Animal models of human babesiosis have provided a basic understanding of the immunological mechanisms that clear, or occasionally exacerbate, Babesia infection and those pathological processes that ...cause disease complications. Human Babesia infection can cause asymptomatic infection, mild to moderate disease, or severe disease resulting in organ dysfunction and death. More than 100 Babesia species infect a wide array of wild and domestic animals, and many of the immunologic and pathologic responses to Babesia infection are similar in animals and humans. In this review, we summarize the knowledge gained from animal studies, their limitations, and how animal models or alternative approaches can be further leveraged to improve our understanding of human babesiosis.
Human babesiosis is a worldwide emerging tick-borne infection that can cause severe complications and death.Animal models have provided insight into the immunology and pathogenesis of three human Babesia pathogens: Babesia microti, Babesia duncani, and Babesia divergens.Both innate and adaptive immunity are important in clearing infection, including the spleen, liver, macrophages, natural killer cells, cytokines, CD4+ T lymphocytes, B lymphocytes and antibody.Theories of babesiosis complications derived from animal model studies include erythrocyte cytoadherence with vascular obstruction and excessive proinflammatory cytokine production.Investment should be made to support translational immunologic and pathogenic studies and to develop further refinements in animal model and non-animal model approaches, such as human organoids, stem-cell-derived three-dimensional (3D) culture systems, direct human challenge, and large genomics.
•Babesiosis: A disease of veterinary importance with an empty drug pipeline.•There is a wide gap in the treatment of animal babesiosis.•Empty anti-Babesia drug pipeline was mainly due to the absence ...of HTS drug system.•SG I fluorescence assay has helped to fulfil the anti-Babesia drug discovery gap.•Establishing of anti- B. gibsoni, B. canis, and B. ovis HTS drug system is required.
Absence of an effective high-throughput drug-screening system for Babesia parasites is considered one of the main causes for the presence of a wide gap in the treatment of animal babesiosis when compared with other hemoprotozoan diseases, such as malaria. Recently, a simple, accurate, and automatic fluorescence assay was established for large-scale anti-Babesia (B. bovis, B. bigemina, B. divergens, B. caballi and T. equi) drug screening. Such development will facilitate anti-Babesia drug discovery, especially in the post-genomic era, which will bring new chemotherapy targets with the completion of the Babesia genome sequencing project currently in progress. In this review, we present the current progress in the various assays for in vitro and in vivo anti-Babesia drug testing, as well as the challenges, highlighting new insights into the future of anti-Babesia drug screening.
There is an innovative approach to discovering and developing novel potent and safe anti-Babesia and anti-Theileria agents for the control of animal piroplasmosis. Large-scale screening of 400 ...compounds from a Malaria Box (a treasure trove of 400 diverse compounds with antimalarial activity has been established by Medicines for Malaria Venture) against the in vitro growth of bovine Babesia and equine Babesia and Theileria parasites was performed, and the data were published in a brief with complete dataset from 236 screens of the Malaria Box compounds. Therefore, in this review, we explored and discussed in detail the in vitro inhibitory effects of 400 antimalarial compounds (200 drug-like and 200 probe-like) from the Malaria Box against Babesia (B.) bovis, B. bigemina, B. caballi, and Theileria (T.) equi. Seventeen hits were the most interesting with regard to bovine Babesia parasites, with mean selectivity indices (SIs) greater than 300 and half maximal inhibitory concentration (IC
s) ranging from 50 to 410 nM. The most interesting compounds with regard to equine Babesia and Theileria parasites were MMV020490 and MMV020275, with mean SIs > 258.68 and >251.55, respectively, and IC
s ranging from 76 to 480 nM. Ten novel anti-B. bovis, anti-B. bigemina, anti-T. equi, and anti-B. caballi hits, MMV666093, MMV396794, MMV006706, MMV665941, MMV085203, MMV396693, MMV006787, MMV073843, MMV007092, and MMV665875, with nanomole levels of IC
were identified. The most interesting hits were MMV396693, MMV073843, MMV666093, and MMV665875, with mean SIs greater than 307.8 and IC
s ranging from 43 to 630 nM for both bovine Babesia and equine Babesia and Theileria parasites. Screening the Malaria Box against the in vitro growth of Babesia and Theileria parasites helped with the discovery of new drugs than those traditionally used, diminazene aceturate and imidocarb dipropionate, and indicated the potential of the Malaria Box in finding new, potent antibabesial drugs.
The in vitro inhibitory effect of two fluroquinolone antibiotics, norfloxacin and ofloxacin, was evaluated in this study on the growth of several Babesia and Theileria parasites with highlighting the ...bioinformatic analysis for both drugs with the commonly used antibabesial drug, diminazene aceturate (DA), and the recently identified antibabesial drugs, luteolin, and pyronaridine tetraphosphate (PYR).
The antipiroplasm efficacy of screened fluroquinolones in vitro and in vivo was assessed using a fluorescence-based SYBR Green I assay. Using atom Pair signatures, we investigated the structural similarity between fluroquinolones and the antibabesial drugs.
Both fluroquinolones significantly inhibited (P < 0.05) the in vitro growths of Babesia bovis (B. bovis), B. bigemina, B. caballi, and Theileria equi (T. equi) in a dose-dependent manner. The best inhibitory effect for both drugs was observed on the growth of T. equi. Atom Pair fingerprints (APfp) results and AP Tanimoto values revealed that both fluroquinolones, norfloxacin with luteolin, and ofloxacin with PYR, showed the maximum structural similarity (MSS). Two drug interactions findings confirmed the synergetic interaction between these combination therapies against the in vitro growth of B. bovis and T. equi.
This study helped in discovery novel potent antibabesial combination therapies consist of norfloxacin/ofloxacin, norfloxacin/luteolin, and ofloxacin/PYR.
Theileria equi, an intraerythrocytic protozoan parasite, causes equine piroplasmosis, a disease which negatively impacts the global horse industry. Genetic manipulation is one of the research tools ...under development as a control method for protozoan parasites, but this technique needs to be established for T. equi. Herein, we report on the first development of a stable transgenic T. equi line expressing enhanced green fluorescent protein/blasticidin S deaminase (eGFP/BSD). To express the exogenous fusion gene in T. equi, regulatory regions of the elongation factor-1 alpha (ef-1α) gene were identified in T. equi. An eGFP/BSD-expression cassette containing the ef-1α gene promoter and terminator regions was constructed and integrated into the T. equi genome. On day 9 post-transfection, blasticidin-resistant T. equi emerged. In the clonal line of T. equi obtained by limiting dilution, integration of the eGFP/BSD-expression cassette was confirmed in the designated B-locus of the ef-1α gene via PCR and Southern blot analyses. Parasitaemia dynamics between the transgenic and parental T. equi lines were comparable in vitro. The eGFP/BSD-expressing transgenic T. equi and the methodology used to generate it offer new opportunities for better understanding of T. equi biology, with the add-on possibility of discovering effective control methods against equine piroplasmosis.
The effect of Zingiber officinale rhizome methanolic extract (ZOR) on the in vitro growth of bovine Babesia (B. bovis, B. bigemina, and B. divergens) and equine piroplasm (B. caballi, and Theileria ...equi) parasites and on the growth of B. microti in mice was evaluated in this study. The possible in vitro synergistic interaction between ZOR and either diminazene aceturate (DA) or potent Medicines for Malaria Venture (MMV) hits from the malaria box was also investigated. In vitro, ZOR reduced the growth of B. bovis, B. bigemina, T. equi, and B. caballi in a dose-dependent manner. B. divergens was the most susceptible parasite to the in vitro inhibitory effect of ZOR. DA and MMV compounds enhanced the in vitro inhibitory antibabesial activity of ZOR. 12.5 mg/kg DA when administrated in combination with ZOR in mice exhibited a significant inhibition (P < 0.05) in B. microti growth better than those observed after treatment with 25 mg/kg DA monotherapy. These findings suggest that ZOR could be a viable medicinal plant for babesiosis treatment, particularly when combined with a modest dose of either DA or powerful anti-B. bigemina MMV hits.
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•ZOR/DA combination inhibited the in vitro and in vivo growth of piroplasm.•MMV hits from malaria box enhanced the in vitro inhibitory effect of ZOR.•ZOR/DA eliminated the B. microti from the blood and spleen tissues of treated mice.•ZOR exhibited a significant delayed inhibitory effect on the growth of B. bigemina.
There are no effective vaccines against Babesia and Theileria parasites; therefore, therapy depends heavily on antiprotozoal drugs. Treatment options for piroplasmosis are limited; thus, the need for ...new antiprotozoal agents is becoming increasingly urgent. Ellagic acid (EA) is a polyphenol found in various plant products and has antioxidant, antibacterial and effective antimalarial activity in vitro and in vivo without toxicity. The present study documents the efficacy of EA and EA-loaded nanoparticles (EA-NPs) on the growth of Babesia and Theileria.
In this study, the inhibitory effect of EA, β-cyclodextrin ellagic acid (β-CD EA) and antisolvent precipitation with a syringe pump prepared ellagic acid (APSP EA) was evaluated on four Babesia species and Theileria equi in vitro, and on the multiplication of B. microti in mice. The cytotoxicity assay was tested on Madin-Darby bovine kidney (MDBK), mouse embryonic fibroblast (NIH/3T3) and human foreskin fibroblast (HFF) cell lines.
The half-maximal inhibitory concentration (IC
) values of EA and β-CD EA on B. bovis, B. bigemina, B. divergens, B. caballi and T. equi were 9.58 ± 1.47, 7.87 ± 5.8, 5.41 ± 2.8, 3.29 ± 0.42 and 7.46 ± 0.6 µM and 8.8 ± 0.53, 18.9 ± 0.025, 11 ± 0.37, 4.4 ± 0.6 and 9.1 ± 1.72 µM, respectively. The IC
values of APSP EA on B. bovis, B. bigemina, B. divergens, B. caballi and T. equi were 4.2 ± 0.42, 9.6 ± 0.6, 2.6 ± 1.47, 0.92 ± 5.8 and 7.3 ± 0.54 µM, respectively. A toxicity assay showed that EA, β-CD EA and APSP EA affected the viability of cells with a half-maximal effective concentration (EC
) higher than 800 µM. In the experiments on mice, APSP EA at a concentration of 70 mg/kg reduced the peak parasitemia of B. microti by 68.1%. Furthermore, the APSP EA-atovaquone (AQ) combination showed a higher chemotherapeutic effect than that of APSP EA monotherapy.
To our knowledge, this is the first study to demonstrate the in vitro and in vivo antibabesial action of EA-NPs and thus supports the use of nanoparticles as an alternative antiparasitic agent.
Due to drug resistance, commonly used anti-
drugs have limited efficacy against babesiosis and inflict severe side effects. Tafenoquine (TAF) was approved by the U.S. Food and Drug Administration in ...2018 for the radical cure of Plasmodium vivax infection and for malaria prophylaxis. Here, we evaluated the efficacy of TAF for the treatment of
infection and elucidated the suspected mechanisms of TAF activity against
parasites. Parasitemia and survival rates of Babesia rodhaini-infected BALB/c and SCID mice were used to explore the role of the immune response in
infection after TAF treatment. Parasitemia, survival rates, body weight, vital signs, complete blood count, and blood biochemistry of B. gibsoni-infected splenectomized dogs were determined to evaluate the anti-
activity and side effects of TAF. Then, to understand the mechanism of TAF activity, hydrogen peroxide was used as an oxidizer for short-term B. rodhaini incubation
, and the expression levels of antioxidant enzymes were confirmed using B. microti-infected mice by reverse transcription-quantitative PCR (qRT-PCR). Acute
and
infections were rapidly eliminated with TAF administration. Repeated administration of TAF or a combination therapy with other antibabesial agents is still needed to avoid a potentially fatal recurrence for immunocompromised hosts. Caution about hyperkalemia should be taken during TAF treatment for
infection. TAF possesses a babesicidal effect that may be related to drug-induced oxidative stress. Considering the lower frequency of glucose-6-phosphate dehydrogenase deficiency in animals compared to that in humans, TAF use on
-infected farm animals and pets is eagerly anticipated.
The effect of MMV665941 on the growth of Babesia microti (B. microti) in mice, was investigated in this study using a fluorescence-based SYBR Green I test. Using atom Pair signatures, we investigated ...the structural similarity between MMV665941 and the commonly used antibabesial medicines diminazene aceturate (DA), imidocarb dipropionate (ID), or atovaquone (AV). In vitro cultures of Babesia bovis (B. bovis) and, Theileria equi (T. equi) were utilized to determine the MMV665941 and AV interaction using combination ratios ranged from 0.75 IC
50
MMV665941:0.75 IC
50
AV to 0.50 IC
50
MMV665941:0.50 IC
50
AV. The used combinations were prepared depending on the IC
50
of each drug against the in vitro growth of the tested parasite. Every 96 h, the hemolytic anemia in the treated mice was monitored using a Celltac MEK-6450 computerized hematology analyzer. A single dose of 5 mg/kg MMV665941 exhibited inhibition in the B. microti growth from day 4 post-inoculation (p.i.) till day 12 p.i. MMV665941 caused 62.10%, 49.88%, and 74.23% inhibitions in parasite growth at days 4, 6 and 8 p.i., respectively. Of note, 5 mg/kg MMV665941 resulted in quick recovery of hemolytic anemia caused by babesiosis. The atom pair fingerprint (APfp) analysis revealed that MMV665941 and atovaquone (AV) showed maximum structural similarity. Of note, high concentrations (0.75 IC
50
) of MMV665941 and AV caused synergistic inhibition on B. bovis growth. These findings suggest that MMV665941 might be a promising drug for babesiosis treatment, particularly when combined with the commonly used antibabesial drug, AV.
Human babesiosis is the most important zoonotic protozoan infection in the world. This is the first report of the cloning, expression, purification, and immunobiochemical characterization of a ...methionine aminopeptidase 1 (MetAP1) protein from
Babesia microti
(
B. microti
). The gene encodes a MetAP1 protein of
B. microti
(BmMetAP1) of approximately 66.8 kDa that includes glutathione S-transferase (GST) tag and shows MetAP activity. BmMetAP1 was detected in a lysate of
B. microti
and further localized in cytoplasm of the
B. microti
merozoite. rBmMetAP1 was found to be immunogenic, eliciting a high antibody titer in mice. Moreover, rBmMetAP1 stimulated the production of IFN-γ and IL-12 but not IL-4. Finally, rBmMetAP1 was able to provide considerable protection to mice against a
B. microti
challenge infection based on a reduction in peak parasitemia levels and earlier clearance of the parasite as compared with control mice. Taken together, these results suggest that rBmMetAP1 confers significant protection against experimental
B. microti
infection and might be considered a potential vaccine target against human babesiosis.
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