The aim of the present study was to evaluate a potential role of different passerine birds species in Mazurian Lake region (northeast Poland) in the spread of Borrelia burgdorferi sensu lato, the spirochaete that causes Lyme disease. A total number of 1254 birds (representing 42 species) were captured during the 3-year study period. Blood samples were collected from birds and analyzed with a nested polymerase chain reaction technique in order to detect fragments of the pathogen DNA. Positive results were obtained in 4.2% of all blood samples. Specifically, B. burgdorferi s.l. were detected in tree pipit (Anthus Erivialis; 21.1% of 19 birds), dunnock (Prunella modularis; 15.8% of 19 birds), chaffinch (Fringilla coelebs; 12.7% of 166 birds), song thrush (Turdus philometos; 9.3% of 54 birds), nuthatch (Sitta euopea; 7.7% of 26 birds), hawfinch (Coccothraustes coccothroustes; 6.7% of 15 birds), robin (Erithacus rebecula; 5.1% of 256 birds), blackbird (Turdus merula; 4.2% of 71 birds) and wren (Troglodytes troglodytes; 3.7% of 27 birds). Additionally, the incidence of the infection was analyzed in relation to the habitat in which the birds resided (mixed coniferous forest or alder swamp forest), months of the study (from April to October), age and sex, but the differences were not statistically significant.