异烟肼对脓肿分枝杆菌L型的诱导作用

Induction of L-forms of Mycobacterium Abscessi with isoniazid

目的探讨异烟肼对脓肿分枝杆菌L型的诱导作用。方法将脓肿分枝杆菌分别接种于含128μg/ml异烟肼和无异烟肼的结核分枝杆菌快速液体培养基中培养。含异烟肼的培养物经0.45μm孔径滤膜过滤,滤液接种于无异烟肼的结核分枝杆菌快速液体培养基内返祖培养。将含异烟肼和无异烟肼的培养物及返祖菌进行细胞壁染色和透射电镜观察其细胞壁完整性,扫描电镜观察其表面微观结构。含异烟肼的培养物转种L型菌琼脂平板培养,无异烟肼的培养物和返祖菌转种营养琼脂平板培养,观察其菌落形态。对返祖菌及诱导前接种菌的16S rDNA进行鉴定。结果在含128μg/ml异烟肼浓度的结核分枝杆菌快速液体培养基中培养的脓肿分枝杆菌细胞壁缺失,形态为球形,L型细菌琼脂平板上菌落呈典型油煎蛋样,而无异烟肼的结核分枝杆菌快速液体培养基中培养的脓肿分枝杆菌细胞壁完整,形态为杆状,营养琼脂平板上呈圆形菌落。128μg/ml异烟肼浓度的结核分枝杆菌快速液体培养基中培养的脓肿分枝杆菌经返祖后,返祖菌细胞壁完整,形态为杆状,营养琼脂平板上菌落也呈圆形。16S rDNA鉴定返祖菌与诱导前接种菌的同源性达100%,为同一种细菌,即脓肿分枝杆菌。结论异烟肼成功诱导脓肿分枝杆菌L型,可用于脓肿分枝杆菌耐药机制和防治研究。

Objective To explore the induction of L-forms of Mycobacterium abscessi using isoniazid. Methods Mycobacterium abscessi were cultured in aqueous culture media in the presence or absence of 128 μg/ml isoniazid. The culture media containing isoniazid were filtered with 0.45 μm membrane, and the filtrate was subcultured in nutrient agar media for reversion. The isoniazid-free and isoniazid-containing media and the reversion bacteria were observed for cell wall integrity by cell wall staining and transmission electron microscopy, and the microstructures of the cell surfaces were observed by scanning electron microscopy. The isoniazid- containing culture was subcultured in L- form agar media, and the isoniazid- free culture and the reversed bacteria in nutrient agar media to observe the colony morphology. The reversed and non- induced bacteria were identified for 16S rDNA. Results The bacteria induced with 128 μg/ml isoniazid showed cell wall defect, presenting with a spherical cell morphology and typical fried egg-like colonies in L-form agar media, while in isoniazid-free cultures, the cells showed intact cell walls with rod-like shapes and round colony morphologies on nutrient agar. The reversed bacteria, showing also intact cell walls with rod-like shapes and round colonies on nutrient agar, had identical 16S rDNA with the non-induced bacteria. Conclusion Isoniazid can induce the L-forms of mycobacterium abscessi for use in studies of multidrug resistance and treatment of the bacteria.