Classic Spotlight: Plate Counting You Can Count On.
Classic Spotlight: Plate Counting You Can Count On.
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经典聚焦：值得信赖的平板计数。
DOI：
10.1128/jb.00711-16
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发表时间：
2016
影响因子：
3.2
通讯作者：
O'Toole,GeorgeA
中科院分区：
文献类型：
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作者：
O'Toole,GeorgeA
在美国的每一堂微生物学入门课上（我希望全世界都是如此），教授或助教都会在课程的早期向下一代微生物学家介绍使用CFU来确定培养物中活菌的概念。教师解释了连续稀释的概念，以及找到稀释度的过程，该稀释度在平板上提供了易于计数的菌落数。大多数教科书和实验室手册通常建议使用产生30到300个菌落（或类似范围）的平板。菌落太少，计数可能不准确;菌落太多，区分平板上的单个菌落既困难又耗时。计数活菌的平板法被认为是常识，因此，当被问及为什么我们使用这种方法时，很容易回答，“嗯，这就是它的工作方式。事实证明，计算殖民地数量曾经是一个有点悬而未决的话题，早在1895年（1）就在出版的著作中得到了解决。Breed和Dotterrer试图在1916年发表在《细菌学杂志》（Journal of Bacteriology）上的一篇论文中解决这个问题（2），该杂志出版的第一年。引言的第一句话很快就提到了论文的中心点，“在进行细菌学计数时非常重要的一点是，在不引入严重错误的情况下，允许在平板上生长的菌落数量的限制。”作者接着指出，“可能每个细菌学工作者在计数时都有这一点，并根据经验有自己的看法;但关于这个问题的公开数据很少。”解决这一点是一个严重的问题，因为这些计数用于分析牛奶的细菌污染。此外，当时已知牛奶中各种微生物的殖民地大小变化很大（某些微生物产生针尖菌落），并且某些微生物可以促进邻近菌落的生长。在他们的报告中，Breed和Dotterrer得出结论，“每个平板具有超过40个和少于200个菌落的平板组”提供了最具重现性的结果，变异性最小。相比之下，作者指出，“很明显，少于20个菌落和多于400个菌落的平板很容易产生很大的差异，这种平板的计数应该被忽略。”他们的研究需要1,435个琼脂平板（这些是可重复使用的玻璃培养皿！），形成了我们今天仍在教授的常识的基础。
In every introductory microbiology class in the United States (and, I expect, the world) at some point early in the course the professor or teaching assistant introduces the next generation of microbiologists to the concept of using CFU to determine the viable bacteria in a culture. Instructors explain the concept of serial dilution, as well as the process of finding that dilution which provides a readily countable number of colonies on a plate. Most textbooks and lab manuals typically recommend using plates that yield somewhere between 30 and 300 colonies (or some similar range). Too few colonies, and the count may not be accurate; too many colonies, and it is difficult as well as time-consuming to distinguish the individual colonies on a plate. The plating method for counting viable bacteria is considered general knowledge, and as such, when asked why we use this method it is easy to respond,“Well, this is just the way that it’s done.” It turns out that counting colonies was once a somewhat unsettled topic and was addressed in published works as far back as 1895 (1). Breed and Dotterrer sought to resolve this issue in a paper published in the Journal of Bacteriology in 1916 (2), the first year of the journal’s publication. The first sentence of the introduction comes to the central point of the paper quickly by stating,“A point which is of much importance in making bacteriological counts is the limit in the number of colonies that may be allowed to grow on a plate without introducing serious errors.” The authors then note,“Probably every bacteriological worker has this point in mind in making counts and has his own opinion based on experience; but there are few published data on the subject.” Resolving this point was a serious matter, as these counts were used in analyzing milk for bacterial contamination. Furthermore, it was known at the time that the colony sizes of the various microbes in milk varied a great deal (with some organisms yielding pinpoint colonies) and that some organisms could enhance the growth of neighboring colonies. In their report, Breed and Dotterrer conclude that “the group of plates having more than 40 and less than 200 colonies per plate” provided the most reproducible findings with the least variability. In contrast, the authors state that “it is at once clear that plates having less than 20 and more than 400 colonies are so apt to be widely discrepant that counts from plates of this sort should be disregarded.” Their study, which required 1,435 agar plates (these were the reusable glass petri plates!), forms the foundation of the general knowledge we still teach today.