Project description:Background: Fasciola hepatica infection still remains one of the helminthic neglected tropical diseases (NTDs). It has a huge worldwide distribution, affecting mainly cattle and, sometimes, human beings. In addition to data reported about the immunological response induced by helminthic infections and that induced by Fasciola hepatica, little is known about the gene expression profile in its organ target, the liver, which is where adult worms are established and live for long periods of time, causing its characteristic pathology. In the present work, we study both the early and late gene expression profiles in the livers of mice infected with Fasciola hepatica metacercariae using a microarray-based methodology. Methodology: A total of 9 female-6-week-old BALB/c mice (Charles River Laboratories, Barcelona, Spain) weighing 20 to 35 g were used for the experiments. Two groups of BALB/c mice were orally infected with seven F. hepatica metacercariae, and the other group remained untreated and served as a control. Mice were humanely euthanized and necropsied for liver recovery, histological assessment of hepatic damage, RNA isolation, microarray design and gene expression analysis on the day of infection (t0), seven days post-infection (t7) and twenty-one days post-infection (t21). Results: We found that Fasciola hepatica infection induces the differential expression of 128 genes in the liver in the early stage of infection and 308 genes in the late stage, and most of them are up-regulated. The Ingenuity Pathway Analysis revealed significant changes in the pathways related to metabolism, biosynthesis and signaling as well as genes implicated in inducing liver-toxicity, injury and death. Conclusion: The present study provides us insights at the molecular level about the underlying mechanisms used by Fasciola hepatica, leading to liver damage and its subsequent pathophysiology. The expression pattern obtained here could also be used to explain the lack of association between infection with F. hepatica and cholangiocarcinoma. However, more studies should be performed to confirm this hypothesis. We used three experimental groups each containing 3 mice. Group 1 remains untreated and served as control. Group 2 was infected with Fasciola hepatica metacercariae on day 0 and humanely necropsied at 7 days post-infection. Group 3 was infected with Fasciola hepatica metacercariae on day 0 and humanely necropsied at 21 days post- infection. At the time of necropsy, liver of each mice were removed and the RNA was isolated. We compared the gene expression profile in the liver of mice infected with Fasciola hepatica.
Project description:Purpose: The miRnome of the liver fluke, Fasciola hepatica, has historically been assembled with limited omic's resources using specific life stages. The goals of this study is to quanity the known and published miRNAs and determine novel miRNAs across three intra-mammlian life stages. Methods: Total RNA was extracted from newly ecysted juveniles 24h post excystment, juveniles at 21d post infection in rats and adult worms during hepatic infection in sheep. Total RNA of each sample was prepared for miRNA sequencing library. Sequenced reads were cleaned and quanitified for F. hepatica mature miRNAs from miRBase version 21 and published miRNAs from Ricafrente et al 2021 using Bowtie tool. Cleaned reads were additionally analysed for novel miRNAs using MiRDeep2 tool in conjunction with the F. hepatica genome and mature miRNAs from F. hepatica, S. japonicum, S. mansoni and C.elegans. Known/published and novel miRNA read counts were normalised to CPM. Results: Of all miRNAs known/published and novel, 124 miRNAs now make up the F. hepatica miRnome. Each life stage exhibited a unique miRNA profile, from which NEJs were the most differentially expressed compared to juveniles and adult. Conclusions: Our study is the first to comparatively assess the miRNA profiles of the intrammalian life stages of F.hepatica simultaneously, from which the miRnome has now been expanded from 77 to 124.