Rapid Isolation and Susceptibility Testing of Leptospira spp. Using a New Solid Medium, LVW Agar
Rapid Isolation and Susceptibility Testing of Leptospira spp. Using a New Solid Medium, LVW Agar
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DOI：
10.1128/aac.01812-12
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发表时间：
2013-01-01
影响因子：
4.9
通讯作者：
Peacock, Sharon J.
中科院分区：
文献类型：
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作者：
Wuthiekanun, Vanaporn;Amornchai, Premjit;Peacock, Sharon J.
致病性钩端螺旋体属，钩端螺旋体病的病原体是生长缓慢的革兰氏阴性螺旋体。从临床样本中分离钩端螺旋体并进行抗菌药物敏感性测试困难且耗时。在这里，我们描述了一种新的固体培养基，有利于更快速的增长钩端螺旋体的发展。以及使用该培养基评价Etest测定钩端螺旋体病常用药物的抗菌MIC的性能。通过评估多种因素对问号钩端螺旋体NR-20157菌株生长速率的影响来开发培养基。这些包括基础琼脂的类型，兔血清（RS）的浓度，以及在培养的初始阶段的CO2孵育的浓度和持续时间。NR-20157的最高生长速率是使用补充有10% RS的Noble琼脂基质（称为LVW琼脂），在30 ℃下在5% CO2中初始孵育2天，然后在30 ℃下在空气中连续培养。这些条件被用来开发三个物种，L。问号线虫（NR-20161）、L. kirschnerii（NR-20327）和L. borgpetersenii（NR-20151）。在第7天读取所有样品的MIC。然后对109株致病性钩端螺旋体进行Etest。青霉素G、强力霉素、头孢噻肟、头孢曲松和氯霉素的MIC 90值分别为0.64单位/ml和0.19、0.047、0.5和2 μ g/ml。LVW琼脂的使用能够快速生长、分离单菌落和进行钩端螺旋体属的简单抗菌药物敏感性测试，为基础和应用研究的新领域提供了机会。
Pathogenic Leptospira spp., the causative agents of leptospirosis, are slow-growing Gram-negative spirochetes. Isolation of Leptospira from clinical samples and testing of antimicrobial susceptibility are difficult and time-consuming. Here, we describe the development of a new solid medium that facilitates more-rapid growth of Leptospira spp. and the use of this medium to evaluate the Etest's performance in determining antimicrobial MICs to drugs in common use for leptospirosis. The medium was developed by evaluating the effects of numerous factors on the growth rate of Leptospira interrogans strain NR-20157. These included the type of base agar, the concentration of rabbit serum (RS), and the concentration and duration of CO2 incubation during the initial period of culture. The highest growth rate of NR-20157 was achieved using a Noble agar base supplemented with 10% RS (named LVW agar), with an initial incubation at 30 degrees C in 5% CO2 for 2 days prior to continuous culture in air at 30 degrees C. These conditions were used to develop the Etest for three species, L. interrogans (NR-20161), L. kirschnerii (NR-20327), and L. borgpetersenii (NR-20151). The MICs were read on day 7 for all samples. The Etest was then performed on 109 isolates of pathogenic Leptospira spp. The MIC90 values for penicillin G, doxycycline, cefotaxime, ceftriaxone, and chloramphenicol were 0.64 units/ml and 0.19, 0.047, 0.5, and 2 mu g/ml, respectively. The use of LVW agar, which enables rapid growth, isolation of single colonies, and simple antimicrobial susceptibility testing for Leptospira spp., provides an opportunity for new areas of fundamental and applied research.