3种方法检测野鼠血吸虫感染的效果评价

doi:10.12140/j.issn.1000-7423.2025.02.006

中国寄生虫学与寄生虫病杂志 ›› 2025, Vol. 43 ›› Issue (2): 186-191.doi: 10.12140/j.issn.1000-7423.2025.02.006

3种方法检测野鼠血吸虫感染的效果评价

唐琦¹()(), 吕超¹, 王熙¹, 郭苏影¹, 许晓娟², 朱海², 李银龙¹, 林伟娜¹, 周新杰¹, 冯婷¹, 许静¹, 秦志强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）；传染病溯源预警与智能决策全国重点实验室；国家卫生健康委员会寄生虫病原与媒介生物学重点实验室；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，上海 200025
2 安徽省疾病预防控制中心，安徽合肥 230601

收稿日期:2025-02-07 修回日期:2025-03-19 出版日期:2025-04-30 发布日期:2025-04-27
通讯作者: *秦志强（ORCID：0000-0002-1130-468）男，博士，研究员，从事血吸虫感染免疫机理与分子诊断。E-mail：qinzq@nipd.chinacdc.cn
作者简介:唐琦（ORCID：0009-0008-2137-7524），女，硕士研究生，从事寄生虫监测与预警。E-mail：15970005361@163.com

Efficiency evaluation of three assays for detection of Schistosoma japonicum infections in wild mice

TANG Qi¹()(), LV Chao¹, WANG Xi¹, GUO Suying¹, XU Xiaojuan², ZHU Hai², LI Yinlong¹, LIN Weina¹, ZHOU Xinjie¹, FENG Ting¹, XU Jing¹, QIN Zhiqiang¹^,^*()()

1 National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Chinese Center for Tropical Diseases Research; National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases; NHC Key Laboratory on Parasite and Vector Biology; WHO Collaborating Centre for Tropical Diseases; National Center for International Research on Tropical Diseases, Ministry of Science and Technology, Shanghai 200025, China
2 Anhui Provincial Center for Disease Control and Prevention, Hefei 230601, Anhui, China

Received:2025-02-07 Revised:2025-03-19 Online:2025-04-30 Published:2025-04-27
Contact: * E-mail：qinzq@nipd.chinacdc.cn

摘要/Abstract

摘要：

目的评价肝脏匀浆涂片镜检（病原学检测）、实时荧光定量PCR（qPCR）和环介导等温扩增（LAMP）（核酸检测）等3种方法检测野鼠日本血吸虫感染情况的效果，为野鼠血吸虫感染的监测提供较优的实验室检测方法。方法从安徽省东至县收集115份野鼠肝脏样品（村A 63份，村B 52份），分别进行肝脏匀浆涂片镜检、qPCR和LAMP方法检测，统计并比较样品的阳性率；以镜检检测结果为标准，对qPCR和LAMP检测结果进行一致性分析，并对病原学检测与核酸检测结果一致性较低的样品随机抽取70%进行DNA测序验证。结果肝脏匀浆涂片镜检共计发现54份阳性样品，阳性率为46.96%（54/115），其中村A和村B的阳性率分别为46.03%（29/63）和48.08%（25/52）（χ² = 0.001, P > 0.05）。qPCR检测样品的阳性率为69.57%（80/115），高于镜检阳性率（χ² = 11.175，P < 0.01）。村A的阳性率（66.67%，42/63）与村B的阳性率（73.08%，38/52）之间的差异无统计学意义（χ² = 0.340，P > 0.05）。LAMP检测样品的阳性率为56.52%（65/115），与镜检结果差异无统计学意义（χ² = 1.741，P > 0.05）。此外，LAMP检测村A的样品阳性率（71.43%，45/63）高于村B（38.46%，20/52）（χ² = 11.293，P < 0.01）。一致性分析结果显示，qPCR检测结果和镜检结果一致性较好（Kappa值 = 0.524 4），LAMP检测结果和镜检结果一致性较低（Kappa值 = 0.154 5）。从村A的24份镜检阴性但LAMP阳性的样品中随机抽取17份样品进行测序，其中15份与日本血吸虫28S核糖体DNA（GenBank登录号为JF721395.1）序列一致性≥ 98%，鉴定为阳性。结论 3种检测方法检测野鼠日本血吸虫感染敏感性存在差异，qPCR检测的阳性率最高，与镜检一致性较好；LAMP法操作便捷，但与镜检一致性较低。

关键词: 日本血吸虫, 野鼠, 肝脏匀浆镜检, 实时荧光定量PCR, 环介导等温扩增

Abstract:

Objective To evaluate the performance of liver homogenate smear microscopy (parasitological detection), real-time quantitative reverse transcription PCR (qPCR) assay and loop-mediated isothermal amplification (LAMP) for detection of Schistosoma japonicum infections in wild rodents, so as to provide an optimal laboratory diagnostic assay for surveillance of S. japonicum infection in wild rodents. Methods A total of 115 wild rodents liver samples (63 samples from Village A and 52 samples from Village B) were collected from Dongzhi County, Anhui Province, and detected by liver homogenate smear microscopy, qPCR assay and LAMP, respectively. The positive rates of the samples were calculated and compared. The consistency between qPCR and LAMP for detection of the liver samples was evaluated with microscopy as a standard, and 70% of liver samples with a low consistency between the parasitological assay and nucleic acid tests were randomly selected for validation with DNA sequencing. Results Liver homogenate smear microscopy detected 54 positive liver samples, with a positive rate of 46.96% (54/115), and the positive rate of liver samples was 46.03% (29/63) in Village A and 48.08% (25/52) in Village B (χ² = 0.001, P > 0.05). qPCR assay tested a 69.57% (80/115) positive rate of liver samples, which was higher than liver homogenate smear microscopy (χ² = 11.175, P < 0.01), and there positive rate was 66.67% (42/63) in Village A and 73.08% (38/52) in Village B (χ² = 0.340, P > 0.05). LAMP tested a 56.52% (65/115) positive rate of liver samples, which was comparable to liver homogenate smear microscopy (χ² = 1.741, P > 0.05), and the positive rate was higher in Village A(71.43%, 45/63) than in Village B (38.46%, 20/52) (χ² = 11.293, P < 0.01). There was a high consistency between qPCR assay and liver homogenate smear microscopy (Kappa value = 0.524 4), and a low consistency between LAMP and liver homogenate smear microscopy (Kappa value = 0.154 5). Among the 24 liver samples from Village A that were negative for liver homogenate smear microscopy but positive for LAMP, 17 samples were randomly selected for DNA sequencing, and 15 of these samples showed a sequence identity of 98% and higher with the 28S ribosomal DNA of S. japonicum (GenBank accession No.: JF721395.1) and were therefore identified as positives. Conclusion Three assays have diverse sensitivities for detection of S. japonicum infections in wild rodents. qPCR assay has the highest positive rate for detection of S. japonicum infections in wild rodents and a high consistency with liver homogenate smear microscopy, while LAMP is convenient to perform but has a low consistency with liver homogenate smear microscopy.

Key words: Schistosoma japoniucm, Wild rodents, Microscopic examination of liver homogenates, Real-time quantitative reverse transcription PCR, Loop mediated isothermal amplification

中图分类号:

R532.21

唐琦, 吕超, 王熙, 郭苏影, 许晓娟, 朱海, 李银龙, 林伟娜, 周新杰, 冯婷, 许静, 秦志强. 3种方法检测野鼠血吸虫感染的效果评价[J]. 中国寄生虫学与寄生虫病杂志, 2025, 43(2): 186-191.

TANG Qi, LV Chao, WANG Xi, GUO Suying, XU Xiaojuan, ZHU Hai, LI Yinlong, LIN Weina, ZHOU Xinjie, FENG Ting, XU Jing, QIN Zhiqiang. Efficiency evaluation of three assays for detection of Schistosoma japonicum infections in wild mice[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2025, 43(2): 186-191.

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引物名称 Primer name	引物序列 (5′→3′) Primer sequence (5'→3')	扩增片段长度/bp Length of amplified fragment/bp
PCR		405
Sj28_Forward	GGTTTGACTATTATTGTTGAGC
Sj28_Reverse	TCTCACCTTAGTTCGGACTGA
qPCR		165
SjR2_Forward	TGCCAGAGCGATGACCATAAAC
SjR2_Reverse	GCAATGCCTCGCAGTGTGATAG
LAMP		217
Sj28s-F3	GCTTTGTCCTTCGGGCATTA
Sj28s-B3	GGTTTCGTAACGCCCAATGA
Sj28s-FIP	ACGCAACTGCCAACGTGACATACTGGTCGGCTTGTTACTAGC
Sj28s-BIP	TGGTAGACGATCCACCTGACCCCTCGCGCACATGTTAAACTC

3种方法检测野鼠血吸虫感染的效果评价

Efficiency evaluation of three assays for detection of Schistosoma japonicum infections in wild mice

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样品来源Sample source	野鼠肝脏数/份No. rodent liver	肝脏匀浆镜检 Liver homogenate microscopy		qPCR		LAMP
样品来源Sample source	野鼠肝脏数/份No. rodent liver	阳性数/份No. positive	阳性率/% Positive rate/%	阳性数/份No. positive	阳性率/% Positive rate/%	阳性数/份No. positive	阳性率/% Positive rate/%
村A Village A	63	29	46.03	42	66.67	45	71.43
村B Village B	52	25	48.08	38	73.08	20	38.46
合计 Total	115	54	46.96	80	69.57	65	56.52

肝脏匀浆镜检 Liver Homogenate microscopy	qPCR			LAMP
肝脏匀浆镜检 Liver Homogenate microscopy	+	-	合计 Total	+	-	合计 Total
+	53	1	54	35	19	54
-	27	34	61	30	31	61
合计 Total	80	35	115	65	50	115

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