Adaptation to fluctuating environments in a selection experiment with Drosophila melanogaster

A fundamental question in life‐history evolution is how organisms cope with fluctuating environments, including variation between stressful and benign conditions. For short‐ lived organisms, environments commonly vary between generations. Using a novel experimental design, we exposed wild‐derived Drosophila melanogaster to three different selection regimes: one where generations alternated between starvation and benign conditions, and starvation was always preceded by early exposure to cold; another where starvation and benign conditions alternated in the same way, but cold shock sometimes preceded starvation and sometimes benign conditions; and a third where conditions were always benign. Using six replicate populations per selection regime, we found that selected flies increased their starvation resistance, most strongly for the regime where cold and starvation were reliably combined, and this occurred without decreased fecundity or extended developmental time. The selected flies became stress resistant, displayed a pronounced increase in early life food intake and resource storage. In contrast to previous experiments selecting for increased starvation resistance in D. melanogaster, we did not find increased storage of lipids as the main response, but instead that, in particular for females, storage of carbohydrates was more pronounced. We argue that faster mobilization of carbohydrates is advantageous in fluctuating environments and conclude that the phenotype that evolved in our experiment corresponds to a compromise between the requirements of stressful and benign environments

Characterization of reproductive dormancy in male Drosophila melanogaster

Insects are known to respond to seasonal and adverse environmental changes by entering dormancy, also known as diapause. In some insect species, including Drosophila melanogaster, dormancy occurs in the adult organism and postpones reproduction. This adult dormancy has been studied in female flies where it is characterized by arrested development of ovaries, altered nutrient stores, lowered metabolism, increased stress and immune resistance and drastically extended lifespan. Male dormancy, however, has not been investigated in D. melanogaster, and its physiology is poorly known in most insects. Here we show that unmated 3-6 h old male flies placed at low temperature (11°C) and short photoperiod (10 Light:14 Dark) enter a state of dormancy with arrested spermatogenesis and development of testes and male accessory glands. Over three weeks of dormancy we see a dynamic increase in stored carbohydrates and an initial increase and then a decrease in lipids. We also note an up-regulated expression of genes involved in metabolism, stress responses and innate immunity. Interestingly, we found that male flies that entered reproductive dormancy do not attempt to mate females kept under non-diapause conditions (25°C, 12L:12D), and conversely non-dormant males do not mate females in dormancy. In summary, our study shows that male D. melanogaster can enter reproductive dormancy. However, our data suggest that dormant male flies deplete stored nutrients faster than females, studied earlier, and that males take longer to recover reproductive capacity after reintroduction to non-diapause conditions

Goldfish can recover after short-term exposure to 2,4- dichlorophenoxyacetate: Use of blood parameters as vital biomarkers

This study investigated the effects of 2,4-dichlorophenoxyacetic acid (2,4-D), a widely used herbicide, on the metabolism of goldfish, Carassius auratus, using only vital (non-lethal) approaches. After 96 h exposure to 1, 10 or 100 mg/L of 2,4-D selected hematological (total hemoglobin and hematocrit) and biochemical (glucose content, aspartate transaminase and acetylcholinesterase activities) parameters were unchanged in blood of exposed fish. At 100 mg/L of 2,4-D lymphocyte numbers decreased by 8%, whereas promyelocyte and metamyelocyte numbers increased by 7- and 2-fold, respectively. Exposure to 100 mg/L of 2,4-D also elevated carbonyl protein levels (by 2-fold), triglyceride content (by 43%) and alanine transaminase activity (by 46%) in goldfish plasma. All of these hematological and biochemical parameters reverted to control values after a 96 h recover

Oxidative stress as a mechanism for toxicity of 2,4-dichlorophenoxyacetic acid (2,4-D): Studies with goldfish gills

The effects of exposure to the widely used herbicide, 2,4- dichlorophenoxyacetic acid (2,4-D), at environmentally permitted (1 mg L -1), slightly toxic (10 mg L-1), and highly toxic (100 mg L-1) concentrations were analyzed in gills of goldfish, Carassius auratus, a popular fish model for ecotoxicological research. Fish were exposed to the pesticide in water for 96 h and an additional group of fish were treated by the highest 2,4-D concentration and then allowed to recover for further 96 h. Among markers of oxidative stress, goldfish exposure to 2,4-D did not affect carbonyl protein levels in the gills, but fish exp

Catalase activity as a potential vital biomarker of fish intoxication by the herbicide aminotriazole

The objective of this study was to investigate the effects of the herbicide 3-amino-1,2,4-triazole (AMT) on the activities of catalase and lactate dehydrogenase (LDH) in blood (plasma and erythrocytes) and eight solid tissues of goldfish, Carassius auratus. Injection of goldfish with AMT (0.5. mg/gww AMT in 0.9% NaCl) resulted in a significant decrease in catalase activity 24. h post-injection in most tissues investigated. In white and red muscle, kidney, heart, liver, brain and erythrocytes the activity of catalase decreased by 61%, 69%, 64%, 48%, 4

Effects of diapause conditions on gut-related structures in <i>Canton S</i> flies.

<p><b>A</b> The intestinal epithelium appears to age more slowly during diapause. The epithelial cells (EC) were marked with NP1-Gal4 driven expression of GFP (green) in combination with nuclear staining with Hoechst 33342 (blue). The insets display enlarged view of nuclear staining. The flies tested were kept under control and diapause conditions as described earlier for 3 h (3 h N) or 3 days normal conditions (3 dN), 3–9 weeks normal conditions (3–9 w N), 3 days (3 d D) or 3–9 weeks diapause conditions (3–9 w D) and finally for 1–9 weeks recovery conditions after 6 weeks of diapause (1–9 w R). The age-associated changes in growth of EC size and disruption of the EC monolayer in the midgut are delayed by at least 3 weeks in diapausing flies. The yellow asterisks indicate small polyploid cells (sign of intestinal dysplasia). <b>B</b> The length of the midgut was measured during diapause (conditions as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113051#pone-0113051-g002" target="_blank">Fig. 2A</a>). The midgut is significantly longer in flies kept 1-week at normal conditions (C1), know to feed properly, than in 3–6 h old flies (C0). In diapausing flies (D1–D12) the midgut is shorter than in C1 controls and then becomes significantly longer after recovery from diapause (R1′, R1–2). Data are presented as means ± S.E.M, <i>n</i> = 6–9 randomly selected flies for each sample point. Significance of differences from the 1-week control (C1) is indicated, as well as between groups indicated by connectors, * <i>p</i><0.05, ** <i>p</i><0.01, *** <i>p</i><0.001, N.S. not significantly different (ANOVA followed with Tukey test). <b>C</b> The width of the midgut did not change much during diapause, except a significant increase after 1-week recovery from diapause (R1′, R1). Data are presented as means ± S.E.M, <i>n</i> = 5–8 randomly selected flies for each sample point. Significance of differences from the 1-week control (C1) is indicated, as well as between groups indicated by connectors, ^#<i>p</i><0.05 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test).</p

Effects of diapause and other conditions on <i>w¹¹¹⁸</i> flies.

<p>Macronutrient composition in hemolymph and body of flies kept for 1–12 weeks at 11°C and 10L:14D, light/dark (D1–D12) and recovery (R) from diapause for 1 and 1–2 weeks at 25°C and 12L:12D after 3 weeks (R1′) or 6 weeks (R1 and R2) of diapause. Virgin flies, kept for 1 week at non-diapausing conditions (C1) and recently hatched 3–6 h old flies (C0) were used as a controls. Data are presented as means ± S.E.M, <i>n</i> = 3–5 independent replicates with 10–15 flies in every replicate. Significantly different either from the control (C1) with *<i>p</i><0.05, ** <i>p</i><0.01, *** <i>p</i><0.001 or from the indicated group as assessed by ANOVA, followed by Tukey test or alternatively with ^#<i>p</i><0.05, ^##<i>p</i><0.01, ^###<i>p</i><0.001 as assessed by Kruskal-Wallis test followed with Wilcoxon pairwise comparison. N.S. – values are not significantly different. Vs, versus.</p><p>Effects of diapause and other conditions on <i>w¹¹¹⁸</i> flies.</p

Selective effects of diapause conditions on expression of innate immune genes.

<p>The relative expression of four immune genes was determined in six groups of female <i>Canton S</i> flies: virgin 3–6 h old flies (C0), one week old uninfected and non-diapausing flies (C1), uninfected flies kept for 3 weeks either at 11°C and 10L:14D (diapause, D3) or at 25°C and 12L:12D, (normal conditions, N3) and infected flies (cross hatched bars) kept under diapause (D3) or non-diapause conditions (N3) for 3 weeks. Infected flies were injected with a suspension of <i>Micrococcus luteus</i> and <i>Escherichia coli</i> and kept for an additional 3 hours before freezing and RNA extraction. Data are presented as means ± S.E.M, <i>n</i> = 3–4 replicates with 10–15 flies in each. Significance compared to the newly hatched control (C1) which was set at one, or as indicated by connectors: * <i>p</i><0.05, ** <i>p</i><0.01, *** <i>p</i><0.001, N.S. – not significant (ANOVA followed with Tukey test) or ^#<i>p</i><0.05, ^##<i>p</i><0.01 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test). <b>A</b> The <i>Drosomycin</i> expression was significantly upregulated in flies diapausing for 3 weeks (D3) in both infected and uninfected flies compared to non-diapausing 1-week old (C1) and 3-week old flies (N3). Infection further increased transcripts in both N3 and D3 flies. <b>B</b><i>Cecropin A1</i> was significantly upregulated during diapause (D3) versus normal conditions (C1), but not N3, in uninfected flies only if comparing C1 to D3. Infection drastically increased transcripts in both diapausing and nondiapausing flies. <b>C</b> The <i>peptidoglycan recognition protein SB1</i> (<i>PGRP-SB1</i>) transcript level are not affected by diapause, only infection increased it. <b>D</b><i>Diptericin</i> also increased only due to infection. For the investigated immune genes we did not find a differences in expression levels between 1-week old (C1) and newly eclosed (C0) flies. Similar results were obtained with flies that were reared on food supplemented with antibiotics (see Fig. S2 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113051#pone.0113051.s002" target="_blank">File S1</a>).</p