Influence of lighting environment on social preferences in sticklebacks from two different photic habitats. II. Shoaling and mate preferences of lab-bred fishes.

Different environmental conditions may lead to diverse morphological, behavioral, and physiological adaptations of different populations of the same species. Lighting conditions, for example, vary vastly especially between aquatic habitats, and have been shown to elicit adaptations. The availability of short-wave ultraviolet (UV) light is especially fluctuating, as UV wavelengths are attenuated strongly depending on water properties. The island of North Uist, Scotland, comprises 2 differential habitat types, tea-stained and clear-water lakes, varying considerably in UV transmission. In previous studies, wild-caught 3-spined stickleback Gasterosteus aculeatus populations (3 populations of each habitat type) were tested with respect to their shoaling and mate preferences for fish viewed under UV-present and UV-absent conditions. The results revealed a habitat-dependent preference of UV cues during shoal choice (tea-stained populations: preference for UV-absent condition in tea-stained water; clear-water populations: no preference in clear-water) but an overall preference for UV-present conditions during mate choice. To assess genetic influences on these behavioral patterns, similar experiments were conducted with lab-bred F1-generations of the same stickleback populations that were raised in a common environment (i.e. standardized clear-water conditions). Offspring of sticklebacks from tea-stained lakes tended to prefer shoals viewed under UV-absent conditions (only in tea-stained water), while sticklebacks from clear-water lakes showed a significant preference for the shoal viewed under UV-present conditions in clear-water but not in tea-stained water. Mate-preference experiments demonstrated that females from the tea-stained lakes significantly preferred and females from the clear-water lakes preferred by trend the male viewed under UV-present conditions in the clear-water treatment. The results for both shoaling- and mate-preference tests were largely similar for wild-caught and lab-bred sticklebacks, thus hinting at a genetic basis for the preference patterns.

Influence of lighting environment on social preferences in sticklebacks from two different photic habitats. I. mate preferences of wild-caught females.

Ultraviolet (UV) A signals (320-400 nm) are important in mate choice in numerous species. The sensitivity for UV signals is not only assumed to be costly, but also expected to be a function of the prevailing ecological conditions. Generally, those signals are favored by selection that efficiently reach the receiver. A decisive factor for color signaling is the lighting environment, especially in aquatic habitats, as the visibility of signals, and thus costs and benefits, are instantaneously influenced by it. Although ecological aspects of color signal evolution are relatively well-studied, there is little data on specific effects of environmental UV-light conditions on signaling at these shorter wavelengths. We studied wild-caught gravid female 3-spined sticklebacks Gasterosteus aculeatus of 2 photic habitat types (tea-stained and clear-water lakes), possessing great variation in their UV transmission. In 2 treatments, tea-stained and clear-water, preferences for males viewed under UV-present (UV+) and UV-absent (UV-) conditions were tested. A preference for males under UV+ conditions was found for females from both habitat types, thus stressing the significance of UV signals in stickleback's mate choice decisions. However, females from both habitat types showed the most pronounced preferences for males under UV+ conditions under clear-water test conditions. Moreover, reflectance measurements revealed that the carotenoid-based orange-red breeding coloration in wild-caught males of both habitat types differed significantly in color intensity (higher in clear-water males) and hue (more red shifted in clear-water males) while no significant differences in UV coloration were found. The differential reflection patterns in longer wavelengths suggest that sticklebacks of both habitat types have adapted to the respective water conditions. Adaptations of UV signals in a sexual context to ambient light conditions in both behavior and coloration seem less evident.

Nest-site selection in a fish species with paternal care.

Fish that perform paternal care may increase their fitness by choosing nest sites that enhance survival and development of embryos. We studied nest-site choice with respect to dissolved oxygen concentration and water temperature in males of the three-spined stickleback (Gasterosteus aculeatus), a small fish species with exclusive male parental care that usually breeds in the littoral zone of freshwaters of the Northern hemisphere. Fathers oxygenate the embryos by fanning movements of their pectoral fins. We expected choice for conditions at potential nest sites that would benefit offspring development, i.e., higher temperature and higher dissolved oxygen concentration. In the laboratory, we offered males a choice between two potential nest sites that differed in dissolved oxygen concentration or water temperature. Males preferred to build a nest at sites with a higher dissolved oxygen level or higher temperature and thus chose sites that would promote embryo development.

Anticipatory plastic response of the cellular immune system in the face of future injury: chronic high perceived predation risk induces lymphocytosis in a cichlid fish.

Vertebrate cellular immunity displays substantial variation among taxa and environments. Hematological parameters such as white blood-cell counts have emerged as a valuable tool to understand this variation by assessing the immunological status of individuals. These tools have long revealed that vertebrate cellular immune systems are highly plastic and respond to injury and infection. However, cellular immune systems may also be able to anticipate a high risk of injury from environmental cues (e.g., predation-related cues) and respond plastically ahead of time. We studied white blood-cell (leukocyte) profiles in African cichlids Pelvicachromis taeniatus that were raised for 4 years under different levels of perceived predation risk. In a split-clutch design, we raised fish from hatching onwards under chronic exposure to either conspecific alarm cues (communicating high predation risk) or a distilled water control treatment. Differential blood analysis revealed that alarm cue-exposed fish had twice as many lymphocytes in peripheral blood as did controls, a condition called lymphocytosis. The presence of a higher number of lymphocytes makes the cellular immune response more potent, which accelerates the removal of invading foreign antigens from the bloodstream, and, therefore, may be putatively beneficial in the face of injury. This observed lymphocytosis after long-term exposure to conspecific alarm cues constitutes first evidence for an anticipatory and adaptive plastic response of the cellular immune system to future immunological challenges.

Differential investment in pre- and post-mating male sexual traits in response to prolonged exposure to ambient UVB radiation in a fish.

Increasing UVB radiation (UVB) reaching earth's surface following stratospheric ozone depletion is linked to serious consequences for organisms. While studies have focused on direct cytocidal and immunomodulatory effects of UVB, indirect consequences for fitness-related life-history traits are largely unexplored, although knowledge is needed to understand organismal responses to climate change. The present study investigates the effects of developmental exposure to enhanced, but ecologically relevant, ambient UVB levels on growth (body size), parental behavior (nest-building), fitness-enhancing traits of pre-mating (sexual ornamentation) and post-mating (sperm traits) sexual selection as well as their interrelationships in male three-spined sticklebacks (Gasterosteus aculeatus). Moreover, potentially underlying direct UVB effects, testicular antioxidant capacity (TAC) and testes and skin melanization were quantified. Juvenile full-siblings were split into two groups and raised until adulthood in a semi-natural set-up under light conditions including either natural (UVBnormal) or elevated UVB levels (UVBenhanced). When becoming reproductive, males were kept individually before data collection took place at their reproductive peak. The results showed that males from the UVBenhanced-group were smaller than their brothers from the UVBnormal-treatment, whereas no treatment differences were observed for sexual ornamentation, sperm traits and TAC. Moreover, UVB-stressed males built their nests faster and the relationship between body size and nest size was negative for UVBenhanced-males while being positive for UVBnormal-males. These results demonstrate that, depending on physical state, UVB-stressed males adjusted their behavior to some extent. Additionally, a trade-off between the investment in pre- and post-mating traits was found for UVBenhanced-males, i.e. the intensity of their breeding coloration was negatively correlated with sperm number whereas this relationship was reversed for UVBnormal-males, thus showing an interaction between pre- and post-mating traits regarding exposure treatment. The interaction provides first experimental evidence that differential allocation to energetically demanding pre- and post-mating components of male fitness is triggered by a key environmental stressor of climate change.

Impact of social rearing-environment on performance in a complex maze in females of a cichlid fish.

Spatial orientation is an important skill as it improves, for example, foraging, localisation of recourses, predator avoidance or navigation. Habitat complexity positively affects spatial abilities in various fish species with a more complex environment promoting learning ability. However, to what extent a complex social environment affects cognitive abilities in fishes has received less attention. Here, we investigated differences in maze performance of adult females of the West African cichlid fish Pelvicachromis taeniatus, which had been reared and maintained either in a group or in isolation from an early age on. Fish had to master the route through a maze in order to gain a food reward. Our results indicate marked differences in performance contingent upon social rearing-environment: isolation fish ran successful trials (i.e. locating the food reward) significantly more often than group fish and were faster during trials, also in a reversed maze. However, the number of mistakes did not differ between isolation and group fish and the time needed to relocate the food reward did not diminish with elapsed training days. In a second experiment, the activity of group and isolation fish was analysed in an open field test. Here, isolation fish were less active than group fish. We discuss different possibilities for performance differences of group and isolation fish including enhanced cognitive abilities of isolation fish, motivational/emotional differences and hyperactivity.

Mate assessment behavior is correlated to learning ability in female threespine sticklebacks.

In many species, males signal quality with elaborate traits, but females often show inter-individual variation in preference for these traits. Choosing a mate requires multiple cognitive steps; therefore, cognitive style (how an individual processes information) likely influences the perception of sexual signals and ability to choose a high-quality mate. An important component of cognitive style is flexibility; cognitively flexible individuals are more perceptive to shifts in cues. We hypothesized that cognitively flexible individuals would acquire more information about potential mates, better discern between two quality-signaling traits, and thus be more discriminatory. Here, we show that mate assessment is correlated to other cognitive traits. Although we did not detect an effect of cognitive style on mate preference or discrimination, we found that female threespine sticklebacks Gasterosteus aculeatus that spent more time assessing potential mates (more responsive) in a dichotomous mate choice task reached both the initial and reversal learning criterion in a spatial learning task with fewer errors. However, these highly responsive females made more consecutive mistakes immediately at the beginning of the reversal phase, suggesting that they did not quickly adapt to the environmental change but instead rapidly formed strict routines during the learning task that were eventually reversible after repeated errors. Furthermore, we found evidence for condition-dependent mate preference, with larger females preferring the high-quality male. These are among the first results that illustrate how cognitive traits might influence mate choice, which has implications for the strength and direction of sexual selection.

Sperm size is negatively related to relative testis size in West African riverine cichlid fishes.

Fishes show a great diversity of mating systems and fertilization mechanisms. This diversity creates an enormous potential for sperm competition. Typically, monogamous species face a low risk of sperm competition and invest less into sperm, and thus show smaller relative testis mass compared to polygamous species with high sperm competition. In cichlids, sperm competition risk is very variable. In lacustrine East African cichlids, large sperm are interpreted as an adaptation to sperm competition, as in those species sperm length correlates with sperm swimming speed. The aim of the present study was to examine variation in sperm and testis traits of substrate breeding cichlids from West African river systems and its relationship to sperm competition. Therefore, sperm traits (total sperm size, flagellum-, midpiece-, and head size) and sperm number were related to the gonadosomatic index (GSI), an indicator of sperm competition, in eight species of two large informal tribes, the chromidotilapiines and the haplotilapiines. We found significant differences between species in all examined sperm traits, sperm number, and GSI with pronounced differences between chromidotilapiines and haplotilapiines. We used a generalized least-squares approach to control for non-independence of data. GSI was positively correlated with sperm number but negatively correlated with total sperm size (also negatively with the flagellum and head size but not significantly with midpiece size). Sperm number and sperm size were negatively correlated suggesting a trade-off between sperm size and quality. Our results suggest that large sperm can evolve in species with relatively low sperm expenditure and probably in absence of high sperm competition between males.

Enhanced ambient UVB radiation affects post-mating, but not pre-mating sexual traits in a fish.

Organisms inhabiting shallow aquatic habitats currently experience increasing levels of solar ultraviolet B radiation (UVB). UVB causes damage on cellular and molecular levels and can affect associated life-history traits either through direct exposure or indirectly through oxidative stress generation. We examined UVB effects on pre- and post-mating sexual traits in three-spined stickleback fish (Gasterosteus aculeatus). Adult, reproductively non-active males were assigned to two exposure treatments under semi-natural conditions in an outdoor experiment; one group received natural radiation (UVBnormal) whilst the other group received additional UVB (UVBenhanced). After two months, colour metrics were used to quantify male breeding colouration as pre-mating trait. At the post-mating stage, sperm morphology, number and movement as well as testes mass were determined. Males did not significantly differ in sexual ornamentation between treatments, but UVBenhanced fish had smaller testes as well as fewer and shorter sperm than UVBnormal fish. Sperm movement was not significantly different between treatments. However, in UVBenhanced males, linear and progressive movement of sperm was positively correlated with sperm morphology (head-to-tail length ratio), whereas in UVBnormal males this relationship was negative (but not significant). Additionally, there was a significant treatment by body condition interaction concerning head-to-tail length ratio, i.e. head-to-tail length ratio increased with condition in UVBnormal males whereas there was no relationship in UVBenhanced fish. Our findings reveal that increased UVB levels influence post-mating fitness-relevant traits in males whilst having no significant impact on pre-mating sexual traits, suggesting selective UVB-effects at the gamete level with consequences for reproductive performance.

Shoal sex composition and predation risk influence sub-adult threespine stickleback shoaling decisions.

Group living reduces individual predation risk most effectively when group members are behaviorally and phenotypically similar. Group preferences are influenced by the individual, the members of the shoal, and the environmental conditions. While shoaling behavior has been studied extensively in the threespine stickleback (Gasterosteus aculeatus), it is unclear whether the sex of shoal mates influences the shoal preference of non-reproductive males and females and how this changes under increasing predation risk. Although non-reproductively active sticklebacks are sexually monochromatic in appearance, sex-related differences may result in sexual segregation when shoaling. Here we show that male and female sub-adult threespine sticklebacks had contrasting preferences for shoal mate sex, and that this preference was dependent on the level of predation risk during standardized experimental choice tests. In detail, test fish shoal with the opposite sex within low predation risk trials and with same-sex shoals within high predation risk trials. This difference might be linked to activity patterns; test males were more active than females. Our results demonstrate that differences between the sexes in a species with a sexually monochromatic non-reproductive stage can result in sex-related shoaling preferences. Most studies examining sexual segregation focus on sexually dimorphic species, but these results highlight the potentially widespread occurrence of sexual segregation beyond the sexually dimorphic reproductive stage.

Neglected Patterns of Variation in Phenotypic Plasticity: Age- and Sex-Specific Antipredator Plasticity in a Cichlid Fish.

The ability of organisms to plastically respond to changing environments is well studied. However, variation in phenotypic plasticity during ontogeny is less well understood despite its relevance of being an important source of phenotypic variation in nature. Here, we comprehensively study ontogenetic variation in morphological antipredator plasticity across multiple traits in Pelvicachromis taeniatus, a western African cichlid fish with sexually dimorphic ornamentation. In a split-clutch design, fish were raised under different levels of perceived predation risk (conspecific alarm cues or distilled water). Morphological plasticity varied substantially across ontogeny: it was first observable at an early juvenile stage where alarm cue-exposed fish grew faster. Subsequently, significant plasticity was absent until the onset of sexual maturity. Here, alarm cue-exposed males were larger than control males, which led to deeper bodies, longer dorsal spines, larger caudal peduncles, and increased eye diameters. Sexual ornamentation emerged delayed in alarm cue-exposed males. In later adulthood, the plastic responses receded. Despite small effect sizes, these responses represent putative adaptive plasticity, as they are likely to reduce predation risk. In females, we did not observe any plasticity. In accordance with theory, these results suggest fine-tuned expression of plasticity that potentially increases defenses during vulnerable developmental stages and reproductive output.

Long-term UVB exposure promotes predator-inspection behaviour in a fish.

Ultraviolet-B radiation (UVB) reaching the earth's surface has increased due to human-caused stratospheric ozone depletion. Whereas the harmful effects of UVB on aquatic organisms are well studied at the molecular and cellular level, recent studies have also begun to address behavioural changes caused by sublethal amounts of UVB. However, the behavioural consequences of long-term exposure to ecologically relevant UVB levels over several life stages are virtually unknown, particularly with regard to predator-prey behaviour. We found increased predator-inspection behaviour together with a smaller body length in three-spined sticklebacks (Gasterosteus aculeatus) after fish were exposed for about seven months to natural sunlight conditions with enhanced UVB, compared with full siblings exposed to natural sunlight only. The observed change in antipredator behaviour may reflect a direct behavioural response mediated through UVB-induced oxidative stress during development. Alternatively, the smaller body size in UVB-exposed fish may result in an increased inspection effort allowing them to spend more time foraging. Our findings suggest that, within the scope of environmental change, UVB radiation constitutes an important stress factor by eliciting behavioural responses that influence crucial ecological processes, such as predator-prey interactions.

Rapid molecular sexing of three-spined sticklebacks, Gasterosteus aculeatus L., based on large Y-chromosomal insertions.

There is a need for rapid and reliable molecular sexing of three-spined sticklebacks, Gasterosteus aculeatus, the supermodel species for evolutionary biology. A DNA region at the 5' end of the sex-linked microsatellite Gac4202 was sequenced for the X chromosome of six females and the Y chromosome of five males from three populations. The Y chromosome contained two large insertions, which did not recombine with the phenotype of sex in a cross of 322 individuals. Genetic variation (SNPs and indels) within the insertions was smaller than on flanking DNA sequences. Three molecular PCR-based sex tests were developed, in which the first, the second or both insertions were covered. In five European populations (from DE, CH, NL, GB) of three-spined sticklebacks, tests with both insertions combined showed two clearly separated bands on agarose minigels in males and one band in females. The tests with the separate insertions gave similar results. Thus, the new molecular sexing method gave rapid and reliable results for sexing three-spined sticklebacks and is an improvement and/or alternative to existing methods.

Strong neutral genetic differentiation in a host, but not in its parasite.

The genetic diversity and population structure of a parasite with a complex life cycle generally depends on the dispersal by its most motile host. Given that high gene flow is assumed to hinder local adaptation, this can impose significant constraints on a parasite's potential to adapt to local environmental conditions, intermediate host populations, and ultimately to host-parasite coevolution. Here, we aimed to examine the population genetic basis for local host-parasite interactions between the eye fluke Diplostomum lineage 6, a digenean trematode with a multi-host life cycle (including a snail, a fish, and a bird) and its second intermediate host, the three-spined stickleback Gasterosteus aculeatus L. We developed the first microsatellite primers for D. lineage 6 and used them together with published stickleback markers to analyse host and parasite population structures in 19 freshwater lakes, which differ in their local environmental characteristics regarding water chemistry and Diplostomum abundance. Our analyses suggest that one parasite population successfully infects a range of genetically differentiated stickleback populations. The lack of neutral genetic differentiation in D. lineage 6, which could be attributed to the motility of the parasite's definitive host as well as its life cycle characteristics, makes local host-parasite co-adaptations seem more likely on a larger geographical scale than among the lakes of our study site. Our study provides a suitable background for future studies in this system and the first microsatellite primers for a widespread fish parasite.

Distribution of common stickleback parasites on North Uist, Scotland, in relation to ecology and host traits.

Analysing spatial differences among macroparasite communities is an important tool in the study of host-parasite interactions. Identifying patterns can shed light on the underlying causes of heterogeneity of parasite distribution and help to better understand ecological constraints and the relative importance of host and parasite adaptations. In the present study, we aimed to find correlational evidence that the macroparasite distribution patterns on the Scottish island of North Uist, which had been described by de Roij and MacColl (2012), are indicative of local processes rather than an unspecific influence of habitat characteristics. We therefore reinvestigated parasite abundances and tested for associations with habitat characteristics and host traits. Distribution patterns of the most common parasites were largely consistent with the observations of de Roij and MacColl (2012). In accordance with the published results, we found that the most obvious abiotic habitat characteristic varying among the lakes on the island, pH, did not statistically explain parasite abundances (except for eye fluke species inside the lens). Instead, we found that genetic differentiation between host populations, measured as pairwise FST values based on available microsatellite data, was significantly correlated with dissimilarity in parasite community composition. Our results indicate that individual lake characteristics rather than physicochemical variables shape parasite distribution on this island, making it an ideal place to study host-parasite interactions. Furthermore, additionally to geographic distance measures taken from maps, we suggest taking into account connectivity among freshwater habitats, indirectly measured via fish population structure, to analyse spatial distribution patterns.

Predator-induced neophobia in juvenile cichlids.

Predation is an important but often fluctuating selection factor for prey animals. Accordingly, individuals plastically adopt antipredator strategies in response to current predation risk. Recently, it was proposed that predation risk also plastically induces neophobia (an antipredator response towards novel cues). Previous studies, however, do not allow a differentiation between general neophobia and sensory channel-specific neophobic responses. Therefore, we tested the neophobia hypothesis focusing on adjustment in shoaling behavior in response to a novel cue addressing a different sensory channel than the one from which predation risk was initially perceived. From hatching onwards, juveniles of the cichlid Pelvicachromis taeniatus were exposed to different chemical cues in a split-clutch design: conspecific alarm cues which signal predation risk and heterospecific alarm cues or distilled water as controls. At 2 months of age, their shoaling behavior was examined prior and subsequent to a tactical disturbance cue. We found that fish previously exposed to predation risk formed more compact shoals relative to the control groups in response to the novel disturbance cue. Moreover, the relationship between shoal density and shoal homogeneity was also affected by experienced predation risk. Our findings indicate predator-induced, increased cross-sensory sensitivity towards novel cues making neophobia an effective antipredator mechanism.

The dynamics of color signals in male threespine sticklebacks Gasterosteus aculeatus.

Body coloration and color patterns are ubiquitous throughout the animal kingdom and vary between and within species. Recent studies have dealt with individual dynamics of various aspects of coloration, as it is in many cases a flexible trait and changes in color expression may be context-dependent. During the reproductive phase, temporal changes of coloration in the visible spectral range (400-700 nm) have been shown for many animals but corresponding changes in the ultraviolet (UV) waveband (300-400 nm) have rarely been studied. Threespine stickleback Gasterosteus aculeatus males develop conspicuous orange-red breeding coloration combined with UV reflectance in the cheek region. We investigated dynamics of color patterns including UV throughout a male breeding cycle, as well as short-term changes in coloration in response to a computer-animated rival using reflectance spectrophotometry and visual modeling, to estimate how colors would be perceived by conspecifics. We found the orange-red component of coloration to vary during the breeding cycle with respect to hue (theta/R50) and intensity (achieved chroma/red chroma). Furthermore, color intensity in the orange-red spectral part (achieved chroma) tended to be increased after the presentation of an artificial rival. Dynamic changes in specific measures of hue and intensity in the UV waveband were not found. In general, the orange-red component of the signal seems to be dynamic with respect to color intensity and hue. This accounts in particular for color changes during the breeding cycle, presumably to signal reproductive status, and with limitations as well in the intrasexual context, most likely to signal dominance or inferiority.

Context-dependent dynamic UV signaling in female threespine sticklebacks.

Color signals, including ultraviolet (UV) signals, are widespread throughout the animal kingdom and color changes can be influenced by reproductive and motivational state. However, studies on dynamic changes of UV signals are scarce. Three spine sticklebacks (Gasterosteus aculeatus) that show intraspecific UV communication were used to study dynamic UV signaling in females. Reflectance measurements were taken from the distended abdomen, which serves as signal of female fecundity and readiness to spawn for courting males, and the melanized dorsal region. Scans were taken during egg maturation as well as before and after stimulation with a male to investigate context-dependent color changes. We used a physiological model of vision to determine how females might be perceived by conspecifics and quantified chromatic contrasts among both body regions and between body regions and the background for all stages. Females showed a significant increase in abdominal UV intensity during egg maturation and in response to a courting male. Measures of chromatic contrast among body regions (abdomen vs. dorsal region) and against the background (abdomen vs. background) were also increased during egg maturation and in response to the male stimulus (abdomen vs. background). Our results provide evidence for dynamic UV signaling in females in a reproductive context.