Bacterial endotoxins have been associated with chronic inflammation and the development and progression of diabetic nephropathy. We hypothesized that subjects with high serum lipopolysaccharide activity also carry remains of bacterial DNA in their system. Serum-derived bacterial DNA clones were isolated and identified from 10 healthy controls and 14 patients with type 1 diabetes (T1D) using universal primers targeted to bacterial 16S rDNA. A total of 240 clones representing 35 unique bacterial species were isolated and identified. A significant proportion of the isolated bacteria could be assigned to our living environment. Proteobacteria was by far the most prevalent phylum among the samples. Notably, the patients had significantly higher frequencies of Stenotrophomonas maltophilia clones in their sera compared to the healthy controls. Real-time PCR analysis of S. maltophilia and Pseudomonas aeruginosa flagellin gene copy number in the human leukocyte DNA fraction revealed that the overall Pseudomonal bacterial load was higher in older patients with T1D. Serum IgA- and IgG-antibody levels against Pseudomonal bacteria Delftia acidovorans, P. aeruginosa, and S. maltophilia were also determined in 200 healthy controls and 200 patients with T1D. The patients had significantly higher serum levels of IgA antibodies against all three Pseudomonal bacteria. Additionally, the IgA antibodies against Pseudomonal bacteria correlated significantly with serum C-reactive protein. These findings indicate that recurrent or chronic Pseudomonal exposure may increase susceptibility to chronic inflammation in patients with T1D.