【目的】明确入侵害虫红棕象甲Rhynchophorus ferrugineus胞质型肽聚糖识别蛋白RfPGRP-L2在肠道菌群稳态的维持和调控过程中的作用,将为靶向破坏肠道菌群稳态的害虫控制新策略研发提供新的科学依据和作用靶标。【方法】利用生物信息学...【目的】明确入侵害虫红棕象甲Rhynchophorus ferrugineus胞质型肽聚糖识别蛋白RfPGRP-L2在肠道菌群稳态的维持和调控过程中的作用,将为靶向破坏肠道菌群稳态的害虫控制新策略研发提供新的科学依据和作用靶标。【方法】利用生物信息学方法分析RfPGRP-L2的序列特征。利用RT-qPCR分析RfPGRP-L2在健康红棕象甲4龄幼虫不同组织(头、脂肪体、表皮、前肠、中-后肠、血淋巴)以及大肠杆菌Escherichia coli DH5α和金黄色葡萄球菌Staphylococcus aureus经注射(注射1μL OD 600=1.6的菌液)和喂食(取食涂抹1 mL OD 600=1.6的菌液的甘蔗薄片)两种不同方式分别感染后红棕象甲4龄幼虫肠道和脂肪体中的表达量;进行RfPGRP-L2原核表达,利用体外孵育方法检测重组蛋白RfPGRP-L2对大肠杆菌DH5α和金黄色葡萄球菌的凝集和抑菌活性;RNAi干扰RfPGRP-L2后,检测红棕象甲4龄幼虫血淋巴和肠道中大肠杆菌菌落数的变化;利用RT-qPCR分析RNAi干扰RfPGRP-L2后红棕象甲4龄幼虫脂肪体和肠道中抗菌肽基因表达量的变化;利用基于细菌16S rRNA的高通量测序分析RNAi干扰RfPGRP-L2对健康红棕象甲4龄幼虫肠道菌群结构组成的影响。【结果】SMART预测发现红棕象甲RfPGRP-L2基因编码的蛋白中无跨膜结构域也无信号肽,这表明RfPGRP-L2是一种胞质型肽聚糖识别蛋白。RT-qPCR检测发现,RfPGRP-L2主要在健康红棕象甲4龄幼虫血淋巴、肠道和脂肪体等免疫组织中表达;被注射感染大肠杆菌和金黄色葡萄球菌6 h和12 h后,红棕象甲4龄幼虫脂肪体中RfPGRP-L2的表达量分别显著上调;被喂食感染大肠杆菌6 h后,红棕象甲4龄幼虫肠道中RfPGRP-L2的表达量显著增加。重组表达蛋白RfPGRP-L2能引起大肠杆菌和金黄色葡萄球菌发生凝集反应,这说明RfPGRP-L2能够识别这两种细菌。当RfPGRP-L2被干扰后,红棕象甲4龄幼虫对肠道和血淋巴中感染EGFP标记的大肠杆菌的清除能力显著弱于对照组;肠道中抗菌肽基因RfCecropin的表达量显著降低;健康红棕象甲4龄幼虫肠道中细菌的菌落数量显著高于对照组,而且肠道菌群结构组成也发生了明显的变化。【结论】红棕象甲体内胞质型肽聚糖识别蛋白RfPGRP-L2能够通过识别细菌并激活肠上皮细胞中相应的免疫信号通路促进抗菌肽基因的表达,从而介导对肠道菌群稳态的调控。展开更多
After exposure to mixed environments with multiple hosts, the frequency of flight take-offs for Plutella xylostella(L.) is known to increase with a concomitant decrease in female fecundity. The objective of the presen...After exposure to mixed environments with multiple hosts, the frequency of flight take-offs for Plutella xylostella(L.) is known to increase with a concomitant decrease in female fecundity. The objective of the present study was to elucidate the physiological mechanism underlying the increased flight activity and decreased fecundity of P. xylostella in mixed-host environments. We measured ovariole length, and triglyceride and vitellogenin(Vg) content in P. xylostella females after they were released into different host environments consisting of either a single host(Brassica campestris, Brassica oleracea or Brassica juncea) or a mixture of all three hosts. Our results indicated that ovariole length varied significantly between female adults from different host environments. Females from the B. campestris environment had the longest ovarioles, whereas those from the mixed environment had the shortest ones. A negative correlation was found between ovariole length and the flight take-off frequency of P. xylostella adults. Additionally, there were significant differences in the triglyceride content of P. xylostella females from different host environments. Our data revealed that more triglyceride was consumed by P. xylostella female adults from B. oleracea and the mixed environments than those from environments containing only B. campestris or B. juncea. In contrast, the relative Vg content in P. xylostella females from the mixed environment was lower than that in females from the B. campestris-, B. juncea-or B. oleracea-only environments. In conclusion, the mixed environment caused increased consumption of available energy resources(triglyceride) at the cost of retarding ovarian development and decreasing the amount of Vg produced.展开更多
文摘【目的】明确入侵害虫红棕象甲Rhynchophorus ferrugineus胞质型肽聚糖识别蛋白RfPGRP-L2在肠道菌群稳态的维持和调控过程中的作用,将为靶向破坏肠道菌群稳态的害虫控制新策略研发提供新的科学依据和作用靶标。【方法】利用生物信息学方法分析RfPGRP-L2的序列特征。利用RT-qPCR分析RfPGRP-L2在健康红棕象甲4龄幼虫不同组织(头、脂肪体、表皮、前肠、中-后肠、血淋巴)以及大肠杆菌Escherichia coli DH5α和金黄色葡萄球菌Staphylococcus aureus经注射(注射1μL OD 600=1.6的菌液)和喂食(取食涂抹1 mL OD 600=1.6的菌液的甘蔗薄片)两种不同方式分别感染后红棕象甲4龄幼虫肠道和脂肪体中的表达量;进行RfPGRP-L2原核表达,利用体外孵育方法检测重组蛋白RfPGRP-L2对大肠杆菌DH5α和金黄色葡萄球菌的凝集和抑菌活性;RNAi干扰RfPGRP-L2后,检测红棕象甲4龄幼虫血淋巴和肠道中大肠杆菌菌落数的变化;利用RT-qPCR分析RNAi干扰RfPGRP-L2后红棕象甲4龄幼虫脂肪体和肠道中抗菌肽基因表达量的变化;利用基于细菌16S rRNA的高通量测序分析RNAi干扰RfPGRP-L2对健康红棕象甲4龄幼虫肠道菌群结构组成的影响。【结果】SMART预测发现红棕象甲RfPGRP-L2基因编码的蛋白中无跨膜结构域也无信号肽,这表明RfPGRP-L2是一种胞质型肽聚糖识别蛋白。RT-qPCR检测发现,RfPGRP-L2主要在健康红棕象甲4龄幼虫血淋巴、肠道和脂肪体等免疫组织中表达;被注射感染大肠杆菌和金黄色葡萄球菌6 h和12 h后,红棕象甲4龄幼虫脂肪体中RfPGRP-L2的表达量分别显著上调;被喂食感染大肠杆菌6 h后,红棕象甲4龄幼虫肠道中RfPGRP-L2的表达量显著增加。重组表达蛋白RfPGRP-L2能引起大肠杆菌和金黄色葡萄球菌发生凝集反应,这说明RfPGRP-L2能够识别这两种细菌。当RfPGRP-L2被干扰后,红棕象甲4龄幼虫对肠道和血淋巴中感染EGFP标记的大肠杆菌的清除能力显著弱于对照组;肠道中抗菌肽基因RfCecropin的表达量显著降低;健康红棕象甲4龄幼虫肠道中细菌的菌落数量显著高于对照组,而且肠道菌群结构组成也发生了明显的变化。【结论】红棕象甲体内胞质型肽聚糖识别蛋白RfPGRP-L2能够通过识别细菌并激活肠上皮细胞中相应的免疫信号通路促进抗菌肽基因的表达,从而介导对肠道菌群稳态的调控。
基金supported by the Key Program of the National Natural Science Foundation of China(31230061)the special fund for Agro-scientific Research in the Public Interest,China(201103021)+1 种基金the Key Technologies R&D Program of China during the 12th Five-Year Plan period(2012BAD19B06)the Natural Science Foundation of Fujian Province,China(2013J01086)
文摘After exposure to mixed environments with multiple hosts, the frequency of flight take-offs for Plutella xylostella(L.) is known to increase with a concomitant decrease in female fecundity. The objective of the present study was to elucidate the physiological mechanism underlying the increased flight activity and decreased fecundity of P. xylostella in mixed-host environments. We measured ovariole length, and triglyceride and vitellogenin(Vg) content in P. xylostella females after they were released into different host environments consisting of either a single host(Brassica campestris, Brassica oleracea or Brassica juncea) or a mixture of all three hosts. Our results indicated that ovariole length varied significantly between female adults from different host environments. Females from the B. campestris environment had the longest ovarioles, whereas those from the mixed environment had the shortest ones. A negative correlation was found between ovariole length and the flight take-off frequency of P. xylostella adults. Additionally, there were significant differences in the triglyceride content of P. xylostella females from different host environments. Our data revealed that more triglyceride was consumed by P. xylostella female adults from B. oleracea and the mixed environments than those from environments containing only B. campestris or B. juncea. In contrast, the relative Vg content in P. xylostella females from the mixed environment was lower than that in females from the B. campestris-, B. juncea-or B. oleracea-only environments. In conclusion, the mixed environment caused increased consumption of available energy resources(triglyceride) at the cost of retarding ovarian development and decreasing the amount of Vg produced.
