Stier Philipp, Kulozik Ulrich
TUM School of Life Sciences, Technical University of Munich, 85354 Freising, Germany.
Methods Protoc. 2021 Sep 10;4(3):63. doi: 10.3390/mps4030063.
In the food and pharmaceutical industries, evaluating the sterilization performance preceding aseptic production processes is of central importance. In the case of hydrogen peroxide sterilization of solid surfaces, bioindicators (BI) consisting of spores of or are used to validate the effectiveness and efficiency of the inactivation procedure. Commercial production of is commonly performed on agar plates, where cultivation and sporulation conditions are not well-defined. Therefore, the produced BI can vary in their resistance, which in turn creates unacceptable uncertainties in the evaluation of aseptic processes. Submerged production in the bioreactor would allow more control over sporulation conditions, while reducing production time, resistance variability, and avoidance of false-positive or false-negative test results. In addition, submerged production of so far was a challenge to achieve sufficiently high spore concentrations for BI production. This study reports on the development of a method for submerged production of spores (pH 7.0, 57 °C, 30% pO) that can achieve 1.6 × 10 spores/mL with a resistance against 35% HO at 25 °C of D = 73 s. This resistance ranks within the range of commercially available BI, making the results directly transferable to industrial applications.
在食品和制药行业，评估无菌生产过程之前的灭菌性能至关重要。在固体表面过氧化氢灭菌的情况下，由嗜热脂肪芽孢杆菌或枯草芽孢杆菌的孢子组成的生物指示剂（BI）用于验证灭活程序的有效性和效率。嗜热脂肪芽孢杆菌的商业生产通常在琼脂平板上进行，其培养和孢子形成条件并不明确。因此，所生产的生物指示剂的抗性可能会有所不同，这反过来又会在无菌工艺评估中产生不可接受的不确定性。在生物反应器中进行深层培养可以更好地控制孢子形成条件，同时缩短生产时间、降低抗性变异性，并避免假阳性或假阴性测试结果。此外，到目前为止，嗜热脂肪芽孢杆菌的深层培养要达到足够高的孢子浓度以用于生物指示剂生产是一项挑战。本研究报告了一种用于深层培养嗜热脂肪芽孢杆菌孢子（pH 7.0，57°C，30% pO₂）的方法的开发，该方法可实现1.6×10⁹个孢子/mL的产量，其在25°C下对35%过氧化氢的抗性为D = 73秒。这种抗性处于市售生物指示剂的范围内，使得研究结果可直接应用于工业生产。
