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METHODS article
Front. Microbiol.
Sec. Microbiotechnology
Volume 15 - 2024 |
doi: 10.3389/fmicb.2024.1450913
Monitoring of Bacillus spore-forming dynamics through flow cytometry
Provisionally accepted
Zhili Chen
1*
Yuanyuan Lu
2*
Jiazhen Cui
1*
Yuzhong Feng
1*
Haolong Dong
1*
Xuan Huang
2*
Chen Zhu
1*
Xianghua Xiong
1*
Hui Chen
1
Qingyang Wang
1*
Gang Liu
1*- 1 Academy of Military Medical Sciences (AMMS), Beijing, China
- 2 Institutes of Physical Science and Information Technology, Anhui University, Hefei, Anhui Province, China
The plate counting method is a traditional and widely accepted technique for live cell counting, often employed for Bacillus enumeration and spore forming rate calculations. However, this method requires at least 12 hours to generate results, making it unsuitable for real-time monitoring of bacterial growth status and spore transformation rate. Bacillus thuringiensis crystals, produced during sporulation, are widely used as microbial pesticides, with high demand for industrial scale production. Variations in cultivation conditions and harvest timing during large-scale pore production of Bacillus thuringiensis significantly affect spore forming rate, impacting crystallization yield. Nevertheless, there is a lack of real-time monitoring methods for spore conversion rate. Flow cytometry (FCM), a well-established technique for single-cell analysis in eukaryotic cells, has been successfully applied in bacterial detection in environmental and food samples. In this study, we introduced a rapid flow cytometry-based method for determining spore forming rate of Bacillus thuringiensis, with two nucleic acid dyes, SYTO24 and LDS751. The method enables dynamic monitoring of spore, vegetative cell, and viable but non-culturable/dead cell proportions during the whole cultivation process, and spore forming rate could be gained within 30 minutes. Data of spore forming rate by FCM method is consistent with that by plate counting method, offering a faster and more efficient approach for assessing sporulation status in industrial Bacillus thuringiensis microbial pesticide production.
Keywords: Flow Cytometry, Bacillus thuringiensis, Spore forming rate, Plate counting, Microbial pesticide
Received: 18 Jun 2024; Accepted: 08 Oct 2024.
Copyright: © 2024 Chen, Lu, Cui, Feng, Dong, Huang, Zhu, Xiong, Chen, Wang and Liu. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence:
Zhili Chen, Academy of Military Medical Sciences (AMMS), Beijing, China
Yuanyuan Lu, Institutes of Physical Science and Information Technology, Anhui University, Hefei, 230601, Anhui Province, China
Jiazhen Cui, Academy of Military Medical Sciences (AMMS), Beijing, China
Yuzhong Feng, Academy of Military Medical Sciences (AMMS), Beijing, China
Haolong Dong, Academy of Military Medical Sciences (AMMS), Beijing, China
Xuan Huang, Institutes of Physical Science and Information Technology, Anhui University, Hefei, 230601, Anhui Province, China
Chen Zhu, Academy of Military Medical Sciences (AMMS), Beijing, China
Xianghua Xiong, Academy of Military Medical Sciences (AMMS), Beijing, China
Qingyang Wang, Academy of Military Medical Sciences (AMMS), Beijing, China
Gang Liu, Academy of Military Medical Sciences (AMMS), Beijing, China
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