Purpose

Leishmaniasis comprises various clinical forms mainly including cutaneous, muco-cutaneous, and visceral leishmaniasis; caused by Leishmania species. Antimoniate is the first-line treatment but some cases showed no response to treatment in the worldwide. In this study, mitogen-activated protein kinase (MAPK) and aquaglyceroporin 1 (AQP1) gene expressions were assessed in treatment failure clinical isolates of Leishmania major. Also, molecular and phylogenic analyses of the mentioned isolates were performed.

Methods

Samples were obtained from the patients with suspicious CL referred to the laboratory of Diagnosis Center, Gorgan Province, Iran, from October 2016 to December 2019. Detection and identification of the parasite was performed. The genes expressions of MAPK1 and AQP1 were done using SYBR Green real-time PCR. The AQP1 gene from the isolates with treatment failure was sequenced and analyzed using BLAST and multiple alignments. The phylogenic analysis was done using MEGA7. The statistical analysis was done using SPSS 16.0 by non-parametric Mann–Whitney U test.

Results

All clinical isolates were detected L. major. The mean AQP1 and MAPK1 gene expressions in treatment failure isolates were 58.71 and 6.139 fold less than the ones in treatment response isolates, respectively. Based on the AQP1 gene sequence, a nucleotide change of aspartic acid with asparagine at the site 234 was observed. Phylogenic tree analysis showed three groups with the minimum dissimilarity of 0.008 between TF isolates with the standard L. major strains.

Conclusion

We showed that MAPK1 and AQP1 may have critical roles in response to antimoniate in clinical isolates L. major in this study.

中文翻译：

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治疗失败利什曼原虫中的丝裂原活化蛋白激酶和水甘油通道蛋白基因表达

目的

利什曼病包括多种临床形式，主要包括皮肤、粘膜皮肤和内脏利什曼病；由利什曼原虫引起。锑酸盐是一线治疗，但在世界范围内有些病例对治疗没有反应。在这项研究中，在治疗失败的主要利什曼原虫临床分离株中评估了丝裂原活化蛋白激酶 ( MAPK ) 和水甘油通道蛋白 1 ( AQP1 ) 基因的表达。此外，对上述分离株进行了分子和系统发育分析。

方法

样本取自 2016 年 10 月至 2019 年 12 月转诊至伊朗戈尔甘省诊断中心实验室的疑似 CL 患者。进行了寄生虫的检测和鉴定。MAPK1和AQP1的基因表达使用 SYBR Green 实时 PCR 完成。使用 BLAST 和多重比对对来自治疗失败的分离株的AQP1基因进行测序和分析。使用MEGA7进行系统发育分析。统计分析使用 SPSS 16.0 通过非参数 Mann - Whitney U检验完成。

结果

所有临床分离株均检测到L. major。治疗失败分离株的平均AQP1和MAPK1基因表达分别比治疗反应分离株低 58.71 和 6.139 倍。基于AQP1基因序列，在234位点观察到天冬氨酸与天冬酰胺的核苷酸变化。系统发育树分析显示三组 TF 分离株与标准L. major菌株之间的最小差异为 0.008。

结论

在本研究中，我们发现MAPK1和AQP1可能对临床分离株L. major中的锑化物起关键作用。