利什曼原虫属对抗两性霉素 B 和氟康唑的敏感性检测，使用 Sensititre™ YeastOne™ YO9 平台。
Susceptibility testing of Leishmania spp. against amphotericin B and fluconazole using the Sensititre™ YeastOne™ YO9 platform.
机构信息
Institute of Medical Sciences, University of Toronto, Address: 1 King's College Circle, Toronto, Ontario, M5S 1A8, Canada.
Faculty of Arts and Science, University of Toronto, Address: 100 St. George Street, Toronto, Ontario, M5S 3G3, Canada.
出版信息
BMC Infect Dis. 2019 Jul 8;19(1):593. doi: 10.1186/s12879-019-4237-3.
BACKGROUND
Current drug regimens for cutaneous leishmaniasis (CL) include toxic systemic therapies such as amphotericin B (AB) and pentavalent antimonials. Fluconazole (FZ) is a well-tolerated potential oral alternative for the management CL. To date, few objective data exist to guide clinical decision-making when selecting a therapeutic agent a priori, and standardized, clinically-approved drug susceptibility testing platforms for Leishmania spp. have yet to be established. The Sensititre™ YeastOne™ YO9 plate is a commercialized drug susceptibility plate including AB and FZ used for routine testing of non-fastidious yeast. Our objective was to adapt the readily available Sensititre™ YeastOne™ YO9 plate, to determine drug susceptibility profiles of AB and FZ in cultured isolates of Old World and New World Leishmania spp. for the treatment of CL.
METHODS
Promastigotes were cultured in Tobie's medium with Locke's overlay until log phase growth was achieved, inoculated into the Sensititre™ system, and incubated over 96 H. minimum inhibitory concentrations (MICs) were determined colorimetrically, and promastigote death was assessed by conventional microscopy out to 96- h. Colour change correlated to MIC values.
RESULTS
All strains tested exhibited MIC values for FZ that were ≥ 256 μg/mL. New World strains demonstrated reduced susceptibility to AB (0.25 μg/mL - 0.50 μg/mL AB) compared to Old World strains at 0.12 μg/mL AB (p = 0.02). Seventeen (61%) of 28 Viannia isolates versus 82% (27/33) of non-Viannia isolates were resistant at 0.12 μg/mL AB (p = 0.09). For L. V. braziliensis isolates, mean MIC for AB was 0.375 ± 0.14 μg/mL (range 0.25-0.50 μg/mL), while for isolates of L. V. panamensis it was 0.314 ± 0.26 μg/mL (range 0.12-1.0 μg/mL).
CONCLUSIONS
We adapted the Sensititre™ YeastOne™ YO9 plate for testing of Leishmania spp. susceptibility profiles for commonly used antifungals in the treatment of CL, including AB and FZ. Given its current utility in mycology, optimization of the system for potential clinical implementation in parasitology should be pursued. However evaluation of clinically relevant amastigote-stage stages, and higher concentrations of FZ beyond the upper limit concentration of the Sensititre™ YeastOne™ Y09 plate would be required.
背景
目前治疗皮肤利什曼病（CL）的药物方案包括毒性全身治疗，如两性霉素 B（AB）和五价锑。氟康唑（FZ）是一种耐受性良好的潜在口服替代物，可用于管理 CL。迄今为止，很少有客观数据可以指导临床决策，以预先选择治疗药物，并且尚未建立用于利什曼原虫的标准化、临床批准的药物敏感性测试平台。Sensititre™YeastOne™YO9 板是一种商业化的药敏板，包括 AB 和 FZ，用于非挑剔酵母的常规测试。我们的目标是改编现成的 Sensititre™YeastOne™YO9 板，以确定 AB 和 FZ 在治疗 CL 的旧世界和新世界利什曼原虫培养分离物中的药物敏感性谱。
方法
将前鞭毛体在含有 Locke 覆盖物的 Tobie 培养基中培养至对数生长期，接种到 Sensititre™系统中，并孵育 96 小时以上。通过比色法测定最低抑菌浓度（MIC），并用常规显微镜评估至 96 小时的前鞭毛体死亡情况。颜色变化与 MIC 值相关。
结果
所有测试的菌株对 FZ 的 MIC 值均≥256μg/mL。与 Old World 菌株相比，New World 菌株对 AB（0.25μg/mL-0.50μg/mL AB）的敏感性降低，而 Old World 菌株的 AB 敏感性为 0.12μg/mL（p=0.02）。17（61%）例 Viannia 分离株对 0.12μg/mL AB 耐药，而非 Viannia 分离株的耐药率为 82%（27/33）（p=0.09）。对于 L. V. braziliensis 分离株，AB 的平均 MIC 为 0.375±0.14μg/mL（范围 0.25-0.50μg/mL），而对于 L. V. panamensis 分离株的 MIC 为 0.314±0.26μg/mL（范围 0.12-1.0μg/mL）。
结论
我们改编了 Sensititre™YeastOne™YO9 板，用于测试治疗 CL 中常用抗真菌药物的利什曼原虫药敏谱，包括 AB 和 FZ。鉴于其目前在真菌学中的应用，应寻求对系统进行优化，以潜在地在寄生虫学中进行临床实施。然而，需要评估临床上相关的无鞭毛体阶段和高于 Sensititre™YeastOne™Y09 板上限浓度的 FZ 更高浓度。
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