The Oesophageal Squamous Cell Carcinoma Cell Line COLO-680N Fails to Support Sustained Cryptosporidium parvum Proliferation.

Liliana M R Silva,Juan Vélez,Anja Taubert,Faustin Kamena,Arwid Daugschies,Sybille Mazurek,Carlos Hermosilla

doi:10.3390/pathogens11010049

Abstract

Cryptosporidium parvum is an important diarrhoea-associated protozoan, which is difficult to propagate in vitro. In 2017, a report described a continuous culture of C. parvum Moredun strain, in the oesophageal squamous cell carcinoma cell line COLO-680N, as an easy-to-use system for C. parvum propagation and continuous production of oocysts. Here, we report that—using the Köllitsch strain of C. parvum—even though COLO-680N cells, indeed, allowed parasite invasion and early asexual parasite replication, C. parvum proliferation decreased after the second day post infection. Considering recurring studies, reporting on successful production of newly generated Cryptosporidium oocysts in the past, and the subsequent replication failure by other research groups, the current data stand as a reminder of the importance of reproducibility of in vitro systems in cryptosporidiosis research. This is of special importance since it will only be possible to develop promising strategies to fight cryptosporidiosis and its ominous consequences for both human and animal health by a continuous and reliable methodological progress.

Highlights

Cryptosporidium parvum and C. hominis are recognized as protozoan parasite species causing diarrhoea in humans worldwide for a long time, but only recently, the true relevance as the second most important diarrheal pathogen in young children was revealed [1].for immunosuppressed humans, such as HIV patients, and malnourished infants it can become a life-threatening disease [2,3]
C. parvum infections were studied in parallel in COLO680N and HCT-8 cells (HCT-8 cells represent the cell line mostly used in Cryptosporidium-related in vitro cultures) and subjected to three different infection protocols, according to Miller et al (2018) and other publications [14,15]
Similar infection rates were observed in both cell lines (HCT-8 and COLO-680N), applying three different infection protocols (Table 1) and estimating infection rates at 24 hpi by

Summary

Introduction

Cryptosporidium parvum and C. hominis are recognized as protozoan parasite species causing diarrhoea in humans worldwide for a long time, but only recently, the true relevance as the second most important diarrheal pathogen in young children was revealed [1].for immunosuppressed humans, such as HIV patients, and malnourished infants it can become a life-threatening disease [2,3]. Cryptosporidium parvum and C. hominis are recognized as protozoan parasite species causing diarrhoea in humans worldwide for a long time, but only recently, the true relevance as the second most important diarrheal pathogen in young children was revealed [1]. Only one FDA-approved drug, namely nitazoxanide, is available to date, but it lacks efficacy in immunocompromised C. parvum-infected patients [4]. A major problem in Cryptosporidium research is the lack of appropriate in vitro systems that enable continuous C. parvum culture including oocyst generation. Even though some important tools for basic research on C. parvum have recently been achieved by various laboratories around the world, the lack of an easy and reproducible, continuous in vitro culture of this parasite remains the major issue and mainly relies on a failure of sexual

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