细菌冰核提高印度谷螟过冷却点的研究

印度谷螟（Ｐｌｏｄｉａｉｎｔｅｒｐｕｎｃｔｅｌｌａ）是一种不耐结冰的昆虫,在冬季它通过降低过冷却点以避免结冰。现已查明,冰核活性细菌能显著提高植物的过冷却点,导致许多作物在较高的温度下发生霜冻害。本文也评明细菌冰核能显著提高印度谷螟幼虫的过冷却点。对照的平均过冷却点是－１７．６℃;分别用０．１ｇ和１ｇ细菌冰核与１ｋｇ面粉混合后进行处理,平均过冷却点分别比对照提高了１２．８℃和１３．６℃。研究结果支持这样的观点：细菌冰核有可能成为一种在冬季使用的、杀灭不耐结冰害虫的生物制剂。     

过冷却和昆虫的耐寒性

过冷却（超冷,supercooling）现象,由Fahrenheit（1842）发现,即为在一定的条件下,当温度低于水的冰点时,水仍能保持液体状态的现象。1过却点及测定方法Beclune6Cw（1901）利用电热测温,首次发现动物在结冰时的过冷却现象。过冷却点是指在逐步冷却过程中自发地结冰产生时的温度。过冷却的能力是指液体的熔点与过冷却点倍CPsllpereoolingpoint）间的差异,通常用过冷却点来表示。当自发的结冰发生时,通过测定潜热的消政。能够准确地测定SCP。特别是陆生节肢动物的超冷极限,可以方便地以过冷却点的形式测到。过冷却点测定最常用的…     

亚洲柑橘木虱和柚喀木虱过冷却点和体液结冰点测定

亚洲柑橘木虱Diaphorina citri Kuwayama和柚喀木虱Cacopsylla citrisuga YangLi均为柑桔上的重要害虫,前者为黄龙病的主要媒介昆虫,后者也已证实为黄龙病菌的携带者。本文对这两种木虱各龄若虫和成虫的过冷却点和体液结冰点进行了测定,结果表明,柑橘木虱过冷却点和冰点均以1龄若虫最低,平均值分别为-26.32℃和-26.22℃;成虫的过冷却点和冰点最高,分别为-19.60℃和-18.71℃;其它虫态过冷却点和冰点由低到高依次为2龄若虫、4龄若虫、5龄若虫、3龄若虫。柚喀木虱过冷却点和冰点也均以1龄若虫最低,平均值分别为-25.30℃和-25.08℃;5龄若虫的过冷却点和冰点最高,平均分别为-19.50℃和-17.69℃;其它虫态过冷却点和冰点由低到高依次为2龄若虫、3龄若虫、4龄若虫、成虫。两种木虱的比较结果表明,各发育阶段的表现不一致,柚喀木虱的3龄若虫和成虫过冷却点显著低于柑桔木虱,而4龄、5龄若虫则显著高于柑桔木虱;3龄若虫体液结冰点也显著低于柑桔木虱,但1龄、4龄却显著高于柑桔木虱。柑桔木虱雌雄成虫的过冷却点和体液结冰点差异不显著。研究结果表明两种木虱的过冷却点、体液结冰点都较低,因而可能具有较强的耐寒能力。     

丝裂霉素C对Erwinia herbicola表达及运转冰核活性蛋白方式的影响

利用添加丝裂霉素C(MMC)于不同培养基中,并在不同培养条件下对草生欧文氏菌10025A(E．herbicola 10025A)诱导培养,首次证实该菌表达冰核活性蛋白的活性及运转分泌冰核活性蛋白方式是不同。研究结果显示,MMC的加入可以提高细菌培养物的冰核活性,该现象可以解释为MMC对E．herbicola的作用能够激活细胞SOS应急系统的反应,诱导细胞合成部分参与修复DNA损伤的酶和蛋白因子,达到对E．herbicola诱导表达冰核蛋白通过高尔基体形成小液泡,向膜外分泌的方式,同时对E．herbicola细胞分裂后的形态也发生明显的影响。这对研究细胞在恶劣环境的生存机制以及获取该条件下的蛋白具有重要的意义,对低温生物的研究也将产生积极地影响。     

我国冰核活性细菌的优势种类调查与研究

１９８６￣１９９４年,从国内１７个省、市、自治区的６８种植物上分离到了２５０株冰核活性细菌,经鉴定分别属于３个属的１７个种或致病变种。其中出现最多的是菠萝欧文氏菌,共１３３株,占总数的５３．２％,其次是丁香假单胞菌群,共７０株,占２８％,其它种类的冰核活性细菌共４７株,仅占１８．８％。因此,我国冰核活性细菌的优势菌种是Ｅ．ａｎａｎａｓ,其次是Ｐ．ｓｙｒｉｎｇａｅ ｐｖｓ。在低纬度的南方地区中Ｅ．ａ     

一种便携式测定昆虫过冷却点的方法

本文介绍了热敏电阻 +万用电表测定昆虫过冷却点的新方法。通过实验比较发现使用热敏电阻法测定昆虫过冷却点较之热电偶法有如下优点 :(1)测量灵敏度和精确度高 ;(2 )测定昆虫范围广 ;(3)装置体积小 ,不用交流电 ,携带方便 ;(4 )价格较低。     

冷冻液温和季节对鼠尾过冷点的影响

为研究动物对寒冷的适应性,将鼠尾置于冷液浸冻,发现在一定条件下鼠尾组织可发生过冷现象。实验表明,鼠尾组织的过冷点和冷冻液温有关,同一季节冷冻液温越低过冷点越高;而不同季节相同冷冻条件下,冬季鼠尾组织的过冷点明显低于春季。 动物肢体组织的过冷特性是动物的抗寒冷特性,它和组织自身的物理化学性质有关。理沦证明,过冷度(△T)和表面张力(O)、摩尔质量(M),冰点(Ti)、密度(p)、摩尔凝固热(△H)及冰胚临界半径(rk)有关,其关系式为△T=26MTi/p△Hrk.     

细菌冰核基因的应用研究

细菌冰核基因的应用研究已成为生物冰核领域的研究热点,研究涉及细菌细胞表面展示、促冻杀虫、报告基因、病原微生物高敏检测、作物抗寒育种等多个领域,显示良好应用前景。通过对国外在该方面的研究现状的综述,和我们在冰核基因促冻杀虫研究方面重要进展的介绍,对今后我们拟开展的这一研究工作进行了展望。     

扶桑绵粉蚧的过冷却点和体液结冰点测定

本文测定了扶桑绵粉蚧Phenacoccus solenopsis Tinsley除卵期以外其它各虫态的过冷却点和结冰点.结果表明,该虫过冷却点以1龄若虫最低,为-24.02℃;雄虫预蛹次之,为-22.13℃;雄虫蛹、雄虫2龄若虫、雌虫3龄若虫、雄成虫、雌虫2龄若虫、雌成虫过冷却点逐渐升高,分别为-21.08℃、-20.25℃、-19.05℃、-18.42℃、-17.91℃、-16.89℃.体液结冰点也以1龄若虫最低,为-23.2℃;雄虫预蛹次之,为-19.09℃;雄虫蛹、雄虫2龄若虫、雌虫3龄若虫、雌虫2龄若虫、雌成虫、雄成虫体液结冰点逐渐升高,分别为-16.64%、-15.81℃、-13.92℃、-13.20℃、-12.85℃、-12.79℃.试验结果表明扶桑绵粉蚧过冷却点低,耐寒性较强,可能适宜在中国北部更广泛的区域生存.     

Experimental studies on the cold tolerance of Alaskozetes antarcticus

The cold tolerance mechanism of the Antarctic terrestrial mite Alaskozetes antarcticus (Michael) was investigated in cultured animals. Freezing is fatal in this species and winter survival occurs by means of supercooling, which is enhanced by the presence of glycerol in the body. There is an inverse, linear relationship between the concentration of glycerol and the supercooling point, which may be as low as ?30°C. Feeding detracts from supercooling ability by providing ice nucleators in the gut which initiate freezing at relatively high sub-zero temperatures. Experiments on the effects of various environmental factors showed that low temperature acclimation gave rise to increased glycerol concentrations and suppressed feeding, while desiccation also stimulated glycerol production. Photoperiod had no effect on cold tolerance in this species. The juvenile instars of A. antarcticus were found to possess a greater degree of low temperature tolerance than adults.     

Ice nucleation and antinucleation in nature

Plants and ectothermic animals use a variety of substances and mechanisms to survive exposure to subfreezing temperatures. Proteinaceous ice nucleators trigger freezing at high subzero temperatures, either to provide cold protection from released heat of fusion or to establish a protective extracellular freezing in freeze-tolerant species. Freeze-avoiding species increase their supercooling potential by removing ice nucleators and accumulating polyols. Terrestrial invertebrates and polar marine fish stabilize their supercooled state by means of noncolligatively acting antifreeze proteins. Some organisms also depress their body fluid melting point to ambient temperature by evaporation and/or solute accumulation.     

Change in overwintering mechanism of the Cucujid beetle, Cucujus clavipes

Overwintering larvae of the Cucujid beetle, Cucujus clavipes, were freeze tolerant, able to survive the freezing of their extracellular body fluids, during the winter of 1978–1979. These larvae had high levels of polyols (glycerol and sorbitol), thermal hysteresis proteins and haemolymph ice nucleators that prevented extensive supercooling (the supercooling points of the larvae were ? 10°C), thus preventing lethal intracellular ice formation. In contrast, C. clavipes larvae were freeze suspectible, died if frozen, during the winter of 1982–1983, but supercooled to ～ ? 30°C. The absence of the ice nucleators in the 1982–1983 larvae, obviously essential in the now freeze-susceptible insects, was the major detected difference in the larvae from the 2 years. However, experiments in which the larvae were artifically seeded at ? 10°C (the temperature at which the natural haemolymph ice nucleators produced spontaneous nucleation in the 1978–1979 freeze tolerant larvae) demonstrated that the absence of the ice nucleators was not the critical factor, or at least not the only critical factor, responsible for the loss of freeze tolerance in the 1982–1983 larvae. The lower lethal temperatures for the larvae were approximately the same during the 2 winters in spite of the change in overwintering strategy.     

Cryoprotective mechanisms in subtidally cultivated and intertidal blue mussels (mytilus edulis) from the Magdalen Islands, Québec

Centrifuged extracts from subtidally cultivated and intertidal blue mussels have higher supercooling points (up to -5.5°C) in winter than in summer (up to -12.5°C). The concentration of nucleators (as estimated by the dilution factor) is greater in winter than in summer in both groups. The nucleator concentration in the extracts of winter mussels is one to two orders of magnitude higher than that in the haemolymph of Norwegian mussels. Although these extracts show spilule-like growth of ice crystals, they caused no thermal hystersis. The seasonal variation of these cryoprotective mechanisms is similar for intertidal and cultivated mussels. However, in the spring, cultivated mussels have a lower supercooling point and a lower concentration of nucleators than their intertidal counterparts. This suggests that cultivated mussels decrease their cryoprotective capacity earlier than intertidal mussels.     

越北腹露蝗卵的过冷却特性

利用热电偶法测定越北腹露蝗FruhstorferiolatonkinensisWill卵的过冷却点。结果表明,越北腹露蝗卵的过冷却点随季节变化而变动。卵的平均过冷却点从2004年9月初(刚产的卵)的-16.0℃显著下降到2005年1月的-24.7℃,随后卵的过冷却点显著增加到2005年3月的-10.7℃。滞育卵的平均过冷却点(-20.1℃)低于滞育后的平均过冷却点(-12.4℃)。滞育卵在5℃下适应30d后平均过冷却点显著减小到-25.9℃,而在30℃下适应30d后平均过冷却点显著增加到-8.2℃。     

Maintenance of the supercooled state in the gut fluid of overwintering pyrochroid beetle larvae, Dendroides canadensis : role of ice nucleators and antifreeze proteins

  The effect of gut fluid ice nucleators and antifreeze proteins on maintenance of supercooling was explored in fire-colored beetle larvae, Dendroides canadensis, via seasonal monitoring of supercooling points, antifreeze protein activity and ice nucleator activity of gut fluid and/or larvae. During cold hardening in the field, freeze-avoiding larvae evacuated their guts and depressed larval supercooling points. Analysis of gut fluid indicated supercooling points and ice nucleator activity decreased, whereas antifreeze protein activity increased as winter approached. Suspensions of bacteria isolated from guts of feeding larvae collected in spring/summer had higher supercooling points than those from midwinter-collected non-feeding larvae, suggesting bacterial ice nucleators are removed from midwinter gut fluid. The ice nucleation active bacterium Pseudomonas fluorescens was isolated from gut fluid of feeding larvae but was absent in winter. When mixed with purified D.␣canadensis hemolymph antifreeze proteins (structurally similar and/or identical to those in gut fluid), the cumulative ice nucleus spectra of P. fluorescens suspensions were shifted to lower temperatures indicating an inhibitory effect on the bacteria's ice-nucleating phenotype. By extending larval supercooling capacity, both gut clearing and masking of bacterial ice nucleators by antifreeze proteins may contribute to overwintering survival in supercooled insects. Accepted: 8 August 1996     

Mesophyll freezing and effects of freeze dehydration visualized by simultaneous measurement of IDTA and differential imaging chlorophyll fluorescence

Infrared differential thermal analysis (IDTA) and differential imaging chlorophyll fluorescence (DIF) were employed simultaneously to study the two-dimensional pattern of ice propagation in leaves and mesophyll freeze dehydration as detected by a significant increase of basic chlorophyll fluorescence (F(0)). IDTA and DIF technique gave different insights into the freezing process of leaves that was highly species-specific. IDTA clearly visualized the freezing process consisting of an initial fast spread of ice throughout the vascular system followed by mesophyll freezing. While mesophyll freezing was homogeneously in Poa alpina, Rhododendron ferrugineum and Senecio incanus as determined by IDTA, DIF showed a distinct pattern only in S. incanus, with the leaf tips being affected earlier. In Cinnamomum camphora, a mottled freezing pattern of small mesophyll compartments was observed by both methods. In IDTA images, a random pattern predominated, while in DIF images, compartments closer to lower order veins were affected earlier. The increase of F(0) following mesophyll freezing started after a species-specific time lag of up to 26 min. The start of the F(0) increase and its slope were significantly enhanced at lower temperatures, which suggest a higher strain on mesophyll protoplasts when freezing occurs at lower temperatures.     

Gut Colonization by an Ice Nucleation Active Bacterium,Erwinia(Pantoea)ananasReduces the Cold Hardiness of Mulberry Pyralid Larvae

To evaluate the suitability of using ice nucleation active (INA) bacteria for the biological control of insect pests, the supercooling point (SCP) of larvae of mulberry pyralid,Glyphodes duplicalis,and silkworm,Bombyx mori,ingesting INA strains ofErwinia(Pantoea)ananasandPseudomonas syringaewas determined. Mean SCP of the guts of silkworm larvae ingesting INA strains ofE. ananasranged from −2.5 to −2.8°C, being 5°C higher than that in control treatments. Similarly, mean SCP of mulberry pyralid larvae ingesting INA strain ofE. ananas,which can grow well in the gut, was −4.7°C at 3 days after treatment, being 6.5°C higher than that in control treatments. On the other hand, mean SCP of the larvae-ingesting INA strain ofP. syringae,which cannot grow in the gut, was −9.0°C at 3 days after treatment, rising by only 2.5°C higher than that in the control treatments. In addition, more than 80% of the larvae of mulberry pyralid ingesting the INA strain ofE. ananasfroze and eventually died when exposed to −6°C for 18 h, while only 36% of the larvae ingesting the INA strain ofP. syringae,or approximately 20% of the control larvae, froze and died. Thus, the gut colonization by INA strains ofE. ananasreduced remarkably the cold hardiness of the insects. These findings suggest that INA strains ofE. ananascould be effective as a potential biological control agent of insect pests.