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

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60076658%3A12310%2F23%3A43907444" target="_blank" >RIV/60076658:12310/23:43907444 - isvavai.cz</a>

Výsledek na webu

<a href="https://www.frontiersin.org/articles/10.3389/fcimb.2023.1278041/full" target="_blank" >https://www.frontiersin.org/articles/10.3389/fcimb.2023.1278041/full</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3389/fcimb.2023.1278041" target="_blank" >10.3389/fcimb.2023.1278041</a>

Jazyk výsledku

angličtina

Název v původním jazyce

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

Popis výsledku v původním jazyce

Babesia divergens is an emerging tick-borne pathogen considered as the principal causative agent of bovine babesiosis in Europe with a notable zoonotic risk to human health. Despite its increasing impact, considerable gaps persist in our understanding of the molecular interactions between this parasite and its hosts. In this study, we address the current limitation of functional genomic tools in B. divergens and introduce a stable transfection system specific to this parasite. We define the parameters for a drug selection system hdhfr-WR99210 and evaluate different transfection protocols for highly efficient generation of transgenic parasites expressing GFP. We proved that plasmid delivery into bovine erythrocytes prior to their infection is the most optimal transfection approach for B. divergens, providing novel evidence of Babesia parasites&apos; ability to spontaneously uptake external DNA from erythrocytes cytoplasm. Furthermore, we validated the bidirectional and symmetrical activity of ef-tgtp promoter, enabling simultaneous expression of external genes. Lastly, we generated a B. divergens knockout line by targeting a 6-cys-e gene locus. The observed dispensability of this gene in intraerythrocytic parasite development makes it a suitable recipient locus for further transgenic application. The platform for genetic manipulations presented herein serves as the initial step towards developing advanced functional genomic tools enabling the discovery of B. divergens molecules involved in host-vector-pathogen interactions.

Název v anglickém jazyce

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

Popis výsledku anglicky

Babesia divergens is an emerging tick-borne pathogen considered as the principal causative agent of bovine babesiosis in Europe with a notable zoonotic risk to human health. Despite its increasing impact, considerable gaps persist in our understanding of the molecular interactions between this parasite and its hosts. In this study, we address the current limitation of functional genomic tools in B. divergens and introduce a stable transfection system specific to this parasite. We define the parameters for a drug selection system hdhfr-WR99210 and evaluate different transfection protocols for highly efficient generation of transgenic parasites expressing GFP. We proved that plasmid delivery into bovine erythrocytes prior to their infection is the most optimal transfection approach for B. divergens, providing novel evidence of Babesia parasites&apos; ability to spontaneously uptake external DNA from erythrocytes cytoplasm. Furthermore, we validated the bidirectional and symmetrical activity of ef-tgtp promoter, enabling simultaneous expression of external genes. Lastly, we generated a B. divergens knockout line by targeting a 6-cys-e gene locus. The observed dispensability of this gene in intraerythrocytic parasite development makes it a suitable recipient locus for further transgenic application. The platform for genetic manipulations presented herein serves as the initial step towards developing advanced functional genomic tools enabling the discovery of B. divergens molecules involved in host-vector-pathogen interactions.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

<a href="/cs/project/GA21-11299S" target="_blank" >GA21-11299S: Funkční analýza kalcium-dependentních proteinkináz babesií</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2023

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Frontiers in Cellular and Infection Microbiology

ISSN

2235-2988

e-ISSN

2235-2988

Svazek periodika

13

Číslo periodika v rámci svazku

DEC 13 2023

Stát vydavatele periodika

CH - Švýcarská konfederace

Počet stran výsledku

12

Strana od-do

—

Kód UT WoS článku

001136267100001

EID výsledku v databázi Scopus

2-s2.0-85180870111