The Relationship between Population Density and Body Size of the Giant Mountain Crab Indochinamon bhumibol (Naiyanetr, 2001), an Endangered Species of Freshwater Crab from Northeastern Thailand (Potamoidea: Potamidae)

Abstract

The giant mountain crab (Indochinamon bhumibol) is endemic to Loei Province in north-eastern Thailand. Although, little knowledge is available on the factors affecting population dynamics and individual fitness. This study aimed to investigate the relationship between the population density and body size of the crab. The mark–recapture method was conducted in Phu Ruea National Park (PR, small area) and Phu Luang Wildlife Sanctuary (PL, large area) between February 2018 and January 2019. The crabs inhabited mountain streams at 400–1100 m. The average water depth and width were 0.6 m and 8.3 m, respectively. In total, 879 individuals were caught from PL compared to 583 individuals from PR. Male were more abundant than female in both areas. Male were also larger and heavier, with a mean weight of 145 g, mean carapace width of 76 mm and mean carapace length of 75.7 mm. The average number of crabs caught was higher in the rainy season (66.3 individuals). The crab population was larger in PL (256.7 individuals), which is bigger in size than PR and is located farther from urban areas. The average crab density in PL and PR were 1.7 and 1.6 crabs/m², respectively. Population density was significantly correlated with body size parameters: body weight (r² = 0.066, df = 1196, p < 0.001), carapace width (r² = 0.053, df = 1196, p < 0.001) and carapace length (r² = 0.067, df = 1196, p < 0.001). The findings of this study suggest that a large area size and high population density are important to the conservation of the giant mountain crab in mountainous forests.

1. Introduction

Asia is a global center of freshwater crab (phylum Arthropoda, order Decapoda) species diversity. Thailand is the second most species-rich country in Southeast Asia, showing 86% of endemism for freshwater crab species [1]. The freshwater crabs of Thailand belong to the Gecarcinucidae [2] and Potamidae [3] families. At least 101 species have been reported, of which 19 are threatened, 11 are vulnerable and 8 are endangered [1]. The giant mountain crab (Indochinamon bhumibol, family Potamidae) is the largest species of freshwater crab in Thailand [4]. The minimum–maximum weight, carapace width and carapace length of the giant crabs were 31.6–98.2 g, 4.2–6.5 cm and 3.2–5 cm, respectively; large were 12.1–72.2 g, respectively, 3.1–6.3 cm and 2.4–4.8 cm.; and small were 9.4–4.9 g, 2.9–4.8 cm and 2.3–3.7 cm, respectively [5]. It was found in Phu Ruea District, Dan Sai District, Tha Li District, Phu Luang District, Pakchom District, Loei Province; Lomkao District, Phetchabun Province; Sang-khom District, Nong Khai Province; and Nayoong District, Udonthani Province, northeastern Thailand [5]. The International Union for the Conservation of Nature (IUCN) Red List assessed this species as an endangered (EN) species [6] because it is threatened by habitat destruction and pollution. Although the habitats of freshwater crabs in tropical regions are diverse and include rivers, streams, waterfalls, wetlands, karsts and caves, this species is highly limited in its distribution and is mostly found in the waterfalls and stream channels of mountain forests [7]. According to previous studies, water pollution, hunting, habitat destruction in the form of deforestation and increasing agriculture are threats to freshwater crabs in Asia. A previous study of freshwater crab in Thailand emphasized taxonomy and diversity aspects [4,5,8,9,10,11], but information is lacking on the population size, abundance, trends and ecology of giant mountain crabs. This study aimed to investigate the relationship between the population density and body size of I. bhumibol in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR), Loei Province, as they are at the head watershed of Loie Province. Furthermore, the holotype and paratype of I. bhumibol were collected in PL [4]. The findings of this study may enhance the knowledge of the population dynamics and body size patterns of this endemic species to northeastern Thailand, which could lead to suggestions of how such a plan could be formulated and area management strategies to protect it from imminent extinction. Moreover, from a more fundamental standpoint, the relationship between habitat and population density as well as the link between population density and body size require investigation.

2. Materials and Methods

2.1. Study Area

This study was conducted in Phu Luang Wildlife Sanctuary (PL; total area = 897 km²) and Phu Ruea National Park (PR; total area = 120 km²) in Loei Province, Thailand. The observations were conducted monthly between February 2018 and January 2019. Six sampling sites along the main stream in both protected areas were chosen based on the presence of crabs during a preliminary survey between April and August 2017. The six sampling sites in PR were Huay Tiew, Huay Kawak, Huay Sai khao, Huay Phai, Huay Toei and Huay Nam Khan. The six sampling sites in PL were Huay Nam San, Huay Nam Thop, Huay Nam Khai, Huay Nam Loei, Huay Sak and Huay Pla Ba (Figure 1). A total of 12 sampling sites (six in each protected area) were chosen in streams ranging from 6 to 9 m in width. At each sampling site, three line transects were established along the stream; each transect was 50 m in length and approximately 100 m in distance from other transects. All sampling sites were located 400 to 1100 m above sea level (ASL) on the upper slope of the mountain. The soil texture was sandy loam in both areas. The annual average temperature in Loei Province during the study period was 26 °C. The average stream width was 9.7 ± 6.7 m in PL and 6.8 ± 3.8 m in PR. The average depth of water was 0.8 ± 0.8 m in PL and 0.5 ± 0.3 m in PR. The average water temperature was 17.9 ± 3.6 °C in PL and 18 ± 3.1 °C in PR. The average pH was 7.6 ± 1 in PL and 6.8 ± 0.6 in PR (

Table 1. Physical characteristics of the streams and the weather in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR) between February 2018 and January 2019.

Environmental Variables	Study Site	Sand	Silt	Clay	Mean ± Sd	df	F	p
Soil texture (%)	PL	50–70	0	15–20
	PR	50–70	0	15–20
Depth of water (m)	PL				0.8 ± 0.8	47	1.78	0.085
	PR				0.5 ± 0.3
Stream width (m)	PL				9.7 ± 6.7	47	1.81	0.076
	PR				6.8 ± 3.8
Water temperature (°C)	PL				17.9 ± 3.6	47	−0.09	0.932
	PR				18.0 ± 3.1
pH	PL				7.6 ± 1.0	47	3.16	0.003
	PR				6.8 ± 0.6
Air temperature (°C)	PL				26.7 ± 1.4	47	−0.5	0.623
	PR				26.9 ± 1.5

).

2.2. Field Sampling and Body Size Measurement

A monthly field survey was carried out, with one site from PL and another from PR. A different set of PL and PR sites was surveyed the following month. Therefore, each sampling site was surveyed twice during the year, once in the dry season (November to April) and once in the rainy season (May to October). At each sampling site, a 1 × 1 m² plot was demarcated every 5 m along the transect in the mountain streams to collect crabs. Thus, each sampling site consisted of 30 sampling plots. As freshwater crabs are generally active at night, the survey was conducted between 6 pm and 12 pm. A small piece of fresh chicken was used as bait [12]. The capture–mark recapture protocol was applied in the field survey. The morphological characters, carapaces, chelipeds, walking legs, abdomen, chelae, third maxilla, mandibular pals, first gonopods (G1) and second gonopod (G2) were analyzed by using Ng and Naiyanetr [3], Chuensri [13] and Naruse et al. [14]. All giant mountain crabs found along the line transect and sampling plots were hand-collected and counted. Each crab was marked and measured for weight, carapace width and carapace length [15]. The weight was measured using an analytical pocket balance, and the carapace was measured with vernier caliper. Sex and age were determined by the shape of the abdomen [16,17]. A waterproof oil-based paint marker was used to mark on the carapace [15]. After recording all data, the marked crab was released at the site where it was caught. The survey was conducted on two consecutive nights each month. The stream width, water depth, water temperature, pH and air temperature at each sampling sites were recorded.

2.3. Data Analysis

The population of I. bhumibol was estimated using the formula; N = M1(M2 + 1)/(X2 + 1), where N is the population estimate, M1 is the number of crabs captured and marked at the first time, M2 is the total number captured the second time (with and without mark) and X2 is the number of crabs with a mark captured for the second time [18]. Crab density was estimated in each sampling site as the number of individuals that were captured per area (total sampling plots) [19]. The sex ratio of the adult stage was estimated each month as the quotient of the number of males and the total number of individuals in the sample [20].

The abundance of I. bhumibol was compared between the two study sites (PL and PR) using a normalization and an independent sample t-test. One-way ANOVA was used to compare (1) body size parameters (carapace width, carapace length and weight) of all crabs and of both sexes between the 12 sampling sites and (2) population size and population density between the 12 sampling sites and seasons at the p = 0.05 level [20]. The relationship between population density and body size was investigated by correlation and linear regression analysis. All statistical analyses were conducted using SPSS v23.

3. Results

3.1. Morphology

The giant mountain crabs in both PL and PR have carapaces that are broader than long, with distinct cervical grooves. The first gonopod is broad. The terminal segment is very short, separated from the subterminal segment and bent outwards to the main axis of gonopod, with the tip curving downwards, whereas the second gonopod has a distal segment more than half the length of the basal segment. The carapace, chelipeds and walking legs are dark brown in color. The tips of the walking legs are orange (Figure 2). The morphological characters of I. bhumibol were described by Naiyanetr [4]. The crabs are mostly found in water or under stones.

3.2. Abundance, Body Size and Sex Ratio

The total number of crabs caught along the line transect and sampling plots survey in the six sampling sites of PL and six sampling sites of PR were 879 and 583 individuals, respectively (

Table 2. Proportion between male and female individuals of all crabs in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR) between February 2018 and January 2019.

Month	PL				PR
	Male	Female	Unk	Sex Ratio	Male	Female	Unk	Sex Ratio
Feb 2018	43	25	5	1:0.6	19	10	1	1:0.7
Mar 2018	31	0	11	N/A	19	8	8	1:0.7
Apr 2018	24	15	3	1:0.6	11	8	0	1:0.6
May 2018	9	14	3	1:0.4	8	17	16	1:0.3
Jun 2018	38	30	11	1:0.6	12	10	4	1:0.6
Jul 2018	27	44	14	1:0.4	15	10	12	1:0.6
Aug 2018	39	26	16	1:0.6	18	13	6	1:0.6
Sep 2018	37	14	4	1:0.7	22	19	16	1:0.5
Oct 2018	89	125	26	1:0.4	11	16	4	1:0.4
Nov 2018	19	11	0	1:0.6	29	24	29	1:0.6
Dec 2018	20	9	6	1:0.7	61	53	32	1:0.5
Jan 2019	50	41	0	1:0.6	22	16	4	1:0.6

The abbreviation Unk means “unidentified sex”, which was due to our inability to catch the individual and measure it, although we were able to observe it within our sampling plots; N/A means “not analyzed”.

). The average number of crabs caught in PL and PR did not differ significantly, 73.3 ± 57.4 and 48.6 ± 34.7 individuals, respectively (t = 1.3, df = 22, p = 0.216). In both study sites, males at the adult stage were the most abundant. The number of crabs caught in PL was higher than the number in PR, and the number of individuals crabs caught in the rainy season (May to October; 66.3 ± 58.7 individuals) was higher than in the dry season (November to April; 55.6 ± 20 individuals).

The averages of body size parameters including weight (W) (F = 24.8, df = 1179, p = 0.001), carapace width (CW) (F = 28.3, df = 1179, p = 0.001) and carapace length (CL) (F = 27.5, df = 1179, p = 0.001) of the crabs were different between sites (

Table 3. Average body size of I. bhumibol in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR).

PA	Sub-Sites	N			Mean ± Sd
					Weight (g)			Carapace (mm)
								Width			Length
		Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju
PL	HNS	125	120	5	49.4 ± 19.6 de	50.9 ± 18.5 cd	13 ± 9.7 a	46.9 ± 7.7 d	47.7 ± 6.8 de	28.8 ± 7.1 a	37.4 ± 6.5 cd	38.0 ± 5.8 cd	23.2 ± 5.7 a
	HNT	81	73	8	37.9 ± 17.5 bc	40.8 ± 15.9 bc	11.8 ± 5.3 a	43.3 ± 7.1 cd	44.6 ± 6.1 cd	31.1 ± 2.7 ab	34.6 ± 5.8 bc	35.7 ± 5 bc	24.4 ± 2 ab
	HNK2	243	214	29	31.7 ± 15.4 ab	34.5 ± 14.1 ab	10.5 ± 3.6 a	39 ± 7.9 bc	40.7 ± 6.7 bc	27.3 ± 5.7 a	31.5 ± 5.9 b	32.8 ± 4.8 b	22.1 ± 4.5 a
	HNL	51	43	8	47.6 ± 29.4 cde	54.8 ± 26.2 de	8.7 ± 2.3 a	44 ± 12.6 cd	47.6 ± 10.2 de	24.7 ± 2.3 a	35.7 ± 10.2 cd	38.5 ± 8.5 cd	20.4 ± 1.5 a
	HS	92	92	0	51.3 ± 24.5 e	51.3 ± 24.5 cd	0 N/A	47.3 ± 8.6 d	47.3 ± 8.6 d	0 N/A	38.7 ± 7.4 d	38.7 ± 7.4 cd	0 N/A
	HPB	152	136	16	31.6 ± 20.4 ab	33.3 ± 20.7 ab	17.5 ± 10.6 ab	36.6 ± 10 ab	38.1 ± 9.2 ab	24.4 ± 9.5 a	30.9 ± 8.5 b	32.1 ± 8 ab	21 ± 6.9 a
PR	HTi	137	105	32	21.4 ± 10.5 a	24.8 ± 9.4 a	10.4 ± 5.5 a	33 ± 7.4 a	35.4 ± 5.8 a	24.9 ± 6.5 a	26.8 ± 5.2 a	28.5 ± 3.9 a	21.1 ± 4.9 a
	HK	55	35	20	49.9 ± 27.9 de	65.2 ± 22.9 e	22.9 ± 8.5 b	47 ± 10.6 d	52.1 ± 8.6 e	38 ± 7.5 b	38.4 ± 8.6 cd	42.8 ± 6.8 e	30.8 ± 5.8 b
	HSK	72	59	13	44.7 ± 26 cde	52.3 ± 22.3 d	10 a	44.8 ± 12.8 d	49.1 ± 9.7 de	25.1 ± 2.8 a	36.5 ± 11.9 cd	40.1 ± 10 de	20.2 ± 1.6 a
	HP	66	46	20	40.3 ± 26 bcd	51.3 ± 23.9 cd	15 ± 5.1 ab	42.7 ± 10.5 cd	47.8 ± 8.2 de	31 ± 2.9 ab	34.6 ± 8.4 bc	38.6 ± 6.9 cd	25.5 ± 2.4 ab
	HTo	45	41	4	49.7 ± 29 de	53.6 ± 27.5 d	10 a	46.8 ± 11.9 d	48.4 ± 11.3 de	30.7 ± 1.4 ab	37.9 ± 9.8 cd	39.3 ± 9.2 cde	24.1 ± 1.1 ab
	HNK1	61	54	7	50.8 ± 19 de	55.1 ± 15.4 de	17.1 ± 4.8 ab	45.8 ± 9.7 d	48.2 ± 7.3 de	27.5 ± 4.7 a	39.1 ± 7.2 d	40.9 ± 5.4 de	25.8 ± 5.1 ab
		df			1179	1017	161	1179	1017	161	1179	1017	161
		F			24.8	30.1	8.3	28.3	34.8	8.2	27.5	33.5	8.2
		p			<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **

PA = Protected Area; Ad = Adult; Ju = Juvenile; HTi = Huay Tiew; HK = Huay Kawak; HSK = Huay Sai Khao; HP = Huay Phai; HTo = Huay Toei; HNK1 = Huay Nam Khan; HNS = Huay Nam San; HNT = Huay Nam Thop; HNK2 = Huay Nam Khai; HNL = Huay Nam Loei; HS = Huay Sak; HPB = Huay Pla Ba; ** significant difference (p ≤ 0.001); data with different letters in the same column indicate a significant difference (p ≤ 0.05) according to the Duncan test performed; ^N/A = not analyzed.

). Overall, crabs in PL were larger on average than those in PR (Figure 3). The average body weight, carapace width and length of male crabs (n = 649 individuals) were 40.4 ± 24.7 g, 42.1 ± 11.2 mm and 34.13 ± 9.01 mm, respectively. The average body weight, carapace width and length of female crabs (n = 531 individuals) were 36.9 ± 20.4 g, 41 ± 9.5 mm and 33.7 ± 7.9 mm, respectively.

The analysis of variance showed that the body size differed significantly between male and female crabs in the 12 sampling sites at the p = 0.01 level (

Table 4. Body size of male of I. bhumibol in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR).

PA	Sub-Sites	N			Mean ± Sd
					Weight (g)			Carapace (mm)
								Width			Length
		Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju
PL	HNS	78	74	4	50.8 ± 22 cde	52.8 ± 20.7 cd	13.7 ± 11 ab	47.1 ± 8.5 cd	48.1 ± 7.3 cde	28.2 ± 8.1 ab	37.4 ± 7.1 cde	38.2 ± 6.2 cde	22.8 ± 6.5 ab
	HNT	67	60	7	39.1 ± 18.2 bcd	42.3 ± 16.5 bc	12.1 ± 5.6 ab	43.8 ± 7.3 cd	45.2 ± 6.3 cd	31.5 ± 2.6 ab	35 ± 6.0 bcde	36.2 ± 5.1 bcd	24.8 ± 1.9 ab
	HNK2	111	93	18	34.5 ± 17.9 ab	39.1 ± 15.9 abc	11.1 ± 4.3 ab	40.2 ± 9.3 bc	42.6 ± 7.6 bc	27.7 ± 6.7 ab	32.1 ± 6.9 abc	34 ± 5.5 abc	22.5 ± 5.3 ab
	HNL	26	21	5	55.9 ± 33.8 e	67.1 ± 27.4 de	9 ± 2.2 a	46.8 ± 14.2 cd	52 ± 10.2 de	24.9 ± 2.1 a	38.1 ± 10.9 de	42.2 ± 7.5 ef	20.8 ± 1.4 a
	HS	55	55	0	54.5 ± 27.3 de	54.5 ± 27.3 cd	0 N/A	48.2 ± 9.8 d	48.2 ± 9.8 cde	0 N/A	39.1 ± 7.4 e	39.1 ± 7.4 cde	0 N/A
	HPB	75	65	10	32.3 ± 22.4 ab	34.7 ± 22.7 ab	16.5 ± 10.8 ab	36.2 ± 11.5 ab	38.3 ± 10.2 ab	22.6 ± 10.5 a	29.7 ± 8.9 ab	31.3 ± 8.1 ab	19.6 ± 7.5 a
PR	HTi	73	54	19	21.5 ± 10.5 a	25.7 ± 8.3 a	9.7 ± 5.8 ab	32.8 ± 8.1 a	35.7 ± 6.7 a	24.4 ± 5.4 a	26.5 ± 5.6 a	28.9 ± 3.7 a	19.9 ± 5 a
	HK	33	19	14	50.8 ± 32.2 cde	72.6 ± 24.9 e	21.3 ± 8.4 b	46.8 ± 11.5 cd	54.6 ± 8.7 e	36.2 ± 3.2 b	38.3 ± 9.5 e	44.9 ± 6.9 f	29.5 ± 3.1 b
	HSK	34	26	8	43.2 ± 28.2 bcde	53.5 ± 24.3 cd	10 a b	44.1 ± 14.4 cd	50.1 ± 10.8 de	24.9 ± 3.2 a	36.6 ± 14.4 cde	41.5 ± 12.8 def	20.6 ± 1.7 a
	HP	39	24	15	35.3 ± 23.6 abc	47.5 ± 22.5 bc	16 ± 5 ab	40.3 ± 9.1 bc	45.5 ± 7.7 cd	31.8 ± 2.4 ab	32.4 ± 7.2 bcd	36.4 ± 6.2 bcd	25.9 ± 2.2 ab
	HTo	22	19	3	45.4 ± 32.9 bcde	51 ± 31.9 bcd	10 ab	44.6 ± 13.3 cd	46.9 ± 13 cd	30.4 ± 1.6 ab	35.5 ± 10.6 bcde	37.3 ± 10.3 cde	24 ± 1.3 ab
	HNK1	36	33	3	50.9 ± 18 cde	54.4 ± 14.6 cd	13.3 ± 5.7 ab	46.7 ± 10 cd	48.7 ± 7.4 cde	23.8 ± 5.7 a	39.7 ± 7.4 e	41.1 ± 5.6 def	23.5 ± 5.5 ab
		df			648	542	105	648	542	105	648	542	105
		F			12.4	14.9	4	14.1	16.5	6.3	14.9	17.9	5.6
		p			<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **

PA = Protected Area; HTi = Huay Tiew; HK = Huay Kawak; HSK = Huay Sai Khao; HP = Huay Phai; HTo = Huay Toei; HNK1 = Huay Nam Khan; HNS = Huay Nam San; HNT = Huay Nam Thop; HNK2 = Huay Nam Khai; HNL = Huay Nam Loei; HS = Huay Sak; HPB = Huay Pla Ba; Juv = Juvenile; ** significant difference (p ≤ 0.01); data with different letter in the same column indicate a significant difference (p ≤ 0.05) according to the Duncan test performed; ^N/A = not analyzed.

and

Table 5. Body size of female of I. bhumibol in Phu Luang Wildlife Sanctuary (PL) and Phu Ruea National Park (PR).

PA	Sub-Sites	N			Mean ± Sd
					Weight (g)			Carapace (mm)
								Width			Length
		Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju	Total	Ad	Ju
PL	HNS	47	46	1	47 ± 14.8 cde	47.8 ± 13.9 de	10 N/A	46.7 ± 6.4 de	47 ± 6 cde	31.1 N/A	37.3 ± 5.4 cdef	37.6 ± 5.1 cde	24.8 N/A
	HNT	14	13	1	32.1 ± 12.5 abc	33.8 ± 11.2 abcd	10 N/A	40.9 ± 5.8 bcd	41.9 ± 4.7 bc	27.9 N/A	32.4 ± 4.6 abcd	33.2 ± 3.6 abc	21.9 N/A
	HNK2	132	121	11	29.2 ± 12.4 ab	31 ± 11.3 ab	9.5 ± 1.5 ab	38.2 ± 6.4 abc	39.3 ± 5.5 ab	26.7 ± 3.7 a	31 ± 4.9 ab	31.8 ± 4 ab	21.4 ± 2.9 a
	HNL	25	22	3	39 ± 21.3 bcd	43.1 ± 19.1 bcde	8.3 ± 2.8 a	41.1 ± 10.1 bcd	43.3 ± 8.4 bcd	24.5 ± 3.3 a	33.2 ± 9 bcde	35.1 ± 8 bcd	19.8 ± 1.6 a
	HS	37	37	0	46.4 ± 19.1 cde	46.4 ± 19.1 cde	0 N/A	46.1 ± 6.2 de	46 ± 6.2 cde	0 N/A	38 ± 7.4 def	38 ± 7.4 cde	0 N/A
	HPB	77	71	6	31 ± 18.4 ab	32 ± 18.6 abc	19.1 ± 11.1 bc	37 ± 8.7 ab	37.8 ± 8.3 ab	27.5 ± 7.4 a	32 ± 8.1 abc	32.8 ± 8 abc	23.4 ± 4.6 a
PR	HTi	64	51	13	21.4 ± 10.7 a	23.9 ± 10.4 a	11.5 ± 5.1 ab	33.1 ± 6.6 a	35 ± 4.7 a	25.7 ± 8.1 a	27 ± 4.7 a	28.1 ± 4.1 a	22.9 ± 4.5 a
	HK	22	16	6	48.6 ± 20.5 de	56.8 ± 17.4 e	26.6 ± 8.1 c	47.2 ± 9.5 de	49.2 ± 7.7 de	42 ± 12.6 b	38.6 ± 7.5 ef	40.4 ± 5.9 de	33.6 ± 9.4 a
	HSK	38	33	5	46 ± 24.2 cde	51.5 ± 21 e	10 ab	45.3 ± 11.4 de	48.3 ± 8.9 de	25.4 ± 2.3 a	36.5 ± 9.3 bcdef	39.1 ± 7 de	19.6 ± 1.4 a
	HP	27	22	5	47.4 ± 28.5 de	55.4 ± 25.2 e	12 ± 4.4 ab	46.2 ± 11.5 de	50.2 ± 8.3 e	28.3 ± 2.8 a	37.8 ± 9.2 cdef	40.9 ± 7 e	24 ± 2.9 a
	HTo	23	22	1	53.9 ± 24.9 e	55.9 ± 23.6 e	10 N/A	48.8 ± 10.3 e	49.6 ± 9.8 e	31.6 N/A	40.2 ± 8.6 f	41 ± 8.1 e	24.4 N/A
	HNK1	25	11	4	50.6 ± 20.5 de	56.4 ± 16.8 e	20 bc	44.6 ± 9.2 cde	47.3 ± 7.3 cde	30.2 ± 0.2 ab	38.4 ± 7 ef	40.5 ± 5.2 de	27.5 ± 4.9 ab
		df			530	474	55	530	474	55	530	474	55
		F			15.6	19.2	5.8	17.1	22	2.9	15.6	19.4	3.7
		p			<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **	<0.001 **

PA = Protected Area; Ad = Adult; Ju = Juvenile; HTi = Huay Tiew; HK = Huay Kawak; HSK = Huay Sai Khao; HP = Huay Phai; HTo = Huay Toei; HNK1 = Huay Nam Khan; HNS = Huay Nam San; HNT = Huay Nam Thop; HNK2 = Huay Nam Khai; HNL = Huay Nam Loei; HS = Huay Sak; HPB = Huay Pla Ba; ** significant difference (p ≤ 0.001); data with different letter in the same column indicate a significant difference (p ≤ 0.05) according to the Duncan test performed; ^N/A = Not analyzed.

). The body weight of female crabs was significantly different between the two protected areas (W; t = 2.59, df = 529, p = 0.01), whereas the body weight and carapace width of male crabs were significantly different between the two protected areas (W; t = 2.1, df = 647, p = 0.03, CW; t = 2.16, df = 647, p = 0.03). The female crabs in PL had a higher average body weight, carapace width and length than those in PR (Figure 4). In contrast, male crabs in PR had a higher average body weight, carapace width and length than those in PL. The size distribution based on carapace width showed that most of the crabs caught in both populations were in the 30–50 mm size range (Figure 5a). The largest individual caught in this study weas male (n = 4), with a mass of 145 ± 5.8 g, carapace width of 76 ± 2.4 mm and carapace length of 75.7 ± 14.2 mm.

The sex ratio of all samples in both populations during February 2018 to January 2019 was 1:0.6 (male: female). The sex ratio showed monthly variation, with more males than females throughout the year (Table 2). However, the number of male and female crabs did not differ statistically between months in both PR (t = 0.66, df = 22, p = 0.514) and PL (t = 0.54, df = 22, p = 0.596), indicating that there was no seasonality in the proportion of individuals of the different sexes. During a field survey in July 2018, we observed I. bhumibol copulating on the river bank at PL.

3.3. Population Size and Density

The population size of I. bhumibol in Loei Province tended to be associated with the size of protected areas; the crab population was larger in PL, which has a larger area than PR (Figure 5b). The average population size of I. bhumibol was higher in PL than in PR (PL was 263.5 ± 223.9 individuals, and PR was 121.2 ± 79.4 individuals), but no statistically significant difference was observed between all sampling sites (F = 0.78, df = 21, p = 0.651). In this study, the average population size of I. bhumibol was highest in Huay Nam Khai (473.5 ± 478.7 individuals), a sampling site in PL, and lowest in Huay Kawak (73.7 ± 14.6 individuals), a sampling site in PR. The average population size was higher in the rainy season (199 ± 210.1 individuals) than in the dry season (178.9 ± 134.5 individuals), but this difference was not statistically significant (t = −0.28, df = 22, p = 0.782).

Average crab density in PL and PR was 2.3 ± 1.1 crabs/m² and 1.8 ± 0.6 crabs/m², respectively. Generally, sampling sites in PL had a higher population density than those in PR (Figure 6a). The population density of crabs was quite similar throughout the year in both protected areas. The analysis of variance showed that density did not differ significantly between the 12 sampling sites (F = 0.92, df = 23, p = 0.549). The highest population density was found at Huay Nam Khai, a sampling site in PL (2.7 ± 1.9 individuals/m²), and the lowest density was at Huay Nam Loei in PL (1.2 ± 0.1 individuals/m²) (Figure 6b). The average population density of crabs in the rainy season (1.7 ± 0.7 individuals/m²) was higher than that in the dry season (1.5 ± 0.4 individuals/m²) (Figure 6c), but this difference was not significant (t = 0.84, df = 22, p = 0.41). The average population density of male and female crabs in PL was 1.7 ± 0.6 and 1.5 ± 1 individuals/m², respectively (Figure 7a), and in PR, it was 1.4 ± 0.4 individuals/m² and 1.4 ± 0.3 individuals/m², respectively (Figure 7b). However, there was no significant difference in density between the sexes (male; t = −1.39, df = 22, p = 0.177 and female; t = −0.46, df = 22, p = 0.649).

3.4. Relationship between Population Density and Body Size

The relationship between the population density and body size of crabs showed a negative trend in both PL and PR (Figure 8). There were significant correlations between population density and body weight (r = −0.250, df = 1180, p < 0.001), carapace width (r = −0.221, df = 1180, p = 0.01) and carapace length (r = −0.25, df = 1180, p = 0.01). The regression analysis showed a significant relationship between the population density and body size of crabs, including weight (r² = 0.063, df = 1179, p = 0.01), carapace width (r² = 0.049, df = 11179, p = 0.01) and carapace length (r² = 0.063, df = 1179, p = 0.01).

4. Discussion

4.1. Abundance and Body Size

According to Suthamrit et al. [5], I. bhumibol were found in the same areas with Iomon nan and Vietopotamon phuluangnense (Potamidae), such as Na Haew Distric, Nong Hin District, Phu Luang District, Wang Saphung District and Phu Kradueng Distric, Loei, Province. However, they were never captured at the same site due to I. bhumibol occurring at high elevation, while Iomon nan and Vietopotamon phuluangnense were found at lower elevation as freshwater crabs from the highland of Zimbabwe that coexistence in a different flowing environment of water [21]. The abundance of crabs was higher in PL than PR. The number of crabs caught was positively correlated with the size of the protected area, water level, water pH and the distance between the protected areas and urban settlements. The general landscape of Loei Province, where the two protected areas are located, includes the high mountains of the Phetchabun range. Therefore, the soil texture, forest type, ambient temperature and humidity of PR and PL were similar throughout the year. The amount of water, water pH and organic matter in the soil may be related to body size and an abundance of crabs [22]. The larger protected area (PL) may be better able to support giant mountain crabs by providing large amounts of food resources necessary for growth. A previous study found that the number of freshwater crabs was positively correlated with the water level in a natural protected area [13], and water pH < 6 affected the total length and body weight of juveniles, and at pH 4, the postlarvae did not metamorphose [23]. In this study, we found that the average water level at sampling sites in PL (0.8 ± 0.8 m) was higher than the level at sites in PR (0.5 ± 0.3 m). Moreover, sampling sites in PL were located far from agricultural and urban landscapes; therefore, these sites were less disturbed. These factors might have affected the abundance of giant mountain crabs in Loei Province.

The abundance of I. bhumibol did not show seasonal variation. This finding is different from the results of another report on the abundance of freshwater crabs in Asia, which found the highest abundance in the rainy season because high rainfall removes nutrients from upper to lower streams and organic matter from sediments. Food resources are more abundant in the rainy season more than in the dry season [24].

According to the frequency distribution of carapace width, the natural population of this species consisted of more adults than juveniles. The higher number of adults throughout the year indicates that this species has low reproductive activity. This finding differs from the result of an investigation of freshwater crabs in Malaysia, where potamid crabs showed high reproductive activity as the frequency of younger crabs was high [15]. However, observation bias, unaccounted physical factors and crab behavior may explain the differences found between adults and juveniles [25].

The sex ratio differed from the expected (1:1) in both populations, but it was not different between the two protected areas. This finding agrees with results from previous work on freshwater crabs, which found that the sex ratio was one female for every two males [24,26]. Our results showed that the habitat characteristics of PL and PR did not affect the proportion of male and female crabs. The absence of female crabs in March 2018 at PL was probably associated with a period of copulation or females moving to burrows [24].

Our results suggested that male and female crabs have different growth rates; males were larger in terms of weight, carapace width, and length than females. This finding agrees with those of previous studies: males are bigger on average than females [17,20]. A larger size may be advantageous to males in competing with other males for reproduction [15,26]. We found differences in the body size (weight, carapace width and length) of female crabs between the two protected areas. The optimal size of females seems to be conditioned by energy [27]. The differences in food resources and individual growth rates between the two protected areas may have led to the differences in body sizes of females.

4.2. Population Size and Density

The population size and density of I. bhumibol were higher in the large protected area. Population size was also higher in the rainy season but declined during the dry season. There were no seasonal patterns in population size and density [28]. From our experience, hunting for crabs is the primary factor affecting the giant mountain crab population size in Loei Province. During our field study, we found local villagers collecting giant mountain crabs for consumption and trade in both protected areas. Our field observations indicated that hunting in PR was usually more intense than in PL. Nowadays, the growth of highland agriculture in Loei Province may be leading to increasing habitat fragmentation nearby or outside the protected area. Previous work suggested that changes in land used was a factor that affected the habitat of endemic freshwater crab species [29].

4.3. Relationship between Population Density and Body Size

Body size was negatively correlated with population density. A smaller crab based on weight, carapace width and carapace length presented higher density than larger crabs [19]. Smaller crabs need a lower amount of food and a smaller home range than larger crabs.

5. Conclusions

This is the first report on the population parameters and individual body size of I. bhumibol in Loei Province. It should be highlighted that I. bhumibol requires a very specific habitat, such as waterfalls and streams, in mountain forests that have flowing water and an area located 400–1,000 m above sea level. Protected areas are refuges for giant mountain crabs. Crab abundance is related to the size of the protected area, while the body size of crabs is correlated with population density. Our finding provides some ecological information, including the population size and body size characteristics of I. bhumibol, which are useful baseline data for designing adapted measures and area management strategies for the conservation of this endangered species.

The population size of I. bhumibol in PL and PR seems to be large, with a strong cohort of reproductive and almost reproductive individuals, which is a positive criterion when assessing extinction risk. It was assessed as EN on the IUCN Red List due to threats from human-induced habitat loss/degradation and pollution, specifically agriculture and aquaculture, logging and wood harvest [6]. To reassess this species in a less threatened category, it would be necessary to address the status of these threats and demonstrate that they are no longer a problem.

A limitation of this study is that observations were made only in a protected area. To further our understanding of the ecology, population dynamics and behavior of this endemic species and to develop relevant conservation policies, additional research will need to include other potential habitats, as well as investigate distribution patterns, feeding habits, reproductive trends, growth rates and the factors threatening the giant mountain crab both inside and outside protected areas.