Delay of sexual maturation in female house mice by exposure to grouped females or urine from grouped females

The experiments examined the timing, duration and possible enhancement effects of group contact on the delay of sexual maturation produced in prepubertal female house mice by urine from grouped females. One or three days of pheromone stimulation at specified ages during the first 2 weeks after weaning was not sufficient to delay puberty in females caged singly. However, pheromone treatment for 7 days, beginning during the first week after weaning, did significantly delay the onset of first vaginal oestrus relative to control females treated with water. Both the timing and duration of pheromone stimulation appear to be critical factors affecting pheromone-induced delay of sexual maturation. Mean ages at first oestrus for females housed with a group of 7 other females, for 3 or 7 days at specified ages during the first 2 weeks after weaning, did not differ from mean ages recorded with urine stimulation only. Contact with other females does not appear to alter or enhance the delay-of-maturation effect achieved with urine stimulation. In all these respects the maturation-delay pheromone of grouped female mice appears to differ from the puberty-accelerating pheromone of male mice.     

Social environment and reproduction in female pouched mice, Saccostomus campestris

Pouched mice were kept under controlled conditions of illumination (10D:14L) and temperature (22 +/- 2 degrees C). Age at vaginal opening, first oestrus and first conception did not differ significantly between juvenile females raised singly, in single-sex groups of 5, or with an adult male. After the introduction of a male, sterile cycles and/or matings before first conception were experienced by all females whether they were raised singly or in single-sex groups. Of 10 females raised with a male, 4 conceived at their first mating. Onset of puberty in juvenile females raised with an adult female was delayed, while in juvenile females raised with their families (mother, father and litter mates) only vaginal opening was delayed. However, when raised in family groups without the father, vaginal opening as well as first oestrus were delayed. Grouping of females after weaning, with or without a male, did not change the oestrous cycle pattern. Females in these groups cycled independently of each other and the females grouped with a male also mated and conceived independently of each other. Pregnancy was blocked in 7 of 8 females when the stud male was removed and a strange male was introduced. In females exposed to a succession of males, pregnancy was blocked up to 4 times.     

Pregnancy in female house mice exposed to urinary chemosignals from other females.

Seven experiments were performed to investigate pregnancy termination, urinary chemosignals, and litter sex ratio variation in female house mice. Experiments tested the effects of urine from adult and prepubertal females, housed individually or in groups, on successful insemination and litter production by females treated at different times and for different periods during the 3 weeks before mating and during gestation. Treatment of females with urine from adult females housed eight per cage or with urine pooled from eight adult females housed individually for 2 or 3 weeks before mating resulted in fewer successful pregnancies and significantly more female-biased litters. Treatment with urine from adult or prepubertal females housed eight per cage or with urine pooled from eight mice housed individually for the first 6 days of gestation or throughout pregnancy resulted in a significant increase in the rate of pregnancy termination. These treatments resulted in lower body weights at birth and slower growth rates in all males and in some females. Puberty was delayed in female progeny from urine-treated dams in five of seven experiments, and these young females attained first oestrus at greater mean body weights than mice in other treatments. These findings indicate that, in mice, at high population density, communication via a urinary chemosignal can alter reproduction in recipient females. Availability of, and competition for, resources such as food would be greater at higher densities, possibly lowering the probability of reproductive success. Pregnancy termination and delays in reproduction and attainment of sexual maturity might lead to greater successful reproduction at a later time.     

Reproduction in a laboratory colony of the pouched mouse, Saccostomus campestris

Pouched mice ovulate spontaneously and have a 4-day cycle (3-5 days). The variation was caused by prolonged oestrus. The vagina opened at about 34 days of age and the first oestrus was experienced at 44 days of age. Females experienced several sterile cycles before their first conception, which occurred at an age of about 56 days. The gestation period in most cases was 21 days and implantation occurred about 4-5 days after mating. The females were not receptive post partum. Litter size varied from 3 to 13 with a mean of 7.1 in young primiparous females and 7.9 in adult multiparous females, indicating that fecundity did not increase with age or parity in this species. Mortality of young was highest during the first 2 days post partum. The young were not attached to the nipples and were weaned at 25 days of age. Females did not cycle during lactation. After lactation most females exhibited one or two oestrous cycles without mating or became oestrous and mated without conceiving, resulting in a litter interval of about 53 days.     

Effects of donor age on urinary chemosignals that influence the timing of first oestrus in young female house mice, Mus domesticus

The effects of donor age on the effectiveness of puberty-influencing urinary chemosignals in wild house mice was tested in a series of 3 experiments. The chemosignal from male mice that accelerates puberty was present in the urine from about the time of puberty and throughout the normal lifespan, but declined about 1 year of age. Oestrous females released a substance in their urine that accelerates puberty in young females. This substance remained effective from first oestrus until over 1 year of age, although older females were in oestrus less frequently than younger mice. Females that are pregnant or lactating released a puberty-accelerating substance in their urine regardless of age. Production and release of the puberty-delaying chemosignal by grouped females was initiated before puberty and continued throughout the lifespan of the mouse.     

Acceleration and delay of first vaginal oestrus in female mice by urinary chemosignals: Dose levels and mixing urine treatment sources

The effects of varying dose levels and mixing of urine from various types of donor mice on the age of sexual maturation in female mice were tested. Over the range from 0.001 ml/day to 0.01 ml/day, there was no difference in the effectiveness of male urine in producing acceleration of puberty, nor was there any difference over the same dose range for urine from grouped females bringing about a delay of puberty. Urine from pregnant and lactating females brought about earlier puberty when applied in the higher dose amounts but was not effective in altering the age of first oestrus relative to untreated controls at lower doses. These findings concerning dose levels are important for a full understanding of the behavioural consequences of urinary chemosignals. When urine from different sources was mixed, all treatments which involved urine from grouped females produced delays in first oestrus. The second finding has important consequences for a feedback model for population regulation in house mice involving urinary chemosignals that accelerate or delay sexual maturation and thus shorten or lengthen generation time by affecting reproductive behaviour.     

Evidence from the oestrous cycle for male-induced ovulation in Bettongia penicillata (Marsupialia).

Female brush-tailed bettongs (Bettongia penicillata) housed in a breeding group of one male and one to three females had an average gestation period of 21.2 days (n = 58) and parturition was followed within 24 h by oestrus and mating. If a young was not born as a result of the mating, oestrus recurred after about 21.7 days (n = 12). From removal of pouch young to birth was 17.5 days, on average (n = 85), in females that had mated post partum, but most females that were isolated from a male before parturition returned to oestrus about 6.6 days after simultaneous removal of pouch young and return to the male (n = 9). Females housed in female-only groups appeared not to come into oestrus or to do so irregularly. These females, when returned to the male, usually came into oestrus within 10 days. These data provide evidence that, in most females of B. penicillata, ovulation does not occur in the absence of a male and that previously isolated females return to oestrus within 10 days of return to the male.     

Reproductive lifespan and reproductive performance in SPF C3H mice: the onset of reproductive life and production efficiency (author's transl)]

To improve the production system, the onset and the termination of reproductive life of C3Hf/HeMsNrs mice mated immediately after weaning and reared for 400 days of life, were studied. From weaning females mated with a full grown male (group A), the first litter was obtained at a mean age of 47 days, suggesting the first copulation at 26 days of age. The age of males at the first copulation was estimated to be at 44 days of age from the age giving the first litters in weanling males mated with weanling (group B) and full grown (group C) females. The sex ratio of litters delivered by young dams tended to be excess in males. The reproductive performance of dams in later life was not affected by the parturition in earlier age. The production efficiency with weaned youngs per pair during the first 200 days after mating was the highest in group A. It was found from these results that the C3H females attained their sexual maturity at 5 to 6 days after weaning, being available for breeding without any deletion in reproductive performance.     

Long-term reversible suppression of oestrus in bitches with nafarelin acetate, a potent LHRH agonist

Adult cyclic beagle bitches were treated for up to 18 months with nafarelin acetate via subcutaneously implanted osmotic pumps, starting during the first week of a pro-oestrous vaginal discharge. The imminent ovulation appeared to be unaffected by treatment, but doses of 8 or 32 micrograms analogue/day reduced the integrated luteal progesterone values. No new oestrus was detected in 3 bitches during 18 months of treatment with 32 micrograms/day, which resulted in mean plasma levels of 0.4 ng analogue/ml. A return to oestrus was observed in all 3 bitches between 3 and 18 weeks after cessation of treatment: 2 of the bitches mated at those times and produced normal litters. Another 2 bitches were similarly treated with 32 micrograms analogue/day; they were mated at the oestrus at start of treatment and dosing was continued for about 63 days. One of the bitches conceived and produced a normal litter. Nafarelin acetate treatment begun during anoestrus resulted in an induced heat 1-2 weeks after the start of treatment. The induced heat consisted of pro-oestrous vaginal discharge, oestrous vaginal cytology, and ovulation (judged by increased circulating levels of progesterone). Three bitches mated at the induced heat and treated for the normal duration of gestation did not litter. Nafarelin treatment of 3 bitches before puberty did not induce signs of oestrus and prevented the occurrence of oestrus through 18 months of treatment. The first oestrus in these bitches occurred 3.5-4 months after cessation of treatment, but mating at that time did not result in pregnancy. These studies have established the feasibility of and dosage requirement for the use of the LHRH agonist as a contraceptive in the bitch.     

The influence of urine from asymptotic laboratory populations on sexual maturation in prairie deermice

Male and female prairie deermice housed singly from weaning (21 days) until 40 days of age had urine from one of the following sources applied daily to their nostrils: sexually mature laboratory population males or females, and sexually mature breeder males or females from our laboratory colony. Other males and females were treated with water. Records were obtained of the weights of the tests, seminal vesicles and bacula of males and the ovaries and uteri of females at 40 days of age. The major results of this study were as follow: Female urine from sexually mature and inhibited population or isolated mice stimulated the development of the male reproductive organs significantly more than water or male urine. Male urine applied to young males did not differentially affect the weights of their reproductive organs compared with water and thus showed no inhibition effects. The reproductive organs of females treated with male urine did not differ significantly in weight from those of females treated with female urine. Females treated with water developed significantly larger uteri than those treated with urine from sexually mature population females (P≤0.02), inhibited population females (P<0.001) and inhibited population males (P<0.007).     

Male-induced puberty acceleration in young female wild mice: hormonal regulation and source of pheromonal cue

Experiments were designed to examine the influence of adult males on the rate of sexual maturation in young female wild mice. In one experiment, young females were raised in presence of adult males, adult females and in absence of any individual, while in another, they were exposed to urines of: (1) castrated males, (2) spayed females, (3) castrated and TP-treated males, (4) castrated and placebo-injected males. Female maturation as measured by age at vaginal opening and first vaginal oestrus was accelerated by presence of adult males, whereas presence of adult females considerably delayed the vaginal opening and the appearance of first oestrus in young females. In the other set of the experiments, urine from castrated or castrated and placebo-injected males was ineffective in inducing early puberty while urine from spayed females highly delayed the sexual maturation. By contrast, urine from castrated and TP-treated males accelerated the puberty more or less like normal males. The results indicate that male's chemosignal accelerating puberty in young females is present in urine and its production is under the control of androgens. However, the female-originating urinary pheromone which delays the puberty in young females is not regulated by ovarian hormones.     

Intermittent stimulation and delay of puberty by urinary chemosignals and social contact in wild stock female house mice (Mus Domesticus)

A sequence of six experiments using wild stock house mouse (Mus domesticus) tested the effects of intermittent stimulation with either the urinary chemosignal released by grouped female mice or social contact from grouped females on the age of first vaginal oestrus in young females. Weanling female mice were exposed to bedding soiled by grouped females or cages containing grouped females for 15 min periods, then removed for a prescribed period, and placed again in a cage with soiled bedding or grouped females. The nature of the exposure to the puberty delaying effect, the number of total exposures each day, the total length of exposure to the stimulus, and the total time period over which the exposures occurred were varied. None of the treatment regimes employed here with soiled bedding from grouped females resulted in delays in the onset of first oestrus in test females. Young females exposed to grouped females for 6 or 8 exposures in a 4 h period, 6 or 8 exposures in an 8 h period, or 8 exposures in a 12 h period were significantly delayed in attaining puberty relative to control females that were exposed to cages containing clean bedding. These results are in contrast to earlier findings involving chemosignals that accelerate first oestrus wherein young females exhibited the capacity to accumulate the exposures to the urinary chemosignals from males, females in oestrus and pregnant or lactating females. Direct exposure to the grouped females on an intermittent basis can provide stimulation that is cummulative and results in delays in the onset of first oestrus.     

Mating induces puberty in the female musk shrew.

Female musk shrews (Suncus murinus), lacking a behavioral estrous cycle, engage in copulatory behavior whenever tested; even during pregnancy, sexual activity can be displayed. Studies suggest that protracted sexual behavior has a functional significance in this species. Virgin females receiving a series of ejaculations over the course of several days are more likely to ovulate and subsequently deliver than animals given the same number of ejaculations over an interval of a few hours. After the virgin mating only one third of the ovaries placed in culture exhibit increased steroidogenesis. In the present set of studies, the hypothesis that the virgin mating induces the onset of puberty, in a manner similar to that in which male-related pheromones induce estrous in rodents, was tested. On the basis of gestational lengths it is evident that females mated three times (on Days 0, 4, and 8) became pregnant in the vast majority of cases as a result of the second and third matings. When the first and second matings were separated by 25 days, 82% of females ovulated in response to the second mating; less than one third of females mated only once ovulated. Finally females were housed across a screen from males, exposed to male urine, or housed alone prior to the virgin mating. In none of these cases did pre-exposure to male-related cues increase ovulation rates in response to the virgin mating. The results show that the virgin mating primes the neuroendocrine system in such a way that the pubertal ovulation can occur in response to subsequent mating.(ABSTRACT TRUNCATED AT 250 WORDS)     

Social influences on reproductive development and fertility in female Djungarian hamsters (Phodopus campbelli).

Social influences on the sexual maturation of female Djungarian hamsters were investigated in two experiments. In the first experiment females were housed from weaning with an adult male, by themselves, or with a weanling sister. Maturation was accelerated in females housed with males as indicated by younger age at first ovulation, increased rates of ovarian and uterine growth, and lower LH levels at some ages. Maturation was delayed in females housed with sisters compared to those housed alone as measured by time of first ovulation and by lower estradiol levels at some ages. The most marked differences between groups occurred 8 to 12 days after weaning, suggesting that events during this period are particularly important in the social mediation of sexual maturation. In the second experiment the effects of reproductive suppression (caused by living with a sister) on the subsequent fertility of females housed with males were examined. If male-female pairs were housed in clean cages, no effects were observed; however, pairs housed in cages previously soiled by the female and her sister had fewer young surviving until 1 week of age despite no differences in the age of pregnancy onset or in the initial litter size. Thus, even cues present in unrenewed soiled bedding may have subtle but long lasting effects on reproductive function.     

Some Factors Influencing Pregnancy Rate and Subsequent Litter Size in Primiparous Sows

The pregnancy rate and the subsequent litter size were studied in 332 Swedish Yorkshire primiparous sows, fed according to a commercial Swedish feeding regime during lactation. The sows were weighed and backfat depth was recorded at the first farrowing, at weaning, and at mating. Oestrous detection was performed once daily after weaning, and the interval from weaning to first oestrus (IWO) was recorded. Blood samples for determination of plasma progesterone were drawn regularly after the first weaning. Statistical analyses were only performed on sows with an IWO of 3-8 days. Of these 206 sows were mated on their first (OE1 sows) and 87 sows on their second (87 OE2 sows) oestrus after weaning. The pregnancy rate was 85.4% for OE1 sows and 75.9% for OE2 sows (p=0.048). There was no significant difference in pregnancy rate between OE1 sows with an IWO of 3-5 days and OE1 sows with an interval of 6-8 days. OE2 sows with an IWO of 6-8 days, on the other hand, had a significantly lower pregnancy rate compared with OE2 sows with an interval of 3-5 days. The pregnancy rate in sows that lost more than 30 kg during the first lactation period did not differ from that of sows losing less than 30 kg. In sows with a first litter size of more than 9 piglets alive at birth, the pregnancy rate decreases significantly if mating is delayed until the second oestrus after weaning. OE2 sows had a significantly larger second litter size at birth than OE1 sows (+ 2.0). The litter size at six weeks did not, on the other hand, differ significantly (+ 0.4). There was a positive correlation between the IWO and 2nd litter size, although significant only for OE1 sows between the IWO and litter size alive at birth. In the OE1 group, sows losing 20 kg or less during lactation had significantly larger second litters at birth than the sows losing 21-30 kg, but not significantly larger than the sows losing more than 30 kg. One piglet more, at birth, in the first litter resulted in 0.25 piglet more in the second litter. For sows with a large first litter there was a low probability of also having a large second litter.     

The influence of conditions at the time of mating on reproduction of commercial pigs

The objective of this study was to compare a new mating system, called the Detection-Mating Area (DM Area), and a conventional mating system on the long-term reproductive performance of commercial pigs. The DM Area treatment basically involved detecting oestrous females in an arena closely surrounded on two sides by boars and mating these females in this arena. This mating system was designed to improve the physical and sexual environments of the pigs at mating. In contrast, the conventional treatment involved conducting oestrus detection and mating in the boar's accommodation pen. The study was conducted over an 18-month period at a commercial farm that housed 2400 breeding female pigs.

In order to control for the effects of the stockperson, an analysis was conducted on the reproductive performance of female pigs in which one stockperson assisted the matings in both treatments over a 12-month period. Gilts mated in the DM Area treatment had a higher (P<0.05) total litter size and a higher (P<0.01) litter size born alive than gilts mated in the conventional treatment (10.31 vs. 8.96 and 9.50 vs. 8.29, respectively). Although gilts in the DM Area treatment had a higher farrowing rate (93.2 vs. 87.9%), this difference was not significant (P>0.05). There were no significant (P>0.05) differences found between the reproductive performance of sows in the two treatments; however this comparison was confounded by sows in the DM Area treatment having a lower number of matings per oestrus than those in the other treatment. Observations on the sexual behaviour of pigs at 145 matings indicated that the boars in the DM Area treatment displayed a higher (P<0.05) number of bouts of courting behaviour directed towards the female than boars in the conventional treatment (8.3 and 6.4, respectively). These very limited observations on sexual behaviour suggest that changes in the courting behaviour of bears may be associated with changes in litter size. While there was some indication from the results of progesterone analysis of blood samples taken from unmated gilts that there may have been some differences between treatments in the sexual age of gilts at matings, these differences are unlikely to explain the differences in litter size between treatments. Further research is required to identify the component (s) of the DM Area treatment that are responsible for the improvement in litter size in gilts and to further examine the effects of the two treatments on the reproductive performance of sows.     

The production of and sensitivity to cues that delay puberty and prolong subsequent oestrous cycles in female mice are influenced by prior intrauterine position

Four experiments were conducted with CF-1 house mice (Mus domesticus) to examine the relationship between a female's prior intrauterine position (2M = between 2 male fetuses, 1M = next to one male fetus, 0M = not next to a male fetus) and the timing of puberty and length of subsequent oestrous cycles under a variety of housing conditions. The results of Exp. 1 confirmed that the presence of males was required for females to enter puberty and exhibit regular oestrous cycles, regardless of prior intrauterine position. When housed individually after weaning either with a male in the cage or separated by a wire mesh partition, 0M-females ovulated and mated at a younger age and had shorter post-pubertal oestrous cycles than did 2M-females (1M-females were intermediate between 0M- and 2M-females). Other females were also housed after weaning with a male or separated by a wire mesh partition from a male in a variety of social environments (4 0M-females and 1 2M-female, 4 2M-females and 1 0M-female, 5 1M-females, 3 0M-females and 3 2M-females). The objective of these different housing conditions was to determine whether prior intrauterine position influenced the transmission of and/or sensitivity to cues that delay puberty and prolong subsequent oestrous cycles. Relative to 2M-females, 0M-females both produced more potent cues and were more sensitive to the cues (again, 1M-females were intermediate between 0M- and 2M-females). Specifically, in the presence of other females, puberty was delayed and post-pubertal oestrous cycles were prolonged to the greatest extent in 0M-females. Prior intrauterine position is therefore a source of individual variation in the production of and sensitivity to cues that modulate the timing of puberty and the length of subsequent oestrous cycles in female mice. The findings suggest that prenatally-androgenized 2M-females may have a reproductive advantage over other females at high population densities.     

Methoprene-induced matings of young Queensland fruit fly males are effective at inducing sexual inhibition in females

Pre-release dietary treatment with methoprene, a juvenile hormone analogue, decreases the age at which male Queensland fruit flies mature and hence may decrease the post-release delay until released sterile flies participate in sterile insect technique (SIT) programmes. However, if matings of young methoprene-treated males are not effective at inducing sexual inhibition in their mates, then this treatment may not enhance SIT. The present study investigates efficacy of matings of methoprene-treated males at inducing sexual inhibition in their mates. Methoprene incorporated into a diet of sugar and yeast hydrolysate (w/w 3:1) for 48 hr after emergence resulted in significantly increased male mating propensity when flies were <10 days of age, but not when older, and longer copulations. Copula latency did not vary with methoprene treatment but did decrease with age. The matings of young methoprene-treated males were effective at inducing sexual inhibition in their mates, matching the efficacy of untreated mature males. Regardless of treatment, females had reduced tendency to remate if their first mate was 15 days of age than if their first mate was younger (6, 8 days) or older (20, 25, 30 days). Females mated by methoprene-treated males that did remate tended to remate later in the day than females mated by untreated males. Also, second copula durations of females first mated by a 6- to 10-day-old male were shorter if the male was methoprene treated. These patterns in remating females may indicate greater efficacy of the initial mating of methoprene-treated males. Overall, we find that the additional matings of young methoprene-treated male Queensland fruit flies are effective at inducing sexual inhibition in their mates. This finding supports the incorporation of methoprene into pre-release diet for SIT.     

Timing of lactational oestrus in intermittent suckling regimes: consequences for sow fertility

Three intermittent suckling (IS) regimes were evaluated for their effects on lactational oestrus and subsequent fertility. Control sows were weaned (CW; n = 38) at d 26 ± 2 of lactation. In IS19-7D (n=40) and IS19-14D (n=42) sows, IS started at d 19 ± 1 of lactation and sows were weaned 7 or 14 d later. In IS26-7D (n=41), IS started at d 26 ± 1 of lactation and sows were weaned 7d later. During IS, sows were separated from their piglets for 10h/day. Oestrus detection was performed twice daily without a boar and ovulation was confirmed by ultrasound once a week. In IS19-7D, IS19-14D and IS26-7D, respectively, 50%, 64% and 61% of the sows showed oestrus and ovulation during IS (P>0.05), and, of the remaining sows, 100%, 93%, and 69% showed oestrus in the first week after weaning. In CW sows, 95% showed oestrus in the first week after weaning. Parity 1 sows were considerably less likely than older parities (23% vs. 68%) to show oestrus in lactation. Pregnancy rate of the first post partum oestrus (during lactation or after weaning) was 89% (CW), 92% (IS19-7D), 80% (IS19-14D) and 77% (IS26-7D) (P>0.05) and subsequent litter size was 14.5 ± 0.5, 14.5 ± 0.6, 15.3 ± 0.5 and 15.2 ± 0.8, respectively (P>0.05). Sows mated during lactation had similar pregnancy rate and litter size to those mated after weaning. Hence, ongoing lactation for the first 2-9 d of pregnancy did not negatively affect fertility. A total of 50-64% of IS sows showed lactational oestrus, regardless of the stage of lactation. Pregnancy rates and litter size were similar to control sows, and were not affected by stage of lactation at mating.     

Effect of age at first mating on primary sex-ratio of the two-spotted spider mite

This paper determines the influence of initial age at mating on the primary sex-ratio of the two-spotted spider miteTetranychus urticae Koch. The specific question asked is whether females that mate at ages beyond ecdysis, or are inseminated at ecdysis by virgin males of set ages, have daughter productions different from control females mated at ecdysis by randomly aged males.Age of initial female mating did not effect survival. Females mated at ecdysis produced more daughters than those mated at age 5 or 10 days. However, daughter production was equal for all female treatments in the post-mating period. All treatments stopped producing daughters at approximately the same age, regardless of when mated. Son production was lowest for females mated at ecdysis. However, son production by females mated when 5 or 10 days old was lower than that of females mated at ecdysis.Male age at mating did not effect female survival. Females exposed to virgin males either 1 or 7 days old produced more daughters than those exposed to males either 11 or 20 days of age. However, a substantial proportion of females in these latter cohorts were not mated. If only mated females are considered, daughter production is the same between all cohorts.Delayed age at first mating influenced sex-ratio only via the females, specifically by changes in daughter production (as opposed to changes in survivorship). Daughters that would have been produced between ecdysis and mating were never recovered by females that mated at ages beyond ecdysis. Thus, these daughters were lost from a sex-ratio viewpoint. Initial age of male mating did not change sex-ratio.