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Background & Aims Dysbiosis of the gut microbiota in response to an energy-rich Western diet and the potential leak of bacteria and/or bacterial products from the intestine to the liver is perceived as a potential risk factor for the development of non-alcoholic fatty liver disease (NAFLD). We investigated the microbiome in liver biopsies from healthy lean and obese individuals and compared it with their blood microbiome. Methods We examined liver biopsies from 15 healthy lean and 14 obese individuals (BMI of 18.5–25 and 30–40 kg/m2, respectively). Bacterial 16S ribosomal DNA (rDNA) was analysed by quantitative polymerase chain reaction (qPCR) and 16S metagenomic sequencing targeting the hypervariable V3–V4 region. Metagenomic analysis was performed using the linear discriminant analysis effect size (LEfSe) algorithm. Data are medians with IQRs in brackets. Results Histology revealed hepatic steatosis in 13 obese individuals and in 2 lean individuals. A robust signal from qPCR revealed significantly higher amounts of bacterial rDNA copies in liver samples from obese individuals compared with those from lean individuals (148 [118–167] vs. 77 [62–122] 16S copies/ng DNA, p, Graphical abstract, Highlights • Liver tissue from healthy lean and obese individuals contains bacterial rDNA. • Significantly higher amounts of bacterial rDNA and lower alpha diversity were found in liver samples from obese individuals. • Proteobacteria-enriched metataxonomic signature was found in liver samples from obese individuals. • There is strong correlation between the amount of bacterial rDNA in liver samples and the fatty liver index. • Differences in the liver microbiome between lean and obese individuals are not reflected in the blood microbiome.