Lillehaug D
Department of Biotechnological Sciences, Agricultural University of Norway, As, Norway.
J Appl Microbiol. 1997 Jul;83(1):85-90. doi: 10.1046/j.1365-2672.1997.00193.x.
This report summarizes the results from an effort to optimize the double-agar plate assay for visualization of the plaques made by six temperate bacteriophages induced from industrial strains of Lactococcus lactis. Among the several parameters found to influence the plaque assay, the effect of incorporating glycine into the growth medium was most striking, resulting in extensive increase in the plaque size of all of the 13 phage-host pairs tested. Notable effects on the plaque size of other factors such as the procedure for sterilization of the agar medium, the volume and softness of the top and bottom layers, and the number and growth stage of the bacterial cells added to the lawn, were also observed. By exploiting these findings in an optimized procedure for plaque assaying, several indicator strains were identified which were unable to support the development of plaques on standard double-agar plates. Since bacterial hosts usually are identified by their ability to support the development of plaques, this observation suggests that the severe difficulty experienced in identifying lactococcal starter strains that are sensitive towards a temperate phage, partly is a problem of methodology.
本报告总结了优化双层琼脂平板测定法的相关结果，该方法用于观察从乳酸乳球菌工业菌株诱导出的六种温和噬菌体形成的噬菌斑。在发现的几个影响噬菌斑测定的参数中，向生长培养基中添加甘氨酸的效果最为显著，使得所测试的13对噬菌体 - 宿主组合的噬菌斑大小均大幅增加。还观察到其他因素对噬菌斑大小有显著影响，如琼脂培养基的灭菌程序、上层和下层的体积与软硬度，以及添加到菌苔中的细菌细胞数量和生长阶段。通过在优化的噬菌斑测定程序中利用这些发现，鉴定出了几种指示菌株，它们在标准双层琼脂平板上无法形成噬菌斑。由于细菌宿主通常是通过其支持噬菌斑形成的能力来鉴定的，这一观察结果表明，在鉴定对温和噬菌体敏感的乳酸乳球菌发酵剂菌株时遇到的严重困难，部分是方法学问题。
