Linköping University Medical Dissertations No. 1315 Tick-Borne Infections in Humans Aspects of immunopathogenesis, diagnosis and co-infections with Borrelia burgdorferi and Anaplasma phagocytophilum Marika Nordberg Division of Infectious Diseases and Clinical Immunology Department of Clinical and Experimental Medicine Faculty of Health Sciences Linköping University, Sweden Linköping 2012 Marika Nordberg, 2012 Cover photo: Mari-Anne Åkeson, Bengt-Arne Fredriksson, Linköping University, Sweden. Published articles have been reprinted with the permission of the respective copyright holders. Printed in Sweden by LiU-Tryck, Linköping, Sweden, 2012 ISBN 978-91-7519-852-1 ISSN 0345-0082 To my family worldwide För att man ska kunna flyga måste modet vara aningen större än rädslan och en gynnsam vind råda Ur: Instruktion för skalbaggar/ Margareta Ekstrand “The Borrelia war” by Aaron Nordberg, seven years old 2009. The immune system fight- ing against the Borrelia bacteria. Contents CONTENTS ABSTRACT .................................................................................................................... 1 SAMMANFATTNING PÅ SVENSKA ......................................................................... 3 LIST OF PAPERS ........................................................................................................... 5 ABBREVIATIONS ......................................................................................................... 6 INTRODUCTION ........................................................................................................... 7 Framework and context ............................................................................................ 7 Tick-borne infections ................................................................................................ 8 Tick – the vector ....................................................................................................... 9 Lyme borreliosis ..................................................................................................... 10 Borrelia burgdorferi sensu lato – the pathogen ..................................................... 11 Epidemiology.......................................................................................................... 12 Tick-borne infections on the Åland Islands ...................................................... 13 Clinical manifestations of Lyme borreliosis .......................................................... 14 Erythema migrans ............................................................................................ 15 Lymphocytoma .................................................................................................. 17 Acrodermatitis chronica atrophicans ............................................................... 17 Neuroborreliosis ............................................................................................... 18 Lyme arthritis ................................................................................................... 19 Other Lyme borreliosis manifestations ............................................................ 19 Laboratory methods of detecting B. burgdorferi .................................................... 20 Direct detection ................................................................................................ 20 Indirect detection .............................................................................................. 21 Treatment ................................................................................................................ 23 Persisting symptoms after treatment ................................................................ 24 Human granulocytic anaplasmosis (HGA) ............................................................. 25 Anaplasma phagocytophilum – the pathogen ......................................................... 25 Epidemiology.......................................................................................................... 27 Clinical manifestations of HGA ............................................................................. 28 Methods of detecting A. phagocytophilum ............................................................. 29 Contents Direct detection ................................................................................................ 29 Indirect detection .............................................................................................. 30 Treatment ................................................................................................................ 31 Differences between Europe and the USA ............................................................. 32 Tick-borne encephalitis (TBE) ............................................................................... 32 Epidemiology.......................................................................................................... 33 Clinical manifestations of TBE .............................................................................. 34 Co-infections and other tick-borne infections ........................................................ 34 Co-infections and immunology ......................................................................... 35 Immunology ............................................................................................................ 36 Innate immune system ...................................................................................... 36 Adaptive immune system .................................................................................. 38 T helper cell subpopulations ............................................................................ 38 Cytotoxic mechanisms ...................................................................................... 40 Cytokines and chemokines ................................................................................ 42 Immunology of Lyme borreliosis ...................................................................... 44 Immunology of human granulocytic anaplasmosis .......................................... 46 HYPOTHESIS ............................................................................................................... 49 AIMS ............................................................................................................................. 50 MATERIALS AND METHODS .................................................................................. 51 Subjects (paper I-IV) .............................................................................................. 51 Patients and controls ........................................................................................ 51 Case definitions used in this thesis ................................................................... 55 Excluded subjects ............................................................................................. 57 Controls ............................................................................................................ 58 Evaluation of data ............................................................................................ 58 Methods .................................................................................................................. 59 Borrelia antibody analyses in serum (Papers I-IV) ......................................... 59 CSF analyses (Papers II, IV) ............................................................................ 59 TBEV serology .................................................................................................. 59 Preparation of cells in blood and CSF (Papers I, II, IV) ................................. 60 Preparation of Borrelia outer surface protein antigen (Papers I, II, IV) ........ 60 ELISPOT- enzyme-linked immunospot assay (Papers I, II, IV) ....................... 60 Contents Indirect immunofluorescence assay (Paper III) ............................................... 64 Polymerase chain reaction (Paper III) ............................................................ 66 Statistics .................................................................................................................. 68 Ethical considerations ............................................................................................. 68 RESULTS AND DISCUSSION .................................................................................... 69 Lyme neuroborreliosis; clinical manifestations, immunopathogenesis and diagnostic aspects (Papers II, IV) ........................................................................... 69 Patient characteristics (Papers II, IV) ............................................................. 69 Clinical manifestations of Lyme neuroborreliosis (Paper II) .......................... 69 Immune responses in adults with Lyme neuroborreliosis (Paper II) ............... 72 IL-17 in CSF in patients diagnosed with LNB (Paper II) ................................ 75 ELISPOT assay in laboratory diagnosis of Lyme neuroborreliosis in clinical practice (Paper IV) .............................................................................. 77 Diagnostic performance of ELISPOT-test ....................................................... 78 Methodological considerations – ELISPOT assay ........................................... 79 Tick-borne co-infections; epidemiology and immunological mechanisms (Papers I, III) ..........................................................................................................