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BackgroundBovine babesiosis, mainly caused by Babesia bovis and B. bigemina, is a huge threat to the livestock industry. In Indonesia, the current distribution of the disease is unknown due to a lack of scientific study.MethodsIn the present study, 487 blood samples were collected from cattle with different breeding and age groups in a broad geographical area across the archipelago. The presence of antibodies and current infections of B. bovis and B. bigemina were determined using enzyme-linked immunosorbent assay (ELISA), immunochromatographic test (ICT), and nested PCR (nPCR) targeting B. bovis SBP-4 and B. bigemina RAP-1a genes. Sequence analysis was performed to the amplicon of B. bovis SBP-4, B. bigemina RAP-1a, and internal transcribed spacer (ITS) region of ribosomal RNA of both Babesia species.ResultsIn total, B. bovis positives were detected by ELISA, single-ICT, dual-ICT and nPCR in 340 (69.8%), 317 (65.1%), 307 (63.0%) and 247 (50.7%) samples, respectively. For B. bigemina, the positive samples were detected in 134 (27.5%), 130 (26.7%), 127 (26.1%) and 93 (19.1%), respectively. Furthermore, mixed infections were found in 125 (25.7%), 113 (23.2%), 109 (22.4%) and 52 (10.7%) samples, respectively, which occurred only by chance and were not influenced by additional factors. The obtained nucleotide sequences of B. bovis SBP-4 and B. bigemina RAP-1a genes showed a high homology with other isolates from different countries. Further nucleotide sequence analysis using ITS region showed a great genetic diversity of B. bovis isolates among sampling locations; a lower diversity was found in B. bigemina ITS isolates.ConclusionsThese data revealed the current distribution of B. bovis and B. bigemina infection in cattle in Indonesia. The rate of infection varied among sampling locations, cattle breeds and age groups. Furthermore, B. bovis ITS isolates from Indonesia were found to be more genetically diverse than B. bigemina ITS isolates. The data presented in this study are necessary to develop an effective strategy for controlling the disease in the country.Electronic supplementary materialThe online version of this article (10.1186/s13071-017-2502-0) contains supplementary material, which is available to authorized users.

Heat shock protein 90 (Hsp90) is a chaperone protein that stabilizes cells during stress or non-stress responses. Previous reports have shown that Hsp90 is a potential drug target to suppress the multiplication of several protozoan parasites. In this study, 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), an Hsp90 inhibitor, was evaluated for its inhibitory effect on five in vitro cultures of Babesia and Theileria species, including B. bovis, B. bigemina, B. divergens, B. caballi, and T. equi, and on the multiplication of a B. microti–infected mouse model. 17-DMAG showed the inhibitory effect in all of the species tested. The half maximum inhibition concentration (IC50) of 17-DMAG on B. bovis, B. bigemina, B. divergens, B. caballi, and T. equi was 77.6 ± 2.9, 62.4 ± 1.9, 183.8 ± 3.2, 88.5 ± 9.6, and 307.7 ± 7.2 nM, respectively. The toxicity assay on MDBK and NIH/3T3 cell lines showed that 17-DMAG affected the viability of cells with an IC50 of 15.5 ± 4 and 8.8 ± 2 μM, respectively. Since the IC50s were much lower on the parasites than on the host cell lines, the selectivity index were high for all tested species. Furthermore, the two-drug combination of 17-DMAG with diminazene aceturate (DA) and atovaquone (AV) showed synergism or addition on in vitro cultures of Babesia and Theileria parasites. In the mouse model, 17-DMAG at a concentration of 30 mg/kg BW effectively inhibited the multiplication of B. microti. Moreover, if combined with DA or AV, 17-DMAG showed a comparable inhibition at the half dose. Taken together, these results indicate that 17-DMAG is a potent drug for treating piroplamosis. The data warrant further evaluation of 17-DMAG as an antibabesial drug and as an option in combination with atovaquone for the treatment of human babesiosis.

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The present study evaluated the growth-inhibitory effects of clofazimine, currently used for treating leprosy, against Babesia bovis, B. bigemina, B. caballi, and Theileria equi in in vitro culture and against Babesia microti in mice. The 50% inhibitory concentrations (IC 50 s) of clofazimine against the in vitro growth of B. bovis, B. bigemina, B. caballi, and T. equi were 4.5, 3, 4.3, and 0.29 M, respectively. In mice infected with B. microti, treatment with 20 mg/kg of body weight of clofazimine administered orally resulted in a significantly lower peak parasitemia (5.3%) than that in the control group (45.9%), which was comparable to the subcutaneous administration of 25 mg/kg diminazene aceturate, the most widely used treatment for animal piroplasmosis. Although slight anemia was observed in both clofazimine-and diminazene aceturate-treated infected mice, the level and duration of anemia were lower and shorter, respectively, than those in untreated infected mice. Using blood transfusions and PCR, we also examined whether clofazimine completely killed B. microti. On day 40 postinfection, when blood analysis was performed, parasites were not found in blood smears; however, the DNA of B. microti was detected in the blood of clofaziminetreated animals and in several tissues of clofazimine-and diminazene aceturate-treated mice by PCR. The growth of parasites was observed in mice after blood transfusions from clofazimine-treated mice. In conclusion, clofazimine showed excellent inhibitory effects against Babesia and Theileria in vitro and in vivo, and further study on clofazimine is required for the future development of a novel chemotherapy with high efficacy and safety against animal piroplasmosis and, possibly, human babesiosis.