Determining the microbial species content in brain tissue

The aetiology of Alzheimer disease (AD) and Parkinson Disease (PD) are unknown and tend to manifest at a later stage in life; even though these diseases have different pathogenic mechanisms, they are both characterized by neuroinflammation in the brain. Links between bacterial and viral infection and AD/PD has been suggested in several studies, however, few have attempted to establish a link between fungal infection and AD/PD. In this study we develop and describe a nanopore-based sequencing approach to characterise the presence or absence of fungi in both human brain tissue and cerebrospinal fluid (CSF). This approach detects fungal DNA in human brain and CSF samples even at low levels, whereas our quantitative polymerase chain reaction (qPCR) assay (FungiQuant) was unable to detect fungal DNA in the same samples. Comparison against kit-controls showed ubiquitous low-level fungal contamination that we observed in healthy human brains and CSF as well AD/PD brains and CSF. We use this technique to demonstrate the presence of fungal DNA in healthy human brains as well as AD/PD brains, with Alternaria spp., Colletotrichum graminicola, and Filobasidium floriforme as the most prominent species. In addition, antibiotic resistant Pseudomonas spp. was identified within the brain of an AD patient. Our method will be broadly applicable to investigating potential links between microbial infection and AD/PD.