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Animal Behaviour 78 (2009) 747–753

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Animal Behaviour

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Potential reproductive rate of a sex-role reversed pipeﬁsh over several bouts of mating

Sunny K. Scobell*, Adam M. Fudickar 1, Rosemary Knapp 2

Department of Zoology, University of Oklahoma article info The potential reproductive rate (PRR, the rate at which each sex could reproduce if given unlimited Article history: mates) has proven to be a useful tool in predicting the direction and strength of sexual selection. We Received 5 February 2009 conducted a 2-month study of the PRR in the polyandrous gulf pipefish, Syngnathus scovelli, a year-round Initial acceptance 19 March 2009 breeder. In this sex-role reversed species, the female transfers eggs to a male’s brood pouch during Final acceptance 28 May 2009 mating and thus renders him unavailable to mate for 2 weeks. We predicted females would have a higher Published online 4 August 2009 PRR than males and that the rates in both sexes would change over successive breeding bouts in rela- MS. number: A09-00073 tionship to previous reproductive output. Females did have a higher overall PRR than males for the entire study period. However, PRR was not constant across individual breeding bouts. For each sex, the PRRs Keywords: from the first and third bouts of mating were significantly higher than the PRR of second mating bout. gulf pipefish Our results are consistent with individuals making trade-offs between current and future reproductive life-history theory investment. We also discuss how ovarian morphology may contribute to elevated female PRR in this ovarian morphology pipefish species. To our knowledge, this is the first study of PRR in a North American pipefish. potential reproductive rate The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. sex-role reversal sexual selection Syngnathidae Syngnathus scovelli

When studying mating behaviour, researchers usually want to However, PI (defined as any effort that increases offspring survival know which sex is under greater sexual selection pressure and to at the expense of the parent’s ability to invest in other offspring) can what degree. This information is useful in determining the sex that be difficult to measure, particularly in species where both sexes should be competitive for mates and the sex that should be choosy. have some form of investment (Clutton-Brock 1991). Emlen & Oring The strength of sexual selection acting on each sex can also be (1977) proposed that a bias in the operational sex ratio (OSR) away correlated with territoriality, the degree of sexual dimorphism and/ from 1:1 could result in an increase in the intensity of sexual or ornamentation and variance in mating success (Andersson selection on the sex that is more abundant. The OSR, which is the 1994). However, measuring the strength of sexual selection acting average ratio of sexually active females to males in a population at on each sex has been difficult historically, and determining which any given time, has been effective in estimating the opportunity for methodology is best has been the subject of debate. sexual selection in several species (Vincent et al. 1994; Kvarnemo & Many researchers have measured sex differences in parental Ahnesjo¨ 1996; Dearborn et al. 2001; Jones et al. 2001). However, investment and/or the operational sex ratio, two indirect measures obtaining an accurate OSR for a population may not be feasible for of the strength of sexual selection. Trivers (1972) proposed that the a particular species being studied and could also vary greatly sex that has relatively greater parental investment (PI) should be temporally (Forsgren et al. 2004), which could in turn have conse- choosy, leaving the sex that has lower PI to compete for mates. quences for conclusions about the strength of sexual selection. In situations where it is not practical to estimate the PI or OSR, the potential reproductive rate can be used to predict the intensity * Correspondence and present address: S. K. Scobell, Department of Biology, of sexual selection. The potential reproductive rate (PRR) is the rate Texas A&M University, 3258 TAMU, College Station, TX 77843-3258, U.S.A. at which each sex in a population could reproduce if given unlim- E-mail address: [email protected] (S.K. Scobell). ited mates (Clutton-Brock & Vincent 1991) and assumes no differ- 1 A. M. Fudickar is now at the Department of Migration and Immuno-ecology, ential immigration or mortality between the sexes (Kokko & Max Planck Institute for Ornithology, Radolfzell, Germany. 2 R. Knapp is at the Department of Zoology, University of Oklahoma, 730 Van Monaghan 2001). The PRR has proven to be a useful tool in pre- Vleet Oval, Room 314, Norman, OK 73019, U.S.A. dicting the direction and strength of sexual selection (fish:

0003-3472/$38.00 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. doi:10.1016/j.anbehav.2009.05.036 Author's personal copy

748 S.K. Scobell et al. / Animal Behaviour 78 (2009) 747–753

Kvarnemo 1994; Kvarnemo & Ahnesjo¨ 1996; Masonjones & Lewis The male fertilizes the eggs and then broods them for about 14 days 2000; Ahnesjo¨ et al. 2001; Wilson 2009; insects: Kvarnemo & (Brown 1972). Thus, male gulf pipefish contribute significantly more Simmons 1998; Wiklund et al.1998; mammals: Preston et al. 2005). time to parental care than do females. The gulf pipefish is also It is usually measured under controlled laboratory conditions sexually dimorphic and has a polyandrous mating system. Females where competition for mates is eliminated and environmental are typically larger than males and have a sexually dimorphic variables are kept constant (i.e. temperature, food, shelter, lack of ornament, a silver bar on each bony ring of the trunk (Reid 1954). predation, etc.). Thus, the PRR controls for many of the confounding Jones et al. (2001) found that mated females in the field had a larger variables that can influence reproductive function and gives an mean snout–vent length, body depth and body mass and were more estimate of the baseline reproductive output for each sex under ornamented than unmated females. They also found that the stan- a similar, constant environment. dardized variance in mating success as measured by microsatellite PRR is typically measured in one of two ways: the number of markers was at least seven times greater in females than in males. offspring (or potential offspring) produced per unit time (Clutton- Despite morphological and genetic evidence supporting Brock & Vincent 1991; Ahnesjo¨ et al. 2001; Wilson 2009) or the a female-biased PRR, it is not known whether male and female gulf ‘time in/time out’ model (Clutton-Brock & Parker 1992). Clutton- pipefish do indeed differ in their respective potential reproductive Brock & Vincent (1991) defined the PRR as the maximum number of rates. If the assumptions underlying sexual selection theory are offspring that each sex produced per unit time. Later, Ahnesjo¨ et al. correct, they should be supported both in species with conven- (2001) measured PRR by counting the number of ‘potential’ tional mating systems and in those that are sex-role reversed offspring produced per day over the course of the breeding season. (Clutton-Brock 2009). The present study was designed to test the They argued that the female’s PRR should be based on the number hypothesis that the sex differences in body size, ornamentation and of eggs released during mating and that the male’s PRR should be parental roles of this species are accompanied by a sex difference in based on the number of eggs fertilized. For the ‘time in/time out’ PRR. The 2-month study period allowed us to measure PRR over model, Clutton-Brock & Parker (1992) defined ‘time out’ as the time multiple breeding bouts. We predicted that females would have period in which an adult is not able to mate, and they defined ‘time a higher PRR than males and that the rates in both sexes would in’ as the time period when an animal is able to mate if given change over successive breeding bouts in relationship to previous a receptive partner. Both methods (‘time in/time out’ and number of reproductive output. offspring/unit time) have proven useful and which method is used often depends on the mating system of the species being studied. METHODS The PRR has been used to study the mating system of several species of syngnathids (pipefish and sea horses). The family Syn- Animals gnathidae is unique among fishes in that male pregnancy is found in all species and some, but not all species, are sex-role reversed We collected sexually mature S. scovelli adults (N ¼ 116) from (Vincent et al. 1992). In species where the PRR of females exceeds sea grass beds in Sarasota Bay, Florida on 6 June 2004. Sexually that of males, the mating system is typically sex-role reversed and mature males have a developed ventral brood pouch and females females compete for access to males. Nerophis ophidion and Syn- have complete silver bars on the trunk (Brown 1972; Jones & Avise gnathus typhle females were estimated to be able to fill the pouch of 1997). Pipefish were collected with push nets (1 m2, with 2 mm2 one male completely and still have a 41% surplus of eggs remaining mesh) at a depth of roughly 1 m. Fish were shipped overnight to the to give to another male (Berglund et al. 1989). Ahnesjo¨ (1995) later University of Oklahoma where they were housed in temporary, calculated that S. typhle had a female-biased of PRR 1.8:1 (females nonreproductive groups in aquaria that did not share a common to males) in warm water and 2.3:1 in cold water. Both N. ophidion water system. Most fish were allowed to acclimate to laboratory and S. typhle are pipefish that have sex-role reversed mating conditions for 10 days before the study began. Five focal females systems in which females compete for access to males (Berglund & spent an additional 10–18 days acclimating while we waited for Rosenqvist 2003). In species where the PRR of males and females a sufficient number of field-pregnant males to return to breeding approaches unity, the mating system is typically monogamous with condition to pair with them. conventional sex roles (but see Sogabe & Yanagisawa 2007). Vin- cent et al. (1994) were the first to show that despite their preg- Animal Housing nancy, male Sri Lankan sea horses, Hippocampus fuscus, had a higher PRR and displayed more competitive and courtship Fish were housed in 75-litre aquaria divided with opaque behaviour than females. Similarly, Masonjones & Lewis (2000) Plexiglas either in half or into a small (18.75-litre) and a large found that dwarf sea horses, Hippocampus zosterae, have a PRR of (56.25-litre) compartment (depending on treatment, see below). 1:1.2 and also show a conventional, monogamous mating system. Aquaria contained both biological sponge filters and undergravel However, the PRR does not always predict a species’ mating filters and were supplemented with saltwater bacteria once per behaviour. Sogabe & Yanagisawa (2007) studied the PRR of the week. Aquaria contained crushed coral and sand substrate, plastic monogamous messmate pipefish, Corythoichthys haematopterus, plants that mimic natural sea grass (Thallassia sp.) and 15 cm long and found that there was no difference in PRR between the sexes, by 3 cm wide sections of grey PVC tubing cut in half for cover. Fish but females were more active in courtship than males. The were fed newly hatched and adult Artemia enriched with Selco observed behavioural differences were associated with a female- (Aquatic Ecosystems, Apopca, FL, U.S.A.) twice daily. The feeding biased OSR for this population. Further studies are needed in this regime was sufficient to maintain a body mass to total length (TL) family to determine how PRR is related to the mating system. ratio that was similar to that found in the field (S.K.S., unpublished We conducted a study of the PRR of the sex-role reversed gulf data). Fish were kept at 25 C and on a 13:11 h light:dark cycle to pipefish, Syngnathus scovelli, over a 2-month period. The gulf pipe- mimic natural conditions in Sarasota Bay during the summer. fish (Evermann & Kendall 1896) is a common syngnathid found throughout the Gulf of Mexico and breeds year round in Florida, Study Design U.S.A. (Reid 1954; Joseph 1957; Brown 1972). Gulf pipefish males, like many members of the family Syngnathidae, have a pouch into We conducted a 61-day study where we provided focal male which the female deposits her eggs (her only known contribution). and female gulf pipefish with a surplus of mates and counted the Author's personal copy

S.K. Scobell et al. / Animal Behaviour 78 (2009) 747–753 749 number of potential offspring each sex could produce. At the start Two focal females did not mate during the study, probably of the study, 12 male and 12 female focal subjects were chosen to because of reproductive suppression (regression of the gonads in represent the range of body sizes for adult breeding individuals in group-housed females; e.g. Rosenqvist 1990) during acclimation, the population (TL ranges: males 119–145 mm, N ¼ 68; females and we removed them from analyses. This left 10 females in all 112–169 mm, N ¼ 48). Focal animals’ TL (134.7 2.6 mm) did not analyses with the exception of analyses involving gonad mass. For differ from that of the population sample (132.4 1.2 mm; Mann– these analyses, sample sizes were reduced (males: N ¼ 10, Whitney U test: U ¼ 1067.0, N1 ¼ 24, N2 ¼ 92, P ¼ 0.58). Focal females: N ¼ 7) because of missing gonad data for several animals were provided with similarly sized (within approximately animals. 5 mm TL) opposite-sex partners because size-assortative mating The methods used in this study comply with the current laws of has been documented in several species of syngnathids (Vincent & the United States of America, in which the studies were performed, Sadler 1995; Jones et al. 2003; Silva et al. 2008). and were approved by the University of Oklahoma Institutional The number of partner males or females that each sex could Animal Care and Use Committee (R02-013). mate with in 1 day was determined in a pilot study in the autumn of 2003. Females transferred eggs to the pouches of a maximum of Analyses two males per day; males only accepted eggs from one female on the first day of the pregnancy. In the current study, to ensure that We calculated the PRR by measuring the potential offspring focal animals had a surplus of mates, each focal female was housed produced per unit time. The gulf pipefish has a courtship period with three breeding males and each focal male was housed with that can be extremely short. Under laboratory conditions, the time two breeding females. Focal males were housed in larger from introduction of a partner to mating can be less than 1 min. compartments (56.8 litres) than focal females (37.5 litres) to reduce Thus, individual mating events can be easily missed without 24 h animal density and thus reduce female–female competition observation, which was not logistically possible in this study. For (between a focal male’s two partner females; S.K.S., personal each subject, we calculated total fecundity over the study period. observation) as this female competition was not the focus of the To obtain the best measure of the focal animal’s fecundity for present study and could have interfered with assessment of PRR. each pregnancy, we compared the mid-pregnancy counts to the On the four occasions when a focal male failed to get pregnant, one pouch contents following birth, and then used whichever count of his partner females was removed (the suspected subordinate) was higher in our analyses. This approach minimized the error of and replaced with a new partner female. This usually resulted in the mid-pregnancy counts where some eggs, or occasionally an a new pregnancy the next day (N ¼ 3). Male–male aggression has entire row of eggs, could be occluded from the scorer’s view. We not been observed in this species, so we were able to house the calculated the PRR as each animal’s fecundity/day. However, as focal females and their partner males in smaller compartments PRR is a measure of an individual’s potential offspring, we than those used for the focal males and their partner females. As calculated the PRR for females as the number of eggs transferred/ the study progressed and the number of available breeding partner day (for mid-pregnancy count) or all pouch contents (for count at males became limited (due to many males being pregnant), females birth), and for males, we calculated the PRR as the number of were housed with at least two breeding males, the maximum fertilized eggs/day (for mid-pregnancy count) or newborn þ number of males that a female could mate with in one day. underdeveloped young (for count at birth), whichever was greater. All focal and partner males were checked daily between 1600 We used Student’s t test to determine whether there was a sex and 2000 hours for new pregnancies. Pregnancies were easily difference in mean PRR between males and females for the entire observed as the orange eggs are clearly visible though the skin of study period. the male’s pouch. The day that the pregnancy was detected was For the repeated measures analysis, we divided the PRR data designated day 0. Pregnant focal males remained in their home over the 61-day study into four breeding bouts. A breeding bout tank with both females. Partner males, after becoming pregnant, was defined as the time that it took for males to complete a preg- were moved to an adjacent 37.5-litre compartment, and a new nancy (from mating to birth and thus return to breeding condition, partner male in breeding condition was placed in with that focal N ¼ 12, mean SE ¼ 15.0 1 days). The PRR for each of the four female. For all pregnant males (focal and partner), at mid-preg- breeding bouts was compared for males and females using nancy we counted the eggs transferred by the female, and then, repeated measures mixed model analysis (see Brown & Prescott following birth, we counted the total number of offspring. To obtain 1999). Because most populations of S. scovelli show sexual size the mid-pregnancy count, we counted the number of fertilized eggs dimorphism (Reid 1954; Brown 1972; Jones et al. 2001), we used and any unfertilized eggs on or around day 7 of the pregnancy; at a principal components analysis (using PC-ORD, MJM Software this time, embryos are pigmented and fertilized eggs can be Design, Gleneden Beach, OR, U.S.A.) to obtain a composite score for distinguished from unfertilized eggs. The males were lightly each focal animal’s body mass, depth, TL and SVL. Axis 1 explained anaesthetized using a 0.05% solution of 2-phenoxyethanol (2-PE) 91.6% of the total variance in focal animals’ body size. We fitted the and the contents of the pouch were counted through the trans- body size PCA axis 1 scores and subjects (nested within sex) as parent skin folds. On day 12 of pregnancy, pregnant males were random, so the effects of sex, breeding bout, PCA, and the interac- placed in breeding nets containing a plastic plant and a PVC shelter tion between sex*bout and sex*PCA on PRR could be assessed in their home aquaria. Breeding nets and the male’s pouch were across focal animals. We used least squares means with post hoc checked daily to determine whether the male had given birth. On Tukey–Kramer tests for sex and breeding bout. the day of birth, the pouch contents (newborn, underdeveloped To assess the relationship between morphological measure- (dead) offspring and unfertilized eggs) were counted, newborn ments and PRR, we used linear regression analysis to determine were euthanized with an overdose of 2-PE, and all pouch contents whether TL, somatic mass (total mass gonad mass) or gonad were then stored in 10% buffered formalin. After the 61-day study mass affected the PRR for each sex. We used correlation analysis to period, we recorded TL, snout–vent length (SVL), body depth just determine the relationship between TL, somatic mass and gonad anterior to the dorsal fin, and total body mass for the focal male and mass for each sex. females. We euthanized focal males and females with an overdose The mixed model analysis was conducted using SAS 8.01 (SAS of 2-PE and dissected the gonads, which we preserved in 10% Institute, Inc., Cary, NC, U.S.A.). All other analyses were conducted formalin and later weighed to the nearest 0.1 mg. using SPSS 15.0 (SPSS, Inc., Chicago, IL, U.S.A.). Author's personal copy

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RESULTS because focal males were pregnant within 2.0 0.7 days of giving birth (median ¼ 0.0 days; most males got pregnant the day after Overall PRR giving birth). Females had a mean PRR 1.95 times greater than that of males over the entire study period of 61 days. Focal females had Females mated on average 2.8 times more often than males over a mean PRR of 6.5 1.1 offspring/day, whereas focal males had the course of our study (mean SE; females: 11.3 1.2, males: a mean PRR of 3.4 0.3 offspring/day (Student’s t test: t20 ¼2.99, 4.1 0.2; Mann–Whitney U test: U ¼ 0.0, N1 ¼ 10, N2 ¼ 12, P ¼ 0.01; Fig. 1c). P < 0.001; Fig. 1a). Accordingly, females had a shorter mean interbrood interval (number of days between matings ¼ 6.3 0.8 days) PRR over Four Breeding Bouts than males (16.1 0.6 days, Student’s t test: t20 ¼ 10.09, P < 0.001; Fig.1b). The males’ longer interbrood interval was due mainly to the The mean PRR of males and females did not remain constant length of pregnancy, rather than limited by female behaviour, over the four breeding bouts (Fig. 2). A repeated measures mixed model analysis showed that the interactions of sex*bout and sex*PCA were not significant (sex*bout: F3,57 ¼ 0.52, P ¼ 0.67, 14 sex*PCA: F2,17 ¼ 2.18, P ¼ 0.14); these terms were removed from the (a) * model and it was rerun. The main effects of sex and breeding bout 12 both affected PRR (sex: F1,20 ¼ 21.79, P ¼ 0.0001; bout: F3,63 ¼ 3.10, P ¼ 0.03). Post hoc analyses showed that the PRR of breeding bout 1 10 was significantly greater than the PRR for breeding bout 2 (least squares means with Tukey–Kramer adjustment: t63 ¼ 2.92, 8 P ¼ 0.005). The decrease in PRR from the first to second breeding bout tended to be greater for females (4.0 1.4) than males 6 (1.9 0.7) (Student’s t test: t20 ¼ 4.28, P ¼ 0.052). The PRR for breeding bout 2 was also significantly lower than the PRR for 4 breeding bout 3 (t63 ¼2.21, P ¼ 0.03). However, the change in PRR Number of pregnancies Number between the second and third breeding bouts did not differ 2 significantly between females (3.2 1.0) and males (1.3 0.6) (Student’s t test: t20 ¼ 0.70, P ¼ 0.41). 0 For both male and female focal animals, we noticed a common pattern of fecundity over the four breeding bouts. Most individuals 18 had high fecundity during the first and third breeding bouts, but (b) 16 * much lower fecundity in the second breeding bout (see Fig. 3). Not all individuals showed this pattern, but all showed at least one 14 ‘boom’ and then ‘bust’ in fecundity over consecutive breeding bouts 12 during the study.

10 PRR and Body Size 8 Although the ranges of male and female body size measure- 6 ments overlapped for the focal animals in this study, females in our sample, as in other studied populations (Reid 1954; Joseph 1957; Interbrood interval (days) 4

0 12 P = 0.005 P = 0.03 8 10 (c) *

8 6

4 4 Mean PRR (offspring/day) PRR (offspring/day) 2 2 1.90:1 1.77:1 2.05:1 2.03:1 0 12345 0 Females Males Breeding bout

Figure 2. Potential reproductive rates (PRR) for female (C) and male (B) gulf pipeﬁsh Figure 1. Mean SE (a) number of pregnancies, (b) interbrood interval and (c) over four breeding bouts (mean SE). The ratio of female to male PRR is shown for potential reproductive rate (PRR) of female and male gulf pipeﬁsh over the 61-day each breeding bout. Brackets and associated P values denote differences in the PRR study. N ¼ 10 females, N ¼ 12 males. *P < 0.01. between the indicated time periods. Author's personal copy

S.K. Scobell et al. / Animal Behaviour 78 (2009) 747–753 751

140 16

120 14 12 100 10 80 8 60 6

40 PRR (offspring/day) 4

20 2 Fecundity (offspring/pregnancy)

0 0 21 Jun 2004 5 Jul 2004 19 Jul 2004 2 Aug 2004 50 100 150 200 250 300 Date of mating Ovary mass (mg) Figure 4. Potential reproductive rate (PRR) versus ovarian mass for focal gulf pipefish Figure 3. Representative fecundity data from an individual female and an individual females. male gulf pipefish. Each data point represents a single pregnancy over the course of the study. Female 19 (C) and male 11 (B) best exemplified the ‘boom’ and ‘bust’ mating patterns observed in the focal animals as well as the sex differences in mating court males and compete with each other for access to mates (Jones frequency. & Avise 1997; Clutton-Brock 2009), as we have observed in the laboratory. Brown 1972), were generally larger than males (Table 1). However, Although it has been suggested previously (Ahnesjo¨ et al. 2001), neither female TL (linear regression: R2 ¼ 0.01, P ¼ 0.81) nor the present study is, to our knowledge, one of the few studies to somatic mass (R2 < 0.01, P ¼ 0.91) was related to the PRR. Ovarian measure PRR over several breeding bouts in a vertebrate. The PRR of mass also did not correlate with female TL (Spearman rank corre- gulf pipefish males, and particularly females, varied over the four lation: rS ¼ 0.07, N ¼ 7, P ¼ 0.88) or somatic mass (rS ¼ 0.11, N ¼ 7, breeding bouts. The mean PRR was highest for both sexes during P ¼ 0.82), but females with a greater ovarian mass also had a higher the first breeding bout. Then, both sexes showed a decrease in PRR (R2 ¼ 0.77, P ¼ 0.01) (Fig. 4). For males, testes mass increased mean PRR during the second breeding bout. This general pattern of with both TL (Spearman rank correlation: rS ¼ 0.94, N ¼ 10, a boom followed by a bust in reproductive investment was seen for P < 0.01) and somatic mass (rS ¼ 0.95, N ¼ 10, P < 0.01). However, most of our focal animals. The change in male and female PRR over larger males and males with larger testes did not have a greater PRR the study is consistent with life-history theory predictions that (TL: R2 < 0.01, P ¼ 0.82; somatic mass: R2 < 0.01, P ¼ 0.99; testes individuals that mate multiply make trade-offs between current mass: R2 < 0.01, P ¼ 0.96). and future reproductive investment given a limited lifetime energy budget and potential availability of mates (Stearns 1992; Zera & DISCUSSION Harshman 2001; Roff 2002; Heubel et al. 2008). However, it is also possible that this pattern of reproduction was a result of the labo- Our study documented that the PRRs of male and female gulf ratory environment. Future studies of this species in the field and in pipefish differ, with females having an overall PRR almost twice the laboratory will help clarify the factors that contribute to the that of males. As in other sex-role reversed species of syngnathids variation in PRR that we observed across multiple bouts of mating. (Berglund et al. 1989; Kvarnemo & Ahnesjo¨ 1996), female gulf Saltwater populations of gulf pipefish generally show sexual size pipefish are able to mate more often and with a shorter interbrood dimorphism (Reid 1954; Joseph 1957; Brown 1972). Our sample interval than are males. The gulf pipefish mating system is from the Sarasota Bay population also showed sexual size dimor- consistent with sexual selection theory in that the sex that has the phism, with females, on average, larger than males. Larger female higher PRR, females, also has the higher variance in mating success body size is common in fish and is often related to a fecundity (Jones et al. 2001) and is the more ornamented sex (Reid 1954; advantage (Andersson 1994). When each sex was examined indi- Brown 1972; Jones et al. 2001; Clutton-Brock 2009). Assuming a 1:1 vidually, neither male or female total length nor somatic mass adult sex ratio and no differential mortality, we predict that gulf affected the PRR. However, males and females did differ with pipefish have a female-biased OSR in the field (Jones & Avise 1997; respect to the relationship of gonad size to PRR. Females that had Ahnesjo¨ et al. 2001; Jones et al. 2001) and, accordingly, that females larger ovaries had a higher PRR, whereas males with larger testes showed no concomitantly greater PRR. A female with larger ovaries probably has a greater fitness advantage over a female with smaller Table 1 ovaries. Females with larger ovaries can transfer more eggs, and Comparison of morphological measurements of focal male and female gulf pipefish because males in this species typically only accept eggs from one following the study period of 61 days (mean SE) female (Jones & Avise 1997; Jones et al. 2001), they would be pre- Females Males tP dicted to choose the female that can transfer the greater number of Sample size (N)1012 eggs (Rosenqvist 1990). As a result of male pregnancy, males with Snout–vent length (mm) 59.82.0 49.41.0 5.01 <0.01** bigger testes probably do not gain a significantly greater fitness Total length (mm) 140.24.8 127.62.5 2.44 0.02* Depth of trunk (mm) 7.20.3 5.50.2 4.45 <0.01** advantage over males with smaller testes. Males in this species are Body mass (g) 2.30.2 1.80.1 1.98 0.06 thought to fertilize the eggs once they are in the male’s pouch, Gonad mass (mg)y 116.930.9 5.40.5 4.37 0.01* similar to fertilization in S. schlegeli (Watanabe et al. 2000). There *P < 0.05; **P < 0.01. has been no record of multiple paternity in S. scovelli (Jones & Avise y Sample size for gonad mass: females N ¼ 7, males N ¼ 10. 1997; Jones et al. 2001). Thus, there appears to be no sperm Author's personal copy

752 S.K. Scobell et al. / Animal Behaviour 78 (2009) 747–753 competition within the brood pouch and sperm probably do not Stanley and Dave Jenkins at the Mote Marine Laboratory for assis- have to fertilize eggs in the external environment. This speculation tance with animal collection and aquaculture techniques. Funding is also supported by the very small size of testes in this species. Our to S.K.S. from the University of Oklahoma’s Graduate Student results are consistent with sexual selection acting more strongly on Senate, the Department of Zoology’s Adams Memorial Scholarship, female ovary size, with relaxed selection on male testis size Sigma Xi and the PADI Foundation supported this study. This study (Clutton-Brock 2009). also benefited from the funds awarded to R.K. from the University The difference in PRR between the sexes in gulf pipefish is of Oklahoma Research Council. presumably influenced by the differing constraints of their reproductive physiology. Males are constrained by the time that it takes to brood offspring, whereas females are only constrained by the References time that it takes to produce another batch of mature eggs. In our study, a male’s ‘time out’ was consistently 14–15 days for each Ahnesjo¨,I.1995. Temperature affects male and female potential reproductive rates differently in the sex-role reversed pipefish, Syngnathus typhle. Behavioral pregnancy. Females, in contrast, were able to remate rapidly. For Ecology, 6, 229–233. example, one female mated with eight males within a 2-week Ahnesjo¨, I., Kvarnemo, C. & Merilaita, S. 2001. Using potential reproductive rates period, delivering full clutches of eggs (enough to fill the male’s to predict mating competition among individuals qualified to mate. Behavioral Ecology, 12, 397–401. brood pouch) to five of them. To be able to mate multiply in rather Andersson, M. 1994. Sexual Selection. Princeton, New Jersey: Princeton University rapid succession, females must have a reproductive system that Press. facilitates successive rounds of ovulation in a short time period. The Begovac, P. C. & Wallace, R. A. 1987. Ovary of the pipefish, Syngnathus scovelli. Journal of Morphology, 193, 117–133. ovary of the gulf pipefish is unusual because it is structured in Begovac, P. C. & Wallace, R. A. 1988. Stages of oocyte development in the pipefish, a conveyor belt fashion (Begovac & Wallace 1987). As one set of Syngnathus scovelli. Journal of Morphology, 197, 353–369. eggs is ovulated into the lumen, the next set moves into its place Berglund, A. & Rosenqvist, G. 2003. Sex role reversal in pipefish. Advances in the Study of Behavior, 32, 131–167. and completes the final stage of maturation (Begovac & Wallace Berglund, A., Rosenqvist, G. & Svensson, I. 1988. Multiple matings and paternal 1988). The anatomy suggests that female gulf pipefish may have brood care in the pipefish Syngnathus typhle. Oikos, 51, 184–188. a very short period of time between the ovulation of each set of Berglund, A., Rosenqvist, G. & Svensson, I. 1989. Reproductive success of females oocytes. However, we are unaware of any published studies on the limited by males in two pipefish species. American Naturalist, 133, 506–516. Brown, J. D. 1972. A comparative life history study of four species of pipefishes length of the female reproductive cycle or the hormonal mediation (family Syngnathidae) in Florida. Ph.D. thesis, Univeristy of Florida, Gainesville. of maturation of oocytes and ovulation in this species. Brown, H. & Prescott, R. 1999. Applied Mixed Models in Medicine. West Sussex: Recent research does suggest a close link between ovarian J. Wiley. Clutton-Brock, T. H. 1991. The Evolution of Parental Care. Princeton, New Jersey: morphology and PRR within the family Syngnathidae. Sea horses Princeton University Press. and the monogamous pipefish C. haematopterus have ovaries that Clutton-Brock, T. H. 2009. Sexual selection in females. Animal Behaviour, 77, 3–11. differ in morphology from those of the gulf pipefish. Recently, Clutton-Brock, T. H. & Parker, G. A. 1992. Potential reproductive rates and the operation of sexual selection. Quarterly Review of Biology, 67, 437–456. Sogabe et al. (2008) found that C. haematopterus females have Clutton-Brock, T. H. & Vincent, A. C. J. 1991. Sexual selection and the potential ovaries with two germinal ridges, similar to ovaries in sea horses reproductive rates of males and females. Nature, 351, 58–60. (Selman et al. 1991),butnottoovariesofotherpipefishinthe Dearborn, D. C., Anders, A. D. & Parker, P. G. 2001. Sexual dimorphism, extrapair fertilizations, and operational sex ratio in great frigatebirds (Fregata minor). genus Syngnathus. These authors suggested that this structural Behavioral Ecology, 12, 746–752. difference might relate to the functional difference in reproduc- Emlen, S. T. & Oring, L. W. 1977. Ecology, sexual selection, and the evolution of tion between polyandrous/polygynandrous Syngathus pipefishes mating systems. Science, 197, 215–223. Evermann, B. W. & Kendall, W. C. 1896. Description of a new species of pipefish with female-biased PRRs versus the monogamous sea horses and (Siphostoma scovelli) from Corpus Christi, Texas. Proceedings of the U.S. National C. haematopterus pipefish with PRRs that approach unity. The Museum, 18, 113–115. group-synchronous ovaries of the monogamous sea horses and Forsgren, E., Amundsen, T., Borg, A. A. & Bjelvenmark, J. 2004. Unusually dynamic C. haematopterus result in a longer ‘time out’ for these females sex roles in a fish. Nature, 3429, 551–554. Foster, S. J. & Vincent, A. C. J. 2004. Life history and ecology of seahorses: impli- (Hippocampus spp.: 9–45 days, Foster & Vincent 2004; C. haema- cations for conservation and management. Journal of Fish Biology, 65, 1–61. topterus:10–19days,Sogabe et al. 2007)thanforSyngnathus Heubel, K. U., Lindstro¨m, K. & Kokko, H. 2008. Females increase current repro- females (48 h, Berglund et al. 1988; present study) that have an ductive effort when future access to males is uncertain. Biology Letters, 4, 224–227. asynchronous-type ovary (Wallace & Selman 1981; Begovac & Jones, A. G. & Avise, J. C. 1997. Microsatellite analysis of maternity and the mating Wallace 1988). It is possible that these two types of ovarian system in the gulf pipefish Syngnathus scovelli, a species with male pregnancy morphologies dictate the speed at which females of these species and sex-role reversal. Molecular Ecology, 6, 203–213. Jones, A. G., Walker, D. & Avise, J. C. 2001. Genetic evidence for extreme polyandry can produce mature eggs, which would directly affect the PRR. and extraordinary sex-role reversal in a pipefish. Proceedings of the Royal Society Further studies are needed to determine how the reproductive B, 268, 2531–2535. physiology of the ovarian cycle relates to these two distinct Jones, A. G., Moore, G. I., Kvarnemo, C., Walker, D. & Avise, J. C. 2003. Sympatric speciation as a consequence of male pregnancy in seahorses. Proceedings of the ovarian morphologies and whether ovarian morphology and/or National Academy of Sciences, U.S.A., 100, 6598–6603. physiology produce functional constraints that affect the PRR in Joseph, E. B. 1957. A study of the systematics and life history of the gulf pipefish these species. Syngnathus scovelli (Evermann and Kendall). Ph.D. thesis, Florida State Univer- sity, Tallahassee. Kokko, H. & Monaghan, P. 2001. Predicting the direction of sexual selection. Acknowledgments Ecology Letters, 4, 159–165. Kvarnemo, C. 1994. Temperature differentially affects male and female reproduc- We thank P.L. Schwagmeyer and Edith Marsh-Matthews for tive rates in the sand goby: consequences for operational sex ratio. Proceedings of the Royal Society B, 256, 151–156. their help with the analysis and comments on the manuscript. We Kvarnemo, C. & Ahnesjo¨,I.1996. The dynamics of operational sex ratios and also thank Ingo Schlupp, Celeste Wirsig and Debra Bemben for help competition for mates. Trends in Ecology & Evolution, 11, 404–408. with many aspects of this research, and Adam Jones, Anders Kvarnemo, C. & Simmons, L. W. 1998. Male potential reproductive rate influences mate choice in a bushcricket. Animal Behaviour, 55, 1499–1506. Berglund and an anonymous referee for helpful comments on an Masonjones, H. D. & Lewis, S. M. 2000. Differences in potential reproductive rates earlier version of the manuscript. Navin Chowhudry, Joe Kelly, of male and female seahorses related to courtship roles. Animal Behaviour, 59, Claire Lukeman, Amber Mielke and Lori Deunk Garman provided 11–20. Preston, B. T., Stevenson, I. R., Pemberton, J. M., Coltman, D. W. & Wilson, K. much assistance with animal husbandry. We also thank Leila 2005. Male mate choice influences promiscuity in Soay sheep. Proceedings of the Wright for assistance in the field. We are especially grateful to John Royal Society B, 272, 365–373. Author's personal copy

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Reid, G. K. 1954. An ecological study of the Gulf of Mexico fishes, in the vicinity of Stearns, S. C. 1992. The Evolution of Life Histories. Oxford: Oxford University Press. Cedar Key, Florida. Bulletin of Marine Science of the Gulf and Caribbean, 4, 1–94. Trivers, R. L. 1972. Parental investment and sexual selection. In: Sexual Selection and Roff, D. A. 2002. Life History Evolution. Sunderland, Massachusetts: Sinauer. the Descent of Man (Ed. by B. Campbell), pp. 136–179. Chicago: Aldine. Rosenqvist, G. 1990. Male mate choice and female–female competition for mates in Vincent, A. C. J. & Sadler, L. M. 1995. Faithful pair bonds in wild seahorses, the pipefish Nerophis ophidian. Animal Behaviour, 39, 1110–1115. Hippocampus whitei. Animal Behaviour, 50, 1557–1569. Selman, K., Wallace, R. A. & Player, D. 1991. Ovary of the seahorse, Hippocampus Vincent, A. C. J., Ahnesjo¨, I., Berglund, A. & Rosenqvist, G. 1992. Pipefishes and erectus. Journal of Morphology, 209, 285–304. seahorses: are they all sex role reversed? Trends in Ecology & Evolution, 7,237–241. Silva, K., Vieira, M. N., Almada, V. C. & Monteiro, N. M. 2008. Can the limited Vincent, A. C. J., Ahnesjo¨, I. & Berglund, A. 1994. Operational sex ratios and marsupium space be a limiting factor for Syngnathus abaster females? Insights behavioral sex differences in a pipefish population. Behavioral Ecology and from a population with size-assortative mating. Journal of Animal Ecology, 77, Sociobiology, 34, 435–442. 390–394. Wallace, R. A. & Selman, K. 1981. Cellular and dynamic aspects of oocyte growth in Sogabe, A. & Yanagisawa, Y. 2007. Sex-role reversal of a monogamous pipefish teleosts. American Zoologist, 21, 325–343. without higher potential reproductive rate in females. Proceedings of the Royal Watanabe, S., Hara, M. & Watanabe, Y. 2000. Male internal fertilization and Society B, 274, 2959–2963. introsperm-like sperm of the seaweed pipefish (Syngnathus schlegeli). Zoological Sogabe, A., Matsumoto, K. & Yanagisawa, Y. 2007. Mate change reduces the Science, 17, 759–767. reproductive rate of males in a monogamous pipefish Corythoichthys haema- Wiklund, C., Kaitala, A. & Wedell, N. 1998. Decoupling of reproductive rates and topterus: the benefit of long-term pair bonding. Ethology, 113, 764–771. parental expenditure in a polyandrous butterfly. Behavioral Ecology, 9, 20–25. Sogabe, A., Matsumoto, K., Ohashi, M., Watanabe, A., Takata, H., Wilson, A. B. 2009. Fecundity selection predicts Bergmann’s rule in syngnathid Murakami, Y., Omori, K. & Yanagisawa, Y. 2008. A monogamous pipefish fishes. Molecular Ecology, 18, 1263–1272. has the same type of ovary as observed in monogamous seahorses. Biology Zera, A. J. & Harshman, L. G. 2001. Physiology of life history trade-offs in animals. Letters, 4, 362–365. Annual Review of Ecology and Systematics, 32, 95–126.