Physiological response of eight Mediterranean maquis species to low air temperatures during winter

We analyzed the physiological response of the Mediterranean evergreen species (Arbutus unedo L., Cistus incanus L., Erica arborea L., Erica multiflora L., Phillyrea latifolia L., Pistacia lentiscus L., Quercus ilex L., and Rosmarinus officinalis L.) to winter low air temperatures. In occasion of two cold events, in February 2005 (T _min = 1.8 °C), and January 2006 (T _min = 3.1 °C and minimum T _air = −0.40 °C during the nights preceding the measurements), R. officinalis, C. incanus, and E. multiflora had the highest net photosynthetic rate (P _N) decrease (73 %, mean value) with respect to the winter P _N maximum, followed by A. unedo (62 %), P. latifolia and P. lentiscus (54 %, mean value), E. arborea (49 %), and Q. ilex (44 %). Among the considered species, Q. ilex was able to maintain P _N near the maximum for 150 min during the day, A. unedo, P. lentiscus, E. arborea, P. latifolia, E. multiflora, and R. officinalis for 60 min, and C. incanus for 30 min. The calculated mean winter daily P _N ranged from 7.9±0.6 (Q. ilex) to 2.8±0.5 (R. officinalis) μmol(CO₂) m⁻² s⁻¹. During the study period, chlorophyll (Chl) content decreased by 36 % on an average in the two cold events, and the carotenoid (Car) to Chl ratio increased by 133 % in Q. ilex, having the highest value in January 2006. Principal component analysis underlined the highest cold resistance of Q. ilex by high P _N and high Car/Chl ratio. On the contrary, R. officinalis and C. incanus had the lowest cold resistance by the highest P _N decrease and the lowest Car/Chl (C. incanus). Thus, winter stress could be an additional limitation to Mediterranean evergreen species production, and the capacity of the species to maintain P _N near 90–100 % during winter is determinant for biomass accumulation.     

A common strategy of cold hardiness in ants of the genus <Emphasis Type="Italic">Myrmica</Emphasis> (Formicidae,Hymenoptera) in Northeast Asia

The biotopic distribution, nest structure, wintering conditions, and cold hardiness of four ecologically contrasting ant species (Myrmica angulinodis, M. kamtschatica, M. bicolor, and M. transsibirica) are considered. The cold hardiness of these species is typical of the genus: the supercooling points vary from −28 to −31°C; cold hardiness levels (LT50%) are higher by 5°C. At this level of cold hardiness, ants can be practically ubiquitous across the whole Hypoarctic (Berman et al., 2007). However, the above Myrmica species are strictly segregated (M. kamtschatica occurs in moss bogs, M. angulinodis and M. transsibirica, on dry and warm south slopes, and M. bicolor, in sandy-gravel floodplains), probably due to different requirements for weather conditions in summer and depth of ground thawing. At present, the excess cold hardiness common to the species in question (exceeding the nest temperature by 5–10°C in different years) is not adaptive and may be considered as preadaptive. It could have been acquired during ancient cold epochs or inherited by the genus as a concomitant result of adaptation not to low temperatures but, for instance, to aridity. These Myrmica species do not undergo selection for resistance to negative temperatures since their current level of cold hardiness is excessive, considering the possible wintering temperatures.     

柴胡幼苗越冬抗寒性及其相关生理指标筛选

以北柴胡、地产柴胡、三岛柴胡的根为试验材料,以早春植株成活率为越冬抗寒性指标,测定越冬期不同柴胡品种根系生理生化特征,进行柴胡抗寒性综合评价,并对主要抗寒生理指标进行通径分析和相关性分析,以探讨冬季自然低温条件下柴胡生理特性与抗寒性的关系,筛选适合柴胡越冬抗寒性鉴定的生理指标。结果表明:(1)北柴胡、地产柴胡、三岛柴胡返青期的返青率依次为80%、50%、10%,且各品种间差异显著。(2)3个品种柴胡越冬期根系可溶性糖含量、游离脯氨酸含量、可溶性蛋白含量、根系活力与返青率大小顺序一致,其中北柴胡根系游离脯氨酸平均含量分别为地产柴胡和三岛柴胡的2.44倍和4.49倍,且可溶性蛋白含量显著高于地产柴胡、三岛柴胡,但3个品种柴胡的相对电导率、丙二醛含量水平与返青率大小顺序相反。(3)自然越冬过程中,各柴胡品种的根系活力均呈下降趋势,且以三岛柴胡根系活力下降幅度最大,达80%,而北柴胡根系活力下降幅度最小为52.04%。(4)越冬期3个柴胡品种的越冬抗寒性综合排序为:北柴胡地产柴胡三岛柴胡;柴胡根中脯氨酸含量、可溶性蛋白含量与综合评价值呈极显著正相关关系,相对电导率与综合评价值呈显著负相关关系;生理指标决策系数的大小顺序依次为脯氨酸含量可溶性蛋白含量电导率。研究表明,越冬期柴胡根系游离脯氨酸含量是影响其抗寒性的主要影响因素,其含量可作为评价柴胡抗寒性的指标。     

The Effect of Local Cooling of Cucumber and Wheat Seedlings on Various Kinds of Stress Resistance of Their Leaves and Roots

Shoots and roots of wheat (Triticum aestivum L., cold-resistant species) and cucumber (Cucumis sativus L., cold-sensitive species) were chilled at 2°C or 10°C, respectively, for 7 h. The changes in cold, heat, and salt resistance in treated leaf and root cells were recorded. Local cooling of the leaf resulted in an increase of its cold and salt tolerance, but its heat tolerance remained unchanged. At the same time, cold tolerance of the root slightly increased as a result of local cooling, but its heat and salt tolerance decreased. Cooling of the shoot did not affect the cold and heat tolerance of root cells but caused a decrease in their salt tolerance. Finally, in the leaf maintained at a normal temperature, there was an increase in all kinds of stress resistance as a result of root cooling. We discuss the possibility of an unspecific change in stress resistance caused by metabolic shifts. These shifts are induced by a signal, which is transmitted inside the plant into plant organs located at a considerable distance from the chilled ones.     

Cytogenetic investigations in Cucurbitaceae I. Interspecific hybridization in Luffa

In two interspecific hybrids of Luffa, i.e., L. acutangula x L. graveolens and L. echinata x L. graveolens, various types of chromosome associations, from univalents to quadrivalents were observed. The hydrid L. echinata x L. graveolens showed a much higher frequency of univalents than the other hybrid, but the pairing data of both indicate partial homology among most of the chromosomes of the three species. The study of these F₁ hybrids has been utilised for establishing the phylogenetic relationship among the various species and to trace the trend of evolution in the genus Luffa. The two wild species, L. graveolens and L. echinata, are closely related, whereas, L. graveolens is more distantly related to L. acutangula than to L. echinata. It has also been concluded that dioecism in L. echinata is a derived feature, established after diverging from the monoecious prime species L. graveolens.This research has been financed in part by a grant made by the United States Department of Agriculture under PL 480.     

Overexpression of ethylene response factor <Emphasis Type="Italic">TERF2</Emphasis> confers cold tolerance in rice seedlings

Rice (Oryza sativa L.) is a warm-season plant exposed to various stresses. Low temperature is an important factor limiting extension of rice cultivation areas and productivity. Previously, we have demonstrated that tomato ERF protein TERF2 enhances freezing tolerance of transgenic tobacco and tomato plants. Herein, we report that overexpression of TERF2 enhances transgenic rice tolerance to cold without affecting growth or agronomic traits. Physiological assays revealed that TERF2 could not only increase accumulation of osmotic substances and chlorophyll, but also reduce reactive oxygen species (ROS) and malondialdehyde (MDA) content and decrease electrolyte leakage in rice under cold stress. Further analysis of gene expression showed that TERF2 could activate expression of cold-related genes, including OsMyb, OsICE1, OsCDPK7, OsSODB, OsFer1, OsTrx23, and OsLti6, in transgenic rice plants under natural condition or cold stress. Thus, our findings demonstrated that TERF2 modulated expression of stress-related genes and a series of physiological adjustments under cold stress, indicating that TERF2 might have important regulatory roles in response to abiotic stress in rice and possess potential utility in improving crop cold tolerance.     

沙棘PEPCK基因的克隆及表达研究

以沙棘为研究对象,通过RACE、基因克隆和定量PCR技术,克隆并分析沙棘磷酸烯醇式丙酮酸羧激酶基因(PEPCK)的序列信息,以及进化树构建和氨基酸的二级结构分析,比较NaCl胁迫后PEPCK基因的表达变化,为揭示PEPCK基因在沙棘抗性方面的功能研究奠定基础。结果显示:(1)成功克隆了沙棘PEPCK基因,该基因开放阅读框(ORF)为2 001bp,编码666个氨基酸。(2)序列分析表明,沙棘PEPCK与蓖麻、黄顶菊、核桃等序列一致性高达85%以上;进化上沙棘PEPCK基因与蓖麻和狭叶羽扇豆较近,与烟草、拟南芥进化关系较远。(3)在300mmol·L~(-1) NaCl盐胁迫下,沙棘PEPCK基因表达在胁迫组中显著上升,胁迫后7d时为未处理的2.5倍,15d时达到未处理的3.2倍。研究表明,沙棘PEPCK基因在沙棘适应外源盐胁迫的过程中可能发挥了重要作用。     

CARBOHYDRATE MOBILIZATION AND MOVEMENT IN ALPINE PLANTS

Starch retention and disappearance from leaves and carbon movement under various temperatures were studied in two alpine species, Oxyria digyna (L.) Hill and Phleum alpinum L., and in two low-elevation species, Helianthus annuus L. and Elymus canadensis L. The alpine species exhibited starch disappearance from the leaf following cool night temperatures, whereas starch retention was noted under similar conditions for the low-elevation species. The alpine species, Oxyria, exhibited the highest rates of starch disappearance from the leaf under cool temperatures as well as the highest carbohydrate translocation under cool temperatures. The low-elevation species had low rates of starch disappearance and carbohydrate translocation under low temperatures, but exhibited relatively higher rates with an increase in temperature. Such a mechanism whereby alpine species can maintain relatively high rates of translocation under cold temperature represents a major form of physiological adaptation to the short, cool, growing season in the alpine tundra.     

Seasonal incidence of and antibiotic resistance among Aeromonas species isolated from domestic wastewater before and after treatment in stabilization ponds

The efficiency of stabilization pond treatment of domestic wastewater in removing culturable cells of motile Aeromonas and its influence on the incidence of resistance to seven antibiotics were investigated in this study. Removal efficiency was higher (P < 0.001) in the warm months (98.8%) than in the cold months (97%). Among the 264 isolates, 163 were Aeromonas caviae, 24 were A. hydrophila, and 54 were A. sobria. Twenty-three isolates could not be identified to the species level. In the influent, A. caviae dominated in both cold and warm months. In the water samples originating from the influent, A. sobria was present at higher percentages in the warm period. All the isolates were resistant to amoxicillin and most of them (73%) exhibited resistance to cephalothin. Of the three species tested, A. sobria was more susceptible to antibiotics than either A. caviae or A. hydrophila. The most striking difference among the species was seen in resistance to cephalothin. There were 91 % of A. caviae strains and 96% of A. hydrophila isolates that were resistant to cephalothin. However, only 9% of A. sobria strains exhibited resistance to this drug. The high incidence of resistance in raw sewage was connected with a high proportion of A. caviae, whereas in the water samples collected from the effluent during the warm months, a high proportion of A. sobria decreased the total amount of multiple-resistant bacteria. Results demonstrated the need for identification to the species level.Offprint requests to: L. Hassani.     

Phylloquinone (Vitamin K1) Levels in Leaves of Plant Species Differing in Susceptibility to 2,4-Dichlorophenoxyacetic Acid

Plant species differing in susceptibility to 2,4-dichlorophenoxyacetic acid (2,4-D) were examined as to the level of phylloquinone (K) in the leaves. The K level was found to be considerably higher in several plants resistant or moderately resistant to 2,4-D (Aegopodium podagraria L., Galium mollugo L., Lamium album L., Matricaria chamomilla L., Oxalis acetosella L., Stellaria media (L.) Vill., Viola arvensis Murr.) than in Chenopodium album L. and Sinapsis arvensis L. which are susceptible to 2,4-D. Capsella bursa-pastoris (L.) Med. and Plantago major L. (both moderately susceptible) and a few resistant or moderately resistant species (Agropyron repens (L.) PB., Anthriscus silvestris (L.) Hoffm., Triticum aestivum L.) had intermediate K levels. Cirsium arvense (L.) Scop. (susceptible) had a relatively high level of K. It is possible that a high K level in the plant can be of importance in the resistance to 2,4-D.     

Temperature drop, dry matter accumulation and cold resistance of young cucumber plants

The study aimed to provide information on the involvement of plant cold resistance in a whole cucumber (Cucumis sativus L.) plant response to temperature drop treatments. The main objective was to explain the effects of a temperature drop on dry matter production and plant morphogenesis. The experimental design included temperature drops to 12°C for 0, 1, 2, 4 and 6 hours at the beginning, in the middle and at the end of the night. The overall average temperature for all treatments was 20°C. A 2-hour temperature drop resulted in the most significant reduction in plant dry weight and leaf petiole length, but the highest degree of cold resistance. With the extension of a temperature drop duration to 4--6 hours, dry matter production was similar to that in the control plants, leaf petiole length increased, but not up to the control level and cold resistance was reduced, but was still higher compared to that of the control plants. Changes in dry matter production, leaf petiole length and cold resistance, depending on the duration of the temperature drop, were interrelated and their patterns varied little with the timing (position effect) of the temperature drop. It is concluded that plant cold resistance mechanisms play an important role in a whole plant response to temperature drop treatments. Thus, plant response to temperature drop treatments can be considered as a response to stress.     

基于rbcL和matK序列探讨马鞭草科部分植物的系统学位置

为探究适用于马鞭草科植物的DNA条形码及该类群的系统分类关系,对豆腐柴(Premna microphylla)的叶绿体基因ycf6-psbM、trnV-atpE、rbcL、trnL-F、psbM-trnD、atpB-rbcL、trnC-ycf6、trnH-psbA、rpl36-infA-rps8和核基因ITS序列进行了PCR扩增和测序,结果表明仅rbcL、trnl-F、trnH-psbA序列的PCR扩增以及测序效果较好,而ITS不能得到明显的扩增条带,ycf6-psbM不能成功测序,其它序列存在有部分双峰或噪值高等问题。根据DNA条形码标准,rbcL序列是所有测试条码中相对最适合的。应用rbcL和matK序列对马鞭草科(Verbenaceae)豆腐柴属、牡荆属(Vitex L.)、马鞭草属(Verbena L.)和大青属(Clerodendrum L.)等4属与唇形科宝盖草属(Lamium L.)、水苏属(Stachys L.)、鼠尾草属(Salvia L.)和香科科属(Teucrium L.)等4属的分类和系统发育关系进行分析,以紫草科Lithospermum multiflorum L.为外群,最大简约法对2个片段的单独和联合矩阵分别构建系统发育树。豆腐柴属和大青属应从马鞭草科划入唇形科,马鞭草属仍归于马鞭草科,而牡荆属的系统学位置还需更多的证据。     

Role of the Rice BAHD Acyltransferase Gene OsAt10 in Plant Cold Stress Tolerance

The rice (Oryza sativa L.) BAHD acyltransferase gene OsAt10 affects growth and metabolism of cells and regulates cell response to environmental stress. However, influence of the OsAt10 gene on low-temperature stress tolerance has not been evaluated in plant cells. Here, cell suspension cultures of plant species Arabidopsis (Arabidopsis thaliana L.), cotton (Gossypium hirsutum L.), white pine (Pinus strobus L.), and rice (Oryza sativa L.) were used to generate transgenic cell lines via Agrobacterium tumefaciens-mediated genetic transformation to examine the effects of OsAt10 on cold stress tolerance. OsAt10 transgenic cell lines of A. thaliana, G. hirsutum, P. strobus, and O. sativa were confirmed by molecular analyses including Southern blotting ND northern blotting, following by physiological and biochemical analyses under cold stress. The experimental results demonstrated that growth rate, cell viability, lipid peroxidation, ion leakage, antioxidative enzyme activity, polyamines level, and cell morphology were changed in transgenic cells under cold stress, compared to the controls. In transgenic A. thaliana cells, overexpression of the OsAt10 gene increases expression of polyamines biosynthesis genes under cold stress. In transgenic A. thaliana plants, overexpression of the OsAt10 gene increased cold stress tolerance by regulating expression of stress marker genes, TBARS content, ion leakage level, antioxidant enzymes activity, and polyamines content, indicating that the OsAt10 gene could be economically important for improving low-temperature stress tolerance in plants.

     

Components of CO<Subscript>2</Subscript> exchange in leaves of C<Subscript>3</Subscript> species with different ability of starch accumulation

Using a radiogasometric method the rates of photorespiratory and respiratory decarboxylations of primary and stored photosynthates in the leaves of two groups of C₃ species, differing in the ability of starch accumulation, were determined. One group included starch-accumulating (SA) species with rates of starch synthesis on the average 38 % the rate of photosynthesis [Solanum tuberosum L., Arabidopsis thaliana (L.) Heynh, Helianthus annuus L., and Plantago lanceolata L.]. The second group represented starch-deficient (SD) species with rates of starch synthesis less than 8 % the rate of photosynthesis (Secale cereale L., Triticum aestivum L., Hordeum vulgare L., and Poa trivialis L.). In SA species the rate of respiration in the dark was significantly higher than in SD species. No differences were found in the rates of photosynthesis, photorespiration, and respiration under irradiation. Thus, the degree of inhibition of respiration by irradiation was in SA species higher than in SD species. It is concluded that starch does not provide substrates for respiratory and photorespiratory decarboxylations in irradiated photosynthesizing leaves.     

Identification of two loci for resistance to clubroot (<Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis> Woronin) in <Emphasis Type="Italic">Brassica rapa</Emphasis> L.

In an analysis of 114 F₂ individuals from a cross between clubroot-resistant and susceptible lines of Brassica rapa L., 'G004' and 'Hakusai Chukanbohon Nou 7' (A9709), respectively, we identified two loci, Crr1 and Crr2, for clubroot (caused by Plasmodiophora brassicae Woronin) resistance. Each locus segregated independently among the F₂ population, indicating that the loci reside on a different region of chromosomes or on different chromosomes. Genetic analysis showed that each locus had little effect on clubroot resistance by itself, indicating that these two loci are complementary for clubroot resistance. The resistance to clubroot was much stronger when both loci were homozygous for resistant alleles than when they were heterozygous. These results indicate that clubroot resistance in B. rapa is under oligogenic control and at least two loci are necessary for resistance.Communicated by H.C. Becker     

Proteome expression patterns in the stress tolerant evergreen <Emphasis Type="Italic">Ammopiptanthus nanus</Emphasis> under conditions of extreme cold

Low temperature is one of the important environmental changes that affect plant growth. The cold resistance capabilities of evergreen plants are the result of long-term adaptation to extreme environmental conditions. To investigate the responses of Ammopiptanthus nanus, a rare stress-tolerant evergreen plant, to extreme cold stress, we analyzed the proteome expression patterns of stressed plants; this is the first study to report these patterns for A. nanus. We collected adult A. nanus leaves under two conditions of cold stress: extreme cold (−29°C) and relatively less extreme cold (−5°C). Total crude proteins were extracted from leaf blades, separated by two-dimensional gel electrophoresis, and stained with Coomassie brilliant blue. Of the 500 protein spots detected in each of the samples, eight of the spots that exhibited clear changes under the different conditions were identified by MALDI-TOF analyses. Our results suggest that cold stress-related proteins may play diverse roles in the resistance to multiple environmental stresses.     

LIMITS TO THE SOUTHERN BORDER OF DROSOPHILA SERRATA: COLD RESISTANCE,HERITABLE VARIATION,AND TRADE-OFFS

There are a number of evolutionary hypotheses about why species distributions are limited, but very little empirical information to test them. We present data examining whether the southern distribution of Drosophila serrata is limited by cold responses. Species comparisons were undertaken for cold resistance, development time, and viability at 15°C and 25°C for D. serrata and other species with a more southerly distribution (D. melanogaster, D. simulans, and D. immigrans). Relative to the other species, D. serrata had a long development time at both temperatures and a low level of cold resistance. Using isofemale lines collected in different seasons, central and marginal populations were compared for cold resistance, as well as development time and viability at 14°C. The border population had a relatively higher resistance to cold shock in postwinter collections, but there was no population differentiation for prewinter collections or for the other traits. The presence of variation among isofemale lines within the border populations suggests that genetic variation as measured in the laboratory is unlikely to limit range expansion. Population cages were used in the field to determine if D. serrata persisted over winter at borders. Although all cages yielded adult offspring at northern sites, only a few produced offspring at or just south of the border. In contrast, all cages with D. simulans produced adult offspring, suggesting that climatic factors limited D. serrata numbers. Offspring from surviving adults showed a phenotypic trade-off between fecundity and cold resistance. Comparisons of the cold resistance of field males and females with their laboratory-reared offspring provided evidence for heritable variation in field-reared flies. Overall, the results suggest that cold stress is important in limiting the southern distribution of D. serrata, but it seems unlikely that a lack of genetic variation restricts range expansion.     

Genomic analyses provide insights into the polyploidization-driven herbicide adaptation in Leptochloa weeds

Polyploidy confers a selective advantage under stress conditions; however, whether polyploidization mediates enhanced herbicide adaptation remains largely unknown. Tetraploid Leptochloa chinensis is a notorious weed in the rice ecosystem, causing severe yield loss in rice. In China, L. chinensis has only one sister species, the diploid L. panicea, whose damage is rarely reported. To gain insights into the effects of polyploidization on herbicide adaptation, we first assembled a high-quality genome of L. panicea and identified genome structure variations with L. chinensis. Moreover, we identified herbicide-resistance genes specifically expanded in L. chinensis, which may confer a greater herbicide adaptability in L. chinensis. Analysis of gene retention and loss showed that five herbicide target-site genes and several herbicide nontarget-site resistance gene families were retained during polyploidization. Notably, we identified three pairs of polyploidization-retained genes including LcABCC8, LcCYP76C1 and LcCYP76C4 that may enhance herbicide resistance. More importantly, we found that both copies of LcCYP76C4 were under herbicide selection during the spread of L. chinensis in China. Furthermore, we identified another gene potentially involved in herbicide resistance, LcCYP709B2, which is also retained during polyploidization and under selection. This study provides insights into the genomic basis of the enhanced herbicide adaptability of Leptochloa weeds during polyploidization and provides guidance for the precise and efficient control of polyploidy weeds.     

The cold resistance of Macaronesian Sempervivoideae

Summary Cold resistance of twenty-seven species of Macaronesian Sempervivoideae uniformly cultivated under cool moderate but not hardening conditions was measured. The resistance limits of all the tested species ranged between-4 and-10°C. Cold stress response was principially different: Cold resistance of about half of the tested species was due to freezing point lowering. This response type, avoidance of freezing, in which any ice formation in the leaves leads to injury, was found in the most cold resistant species (Aeonium spathulatum, several Aichryson species). The other species developed tolerance to freezing, thus resembling the behaviour of the hardy Eurasian Sempervivum species. Several Aeonium and Monanthes species resist to reasonable lower temperatures than normally occur in their natural habitats. The species-specific differences in resisting cold stress may originate from different abilities to tolerate cellular freeze dehydration. The Sempervivum alliance illustrates well the two evolutionary strategies of cold tolerance: Avoidance mechanisms, like lowering the osmotic potential, are typical for species colonizing higher altitudes with moderate frosts. For species extending their distribution area into higher latitudes with more severe frosts, however, freezing tolerance is necessary.Cordially dedicated to Prof. Dr. Michael Evenari