Figure 1. Control of pupation height behavior by the internal microenvironment of the culture vial.

(A) Schematic representation of a Drosophila culture vial and the internal humidity gradient generated within the microenvironment of the plugged vial. Higher humidity at the bottom near the food surface is indicated as darker shading decreasing to low humidity at the top of the vial. Pupae are shown as black ovals within the designated zones indicated as height(cm) above the vial bottom used to calculate the pupation height index(PHI). PHI = ((#pupae>3 cm)-(#pupae<3 cm))/total # pupae. (BC) Pupation height behavior for ppk1-GAL4/+ control larvae represented by the indicated Pupation Height Index; (B) Pupation height is strongly influenced by external humidity. With constant larval density(50 larvae/vial), low external humidity(25%) causes larvae to pupate at sites lower in the vial indicated as a downward deflection. At higher humidity(50% and 75%), larvae choose to predominantly pupate at higher sites within the vial indicated as an upward deflection. (C) Pupation height is influenced by larval density. With vials grown at constant external humidity(50%), low larval density(25 larvae/vial) causes a low pupation height. Higher larval density causes a drastic increase in pupation height resulting from the altered microenvironment as larvae exit the food source. Error bars represent SEM with n≥15 for all values. ***P<.0001, one-way ANOVA with Tukey posttest.