Female Sticklebacks Research Articles

Detection of environmental androgens: A novel method based on enzyme‐linked immunosorbent assay of spiggin, the stickleback (Gasterosteus aculeatus) glue protein

We report the development and validation of a novel in vivo biomarker test for waterborne androgens. During breeding, male sticklebacks (Gasterosteus aculeatus) manufacture a glue protein, spiggin, in their kidneys that they use to build their nests. Spiggin production is under the control of androgens. Until now, however, it has only been possible to quantify its production by measurement of the height of kidney epithelial cells. In the present study, we report the development of an enzyme-linked immunosorbent assay (ELISA) for spiggin and demonstrate its application to the measurement of spiggin in the kidneys of female sticklebacks that have been exposed to androgens in water. Results from the ELISA procedure revealed a strong correlation with measurement of kidney epithelial cell height (r2 = 0.93). However, the ELISA was much quicker and had a considerably higher response range (100,000-fold vs fourfold). Clear, graded responses in spiggin production were obtained by exposing intact females to increasing concentrations of 17a-methyltestosterone and 5alpha-dihydrotestosterone over three-week test periods. The lowest effective concentrations for these two steroids were 100 ng/L and 3 micorg/L, respectively. Female sticklebacks that were exposed to pulp mill effluent also produced spiggin in their kidneys. Possession of an androgen-regulated protein by the female stickleback makes it a unique bioassay organism for detecting androgenic contamination in the aquatic environment.

Do male sticklebacks prefer females with red ornamentation?

In general, females are less ornamented than males and until recently female ornaments have been regarded as non-adaptive correlated effects of selection on males. This view is challenged and the alternative hypothesis that females' ornaments have evolved independently of male showiness, by intersexual selection for example, is supported. I tested these hypotheses experimentally using threespine sticklebacks (Gasterosteus aculeatus) from a population in which both sexes have red pelvic spines. Two females differing in red spine colour extravagance were presented simultaneously to a male under white and green light, and the male's courtship activity towards each female was quantified. Green light prevented the use of red cues by sticklebacks. The results show that red colour on the pelvic spines of female sticklebacks has value as a signal to males in this population, and males actually courted females with drab pelvic spines more than females whose pelvic spines had a redder hue, but only when illuminated by white light. The interpretation that best accounts for this result is that spine colour has a strong function in male–male aggressive interactions. This interpretation favours the hypothesis of non-adaptive correlation that has been proposed to explain the evolution of red spines in female sticklebacks.

Male preference for a subtle posture cue that signals spawning readiness in female sticklebacks

We presented territorial male three-spined sticklebacks, Gasterosteus aculeatus, two stationary dummies simultaneously, each simulating an equally gravid female. Both dummies featured the head-up orientation indicative of a sexually receptive female but differed in more subtle aspects of posture. In one dummy the back and tail were moulded in a relaxed straight posture and the other was moulded with the raised tail and concave back posture (lordosis), which mating tests revealed to be a strong predictor of spawning readiness in females. Males courted both dummies but directed more to the lordosis posture. Male sticklebacks thus recognize and respond appropriately to relatively subtle features of female posture in addition to other cues such as the head-up angle and dorsal barring reported in earlier studies. Physiological or morphological constraints may make the lordosis posture a reliable, and hence relatively honest, indicator of spawning readiness in females. Males may therefore enhance their fitness by preferentially courting females that assume this posture. Copyright 2002 The Association for the Study of Animal Behaviour. Published by Elsevier Science Ltd. All rights reserved.

Modelling the evolution of female choice strategies under inbreeding conditions.

Recently, many mate choice studies have discussed the role of genetic compatibility and inbreeding for the evolution of mate choice. With population genetic simulations I compared the potential advantage of three different female choice strategies under inbreeding conditions. Females were assumed to benefit indirectly via a preference for (i) complementary males, (ii) males with few detrimental mutations, and (iii) non-inbred males. Probably related to the reduced inbreeding depression in offspring of choosy females, the choice-allele increased for all three strategies. However, the advantage of the strategies differed widely. Choice of males with fewer mutations provided a comparatively large advantage, choice of complementary males led to a reasonable advantage, and choice of non-inbred males only resulted in a minor advantage of female choice. My results show that complementary mate choice can be almost as beneficial as conventional good-genes choice of mates with lower genetic load. Compared to the two other mate choice strategies, choice of non-inbred males is less likely to contribute to the evolution of costly mate choice. The results of a recent study showing that female sticklebacks prefer males with a larger number of MHC-loci is thus unlikely to be related to an indirect benefit of choosing non-inbred males.

Female preferences for single and combined traits in computer animated stickleback males

In many animal species, males have more than one secondary sexual character. Apart from theoretical considerations about the evolution of multiple traits, there are almost no empirical studies on female mate choice decisions based on combinations of traits as opposed to decisions based on single traits. Because three-spined sticklebacks are exceptionally well suited to be tested with computer animation technique, which itself fills the gap of adequate test paradigms for multiple traits, we tested female sticklebacks for their preferences for both single and combined male traits. We used virtual stickleback males that differed either in red throat coloration, courtship intensity, body size, or in combinations of these traits. The virtual male with increased redness was found to be preferred by females, whereas the male courting more intensely was not. The tests for combinations of traits revealed the more pronounced female preferences, the more traits were available to the females to judge male quality. Key words: computer animation, courtship, redness, female mate preference, Gasterosteus aculeatus, multiple traits. [Behav Ecol 12:681–685 (2001)]

Female sticklebacks count alleles in a strategy of sexual selection explaining MHC polymorphism.

The origin and maintenance of polymorphism in major histocompatibility complex (MHC) genes in natural populations is still unresolved. Sexual selection, frequency-dependent selection by parasites and pathogens, and heterozygote advantage have been suggested to explain the maintenance of high allele diversity at MHC genes. Here we argue that there are two (non-exclusive) strategies for MHC-related sexual selection, representing solutions to two different problems: inbreeding avoidance and parasite resistance. In species prone to inadvertent inbreeding, partners should prefer dissimilar MHC genotypes to similar ones. But if the goal is to maximize the resistance of offspring towards potential infections, the choosing sex should prefer mates with a higher diversity of MHC alleles. This latter strategy should apply when there are several MHC loci, as is the case in most vertebrates. We tested the relative importance of an 'allele counting' strategy compared to a disassortative mating strategy using wild-caught three-spined sticklebacks (Gasterosteus aculeatus) from an interconnected system of lakes. Here we show that gravid female fish preferred the odour of males with a large number of MHC class-IIB alleles to that of males with fewer alleles. Females did not prefer male genotypes dissimilar to their own.

Effects ofSchistocephalus solidusinfection on brain monoaminergic activity in female three-spined sticklebacksGasterosteus aculeatus

The three-spined stickleback Gasterosteus aculeatus is an intermediate host of the tapeworm Schistocephalus solidus. Changes in predator avoidance, foraging and shoaling behaviour have been reported in sticklebacks infested with S. solidus, but the mechanisms underlying parasite-induced behavioural changes are not understood. Monoamine neurotransmitters are involved in the control of behaviour and central monoaminergic systems are sensitive to various stressors. Thus, the behavioural effects of S. solidus infestation might be a reflection of changes in brain monoaminergic activity in the stickleback host. The concentrations of 5-hydroxytryptamine (5-HT), dopamine (DA), norepinephrine (NE) and their metabolites 5-hydroxy-indoleacetic acid (5-HIAA), homovanilic acid (HVA) and 3-methoxy-4-hydroxyphenylglycol (MHPG) were measured in the telencephalons, hypothalami and brainstems of parasitized and non-parasitized female sticklebacks held in the laboratory. The ratios of 5-HIAA:5-HT were significantly elevated in both the hypothalami and brainstems of infected sticklebacks. The concentrations of 5-HT and NE were significantly reduced in the telencephalons of infected fish as compared with controls, but there was no elevation of metabolite concentrations. The results are consistent with chronic stress in infected fish, but may also reflect other alterations of neuroendocrine status resulting from parasite infection.

Hormones and endocrine disrupters in aquatic environment

The UK's programme on endocrine disruption in the marine environment (EDMAR) was set up in 1998 to investigate in more detail the implications of earlier observations of strongly oestrogenic effects in flounder (Platichthys flesus) from several UK estuaries. Much of the work has focused on fish, including further studies of flounder, as well as investigations of sand gobies (Pomatoschistus minutus and P. lozanoi) and viviparous blennies (Zoarces viviparous). Some work has also begun to investigate androgenic effects in the stickleback (Gasterosteus aculeatus). There have in addition been detailed chemical studies to identify the sources and types of oestrogenic substances. The additional work with flounder has confirmed the original findings that many industrialised UK estuaries are sufficiently oestrogenic to cause strong induction of yolk protein (vitellogenin or VTG) in male fish (by up to a million‐fold in excess of controls), as well as induction of intersex (ovotestis) in some male individuals (15–20% in some areas). Although the levels of activity at some sites have not changed significantly over 3 years, VTG titres have decreased at other sites, perhaps due to improvements in sewage treatment, although these findings need confirmation over a longer timespan. Experiments with male flounder showed that they did not produce VTG when caged in oestrogen‐contaminated estuaries, but experienced mild VTG induction when fed on mussels (Mytilus edulis) which had been held in an oestrogen‐contaminated estuary (the Tees) for 3 months. This shows that at least some oestrogenic substances have the potential for transmission through the food chain. Studies of the viviparous blenny have shown that, like in flounder, males are producing VTG (measured as VTG mRNA) in some estuaries, and up to 25% of male fish are showing intersex (ovotestis). Studies are under way to investigate whether this is affecting the production of fry in this species. Viviparous blennies are ideal for such work because the fry produced by each individual wild female can be directly counted and measured. Sand gobies from oestrogen‐contaminated estuaries did not show either induction of VTG mRNA or intersex, even though the former could be induced in the laboratory through exposure to reference oestrogens. However, high proportions of male gobies from oestrogen‐contaminated locations such as the Mersey and Tees showed a condition of the urogenital papilla (UP) in which the terminal part of this organ had grown villae resembling those seen on female papillae. The UP is used by both sexes to deposit gametes, and it is possible that the apparent feminisation seen in oestrogen‐ exposed males could interfere with normal reproduction. Experiments are in progress to attempt to replicate the condition (known as Male Intermediate Papilla Syndrome ‐ MIPS) in oestrogen‐exposed fish. The EDMAR programme has developed a biomarker of androgenic exposure, the glue protein spiggi, which is induced in female sticklebacks when they are exposed to exogenous androgens. Preliminary work has shown that there is only limited androgenic activity induced in female fish held in the Tees. However, a range of natural androgens has been detected in certain sewage discharges. Work is also progressing to investigate the possible effects of estuarine endocrine disrupters on smoltification in salmon. Finally, the yeast oestrogen screen (YES) has been used in a Toxicity Identification and Evaluation (TIE) scheme to identify substances probably causing these effects. The majority of oestrogenic activity in sewage effluents and estuarine waters has been attributable to oestradiol, although a small proportion is being caused by other natural steroids, as well as synthetic substances such as nonylphenol. However, the overwhelming majority of the activity in the investigated estuaries (Tees and Tyne) is to be found in the sediments. Most of the sediment‐bound activity remains to be identified, but little appears to be attributable to oestradiol. Although the EDMAR programme will only reach completion at the end of 2001, it is already apparent that oestrogenic effects are very widespread in 4 fish species from several industrialised UK estuaries. It appears that the sediments can become heavily contaminated with certain oestrogens, and it seems likely that fish become contaminated at least partly via consumption of benthic invertebrates. However, the full ecological implications of the effects observed in fish have not yet been fully elucidated.

Melatonin and Maturation Pace in Female Three-Spined Stickleback, Gasterosteus aculeatus

In many animals sexual maturation is controlled by the photoperiod. In mammals, the photoperiodic message is mediated via melatonin, but it is unclear whether this also applies to fishes. Administration of melatonin via the water on a schedule aimed at mimicking a short nonstimulatory photoperiod cycle has been found not to inhibit maturation in male three-spined sticklebacks, Gasterosteus aculeatus, kept under stimulatory long photoperiods. To study whether melatonin affects maturation pace, adult female sticklebacks kept under stimulatory photoperiodic regimes of Light:Dark (LD) 24:0 h (Exp. 1) or LD 16:8 h (Exp. 2) and 18° were treated with melatonin (0, 20, or 80 μg/L water) via the water for 16 h per day. In addition, females were also kept under a nonstimulatory short photoperiod (LD 8:16). The time at which full maturation was achieved (running roe) was noted and the ovaries of nonovulated fish were studied histologically. Most fish under LD 24:0 and LD 16:8 matured (maturation rates in Exp. 1 and 2, respectively: control, 100 and 86%; low-dose melatonin, 83 and 93%; high-dose melatonin, 90 and 75%), whereas almost all females kept under LD 8:16 remained immature (maturation rates: 0% in Exp. 1 and 3% in Exp. 2). There was no difference in maturation pace, proportion of fish maturing, or relative ovarian weights between controls and melatonin-treated fish kept under LD 16:8. Furthermore, there was no difference in proportion of fish maturing or relative ovarian weights between controls and melatonin-treated fish kept under LD 24:0. However, LD 24:0 controls matured significantly earlier than fish receiving the high melatonin dose. Thus, there was an inhibitory effect of the high melatonin dose on maturation pace under LD 24:0. Nevertheless, this effect was small compared to the inhibitory effect of LD 8:16 treatment, suggesting that at least in this season, the major part of the photoperiodic effects in the stickleback is mediated via mechanisms other than circulating melatonin.

Are nest characters of importance when choosing a male in the fifteen-spined stickleback ( Spinachia spinachia )?

Male fifteen-spined sticklebacks (Spinachia spinachia) build their nests by wrapping epiphytic algae around macroalgae and securing them with secretional threads. In this study, I show that female fifteen-spined stickleback prefer males which have built their nests above the surrounding vegetation. High-located nests are safer, because they are less likely to attract egg-predators such as shore crabs (Carcinus maenas). Thus, I found that shore crabs more rapidly find eggs in nests built close to the bottom than in nests well above the it. Moreover, male-male competition could be an additional explanation as to why males build nests high up. Thus, my field results suggest that the closer the males were to a neighbour, the higher they built. Larger males were also found to have larger territories but male size did not correlate with the height of the nest above the surrounding vegetation. Females showed no preference for nest size, regardless of the presence of egg-predators. However, they tended to choose nests that had no egg-predators in the vicinity over nests with egg-predators close by. Female choosiness for certain nest characters may provide one explanation for the evolution of male care in this species.

Free choice by female sticklebacks: lack of preference for male dominance traits

To experimentally assess if a male sexual trait has an influence on female choice and (or) male-male competition, it is essential to separate these two evolutionary forces. Female choice may be obscured by the overriding influence of male-male competition, and it may therefore appear that female interests coincide with male dominance. We approached this problem using the threespine stickleback, Gasterosteus aculeatus, by leashing the males, allowing the female to fully interact and mate with each male, while physical interactions between the males were prevented. Our results show that in this situation, females display no preference for dominant males and no, or only a weak, preference for more colorful males. When the males could interact freely with each other, females "chose" the dominant and the more colorful males, but in this case the dominant male always deprived the female of the opportunity to interact with the subordinate male. When the females were separated physically from the males by glass walls, they demonstrated a preference for males with the most intense red color. This suggests that if females are given only a limited number of cues on which to base their choice (by introducing glass walls), they are attracted to the redness of the males, while in a situation involving free physical and sensory interaction, other factors become more important for females' choice if they have the possibility to make a free choice in the absence of forceful male-male interactions.

Free choice by female sticklebacks: lack of preference for male dominance traits

To experimentally assess if a male sexual trait has an influence on female choice and (or) male-male competition, it is essential to separate these two evolutionary forces. Female choice may be obs ...

Virulence of the cestode Schistocephalus solidus and reproduction in infected threespine stickleback, Gasterosteus aculeatus

We investigated the relationship between reproduction in the threespine stickleback (Gasterosteus aculeatus) and parasitism by plerocercoids of the cestode Schistocephalus solidus in Walby Lake, Alaska, by quantifying stickleback reproduction and parasite infection using 1655 fish from four samples collected in 1990-1996. Stickleback in Walby Lake largely spawned during May and June as 2-year-olds in the second spring-summer after hatching, as was the case with other stickleback populations we studied in south-central Alaska. Contrary to an earlier hypothesis that S. solidus has been selected to delay its deleterious effects on threespine stickleback, i.e., limit its infection levels, until after the stickleback have reproduced, substantial levels of parasitic infection co-occurred with the stickleback reproductive period. Chi-squared analyses of individual samples suggested that in May, infected females were as capable of producing clutches of eggs as uninfected females but in June, S. solidus inhibited clutch production. An overall analysis, however, failed to support the hypothesis that the effect of S. solidus on clutch production differed between early and late periods of the spawning season. We concluded that S. solidus inhibits the ability of female stickleback in Walby Lake to produce a clutch, and that there was no differential effect on clutch production with season. Nonetheless, 77% of all infected females produced clutches. These results contrast with those of one study in which it was found only 9% of infected females became gravid (ripe) and another report that 23% of infected females were able to mature. We offer hypotheses for the co-occurrence of stickleback reproduction and substantial parasitism at the population level and for the ability of a large proportion of infected females to produce clutches. Our results suggest that the host-parasite relationship is more complex than was previously realized.

Effect of variable food levels on reproductive performance of breeding female three‐spined sticklebacks

The study examined the response to variation in food ration within a breeding season on components of the reproductive performance of mature female three‐spined sticklebacks Gasterosteus aculeatus, a batch spawner. The main effect of food availability was on the duration of the inter‐spawning interval (ISI). Low ration resulted in an increase in the number of days until the next spawning. A low ration during one inter‐spawning interval increased the length of the next ISI, even if there was a high ration in the latter. Females that experienced high rations for two successive ISIs showed an increase in batch fecundity over the successive spawnings. There was no tendency for fish on low rations to produce bigger eggs or eggs with higher lipid concentration. Females on low ration had a lower carcass dry matter concentration and a lower liver dry weight immediately after their third spawning. Female sticklebacks responded to a low ration with little change in egg number or size, but a slight reduction in physiological condition. Egg characteristics and, to a lesser extent, batch fecundity are insensitive to current rate of food consumption, but the interval between spawnings is sensitive to both current and previous rates of consumption.

Condition-related mate choice in sticklebacks

Exactly how the high genetic variance of traits involved in sexual selection through female mate choice is maintained is a much debated issue1,2,3. Theoretical models attempt to explain the high genetic variance of sexual traits, among others, by the evolution of condition dependence1,4, in which expression of the trait depends on physical condition. If it is costly to be choosy, then these models should also apply to female mate preference and predict that female condition affects the choice of mate. Here we allow female laboratory-bred three-spined sticklebacks, Gasterosteus aculeatus, to choose between computer animations of courting males5, and find that the choice of mate correlates well with female physical condition.

Empirical evidence of a trade–off between reproductive effort and expectation of future reproduction in female three-spined sticklebacks

Optimal life-history models generally predict that the reproductive effort of iteroparous organisms may increase with age, as their expectation of future reproduction decreases. The population of three–spined sticklebacks ( Gasterosteus aculeatus ) in the Camargue (Rhone River Delta, France) is annual, all adults dying after their first breeding season. As the three–spined stickleback is a multiple spawner, we tested the hypothesis that reproductive effort may increase during the breeding season on field data. From 1987 to 1998, 653 female sticklebacks were collected in the field during the breeding seasons. The body size, body weight and weights of the liver, gonads and carcass were measured for these individuals. Only gravid females with mature eggs (176 fish) were included in the analysis. Considering the female three–spined stickleback as a capital breeder, the energetic resources available for allocation between soma and gonads were estimated by its body weight. Somatic condition decreased during the breeding season and reproductive effort (gonad weight relative to body weight) increased. These patterns did not vary significantly between years. These observed variations in reproductive effort during the breeding season can be interpreted as empirical evidence of a trade–off between reproductive effort and expectation of future reproduction.

Female fifteen-spined sticklebacks prefer better fathers

We studied how male fifteen-spined sticklebacks,Spinachia spinachia, vary in paternal competence, whether males advertise their competence and whether females prefer better fathers. In this species the male alone provides care for the offspring through nest building, fanning, cleaning and protecting the eggs. We found no female preference for larger males. Instead, females preferred males that during the subsequent paternal phase fanned their nests in shorter fanning bouts. Such males enjoyed a significantly higher hatching success because they fanned more often than males with longer fanning bouts. Males that fanned for short bouts during the paternal phase were also able to increase their fin beat rate. Frequent fanning and high fin beat rates may improve the flow of oxygen to the eggs. Beat rate may be a condition-dependent trait, because males that lost more weight were unable to increase their fin beat rate. During courtship, males perform behaviours such as displacement fanning and body shaking. Females preferred males showing more frequent body shakes during courtship. Body shake frequency correlated positively with hatching success, and negatively with mean fanning bout duration during the paternal phase. The results indicate that femaleS.spinachiashow a preference for direct benefits in terms of better paternal care, and that males may signal this ability to females by shaking their body during courtship.

Effects of gravidity on habitat use and antipredator behaviour in three‐spined sticklebacks

Habitat use was examined in six Alaskan populations of three‐spined sticklebacks Gasterosteus aculeatus. In three lakes with predatory fishes, gravid female sticklebacks remained closer to refuge than did non‐gravid females, while those in lakes devoid of piscivores did not display this shift in behaviour. Gravid females in a lake with predatory rainbow trout Oncorhynchus mykiss used different evasive manoeuvres than did non‐gravid females during in situ encounters with a model rainbow trout. Gravid females were more likely to incorporate a protean element in their flight moffements than were non‐gravid females. Offerall, these data suggest that gravid female three‐spined sticklebacks modify habitat use and escape behaviour to reduce vulnerability to predators.

Different Carotenoids and Potential Information Content of Red Coloration of Male Three-Spined Stickleback

Female sticklebacks (Gasterosteus aculeatus) use the red coloration of males as a criterion for mate choice. Redder males are more attractive. However, males often differ not only in the intensity of their coloration (from dull to bright red) but also in color quality (from yellowish to purple-red). We investigated whether the red coloration of the stickleback is actually a multiple signal made by several pigments. We kept wild caught males singly in tanks until they had built a nest and were ready to accept females. Then, we took standard photographs and measured their colors by spectrometer analyses of the slides and by descriptions of human observers. These two measurements were highly correlated. When analyzing the carotenoid content of the sticklebacks' skin we found two groups of carotenoids (astaxanthin and tunaxanthin/lutein) that were quantified for each individual. The differences in color observed in the fish are correlated to this pigment quantification. Redder fish have more astaxanthin in their skin than yellowish fish, while the color of the yellowish fish appears to be made by tunaxanthin/lutein. Our results suggest that the red coloration of sticklebacks is a multiple trait that is made of at least two different carotenoids. This opens the possibility that male sticklebacks signal more detailed information to females than a one-dimensional trait would allow.

Female sand gobies prefer good fathers over dominant males

Female preference for males successful in male–male competition is generally assumed to result in mating with high quality males. Here I report results from an experiment disentangling the effects of intra– and intersexual selection in the sand goby, Pomatoschistus minutus , a marine fish that exhibits paternal care. I show that large males are successful in male–male competition, but contrary to what one would expect, dominants are not preferred by females and are not better at taking care of the eggs. Female preference, however, correlated with the subsequent hatching success of the eggs. Thus, female choice selects for good parenting. Hence, direct benefits in the form of superior paternal care can explain female choice in this species, supporting a good parent process of sexual selection. However, choosing on the outcome of male–male competition does not enable females to mate with the ‘best’ males.