苜蓿耐药根瘤菌筛选及荧光蛋白标记对根瘤菌耐药性的影响

为明确荧光蛋白标记技术对根瘤菌耐药性的影响,采集6个品种的苜蓿根瘤进行根瘤菌的分离、纯化和初步鉴定,并筛选其中对3种稀土盐抑菌剂La（NO3）3·6H2O,LaCl3和Ce（NO3）3·6H2O以及2种植物源抑菌剂苦参碱和除虫菊素具有耐受性的菌株,得到一株编号为LH3436的根瘤菌,对0.6mg/mL苦参碱具有天然耐受性。将LH3436和对照根瘤菌R.12531以及参比根瘤菌R.GN5进行荧光蛋白标记后,观察其对抑菌剂耐受性的变化。发现各初筛菌株在无氮培养基上的生长状况对非根瘤菌的筛除率最高,所有初筛菌株均为革兰氏染色阴性的快生型产酸菌,全部难以利用柠檬酸盐,而可以利用蔗糖和可溶性淀粉,但对乳糖、葡萄糖和果糖等碳源的利用存在差异性;试验初筛菌株对植物源类抑菌剂的相对耐受性好,而对稀土盐类抑菌剂的相对耐受性差,将筛选根瘤菌进行荧光蛋白标记后,其对抑菌剂的耐受性未发生改变。

Screening of antimicrobial resistant alfalfa rhizobia and effect of fluorescent protein marker on rhizobia resistance

In order to clarify the effect of fluorescent protein marker technology on rhizobia resistance to an timicrobials,the nodules of 6 alfalfa varieties were collected for rhizobia isolation and identification. The prelimi nari and ly C isolated rhizobia were tested in terms of their resistance to 3 rare earth salts （La（NO3）3 · 6H2 O, LaCla e（NO3） 3 · 6H2O） and 2 botanical antimicrobials （matrine and pyrethrin）. The obtained rhizobium LH3436 was resistant to 0. 6 mg/mL matrine. LH3436 and the control rhizobia R. 12531 and R. GN5 were marked with fluorescence protein and their resistance variance to antimicrobials was tested. The results indicated that the growth status of the preliminari ing rate,all the preliminarily screened r ly hi screened rhizobia on nitrogen free medium showed the highest screen zobia were gram negative and fast growing acid forming bacteria, all were incapable of using citrate,but could use sucrose and soluble starch, differences existed among the rhizobia in the utilization of lactose,glucose and fructose. All the screened rhizobia showed relatively better tolerance to botanical antimicrobials, but poorer tolerance to rare earth salts. Fluorescence protein marker technology did not show influence on rhizobia resistance to antimicrobials.