

**77**

**6. Conclusions**

*The Effects of Infection on Mosquito Rhythmic Behavior DOI: http://dx.doi.org/10.5772/intechopen.89409*

crucial modifications in clock regulation.

**5.1 The pacemaker neurons in insects**

between arbovirus infection and the pacemaker cells.

It is known that the virus-host interaction has a crucial importance in the spreading of a pathogen, since mutations in the viral genome or the genetic

main activator and presents a peak mRNA expression in antiphase to *per* and *tim*, whereas *cyc* presents a constitutive mRNA expression. On the contrary, *A. aegypti* Clk mRNA expression is arrhythmic, while *cyc* presents a very robust mRNA cycling pattern [8, 80]. Some years later, when *Clk* and *cyc* nucleotides and corresponding proteins were characterized, it was observed that the brain and muscle ARNT-like (BMAL) C-terminus region (BCTR) activation domain is missing in *Drosophila* CYC protein but is present in *A. aegypti* CYC protein [81]. These data can partially explain the variations in *Clk* and *cyc* expression pattern and suggest that there is a dissimilarity in circadian regulation between the two species. The second difference is related to the presence of the *cry2* gene and its bimodal mRNA expression pattern. As mentioned above, cry2 gene is present in *A. aegypti* and other *Diptera* but is absent in *D. melanogaster* [77]. According to putative clock models proposed by Yuan et al. in 2007, the presence or absence of *cryptochrome* genes may lead to

Clock genes are expressed in specific groups of neurons called pacemaker neurons, in the central nervous system of the organism, and are identified as pacemakers due to PER expression [82]. However, the distribution of these cells in the brain can vary from species to species; while in *Manduca sexta*, the pacemaker is located in the dorsal protocerebrum, in *Pachnoda marginata* it is located in the proximal optic lobe [82]. In *Drosophila*, around 150 clock neurons are located in the lateral protocerebrum, close to optic lobe and in the dorsal protocerebrum. The lateral neurons are subdivided in ventrolateral neurons (LNvs), dorsolateral neurons (LNds), lateral posterior neurons (LPNs), and dorsal neurons (DNs) [83]. Each group expresses the clock genes and communicates with each other according to different neurotransmitters, such as pigment-dispersing factor (PDF), which is the most well known [84]. Depending on the combination between neurotransmitters and hormone signaling among the neurons and clock genes' expression during the time of day, distinct neuronal clusters are responsible for different behaviors, such as feeding (which is regulated by the dorsolateral neurons) or temperature preference (regulated by dorsal neurons). On the other hand, the locomotor behavior, for instance, recruits all groups of neurons, which interact with each other to generate activity and rest along the 24-h day [72]. In *A. aegypti* there is not yet a map of the pacemaker neurons. However, it is described that some clustered neurons present a cycling expression of *cyc* and *per* mRNAs in antiphase, which strongly indicates that the areas where pacemaker neurons are found are similar to those observed in *D. melanogaster* [85]. Regarding the arbovirus infection, it was observed that both DENV and ZIKV are able to impact the neurons' spike activity in opposite manners. While ZIKV infection leads to hyperexcitation in a primary neuron culture, DENV2 infection does not seem to alter the spikes. Moreover, ZIKV reaches a plateau in replication around 2 dpi, whereas DENV2 initially decreases its replication and follows an increase in virus titers until 3 dpi [57]. It was already observed that ZIKV presents a strong neurotropism in mosquitoes [57], but it was not possible to associate the infected cells with clock neurons. Additional studies are necessary to establish the relationship

*The Effects of Infection on Mosquito Rhythmic Behavior DOI: http://dx.doi.org/10.5772/intechopen.89409*

*Vector-Borne Diseases - Recent Developments in Epidemiology and Control*

reasonable that this gene also plays this role in mosquitoes as well.

In a general manner, the circadian expression pattern of the main clock genes in *A. aegypti* under light dark cycles (LD12:12, which means 12 h of light followed by 12 h of dark) and constant temperature presents some similarities to what is observed in the *D. melanogaster* model. Genes *per* and *tim* present a cyclic mRNA expression with a peak in ZT 17, in the middle of the dark phase, and *vri* mRNA peak expression occurs some hours earlier than *Pdp1* mRNA peak expression (ZT 11 × ZT 17, respectively). However, two striking differences in relation to the *Drosophila* clock can be seen. The first one is related to the expression of the genes that encode the transcriptional activators, *Clk* and *cyc.* In Drosophila, *Clk* is the

*The molecular circadian clock of Drosophila melanogaster. The heterodimer CLK-CYC plays a crucial role in the maintenance of the 24-h cycle, integrating the three regulatory feedback loops. In the first loop, CLK-CYC binds to the E-box regions in per and tim promoters and activates their transcription. Once in the cytoplasm, they are translated into proteins and suffer posttranslational modifications, caused by kinases and phosphatases. Once they can accumulate, the heterodimer PER-TIM enters in the nucleus and represses its own transcription. In the second loop, CLK-CYC heterodimer binds to E-boxes and activates* vri *and* Pdp*1ε transcription. VRI binds to V/P boxes and inhibits* Clk *transcription, while PDP1ε accumulates a few hours later and shifts VRI from V/P box and activates Clk transcription. Finally, the third loop is related to the activation of* clockwork orange *gene and the inhibition of CLK-CYC by this gene product. The light synchronization is fired by a conformational change of CRY, which allows it to bind to TIM and leads to its degradation via proteasome, reinitiating the cycle. Straight arrows indicate following steps in the feedback loops; curved arrows indicate activation interactions; blocked line indicates inhibitory interaction. Colored forms represent proteins and wavy* 

dependent upon CRYPTOCHROME (CRY), which is a photoreceptor that induces TIM phosphorylation and leads it to degradation via proteossome [74, 75]. Other stimuli act as *Zeitgebers*, such as temperature and food, and their importance varies from species to species. In *A. aegypti*, temperature cycles are a very strong environmental cue, although the molecular mechanisms for entrainment are still unknown [76]. Molecular studies regarding the circadian clock in *A. aegypti* have been purely descriptive, because of the lack of genetic tools to unravel the function of clock genes in this species. Moreover, the phylogenetic distance between *Aedes aegypti* and *Drosophila melanogaster* implies significant differences between the two species in several biological aspects, beyond the circadian expression pattern. One significant difference is the presence, in *A. aegypti*, of a second cryptochrome gene, called *cryptochrome 2* (*cry2*), which does not exist in *D. melanogaster* [77]. This orthologue in mammals is a transcriptional repressor [78], and some studies done in *Danaus plexippus* confirmed this function in monarch butterflies [79]. Therefore, it is

**76**

*lines represent the mRNA. Based on [71, 72].*

**Figure 6.**

main activator and presents a peak mRNA expression in antiphase to *per* and *tim*, whereas *cyc* presents a constitutive mRNA expression. On the contrary, *A. aegypti* Clk mRNA expression is arrhythmic, while *cyc* presents a very robust mRNA cycling pattern [8, 80]. Some years later, when *Clk* and *cyc* nucleotides and corresponding proteins were characterized, it was observed that the brain and muscle ARNT-like (BMAL) C-terminus region (BCTR) activation domain is missing in *Drosophila* CYC protein but is present in *A. aegypti* CYC protein [81]. These data can partially explain the variations in *Clk* and *cyc* expression pattern and suggest that there is a dissimilarity in circadian regulation between the two species. The second difference is related to the presence of the *cry2* gene and its bimodal mRNA expression pattern. As mentioned above, cry2 gene is present in *A. aegypti* and other *Diptera* but is absent in *D. melanogaster* [77]. According to putative clock models proposed by Yuan et al. in 2007, the presence or absence of *cryptochrome* genes may lead to crucial modifications in clock regulation.

#### **5.1 The pacemaker neurons in insects**

Clock genes are expressed in specific groups of neurons called pacemaker neurons, in the central nervous system of the organism, and are identified as pacemakers due to PER expression [82]. However, the distribution of these cells in the brain can vary from species to species; while in *Manduca sexta*, the pacemaker is located in the dorsal protocerebrum, in *Pachnoda marginata* it is located in the proximal optic lobe [82]. In *Drosophila*, around 150 clock neurons are located in the lateral protocerebrum, close to optic lobe and in the dorsal protocerebrum. The lateral neurons are subdivided in ventrolateral neurons (LNvs), dorsolateral neurons (LNds), lateral posterior neurons (LPNs), and dorsal neurons (DNs) [83]. Each group expresses the clock genes and communicates with each other according to different neurotransmitters, such as pigment-dispersing factor (PDF), which is the most well known [84]. Depending on the combination between neurotransmitters and hormone signaling among the neurons and clock genes' expression during the time of day, distinct neuronal clusters are responsible for different behaviors, such as feeding (which is regulated by the dorsolateral neurons) or temperature preference (regulated by dorsal neurons). On the other hand, the locomotor behavior, for instance, recruits all groups of neurons, which interact with each other to generate activity and rest along the 24-h day [72].

In *A. aegypti* there is not yet a map of the pacemaker neurons. However, it is described that some clustered neurons present a cycling expression of *cyc* and *per* mRNAs in antiphase, which strongly indicates that the areas where pacemaker neurons are found are similar to those observed in *D. melanogaster* [85]. Regarding the arbovirus infection, it was observed that both DENV and ZIKV are able to impact the neurons' spike activity in opposite manners. While ZIKV infection leads to hyperexcitation in a primary neuron culture, DENV2 infection does not seem to alter the spikes. Moreover, ZIKV reaches a plateau in replication around 2 dpi, whereas DENV2 initially decreases its replication and follows an increase in virus titers until 3 dpi [57]. It was already observed that ZIKV presents a strong neurotropism in mosquitoes [57], but it was not possible to associate the infected cells with clock neurons. Additional studies are necessary to establish the relationship between arbovirus infection and the pacemaker cells.

### **6. Conclusions**

It is known that the virus-host interaction has a crucial importance in the spreading of a pathogen, since mutations in the viral genome or the genetic

### *Vector-Borne Diseases - Recent Developments in Epidemiology and Control*

background of a mosquito population can enhance or even inhibit the replication of the virus in the mosquito. Beyond this genetic interaction, behavior is also directly related to the vectorial capacity of *A. aegypti*. Reports about the influence of the viral infection on several mosquitoes' behaviors have been increasing along the years. However, we still do not know the way arboviruses modulate the expression of the core clock genes that control behavior. It is even possible that infection does not have a direct effect on the core clock genes themselves but possibly on the genes regulated by them, leading to alterations in behavior and, consequently, impacts on the vectorial capacity. Improving the knowledge on behavioral traits that are susceptible to infection-driven changes (e.g., time-of-day biting activity, time-ofday mating behavior, time-of-day oviposition behavior) can increase the efficacy of strategies of vector control.
