荧光定量PCR在疟疾实验室诊断中的应用
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摘要
分别采用镜检、巢式PCR和荧光定量PCR等3种检测方法对收集到的79份疟疾血样进行检测,并对检测结果进行比较分析。结合患者临床症状和3种检测结果, 79份血样最终确诊74份为疟原虫阳性,阳性率为93.7%(74/79),其中镜检阳性检出率为82.3%(65/79),巢式PCR阳性检出率为82.3%(65/79),荧光定量PCR阳性检出率为93.7%(74/79),荧光定量PCR阳性检出率高于其他两种方法(P <0.05)。镜检和巢式PCR的一致率为78.4%(58/74),镜检和荧光定量PCR的一致率为63.5%(47/74),巢式PCR和荧光定量PCR的一致率为83.8%(62/74)。相比镜检和巢式PCR,荧光定量PCR敏感性和检出率更高,且用时更短。
Seventy-nine malaria samples were examined by microscopy, nested PCR and fluorescence quantitative PCR, and the detection rate was analyzed among the three methods. After combining clinical symptoms of the patients and results of the three methods, 74 of the 79 samples were confirmed to be malaria infection, with a positive rate of 93.7%( 74/79). Specifically, the detection rate of microscopy, nested PCR and fluorescence quantitative PCR were 82.3%(65/79), 82.3%65/79) and 93.7%(74/79), respectively. The concordance rate between microscopy and nested PCR was 78.4%(58/74), and that between microscopy and fluorescence quantitative PCR was 63.5%( 47/74), between nested PCR and fluorescence quantitative PCR was 83.8%( 62/74). In addition, the detection rate of fluorescence quantitative PCR was significantly higher than that of the nested PCR(P < 0.05).Compared with microscopy and nested PCR, the fluorescence quantitative PCR has higher sensitivity and positive detection rate, and is less time-consuming.
Seventy-nine malaria samples were examined by microscopy, nested PCR and fluorescence quantitative PCR, and the detection rate was analyzed among the three methods. After combining clinical symptoms of the patients and results of the three methods, 74 of the 79 samples were confirmed to be malaria infection, with a positive rate of 93.7%( 74/79). Specifically, the detection rate of microscopy, nested PCR and fluorescence quantitative PCR were 82.3%(65/79), 82.3%65/79) and 93.7%(74/79), respectively. The concordance rate between microscopy and nested PCR was 78.4%(58/74), and that between microscopy and fluorescence quantitative PCR was 63.5%( 47/74), between nested PCR and fluorescence quantitative PCR was 83.8%( 62/74). In addition, the detection rate of fluorescence quantitative PCR was significantly higher than that of the nested PCR(P < 0.05).Compared with microscopy and nested PCR, the fluorescence quantitative PCR has higher sensitivity and positive detection rate, and is less time-consuming.
引文
[1] Lee PC, Chong ET, Anderios F, et al. Molecular detection of human Plasmodium species in Sabah using PlasmoNexTM multiplex PCR and hydrolysis probes real-time PCR[J]. Malar J, 2015, 14:28.
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[4]李美,夏志贵,汤林华.检测4种人体疟原虫多重PCR体系的建立和应用[J].中国寄生虫学与寄生虫病杂志, 2015, 33(2):91-95.
[5]黄雨婷,佘丹娅,卢丽丹,等.单管单轮多重PCR检测4种疟原虫混合血样[J].中国寄生虫学与寄生虫病杂志, 2015, 33(3):200-205.
[6]李欢. SYBR Green实时PCR检测恶性疟原虫与间日疟原虫方法的建立[D].长沙:中南大学, 2014.
[7]张金飞,周燕珍.三种不同检测方法在疟疾诊断中的应用比较[J].中国地方病防治杂志, 2016, 31(2):201-202.
[8]雷露,刘学升,毛玲玲,等. 3种实验室检测方法在疟疾诊断中的应用比较[J].中国媒介生物学及控制杂志, 2015, 26(2):206-207.
[9] Barker RH Jr, Banchongaksorn T, Courval JM, et al. A simple method to detect Plasmodium falciparum directly from blood samples using the polymerase chain reaction[J]. Am J Trop Med Hyg, 1992, 46(4):416-426.
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[11]周瑞敏,刘颖,钱丹,等. 2012年河南省疟疾标本的实验室检测分析[J].中国病原生物学杂志, 2014, 9(2):177-179.
[12]鲁少平,邓中平,龙妍娇,等.运用实时荧光RT-PCR方法检测疟原虫的研究[J].实用预防医学, 2015, 22(10):1171-1173.
[13]焦炳欣,华文浩,陈志海,等.三种检测方法在疟疾诊断和疗效中的应用评估[J].中国医药导报, 2012, 9(270):98-99.
[2] Koukouikila-Koussounda F, Malonga V, Mayengue PI, et al.Genetic polymorphism of merozoite surface protein 2 and prevalence of K76T pfcrt mutation in Plasmodium falciparum field isolates from Congolese children with asymptomatic infections[J]. Malar J, 2012, 11:105.
[3]孙菲,万向阳,李欢,等.多重巢式PCR检测恶性疟和间日疟原虫[J].临床检验杂志, 2014, 32(8):565-567.
[4]李美,夏志贵,汤林华.检测4种人体疟原虫多重PCR体系的建立和应用[J].中国寄生虫学与寄生虫病杂志, 2015, 33(2):91-95.
[5]黄雨婷,佘丹娅,卢丽丹,等.单管单轮多重PCR检测4种疟原虫混合血样[J].中国寄生虫学与寄生虫病杂志, 2015, 33(3):200-205.
[6]李欢. SYBR Green实时PCR检测恶性疟原虫与间日疟原虫方法的建立[D].长沙:中南大学, 2014.
[7]张金飞,周燕珍.三种不同检测方法在疟疾诊断中的应用比较[J].中国地方病防治杂志, 2016, 31(2):201-202.
[8]雷露,刘学升,毛玲玲,等. 3种实验室检测方法在疟疾诊断中的应用比较[J].中国媒介生物学及控制杂志, 2015, 26(2):206-207.
[9] Barker RH Jr, Banchongaksorn T, Courval JM, et al. A simple method to detect Plasmodium falciparum directly from blood samples using the polymerase chain reaction[J]. Am J Trop Med Hyg, 1992, 46(4):416-426.
[10] Win TT, Jalloh A, Tantular IS, et al. Molecular analysis of Plasmodium ovale variants[J]. Emerging Infect Dis, 2004, 10(7):1235-1240.
[11]周瑞敏,刘颖,钱丹,等. 2012年河南省疟疾标本的实验室检测分析[J].中国病原生物学杂志, 2014, 9(2):177-179.
[12]鲁少平,邓中平,龙妍娇,等.运用实时荧光RT-PCR方法检测疟原虫的研究[J].实用预防医学, 2015, 22(10):1171-1173.
[13]焦炳欣,华文浩,陈志海,等.三种检测方法在疟疾诊断和疗效中的应用评估[J].中国医药导报, 2012, 9(270):98-99.
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