The possibility that Alzheimer’s disease (AD) has a microbial aetiology has been proposed by several researchers. Here, we provide evidence that tissue from the central nervous system (CNS) of AD patients contain fungal cells and hyphae. Fungal material can be detected both intra- and extracellularly using specific antibodies against several fungi. Different brain regions including external frontal cortex, cerebellar hemisphere, entorhinal cortex/hippocampus and choroid plexus contain fungal material, which is absent in brain tissue from control individuals. Analysis of brain sections from ten additional AD patients reveals that all are infected with fungi. Fungal infection is also observed in blood vessels, which may explain the vascular pathology frequently detected in AD patients. Sequencing of fungal DNA extracted from frozen CNS samples identifies several fungal species. Collectively, our findings provide compelling evidence for the existence of fungal infection in the CNS from AD patients, but not in control individuals.

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Young parkin null (pk)/)) mice have subtle abnormalities of behaviour, dopamine (DA) neurotransmission and free radical production, but no massive loss of DA neurons. We investigated whether these findings are maintained while ageing. Pk)/) mice have reduced life span and age-related reduced exploratory behaviour, abnormal walking and posture, and behaviours similar to those of early Parkinson's disease (PD), reduced number of nigrostriatal DA neurons and proapoptotic shifts in the survival/death proteins in midbrain and striatum. Contrary to young pk)/) animals 24-month-old pk)/) mice do not have compensatory elevation of GSH in striatum, glutathione reductase (GR) and glutathione peroxidase (GPx) activities are increased and catalase unchanged. Aged pk)/) mice accumulate high levels of tau and fail to up-regulate CHIP and HSP70. Our results suggest that aged pk)/) mice lack of the compensatory mechanisms that maintain a relatively normal DA function in early adulthood. This study could help to explain the effects of ageing in patients with genetic risks for Parkinson's disease.

Among neurogenerative diseases, amyotrophic lateral sclerosis (ALS) is a fatal illness characterized by a progressive motor neuron dysfunction in the motor cortex, brainstem and spinal cord. ALS is the most common form of motor neuron disease; yet, to date, the exact etiology of ALS remains unknown. In the present work, we have explored the possibility of fungal infection in cerebrospinal fluid (CSF) and in brain tissue from ALS patients. Fungal antigens, as well as DNA from several fungi, were detected in CSF from ALS patients. Additionally, examination of brain sections from the frontal cortex of ALS patients revealed the existence of immunopositive fungal antigens comprising punctate bodies in the cytoplasm of some neurons. Fungal DNA was also detected in brain tissue using PCR analysis, uncovering the presence of several fungal species. Finally, proteomic analyses of brain tissue demonstrated the occurrence of several fungal peptides. Collectively, our observations provide compelling evidence of fungal infection in the ALS patients analyzed, suggesting that this infection may play a part in the etiology of the disease or may constitute a risk factor for these patients.

Abnormal deposition of protein tau takes place in the brain of patients with several neurodegenerative diseases. Few of these patients present frontotemporal dementia with parkinsonism and amyotrophy (FTDPA-17), an autosomal dominant tauopathy related to mutations of the gene that codes for protein tau, localized in chromosome 17. The great majority of patients with tauopathies such as Alzheimer's disease, sporadic frontotemporal dementia or progressive supranuclear palsy do not show a Mendelian pattern of inheritance. We have occasionally seen tauopathies in patients with parkin mutations and, therefore, hypothesized that the protein tau interacts with parkin. We have tested that hypothesis in mice with combined genetic modifications of tau (over-expression of human tau with three mutations known to produce FTDPA-17) and parkin (deleted) proteins. Homozygote parkin null or over-expressing mutated-human tau mice have subtle behavioral and molecular abnormalities but do not express a clinical phenotype of neurodegenerative disease. Mice with combined homozygous mutations of these two genes show progressively abnormal walking already noticeable at 3 months of age, loss of dopamine and dopamine markers in striatum, nuclear tau immunoreactive deposits in motor neurons of the spinal cord, abnormal expression of glial markers and enhanced levels of pro-apoptotic proteins; findings that were absent or less pronounced in homozygote animals with deletions of parkin or over-expression of tau. The double transgenic mice do not express normal mechanisms of adaptation to stress such as increased levels of GSH and Hsp-70. In addition, they have reduced levels of CHIP-Hsc70, a complex known to attenuate aggregation of tau and to enhance ubiquitination of phosphorylated tau. We have found high levels of phosphorylated tau in parkin-/-+tau(VLW) mice and a relative decrease of the inactivated pSer9 to total GSK-3 levels. Our data reveal that there are interactions between tau and parkin that could be relevant for the pathogenesis and treatment of tauopathies. Similarly, we hope that the double transgenic parkin-/-+tau(VLW) mice could be useful for testing of compounds with putative therapeutic value in human tauopathies.

Deposition of proteins leading to amyloid takes place in some neurodegenerative diseases such as Alzheimer's disease and Huntington's disease. Mutations of tau and parkin proteins produce neurofibrillary abnormalities without deposition of amyloid. Here we report that mature, parkin null, over-expressing human mutated tau (PK(-/-)/Tau(VLW)) mice have altered behaviour and dopamine neurotransmission, tau pathology in brain and amyloid deposition in brain and peripheral organs. PK(-/-)/Tau(VLW) mice have abnormal behaviour and severe drop out of dopamine neurons in the ventral midbrain, up to 70%, at 12 months and abundant phosphorylated tau positive neuritic plaques, neuro-fibrillary tangles, astrogliosis, microgliosis and plaques of murine beta-amyloid in the hippocampus. PK(-/-)/Tau(VLW) mice have organomegaly of the liver, spleen and kidneys. The electron microscopy of the liver confirmed the presence of a fibrillary protein deposits with amyloid characteristics. There is also accumulation of mouse tau in hepatocytes. These mice have lower levels of CHIP-HSP70, involved in the proteosomal degradation of tau, increased oxidative stress, measured as depletion of glutathione which, added to lack of parkin, could trigger tau accumulation and amyloidogenesis. This model is the first that demonstrates beta-amyloid deposits caused by over-expression of tau and without modification of the amyloid precursor protein, presenilins or secretases. PK(-/-)/Tau(VLW) mice provide a link between the two proteins more important for the pathogenesis of Alzheimer disease.

The identification of biomarkers for Alzheimer's disease is important for patient management and to assess the effectiveness of clinical intervention. Cerebrospinal fluid (CSF) biomarkers constitute a powerful tool for diagnosis and monitoring disease progression. We have analyzed the presence of fungal proteins and DNA in CSF from AD patients. Our findings reveal that fungal proteins can be detected in CSF with different anti-fungal antibodies using a slot-blot assay. Additionally, amplification of fungal DNA by PCR followed by sequencing distinguished several fungal species. The possibility that these fungal macromolecules could represent AD biomarkers is discussed.

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