快速诊断靶蛋白恶性疟原虫富组氨酸蛋白Ⅱ和疟原虫乳酸脱氢酶的热稳定性

doi:10.12140/j.issn.1000-7423.2021.02.022

中国寄生虫学与寄生虫病杂志 ›› 2021, Vol. 39 ›› Issue (2): 245-248.doi: 10.12140/j.issn.1000-7423.2021.02.022

快速诊断靶蛋白恶性疟原虫富组氨酸蛋白Ⅱ和疟原虫乳酸脱氢酶的热稳定性

李美¹(), 涂宏¹, 夏志贵¹^,^*(), 王真瑜², 周何军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）,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,国家级热带病国际联合研究中心,上海 200025
2 上海疾病预防控制中心,上海 200003

收稿日期:2020-06-03 修回日期:2020-08-06 出版日期:2021-04-30 发布日期:2021-04-30
通讯作者: 夏志贵
作者简介:李美（1976-）,女,博士,研究员,从事疟疾原虫学研究。E-mail： limei@nipd.chinacdc.cn

Thermal stability of diagnostic targets Plasmodium falciparum histidine rich protein Ⅱ and Plasmodium lactate dehydrogenase in rapid detection

LI Mei¹(), TU Hong¹, XIA Zhi-gui¹^,^*(), WANG Zhen-yu², ZHOU He-jun¹

1 National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention (Chinese Center for Tropical Diseases Research); NHC Key Laboratory of Parasite and Vector Biology; WHO Collaborating Centre for Tropical Diseases; National Center for International Research on Tropical Diseases, Shanghai 200025, China
2 Shanghai Center for Diseases Control and Prevention, Shanghai 200003, China

Received:2020-06-03 Revised:2020-08-06 Online:2021-04-30 Published:2021-04-30
Contact: XIA Zhi-gui

摘要/Abstract

摘要：

在新型冠状病毒肺炎流行期间,通常需将血样进行56 ℃病毒灭活处理后再行病原体检测。为了解56 ℃灭活对血样中疟原虫抗原稳定性的影响,本研究收集2017—2019年上海市疾病预防控制中心上报的疟疾患者血样71份,其中恶性疟血样38份、三日疟血样8份、卵形疟血样11份、间日疟血样14份。应用快速诊断检测（RDT）试剂盒对56 ℃孵育30 min前后血样中恶性疟原虫富组氨酸蛋白Ⅱ（PfHRPⅡ）和疟原虫乳酸脱氢酶（pLDH）的热稳定性进行测定。结果显示,热处理前T1检测线（检测目标PfHRPⅡ）呈阳性的38份恶性疟血样,热处理后35份仍呈T1阳性（92.11%,35/38, χ ² = 3.123,P > 0.05）;热处理前T2检测线（检测目标pLDH）阳性的54份血样（26份恶性疟血样、6份三日疟血样、10份卵形疟血样和12份间日疟血样）,经热处理后T2均呈阴性（阳性率为0,0/54,χ ² = 87.755, P < 0.01）。表明在56 ℃孵育30 min条件下,PfHRPⅡ的稳定性较好;pLDH不稳定,全部降解或失活。因此,应用RDT检测热处理后的血样,恶性疟血样的检测结果不受影响,但非恶性疟血样会被漏诊。

关键词: 疟原虫抗原, 富组氨酸蛋白Ⅱ, 疟原虫乳酸脱氢酶, 热稳定性

Abstract:

During the COVID-19 epidemic, blood samples are usually processed at 56 ℃ to attenuate the virus before pathogen detection. 71 blood samples of malaria patients reported by Shanghai Center for Disease Control and Prevention in 2017—2019 were collected, including 38 with Plasmodium falciparum infection, 8 P. malariae, 11 P. ovale and 14 P. vivax. The effect of inactivation on the thermal stability of P. falciparum histidine rich protein Ⅱ (PfHRPⅡ) and Plasmodium lactate dehydrogenase (pLDH) in blood samples was assessed before and after incubation at 56 ℃ for 30 min using the rapid diagnostic test (RDT) kit. The results showed that among the 38 P. falciparum T1-positive (PfHRPⅡ) blood samples before heat treatment, 35 samples remained to be T1-positive (92.11%, 35/38, χ ² = 3.123, P > 0.05) after heat treatment; while 54 blood samples (26 P. falciparum, 6 P. vivax, 10 P. ovale and 12 P. vivax) that were T2-positive (pLDH) before heat treatment turned to be T2-negative (positive rate 0, 0/54, χ ² = 87.755, P < 0.01) after heat treatment. It was demonstrated that PfHRPⅡ is stable during incubation at 56 ℃ for 30 min, while pLDH is unstable and degraded or inactivated during the heating. Therefore, the detection results of P. falciparum will not be affected by RDT, but diagnosis of the parasites other than P. falciparum in blood samples may be missed.

Key words: Plasmodium antigens, Plasmodium falciparum histidine-rich protein Ⅱ, Plasmodium lactate dehydrogenase, Thermal stability

中图分类号:

R382.31

李美, 涂宏, 夏志贵, 王真瑜, 周何军. 快速诊断靶蛋白恶性疟原虫富组氨酸蛋白Ⅱ和疟原虫乳酸脱氢酶的热稳定性[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(2): 245-248.

LI Mei, TU Hong, XIA Zhi-gui, WANG Zhen-yu, ZHOU He-jun. Thermal stability of diagnostic targets Plasmodium falciparum histidine rich protein Ⅱ and Plasmodium lactate dehydrogenase in rapid detection[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2021, 39(2): 245-248.

图/表 1

参考文献 20

[1]	Lei ZL, Wang LY. Control situation and primary task of key parasitic diseases in China[J]. Chin J Parasitol Parasit Dis, 2012,30(1):1-5. (in Chinese)
	( 雷正龙, 王立英. 全国重点寄生虫病防治形势与主要任务[J]. 中国寄生虫学与寄生虫病杂志, 2012,30(1):1-5.)
[2]	Zhang L, Feng J, Zhang SS, et al. The progress of national malaria elimination and epidemiological characteristics of malaria in China in 2017[J]. Chin J Parasitol Parasit Dis, 2018,36(3):201-209. (in Chinese)
	( 张丽, 丰俊, 张少森, 等. 2017年全国消除疟疾进展及疫情特征分析[J]. 中国寄生虫学与寄生虫病杂志, 2018,36(3):201-209.)
[3]	Zhang L, Feng J, Zhou SS. The progress of national malaria elimination and epidemiological characteristics of malaria in China in 2018[J]. 2019,37(3):241-247. (in Chinese)
	( 张丽, 丰俊, 周水森, 等. 2018年全国疟疾疫情特征及消除疟疾工作进展[J]. 中国寄生虫学与寄生虫病杂志, 2019,37(3):241-247.)
[4]	Zhang L, Feng J, Xia ZG. The progress of national malaria elimination and epidemiological characteristics of malaria in China in 2018[J]. 2020,38(2):133-138. (in Chinese)
	( 张丽, 丰俊, 夏志贵, 等. 2019年全国疟疾疫情特征及消除疟疾工作进展[J]. 中国寄生虫学与寄生虫病杂志, 2020,38(2):133-138.)
[5]	World Health Organization. WHO coronavirus disease (COVID-19) Dashboard[EB/OL]. (2021-04-12) [2021-04-12]. https://covid19.who.int/
[6]	Zou Y, Zheng YS, Cao J. Suggestions for standardizing the process of malaria diagnosis and treatment during the epidemic control of coronavirus disease (COVID-19)[J]. Chin J Parasitol Parasit Dis, 2020,38(1):1-4. (in Chinese)
	( 邹洋, 郑以山, 曹俊. 新型冠状病毒肺炎防控期间规范疟疾诊疗流程的建议[J]. 中国寄生虫学与寄生虫病杂志, 2020,38(1):1-4.)
[7]	Hussein M, Ali O, Homeida A, et al. Malaria and COVID-19: unmasking their ties[J]. Malar J, 2020,19(1):457. doi: 10.1186/s12936-020-03541-w
[8]	Chiodini J. COVID-19 and the impact on malaria[J]. Travel Med Infect Dis, 2020,35:101758. doi: S1477-8939(20)30245-3 pmid: 32479815
[9]	National Health Commission of the People’s Republic of China. The technical guidelines for novel coronavirus pneumonia diagnosis and treatment (version 7) [EB/OL]. (2020-03-04) [2020-06-03]. http://www.nhc.gov.cn/xcs/zhengcwj/202003/46c929-4a7dfe4cef80dc7f5912eb1989.shtml (in Chinese)
	( 中华人民共和国国家卫生健康委办公厅. 新型冠状病毒肺炎诊疗方案(试行第七版)[EB/OL]. (2020-03-04) [2020-06-03]. http://www.nhc.gov.cn/xcs/zhengcwj/202003/46c9294a7dfe4cef80-dc7f5912eb1989.shtml
[10]	WHO. Results of WHO product testing of malaria RDTs: round 7 (2015—2016) [M]. Geneva: WHO Press, 2017: 1-9.
[11]	WHO. Malaria Rapid Diagnostic Test Performance-Summary results of WHO malaria RDT product testing: Round 1-3 (2008—2011)[M]. Geneva: WHO Press, 2009: 1-16.
[12]	Jimenez A, Rees-Channer RR, Perera R, et al. Analytical sensitivity of current best-in-class malaria rapid diagnostic tests[J]. Malar J, 2017,16(1):128. doi: 10.1186/s12936-017-1780-5
[13]	Wang XY, Shi WZ. Protein chemistry and its application[M]. Beijing: China Textile Press, 2015: 45-64. (in Chinese)
	( 王雪燕, 师文钊. 蛋白质化学及其应用[M]. 北京: 中国纺织出版社, 2015: 45-64.)
[14]	Stillman JH, Somero GN. A comparative analysis of the evolutionary patterning and mechanistic bases of lactate dehydrogenase thermal stability in porcelain crabs, genus Petrolisthes[J]. J Exp Biol, 2001,204(Pt 4):767-776. pmid: 11171359
[15]	Wilson RJ McGregor IA Wilson RJ. The stability and fractionation of malarial antigens from the blood of Africans infected with Plasmodium falciparum[J]. Int J Parasitol, 1973,3(4):511-520. pmid: 4199255
[16]	Wu CP, Liu X, Li QW, et al. Structure and function of histidine-rich glycoprotein[J]. Chin J Biochem Mol Biol, 2012,28(1):1-8. (in Chinese)
	( 武彩萍, 刘欣, 李庆伟, 等. 富组氨酸糖蛋白(HRG)的结构与功能[J]. 中国生物化学与分子生物学报, 2012,28(1):1-8.)
[17]	Wilson RJ McGregor IA Hall PJ. Persistence and recurrence of S-antigen in Plasmodium falciparum infections in man[J]. Trans R Soc Trop Med Hyg, 1975,69(5/6):460-467. doi: 10.1016/0035-9203(75)90098-X
[18]	Dalrymple U, Arambepola R, Gething PW, et al. How long do rapid diagnostic tests remain positive after anti-malarial treatment?[J]. Malar J, 2018,17(1):228. doi: 10.1186/s12936-018-2371-9
[19]	Jiang L, Wang ZY, Ma XJ, et al. Comparative analysis of nucleotide sequences of lactate dehydrogenase (LDH) gene and LDH epitopes of Plasmodium vivax and Plasmodium falciparum[J]. Chin J Parasitol Parasit Dis, 2010,28(2):103-107. (in Chinese)
	( 江莉, 王真瑜, 马晓疆, 等. 间日疟原虫和恶性疟原虫乳酸脱氢酶基因的序列和重组抗原表位分析[J]. 中国寄生虫学与寄生虫病杂志, 2010,28(2):103-107.)
[20]	Liu ZY, Li XW, Zhou Y, et al. Impact of heating for virus inactivation on result of test of biochemical and inflammatory indexes[J]. Chin J Nosocomiol, 2020,30(10):1485-1489. (in Chinese)
	( 刘兆宇, 李学武, 周月, 等. 加热灭活病毒法对生化及炎症指标检测结果的影响[J]. 中华医院感染学杂志, 2020,30(10):1485-1489.)

血样	检测血样数	T1阳性血样数（占比/%）		χ²	P	T2阳性血样数（占比/%）		χ²	P	合计阳性血样数（占比/%）		χ²	P
血样	检测血样数	热处理前	热处理后	χ²	P	热处理前	热处理后	χ²	P	热处理前	热处理后	χ²	P
恶性疟	38	38(100)^a	35(92.1)	3.123	> 0.05	26(68.4)	0(0)	39.520	< 0.01	38(100)	35(92.1)	3.123	> 0.05
三日疟	8	0(0)	0(0)	-	-	6(6/8)	0(0)	9.600	< 0.01	6(6/8)	0(0)	9.600	< 0.01
卵形疟	11	0(0)	0(0)	-	-	10(10/11)	0(0)	7.619	< 0.01	10(10/11)	0(0)	7.619	< 0.01
间日疟	14	0(0)	0(0)	-	-	12(12/14)	0(0)	9.231	< 0.01	12(12/14)	0(0)	9.231	< 0.01
合计	71	38(50)	35(46.0)	0.237	> 0.05	54(71.0)	0(0)	83.755	< 0.01	66(86.8)	35(46.0)	16.189	< 0.01

快速诊断靶蛋白恶性疟原虫富组氨酸蛋白Ⅱ和疟原虫乳酸脱氢酶的热稳定性

Thermal stability of diagnostic targets Plasmodium falciparum histidine rich protein Ⅱ and Plasmodium lactate dehydrogenase in rapid detection

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 1

参考文献 20

相关文章 10

编辑推荐

Metrics

本文评价

[1]	杨英超，张瑾，王国柱，薄淑英，张影，辛晓芳*. 恶性疟原虫组氨酸富集蛋白Ⅱ、Ⅲ的序列多态性分析[J]. 中国寄生虫学与寄生虫病杂志, 2014, 32(5): 10-373-376.
[2]	刘荣珍,石裕明. 应用猴抗食蟹猴疟原虫抗体检测间日疟抗原[J]. 中国寄生虫学与寄生虫病杂志, 1993, 11(4): 307-307.
[3]	刘克义,高金桐. 固相放射免疫分析检测食蟹猴疟原虫抗原[J]. 中国寄生虫学与寄生虫病杂志, 1989, 7(2): 157-158.
[4]	高琪,杨仲炎,张炜琪,杨存性. 用单克隆抗体——酶联免疫吸附试验双抗体夹心法检测人体疟原虫抗原的研究[J]. 中国寄生虫学与寄生虫病杂志, 1988, 6(S1): 13-14.
[5]	王秀珍,黎世涛,周肇西. 酶标记抗原斑点试验检测间日疟抗体的研究[J]. 中国寄生虫学与寄生虫病杂志, 1988, 6(S1): 13-13.
[6]	杨存性,高琪,王祥荣,张伟琪,罗国湘. 用单克隆抗体检测疟原虫抗原的研究[J]. 中国寄生虫学与寄生虫病杂志, 1987, 5(4): 248-250.
[7]	黄文洲,罗曼珍,周铭贤,陈辅群,宣圣珍. 间接荧光抗体试验使用两种不同疟原虫抗原进行疟疾反复横向调查[J]. 中国寄生虫学与寄生虫病杂志, 1987, 5(2): 118-120.
[8]	肖贤琦,陈昌源,裴速建. 用间接荧光抗体试验及猴疟抗原查出一例三日疟供血者[J]. 中国寄生虫学与寄生虫病杂志, 1985, 3(3): 180-180.
[9]	黄文洲,罗曼珍,周铭贤,陈辅群,宣圣珍. 间接荧光抗体试验使用两种不同疟原虫抗原进行疟疾单次横向调查[J]. 中国寄生虫学与寄生虫病杂志, 1985, 3(3): 181-183.
[10]	季始荣,王克泰,张志良,张筑安,吴怀君,王龙英. 以体外培养的恶性疟原虫为抗原作间接荧光抗体试验检测间日疟[J]. 中国寄生虫学与寄生虫病杂志, 1983, 1(4): 5-5.