Centro Nacional de Secuenciación Genómica-CNSG, Sede de Investigación Universitaria-SIU, Medellín, Antioquia, Colombia.
Departamento de Microbiología y Parasitología, Facultad de Medicina, Universidad de Antioquia, Medellín, Antioquia, Colombia.
PLoS One. 2022 Jul 8;17(7):e0270995. doi: 10.1371/journal.pone.0270995. eCollection 2022.
Multilocus Sequence Typing has become a useful tool for the study of the genetic diversity and population structure of different organisms. In this study, a MLST approach with seven loci (CP47, MS5, MS9, MSC6-7, TP14, and gp60) was used to analyze the genetic diversity of Cryptosporidium hominis and Cryptosporidium parvum isolated from 28 Colombian patients. Five Cryptosporidium species were identified: C. hominis, C. parvum, Cryptosporidium felis, Cryptosporidium meleagridis, and Cryptosporidium suis. Unilocus gp60 analysis identified four allelic families for C. hominis (Ia, Ib, Id, and Ie) and two for C. parvum (IIa and IIc). There was polymorphic behavior of all markers evaluated for both C. hominis and C. parvum, particularly with the CP47, MS5, and gp60 markers. Phylogenetic analysis with consensus sequences (CS) of the markers showed a taxonomic agreement with the results obtained with the 18S rRNA and gp60 gene. Additionally, two monophyletic clades that clustered the species C. hominis and C. parvum were detected, with a higher number of subclades within the monophyletic groups compared to those with the gp60 gene. Thirteen MLG were identified for C. hominis and eight for C. parvum. Haplotypic and nucleotide diversity were detected, but only the latter was affected by the gp60 exclusion from the CS analysis. The gene fixation index showed an evolutionary closeness between the C. hominis samples and a less evolutionary closeness and greater sequence divergence in the C. parvum samples. Data obtained in this work support the implementation of MLST analysis in the study of the genetic diversity of Cryptosporidium, considering the more detailed information that it provides, which may explain some genetic events that with an unilocus approach could not be established. This is the first multilocus analysis of the intra-specific variability of Cryptosporidium from humans in South America.
多位点序列分型已成为研究不同生物遗传多样性和种群结构的有用工具。在这项研究中，采用了一种包含 7 个基因座（CP47、MS5、MS9、MSC6-7、TP14 和 gp60）的 MLST 方法，分析了 28 名哥伦比亚患者分离的隐孢子虫同源物和微小隐孢子虫的遗传多样性。鉴定出 5 种隐孢子虫：人隐孢子虫、微小隐孢子虫、猫隐孢子虫、火鸡隐孢子虫和猪隐孢子虫。单基因座 gp60 分析鉴定出人隐孢子虫的 4 个等位基因家族（Ia、Ib、Id 和 Ie）和微小隐孢子虫的 2 个（IIa 和 IIc）。CP47、MS5 和 gp60 标记的所有评估标记都表现出多态性，尤其是人隐孢子虫和微小隐孢子虫。基于标记共识序列的系统发育分析与 18S rRNA 和 gp60 基因的结果具有分类学一致性。此外，检测到人隐孢子虫和微小隐孢子虫的两个单系进化枝，与 gp60 基因相比，单系群内的亚群数量更多。人隐孢子虫鉴定出 13 个单倍型，微小隐孢子虫鉴定出 8 个。检测到单倍型和核苷酸多样性，但只有后者受到从 CS 分析中排除 gp60 的影响。基因固定指数显示人隐孢子虫样本之间的进化关系较近，而微小隐孢子虫样本之间的进化关系较远，序列差异更大。本工作获得的数据支持在隐孢子虫遗传多样性研究中实施 MLST 分析，考虑到它提供的更详细信息，这可能解释了一些在单基因座方法中无法建立的遗传事件。这是首次在南美洲对人类隐孢子虫的种内变异性进行多位点分析。
