热处理和抗生素对桉树固有内生细菌去除的研究

为获得无内生细菌干扰的尾叶桉组培苗,本试验采用尾叶桉种子干热和湿热处理、组培苗热处理结合茎尖培养及组培苗抗生素处理3种方法去除内生细菌,并用透明染色法和血球计数法对其体内内生细菌的分布及数量的变化开展了研究.结果表明:经种子热处理桉树种苗根部内生细菌含量略高于对照,而茎、叶部略低于对照组；组培苗恒温37℃热处理第20、...

Elimination of indigenous endophytic bacteria in Eucalyptus urophylla by heat and antibiotics treatment

In order to obtain endophytic bacteria free seedlings of Eucalyptus urophylla,three methods were applied to eliminate or inhibit the indigenous endophytic bacteria in E.urophylla,i.e.,heat treatment of the seeds of E.urophylla at stable and variated high temperatures,heat treatment of seedlings from tissue culture of E.urophylla combined with apical meristem culture,and antibiotics treatment of the seedlings grown in tissue culture.A differential staining procedure was used to detect the existence of endophytic bacteria and hemacytometer counting method was used to study the bacterial population.The results showed that there was no obvious effect in the elimination of endophytic bacteria by heat treatment of the seeds,and the bacteria in root of treated seedlings had a bit more population than that of control.However,the bacteria in stem and leaf of treated seedlings had less population than those of control.After the heat treatment of seedlings combined with shoot-tip culture,the endophytic bacteria can be removed effectively as the temperature increased daily,with less number compared with control.And when it reached 40 ℃,the bacterial growth was inhibited largely.Heat treatment at 37 ℃ reduced the number of indigenous bacteria at 20th and 21st day,with a population of 5.33×108 CFU/g and 4.02×108 CFU/g,respectively,and only about one seventh of the number of control at 3.43×109 CFU/g.When the temperature for heat treatment reached 40 ℃,the number of bacteria decreased to 1.63×108 CFU/g,occupied only one forty-seventh of the bacteria number of control at 7.70×109CFU/g,indicating that treatment with higher temperature at 37 ℃ and 40 ℃ could significantly reduce the number of bacteria in seedlings grown in vitro culture.The population of the endophytic bacteria of E.urophylla increased obviously after growing in the MS supplemented with antibiotics,i.e.,oxytetracycline,tetracycline and cephamycins,and the reason for this phenomenon is not yet known.Based on these results,it can be concluded that proper high temperature combined with apical meristem culture could effectively reduce the number of indigenous endophytic bacteria in the tissue of E.urophylla,but not completely eliminate them.The stain procedure could detect the distribution of bacteria in plant tissue much more quickly and effectively.