QTL Determining Diel Flight Activity in Male Culex pipiens Mosquitoes.

Members of the Culex pipiens complex differ in physiological traits that facilitate their survival in diverse environments. Assortative mating within the complex occurs in some regions where autogenous (the ability to lay a batch of eggs without a blood meal) and anautogenous populations are sympatric, and differences in mating behaviors may be involved. For example, anautogenous populations mate in flight/swarms, while autogenous populations often mate at rest. Here, we characterized flight activity of males and found that anautogenous strain males were crepuscular, while autogenous strain males were crepuscular and nocturnal, with earlier activity onset times. We conducted quantitative trait locus (QTL) mapping to explore the genetic basis of circadian chronotype (crepuscular vs. crepuscular and nocturnal) and time of activity onset. One major-effect QTL was identified for chronotype, while 3 QTLs were identified for activity onset. The highest logarithm of the odds (LOD) score for the chronotype QTL coincides with a chromosome 3 marker that contains a 15-nucleotide indel within the coding region of the canonical clock gene, cryptochrome 2. Sequencing of this locus in 7 different strains showed that the C-terminus of CRY2 in the autogenous forms contain deletions not found in the anautogenous forms. Consequently, we monitored activity in constant darkness and found males from the anautogenous strain exhibited free running periods of ~24 h while those from the autogenous strain were ~22 h. This study provides novel insights into the genetic basis of flight behaviors that likely reflect adaptation to their distinct ecological niches.

Genome-wide profiling of 24 hr diel rhythmicity in the water flea, Daphnia pulex: network analysis reveals rhythmic gene expression and enhances functional gene annotation.

BACKGROUND: Marine and freshwater zooplankton exhibit daily rhythmic patterns of behavior and physiology which may be regulated directly by the light:dark (LD) cycle and/or a molecular circadian clock. One of the best-studied zooplankton taxa, the freshwater crustacean Daphnia, has a 24 h diel vertical migration (DVM) behavior whereby the organism travels up and down through the water column daily. DVM plays a critical role in resource tracking and the behavioral avoidance of predators and damaging ultraviolet radiation. However, there is little information at the transcriptional level linking the expression patterns of genes to the rhythmic physiology/behavior of Daphnia. RESULTS: Here we analyzed genome-wide temporal transcriptional patterns from Daphnia pulex collected over a 44 h time period under a 12:12 LD cycle (diel) conditions using a cosine-fitting algorithm. We used a comprehensive network modeling and analysis approach to identify novel co-regulated rhythmic genes that have similar network topological properties and functional annotations as rhythmic genes identified by the cosine-fitting analyses. Furthermore, we used the network approach to predict with high accuracy novel gene-function associations, thus enhancing current functional annotations available for genes in this ecologically relevant model species. Our results reveal that genes in many functional groupings exhibit 24 h rhythms in their expression patterns under diel conditions. We highlight the rhythmic expression of immunity, oxidative detoxification, and sensory process genes. We discuss differences in the chronobiology of D. pulex from other well-characterized terrestrial arthropods. CONCLUSIONS: This research adds to a growing body of literature suggesting the genetic mechanisms governing rhythmicity in crustaceans may be divergent from other arthropod lineages including insects. Lastly, these results highlight the power of using a network analysis approach to identify differential gene expression and provide novel functional annotation.

Diel flight activity of wild-caught Anopheles farauti (s.s.) and An. hinesorum malaria mosquitoes from northern Queensland, Australia.

BACKGROUND: Species in the Anopheles farauti complex are major malaria vectors in the Asia Pacific region. Anopheline mosquitoes exhibit circadian and diel rhythms in sugar- and blood-feeding (biting), flight activity, oviposition, and in some species, a short-lived dusk/early night associated swarming behaviour during which mating occurs. A behavioural study of wild-caught mosquitoes from Queensland, Australia was conducted to investigate the differences in diel rhythmic flight activity between two cryptic species in several reproductive states. RESULTS: The 24-hour flight activity of individual adult female mosquitoes under light:dark cycle conditions were monitored with a minute-to-minute time resolution using an infrared beam break method. Mosquitoes were analyzed for reproductive state (insemination and parity) and identified to species [An. farauti (s.s.) Laveran and An. hinesorum Schmidt] by PCR analysis. We compared daily total flight activity, timing of activity onset, the peak in early nocturnal activity, and patterns of activity during the scotophase (night). Species-specific differences between An. farauti and An. hinesorum were observed. Compared to An. farauti, An. hinesorum had an earlier onset of dusk activity, an earlier peak in nocturnal activity, and a higher level of activity at the onset of darkness. Small differences between species were also observed in the pattern of the dusk/early-night bouts of activity. A second nocturnal peak in inseminated nulliparous An. hinesorum was also observed during the middle of the scotophase. CONCLUSIONS: The behavioural differences between these two sympatric species of the An. farauti complex might contribute to subtle differences in habitat adaptation, the timing of host-seeking and/or sugar-feeding activity. This study provides baseline data for analysis of populations of mosquitoes from other geographical regions where these species are malaria vectors, such as in the Solomon Islands and Papua New Guinea. This is important as selective pressures due to long-term use of indoor residual spraying of insecticides and insecticide-treated bed nets are shifting the nocturnal profile of biting behaviour of these vectors to earlier in the night.

Light manipulation of mosquito behaviour: acute and sustained photic suppression of biting activity in the Anopheles gambiae malaria mosquito.

BACKGROUND: Host-seeking behaviours in anopheline mosquitoes are time-of-day specific, with a greater propensity for nocturnal biting. We investigated how a short exposure to light presented during the night or late day can inhibit biting activity and modulate flight activity behaviour. RESULTS: Anopheles gambiae (s.s.), maintained on a 12:12 LD cycle, were exposed transiently to white light for 10-min at the onset of night and the proportion taking a blood meal in a human biting assay was recorded every 2 h over an 8-h duration. The pulse significantly reduced biting propensity in mosquitoes 2 h following administration, in some trials for 4 h, and with no differences detected after 6 h. Conversely, biting levels were significantly elevated when mosquitoes were exposed to a dark treatment during the late day, suggesting that light suppresses biting behaviour even during the late daytime. These data reveal a potent effect of a discrete light pulse on biting behaviour that is both immediate and sustained. We expanded this approach to develop a method to reduce biting propensity throughout the night by exposing mosquitoes to a series of 6- or 10-min pulses presented every 2 h. We reveal both an immediate suppressive effect of light during the exposure period and 2 h after the pulse. This response was found to be effective during most times of the night: however, differential responses that were time-of-day specific suggest an underlying circadian property of the mosquito physiology that results in an altered treatment efficacy. Finally, we examined the immediate and sustained effects of light on mosquito flight activity behaviour following exposure to a 30-min pulse, and observed activity suppression during early night, and elevated activity during the late night. CONCLUSIONS: As mosquitoes and malaria parasites are becoming increasingly resistant to insecticide and drug treatment respectively, there is a necessity for the development of innovative control strategies beyond insecticide-treated nets (ITNs) and residual spraying. These data reveal the potent inhibitory effects of light exposure and the utility of multiple photic pulses presented at intervals during the night/late daytime, may prove to be an effective tool that complements established control methods.

Dissociation between diurnal cycles in locomotor activity, feeding behavior and hepatic PERIOD2 expression in chronic alcohol-fed mice.

Chronic alcohol consumption contributes to fatty liver disease. Our studies revealed that the hepatic circadian clock is disturbed in alcohol-induced hepatic steatosis, and effects of chronic alcohol administration upon the clock itself may contribute to steatosis. We extended these findings to explore the effects of chronic alcohol treatment on daily feeding and locomotor activity patterns. Mice were chronically pair-fed ad libitum for 4 weeks using the Lieber-DeCarli liquid diet, with calorie-controlled liquid and standard chow diets as control groups. Locomotor activity, feeding activity, and real-time bioluminescence recording of PERIOD2::LUCIFERASE expression in tissue explants were measured. Mice on liquid control and chow diets exhibited normal profiles of locomotor activity, with a ratio of 22:78% day/night activity and a peak during early night. This pattern was dramatically altered in alcohol-fed mice, marked by a 49:51% ratio and the absence of a distinct peak. While chow-diet fed mice had a normal 24:76% ratio of feeding activity, with a peak in the early night, this pattern was dramatically altered in both liquid-diet groups: mice had a 43:57% ratio, and an absence of a distinct peak. Temporal differences were also observed between the two liquid-diet groups during late day. Cosinor analysis revealed a ∼4-h and ∼6-h shift in the alcohol-fed group feeding and locomotor activity rhythms, respectively. Analysis of hepatic PER2 expression revealed that the molecular clock in alcohol-fed and control liquid-diet mice was shifted by ∼11 h and ∼6 h, respectively. No differences were observed in suprachiasmatic nucleus explants, suggesting that changes in circadian phase in the liver were generated independently from the central clock. These results suggest that chronic alcohol consumption and a liquid diet can differentially modulate the daily rhythmicity of locomotor and feeding behaviors, aspects that might contribute to disturbances in the circadian timing system and development of hepatic steatosis.