Recent Advances in Molecular Genetic Tools for Babesia

Abstract

Development of in vitro culture and completion of genome sequencing of several Babesia parasites promoted the efforts to establish transfection systems for these parasites to dissect the gene functions. It has been more than a decade since the establishment of first transfection for Babesia bovis, the causative agent of bovine babesiosis. However, the number of genes that were targeted by genetic tools in Babesia parasites is limited. This is partially due to the low efficiencies of these methods. The recent adaptation of CRISPR/Cas9 for genome editing of Babesia bovis can accelerate the efforts for dissecting this parasite’s genome and extend the knowledge on biological aspects of erythrocytic and tick stages of Babesia. Additionally, glmS ribozyme as a conditional knockdown system is available that could be used for the characterization of essential genes. The development of high throughput genetic tools is needed to dissect the function of multigene families, targeting several genes in a specific pathway, and finally genome-wide identification of essential genes to find novel drug targets. In this review, we summarized the current tools that are available for Babesia and the genes that are being targeted by these tools. This may draw a perspective for the future development of genetic tools and pave the way for the identification of novel drugs or vaccine targets.

Establishment of a stable transfection and gene targeting system in Babesia divergens

“…Babesia -specific transfection protocols typically involve electroporation of infected RBCs (iRCBs) or, less commonly, transfection of free (purified) merozoites ( Hakimi et al., 2021b ). While the alternative approach based on vector DNA electroporation of uninfected RBCs (uRBCs) cells has proven highly effective in P. falciparum ( Deitsch et al., 2001 ; Skinner-Adams et al., 2003 ; Hasenkamp et al., 2012 ), its application in Babesia species is still unverified.…”

Advances in understanding red blood cell modifications by Babesia

“…Exportomes have fundamental roles in apicomplexan parasite survival and virulence, but studies on the Babesia exportome have lagged behind. The recent completion of genome databases for multiple Babesia species and the development of transfection tools (reviewed in [ 85 ]) have allowed us to begin to describe Babesia exportomes. Several proteins have been identified from bioinformatic or proteomics approaches and verified by expression of the tagged candidate proteins.…”

A Transfected Babesia bovis Parasite Line Expressing eGFP Is Able to Complete the Full Life Cycle of the Parasite in Mammalian and Tick Hosts

“…Transfection of Plasmodium sp also targets EF1α and has been used successfully to transmit transfected parasites throughout the parasite's entire life cycle, including development within the mosquito vector [17]. However, the development of sexual stages of Plasmodium begins within the mammalian host, whereas Babesia gametes develop within the lumen of the tick midgut [22,23]. Importantly, transfection into the Plasmodium EF1α locus has been shown to alter the growth cycle characteristics of transfected parasites, extending the duration of the G1 phase [17].…”