Detecting eukaryotic microbiota with single-cell sensitivity in human tissue

Susanne Lager, Marcus C. de Goffau, Ulla Sovio, Sharon J. Peacock, Julian Parkhill, D. Stephen Charnock-Jones, Gordon C. S. Smith

Fetal growth restriction, pre-eclampsia, and pre-term birth are major adverse pregnancy outcomes. These complications are considerable contributors to fetal/maternal morbidity and mortality worldwide. A significant proportion of these cases are thought to be due to dysfunction of the placenta. However, the underlying mechanisms of placental dysfunction are unclear. The aim of the present study was to investigate whether adverse pregnancy outcomes are associated with evidence of placental eukaryotic infection. We modified the 18S Illumina Amplicon Protocol of the Earth Microbiome Project and made it capable of detecting just a single spiked-in genome copy of Plasmodium falciparum, Saccharomyces cerevisiae, or Toxoplasma gondii among more than 70,000 human cells. Using this method, we were unable to detect eukaryotic pathogens in placental biopsies in instances of adverse pregnancy outcome (n = 199) or in healthy controls (n = 99). Eukaryotic infection of the placenta is not an underlying cause of the aforementioned pregnancy complications. Possible clinical applications for this non-targeted, yet extremely sensitive, eukaryotic screening method are manifest.