Inbreeding does not alter the response to an experimental heat wave in a freshwater snail.

Global climate change affects natural populations of many species by increasing the average temperature and the frequency of extreme weather events (e.g. summer heat waves). The ability of organisms to cope with these environmental changes can, however, depend on their genetic properties. For instance, genetic load owing to inbreeding could alter organisms' responses to climate change-mediated environmental changes but such effects are often overlooked. We investigated the effects of an experimental heat wave (25°C versus 15°C) on life history (reproduction, size) and constitutive immune defence traits (phenoloxidase-like and antibacterial activity of haemolymph) in relation to inbreeding by manipulating the mating type (outcrossing, self-fertilization) in two populations of a hermaphroditic freshwater snail, Lymnaea stagnalis. High temperature increased reproduction and size of snails but impaired their immune function. In one of the two study populations, inbreeding reduced reproductive output of snails indicating inbreeding depression. Furthermore, this effect did not depend on the temperature snails were exposed to. Our results suggest that L. stagnalis snails can be negatively affected by inbreeding but it may not alter their responses to heat waves.

Potential for adaptation to climate change: family-level variation in fitness-related traits and their responses to heat waves in a snail population.

BACKGROUND: On-going global climate change poses a serious threat for natural populations unless they are able to evolutionarily adapt to changing environmental conditions (e.g. increasing average temperatures, occurrence of extreme weather events). A prerequisite for evolutionary change is within-population heritable genetic variation in traits subject to selection. In relation to climate change, mainly phenological traits as well as heat and desiccation resistance have been examined for such variation. Therefore, it is important to investigate adaptive potential under climate change conditions across a broader range of traits. This is especially true for life-history traits and defences against natural enemies (e.g. parasites) since they influence organisms' fitness both directly and through species interactions. We examined the adaptive potential of fitness-related traits and their responses to heat waves in a population of a freshwater snail, Lymnaea stagnalis. We estimated family-level variation and covariation in life history (size, reproduction) and constitutive immune defence traits [haemocyte concentration, phenoloxidase (PO)-like activity, antibacterial activity of haemolymph] in snails experimentally exposed to typical (15 °C) and heat wave (25 °C) temperatures. We also assessed variation in the reaction norms of these traits between the treatments. RESULTS: We found that at the heat wave temperature, snails were larger and reproduced more, while their immune defence was reduced. Snails showed high family-level variation in all examined traits within both temperature treatments. The only negative genetic correlation (between reproduction and antibacterial activity) appeared at the high temperature. However, we found no family-level variation in the responses of most examined traits to the experimental heat wave (i.e. largely parallel reaction norms between the treatments). Only the reduction of PO-like activity when exposed to the high temperature showed family-level variation, suggesting that the cost of heat waves may be lower for some families and could evolve under selection. CONCLUSION: Our results suggest that there is genetic potential for adaptation within both thermal environments and that trait evolution may not be strongly affected by trade-offs between them. However, rare differences in thermal reaction norms across families indicate limited evolutionary potential in the responses of snails to changing temperatures during extreme weather events.

Infection success of Echinoparyphium aconiatum (Trematoda) in its snail host under high temperature: role of host resistance.

BACKGROUND: Extreme weather events such as summer heat waves become more frequent owing to global climate change and are predicted to alter disease dynamics. This is because high temperatures can reduce host immune function. Predicting the impact of climate change on host-parasite interactions is, however, difficult as temperature may also affect parasite infective stages and other host characteristics determining the outcome of interaction. METHODS: Two experiments were conducted to investigate these phenomena in a Lymnaea stagnalis-Echinoparyphium aconiatum (Trematoda) interaction. In the first experiment, the effects of exposure of snails to experimental heat waves [maintenance at 25°C vs. 15°C (control)] with different durations (3 days, 7 days) on the infection success of parasite cercariae was examined. In the second experiment, the infection success was examined under similar conditions, while controlling for the possible temperature effects on cercariae and at least partly also for host physiological changes that take place rapidly compared to alterations in immune function (exposure to cercariae at intermediate 20°C). RESULTS: In the first experiment, increased infection success at 25°C was found independently of the duration of the heat wave. In the second experiment, increased infection success was found only in snails maintained at 25°C for 7 days, a treatment in which snail immune defence is known to be impaired. CONCLUSIONS: These results suggest that the effects of host resistance in determining overall parasite infection success can be overridden by effects of temperature on parasite transmission stages and/or alterations in other host traits than immune defence.

Activation of the immune defence of the freshwater snail Lymnaea stagnalis by different immune elicitors.

Understanding the outcomes of host-parasite interactions in nature is in high demand as parasites and pathogens are important for several ecological and evolutionary processes. Ecological immunology (ecoimmunology) has a key role in reaching this goal because immune defence is the main physiological barrier against infections. To date, ecoimmunological studies largely lean on measuring constitutive immune defences (components of defence that are always active). However, understanding the role of inducible components of immune function is important as the immune system is largely an inducible defence. Measuring such defences can be complicated as different parasites may activate different immune cascades, and expression of different immune traits may not be independent. We examined the suitability of different immune activation techniques for the freshwater snail Lymnaea stagnalis. By experimentally challenging snails with different immune elicitors [injection with snail saline (i.e. wounding), lyophilized Escherichia coli cells, lyophilized Micrococcus lysodeikticus cells, healthy snail gonad, and trematode-infected snail gonad; maintenance in microorganism-enriched water] and measuring phenoloxidase-like and antibacterial activity of their haemolymph, we found increased immune activity against some immune elicitors, but also decreased activity. Our findings suggest potentially complicated relationships among immune traits, and propose suitable techniques for ecological studies in this study system.

An experimental heat wave changes immune defense and life history traits in a freshwater snail.

The predicted increase in frequency and severity of heat waves due to climate change is expected to alter disease dynamics by reducing hosts' ability to resist infections. This could take place via two different mechanisms: (1) through general reduction in hosts' performance under harsh environmental conditions and/or (2) through altered resource allocation that reduces expression of defense traits in order to maintain other traits. We tested these alternative hypotheses by measuring the effect of an experimental heat wave (25 vs. 15°C) on the constitutive level of immune defense (hemocyte concentration, phenoloxidase [PO]-like activity, antibacterial activity of hemolymph), and life history traits (growth and number of oviposited eggs) of the great pond snail Lymnaea stagnalis. We also manipulated the exposure time to high temperature (1, 3, 5, 7, 9, or 11 days). We found that if the exposure to high temperature lasted <1 week, immune function was not affected. However, when the exposure lasted longer than that, the level of snails' immune function (hemocyte concentration and PO-like activity) was reduced. Snails' growth and reproduction increased within the first week of exposure to high temperature. However, longer exposures did not lead to a further increase in cumulative reproductive output. Our results show that short experimental heat waves do not alter immune function but lead to plastic responses that increase snails' growth and reproduction. Thus, although the relative expression of traits changes, short experimental heat waves do not impair snails' defenses. Negative effects on performance get pronounced when the heat waves are prolonged suggesting that high performance cannot be maintained over long time periods. This ultimately reduces the levels of defense traits.

Tbc1d1 mutation in lean mouse strain confers leanness and protects from diet-induced obesity.

We previously identified Nob1 as a quantitative trait locus for high-fat diet-induced obesity and diabetes in genome-wide scans of outcross populations of obese and lean mouse strains. Additional crossbreeding experiments indicated that Nob1 represents an obesity suppressor from the lean Swiss Jim Lambert (SJL) strain. Here we identify a SJL-specific mutation in the Tbc1d1 gene that results in a truncated protein lacking the TBC Rab-GTPase-activating protein domain. TBC1D1, which has been recently linked to human obesity, is related to the insulin signaling protein AS160 and is predominantly expressed in skeletal muscle. Knockdown of TBC1D1 in skeletal muscle cells increased fatty acid uptake and oxidation, whereas overexpression of TBC1D1 had the opposite effect. Recombinant congenic mice lacking TBC1D1 showed reduced body weight, decreased respiratory quotient, increased fatty acid oxidation and reduced glucose uptake in isolated skeletal muscle. Our data strongly suggest that mutation of Tbc1d1 suppresses high-fat diet-induced obesity by increasing lipid use in skeletal muscle.

Endocytosis of the glucose transporter GLUT8 is mediated by interaction of a dileucine motif with the beta2-adaptin subunit of the AP-2 adaptor complex.

The glucose transporter GLUT8 cycles between intracellular vesicles and the plasma membrane. Like the insulin-responsive glucose transporter GLUT4, GLUT8 is primarily located in intracellular compartments under basal conditions. Whereas translocation of GLUT4 to the plasma membrane is stimulated by insulin, the distribution of GLUT8 is not affected by insulin treatment in adipose cells. However, blocking endocytosis by co-expression of a dominant-negative dynamin GTPase (K44A) or mutation of the N-terminal dileucine (LL(12/13)) motif in GLUT8 leads to accumulation of the glucose transporter at the cell surface in a variety of different cell types. Yeast two-hybrid analyses and GST pulldown assays reveal that the LL signal constitutes a binding site for the beta2-adaptin subunit of the heterotetrameric AP-2 adaptor complex, implicating this motif in targeting of GLUT8 to clathrin-coated vesicles. Moreover, yeast two-hybrid assays provide evidence that the binding site for the LL motif maps to the appendage domain of beta2-adaptin. To analyze the biological significance of the LL/beta2 interaction, we utilized RNA interference to specifically knockdown AP-2. Our results show that RNAi-mediated targeting of the mu2 subunit leads to cellular depletion of AP-2, but not AP-1 adaptor complexes in HeLa cells. As a consequence, GLUT8 accumulates at the plasma membrane at comparable levels to those observed in K44A-transfected cells. Conversely, the intracellular localization of mutant GLUT8-LL/AA is restored by replacing the LL motif in GLUT8 with the transferrin receptor-derived mu2-adaptin binding motif YTRF, indicating that for endocytosis both AP-2 binding motifs can substitute for each other. Thus, our data demonstrate that recruitment of GLUT8 to the endocytic machinery occurs via direct interaction of the dileucine motif with beta2-adaptin, and that endocytosis might be the main site at which GLUT8 is likely to be regulated.