,西南大学植物保护学院,重庆 400716Comparison of Metabolic Profile Between Diapause-Destined and Non-Diapause-Destined Pupae of Bactrocera minax
WANG Jia, WANG Pan, FAN Huan, LIU YingHong,College of Plant Protection, Southwest University, Chongqing 400716通讯作者:
责任编辑: 岳梅
收稿日期:2018-11-18接受日期:2019-01-12网络出版日期:2019-03-16
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Received:2018-11-18Accepted:2019-01-12Online:2019-03-16
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王佳,E-mail： aimarjia@126.com。
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王佳, 王攀, 樊欢, 刘映红. 柑橘大实蝇滞育型与非滞育型蛹的代谢谱比较[J]. 中国农业科学, 2019, 52(6): 1021-1031 doi:10.3864/j.issn.0578-1752.2019.06.006
WANG Jia, WANG Pan, FAN Huan, LIU YingHong.
0 引言
【研究意义】柑橘大实蝇（Bactrocera minax）是一种重要的柑橘害虫,近年来对柑橘产业造成了巨大损失[1,2,3]。该虫为一化性害虫,越冬蛹历期可长达6个月,其间发生专性滞育。滞育在协调昆虫与环境之间的关系中具有十分重要的作用,探讨柑橘大实蝇滞育过程中体内物质代谢的变化,不仅有利于了解其生长发育的内在规律,也有助于揭示柑橘大实蝇发生滞育并适应恶劣环境的分子机制。【前人研究进展】一化性昆虫的专性滞育固定发生在特定的虫态下,不需要接受任何外界因子的诱导[4]。柑橘大实蝇在蛹期出现专性滞育,但滞育起始和终止的确切时间长期以来都不明确,而新陈代谢受到强烈抑制是滞育的典型特征,因此可以将呼吸速率作为判断滞育进程的指标[5,6]。前期通过对柑橘大实蝇整个蛹期进行呼吸速率的监测,发现在化蛹一个月以后进入深度滞育状态,大约持续两个月后滞育解除并恢复发育[7]。滞育是一个复杂的生理过程,而各种高通量、大规模的组学检测分析技术为全面、深入地发掘滞育机理提供了便利,并且得到广泛应用[7,8,9,10,11,12,13]。核磁共振技术（nuclear magnetic resonance,NMR）是代谢组学研究的主要方法之一,可全面检测生物体内各种代谢路径的底物和产物小分子代谢物水平在某种刺激或状态下所产生的整体变化（量变以及质变）,来探索生命体的生物过程[14,15]。近年来,代谢组检测在昆虫研究领域中逐渐得到应用,包括昆虫滞育的相关研究[7,9-10,13]。目前,已有研究利用表达谱和代谢谱技术对柑橘大实蝇不同发育阶段的蛹进行比较分析,发掘出在蛹滞育期与滞育前、后有显著变化的基因和代谢物,它们可能参与到滞育的起始或结束过程中[7,8]。在此基础上,进一步将滞育蛹与非滞育蛹进行组学比较分析,能够验证并最终明确与滞育相关的代谢物、基因和蛋白质等。鉴于柑橘大实蝇发生专性滞育的习性,无法通过改变环境来获取非滞育蛹,但可通过在化蛹初期注射20-羟基蜕皮酮（20E）来解除滞育[16,17]。此外,已有的柑橘大实蝇转录组数据能够为后期深入研究滞育相关生理途径的作用机理提供基础[8,18]。【本研究切入点】通过在柑橘大实蝇化蛹初期注射20E和溶剂,分别获取非滞育型蛹（non-diapause-destined,ND）与对照滞育型蛹（diapause-destined,D）,再利用NMR检测技术对两者的代谢组进行组内和组间的对比分析。【拟解决的关键问题】通过代谢组分析明确D型与ND型蛹各自在不同时间点之间的代谢物水平变化情况,以及相同时间点上D型与ND型蛹之间的代谢物浓度变化情况。发掘出与滞育相关的代谢物,为后续深入研究其在滞育中的作用机理打下基础。1 材料与方法
试验于2017—2018年在西南大学和武汉安隆科讯技术有限公司完成。1.1 供试昆虫
于重庆市武隆县一个橘园中（29°20′56″N,107°45′20″E）采集柑橘大实蝇受害果并带回实验室。将受害果逐一剖开,取出柑橘大实蝇老熟幼虫,放置于沙土上使其自然入土化蛹。试验中所使用的蛹全都埋于沙土中,并放置于室外自然环境温度下。定期向土壤中喷水,使土壤湿度保持在适宜蛹存活的40%—70%。1.2 非滞育型蛹与滞育型蛹的获取
柑橘大实蝇蛹滞育类型为专性滞育,自然状态下,所有虫体在化蛹约1个月后都将进入深度滞育状态。为了获取非滞育型蛹,在化蛹后1 d内,将20E粉末（HPLC级,Sigma公司）溶解于10%乙醇溶液中,配制终浓度为1.0 μg·μL-1的20E溶液,随后使用微注射仪（美国Drummond公司）在蛹的尾部背面注射1.0 μL的20E溶液以解除滞育[16];同时,向其余部分蛹体内注射10%乙醇溶剂以获取对照滞育型蛹。最后将所有注射后的蛹埋入沙土中备用。1.3 代谢物提取
在注射后的1、15和30 d,分别收集滞育型蛹（D1、D15和D30）与非滞育型蛹（ND1、ND15和ND30）,将蛹置于研钵中,加入液氮充分研磨至粉末。每个时间点上的处理均设置5个生物学重复,每个重复取1头蛹。在每个重复中取15 mg研磨后的样品粉末,加入500 μL纯水,涡旋振荡1 min;在冰浴中超声波破碎4 s,间隔3 s,循环8次;13 000 r/min、4℃离心5 min后,将上清液于预洗的超滤膜（美国Millipore公司）中以13 000 r/min、4℃离心45 min过滤;取300 μL滤液与-80℃中预冷冻,然后再移至冻干机（美国LABCONCO公司）中冷冻干燥;加入135 μL重水复溶样本,再加入15 μL ACDSS（Anachro Certified DSS Standard Solution）标准溶液,充分混匀;13 000 r/min、4℃离心2 min后,将上清液转移至微量管中,加入400 μL重水,将微量管装入核磁管中,封口备用。1.4 样品的NMR检测
采用NMR谱仪（Agilent DD2 600 MHz spectrometer equipped with a triple-resonance cryoprobe,美国Agilent公司）采集核磁共振谱图。设置采集参数：温度298.15 K,共振频率599.79 MHz,采样数据点32 768个,累加次数128,谱宽7 225.434 Hz。将1H NMR自由感应衰减（free induction decay,FID）信号导入到Chenomx NMR suit（version 8.3,Chenomx,Edmonton,Canada）软件中,自动进行傅立叶转换,调整相位,校正基线。以DSS峰（0.0 ppm）作为全部谱图化学位移的标准,并对其进行反转卷积操作,调整谱图峰形（CSI）。根据NMR谱图中信号的相关信息（如化学位移、峰形、半峰宽、耦合裂分等）,以DSS的浓度和谱峰面积为标准,结合Chenomx自带数据库对谱图的信号逐一比对分析,确定代谢物种类和对应的绝对浓度值。然后把代谢物和对应的绝对浓度值导出到EXCEL表格中,得到变量矩阵,用于后续分析。1.5 数据分析
将变量矩阵作为源数据进行质量归一化处理,导入R平台的PLS工具包,用偏最小二乘判别分析法（partial least squares discriminant analysis,PLS-DA）进行数据分析[19],判定D型与ND型蛹各自在不同时间点之间（D1对D15对D30;ND1对ND15对ND30）,以及相同时间点上D型与ND型蛹之间的代谢谱差异（D1对ND1;D15对ND15;D30对ND30）,通过观察PLS-DA得分图上各处理样品点之间的分离情况,判断代谢谱差异的显著性。样本组之间分离越远,则代谢谱差异越显著,反之则越相似。此外,在PLS-DA分析的基础上计算变量投影重要性值（variable importance in projection,VIP）,VIP值越大说明代谢物在区分样品中所作贡献越大。随后,采用SPSS21.0软件对两种处理组内不同时间点的代谢物浓度进行单因素方差分析;对两种处理组间相同时间点上的代谢物浓度进行t检验。同时满足VIP>1,且P<0.05的代谢物被认定为在比较中具有显著性差异。2 结果
2.1 柑橘大实蝇蛹的NMR图谱分析
根据柑橘大实蝇蛹的代谢物NMR归属图谱,参照Chenomx自带数据库对30张样本谱图的信号进行逐一比对分析,共获得50种代谢物及其浓度。其中,氨基酸及其衍生物20种、有机酸11种、糖类4种、核酸组分5种,其他代谢物10种（图1）。将代谢物的浓度进行质量归一化处理后,得到了每种代谢物在6个不同处理间的浓度比例（图2）。图1
新窗口打开|下载原图ZIP|生成PPT图1柑橘大实蝇蛹代谢物的NMR图谱
图中数字表示所检测代谢物的特征峰位置Numbers in the figure indicate the characteristic peak of the detected metabolites。1：2-羟基异丁酸甲酯2-Hydroxyisobutyrate;2：2-氧戊二酸盐2-Oxoglutarate;3：3-羟基犬尿氨酸3-Hydroxykynurenine;4：醋酸盐Acetate;5：丙氨酸Alanine;6：精氨酸Arginine;7：天冬酰胺Asparagine;8：天冬氨酸Aspartate;9：丁酸盐Butyrate;10：肉碱Carnitine;11：胆碱Choline;12：柠檬酸盐Citrate;13：胞苷Cytidine;14：乙醇Ethanol;15：乙醇胺Ethanolamine;16：甲酸盐Formate;17：延胡索酸盐Fumarate;18：葡萄糖Glucose;19：谷氨酸Glutamate;20：谷氨酰胺Glutamine;21：甘氨酸Glycine;22：鸟嘌呤Guanosine;23：组氨酸Histidine;24：肌苷Inosine;25：异亮氨酸Isoleucine;26：乳酸Lactate;27：亮氨酸Leucine;28：赖氨酸Lysine;29：麦芽糖Maltose;30：甲醇Methanol;31：N-乙酰谷氨酸N-Acetylglutamate;32：胆碱磷酸O-Phosphocholine;33：O-磷酸乙醇胺O-Phosphoethanolamine;34：泛酸盐Pantothenate;35：苯基丙氨Phenylalanine;36：脯氨酸Proline;37：丙酸Propionate;38：丙酮酸Pyruvate;39：琥珀酸Succinate;40：苏氨酸Threonine;41：海藻糖Trehalose;42：葫芦巴碱Trigonelline;43：色氨酸Tryptophan;44：酪氨酸Tyrosine;45：糖胺UDP-N-Acetylglucosamine;46：尿苷二磷酸葡萄糖UDP-glucose;47：鸟苷Uridine;48：缬氨酸Valine;49：肌醇myo-Inositol;50：β-丙氨酸β-Alanine
Fig. 1NMR spectra of metabolites in B. minax pupae
图2
新窗口打开|下载原图ZIP|生成PPT图2柑橘大实蝇蛹代谢物在不同处理中的浓度百分比
D1、D15、D30：注射后1、15和30 d的滞育型蛹Diapause-destined pupae 1, 15 and 30 d after injection;ND1、ND15、ND30：注射后1、15和30 d的非滞育型蛹Non-diapause-destined pupae 1, 15 and 30 d after injection
Fig. 2Percentage of metabolite concentrations in different treatments of B. minax pupae
2.2 代谢谱差异的PLS-DA分析
利用PLS-DA分析D型与ND型蛹在组内不同时间点,以及组间相同时间点的代谢谱差异。共5种对比组合,分别绘制得分图。D型蛹的3个时间点样品在得分图中分离较好,且每个时间点5个样品相对聚拢,表示不同时间点样品代谢谱差异显著,且各处理重复性较好（图3-A）;ND型蛹在注射20E后15 d和30 d样品之间的分离性较差,无显著差异,但两者均与注射后1 d样品之间的分离性较好,差异显著（图3-B）。在3个不同时间点上,D型与ND型蛹样品在得分图中均明显分离,呈现显著差异（图3-C、3-D、3-E）。进一步对5种对比组合的载荷图进行分析发现,大多数代谢物集中在原点附近,表示对差异贡献不大;少数代谢物在载荷1和载荷2方向上都远离原点,表示对差异贡献较大,且距离越远,对差异贡献越大（图4）。图3
新窗口打开|下载原图ZIP|生成PPT图3不同对比组合间的PLS-DA得分图
Fig. 3PLS-DA scores plot for different types of comparisons
A: D1 vs D15 vs D30; B: ND1 vs ND15 vs ND30; C: D1 vs ND1; D: D15 vs ND15; E: D30 vs ND30
图4
新窗口打开|下载原图ZIP|生成PPT图4不同对比组合间的PLS-DA载荷图
Fig. 4PLS-DA loading plot for different types of comparisons
A: D1 vs D15 vs D30; B: ND1 vs ND15 vs ND30; C: D1 vs ND1; D: D15 vs ND15; E: D30 vs ND30
2.3 代谢谱差异的VIP分析
在PLS-DA分析的基础上,计算每个代谢物的VIP值,对VIP>1的代谢物进行方差分析或t检验。在5个对比组合中,共发现21种具有显著差异的代谢物（VIP>1且P<0.05）,其中,D1对D15对D30组合中有10种（图5-A）;ND1对ND15对ND30组合中有12种（图5-B）;D1对ND1组合中有6种（图5-C）;D15对ND15组合中有12种（图5-D）;D30对ND30组合中有8种（图5-E）。在这21种代谢物中,柠檬酸、葡萄糖在5个对比组合中都差异显著;麦芽糖和脯氨酸在4个对比组合中差异显著（麦芽糖：D1对D15对D30、D1对ND1、D15对ND15、D30对ND30;脯氨酸：D1对D15对D30、ND1对ND15对ND30、D15对ND15、D30对ND30）;海藻糖、谷氨酸、天冬酰胺和N-乙酰谷氨酸在3个对比组合中差异显著（海藻糖、谷氨酸：D1对D15对D30、D15对ND15、D30对ND30;天冬酰胺：D1对D15对D30、ND1对ND15对ND30、D1对ND1;N-乙酰谷氨酸：ND1对ND15对ND30、D15对ND15、D30对ND30）。图5
新窗口打开|下载原图ZIP|生成PPT图5不同对比组合间的VIP得分图
图左侧为VIP值排名前15的代谢物Metabolites of the top 15 VIP values are listed on the left;图右侧表示各代谢物在不同处理中的相对浓度The relative concentrations of metabolites are presented on the right。红色圆点表示满足VIP>1,且P<0.05的代谢物Red dots indicate metabolites with VIP>1 and P<0.05
Fig. 5VIP scores plot for different types of comparisons
A: D1 vs D15 vs D30; B: ND1 vs ND15 vs ND30; C: D1 vs ND1; D: D15 vs ND15; E: D30 vs ND30
3 讨论
柑橘大实蝇D型蛹在3个时间上的代谢谱呈现显著差异,这与滞育深度逐渐加强,代谢水平逐渐受到抑制有关。ND15与ND30的代谢谱差异不显著,这可能是由于ND型蛹在这两个时间点上均处于解除滞育后的发育状态,物质代谢相对保持稳定。D型与ND型蛹在不同时间点上的代谢谱均具有显著差异,但在D1对ND1组合中,差异显著的代谢物数量较少,且大多数代谢物在PLS-DA载荷图中分布更集中,距离原点更近,说明此时代谢谱差异小于其他两个时间点。在本研究5个组合中,共有21种代谢物呈现出显著差异,其中柠檬酸、葡萄糖、麦芽糖、脯氨酸、海藻糖、谷氨酸、天冬酰胺和N-乙酰谷氨酸出现显著差异的次数最多,且在相应组合中的VIP值排名靠前。这与麻蝇（Sarcophaga crassipalpis）[9]和棉铃虫（Helicoverpa armigera）[10]中的相关研究结果相似,表明这些物质与昆虫滞育存在普遍联系。在这些物质中,柠檬酸是三羧酸循环的中间产物;葡萄糖经糖酵解后生成丙酮酸,可作为三羧酸循环的起始物质;麦芽糖和海藻糖可分解成葡萄糖,随后进入糖酵解和三羧酸循环;谷氨酸和天冬酰胺经过转化后可分别生成三羧酸循环中间物质2-氧戊二酸和草酰乙酸[20]。此外,三羧酸循环中的2-氧戊二酸、丙酮酸和琥珀酸也在特定组合中呈现显著差异。这些结果表明,柑橘大实蝇滞育对三羧酸循环产生了显著影响。在生命体中,三羧酸循环不仅为生命体提供能量,同时也是糖类、脂类和氨基酸三者之间相互转化的枢纽[20]。已有研究表明,三羧酸循环在昆虫滞育期间受到抑制[9,21-23],这与滞育期间代谢减弱相符合。另外,三羧酸循环的中间物质还能够刺激蜕皮激素分泌,从而调控昆虫的滞育进程[22]。在本研究中,三羧酸循环中的柠檬酸、丙酮酸和2-氧戊二酸在解除滞育15 d和30 d后,含量显著下降。一方面可能是因为蛹恢复发育后需要大量能量,从而增加了其消耗;另一方面可能是其转化为了其他物质,以保证正常发育下的物质平衡。然而,该循环在柑橘大实蝇滞育过程中的作用仍有待进一步研究。
本研究发掘的一些差异显著的代谢物还与抗逆性有关。海藻糖在昆虫体内发挥至关重要的作用,既能作为能量来源物质,也能协助细胞抵御干燥、脱水、低温及氧化等多种环境胁迫[24,25,26]。研究表明,许多昆虫在越冬滞育期间通过积累海藻糖来抵御低温[27,28,29]。脯氨酸也能作为昆虫防冻剂,在低温下避免体液结冰对细胞造成伤害[30,31]。前期研究结果表明,柑橘大实蝇D型蛹体内海藻糖和脯氨酸含量在深度滞育期间显著高于滞育前和滞育后,总体含量呈倒“U”字型变化[7]。在本研究中发现,D型蛹体内海藻糖和脯氨酸含量在化蛹后30 d内呈下降趋势,而在ND型蛹体内呈稳定或上升趋势。结合前后研究结果推断,D型蛹在进入深度滞育前,由于代谢水平逐渐降低,海藻糖和脯氨酸的含量也随之降低;进入深度滞育后,随着气温的降低,海藻糖和脯氨酸的含量也随之上升,以协助细胞抵御低温。而ND型蛹在彻底解除滞育后,海藻糖和脯氨酸的含量与代谢水平一样保持相对稳定。此外,肌醇也能够在低温环境下为多种昆虫提供抗冻保护,尤其是提高一些滞育越冬昆虫的耐寒性[21,32-33]。在本研究中,随着时间推移,D型蛹体内的肌醇含量始终较低,而ND型蛹体内的肌醇含量逐渐上升,这可能有助于提高耐寒性,从而应对冬季低温。
此外,还有一些差异显著的代谢物具有其他重要的功能。3-羟基犬尿氨酸是色氨酸的代谢产物,与眼色素的形成有密切关系[34,35],因此,ND型蛹体内高浓度的3-羟基犬尿氨酸可能与复眼发育需要大量眼色素有关。N-乙酰谷氨酸是尿素循环的强制激活剂,促进新陈代谢中产生的有毒物质氨进入尿素循环,最终转化为无毒的尿素[36]。在注射后15 d和30 d,N-乙酰谷氨酸在D型蛹中浓度高于非滞育型蛹,表明D型蛹体内将有更多的氨进入尿素循环。这可能是由于D型蛹较弱的新陈代谢抑制了氨基酸的合成,导致氨过剩的结果。此外,有研究报道,滞育型棉铃虫幼虫可能通过在体内积累大量尿素达到抵御低温的作用[10]。柑橘大实蝇D型蛹也同样可能利用尿素来提高其耐寒性。
4 结论
在柑橘大实蝇非滞育型蛹（ND）与对照滞育型蛹（D）的5个组合（D1对D15对D30;ND1对ND15对ND30;D1对ND1;D15对ND15;D30对ND30）中,共有21种代谢物浓度至少出现一次显著差异,其中柠檬酸、葡萄糖、麦芽糖、脯氨酸、海藻糖、谷氨酸、天冬酰胺和N-乙酰谷氨酸出现显著差异的次数最多。这些物质能直接或间接进入三羧酸循环,说明柑橘大实蝇滞育期间能量代谢和物质转化产生了显著变化。此外,海藻糖、脯氨酸、N-乙酰谷氨酸和肌醇的显著差异有助于调控柑橘大实蝇滞育期间的耐寒性。研究结果可为后续解析重要代谢物及相关生理途径在柑橘大实蝇滞育中的作用机理提供依据。参考文献 原文顺序
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被引期刊影响因子
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DOI:10.1603/0022-0493-99.5.1651URLPMID:17066795 [本文引用: 1]
Bactrocera minax (Enderlein) (Diptera: Tephritidae) is a major pest of citrus fruit in the region from Nepal through to southwestern China. In tests on wild adult populations of B. minax in a mandarin, Citrus reticulata Blanco, orchard in western Bhutan, both males and females were more attracted to 50-mm-diameter spheres than to 50-mm discs of the same color. Furthermore, they were more attracted to spheres colored with orange or green-yellow mixtures than to similar spheres colored red, yellow, green, blue, black, or white. The UV reflectance from the orange (600-650 nm) and yellow-green mixtures (530 nm) was similar to that proposed byProkopy (1977) as eliciting strong attractant responses in other species of Tephritidae.
DOI:10.3969/j.issn.1674-0858.2009.01.012URL [本文引用: 1]
本文综述了桔大实蝇Bactrocera(Tetradacus)minax Enderlein主要生物学、生态学特点和防治措施的研究进展,包括桔大实蝇在国内的分布情况,对寄主的选择性,各虫态的形态特征、发育历期和不同因子对其生长发育的影响,生活史,种群动态以及农业防治、化学防治、诱杀防治、生物防治、物理防治等不同防治方法的研究和应用情况。对今后的防治和研究提出了一些建议。
DOI:10.3969/j.issn.1674-0858.2009.01.012URL [本文引用: 1]
本文综述了桔大实蝇Bactrocera(Tetradacus)minax Enderlein主要生物学、生态学特点和防治措施的研究进展,包括桔大实蝇在国内的分布情况,对寄主的选择性,各虫态的形态特征、发育历期和不同因子对其生长发育的影响,生活史,种群动态以及农业防治、化学防治、诱杀防治、生物防治、物理防治等不同防治方法的研究和应用情况。对今后的防治和研究提出了一些建议。
DOI:10.1146/annurev.ento.47.091201.145137URL [本文引用: 1]
DOI:10.1016/j.jinsphys.2007.03.018URLPMID:17532002 [本文引用: 1]
Insects in diapause characteristically feed very little or not at all, thus they are largely or totally dependent on energy reserves sequestered prior to the entry into diapause. Fats are the dominant reserve used during this period, but non-fat reserves are also important for some species, especially during certain phases of diapause. Metabolic depression, coupled with the low temperatures of winter, facilitates the economic utilization of reserves during the many months typical of most diapauses. Though many insects store additional lipid prior to the entry into diapause, our review of the literature indicates that this is not always the case. We provide evidence that interactions between nutrient storage and metabolism can influence the decision to enter diapause and determine how long to remain in diapause. In addition, the energy reserves expended during diapause have a profound effect on post-diapause fitness. Though the physiological and biochemical mechanisms that regulate nutrient homeostasis prior to and during diapause remain poorly known, we propose several mechanisms that have the potential to contribute to diapause-associated nutrient homeostasis. Potential players include insulin signaling, neuropeptide F, cGMP-kinase, AMP-activated protein kinase, and adipokinetic hormone.
DOI:10.1016/j.jinsphys.2008.12.013URLPMID:19200436 [本文引用: 1]
Metabolic depression is a highly conserved feature of insect diapause, and an increase in metabolism is a reliable indicator of diapause termination and the initiation of post-diapause development. The trajectory of metabolic rate following diapause termination can guide the identification of important physiological and developmental landmarks during this developmental transition, yet quantitative descriptions of these trajectories are relatively rare. Here we track changes in metabolic rate from diapause through diapause termination and pharate adult development in Rhagoletis pomonella (Diptera: Tephritidae), a univoltine tephritid fly that diapauses in the pupal stage. Using respirometric monitoring we show that diapause termination and subsequent pharate adult development is characterized by a biphasic increase in metabolic rate. Respiration rate initially increases logistically, reaching a plateau that is followed by a final, exponential increase terminating in adult eclosion. We develop a non-linear model describing this pattern with easily interpretable landmarks, and we map visible landmarks of morphogenesis onto the trajectory. The bulk of visible morphogenesis in pharate adult development occurs relatively late in the trajectory during the final, exponential phase, that matches the U-shaped trajectory of non-diapause individuals. These results mirror a qualitative description of the same trajectories of diapause and non-diapause flesh flies ( Sarcophaga). Overall, our results suggest that (1) the course of diapause breakage and post-diapause development may be evolutionarily conserved in higher flies with pupal diapause, and (2) the biphasic trajectory is distinct from a more continuous trajectory observed during direct development.
[本文引用: 5]
DOI:10.7150/ijbs.9438URLPMID:25285037 [本文引用: 3]
Bactrocera minaxis a major citrus pest distributed in China, Bhutan and India. The long pupal diapause duration of this fly is a major bottleneck for artificial rearing and underlying mechanisms remain unknown. Genetic information onB. minaxtranscriptome and gene expression profiles are needed to understand its pupal diapause. High-throughput RNA-seq technology was used to characterize theB. minaxtranscriptome and to identify differentially expressed genes during pupal diapause development. A total number of 52,519,948 reads were generated and assembled into 47,217 unigenes. 26,843 unigenes matched to proteins in the NCBI database using the BLAST search. Four digital gene expression (DGE) libraries were constructed for pupae at early diapause, late diapause, post-diapause and diapause terminated developmental status. 4,355 unigenes showing the differences expressed across four libraries revealed major shifts in cellular functions of cell proliferation, protein processing and export, metabolism and stress response in pupal diapause. When diapause was terminated by 20-hydroxyecdysone (20E), many genes involved in ribosome and metabolism were differentially expressed which may mediate diapause transition. The gene sets involved in protein and energy metabolisms varied throughout early-, late- and post-diapause. A total of 15 genes were selected to verify the DGE results through quantitative real-time PCR (qRT-PCR); qRT-PCR expression levels strongly correlated with the DGE data. The results provided the extensive sequence resources available forB. minaxand increased our knowledge on its pupal diapause development and they shed new light on the possible mechanisms involved in pupal diapause in this species.
DOI:10.1007/s00360-007-0172-5URLPMID:17576567 [本文引用: 4]
Flesh flies can enhance their cold hardiness by entering a photoperiod-induced pupal diapause or by a temperature-induced rapid cold-hardening process. To determine whether the same or different metabolites are involved in these two responses, derivatized polar extracts from flesh flies subjected to these treatments were examined using gas chromatography–mass spectrophotometry (GC–MS). This metabolomic approach demonstrated that levels of metabolites involved in glycolysis (glycerol, glucose, alanine, pyruvate) were elevated by both treatments. Metabolites elevated uniquely in response to rapid cold-hardening include glutamine, cystathionine, sorbitol, and urea while levels of β-alanine, ornithine, trehalose, and mannose levels were reduced. Rapid cold-hardening also uniquely perturbed the urea cycle. In addition to the elevated metabolites shared with rapid cold-hardening, leucine concentrations were uniquely elevated during diapause while levels of a number of other amino acids were reduced. Pools of two aerobic metabolic intermediates, fumarate and citrate, were reduced during diapause, indicating a reduction of Krebs cycle activity. Principal component analysis demonstrated that rapid cold-hardening and diapause are metabolically distinct from their untreated, non-diapausing counterparts. We discuss the possible contribution of each altered metabolite in enhancing the overall cold hardiness of the organism, as well as the efficacy of GC–MS metabolomics for investigating insect physiological systems.
DOI:10.1186/1471-2164-14-751PMID:24180224 [本文引用: 4]
Background Diapause is programmed developmental arrest coupled with the depression of metabolic activity and the enhancement of stress resistance. Pupal diapause is induced by environmental signals and is prepared during the prediapause phase. In the cotton bollworm, Helicoverpa armigera , the prediapause phase, which contains two sub-phases, diapause induction and preparation, occurs in the larval stage. Here, we performed parallel proteomic and metabolomic analyses on H. armigera larval hemolymph during the prediapause phase. Results By two-dimensional electrophoresis, 37 proteins were shown to be differentially expressed in diapause-destined larvae. Of these proteins, 28 were successfully identified by MALDI -TOF/TOF mass spectrometry. Moreover, a total of 22 altered metabolites were found in diapause-destined larval hemolymph by GC-MS analysis, and the levels of 17 metabolites were elevated and 5 were decreased. Conclusions The proteins and metabolites with significantly altered levels play different roles in diapause-destined larvae, including diapause induction, metabolic storage, immune response, stress tolerance, and others. Because hemolymph circulates through the whole body of an insect, these differences found in diapause-destined larvae most likely correspond to upstream endocrine signals and would further influence other organ/tissue activities to determine the insect fact: diapause or development.
DOI:10.1073/pnas.1707281114URLPMID:28720705 [本文引用: 1]
Abstract Insects often overcome unfavorable seasons in a hormonally regulated state of diapause during which their activity ceases, development is arrested, metabolic rate is suppressed, and tolerance of environmental stress is bolstered. Diapausing insects pass through a stereotypic succession of eco-physiological phases termed "diapause development." The phasing is varied in the literature, and the whole concept is sometimes criticized as being too artificial. Here we present the results of transcriptional profiling using custom microarrays representing 1,042 genes in the drosophilid fly, Chymomyza costata Fully grown, third-instar larvae programmed for diapause by a photoperiodic (short-day) signal were assayed as they traversed the diapause developmental program. When analyzing the gradual dynamics in the transcriptomic profile, we could readily distinguish distinct diapause developmental phases associated with induction/initiation, maintenance, cold acclimation, and termination by cold or by photoperiodic signal. Accordingly, each phase is characterized by a specific pattern of gene expression, supporting the physiological relevance of the concept of diapause phasing. Further, we have dissected in greater detail the changes in transcript levels of elements of several signaling pathways considered critical for diapause regulation. The phase of diapause termination is associated with enhanced transcript levels in several positive elements stimulating direct development (the 20-hydroxyecdysone pathway: Ecr , Shd , Broad ; the Wnt pathway: basket , c-jun ) that are countered by up-regulation in some negative elements (the insulin-signaling pathway: Ilp8 , PI3k , Akt ; the target of rapamycin pathway: Tsc2 and 4EBP ; the Wnt pathway: shaggy ). We speculate such up-regulations may represent the early steps linked to termination of diapause programming.
DOI:10.1038/srep11402URLPMID:4650673 [本文引用: 1]
Low temperature induces diapause in locusts. However, the physiological processes and initiation mechanism of diapause are not well understood. To understand the molecular basis of diapause, mics analyses were performed to examine the differences between diapause and non-diapause eggs at both transcriptional and translational levels. Results indicated that a total of 62,241 mRNAs and 212 proteins were differentially expressed. Among them, 116 transcripts had concurrent transcription and translation profiles. Up-regulated genes related to diapause included glutathiones-S-transferase et al., and down-regulated genes including juvenile hormone esterase-like protein et al. KEGG analysis mapped 7,243 and 99 differentially expressed genes and proteins, to 83 and 25 pathways, respectively. Correlation enriched pathways indicated that there were nine identical pathways related to diapause. Gene Ontology analysis placed these genes and proteins into three categories, and a higher proportion of genes related to metabolism was up-regulated than down-regulated. Furthermore, three up-regulated pathways were linked to cryoprotection. This study demonstrates the applicability of high-throughput omics tools to identify molecules linked to diapause in the locust. In addition, it reveals cellular metabolism in diapause eggs is more active than in non-diapause eggs, and up-regulated enzymes may play roles in cryoprotection and storing energy for diapause and post-diapause stages.
DOI:10.1021/pr200796aURLPMID:22149145 [本文引用: 2]
Diapause is a developmental arrest that allows an organism to survive unfavorable environmental conditions and is induced by environmental signals at a certain sensitive developmental stage. In Helicoverpa armigera, the larval brain receives the environmental signals for diapause induction and then regulates diapause entry at the pupal stage. Here, combined proteomic and metabolomic differential display analysis was performed on the H. armigera larval brain. Using two-dimensional electrophoresis, it was found that 22 proteins were increased and 27 proteins were decreased in the diapause-destined larval brain, 37 of which were successfully identified by MALDI-TOF/TOF mass spectrometry. RT-PCR and Western blot analyses showed that the expression levels of the differentially expressed proteins were consistent with the 2-DE results. Furthermore, a total of 49 metabolites were identified in the larval brain by GC-MS analysis, including 4 metabolites at high concentrations and 14 metabolites at low concentrations. The results gave us a clue to understand the governing molecular events of the prediapause phase. Those differences that exist in the induction phase of diapause-destined individuals are probably relevant to a special memory mechanism for photoperiodic information storage, and those differences that exist in the preparation phase are likely to regulate accumulation of specific energy reserves in diapause-destined individuals.
DOI:10.1016/j.trac.2007.12.001URL [本文引用: 1]
DOI:10.3321/j.issn:0253-3820.2009.01.029URLMagsci [本文引用: 1]
代谢组学研究的是生物体系受到内在和外在因素刺激产生的内源性代谢变化,可以对那些能描述代谢循环情况的关键化合物进行定性和定量分析。近几年来,代谢组学已经成为生命科学领域一个重要的、有价值的工具,并在不断创新的分析技术推动下稳步发展。虽然代谢组学本身还存在一些不足,但许多研究者以解决问题为出发点,提出了一些新的研究策略、方法和技术。代谢组学发展呈现出整合一体化,定量化和标准化的趋势。本文对代谢组学的概况,现在的发展情况和未来的趋势进行综述。
DOI:10.3321/j.issn:0253-3820.2009.01.029URLMagsci [本文引用: 1]
代谢组学研究的是生物体系受到内在和外在因素刺激产生的内源性代谢变化,可以对那些能描述代谢循环情况的关键化合物进行定性和定量分析。近几年来,代谢组学已经成为生命科学领域一个重要的、有价值的工具,并在不断创新的分析技术推动下稳步发展。虽然代谢组学本身还存在一些不足,但许多研究者以解决问题为出发点,提出了一些新的研究策略、方法和技术。代谢组学发展呈现出整合一体化,定量化和标准化的趋势。本文对代谢组学的概况,现在的发展情况和未来的趋势进行综述。
DOI:10.1603/EC13539URLPMID:25195444 [本文引用: 2]
The univoltine Bactrocera minax (Enderlein) is a devastating pest of citrus in Nepal, India, Bhutan, and China. To better understand the overwintering pupal diapause, we investigated the juvenile hormone III (JH III) and ecdysteroid 20-hydroxyecdysone (20E) titers across the developmental stages, and the effects of juvenile hormone analog (JHA) and 20E treatments on adult emergence patterns. The results showed that both JH III and 20E levels fluctuated from the late larval stage to the late pupal stage. JHA and 20E treatments at the late larval stage had marginal effect on the adult emergence patterns, with slightly faster adult eclosion and higher eclosion rate. Similarly, JHA treatment at the early pupal stage did not affect the adult emergence. However, 20E treatment at the early pupal stage remarkably shortened the duration of diapause, but lowered the eclosion rate, especially with relatively high dose. These findings demonstrated that 20E treatment at the early pupal stage is a suitable method to hasten the diapause termination of B. minax, which will contribute to the rearing of B. minax in the laboratory as needed for routine experiments.
DOI:10.1038/srep27440URLPMID:27273028 [本文引用: 1]
Abstract The Chinese citrus fruit fly, Bactrocera minax, is an economically important pest of citrus. It exhibits pupal diapause from November to May to combat harsh environmental conditions. Such a long pupal diapause is a barrier for laboratory rearing and development of control strategies against this pest. In the present study, 20-hydroxyecdysone (20E) was used to break pupal diapause of B. minax by topical application. After diapause termination by 20E treated, the pupal ontogenetic processes were observed along the temporal trajectory. The pupal response time to 20E was estimated by detecting the relative expression of 20E responsive genes at different times after 20E-treatment. Results revealed that 20E could effectively terminate the pupal diapause in a dose-dependent manner and significantly shorten the time for 50% adult emergence (Et50). 20E response genes, including ecr, broad and foxo, were up-regulated within 72h, indicating these genes are involved in pupal metamorphosis and diapause termination processes. Morphological changes showed the pupal metamorphosis began ~7 days after 20E-treatment at 2209000900°C. This study does not only pave the way for artificial rearing in the laboratory through manipulating of pupal diapause termination, but also deepens our understanding of the underlying pupal diapause termination mechanism of B. minax.
[本文引用: 1]
DOI:10.1360/jos180001URL [本文引用: 1]
This paper proposes a Petri net extended with price information——Price Petri Net (PPN).Firstly we as- sociate a price with a transition,and give the semantics for PPN in terms of prieed transition systems.Then we propose a method to make teachability analysis for PPN model and discuss the minimum-cost reachability problem.Finally we use PPN to establish the'cost model for a business process.We can draw a conclusion:incorporating price information in Petri net and applying PPN to the domain of business process management are feasible.
[本文引用: 2]
DOI:10.1371/journal.pone.0099948PMID:4057385 [本文引用: 2]
Abstract A strategy known as diapause (developmental arrest) has evolved in insects to increase their survival rate under harsh environmental conditions. Diapause causes a dramatic reduction in the metabolic rate and drastically extends lifespan. However, little is known about the mechanisms underlying the metabolic changes involved. Using gas chromatography-mass spectrometry, we compared the changes in the metabolite levels in the brain and hemolymph of nondiapause- and diapause-destined cotton bollworm, Helicoverpa armigera, during the initiation, maintenance, and termination of pupal diapause. A total of 55 metabolites in the hemolymph and 52 metabolites in the brain were detected. Of these metabolites, 21 and 12 metabolite levels were altered in the diapause pupal hemolymph and brain, respectively. During diapause initiation and maintenance, the number of metabolites with increased levels in the hemolymph of the diapausing pupae is far greater than the number in the nondiapause pupae. These increased metabolites function as an energy source, metabolic intermediates, and cryoprotectants. The number of metabolites with decreased levels in the brain of diapausing pupae is far greater than the number in the nondiapause pupae. Low metabolite levels are likely to directly or indirectly repress the brain metabolic activity. During diapause termination, most of the metabolite levels in the hemolymph of the diapausing pupae rapidly decrease because they function as energy and metabolic sources that promote pupa-adult development. In conclusion, the metabolites with altered levels in the hemolymph and brain serve as energy and metabolic resources and help to maintain a low brain metabolic activity during diapause.
DOI:10.1073/pnas.1212879109URLPMID:22912402 [本文引用: 1]
Developmental arrest, a critical component of the life cycle in animals as diverse as nematodes (dauer state), insects (diapause), and vertebrates (hibernation), results in dramatic depression of the metabolic rate and a profound extension in longevity. Although many details of the hormonal systems controlling developmental arrest are well-known, we know little about the interactions between metabolic events and the hormones controlling the arrested state. Here, we show that diapause is regulated by an interplay between blood-borne metabolites and regulatory centers within the brain. Gene expression in the fat body, the insect equivalent of the liver, is strongly suppressed during diapause, resulting in low levels of tricarboxylic acid (TCA) intermediates circulating within the blood, and at diapause termination, the fat body becomes activated, releasing an abundance of TCA intermediates that act on the brain to stimulate synthesis of regulatory peptides that prompt production of the insect growth hormone ecdysone. This model is supported by our success in breaking diapause by injecting a mixture of TCA intermediates and upstream metabolites. The results underscore the importance of cross-talk between the brain and fat body as a regulator of diapause and suggest that the TCA cycle may be a checkpoint for regulating different forms of animal dormancy.
DOI:10.3969/j.issn.2095-039X.2014.05.004URLMagsci [本文引用: 1]
为分析七星瓢虫滞育相关蛋白质的类别和功能，在滞育调控及滞育后生物学研究的基础上，应用双向电泳、质谱分析（ESI-QUAD-TOF）、生物信息学等蛋白质组学方法，对表达量存在2倍及以上差异且差异显著的蛋白点进行鉴定。应用Mascot软件进行数据库检索，根据数据的匹配程度鉴定蛋白质。所得蛋白质中，参与三羧酸循环的两种关键酶呈现差异性表达，其中苹果酸脱氢酶（gi|212508346）呈上调表达，异柠檬酸脱氢酶（gi|21392222）则呈下调表达。苹果酸脱氢酶的增加，推测与滞育状态下的生理需求相关：一方面与昆虫对NAD的合成和利用有关，另一方面或是七星瓢虫应对滞育环境条件的一种应激方式，与正常个体的代谢通路相比，滞育个体开启了另外的代谢通路以适应环境条件的改变。异柠檬酸脱氢酶作为三羧酸循环中起关键调节作用的限速酶，在滞育个体中下调表达，或反映了三羧酸循环整体速率的降低，表现在滞育七星瓢虫体内维持了低水平的能量代谢。本文的研究结果，为从蛋白质水平深入揭示七星瓢虫滞育调控及其机理提供一定的参考。
DOI:10.3969/j.issn.2095-039X.2014.05.004URLMagsci [本文引用: 1]
为分析七星瓢虫滞育相关蛋白质的类别和功能，在滞育调控及滞育后生物学研究的基础上，应用双向电泳、质谱分析（ESI-QUAD-TOF）、生物信息学等蛋白质组学方法，对表达量存在2倍及以上差异且差异显著的蛋白点进行鉴定。应用Mascot软件进行数据库检索，根据数据的匹配程度鉴定蛋白质。所得蛋白质中，参与三羧酸循环的两种关键酶呈现差异性表达，其中苹果酸脱氢酶（gi|212508346）呈上调表达，异柠檬酸脱氢酶（gi|21392222）则呈下调表达。苹果酸脱氢酶的增加，推测与滞育状态下的生理需求相关：一方面与昆虫对NAD的合成和利用有关，另一方面或是七星瓢虫应对滞育环境条件的一种应激方式，与正常个体的代谢通路相比，滞育个体开启了另外的代谢通路以适应环境条件的改变。异柠檬酸脱氢酶作为三羧酸循环中起关键调节作用的限速酶，在滞育个体中下调表达，或反映了三羧酸循环整体速率的降低，表现在滞育七星瓢虫体内维持了低水平的能量代谢。本文的研究结果，为从蛋白质水平深入揭示七星瓢虫滞育调控及其机理提供一定的参考。
[本文引用: 1]
[本文引用: 1]
DOI:10.3864/j.issn.0578-1752.2018.04.009URL [本文引用: 1]
海藻糖为一种非还原性糖,广泛存在于细菌、藻类、真菌、植物和无脊椎动物中。海藻糖被称为昆虫"血糖",源于该糖为昆虫血淋巴中的主要糖类物质,在昆虫生长、发育、蜕皮等正常生理活动中有着重要的作用。昆虫的海藻糖先由海藻糖合成酶(trehalose-6-phosphate synthase,TPS)生成海藻糖-6-磷酸,然后通过海藻糖磷酸脂酶(trehalose-6-phosphate phosphatase,TPP)最终合成海藻糖,在能量需求时通过海藻糖酶(trehalase,TRE)降解为葡萄糖,用于能量供给。几丁质为昆虫表皮、中肠围食膜和气管系统的主要组成成分,昆虫在发育过程中需要蜕掉旧表皮,形成新的表皮,该过程一直是害虫控制的重要靶标途径。海藻糖酶为昆虫几丁质合成途径的第一个酶,在几丁质合成通路中有着重要的功能。那么海藻糖代谢又是如何调控几丁质合成途径来控制昆虫的蜕皮及几丁质代谢的呢?随着国内外海藻糖代谢相关基因功能研究的深入开展,研究结果表明昆虫海藻糖供给在几丁质合成中具有重要的作用,海藻糖酶分为可溶性和膜结合型两类,可溶性TRE和TPS在不同昆虫种类中具有多个同源基因,表明昆虫海藻糖代谢进化途径多样化。其次,海藻糖代谢直接调控几丁质合成途径,不论是海藻糖合成酶还是海藻糖酶基因的低表达,均能控制海藻糖使其供给不平衡,从而导致几丁质合成途径受阻,特别是几丁质合成酶基因表达降低而造成几丁质含量下降,该调控作用可进一步引起昆虫蜕皮困难、翅发育畸形等,甚至大量死亡。再次,海藻糖酶抑制剂能够抑制可溶性和膜结合型两类海藻糖酶活性、引起几丁质合成通路相关基因及几丁质酶基因表达的显著下降,导致几丁质含量显著降低,产生高比例的昆虫个体死亡。这些结果充分表明,一旦昆虫海藻糖代谢的供给平衡被打破,会直接影响到昆虫的几丁质合成乃至昆虫的蜕皮与发育过程。本文综述前人在海藻糖代谢调控几丁质合成方面的最新研究成果,为将来开发和利用海藻糖酶抑制剂及海藻糖合成酶抑制剂等绿色农药防治害虫提供理论依据。
DOI:10.3864/j.issn.0578-1752.2018.04.009URL [本文引用: 1]
海藻糖为一种非还原性糖,广泛存在于细菌、藻类、真菌、植物和无脊椎动物中。海藻糖被称为昆虫"血糖",源于该糖为昆虫血淋巴中的主要糖类物质,在昆虫生长、发育、蜕皮等正常生理活动中有着重要的作用。昆虫的海藻糖先由海藻糖合成酶(trehalose-6-phosphate synthase,TPS)生成海藻糖-6-磷酸,然后通过海藻糖磷酸脂酶(trehalose-6-phosphate phosphatase,TPP)最终合成海藻糖,在能量需求时通过海藻糖酶(trehalase,TRE)降解为葡萄糖,用于能量供给。几丁质为昆虫表皮、中肠围食膜和气管系统的主要组成成分,昆虫在发育过程中需要蜕掉旧表皮,形成新的表皮,该过程一直是害虫控制的重要靶标途径。海藻糖酶为昆虫几丁质合成途径的第一个酶,在几丁质合成通路中有着重要的功能。那么海藻糖代谢又是如何调控几丁质合成途径来控制昆虫的蜕皮及几丁质代谢的呢?随着国内外海藻糖代谢相关基因功能研究的深入开展,研究结果表明昆虫海藻糖供给在几丁质合成中具有重要的作用,海藻糖酶分为可溶性和膜结合型两类,可溶性TRE和TPS在不同昆虫种类中具有多个同源基因,表明昆虫海藻糖代谢进化途径多样化。其次,海藻糖代谢直接调控几丁质合成途径,不论是海藻糖合成酶还是海藻糖酶基因的低表达,均能控制海藻糖使其供给不平衡,从而导致几丁质合成途径受阻,特别是几丁质合成酶基因表达降低而造成几丁质含量下降,该调控作用可进一步引起昆虫蜕皮困难、翅发育畸形等,甚至大量死亡。再次,海藻糖酶抑制剂能够抑制可溶性和膜结合型两类海藻糖酶活性、引起几丁质合成通路相关基因及几丁质酶基因表达的显著下降,导致几丁质含量显著降低,产生高比例的昆虫个体死亡。这些结果充分表明,一旦昆虫海藻糖代谢的供给平衡被打破,会直接影响到昆虫的几丁质合成乃至昆虫的蜕皮与发育过程。本文综述前人在海藻糖代谢调控几丁质合成方面的最新研究成果,为将来开发和利用海藻糖酶抑制剂及海藻糖合成酶抑制剂等绿色农药防治害虫提供理论依据。
Delia antiqua(Diptera: Anthomyiidae).
DOI:10.1016/j.gene.2015.03.070URLPMID:25841989 [本文引用: 1]
61TPS, TPP and TRE involving in trehalose metabolism were cloned and characterized.61Their enzymatic activities were determined in diapausing Dalia antiqua pupae.61They share a similar expression pattern in summer and winter diapausing pupae.61Glucose and trehalose contents are different between both diapausing pupae.61Trehalose's function was suggested in cold hardiness and desiccation resistance.
DOI:10.1016/0022-1910(94)00124-YURL [本文引用: 1]
The cabbage root fly, Delia radicum L., overwinters as a phanerocephalic pupa (PCP) in a diapause state and its development may be temporarily stopped at the PCP stage even by high summer temperatures. Cold acclimation (5 days at 5 °C) of non-diapausing PCP resulted in an increased cold hardiness (survival of exposure to 61 15 °C for 1 or 20 days) and was associated with an increase of trehalose content (from 6.6 to 10.0 μg/mg wet weight). Non-diapausing pupae in a high-temperature quiescence had a higher osmotic pressure of haemolymph (623 mOsm) and a higher glucose content (3.4 μg/mg) in comparison to normal non-diapausing pupae (550 mOsm, 1.4 μg/mg). Diapausing PCP at the start of diapause showed cold hardiness levels equal to non-diapausing PCP. Their cold hardiness rapidly increased during the first 10–30 days of diapause development attaining a maximum between days 30 and 50 followed by an obvious decrease. Osmotic pressure increased from about 550 mOsm at the start of diapause to 860 mOsm by day 80 and then dropped slowly. Trehalose increased from 4.7 to 11.2 μg/mg during the first 80 days and then declined too. Glucose showed transitional increase of concentration during the first 20 days of diapause development followed by an abrupt drop until about day 50 and repeated the increase toward the diapause termination. Myo-inositol level increased slowly during diapause development showing a plateau after 80 days. Supercooling capacity of the PCP stage was similar irrespective of the physiological state or age of the pupa.
DOI:10.1371/journal.pone.0061745URLPMID:3629207 [本文引用: 1]
Background The codling moth (Cydia pomonella) is a major insect pest of apples worldwide. Fully grown last instar larvae overwinter in diapause state. Their overwintering strategies and physiological principles of cold tolerance have been insufficiently studied. No elaborate analysis of overwintering physiology is available for European populations. Principal Findings We observed that codling moth larvae of a Central European population prefer to overwinter in the microhabitat of litter layer near the base of trees. Reliance on extensive supercooling, or freeze-avoidance, appears as their major strategy for survival of the winter cold. The supercooling point decreases from approximately 6115.3°C during summer to 6126.3°C during winter. Seasonal extension of supercooling capacity is assisted by partial dehydration, increasing osmolality of body fluids, and the accumulation of a complex mixture of winter specific metabolites. Glycogen and glutamine reserves are depleted, while fructose, alanine and some other sugars, polyols and free amino acids are accumulated during winter. The concentrations of trehalose and proline remain high and relatively constant throughout the season, and may contribute to the stabilization of proteins and membranes at subzero temperatures. In addition to supercooling, overwintering larvae acquire considerable capacity to survive at subzero temperatures, down to 6115°C, even in partially frozen state. Conclusion Our detailed laboratory analysis of cold tolerance, and whole-winter survival assays in semi-natural conditions, suggest that the average winter cold does not represent a major threat for codling moth populations. More than 83% of larvae survived over winter in the field and pupated in spring irrespective of the overwintering microhabitat (cold-exposed tree trunk or temperature-buffered litter layer).
[本文引用: 1]
[本文引用: 1]
DOI:10.1016/j.jinsphys.2012.04.003URLPMID:22542495 [本文引用: 1]
The pistachio fruit hull borer, Arimania comaroffi (Ragonot) (Lepidoptera: Pyralidae), is a key pest of pistachio orchards in Iran. This pest passes the winter as diapausing pupae. In this study, some physiological changes in relation to environmental temperature were investigated in field collected pupae. The relationship between supercooling point, cold hardiness and physiological changes of a wild population of this pest was also investigated. The glycogen content decreased with decrease in environmental temperature. Decrease in glycogen content was proportional to increase in total body sugar, trehalose, myo-inositol and sorbitol contents. In January with mean ambient temperature of 5.4°C, glycogen (5mg/g fresh body weight) content was at the lowest level whereas total body sugar (10.3mg/g fresh body weight), trehalose (8.6mg/g fresh body weight), myo-inositol (5.3mg/g fresh body weight) and sorbitol (2.6mg/g fresh body weight) were at the highest levels. Total body sugar, trehalose, myo-inositol and sorbitol contents increased as mean temperature decreased from 22.7°C in October to 5.4°C in January. Total body lipid decreased during overwintering and reached to the lowest level at the end of March. Supercooling points were decreased from October to January and reached to the lowest level (6116°C) in January with minimum ambient temperature of 6110°C. Survival at low temperature after 24h was also greatest in January with 72% survival at 6110°C, 39% survival at 6115°C and 0% survival at 6120°C. Increase in temperature from February onward, was proportional with increase in supercooling points and decrease in survival rate. Regardless of sampling date, all pupae died after 24h at 6120°C, whereas none pupae died after 24h at 615°C. This indicates that this insect is freeze-intolerant.
[本文引用: 1]
DOI:10.1016/S0965-1748(97)00068-4URLPMID:9474782 [本文引用: 1]
An electrochemically active compound was detected in the larvae of Aedes aegypti mosquitoes and progressive accumulation of this compound was observed during larval development. The compound was purified from mosquito larvae using various chromatographic techniques and spectral analysis of the purified compound resulted in its identification as xanthurenic acid. Production of xanthurenic acid results from the transamination of 3-hydroxykynuorenine, and analysis of the biochemical pathway in xanthurenic acid production revealed the presence of a particular transaminase that has a much higher specific activity to 3-hydroxykynurenine than to kynurenine in the mosquito larvae. Concentration of xanthurenic acid is closely related to the level of this transaminase activity. Results suggest that this particular transaminase plays an important role in regulating the level of 3-hydroxykynurenine in the mosquito, A. aegypti during larval development.
DOI:10.1023/A:1004115718597URLPMID:11294610 [本文引用: 1]
The white, scarlet , and brown genes of Drosophila melanogaster encode ABC transporters involved with the uptake and storage of metabolic precursors to the red and brown eye colour pigments. It has generally been assumed that these proteins are localised in the plasma membrane and transport precursor molecules from the heamolymph into the eye pigment cells. However, the immuno-electron microscopy experiments in this study reveal that the White and Scarlet proteins are located in the membranes of pigment granules within pigment cells and retinula cells of the compound eye. No evidence of their presence in the plasma membrane was observed. This result suggests that, rather than tranporting tryptophan into the cell across the plasma membrane, the White/Scarlet complex transports a metabolic intermediate (such as 3-hydroxy kynurenine) from the cytoplasm into the pigment granules. Other functional implications of this new finding are discussed.
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