Sex Reversal in Pairs of Lythrypnus dalli: Behavioral and Morphological Changes

2005·2.9 MB·

by  Edmund W Rodgers

Reference: Biol. Bull. 208: 120-126. (April 2005)
2005 Marine Biological Laboratory
Sex Reversal in Pairs of Lythrypnus dalli: Behavioral
and
Morphological Changes
EDMUND W. RODGERS. SHELIA DRANE, AND MATTHEW S. GROBER*
Department ofBiolos>\, Georgia State Universitv, P.O. Box 4010, Atlanta, Georgia 30302-4010
Abstract. In Lythrypnus dalli. the bluebanded goby, re- inhibition by males and stimulation from other females
productive success is primarily determined by functional (Robertson. 1972; Shapiro, 1979). Without sufficient so-
sex, and functional sex is determined largely by rank in the cial stimulation (i.e., in isolation), fish capable of sex
dominance hierarchy. In most natural social groups of L. reversal might not initiate sex change (Cole and Shapiro,
dalli. one male is at the apex of the hierarchy, and 1 to 7 1995; Carlisle et al.. 2000). Size advantage (Warner et
females are lower in rank. When a male exits the group, a til.. 1975) contributes to the determination of which an-
female ascends to the top of the hierarchy and becomes a imal changes sex. but behavioral interactions are also
male. We have examined this process in a simplified envi- critical determinants (Lutnesky. 1996; Munday. 2002).
ronment a pair of females that allows us to identify Larger size often equates with increased success in ag-
behavior associated with the formation of a dominance gressive encounters and therefore social dominance, pro-
relationship and any other phenotypic changes associated viding a proximate mechanism for the size advantage
with dominance, sex change or both. We found that pairs of hypothesis. In protogynous sex changers, the most repro-
L. dalli females quickly and readily form stable dominance ductively significant resource that dominance affords is
relationships, with the dominant fish changing sex into a "maleness"; thus the reproductive payoff for dominance
male. This dominant animal also rapidly increases in body is extremely large, and females would be highly moti-
size and length of its dorsal fin. In summary, dominant L. vated to increase their aggressive behavior in times of
dalli females change sex in this simplified environment, social instability (i.e.. in the absence ofadominant male).
providing excellent opportunities to examine the early be- The study of Lythrypnn.\ Jalli (Gilbert. 1890) had
havioral and morphological changes associated with domi- previously dealt exclusively with larger social groups
nance and sex change. (>3), and it was not known whether sex change could be
induced in pairs of females. This study used pairs of fish
Introduction to closely examine changes associated with the acquisi-
tion of dominance and, potentially, sex change. In a
Social context often determines how a given individual group environment, dominance relationships are more
responds to behavioral stimuli, with individuals modify- difficult to tease apart owing to multiple interactions
ing their behavior on the basis of the behavior of the between individuals in the group, whereas in pairs there
individuals around them. This is especially true in several are fewer social variables contributing to an animal's
species of protogynous (female to male) sex-changing behavior. We found that one of the pair quickly changes
fish, where sex reversal is socially controlled. In these to male, as evidenced by male-typical behavior and male
species, a unique pattern of behavior stimulates one in- reproductive morphology.
dividual to change sex while inhibiting others (Robert-
son, 1972: Ross el a/., 1983). Two social factors are
thought to be the primary regulators of sex change: Materials and Methods
We use four measures of "maleness" to determine ifany
Received 1 July 2004; accepted 12 December 2004.
*To whom correspondence should be addressed. ofthe fish changed sex: (1) display ofmale-typical courting
[email protected] behavior; (2) male-typical papilla ratio, defined as a length-
120
SEX REVERSAL IN PAIRS OF L DALLI 121
to-width ratio greater than 1.6 (female -1.0 1/w); (3) the During sex reversal, L. dalli undergoes gonadal reorga-
presence of an accessory gonadal structure (AGS); and (4) nization (St. Mary, 1994), which in a number ofsex-chang-
the presence offertilized eggs, an unequivocal indication of ing gobies involves degeneration ofovarian tissue (Sadovy
functional sex change. and Shapiro, 1987) and generation of AGS and testicular
tissue (Cole and Shapiro, 1990). In L. dalli, the presence or
Subjects absence of the AGS is the primary indicator of functional
We selected 32 female specimens of Lythrypnus dalli sex; males have a highly developed AGS and females have
with lengths between 23 and 30 mm for the study. Sex was none. Visual inspection was used to assess the composition
d(Beetherremnitnse.d19b8y3)e.xTamhienapatpiiolnlaorfattihoeinexfteemranlaelsgoefnitthails sppaepciilelsa oAfGSre.prGooduncatdivesetxiswsuaes, iv.ee.r,ifpireedseunsciengofsetagngdsaradndpaprraefsfeinncehios-f
is about 1.0, compared with about 2.0 in the males (St. tology followed by staining with hematoxylin and eosin.
Mary, 1994); the largest ratio at the start ofthis experiment
was 1.3. Fish with female-typical papilla have gonads con-
sisting of more than 95% ovarian tissue (St. Mary, 1994). Pairing
During the experiments, animals were housed in 33-1
At the start of each experiment, animals were removed
aquaria, each with an individual filter system (Marineland), from the holding tank and anesthetized using MS 222. Mass
cpatyacrl2ee0d.-d2Oie1nt.e(1OCgSr;IoufMpeadroiftnwfeiicsLehabwdsaa)is;lycaonuldsliekncegtpetdao(cnCoaamlmi1ef2ro-rchniialailglFhyits/hpdraer&-k asunrdeds,taannddarddilgeitnaglthiwmeargeesrewceorrdeeda.lsPomaptmialklean.ratAinosimwaelrse mweear-e
Game permit # 803034-01) on Santa Catalina Island, Cali- paired by standard length (within 2 ofeach other) and
fornia, in May 2002 (pre-breeding season), using an anes- body mass (within 0.1 g). The fish were individually iden-
thshaaenmtdie-cnmesetotslh.uotAdiosn,seodcfuorqniudnigngatrlhoeduibpnreoefesdufilifnsaghteswea(asSsiocgnomlialnecCltahteeedm,Jiuucnsaeiln)2g0a0tn2h.de ctwihefarienedgpeblyancoetrdhehtiaorvgeebtahanendrieinfngfteocptaat3ot3ne-r1nt,hteaiwnrhkibwecihhtahvdiaiodrs.inngToltweoaPpVapnCeiamrtaulbtseo
to serve as the nest (St. Mary, 1994).
Prior to the experiments, the animals were kept in 180-1
holding tanks.
Observations and behavior
Experimental design
The study is divided into two experiments. In experiment The behavior of each pair was observed twice daily,
1, morphological traits and behavioral interactions were once in the morning and once in the afternoon, for 10 min
quantified between eight pairs of individuals identified as each session. Data were recorded on paper and then
females. External morphological data were collected prior transferred to a computer spreadsheet. The fish were
to pairing (standard length, mass, and papilla ratio, de- given one day to acclimate before observations began.
scribed below). The pairs were observed until eyed eggs (a The observer recorded the number of approaches, dis-
marker of fertilization), indicating functional sex change, placements, jerks, bites, nips, solicitations, and tail-wag-
were seen, or until 14 days, an adequate amount oftime for gles. In addition, the observer noted which animal resided
sex change in this species (Reavis and Grober. 1999), had in the nest tube, if either animal was gravid, and whether
elapsed. Experiment 2 was similar to experiment 1 and was eggs were present in the tube.
designed to confirm a novel observation in experiment 1, An approach is defined as a fish moving within 5 cm of
that dominant fish exhibited disproportional growth of the the other fish. Movement away from the approaching fish is
longest rays ofthe dorsal fin. The behavior ofthe eight pair recorded as a displacement. Jerks are a male-typical behav-
of fish in experiment 2 was not observed closely. In both ior used during courtship (Behrents, 1983), and involve a
experiments, the pairing and morphological measurements saltatory swim motion with movement laterally as well.
(with the exception of initial dorsal fin length) were iden- Becausejerks can be directed either at the female or around
tical (see below). the nest without being directed at a fish, a jerk towards a
At the completion of each experiment, animals were female is scored as both an approach and a jerk. Bites are
given an overdose of tricaine methanesulfate (MS 222), aggressive interactions in whichone fishbitestheotherafter
and their morphological characters were measured again. an approach. In contrast, nips follow ajerk to afemale, with
This included measurement ofthe longest dorsal ray (see a male nipping the tail of the female. This behavior is also
Results). The animals were placed in Bouin's fixative, the part of the courting process. Solicitations are produced by
gonads were dissected out to examine internal morphol- females when they move within 12 cm of the male in his
ogy, and digital photographs ofall the gonads were taken line of view. Tail waggles are displayed by both sexes: the
to aid in analysis. fish remains stationary while moving its tail back and forth.
122 E. W. RODGERS ET AL
Statistics ences in size predicted which individual would change sex.
Significance level in all cases was set at P < 0.05. Mean
Morphological data were analyzed using parametric sta- values standard error of the mean (SEM) are given. All
tistics. Fortraits that had "before" and "after" measures, we analyses were earned out using the Statview 5.01 (SAS
used paired student's Mests (standard length and fin length Institute Inc.) unless otherwise noted.
in experiment 2). For fin length in experiment 1, we used an
unpaired f-test to look at differences between males and
females. We also used a Mest to examine whether papillar Results
morphology was significantly different between new males
and females. The distributions of the frequencies of the In all cases, the pairs readily established a stable domi-
various behaviors fornewly sex-changed males and females nance relationship, defined as one individual (the dominant)
did not meet the criteria for parametric statistics (e.g.. nor- instigating the majority of encounters (Fig. la) and exhib-
mality, homogeneity of variance); thus we used nonpara- iting male courtship behavior (Fig. Ib). while the other
metric methods to examine group differences. All behav- individual (the subordinate) is displaced as a result of the
ioral analyses were conducted with the Wilcoxon signed encounter (Fig. Ic). In all cases the dominant individual
rank test. Values for mean and median were similar, indi- initiated spawning behavior with the subordinate individual
cating a symmetrical distribution. Simple linear regressions that resulted in a clutch of eggs. Of the 8 clutches, 4 were
were employed to examine the relationship between known verified as fertile; in the other 4 cases, the male consumed
interdependent behaviors (e.g., approaches regressedon dis- the clutch before verification. At the completion of the
placements). Though the fish were size-matched, we used experiment, all dominants had a male papillaratioas well as
logistic analysis (SAS) to examine whether small differ- an ACS (Figs. 2, 3). Using our criteria, we demonstrated
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Day
Figure 1. Changesinsex-typical behavioroverthecourseofexperiment. Animalsthatare labeledmaleare
individuals that changed sex i.e., those that would become male, (a) Males approach and females rarely do,
demonstrating dominance, (b) Jerking behavior, a majorcomponent ofmale courting behavior, is exclusively
produced by males, (c) Females were displaced at high rates compared to males, (d) Solicitations, a type of
femalecourting behavior, are exhibitedexclusively by females. Notethe similarpatternofbehaviorin band d.
SEX REVERSAL IN PAIRS OF L. DALL1 123
mean of5.5 0.269 mm (t = 8.475, df = 14, P < 0.0001 ).
To control fordominant fish simply growing more, we used
--o* - SDuobmoirndainntat/eM/alFeemale a ratio of fin length to body length. This ratio was signifi-
cantly different: subordinates had a smaller fin with an
average of4.648 0.137 fin lengths per body length, and
dominants had aratioof2.64 0.143 (t = 10.129, df = 14,
o P < 0.0001).
Experiment 2 verified the marked change in fin length
observed in the first experiment. Future dominants and
'5. subordinates did not have different fin lengths at the begin-
Or.o ning of the experiment (t = 1.669, df = 8; P > 0.05), but
the dominant fish had a significantly longer fin at the end of
the experiment (t = 4.091, df = 8; P = 0.0035). Dominant
individuals exhibited a pronounced lengthening of their
longest dorsal ray, while many of the subordinates showed
a fin shortening.
Initial Final We attempted to minimize size as a major contributing
Figure2. Papillalength/widthratiosatthestartandconclusionforall factor; however, small differences in size did exist between
Pathnai<nmas0lu.sb0o0irn0d1ei)xn.paetersim(efnetma1l.eDs)omaitntahnetesnd(moaflethse)ehxapdearismiegnnitfi(tca=ntl8y.6h4i,gdhfer=rat14i,o ethnecepsaiirnedstaannidmaarlds.leTngothaswseersse wphreedtihcetrivetheosfewshmiaclhl adniiffmearl-
changed sex, we ran a logistical regression. Our results
suggest that the size differences present in this experiment
tsheaxtftrhoemdofemmianlaenttoinmdailvei.dual in a pair of females changed dwoemrienanontta(gWoaoldd prred=ict0o.r03o4f.whPic=h 0a.n8i5m3a2l,wdoful=d b1,ec/3om=e
-0.05006).
Morphological changes
Behavior
All fish initially had a female-typical papilla ratio
(mean SEM: 1.15 0.061). There was no difference in Verification of egg fertility in experiment 1 was diffi-
= =
papilla ratio at the start ofthe experiment (t 0.732, df cult because of consumption of clutches by males (see
14, P > 0.05) between fish that became male (mean below). Four of eight pairs completed sex change in less
SEM: 1.15 0.102) and those that remained female than 14 days, as determined by the presence of fertilized
(mean SEM: 1.17 0.086). After sex change there was eggs. In the four remaining pairs, males exhibited all
a significant change to a male-typical papilla in the fish that other sex-typical characteristics, but consumed at least
demonstrated male-typical behavior(t = 8.64, df = 14. P < one complete clutch of eggs. Because fish that parented
0.0001). Female papilla ratio remained the same. 1.09 and fish that consumed their eggs displayed similar rates
0.052, whereas the ratio in sex changers more than doubled of behavior in all measures, we grouped them in the
to 2.26 0.127 (Fig. 2). behavioral analysis.
All dominant animals exhibited unambiguous male-typi- A clear dominance relationship was evident from the
cal gonad morphology that included the presence of sperm first observational session and persisted unchanged
and an AGS. All subordinate individuals were gravid, and throughout the experiment (Fig. 1). The dominant fish
eggs were clearly visible within the ovary upon inspection. produced male-typical behavior and began courting the
These results were verified histologically (Fig. 3). subordinate. The dominant fish instigated the vast major-
The standard length of dominant individuals changed ity of behavioral interactions, evidenced by high rates of
significantly over the course of the experiment (P < 0.05); approach behavior. Dominants displayed significantly
that ofsubordinates did not (P > 0.05). When we examined higher rates of aggressive and courtship behavior (ap-
the magnitude and direction of the change, we found that proaches and jerks, P < 0.05. Fig. la, b). Subordinates
dominants grew significantly more relative to subordinates displayed significantly more submissive and female-typ-
(paired /-test. / = 4.528, df = 15. P = 0.0003). ical behavior (displacements and solicitations, P < 0.05,
In experiment 1, we noted that the dorsal fin of the Fig. Ic, d). All dominants jerked and subordinates gen-
dominant fish was elongated. Although we had not mea- erally did not jerk, although there were rare individual
sured initial fin length, at the conclusion of the experiment displays over the course of the experiment. Subordinate
we measured the longest dorsal ray. Dominants had a mean fish were displaced at high rates (9.138 per 20 min. Fig.
fin length of 10.47 0.539 mm, while subordinates had a Ic) compared to dominants (0.062 per 20 min). There
124 E. W. RODGERS ET AL
ACS
Figure 3. Gonad morphology ofa typical male (left) and female (right). The cross-section ofeach type is
shown underneath. Note that the male gonad contains mostly testicular tissue, some ovarian tissue, and an
accessory gonadul structure (AGS). The female gonad contains only ovarian tissue and is visibly full ofeggs.
was a significant association between approaches and Other behavior occurred at a far lower frequency, includ-
displacements, such that when a dominant approached, ing bites, nips, and tail waggles. Dominant individuals pro-
the subordinate was usually displaced (R2 := 0.917; P < duced bites and nips. Tail-waggle was the one behaviorthat
0.001 ). The dominant guarded or resided in the tube for did not show an overall sex/dominance bias, but was ex-
most of the time. hibited by both dominants and subordinates.
SEX REVERSAL IN PAIRS OF L. DALL1 125
Discussion ior (approaches, jerks, bites, and nips). Morphological ex-
amination revealed no differences between males that dem-
When two female bluebanded gobies were paired, in our onstrated appropriate care and those that did not. The
experimental conditions, inallcases,onefishestablisheddom- gonads of the two groups were indistinguishable from one
inance over the other, with that fish instigating most of the another, and both groups developed an ACS, the hallmark
encounters and winning nearly all ofthem. The dominant fish of the male sex. There were no differences in the papilla
became male, exhibiting all behavioral and morphological between the two groups, and all males displayed sexually
characters associated with that sex. The subordinate fish re- dimorphic elongation ofthe longest dorsal rays. In addition,
mained female and exhibited all associated sex-typical behav- parenting males had fertile clutches at the same time as egg
ior and morphology. consumption was occurring, suggesting that all ofthe males
In Lythrypnusdalli, body size is sexually dimorphic, with were capable of fertilizing eggs.
males being larger (Wiley, 1976). In our work, dominant In experiments with groups of fish greater than two
individuals grew more than subordinates and changed sex. (Reavis and Grober, 1999), the amount oftime required for
The observed sexual dimorphism in growth rate is consis- full sex change depended on the size of the individuals in
tent with previous data from St. Mary (1994); however, we the group upon male removal. In groups where the size
cannot address whether the increase in growth is a result of asymmetry between the top two females was greater than
sex change or of dominance. Dominance rank has been 10%, sex change occurred more rapidly than in groups
shown to affect the growth rate of individuals within a where the size asymmetry was less than 10%. The observed
hierarchy of sex-changing clown fish (Buston, 2003). We delay was determined to be the result ofan increase in time
alsofound that the dorsal fin was elongated in the dominant, spent resolving the dominance relationship between the top
sex-changed fish. The dorsal fin is known to be sexually two individuals. Individuals of similar size take longer to
dimorphic (Wiley, 1976). but we did not expect the rapidity settle a conflict than individuals of different sizes (Enquist
and magnitude of the changes we observed in the first el al. 1990; Koops and Grant, 1993). In this experiment, we
experiment. Experiment 2 confirmed that the fin lengthened used pairs of size-matched individuals; thus we were sur-
dramatically in the dominant individual over the course of prised to find that the contestants quickly resolved the
the experiment. The dorsal fin shrank in many of the sub- dominance relationship. One possible explanation is that in
ordinate fish, but whether this is a natural process or the a group, individuals may receive contradictory signals. This
result of aggressive encounters with the dominant is un- could result from being dominated by one individual but
known. As with the change in growth rate, we cannot say also receiving positive stimulus by dominating other indi-
whether the fin elongation is caused by the sex-change viduals in the group, thus prolonging conflict resolution. In
process or is modulated by dominance status. Ifthe latter is dyadic contests, the signals are straightforward; one animal
the case, then this trait can potentially be used as an index wins the encounters while the other loses them, thereby
of rank within a hierarchy. shortening the time and probably the numberofinteractions
Simplification ofthe social group to a pair ofanimals, as required to form a stable dominance relationship.
in our study, revealed an interdependence of behavior be- In summary, two females is a sufficient group size to
tween males and females that was not seen in previous induce sex change in L. dalli, as evidenced by one individ-
studies using larger groups offish. This is best illustrated in ual exhibiting male typical gonads, external genitalia, be-
Figure Ib andd, where courtship behavior(solicitations and havior patterns, and often fertilized eggs. When an L. dalli
jerks) clearly shows the same pattern over time. Although female becomes dominant, the first noticeable morphologi-
the precise nature of the interaction is unclear, female be- cal change is the elongation of the dorsal fin. Dominant
havior apparently had an impact on the rate ofmale behav- individuals also grow more rapidly.
ior, an effect that would be difficult to examine in a group
with multiple females. Acknowledgments
Like ReavisandGrober(1999), we used fertilizedeggsas
a marker for complete sex change and allowed the experi- The authors thank Cathleen Drilling for her work in
ment to run for a maximum of 14 days. All tanks had eggs collecting the experimental animals, help with behavioral
in the nest within the 14 days, but only half of those nests observations, and gonad histology; Michael Black for ani-
hadeggsthatremained until they couldbe verified as fertile. mal collection and behavioral observations; and William
In the four other groups, the males did not display paternal Lonergan and Beth Stokes for their help with behavioral
care. These males ate the eggs after day 10 of the experi- observations. We also thank Ryan Earley for his help with
ment, making it impossible to use the presence ofeyed eggs statistical analysis, and Dr. Charles Derby and an anony-
to verify fertility and thus terminate the experiment. The mous reviewer for their comments on an earlier version of
bulk of the data indicates that these eggs were viable. this paper. This work was supported by NSF IBN 9723817
',
Clutch-consuming males exhibited all male-typical behav- to Matthew S. Grober, the Georgia Research Alliance and
126 E. W. RODGERS ET ,\L.
tthhee CNeSnFteragfroereBmeehnatvi#oIraBlN-Ne9u8r7o6s7ci5e4n.ce, an STC programof lM.uuntpdnoeamsyak,yc,aPn.Mt.hLi.d129a90n60g.2e.lfiSsiBhzi.-ed-Cidereneptcetrniodopneyangltesrpeaoxttetceohrfai.npgrCeoo:tpoteegisyatnion1ug:st2h0see9x-g2rc1oh2wa.tnhgeraitne
advantage model. Behav. Ecol. Sociobiol. 52: 247-254.
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EKnoqoupfs41iies97ssq9,5htu0-,e.N4MnaM7t.ni.9anA,.lAa.Oct,a.eassrtasLaeenosidfasmnmasoJeer.mnq,atuWleTan.i.t.niLaAAcjl.onuninavmGsdi.rscbaeteBnsretscgh.mi,acevhn1Y.lt9.id94g3M0a.ca:molnel1t:nW-ee1se4ftrii.s,g.ghhattCniandnag.sNiy.nJm.SmteheZgetoeorrc/yid.cahha7lln1i.d:d WWialre5an(Ay:Gnem,iderl3rb,J0se.i4ercR-txW.a)u3..an1Rl3.T1..grs9oae7Dnbl6es.i.c.etsRiS.o.LnLiR.DyfotieSbhecehigiriyoestpntsnSocouorencise.,1s9dNa0aaaln:ntldd.i63sEH(3i.yGs-sitGt6l.e.3bme81arL.8tte:)iicg1sah6n.o9df-11t9L8h7y4e5t..WhersySptemexirsnchNzaoenrbgtreha