Perkinsus marinus (Perkinsea) is an osmotrophic facultative intracellular marine protozoan responsible for ‘Dermo’ disease in the eastern oyster, Crassostrea virginica. In 1993, three laboratories developed the in vitro culture of P. marinus in the absence of host cells. Two of the culture media and conditions went through several optimization reiterations, with one becoming the standard for the small-scale growth of Perkinsus species. Compared to most intracellular protozoan parasites, the availability of P. marinus to grow in the absence of host cells has provided the basis to explore its use as a heterologous expression system, which could potentially be used for therapeutic applications, such as vaccines. As the genetic toolbox is becoming available, there is also the need for larger-scale cultivation and lower-cost media formulations. Here, we took an industrial approach to scaled-up growth from a small culture flask to bioreactors which required developing new cultivation parameters, including aeration, mixing, pH, temperature control, and media formulation. Our approach also enabled more real-time data collection on growth. The bioreactor cultivation method showed similar or accelerated growth rates of P. marinus compared to culture in T-flasks. Redox measurements indicated sufficient oxygen availability throughout the cultivation. Replacing fetal bovine serum with chicken serum showed no differences in the growth rate and a 60% reduction in the medium cost. This study opens the door to furthering P. marinus as a valid heterologous expression system by showing the ability to grow in bioreactors.

中文翻译：

---

生物反应器中的 Perkinsus marinus：生长和降低成本的生长培养基

Perkinsus marinus (Perkinsea) 是一种渗透营养型兼性细胞内海洋原生动物，导致东部牡蛎（Crassostrea virginica）的“皮肤病”。1993年，三个实验室在没有宿主细胞的情况下开发了P. marinus的体外培养方法。其中两种培养基和条件经过多次优化反复，其中一种成为帕金斯菌小规模生长的标准。与大多数细胞内原生动物寄生虫相比，P. marinus 在没有宿主细胞的情况下生长的可用性为探索其作为异源表达系统的用途提供了基础，该系统可能用于治疗应用，例如疫苗。随着遗传工具箱的出现，还需要更大规模的培养和更低成本的培养基配方。在这里，我们采用工业方法从小型培养瓶扩大到生物反应器，这需要开发新的培养参数，包括通气、混合、pH、温度控制和培养基配方。我们的方法还能够收集更多关于增长的实时数据。与 T 型烧瓶中的培养相比，生物反应器培养方法显示出相似或加速的 P. marinus 生长速率。氧化还原测量表明整个培养过程中有足够的氧气供应。用鸡血清代替胎牛血清，生长速度没有差异，并且培养基成本降低了 60%。这项研究通过展示 P. marinus 在生物反应器中生长的能力，为进一步将 P. marinus 作为有效的异源表达系统打开了大门。