3种检测方法在中缅边境疟疾无症状感染者筛查中的比较

doi:10.12140/j.issn.1000-7423.2020.02.004

中国寄生虫学与寄生虫病杂志 ›› 2020, Vol. 38 ›› Issue (2): 152-158.doi: 10.12140/j.issn.1000-7423.2020.02.004

3种检测方法在中缅边境疟疾无症状感染者筛查中的比较

王笑笑¹^,², 肖回回³, 黄芳¹, 周水森¹^,^*()

1 中国疾病预防控制中心寄生虫病预防控制所,国家热带病研究中心,世界卫生组织热带病合作中心,科技部国家级热带病国际联合研究中心,卫生部寄生虫病原与媒介生物学重点实验室,上海 20025
2 浙江省疾病预防控制中心,杭州310051
3 长沙市疾病预防控制中心,长沙 410004

收稿日期:2019-11-06 出版日期:2020-04-30 发布日期:2020-05-11
通讯作者: 周水森
作者简介:王笑笑（1982-）,女,硕士研究生,副主任医师,从事寄生虫病研究。E-mail： xxwang@cdc.zj.cn
基金资助:
“艾滋病和病毒性肝炎等重大传染病防治”科技重大专项(2018ZX10101002-002);国家卫生计生委2017年澜湄合作专项基金(2020399);上海市自然科学基金(18ZR1443400);美国国立卫生院A美国国家过敏和传染病研究所U19项目(U19AI129386)

Performance comparison of three methods in detecting asymptomatic malaria infection on China-Myanmar border

Xiao-xiao WANG¹^,², Hui-hui XIAO³, Fang HUANG¹, Shui-sen ZHOU¹^,^*()

1 National Institute of Parasite Diseases, Chinese Center for Disease Control and Prevention; Chinese Center for Tropical Diseases Research; WHO Collaborating Center for Tropical Diseases; National Center for international Research on Tropical Diseases, Ministry of Science and Technology; Key Laboratory of Parasite and Vector Biology, Ministry of Health, Shanghai 20025, China
2 Zhejiang Province Center for Disease Control and Prevention, Hangzhou 310051, China
3 Changsha Center for Disease Control and Prevention, Changsha 410004, China

Received:2019-11-06 Online:2020-04-30 Published:2020-05-11
Contact: Shui-sen ZHOU
Supported by:
Supported by the National Major Science and Technology Project--Prevention and Treatment of AIDS, Viral Hepatitis, and Other Major Infectious Diseases(2018ZX10101002-002);National Health Commission Special Project for Lancang-Mekong Cooperation in 2017(2020399);the Natural Science Foundation of Shanghai(18ZR1443400);the National Institutes of Health/National Institute of Allergy and Infectious Diseases U19 project(U19AI129386)

摘要/Abstract

摘要：

目的比较3种不同检测方法对疟疾无症状感染者的检出能力,并了解中缅边境地区的疟疾无症状感染水平。方法 2014年7月在中缅边境地区云南省盈江县选择那邦镇、支那乡和缅甸拉咱安置点等3个调查点,采集调查对象血样,制作厚薄血膜和滤纸血标本,分别采用显微镜观察、荧光定量PCR和超敏PCR检测疟原虫感染情况。结果共采集387份血样,显微镜观察检出6例无症状感染者（间日疟5例、恶性疟1例）,感染检出率为1.6%;荧光定量PCR检出13例无症状感染者（间日疟12例、恶性疟1例）,感染检出率为3.4%;超敏PCR检出38例无症状感染者（间日疟29例、恶性疟9例）,感染检出率为9.8%。以显微镜观察为疟原虫感染诊断的金标准,荧光定量PCR检测无症状感染者的灵敏度为100%,特异性为98.2%;超敏PCR检测无症状感染者的灵敏度为100%,特异性91.6%。超敏PCR结果显示,那邦镇的无症状感染检出率最高,为17.1%（22/129）,其次为缅甸拉咱难民安置点,为10.0%（11/110）,支那乡最低,为3.4%（5/148）,3个调查点的感染检出率差异有统计学意义（P < 0.05）。检出虫种以间日疟原虫为主,占76.3%,恶性疟原虫占23.7%;女性无症状感染检出率为10.7%（23/215）,高于男性的8.7%（15/172）,两者差异无统计学意义（P > 0.05）。不同年龄组以15~29岁的无症状感染检出率最高,为17.5%（10/57）,各年龄组间差异无统计学意义（P > 0.05）。结论超敏PCR对疟疾无症状感染者的检出率高于荧光定量PCR和显微镜观察。中缅边境地区人群中存在一定比例的疟疾无症状感染者。

关键词: 疟疾, 无症状感染者, PCR

Abstract:

Objective To compare the performances of three methods in detecting asymptomatic malaria infection in residents, and assess the prevalence of asymptomatic malaria on China-Myanmar border.Methods Cross-sectional survey was carried out in Nabang town and Zhina town in Yingjiang county of Yunnan Province, China and a resettlement site in Laiza of Myanmar. Finger-prick blood samples were collected to prepare thick/thin blood smears and dried blood spots on filter paper. Malaria parasite infection was examined by light microscopy, real-time fluorescent PCR and ultrasensitive PCR (usPCR).Results A total of 387 blood samples were collected. Light microscopy found 6 participants with asymptomatic malaria infection (5 Plasmodium vivax infection and 1 P. falciparum infection), with a detection rate of 1.6%; real-time fluorescent PCR revealed 13 asymptomatic malaria infection cases (12 P. vivax infection; 1 P. falciparum infection) with a detection rate of 3.4%; and usPCR found 38 asymptomatic malaria infection cases(29 P. vivax infection and 9 P. falciparum infection) with a detection rate of 9.8%. Using light microscopy as the gold standard, the sensitivity and specificity of real-time fluorescent PCR were 98.2% and 100%, and those of usPCR were 91.6% and 100%, respectively. The usPCR results revealed that the detection rate of asymptomatic malaria infections was highest in Nabang town (17.1%, 22/129), followed by the Laiza resettlement site (10.0%, 11/110) and Zhina town (3.4%, 5/148), showing significant differences between the three sites (P < 0.05). Among the parasite species detected, P. vivax accounts for 76.3%, while P. falciparum for 23.7%. The detection rate was higher in females (10.7%, 23/215) than in males (8.7%, 15/172), but with no significant difference (P > 0.05). The age distribution showed that, the detection rate of asymptomatic infection was highest in the examinees aged 15-29 years(17.5%, 10/57), but there were no significant differences between the age groups.Conclusion Of the three methods compared for the ability in detection of asymptomatic malaria infection, the usPCR method presents higher detection rate than light microscopy and real-time fluorescent PCR. There are a certain proportion of asymptomatic malaria infection cases among the residents on China-Myanmar border.

Key words: Malaria, Asymptomatic malaria infection, PCR

中图分类号:

R531.3

王笑笑, 肖回回, 黄芳, 周水森. 3种检测方法在中缅边境疟疾无症状感染者筛查中的比较[J]. 中国寄生虫学与寄生虫病杂志, 2020, 38(2): 152-158.

Xiao-xiao WANG, Hui-hui XIAO, Fang HUANG, Shui-sen ZHOU. Performance comparison of three methods in detecting asymptomatic malaria infection on China-Myanmar border[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2020, 38(2): 152-158.

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方法Method	引物和探针名称 Primers and probes	序列（5′→3′） Sequences (5′→3′)
荧光PCR Fluorescent PCR	Pv-F	ATTGCTTTTGAGAGGTTTTGTTACTTT
	Pv-R	GCTGTAGTATTCAAACACAATGAACTCAA
	Pf-F	CGCTTCTAGCTTAATCCACATAACTG
	Pf-R	AATTTACTCAAAGTAACAAGGACTTCCAAG
超敏PCR Ultrasensitive PCR	Pf-F	CCGACTAGGTGTTGGATGAAAGTGTTAA
	Plas-R	AACCCAAAGACTTTGATTTCTCATAA
	Pf-P	/5Cy5/agcaatctaaaagtcacctcgaaagatgact/3BHQ_2/
	Pv-F	CCGACTAGGCTTTGGATGAAAGATTTTA
	Pv-P	/56-FAM/agcaatctaagaataaactccgaagagaaaattct/3BHQ_2/
	Actin-F	ACCGAGCGCGGCTACAG
	Actin-R	CTTAATGTCACGCACGATTTCC
	Actin-P	/VIC/ttcaccaccacggccgagc/MGB/

调查点 Spots	调查人数No. exam’d	显微镜观察Microscopic examination				荧光PCR Fluorescent PCR				超敏PCR Ultrasensitive PCR
调查点 Spots	调查人数No. exam’d	间日疟检出数 No. P. vivax infected	恶性疟检出数 No. P. falciparum infected	小计Subtotal	感染检出率/%Prevalence/%	间日疟检出数 No. P. vivax infected	恶性疟检出数 No. P. falciparum infected	小计Subtotal	感染检出率/%Prevalence/%	间日疟检出数 No. P. vivax infected	恶性疟检出数 No. P. falciparum infected	小计Subtotal	感染检出率/% Prevalence/%
那邦镇Nabang town	129	2	0	2	1.6	6	0	6	4.7	19	3	22	17.1
支那乡Zhina town	148	0	1	1	0.7	0	1	1	0.7	0	5	5	3.4
拉咱安置点Laiza resettlement site	110	3	0	3	2.7	6	0	6	5.5	10	1	11	10.0
合计Total	387	5	1	6	1.6	12	1	13	3.4	29	9	38	9.8

方法和结果Methods and results		显微镜观察Microscopic examination		小计Subtotal	灵敏度/%Sensitivity/%	特异度/%Specificity/%
方法和结果Methods and results		阳性数No. positive	阴性数No. negative	小计Subtotal	灵敏度/%Sensitivity/%	特异度/%Specificity/%
荧光PCR Fluorescent PCR	阳性数No. positive	6	7	13
荧光PCR Fluorescent PCR	阴性数No. negative	0	374	374	100	98.2
	小计Subtotal	6	381	387
超敏PCR Ultrasensitive PCR	阳性数No. positive	6	32	38
	阴性数No. negative	0	349	349	100	91.6
	小计Subtotal	6	381	387

特征 Feature	感染检出例数No. infected	总例数 Total	比例/% Percent
性别Gender
男Male	15	172	8.7
女Female	23	215	10.7
年龄组Age group
0~4	1	140	0.7
5~14	7	56	12.5
15~29	10	57	17.5
30~44	10	66	15.2
45~60	8	49	16.3
> 60	2	19	2/19
虫种Species
间日疟原虫P. vivax	29	387	7.5
恶性疟原虫P. falciparum	9	387	2.3

3种检测方法在中缅边境疟疾无症状感染者筛查中的比较

Performance comparison of three methods in detecting asymptomatic malaria infection on China-Myanmar border

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