【作者】 李珏闻；

【导师】 骆有庆；

【作者基本信息】 北京林业大学 ， 森林保护， 2014， 博士

【摘要】 青杨脊虎天牛(Xylotrechus rusticus L.)近10年来一直被列为全国林业检疫性有害生物,是我国东北地区重要的林木钻蛀性害虫,主要危害杨属(Populus)、柳属(Salix)和榆属(Ulmus)的健康大径级树木,以幼虫蛀食主干,严重影响树木生长,还极易造成被害木主干风折,威胁路人生命及财产安全。目前,青杨脊虎天牛主要分布于我国高纬度地区,但其暴发区呈现出逐渐向北转移的趋势。一般而言,温度是限制昆虫种群地理分布的主要因素之一。因此,开展青杨脊虎天牛在极端温度胁迫下的响应研究,对其潜在分布区的预警和综合管理具有重要意义。本文在系统观察青杨脊虎天牛危害过程的基础上,通过对其温度胁迫耐受性、温度胁迫下的生理响应,以及蛀干坑道微生境温度变化规律的研究,揭示了该天牛对高、低温胁迫的耐受极限和主要耐受对策,并以此为主导,划分青杨脊虎天牛在我国的潜在适生区。主要结果如下：1.首次系统精细地观察了青杨脊虎天牛的危害特征。通过长期连续的大量观察和解剖,明确了该天牛在不同发育阶段的典型危害特征,即初孵幼虫群集式垂直于主干蛀食,在形成平均深度3.5cm的“刀砍”状危害特征后,中龄幼虫分散转移至韧皮部危害,但进入秋季,老熟幼虫会再次蛀食木质部,并在平均深度4.5cm处越冬、直至春末羽化。同时,以3-D手绘图系统展示了青杨脊虎天牛幼虫在蛀干坑道内的逐步危害过程,以及相应危害阶段树皮表面的典型特征照片。2.明确了青杨脊虎天牛成虫和幼虫的高温耐受极限,并初步揭示了其越夏幼虫高温胁迫下的生理响应能力。结果表明,造成大多数成虫休克、失去行为能力的临界高温区间(38℃-39.5℃)低于成虫高温胁迫2h后的半致死温度HLT50(44.87℃)说明临界高温并非成虫的高温致死上限；但临界高温的上限(41℃)与高温胁迫4h后的半致死温度HLT50(41.90℃)十分接近,因此当临界高温的胁迫时间延长至4h左右,则足以对50%成虫个体造成致死性伤害。另外,在高温胁迫24h后,幼虫的HLT50(43.0℃)比成虫的HLT50(37.79℃)高约5℃,说明幼虫比成虫具有更强的耐热性。由此推断,青杨脊虎天牛种群通过长期对环境条件的适应,以幼虫作为唯一越夏虫态,因此其幼虫在不同高温胁迫时间内的各致死温度(HLT50和HLT99)对其种群适生分布区的评价具有更合理的指导意义。另外,夏季幼虫在两组时间(4h和24h)内的高温胁迫条件下,35℃胁迫后的虫体蛋白质含量与对照组(25℃)相比均显著升高,说明在35℃下有大量热激蛋白产生,从而进行高温胁迫下的生理调节,而虫体含水率和脂肪率变化均不显著,说明水和脂肪能够在35℃进行较为正常的代谢作用,因此35℃是幼虫可承受、并可进行生理响应的胁迫高温；但当温度上升至40℃,虫体蛋白质含量和脂肪率显著降低、含水率显著升高,并且3个生理指标均维持各自近似水平至45℃,说明幼虫经40-C或更高温度胁迫后,将丧失耐热生理调节能力,即为不可进行生理响应的胁迫高温。3.明确了青杨脊虎天牛越冬幼虫的低温耐受极限,并初步揭示了其越冬幼虫的主要生理耐寒机制。结果表明,青杨脊虎天牛越冬幼虫在不同低温下,持续暴露24h(短期胁迫)和768h(长期胁迫)后的半致死温度LLT5o分别为-33.64-C和-30.08℃,均远低于越冬期内过冷却点(SCP)的最低值-14.7℃和平均值-11℃,说明越冬幼虫在体液结冰之后,依然能够长期维持存活状态,由此可推断青杨脊虎天牛属于传统耐寒对策分类中的耐结冰型昆虫。另外,在越冬期内幼虫SCP与环境温度之间呈现出相同的变化趋势,说明越冬幼虫SCP的高低,能够体现其个体耐寒性的强弱。通过越冬期幼虫生理生化指标的测定表明,a.越冬期内,甘油含量的变化趋势与SCP之间表现为显著负相关(P=0.033,R=0.907),由此推断越冬中期增加的甘油为越冬幼虫的主要抗寒物质；b.虫体糖原含量和脂肪含量均在越冬中期大幅度降低,并在越冬后期还原至越冬初期水平,因此推断其为越冬幼虫的主要能源物质,但仅糖原含量与SCP呈现出显著正相关(P=0.006,R=0.971)；c.越冬期幼虫体内的糖原与甘油之间(P=0.046,R=0.885)、糖原与甘露醇(P=0.012,R=0.954)之间存在相互转化关系；d.越冬期幼虫体内含水率变化不显著,推断幼虫可通过“节水”机制来提高虫体耐寒性；e.虫体蛋白质含量在越冬中期显著升高,说明越冬幼虫在冬季受到低温诱导后,其体内合成了与抗寒作用相关的功能性蛋白。4.测定并明确了幼虫坑道内温度在越冬期的变化规律及其特点,并揭示了坑道在越冬幼虫遭遇冬季低温时提供的保护模式。在2012年1月,应用微探头温度自动记录仪定时(1次/30min)测量杨树的模拟坑道内温度。结果表明,坑道内的温度变化呈现出与野外环境温度变化几乎相同的昼夜周期性,以及明显的时滞性；坑道内最低温记录值(-39.58℃)与坑道外(-39.88℃)相比仅高0.3℃,且坑道内1月平均温度仅比坑道外高0.227℃,差异均不显著；在相同时刻,南侧与北侧坑道内的温度差值在0.15℃-1.85℃之间,平均值为0.24℃,差异不显著。故以杨树为寄主的幼虫坑道对昆虫的保护作用并非因为坑道内温度单纯的、持续的比坑道外高,而是因为木材比热较空气大,从而使坑道内温度的升高或降低较坑道外缓和,进而为幼虫适应低温提供了保护性的缓冲作用,并通过这种自然驯化逐渐提高越冬幼虫的耐寒性。5.基本明确了青杨脊虎天牛在我国的高度适生区和中度适生区。经统计分析中国43个气象台站连续5年(2007-2012年)温度参数,应用Arcgis9.3软件,通过反距离权重法将温度差值为等温线图,并以青杨脊虎天牛幼虫分别在温度胁迫24h和32d后的致死温度(LLTso、LLT99、HLT50和HLT99)作为适生分布区划分的临界参考值,结果表明：我国东北大部、华北、西北和西南地区均属于青杨脊虎天牛的高度适生区；福建、浙江和江西全境,江苏、安徽、湖北、湖南和广东的大部分区域,以及东北北部、河南省东南部和内蒙古额济纳旗中部的较小区域,为该天牛的中度适生区；在仅以温度限制种群地理分布时,该天牛在我国不存在非适生区。更多还原

【Abstract】 The grey tiger longicorn beetle, Xylotrechus rusticus L., is one of the most severe wood bores in northeast China and is listed as one of the national forestry quarantine pests in recent10years. The beetles mainly damaged the healthy and mature trees of Populus, Salix and Ulmus. Woods growth were seriously affected after larvae fed on the trunk, for which reason the damaged trees can be easily blown down and the safety of local populace were critically threatened. At present, this pest mainly occurred in high latitudes areas in China, however, the geographic distribution of X. rusticus had expanded to higher latitudes because of global warming. Generally speaking, temperature is a vital factor that limit the geographic distribution of insect. So it’s important to study the extreme temperature tolerance of X. rusticus for early warnning about potential distribution and its integrated control. Thus, based on the detailed observation of damage features, the present study aimed to determine the tolerance to extreme temperatures and the relevant physiological variations, as well as the change regulations of larvae microhabitat temperature during overwintering, thereby characterizing its limiting of tolerance to extreme low and high temperatures and the tolerant strategies, then to assess the suitable geographic distribution and its potential adaptive area in China. The main results are as follows:1. The detailed damage characteristics of X. rusticus in different development stage were systematically observed for the first time. After eggs hatched, the newly larvae would bore the trunk in cluster and leave a damaging feature of a chopped scar with mean depth of3.5cm; till the middle stage, larvae started to feed on phloem; the mature larvae would damage the xylem again in autumn and overwinter in the trunk mean depth of4.5cm until to the emergence of adults. Based on all above, a three dimensional gallery features internal the trunk were drawn refered to different damage stages, and the photos of corresponding damage features on the bark surface were also presented. 2. Thermal tolerance of X. rusticus adults and larvae were defined, and the ability for physiological adjustment that larvae reponse to high temperature stress were also revealed. The results showed that the critical thermal maximum (CTMax) ranged from38℃to39.5℃, during which most adults would lost the ability for of action and then went into shock state. All the values of CTMax were lower than the high lethal temperature of HLT50(44.87℃) after adults under thermal stress for2h, which implied that CTMax was not the upper lethal temperature for the beetles. However, the highest value of CTMax (41℃) was approximately the same with the HLT50(41.90℃) that adults were exposed to high temperature for4h, which indicated that CTMax would be the lethal damage temperature resulting50%mortality of adults if the time of exposure to the CTMax prolonged to4h. In addition, the HLT50(43.0℃)of larvae exposed to high temperature for24h was higher than that of adults (HLT50,37.79℃), which indicated that larva had stronger thermal tolerance than adults. It showed that larva of X. rusticus was the only developed stage over summer because of the long-term natural selection that the species in habitat, so the lethal temperatures of larvae in different time duration was more meaningful for assessing the potential distribution of X. rusticus. In addition, the body protein content of the two groups were both increased significantly when oversummering larvae stressed under35℃for4h and24h, respectively. It implied that there was much heat shock protein synthesized in larvae bodies, in order to progress physiological adjustment response to high temperature stress at35℃. While the body water content and lipid content did not change significantly, in other words, the two physiological substances could metabolize regularly at high temperature of35℃. Therefore,35℃is the endurable high temperature that larva can be tolerant through physiological adjustment. However, when larvae were exposed to high temperature of40℃, all the three physiological substances changed significantly, and maintained the similar level as temperature rising to45℃, though the body protein content and lipid rate decreased whereas the water rate increased. It indicate that oversummering larvae would loss the ability of physiological adjustment when the temperature rise to40℃or more.3. The cold hardiness of overwintering X. rusticus larva was demonstrated, and the physiological and biochemical indices were preliminarily revealed. The lower lethal temperature (LLT50) was respectively-33.64℃and-30.08℃under24h (short-term stress) and768h (long-term stress) whenX. rusticus overwintering larvae were exposed to different low temperatures. And the two values were far lower than the average (-11℃) and minimum (-14.7℃) of SCP, which indicated that the larva could survive after their body fluid freeze for a long time. It implied that X. rusticus belong to freezing-tolerant insect according to the traditional cold hardiness strategy classification. In addition, the similar variation tendency were showed between SCP and the environment temperature during the overwintering period, which implied that SCP could be used to assess the cold hardiness variation tendency during a sampling period and the cold hardiness differences between individuals in a population. However, SCP was not considered as the lower limit of lethal temperature of this species. The determination of physiological and biochemical indices showed:a. glycerol play an important role as major cryoprotectant for the accumulation in mid-overwintering stage and showed a significantly negative correlation with SCP (P=0.033, R=0.907); b. glycogen and lipid were considered as the important energy substances, which decreased obviously in mid-overwintering period, but only glycogen correlated positively with (P=0.006R=0.971); c. transformations between glycogen and glycerol(.P=0.046, R=0.885), as well as glycogen and mannitol (P=0.012, R=0.954) were showing negative correlations; d. water content of larva body changed little during the whole overwintering period, which implied that "Water-saving mechanism" may have been conducted to improve the cold tolerance; e. the protein content of larva body increased significantly in mid-overwintering period. It seemed that functional protein about cold resistance could be synthesized in larva body, which may be triggered by the low temperature.4. The change regulation of temperature in larva gallery during overwintering period was determined and clarified. The internal gallery temperature of poplar during January2012was detected by a micro-detector Auto-temperature recorder (1time per30min). The results showed that temperature changed inside gallery during January presented a significant feature of circadian periodicity and time delay; the minimum temperature of internal gallery (-39.58℃) was only0.3℃higher than that of the external gallery, and the average temperature of internal gallery in January was only0.227℃higher than that of the external, differences of both were not significant; in addition, the temperature difference between northern and southern internal gallery was0.15℃-1.85℃, averagely0.24℃, so the protection provided by the gallery to the larva was depended on the larger specific heat capacity of wood than air rather than the temperature of internal gallery higher than that of the external, this features of gallery enabled a lower change ratio of temperature of internal gallery, providing a protective buffer role for larva, and through this nature accommodation, the larva could improve their cold hardiness gradually.5. Highly and moderately adaptive distribution of X. rusticus were determined. The lowest and highest monthly mean temperature, within5years from2007to2012, were collected from January and July respectively, which were analyzed by meteorological data from43meteorological stations among China. The results showed that the highly adaptive area is northern, northwestern, southwestern China, and most part of northeastern China. The moderately adaptive area is Fujian, Zhejiang and Jiangxi province, most part of Jiangsu, Anhui, Hubei and Guangdong province, north part of northeastern China, southeast part of Henan province, and central part of Ejina County. There is no disadaptive area in China if take temperature as the only population restrict factor.更多还原