Fig 1.
Short photoperiod induces diapause egg production.
(A) Schematic of rearing conditions, developmental timeline, and diapause (DP) egg status. Eggs from the same clutch were common-garden reared under 26°C, 16L:8D conditions. At the pupal stage, animals were separated into three conditions: high temperature, long photoperiod (red); low temperature, long photoperiod (purple); low temperature, short photoperiod (blue). B) Hatching assay to determine DP egg status. % of eggs hatched after exposure to hatching stimulus within 3 weeks of oviposition. Data is shown as mean with SEM (n = 3 replicates; >50 eggs per replicate; one-way ANOVA followed by Tukey’s multiple comparisons test; * = p < 0.05, ** = p < 0.01). C and D) Egg width (C) and length (D) measurement. Data is shown as median with range (n = 2 replicate experiments; 130–190 eggs per replicate; one-way ANOVA followed by Tukey’s multiple comparisons test; *** = p < 0.001, **** = p < 0.0001). Created with Biorender.com.
Fig 2.
Temperature impacts meal size in a 2-choice assay and egg retention.
A) Two-choice assay to measure preference for “no protein” sugar meal (10% sucrose; bottom) versus “protein rich” meal (10% sucrose + 10% BSA; top) using fluorescent dyes to distinguish between meals. Dot size represents total meal size in μL of an individual female and preference index was calculated as (% “BSA+Sucrose” meal—% “Sucrose only” meal). n = 6 experimental replicates, 4–14 females per replicate. Each experimental replicate is from an independently-reared cohort with matched groups for each condition. B-D) Meal size for B) combined “sucrose” and “BSA + sucrose” meals; C) “BSA + sucrose” meal only; D) “sucrose” meal only. Data is shown as median with range (n = 6 experimental replicates, 4–14 females per replicate; Kruskal-Wallis with Dunn’s multiple comparisons test; * = p < 0.05, ** = p < 0.01, *** = p < 0.001, **** = p < 0.0001). E) Sugar meal size in a “no choice” assay. Data is shown as median with range (n = 4–5 experimental replicates, 7–16 females per replicate; Kruskal-Wallis with Dunn’s multiple comparisons test, n.s. = p > 0.05. F) Quantification of body weight per female before and after blood feeding in “no choice” assay. Data is shown as median with range (n = 2 experimental replicates, 14–33 measurements per replicate; Kruskal-Wallis with Dunn’s multiple comparisons test; letters indicate statistically distinct groups p < 0.05). G) Number of eggs laid/total eggs produced per female. Data is shown as mean with SEM (n = 2–5 experimental replicates; n = 13–28 females per replicate; one-way ANOVA with Tukey’s multiple comparisons test; *** = p < 0.001). H) Daily oviposition measured for days 1–14 post blood meal (n = 2 experimental replicates; 21–28 females per replicate).
Fig 3.
Temperature impacts oocyte heterogeneity before blood feeding.
A) Representative images of ovarioles from females at pre-blood meal, and 1, 3, 7, and 14 days after a blood meal housed in the conditions indicated. B) Left diagram shows an example of a primary ovarian follicle. Ovarian development was measured by calculating the percentage of the follicle (green) occupied by the nurse cells (gray). As the follicle matures into an egg, the percentage occupied by the nurse cells decreases. Right graph shows all oocyte developmental stages at each timepoint and environmental conditions. Each timepoint includes measurements from 20–30 ovarioles from 3–7 pairs of ovaries. Data is shown as mean with SEM.
Fig 4.
Short photoperiod increases starvation resistance.
A) Daily survival for adult females provided with access to water only. Data is shown as mean with SEM (n = 2–4 experimental replicates, 24–42 females per replicate; Log-rank (Mantel-Cox) test; *** = p < 0.001, **** = p < 0.0001). B) Total daily locomotor activity over 3 days by females provided access to water only. Data is shown as median with range. (n = 3 experimental replicates, n = 6–16 females/replicate; Kruskal-Wallis with Dunn’s multiple comparisons test; *** = p < 0.001, **** = p < 0.0001) C—F) Relative C) triacylglycerols (TAG), D) glucose, E) glycogen, and F) trehalose levels in whole females (groups of 5) collected after 0 and 4 days of starvation. Data is shown as median with range. n = 3 experimental replicates, 4 groups of 5 females per replicate; two-way ANOVA with Tukey’s multiple comparisons test; letters indicate statistically distinct groups p < 0.05).