Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
Department of Veterinary Tropical Diseases, Faculty of Veterinary Science, University of Pretoria, Onderstepoort, South Africa.
Transbound Emerg Dis. 2020 Mar;67 Suppl 1:35-39. doi: 10.1111/tbed.13348.
Theileriosis is a tick-borne disease caused by intracellular protozoa of the genus Theileria. The most important species in cattle are Theileria annulata and Theileria parva. Both species transform leucocyte host cells, resulting in their uncontrolled proliferation and immortalization. Vaccination with attenuated T. annulata-infected cell lines is currently the only practical means of inducing immunity in cattle. Culture media for Theileria spp. typically contain 10%-20% foetal bovine serum (FBS). The use of FBS is associated with several disadvantages, such as batch-to-batch variation, safety and ethical concerns. In this study, the suitability of serum-free media for the cultivation of Theileria-transformed cell lines was examined. Three commercial serum-free media (HL-1, ISF-1 and Hybridomed DIF 1000) were evaluated for their ability to support growth of the T. annulata A288 cell line. The generation doubling times were recorded for each medium and compared with those obtained with conventional FBS-containing RPMI-1640 medium. ISF-1 gave the shortest generation doubling time, averaging 35.4 ± 2.8 hr, significantly shorter than the 52.2 ± 14.9 hr recorded for the conventional medium (p = .0011). ISF-1 was subsequently tested with additional T. annulata strains. The doubling time of a Moroccan strain was significantly increased (65.4 ± 15.9 hr) compared with the control (47.7 ± 7.5 hr, p = .0004), whereas an Egyptian strain grew significantly faster in ISF-1 medium (43.4 ± 6.5 hr vs. 89.3 ± 24.8 hr, p = .0001). The latter strain also showed an improved generation doubling time of 73.7 ± 21.9 hr in an animal origin-free, serum-free, protein-free medium (PFHM II) compared with the control. Out of four South African T. parva strains and a Theileria strain isolated from roan antelope (Hippotragus equinus), only one T. parva strain could be propagated in ISF-1 medium. The use of serum-free medium may thus be suitable for some Theileria cell cultures and needs to be evaluated on a case-by-case basis. The relevance of Theileria cultivation in serum-free media for applications such as vaccine development requires further examination.
泰勒虫病是一种由泰勒虫属的细胞内原生动物引起的蜱传疾病。在牛中最重要的物种是环形泰勒虫和边缘泰勒虫。这两个物种都会转化白细胞宿主细胞，导致其不受控制的增殖和永生化。使用减毒的感染细胞系进行疫苗接种是目前在牛中诱导免疫的唯一实用手段。用于泰勒虫属的培养基通常含有 10%-20%胎牛血清（FBS）。FBS 的使用存在一些缺点，例如批次间差异、安全性和伦理问题。在这项研究中，研究了无血清培养基用于培养泰勒虫转化细胞系的适宜性。评估了三种商业无血清培养基（HL-1、ISF-1 和 Hybridomed DIF 1000）支持环形泰勒虫 A288 细胞系生长的能力。记录了每种培养基的倍增时间，并与常规含 FBS 的 RPMI-1640 培养基获得的倍增时间进行比较。ISF-1 给出的倍增时间最短，平均为 35.4 ± 2.8 小时，明显短于常规培养基记录的 52.2 ± 14.9 小时（p =.0011）。随后，ISF-1 与其他环形泰勒虫株进行了测试。与对照相比，摩洛哥株的倍增时间显著增加（65.4 ± 15.9 小时）（p =.0004），而埃及株在 ISF-1 培养基中生长明显加快（43.4 ± 6.5 小时与 89.3 ± 24.8 小时，p =.0001）。后者株在无动物来源、无血清、无蛋白的培养基（PFHM II）中的倍增时间也有所改善，为 73.7 ± 21.9 小时，而对照为 101.2 ± 20.5 小时。在来自南非的 4 株边缘泰勒虫株和一株来自旋角羚羊（Hippotragus equinus）的泰勒虫株中，只有一株边缘泰勒虫株能够在 ISF-1 培养基中繁殖。因此，无血清培养基的使用可能适用于一些泰勒虫细胞培养，需要根据具体情况进行评估。无血清培养基中泰勒虫的培养对于疫苗开发等应用的相关性需要进一步研究。
