Departamento de Bioquímica e Imunologia, Universidade Federal de Minas Gerais, Avenida Antônio Carlos 6627, Belo Horizonte, MG CEP 31.270-901, Brazil.
Departamento de Análises Clínicas e Toxicológicas, Universidade Federal de Minas Gerais, Avenida Antônio Carlos 6627, Belo Horizonte, MG CEP 31.270-901, Brazil.
Genomics. 2023 Sep;115(5):110661. doi: 10.1016/j.ygeno.2023.110661. Epub 2023 May 30.
We report the sequencing and assembly of the PH8 strain of Leishmania amazonensis one of the etiological agents of leishmaniasis. After combining data from long Pacbio reads, short Illumina reads and synteny with the Leishmania mexicana genome, the sequence of 34 chromosomes with 8317 annotated genes was generated. Multigene families encoding three virulence factors, A2, amastins and the GP63 metalloproteases, were identified and compared to their annotation in other Leishmania species. As they have been recently recognized as virulence factors essential for disease establishment and progression of the infection, we also identified 14 genes encoding proteins involved in parasite iron and heme metabolism and compared to genes from other Trypanosomatids. To follow these studies with a genetic approach to address the role of virulence factors, we tested two CRISPR-Cas9 protocols to generate L. amazonensis knockout cell lines, using the Miltefosine transporter gene as a proof of concept.
我们报告了利什曼原虫亚马逊亚种 PH8 株的测序和组装，该种是利什曼病的病原体之一。在结合了长 Pacbio 读段、短 Illumina 读段和与 Leishmania mexicana 基因组的同线性数据后，生成了 34 条染色体的序列，其中包含 8317 个注释基因。鉴定并比较了编码三种毒力因子（A2、变形蛋白和 GP63 金属蛋白酶）的多基因家族，以及它们在其他利什曼原虫物种中的注释。由于这些因子最近被认为是对疾病建立和感染进展至关重要的毒力因子，我们还鉴定了 14 个编码参与寄生虫铁和血红素代谢的蛋白的基因，并与其他原生动物的基因进行了比较。为了通过遗传方法研究这些毒力因子的作用，我们测试了两种 CRISPR-Cas9 方案，使用米替福新转运蛋白基因作为概念验证，以生成 L. amazonensis 敲除细胞系。
