Cryptosporidium parvum Infection in SCID Mice Infected with Only One Oocyst: qPCR Assessment of Parasite Replication in Tissues and Development of Digestive Cancer

Sadia Benamrouz,Valerie Conseil,Eric Viscogliosi,Karine Guyot,Baptiste Delaire,Thierry Chassat,Pierre Gosset,Eduardo Dei-Cas,Colette Creusy,Magali Chabé,Sophie Gazzola,Gabriela Certad,Anthony Mouray,Gordon Langsley

doi:10.1371/journal.pone.0051232

Abstract

Dexamethasone (Dex) treated Severe Combined Immunodeficiency (SCID) mice were previously described as developing digestive adenocarcinoma after massive infection with Cryptosporidium parvum as soon as 45 days post-infection (P.I.). We aimed to determine the minimum number of oocysts capable of inducing infection and thereby gastrointestinal tumors in this model. Mice were challenged with calibrated oocyst suspensions containing intended doses of: 1, 10, 100 or 105 oocysts of C. parvum Iowa strain. All administered doses were infective for animals but increasing the oocyst challenge lead to an increase in mice infectivity (P = 0.01). Oocyst shedding was detected at 7 days P.I. after inoculation with more than 10 oocysts, and after 15 days in mice challenged with one oocyst. In groups challenged with lower inocula, parasite growth phase was significantly higher (P = 0.005) compared to mice inoculated with higher doses. After 45 days P.I. all groups of mice had a mean of oocyst shedding superior to 10,000 oocyst/g of feces. The most impressive observation of this study was the demonstration that C. parvum-induced digestive adenocarcinoma could be caused by infection with low doses of Cryptosporidium, even with only one oocyst: in mice inoculated with low doses, neoplastic lesions were detected as early as 45 days P.I. both in the stomach and ileo-caecal region, and these lesions could evolve in an invasive adenocarcinoma. These findings show a great amplification effect of parasites in mouse tissues after challenge with low doses as confirmed by quantitative PCR. The ability of C. parvum to infect mice with one oocyst and to develop digestive adenocarcinoma suggests that other mammalian species including humans could be also susceptible to this process, especially when they are severely immunocompromised.

Highlights

Cryptosporidium species are worldwide spread apicomplexan parasitic protists that infect mostly the gastrointestinal tract of fish, amphibians, reptiles, birds and more than 150 species of mammals including human beings [1]
In order to better describe our animal model, we explored the potential ability of freshly isolated Cryptosporidium oocysts to induce both patent infection and gastrointestinal neoplastic changes when administered at very low dose
analysis of variance (ANOVA) analysis of the whole data set showed that the day P.I. and the inoculum size significantly influence the geometric means of oocyst sheding (P = 0.02 and 0.005, respectively): at 75 days P.I., mice inoculated with intended doses of 1, 10, 100 and 105 oocysts had a multiplication of 3.3, 3.25, 2.26 and 1.45 log respectively compared to initial inoculum

Summary

Introduction

Cryptosporidium species are worldwide spread apicomplexan parasitic protists that infect mostly the gastrointestinal tract of fish, amphibians, reptiles, birds and more than 150 species of mammals including human beings [1]. More than 20 Cryptosporidium species are regarded as valid [3], and two major species, Cryptosporidium parvum and C. hominis, are responsible for most human cases of cryptosporidiosis [4]. A high infectious power of Cryptosporidium isolates from human or animal origin was reported [5]. In healthy volunteers with no serologic evidence of past infection with Cryptosporidium, an oral dose of 30 C. parvum oocysts caused infection [5]. The same group reported that Cryptosporidium hominis ID50 was 10 oocysts [4]. It was reported that a single oocyst of C. meleagridis can produce a patent infection in steroid-treated C57BL/6 mice [6]

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