Islam D, Lindberg A A
Laboratory Science Division, International Center for Diarrhoeal Disease Research, Bangladesh, Dhaka.
J Clin Microbiol. 1992 Nov;30(11):2801-6. doi: 10.1128/jcm.30.11.2801-2806.1992.
A combination of immunomagnetic separation (IMS) and a polymerase chain reaction (PCR) procedure was used for direct isolation and identification of Shigella dysenteriae type 1 and Shigella flexneri from feces. Immunomagnetic particles were coated with monoclonal antibody MASFB, which is specific for a common epitope of the O polysaccharides of S. dysenteriae type 1 and S. flexneri. Bacteria bound to the beads were boiled in water, and target DNA was amplified with a primer pair specific for a gene coded on the invasion-associated locus (ial) of the large virulence plasmid of all four Shigella spp. and enteroinvasive strains of Escherichia coli. A 320-bp DNA fragment was generated and detected by an alkaline phosphatase-conjugated probe. Nonviable cells were also captured and detected by this technique. The method is simple and fast (7 h) and has a detection limit of ca. 10 Shigella organisms per g in fecal samples. The combined IMS-PCR assay correctly identified all 57 samples carrying S. dysenteriae type 1 and 68 samples carrying S. flexneri from 238 fecal specimens and also permitted detection of 17 samples carrying Shigella spp. from 113 specimens from diarrheal patients in whom shigellae were not detected by conventional culture.
采用免疫磁珠分离（IMS）与聚合酶链反应（PCR）相结合的方法，直接从粪便中分离并鉴定1型痢疾志贺菌和福氏志贺菌。免疫磁珠包被有单克隆抗体MASFB，该抗体对1型痢疾志贺菌和福氏志贺菌O多糖的共同表位具有特异性。将结合在磁珠上的细菌在水中煮沸，然后用一对引物扩增靶DNA，该引物对所有四种志贺菌属和肠侵袭性大肠杆菌大毒力质粒上侵袭相关位点（ial）编码的基因具有特异性。通过碱性磷酸酶偶联探针产生并检测到一个320 bp的DNA片段。该技术也能捕获并检测无活力的细胞。该方法简单快速（7小时），粪便样本中的检测限约为每克10个志贺菌。联合IMS-PCR检测法从238份粪便标本中正确鉴定出所有57份携带1型痢疾志贺菌的样本和68份携带福氏志贺菌的样本，还能从113份腹泻患者标本中检测出17份携带志贺菌属的样本，而这些标本用传统培养法未检测到志贺菌。
