Oviposition by Anopheles gambiae

Anopheles gambiae eggsThe factors and behavioral processes by which Anopheles gambiae female mosquitoes select sites for egg laying are poorly known but ultimately dictate the sites of larval development. They further determine the necessary range of movement of females so that they may locate and utilize appropriately acceptable sites.  Oviposition traps could be designed to take advantage of attractant factors.  Therefore, we conducted a range of experiments designed to elucidate those important factors and processes.  We originally postulated that heterotrophic microbial metabolic activity in puddle habitats of Anopheles gambiae larvae produces unique volatile organic compounds that attract and arrest gravid females at ovipositional sites and release egg deposition. However, this hypothesis received little support.  Direct tests of the chemical specialization hypothesis involved a standardized bioassay in which known amounts of a wide range of volatile chemicals representing heterotrophic bacterial activity were added in appropriate soluble form to wet, white filter papers placed on top of a 9 cm petri dish filled with wet cotton balls.  Phenol, p-cresol, and p-ethylphenol increased oviposition over controls by ca. two-fold. Indole and skatole reduced egg output significantly, but less than two-fold.  Heterotrophic bacteria cultured from Anopheles gambiae larval habitats in Kenya were tested for attractiveness in a newly designed bioassay system that isolated odors from contact effects.  Bacterial odors were repellent to gravid females; oviposition response was 8-fold greater when bacteria were absent compared to when bacteria were present in cultured and actively growing form on media in the lower plate of the apparatus.  Later tests revealed that odors from autotrophic bacteria (Microcystis aeruginosa, a cyanobacterium or “blue-green alga”) stimulated a mild, positive oviposition response.


Given only moderately positive, and sometimes negative responses, to odors from volatile chemicals and bacteria, we broadened our thinking to consider visual, tactile, odor, and taste cues of substrates.  A robust behavioral analysis of females revealed 9 behavioral categories associated with oviposition.  Experienced females found a site faster and laid eggs more quickly than did naïve females.  A dark and moist substrate was found to be the most stimulatory to oviposition.  Merely darkening the bottom of a dish of water increased egg laying by > 9 fold. Moreover, Huang, et al 2007An. gambiae females deposited as many or more eggs on moist mud as they did on free water.  Egg output progressively diminished with diminishing moisture content of mud.  We adapted a Spectrum Technologies leaf-wetness meter to quantify soil-moisture content in the lab and field and related it to probability of egg deposition by An. gambiae using a standard curve inter-relating egg output against reading of the soil-wetness meter.  This finding provides us with a tool for predicting suitably moist soils for egg laying in nature.  Additional experiments designed to elucidate the significance of darkness of ovipositional substrate, contrasted with background, showed that contrast and darkness of substrate were very important in egg laying, and that An. gambiae females could not discriminate color (hue) of substrate but could discriminate brightness and contrast.  With this accumulated knowledge, we designed a field ovipositional sampler incorporating elements of dark and contrasting substrate, and moisture, and tested it successfully in western Kenya. 



Papers of interest:


Huang J, Walker ED, Giroux P, Vulule J, Miller JR. Ovipositional site selection by Anopheles gambiae: influences of substrate moisture and texture.  Med Vet Entomol 2005; 19:442-450.  PMID: 16336309


Huang J, Walker ED, Vulule JM, Miller JR. Daily temperature profiles in and around western Kenyan larval habitats of Anopheles gambiae as related to egg mortality.  Malaria J 2006; 5:87.  PMID: 17038186


Huang J, Walker ED, Otienoburu PE, Amimo F, Vulule JM, Miller JR. Laboratory tests of oviposition by the African malaria mosquito, Anopheles gambiae, on dark soil as influenced by presence or absence of vegetation.  Malaria J 2006; 5:88.  PMID: 17038187


Miller JM, Huang J, Vulule J, Walker E. Life on the edge: African malaria mosquito larvae are amphibious.  Naturwissenschaften 2006; 94:195-9.  PMID: 17139499


Fritz ML, Huang J, Walker ED, Bayoh MB, Vulule JM, Miller JR.  Automated measurement of ovipositional periodicity of caged Anopheles gambiae individuals.  J Circadian Rhythms 2008, 6:2.  doi:10.1186/1740-3391-6-2.  PMID: 18221544


Huang J, Walker ED, Vulule JM, Miller JR.  The influence of darkness and visual contrast on oviposition by Anopheles gambiae (Diptera: Culicidae).  Physiol Entomol 2007; 32: 32-40. 


Otienoburu P, Bayoh MN, Gimnig JE, Huang J, Walker ED, Otieno MF, Vulule JM, Miller JR.  Anopheles gambiae oviposition as influenced by type of water infused into black and red soils in western Kenya.  Intl J Trop Insect Sci 2007; DOI: 10.1017/S1742758407706902.


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