Field contamination of feces of Echinococcus definitive hosts and its characteristics in Seda County, Sichuan Province

doi:10.12140/j.issn.1000-7423.2021.05.018

Abstract

Abstract:

Objective To understand the feces contamination and characteristics of definitive hosts of Echinococcus in the field in Seda County, Sichuan Province, to provide a basis for a prevention and control strategy of the source of infection in the field. Methods In October 2020, survey spots were selected every 1-2 km apart along the main traffic routes and field grazing routes in Seda County using mechanical sampling method to record their geographic location, number of feces found, pasture type, topographic type, vegetation type, vegetation coverage, mean height of grass, small mammal densities, and feces type of definitive hosts. Canine fecal samples were collected from survey spots, and the DNA was extracted from fthe samples for detecting the infection of Echinococcus by PCR. PCR-positive products were sent for sequencing, and the sequences were compared against the NCBI database to determine the species of Echinococcus. SPSS 18.0 software was used to statistically analyze the field contamination of definitive hosts’ feces and its characteristics. Results In total, 40 survey spots were selected, among them 34 (85.00%) were found having canine feces, while Echinococcus-positive feces were detected in 17 survey spots (42.50%). A total of 227 canine fecal samples were collected and examined by PCR, revealing the positive rates of Echinococcus spp., E. shiquicus(Es) and E. multilocularis (Em) were 18.06% (41/227), 14.10% (32/227), and 4.85% (11/227), respectively. Among these samples, two were of mixed infection of Es and Em, while no Eg-positive feces was found. The fecal-positive rate of Echinococcus in winter and summer pastures was 19.90% (40/201) and 3.85% (1/26), respectively (P > 0.05). The fecal-positive rate of Es in winter pasture was 15.92% (32/201) higher than that in summer pasture (0, 0/26) (P < 0.05). There was no significant differences in fecal-positive rates of Echinococcus, Es, and Em on plain, hillside, and benchland sites(P > 0.05). Among the different vegetation types, there were no significant differences in the positive rates of Echinococcus, Es, and Em in fecal samples between altitude cold meadow and altitude cold bush-meadow. The feces density in altitude cold meadow (7.17/10 000 m ²) was higher than that of altitude cold bush-meadow (3.44/10 000 m²) (Z = -2.376, P < 0.05). There was no significant difference in the fecal-positive rates of Echinococcus, Es, and Em in areas with sparse, relatively sparse, medium, relatively dense, and dense vegetation coverage (P > 0.05). Among different mean grass heights, theere was no significant difference in the fecal-positive rates of Echinococcus, Es, and Em at mean grass heights ≤ 5 cm, 6-10 cm, 11-15 cm and ≥ 16 cm(P > 0.05). The small mammals density 401-600 heads/10 000 m² rendered the feces density 12.25/10 000 m², which was higher than that at the mammal density ≤ 200 heads/10 000 m ² (3.32/10 000 m²) (Z = -2.738, P < 0.008). The fecal-positive rates of Es at the mammal density≥ 601 heads/10 000 m² was 29.73% (11/37), which was higher than the positive rate 5.48% (4/73) at the density ≤ 200 heads/10 000 m ² (χ² = 12.261, P < 0.008). The fox fecal-positive rates of Echinococcus and Es were 25.95% (34/131) and 22.14% (29/131), respectively, which were both higher than that of canine feces (7.45%, 7/94; 3.19%, 3/94) (χ ² = 12.580, 16.103, P < 0.05). There was no significant difference in the positive rate of Em between dog feces (5.32%, 5/94) and fox feces (4.58%, 6/131) (P > 0.05). Conclusion The field surveyed in Seda County, Sichuan Province is highly contaminated by Echinococcus of definitive hosts’ feces, showing foxes as the main source of infection, high Em positive rate in dog feces, in Seda County, Sichuan Province. It was demonstrated that winter pasture, altitude cold meadow, and areas with high densities of small mammals may the key areas for control of sources of infection in the wild.

Key words: Echinococcosis, Definitive host, Fecal contamination, PCR, Seda county

CLC Number:

R532.32

FU Mei-hua, WANG Xu, HAN Shuai, YU Wen-jie, YANG Yi, GAO Ming-jun, LIU Jian-feng, GUAN Ya-yi, WANG Ying, LI Chun-yang, SHI Dan-dan, WU Wei-ping. Field contamination of feces of Echinococcus definitive hosts and its characteristics in Seda County, Sichuan Province[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2021, 39(5): 687-695.

Figures/Tables 3

Table 1

Table 2

Fecal contamination of definitive hosts in different environmental types and small mammals densities in Seda County, Sichuan Province in 2020

变量 Variables	分组 Group	调查点个数 No. survey spot	粪便份数 No. fecal sample	阳性粪便份数 No. positive fece	粪便密度^a/份(10 000 m²)^-1 Density of feces^a/No. (10 000 m²) ^-1	阳性粪便密度^a/份·(10 000 m²) ^-1 Density of positive feces^a/No. (10 000 m²) ^-1	粪便棘球绦虫阳性率^b/% Fecal positive rate^b/%	Es阳性粪便数^b（占比/%） No. Es positive fece^b (Ratio/%)	Em阳性粪便数^b（占比/%） No. Em positive fece^b (Ratio/%)
牧场类型 Type of pasture	冬季牧场 Winter pasture	37	201	40	5.43	1.08	19.90	32 (15.92)	10 (4.98)
牧场类型 Type of pasture	夏季牧场Summer pasture	3	26	1	8.67	0.33	3.85	0 (0)	1 (3.85)
	组间检验	df			1	1	-	-	-
	Intergroup test	Z/χ²			-0.078	-0.574	-	-	-
		P			> 0.05	> 0.05	> 0.05	< 0.05	> 0.05
地形类别 Type of terrain	平坝 Plain	19	88	19	4.63	1.00	21.59	13 (14.77)	7 (7.95)
地形类别 Type of terrain	山坡 Hillside	13	73	10	5.62	0.77	13.70	8 (10.96)	3 (4.11)
	河滩 Benchland	8	66	12	8.25	1.50	18.18	11 (16.67)	1 (1.52)
	组间检验	df			2	2	2	2	-
	Intergroup test	χ²			0.116	0.384	1.680	0.987	-
		P			> 0.05	> 0.05	> 0.05	> 0.05	> 0.05
植被类型Vegetation type	高寒草甸Alpine meadow	24	172	34	7.17	1.42	19.77	28 (16.28)	7 (4.07)
植被类型Vegetation type	高寒灌木丛草甸Alpine shrub meadow	16	55	7	3.44	0.44	12.73	4 (7.27)	4 (7.27)
	组间检验	df			1	1	1	1	-
	Intergroup test	χ²			-2.376	-1.542	1.396	2.791	-
		P			< 0.05	> 0.05	> 0.05	> 0.05	> 0.05
植被覆盖度^c Vegetation coverage^c	Ⅰ	2	24	1	12.00	0.50	4.17	0 (0)	1 (4.17)
	Ⅱ	2	2	0	1.00	0	0	0 (0)	0 (0)
	Ⅲ	4	35	3	8.75	0.75	8.57	1 (2.86)	3 (8.57)
	Ⅳ	13	84	20	6.46	1.54	23.81	17 (20.24)	4 (4.76)
	Ⅴ	19	82	17	4.32	0.89	20.73	14 (17.07)	3 (3.66)
	组间检验	df			4	4	-	-	-
	Intergroup test	χ²			6.127	7.130	-	-	-
		P			> 0.05	> 0.05	> 0.05	< 0.05	> 0.05
平均草高度/cm Mean grass height/cm	≤ 5	9	86	16	9.56	1.78	18.60	12 (13.95)	4 (4.65)
	6~10	19	63	9	3.32	0.47	14.29	7 (11.11)	4 (6.35)
	11~15	6	56	10	9.33	1.67	17.86	9 (16.07)	1 (1.79)
	≥ 16	6	22	6	3.67	1.00	27.27	4 (18.18)	2 (9.09)
	组间检验	df			3	3	3	3	-
	Intergroup test	χ²			4.459	2.813	1.887	0.949	-
		P			> 0.05	> 0.05	> 0.05	> 0.05	> 0.05
小型哺乳动物密度/ 个·（10 000 m²）^-1 The density of small mammals/No. (10 000 m²) ^-1	≤ 200	22	73	9	3.32	0.41	12.33	4 (5.48)	6 (8.22)
	201~400	12	68	14	5.67	1.17	20.59	10 (14.71)	4 (5.88)
	401~600	4	49	7	12.25	1.75	14.29	7 (14.29)	1 (2.04)
	≥ 601	2	37	11	18.50	5.50	29.73	11 (29.73)	0 (0)
	组间检验	df			3	3	3	3	-
	Intergroup test	χ²			13.000	7.278	5.790	11.966	-
		P			< 0.05	> 0.05	> 0.05	< 0.05	> 0.05

Table 2

Table 3

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目的基因Target genes	引物序列（5′→3′） Primers squence (5′→3′)	产物长度/bp Amplicon length/bp
nad1（Eg)	FP: 5′-GTTTTTGGCTGCCGCCAGAAC-3′	226
	RP: 5′-AATTAATGGAAATAATAACAAACTTAATCAACAAT-3′
cox1（带科绦虫) cox1（Taeniidae)	FP: 5′-TTGAATTTGCCACGTTTGAATGC-3′	874
cox1（带科绦虫) cox1（Taeniidae)	RP: 5′-GAACCTAACGACATAACATAATGA-3′
cox1（Em)	FP: 5′-GTCATATTTGTTTAAGTATAAGTGG-3′	243
	RP: 5′-CACTCTTATTTACACTAGAATTAAG-3′
nad1（Es)	FP: 5′-TTATTCTCAGTCTCGTAAGGGTCCG-3′	442
	RP: 5′-CAATAACCAACTACATCAATAATT-3′

变量 Variables	对比组 Comparative group	粪便密度 Fecal density		Es阳性粪便数/% No. Es positive fece/%
变量 Variables	对比组 Comparative group	Z	P	χ²	P
植被覆盖度 Vegetation coverage	Ⅰ vs. Ⅱ	-	-	-	-
植被覆盖度 Vegetation coverage	Ⅰ vs. Ⅲ	-	-	-	> 0.005^a
	Ⅰ vs. Ⅳ	-	-	-	> 0.005^a
	Ⅰ vs. Ⅴ	-	-	-	> 0.005^a
	Ⅱ vs. Ⅲ	-	-	-	> 0.005^a
	Ⅱ vs. Ⅳ	-	-	-	> 0.005^a
	Ⅱ vs. Ⅴ	-	-	-	> 0.005^a
	Ⅲ vs. Ⅳ	-	-	5.814	> 0.005^a
	Ⅲ vs. Ⅴ	-	-	-	> 0.005^a
	Ⅳ vs. Ⅴ	-	-	0.274	> 0.005^a
小型哺乳动物密度/ 个·（10 000 m²）^-1 The density of small mammals /No.·(10 000 m²)^-1	≤ 200 vs. 201~400	-1.748	> 0.008^b	3.351	> 0.008^b
	≤ 200 vs. 401~600	-2.738	< 0.008^b	-	> 0.008^b
	≤ 200 vs. ≥ 601	-2.124	> 0.008^b	12.261	< 0.008^b
	201~400 vs. 401~600	-1.950	> 0.008^b	0.004	> 0.008^b
	201~400 vs. ≥ 601	-1.930	> 0.008^b	3.380	> 0.008^b
	401~600 vs. ≥ 601	-0.926	> 0.008^b	3.038	> 0.008^b