植物乳杆菌促进黑腹果蝇生长发育

Lactobacillus plantarum promotes the growth and development of Drosophila melanogaster

【目的】检测乳酸菌对果蝇发育历期的影响,进一步探讨其对果蝇促生长的分子机制。【方法】利用选择性培养基MRS从黑腹果蝇Drosophila melanogaster体内分离乳酸菌,利用革兰氏染色、生化方法及16S rRNA基因进行鉴定;通过体内定植和世代传递实验验证该菌是黑腹果蝇的共生菌;采用悉生模型检测乳酸菌对黑腹果蝇发育的促生长作用;利用实时定量PCR技术检测黑腹果蝇体内促前胸腺激素基因PTTH和胰岛素通路相关基因InR的表达水平;利用葡萄糖试剂盒检测血淋巴液葡萄糖浓度。【结果】从黑腹果蝇中分离到的菌株鉴定为植物乳杆菌Lactobacillus plantarum FY1菌株(Gen Bank登录号:KY038178),可在黑腹果蝇肠道内定植,每个肠道定植量约为104CFU,并能在世代间稳定传递。FY1菌株体外发酵可降低p H值至5.2,可诱导无菌果蝇卵至蛹发育时间由20.0 d缩短至6.9 d,卵至成虫发育时间由30.0 d缩短至10.7 d,其生长速率是无菌果蝇的约2倍。实时定量PCR结果表明,FY1菌株显著地提前了PTTH表达高峰期,同时降低果蝇中InR表达水平,血淋巴液葡萄糖浓度从5.1 mg/mL降低至2.7 mg/mL。【结论】植物乳杆菌是黑腹果蝇的一种益生菌,推测能通过胰岛素信号通路促进宿主黑腹果蝇的生长和发育。

【Aim】This study aims to assay the effect of lactic acid bacteria on the developmental duration of fruit flies,and to further exploit their molecular mechanisms of growth stimulation of fruit flies.【Methods】Lactic acid bacteria were isolated from D.melanogaster using lactic acidic bacteria-selective medium MRS and identified by Gram staining,biochemical and 16 S rRNA gene techniques.The symbiotic relationship was assayed by in vivo colonization and generation transfer experiments.The growth promoting effect of lactic acid bacteria on D.melanogaster was assessed in germ-free and gnotobiotic model.The expression levels of prothoracicotropic hormone gene（PTTH） and insulin-like receptor gene（InR） were examined using real-time PCR.The glucose concentration in haemolymph was examinedusing commercial kit with spetrometry.【Results 】 The isolated bacterium from D.melanogaster was identified as Lactobacillus plantarum strain FY1（Gen Bank accesson no.：KY038178）,which could colonize in the guts of flies with a mount of 10~4CFU/gut and be transferred from the parental generation to their offspring.L.plantarum FY1 decreased the pH value of culture media to 5.2 in vitro,and shortened the egg-to-puparium duration of D.melanogaster from 20.0 d to 6.9 d and the egg-to-adult duration from 30.0 d to 10.7 d,respectively,resulting in 2-fold growth rate of flies compared with that of germ-free siblings.Real-time PCR result revealed that L.plantarum FY1 significantly stimulated the expression peak of PTTH gene in advance,while decreased the expression level of InR gene in D.melanogaster,and reduced the glucose concentration in haemolymph from 5.1 mg/m L to 2.7 mg/m L.【Conclusion】L.plantarum is a type of commensal bacteria of D.melanogaster,and might promote the growth and development of the host via insulin signal pathway.