Elvira-Recuenco Margarita, van Vuurde Jim W L
Dep. Protección Vegetal, INIA, Ctra. La Coruña Km. 7.5, 28040 Madrid, Spain.
Microbiol Res. 2003;158(4):271-9. doi: 10.1078/0944-5013-00205.
The L-form of Pseudomonas syringae pv. phaseolicola has been proved to induce resistance to bean halo blight. Various procedures were tested to induce the L-form of Pseudomonas syringae pv. pisi for its potential use as biocontrol agent of pea bacterial blight. Cell-wall deficient cells were induced in a liquid medium with penicillin following a protocol described for P. s. pv. phaseolicola. Cell growth on solid induction medium developed as typical granular and vacuolated structures, and characteristic colonies were observed in the first transfer. However, there was poor growth in subsequent transfers and some reversion to the parental type. To improve the induction procedure, the following new procedures were applied: (1) viability of cells was monitored during induction. The optimum induction time in liquid medium with penicillin was lower for pv. pisi than for pv. phaseolicola. Viability of L-forms in solid induction medium with penicillin was low and decreased in time. (2) the inducer ticarcillin was combined with clavulanic acid, which prevented the reversion to the parental type and (3) a range of concentrations of penicillin and ticarcillin/clavulanic acid was applied by the spiral gradient endpoint method for calculation of minimum inhibitory concentrations (MIC). Based on the results from these tests an induction method for P. s. pv. pisi L-form is proposed and the relevance of L-form is discussed for practice.
丁香假单胞菌菜豆致病变种的L型已被证明可诱导对菜豆晕疫病的抗性。人们测试了各种方法来诱导豌豆假单胞菌致病变种的L型，以探究其作为豌豆细菌性疫病生物防治剂的潜力。按照针对菜豆假单胞菌致病变种描述的方案，在含有青霉素的液体培养基中诱导细胞壁缺陷型细胞。在固体诱导培养基上的细胞生长形成典型的颗粒状和空泡状结构，并且在首次传代时观察到特征性菌落。然而，后续传代中的生长较差，并且有些回复为亲本类型。为了改进诱导程序，应用了以下新程序：（1）在诱导过程中监测细胞活力。对于豌豆假单胞菌致病变种，在含有青霉素的液体培养基中的最佳诱导时间比菜豆假单胞菌致病变种短。在含有青霉素的固体诱导培养基中，L型的活力较低且随时间下降。（2）将诱导剂替卡西林与克拉维酸联合使用，可防止回复为亲本类型；（3）通过螺旋梯度终点法应用一系列浓度的青霉素和替卡西林/克拉维酸来计算最小抑菌浓度（MIC）。基于这些测试结果，提出了一种诱导豌豆假单胞菌致病变种L型的方法，并讨论了L型在实际应用中的相关性。
