Comparison of Chiggers (Acari: Trombiculidae, Leeuwenhoekiidae) on Two Sibling Mouse Species, Apodemus draco and A. ilex (Rodentia: Muridae), in Southwest China

Simple Summary

Chigger mites (Acari) are common ectoparasites and the exclusive vector of scrub typhus. South China field mouse (Apodemus draco) and Lantsang field mouse (A. ilex) are two sibling rodent species. The investigation in southwest China (2001–2015) showed that chigger infestations on two mouse species were quite different, including different species composition, overall infestation, community parameters and dominant chigger species. There were 36 chigger species found on A. draco and 11 on A. ilex, and the overall mean intensity of chiggers on A. draco (MI = 4.26) was higher than that on A. ilex (MI = 3.91, p < 0.05). Dominant chiggers were unevenly distributed among different individuals of mouse hosts, and chigger infestation showed sex bias on different mouse sexes. The species abundance of the chigger community on A. draco was revealed as a log-normal distribution pattern.

Abstract

Chigger mites (Acari) are common ectoparasites on rodents, and they are the exclusive vector of scrub typhus. South China field mouse (Apodemus draco) and Lantsang field mouse (A. ilex) are two sibling rodent species. Based on field investigations in southwest China (2001–2015), this paper compared the infestation of these two mouse species with chiggers. Of 42 chigger species identified from two mouse species, 36 were found on A. draco, 11 on A. ilex and 5 common species on both mice. Jaccard similarity index (J = 0.12, J < 0.25) showed a very different species composition of chiggers on two mouse species, and some parameters of the chigger community were also different. The overall mean intensity of chiggers on A. draco (MI = 4.26) was higher than that on A. ilex (MI = 3.91, p < 0.05). The dominant chigger species on A. draco were Trombiculindus yunnanus, Leptotrombidium scutellare (a major vector species in China) and L. sinicum with a total constituent ratio C_r = 42.9% (106/247). Leptorombidium sinicum and L. scutellare independently occurred on A. draco with an association coefficient V = 0.09 (V ≈ 0). The dominant chigger species on A. ilex were L. rusticum, L. densipunctatum and L. gongshanense, with a total C_r = 58.14% (25/43). Leptorombidium rusticum and L. densipunctatum on A. ilex had a slight positive association (V = 0.49, 0.5 < V < 1). All dominant chigger species were unevenly distributed among different individuals of two mouse species. Chigger infestation showed sex bias on different sexes of two mouse species. The species abundance of the chigger community on A. draco was revealed as a log-normal distribution pattern.

1. Introduction

Chigger mites belong to the families Trombiculidae and Leeuwenhoekiidae in the subclass Acari [1,2]. It is estimated that about 3013 species of chigger mites have been recorded worldwide [3,4], and over 510 species have been reported in China [3,4,5]. The life cycle of chigger mites is complex, with seven basic stages: the egg, deutovum (prelarva), larva, nymphochrysalis, nymph, imagochrysalis and adult (male and female). The larva (chiggers) is the only ectoparasitic stage in rodents and other animal hosts [1,6,7]. In addition to directly biting humans to cause chigger dermatitis, chiggers are the exclusive vector of scrub typhus (tsutsugamushi disease), which is the most important medical significance of the mites. In addition, some chigger species, e.g., Leptotrombidium scutellare (Nagayo, Miyagawa, Mitamura, Tamiya and Tenjin, 1921), can be potential vectors of hemorrhagic fever with renal syndrome, HFRS [4,8,9]. With the spread of scrub typhus worldwide in recent years, the prevalence of the disease in China is also on the rise [10,11].

Rodents account for more than 40% of all mammals in the world, and they are widely distributed in different environments [12,13]. In addition to being harmful to agriculture and forestry, rodents can also carry a variety of pathogens of some zoonotic diseases such as scrub typhus, leptospirosis, tularemia, HFRS and plague [3,12,13]. South China field mouse (Apodemus draco Barrett-Hamilton, 1900) and Lantsang field mouse (A. ilex Thomas, 1922) belong to the genus Apodemus of the family Muridae in the order Rodentia, and they are two sibling species of mice with similar morphology [14,15,16]. These two sibling rodent species were once regarded as the same species, and A. ilex was once considered a subspecies of A. draco [16,17,18]. A series of previous studies on these two mouse species have involved their taxonomic status and morphological characteristics [14,15,19,20], relative fatness and seasonal changes in digestive tract length [21,22], eco-physiology [23], karyotype [24,25] and molecular differentiation [16,26], but there is little research literature on their ectoparasites including chiggers [18,20]. To date, there have been no specific reports on chiggers of these two sibling mouse species in southwest China.

The southwest China involved in the present paper covers five provincial regions, Yunnan, Guizhou, Sichuan, Chongqing and Tibet (Xizang Autonomous Region), with a vast territory, accounting for 24.5% of China’s land area [27]. Recent studies have shown that there are obvious differences in the geographical distribution of A. draco and A. ilex in southwest China. The former is mainly distributed east of the Jinsha River (roughly equivalent to the east of 101°50′ E), while the latter is mainly distributed west of the Lantsang River (roughly equivalent to the west of 100°45′ E) [16,17,28]. Based on previous field investigations in southwest China from 2001 to 2015, this paper retrospectively studied the species composition, infestation status and some ecological characteristics (community structure and species abundance distribution) of chiggers on these two sibling species of Apodemus mice for the first time. As an exploration of some unknown scientific issues, this study aims to compare the difference in chigger infestation on these two sibling mouse species, enrich the knowledge of these two mouse species and their ectoparasites, and provide more scientific information for subsequent related research.

2. Materials and Methods

2.1. Field Survey Sites

The original data came from the field investigations in 91 survey sites of five provincial regions in southwest China from 2001 to 2015 [29,30]. Among the 91 sites investigated, there are 8 sites where A. draco was captured, and 3 sites where A. ilex was captured, totaling 11 sites (Figure 1).

2.2. Collection and Identification of Chiggers and Their Animal Hosts

Rodents and other small mammals (animal hosts) were captured with mousetraps, and chiggers on their body surfaces were routinely collected and fixed. Each host was identified into species according to its morphological appearance (body size, shape and coat color), various measurements (body length, body weight, tail length, ear height, hind foot length, etc.) and other morphological features [31,32]. The collected chiggers were mounted with Hoyer’s medium and made into slide specimens. After dehydration, drying and transparency, each chigger was identified into species under an optical microscope (Olympus Corporation, Tokyo, Japan) [1,3,33,34,35]. Based on the identification results of chiggers and their animal hosts, A. draco and A. ilex, together with their chiggers, were chosen as the target of present study. The capture and use of animals were officially approved by the local wildlife affairs authority and the Animal Ethics Committee of Dali University, and the ethics approval number is DLDXLL2020-1104.

2.3. Infestation Statistics Analysis

The species and numbers of all chiggers on the body surface of each A. draco and A. ilex were counted, respectively. The constituent ratio (C_r), infestation prevalence (P_M), mean abundance (MA) and mean intensity (MI) were adopted to calculate the infestation of mice with chiggers [7,34,36].

$$C_r=\frac{N_i}{N}\times 100\%$$

(1)

$$P_M=\frac{H_i}{H}\times 100\%$$

(2)

$$MA=\frac{N_i}{H}$$

(3)

$$MI=\frac{N_i}{H_i}$$

(4)

In the above formulae, N_i = the number of a certain chigger species (species i) on a certain species of host, N = the total number of all the chigger species, H = the total number of hosts captured, H_i = the number of hosts infested with chiggers.

2.4. Basic Community Structure Statistics

Species richness (S), Shannon–Wiener diversity index (H), Simpson dominance index (D) and Pielou evenness index (E) were used for chigger community statistics. Jaccard similarity coefficient (J) was used to analyze the similarity of community species composition [36,37].

$$H=-\sum _{i=1}^S\left(\frac{N_i}{N}\right)\ln \left(\frac{N_i}{N}\right)$$

(5)

$$D=1-{\sum _{i=1}^S\left(\frac{N_i}{N}\right)}^2$$

(6)

$$E=\frac{H}{\mathrm{l}\mathrm{n}S}$$

(7)

$$J=\frac{M}{G+F-M}$$

(8)

In the above formulae, N = the total number of all the chigger species, N_i = the number of a certain chigger species (species i) on a certain species of host, S = species richness (the number of species), G = the number of chigger species in community A, F = the number of chigger species in community B, M = the number of common species existed in both community A and B. 0.00 < J < 0.25 means extremely dissimilar, 0.25 ≤ J < 0.50 means moderately dissimilar, 0.50 ≤ J < 0.75 means moderately similar, and 0.75 ≤ J < 1.00 means extremely similar.

2.5. Measurement of Spatial Distribution Patterns

The spatial distribution of dominant chigger species among different individuals of mouse hosts was determined by the indexes of diffusion (C), mean crowding (m*) and clumping index (I). The calculation formulae and judgment criteria are listed in

Table 1. Index formulae and judgment criteria for spatial distribution patterns.

Name	Formula	Aggregation Distribution	Random Distribution	Uniform Distribution
C	$C={\sigma }^2/m$	>1	=1	<1
m*	$m^{*}=m+{\sigma }^2/m-1$	>m	=m	<m
I	$I=n\times \frac{\left(\sum m^2-\sum m\right)}{\left[{\left(\sum m\right)}^2-\sum m\right]}$	>1	=1	<1

$\mathrm{A}\mathrm{n}\mathrm{n}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}:{\sigma }^2=$ variance, m = mean and n = the total number of host samples.

[38,39].

2.6. Measurement of Interspecific Association

The association coefficient (V) was used to analyze the interspecific relationship between any two dominant chigger species on A. draco and A. ilex, and Chi-square test was used to test the statistical significance of V [5,40].

$$V=\frac{ad-bc}{\sqrt{(a+b)(c+d)(a+c)(b+d)}}$$

(9)

In the above formula, V = association coefficient between any two chigger species, X and Y, on a certain host species; a = host individuals on which both chigger species X and Y concurrently appear; b = host individuals on which chigger species Y appears, but chigger species X does not appear; c = host individuals on which chigger species X appears, but chigger species Y does not appear; and d = host individuals on which neither chigger species X nor Y appears. When 0 < V ≤ 1 and p < 0.05, the interspecific relationship between chigger species X and Y is determined as positive association, and when −1 ≤ V < 0 and p < 0.05, negative association. When V = 0 or V ≈ 0, it can be considered that chigger species X and Y independently occur on the host.

2.7. Species Abundance Distribution

X-axis (indicating individuals of chiggers) was labeled with log intervals based on log₃M, and Y-axis (representing the number of chigger species) was marked with arithmetic scales. Based on following formulae, Preston’s lognormal model was used to fit the theoretical curve of species abundance distribution of chigger community with the calculation of fitting goodness, R² [34,41,42,43].

$$S\left(R\right)=S_0{e}^{{-\left[a\left(R-R_0\right)\right]}^2}(e=2.71828\dots )(\mathrm{Preston}’\mathrm{s}\mathrm{lognormal}\mathrm{model})$$

(10)

$$R^2=1-\sum _{R=0}^m{\left[S^{\prime }\left(R\right)-S\left(R\right)\right]}^2/\sum _{R=0}^m{\left[S^{\prime }\left(R\right)-\stackrel{-}{S}\left(R\right)\right]}^2$$

(11)

$$\stackrel{-}{S}\left(R\right)=\frac{1}{m}\sum _{R=0}^m{S}^{\prime }\left(R\right)$$

(12)

In the above formulas, S(R) = the theoretical number of chigger species at the R-th log interval, S₀ = the number of chigger species at the R₀ log interval, m = the number of log intervals, R₀ = the mode log interval, S’(R) = the actual chigger species at R-th log interval and $\stackrel{-}{S}\left(R\right)$ = the average chigger species for each log interval. The value of α was determined according to the best-fitting goodness, R².

3. Results

3.1. Species Composition and Community Structure of Chiggers on Two Mouse Species

Among the 91 sites investigated in the five provincial regions of southwest China, there are 8 sites where A. draco was captured, and 3 sites where A. ilex was captured, totaling 11 sites. The 11 survey sites are as follows: Dali (DL), Daocheng (DC), Fuyuan (FY), Gengma (GM), Gongshan (GS), Miyi (MY), Muli (ML), Weixi (WX), Yanyuan (YY), Yongde (YD) and Zhijin (ZJ) (Figure 1).

Among the 11 survey sites shown in Figure 1, 567 A. draco were captured in 8 sites, and 154 A. ilex were captured in 3 sites, with a total of 721 hosts (567 + 154). From the body surface of 721 hosts, 313 chiggers were collected. Of the 313 chiggers collected, 290 ones were identified as 42 species and 7 genera in 3 subfamilies under 2 families, and 23 mites were unidentified because of broken bodies, dirt-covered bodies, blurred structures or suspected new species. The 23 unidentified chiggers were not included in the statistical analysis of this study. Among the 42 chigger species identified, there are 36 species on A. draco, 11 species on A. ilex, and 5 common species on both Apodemus (

Table 2. Distribution of identified 42 chigger species on two mouse species (Apodemus draco and A. ilex) at 11 survey sites of southwest China (2001–2015).

Chigger Names	Distribution on Hosts		Distribution at Survey Sites	Chigger Names	Distribution on Hosts		Distribution at Survey Sites
	A. draco	A. ilex			A. draco	A. ilex
Family Trombiculidae [1,2]				Trombiculindus yunnanus Wang et Yu, 1965 [1]	+	+	YY, GS
Subfamily Trombiculinae [1,2]				T. cuneatus Traub et Evans, 1951 [1]	-	+	GS
Leptotrombidium scutellare (Nagayo et al., 1921) [1,2]	+	+	GS, WX	T. nujiange Wen et Xiang, 1984 [1]	-	+	GS
L. sinicum Yu et al., 1981 [1,2]	+	-	WX	T. bambusoides Wang et Yu, 1965 [1]	+	+	ZJ, MY, YY, GS
L. rusticum Yu et al., 1986 [1,2]	-	+	GS	Neotrombicula japonica (Tanaka et al., 1930) [1]	+	-	MY
L. wangi Yu et al., 1986 [1,2]	+	-	WX	N. tongtianhensis Yang et al., 1995 [1]	+	-	WX
L. densipunctatum Yu et al., 1982 [1,2]	+	+	GS, WX	N. aeretes Hsu et Yang, 1985 [1]	+	-	WX
L. robustisetum Yu et al., 1983 [1,2]	+	-	WX	Helenicula simena (Hsu et Chen, 1957) [1]	+	-	MY, YY, ML
L. laojunshanense Yu et al., 1986 [1,2]	+	-	WX	H. hsui Zhao, 1990 [1]	+	-	YY
L. baoshui Wen et Xiang, 1984 [1,2]	+	-	ML	Subfamily Gahrliepiinae [1]
L. kawamurai (Fukuzumi et Obata, 1953) [1,2]	+	-	YY
L. alpinum Yu et Yang, 1986 [1,2]	+	-	YY, WX	Walchia kor (Chen et Hsu, 1957) [1]	+	-	ML
L. gongshanense Yu et al., 1981 [1,2]	+	+	YY, GS	W. zangnanica Wu et Wen, 1984 [1]	+	-	MY
L. muntiaci Wen et Xiang, 1984 [1,2]	+	-	ML	W. tianguangshanensis Zhao et al., 1980 [1]	-	+	GM
L. jinmai Wen et Xiang, 1984 [1,2]	+	-	YY	W. ewingi (Fuller, 1949) [1]	-	+	GS
L. eothenomydis Yu et Yang, 1986 [1,2]	+	-	ZJ, YY, WX	Gahrliepia myriosetosa Wang, 1964 [1]	+	-	ML
L. nyctali Wen et Sun, 1984 [1,2]	+	-	YY	G. linguipelta Jeu et al., 1983 [1]	+	-	YY, WX
L. suense Wen, 1984 [1,2]	+	-	ML	G. longipedalis Yu et Yang, 1986 [1]	+	-	WX
L. neotebraci Xiang et Wen, 1986 [1,2]	+	-	ML	G. radiopunctata Hsu et al., 1965 [1]	-	+	GM
L. rupestre Traub et Nadchatram, 1967 [1,2]	+	-	ML	G. megascuta Hsu et al., 1965 [1]	+	-	WX
L. yongshengense Yu et Yang, 1986 [1,2]	+	-	MY, ML	Family Leeuwenhoekiidae [1]
L. longimedium Wen et Xiang, 1984 [1,2]	+	-	ML	Subfamily Leeuwenhoekiinae [1]
L. sinotupaium Wen et Xiang, 1984 [1,2]	+	-	ML	Chatia alpine Shao et Wen, 1984 [1]	+	-	YY
L. bambicola Wen et Xiang, 1984 [1,2]	+	-	ML	C. acrichela Wen et al., 1984 [1]	+	-	YY

Annotation: “+”= The host surface contains this chigger mite. “-”= The host body surface does not contain this chigger mite.

). Jaccard similarity index (J) shows that the species composition of chiggers on two mouse species of Apodemus is very different, with J = 0.12 (J < 0.25, extremely dissimilar). There are some differences in the community parameters of chiggers on two mouse species. The species richness (S = 36) and Shannon–Wiener diversity index (H = 2.89) of chigger community on A. draco are higher than those on A. ilex (S = 11, H = 2.10), but Pielou evenness index and Simpson dominance index (E = 0.81, D = 0.09) on A. draco are lower than those on A. ilex (E = 0.88, D = 0.15).

3.2. Overall Infestation and Dominant Species of Chiggers on Two Mouse Species

There were consistent differences in the overall infestation of chiggers in the two mouse species. Of all A. draco mouse hosts, 58 mice of them were infested with chiggers with 10.23% of overall infestation (P_M = 10.23%, 58/567), 0.44 mites/per mouse of mean abundance (MA = 0.44) and 4.26 mites/per mouse of mean intensity (MI = 4.26), which were higher than the corresponding indices on A. ilex (P_M = 7.14%, MA = 0.28 mites/mouse, MI = 3.91 mites/mouse). The difference in overall mean intensity (MI) of chiggers on two mouse species was statistically significant (p < 0.05), but there was no statistical significance in the differences in overall prevalence (P_M) and overall mean abundance (MA) of the mites on the mice (p > 0.05). The dominant chigger species on the two mouse species were also different. The dominant chigger species on A. draco are Trombiculindus yunnanus Wang and Yu, 1965, L. scutellare and L. sinicum Yu, Yang and Gong, 1981 (total C_r = 42.9%, 106/247). The dominant chigger species on A. ilex, however, are L. rusticum Yu, Yang and Gong, 1986, L. densipunctatum Yu, Yang and Gong, 1982 and L. gongshanense Yu, Yang and Gong, 1981 (total C_r = 58.14%, 25/43) (

Table 3. Dominant chigger species on two species of mice (Apodemus draco and A. ilex) in southwest of China (2001–2015).

Names of Mouse Hosts	Names of Dominant Chigger Species	Individuals and Constituent Ratios (Cr) of Chiggers		Overall Infestations of Chiggers on the Hosts
		Individuals	Cr (%)	PM (%)	MA	MI
A. draco (n = 567)	T. yunnanus	50	20.24	0.35	0.09	25.00
	L. scutellare	34	13.77	3.02	0.06	2.00
	L. sinicum	22	8.91	0.89	0.04	4.40
A. ilex (n = 154)	L. rusticum	12	27.91	2.60	0.08	3.00
	L. densipunctatum	8	18.60	2.60	0.05	2.00
	L. gongshanense	5	11.63	1.30	0.03	2.50

). The diagnostic characteristics of the six dominant chigger species on two mouse species are listed in

Table 4. The diagnostic characteristics of the six dominant chigger species on two mouse species, Apodemus draco and A. ilex.

Names of Mouse Hosts	Names of Dominant Chigger Species	Diagnostic Characteristics
A. draco	T. yunnanus	fPp = N/N/BNN; PC = 3; Gn = 2; Sc: PL > AM> AL; PL/SB; fCx = 1.1.1; fSt = 2.2; fD = 2H + 8-6-6-6-2-2; DS = 32; VS = 35; NDV = 67; Ip = 1059; AW 88, PW 99, SB 41, ASB 32–33, PSB 23, SD 55, AP 23, AM 55, AL 42, PL 65 S 75, H-, Dmin-, Dmax-, Vmin-, Vmax-, pa 353, pm 323, pp 383.
	L. scutellare	fPp = N/N/BNN; PC = 3; Gn = 2; Sc: PL > AM> AL; SB-PL; fCx = 1.1.1; fSt = 2.2; fD = 2H-10-[10-2]-[12-4]-8-6-2; DS = 56; VS = 38; NDV = 94; Ip = 858; AW 71–72, PW 79–82, SB 30–32, ASB 29–32, PSB 15–17, SD 44 49, AP 28–29, AM 57–59, AL 50–51, PL 62–64, S 78–86, H 62, Dmin 44, Dmax 57–69, Vmin 34, Vmax 50, pa 295, pm 256, pp 307.
	L. sinicum	fPp = N/N/BNN; Pc = 3; Gn = 2; fSc: PL > AM > AL; PL/SB; fCx = 1.1.1; fSt = 2.2; fD = 2H-9-8-8-8-2-3; DS = 38 41; VS = 36–39; NDV = 78; Ip = 824–893; AW 60–67, PW 73–76, SB 25–30, ASB 35–37, PSB 14–15, SD 49–52, AP 30–34, AM 60–67, AL 51–60, PL 70–77, S 73–79, H 68–74, Dmin 50–55, Dmax 68–72, Vmin 35–42, Vmax 45–56, pa-, pm-, pp-.
A. ilex	L. rusticum	fPp = N/N/BNN; Pc = 3; Gn = 2; fSc: PL > AM >> AL; PL/SB; fCx = 1.1.1; St = 2.2; D = 2H-8-6-6-4-2; DS = 30; VS = 26–30; NDV = 56; Ip = 685–725; AW 64–70, PW 71–80, SB 31–37, ASB 24–28, PSB 13–16, SD 40, AP 18–22, AM 41 = 55, AL 30–38, PL 53–65, S 55–70, H 48–58, Dmin 38–48, Dmax 51–61, Vmin 20–30, Vmax 43–54, pa 236, pm 217, pp 246.
	L. densipunctatum	fPp = N/N/BNN; Pc = 3; Gn = 2; fSc: PL > AM > AL; SB-PL; fCx = 1.1.1; (St = 2.2; fD = 2H1-8-6-6-4-2-2; DS = 28–30; VS = 27–35; NDV = 60; Ip = 776; AW 68–79, PW 80–88, SB 33–38, ASB 29–33, PSB 13–15, SD 41–48, AP 25–31, AM 58–71, AL 43–50, PL 65–74, S 80–87, H 60–73, Dmin 45–55, Dmax 50–65, Vmin 31–35, Vmax 45–57, pa 243–293, pm 220–293, pp 257–304.
	L. gongshanense	fPp = N/N/BNN; Pc = 3; Gn = 2; fSc: AM > PL > AL; PL/SB; fCx = 1.1.1; fSt = 2.2; fD = 2H-8-6-6-4-2; DS = 28–30; VS = 22–25; NDV = 53; Ip = 675–710; AW 65–70, PW 73–78, SB 30–33, ASB 25–27, PSB 12–14, SD 37–40, AP 22–23, AM 64–76, AL 38–42, PL 60–66, S 78–84, H 54–61, Dmin 42–47, Dmax 52–57, Vmin 37–40, Vmax 42–47, pa-, pm-, pp-.

, and the corresponding abbreviations refer to the relevant taxonomic literature [1,2]. The photos of the representative dominant mite species are shown in Figure 2, Figure 3, Figure 4 and Figure 5.

The calculation of the Jaccard similarity index showed that the species composition of chiggers on different sexes (males and females) of two mouse species was quite different (moderately dissimilar), J < 0.5 (A. draco: J = 0.27, A. ilex: J = 0.45). The infestation indexes (P_M, MA, MI) of chiggers were also different between the two sexes of the mice, but the differences are of no statistical significance (

Table 5. Infestation differences of chiggers on two sexes of hosts, Apodemus draco and A. ilex.

Host Names	Host Sexes	Number of Hosts	Species, Individuals and Constituent Ratios (Cr) of Chiggers			Overall Infestations of Chiggers on the Hosts
			Species	Individuals	Cr (%)	PM (%)	MA	MI
A. draco	Female	184	17	57	23.85	10.33	0.31	3.00
	Male	378	25	182	76.15	9.79	0.48	4.92
	Total	562	-	239	100.00	20.12	0.79	7.92
A. ilex	Female	86	10	32	74.42	8.14	0.37	4.57
	Male	68	6	11	25.58	5.88	0.16	2.75
	Total	154	-	43	100.00	14.02	0.53	7.32

Annotation: There were five individuals (n = 5) of A. draco without sex records, and these five mice were not included in the calculation of the above table.

).

The association coefficient (V) between L. sinicum and L. scutellare on A. draco was close to 0 (V = 0.09, V ≈ 0), and that between L. rusticum and L. densipunctatum on A. ilex was close to 0.5 (V = 0.49) (

Table 6. Interspecific association coefficient (V) between two dominant chigger species, Leptotrombidium sinicum and L. scutellare, on Apodemus draco in southwest China (2001–2015).

Dominant Chigger Species		L. sinicum (Species X)
		+	-	Total
L. scutellare (species Y)	+	1 (a)	16 (b)	17 (a + b)
	-	4 (c)	548 (d)	552 (c + d)
Total		5 (a + c)	564 (b + d)	569 (n)
Association coefficient		V = 0.09
Chi-square		χ2 = 5.037
Significance		p < 0.05

Annotation: “+” = The host surface contains this chigger mite. “-” = The host body surface does not contain this chigger mite.

and

Table 7. Interspecific association coefficient (V) between two dominant chigger species, Leptotrombidium rusticum and L. densipunctatum, on Apodemus ilex in southwest China (2001–2015).

Dominant Chigger Species		L. rusticum (Species X)
		+	-	Total
L. densipunctatum (species Y)	+	2 (a)	2 (b)	4 (a + b)
	-	2(c)	152 (d)	154 (c + d)
Total		4(a + c)	154 (b + d)	158 (n)
Association coefficient		V = 0.49
Chi-square		χ2 = 37.475
Significance		p < 0.05

Annotation: “+” = The host surface contains this chigger mite. “-” = The host body surface does not contain this chigger mite.

).

All the calculated indexes of dominant chigger species on two mouse species for spatial distribution patterns were higher than the border values (C > 1, m* > m, I > 1) of determining aggregated distribution (Table 1 and

Table 8. Spatial distribution indexes of dominant chigger species on two mouse species (Apodemus draco and A. ilex) in southwest China (2001–2015).

Names of Mouse Hosts	Names of Dominant Chigger Species	C	I	m*
A. draco	T. yunnanus	26.40	25.40	25.49
	L. scutellare	4.18	3.18	3.24
	L. sinicum	14.90	13.90	13.94
A. ilex	L. rusticum	4.95	3.95	4.03
	L. densipunctatum	2.71	1.72	1.77
	L. gongshanense	3.38	2.39	2.42

).

3.3. Species Abundance Distribution of Chigger Community

Of the 36 species and 247 identified chiggers on A. draco, the number of chigger individuals at Log interval 4 was the highest, but the number of species was minimal. At Log interval 0 (R₀ = 0), there was only one individual chigger, but the number of chigger species was the largest (S₀ = 13). The species abundance distribution of the chigger community on A. draco was successfully fitted by Preston’s lognormal model with α = 0.36 and R² = 0.86. The theoretical curve equation was (R) = $13e^{{-\left[0.36\left(R-0\right)\right]}^2}$ (S₀ = 13, R₀ = 0) (

Table 9. Fitting results of species abundance distribution of chigger community on Apodemus draco in southwest China (2001–2015).

Log Intervals Based on log3M	Individual Ranges of Chiggers in Each Log Interval	Midpoint Values of Each Individual Range	A. draco
			Actual Chigger Species	Theoretical Chigger Species
0	0–1	1	13	13
1	2–4	3	9	11.42
2	5–13	9	9	7.74
3	14–40	27	4	4.04
4	41–121	81	1	1.63

, Figure 6).

4. Discussion

Previously A. draco and A. ilex were regarded as the same species, and A. ilex was once considered a subspecies of A. draco [18,44,45]. Recent studies have proved that A. draco and A. ilex are two independent species of rodents, and they are two sibling species. Although A. draco and A. ilex are quite similar in morphology, they still have some differences in morphology, molecular characteristics and geographical distribution areas [16,17,28]. Apodemus draco is mainly distributed in countries and regions such as China, north Myanmar and northeast India. In China, A. draco is mainly distributed in Yunnan, Tibet and Fujian provincial regions. The distribution range of A. ilex is narrow and mainly distributed south of the Yangtze River in south China and west of the Lantsang River in southwest China [16,17,28]. The present study revealed that A. draco and A. ilex were distributed in southwest China, but they were not the dominant rodent species in this area (only 567 A. draco and 154 A. ilex were captured). The distribution range of two mouse species in southwest China was obviously different. Apodemus draco was mainly captured east of the Jinsha River, and A. ilex was mainly captured west of the Lantsang River (Figure 1). This result is consistent with some previous research reports [16,28].

The results of this study showed that A. draco and A. ilex were not only obviously different in distribution areas but also different in chigger infestations, including species composition, community structure, infestation status and dominant chigger species. The Jaccard similarity index (J) used in this study is an index reflecting the similarity of species composition of any two communities. When 0.00 < J < 0.25, the species composition of two communities is extremely dissimilar, 0.25 ≤ J < 0.50 means moderately dissimilar, 0.50 ≤ J < 0.75 means moderately similar, and 0.75 ≤ J < 1.00 means extremely similar [46]. The results reveal that the species composition of the chigger community on two sibling species of Apodemus are very different with J = 0.12 (J < 0.25), and the community parameters (S, H, E and D) are also different. The overall infestation prevalence (P_M), mean abundance (MA) and mean intensity (MI) of chiggers on A. draco are higher than those on A. ilex. The dominant chigger species on A. draco are T. yunnanus, L. scutellare and L. sinicum, which are obviously different from the dominant mite species on A. ilex (L. rusticum, L. densipunctatum and L. gongshanense) (Table 3). Previous studies have shown that different small mammal species have different susceptibilities to the infestation of ectoparasites, including chiggers, which leads to differences in species composition, infestation status and dominant parasite species on different species of animal hosts [1,47]. Chevrier’s field mouse (A. chevrieri Miline-Edwards, 1868) is a mouse species in the same genus (Apodemus) as A. draco and A. ilex, and it is one of the dominant rodent species in southwest China [30]. A special study on chiggers of A. chevrieri in southwest China showed that its overall infestation prevalence (P_M = 31.95%), mean abundance (MA = 6.32 mites/mouse) and mean intensity (MI = 19.77 mites/mouse) with chiggers were significantly higher than the corresponding infestation indexes on the two mouse species of Apodemus in this study. The dominant chigger species on A. chevrieri are L. scutellare, L. densipunctatum and L. cricethrionis Wen, Sun and Sun, 1984, which are also obviously different from A. draco and A. ilex in this study [30]. The above differences reflect the different susceptibility of different mouse host species to chigger infestation. In this study, there are a series of differences in species composition, infestation status and dominant species composition of chiggers between two mouse species of Apodemus, which further verify the different susceptibility of different hosts to chiggers and the different preference of chiggers to different hosts. From the aspect of ectoparasites, the differences in species composition, infestation status and dominant species composition of chiggers between two sibling mouse species also support that A. draco and A. ilex belong to two independent species [16,28]. As one of the dominant chigger species of A. draco, L. scutellare is not only the second major vector of scrub typhus in China but also a potential vector of hemorrhagic fever with renal syndrome [9,48]. The occurrence of L. scutellare on the body surface of A. draco may increase the potential risk of spreading the pathogen of scrub typhus, Orientia tsutsugamushi, from rodents to humans.

The calculation results of the Jaccard similarity index (J) showed that the species composition of chiggers on different sexes of two mouse species, A. draco and A. ilex, were quite different (J < 0.5, moderately dissimilar). Besides, the infestation indexes (P_M, MA, MI) of chigger on different sexes of two mouse species were also different (Table 5). These results indicate that there is sex bias in chigger infestation between male and female hosts [29,36]. Sex bias is prevalent in parasite infections (including ectoparasite infestations), and many parasites are more likely to choose to parasitize in male hosts [4,49,50], but the preference of chiggers to male hosts is not obvious in this paper (Table 5), which may need further studies.

In this study, the interspecific relationship between the dominant chigger species on two mouse species was measured by association coefficient (V). The results showed that the association coefficient (V) between L. sinicum and L. scutellare on A. draco was close to 0 (V = 0.09, V ≈ 0). The V between L. rusticum and L. densipunctatum on A. ilex was close to 0.5 (V = 0.49, V ≈ 0.5). The association coefficient (V) used in this paper is one of the simple and practicable methods to judge the interspecific relationship between any two species in a certain community. The range of V is from 0 to positive and negative 1, that is, [1, ±1]. When V approaches 0, the distribution of the two species is independent of each other. When V is positive and close to 1, it means that two species have a tendency to coexist in a certain environment or on a certain species of host for parasites. When the V is negative and close to -1, it means that there is a mutually exclusive relationship between the two species [5,40]. This study implies that the distribution of L. sinicum and L. scutellare on A. draco seems independent of each other without obvious interspecific dependency. A low degree of interdependence, however, exists between L. rusticum and L. densipunctatum on A. ilex, and these two chigger species tend to choose the same individuals of A. ilex at the same time, but the degree of interdependence is still relatively low (V < 0.5).

The results showed that all the distribution indexes calculated were larger than the critical value (C > 1, m* > m, I > 1) of determining the aggregated distribution (Table 1, Table 8), and this indicates that the dominant chigger species are of aggregated distribution among different individuals of their corresponding mouse host, A. draco and A. ilex. This aggregated distribution further indicates that the distribution of dominant chigger species among different individuals of their hosts is very uneven. Some host individuals may harbor a large number of chiggers on their body surface, while some other hosts may have no or only a few chiggers. The aggregated distribution of ectoparasites, including chiggers, suggests that there is an intraspecific relationship of mutual attraction and interdependence between different individuals of the same parasite species. This mutual attraction and interdependence within a certain species are conducive to the survival, mating and reproduction of the population [5,51].

The species abundance distribution aims to reveal the relationship between the number of species and the number of individuals in a community, which reflects the proportion structure of common and rare species in the community [33,42,43]. In the present study, the species abundance distribution of the chigger community on A. draco was successfully fitted by Preston’s lognormal model, which shows that most chigger species are rare species with few individuals, while few mite species are dominant species with abundant individuals. With the increase of chigger individuals, the number of chigger species gradually decreased (Table 9, Figure 6). The result is consistent with some previous reports on the species abundance of chiggers [36,43]. Due to the small number of individuals and species of chiggers and mice collected, however, Preston’s lognormal model is not applicable to the chigger community on A. ilex. In ecological practice, if the species abundance distribution of a specific community is successfully fitted by Preston’s log-normal distribution model, the number of expected total species in the community can be roughly estimated, but this estimation must be based on a large number of samples [52]. Due to the small number of host samples (only 567 A. draco and 154 A. ilex captured) and a small number of chiggers (total 290), the total number of chigger species was not estimated in this paper, which remains to be conducted in further studies.

It must be pointed out that the present study is just a preliminary comparison of chiggers on two sibling mouse species due to the small number of host samples, especially A. ilex, and some results may still be unstable. With the expansion of survey areas and the increase of host samples in future research, some results may fluctuate and change to some extent.

5. Conclusions

In southwest China, the susceptibility of two sibling mouse species (A. draco and A. ilex) to chigger infestation is quite different, with different species composition and community structure, different infestation status and different dominant chigger species. The results support that A. draco and A. ilex are two independent rodent species from the aspect of chigger infestation.