Nucleic acid test to diagnose cryptosporidiosis: lab assessment in animal and patient specimens.
Nucleic acid test to diagnose cryptosporidiosis: lab assessment in animal and patient specimens.
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DOI：
10.1021/ac403750z
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发表时间：
2014-03-04
影响因子：
7.4
通讯作者：
Richards-Kortum, Rebecca
中科院分区：
文献类型：
--
作者：
Crannell, Zachary Austin;Castellanos-Gonzalez, Alejandro;Irani, Ayesha;Rohrman, Brittany;White, Arthur Clinton;Richards-Kortum, Rebecca
腹泻病在世界范围内造成的发病率和死亡率高于人类免疫缺陷病毒（HIV）、疟疾或结核病。鉴于有效治疗持续性腹泻病需要了解致病微生物，诊断测试是至关重要的。隐孢子虫属的原生动物寄生虫越来越被认为是腹泻发病率的重要组成部分。我们提出了一种新的核酸检测从粪便样本中提取的DNA中的隐孢子虫物种的存在。该检测使用等温扩增技术重组酶聚合酶扩增（RPA），在30分钟内将从粪便中提取的痕量病原体DNA扩增至可检测水平;然后在简单的侧向流条上目视检测产物。使用来自加标病原体的人粪便样品的DNA开发并优化了基于RPA的隐孢子虫检测（RPAC检测）。然后使用从受感染小鼠的粪便中提取的DNA进行测试，其中它正确地鉴定了28个粪便样本中27个的存在或不存在。最后，使用从感染患者粪便中提取的DNA进行测试，在21个粪便样本中，它正确识别了21个样本的存在或不存在。该测定被整合到一个可折叠的纸和塑料装置中，该装置仅使用移液管、移液管尖端和加热器就能进行DNA扩增。集成检测的性能与聚合酶链反应（PCR）相当或更好，无需使用热循环设备。该平台可以很容易地适用于检测来自多种病原体的DNA。
Diarrheal diseases cause more morbidity and mortality around the world than human immunodeficiency virus (HIV), malaria, or tuberculosis. Given that effective treatment of persistent diarrheal illness requires knowledge of the causative organism, diagnostic tests are of paramount importance. The protozoan parasites of the genus Cryptosporidium are increasingly recognized to be responsible for a significant portion of diarrhea morbidity. We present a novel nucleic acid test to detect the presence of Cryptosporidium species in DNA extracted from stool samples. The assay uses the isothermal amplification technique recombinase polymerase amplification (RPA) to amplify trace amounts of pathogen DNA extracted from stool to detectable levels in 30 min; products are then detected visually on simple lateral flow strips. The RPA-based Cryptosporidium assay (RPAC assay) was developed and optimized using DNA from human stool samples spiked with pathogen. It was then tested using DNA extracted from the stool of infected mice where it correctly identified the presence or absence of 27 out of 28 stool samples. It was finally tested using DNA extracted from the stool of infected patients where it correctly identified the presence or absence of 21 out of 21 stool samples. The assay was integrated into a foldable, paper and plastic device that enables DNA amplification with only the use of pipets, pipet tips, and a heater. The performance of the integrated assay is comparable to or better than polymerase chain reaction (PCR), without requiring the use of thermal cycling equipment. This platform can easily be adapted to detect DNA from multiple pathogens.
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影响因子：
9.4
作者：
Agnamey, Patrice;Sarfati, Claudine;Derouin, F.
通讯作者：
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影响因子：
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作者：
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DOI：
10.4269/ajtmh.2007.76.713
发表时间：
2007-04-01
影响因子：
3.3
作者：
Haque, Rashidul;Roy, Shantanu;Petri, William A., Jr.
通讯作者：
Petri, William A., Jr.
影响因子：
6.1
作者：
Govindarajan, A. V.;Ramachandran, S.;Boehringer, K. F.
通讯作者：
Boehringer, K. F.
DOI：
10.4269/ajtmh.2009.80.824
发表时间：
2009-05-01
影响因子：
3.3
作者：
Mondal, Dinesh;Haque, Rashidul;Petri, William A., Jr.
通讯作者：
Petri, William A., Jr.