Retrospective study of the epidemiological risk and serological diagnosis of human babesiosis in Asturias, Northwestern Spain

Abstract

Background: Babesiosis is a globally growing tick-borne disease in humans. Severe babesiosis caused by Babesia divergens has been reported in two patients from Asturias (Northwestern Spain), suggesting an undetected risk for the disease. To analyze this risk, we retrospectively evaluated the seroprevalence of babesiosis in the Asturian population from 2015 through 2017, a period covering the intermediate years in which these two severe cases occurred.

Methods: Indirect fluorescent assay (IFA) and Western blot (WB) were performed to detect B. divergens IgG antibodies in 120 serum samples from Asturian patients infected with the tick-transmitted spirochete Borrelia burgdorferi sensu lato, a condition that indicates exposure to tick bites.

Results: This retrospective study confirmed a B. divergens seroprevalence rate of 39.2% according to IFA results. B. divergens incidence was 7.14 cases/100,000 population, exceeding previously reported seroprevalence rates. No differences in epidemiology and risk factors were found between patients infected solely with B. burgdorferi s.l. and those infected with B. burgdorferi s.l. and with IgG antibodies against B. divergens. This last group of patients lived in Central Asturias, had a milder clinical course and, according to WB results, developed different humoral responses against B. divergens.

Conclusions: Babesia divergens parasites have circulated for several years in Asturias. Epidemiological evidence of babesiosis makes Asturias an emerging risk area for this zoonosis. Human babesiosis could also be relevant in other Spanish and European regions affected by borreliosis. Hence, the potential risk of babesiosis on human health in Asturias and other European forest regions needs to be addressed by the health authorities.

Keywords: Babesia divergens; Epidemiological risk; Serological diagnosis; Ticks; Vector-borne diseases.

Fig. 1

Serological detection of specific anti-B. divergens IgG antibodies by Western blot (WB). Panel a: WB was prepared by using B. divergens (Rouen 87) and non-infected human erythrocyte protein extracts. Blots were incubated with sera of patients infected with B. burgdorferi s.l. and positive and negative controls. Lanes 1–40: different B. divergens proteins, ranging from 25 to 75 kDa, that were identified from serum patient samples. Protein bands differed in number, frequency, size and intensity between patients. Lanes 41–47: some of the serum samples from patients infected with B. burgdorferi s.l. that did not recognize B. divergens proteins. Lanes 48–53 and lines C1+, C2+ and C3+: Some of the serum samples and positive controls that did not recognize erythrocyte proteins. Serum samples that reacted against a native B. divergens protein with ~ 37 kDa (*). Serum samples that reacted against native B. divergens proteins of ~ 48–63 kDa (+). Panel b: WB was prepared by using the recombinant GST-rBd37 (Rouen 87) protein and the GST fusion partner for Bd37. Specificity from the patients’ sera against recombinant Bd37 from B. divergens was observed by WB. Lanes A and B: GST-rBd37 (59.5 kDa) and GST (26 kDa) recombinant proteins, respectively, subjected to SDS-PAGE. The number of the lanes indicates which serum sample from Panel a recognized the GST-rBd37 protein. Positive control 1 (C1+): serum sample from an Asturian patient [23]. Positive control 2 (C2+): serum sample from a non-Asturian Spanish patient co-infected with HIV and B. divergens [25]. Positive control 3 (C3+): serum sample from a Finnish patient doubly infected with B. burgdorferi s.l and B. divergens. Negative control (C−) consisted of a human sera pool negative for different infectious diseases, including toxoplasmosis, malaria and babesiosis. None of the sera reacted with the recombinant GST protein

Fig. 2

Geographical distribution of patients who had antibodies against B. divergens. Map shows the location of Asturias in Spain and the Asturian councils where B. divergens infected patients lived from 2015 through 2017. The number of infected patients appears associated to each council. The map also shows the councils where severe (*) and fatal (+) human babesiosis cases occurred in Asturias [16, 17]. The color blocks represent the range of incidence of babesiosis per 100,000 population in each council