First detection probability, comparative analysis of ecological disparities and population dynamics of Rana pseudodalmatina (Eiselt &amp; Schmidtler, 1971) in the Hyrcanian forests: implications for conservation planning

doi:10.21203/rs.3.rs-3991978/v1

The Hyrcanian forests have a diverse fauna and flora with many endemic species of plants and animals. Rana pseudodalmatina is one of the endemic species in this forest. The phylogenetic and morphologic analysis indicates the existence of the western and the eastern clades in R. pseudodalmatina. The life history of an organism comprises its growth, development, age, and reproduction. Differences in the life history of the same species can attributed to different habitat quality or climate. The probability of encountering a wood frog in the western half of the Hyrcanian forests of the southern shores of the Caspian Sea is much higher than in the forests of the eastern sea. This increase in the detection probability can attributed to higher precipitation levels in the west of the Caspian Sea compared to the east. We observed that the Eastern populations tend to have larger sizes. In addition, the individuals of the western branch have a lower mean age (5.29 ± 0.74 years) than the eastern ones (8.36 ± 0.08 years). According to our results, females were larger (54.64 ± 0.53 mm) and older (7.68 ± 0.24 years) than males (48.13 ± 0.39 mm; 5.89 ± 0.39 years) in both eastern and western clades. So, the difference has been attributed to the variation in precipitation between the east and west of the sea, leading to different living conditions for R. pseudodalmatina. Hence, any conservation planning to protect this native species of the Hyrcanian forests must consider inter and intra-population differences in the east and west of the Hyrcanian forests.

Female-biased SSD age structure life history the Hyrcanian frog climate data

The Hyrcanian forests, which have survived the Cenozoic geology and the Ice Age due to the northern temperate climate in northern Iran, are a unique forest mass along the southern shores of the Caspian Sea, dating back to the Triassic period, which has been registered as Iran's second natural heritage in the UNESCO World Heritage List (Akhani et al. 2010). These forests are spread in some parts of the five provinces of Semnan, Golestan, Gilan, Mazandaran, and North Khorasan with an area of 1.9 million hectares (approximately 800 km long and 20 to 70 km wide) from Astara in the north of Gilan to Glidagh in the east of Golestan province (Akhani et al. 2010; Surfira 2019; Misir et al. 2012). These forests have a very diverse fauna and flora. The forests of northern Iran were one of the main refugium in the ice ages, so many endemic species of plants and animals can be seen in these forests (Akhani et al. 2010; IUCN 2008).

The Iranian long-legged wood frog, Rana pseudodalmatina Eiselt & Schmidtler, 1971 is one of the endemic species in the Hyrcanian forests (Veith et al. 2003). There is considerable genetic diversity among the wood frog populations (Najibzadeh et al. 2017a). The phylogenetic and morphologic analysis clearly indicates the existence of two major haplogroups in R. pseudodalmatina (Najibzadeh et al. 2017a). The eastern clade haplotypes occur throughout localities east of the Hyrcanian forest, whereas haplotypes belonging to the western clade are almost completely confined to localities west of the Hyrcanian forest (Najibzadeh et al. 2017a).

The life history of an organism comprises its growth, development, age, and reproduction. Differences in the life history of the same species can be attributed to different habitat quality or climate (Hastings 1997). An effective method to determine the age of anurans is skeletochronology. In temperate climates, anurans produce annual growth patterns that include a broad growth annulus during the warm season and a Line of Arrested Growth (LAG) during the cooler season (McCreary et al. 2008). Counting the LAGs can help estimate the age of an individual anuran because it corresponds to an annual cycle of increased and decreased activity (Morrison et al. 2004; Sinsch 2015). Investigation of the growth rate and distribution rate in amphibians provides an important source of information on life-history traits (Amado et al. 2019). The trend characteristics such as age structure, age at sexual maturity, longevity, and detection probability in populations can potentially be used to assess the lifespan of amphibians (Altunışık 2018). The aim of this research is to examine and compare some life history traits, including age estimation, age at sexual maturity, longevity, the correlation between body size and age, the correlation between some cofactors such as precipitation and temperature with age and the study of detection probability of species and their correlation with age, between populations in the eastern and western clades. In addition, this is the first attempt from Iran to determine the characteristics of the population dynamic of R. pseudodalmatina and provide new insights into this endemic species.

Study area

A total of 151 adult specimens (74 males, 77 females) of R. pseudodalmatina from 14 different stations located in the distribution area from north of Iran were analyzed for study of the life history traits. Field work and sampling were carried out in seven sampling sites in the western of the Hyrcanian forest and seven sampling sites in the eastern of the Hyrcanian forest at the southern edge of the Caspian Sea (Table S1; Fig. 1). In addition, climate data on rainfall and temperature of the studied sites from the past 22 years were extracted from website https://www.meteoblue.com based on coordinate data of the sampling sites (Table S1).

Table 1

Discriminate data of 14 sampling sites of the two clades of the Hyrcanian frogs, *Rana pseudodalmatina.*
Clades	Sex	n	Mean Age (yrs)	Range. Age	Mean Age (yrs)	Mean. SVL (mm)	Range. SVL (mm)	Mean SVL (mm)	Mean annual temperature (°C)	Mean Annual rainfall (mm)
Western clade	Female	40	6.22 ± 0.13	4–11	5.29 ± 0.74^a	49.48 ± 0.56	44.91–65.56	46.19 ± 0.56^a	15.24	1485.57
Western clade	Male	35	4.22 ± 0.11	3–6	5.29 ± 0.74^a	42.43 ± 1.06	36.89–49.20	46.19 ± 0.56^a	15.24	1485.57
Eastern Clade	Female	38	9.15 ± 0.36	6–12	8.36 ± 0.08^b	59.80 ± 0.54	52.24-68	56.82 ± 0.54^b	18.09	601.48
Eastern Clade	Male	38	7.57 ± 0.67	4–10	8.36 ± 0.08^b	53.84 ± 0.49	47.6-60.37	56.82 ± 0.54^b	18.09	601.48
Total	Female + Males	151	6.84 ± 0.61	3–12	6.84 ± 0.61	51.54 ± 0.58	36.89-68	51.54 ± 0.58	-	-
Non-identical letters indicate statistical significant.

Detection probability

In order to compare the detection probability of the Hyrcanian frogs in the eastern and western clades, we evaluated one breeding habitat in each clade - the Rasht breeding habitat in the west and the Minudasht breeding habitat in the east.

Capture mark–recapture (CMR) methods are common for estimating population size in many species (Roy et al. 2014). Most of the models that can estimate the detection probability in each region depend on various factors such as migration rates or require long-term monitoring (Tanadini and Schmidt 2011; Hickey and Sollmann 2018). The repeated capture–re-capture (RCR) method is a method it is catching, marking and releasing, which can be used to quickly determine the probability of the presence of individuals in each area. This method is also simulated by simulators, and the results are completely consistent with reality (Pesarakloo et al. 2019). The RCR plan involves repetition of capturing, counting, marking (toe clipping in this study), and releasing the animals in the habitat for K times in short periods. To estimate the detection probability of R. pseudodalmatina we use the following formula:

$$\widehat{\text{p}}=\frac{\sum _{\text{i}=1}^{\text{K}}{\text{U}}_{\text{i}}}{\sum _{\text{i}=1}^{\text{K}}{\text{C}}_{\text{i}}} (\text{P}\text{e}\text{s}\text{a}\text{r}\text{a}\text{k}\text{l}\text{o}\text{o} \text{e}\text{t} \text{a}\text{l}. 2019)$$

Age determination

Like other amphibians, frogs in temperate climates each year of growth during the warm seasons, and the subsequent slowing of growth during the cooler seasons create a repeated pattern in their bone called Lines of Arrested Growth (LAGs). Because this growth pattern predominately corresponds to an annual cycle, the LAG count can be taken as an individual's age estimation (Morrison et al. 2004). Age estimation was performed using the standard method of skeletochronologycal analysis according to (Castanet and Smirina 1990) by counting the lines of arrested growth (LAGs) in stained cross sections of the phalangeal bones (Jin et al. 2017). From each specimen, the second segment was cut from the longest toe (4th) of the right or left hindlimb and placed in formaldehyde (10%) for histological analysis. The phalanges were decalcified in nitric acid (5%) and after dehydration, the serial sections (18–20 µm) from the mid-diaphyseal part were cut using a freezing microtome and then stained with Haematoxylin-eosin.

The cross sections fixed under a light microscope (Olympus BX51) and were independently check outed by two observers for age estimation. Because resorption of the innermost resting line follow-up endosteal resorption, sections whit having the smallest medullar cavity were selected. Longevity (maximum lifespan) were estimated according to Otero et al. (2017): longevity is maximum number of LAGs counted in adults. To estimate age at sexual maturity using three combined methods: 1. according to Ryser (1998), the criterion was the closest distance between two subsequent LAG; 2. Age at sexual maturity defined by the minimum number of LAGs counted in breeding individuals (Otero et al. 2017); and 3. Arısoy and Başkale (2019) assumed that the age at first reproduction is the lowest age recorded amongst the breeding individuals. In this study, we utilized the method introduced by Otero et al. (2017) to determine sexual maturity.

Data analysis

In this study, we wrote a computer program in R language (R is an open source and free statistical software, which is available at www.r-project.org) to Data analysis.

Shapiro-Wilk test was performed to assess the normality distribution of the data

Sexual size dimorphism was computed according to Lovich and Gibbons (1992) SDI defined by: (mean length of larger sex/mean length of smaller sex) ± 1, when the females are larger, the result is arbitrarily defined as positive (+ 1) and negative (-1), if males are larger sex. Body size or Snout-vent length (SVL) was measured with a digital caliper to the nearest 0.01.

One-way ANOVA was performed to compare the mean of the life history parameters (SVL, age) between the frogs of the western and eastern clades and the females and males of R. pseudodalmatina from sampling sites.

Multivariate analysis carried out using PAST software (version 2.20, folk.uio.no/ohammer/past). Principle component analysis (PCA) was performed to check variance for each of the study groups. Spearman's correlation coefficient was used to investigate the relationship between the SVL/age of sexes and the frogs of the western and eastern clades. We used linear mixed regression models to assess the impact of climate data (mean annual temperature and mean annual rainfall) on the average of SVL, age and detection probability of the Hyrcanian frogs in the eastern and western clades.

Detection probability

This study used the RCR sampling plan with the stopping criteria of 10 successive zeros for new capture counts${c_i}$. By these criteria, we stopped the RCR plan in the Rasht site (of the eastern clade) at 38 iterations and in the Minudasht site (of the western clade) at 24 iterations. The series of $\{ {c_i}\} _{{i=1}}^{K}$and $\{ {u_i}\} _{{i=1}}^{K}$are displayed in Fig. 2a, b. We can see that the series of newly captured animals decreases over time, while the series of all captured animals $\{ {c_i}\} _{{i=1}}^{K}$are stationary. According to the data of this study, the estimated probability of detecting the Hyrcanian frog in the eastern clade is p̂ = 0.042, with a standard error of 0.0032. Similarly, in the western clade, the estimated detection probability is p̂ = 0.062 with a standard error of 0.0051. Although the main objective of this study was not to estimate forest frog populations, the probability of their presence obtained in this study can be used to estimate their numbers in future studies.

3.2. Growth estimation and age determination

The study found that the mean body size (SVL) for the western and eastern clades was (46.19 ± 0.56 mm, 56.82 ± 0.54 mm, respectively). The age was determined by observing the Hematoxynophilic lines in the light microscope. In all cross sections, the lines were clearly marked and results of age estimating obtained by direct observation in the light microscope (Fig. 3(. The mean age of the western and eastern clades was (5.29 ± 0.74 years, 8.36 ± 0.08 years, respectively). More information on the variables measured is provided in Table 1. There was a significant difference in SVLs and age structure between the western and eastern clades (F = 101.9, p = 3.47×10^− 6; F = 137.5, p = 1.48×10^− 6, respectively).

Two principal components explained 97% of the variation in the life history data set of SVL and age parameters, and the PCA1/2-scores plot showed body size and age in two distinct clades (Fig. 4a).

Additionally, the study revealed that the average total of females (in two western and eastern clades) was larger (females = 54.64 ± 0.55, male = 48.35 ± 0.77) and older (females = 7.68 ± 0.24 years, males = 5.89 ± 0.39 years) than the total of males in both clades (Fig. 4b, Table 1), indicating a difference in body length and age structure between the sexes. The SDI was 0.086, indicating a difference in body length between the sexes (i.e., female-biased SSD).

The breeding females of the western clade of R. pseudodalmatina reach sexual maturity at around four years of age, while males reach maturity at three years. In contrast, the females of the eastern clade reach sexual maturity at around sex years of age, and their males reach maturity at around four years of age. In addition, the maximum lifespan of females is 12 years, while males have a maximum lifespan of 10 years (Table 1).

A significant positive relation was detected between age and body size (SVL) in frogs of the western and eastern clades and both sexes of population of R. pseudodalmatina (Spearman’s correlation; r = 0.88, p < 0.05). The growth curves represented good fit to correlation between body size and age, as it is presented in Fig. 5a,b.

The effects of climatic factors on the Hyrcanian frog

The results of the Linear mixed regression analysis showed a significant effect of the climatic factors of temperature and rainfall on the SVL, age structure and detection probability of the Hyrcanian frogs (p < 0.01) (Table 2). The results revealed that with increasing temperature, SVL and age increase, but the detection probability of the frog decreases. With more rainfall, SVL and age of the frogs decrease, but the detection probability of the frog increases. This means that temperature has a positive relationship with SVL and age, but it has a negative relationship with the detection probability of the Hyrcanian frog. Rainfall has a positive relationship with the detection probability, but a negative relationship with SVL and age.

Table S1

General information about Hyrcanian frogs collected from 14 sampling sites from two western and eastern clades.
NO	Clade	Locality		SVL	Sex	Age	Long	Lat	Mean annual temperature	Mean Annual rainfall
1	Western	1	Anzali	48.34	Female	7	49.11	37.38	1377.86	16.65
2	Western		Anzali	45.87	Female	5	49.11	37.38	1377.86	16.65
3	Western		Anzali	45.31	Female	6	49.11	37.38	1377.86	16.65
4	Western		Anzali	49.27	Female	7	49.11	37.38	1377.86	16.65
5	Western		Anzali	45.08	Female	5	49.11	37.38	1377.86	16.65
6	Western		Anzali	48.68	Female	6	49.11	37.38	1377.86	16.65
7	Western		Anzali	43.81	Male	3	49.11	37.38	1377.86	16.65
8	Western		Anzali	44.16	Male	6	49.11	37.38	1377.86	16.65
9	Western		Anzali	40.91	Male	3	49.11	37.38	1377.86	16.65
10	Western		Anzali	41.59	Male	3	49.11	37.38	1377.86	16.65
11	Western	2	Astara	46.15	Female	6	48.83	38.34	1365.04	12.65
12	Western		Astara	45.69	Female	6	48.83	38.34	1365.04	12.65
13	Western		Astara	47.1	Female	6	48.83	38.34	1365.04	12.65
14	Western		Astara	46.49	Female	6	48.83	38.34	1365.04	12.65
15	Western		Astara	45.37	Female	5	48.83	38.34	1365.04	12.65
16	Western		Astara	42.41	Male	5	48.83	38.34	1365.04	12.65
17	Western		Astara	41.28	Male	3	48.83	38.34	1365.04	12.65
18	Western		Astara	43.18	Male	6	48.83	38.34	1365.04	12.65
19	Western		Astara	42.19	Male	5	48.83	38.34	1365.04	12.65
20	Western		Astara	41.68	Male	5	48.83	38.34	1365.04	12.65
21	Western	3	Rasht	50.27	Female	7	49.51	37.15	1477.96	16.06
22	Western		Rasht	47.84	Female	4	49.51	37.15	1477.96	16.06
23	Western		Rasht	46.38	Female	6	49.51	37.15	1477.96	16.06
24	Western		Rasht	49.37	Female	7	49.51	37.15	1477.96	16.06
25	Western		Rasht	50.95	Female	8	49.51	37.15	1477.96	16.06
26	Western		Rasht	46.68	Female	6	49.51	37.15	1477.96	16.06
27	Western		Rasht	45.32	Female	6	49.51	37.15	1477.96	16.06
28	Western		Rasht	46.02	Female	6	49.51	37.15	1477.96	16.06
29	Western		Rasht	44.53	Male	5	49.51	37.15	1477.96	16.06
30	Western		Rasht	42.86	Male	5	49.51	37.15	1477.96	16.06
31	Western		Rasht	41.67	Male	5	49.51	37.15	1477.96	16.06
32	Western		Rasht	40.75	Male	4	49.51	37.15	1477.96	16.06
33	Western		Rasht	41.34	Male	5	49.51	37.15	1477.96	16.06
34	Western		Rasht	42.42	Male	5	49.51	37.15	1477.96	16.06
35	Western	4	Asalem	51.34	Female	9	48.92	37.7	1884.73	13.36
36	Western		Asalem	65.56	Female	11	48.92	37.7	1884.73	13.36
37	Western		Asalem	48.2	Female	6	48.92	37.7	1884.73	13.36
38	Western		Asalem	44.91	Female	4	48.92	37.7	1884.73	13.36
39	Western		Asalem	43.24	Male	4	48.92	37.7	1884.73	13.36
40	Western		Asalem	39.01	Male	3	48.92	37.7	1884.73	13.36
41	Western		Asalem	41.87	Male	4	48.92	37.7	1884.73	13.36
42	Western		Asalem	36.89	Male	3	48.92	37.7	1884.73	13.36
43	Western		Asalem	41.01	Male	5	48.92	37.7	1884.73	13.36
44	Western	5	Amlash	45.21	Female	5	50.16	37.09	1334.77	16.69
45	Western		Amlash	53.21	Female	7	50.16	37.09	1334.77	16.69
46	Western		Amlash	48.21	Female	5	50.16	37.09	1334.77	16.69
47	Western		Amlash	50.78	Female	6	50.16	37.09	1334.77	16.69
48	Western		Amlash	50.92	Female	6	50.16	37.09	1334.77	16.69
49	Western		Amlash	43.59	Male	4	50.16	37.09	1334.77	16.69
50	Western		Amlash	37.49	Male	3	50.16	37.09	1334.77	16.69
51	Western		Amlash	41.17	Male	4	50.16	37.09	1334.77	16.69
52	Western		Amlash	40.31	Male	4	50.16	37.09	1334.77	16.69
53	Western	6	Langroud	54.37	Female	7	50.16	37.17	1335.16	16.69
54	Western		Langroud	50.98	Female	6	50.16	37.17	1335.16	16.69
55	Western		Langroud	48.51	Female	5	50.16	37.17	1335.16	16.69
56	Western	6	Langroud	50.43	Female	6	50.16	37.17	1335.16	16.69
57	Western		Langroud	44.39	Male	4	50.16	37.17	1335.16	16.69
58	Western		Langroud	41.35	Male	4	50.16	37.17	1335.16	16.69
59	Western		Langroud	38.28	Male	4	50.16	37.17	1335.16	16.69
60	Western		Langroud	42.45	Male	4	50.16	37.17	1335.16	16.69
61	Western		Langroud	44.31	Male	5	50.16	37.17	1335.16	16.69
62	Western		Langroud	42.87	Male	4	50.16	37.17	1335.16	16.69
63	Western	7	Rudsar	56.26	Female	7	50.28	36.99	1613.14	14.45
64	Western		Rudsar	54.36	Female	6	50.28	36.99	1613.14	14.45
65	Western		Rudsar	50.75	Female	8	50.28	36.99	1613.14	14.45
66	Western		Rudsar	55.21	Female	9	50.28	36.99	1613.14	14.45
67	Western		Rudsar	47.68	Female	5	50.28	36.99	1613.14	14.45
68	Western		Rudsar	50.84	Female	5	50.28	36.99	1613.14	14.45
69	Western		Rudsar	53.63	Female	6	50.28	36.99	1613.14	14.45
70	Western		Rudsar	51.84	Female	5	50.28	36.99	1613.14	14.45
71	Western		Rudsar	49.2	Male	5	50.28	36.99	1613.14	14.45
72	Western		Rudsar	46.38	Male	4	50.28	36.99	1613.14	14.45
73	Western		Rudsar	44.97	Male	4	50.28	36.99	1613.14	14.45
74	Western		Rudsar	45.38	Male	4	50.28	36.99	1613.14	14.45
75	Western		Rudsar	46.14	Male	4	50.28	36.99	1613.14	14.45
76	Eastern	8	Loveh	54.36	Female	6	55.65	37.35	532.4	18.15
77	Eastern		Loveh	53.8	Female	7	55.65	37.35	532.4	18.15
78	Eastern		Loveh	56.48	Female	8	55.65	37.35	532.4	18.15
79	Eastern		Loveh	56.93	Female	8	55.65	37.35	532.4	18.15
80	Eastern		Loveh	61.28	Female	9	55.65	37.35	532.4	18.15
81	Eastern		Loveh	56.81	Female	7	55.65	37.35	532.4	18.15
82	Eastern		Loveh	52.3	Male	6	55.65	37.35	532.4	18.15
83	Eastern		Loveh	50.28	Male	6	55.65	37.35	532.4	18.15
84	Eastern		Loveh	55.4	Male	7	55.65	37.35	532.4	18.15
85	Eastern		Loveh	60.37	Male	8	55.65	37.35	532.4	18.15
86	Eastern		Loveh	53.18	Male	6	55.65	37.35	532.4	18.15
87	Eastern	9	Minudasht	63.28	Female	7	55.37	37.17	379.83	19.15
88	Eastern		Minudasht	61.51	Female	9	55.37	37.17	379.83	19.15
89	Eastern		Minudasht	56.46	Female	8	55.37	37.17	379.83	19.15
90	Eastern		Minudasht	55.28	Female	7	55.37	37.17	379.83	19.15
91	Eastern		Minudasht	52.37	Female	6	55.37	37.17	379.83	19.15
92	Eastern		Minudasht	62.15	Female	9	55.37	37.17	379.83	19.15
93	Eastern		Minudasht	62.38	Female	10	55.37	37.17	379.83	19.15
94	Eastern		Minudasht	57.54	Female	9	55.37	37.17	379.83	19.15
95	Eastern		Minudasht	54.37	Male	9	55.37	37.17	379.83	19.15
96	Eastern		Minudasht	49.62	Male	7	55.37	37.17	379.83	19.15
97	Eastern		Minudasht	48	Male	7	55.37	37.17	379.83	19.15
98	Eastern		Minudasht	53.44	Male	9	55.37	37.17	379.83	19.15
99	Eastern		Minudasht	50.98	Male	9	55.37	37.17	379.83	19.15
100	Eastern		Minudasht	51.06	Male	8	55.37	37.17	379.83	19.15
101	Eastern		Minudasht	54.38	Male	8	55.37	37.17	379.83	19.15
102	Eastern	10	Fazelabad	65.01	Female	11	54.77	36.84	684.55	18
103	Eastern		Fazelabad	63.74	Female	11	54.77	36.84	684.55	18
104	Eastern		Fazelabad	64.94	Female	10	54.77	36.84	684.55	18
105	Eastern		Fazelabad	57.33	Female	9	54.77	36.84	684.55	18
106	Eastern		Fazelabad	57.88	Female	9	54.77	36.84	684.55	18
107	Eastern		Fazelabad	60.11	Female	10	54.77	36.84	684.55	18
108	Eastern		Fazelabad	56.89	Male	8	54.77	36.84	684.55	18
109	Eastern		Fazelabad	53.46	Male	7	54.77	36.84	684.55	18
110	Eastern		Fazelabad	57.82	Male	8	54.77	36.84	684.55	18
111	Eastern		Fazelabad	55.67	Male	7	54.77	36.84	684.55	18
112	Eastern		Fazelabad	58.3	Male	8	54.77	36.84	684.55	18
113	Eastern		Fazelabad	52.87	Male	8	54.77	36.84	684.55	18
114	Eastern	11	Kordkuy	57.24	Female	9	54.13	36.74	532.93	19.1
115	Eastern		Kordkuy	61.25	Female	10	54.13	36.74	532.93	19.1
116	Eastern		Kordkuy	64.86	Female	11	54.13	36.74	532.93	19.1
117	Eastern		Kordkuy	59.13	Female	10	54.13	36.74	532.93	19.1
118	Eastern		Kordkuy	56.38	Female	10	54.13	36.74	532.93	19.1
119	Eastern		Kordkuy	55.74	Male	10	54.13	36.74	532.93	19.1
120	Eastern		Kordkuy	48.26	Male	5	54.13	36.74	532.93	19.1
121	Eastern		Kordkuy	47.6	Male	4	54.13	36.74	532.93	19.1
122	Eastern		Kordkuy	52.31	Male	7	54.13	36.74	532.93	19.1
123	Eastern	12	Behshahr	56.8	Female	9	53.59	36.66	632.01	17.63
124	Eastern		Behshahr	61.27	Female	10	53.59	36.66	632.01	17.63
125	Eastern		Behshahr	55.93	Female	9	53.59	36.66	632.01	17.63
126	Eastern		Behshahr	52.24	Female	8	53.59	36.66	632.01	17.63
127	Eastern		Behshahr	57	Female	10	53.59	36.66	632.01	17.63
128	Eastern		Behshahr	54.32	Male	9	53.59	36.66	632.01	17.63
129	Eastern		Behshahr	48.21	Male	5	53.59	36.66	632.01	17.63
130	Eastern		Behshahr	51.46	Male	9	53.59	36.66	632.01	17.63
131	Eastern		Behshahr	51.55	Male	7	53.59	36.66	632.01	17.63
132	Eastern		Behshahr	53.07	Male	9	53.59	36.66	632.01	17.63
133	Eastern	13	Neka	68	Female	12	53.36	36.62	779.9	17.08
134	Eastern		Neka	64.23	Female	10	53.36	36.62	779.9	17.08
135	Eastern		Neka	65.67	Female	9	53.36	36.62	779.9	17.08
136	Eastern		Neka	56.34	Female	7	53.36	36.62	779.9	17.08
137	Eastern		Neka	57.5	Male	8	53.36	36.62	779.9	17.08
138	Eastern		Neka	59.64	Male	10	53.36	36.62	779.9	17.08
139	Eastern		Neka	52.61	Male	9	53.36	36.62	779.9	17.08
140	Eastern		Neka	54.37	Male	8	53.36	36.62	779.9	17.08
141	Eastern		Neka	56.01	Male	8	53.36	36.62	779.9	17.08
142	Eastern	14	Sari	67.31	Female	12	52.6	36.33	780.85	17.06
143	Eastern		Sari	62.38	Female	11	52.6	36.33	780.85	17.06
144	Eastern		Sari	65.59	Female	11	52.6	36.33	780.85	17.06
145	Eastern		Sari	61.25	Female	10	52.6	36.33	780.85	17.06
146	Eastern		Sari	56.31	Male	7	52.6	36.33	780.85	17.06
147	Eastern		Sari	57.51	Male	6	52.6	36.33	780.85	17.06
148	Eastern		Sari	55.87	Male	7	52.6	36.33	780.85	17.06
149	Eastern		Sari	55.55	Male	7	52.6	36.33	780.85	17.06
150	Eastern		Sari	56.25	Male	8	52.6	36.33	780.85	17.06
151	Eastern		Sari	53.67	Male	9	52.6	36.33	780.85	17.06

Table 2

The influences of climate data on SVL, age and detection probability of the Hyrcanian frogs (*Rana pseudodalmatrina*) in the eastern and western clades using Linear mixed regression.
	Coefficient	Std.err.	t- value	p-value
SVL^a
Constant	70.38	19.647	3.5822	0.0043*
Mean annual temperature	2.0108	0.66115	3.0413	0.0102*
Mean annual rainfall	-0.01022	0.00202	-5.0611	0.0002*
Age ^b
Constant	14.362	5.269	2.7258	0.0197*
Mean annual temperature	0.5615	0.19413	2.8924	0.0135*
Annual rainfall	-0.003	0.000555	-5.4083	0.0001*
Detection probability^c
Constant	0.1177	0.01713	6.8709	< 0.0001*
Mean annual temperature	-0.00395	0.001024	-3.8598	0.0002*
Annual rainfall	2.02E-05	2.20E-06	9.1617	< 0.0001*
^a Dependent variables: SVL_; ^b Dependent variables :Age_; ^c Dependent variables: Detection probability

As the results of this study showed, the probability of encountering a forest frog in the western half of the forests of the southern shores of the Caspian Sea is much higher than in the forests of the eastern sea. It could mean that frogs are more abundant and dense in the western half of the sea. In other words, the living conditions in the west of the sea are better for the forest frog than in the east of the Hyrcanian forests. Objective observations confirm that there are more reproductive habitats in the west of the Hyrcanian forest than in the east of the Hyrcanian forests. The study on the Hyrcanian forests revealed that the precipitation and temperature in the east and west regions are significantly different. The mean precipitation in the west is double the amount in the east, and the mean temperature in the west is about 3°C colder than in the east. Previous studies on amphibian distribution around the world have shown that more precipitation and lower temperatures lead to species richness and increased abundance of amphibians (Ochoa-Ochoa et al. 2019; Walls et al. 2013; Ficetola et al. 2016). The results of this study also confirm the above.

Sexual size dimorphism has been studied many times by researchers on different species of amphibians (e.g. Monnet et al. 2002; Liao et al. 2013; Rastegar-Pouyani et al. 2013; Yan et al. 2023; Dursun et al. 2023). According to the difference in Snout-vent length (SVL) in amphibians, there are three patterns for sexual dimorphism. Female-biased when females are larger than males, male-biased when males are larger than females, and unbiased when males are equal to females (Zhang et al. 2016). For example, In Rana holtzi Werner (1898), SVL ranged from 46.4 to 66.8mm for males and 39.2 to 66mm for females (Guarino et al. 2008). Our results in in populations of both clades showed that the size of female frogs is larger than male ones. A significant difference observed between females and males concerning mean SVL. The data obtained from the current study confirm the general patterns of body size variation in amphibians and suggest that females are larger than males (Reinhard et al. 2015; Zhang et al. 2016).

Upon comparing the mean body size between Eastern and Western samples, it was found that Eastern samples tend to be larger. At first, this may seem counterintuitive because one would expect the Western clade of the forest frog to have more favorable living conditions, resulting in higher SVLs and life expectancy. However, the results suggest otherwise. Field observations showed that the reproductive habitats in the west of the Hyrcanian forests are more than in the east, resulting in more frequent breeding. In the east of the Hyrcanian forests, breeding habitats are few and far apart, and you have to traverse several kilometers on the forest floor to find a specimen. The studies of Najibzadeh et al. (2017b) showed that the Hyrcanian frogs sometimes travel kilometers on the forest floor to find breeding habitats. Our observation has revealed that as summer approaches, the breeding habitats located in the east gradually dry up, leading to a significant loss of tadpoles. Pesarakloo et al. (2012) conducted a study on the reproductive biology of the Hyrcanian frog in the Minudasht habitat. According to their findings, the frogs in the Minudasht habitat have adapted to the winter cold to avoid the loss of larvae in late spring due to the drying of the aquatic habitat. They engage in reproductive activities during winter and spawn in March. On the other hand, the marsh frog, Pelophylax sp, living in ponds and outside the forest hibernates during this time. So, one of the important reasons explaining the larger body size of Eastern samples than Western samples is age differences (Monnet and Cherry 2002; Young 2005; Liao et al. 2011) and survival (Vargas-Salinas 2006). To prove this supplication, one should refer to the results of age measurement. The results of the present studies show that the samples of the western branch have a lower mean age (5.29 ± 0.74 years) than the eastern samples (8.36 ± 0.08 years). On the other hand, there is a direct relationship between age and body size. A general positive correlation between body size and age is well-documented among amphibians (e.g. Halliday and Verrell 1988; Cogâlniceanu and Miaud 2003; Bülbül et al. 2018).

According to our results, females were larger (54.64 ± 0.53 mm) and older (7.68 ± 0.24 years) than males (48.13 ± 0.39 mm; 5.89 ± 0.39 years) in both eastern and western clades. The China Wood Frog (Rana chensinensis) the maximum observed longevity was 5 years in males and 4 years in females, respectively (Chen et al. 2011). Age at sexual maturity in both sexes was 1 years (Chen et al. 2011). In the frog endemic to the Middle Taurus Mountain Range, R. holtzi, Ages ranged from 4 to 6 years for males, and from 4 to 7 for females (Guarino et al. 2008). The life expectancy of females and males of P. caralitanus was estimated at 6.61 and 6.15 years, respectively (Arısoy and Başkale 2019). The mean survival rates of P. caralitanus females and males were calculated as 0.84 and 0.83, respectively.

Survival rates and life expectancy are estimated at 0.79 and 5.26 years in Ommatotriton vittatus, respectively (Altunışık 2018). The age at maturity of O. nesterovi, O. ophryticus, and O. vittatus were determined as 4–6 years, 3–5 years, and 3 years, respectively (Ozcan and Uzum 2015; Kuzmin 1999; Altunışık 2018). In another example, Altunışık et al. (2017) reported the maximum observed lifespan was eight years for females (mean 4.28 years, range 3–8 years) and seven years for males (mean 3.08 years, range 2–7 years). Altunışık et al. (2017) continued age at maturity was estimated as two years for males and three years for females in B. variabilis.

The study conducted by Najibzadeh et al. (2017a) found differences in wood frog samples from the east and west of the Hyrcanian forests using molecular and morphological data. This study also showed that the two clades are significantly different in terms of life history traits. This difference has been attributed to the variation in precipitation between the east and west of the sea, leading to different living conditions for the species. Hence, any conservation planning to protect this native species of Hyrcanian forests must consider population differences in the east and west of the Hyrcanian forests and intra-population differences such as gender and age.

Acknowledgements

The author gratefully thanks Hasan Pesarakloo, Amireza Zarei for their support during sampling activities. The authors acknowledge Arak University for supporting with all sorts of laboratory facilities.

Author contributions

M.N. designed and conducted the study, analyzed data, and wrote the manuscript.

Data availability

All Data will be provided to readers on request ([email protected]). Data directly linked to the article are available in Supporting Information.

Competing interests The authors have no relevant financial or non-financial interests to disclose.

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Supplementarydata.docx

First detection probability, comparative analysis of ecological disparities and population dynamics of Rana pseudodalmatina (Eiselt & Schmidtler, 1971) in the Hyrcanian forests: implications for conservation planning

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Abstract

Figures

Introduction

Material and methods

Study area

Detection probability

Age determination

Data analysis

Results

Detection probability

3.2. Growth estimation and age determination

The effects of climatic factors on the Hyrcanian frog

Discussion

Conclusion

Declarations

References

Additional Declarations

Supplementary Files

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Journal Publication

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