Calderaro Adriana, Villanacci Vincenzo, Conter Mauro, Ragni Patrizia, Piccolo Giovanna, Zuelli Claudia, Bommezzadri Simona, Guégan Rozenn, Zambelli Claudia, Perandin Francesca, Arcangeletti Maria Cristina, Medici Maria Cristina, Manca Nino, Dettori Giuseppe, Chezzi Carlo
Department of Pathology and Laboratory Medicine, Section of Microbiology, University of Parma, Viale A. Gramsci 14, 43100 Parma, Italy.
Res Microbiol. 2003 Mar;154(2):145-53. doi: 10.1016/S0923-2508(02)00014-1.
This study reports for the first time the detection of Brachyspira aalborgi in faeces and rectal biopsies of a female suffering for 3-4 months of abdominal pain with long-standing mucosal diarrhoea, rectal bleeding and suspected carcinoma of the rectum. After pre-treatment of samples (faeces and biopsies) with a liquid medium (trypticase soy broth-TSB) containing foetal calf serum (FCS, 10%) and spectinomycin and rifampicin (TSB-SR) the first detection of B. aalborgi isolate HBS1 was observed after 48 h in the primary plates of selective blood agar modified medium (BAM) containing spectinomycin and rifampicin (BAM-SR), where growth zones were signalled by a small weakly beta-haemolytic halo. Attempts to subculture spirochaetes in agar media failed. The new HBS1 isolate was only propagated in TSB broth and at electron microscopy it showed 4 endoflagella inserted at each tapered end. The phenotypic characterization of HBS1 demonstrated absence of hippurate hydrolysis, indole production, alpha-galactosidase, alpha- and beta-glucosidase activities in accordance with the B. aalborgi type strain. Rapid identification of B. aalborgi isolate HBS1 was performed directly from faeces and rectal biopsies and subsequently from pure cultures by a genetic method based on 16S DNA restriction fragment length polymorphism (RFLP)-polymerase chain reaction (PCR). The sequence of 16S DNA amplicon of the isolate HBS1 was found 99.2% corresponding to that of the B. aalborgi type strain. Our results encourage further investigations for the development of a suitable selective agar medium for the isolating and cultivating B. aalborgi from human specimens.
本研究首次报告了在一名患有3 - 4个月腹痛、长期黏膜腹泻、直肠出血且疑似直肠癌的女性的粪便和直肠活检组织中检测到奥尔堡短螺旋体。在用含有胎牛血清（FCS，10%）、壮观霉素和利福平的液体培养基（胰蛋白酶大豆肉汤 - TSB）对样本（粪便和活检组织）进行预处理后，在含有壮观霉素和利福平的选择性血琼脂改良培养基（BAM）的原代平板中，48小时后首次观察到奥尔堡短螺旋体菌株HBS1的生长，在生长区域可见一个小的弱β - 溶血晕圈。在琼脂培养基中继代培养螺旋体的尝试失败。新的HBS1菌株仅在TSB肉汤中繁殖，电子显微镜观察显示其在每个锥形末端插入有4根内鞭毛。HBS1的表型特征表明其缺乏马尿酸盐水解、吲哚产生、α - 半乳糖苷酶、α - 和β - 葡萄糖苷酶活性，与奥尔堡短螺旋体模式菌株一致。通过基于16S DNA限制性片段长度多态性（RFLP） - 聚合酶链反应（PCR）的基因方法，直接从粪便和直肠活检组织以及随后从纯培养物中快速鉴定出奥尔堡短螺旋体菌株HBS1。发现分离株HBS1的16S DNA扩增子序列与奥尔堡短螺旋体模式菌株的序列有99.2%的对应性。我们的结果鼓励进一步开展研究，以开发一种适合从人类标本中分离和培养奥尔堡短螺旋体的选择性琼脂培养基。
