Instituto de Agrobiotecnología y Biología Molecular (IABIMO) INTA - CONICET, De Los Reseros y Dr. Nicolás Repetto s/n, P.O. Box 25 (B1712WAA), Castelar, Buenos Aires, Argentina.
Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164, USA.
Int J Parasitol. 2021 Jul;51(8):643-658. doi: 10.1016/j.ijpara.2020.12.010. Epub 2021 Mar 20.
Bovine babesiosis is a tick-borne disease caused by apicomplexan parasites of the Babesia genus that represents a major constraint to livestock production worldwide. Currently available vaccines are based on live parasites which have archetypal limitations. Our goal is to identify candidate antigens so that new and effective vaccines against Babesia may be developed. The perforin-like protein (PLP) family has been identified as a key player in cell traversal and egress in related apicomplexans and it was also identified in Babesia, but its function in this parasite remains unknown. The aim of this work was to define the PLP family in Babesia and functionally characterize PLP1, a representative member of the family in Babesia bovis. Bioinformatic analyses demonstrate a variable number of plp genes (four to eight) in the genomes of six different Babesia spp. and conservation of the family members at the secondary and tertiary structure levels. We demonstrate here that Babesia PLPs contain the critical domains present in other apicomplexan PLPs to display the lytic capacity. We then focused on the functional characterization of PLP1 of B. bovis, both in vitro and in vivo. PLP1 is expressed and exposed to the host immune system during infection and has high hemolytic capacity under a wide range of conditions in vitro. A B. bovis plp1 knockout line displayed a decreased growth rate in vitro compared with the wild type strain and a peculiar phenotype consisting of multiple parasites within a single red blood cell, although at low frequency. This phenotype suggests that the lack of PLP1 has a negative impact on the mechanism of egression of the parasite and, therefore, on its capacity to proliferate. It is possible that PLP1 is associated with other proteins in the processes of invasion and egress, which were found to have redundant mechanisms in related apicomplexans. Future work will be focused on unravelling the network of proteins involved in these essential parasite functions.
牛巴贝斯虫病是一种由巴贝斯虫属的顶复门寄生虫引起的蜱传疾病，它是全球畜牧业生产的主要制约因素。目前可用的疫苗基于活寄生虫，但具有典型的局限性。我们的目标是确定候选抗原，以便开发针对巴贝斯虫的新型有效疫苗。穿孔素样蛋白 (PLP) 家族已被确定为相关顶复门生物中细胞穿越和逸出的关键因素，它也在巴贝斯虫中被发现，但它在该寄生虫中的功能仍不清楚。这项工作的目的是确定巴贝斯虫中的 PLP 家族，并对巴贝斯牛的代表性成员 PLP1 进行功能表征。生物信息学分析表明，在六个不同的巴贝斯虫种的基因组中存在可变数量的 plp 基因（4 到 8 个），并且家族成员在二级和三级结构水平上保持保守。我们在这里证明，巴贝斯虫 PLPs 包含其他顶复门 PLPs 中存在的关键结构域，以显示裂解能力。然后，我们专注于 B. bovis 的 PLP1 的功能表征，包括在体外和体内。PLP1 在感染期间表达并暴露于宿主免疫系统中，并且在体外多种条件下具有高溶血能力。与野生型菌株相比，B. bovis plp1 敲除株在体外的生长速度较慢，并且表现出一种独特的表型，即在单个红细胞内有多个寄生虫，但频率较低。这种表型表明 PLP1 的缺乏对寄生虫逸出机制有负面影响，因此对其增殖能力有负面影响。PLP1 可能与入侵和逸出过程中的其他蛋白质相关联，在相关的顶复门生物中，这些蛋白质的机制具有冗余性。未来的工作将集中于揭示参与这些重要寄生虫功能的蛋白质网络。
