Abstract

Background

Cryptosporidiosis is a parasitic disease associated with potentially fatal diarrhea. The most used method in Cryptosporidium subtyping is based on the glycoprotein gene gp60. Each infection can represent a parasite population, and it is important to investigate the influence on transmission and virulence, as well as any impact on public health investigations. However, an easy-to-use method for detection is lacking.

Methods

Here we report on the use of the bioinformatic program TIDE for deconvolution of gp60 chromatograms. A combination of single oocyst analysis and cloning successfully confirmed the within-sample parasite population diversity. Retrospective sample analysis was conducted on archived chromatograms.

Results

For Cryptosporidium parvum, 8.6% multistrain infections (13 of 152) obscured by currently used consensus base calling were detected. Importantly, we show that single oocysts can harbor a mixed population of sporozoites. We also identified a striking dominance of unappreciated polymerase stutter artefacts in all 218 chromatograms analyzed, challenging the uncritical use of gp60 typing.

Conclusions

We demonstrate the value of a new, easy-to-use analytical procedure for critical characterization of C. parvum and Cryptosporidium hominis in epidemiological investigations, also applicable retrospectively. Our findings illuminate the hidden parasite diversity with important implications for tracing zoonotic and person-to-person transmissions.

中文翻译：

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通过 gp60 分型对隐孢子虫感染的 TIDE 分析揭示了模糊的混合感染

摘要

背景

隐孢子虫病是一种与可能致命的腹泻相关的寄生虫病。隐孢子虫亚型中最常用的方法是基于糖蛋白基因gp60。每种感染都可以代表一个寄生虫种群，调查对传播和毒力的影响以及对公共卫生调查的任何影响非常重要。然而，缺乏一种易于使用的检测方法。

方法

在这里，我们报告了使用生物信息学程序 TIDE 对gp60色谱图进行反卷积的情况。单个卵囊分析和克隆的结合成功地证实了样本内寄生虫种群的多样性。对存档的色谱图进行了回顾性样品分析。

结果

对于小隐孢子虫，检测到 8.6% 的多菌株感染（152 例中的 13 例）被当前使用的共识碱基调用所掩盖。重要的是，我们表明单个卵囊可以含有混合的子孢子种群。我们还在所分析的所有 218 个色谱图中发现了未被发现的聚合酶口吃伪影的显着优势，这对不加批判地使用gp60分型提出了挑战。

结论

我们展示了一种新的、易于使用的分析程序在流行病学调查中对C. parvum和人隐孢子虫的关键表征的价值，也可追溯适用。我们的研究结果阐明了隐藏的寄生虫多样性，对追踪人畜共患病和人与人之间的传播具有重要意义。