检查 大多数针对锥虫亚属的引物具有特异性并且源于卫星序列或多拷贝基因,检测限值为 1-40 个锥虫/mL ,且敏感性较高。[99]Moser DR, Cook GA, Ochs DE, et al. Detection of Trypanosoma congolense and Trypanosoma brucei subspecies by DNA amplification using the polymerase chain reaction. Parasitology. 1989 Aug;99 Pt 1:57-66.http://www.ncbi.nlm.nih.gov/pubmed/2797872?tool=bestpractice.com[100]Becker S, Franco JR, Simarro PP, et al. Real-time PCR for detection of Trypanosoma brucei in human blood samples. Diagn Microbiol Infect Dis. 2004 Nov;50(3):193-9.http://www.ncbi.nlm.nih.gov/pubmed/15541605?tool=bestpractice.com[101]Kabiri M, Franco JR, Simarro PP, et al. Detection of Trypanosoma brucei gambiense in sleeping sickness suspects by PCR amplification of expression-site-associated genes 6 and 7. Trop Med Int Health. 1999 Oct;4(10):658-61.http://onlinelibrary.wiley.com/doi/10.1046/j.1365-3156.1999.00465.x/fullhttp://www.ncbi.nlm.nih.gov/pubmed/10583899?tool=bestpractice.com[102]Deborggraeve S, Claes F, Laurent T, et al. Molecular dipstick test for diagnosis of sleeping sickness. J Clin Microbiol. 2006 Aug;44(8):2884-9.http://jcm.asm.org/content/44/8/2884.fullhttp://www.ncbi.nlm.nih.gov/pubmed/16891507?tool=bestpractice.com 诊断特异性达到 97% 或以上,但这些聚合酶链反应无法区分冈比亚锥虫和罗得西亚锥虫感染,并且阳性反应可能与布氏锥虫、伊氏锥虫和马疫锥虫有关。
冈比亚锥虫和罗得西亚锥虫感染之间的差异可以通过使用适用于罗得西亚锥虫病的抗血清相关基因 (serum resistance-associated gene, SRA) 检测,[103]Radwanska M, Chamekh M, Vanhamme L, et al. The serum resistance-associated gene as a diagnostic tool for the detection of Trypanosoma brucei rhodesiense. Am J Trop Med Hyg. 2002 Dec;67(6):684-90.http://www.ncbi.nlm.nih.gov/pubmed/12518862?tool=bestpractice.com[104]Welburn SC, Picozzi K, Fevre EM, et al. Identification of human-infective trypanosomes in animal reservoir of sleeping sickness in Uganda by means of serum-resistance-associated (SRA) gene. Lancet. 2001 Dec 15;358(9298):2017-9.http://www.ncbi.nlm.nih.gov/pubmed/11755607?tool=bestpractice.com[105]Picozzi K, Fèvre EM, Odiit M, et al. Sleeping sickness in Uganda: a thin line between two fatal diseases. BMJ. 2005 Nov 26;331(7527):1238-41.http://www.bmj.com/content/331/7527/1238.fullhttp://www.ncbi.nlm.nih.gov/pubmed/16308383?tool=bestpractice.com 以及可使用冈比亚锥虫病特异性糖蛋白 (TgsGP) 基因检测进行冈比亚锥虫病的检测。[105]Picozzi K, Fèvre EM, Odiit M, et al. Sleeping sickness in Uganda: a thin line between two fatal diseases. BMJ. 2005 Nov 26;331(7527):1238-41.http://www.bmj.com/content/331/7527/1238.fullhttp://www.ncbi.nlm.nih.gov/pubmed/16308383?tool=bestpractice.com[106]Radwanska M, Claes F, Magez S, et al. Novel primer sequences for polymerase chain reaction-based detection of Trypanosoma brucei gambiense. Am J Trop Med Hyg. 2002 Sep;67(3):289-95.http://www.ajtmh.org/content/journals/10.4269/ajtmh.2002.67.289#html_fulltexthttp://www.ncbi.nlm.nih.gov/pubmed/12408669?tool=bestpractice.com 这些单拷贝基因检测的敏感性较低。当前缺乏标准化和比较研究。脑脊液聚合酶链反应阳性的分期显著性依然不明。[107]Jamonneau V, Solano P, Garcia A, et al. Stage determination and therapeutic decision in human African trypanosomiasis: value of PCR and IgM quantification on the cerebrospinal fluid of sleeping sickness patients in Côte d'Ivoire. Trop Med Int Health. 2003 Jul;8(7):589-94.http://onlinelibrary.wiley.com/doi/10.1046/j.1365-3156.2003.01079.x/fullhttp://www.ncbi.nlm.nih.gov/pubmed/12828540?tool=bestpractice.com对于治疗结局的评估,经证明 PCR 是不可靠的。[108]Kirchhoff LV. Use of a PCR assay for diagnosing African trypanosomiasis of the CNS: a case report. Cent Afr J Med. 1998 May;44(5):134-6.http://www.ncbi.nlm.nih.gov/pubmed/9810412?tool=bestpractice.com[109]Truc P, Jamonneau V, Cuny G, et al. Use of polymerase chain reaction in human African trypanosomiasis stage determination and follow-up. Bull World Health Organ. 1999;77(9):745-8.http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2557728/pdf/10534898.pdfhttp://www.ncbi.nlm.nih.gov/pubmed/10534898?tool=bestpractice.com[110]Deborggraeve S, Lejon V, Ekangu RA, et al. Diagnostic accuracy of PCR in gambiense sleeping sickness diagnosis, staging and post-treatment follow-up: a 2-year longitudinal study. PLoS Negl Trop Dis. 2011 Feb 22;5(2):e972http://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0000972http://www.ncbi.nlm.nih.gov/pubmed/21364966?tool=bestpractice.com
更新的分子技术包括环介导等温扩增 (loop-mediated isothermal amplification, LAMP)、依赖核酸序列扩增 (NASBA)、寡核苷酸色谱分析法和逆转录酶实时聚合酶链反应,可用于检测锥虫亚属特异性剪接前导序列 RNA,[102]Deborggraeve S, Claes F, Laurent T, et al. Molecular dipstick test for diagnosis of sleeping sickness. J Clin Microbiol. 2006 Aug;44(8):2884-9.http://jcm.asm.org/content/44/8/2884.fullhttp://www.ncbi.nlm.nih.gov/pubmed/16891507?tool=bestpractice.com[111]Kuboki N, Inoue N, Sakurai T, et al. Loop-mediated isothermal amplification for detection of African trypanosomes. J Clin Microbiol. 2003 Dec;41(12):5517-24.http://jcm.asm.org/content/41/12/5517.fullhttp://www.ncbi.nlm.nih.gov/pubmed/14662933?tool=bestpractice.com[112]Njiru ZK, Mikosza AS, Matovu E, et al. African trypanosomiasis: sensitive and rapid detection of the sub-genus Trypanozoon by loop-mediated isothermal amplification (LAMP) of parasite DNA. Int J Parasitol. 2008 Apr;38(5):589-99.http://www.ncbi.nlm.nih.gov/pubmed/17991469?tool=bestpractice.com[113]Mugasa CM, Schoone GJ, Ekangu RA, et al. Detection of Trypanosoma brucei parasites in blood samples using real-time nucleic acid sequence-based amplification. Diagn Microbiol Infect Dis. 2008 Aug;61(4):440-5.http://www.ncbi.nlm.nih.gov/pubmed/18486402?tool=bestpractice.com[114]Matovu E, Mugasa CM, Ekangu RA, et al. Phase II evaluation of sensitivity and specificity of PCR and NASBA followed by oligochromatography for diagnosis of human African trypanosomiasis in clinical samples from D.R. Congo and Uganda. PLoS Negl Trop Dis. 2010 Jul 6;4(7):e737.http://www.ncbi.nlm.nih.gov/pubmed/20625557?tool=bestpractice.com[115]Njiru ZK, Traub R, Ouma JO, et al. Detection of group 1 trypanosoma brucei gambiense by loop-mediated isothermal amplification. J Clin Microbiol. 2011 Apr;49(4):1530-6.http://jcm.asm.org/content/49/4/1530.longhttp://www.ncbi.nlm.nih.gov/pubmed/21307218?tool=bestpractice.com[116]Wastling SL, Picozzi K, Kakembo AS, et al. LAMP for human African trypanosomiasis: a comparative study of detection formats. PLoS Negl Trop Dis. 2010 Nov 2;4(11):e865.http://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0000865http://www.ncbi.nlm.nih.gov/pubmed/21072228?tool=bestpractice.com[117]González-Andrade P, Camara M, Ilboudo H, et al. Diagnosis of trypanosomatid infections: targeting the spliced leader RNA. J Mol Diagn. 2014 Jul;16(4):400-4.http://www.ncbi.nlm.nih.gov/pubmed/24814957?tool=bestpractice.com 但需要适当的标准化和验证。[118]Mugasa CM, Adams ER, Boer KR, et al. Diagnostic accuracy of molecular amplification tests for human African trypanosomiasis - systematic review. PLoS Negl Trop Dis. 2012 Jan;6(1):e1438.http://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0001438http://www.ncbi.nlm.nih.gov/pubmed/22253934?tool=bestpractice.com[119]Deborggraeve S, Büscher P. Molecular diagnostics for sleeping sickness: what is the benefit for the patient? Lancet Infect Dis. 2010 Jun;10(6):433-9.http://www.ncbi.nlm.nih.gov/pubmed/20510283?tool=bestpractice.com[120]Deborggraeve S, Büscher P. Recent progress in molecular diagnosis of sleeping sickness. Expert Rev Mol Diagn. 2012 Sep;12(7):719-30.http://www.ncbi.nlm.nih.gov/pubmed/23153239?tool=bestpractice.com 在首次评估冈比亚锥虫型人类非洲锥虫病的临床样本时,Loopamp 布氏锥虫检测试剂盒 (Loopamp Trypanosoma brucei Detection Kit)(品牌名称)的检测效果(根据锥虫亚属重复插入可移动 DNA 元件的检测结果)与常规聚合酶链反应相似。[121]Mitashi P, Hasker E, Mumba Ngoyi D, et al. Diagnostic accuracy of Loopamp Trypanosoma brucei detection kit for diagnosis of human African trypanosomiasis in clinical samples. PLos Negl Trop Dis. 2013 Oct 17;7(10):e2504.http://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0002504http://www.ncbi.nlm.nih.gov/pubmed/24147176?tool=bestpractice.com
聚合酶链反应或环介导等温扩增反应阳性表示患有人类非洲锥虫病,但是应进行进一步的寄生虫学检查,以便确诊。