Investigation into Toxoplasma gondii infection in relation to expression of neurodegenerative disease markers in natural mice populations
Abstract
Toxoplasma gondii is a highly prevalent opportunistic protozoan parasite that has a remarkable capacity to infect humans and a range of warm-blooded animals. This research had two broad aims. Firstly, to investigating whether detection of T. gondii in cadaveric brain tissue of patients, who had been previously diagnosed with Alzheimer’s Disease (AD), could be used to investigate the role of this parasite in AD. Secondly to investigate T. gondii infection in naturally infected wood mice (Apodemus sylvaticus) populations and the parasite’s role in the expression of a range of key autophagic and inflammatory genes associated with neurodegenerative diseases in humans.
Human brain tissue from the hippocampus and frontal lobe regions was obtained from 45 cadaveric specimens and analysed for the presence of Toxoplasma gondii cysts using visual microscopy. The human brain tissue did not show infection with T. gondii as judged by the absence of cysts found within the parenchyma of any of the subjects. Given the expected 10% prevalence in the UK and the small volume of brain tissue used and it was concluded that this was insufficient tissue material to investigate the association in this way. Due to the amount of brain tissue needed for scaling up and the unlikely capability of sourcing enough brain tissue, this approach was considered unfeasible.
A population of 21 A. sylvaticus (wood mice) were captured from a local peri-aquatic woodland ecosystem, euthanised followed by extraction and analysis of brain tissue. Toxoplasma infection was diagnosed using nested PCR with specific markers for B1, SAG1 and SAG2. Additionally, PCR and Western blotting were carried out to permit detection of the presence and expression of various autophagic and inflammatory genes/proteins.
As a result, this naturally infected murine population was used as model system for studying the interaction between infection and host brain gene expression. T. gondii was detected in five samples (5/21) of A. sylvaticus reflecting an infection rate of 23.8% (95% confidence interval: 10.2; 45.5%), which is a comparable prevalence to other recent studies in wood mice but considered to be a high for a species resident among a relatively feline-free habitat. Among the autophagy and inflammatory genes, PCR detection and DNA sequencing identified Apodemus homologues of human Beclin1, Lc3 and Lamp2, TLR2, Tau and APP in the murine brain tissue. Expression of various autophagy and inflammatory proteins were examined by western blotting. There were significantly increased levels of TLR2, LC3, Socs1, Arg1, in T. gondii infected mice compared to non-infected mice, but decreased levels of NMDAR1, Lamp2, P62, Beclin-1 and iNOS. No significant difference was observed in the levels of Tau and APP between infected and non-infected mice. The findings of this study are novel and insightful as they have revealed that the brain tissues of T. gondii infected mice, in the context of a natural environment with natural infection, are undergoing changes in autophagic and inflammatory processes, often associated with neurodegenerative disease, in response to infection.. Overall, this study concludes that wild rodents could be used as a model natural population to explore gene expression relevant to human neurodegenerative diseases. This may pave the way to a greater understanding of the interactions of T. gondii with the brain and lead to new insights into its effect on neurological diseases.