{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T02:43:59Z","timestamp":1760237039082,"version":"build-2065373602"},"reference-count":30,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2020,2,10]],"date-time":"2020-02-10T00:00:00Z","timestamp":1581292800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["18F18403"],"award-info":[{"award-number":["18F18403"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Grants-in-Aid for Scientific Research","award":["18H02336"],"award-info":[{"award-number":["18H02336"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pathogens"],"abstract":"<jats:p>The development of genetic manipulation techniques has been reported in many protozoan parasites over the past few years. However, these techniques have not been established for Babesia microti. Here, we report the first successful transient transfection of B. microti. The plasmids containing the firefly luciferase reporter gene were transfected into B. microti by an AMAXA 4D Nucleofection system. Twenty-four-hour synchronization, the 5\u2032-actin promoter, program FA100, and 50 \u03bcg of plasmid DNA constituted the best conditions for the transient transfection of B. microti. This finding is the first step towards a stable transfection method for B. microti, which may contribute to a better understanding of the biology of the parasite.<\/jats:p>","DOI":"10.3390\/pathogens9020108","type":"journal-article","created":{"date-parts":[[2020,2,11]],"date-time":"2020-02-11T09:25:21Z","timestamp":1581413121000},"page":"108","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["Transient Transfection of the Zoonotic Parasite Babesia microti"],"prefix":"10.3390","volume":"9","author":[{"given":"Mingming","family":"Liu","sequence":"first","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shengwei","family":"Ji","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5250-6172","authenticated-orcid":false,"given":"Mohamed Abdo","family":"Rizk","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"},{"name":"Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Mansoura University, Mansoura 35516, Egypt"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Paul Franck","family":"Adjou Moumouni","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2411-8401","authenticated-orcid":false,"given":"Eloiza May","family":"Galon","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jixu","family":"Li","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yongchang","family":"Li","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Weiqing","family":"Zheng","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"},{"name":"Nanchang Center for Disease Control and Prevention, Nanchang 330038, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Byamukama","family":"Benedicto","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Maria Agnes","family":"Tumwebaze","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Masahito","family":"Asada","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xuenan","family":"Xuan","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2020,2,10]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Parveen, N., and Bhanot, P. (2019). Babesia microti-Borrelia Burgdorferi Coinfection. Pathogens, 8.","DOI":"10.3390\/pathogens8030117"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Bloch, E.M., Kumar, S., and Krause, P.J. (2019). Persistence of Babesia microti Infection in Humans. Pathogens, 8.","DOI":"10.3390\/pathogens8030102"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"357","DOI":"10.1016\/j.idc.2015.02.008","article-title":"Babesiosis","volume":"29","author":"Vannier","year":"2015","journal-title":"Infect. Dis. Clin. North Am."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"35284","DOI":"10.1038\/srep35284","article-title":"Genome-wide diversity and gene expression profiling of Babesia microti isolates identify polymorphic genes that mediate host-pathogen interactions","volume":"6","author":"Silva","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"e00109-19","DOI":"10.1128\/mSphere.00109-19","article-title":"Genome Editing of Babesia bovis Using the CRISPR\/Cas9 System","volume":"4","author":"Hakimi","year":"2019","journal-title":"mSphere"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1186\/s13071-016-1439-z","article-title":"Establishment of transient and stable transfection systems for Babesia ovata","volume":"9","author":"Hakimi","year":"2016","journal-title":"Parasites Vectors"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"6096","DOI":"10.1038\/s41598-018-23010-4","article-title":"Stable transformation of Babesia bigemina and Babesia bovis using a single transfection plasmid","volume":"8","author":"Silva","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"56","DOI":"10.1016\/j.molbiopara.2017.07.003","article-title":"Transient transfection of intraerythrocytic Babesia gibsoni using elongation factor-1 alpha promoter","volume":"216","author":"Liu","year":"2017","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"101279","DOI":"10.1016\/j.ttbdis.2019.101279","article-title":"Transient transfection of Babesia ovis using heterologous promoters","volume":"10","author":"Rosa","year":"2019","journal-title":"Ticks Tick-borne Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"330","DOI":"10.1016\/j.ttbdis.2017.11.008","article-title":"Identification and characterization of interchangeable cross-species functional promoters between Babesia gibsoni and Babesia bovis","volume":"9","author":"Liu","year":"2018","journal-title":"Ticks Tick-borne Dis."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1016\/j.vetpar.2011.05.032","article-title":"Emerging perspectives in the research of bovine babesiosis and anaplasmosis","volume":"180","author":"Suarez","year":"2011","journal-title":"Veter- Parasitol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"722","DOI":"10.1016\/j.ttbdis.2019.01.006","article-title":"Transgenic Babesia bovis lacking 6-Cys sexual-stage genes as the foundation for non-transmissible live vaccines against bovine babesiosis","volume":"10","author":"Alzan","year":"2019","journal-title":"Ticks Tick-borne Dis."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1016\/j.ijpara.2018.05.015","article-title":"Up-regulated expression of spherical body protein 2 truncated copy 11 in Babesia bovis is associated with reduced cytoadhesion to vascular endothelial cells","volume":"49","author":"Lau","year":"2019","journal-title":"Int. J. Parasitol."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Hussein, H.E., Bastos, R.G., Schneider, D.A., Johnson, W.C., Adham, F.K., Davis, W.C., Laughery, J.M., Herndon, D.R., Alzan, H.F., and Ueti, M.W. (2017). The Babesia bovis hap2 gene is not required for blood stage replication, but expressed upon in vitro sexual stage induction. PLOS Neglected Trop. Dis., 11.","DOI":"10.1371\/journal.pntd.0005965"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"260","DOI":"10.1186\/s13071-018-2853-1","article-title":"Establishment of a stable transfection system for genetic manipulation of Babesia gibsoni","volume":"11","author":"Liu","year":"2018","journal-title":"Parasites Vectors"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"2236","DOI":"10.1056\/NEJMoa1600897","article-title":"Screening for Babesia microti in the U.S. blood supply","volume":"375","author":"Moritz","year":"2016","journal-title":"N. Engl. J. Med."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Lim, P.L., Chavatte, J.M., Vasoo, S., and Yang, J. (2020). Imported Human Babesiosis, Singapore, 2018. Emerg. Infect. Dis., 26.","DOI":"10.3201\/eid2604.200025"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"6624","DOI":"10.1128\/AEM.02373-16","article-title":"Ixodes persulcatus Ticks as Vectors for the Babesia microti U.S. Lineage in Japan","volume":"82","author":"Morikawa","year":"2016","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1445","DOI":"10.3201\/eid2208.150195","article-title":"Human Babesiosis, Bolivia, 2013","volume":"22","author":"Gabrielli","year":"2016","journal-title":"Emerg. Infect. Dis."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"9102","DOI":"10.1093\/nar\/gks700","article-title":"Sequencing of the smallest Apicomplexan genome from the human pathogen Babesia microti","volume":"40","author":"Cornillot","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"248","DOI":"10.3389\/fcimb.2018.00248","article-title":"The Complexity of Piroplasms Life Cycles","volume":"8","author":"Jalovecka","year":"2018","journal-title":"Front. Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Schreeg, M.E., Marr, H.S., Tarigo, J.L., Cohn, L.A., Bird, D.M., Scholl, E.H., Levy, M.G., Wiegmann, B.M., and Birkenheuer, A.J. (2016). Mitochondrial genome sequences and structures aid in the resolution of Piroplasmida phylogeny. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0165702"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"477","DOI":"10.1038\/nature14651","article-title":"Genetic modification of the diarrhoeal pathogen Cryptosporidium parvum","volume":"523","author":"Vinayak","year":"2015","journal-title":"Nature"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"60","DOI":"10.1016\/j.molbiopara.2005.09.007","article-title":"High efficiency transfection of Plasmodium berghei facilitates novel selection procedures","volume":"145","author":"Janse","year":"2006","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"346","DOI":"10.1038\/nprot.2006.53","article-title":"High-efficiency transfection and drug selection of genetically transformed blood stages of the rodent malaria parasite Plasmodium berghei","volume":"1","author":"Janse","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"955","DOI":"10.1292\/jvms.57.955","article-title":"A Short Term in Vitro Cultivation of Babesia rodhaini and Babesia microti","volume":"57","author":"Shikano","year":"1995","journal-title":"J. Veter- Med. Sci."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"19974","DOI":"10.1074\/jbc.AC118.005771","article-title":"Establishment of a continuous in vitro culture of Babesia duncani in human erythrocytes reveals unusually high tolerance to recommended therapies","volume":"293","author":"Abraham","year":"2018","journal-title":"J. Boil. Chem."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Asada, M., Yahata, K., Hakimi, H., Yokoyama, N., Igarashi, I., Kaneko, O., Suarez, C.E., and Kawazu, S.-I. (2015). Transfection of Babesia bovis by Double Selection with WR99210 and Blasticidin-S and Its Application for Functional Analysis of Thioredoxin Peroxidase-1. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0125993"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1004","DOI":"10.3855\/jidc.5500","article-title":"Inhibitory effects of 19 antiprotozoal drugs and antibiotics on Babesia microti infection in BALB\/c mice","volume":"9","author":"Yao","year":"2015","journal-title":"J. Infect. Dev. Ctries."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"4760","DOI":"10.1038\/srep04760","article-title":"A rapid and robust selection procedure for generating drug-selectable marker-free recombinant malaria parasites","volume":"4","author":"Manzoni","year":"2014","journal-title":"Sci. Rep."}],"container-title":["Pathogens"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-0817\/9\/2\/108\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T08:56:26Z","timestamp":1760172986000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-0817\/9\/2\/108"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,2,10]]},"references-count":30,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2020,2]]}},"alternative-id":["pathogens9020108"],"URL":"https:\/\/doi.org\/10.3390\/pathogens9020108","relation":{},"ISSN":["2076-0817"],"issn-type":[{"type":"electronic","value":"2076-0817"}],"subject":[],"published":{"date-parts":[[2020,2,10]]}}}