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Abstract: There has been an emergence of tick-borne encephalitis (TBE) cases in western Sweden over the past 10 years. Human cases cluster in distinct regions around major bodies of water, raising the question of TBE prevalence in ticks within these defined localities. This study was based on a collection of 7120 questing nymphs, 510 questing adults, and 132 fed female ticks from cows in 4 suspected TBE foci, based on human cases, and 2 non-endemic areas of western Gotaland. All tick pools were screened with Real-Time RT-PCR targeting the non-coding 3′-region. Prevalence in ticks in the endemic areas ranged from 0.1% to 0.42%, which is comparable with other more established TBE endemic regions in Europe. Of the 18 positive pools, viral copy numbers ranged from 500 to 3.7×109 copies/pool. Sequence data from a TBE patient in western Gotaland confirmed that the western European subtype of TBEV has spread to western Sweden. [Copyright &y& Elsevier]