Gökmen Tülin Güven, Soyal Ayben, Kalayci Yıldız, Önlen Cansu, Köksal Fatih
Cukurova University, Ceyhan Veterinary Faculty, Department of Microbiology. 01330, Adana, Turkey. E-mail:
Cukurova University, Medical Faculty, Department of Microbiology. 01330 Adana, Turkey. E-mails:
Rev Inst Med Trop Sao Paulo. 2016 Sep 22;58:64. doi: 10.1590/S1678-9946201658064.
Leptospirosis is still one of the most important health problems in developing countries located in humid tropical and subtropical regions. Human infections are generally caused by exposure to water, soil or food contaminated with the urine of infected wild and domestic animals such as rodents and dogs. The clinical course of leptospirosis is variable and may be difficult to distinguish from many other infectious diseases. The dark-field microscopy (DFM), serology and nucleic acid amplification techniques are used to diagnose leptospirosis, however, a distinctive standard reference method is still lacking. Therefore, in this study, we aimed to determine the presence of Leptospira spp., to differentiate the pathogenic L. interrogans and the non-pathogenic L. biflexa, and also to determine the sensitivity and specificity values of molecular methods as an alternative to conventional ones. A total of 133 serum samples, from 47 humans and 86 cattle were evaluated by two conventional tests: the Microagglutination Test (MAT) and the DFM, as well as three molecular methods, the 16S rRNA-PCR followed by Restriction Fragment Lenght Polymorphism (RFLP) of the amplification products 16S rRNA-PCR-RFLP, LipL32-PCR and OmpL1-PCR. In this study, for L. interrogans, the specificity and sensitivity rates of the 16S rRNA-PCR and the LipL32-PCR were considered similar (100% versus 98.25% and 100% versus 98.68%, respectively). The OmpL1-PCR was able to classify L. interrogans into two intergroups, but this PCR was less sensitive (87.01%) than the other two PCR methods. The 16S rRNA-PCR-RFLP could detect L. biflexa DNA, but LipL32-PCR and OmpL1-PCR could not. The 16S rRNA-PCR-RFLP provided an early and accurate diagnosis and was able to distinguish pathogenic and non-pathogenic Leptospira species, hence it may be used as an alternative method to the conventional gold standard techniques for the rapid disgnosis of leptospirosis.
钩端螺旋体病仍然是位于潮湿热带和亚热带地区的发展中国家最重要的健康问题之一。人类感染通常是由于接触被感染的野生动物和家畜（如啮齿动物和狗）尿液污染的水、土壤或食物所致。钩端螺旋体病的临床病程多变，可能难以与许多其他传染病区分开来。暗视野显微镜检查（DFM）、血清学和核酸扩增技术用于诊断钩端螺旋体病，然而，仍然缺乏独特的标准参考方法。因此，在本研究中，我们旨在确定钩端螺旋体属的存在，区分致病性问号钩端螺旋体和非致病性双曲钩端螺旋体，并确定分子方法作为传统方法替代方法的敏感性和特异性值。通过两种传统检测方法：显微凝集试验（MAT）和DFM，以及三种分子方法，即16S rRNA-PCR，随后对扩增产物进行限制性片段长度多态性分析（RFLP）（16S rRNA-PCR-RFLP）、LipL32-PCR和OmpL1-PCR，对来自47名人类和86头牛的总共133份血清样本进行了评估。在本研究中，对于问号钩端螺旋体，16S rRNA-PCR和LipL32-PCR的特异性和敏感性率被认为相似（分别为100%对98.25%和100%对98. 68%）。OmpL1-PCR能够将问号钩端螺旋体分为两个组间，但该PCR的敏感性（87.01%）低于其他两种PCR方法。16S rRNA-PCR-RFLP可以检测到双曲钩端螺旋体DNA，但LipL32-PCR和OmpL1-PCR不能。16S rRNA-PCR-RFLP提供了早期准确的诊断，能够区分致病性和非致病性钩端螺旋体物种，因此它可以用作传统金标准技术的替代方法，用于快速诊断钩端螺旋体病。
