THE EFFECT OF PERCEIVED INTRUDER PROXIMITY AND RESIDENT BODY SIZE ON THE AGGRESSIVE RESPONSES OF MALE GREEN FROGS, RANA CLAMITANS (ANURA: RANIDAE)

Behav Ecolog Sociobiol 2005

Patrick C. OwenandNoah M. Gordon

Introduction Graded response A variety of animals modify or grade their

aggressive responses in a continuous fashion.

Can prevent encounter from escalating into fight.

Costsreduced mating success, greater energetic expenditure, or greater risk of predation.

(Archer 1988; Bond 1989a; Hauser and Nelson 1991; Bradbury and Vehrencamp 1998; Hurd and Enquist 2001)

(Martins 1994; Adams and Mesterton-Gibbons 1995; Hurd and Enquist 2001)

2

These aggressive responses may vary with the size

(Archer 1988; Andersson 1994; Pusey and Packer 1997).

Graded responses may function as signals of1) aggressive motivation2) fighting ability

Studies in amphibians have suggested that graded variation may serve in both of these roles.

(Bond 1989a, b, 1992; Hauser and Nelson 1991; Hurd and Enquist 2001; Lange and Leimar 2003)

(Wells 1988; Schwartz 2001; Gerhardt and Huber 2002)

3

These responses most often take the form of discrete call types that can vary continuously

Modified item Reference

Duration of notes Schwartz and Wells 1984; Wells and Schwartz 1984; Littlejohn and Harrison 1985; Wells and Bard 1987; Martins et al. 1998; Wogel et al. 2004

The number of pulses of notes

Pengilley 1971; Arak 1983b; Schwartz 1989; Wagner 1989b; Grafe 1995;Wogel et al. 2004

The dominant frequency

Lopez et al. 1988; Wagner 1989a; Grafe 1995

Calling rate Given 1987; Stewart and Bishop 1994; Grafe 1995; Jehle and Arak 1998; Grafe et al. 2000; Narins et al. 2000; Grafe 2003

Call intensity Lopez et al. 1988

Response threshold Rose and Brenowitz 1991; Brenowitz et al. 2001

Diversity of call types

Christensen-Dalsgaard et al. 2002 4

Some anurans may signal body size or fighting ability with these call responses (Davies and Halliday 1978; Arak 1983a; Ramer et al. 1983; Robertson 1986; Given 1987; Wagner 1989a; Bee et al. 1999, 2000),

Less attention has been focused on their role in signaling aggressive motivation (Burmeister et al. 2002).

The graded nature of non-vocalization responses such as movements or visual displays has not been investigated.

5

We examined graded responses in green frogs (Rana clamitans,Ranidae). Eastern North America Part of the Rana catesbeiana species group

(Austin et al. 2003). Prolonged summer breeders (Wells 1977a)

Resource defense mating system in which males defend oviposition sites (Wells 1977b).

The quality of the oviposition site is more important in mate choice than is calling behavior.

6

Aggressive behavior is well documented in green frogs (Jenssen and Preston 1968; Wells 1978; Ramer et al. 1983; Bee and Perrill 1996; Owen and Perrill 1998; Bee et al. 1999, 2000)

Aggressive encounters escalate when an intruding male approaches and enters the territory of a resident.

Encounters sometimes result in wrestling matches. Residents typically have the advantage Larger males usually defeat smaller males (Wells 1978).

Green frog displays consist of a combination of calls, movements, and other visual stimuli such as prominently displayed yellow throats and elevated body posture (Wells 1978).

7

Green frog advertisement (type I) and aggressive (type III) calls are similar (Wells 1978).

The main consistent differences are that aggressive calls are lower in dominant frequency, and longer than advertisement calls (Bee and Perrill 1996).

8

type IIItype I

lower

longer

Advertisement and aggressive calls are not really two discretely different signal types, but a single signal that varies continuously along a continuum.

9

http://www.sonic.net/~ric/go/vgame1.htm

A third, discretely different call sometimes given during the course of aggressive encounters is the type II “high-intensity” advertisement call (Wells 1978).

The function of this call has never been satisfactorily determined (Bee and Perrill 1996).

10

Dominant frequency In advertisement calls are stable(Bee et al.

2001), but lower and more variable in aggressive calls

May function in signaling resident size to intruders (Bee et al. 1999), and smaller green frogs may bluff their size(Bee et al. 2000).

Frequency alteration information about fighting ability. information about signaler motivation,

but this has not been previously investigated.

11

Objectives

To determine whether these aggressive responses are graded according to increases in stimulus level and to examine how these responses vary with resident body size.

We investigated the components of aggressive responses rates of response calls temporal (duration) and spectral (dominant

frequency) characteristics We also examined rates of response

movements.

12

The null expectation1. Resident would not respond differently to

stimuli representing different level of threat

Progressive changes in responses would provide evidence that aggressive responses are graded as a function of the perceived level of threat

2. Frogs would give the same level of response regardless of body size

If larger males give proportionally greater responses to stimuli, then this suggests that larger males employ potentially more costly responses than do smaller males

13

Methods

We conducted experiments in a large pond in Brooklyn, Connecticut, USA, during the summer of 1999.

Time: 2200 to 0300 h Measurement: SVL and body mass Individual recognition

spot patterns and scars were recorded avoid pseudo-replication.

Water temperature did not vary a lot.14

Stimulus call intensity We recorded the intensity of four

calls for each subject, measured the distance

mean sound intensity (y) as a function of distance (x) in cm (n=20, r2=0.884, P<0.0005, y=−0.07x+96.815).

territory sizes were typically 1–2m in radius.

Simulate an encroaching intruder15

Distance (m)

2 1 0.5

Intensity (dB)

83 90 97

Playback Speaker mounted on a Styrofoam platform. 1m from a calling male. Calls were recorded using a tape cassette

recorder. Average characteristics

327 Hz in dominant frequency 250 ms in duration

presented at a rate of 2 calls per min). Our focus for this experiment was on

responses to an “average” intruder. Battery power can affect stimulus

properties.16

Playback We used a graded playback design

similar to Schwartz (1989) and Wagner (1989b) with the following modifications. Pre-experimental period of at least 5 min. If the male interrupted during the pre-

experimental or experimental periods, then we stopped

We ultimately excluded five males However, relocate disturbed males was

not an ideal solution (Gordon 2004).

17

Playback

Record 3-5 calls→S1→S2→S3

Each experiment lasted for at least 35 min.

A balanced design that controlled for possible carry-over effects (Quinn and Keough

2002). Movements were assessed visually.

18

Pre-experimental period Experimental period

Sound analyses Digitized recordings at 48 kHz (16

bit). Only the first note of multi-note calls

was analyzed (Bee and Perrill 1996). Call variables included

dominant frequency call note duration Response latency (reaction time).

19

Repeated measures multivariate analysis of variance (MANOVA) and covariance (MANCOVA)

dependent variables mean dominant frequency mean note duration calling rate (calls/5-min period) movement rate (moves/5-min period) calling latency, and movement latency

SVL was used as the covariate.

20

We used the rank transform approach in which response data were converted to ranks within each variable, and the resulting ranked data were analyzed using standard parametric methods (Quinn and Keough 2002).

Linear contrasts were used to test the hypothesis of response change trends for each significant ANOVA.

Critical significance levels of ANOVAs and linear contrasts were adjusted using the sequential Bonferroni method (Quinn and Keough 2002).

21

Results

Aggressive response 17 individuals responded to the stimulus with a

combination of calls and movements. Gave a response call first (69% of broadcasts), the first

response movement was given an average of 21.1 s later. When a movement was given first (31% of broadcasts), the

first response call was given an average of 15.3 s later. In one instance the male mounted the speaker platform

and attempted to wrestle with the speaker. Normally, the frogs approached no closer than the water in

front of the speaker platform. Returned to their original positions by the end of each

intervening 5-min silent period

22

Type II high-intensity advertisement calls occasionally given by 10 males. Three of these individuals gave type II calls in

response to our broadcasts The other seven individuals gave type II calls in

response to other frogs what were producing type II calls.

Most type II calls were produced during broadcast periods (83 dB = 6 calls, 90 dB = 5 calls, 97 dB = 5 calls, silent periods = 2 calls).

Only 17 of 940 total response calls for all individuals were type II calls.

23

Male body size (SVL) of the subjects ranged from 69 to 91 mm encompassing the range of sizes in previous studies that addressed fighting ability (Wells 1977b, 1978; Ramer et al. 1983;

Bee et al. 1999, 2000).

24

A preliminary analysis using Spearman rank-order correlations indicated that SVL was significantly correlated with some of the response variable difference scores at some broadcast intensities.

25

Broadcast level (dB)

Move rate

Call rate

Call duration

Dominant frequency

Move latency

Call latency

83 0.371 0.162 -0.203 0.147 -0.402 0.086

90 0.477 0.148 0.067 0.391 -0.055 -0.083

97 0.522 0.419 0.125 0.268 0.124 -0.187

A repeated measures MANCOVA was performed with SVL as the covariate.

In contrast to the analysis of Bee et al. (1999), SVL did not have a significant effect on response level (F1,15=3.74, P=0.072),

and including SVL as a covariate only resulted in a decrease of 0.9% in total unexplained variation compared to a MANOVA (see below) on the same data.

Thus, SVL was not included in further analysis of stimulus level effect.

26

Repeated measures ANOVA on response level

27

movement rate(F2,32=18.42, Pcrit=0.050,

P<0.0001)

calling rate (F2,32=10.071, Pcrit=0.013,

P=0.001)

Stimulus levels did not have a significant effect on response latencies

28

callsF2,32=0.14, Pcrit=0.001, P=0.849

movements F2,32=0.14, Pcrit=0.001, P=0.849

stimulus level had a significant effect on call dominant frequency (F2,32=6.66,

Pcrit=0.006, P=0.004; Fig. 4A) and on call duration (F2,32=13.45, Pcrit=0.025, P<0.0001; Fig. 4B).

29

Linear contrasts indicated significant changes for movement rate (F1,16=37.09, Pcrit=0.050, P<0.0001)

call duration (F1,16=13.452, Pcrit=0.025, P<0.0001)

calling rate (F1,16=10.07, Pcrit=0.013, P=0.001)

call dominant frequency (F1,16=13.649, Pcrit=0.006, P=0.002)

Repeated measures MANOVA showed that the ordering of stimuli did not significantly influence response levels (F17,272=0.42, P=0.832).

30

DiscussionGraded Response Male green frogs graded their aggressive

responses as a function of stimulus intensity. With increasing intensity, males increased both their calling and movement rates

Latencies for these responses did not significantly change with increasing stimulus intensity.

In some frogs different types of responses have different latencies (Martins et al. 1998; Christensen-Dalsgaard

et al. 2002). However, in green frogs the latencies did not

greatly differ according to response type. Although most individuals gave a call before the first movement.

31

Graded Response As calling rate increased, call

duration and dominant frequency decreased

Longer calls with a dominant frequency at least 20–30 Hz below that of advertisement calls to be aggressive calls (Bee and Perrill 1996; Owen and Perrill 1998).

32

However, there was no obvious clustering that would indicate two distinct call types according to those measures (Fig. 5).

33

Thus, advertisement (type I) and aggressive (type III) calls are graded continuum.

Similar in Acris crepitans (Wagner 1989a, b)

and Hyperolius marmoratus (Grafe 1995).

34

type II high-intensity advertisement calls not commonly employed in close range

aggressive interactions. in most instances they appeared to be given in

response to type II calls from other males in the chorus.

In contrast, the other types of response calls and movements were exclusively directed at stimulus broadcasts or real intruders.

Wells (1978) and Ramer et al. (1983) suggested that type II calls played an intermediate role in the escalation of aggressive encounters.

However, in the context of this study, type II calls do not appear to be used as a close-range threat display, and their function may have more to do with long-range communication. 35

Movement respond Most previous studies of graded responses in frogs

have demonstrated that males increase their calling rate with increasing stimulus intensity (Brzoska et al. 1982; Schwartz and Wells 1984; Wells and Schwartz 1984; Schwartz and Wells 1985; Given 1987; Schwartz 1989; Stewart and Bishop 1994; Grafe 1995;

Jehle and Arak 1998; Grafe et al. 2000). Occasional phonotaxis toward an intruder stimulus

has been previously observed in a number of species, but this occurs infrequently during encounters (Brzoska et al. 1982; Harrison and Littlejohn 1985; Littlejohn and Harrison 1985; Brenowitz 1989; Wagner 1989b; Stewart and

Bishop 1994; Jehle and Arak 1998; Grafe et al. 2000). Most of our subject green frogs approached our

intruder stimulus, and they appear more likely to use movements as a threat display than other species of frogs.

36

Graded changes in temporal properties males generally increased the duration of

the calls or call notes in response to increasing stimulus intensity (Schwartz and Wells 1984; Wells and Schwartz 1984; Littlejohn and Harrison 1985; Schwartz and

Wells 1985; Schwartz 1989; Wagner 1989b; Grafe 1995). In contrast, we found that green frogs

decreased the duration of call, also differs from the results of Bee and Perrill (1996).

A possible explanation increase call duration in response to calls of a

more distant intruder shorten call duration if that intruder

approaches.37

Graded changes in spectra properties Several species of frogs are known to alter

dominant frequency during aggressive interations (Lopez et al. 1988; Wagner 1992; Bee and Perrill 1996; Howard and Young 1998; Bee et al. 1999, 2000; Given 1999; Bee and Bowling

2002). Only a few species are known to do this in a

graded context. Leptodactylus albilabris (Lopez et al. 1988) tended to

increase or decrease frequency to match that of a stimulus.

In contrast, both Acris crepitans (Wagner 1989a) and Hyperolius marmoratus (Grafe 1995) decreased their dominant frequency regardless of the frequency of the stimulus.

38

Graded changes in spectra properties Green frogs responded in a similar manner

by progressively decreasing the dominant frequency

Dominant frequency changes on the order of 50 Hz play a potentially important behavioral role in green frog communication (Bee et al. 1999, 2000).

For all calls, the mean difference between highest and lowest frequencies was 43 Hz (range 8–68 Hz)

For response calls alone, the mean difference was 26 Hz (range 2–50 Hz).

39

the mechanism for lowering dominant frequency does not related to the shortening of call note duration.

40

Cost and benefit in graded response Potential cost- Reduced mating

success Females of other frog species prefer

advertisement to aggressive calls (Wells

1988; Gerhardt and Huber 2002). In green frogs, aggressive (response)

calls are produced at lower sound intensities (Bee and Perrill 1996)

This may decrease the chance of long-range detection by females.

41

Since advertisement and aggressive calls form a graded continuum in green frogs, graded response calls could be “bi-functional” (Wells 1988; Schwartz 2001).

Female choice in green frogs is based on territorial quality.

Thus, males may not suffer reduced mating success

42

Potential cost- energetic costs Energetic costs are associated with aggressive

responses in green frogs, and these costs increase as responses increase in intensity.

The energetic cost of advertisement calling is probably very low, especially when compared to species of hylid treefrogs(Wells 2001).

However, aggressive response behaviors in green frogs are more costly than advertisement behaviors.

Males giving the most intense aggressive responses produced their calls at rates more than three times.

43

The increase in movement rates was even more dramatic.

Movements are most likely primarily fueled by anaerobic metabolism (Gatten et al.

1992), and green frogs lack the large lipid stores in their trunk muscles (Wells 2001).

Thus, high rates of movement or calling cannot be sustained for long periods.

44

Potential cost- predation risk Elevated levels of responses may make male green

frogs more visible to predators. Predation was the most obvious cost in our study. A wide variety of vertebrates are known to prey upon

adult green frogs (Martof 1956), and predators are attracted to calls in other species of frogs (Gerhardt and Huber 2002).

Two male green frogs did not appear in our final analysis of graded responses. While responding to our playbacks, one male was attacked

and injured by a water snake (Nerodia sipedon), The other male was devoured by a female bullfrog (Rana

catesbeiana). In contrast, we did not witness any attacks on males

that were merely advertising during our experimental trials.

45

Graded responses are favored if they are on average less costly than producing discrete “all-or-nothing” responses.

If graded responses reduce the chances of an encounter escalating into a fight (i.e. they act as signals), then the responses may also function as threat displays.

46

Trade off Stronger responses

less appealing to females more energetically costly attract predators

Weaker responses less likely to repel an intruder.

47

Since male green frogs defend calling positions that are used as oviposition sites by females, and females enter breeding aggregations sporadically throughout the breeding season(Wells 1977b;

Gordon 2004). Thus, the cost of losing a territory in an

encounter is potentially very high for a resident.

Graded responses may represent a compromise in which the costs of producing the response are balanced with the potential costs of territory loss.

48

Signaling system Graded signaling system potentially

provides males with the ability to convey information on motivation to fight or on fighting ability itself (Hurd and

Enquist 2001). Spectral components of calls may be

used to assess fighting ability in several anurans including green frogs (Davies and Halliday 1978; Arak 1983a; Ramer et al. 1983; Dyson and Passmore 1992; Wagner 1992; Bee et al. 1999,

2000; Burmeister et al. 2002).49

Signaling system - body size or fighting ability In this experiment, some of the difference

scores for responses were significantly and positively correlated with body size (Table 1)

The intensity of these responses between individuals potentially could be used by opponents to assess body size or fighting ability.

Movement rates were most strongly correlated with body size in this experiment

In the current experiment, body size was not a significant predictor of male response level

50

Signaling system - motivation to fight Alternatively, the differential changes in response

levels within individuals may signal motivation to fight (Hauser and Nelson 1991; Hurd and Enquist 2001; Burmeister et

al. 2002; Lange and Leimar 2003). Motivation is expected to vary over a much shorter

time scale than fighting ability. Since the responses of the frogs in our study were

highly variable during the 35-min timeframe, this variation more likely reflects motivation to fight rather than just simply fighting ability.

An intruder potentially could use pre-response or initial response levels of calling and movements as well as the dominant frequency of advertisement calls to assess fighting ability and size of a resident.

51

The intruder then could use these changes in the number of calls, movements, and other information as the encounter escalated to assess the aggressive motivation of the resident.

Calls are well documented as threat displays, and presumably movements (and possibly other visual displays) serve in this capacity as well.

However, a definitive test of the role of motivation in these responses requires assessing threat displays as well as the responses to them in both signalers and receivers (Hauser and Nelson 1991).

52

Our data show that male green frogs increase their aggressive responses in relation to perceived intruder threat.

However, the exact costs involved and the functional significance of these responses remains uncertain.

No studies have quantified the physiological costs of graded responses in frogs.

Only a few studies of frogs have examined whether differences in graded aggressive calls are detected by receivers (Schwartz 1989, 1994; Wells 1989;

Burmeister et al. 1999, 2002), and other kinds of graded responses such as movements have not been investigated.

53

It is likely that graded responses in green frogs reduce energetic and other potential costs of aggression.

However, it is not known whether they serve as graded threat displays to conspecific males.

Future work will examine whether these observed changes in aggressive responses are perceived by receivers and represent information that can be used to assess fighting ability or aggressive intent.

54

YOU WILL ONLY REGRET HOW LITTLE YOU HAVE LEARNT WHEN YOU NEED TO UTILIZE WHAT YOU HAVE LEARNT

The end

55

Dominant frequency changes were quite variable among individuals (see Appendix).

56

57

Response thresholds Green frogs in our study may have had thresholds

for aggressive responses, and it is possible that our stimulus broadcasts did not exceed the aggressive thresholds of the frogs in all instances.

Thus, the overall picture of graded responses could be an artifact of proportionally more frogs giving responses at each successive stimulus level due to individual differences in aggressive thresholds.

Studies of both Pacific treefrogs (Hyla regilla) and spring peepers (Pseudacris crucifer) used the production of aggressive calls that were discrete in structure from advertisement calls to determine the exact onset of an aggressive response (Brenowitz et al. 2001; Marshall et al. 2003).

58

In almost all cases in our study, individual green frogs reacted to the stimulus at all broadcast levels with movements and call responses (see Appendix).

Since advertisement and aggressive calls in this species are continuous in structure, they are not ideal for determining the threshold onset of an aggressive response.

Movements could be a better indicator since the frogs rarely moved during non-broadcast periods.

Four individuals did not move in response to the 83 dB stimulus, and of those four, three did move in response to the 90 dB stimulus.

59

All of our subject frogs moved in response to the 97 dB stimulus.

However, in five of the six total cases in which the frogs were not moving, the calling rate was elevated or the dominant frequency and duration of calls had decreased compared to the baseline.

Thus, there were really only two instances where our stimulus had not exceeded the aggressive thresholds of the subjects for at least one measure of response, and the instances where we did not exceed their thresholds probably exert little influence on the overall interpretation of individuals showing graded responses.

60

This issue is not easily resolved within the context of a balanced experimental design, and additional work is needed to determine whether green frogs have aggressive thresholds and whether these thresholds act in concert with graded response levels during the escalation of encounters.

61