Effects of 17alpha-ethynylestradiol on sexual development of male western mosquitofish (Gambusia affinis)

Juvenile male western mosquitofish (Gambusia affinis) were exposed to different concentrations of 17alpha-ethynyl estradiol (EE2) in the diet during the period of sexual maturation. A clear inhibiting influence of EE2 on sexual development was apparent. The proportion of males in each treatment group that failed to complete gonopodial development during the 150-day observation period increased significantly with EE2 concentration. There were significant nonlinear trends toward shorter gonopodia in groups exposed to higher EE2 concentrations. Vitellogenin (VTG) was detectable in the blood of all fish exposed to 1.0 or more micro/g food and the concentration increased dramatically with increasing EE2 exposure. A significant negative association was seen between EE2 concentration and spermatophore counts. This study has demonstrated deleterious effects of EE2 exposure on sexual maturation and several indirect measures of reproductive fitness. It supports the biological relevance of vitellogenin in the blood and reduced gonopodium length as biomarkers for estrogen exposure and endocrine disruption in mosquitofish.     

Regional ecological risk assessment for the introduction of Gambusia affinis (western mosquitofish) into Montana watersheds

Qualitative risk assessment methodologies were used to assess the risk of establishment and consequent impacts on native minnows and species of concern (SOC) associated with the intentional or unintentional introduction of the mosquito biological control agent, Gambusia affinis, to various Montana watersheds. Gambusia affinis introductions for mosquito control have been made throughout the world; some introductions have resulted in deleterious effects on native species. We used average January water temperatures, the presence of dams, and the presence of native minnows and SOC to define endpoints for our assessment. Our results suggest that a section of the Missouri River running between Wolf Creek and Landusky had the highest overall risk score, which corresponds to the highest likelihood of establishment and effects on native minnows and SOC. We also demonstrate how rivers with the highest temperatures are not necessarily at the highest risk of having deleterious effects on minnows and SOC; conversely, these rivers were found to be at highest risk of a population of G. affinis establishing.     

Impacts of mosquitofish (Gambusia affinis) predation on plankton communities

An investigation of the effects of mosquitofish (Gambusia affinis) predation was conducted in 12 experimental ponds in southern California over a period of 10 months.Gambusia essentially eliminatedDaphnia pulex andCeriodaphnia sp. populations, reducedDiaptomus pallidus andKeratella quadrata populations, had little impact onCyclops vernalis, and caused large increases inK. cochlearis, Polyarthra sp.,Synchaeta sp., andTrichocerca spp. populations and in total phytoplankton.Gambusia caused a decrease in the PIE (probability of interspecific encounter) of the planktonic crustaceans and an increase in the PIE of the planktonic rotifers. Hemiptera, such as neustonicMicrovelia sp. and nektonicBuenoa sp. andNotonecta sp., andHyla regilla tadpoles were absent from fish ponds but sometimes abundant in control ponds.Gambusia caused higher pH and oxygen levels, presumably via its effect on the phytoplankton. The impact ofGambusia on the pond ecosystems was less in winter, when fish numbers and feeding rates were low, than in summer. Results of other fish-plankton studies are summarized in tabular form. A model is proposed to account for variation in the calanoid/cyclopoid ratio; evidence is summarized suggesting that in general calanoids are more susceptible to predation by predaceous zooplankters while cyclopoids are more susceptible to fish predation. Some parallels are drawn between the effects ofGambusia predation and those of insecticide treatments.     

Geographical and temporal variation of multiple paternity in invasive mosquitofish (Gambusia holbrooki,Gambusia affinis)

Multiple paternity (MP) increases offspring's genetic variability, which could be linked to invasive species' evolvability in novel distribution ranges. Shifts in MP can be adaptive, with greater MP in harsher/colder environments or towards the end of the reproductive season, but climate could also affect MP indirectly via its effect on reproductive life histories. We tested these hypotheses by genotyping N = 2,903 offspring from N = 306 broods of two closely related livebearing fishes, Gambusia holbrooki and Gambusia affinis. We sampled pregnant females across latitudinal gradients in their invasive ranges in Europe and China, and found more sires per brood and a greater reproductive skew towards northern sampling sites. Moreover, examining monthly sampling from two G. affinis populations, we found MP rates to vary across the reproductive season in a northern Chinese, but not in a southern Chinese population. While our results confirm an increase of MP in harsher/more unpredictable environments, path analysis indicated that, in both cases, the effects of climate are likely to be indirect, mediated by altered life histories. In both species, which rank amongst the 100 most invasive species worldwide, higher MP at the northern edge of their distribution probably increases their invasive potential and favours range expansions, especially in light of the predicted temperature increases due to global climate changes.     

The western mosquitofish (Gambusia affinis affinis): a new laboratory animal resource for the study of sexual dimorphism in neural circuits

The western mosquitofish (Gambusia affinis affinis) is a useful model for the study of sexual dimorphism and the neural circuits associated with sexual differentiation. This is largely because of its anal fin, which undergoes radical postnatal transformation in males. Understanding the neural mechanisms involved in this process may also help elucidate basic principles of the nervous system. The authors describe the mosquitofish as a model for research and present guidelines for the care and use of this species.     

Deleterious effects by mosquitofish (Gambusia affinis) on the endangered fire salamander (Salamandra infraimmaculata)

Invasive fishes, via competition and predation, may be an important factor in endangering populations of amphibians and other aquatic fauna. The mosquitofish, Gambusia affinis , commonly stocked into amphibian breeding sites for mosquito control worldwide, has recently been found in some breeding sites of the endangered fire salamander Salamandra infraimmaculata in northern Israel. A comparison of Salamandra larvae in natural pools varying in Gambusia densities suggested that Gambusia negatively affects Salamandra by causing damage to its appendages, in particular, its tailfin (as manifested in a smaller tail:body ratio). Comparison of the same pool before and after mosquitofish introduction suggested strong negative effects on larval performance and number of emerging metamorphs. To explicitly test the hypothesis that Gambusia negatively affects Salamandra larval development, growth and survival, we conducted a replicated outdoor mesocosm experiment. In this experiment, we also tested how habitat structural complexity, in the form of radiating cords simulating artificial vegetation, might mediate damage caused by Gambusia . We used a two-by-two factorial design: the presence or absence of increased structural complexity crossed with the presence or absence of mosquitofish. After a short period, mosquitofish strongly reduced larval wet weight and survival, and increased body damage in the form of a reduced larval tail:body ratio and partially missing gills and limbs. No Salamandra larvae exposed to mosquitofish survived to metamorphosis in either habitat type. We conclude that Gambusia strongly and negatively affects Salamandra larvae and that the introduction of Gambusia into sites containing Salamandra is inconsistent with the goal of preserving this endangered urodele.     

The effects of induced masculinization on reproductive and aggressive behaviors of the female mosquitofish,Gambusia affinis affinis

Synopsis FemaleGambusia affinis affinis were masculinized with the degraded products of 65% stigmastanol-30% B-sitosterol, a phytosterol. The masculinized females were paired with non-treated males, non-treated females, and other masculinized females. The pairs were analyzed for reproductive and aggressive behaviors exhibited. The behavioral patterns of these pairs were compared statistically to the behavior patterns of the following non-treated pairs: male with male, female with female, and male with female. The masculinized females behaved like males in that they followed, swung, and thrust their gonopodia at non-treated females and larger masculinized females. However, the male-like behavior of the masculinized females was not as intense as that of normal males. Also, the effect of masculinization on the behavior of the treated female was context-dependent as seen when placed with a male or a smaller masculinized female. Under these conditions the masculinized females behaved like typical females and exhibited no masculinized behaviors. The effects of anal spots and size differences are discussed as possible explanations for the variability in behaviors exhibited. The masculinized females displayed no change in aggressive behaviors.     

Gonopodial morphogenesis in female mosquitofish,Gambusia affinis affinis,masculinized by exposure to degradation products from plant sterols

Synopsis Female mosquitofish,Gambusia affinis affinis, were masculinized by exposure to degradation products (presumably steroids) of the plant sterol, stigmastanol. Masculinization was indicated by the induction of a male-like gonopodium in each specimen. The morphogenetic stages of gonopodial development are discussed as are the anatomical specializations produced in the mature gonopodial tip.     

Putative chemical inhibition of development by conspecifics in mosquitofish, Gambusia affinis

The objective of this study was to test for a chemical influence of adult Gambusia affinis on the development and maturation of juveniles. Sixty juveniles (<1 day old) were reared (from a total of 157 trials) individually up to 115d in 3liter containers that circulated water bathing either (1) two adult males, (2) two adult females, (3) an adult male and an adult female, or (4) conspecific-free tank water. Randomly chosen juveniles were sampled over time, sacrificed, sectioned, and measurements were taken of standard length and gonadal parameters such as gonad size, number of spermatogenic cysts, or number of oocytes. Measurements were regressed as functions of time, and analyzed for significant differences by ANCOVA. A total of 32 males and 28 females were examined. Males showed no significant differences in any of the parameters, but females showed reduced growth rate and smaller ovary size when reared in water bathing two adult females. Mortality rate in these young fish was high, but Chi-square analysis showed that mortality rate was significantly higher when they were raised in water bathing at least one adult female. Females are known to influence female development in other vertebrates; our findings suggest this may also be true for a fish, and that the interaction may be through chemical communication.     

Tolerance of low oxygen stress in insecticide-resistant and susceptible populations of mosquitofish (Gambusia affinis)

The tolerance of low oxygen stress in insecticide-resistant and susceptible populations of mosquitofish ($\text{Gambusia}$$\text{affinis}$) was investigated. Dissolved oxygen was reduced chemically with formaldehyde and sodium sulfite treatment. Available oxygen was reduced physically through barriers, both by preventing oxygen exchange at the air-water interface and by preventing the fish from contacting the surface of the water. In all cases, the tolerance to low oxygen stress was reduced in the insecticide-resistant population compared to the susceptible population. The results indicate that although tolerance of pesticides in the resistant population has been dramatically increased by environmental selective pressures, tolerance to other stresses has not been concomitantly increased.     

Regulation of pro-gonadotropin-releasing hormone gene expression by sex steroids in the brain of male and female rats

The fine modulation of gonadotropin gene expression and secretion is well recognized to be regulated by sex steroids through their direct action both at the anterior pituitary level and on the pulsatile pattern of GnRH secretion at the hypothalamic level. Since the influence of sex steroids on hypothalamic GnRH mRNA levels remains to be elucidated, quantitative in situ hybridization was used to study the effect of sex steroids on cellular levels of pro-GnRH mRNA in adult rats of both sexes. The effects of 14-day gonadectomy as well as administration of 17 beta-estradiol (E2, 0.25 micrograms) or dihydrotestosterone (DHT, 100 micrograms) twice a day during 14 days to gonadectomized animals were evaluated. In addition, the effect of progesterone (P, 2 mg, twice daily) alone or in the presence of E2 was also studied in ovariectomized animals. Hybridization was performed using a 35S-labeled cDNA probe encoding rat pro-GnRH and the corresponding mRNA levels were assessed by counting the number of silver grains overlying labeled neurons. In male rats, castration induced a highly significant 65% increase (compared to intact rats) in the mean number of grains per neuron. Administration of E2 or DHT to castrated animals completely prevented the post castration rise in pro-GnRH mRNA levels. In female animals, the effect of ovariectomy was less striking than in the male, a 25% increase (P less than 0.001) being observed. Treatment with E2 or DHT also completely prevented the increase in pro-GnRH mRNA levels induced by ovariectomy. Moreover, treatment with P in ovariectomized animals markedly potentiated the inhibitory effect of E2 on pro-GnRH mRNA levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fat cycling in the mosquitofish (Gambusia affinis): fat storage as a reproductive adaptation

Summary We argue, based on reviewed literature covering reptiles, amphibians, birds, and fish, that fat storage may represent a life history adaptation because it enables an organism to shift in time when resources are allocated to reproduction. We applied these arguments to fat and population cycles in three populations of the mosquito fish, Gambusia affinis. For males, there appeared to be a constant size at maturation during the reproductive season. Mature males became scarce late in the summer. At the same time, immature males delayed maturity and attained much larger sizes; they matured in large numbers in the fall. The amount of stored fat tended to be equal for immature and mature males at all times except in the late summer. In the August samples, when mature males were relatively rare, they also had the lowest level of fat reserves. It appears that the older generation of mature males did not store fat and did not overwinter. At the same time, immature males registered a two to three fold increase in fat reserves. These differences in fat content between mature and immature males disappeared by September, probably because of the recruitment of a new generation of mature males. The reserves were gradually utilized during the winter. Females reproduced from the late spring through mid- to late-summer. They stopped reproducing in the late summer, when there was ample time to produce an additional litter of young. There was an inverse relationship between resources devoted to reproduction and fat reserves. As reproductive allotment decreased in the late summer, fat reserves increased. The magnitude of the change in fat reserves was similar to that displayed by males. The reserves were depleted over the winter. Significant reserves remained at the beginning of the reproductive season the following spring. Reproducing females utilized the remaining reserves significantly more rapidly than non-reproducing females. An analysis of resource availability revealed an overall decrease in food availability in the late summer, coincident with the increase in fat reserves. These cycles are therefore not attributable to changes in resource availability. They instead indicate a change in how resources are allocated by the fish. The trends in the data indicate that fat reserves are used to shift investment in reproduction from the late summer to the following spring. In males, deferring maturity, rather than maturing in August, allows them to store the necessary reserves to survive the winter so that they can mate the following spring. In females, a subset of the fat reserves is intended for producing the first clutch of eggs the following spring. The female pattern corresponds to those reported for a diversity of organisms. The possible advantages of shifting reproductive effort from the fall to the following spring include higher fecundity and higher offspring fitness. The limitations of the methodology and potential directions for future research are discussed.     

Effects of steroid hormones on reproduction- and detoxification-related gene expression in adult male mosquitofish, Gambusia affinis

The molecular mechanisms that mediate fish reproduction and detoxification in response to steroid hormones were studied by using adult male western mosquitofish (Gambusia affinis) as sentinel species. The expression patterns of three vitellogenins (VtgA, VtgB and VtgC), two estrogen receptors (ERα and ERβ), two androgen receptors (ARα and ARβ), metallothionein (MT) and cytochrome P450 1A (CYP1A) in the liver and testis of adult male mosquitofish were assessed through exposure treatments with progesterone (P), testosterone (T) and 17β-estradiol (E2), alone and in combination for eight days. The results showed that expression patterns of Vtg subtype, ER subtype, AR subtype, MT and CYP1A genes in male mosquitofish varied according to tissue and specific hormone stress. Vtg subtype mRNA expression was induced in the liver in E2-added treatments, and an up-regulation of ERα mRNA expression was also observed. In addition, hormone treatments increased three Vtg subtype mRNA expression levels in the testis, at least to some extent. All hormone treatments significantly inhibited ERα, ERβ and ARβ mRNA expression in the testis. Some of hormone treatments could affect MT and CYP1A gene expression in mosquitofish. In general, multiple hormone treatments showed different effects on target gene expression compared with corresponding hormone alone. The results from the present study provided valuable information on the toxicological effects of steroid hormones in mosquitofish.     

Population development of the mosquitofish,Gambusia affinis,in rice fields

Synopsis Development of mosquitofish,Gambusia affinis, stocks in rice fields following stocking for mosquito control is poorly understood and highly variable. To characterize population development and explain observed variability, size distributions and total numbers of mosquitofish stocks were followed from stocking through the end of the summer rice season in several experimental rice paddies. Instead of the highly variable, logistic growth implied by previous studies in which only adult fish were sampled, we observed a consistent development of size structure between years, with some variability in specific demographic processes. Population development consists of: (1) an initial peak in numbers due to a pulse of recruitment, (2) a period of low recruitment and constant or declining numbers, and (3) a second peak caused by a second pulse of recruitment. Timing of the initial peak appears to depend on rice height. Differences in the rate of decline following the peak are apparently due to different mortality rates. The second peak in reproduction is probably due to a second parturition by stocked females, possibly followed by first reproduction by young born earlier. Since population development in rice fields following stocking in the spring is similar to development of natural populations in temperate regions in which there is no reproduction during the winter, results obtained here are relevant to natural as well as artificially stocked systems.     

Environmental regulation of annual reproductive cycle in the mosquitofish, Gambusia affinis

To clarify the environmental factors regulating the annual reproductive cycle of the female mosquitofish, Gambusia affinis, a viviparous teleost, histological changes of the ovary in natural population, and laboratory experiments were examined. The results, extending over two years, suggested that ovarian recrudescence is initiated by the rise in temperature during spring and that ovarian regression is caused by the shorter daylength during late summer. The first rearing experiments using four photoperiod-temperature groups to investigate the factors triggering the onset of reproduction revealed that females with regressing ovaries began reproduction with the rise of temperature regardless of the photoperiod during spring. The results of the second experiment using three different temperature groups indicated that vitellogenesis occurred at over 14 degrees C and pregnancy at over 18 degrees C. The third experiment with four photoperiod-temperature groups was arranged to investigate the factors in the cessation of reproduction. Sexually active females ceased vitellogenesis of the next clutch of oocytes due to the shorter daylength regardless of temperatures during late summer; however, temperature seemed to influence the rate of embryo development. The critical photoperiod is estimated at about 12.5 hr. In nature, it is supposed that vitellogenesis starts when the temperature rises to about 14 degrees C, and final maturation of oocytes occurs when the temperature reaches about 18 degrees C during spring. Then, vitellogenesis of the next clutch of oocytes ceases when the daylength becomes shorter than 12.5 hr during late summer; the last gestation proceeds at a rate dependent on the temperature, and finally reproduction ends by the last parturition. J. Exp. Zool. 286:204-211, 2000.     

Quantification of vitellogenin mRNA induction in mosquitofish (Gambusia affinis) by reverse transcription real-time polymerase chain reaction (RT-PCR)

A method to quantify induction of vitellogenin (Vtg) mRNA in adult male mosquitofish was developed. Male mosquitofish were exposed to 0, 1, 20 and 250 ng l^-1 17β-oestradiol (E₂) for 4 and 8 days in static exposures, and liver Vtg mRNA and 18S rRNA expression were quantified in duplex RT-PCR. Liver 18S rRNA expression was very consistent among individuals, and there was a highly significant increase in Vtg mRNA expression after exposure of mosquitofish for just 4 days at 250 ng l^-1 E₂. Lower doses did not induce Vtg mRNA expression even at 4 or 8 days. This method could be used as a rapid test to detect exposure of mosquitofish to oestrogenic chemicals. Further work is needed to determine if increased Vtg mRNA levels in male mosquitofish induce Vtg synthesis, and to determine the usefulness of the method in field sampling.     

Quantification of vitellogenin mRNA induction in mosquitofish (Gambusia affinis) by reverse transcription real-time polymerase chain reaction (RT-PCR)

Abstract

A method to quantify induction of vitellogenin (Vtg) mRNA in adult male mosquitofish was developed. Male mosquitofish were exposed to 0, 1, 20 and 250 ng l^?1 17β-oestradiol (E₂) for 4 and 8 days in static exposures, and liver Vtg mRNA and 18S rRNA expression were quantified in duplex RT-PCR. Liver 18S rRNA expression was very consistent among individuals, and there was a highly significant increase in Vtg mRNA expression after exposure of mosquitofish for just 4 days at 250 ng l^?1 E₂. Lower doses did not induce Vtg mRNA expression even at 4 or 8 days. This method could be used as a rapid test to detect exposure of mosquitofish to oestrogenic chemicals. Further work is needed to determine if increased Vtg mRNA levels in male mosquitofish induce Vtg synthesis, and to determine the usefulness of the method in field sampling.     

Regulation of adult bone turnover by sex steroids

Recent reports reveal increasing complexity of mechanisms underlying the bone sparing effects of sex steroids. This review focuses on mechanisms by which sex steroids attenuate endocortical and trabecular adult bone turnover, perhaps their most important property as bone mass regulators. Clearly, estrogen withdrawal increases osteoclast number and bone resorption; however, important open questions are the extent to which osteoblasts and their precursors are involved, and the relative contributions of the RANK/RANKL/OPG system, Fas ligand and Runx2. In addition to reviewing these aspects of estrogen action, we also discuss proskeletal effects of androgens on the adult male skeleton, including aromatization to estrogens and male‐specific mechanisms. Detailed understanding of skeletal site‐ and gender‐dependent mechanisms by which sex steroids protect the adult skeleton will provide the foundation for improved risk assessment, prevention and management of osteoporosis. J. Cell. Physiol. 224: 305–310, 2010. © 2010 Wiley‐Liss, Inc.