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Analysis of the Babesia microti proteome in infected red blood cells by a combination of nanotechnology and mass spectrometry.

ABSTRACT: Proteomics of Babesia microti has lagged behind other apicomplexans despite recent genome and transcriptome studies. Here, we used a combination of nanotechnology and mass spectrometry to provide a proteomic profile of B. microti acute infection. We identified ∼500 parasite proteins in blood with functions such as transport, carbohydrate and energy metabolism, proteolysis, DNA and RNA metabolism, signaling, translation, lipid biosynthesis, and motility and invasion. We also identified surface antigens with roles in the immune response to the parasite. This first evaluation of the B. microti proteome in erythrocytes provides information for the study of intracellular survival and development of diagnostic tools using mass spectrometry.

SUBMITTER: Magni R

PROVIDER: S-EPMC10548858 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Analysis of the Babesia microti proteome in infected red blood cells by a combination of nanotechnology and mass spectrometry.

Magni Ruben R Luchini Alessandra A Liotta Lance L Molestina Robert E RE

International journal for parasitology 20181101 2

Proteomics of Babesia microti has lagged behind other apicomplexans despite recent genome and transcriptome studies. Here, we used a combination of nanotechnology and mass spectrometry to provide a proteomic profile of B. microti acute infection. We identified ∼500 parasite proteins in blood with functions such as transport, carbohydrate and energy metabolism, proteolysis, DNA and RNA metabolism, signaling, translation, lipid biosynthesis, and motility and invasion. We also identified surface an ...[more]

PMID: 30391228

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Project description:Babesiosis is a zoonosis and an important blood-borne human parasitic infection that has gained attention because of its growing infection rate in humans by transfer from animal reservoirs. Babesia represents a potential threat to the blood supply because asymptomatic infections in man are common, and blood from such donors can cause severe disease in certain recipients. Extracellular vesicles (EVs) are vesicles released by cells that contain a complex mixture of proteins, lipids, glycans, and genetic information that have been shown to play important roles in disease pathogenesis and susceptibility, as well as cell-cell communication and immune responses. In this article, we report on the identification and characterization of EVs released from red blood cells (RBCs) infected by two major human Babesia species-Babesia divergens from in vitro culture and those from an in vivo B. microti mouse infection. Using nanoparticle tracking analysis, we show that there is a range of vesicle sizes from 30 to 1,000 nm, emanating from the Babesia-infected RBC. The study of these EVs in the context of hemoparasite infection is complicated by the fact that both the parasite and the host RBC make and release vesicles into the extracellular environment. However, the EV frequency is 2- to 10-fold higher in Babesia-infected RBCs than uninfected RBCs, depending on levels of parasitemia. Using parasite-specific markers, we were able to show that ~50%-60% of all EVs contained parasite-specific markers on their surface and thus may represent the specific proportion of EVs released by infected RBCs within the EV population. Western blot analysis on purified EVs from both in vivo and in vitro infections revealed several parasite proteins that were targets of the host immune response. In addition, microRNA analysis showed that infected RBC EVs have different microRNA signature from uninfected RBC EVs, indicating a potential role as disease biomarkers. Finally, EVs were internalized by other RBCs in culture, implicating a potential role for these vesicles in cellular communication. Overall, our study points to the multiple functional implications of EVs in Babesia-host interactions and support the potential that EVs have as agents in disease pathogenesis.

Project description:IntroductionBabesia microti (B. microti) is the dominant species responsible for human babesiosis, which is associated with severe hemolytic anemia and splenomegaly because it infects mammalian erythrocytes. The actual prevalence of B. microti is thought to have been substantially underestimated.MethodsIn this study, Bagg's albino/c (BALB/c) mice were intraperitoneally injected with B. microti-infected erythrocytes, and parasitemia was subsequently measured by calculating the proportion of infected erythrocytes. The ultrastructure of infected erythrocytes was observed using scanning and transmission electron microscopes. Quantifying phosphatidylserine (PS) exposure, oxidative stress, intracellular Ca2+, and erythropoiesis of erythrocytes were done using flow cytometry. The physiological indicators were analyzed using a Mindray BC-5000 Vet automatic hematology analyzer.ResultsOf note, 40.7 ± 5.9% of erythrocytes changed their structure and shrunk in the B. microti-infected group. The percentage of annexin V-positive erythrocytes and the levels of reactive oxygen species (ROS) in the erythrocytes were higher in the B. microti-infected group than in the control group at 10 dpi. Significant splenomegaly and severe anemia were also observed following B. microti infection. The parasitemia level in the B. microti-infected splenectomized group was higher than that of the B. microti-infected sham group. The population of early erythroblasts increased, and the late erythroblasts decreased in both the bone marrow and spleen tissues of the B. microti-infected group at 10 dpi.DiscussionPS exposure and elevated ROS activities were hallmarks of eryptosis in the B. microti-infected group. This study revealed for the first time that B. microti could also induce eryptosis. At the higher parasitemia phase, the occurrence of severe anemia and significant changes in the abundance of erythroblasts in B. microti-infected mice group were established. The spleen plays a critical protective role in controlling B. microti infection and preventing anemia. B. microti infection could cause a massive loss of late erythroblasts and induce erythropoiesis.

Dataset Information

Analysis of the Babesia microti proteome in infected red blood cells by a combination of nanotechnology and mass spectrometry.

Publications

Analysis of the Babesia microti proteome in infected red blood cells by a combination of nanotechnology and mass spectrometry.

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