Culture-Independent Detection and Identification of Leptospira Serovars.
Culture-Independent Detection and Identification of Leptospira Serovars.
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DOI：
10.1128/spectrum.02475-22
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发表时间：
2022-12-21
影响因子：
3.7
通讯作者：
中科院分区：
文献类型：
--
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致病性钩端螺旋体是钩端螺旋体病的病原体，包括> 200种血清型（称为血清型）。大多数有一个有限的寄主范围，和一些，例如，Copenhageni血清型是世界性的，并且由于它们倾向于在人类中产生严重的、致命的疾病而具有公共卫生重要性。现有的血清分型方法，如多位点序列分型，核心基因组序列分型，脉冲场凝胶电泳，交叉凝集吸收试验是繁琐和昂贵的，并需要在培养基中分离的生物体，一个漫长的和令人难以置信的效率低下的过程，排除了他们在前瞻性研究或疫情调查中的使用。无法获得能够区分钩端螺旋体血清型的不依赖培养的测定方法，这仍然是该领域的一个关键空白。在这里，我们已经开发了一种简单而特异的实时qPCR测定法-靶向钩端螺旋体-编码推定的多糖翻转酶的独特基因-其提供种内、型别定义（即，流行病学上有用的）信息，并且改进了优选的基于lipL32的基于qPCR的诊断测试的灵敏度。被称为RAgI（"rage one"）的检测方法快速且经济实惠，并且可靠且特异地检测培养物、血清和尿液中的I组致病性钩端螺旋体，即使是遗传相关但低毒力的II组致病性（以前为"中间"）或非致病性钩端螺旋体也没有可检测的脱靶扩增。它保留了100%的诊断特异性，当测试困难的样品类型，包括现场收集的狗尿样和环境样品含有各种复杂的微生物物种财团。该检测在临床环境中以及常规流行病学和环境监测研究中具有相当大的前景。钩端螺旋体病是由一组不同的致病螺旋体引起的，包括200多种不同的血清型。其中一些已被广泛报道，由于它们容易在人类中产生严重、致命的疾病，因此具有公共卫生重要性。除了繁琐和昂贵之外，目前的血清分型方法还需要在培养基中分离微生物这是一个旷日持久且效率极低的过程，这使得它们在临床上毫无用处，并限制了它们在前瞻性研究或疫情调查中的应用。无法获得能够区分钩端螺旋体血清型的不依赖培养的测定方法，这仍然是该领域的一个关键空白。11108 qPCR检测通过对尿液和血清样本中的钩端螺旋体进行直接分类和血清型分类，克服了这一进展障碍，因此是首个基于qPCR的人类钩端螺旋体病预后检测。
Pathogenic Leptospira, the causative agents of leptospirosis, comprise >200 serotypes (called serovars). Most have a restricted reservoir-host range, and some, e.g., serovar Copenhageni, are cosmopolitan and of public health importance owing to their propensity to produce severe, fatal disease in humans. Available serotyping approaches—such as multilocus sequence typing, core genome sequence typing, pulsed-field gel electrophoresis, and the cross-agglutination absorption test—are tedious and expensive, and require isolation of the organisms in culture media—a protracted and incredibly inefficient process—precluding their use in prospective studies or outbreak investigations. The unavailability of culture-independent assays capable of distinguishing Leptospira serotypes remains a crucial gap in the field. Here, we have developed a simple yet specific real-time qPCR assay—targeting a Leptospira-unique gene encoding a putative polysaccharide flippase—that provides intraspecies, serotype-defining (i.e., epidemiologically useful) information, and improves upon the sensitivity of preferred lipL32-based qPCR-based diagnostic tests. The assay, dubbed RAgI (“rage one”), is rapid and affordable, and reliably and specifically detects group I pathogenic Leptospira in culture, serum, and urine, with no detectable off-target amplification—even of the genetically related but low virulence group II pathogenic (formerly “intermediate”) or nonpathogenic Leptospira. It retained 100% diagnostic specificity when tested against difficult sample types, including field-collected dog urine samples and environmental samples containing varied and complex microbial species-consortia. This assay holds considerable promise in the clinical setting, and for routine epidemiological and environmental surveillance studies. IMPORTANCE Leptospirosis is caused by a diverse group of pathogenic spirochetes comprising over 200 different serotypes. Some are widely reported and of public health importance owing to their propensity to produce severe, fatal disease in humans. Apart from their tedium and expense, current serotyping approaches require isolation of the organisms in culture media—a protracted and incredibly inefficient process—rendering them useless clinically and limiting their utilization in prospective studies or outbreak investigations. The unavailability of culture-independent assays capable of distinguishing Leptospira serotypes remains a crucial gap in the field. The 11108 qPCR-assay overcomes this barrier to progress via direct taxonomic and serotype classification of Leptospira from urine and serum samples, and hence, is the first qPCR-based prognostic test for human leptospirosis.
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影响因子：
3.8
作者：
Jayasundara D;Senavirathna I;Warnasekara J;Gamage C;Siribaddana S;Kularatne SAM;Matthias M;Mariet JF;Picardeau M;Agampodi S;M Vinetz J
通讯作者：
M Vinetz J
影响因子：
3.7
作者：
Ahmed AA;Goris MGA;Meijer MC
通讯作者：
Meijer MC
影响因子：
3.2
作者：
Caimi, K.;Repetto, S. A.;Ruybal, P.
通讯作者：
Ruybal, P.
影响因子：
3.7
作者：
Naze F;Desvars A;Picardeau M;Bourhy P;Michault A
通讯作者：
Michault A
DOI：
10.1016/j.diagmicrobio2015.03.024
发表时间：
2015-07-01
影响因子：
2.9
作者：
Galloway, Renee L.;Hoffmaster, Alex R.
通讯作者：
Hoffmaster, Alex R.