ABSTRACT

Cutaneous leishmaniasis (CL) is endemic in Sri Lanka. Giemsa-stained slit-skin-smears (SSS-Giemsa) and histology are routinely used in diagnosis with a sensitivity of 40–70%. PCR currently has limited accessibility. Therefore, we assessed the sensitivity and specificity of a previously described fluorescence in situ hybridization assay, on skin smears and biopsy samples to overcome the limitations encountered with routine diagnostic methods.

Samples from a total of 123 suspected CL patients were collected and subjected to SSS-Giemsa, fluorescence in situ hybridization (FISH) on slit skin smears (SSS-FISH), formalin-fixed-paraffin-embedded-tissues stained with Hematoxylin & Eosin staining (FFPE-H&E) and FISH on formalin-fixed-paraffin-embedded-tissues (FFPE-FISH). Negative controls of 61 patient samples were collected from a CL non-endemic area and subjected to the same procedures. The gold standard PCR was used as a comparator. For FISH, two previously described cyanine 3 tagged Leihsmania genus-specific probes were used.

Compared to PCR, SSS-Giemsa, SSS-FISH, FFPE-H&E, and FFPE-FISH had sensitivities of 76.5%, 79.1%, 50.4% and 80.9%, respectively. Routine diagnostic tests (SSS-Giemsa and FFPE-H&E) had a specificity of 100%. SSS-FISH and FFPE-FISH had specificities of 96.7% and 93.4%, respectively. FFPE-FISH had a statistically significant higher diagnostic performance than FFPE-H&E (p < 0.001). The relative performance of SSS-Giemsa, SSS-FISH and FFPE-FISH was similar (p > 0.05 for all comparisons).

We conclude that FFPE-FISH is a more accurate diagnostic tool than FFPE–H&E. SSS-FISH did not have an additional advantage over SSS-Giemsa in diagnosis. However, SSS-FISH could be recommended as a minimally invasive method in studies assessing wound healing where immunological probes are used.

摘要

皮肤利什曼病（CL）在斯里兰卡流行。吉姆萨染色裂隙皮肤涂片 (SSS-Giemsa) 和组织学常规用于诊断，灵敏度为 40-70%。PCR 目前的可及性有限。因此，我们评估了先前描述的荧光原位杂交测定对皮肤涂片和活检样本的敏感性和特异性，以克服常规诊断方法遇到的局限性。

共收集 123 名疑似 CL 患者的样本，并进行 SSS-Giemsa、裂隙皮肤涂片荧光原位杂交 (SSS-FISH)、福尔马林固定石蜡包埋组织苏木精和伊红染色(FFPE-H&E) 和福尔马林固定石蜡包埋组织上的 FISH (FFPE-FISH)。61 名患者样本的阴性对照是从 CL 非流行区收集的，并进行相同的程序。使用金标准 PCR 作为比较器。对于 FISH，使用了两种先前描述的花青 3 标记的莱氏曼原虫属特异性探针。

与 PCR 相比，SSS-Giemsa、SSS-FISH、FFPE-H&E 和 FFPE-FISH 的灵敏度分别为 76.5%、79.1%、50.4% 和 80.9%。常规诊断测试（SSS-Giemsa 和 FFPE-H&E）的特异性为 100%。SSS-FISH 和 FFPE-FISH 的特异性分别为 96.7% 和 93.4%。FFPE-FISH 的诊断性能显着高于 FFPE-H&E (p < 0.001)。SSS-Giemsa、SSS-FISH 和 FFPE-FISH 的相对性能相似（所有比较的 p > 0.05）。

我们的结论是，FFPE-FISH 是比 FFPE-H&E 更准确的诊断工具。SSS-FISH 在诊断方面并不比 SSS-Giemsa 具有额外的优势。然而，在使用免疫探针评估伤口愈合的研究中，可以推荐 SSS-FISH 作为一种微创方法。