Human Babesiosis Research Articles

507. Peripheral blood RNA signatures associated with human babesiosis, quality of life and neurological symptoms

Abstract Background Human babesiosis is an emerging tick-borne disease in the US that is primarily caused by the protozoan Babesia microti (Bm). Although Babesiosis can cause life-threatening infections, the pathophysiology including host and parasite determinants associated with disease severity are largely unknown. The aim of this study is to identify parasite and host signatures of human babesiosis and correlate these with disease severity and long-term symptoms. Methods We enrolled prospectively patients with acute babesiosis (microcopy positive blood smear for Bm, confirmed by PCR) at Stony Brook University Hospital during 2020-2021. Blood samples and surveys were completed at initial diagnosis (Visit=1;V1) and at months 1, 6 and 12 (V2,V3,V4). Total RNA from whole peripheral blood was isolated using the PAXgene Blood RNA Kit from individuals infected with Bm and healthy controls. Quality of life (QOL) surveys were administered to patients. Results Peripheral blood transcriptome analysis of 17 patients (median age: 63 years, range 42-78; 24% female) who presented with babesiosis were distinctly different from uninfected individuals (n=9). Babesiosis was associated with a peripheral blood Type I and Type II Interferon transcriptional signature. Other dominant transcriptional signatures included those involved in activation of mononuclear cells, oxidative phosphorylation, inducible nitric oxide synthase and regulation of homeostasis as well as inducible nitric oxide synthase. Ingenuity Pathway Analysis for clinical chemistry and hematology also highlight changes associated with cardiotoxicity, hepatotoxicity, and nephrotoxicity as well as increased levels of red blood cells. Patients with babesiosis showed a persistent deterioration of QOL in cognition [mean V1=33.1; V3=36.3] and satisfaction [mean V1=28.4; V3=34]. Also a decrease in QOL over time despite effective treatment [mean at V1 82 (SD: 5.9), V2 80 (8.1), V3 76 (4.7) (p >0.05). Heat Map Summary. Volcano Plot Gene Pathways Analysis Conclusion Babesiosis is associated with a marked alteration in the peripheral blood transcriptome of patients that may also provide insight into both the pathophysiology of the disease including severity and complications. Furthermore, even after successful treatment, babesiosis may result in long-term impacts on quality of life indicators. Disclosures All Authors: No reported disclosures.

367. Procalcitonin as a Potential Biomarker in the Study of Human Babesiosis

Abstract Background Procalcitonin (Pct) has been gaining momentum as a potential biomarker during parasitic infections as studies have demonstrated that Pct levels are significantly elevated during certain forms of protozoal sepsis. Given this, the goal of this pilot study is to determine the relationship between Pct levels and parasitemia (Par) during infection with Babesia microti and to elucidate the potential for Pct to be used as a biomarker in the study of babesiosis. Methods Babesia cases were collected from Stony Brook University Hospital (SBUH) and South Hampton Hospital (SHH) from 2012 to 2019. Median values of maximum Par and Pct throughout hospitalization were recorded and cases were referenced for admission to the intensive care unit (ICU). Correlation between maximum Par and Pct values were examined with the nonparametric Spearman rank correlation and fractional polynomials were used to assess the functional form of the correlation. Receiver-operator characteristic (ROC) curve analyses were used to identify cutoff points for Pct and maximum Par as markers for prediction of ICU admission. Results A total of 33 patients met the inclusion criteria for acute babesiosis. In fractional polynomial analysis, log-transformed Pct levels had a linear correlation with log-transformed maximum Par, r=0.556 (P=0.001, Fig 1A). In ROC curve analysis, a cut off level of ≥ 1.2 ng/mL for Pct had optimal prediction characteristics for ICU admission (sensitivity 62.5%, specificity 88%; correct classification 82%) with a relevant AUC of 0.77 (95% CI: 0.58–0.89; P=0.005, Fig 1B). A cut off level of ≥ 3.6% for maximum Par had optimal prediction characteristics for ICU admission (sensitivity 75%, specificity 92%, correct classification 88%) with a relevant AUC of 0.75 (95% CI 0.58—0.89; P= 0.005, Fig 1B). Comparison of AUC for Pct and Par yielded no significant difference (P=0.8). Figure 1A. Linear correlation between log10-parasitemia and log10- procalcitonin levels. The linear correlation of the log-transformed variables was r=0.556 (P=0.001), similar to the nonparametric (Spearman) correlation coefficient (ρ=0.567). The red dots represent patients who were admitted to the intensive care unit. Figure 1B. Receiver-operator characteristic curve of parasitemia for prediction of intensive care unit admission (0.75; 95%CI: 0.58—0.89; P=0.005); optimal cut-off is 3.6%. Receiver-operator characteristic curve of procalcitonin for prediction of intensive care unit admission (C=0.77; 95%CI: 0.58–0.89; P=0.005); optimal cut-off is 1.2 ng/mL. Conclusion Our analysis demonstrated a positive linear correlation between Pct and maximum Par, which is the current standard marker for severe disease. Also, our ROC analyses estimate that Pct values ≥ 1.2 ng/mL and Par ≥ 3.6% suggests severe disease and need for ICU admission (Fig 1A, 1B). Given this, our findings suggest that Pct may have potential for predicting severe disease during babesiosis. Disclosures All Authors: No reported disclosures.

Epitope profiling of monoclonal antibodies to the immunodominant antigen BmGPI12 of the human pathogen Babesia microti.

The significant rise in the number of tick-borne diseases represents a major threat to public health worldwide. One such emerging disease is human babesiosis, which is caused by several protozoan parasites of the Babesiagenus of which B. microti is responsible for most clinical cases reported to date. Recent studies have shown that during its intraerythrocytic life cycle, B. microti exports several antigens into the mammalian host using a novel vesicular-mediated secretion mechanism. One of these secreted proteins is the immunodominant antigen BmGPI12, which has been demonstrated to be a reliable biomarker of active B. microti infection. The major immunogenic determinants of this antigen remain unknown. Here we provide a comprehensive molecular and serological characterization of a set of eighteen monoclonal antibodies developed against BmGPI12 and a detailed profile of their binding specificity and suitability in the detection of active B. microti infection. Serological profiling and competition assays using synthetic peptides identified five unique epitopes on the surface of BmGPI12 which are recognized by a set of eight monoclonal antibodies. ELISA-based antigen detection assays identified five antibody combinations that specifically detect the secreted form of BmGPI12 in plasma samples from B. microti-infected mice and humans but not from other Babesia species or P. falciparum.

Phosphatidylinositol 4-kinase is a viable target for the radical cure of Babesia microti infection in immunocompromised hosts.

Human babesiosis is a global emerging tick-borne disease caused by infection with intra-erythrocytic parasites of the genus Babesia. With the rise in human babesiosis cases, the discovery and development of new anti-Babesia drugs are essential. Phosphatidylinositol 4-kinase (PI4K) is a widely present eukaryotic enzyme that phosphorylates lipids to regulate intracellular signaling and trafficking. Previously, we have shown that MMV390048, an inhibitor of PI4K, showed potent inhibition against Babesia species, revealing PI4K as a druggable target for babesiosis. However, twice-administered, 7-day regimens failed to clear Babesia microti parasites from the immunocompromised host. Hence, in this study, we wanted to clarify whether targeting PI4K has the potential for the radical cure of babesiosis. In a B. microti-infected SCID mouse model, a 64-day-consecutive treatment with MMV390048 resulted in the clearance of parasites. Meanwhile, an atovaquone (ATO) resistant parasite line was isolated from the group treated with ATO plus azithromycin. A nonsynonymous variant in the Y272C of the cytochrome b gene was confirmed by sequencing. Likewise, MMV390048 showed potent inhibition against ATO-resistant parasites. These results provide evidence of PI4K as a viable drug target for the radical cure of babesiosis, which will contribute to designing new compounds that can eradicate parasites.

Procalcitonin as a Potential Biomarker in the Study of Babesiosis Caused by B. microti.

Procalcitonin is gaining momentum in the study of protozoal sepsis, but its utility as a biomarker has yet to be fully discovered in human babesiosis. A total of 33 cases of acute babesiosis dating between 2012 and 2019 were retrospectively collected from Stony Brook University Hospital (SBUH) and Stony Brook South Hampton Hospital (SHH), both of which are located on Long Island, NY. Cases were cross-referenced for the need for ICU admission, and the procalcitonin levels were measured by the use of BRAHMS Elecsys assay at SBUH and BRAHMS Architect assay at SHH. Our study demonstrated that the log-transformed procalcitonin levels had a linear correlation with log-transformed maximum parasitemia, which suggests that procalcitonin directly correlates with parasitemia values. Furthermore, when comparing values that predict ICU admission, our ROC analysis of procalcitonin demonstrated similar AUC values to the percentage of parasitemia, suggesting that procalcitonin may assist in determining the severity of disease. We demonstrate that procalcitonin may directly correlate with the parasitemia percentage and have prognostic capabilities, which suggests that procalcitonin may have biomarker potential in human babesiosis.

Nested qPCR assay to detect Babesia duncani infection in hamsters and humans.

Human babesiosis is caused by Babesia duncani that is transmitted through tick bites, blood transfusions, and transplacental transmission. Despite its health burden, diagnostic assays for this pathogen are either unsuitable for clinical applications or have a low detection efficiency; therefore, it remains undetected during transfusion and utilization of blood and blood-component transfusions. This study used a molecular approach via nested quantitative polymerase chain reaction (qPCR) by designing primers and probes corresponding to the variable regions of B. duncani 18S rRNA gene to specifically detect B. duncani DNA in experimentally infected LVG Golden Syrian hamster (n = 70) and human (n = 492; tick bite patients from Gansu Province, China) blood samples. Moreover, comparative analyses of this technique with previously reported nested PCR and microscopy were conducted. The newly optimized diagnostic technique exhibited no cross-reactivity with genomic DNA or plasmids containing the 18S rRNA gene of other zoonotically important Babesia spp., including B. microti, B. divergens, B. crassa, and B. motasi Hebei. The detection limit of nested qPCR was approximately one plasmid copy in 20 μL or one infected red blood cell in 200 μL whole blood. The specificity and sensitivity of the method were 100% and 98.6%, respectively. Comparative analyses revealed that nested qPCR detected B. duncani had relatively higher efficacy and specificity than microscopic examination and nested PCR. The 492 human blood samples were negative for B. duncani infection. Thus, the present study provides an improved diagnostic assay for the efficient and effective detection and analysis of B. duncani infections and its prevalence in infection-prone areas.

BABESIA MICROTI INFECTION WITH LOW-GRADE PARASITEMIA CAUSING ARDS IN AN IMMUNOCOMPETENT PATIENT

BABESIA MICROTI INFECTION WITH LOW-GRADE PARASITEMIA CAUSING ARDS IN AN IMMUNOCOMPETENT PATIENT

Tick-Borne Pathogens Anaplasma phagocytophilum, Babesia odocoilei, and Borrelia burgdorferi Sensu Lato in Blacklegged Ticks Widespread across Eastern Canada

Blacklegged ticks, Ixodes scapularis, can transmit single or multiple infections during a tick bite. These tick-borne, zoonotic infections can become chronic and cause insidious diseases in patients. In the present tick-pathogen study, 138 (48.9%) of 282 ticks collected from 17 sites in 6 geographic area in eastern Canada harbored various combinations of Borrelia burgdorferi sensu lato (Lyme disease), Anaplasma phagocytophilum (human anaplasmosis), and Babesia spp. (human babesiosis). Overall, 167 microbial infections were detected and, of these, 25 ticks had co-infections and two ticks had polymicrobial infections. The prevalence of Babesia spp. was 15.2%, and the ratio of Babesia odocoilei to Babesia microti was 41 to 1 with this sole B. microti being detected in Nova Scotia. Notably, we provide the first documentation of B. odocoilei in the Maritimes. Eastern Ontario had an infection prevalence for B. odocoilei of 25%-the highest among the areas surveyed in this study. By far, the predominant Babesia sp. was B. odocoilei. Based on our findings, health-care practitioners need to recognize that I. scapularis ticks removed from patients may be carrying multiple tick-borne pathogens.

In vivo activity and atom pair fingerprint analysis of MMV665941 against the apicomplexan parasite Babesia microti, the causative agent of babesiosis in humans and rodents

ABSTRACT 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 IC50 MMV665941:0.75 IC50 AV to 0.50 IC50 MMV665941:0.50 IC50 AV. The used combinations were prepared depending on the IC50 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 IC50) 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.

Specific and Sensitive Diagnosis of Babesia microti Active Infection Using Monoclonal Antibodies to the Immunodominant Antigen BmGPI12.

The apicomplexan pathogen Babesia microti is responsible for most cases of human babesiosis worldwide. The disease, which presents as a malaria-like illness, is potentially fatal in immunocompromised or elderly patients, making the need for its accurate and early diagnosis an urgent public health concern. B. microti is transmitted primarily by Ixodes ticks but can also be transmitted via blood transfusion. The parasite completes its asexual reproduction in the host red blood cell, where each invading merozoite develops and multiplies to produce four daughter parasites. While various techniques, such as microscopy, PCR, and indirect fluorescence, have been used over the years for babesiosis diagnosis, detection of the secreted B. microti immunodominant antigen BmGPI12 using specific polyclonal antibodies was found to be the most effective method for the diagnosis of active infection and for evaluation of clearance following drug treatment. Here, we report the development of a panel of 16 monoclonal antibodies against BmGPI12. These antibodies detected secreted BmGPI12 in the plasma of infected humans. Antigen capture assays identified a combination of two monoclonal antibodies, 4C8 and 1E11, as a basis for a monoclonal antibody-based BmGPI12 capture assay (mGPAC) to detect active B. microti infection. Using a collection of 105 previously characterized human plasma samples, the mGPAC assay showed 97.1% correlation with RNA-based PCR (transcription-mediated amplification [TMA]) for positive and negative samples. The mGPAC assay also detected BmGPI12 in the plasma of six babesiosis patients at the time of diagnosis but not in three matched posttreatment samples. The mGPAC assay could thus be used alone or in combination with other assays for accurate detection of active B. microti infection.

Cardiac Complications of Human Babesiosis.

Human babesiosis is a worldwide emerging tick-borne disease caused by intraerythrocytic protozoa. Most patients experience mild to moderate illness, but life-threatening complications can occur. Although cardiac complications are common, the full spectrum of cardiac disease and the frequency, risk factors, and outcomes in patients experiencing cardiac complications are unclear. Accordingly, we carried out a record review of cardiac complications among patients with babesiosis admitted to Yale-New Haven Hospital over the last decade to better characterize cardiac complications of babesiosis. We reviewed the medical records of all adult patients with babesiosis admitted to Yale-New Haven Hospital from January 2011 to October 2021, confirmed by identification of Babesia parasites on thin blood smear and/or by polymerase chain reaction. The presence of Lyme disease and other tick-borne disease coinfections were recorded. Of 163 enrolled patients, 32 (19.6%) had ≥1 cardiac complication during hospitalization. The most common cardiac complications were atrial fibrillation (9.4%), heart failure (8.6%), corrected QT interval prolongation (8.0%), and cardiac ischemia (6.8%). Neither cardiovascular disease risk factors nor preexisting cardiac conditions were significantly associated with the development of cardiac complications. The cardiac complication group had a greater prevalence of high-grade parasitemia (>10%) (P < .001), longer median length of both hospital (P < .001) and intensive care unit stay (P < .001), and a higher mortality rate (P = .02) than the group without cardiac complications. Cardiac complications of acute babesiosis are common and occurred in approximately one-fifth of this inpatient sample. Further investigation is needed to elucidate the relationship between babesiosis severity and cardiac outcomes.

Comparative genomic analysis of Babesia duncani responsible for human babesiosis

BackgroundHuman babesiosis, caused by parasites of the genus Babesia, is an emerging and re-emerging tick-borne disease that is mainly transmitted by tick bites and infected blood transfusion. Babesia duncani has caused majority of human babesiosis in Canada; however, limited data are available to correlate its genomic information and biological features.ResultsWe generated a B. duncani reference genome using Oxford Nanopore Technology (ONT) and Illumina sequencing technology and uncovered its biological features and phylogenetic relationship with other Apicomplexa parasites. Phylogenetic analyses revealed that B. duncani form a clade distinct from B. microti, Babesia spp. infective to bovine and ovine species, and Theileria spp. infective to bovines. We identified the largest species-specific gene family that could be applied as diagnostic markers for this pathogen. In addition, two gene families show signals of significant expansion and several genes that present signatures of positive selection in B. duncani, suggesting their possible roles in the capability of this parasite to infect humans or tick vectors.ConclusionsUsing ONT sequencing and Illumina sequencing technologies, we provide the first B. duncani reference genome and confirm that B. duncani forms a phylogenetically distinct clade from other Piroplasm parasites. Comparative genomic analyses show that two gene families are significantly expanded in B. duncani and may play important roles in host cell invasion and virulence of B. duncani. Our study provides basic information for further exploring B. duncani features, such as host-parasite and tick-parasite interactions.

Babesiosis in Southeastern, Central and Northeastern Europe: An Emerging and Re-Emerging Tick-Borne Disease of Humans and Animals.

There is now considerable evidence that in Europe, babesiosis is an emerging infectious disease, with some of the causative species spreading as a consequence of the increasing range of their tick vector hosts. In this review, we summarize both the historic records and recent findings on the occurrence and incidence of babesiosis in 20 European countries located in southeastern Europe (Bosnia and Herzegovina, Croatia, and Serbia), central Europe (Austria, the Czech Republic, Germany, Hungary, Luxembourg, Poland, Slovakia, Slovenia, and Switzerland), and northern and northeastern Europe (Lithuania, Latvia, Estonia, Iceland, Denmark, Finland, Sweden, and Norway), identified in humans and selected species of domesticated animals (cats, dogs, horses, and cattle). Recorded cases of human babesiosis are still rare, but their number is expected to rise in the coming years. This is because of the widespread and longer seasonal activity of Ixodes ricinus as a result of climate change and because of the more extensive use of better molecular diagnostic methods. Bovine babesiosis has a re-emerging potential because of the likely loss of herd immunity, while canine babesiosis is rapidly expanding in central and northeastern Europe, its occurrence correlating with the rapid, successful expansion of the ornate dog tick (Dermacentor reticulatus) populations in Europe. Taken together, our analysis of the available reports shows clear evidence of an increasing annual incidence of babesiosis across Europe in both humans and animals that is changing in line with similar increases in the incidence of other tick-borne diseases. This situation is of major concern, and we recommend more extensive and frequent, standardized monitoring using a “One Health” approach.

Errata Abound in Human Babesiosis Communique: Retort to Kumar, O'Bryan and Krause

Kumar et al. [1] hereafter called authors, indicate that Babesia microti is the main Babesia species in North America, but skip Babesia odocoilei as the predominant Babesia sp. continent-wide [2-6]. They speculate that babesiosis will merge between the Midwest and the East Coast, but sidestep the fact the B. odocoilei already fills this geographic gap [2-6]. In eastern and central North America, acarology research teams have reported B. odocoilei-infected vector ticks on avian and mammalian hosts and by flagging low-level vegetation [2-8]. One research team found that half of the Babesia spp. detected in adults of blacklegged ticks, Ixodes scapularis, in Maine harbored B. microti, whereas the other half were B. odocoilei [3]. Notably, the Babesia species detected in each of the other three states (i.e., Indiana, Pennsylvania, and Wisconsin) were B. odocoilei.

Establishment of a Transient and Stable Transfection System for Babesia duncani Using a Homologous Recombination Strategy.

Genetic modification provides an invaluable molecular tool to dissect the biology and pathogenesis of pathogens. However, no report is available about the genetic modification of Babesia duncani, a pathogen responsible for human babesiosis that is widespread in North America, suggesting the necessity to develop a genetic manipulation method to improve the strategies for studying and understanding the biology of protozoan pathogens. The establishment of a genetic modification method requires promoters, selectable markers, and reporter genes. Here, the double-copy gene elongation factor-1α (ef-1α) and its promoters were amplified by conventional PCR and confirmed by sequencing. We established a transient transfection system by using the ef-1αB promoter and the reporter gene mCherry and achieved stable transfection through homologous recombination to integrate the selection marker hDHFR-eGFP into the parasite genome. The potential of this genetic modification method was tested by knocking out the thioredoxin peroxidase-1 (TPX-1) gene, and under the drug pressure of 5 nM WR99210, 96.3% of the parasites were observed to express green fluorescence protein (eGFP) by flow cytometry at day 7 post-transfection. Additionally, the clone line of the TPX-1 knockout parasite was successfully obtained by the limiting dilution method. This study provided a transfection method for B. duncani, which may facilitate gene function research and vaccine development of B. duncani.

Call for Caution to Consider Babesia bovis and Babesia bigemina as Anthropozoonotic Agents in Colombia. Comment on Kumar et al. The Global Emergence of Human Babesiosis. Pathogens 2021, 10, 1447.

Currently, six species and two genetic variants within Babesia genus have been confirmed as human pathogens. Babesia bovis and Babesia bigemina are causative agents of bovine babesiosis, and, in spite of the worldwide distribution of those species and their vectors, no description of related human cases has been reported. As a contribution, we would like to address the articles which claim the alleged role of B. bovis and B. bigemina as anthropozoonotic pathogens in Colombia.

A Novel Thioredoxin-Like Protein of Babesia microti Involved in Parasite Pathogenicity.

Babesiosis poses a serious threat to immunocompromised individuals and the major etiological species of Babesia for human babesiosis is Babesia microti. Merozoites are a critical stage in the life cycle of Babesia microti. Several merozoite proteins have been demonstrated to play important roles in this process; however, most of the merozoite proteins of B. microti remain unknown. In the present study, we identified a novel merozoite protein of B. microti with similar structure to the thioredoxin (Trx)-like domain of the Trx family, which was named as B. microti Trx-like protein (BmTLP). Western blot assays demonstrated that this protein was expressed by B. microti during the erythrocytic infection process, and its expression peaked on day 7 post-infection in vivo. Immunofluorescence assay further showed that this protein is mainly expressed in B. microti merozoites. BmTLP hold both heparin- and erythrocyte-binding properties, which are critical functions of invasion-related proteins. Immunization with recombinant BmTLP imparted significant protection against B. microti infection in mice. Taken together, these results suggest that the novel merozoite protein, BmTLP, is an important pathogenic molecule of B. microti and may be a possible target for the design of babesiosis control strategy.

Babesia crassa-Like Human Infection Indicating Need for Adapted PCR Diagnosis of Babesiosis, France.

Human babesiosis in Europe is caused by multiple zoonotic species. We describe a case in a splenectomized patient, in which a routine Babesia divergens PCR result was negative. A universal Babesia spp. PCR yielded a positive result and enabled classification of the parasite into the less-described Babesia crassa–like complex.

The Piroplasmida Babesia, Cytauxzoon, and Theileria in farm and companion animals: species compilation, molecular phylogeny, and evolutionary insights.

The order Piroplasmida, including the genera Babesia, Cytauxzoon, and Theileria is often referred to as piroplasmids and comprises of dixenous hemoprotozoans transmitted by ticks to a mammalian or avian host. Although piroplasmid infections are usually asymptomatic in wild animals, in domestic animals, they cause serious or life-threatening consequences resulting in fatalities. Piroplasmids are particularly notorious for the enormous economic loss they cause worldwide in livestock production, the restrictions they pose on horse trade, and the negative health impact they have on dogs and cats. Furthermore, an increasing number of reported human babesiosis cases are of growing concern. Considerable international research and epidemiological studies are done to identify existing parasite species, reveal their phylogenetic relationships, and develop improved or new drugs and vaccines to mitigate their impact. In this review, we present a compilation of all piroplasmid species, isolates, and species complexes that infect domestic mammals and which have been well defined by molecular phylogenetic markers. Altogether, 57 taxonomic piroplasmid entities were compiled, comprising of 43 piroplasmid species, 12 well-defined isolates awaiting formal species description, and two species complexes that possibly mask additional species. The extrapolation of the finding of at least 57 piroplasmid species in only six domestic mammalian groups (cattle, sheep, goat, horse, dog, and cat) allows us to predict that a substantially higher number of piroplasmid parasites than vertebrate host species exist. Accordingly, the infection of a vertebrate host species by multiple piroplasmid species from the same and/or different phylogenetic lineages is commonly observed. Molecular phylogeny using 18S rRNA genes of piroplasmids infecting domestic mammals results in the formation of six clades, which emerge due to an anthropocentric research scope, but not due to a possibly assumed biological priority position. Scrutinizing the topology of inferred trees reveals stunning insights into some evolutionary patterns exhibited by this intriguing group of parasites. Contrary to expectations, diversification of parasite species appears to be dominated by host-parasite cospeciation (Fahrenholz's rule), and, except for piroplasmids that segregate into Clade VI, host switching is rarely observed. When only domestic mammalian hosts are taken into account, Babesia sensu lato (s.l.) parasites of Clades I and II infect only dogs and cats, respectively, Cytauxzoon spp. placed into Clade III only infect cats, Theileria placed into Clade IV exclusively infect horses, wheras Theileria sensu stricto (s.s.) of Clade V infects only cattle and small ruminants. In contrast, Babesia s.s. parasites of Clade VI infect all farm and companion animal species. We outline how the unique ability of transovarial transmission of Babesia s.s. piroplasmids of Clade VI facilitates species diversification by host switching to other host vertebrate species. Finally, a deterioration of sequence fidelity in databases is observed which will likely lead to an increased risk of artifactual research in this area. Possible measures to reverse and/or avoid this threat are discussed.

Major Surface Antigens in Zoonotic Babesia.

Human babesiosis results from a combination of tick tropism for humans, susceptibility of a host to sustain Babesia development, and contact with infected ticks. Climate modifications and increasing diagnostics have led to an expanded number of Babesia species responsible for human babesiosis, although, to date, most cases have been attributed to B. microti and B. divergens. These two species have been extensively studied, and in this review, we mostly focus on the antigens involved in host–parasite interactions. We present features of the major antigens, so-called Bd37 in B. divergens and BmSA1/GPI12 in B. microti, and highlight the roles of these antigens in both host cell invasion and immune response. A comparison of these antigens with the major antigens found in some other Apicomplexa species emphasizes the importance of glycosylphosphatidylinositol-anchored proteins in host–parasite relationships. GPI-anchor cleavage, which is a property of such antigens, leads to soluble and membrane-bound forms of these proteins, with potentially differential recognition by the host immune system. This mechanism is discussed as the structural basis for the protein-embedded immune escape mechanism. In conclusion, the potential consequences of such a mechanism on the management of both human and animal babesiosis is examined.