Anuran Population Monitoring: Comparison of the North

American Amphibian Monitoring Program's Calling Index

with Mark-Recapture Estimates for Rana clamitans
Author(s): Gerald L. Nelson and Brent M. Graves
Source: Journal of Herpetology, 38(3):355-359. 2004.
Published By: The Society for the Study of Amphibians and Reptiles
DOI: http://dx.doi.org/10.1670/22-04A
URL: http://www.bioone.org/doi/full/10.1670/22-04A

BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the

biological, ecological, and environmental sciences. BioOne provides a sustainable online
platform for over 170 journals and books published by nonprofit societies, associations,
museums, institutions, and presses.
Your use of this PDF, the BioOne Web site, and all posted and associated content
indicates your acceptance of BioOnes Terms of Use, available at www.bioone.org/page/
terms_of_use.
Usage of BioOne content is strictly limited to personal, educational, and non-commercial
use. Commercial inquiries or rights and permissions requests should be directed to the
individual publisher as copyright holder.

BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit
publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to
critical research.

Journal of Herpetology, Vol. 38, No. 3, pp. 355359, 2004

Copyright 2004 Society for the Study of Amphibians and Reptiles

Anuran Population Monitoring: Comparison of the

North American Amphibian Monitoring Programs Calling
Index with Mark-Recapture Estimates for Rana clamitans
GERALD L. NELSON

AND

BRENT M. GRAVES1

Department of Biology, Northern Michigan University, Marquette, Michigan 49855, USA

ABSTRACT.The North American Amphibian Monitoring Program uses a ranked, categorical calling index to
estimate anuran abundance. However, there are few data assessing the assumption that calling index values
are accurate indicators of population sizes or to suggest ranges of population sizes associated with specific
values of the calling index. This study compared mark-recapture population estimates and call rates to calling
index values for Green Frogs (Rana clamitans) in the Upper Peninsula of Michigan. Mean mark-recapture
population size estimates and mean calls per minute were greater in ponds with larger calling index values.
Similarly, calls per minute increased with increasing population size, although rate of increase declined at
high population sizes possibly caused by a higher proportion of noncalling satellite males at high densities.
Sex ratios in breeding habitat along the edges of ponds were increasingly male biased as population size
increased. These data support the assumption that calling index values are useful indicators of abundance of
R. clamitans.

Apparent reduction, and even extirpation, of

amphibian populations in seemingly pristine
habitats have alarmed the scientific community,
and led to intensified research on amphibian
abundance (Blaustein and Wake, 1990; Pechmann
and Wilbur, 1994). Vitt et al. (1990) suggested that
amphibian population declines indicate a stressed
environment associated with decreased water
quality and ecosystem biodiversity. The complex
life cycles, aquatic eggs, and permeable skin of
many amphibians are thought to make them
especially sensitive to environmental contaminants (Blaustein, 1994). Amphibians are considered bioindicators of environmental quality
(Power et al., 1989; Licht and Grant, 1997;
Lannoo, 2000). Additionally, amphibians play
important roles in a variety of ecological communities (Fisher and Shaffer, 1996; Hayes et al.,
2002). Hence, information concerning amphibian
populations may be important for understanding
and managing ecological systems.
Various amphibian monitoring programs have
been implemented across the United States and
Canada to obtain baseline data on population
densities and distributions. Such long-term data
will allow researchers to determine whether
apparent amphibian population declines reflect
normal fluctuations or temporally and geographically consistent trends. Many of these programs
enlist volunteers to assess anuran abundance
using a calling index (CI; Lepage et al., 1997).
Initially, methods varied between states and
1

Corresponding Author. E-mail: bgraves@nmu.edu

monitoring groups, making compilation and

comparison of data difficult. In 1996, the need
for a unified protocol prompted 44 states and
provinces, under the auspices of the North
American Amphibian Monitoring Program
(NAAMP), to adopt the Wisconsin survey protocol (Heyer et al., 1994; Sargent, 2000).
The NAAMP protocol assigns categorical CI
values to anuran populations on the basis of
vocalization frequency (Sargent, 2000). The CI is
simple, allowing data on anuran population
densities to be collected by trained volunteers.
Increasing values of CI are assumed to be
positively correlated with anuran abundance
(Sargent, 2000). However, there is little evidence
to support this claim (Zimmerman, 1994; Driscoll,
1998), and there are no data indicating the range
of population densities associated with specific
CI values. Calling rates (not CI values) have been
shown to be correlated with population density
among Bufo fowleri and Rana catesbeiana (Shirose
et al., 1997), as well as several species of
Eleutherodactylus (Fogarty and Vilella, 2001).
Crouch and Paton (2002) estimated population
sizes associated with each CI value for seven
anuran species, although these were subjective,
rather than quantitative, statistical estimates.
The objectives of this study were to determine
whether NAAMP CI values are positively correlated with other population estimates for Rana
clamitans and to estimate population sizes associated with each CI value. Additionally, we
investigated the relationship between categorical
CI values and call rates. The latter are slightly
more time consuming to obtain but may provide

356

G. L. NELSON AND B. M. GRAVES

TABLE l. Means, SE, and 95% confidence intervals

for indicators of Rana clamitans population sizes in
ponds categorized by calling index (see text for
abbreviations).
Calling Number of Population
index
ponds
indicator

0
0
0
1
1
1
2
2
2

5
5
5
25
25
25
12
12
12

C/M
MRtotal
MRmale
C/M
MRtotal
MRmale
C/M
MRtotal
MRmale

Mean

SE

Confidence
interval

0.00
0.50
0.00
8.86
10.77
6.64
30.53
37.83
27.43

0.00
0.50
0.00
1.39
1.50
0.98
2.75
6.40
5.73

00
02
00
612
814
59
2536
2551
1639

a more accurate indicator of population density.

We also monitored sex ratios in habitats where CI
values were obtained and examined the relationship between CI values, numbers of males in
populations, and total population sizes. This is
important because only males call, although
densities of both sexes are important for monitoring population trends.
MATERIALS AND METHODS
Populations of R. clamitans were studied in
Delta and Menominee Counties in the Upper
Peninsula of Michigan. Male R. clamitans call and
establish breeding territories in permanent bodies
of water (Wells, 1977). Single-note advertisement
calls are produced throughout the day, but call
rates are highest at night (Wells, 1978). To reduce
ecological variation between ponds, only humanmade ponds with perimeters
450 m were
included in the study. However, ponds were
situated in a variety of habitats including cedar
swamps, golf courses, meadows, marshes, and
adjacent to residential yards. Forty-two ponds
were studied between 11 June and 14 July 2002
(GPS coordinates are available in Nelson, 2003).
Population sizes were estimated using (1)
calling index (CI), (2) calls per minute (C/M),
and (3) mark-recapture (MR) techniques. Calling
index values were assigned, and C/M were
quantified under the restrictions of phenology,
wind speed, temperature, humidity, and time
specified by the NAAMP (Sargent, 2000). Calling
data collection began after 2000 h and was
completed before 2400 h. Calling index values
were assigned using NAAMP definitions (Sargent, 2000). A CI of 0 was assigned if no frogs
were heard calling. A CI value of 1 was assigned
when vocalizations were heard, but there was no
temporal overlap between calls. A CI of 2 was
recorded when vocalizations overlapped temporally, but there were some periods of time when
no vocalizations were heard. A CI value of 3 was

recorded when vocalizations were heard continuously. Calling index values were obtained
independently by two observers. If there was
a discrepancy, observations were repeated until
there was consensus. Calls per minute were
quantified by counting the number of calls
during five, one-minute intervals. The mean of
these five samples was used as the C/M value for
a given pond.
Mark-recapture population estimates were
initiated on the day following CI and C/M
estimation at a given pond. One person waded or
canoed the pond periphery and located frogs
visually. A second surveyor, located a few meters
inland from the first, recorded data and aided in
capture of frogs that moved in that direction.
Frogs were captured by hand or with a net. Only
frogs of adult size (i.e. . 5.7 cm; Conant and
Collins, 1991) were included in the count. In
summer, adult frogs were distinguishable from
newly metamorphosed juveniles by a large difference in size (Martof, 1956); juveniles were not
sighted commonly (Martof, 1956; Wells, 1977).
All captured R. clamitans were marked by toeclipping (Martof, 1953). Digit wounds were
disinfected with antiseptic (Bactine; Bayer;
Pittsburgh, PA), and scissors were dipped in
ethanol before each frog was marked. Male frogs
were identified on the basis of ventral coloration,
size of the tympanum, and presence of nuptial
pads on the forelimbs. Each frog was released at
its point of capture. Recapture efforts occurred 24
h later using the same methods. Mark-recapture
population estimates (MRtotal) were derived from
the Peterson index, with Baileys formula applied
when the number of frogs recaptured in a pond
was , 10 (Heyer et al., 1994). The number of
males in the population (MRmales) was determined similarly but using only data from
captured males.
The programs Prophet (Market Miner Inc.,
2001) and JMP (SAS Institute, 1997) were used for
statistical analyses. All data were normalized by
log (x) 10 transformations, after which normality and homoscedasticity were confirmed
prior to analysis.
RESULTS
More than 12 male frogs were estimated to be
present in nine of the 42 ponds, yet no pond had
a CI of 3 (Table 1). Mean MRtotal (F2,39 5 30.76;
P , 0.001), and MRmales (F2,39 5 29.5; P , 0.001)
increased with increasing CI values. Both MRtotal
and MRmales differed significantly (Tukeys multiple comparisons; P , 0.05) between all CI pairs.
Similarly, mean C/M increased with increasing
CI, and these differences were significant (F2,39 5
41.62; P , 0.001). Furthermore, there was
a curvilinear relationship between MRmale population size and C/M, with an asymptote near 35

ANURAN CALLING INDEX

FIG. 1. Relationship between male population size

of Rana clamitans and calls per minute (r2 5 0.56;
F1,40 5 51.5; P , 0.001).

C/M (Fig. 1; C/M 5 1.89 1.031 log (MRmales);

r2 5 0.56; F1,40 5 51.5; P , 0.001).
Mark-recapture estimates of male population
sizes were positively correlated with MRtotal (r2
5 0.918; F1,40 5 453.3; P , 0.001), presumably
because the majority (72%) of frogs captured
around pond perimeters in midsummer were
males. More substantively, a regression comparing MRmales to number of females (i.e., MRtotal
minus MRmales) indicated a curvilinear relationship (Fig. 2; female frogs 5 1.04 0.599 log (male
frogs); r2 5 0.60; F1,40 5 60.4; P , 0.0001).
Although up to 54 males were estimated to be
present in a single pond, maximum number of
females in a pond was 12. Overall sex ratio was
approximately 2.5:1 (399 males:159 females),
which is significantly different from 1:1 (v2 5
54.0; df 5 1; P , 0.001).
DISCUSSION
The NAAMPs calling survey is an important
tool for monitoring anuran populations. For CI to
be a useful estimate of population size, a multispecies comparison of CI values with more
rigorous population estimation methods (i.e.,
MR) is necessary (Shirose et al., 1997). Results
presented here demonstrate that population sizes
were larger in ponds with higher CI values and
identify probable population sizes associated
with each CI value (Table 1). In general, only
male anurans call. Therefore, a correlation between male population size and total population
size is necessary for quantifications of calling to
be useful estimates of population size, which we
demonstrated. Furthermore, mean C/M differed
significantly between ponds assigned to each CI
value, and C/M was correlated with population
size. Thus, higher values of the CI indicate higher
calling rates, higher calling rates indicate higher
male population sizes, and higher male popula-

357

FIG. 2. The relationship between female population

size and male population size for Rana clamitans (r2 5
0.60; F1,40 5 60.4; P , 0.0001).

tion sizes indicate higher total population size.

This series of correlations, as well as the direct
association between increasing population size
and increasing CI value, supports use of the
NAAMP survey protocol as a method to assess
anuran population densities.
Despite the demonstrated utility of the methods used in this study, care should be taken in
generalization of our conclusions. Our data were
derived from a single species, and only in
human-made ponds with perimeters
450 m
in the southwestern Upper Peninsula of Michigan. Extrapolation to other species, locales, and
habitat types, especially with regard to population means and ranges associated with specific CI
values, would be inappropriate. The danger of
generalization between species is illustrated by
the absence of CI values of 3 in this study.
Because the call of R. clamitans is a single syllable,
it is unlikely that there would ever be continuous
temporal overlap of calls for 35 min, as required
by the NAAMP calling index protocol. Thus, a CI
value of 2 in a population of R. clamitans is likely
to reflect a very different range of population
densities than it would, for example, in a population of Bufo americanus, in which individual
vocalizations may have durations of 30 sec or
more.
Sargent (2000) stated that CI values of 0, 1, 2,
and 3 indicate populations of 0, 15, 612, and .
12 males, respectively, with the implication that
this would be true for all anuran species. Lepage
et al. (1997) suggested that CI values of 0, 1, 2,
and 3 indicate male populations of 0, 1, 5, and 15,
respectively, again with reference to multiple
species. Crouch and Paton (2002) suggested male
population sizes of 0, 110, 1135, and . 35 for
these CI values, with regard to seven anuran
species in Rhode Island. However, none of these
predictions were based on quantitative data. Confidence intervals for population sizes in ponds
categorized under each CI value generated in this

358

G. L. NELSON AND B. M. GRAVES

study indicate an underestimation of male

population size by Lepage et al. (1997) and
Sargent (2000), but our estimates are similar to
those provided by Crouch and Paton (2002).
Our data also show that use of call rates as
indicators of population size becomes increasingly tenuous as density increases. At least two
factors may limit the utility of this method. First,
in a number of species, a proportion of males are
unable to obtain territories as density increases,
and such frogs exhibit noncalling satellite behavior (Arak, 1988; Ovaska and Hunte, 1992). As the
proportion of noncalling males in the population
increases, the validity of population size estimates based on vocalizations decreases. Our data
are consistent with this possibility, and this is
reflected in the decreasing slope of the relationship between C/M and number of males (Fig. 1).
The curve should approach an asymptote at a C/
M value associated with the occupation of all
available territories. A second cause of nonlinearity between calling and population size is
that males comprised a larger proportion of the
population as total population size increased
(Fig. 2). This pattern has not been reported
previously, and we made no observations that
might indicate its cause. Both factors produce
a nonlinear relationship between population
density and calling rates, although the two
would have opposite effects on total population
size estimates if one assumes equal proportions
of males and females in populations. That is, the
first factor would lead to underestimation of the
number of males, and consequently, females,
whereas the latter factor would lead to overestimation of females.
Almost three-fourths of all frogs captured in
the study were males, which is similar to the sex
ratio reported by Martof (1956) for R. clamitans in
breeding habitats in southern Michigan. This
may simply reflect a tendency of males to remain
in the breeding area along the edges of ponds
throughout the prolonged breeding season,
whereas females occupy habitat away from the
ponds except during mate choice and oviposition
(Martof, 1956; Wells, 1977). Hence, proportions of
males and females in our study likely reflect the
operational sex-ratio, rather than the population
sex-ratio. This finding violates an assumption of
the Peterson index, which requires that individuals captured must be an unbiased sample from
the entire population (Heyer et al., 1994). A
population sex ratio of 1:1 is common among
anurans (Martof, 1956; Duellman and Trueb,
1986). If we assume a 1:1 sex ratio in the
populations studied here, 60% of the female
population was not sampled. Thus, numbers of
frogs estimated using MR methods in this study
must be assumed to represent the proportion of
the population using shoreline habitat, rather

than the entire population. Mark-recapture population estimates also assume that marked
individuals are distributed randomly in the
population prior to recapture. This assumption
was probably violated as well, because male R.
clamitans are territorial. Therefore, individuals in
accessible territories would be more likely to be
captured and recaptured than individuals occupying less accessible or obvious sites.
Our data suggest that population monitoring
using the NAAMP calling index provides useful
information concerning the relative abundance of
anurans. Although the categorical data provided
by the CI are not as precise as continuous data
associated with counts of calls per minute, the
former is simpler for volunteers to quantify, and
there is a good correlation between the two
variables. Additionally, higher values of the
calling index are indicative of higher population
densities in small human-made ponds within
a range of habitat types occupied by R. clamitans.
However, extrapolation beyond the species and
ecological conditions of this study should be
made with considerable caution. Further research should test for such a relationship across
a greater range of taxa, geographic locations, and
ecological conditions.
Acknowledgments.We thank J. Bird and J.
Bruggink for helpful suggestions throughout this
project. Funding was provided by citizens who
contributed to the Nongame Wildlife Fund of the
Michigan Department of Natural Resources and
an Excellence in Education Award from Northern Michigan University. This work represents
a portion of the masters thesis of the senior
author.
LITERATURE CITED
ARAK, A. 1988. Callers and satellites in the Natterjack
Toad: evolutionarily stable decision rules. Animal
Behaviour 36:416432.
BLAUSTEIN, A. R. 1994. Chicken Little or Neros fiddle?
A perspective on declining amphibian populations.
Herpetologica 50:8597.
BLAUSTEIN, A. R., AND D. B. WAKE. 1990. Declining
amphibian populations: a global phenomenon?
Trends in Ecology and Evolution 5:203204.
CONANT, R., AND J. T. COLLINS. 1991. A Field Guide to
Reptiles and Amphibians: Eastern and Central
North America. 3rd ed. Houghton Mifflin, Boston,
MA.
CROUCH III, W. B., AND P. W. C. PATON. 2002. Assessing
the use of call surveys to monitor breeding anurans
in Rhode Island. Journal of Herpetology 36:185
192.
DRISCOLL, D. A. 1998. Counts of calling males as
estimates of population size in the endangered
frogs Geocrinia alba and G. vitellina. Journal of
Herpetology 32:475481.
DUELLMAN, W. E., AND L. TRUEB. 1986. Biology of
Amphibians. McGraw-Hill Book Co., New York.

ANURAN CALLING INDEX

FISHER, R. N., AND H. B. SHAFFER. 1996. The decline of
amphibians in Californias Great Central Valley:
historical patterns and anthropogenic causes. Conservation Biology 10:414425.
FOGARTY, J. H., AND F. J. VILELLA. 2001. Evaluating
methodologies to survey Eleutherodactylus frogs in
montane forests of Puerto Rico. Wildlife Society
Bulletin 29:948955.
HAYES, T. B., A. COLLINS, M. LEE, M. MENDOZA, N.
NORIEGA, A. A. STUART, AND A. VONK. 2002.
Hermaphroditic, demasculinized frogs after exposure to herbicide atrazine at low ecologically
relevant doses. Proceedings of the National Academy of Science 99:54765480.
HEYER, W. R., M. D. DONNELLY, R. W. MCDIARMID, L. C.
HAYEK, AND M. S. FOSTER (eds.). 1994. Measuring
and Monitoring Biological Diversity, Standard
Methods for Amphibians. Smithsonian Institution
Press, Washington, DC.
LANNOO, M. J. 2000. Conclusions drawn from the
malformity and diseases section, Midwest Declining Amphibians Conference, 1998. Journal of the
Iowa Academy of Science 107:212216.
LEPAGE, M., R. COURTOIS, AND C. DAIGLE. 1997. Surveying calling anurans in Quebec using volunteers. In
D. M. Green (ed.), Amphibians in decline: Canadian
studies of a global problem. Herpetological Conservation 1:128140.
LICHT, L. E., AND K. P. GRANT. 1997. The effects of
ultraviolet radiation on the biology of amphibians.
American Zoologist 37:137145.
MARTOF, B. S. 1953. Home range and movements of the
Green Frog, Rana clamitans. American Midland
Naturalist 34:529543.
. 1956. Factors influencing size and composition
of populations of Rana clamitans. American Midland Naturalist 56:224245.
NELSON, G. 2003. A Comparison of the North American
Amphibian Monitoring Programs Calling Index
with Population Estimates of Green Frogs (Rana
clamitans) in the Upper Peninsula of Michigan.

359

Unpubl. masters thesis, Northern Michigan Univ.,

Marquette.
OVASKA, K., AND W. HUNTE. 1982. Male mating behavior
of the frog Eleutherodactylus johnstonei (Leptodactylidae) in Barbados, West Indies. Herpetologica
48:4049.
PECHMANN, J. H. K., AND H. M. WILBUR. 1994. Putting
declining amphibian populations in perspective:
natural fluctuations and human impacts. Herpetologica 50:6584.
POWER, T., K. L. CLARKE, A. HARFENIST, AND D. B.
PEAKALL. 1989. A Review and Evaluation of the
Amphibian Toxicological Literature. Canadian
Wildlife Service Headquarters, Technical Report
61, Ottawa, ON.
SARGENT, L. 2000. Frog and toad population monitoring
in Michigan. Journal of the Iowa Academy of
Science 107:195199.
SHIROSE, L. J., C. A. BISHOP, D. M. GREEN, C. J.
MACDONALD, R. J. BROOKS, AND N. J. HILBERT. 1997.
Validation tests of an amphibian call count survey
technique in Ontario, Canada. Herpetologica
53:312320.
VITT, L. J., J. P. CALDWELL, H. M. WILBUR, AND D. C.
SMITH. 1990. Amphibians as harbingers of decay.
BioScience 40:418.
WELLS, K. D. 1977. Territoriality and male mating
success in the Green Frog (Rana clamitans). Ecology
58:750762.
. 1978. Territoriality in the Green Frog (Rana
clamitans): vocalizations and agonistic behaviour.
Animal Behaviour 26:10511063.
ZIMMERMAN, B. L. 1994. Audio strip transects. In R.
Heyer, M. A. Donnelly, R. W. McDiarmid, L. Hayek,
and M. S. Foster (eds.), Measuring and Monitoring
Biological Diversity. Standard Methods for Amphibians, pp. 9297. Smithsonian Institute Press,
Washington, DC.
Accepted: 4 May 2004.