Hormonal induction of undescribed males resolves cryptic species of cladocerans

Cyclic parthenogens have a mixed breeding system with both meiotic and ameiotic eggs. Although investment in sexual stages is often synchronized with seasonal cycles, the degree of investment is a quantitative trait associated with habitat instability. Populations of cyclic parthenogens from stable environments, such as large lakes and oceans, generally show reduced or undetectable investment in males. Indeed, males of many species of lacustrine cyclic parthenogens are unknown to science. Methyl farnesoate (MF), a crustacean juvenile hormone, has been implicated as an inducer of male formation in Daphnia magna (Crustacea: Cladocera), a denizen of unstable habitats with marked sexual recruitment. Here, we show experimentally that MF induces male production in four distantly related lacustrine species of cladocerans under growth conditions unfavourable for male production. The males of three species are new to science. Unlike females, the anatomy of the previously unknown males of Bosmina (Lunobosmina) oriens permitted ready morphological diagnosis of sibling species and subfossils. The results suggest that the role for MF in the sex determination of cladocerans is general.     

Sex allocation variation in Daphnia pulex

Daphnia (Crustacea: Cladocera) reproduce by cyclical parthenogenesis in which the sex of offspring is environmentally determined. Although numerous studies have demonstrated that factors such as crowding and short-day photoperiod stimulate male production, there is limited information on variation in allocation to male and female offspring for any species of Daphnia . The present study assessed the presence or absence of male production in 96 isofemale lines (clones) from each of eight populations of Daphnia pulex . An average of 37% (range 18–51%) of clones failed to produce males under crowded conditions in the laboratory. A subset of 14 of these non-male-producing clones also failed to produce males under short-day photoperiod (8L:16D). Three male-producing clones were within-clone mated as well as crossed to three non-male-producing clones to study the inheritance of the failure to produce males. The average frequency of non-male-producing F ₁ progeny was significantly higher (58%, N = 486) among the outcrossed progeny than the inbred progeny (5%, N = 86). In addition, when sixteen of the male-producing outcrossed progeny were within-clone mated, only 7% ( N = 106) of the resulting F ₂ progeny failed to produce males. These results are consistent with a genetic basis for the absence of male production. Average survival of the progeny from the nine outcrossed matings was more than twice (67%) that of the inbred progeny from the three within-clone matings (30%), suggesting that within-clone mating would result in significant inbreeding depression. We present a model that suggests that even low levels of inbreeding could allow non-male-producing females to be maintained in a population. The co-occurrence of non-male-producing females and females that produce both males and females in Daphnia pulex bears a similarity to the gynodioecious breeding system found in some plant species.     

Photoperiodic effect on size-related metamorphosis in the milkweed leaf beetle, Labidomera clivicollis

ABSTRACT. Photoperiod significantly affected size-related aspects of metamorphosis in the milkweed leaf beetle Labidomera clivicollis (Kirby) (Chrysomelidae). Larvae reared under short-day photoperiod successfully pupated at lower weights than larvae reared under long-day photoperiods. This short-day effect was observed to an equivalent extent in both males and females; but within photoperiod treatments, males consistently pupated at lower weights than females. It is likely that the short-day effect on metamorphosis is associated with photo-periodically induced changes in larval corpora allata activity or juvenile hormone titre. Ecologically, the short-day effect on metamorphosis may enhance the moulting success of larvae which encounter food shortages during autumn.     

Methyl farnesoate synthesis is necessary for the environmental sex determination in the water flea Daphnia pulex

Sex-determination systems can be divided into two groups: genotypic sex determination (GSD) and environmental sex determination (ESD). ESD is an adaptive life-history strategy that allows control of sex in response to environmental cues in order to optimize fitness. However, the molecular basis of ESD remains largely unknown. The micro crustacean Daphnia pulex exhibits ESD in response to various external stimuli. Although methyl farnesoate (MF: putative juvenile hormone, JH, in daphnids) has been reported to induce male production in daphnids, the role of MF as a sex-determining factor remains elusive due to the lack of a suitable model system for its study. Here, we establish such a system for ESD studies in D. pulex. The WTN6 strain switches from producing females to producing males in response to the shortened day condition, while the MFP strain only produces females, irrespective of day-length. To clarify whether MF has a novel physiological role as a sex-determining factor in D. pulex, we demonstrate that a MF/JH biosynthesis inhibitor suppressed male production in WTN6 strain reared under the male-inducible condition, shortened day-length. Moreover, we show that juvenile hormone acid O-methyltransferase (JHAMT), a critical enzyme of MF/JH biosynthesis, displays MF-generating activity by catalyzing farnesoic acid. Expression of the JHAMT gene increased significantly just before the MF-sensitive period for male production in the WTN6 strain, but not in the MFP strain, when maintained under male-inducible conditions. These results suggest that MF synthesis regulated by JHAMT is necessary for male offspring production in D. pulex. Our findings provide novel insights into the genetic underpinnings of ESD and they begin to shed light on the physiological function of MF as a male-fate determiner in D. pulex.     

Sex steroid hormones enhance immune function in male and female Siberian hamsters

Immune function is better in females than in males of many vertebrate species, and this dimorphism has been attributed to the presence of immunosuppressive androgens in males. We investigated the influence of sex steroid hormones on immune function in male and female Siberian hamsters. Previous studies indicated that immune function was impaired in male and female hamsters housed under short-day photoperiods when androgen and estrogen concentrations were virtually undetectable. In experiment 1, animals were gonadally intact, gonadectomized (gx), or gx with hormone replacement. Females exhibited the expected increase in antibody production over males, independent of hormone treatment condition, whereas male and female gx animals exhibited decreased lymphocyte proliferation to the T cell mitogen, phytohemagglutinin (PHA) compared with intact animals, and this effect was reversed in gx hamsters following testosterone and estradiol treatment, respectively. In experiment 2, testosterone, dihydrotestosterone, and estradiol all enhanced cell-mediated immunity in vitro, suggesting that sex steroid hormones may be enhancing immune function through direct actions on immune cells. In experiment 3, an acute mitogen challenge of lipopolysaccharide significantly suppressed lymphocyte proliferation to PHA in intact males but not females, suggesting that males may be less reactive to a subsequent mitogenic challenge than females. Contrary to evidence in many species such as rats, mice, and humans, these data suggest that sex steroid hormones enhance immunity in both male and female Siberian hamsters.     

Hormone-pheromone relationships of male Tenebrio molitor

Males of the beetle Tenebrio molitor produce a volatile sex pheromone which attracts females of the same species. The pheromone level peaks 8 days after emergence and then reaches a plateau. Elimination of endocrine centers by decapitating male adults 24 hr following adult ecdysis did not impair pheromone production. Treatment of decapitated males with juvenile hormone analogues did not make any detectable difference in the levels of pheromone activity. However, undecapitated males treated with juvenile hormone analogue showed a significant increase in pheromone activity when compared with those that had been decapitated and subsequently treated with juvenile hormone analogue. This observation is discussed in the light of published research on the effect of juvenile hormone on pheromone activity of females of T. molitor.     

Parental control of sex ratio in Gammarus duebeni,an organism with environmental sex determination

Parental sex ratio control was investigated in Gammarus duebeni, an amphipod with an environmentally mediated sex determining system. The effect on the F2 generation sex ratio of the photoperiodic conditions experienced by a) the P generation during and after copulation, b) the F1 generation before and after sex determination, and c) the F2 generation themselves during the period of sex determination, was tested. The photoperiodic conditions the F2 generation experienced during the period of sex determination had a significant effect on their sex ratio (more males were produced under long-day than under short-day conditions), but the photoperiodic conditions experienced by the F1 generation males and females or the P generation on the F1 male's side had no effect on the F2 sex ratio. However, the photoperiodic conditions the P generation on the F1 female's side experienced significantly affected the F2 sex ratio. When these animals experienced long-day conditions the F2 generation became female biased and when they experienced short-day conditions, male biased. It is proposed that the mechanism of control operates through the F1 generation mothers whilst in an embryonic stage of development in the P generation mother's brood pouch. The photoperiodically mediated effects of the embryonic F1 generation mother and the F2 generation on sex determination are additive. A mechanism by which both F1 generation maternal and F2 generation sex ratio control could operate in the field is proposed.     

Photoperiodic and hormonal control of reproduction in male beetles, Pterostichus nigrita

After metamorphosis the development of males of Pterostichus nigrita is controlled by photoperiods. Development is free of any dormancy in short day conditions (photoperiods with less than 16·9 to 15·1 hr of light per day). This type of dormancy can be called ‘photoperiodic quiescence’. Short days promote the aggregation of spermatozoa to spermiozeugma. Long day conditions suppress the development of spermiozeugma reversibly. The optimum temperature for the production of spermiozeugma is 15°C. Photoperiods must be perceived by means of the compound eyes. Extirpation of both eyes results in a development of the gonads just as occurs in continuous darkness.Short days activate the corpora allata. The volume of these glands is correlated with the state of sexual maturity (formation of spermiozeugma, growth and filling of the accessory glands). Intra- and intersexual transplantations of active corpora allata into undeveloped male beetles maintained in long day photoperiods replace the short day influence.Very probably the gonadotropic hormone is a juvenile hormone. By injection of 10,11-farnesyl-methyl-ester or a synthetic juvenile hormone, sexual maturity can be initiated.     

Two-step photoperiodic and hormonal control of reproduction in the female beetle, Pterostichus nigrita

The carabid P. nigrita is a short-day/long-day insect. Short days with less than 15.0 hr of light per day induce pre-vitellogenesis. Vitellogenesis only takes place in subsequent long days with more than 13,1 hr of light. In the range from ld 13:11 to ld 16:8 egg production is possible without a change in photoperiod. Pre-vitellogenesis is induced by the corpus allatum hormone. Implantation of active corpora allata or injection of juvenile hormone substitutes for short-day photoperiods. Histological investigations and implantation experiments revealed that for vitellogenesis some neurosecretion is necessary. Ecdysterone cannot replace this neurosecretion.     

Enhancing male sexual success in a lekking fly (Anastrepha suspensa Diptera: Tephritidae) through a juvenile hormone analog has no effect on adult mortality

While defending lek-territories, male Anastrepha suspensa (Loew) produce chemical, acoustic and visual courtship signals. In the laboratory and under semi-natural conditions, topical application of the juvenile hormone analog methoprene doubles pheromone production and subsequently doubles sexual success. However, sexual signals and interactions are likely to be physiologically expensive and so result in higher male mortality. Comparison of males kept in isolation for 35 days, but provided daily with a potential mate or a rival male, revealed that both male- and female-interactors shortened focal-male lifespan. In addition, focal males were either treated with methoprene or not, then either provided with protein in their sucrose-based diet or not. Protein proved to similarly double sexual success and also resulted in longer male life spans in all of the interactor-categories. However, there was no evidence that methoprene induced hypersexuality resulted in higher rates of mortality, i.e., the longevity of males treated with methoprene did not significantly differ from untreated males in the same interactor/diet categories. This apparent lack of costs to a putatively sexually selected signal is unexpected but presents an opportunity to increase the sexual competence of sterile flies with few consequences to their survival following mass-release.     

Regulation of methyl farnesoate production by mandibular organs in the crayfish, Procambarus clarkii: a possible role for allatostatins

Decapod crustaceans do not appear to produce juvenile hormone, but rather its immediate precursor, methyl farnesoate (MF). Both MF and its immediate precursor, farnesoic acid (FA) are produced by the mandibular organs (MO) in crustaceans. The MO are homologous to the insect corpora allata (CA), the site of juvenile hormone biosynthesis. However, the FGLamide allatostatin (ASTs) peptides, of which there are about 60 distinct forms reported from crustaceans, have previously been found to have no effect on MO activity in crustaceans. We have identified by immunocytochemistry the presence of FGLamide-like AST immunoreactivity in neurosecretory cells throughout the CNS as well as in neurohaemal structures such as the sinus gland and pericardial organs. The ASTs are likely delivered to the MO hormonally and/or by local neurohaemal release. Using MO from adult males, we have found wide variability between animals in the in vitro rates of MF and FA biosynthesis. Treatment with Dippu-ASTs has a statistically significant stimulatory effect on MF synthesis, but only in MO that are initially producing MF at lower rates. No effect on FA production was observed, suggesting that the FGLamide ASTs exert their effect on the o-methyl transferase, the enzyme responsible for the conversion of FA to MF.     

Control of diapause induction by a change in photoperiod in Melanoplus sanguinipes

The effect of photoperiod on diapause induction, developmental time and body size was examined in Melanoplus sanguinipes, the lesser migratory grasshopper. Contrary to what is found in most insects, facultative diapause-egg production is found to be controlled by changing rather than constant photoperiods. In addition, developmental time is shown to be faster under short-day photoperiods than long-day photoperiods. And finally, body size is larger under long-day photoperiods and smaller under short-day photoperiods. Implications of these results for the regulation of the seasonal life cycle of M. sanguinipes in the field are discussed.     

Variation in allocation to sexual and asexual reproduction among clones of cyclically parthenogenetic Daphnia pulex (Crustacea: Gladocera)

Organisms reproducing by cyclical parthenogenesis combine the benefits of both sexual and asexual reproduction within the same life cycle. Few studies have examined the evolution of variation in the pattern of investment in parthenogenetic compared to sexual reproduction. Seven clones of Daphnia pulex (Crustacea: Cladocera) varying in allocation to sexual reproduction, as measured by the production of males, were raised in isolation and together in a microcosm to study the pattern of sexual reproduction and the effect of this variation on clone fitness. Sex allocation for clones raised together a microcosm was similar to their allocation when raised in isolation, suggesting a genetic basis to the variation. Three clones showed a cost of producing males that lead to their extinction after about 30 days due to the lack of females required for the clones to persist by parthenogenetic reproduction. The remaining four clones persisted until the end of the 72-day experiment. Clones with little or no allocation to males showed no increased allocation to sexual females. The seven clones showed a greater variation in estimated fitness through male and female function than in total estimated fitness. The clone with the greatest total fitness gained most of its fitness through male function but also had a relatively high fitness through female function. Although one clone produced only females it had the next highest fitness. The three clones that went extinct because of a high investment in males had estimated fitness as high as some clones that persisted in the microcosm because of a higher investment in parthenogenetic reproduction. The similarity in total fitness among clones suggests that Daphnia pulex populations in temporary habitats maintain a sex polymorphism where different genotypes vary-in functional gender ranging from female to primarily male.     

Sexual differentiation and hormonal regulation of the laryngeal synapse in Xenopus laevis

In Xenopus laevis frogs, sex differences in adult laryngeal synapses contribute to sex differences in vocal behavior. This study explores the development of sex differences in types of neuromuscular synapses and the development and hormone regulation of sex differences in transmitter release. Synapses in the juvenile larynx have characteristics not found in adults: juvenile muscle fibers can produce subthreshold or suprathreshold potentials in response to the same strength of nerve stimulation and can also produce multiple spikes to a single nerve stimulus. Juvenile laryngeal muscle also contains the same synapse types (I, II, and III) as are found in adult laryngeal muscle. The distribution of laryngeal synapse types in juveniles is less sexually dimorphic than the distribution in adults. Analysis of quantal content indicates that laryngeal synapses characteristically release low amounts of transmitter prior to sexual differentiation. Quantal content values from male and female juveniles are similar to values for adult males and are lower than values for adult females. When juveniles are gonadectomized and treated with exogenous estrogen, quantal content values increase significantly, suggesting that this hormone may increase transmitter release at laryngeal synapses during development. Gonadectomy alone does not affect quantal content of laryngeal synapses in either sex. Androgen treatment decreases quantal content in juvenile females but not males; the effect is opposite to and smaller than that of estrogen. Thus, muscle fiber responses to nerve stimulation and transmitter release are not sexually dimorphic in juvenile larynges. Transmitter release is strengthened, or feminized, by the administration of estradiol, an ovarian steroid hormone. © 1995 John Wiley & Sons, Inc.     

Sex differences in hormonal responses to social conflict in the monogamous California mouse

Monogamous species are usually considered to be less likely to exhibit sex differences in behavior or brain structure. Most previous studies examining sex differences in stress hormone responses have used relatively sexually dimorphic species such as rats. We examined the stress hormone responses of monogamous California mice (Peromyscus californicus) to resident-intruder tests. We also tested males and females under different photoperiods, because photoperiod has been shown to affect both aggression and stress hormone responses. Females, but not males showed a significant increase in corticosterone levels immediately following a resident-intruder test. Males but not females showed elevated corticosterone levels under short days. Females tested in aggression tests also showed a significant increase in plasma oxytocin levels, but only when housed in long days. This was consistent with our observation that females but not males had more oxytocin positive cells in the paraventricular nucleus (PVN) when housed under long days. Our data show that sex differences in glucocorticoid responses identified in other rodents are present in a monogamous species.     

The In Vitro Flowering of Kalanch?e blossfeldiana Poellniz. II. The Effects of Growth Regulators and Gallic Acid

Nodal explants of Kalanch?e blossfeldiana Poellniz. were culturedin vitro on a low nutrient hormone-free medium. Flowering wasachieved in response to short-day inductive cycles. This systemwas used to test a variety of culture conditions and media onthe flowering response. Plant hormones added to the medium influencedvarious facets of the flowering response when induction occurredby short-day photoperiods. No hormone treatment resulted inflower induction under non-inductive conditions, although vegetativegrowth was affected. The phenolic gallic acid appears not tobe a specific inhibitor of flowering. (Received February 19, 1990; Accepted May 15, 1990)     

Frequency and inheritance of non‐male producing clones in Daphnia magna: evolution towards sex specialization in a cyclical parthenogen?

In Daphnia (Cladocera, Crustacea), parthenogenetic reproduction alternates with sexual reproduction. Individuals of both sexes that belong to the same parthenogenetic line are genetically identical, and their sex is determined by the environment. Previously, non-male producing (NMP) genotypes have been described in species of the Daphnia pulex group. Such genotypes can only persist through phases of sexual reproduction if they co-occur with normal (MP) genotypes that produce both males and females, and thus the breeding system polymorphism is similar to gynodioecy (coexistence of females with hermaphrodites), which is well known in plants. Here we show that the same breeding system polymorphism also occurs in Daphnia magna, a species that has diverged from D. pulex more than 100 MY ago. Depending on the population, between 0% and 40% of D. magna females do not produce males when experimentally exposed to a concentration of the putative sex hormone methyl farnesoate that normally leads to male-only clutches. Natural broods of these NMP females never contained males, contrasting with high proportions of male offspring in MP females from the same populations. The results from a series of crossing experiments suggest that NMP is determined by a dominant allele at a single nuclear locus (or a several closely linked loci): NMP × MP crosses always yielded 50% NMP and 50% MP offspring, whereas MP × MP crosses always yielded 100% MP offspring. Based on cytochrome c oxidase subunit I-sequences, we found that NMP genotypes from different populations belong to three highly divergent mitochondrial lineages, potentially representing three independent evolutionary origins of NMP in D. magna. Thus, the evolution of NMP genotypes in cyclical parthenogens may be more common than previously thought. Moreover, MP genotypes that coexist with NMP genotypes may have responded to the presence of the latter by partially specializing on male production. Hence, these populations of D. magna may be a model for an evolutionary transition from a purely environmental to a partially genetic sex determination system.     

Photoperiod influences reproduction in the prairie vole (Microtus ochrogaster)

Four experiments examined the role of photoperiod in the regulation of seasonal breeding in the prairie vole. Adult male voles maintained in short (8L:16D) as compared to long (16L:8D) photoperiods for 10 wk had reduced testicular and seminal vesicle weights, but fertility was not impaired. Male prairie voles reared from birth until 35 days of age in short as compared to long photoperiods also had reduced testicular and seminal vesicle weights, as well as diminished fertility. The incidence of pregnancy did not differ between long- and short-day female voles paired for 6 days with long- or short-day males (93%, 86%, 89%, and 88%, respectively). Photoperiod did not affect the incidence or the timing of postpartum pregnancies in long- or short-day females paired with long-day males through the birth of several litters. Adult male prairie voles retain only marginal sensitivity to short photoperiods, maturing males are highly responsive to short days, and adult females are insensitive to photoperiod. These data suggest that termination of the breeding season in the autumn may be due to the lack of fecund males in the population.     

The effects of photoperiod on androgen-induced reproductive behavior in male ring doves, Streptopelia risoria

The effect of photoperiod on reproductive behavior in male ring doves (Streptopelia risoria) was studied in androgen-injected castrates maintained under long (16L:8D) or short (86:16D) photoperiods. Behavioral recordings were made over a 2-week period during which each male was paired with a female for 6.5 hr/day. There was some indication that males held on long days display higher levels of courtship activity during the initial period following pairing, but the evidence was not conclusive. Day length had no effect upon copulatory behavior. Long-day males exhibited consistently higher levels of nest building than short-day males, indicating that photoperiod affects building through some mechanism other than changes in endogenous androgen levels.