Infection of Blastocystis and Cyclospora cayetanensis in residents in Eryuan County, Yunnan Province and their molecular characterization

doi:10.12140/j.issn.1000-7423.2024.02.016

Abstract

Abstract:

Objective To investigate the prevalence and molecular characterizations of Blastocystis and Cyclospora cayetanensis in residents in Eryuan County, Yunnan Province to provide reference for parasitic disease control and prevention. Methods From July to August 2022, permanent residents over the age of 1 were enrolled for the survey by simple random sampling from the five villages in Eryuan County, Yunnan Province. Fresh fecal samples and their social demographic information were collected. The fecal genomic DNA was extracted, and the ribosomal small subunit RNA (SSU rRNA) genes of Blastocystis and C. cayetanensis were amplified by PCR and nested PCR, respectively, in order to analyze the prevalence in the residents. The positive products were sequenced bidirectionally, the species and gene subtypes were identified by BLAST alignment in NCBI. The phylogenetic trees were constructed using the neighbor-Joining method by MEGA 11.0 software. SPSS 21.0 software was used for statistical analysis, and the infection rates were compared by χ² test or Fisher’s exact probability method. Results A total of 100 fecal samples were obtained from residents, of which the Blastocystis infection rate was 14.0% (14/100). Among them, the Blastocystis infection rates in male and female were 14.6% (6/41) and 13.6% (8/59), respectively. Residents over 60 years old and the family size of 6 persons and above had the highest infection rate, which was 18.5% (5/27) and 3/15, respectively. The infection rates were 2/11 and 17.0% (8/47) for those with no education and only elementary school education, respectively. All the infected residents were farmers. There was no statistically significant difference in Blastocystis infection rate by gender, age, family size, education level and occupation (χ² = 0.023, 2.730, 2.235, 1.404, 1.668; all P > 0.05). The C. cayetanensis infection rate was 2.0% (2/100). All of the infected residents were female farmers. No mixed infection of Blastocystis and C. cayetanensis was detected. Both 7 ST1 and ST3 sequences were amplified from 14 Blastocystis-positive samples, of which 7 ST1 sequences had 25 nucleotide differences. Five sequences were 100% homology to the Blastocystis sequences (GenBank: ON932511, KU147348, MK801408, MW728079, OR754904); 2 were new sequences (GenBank: PP439288-PP439289), with 99.8% homology to the Blastocystis sequences (GenBank: OP725964, KU147333). There was 1 nucleotide difference within 7 ST3 sequences, with 100% homology to the Blastocystis sequences (GenBank: KU147372, MK801366). The 2 sequences of C. cayetanensis obtained were identical and showed 100% homology to the C. cayetanensis sequence (GenBank: KY770755). The phylogenetic trees showed that Blastocystis in this study clustered in large branches with gene subtype ST1 and ST3, respectively, and C. cayetanensis clustered in the same branch with C. cayetanensis of human origin. Conclusion Blastocystis and C. cayetanensis were detected in the residents of Eryuan County, Yunnan Province. The infection rate of Blastocystis was high, with the subtypes of ST1 and ST3, and the ST1 of Blastocystis reveals higher genetic variation.

Key words: Blastocystis, Cyclospora cayetanensis, Infection, Molecular characterization, Eryuan County

CLC Number:

R531.9

WANG Yaxue, JIAN Jinhua, LIU Hua, QIN Yuan, PENG Xiaoxue, SU Yaxin, SHEN Yujuan. Infection of Blastocystis and Cyclospora cayetanensis in residents in Eryuan County, Yunnan Province and their molecular characterization[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2024, 42(2): 242-250.

Figures/Tables 4

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References 47

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人口学特征 Demographic characteristic	检查人数 No. examined	阳性例数（感染率/%) No. positive (Infection rate/%)	亚型（数量） Subtype (n)	χ² 值 χ² value	P值 P value
性别 Gender				0.023	0.879
男性 Male	41	6 (14.6)	ST1 (3), ST3 (3)
女性 Female	59	8 (13.6)	ST1 (4), ST3 (4)
年龄/岁 Age/Year				2.730	0.384^a
< 5	0	0
5~17	13	0 (0)
18~35	4	0 (0)
36~60	56	9 (16.1)	ST1 (5), ST3 (4)
> 60	27	5 (18.5)	ST1 (2), ST3 (3)
家庭人口数 Family size				2.235	0.568^a
3人及以下 Three persons and below	22	2 (9.1)	ST1 (1), ST3 (1)
4人 Four persons	31	6 (19.4)	ST1 (3), ST3 (3)
5人 Five persons	32	3 (9.4)	ST1 (1), ST3 (2)
6人及以上 Six persons and above	15	3/15	ST1 (2), ST3 (1)
民族 Ethnic group				0.434	0.596
白族 Bai	92	14 (15.2)	ST1 (7), ST3 (7)
汉族 Han	8	0 (0)
职业 Occupation				1.668	0.197
农民 Farmer	85	14 (16.5)	ST1 (7), ST3 (7)
非农民^b Non-farmer^b	15	0 (0)
教育水平 Educational level				1.404	0.788^a
未接受教育 Uneducated	11	2/11	ST1 (1), ST3 (1)
小学 Elementary school	47	8 (17.0)	ST1 (3), ST3 (5)
初中 Junior high school	33	3 (9.1)	ST1 (2), ST3 (1)
高中及以上 Senior high school and above	9	1/9	ST1 (1)
总计 Total	100	14 (14.0)	ST1 (7), ST3 (7)

分离株编号 Isolate number	GenBank号 GenBank ID	核苷酸位点Nucleotide positions
分离株编号 Isolate number	GenBank号 GenBank ID	36	37	186	188	194	197	198	199	200	209	210	211	214	215	216	217	219	220	221	222	223	224	243	268	281
EY96	PP439288^a	C	A	A	C	-	T	G	A	G	C	T	C	G	A	C	A	T	A	A	G	T	C	A	T	A
EY21	OP725948	A	C	A	C	-	T	G	A	G	C	T	C	G	A	C	A	T	A	A	G	T	C	G	C	C
EY30	PP439289	A	C	G	-	C	A	G	T	A	T	A	G	A	G	A	T	A	T	C	A	C	T	G	T	C
EY75	KU147348	C	A	A	C	-	T	G	A	G	C	T	C	G	A	C	A	T	A	A	G	T	C	G	T	G
EY77	MK801408	C	A	G	-	C	A	A	T	G	T	A	G	A	G	A	T	A	T	C	A	C	T	G	T	C
EY78	MW728079	C	A	A	C	-	T	G	A	G	C	T	C	G	A	C	A	T	A	A	G	T	C	G	T	C
EY81	OR754904	A	C	A	C	-	T	G	A	G	C	T	C	G	A	C	A	T	A	A	G	T	C	G	C	C

年份 Year	地区 Region	人群 Population	检测方法 Detection method	检查数No. examined	阳性数No. positive	感染率/% Infection rate/%	亚型（数量） Subtype (n)	参考文献 Reference
1994	云南省 Yunnan Province	常住居民 Permanent residents	碘液涂片 Iodine smear	53 061	2 390	4.5	-	[15]
2007	大理 Dali	普通村民 Ordinary villagers	体外培养 + PCR In vitro cul-ture + PCR	407	75	18.4	ST3 (38), ST1 (22), ST2(6), ST1/3 (5), ST2/3 (1), 未知 Unknown (3)	[16]
2007	西双版纳 Xishuangbanna	普通村民 Ordinary villagers	体外培养 + PCR In vitro cul-ture + PCR	239	78	32.6	ST3 (55), ST1 (16), ST2 (1), ST4 (1), ST1/2 (1), ST1/3 (1), 未知 Unknown (3)	[17]
2008	西双版纳 Xishuangbanna	普通村民 Ordinary villagers	涂片 Smear	215	43	20.0	-	[18]
2016	昆明 Kunming	腹泻儿童 Diarrheal children	PCR	850	26	3.1	-	[19]
2016	昆明 Kunming	健康儿童 Healthy children	PCR	170	1	0.6	-	[19]
2016	昆明 Kunming	腹泻患者 Diarrheal patients	PCR	1 121	47	4.2	ST1 (46), ST2 (1)	[20]
2016	昆明 Kunming	非腹泻患者 Non-diarrheal patients	PCR	319	11	3.5	ST1 (11)	[20]
2018	腾冲 Tengchong	HIV携带者 HIV carriers	PCR	324	12	3.7	ST1 (3), ST4 (3), ST7 (3), ST3 (2), ST12 (1)	[21]
2019	西双版纳 Xishuangbanna	瑶族村民 Yao villagers	PCR	289	13	4.5	ST3 (8), ST1 (3), ST4 (1), 未知 Unknown (1)	[22]
2021	大理 Dali	在校大学生 College students	PCR	426	6	1.4	ST1 (6)	[23]
合计 Total				57 421	2 702	4.7	ST1 (107), ST3 (103), ST2 (8), ST4 (5), ST1/3 (6), ST1/2 (1), ST2/3 (1), 未知 Unknown (7)