Lyme disease caused by B. burgdorferi and B. garinii has been traditionally treated with antibiotics. However, to eradicate all three morphological forms of Borrelia species such as spirochetes, rounded bodies and biofilm more effective and safe treatment plan is required. In the instant application, Vitamins and Phytobiologicals have been tested for as a short term and long term treatment in vitro and on biofilm. The combination of tested compounds in form of Mix AO seems to be very beneficial in treating Lyme disease caused by B. burgdorferi and B. garinii.