Population Dynamics, Reproduction, and Life History Traits of Taurus Frog, Rana Holtzi Werner, 1898 (Anura: Ranidae) in Karagöl (Ulukışla, Niğde), Turkey

HERPETOLOGICA ROMANICA Vol. 6, 2012, pp.1-40 ISSN: 1842-9203 Article No. 121101

Population dynamics, reproduction, and life history traits of Taurus Frog, Rana holtzi Werner, 1898 (Anura: Ranidae) in Karagöl (Ulukışla, Niğde), Turkey

Mehmet Zülfü YILDIZ1,* and Bayram GÖÇMEN2

1. Harran University, Faculty of Arts and Sciences, Department of Biology, Zoology Section, Osmanbey Campus, 63300 Şanlıurfa, Turkey 2. Ege University, Faculty of Science, Department of Biology, Zoology Section, 35100 Bornova-Izmir, Turkey. * Corresponding author, M.Z. Yildiz, Tel: +90 414 318 35 58, Fax: +90 414 318 35 41, Mobile Phone: +90 506 531 46 63, E-mail: [email protected]

Abstract. In this study, Taurus Frog’s, Rana holtzi’s, population structure, population size, and population density were investigated using mark-recapture method at Karagöl (Ulukışla, Niğde) population. Population age, the age of maturity, and life span were determined using skeletochronology. Furthermore, reproduction ecology, reproduction and activity periods and environmental conditions, their possible effects on population, and precautions that should be taken were determined. Mean snout vent length (SVL) of males was of 49.65±4.91 mm females was 45.74±6.03 mm and juveniles was 33.68±3.49 mm. Male individuals’ mean weight was concluded as 11.78±3.81 g while females’ was 9.23±4.22 g and juveniles’ was 3.66±1.03 gr. During three years, which covered 2007–2009, a total of 5151 individuals (1344 ♂♂, 1828 ♀♀ and 1979 juvenile) were marked. According to Jolly-Seber formula, population size was calculated to be 17563 (SE=1346.93, Min-Max= 14923–20203); survival rate ranged from 0.37 and 1 with a mean of 0.83±0.25; capture rate ranged from 0.07 to 0.39 with a mean of 0.16±0.096. Population density was determined as 0.29 (individual/m2). Mean age of males was 4.87±0.97, of females was 4.29±1.38, and of juveniles was 2.22±0.99 in Karagöl population .Among the examined specimens, the oldest age found as IX belonging to only one female individual. Egg clutches of approximately 464±277 eggs were measured and they hatched in 5-8 days depending on environmental factors. From 20th Gosner phase all tadpoles’ morphological features were determined in all Gosner phases.

Key Words: Rana holtzi, Taurus frog, Bolkar Mountains, population dynamics, mark- recapture, breeding ecology, skeletochronology.

©Romanian Herpetological Society,Cluj-Napoca / Oradea, Romania, 2012 Herpetol. Rom, 6, http://herpetofauna.uv.ro/herprom.html 2012, Romania 2 Yıldız, M.Z. & Göçmen, B.

INTRODUCTION

The global amphibian decline phenomenon (Beebee and Griffiths 2005), associated with population decreases and extinctions in many regions of the world (Houlahan et al. 2000), has enormously increased the need, and for ecological studies of amphibian, especially those addressing population dynamics. Nearly 168 species were believed to have gone extinct and at least 2,469 more had populations that were declining (AmphibiaWeb 2012). In many studies, reduction of amphibian population and necessity of ecological studies were emphasized (Pechman 1994, Meyer et al. 1997, Alfold and Richards 1999, Broadman et al. 2006). The estimation of basic demographic parameters and the identification of their determinants are essential to improve the understanding of population dynamics and life histories of amphibians (Schmidt et al. 2002). Many recent advances have been made in population modeling analysis using capture–recapture models to ac- curately estimate demographic parameters such as population size and separate estimates of survival and capture probabilities, providing important tools for the identification of population trends (Lebreton et al. 1992, Schmidt 2003). Taurus frog, Rana holtzi Werner, 1898 is a brown frog species endemic to the Bolkar Mountains in the Taurus range of southern Turkey. It is known from only in two lakes: Karagöl Lake (2588 m a.s.l) approximately 60 hectare, Çinigöl Lake (2673 m a.s.l) about 22.5 hectares and some springs near Eğrigöl Lake (2777 m a.s.l). Rana holtzi was listed as a vulnerable species under the category of endangered species by the International Union for the Conversation of Nature (IUCN) for the first time in 1990. However, due to a serious decrease in population size it was listed as rare in 1994, as Endangered in 1996. Finally, since 2008 it has been listed as Critical Endangered. Several studies have been carried out on its taxonomy (Baran 1969, Veith et al. 2003), morphology and serology (Çevik et al. 2006), population size (Baran et al. 2001, Kaya et al. 2005, Göçmen et al. 2008, Kaya et al. 2010), and age structure (Miaud et al. 2007, Guarino and Erişmiş 2008). Although a few studies have been carried out about R. holtzi, there is no comprehensive and longitudi- nal study about its population dynamics, reproductive and life history traits. In this context, the main aim of this study is to determine its population size, structure and density, population age, age of maturity, life span, reproductive ecology, and environmental conditions, their possible effects on population, and the precautions that should be taken.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 3

MATERIAL AND METHODS

Study area The Bolkar Mountains are located in the eastern part of the central Taurus Ranges, extend- ing southwest and northeast for about 40-50 km width and 150 km length, a massif in southern Anatolia. Karagöl Lake (37° 24' 10.12'' N, 34° 33' 28.36'' E, 2588 m a.s.l.) is located in the Bolkar Mountains at the intersection of Maden village, Ulukışla, and Niğde province. It is a tectonic lake covering area approximately 60 ha, 430 m in length 80-160 m in with and 12 m at maximum depth. Karagöl Lake is supplied by snow melt and two springs, one is perma- nent and the others are temporary, and another spring may be connected to Çinigöl Lake in south east. Southwest coast of Lake is surrounded by high and steep rocks, that is why southwest side has poor vegetation and lake is often deeper. The other sides of the lake con- sist of flat and shallow areas and are covered with dense alpine vegetation. Çinigöl Lake (37° 23' 56.95'' N, 34° 33' 15.00'' E, 2673 m a.s.l.) is located on the southwest of Karagöl Lake and 85 m higher. Its length is 210 m and width ranges from 60-150 m. It is surrounded by high mountains; therefore, less sunlight reaches the surface of Çinigöl Lake. Çinigöl Lake does not have any vegetation except near 20 meter on the southwest side. Data logger (Hobo Micro Station, Onset Computer Corporation, USA) was placed near Karagöl Lake and programmed to record air temperature, water temperature, air pressure, and relative humidity per hour. It recorded the data from 17th May 2007 to the end of study. Data were transferred to computer and calculated mean, minimum, and maximum value for each parameter. During Mark-recapture study day, pH (WTW 315i pH) and Oxygen parameters (WTW 340i) were measured. Calcium (mg/l), silicon (mg/l), phosphate (mg/l), nitrite (mg/l), nitrate (mg/l), ammonium (mg/l), ferrous (mg/l), and free chlorine (mg/l) were determined with Merckaquant kit.

Field Surveys Mark-Recapture Studies: Mark-recapture studies were performed for each of three seasons (spring, summer, and autumn) each year during May 2007-November 2009 to calculate population size and other relevant parameters (capture probability, survival rate etc.). Specimens were collected by dip net or hand. As work areas were located in highlands, activity starts at the end of the spring season (around the end of May). R. holtzi is a critical endangered species, we waited until eggs had hatch (until June or July) as not to affect the population negatively during the spring surveys. Therefore, mark-recapture studies in spring season were carried out between June and first week of July. The field surveys in summer were carried out in August while in autumn they were performed between Sep- tember and early October. The sex of mature marked frogs was determined based on by the presence or absence of nuptial callus tissue. Snout-vent length (SVL, mm) was measured by digital calipers (Mitu- tuyo 500-181 U) with an accuracy of 0.02 mm and weight (W, g) was measured with a sensi- tive balance (Persica, BJ 410 C) with 0.01 g accuracy after drying with blotting paper. Specimens were marked with toe clipping in 2007 according to Donnelly method (Don- nelly et al. 1994) three times in each season within two hours collecting time, then VI Nu-

Herpetol. Rom, 6, 2012 4 Yıldız, M.Z. & Göçmen, B. meric Tags were used and individuals were marked six times each season. Because VI Al- pha Numeric Tags took a long time so collecting time was reduced to 30 min for the animal’s safety not to keep waiting individuals in cloth bag for such a long time. Only individuals were 30 mm or bigger were marked. VI numeric tags were injected right hind limb third toe (Fig. 1). Before marking, to prevent the individual from feeling the pain, anestol pomade (5% lidakoin HCl, Sandoz İlaç San ve Tic Aş.) was applied topically to prevent infec- tion, Fucidin H pomade (fusidic acid+hydrocortisone acetate, Abdi İbrahim İlaç San. ve Aş.) was applied to injection area after the individual was marked,. In recaptured individuals the injection area healed in two weeks.

Figure 1. Appearance of VI alpha numeric tags injected in third toe.

Photos of all marked specimens were taken with Nikon Coolpix L1 camera and photo- graphs were organized to tag numbers; thus, the changes observed individually.

Activity period survey In order to determine activity period, field surveys were carried out in 15 day intervals between April and June with three researchers to Karagöl Lake. Surveys were carried out at between 12:00-14:00 pm. The beginning of activity period was determined as first day that any individual (tadpoles, juvenile or adult) was observed in or around the lakes, date were accepted the beginning of activity period. The date when no individuals were found in X days was accepted as the end of activity period.

Reproductive ecology The breeding period usually started 10-15 days after the activity period beginning. During breeding period we camped at Karagöl Lake to determine breeding phenology. Observa- tions were carried out both day and night. Mobile flashlight was used in night surveys. Amplexus individuals were collected, marked and after measuring SVL and weight they released to the lake again. When reproductive period was over, amplexus individuals were

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 5 collected during night surveys and weighted again. Reproductive investment was calculated to measure the difference before and after reproductive period weights. Formula, Ri=

(Wp-Ws/Wp)*100 (Ri= Reproductive investment, Wp= weight before reproductive, Wp= weight after reproductive). Egg clutches were calculated with volume methods (Surova and Cherdantsev 1987, Cherdantsev et al. 1997). Egg diameter was measured at 10th Gosner stage at 921 eggs from 25 different clutches, under the stereomicroscopes with micrometric ocular. 20 clutches volumes were determined and after hatchling unfertilized and undeveloped eggs were counted. Two newly laid clutches were moved from Karagöl Lake to amphibian and reptilian ecology laboratory (Ege University, Department of Biology, Zoology Section) and raised in aquarium (45X35X15cm) with ventilation system at 18°C temperature. All stages were iden- tified according to Gosner (1960). Tadpoles were fed ad labitum green algae during 25th and 35th Gosner stage then fed with boiled spinach and banana in addition. Water was changed daily. Tadpoles were anaesthetized with 0.4 % mg/l MS–222 (Tricaine methanesulfonate) at pH 7.0 (adjusted with sodium bicarbonate) and their photos were taken with Olympus SZ- 60 camera connected with Olympus C-7070 stereo-microscope. Measured tadpoles characters: Snout-vent length (SVL), tail length (TL) femur (FU), tibia length (TbU), Tail muscle width (TMW), Tail muscle high (TMH), internostril distance (IND), interorbital distance (IOD). Metamorphic individuals were released to Karagöl Lake. 15-20 tadpoles were fixed from Karagöl Lake after each field survey to compare with laboratory raised tadpoles. To calculate tadpoles’ density, quadrate method (Heyer et al. 1994) was used. Reproduc- tive sites were divided into 0.5m square and tadpoles were counted in one litter containers from randomly selected areas in Karagöl at 12:00 pm. In order to determine sexual maturity age, secondary sexual characters, nuptial pad (callus tissue), and age study were used. When individuals reach sexual maturity, growing decreases and thus distance between LAGs (line arrested growth) diminishes (Francillin- Vieillot et al. 1990, Leclair et al. 2005).

Population Density and Fluctuation Mean population density (d) was calculated dividing the super population size (N*)- calculated for 3 years- to Karagöl Lake’s area (a, m2) (d= N*/a). Population fluctuation (λ) was determined by calculating the difference between the normalized population sizes

[λ=Log (N(i+1)+1)-Log (N(i)+1)] (Houlahan et al. 2000).

Age Study In order to calculate population age structure, the second phalanges of the right hind leg fourth toe were used. Phalanx samples were collected from 138 randomly selected individuals (47 ♂♂, 42 ♀♀ and 49 Juveniles) in Karagöl Lake and stored in 70 % ethanol until trans- port to laboratory. After the skin was removed from phalanges, bone samples were decalci- fied with 5 % nitric acid then embedded in paraffin with routine histological procedures. 10- 12 μm thickness transverse sections were cut with rotary microtome and samples were stained with Harris Hematoxylin (Hematoxylen 5 min Eosin 3 min). LAGs were counted under light and phase-contrast microscopes.

Herpetol. Rom, 6, 2012 6 Yıldız, M.Z. & Göçmen, B.

Statistical Analysis Student t test was used to compare snout-vent length and weight between sexes and morphological characters in tadpoles raised in laboratory and collected from Karagöl Lake. All metric and meristic data were transformed with Log10(X+1) to fit normal dispersal before comparing while arctangent transformation was applied to rates. All analyses were made with 95% confidence interval. The sex ratio was calculated by matching the number of male individuals with the number of female individuals. Chi-square test was used to compare seasonal sex ratio. Additionally, to determine differences sexual dimorphism index (SDI: SVL ♀♀/ SVL ♀♀ or W ♀♀/ W ♀♀) was used (Gibbons and Lovich 1990). Regression analysis was used to determine SVL-reproductive investment, size-weight, age-weight, and age-size correlations. Pearson correlation analysis was used to determine population size and seasonal meteorological data. Karagöl population was accepted as closed population for seasonal calculating parameters related population because each season 3-6 days mark-recapture was performed. Thus, program CAPTURE was used to calculate seasonal population size (N), capture probability (p), and standard error (SE) (Pollock 1974, Otis et al. 1978). To find the most appropriate formula calculating the population size, goodness of fit (GOF) test was applied. According to test results, model heterogeneity (h) was used (Otis et al. 1978, Williams et al. 2002). For calculation annual (for 9-18 times for years) or three years data (for 9 times by combination), Karagöl population was accepted open population thus Jolly-Seber (JS) and Cormack-Jolly- Seber (CJS) formula were used (Cormack 1964, Jolly 1965, Seber 1965, Lebreton et al. 1992, Coach and White 2009) via program POPAN (Schwarz and Arnason 1996) and program MARK (Coach and White 2009). The optimum model was specified according to the lowest value of Akaike’s Information Kriteria [AICc] (Burnham and Anderson 2002; Coach and White 2009). First, Gof test was carried out by Program Release (Burnham et al. 1987) via program MARK. As a result of this test, if Test 2 and Test 3 are meaningful statistically it means that you are not able to use time-dependent model in terms of capture probability and survival rate [Ф (t), p (t)] to calculate population size (Coach and White 2009). Second, optimum model analysis was carried out by CJS formula. To determine optimum model was regarded GOF result and the lowest value of AICc together (Burnham and Anderson 2002, Coach and White 2009). After determining optimum model, survival rate, capture probability, and survival rate for marked population were calculated. Population size (N) capture probability (p) and survival rate and population gain (PENT) were calculated using JS formula via program POPAN.

RESULTS

Study area The highest air temperature recorded was 25.44°C while the lowest air temperature was 4.35°C. Temperature data was recorded by data logger as shown in Fig. 2. The highest water temperature recorded was 25.55°C while the lowest air temperature

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 7 was -0.31°C (Fig. 2). Air pressure and relative humidity were showed in diagrams (Fig. 3 and Fig. 4).

Air Temp (°C) Water Temp (°C)

20,00

15,00

10,00

(°C) 5,00

0,00 I II III IV V VI VII VIII IX X XI XII -5,00 Months

Figure 2. Water and air temperature were recorded by data logger from May 2008 to November 2009.

Relative Humidity (%)

100,00 90,00 80,00 70,00 60,00 50,00 (%) 40,00 n 30,00 20,00 10,00 0,00 I II III IV V VI VII VIII IX X XI XII Months

Figure 3. Relative humidity was recorded by data logger from May 2008 to November 2009.

Inorganic substances were recorded in Karagöl Lake during all four seasons and results showed that calcium ranges 23-32 mg/l; silicon ranges 02.-05 mg/l; phosphate was under 0.25 mg/l; nitrite ranges 0-0.12 mg/l; nitrate ranges 5-10 mg/l; ammonium was 0.05 mg/l; free chlorine was under 0.3 mg/lt. In Çinigöl Lake, calcium ranges 22-30 mg/l; silicon ranges 0.25.-05 mg/l; phosphate ranges 0-

Herpetol. Rom, 6, 2012 8 Yıldız, M.Z. & Göçmen, B.

0.25 mg/l; nitrite ranges 0-0.5 mg/l; nitrate was 5 mg/l; ammonium ranges 0-0.05 mg/l; free chlorine was under 0.3 mg/lt. Conductivity, pH, oxygen saturation (mbar), and oxygen ratio (%) are shown in Table 1.

Pressure (mbar)

752,00 750,00 748,00 746,00 744,00 742,00

(mbar) 740,00 738,00 736,00 734,00 732,00 I II III IV V VI VII VIII IX X XI XII Months

Figure 4. Air pressure was recorded by data logger from May 2008 to November 2009.

Table 1. Summary case of conductivity, pH, oxygen saturation (mbar), and oxygen ratio (%) recorded in Karagöl Lake.

Oxygen Saturation Oxygen Ratio pH Conductivity (mbar) (%) N 60 60 33 36 Mean 8.29 82.25 159.91 94.74 SD 0.17 9.53 71.56 48.84 Min 7.77 59.10 17.10 11.30 Max 8.53 98.80 301.00 198.40

It was determined that there is a positive correlation between daily captured individuals’ counts and oxygen saturation (P= 0.004) and oxygen ratio (P= 0.011). In addition, there is a positive correlation between the seasonal population size and water temperature (P= 0.05) and conductivity (P= 0.05). Water level was the highest in May and June because of the melting snow and ice in Karagöl Lake, it gradually decreased in summer season approximately 25 cm until autumn. Poligonium amphibium, covers a large part of the lake come out after water level decline (Fig. 5). It is clear that P. amphibium will gradually cause the lake to dry in a long time.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 9

Figure 5. A view of the east of Karagöl Lake with P. amphibium covering the lake (12 July 2008).

Water level of Çinigöl Lake was reduced about 2.5 m from spring to autumn. The lakeshore was deep resulting in lower water temperature. There was no shallow area around Çinigöl Lake to lay eggs and it is possible that low water temperature caused embryonic development to slow down and to raise tadpoles. On the other hand, activity period was as short as 3-4 months. Due to these reasons, Çinigöl’s R. holtzi population growth was inhibited (see section to population size and related parameters).

Morphology During the study, 5071 individual’s snout-vent length and weight were measured. Mean SVL recorded for males was 49.65 mm (n=1309) while mean SVL for females was 45.74 mm (n=1701) (Table 2). It was found out that male SVL were significantly greater than females statistically (t-test, t=19.074, df=3008 P=0.000, SDI=0.92). Mean weight recorded for males was 11.78 g (n=1024) while mean weight recorded for females was 9.23 mm (n=1415). It was also found that males were significantly heavier than females statistically (t-test, t=15.349, df=2436, P=0.000, SDI=0.78).

Population size and related parameters For three years, 2407 individuals (692 ♂♂, 816 ♀♀ and 899 juvenile) in 2007, 1147 individuals (301 ♂♂, 399 ♀♀ and 447 juvenile) in 2008, and 1597 individuals (351

Herpetol. Rom, 6, 2012 10 Yıldız, M.Z. & Göçmen, B.

Table 2. Some merıstic data and derived ratio from investigated specimens.

W (g) TL/FL FL (mm) HL/HW TL (mm) HL (mm) HW (mm) SVL (mm) SVL/FL+TL SVL/FL+TL

N 1309 1024 150 150 150 150 150 150 150 Ort 49.65 11.78 18.01 18.97 27.78 28.73 0.90 0.95 1.04 SE 0.14 0.12 0.33 0.24 0.37 0.36 0.01 0.01 0.01 ♂♂ SD 4.91 3.81 2.62 1.90 2.95 2.80 0.05 0.11 0.05 Min 37.69 4.00 12.98 14.39 20.20 21.19 0.80 0.73 0.93 Max 68.95 28.00 23.91 22.34 33.62 33.62 0.99 1.21 1.23 N 1701 1415 155 155 155 155 155 155 155 Ort 45.74 9.23 17.09 18.55 26.03 27.09 0.95 0.92 1.04 SE 0.15 0.11 0.43 0.47 0.58 0.53 0.01 0.02 0.01 ♀♀ SD 6.03 4.22 2.45 2.68 3.33 3.03 0.06 0.09 0.05 Min 37.52 3.00 12.61 14.57 20.40 22.08 0.86 0.75 0.96 Max 72.34 31.73 22.54 27.00 36.44 35.24 1.12 1.09 1.17 N 2061 1733 103 103 103 103 103 103 103 Ort 33.68 3.66 12.35 12.77 17.06 16.95 0.89 0.99 0.99 SE 0.08 0.02 0.60 1.60 1.61 1.58 0.00 0.10 0.00 Juvenile SD 3.49 1.03 1.03 2.78 2.79 2.73 0.01 0.18 0.00 Min 17.18 1.00 11.34 9.60 13.91 13.87 0.88 0.83 0.99 Max 41.98 7.06 13.40 14.77 19.21 19.08 0.90 1.18 1.00

♂♂, 613 ♀♀ and 633 juvenile) in 2009, in total 5151 individuals (1344 ♂♂, 1828 ♀♀ and 1979 juvenile), were marked. As a result of the capturing program, the lowest seasonal population size calculated was the spring season in 2007 (Table 3). When mark-recapture studies were carried out in spring season, it was colder than the other years’ spring seasons and it was windy. On the other hand, it was performed at an earlier date than the other years. That might be the reason why the population size was calculated lower than the other spring seasons. When Population sizes were compared with the 2007 summer population size, it is seen that it is greater than the other years’ summer seasons’ population size. First year, toe clipping method was used and sampling was performed for two hours. In subsequent

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 11

years, VI alpha numeric tags were used and thus sampling was performed for 30 min (see materials and methods). For this reason, 2007 summer population size may be greater than the other years.

Table 3. Years’ seasonal population size (N), standard error (SE) confidence interval (CI) and recapture probability (p) calculated by the program capture according to model h.

Year Season N SE CI p Spring 813 28.37 762–872 0.17 Summer 3309 54.56 3207–3420 0.22 2007 Autumn 960 30.54 904–1023 0.18 Spring 2037 73.63 1901–2189 0.06 Summer 2151 74.37 2013–2304 0.05 2008 Autumn 1059 52.77 963–1169 0.06 Spring 3130 90.37 2961–3314 0.06 Summer 2526 80.42 2375–2690 0.05 2009 Autumn 1687 66.22 1565–1824 0.06

Spring season mark-recapture studies were performed between June and July not to negatively affect breeding in 2008 and 2009. Hence, population size in spring and summer for these years was close to each other. In addition, in summer season water level decreased and Poligonium amphibium appeared in large parts of the lake making sampling difficult. Thus, summer season population size was calculated lower than the spring season population size. According to the annual population size, results showed that the highest population size calculated was in 2009 while the lowest population size calculated was in 2008 (Table 4). Final population size calculated for three years was 17563 individuals and population density calculated was 0.29 (individual/m2). In Çinigöl population size was counted directly as 155, 157, and 158 in 2007, 2008, and 2009 respectively. Population size increased 0.316 rate in 2009 compared to the size in 2008 and 0.232 rate compared to the size in 2007; whereas, it declined to -0.084 rate in 2008 compared to the size in 2007.

Population structure Rana holtzi’s Karagöl population consisted of 31.57% males, 36.78% females, and 31.65% juveniles. Juveniles were observed to be activated in spring later than and hibernated in autumn earlier than adult individuals. The number of juveniles reached its peak in summer season. Sex ratio of marked specimens either included

Herpetol. Rom, 6, 2012 12 Yıldız, M.Z. & Göçmen, B.

Table 4. Population size (N), survival rate (Ф1), capture probability (p), and population gain (PENT) calculated annually and totally (2007-2009) by jolly Seber formula.

N Ф1 p PENT Mean 6137.74 0.74 0.16 0.12 SE 1529.84 0.10 0.03 0.08 2007 Min 3139.27 0.25 0.03 0.00 Max 9136.22 1.00 0.33 0.59 Mean 5055.08 0.87 0.09 0.05 SE 342.28 0.06 0.04 0.03 2008 Min 4384.22 0.06 0.02 0.00 Max 5725.95 1.00 0.63 0.46 Mean 10473.91 0.85 0.11 0.06 SE 1007.27 0.06 0.06 0.04 2009 Min 8499.65 0.03 0.01 0.00 Max 12448.17 1.00 1.00 0.72 Mean 17563.23 0.83 0.16 0.11 SE 1346.93 0.09 0.03 0.03 Total Min 14923.24 0.37 0.07 0.00 Max 20203.21 1.00 0.39 0.25 more females or was equal. Although in all spring seasons female ratio was higher than male’s, sex ratio was statistically significant only in 2008 and in total per years (Table 5). It was observed that female individuals were more timid than males but breeding period continued in spring seasons; therefore, all individuals were active and the number of females was high. In summers, female and male numbers were close to each other because of female individuals’ timid behavior. It was observed that pregnant females tended to hibernate earlier than males. Thus, sex ratio shifted to including more males or being equal.

Table 5. Seasonal sex ratio (Male: Female) in Karagöl. Bold characters refers to statistically significant sex ratio.

Spring Summer Autumn Annual 2007 1:1.11 1:1.10 1:1.06 1:1.09 2008 1:1.50 1:1.04 1:0.79 1:1.12 2009 1:1.54 1:1.22 1:1.04 1:1.30 Total 1:1.43 1:1.11 1:0.97 1:1.17

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 13

Population age 138 randomly selected individuals were used to determine the age structure of Karagöl’s R. holtzi population (Table 6). Lag (Line of Arrested Growth) was stained dense purple in all bone samples but their optic sharpness was different. It was observed that lags were clearly distinguishable in 112 (81.4%) samples, they looked indistinct in 14 (5.9%) samples and it was quite difficult to distinguish them in 14 (5.9%) samples. Double resting LAGs were observed in 12 (8.6%) samples. Meta- morphosis line was observed only in animals 1-2 years of age as light and weak

Table 6. Age-dependent SVL (mm) distribution in Karagöl population.

Sex Age n % Mean SE SD Min Max III 3 2.2 45.13 3.029 5.246 39.18 49.09 IV 9 6.5 46.66 1.416 4.247 38.64 51.51 V 31 22.5 51.38 0.790 4.400 43.03 59.33 ♂♂ VI 1 0.7 59.90 59.9 59.9 VII 1 0.7 63.73 63.73 63.73 VIII 2 1.4 57.83 0.150 0.212 57.68 57.98 Total 47 34.1 50.79 0.802 5.497 38.64 63.73 III 17 12.3 43.46 1.039 4.282 37.93 50.4 IV 5 3.6 46.84 1.103 2.466 44.24 50 V 16 11.6 48.94 1.252 5.008 39.53 56.85 ♀♀ VI 2 1.4 56.36 2.085 2.949 54.27 58.44 VIII 1 0.7 58.27 58.27 58.27 IX 1 0.7 64.20 64.2 64.2 Total 42 30.4 47.41 0.954 6.182 37.93 64.2 I 19 13.8 22.09 0.404 1.762 20.1 25.79 Juveniles III 30 21.7 34.81 0.360 1.974 30.99 37.97 Total 49 35.5 29.88 0.934 6.538 20.1 37.97 I 19 13.8 22.09 0.404 1.762 20.1 25.79 III 50 36.2 38.37 0.761 5.379 30.99 50.4 IV 14 10.1 46.73 0.963 3.602 38.64 51.51 V 47 34.1 50.55 0.687 4.709 39.53 59.33 Total VI 3 2.2 57.54 1.687 2.922 54.27 59.9 VII 1 0.7 63.73 63.73 63.73 VIII 3 2.2 57.98 0.170 0.295 57.68 58.27 IX 1 0.7 64.20 64.2 64.2 Total 138 100.0 42.34 0.950 11.157 20.1 64.2

Herpetol. Rom, 6, 2012 14 Yıldız, M.Z. & Göçmen, B. line. Endosteal resorption caused two or three years old to lose their metamorphosis line completely. In the following years, it also caused the absorption of first or second LAGs partially or completely which was observed in 82 (59.4%) samples. Endosteal bone was observed in 116 (84.1 %) of bone samples, but not in 23 (15.9%). Mean age calculated for males was 4.87±0.99, for females 4.29±1.38, and for juveniles 2.22±0.99. The maturity age ranged 3-5 for both sexes. But it was determined that females reached maturity earlier than males statistically (Man Whitney- U test, Z= -2.348 P=0.019). The oldest individual detected was a 9 years old female. Mean SVL was 50.79±5.5 for males and 47.41±6.18 for female individuals (Table 6). Positive correlation was observed between Age-SVL (r= 0.87, F=529.615, df=1, P=0.000) (Fig. 6) and Age-W (r=0.8, F=327.487, df=1, P=0.000) (Fig. 7, Table 7).

y = 2,1948x0,513 Age (Year)-SVL (cm) R2 = 0,8732 8 7 6 5 4 3 SVL (cm) SVL 2 1 0 012345678910 Age

Figure 6. Age-SVL curve in Karagöl population.

Age (Year) -W (gr) y = 1,3761x1,2592 R2 = 0,8115 25

W (gr) 10

0 012345678910 Age

Figure 7. Age-W curve in Karagöl population.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 15

Table 7. Age-dependent W (g) distribution in Karagöl population.

Sex Age n % Mean SE SD Min Max III 3 2.2 8.47 1.764 3.056 5.17 11.2 IV 9 6.5 9.49 0.886 2.657 5.37 14.11 V 31 22.5 12.06 0.548 3.050 7.53 18.75 ♂♂ VI 1 0.7 21.79 21.79 21.79 VII 1 0.7 22.70 22.7 22.7 VIII 2 1.4 16.61 0.180 0.255 16.43 16.79 Total 47 34.1 11.97 0.569 3.904 5.17 22.7 III 17 12.3 6.68 0.519 2.141 3.9 10.39 IV 5 3.6 10.92 1.111 2.483 7.45 13.99 V 16 11.6 9.94 0.742 2.969 5.44 15.71 ♀♀ VI 2 1.4 17.48 1.860 2.630 15.62 19.34 VIII 1 0.7 13.45 13.45 13.45 IX 1 0.7 22.18 22.18 22.18 Total 42 30.4 9.47 0.633 4.101 3.9 22.18 I 19 13.8 1.57 0.082 0.359 1.12 2.4 Juveniles III 30 21.7 3.79 0.104 0.569 2.82 4.82 Total 49 35.5 2.93 0.171 1.196 1.12 4.82 I 19 13.8 1.57 0.082 0.359 1.12 2.4 III 50 36.2 5.05 0.306 2.164 2.82 11.2 IV 14 10.1 10.00 0.694 2.598 5.37 14.11 V 47 34.1 11.34 0.461 3.157 5.44 18.75 Total VI 3 2.2 18.92 1.794 3.107 15.62 21.79 VI 1 0.7 22.70 22.7 22.7 VIII 3 2.2 15.56 1.058 1.833 13.45 16.79 IX 1 0.7 22.18 22.18 22.18 Total 138 100.0 8.00 0.433 5.091 1.12 22.7

Breeding Ecology Male skin looked shiny and smooth in the reproduction period with stored len- fomatic secretion beneath the skin (Fig. 8). In addition, nuptial pads developed to grasp females for amplexus. Skin pattern got pale and pink color appeared on ventral and dorsal sides. Pregnant females had swollen abdomen and pink color appeared on the ventral side than the dorsal side (Fig. 9). When the ice covering Karagöl Lake’s surface started to melt, individuals appeared active but reproduction period did not start until the ice completely disap- peared. Reproduction period was on 26th May, 17th May, and 1st June in 2007, 2008,

Herpetol. Rom, 6, 2012 16 Yıldız, M.Z. & Göçmen, B.

Figure 8. a) a male specimen in reproduction period; b) the same specimen after reproduction period.

Figure 9. a) a female specimen in reproduction period; b) the same specimen after reproduction period. and 2009 respectively. As a result of this observation, reproduction period starts in the second week of May or first week of June. Spawning time lasted about 8-10 days but 70% egg clutches were laid in the middle 4-5 days. Three reproduction sites in Karagöl were determined. First one was on the northeastern part of the lake that was shallow (deep ranged 5-50 cm) and had dense vegetation. Data logger

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 17

was placed in this area and it recorded meteorological parameters during the reproduction period. Water temperature and air temperature ranged 3.56–28.47 °C and 3.15–31.51 °C in 2008 and 3.83–19.58 °C and -0.85–25.57 °C in 2009. Second one was on the southeastern part that was the shallowest (deepness ranges 5-20 cm) among the three reproduction sites. It extends nearly on a 20 m2 area and looked like a small lagoon. This site was preferred by most of the R. holtzi individuals as it was observed that most of them clutches on this site (Fig. 10). Third one was on the southwestern part of the Lake; Depth ranged from 50-80 cm. Poligonium amphibium was widespread around this site. Egg clutches were observed 20-200 cm from the lake shore and at least 60 cm in depth clinging to the P. amphibium specimen. Am- plexus was observed during day and night surveys in the Lake water but all egg clutches were laid during the night. Males were observed as overeager to perform amplexus as they were observed engaging it with juveniles, a piece of bread, a dead mouse, male to male, a female with two males, and as a group (2–3 ♀♀, 6– 7♂♂).

Figure 10. Some egg clutches was observed in the second reproduction site (arrow shows newly laid clutches).

305 egg clutches were counted (94 clutches in the first reproduction site, 148 clutches in the second one and 63 in the third one) in 2008 and 504 clutches (134

Herpetol. Rom, 6, 2012 18 Yıldız, M.Z. & Göçmen, B. clutches in the first reproduction site, 260 clutches in the second one and 110 in the third one) in 2009. Animal and vegetative poles of eggs could be distinguished by naked eye when clutches were recently laid. They looked highly dense until water diffused into egg jelly envelopes. The color of clutches was getting dull and jelly envelopes were getting yellow as the development progresses. Mean clutches size calculated was 464±277eggs (ranges: 46-1758 eggs). Egg size without jelly envelopes was 2.48±0.09 mm (ranges: 2.30-2.66 mm) and with jelly envelopes was 5.15±0.21 mm (ranges 3.75-6.84 mm). Fertilization success was 58.77% (ranges: 43-68%) and hatchling was 37.23% (28-53%). Reproductive investment calculated for females was 28.41±14.51% (ranges: 4.48- 61.52) and 22.4±7.90% (ranges 1.89-37.81) for males and positive correlation was found between SVL and reproductive investment (r=0.240 F=6.965, df=1, P=0.009) (Fig. 11).

Figure 11. Reproductive investment-SVL curve in Karagöl population.

Morphology and growth of tadpoles Rana holtzi’s individuals saw shallow areas (range: 5-40 cm) as reproductive sites. The mean water temperature of this area was measured 12.49±5.16°C (ranges 3.15- 31.57°C) and 10.83±3.78°C (ranges 3.83-19.58) in 2008 and 2009 respectively. Both in 2008 and 2009 years’ hatchling time ranged between 5-8 days depending on temperature. Hatching stage was found as 20th or 21st Gosner stage. Total length was measured 7.70±0.40 in 20th Gosner stage. The color of body was blackish brown and blood circulation was seen in external gill filaments in this stage under the ste- reomicroscope. Gill filaments consisted of two branches, each having 5-6 filaments on either side (Fig. 12). They hung on jell envelopes with adhesive organs (oral

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 19

suckers). All Gosner stages of tadpoles are shown in Fig. 12 and measured characters were given in Table 6. In the 21st Gosner stage gill filaments continued to grow, cornea appeared, and melanin pigmentation started to dissolve. On the right side, operculum started to develop and cover gill filaments in the 22nd Gosner stage. Tadpoles had adhesive organs, and cornea continued developing in the 23rd stage. Although right operculum was covered with gill filaments completely, on the left side operculum filaments could still be seen in the 24th stage. Oral sucker disap- peared, mouth developed, and feeding was observed in the 24th stage. In the next stage, eye, oral disc, and labial rows developed. Silver gray guanophores appeared and until the 25th stage opaque tail changed to transparent. SVL and TL were found 5.25± 1.92 and 7.68±2.49 mm in the 25th stage respectively (Table 8). Spiracu- lum developed only on the left side. Three different mouth types were observed of R. holtzi tadpoles (Fig. 13). Type A was the most common in tadpoles (55.95%). Type A had two supralabial and three sublabial rows [mouth type formula = 2(2)/3]. Type B (38.10%) had three supralabial and three sublabial rows [mouth type formula =.3(2, 3)/3]. Type C (5.95%) was distinguishable from Type B by first sublabial row to be interrupted [mouth type formula =.3(2, 3)3(1)]. Tadpole development of R. holtzi was compatible with the data provided by Gosner (1960). Therefore, all stage identification did not repeat itself here. Some observation was provided here that was not previously recorded. Ventral size of tadpoles had been transparent and all visceral organs could be seen in the 33rd stage. Guanophores became widespread on the dorsal side and tail in this stage. Lipophores (xantphores, yellow, green or orange color) were observed in the 39th stage. Metamorphosis was completed between 45-129 days in Karagöl Lake after hatchling. First metamorphosed froglets were observed in the second reproduction site which was the shallowest, and their SVL was measured 13.33±0.28 mm in July. In this reproduction site, larvae of Dytiscus marginalis, great diving beetle, and of Coenagrion puella, the azure damselfly, were found. It was known that these insect larvae feed on eggs and tadpoles. Besides these larvae, Cy- prinus carpio Linnaeus, 1758 may feed on R. holtzi tadpoles. Tadpoles completed metamorphosis in August had 14.52±2.38 mm SVL and reached 20.70±1.81 mm SVL in September; therefore, their size increased 50.25% at the end of September. Froglets that had metamorphosed in a short time period also had smaller SVL than froglets that had taken a longer time to metamorphose. On the other hand, they had a long feeding time to grow up until hibernation. Tadpoles’ density of first, second and third reproduction sites were calculated 2-4, 10-15, and 7-10 individuals respectively. Tadpoles were observed to feed on, undeveloped eggs, anomaly tadpoles that could not move, dead frogs, and pieces of chicken thrown into the lake in addition

Herpetol. Rom, 6, 2012 20 Yıldız, M.Z. & Göçmen, B.

Figure 12. All Gosner stage was observed in R. holtzi tadpoles.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 21

Figure 12. (continued)

Herpetol. Rom, 6, 2012

Table 8. Some metric measurement of R. holtzi’s tadpoles depending on Gosner stage.

20th Gosner stage 21st Gosner stage 22nd Gosner stage N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 12 7.70 0.40 7.02 8.08 21 4.12 0.12 3.73 4.45 20 4.50 0.18 4.25 4.68 TL 21 6.35 0.59 5.66 6.77 20 5.58 0.58 5.10 6.35 23rd Gosner stage 24th Gosner stage 25th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max IND 18 4.19 0.28 3.99 4.45 10 4.85 0.21 4.39 5.11 14 5.25 1.92 3.45 10.92 IOD 18 7.57 0.25 7.31 7.76 10 7.89 0.24 7.51 8.27 12 7.68 2.49 5.40 14.71 26th Gosner stage 27th Gosner stage 28th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 26 7.67 2.50 4.71 11.55 17 8.24 2.75 5.63 13.84 14 7.96 1.32 5.75 10.00 TL 25 11.15 2.87 6.13 16.20 14 10.97 2.58 7.32 16.61 14 11.88 1.99 9.57 15.63 TMW 8 0.61 0.03 0.55 0.65 9 0.85 0.25 0.60 1.25 8 0.93 0.21 0.64 1.25 TMH 10 1.11 0.10 1.00 1.25 10 1.30 0.29 1.00 1.88 9 1.49 0.27 1.13 2.00 IND 9 1.40 0.17 1.25 1.80 8 1.58 0.29 1.25 2.13 9 1.83 0.44 1.17 2.50 IOD 10 2.02 0.13 1.80 2.25 8 2.38 0.43 2.00 3.25 9 2.73 0.48 2.13 3.50

Table 8. (continued)

29th Gosner stage 30th Gosner stage 31st Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 7 9.20 2.08 5.72 11.53 3 10.25 0.86 9.75 11.24 14 11.50 1.07 8.94 13.51 TL 7 12.30 2.56 8.86 17.07 3 15.36 1.14 14.16 16.42 14 18.21 3.68 13.62 27.22 TMW 3 1.28 0.60 0.76 1.94 2 1.13 0.18 1.00 1.25 10 1.69 0.35 1.00 2.20 TMH 3 1.81 0.55 1.38 2.42 2 1.63 0.00 1.63 1.63 10 2.36 0.48 1.75 3.23 IND 3 1.88 0.28 1.60 2.15 2 2.06 0.09 2.00 2.13 10 2.32 0.24 1.88 2.69 IOD 3 3.10 1.05 2.38 4.31 2 2.69 0.44 2.38 3.00 10 3.85 0.51 2.88 4.55 32nd Gosner stage 33rd Gosner stage 34th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 4 11.36 0.91 10.63 12.68 6 12.35 1.19 10.59 14.19 10 12.75 1.22 11.25 14.73 TL 4 18.70 1.82 16.26 20.64 6 18.97 0.98 17.50 19.87 10 19.20 2.49 15.05 23.09 TMW 4 1.38 0.10 1.25 1.50 5 1.50 0.16 1.37 1.77 10 1.72 0.50 1.12 2.65 TMH 4 2.10 0.19 1.88 2.27 5 2.22 0.22 1.88 2.49 10 2.57 0.53 1.85 3.67 IND 4 2.32 0.12 2.25 2.50 5 2.16 0.19 1.86 2.38 10 2.37 0.13 2.14 2.54 IOD 4 3.87 0.02 3.84 3.88 5 3.92 0.19 3.73 4.13 10 4.26 0.25 3.80 4.75

Table 8. (continued)

35th Gosner stage 36th Gosner stage 37th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 4 14.36 1.72 12.67 16.20 14 15.67 1.87 11.75 18.65 3 17.26 2.93 14.28 20.14 TL 4 25.46 0.86 24.54 26.38 14 23.51 3.55 19.07 29.72 3 28.98 2.90 27.28 32.33 TMW 4 2.36 0.55 1.67 3.00 6 2.32 0.44 1.88 3.07 1 2.48 . 2.48 2.48 TMH 4 3.54 0.73 2.74 4.50 6 3.48 0.76 2.63 4.50 2 3.54 0.58 3.13 3.95 IND 4 2.55 0.18 2.34 2.75 6 2.61 0.34 2.25 3.11 2 2.89 0.19 2.75 3.02 IOD 4 4.61 0.59 3.74 5.00 6 5.02 0.41 4.38 5.47 2 4.91 0.76 4.38 5.45 FU 2 2.02 0.78 1.47 2.57 3 2.43 0.66 1.80 3.13 TbL 1 1.43 . 1.43 1.43 3 1.76 0.25 1.50 2.00 38th Gosner stage 39th Gosner stage 40th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 5 16.78 2.05 14.84 19.20 8 16.92 1.99 13.88 18.90 7 18.36 2.01 15.66 21.63 TL 5 30.19 4.51 23.27 35.36 8 30.03 7.04 16.48 35.73 7 31.83 5.23 23.37 37.51 TMW 5 3.06 0.87 2.26 4.11 6 3.48 0.51 2.63 3.98 5 3.62 0.32 3.11 3.88 TMH 5 4.32 1.23 3.21 6.16 6 5.07 0.90 3.50 6.16 5 4.66 0.43 3.96 5.00 IND 5 2.93 0.31 2.60 3.31 6 2.89 0.19 2.63 3.12 5 2.83 0.41 2.24 3.13 IOD 5 5.55 0.52 5.00 6.33 6 5.67 0.51 4.98 6.16 5 5.95 0.52 5.38 6.38 FU 1 2.82 . 2.82 2.82 3 2.98 0.11 2.85 3.05 5 4.32 1.11 3.02 5.63 TbL 1 2.21 . 2.21 2.21 3 2.78 0.78 2.15 3.65 5 4.66 1.01 3.75 6.25

Table 8. (continued)

41st Gosner stage 42nd Gosner stage 43rd Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 9 19.56 1.92 17.65 24.23 15 16.10 1.90 13.78 19.32 18 15.67 1.90 11.60 17.93 TL 9 32.94 3.75 29.44 38.92 15 27.85 5.72 17.82 40.11 18 19.23 8.12 6.54 33.30 TMW 5 3.98 0.52 3.13 4.50 11 3.28 0.62 2.19 4.45 5 3.04 0.48 2.57 3.75 TMH 5 5.42 0.63 4.38 5.94 11 4.13 0.94 3.46 5.89 4 3.38 0.38 3.05 3.88 IND 4 2.62 0.10 2.48 2.71 11 2.19 0.30 1.84 2.63 13 2.25 0.38 1.38 2.75 IOD 4 6.29 0.38 6.00 6.84 11 5.35 0.52 4.64 6.33 13 5.25 0.47 4.25 5.94 FU 5 6.45 1.00 5.02 7.66 11 6.58 0.87 5.62 8.10 12 7.17 1.07 4.75 8.37 TbL 5 6.65 1.02 5.28 7.87 11 6.67 1.21 5.33 8.80 12 7.01 1.40 4.15 8.87 44th Gosner stage 45th Gosner stage 46th Gosner stage

N Mean SD Min Max N Mean SD Min Max N Mean SD Min Max SVL 8 15.99 2.77 11.30 19.02 4 14.38 2.22 12.79 17.57 15 14.52 2.38 11.88 18.73 TL 6 7.02 3.21 4.10 12.54 IND 5 2.22 0.25 1.88 2.52 2 2.06 0.26 1.88 2.24 11 1.90 0.21 1.63 2.31 IOD 4 5.05 0.91 4.13 5.86 2 4.75 0.35 4.50 5.00 11 4.92 0.76 4.00 6.03 FU 5 7.02 1.28 5.00 8.32 2 6.40 1.98 5.00 7.80 11 6.16 1.42 4.63 8.34 TbL 5 7.29 2.30 3.63 9.47 2 6.57 2.21 5.00 8.13 11 6.09 1.71 4.38 8.26

26 Yıldız, M.Z. & Göçmen, B.

Figure 13. Mouth type of R. holtzi’s tadpoles. the mosses. Thus, cannibalism was observed first in R. holtzi tadpoles. Although cannibalism was commonly seen between the 25th and 30th Gosner stage, it was rarely observed until the 40th stage.

Activity periods Activity period began on 15th May and continued until 26th October for 5 months and 11 days in 2007. It began on 2nd May and continued until 10th October for 6 months and 8 days in 2008, and between 23rd May and 3rd November for 5 months and 11 days in 2009. As a result, depending on the meteorological factors activity period goes on for 5-6 months in Karagöl Lake.

Behavior When it was cold in spring and autumn, R. holtzi individuals would be activated after sun light hit the lake surface and shores. They were found to bask just on the lakeshore. If it is rainy or there is high humidity, they travel about 50 m in land. But they were observed around wet grass near the spring on the northern west side of the lake every time. They stayed in the lake water during the nights. Juve- niles got active later than adults in spring and hibernated earlier in autumn. Specimens were searched under the lake surface covered with ice and found that they hibernated under the stone in the lake water not out. When they were handed, all individuals had an urination reflex. In addition to playing dead and arching their back, another behavior was observed that when they were frightened they lower their body, turn out of their forelimb’ palm and put it behind or on their

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 27

closed eye (Fig. 14). Showing the red palm and looking strange with an enlarged head may be an attempt to frighten the enemy. This behavior known as Unken reflex (Hödl & Amezquita 2001). Although Unken reflex was commonly observed in females, it was also done by males. Unken reflex was recorded for R. holtzi for the first time.

Figure 14. Unken reflex shown by Taurus frog.

Threats and affecting factors to Taurus frog The major problem affecting the Taurus frog is a carp species, Cyprinus carpio in- troduced to the lake in 1990. Carp is an omnivore species therefore it may feed on eggs, tadpoles or adults of the Taurus frog. A fisherman expressed that he wit- nessed a carp attacked an adult Taurus frog on the lakeshore. The Ministry of En- vironment and Forestry encouraged fishery and supported them but it was observed that some fishermen try to feed carps with bread. On the other hand, when they casted nets, they accidentally damaged egg clutches in the breeding season. Some fisherman expressed that they release juvenile fishes they collect from other lakes nearby and try to reproduce them. It must be the reason why only one Capo- etta specimen was caught by the fishermen in Karagöl Lake. Karagöl Lake is a fa- mous area known by mountaineers and local people. Many visitors traveling to and camp around Karagöl Lake has caused environmental pollution. It is clear that pollution will affect amphibian population over time. In addition, some mountaineers crushed frogs when climbing at the night. Another problem as stated above, aquatic larvae of Dytiscus marginalis and Coenagrion puella may feed on eggs and tadpoles and thus affecting the population. The last but certainly not least prob-

Herpetol. Rom, 6, 2012 28 Yıldız, M.Z. & Göçmen, B. lems, if precautions are not taken Poligonium amphibium will dry the lake up and cause the extinction of Taurus frog in the long run.

DISCUSSION

Distribution and Morphology Rana holtzi was described from Karagöl Lake (2588 a.s.l.) in 1898 by Werner. Ar- palık frog population, 10 km away from Karagöl Lake at 200m a.s.l., included Rana macrocnemis Boulenger, 1885 by Baran (1969). Distribution of R. holtzi was known only in Karagöl Lake and Çinigöl Lake (Ulukışla, Niğde) until it was recorded in Eğrigöl Lake (Mersin) at 2777 a.s.l. by Baran et al. 2007. Eğrigöl Lake is a temporary lake. No Taurus frog individuals were observed in Eğrigöl Lake in 2008 excursion but they were found in four springs that are disconnected with Eğrigöl Lake. A sympatric amphibian species, Pseudoepidalea variabilis (Pallas, 1769), was observed in Karagöl Lake with R. holtzi. Except P. variabilis no other amphibian species were seen in Karagöl Lake and Çinigöl Lake. Werner (1898) reported that SVL of Taurus frog was 39-45 mm. Baran (1969) found that SVL ranged about 41-61 mm for males and 41-74.5 mm for females and SVL/FL+TL ratio, 0.91 (0.85–0.98) for males and 0.94 (0.86–1.00) for females; Head Length (HL)/Head Width (HW), 0.90 (0.81–0.95) for males and 0.88 (0.77–0.95) for females (n=135). Başoğlu and Özeti (1973) informed that males had 60 mm and females had 74 mm for R. holtzi. Çevik et al. (2006) reported that SVL 44.5±5.25 mm (n=48) for all individuals and they found no statistically significant difference between sexes. Baran et al. (2007) reported for Mersin R. holtzi population SVL 46.3±5.59 mm (n= 14, ranges 40.70–56.94) for males and 43.90±5.04 mm (n=11, ranges 40.24–56.38) for females; SVL/FL+TL ratio, 0.95±0.3 (ranges 0.90–1.00) for males, 0.96±0.03 (ranges 0.91–1.00) for females; HL/HW, 0.96±0.06 (range 0.87– 1.05) for males and 0.89±0.05 (0.81–0.96) for females. They found no statistically significant difference between sexes. Miaud et al. (2007) reported that males were larger than females in R. holtzi Karagöl population. In this study, mean SVL was calculated as 49.65±4.91 mm (n= 1309 ranges 37.69–68.95) for males and 45.74±6.03 mm (n= 1701, ranges 37.52–72.34) for females and 33.68±3.(n= 2061, ranges 49 17.18–41.98) for juveniles. Significant differences were found between sexes. SVL/FL+TL ratio was calculated as 0.90±0.05 (ranges 0.80–0.99) for males, 0.95±0.06 (ranges 0.86–1.12) for females; HL/HW ratio as 0.95±0.11 (ranges 0.73– 1.21) for males and 0.92±0.09 (ranges 0.75–1.09) for females. Although HL/HW ratio was found not to differ between sexes, females had significantly higher

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 29

SVL/FL+TL ratio than males. In each of previous studies, small specimen size was surveyed. That might be the reason why they did not find any difference between sexes. The SVL data to our result was not compatible with Başoğlu and Özeti (1973). Even though Baran et al. (2007) reported that there was no significant difference between sexes, mean SVL of males were higher than of those females. SVL/FL+TL ratio was compatible with the data given in previous studies. In most of the anurans females are bigger than males but it was known bigger males in especially some species which male have physical competition for mating (Shine 1979). Werner (1898) noted that Taurus frog had rather warty skin whereas Baran (1969) reported that investigated specimens had thin and smooth skin without warts. It was pointed out in many studies that R. holtzi had smooth or few warts on skin (Baran and Atatür, 1998; Budak and Göçmen, 2005; Çevik et al. 2006; Baran et al. 2007). Başoğlu and Özeti (1973) reported that most of the specimens did not have warts except for some females that had a few. According to our results, while males had bright and smooth skin during reproduction periods they had few warts or none at all after the reproduction period (Fig. 8). Females always had few warts (Fig. 9). Ground color was usually yellowish green or light brown and rarely pinkish or greenish gray. Dark green, brown, gray, or blackish spots were seen on the ground. Even in some studies it was reported that around of these dark spots there was a light strip, it was not observed on any of the specimens in this pattern (Baran 1969, Başoğlu and Özeti 1973, Baran and Atatür 1998, Çevik et al. 2006). Although Werner (1898) and Çevik et al. (2006) did not report any vertebral strip pattern on R. holtzi specimens, Baran observed vertebral strip on three individuals of the investigated 135 specimens. In this study, it was found that 4.4% of individuals (n=1000) had vertebral strip pattern.

Population structure and dynamics Daily activities of Taurus frog changed according to the seasons. Most of the individuals were observed in water and in shallow areas during the breeding season (the end of May and early June). Low temperature and wind inhibited activity of the frogs out of water. It was known that wind increases evaporation rate on skin thus cause dehydration and inhibit activity of amphibians (Porter 1972). Most of the individuals were adults in this season. After the breeding season, water was at its maximum level and when the sun light hit the lake surface and its shores, R. holtzi individuals would be activated and generally they were found just on the lakeshores. Whereas Baran (1969) reported that they constituted large herds on the grass, they were observed only on wet grass formed by the spring water in the

Herpetol. Rom, 6, 2012 30 Yıldız, M.Z. & Göçmen, B. north-western side of the lake water during three years. Otherwise, Taurus frog was observed 50 m away from the lake only on the rainy days. Ranid frog was known as real water frogs that depend on a water system and do not go away from water (Porter 1972, Başoğlu and Özeti 1973, Budak and Göçmen 2005). Addition- ally, it was reported that mountain frogs always depended on water, too (Tarkhnishvili and Gokhelashvili 1999). With the decrease of water level in summer, shelters appeared around the lakeshore and frogs spend their times in these cavities during daytime while they were found in water during nights. Baran et al. (2001) reported that R. holtzi hibernated under the meadow pad and repeated this expression in their subsequent studies (Baran et al. 2007, Miaud et al. 2007) but it was determined that they hibernated under the stone in the lake water not on land. It was previously reported that mountain frogs distributed in high mountains hibernate in lake water (Terentjev and Chernov 1965, Başoğlu and Özeti 1973). Çiçek et al (2011) declared that R. macrocnemis specimens were found just on the lakeshore during the whole year and night activity was more than daytime activity in Lake District’ Uludağ. They observed that while 10-20 individuals were active in Kilimli Lake (Uludağ) in daytime, 40-50 individuals were active in night time. In other lakes, in Lake District on Uludağ it was found that 3-5 individuals were active in daytime whereas 30-60 individuals in night time. Individuals of Taurus frog were found more (7-10/individuals m2) on grass and high vegetation areas than deeper ones and areas surrounded by high rocks (1-2/individuals m2). Baran (1969) did not provide any number on the population size of R. holtzi but he stated that Taurus frog constitutes highly dense flocks. Baran et al. (2001) counted the number of individuals directly, compared with the lake area (60 de- cars), and then calculated the population size as 30000 and expressed that population decreased to 60-70% ratio. However, population size was not known; therefore, reduction ratio given only depends on personal opinion. Çevik et al. (2005) performed mark-recapture in three days and calculated population size as ranging from 725 to 1432 individuals. However, according to their data and population formula used, population size ranged from 725 to 1597. Probably, they made a calculation error. At the end of our study that was performed between 2007 and 2009, population size calculated 17563 individuals. It was determined that population size increased compared to the population size was given Çevik et al. (2005). Mean population size in Çinigöl Lake was calculated 157 individuals. Çicek et al. (2011) calculated survival rate of R. macrocnemis that was 0.74-0.83 for Kirazlıyayla; 0.79–0.88 for Sarıalan; 0.78–0.98 for hotel district; Capture probability: 0.21–0.33 for Kirazliyayla; 0.25–0.35 for Sarialan; ranged 0.22–0.29 and 0.21– 0.93 for hotel district. They noticed that as the altitude increased the survival rate

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 31

and capture probability increased. However, in some studies it was reported that depending on the increase in the altitude, survival rate and capture probability decreased (Tarkhnishvili and Gokhelashvili 1999). Başkale (2009) reported that survival rate for P. bedriagae Eagean population was 0.52 for females; 0.83 for male in Karagöl Lake (İzmir); 0.51 for males and females in Sülüklü Lake. Beebee and Grif- fiths (2000) found out that survival rate ranged between 0.25–0.50 for R. temporaria individuals. Vasconcellos et al. (2009) reported that survival rate for Bufo schneideri females was 0.35 and for males was 0.37 and for B. rubescens was 0.38 and 0.34 for females and males in Brazil, respectively. Peter (2001) reported that survival ratio of Rana lessonae (0.72–0.84) was higher than R. esculenta’s survival ratio (0.53–0.70). Çevik et al (2005) calculated survival rate as 0.12 and population gain 0.92 for Tau- rus frog Karagöl Lake population. According to this study, survival rate in Karagöl Lake R. holtzi Karagöl population was 0.74 in 2007; 0.87 in 2008; 0.85 in 2009 and 0.83 in three years and capture ratio were calculated at 0.16, 0.09, 0.11 and 0.16 in 2007, 2008, 2009 and for three years, respectively. Calculated survival rate was quite high when compared with Çevik et al. (2005). Mean population decline ratio was determined 3.1% for Rana arvalis, R. sylvatica, R. temporaria, Bufo bufo, B. clamita and Hyla arborea. The highest decline rate was calculated for R. sylvatica and the lowest for H. arborea (Gren 2003). It was determined that in Karagöl Lake Taurus frog population declined –0.084 ratio between 2007–2008 whereas it increased 0.316 and 0.232 between 2008-2009 and 2007-2009 years, respectively. As a result, population showed tendency to grow. Population decline ratio was determined for R. macrocnemis Uludağ populations ranged from –0.004 to –0.081 in 2007-2008 years (Çicek et al. 2011). Pelophylax bedriagae population declined from 403 to 76 between 2007 and 2008 in Karagöl (Izmir), population declined from 2573 individuals to 827 individuals in Sülüklü Lake in the same years (Başkale 2009). Başkale (2009) also observed population decline for P. bedriagae in İkizgol Lake and Göztepesi Lake. Rainfall ratio was at its lowest level (504.3 mm) for the last 38 years in 2008, therefore it is clear that all amphibian population living in Anatolia was negatively affected (meteorology, 2010).

Population Age Miaud et al. (2007) determined that median age for juveniles was 3.5 (range: 1-5); median age for males and females was similar at 6 (range: 4–8) and longevity reached 8 for males and 10 for females within the R. holtzi population. Guarino and Erişmiş (2008) calculated that mean age was 5.04±0.68 (4–6) for males, 5.35±1.00 (4– 7) for females and ranged from 1 to 9 years old at Karagöl Lake R. holtzi population. Results of this study showed that mean age of males was 4.87±0.97 (range: 3–

Herpetol. Rom, 6, 2012 32 Yıldız, M.Z. & Göçmen, B.

8), of females were 4.29±1.38 (range: 3–9) and longevity reached 9 years in a female. The results were in accordance with the data was given in previous studies Miaud et al. 2007, Guarino and Erişmiş 2008) Esteban et al. (1999) showed that mean age of males was 3.63±1.43, of females was 2.91±1.37 and ranged from 1 to 6 years in R. sharica Morocco population. Kumbar and Pancharatna (2001) determined that age ranged from 1 to 5 and longevity reached 5 in Microhyla ornate population. In R. camerani’s Tabatskuri Lake (2000 m a.s.l.) the mean age of males was found 2.63, of females 2.84 and ranged 2- 4 ages in all individuals (Gokhelashvili and Tarkhnishvili, 1994). Erişmiş (2005) found that ages ranged from 2 to 9 in Beyşehir Lake for P. bedriagae population. Lai et al. (2005) investigated the effect of altitude on population age and longevity in R. swinhoana population in Taiwan. They found that mean age of males at low elevation was 3.73±1.10, of females was 4.71±1.01, while at high elevation mean age of males was 5.12±1.28 and of females was 6.55±1.82. As a result, individuals living on high elevation had higher age and longevity ratios than living on low elevation populations. Çicek et al. (2011) calculated age composition in R. macrocnemis’s Uludağ populations and found that mean age of males 4.65±0.208, of females was 4.72±0.202 and detected increase at mean age and longevity depending on the elevation. In high elevation, lacking predator species, growing slower, low elevation, and high maturity age caused to increase mean age and longevity on high elevations (Morrison, 2001; Morrison et al., 2004; Lai et al., 2005. Age of maturity in amphibians depends on the living area, elevation, temperature, and activity period length (Berven 1982, Duelman and Trueb 1986, Stebbins and Cohen 1995, Morrison 2001, Lai et al. 2005). The highest age of maturity was determined at high elevations (Duelman and Trueb 1986). It was reported that age of maturation for R. pretiosa was 1 or 2 years, that it breeds every year at low elevations; although, its age of maturity was 5-6 years and it breeds once in every 2-3 years at high elevations (Licht 1975). The higher age of maturation depending on higher elevation was also reported for R. swinhoana (Lai et al. 2005) R. macrocnemis (Gokhelashvili and Tarkhnishvili 1994, Çiçek et al. 2011), B. bufo (Hemelaar and Van Gelder 1980), B. calamita and R. pipens (Duelman and Trueb 1986). Age of maturity of R. macrocnemis at 2100 m population was reported 4-6 years (Çicek et al. 2011), of R. swinhoana population in Lishing–Taiwan at 1600 m was 4 years (Lai, 2005), of R. camerani in Tabatskari at 1900 m was 2-3 years (Gokhelashvili and Tarkhnishvili 1994). Age of maturity in Karagöl Lake population was reported 4-5 years by Miaud et al (2007) and 4 years by Guarino and Erişmiş (2008). In Karagöl Lake population, age of maturity was determined to range 3-5 years and females reached maturity earlier than males.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 33

Miaud et al. (2007) found out that with 83.9% in males and 71.7% in females fit the age-body length relationship of sampled individuals to the von Bertalanfyy growth model and that there was no difference between sexes regarding the growing pattern in Taurus frog. In another study, it was reported that correlation between SVL and age was significant in females where in males it was not significant in R. holtzi (Guarino and Erişmiş 2008). According to the obtained data, positive correlation was determined between age and SVL and age and weight in R. holtzi population but range of SVL and weight depending on age overlapped. In amphibians generally there were positive correlations between age and SVL (Esteban et al. 1999, Erişmiş 2005, Matthews and Miaud 2007) but it was also reported that there was a weak positive correlation between age and SVL (Ischenko 1989, 1996, Gokhelashvili and Tarkhnishvili 1994). However, this does not mean that the long- est individuals had the oldest individuals in population. The growing rate depended on temperature (Berven 1982), habitat (Duelman and Trueb 1974), activity period length (Hemelaar and Gelder 1980), food supply (Berven and Chadra 1988), and metamorphosis length (Ryser 1996).

Breeding ecology Breeding period begins with rainfall in subtropical area (Duelman and Trueb 1974) while amphibians living in tropical area that have high rainfall can reproduce year round (Crump 1974). Although breeding period depends on precipitation, the main factor is temperature in amphibians living in temperate climates (Duelman and Trueb 1974). Temperature is the major factor that limited the reproduction period. Reproduction period of R. sylvatica lasts 5-6 months in low areas whereas in highlands it lasts only 1-2 weeks (Herreid and Kinney 1967). Reproduction period length also affects the number of clutches that lay in a year. A female of Hyla rosenbergi was reported to lay 6 clutches in a year (Kluge 1981). Most of the species reproduce once a year in temperate climates because of the short reproduction period (Berven 1982). However, the females of R. catesbeiana in North America and Bufo valliceps in Mexico were reported to laid twice a year (Blair 1960). Amphibians living in drought and low temperature areas can reproduce once in two years or in more lengthy periods. Plethodon glutinosus and P. wehlei were reported to reproduce once in two years and Notophthalmus viridescens did not reproduce for 5 years (Bull and Shine 1979, Gill 1985). Breeding period of R. macrocnemis began by the snow melting and reproduction period length depending on elevation (Efendiev and Ishchenko 1974, Tarkhnishvili and Gokhelashvili 1999). It was reported that Taurus frog reproduced in May (Baran et al. 2001).

Herpetol. Rom, 6, 2012 34 Yıldız, M.Z. & Göçmen, B.

It was determined that reproduction period began between the second week of May and early June and lasted 8-10 days but most of the clutches were laid in 4-5 days in Karagöl Lake. It was observed that Taurus frog reproduced once a year due to the short activity and breeding period. Clutch size depends on the environmental factors and breeding count a year at ectothermic animals (Crump, 1974). There is a positive correlation between body size and clutch size in anurans (Salthe and Duelman 1973; Kaplan and Salthe 1979). Clutch size for Taurus frog ranged from 46 to 1758 and mean was 464±277 eggs, and egg size was measured 2.48±0.02 mm. Clutch size in R. arvalis was reported as 724–1029 in Moscow population (Cherdantsev et al. 1997), of R. dalmatina was 950 in Bonn population, of R. temporaria was 1766 (Hatchel et al. 2005), of R. macrocnemis was 987 in Uludağ population (Çiçek et al. 2011). Egg size of R. dalmatina was reported as 2.14 mm in Bonn population, of R. temporaria was 2.08 mm (Hachtel et al. 2005), of R. macrocnemis was 2.39 mm in Uludağ population (Çiçek et al. 2011). Egg size of Taurus frog was close to the size of R. macrocnemis. Phyllomedusa azurea females were reported to lose 29.1% of their ovarian weight and of P. sauvagi lost 21.1%. Lai et al (2005) observed that female of R. sauteri lost 22.5 % of their weight in Chihchung (2360 m a.s.l.) and Mienyueh (2350 m a.s.l.), 25.7 % in Chitou (1100 m a.s.l.) and they noticed that reproductive investment decreased depending on the increase in elevation. Çicek et al. (2011) declared that females of R. macrocnemis lost 12.4–42.22% their weight. It was calculated that females of R. holtzi lost 28.41% of their weight after breeding. Temperature is the major factor affecting the development embryonic stage and growing stage of tadpoles in species; although, it also depends on some environmental and genetic factors (Moore 1939). Development of eggs and tadpoles increases with the temperature up to the tolerance limits. It was reported that with the increase in temperature, development increased in R. pipiens, R. sylvatica, R. palustris (Moore 1939), Ambystoma maculatum (Voss 1993), R. macrocnemis (Çiçek et al. 2011). It was observed that temperature of the breeding site ranged from 3°C to 28°C, hatchling occurred in 5-8 days, and tadpoles completed metamorphosis between 45 and 129 days depending on the temperature. Though Baran et al. (2001) reported that tadpoles of R. holtzi did not complete metamorphosis until Septem- ber; it was observed that they did not complete all developmental stages until the middle of October. The major factors that affect tadpoles’ development are food supply, density, and temperature (Duellman and Trueb 1974). These factors affect metamorphosis length altogether (Berven and Chadra, 1988). Generally in highlands, tadpoles’ density and lack of food supply decrease the development rate; therefore, Tadpoles stage lasts longer (Smith-Gill and Berven 1979, Berven and

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 35

Chadra 1988). That is why tadpoles in highlands are larger than the ones in low elevation. However, activity period is short in highlands, tadpoles have some ad- aptation on living there (Altig and McDiarmid 1999). Nagai et al. (1971) investigated the effect of nutrient contents for tadpoles’ development rate and they found that the herbivore and cannibal tadpoles reached largest metamorphosis froglet. It was observed in laboratory and in Karagöl Lake that tadpoles of R. holtzi were herbivore and cannibals in early Gosner stage (25-30 Gosner stage). It was reported that SVL of R. pseudodalmatina’s tadpoles were 7.1±0.6, and of R. macrocnemis was 4.74 mm (Çicek et al. 2011). In this study, SVL of R. holtzi tadpoles were measured 5.25±1.92 mm. It was known that when Tadpoles completed their metamorphosis in a short time, they would have short SVL whereas Tadpoles completed their metamorphosis in a long time, they would have long SVL (Altig and McDiarmid 1999). It was observed that tadpoles of Taurus frog completed their metamorphosis in 45 days, they were measured as 13.33±0.28 mm in the second reproduction site while they reached 20.04±1.44 mm in September metamorphic froglets The second reproduction site was shallow and had high vegetation; therefore, probably temperature was high in this site than in the other two reproduction sites. In addition, larvae of Dytiscus marginalis were observed in this site and it was drought until August. It was known that drought and presence of predators increase the development rate of tadpoles (Altig and McDiarmid 1999).

ACKNOWLEDGEMENT. This study is a part of a larger research study, Ph.D. thesis of first author. We would like to express our gratitude to Bahadır AKMAN, Deniz YALÇINKAYA (Biology Department of Ege University, Izmir) and Naşit IGCI (Biotechnology institute of Ankara University) for their help in the field and Dr. Kerim ÇİÇEK (Biology Department of Ege University, Izmir) for his help in statistical analysis, and Yehudah L. WERNER (Professor Emeritus, Department of Evolution, Systematics and Ecology, The Hebrew University of Jerusalem) for invaluable suggestions and improving the English of the manuscript. We are grateful to TUBITAK (Project No: 107T167), HÜBAK (Project No: Hübak 1111), and EBILTEM (Project No: 2007BIL004) for supporting this project.

REFERENCES

Alford, R.A., Richards, S.J. (1999): Global Amphibian Declines: a problem in applied ecology, Annual Review of Ecology and Systematics 30: 133–165. Altig, R., McDiarmid, W.R. (1999): Tadpoles: the biology of anuran larvae, University Of Chicago Press, Chicago.

Herpetol. Rom, 6, 2012 36 Yıldız, M.Z. & Göçmen, B.

AMPHIBIAWEB, (2012): “Information on amphibian biology and conservation. [web application]. 2009. Berkeley, California: AmphibiaWeb” http://amphibiaweb.org/. (Erişim tarihi: 24 Mart 2012). Baran, I. (1969): Anadolu dağ kurbağaları üzerinde sistematik araştırma, Ege Üniversitesi Fen Fakültesi İlmi Raporlar serisi No. 80, Bornova–İzmir. Baran, İ., Atatür, M.K. (1998): Turkish Herpetofauna (Amphibians and Reptiles), Ministry of Environment, Ankara. Baran, I., Balık, S., Kumlutaş, Y., Tok, C.V., Olgun, K., Durmuş, H., Türkozan, O., Ilgaz, Ç., Üret, F. (2001): Rana holtzi (Toros Kurbağası)’nin Biyolojik ve Ekolojik Yönden Araştırılması ve Koruma Stratejisinin Saptanması. IV. Ulusal Ekoloji ve Çevre Kongresi, Bodrum, 213–218. Baran, İ., Ilgaz, C., Kumlutaş, Y., Olgun, K., Avcı, A., İret, F. (2007): On new populations of Rana holtzi and Rana macrocnemis (Ranidae: Anura). Turkish Journal of Zoology 31: 241– 247. Başkale, E. (2009): Ege bölgesindeki bazı göllerde yaşayan amfibi türlerine ait populasyonların gözlenmesi, populasyon büyüklüklerinin hesaplanması ve habitat özelliklerinin belirlenmesi, Doktora tezi, Ege üniversitesi Fen Bilimleri Enstitüsü. Başoğlu, M., Özeti, N. (1973): Türkiye Amfibileri (The Amphibians of Turkey; Taxonomic list, Distribution, Key for Identification pp. 127–138), Ege Üniv. Fen Fak. Kitaplar Serisi No. 50, Bornova–Izmir. Beebee, T.J.C., Griffiths. R. A. (2005): The amphibian decline crisis: a watershed for conservation biology? Biological Conservation 125: 271–285. Berven, K.A. (1982): The genetic basis of altitudinal variation in the wood frog Rana sylvatica I. An experimental analysis of larval development. Oecologia 52: 360–369. Berven, K.A., Chadra, B.G. (1988): The relationship among egg size, density and food level on larval development in the wood frog (Rana sylvatica) Oecologia 75: 67–72. Blair, W. (1960): A breeding population of the Mexican toad (Bufo valliceps) in relation to its environment. Ecology 41: 165–174. Brodman, R., Parrish, M., Kraus, H. Cortwright, S. (2006): Amphibian biodiversity recovery in a large–scale ecosystem restoration. Herpetological Conservation and Biology 1: 101– 108. Budak, A., Göçmen, B. (2005): Herpetology, Ege üniversitesi Fen Fakültesi Yayınları, No: 194, Bornova, Izmir. Bull, J.B., Shine, R. (1979): Iteroparous animals that skip opportunities for reproduction, American Naturalist 114: 296–303. Burnham, K.P. Anderson, D.R. (2002): Model Selection and Inference: A Practical Information–Theoretic Approach. Springer Verlag, Berlin. Çevik , I.E., Arıkan, H., Kaya, U., Atatür, M.K. (2006): Comparative morphological and serological studies of three Anatolian Mountain frogs, Rana macrocnemis, R. camerani and R. holtzi (Anura, Ranidae). Amphibia–Reptilia 27: 63–71. Cherdantsev, V.G., Lyapkov, S.M., Cherdantseva, E.M. (1997): Mechanisms accounting for the pattern of fecundity formation in the frog, Rana arvalis Nilss. Russian Journal of Zoology 1: 30–40.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 37

Çicek, K., Mermer, A., Tok, C.V. (2011): Population Dynamics of Rana macrocnemis Boulenger, 1885 at Uludağ, Western Turkey. Zoology in the Middle East 53: 41–60. Cooch, E., White, G.C. (2009): Program Mark, A Gentle introduction, 8. Ed. http://www.phidot.org/software/mark/docs/book/, (Erişim tarihi: 17 Şubat 2010). Cormack, R.M. (1964): Estimates of survival from the sighting of marked animals. Biometrika 51: 429–438. Crump, M.L. (1974): Reproductive strategies in a tropical anuran community.University of Kansas Museum of Natural History Miscellaneous Publications 61: 1–68. Crump, M.L. (1984): Intraclutch egg size variability in Hyla crucifer (Anura: Hylidae). Copeia 2: 302–308. Donnelly, M.A., Guyer, C., Juterbock, J.E., Alford, R. (1994): Techniques for marking amphibians, 277–284, Measuring and monitoring biological diversity, standard methods for amphibians, Heyer, W.R., Donnelly, M.A., McDiarmid, R.W., Hayek, L.C. and Foster, M.S. (Eds), Smithsonian Institution Press, Washington, DC. Duellman, W.E., Trueb, L. (1986): Biology of amphibians. The Johns Hopkins University Press, London. Efendiev, S.M., Ischenko, V.G. (1974): Patterns of reproduction in the North Caucasus mountain populations of Rana macrocnemis Blgr. Ekologiya (Sverdlovsk) 6: 80–83. Erişmiş, U.C. (2005): Göller bölgesi Rana ridibunda (Anura: Ranidae) Populasyonlarında Yaş– Boy, Yaş–Ağırlık ve Boy–Ağırlık ilişkilerinin araştırılması (Doktora Tezi), Ege Üniversitesi Fen Bilimleri Enstitüsü. Esteban, M., García–París, M., Buckley, D., Castanet, J. (1999): Bone growth and age in Rana sharia, a water frog living in a desert environment, Annales Zoologici Fennici 36: 53–62. Francillon–Vieillot, H., Arntzen, J.W., Géraudie, J. (1990): Age, growth and longevity of sympatric Triturus cristatus, T. marmoratus and their hybrids (Amphibia, Urodela): a skeletochronological comparison Journal of Herpetology 24: 13–22. Gibbons, L.L., Lovich, J. E. (1990): Sexual dimorphism in turtles with emphasis on the slider turtle (Trachemys scripta). Herpetological Monographs 4: 1–29. Gill, D. (1985): Interpreting breeding patterns from census data: a solution to the Husting dilemma. Ecology 66: 344–354. Göçmen, B. Yıldız, M. Z., Yalçınkaya, D., Akman, B. (2008): Bolkar Dağlarında (Ulukışla, Niğde) Yaşayan Toros Kurbağası Rana holtzi Werner, 1898 Populasyon büyüklüğünün son durumu hakkında. Ulusal Ekoloji ve Çevre Kongresi 20-23 Ekim, Girne-Kıbrıs. Gokhelashvili, R.K., Tarkhnishvili, D.N. (1994): Age structure of six Georgian anuran populations and its dynamics during two consecutive years (Anura). Herpetozoa 7: 11-18. Gosner, K.L. (1960): A simplified table for staging anuran embryos and larvae with notes on identification. Herpetologica 16: 183–190. Green, D.M. (2003): The ecology of extinction: population fluctuation and decline in amphibians. Biological Conservation 111: 331–343. Guarino, F.M., Erişmiş, U.C. (2008): Age determination and growth by skeletochronology of Rana holtzi, an endemic frog from Turkey. Italian Journal of Zoology 75: 237–242. Gürses, M.K., Gemici, Y., Özkurt, N., Gülbaba, A.G., Özkurt, A., Tüfekçi, S. (1996): Bolkar dağları Karaçam (Pinus nigra Arn. var. pallasiana Schneid.) populasyonlarında biyolojik

Herpetol. Rom, 6, 2012 38 Yıldız, M.Z. & Göçmen, B.

çeşitlililk üzerine araştırmalar, Doğu Akdeniz Ormancılık Araştırma Enstitüsü, DOA dergisi 2: 1–18. Hachtel, M., Ortmann, D., Kupfer, A., Sander, U., Schmidt, P., Weddeling, K. (2005): Return rates and long–term capture history of amphibians in an agricultural landscape near Bonn (Germany), Herpetologia Petropolitana: Proceedings of the 12th Ordinary General Meeting of the Societas Europaea Herpetologica, Ananjeva, N. & Tsınenko, O. (Eds), Russian Journal of Herpetology 12: 146–149. Hemelaar, A.S.M., van Gelder, J.J. (1980): Annual growth rings in phalanges of Bufo bufo (Anura, Amphibia) from the Netherlands and their use for age determination. Netherlands Journal of Zoology 30: 129–135. Herreid, C. F. II., Kinney, S. (1967): Temperature and development of the wood frog Rana sylvatica, in Alaska. Ecology 48: 579–590. Houlahan, J. E., Findlay, C. S., Schmidt, B. R., Meyer, A. H. Kuzmin, S.L. (2000): Quantitative evidence for global amphibian population declines. Nature 404: 752–755. Hödl, W. and Amezquita, A. (2001): Visual signaling in anuran amphibians. pp.121-141. In: Ryan, M.J. (ed), Anuran communication. Smithsonian lust. Press, Washington. Ishchenko, V.G. (1989): Population biology of amphibians, Soviet Scientific Review series F, Physiology General Biology 3: 119–155. Ishchenko, V.G. (1996): Problems of the demography and declining populations of some problems Eurasiatic Brown Frogs. Russian Journal of Zoology 3: 143–151. Jolly, G.M. (1965): Explicit estimates from capture–recapture data with both death and immigration stochastic model. Biometrika 52: 225–247. Kaplan, R. H., Salthe, S. N. (1979): The allometry of reproduction: an empirical view in salamanders. American Naturalist 113: 671–689. Kaya, U., Çevik, İ.E., Erişmiş, U.C. (2005): Population status of the Taurus Frog, Rana holtzi Werner, 1898, in its terra typica: Is there a decline? Turkish Journal of Zoology 29: 317–319. Kaya, U., Baskale, E., Cevik, I.E, Kumlutas, Y., Olgun, K (2010): Population sizes of Taurus frog, Rana holtzi, in two different localities, Karagol and Egrigol: new estimations, decline, and a warning for their conservations. Russian Journal of Herpetology 17:247-250. Kluge, A. G. (1981): The life history, social organization, and parental behavior of Hyla rosenbergi Boulenger, a nest–building gladiator frog. Miscellaneous Publications of the Museum of Zoology, University of Michigan 160: 1–170. Kumbar, S.M., Pancharatna K. (2001): Determination of age, longevity and age at reproduction of the frog Microhyla ornata by skeletochronology. Journal of Biosciences 26: 265–70. Lai, Y.C., Lee, T.H., Kam, Y.C. (2005): A skeletochronological study on a subtropical, riparian ranid (Rana swinhoana) from different elevations in Taiwan. Zoological Science 22: 653–658. Lebreton, J. D., Burnham K. P., Clobert, J., Anderson, D.R. (1992): Modeling survival and testing biological hypotheses using marked animals: a unified approach with case studies. Ecological Monographs 62: 67–118.

Herpetol. Rom, 6, 2012 Population dynamics, reproduction, and life history traits of Rana holtzi 39

Leclair, M.H., Leclair, R.Jr., Gallant, J. (2005): Application of skeletochronology to a population of Pelobates cultripes (Anura: Pelobatidae) from Portugal. Journal of Herpetology 39: 199–207. Licht, L.E. (1975): cooperative life history of the western spotted frog, Rana pretiosa, from low and high elevation populations. Canadian journal of Zoology 53: 1258–1269. Matthews K.R., Miaud, C. (2007): A skeletochronological study of the longevity and age structure of the mountain yellow–legged frog, Rana muscosa, in the Sierra Nevada, California. Copeia 986–993. Metoroloji, (2010): Yıllık Toplam Yağış Verileri, (http://www.dmi.gov.tr/), (Erişim tarihi: 24 Haziran 2010). Meyer, A.H., Schmidt, B.R., Grossenbacher, K. (1998): Analysis of three amphibian populations with quarter century long time–series, Proceedings of the Royal Society, London 265: 523–528. Miaud, C., Üzüm, N., Avcı, A., Olgun, K. (2007): Age, Size and growth of the endemic Anatolian mountain frog Rana holtzi from Turkey. Herpetological journal 17: 167–173. Moore, J.A. (1939): Temperature tolerance and rates of development in the eggs of amphibian. Ecology 20: 459–478. Morrison, C. Hero, J.M. (2004): Geographic variation in life–history characteristics of amphibians: a review. Journal of Animal Ecology 72: 270–279. Morrison, F. C. (2001): Altitudinal Variation in the Life History of Anurans in Southeast Queensland. Ph.D. Dissertation. Griffith University, Gold Coast, Australia. Nagai, Y., Nagai, S., Nishikawa, T. (1971): The nutritional efficiency of cannibalism and an artificial feed for the growth of tadpoles of Japanese toad (Bufo vulgaris sp). Agricultural and Biological Chemistry 35: 697–703. Otis, D.L., Burnham, K.P., White, G.C., Anderson, D.R. (1978): Statistical inference from capture data on closed animal populations. Wildlife Monographs 62. Pechmann, J.H.K., Wilbur, H.M. (1994): Putting declining amphibian populations in perspective: Natural fluctuations and human impacts. Herpetologica 50: 65–84. Peter, A–K. H. (2001): Survival in adults of water frog Rana lessonae and its hybridogenetic associate R. esculenta. Canadian Journal of Zoology 79: 652–661. Pollock, K.H. (1974): A tag–recapture model allowing for unequal survival, and catchability. Biometrics 30: 77–87. Ryser, J. (1996): Comparative life histories of a low– and a high–elevation population of the common frog Rana temporaria. Amphibia–Reptilia 17: 183–195. Salthe, S. N., Duellman, W. E. (1973): Quantitative constraints associated with reproductive mode in Anurans In: Evolutionary biology of the anurans: contemporary research on major problems. (Ed J. L.Vial), University of Missouri Press, Columbia. Schmidt, B.R. (2003): Count data, detection probabilities, and the demography, dynamics, distribution, and decline of amphibians. Comptes Rendus Biologies 326:119–124. Schmidt, B.R., Schaub, M., Anholt, B.R. (2002): Why you should use capture–recapture methods when estimating survival and breeding probabilities: on bias, temporary emigration, over dispersion, and common toads. Amphibia–Reptilia 23:375–388.

Herpetol. Rom, 6, 2012 40 Yıldız, M.Z. & Göçmen, B.

Schwarz, C.J., Arnason, N. (1996): A general methodology for the analysis of Capture-recapture experiments in open populations. Biometrics 52: 860–873. Seber, G.A.F. (1965): A note on the multiple recapture census. Biometrika 52: 249–259. Shine, R. (1979): Sexual selection and sexual dimorphism in the amphibia. Copeia 297–306. Smith–Gill, S.J, Berven, K.A. (1979): Predicting amphibian metamorphosis. American Naturalist 113: 563–585. Stebbins, R.C., Cohen, N.W. (1995): A natural history of amphibians, Princeton Univ. Press, Princeton. Surova, G.S., Cherdantsev, V.G. (1987): Embryonic morphs in the populations of brown frogs egg size and rates of larval growth in Moscow district USSR Rana temporaria and Rana arvalis. Zoologichesky Zhurnal 66: 1864–1872. Tarkhnishvili, D.N., Gokhelashvili, R.K. (1999): The amphibians of the Caucasus (Advances in Amphibian Research in the Former Soviet Union). Pensoft Publications, Sofia. Terentjev, P.V., Chernov, S.A. (1965): Key to amphibians and reptiles of the USSR, Israel Program for Scientific Translations, Jerusalem. Veith, M., Schmidtler, J.F., Kosuch, J., Baran, I., Seitz, A. (2003): Palaeoclimatic changes explain Anatolian mountain frog evolution: a test for alternating vicariance and dispersal events. Molecular Ecology 12: 185–199. Voss, S. R. (1993): Effect of temperature on body size, developmental stage, and timing of hatching in Ambystoma maculatum. Journal of Herpetology 27: 329–333. Werner, F. (1898): Über einige neu Reptilien und einen neuen Frosch aus dem Cilicischen Tarus. Zoologischer Anzeiger 21: 217-223. Williams, B.K., Nichols, J.D., Conroy, M.J. (2002): Analysis and Management of Animal Populations. Academic Press, San Diego.

Herpetol. Rom, 6, 2012