Human granulocytic anaplasmosis in Croatia and new insights about anaplasma and ehrlichia species

Abstract

Anaplasma phagocytophilum je emergentni patogen kojeg u Europi prenose krpelji Ixodes ricinus koji su vektori i za virus krpeljnog encefalitisa, Borrelia sensu lato, Babesia i neke vrste Rickettsia spp. A. phagocytophilum je obvezno unutarstanična Gram-negativna bakterija koja ima tropizam za granulocite i uzrokuje humanu granulocitnu anaplazmozu (HGA). Dijagnoza HGAtemelji se na kliničkoj procjeni i mora se potvrditi serološki dokazom serokonverzije ili četverostrukog porasta titra protutijela IgG ili određivanjem DNK. Protutijela su često negativna u početnoj fazi akutne bolesti i obavezno treba testirati parne serume u razmaku od 2–4 tjedna. U ranoj fazi bolesti kada je serologija još negativna značajniji je nalaz DNK iz uzorka krvi s antikoagulansom, ali dostupnost PCR je ograničena. Morule u razmazu periferne krvi obojene po Giemsi značajne su za rano postavljanje dijagnoze, ali se u HGA pronalaze jako rijetko. Od 2009. do 2012.g. u Klinici za infektivne bolesti u Zagreb testirali smo protutijela IgM i IgG protiv A. phagocytophilum u 496 seruma od 425 bolesnika. Parne serume imalo je samo 68 bolesnika. Pozitivna protutijela na A. phagocytophilum nađena su u 160 (37,6%) bolesnika. Tri bolesnika su zadovoljila kriterije za akutnu HGA. Prisutnost IgG u titru 256 ili većem, što se definira kao moguća HGA, nađena je u 40 bolesnika. Samo IgM imalo je 17, a IgM i IgG anti-A. phagocytophilum 16 bolesnika. Anti-A. phagocytophilum IgG u titru 64 ili 128 imalo je u 84 bolesnika. Podaci potvrđuju prisutnost infekcije A. phagocytophilum u Hrvatskoj premda većina akutnih infekcija ostaje nedokazana, većinom kao samoizlječive bolesti. HGA se treba uključiti u diferencijalnu dijagnozu bolesnika sa simptomima sličnima gripi u područjima gdje se nalazi Ixodes ricinus, posebno u vrijeme aktivnosti krpelja. U životinja i krpelja u Hrvatskoj utvrđena je prisutnost i drugih patogenih članova porodice Anaplasmataceae zbog čega dodatno treba misliti o HGA uz veća dijagnostička nastojanja za postavljanje dijagnoze.Anaplasma phagocytophilum is an emergent tick-born pathogen in Europe transmitted by Ixodes ricinus ticks which may also transmit tick-borne encephalitis virus, Borrelia sensu lato, Babesia and some Rickettsia spp. A. phagocytophilum is an obligate intracellular Gram- negative bacteria that has tropism for granulocytes and causes human granulocytic anaplasmosis (HGA). The diagnosis of HGA relies on clinical suspicion and must be confirmed with seroconversion or 4-fold increase in antibody titre or by DNA determination. Antibodies are often negative in the initial phase of acute illness and paired sera taken 2–4 weeks later are obligatory. PCR on anticoagulated blood could be more efficient tool in this phase but it is of limited availability. Morulae in Giemsa-stained peripheral blood smears may provide early diagnosis but could be observed very rarely in HGA. From 2009 till 2012, we tested 496 sera from 425 patients for IgM and IgG anti-A. phagocytophilum antibodies at the University Hospital for Infectious Diseases in Zagreb. Paired sera were sent for only 68 patients. Positive antibodies against A. phagocytophilum were found in 160 (37.6%) patients. Three patients fulfilled the criteria for acute HGA. The presence of IgG in titres 256 or higher defined as probable HGA was found in 40 patients. Only IgM, and IgM and IgG anti-A. phagocytophilum antibodies were nadetected in 17 and 16 patients, respectively. Anti-A. phagocytophilum IgG in titres 64 or 128 was found in 84 patients. The data show that A. phagocytophilum infections are present in Croatia, although most of the acute infections remain unconfirmed as self-resolved diseases. HGA should be included in the differential diagnosis of patients with flu-like illness in regions with Ixodes ricinus, especially during tick-activity season. Different members of Anaplasmataceae family were found in animals and ticks in Croatia, which is why better awareness of HGAand diagnostics effort are needed