Yoshimatsu Shoji, Kato-Matsumaru Tomoko, Aono Akio, Chikamatsu Kinuyo, Yamada Hiroyuki, Mitarai Satoshi
Department of Epidemiology and Clinical Research, Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association, Japan.
Department of Mycobacterium Reference and Research, Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association, Japan; Department of Basic Mycobacteriosis, Nagasaki University Graduate School of Biomedical Sciences, Japan.
Int J Mycobacteriol. 2015 Sep;4(3):245-9. doi: 10.1016/j.ijmyco.2015.05.014. Epub 2015 Jun 25.
A concentration of specimen is recommended for the effective recovery of Mycobacterium tuberculosis (MTB), but the bacteriological efficiency is not well evaluated. The present study evaluated the factors contributing to concentration efficiency of centrifugation and bead-based technique (TB-Beads; Microsens, UK) to recover MTB by using simple in vitro specimens.
Four specimens were prepared (6.5×10(3); 8.1×10(4); 7.9×10(5); and 6.4×10(6)cfu/mL) of different concentrations with or without 5×10(4) of THP-1 cells (RIKEN BRC, Japan). Specimens were subjected to centrifugation at 2000, 3000, and 4000g for 15min, and to TB-Beads. The concentration and recovery rate were calculated to evaluate the efficiency of each method.
The specimens containing a higher number of bacteria and THP-1 cells had a tendency to yield a higher concentration and recovery rate (p=0.001-0.083). MTB was recovered more efficiently with THP-1 cells from the 6.5×10(3)cfu/mL specimen by centrifugation (p⩽0.001) than without them; 24.7-54.4% of MTB were recovered with THP-1 cells by centrifugation at 3000g for 15min, while the recovery using TB-Beads was a maximum of 12.7%.
The efficiency of centrifugation depends on the bacterial density and the co-existence of THP-1 cells. The efficiency of TB-Beads was not as high as centrifugation.
为有效回收结核分枝杆菌（MTB），推荐对标本进行浓缩，但细菌学效率尚未得到充分评估。本研究使用简单的体外标本，评估了影响离心法和基于磁珠的技术（TB磁珠；英国Microsens公司）回收MTB浓缩效率的因素。
制备了四种不同浓度（6.5×10³；8.1×10⁴；7.9×10⁵；和6.4×10⁶cfu/mL）的标本，分别添加或不添加5×10⁴个THP-1细胞（日本理化学研究所生物资源中心）。标本分别以2000、3000和4000g离心15分钟，并使用TB磁珠处理。计算浓缩率和回收率以评估每种方法的效率。
含有较多细菌和THP-1细胞的标本往往具有较高的浓缩率和回收率（p=0.001-0.083）。通过离心从6.5×10³cfu/mL的标本中回收MTB时，添加THP-1细胞比不添加时更有效（p≤0.001）；在3000g离心15分钟的情况下，添加THP-1细胞可回收24.7-54.4%的MTB，而使用TB磁珠的最大回收率为12.7%。
离心效率取决于细菌密度和THP-1细胞的共存情况。TB磁珠的效率不如离心法高。
