Transcriptome analysis reveals unique metabolic features in the Cryptosporidium parvum Oocysts associated with environmental survival and stresses

Abstract

BackgroundCryptosporidium parvum is a globally distributed zoonotic parasite and an important opportunistic pathogen in immunocompromised patients. Little is known on the metabolic dynamics of the parasite, and study is hampered by the lack of molecular and genetic tools. Here we report the development of the first Agilent microarray for C. parvum (CpArray15K) that covers all predicted ORFs in the parasite genome. Global transcriptome analysis using CpArray15K coupled with real-time qRT-PCR uncovered a number of unique metabolic features in oocysts, the infectious and environmental stage of the parasite.ResultsOocyst stage parasites were found to be highly active in protein synthesis, based on the high transcript levels of genes associated with ribosome biogenesis, transcription and translation. The proteasome and ubiquitin associated components were also highly active, implying that oocysts might employ protein degradation pathways to recycle amino acids in order to overcome the inability to synthesize amino acids de novo. Energy metabolism in oocysts was featured by the highest level of expression of lactate dehydrogenase (LDH) gene. We also studied parasite responses to UV-irradiation, and observed complex and dynamic regulations of gene expression. Notable changes included increased transcript levels of genes involved in DNA repair and intracellular trafficking. Among the stress-related genes, TCP-1 family members and some thioredoxin-associated genes appear to play more important roles in the recovery of UV-induced damages in the oocysts. Our observations also suggest that UV irradiation of oocysts results in increased activities in cytoskeletal rearrangement and intracellular membrane trafficking.ConclusionsCpArray15K is the first microarray chip developed for C. parvum, which provides the Cryptosporidium research community a needed tool to study the parasite transcriptome and functional genomics. CpArray15K has been successfully used in profiling the gene expressions in the parasite oocysts as well as their responses to UV-irradiation. These observations shed light on how the parasite oocysts might adapt and respond to the hostile external environment and associated stress such as UV irradiation.

UV irradiation responses in Giardia intestinalis

“…It has been reported that human cells submitted to UV stress in general show low alternation in transcription levels (Rieger and Chu, 2004). This was also observed for E. histolytica (Weber et al, 2009) and C. parvum (Zhang et al, 2012) and now also in this study. The question is if this low alteration in transcription will be reflected on the protein synthesis in the cell.…”

“…The chosen recovery time was based on a similar study in E. histolytica where it was shown that the transcriptional response was most evident after 3 hours (Weber et al, 2009). In addition, our chosen recovery time falls in between those used in a study on UV-C treatment on Cryptosporidium oocyst in which significant transcriptional regulation at both 0.5 hours and 5 hours post-radiation (Zhang et al, 2012) was shown. Our transcriptional analysis showed that 108 genes in trophozoites and 202 genes in cysts changed their expression upon UV-C irradiation more than 1.5-fold and 85 genes showed more than 2-fold changes.…”

“…The response to UV-C treatment of three other protozoan parasites, Entamoeba histolytica, Cryptosporidium parvum and Toxoplasma gondii, has recently been published (Ware et al, 2010;Weber et al, 2009;Zhang et al, 2012). In E. histolytica, trophozoites subjected to UV-C treatment were studied using microarrays and the expression of genes encoding Fe-S clusters-containing proteins and DNA repair proteins were reported to be up-regulated, whereas genes associated to the cytoskeleton were repressed (Weber et al, 2009).…”

“…The overall expression analysis revealed few genes with large gene expression differences (5 genes >5-fold up or down) between UV irradiated and control trophozoites and cysts (Supplementary File S1). To study changes in the pattern of gene expression as a response to UV irradiation we selected a cutoff value at ≥1.5-fold change between irradiated and control cells, in line with earlier studies of Cryptosporidium oocysts (Zhang et al, 2012). We found that 108 and 202 genes in trophozoites and cysts, respectively had a significant (≥1.5-fold) differential expression due to UV treatment.…”

“…Entamoeba histolytica and Cryptosporidium parvum, pathogenic intestinal protozoan parasites, have both been shown to induce gene expression changes following UV irradiation (Weber et al, 2009;Zhang et al, 2012). Here we studied if also G. intestinalis can respond to UV irradiation with gene expression changes.…”