Role of lutoid membrane transport and protein synthesis in the regeneration mechanism of latex in different <Emphasis Type="Italic">Hevea</Emphasis> clones

Natural rubber (cis-1,4 polyisoprene) is synthesised in the milky cytoplasm, the latex, of specialized cells called laticifers in the bark tissues of the rubber tree (Hevea brasiliensis). Regeneration mechanism of latex after each tapping (controlled wounding of the bark) was studied in relation to lutoid membrane enzymes and protein synthesis in twelve rubber clones with varying yield potentials during the peak rubber yielding season. High activity of membrane enzymes and better availability of biochemical energy [ATP] were observed in clones viz; RRII 105, RRIM 600, PB 260, RRII 422 and RRII 430. The highest protein biosynthetic capacity was noticed in clone PB 260 and RRIM 600. However, high ATP content, increased invertase activity and protein biosynthesis were observed in the medium yielding clone GT1 compared to clones with low rubber yield potential. Very low sugar content and increased invertase activity in the latex of clone PB 260 indicated intense latex metabolism with high protein turnover that implies fast recouping of the cellular metabolites lost during latex harvesting. Clone PB 217 was characterized by very high sucrose and low ATP concentration and ATPase activity in latex indicating slow metabolism and hence be suitable for inducing latex metabolism using ethylene stimulant. Low rubber yielding clones such as RRII 33 and RRII 38 were consistently recorded a high sucrose content but very low activity of membrane enzymes, reduced ATP concentration and low protein biosynthesis in latex. Among the recently released modern clones (RRII 400 series), latex regeneration capacity was higher in RRII 422 and RRII 430. The significance of lutoid membrane transport and protein synthesis is discussed in relation to general latex metabolism of these rubber clones. The outcome of this study would be helpful to design suitable latex harvesting systems and yield stimulation methods for optimizing latex production in each clone based on metabolic profiling.     

Influence of soil,plant and meteorological factors on water relations and yield inHevea brasiliensis

Influence of factors governing the soil-plantatmosphere system on components of water relations and yield was studied in two clones of rubber tree,Hevea brasiliensis, viz. RRII 105 and RRII 118. Clonal variations were evident in yield and yield components and associated physiological parameters in response to soil moisture status and meteorological factors. Observations made during different seasons indicatedvariations in yield are attributed to differences in plugging index and initial flow rates, to the major yield components and also variations in components of water relations as influenced by meteorological factors. Among the two clones, RRII 105 was found to be fairly drought tolerant compared to RRII 118. RRII 105 was found to respond well to dry weather through higher stomatal resistances, higher leaf water potentials, lowered transpirational water loss and lower relative transpiration ratios, while RRII 118 was susceptible to stress situations.     

SNP Identification from RNA Sequencing and Linkage Map Construction of Rubber Tree for Anchoring the Draft Genome

Hevea brasiliensis, or rubber tree, is an important crop species that accounts for the majority of natural latex production. The rubber tree nuclear genome consists of 18 chromosomes and is roughly 2.15 Gb. The current rubber tree reference genome assembly consists of 1,150,326 scaffolds ranging from 200 to 531,465 bp and totalling 1.1 Gb. Only 143 scaffolds, totalling 7.6 Mb, have been placed into linkage groups. We have performed RNA-seq on 6 varieties of rubber tree to identify SNPs and InDels and used this information to perform target sequence enrichment and high throughput sequencing to genotype a set of SNPs in 149 rubber tree offspring from a cross between RRIM 600 and RRII 105 rubber tree varieties. We used this information to generate a linkage map allowing for the anchoring of 24,424 contigs from 3,009 scaffolds, totalling 115 Mb or 10.4% of the published sequence, into 18 linkage groups. Each linkage group contains between 319 and 1367 SNPs, or 60 to 194 non-redundant marker positions, and ranges from 156 to 336 cM in length. This linkage map includes 20,143 of the 69,300 predicted genes from rubber tree and will be useful for mapping studies and improving the reference genome assembly.     

Seasonal suitability of three rubber tree clones to Calacarus heveae (Acari, Eriophyidae)

The suitability of rubber tree clones to Calacarus heveae was inferred from the life cycle, reproduction and survivorship of this mite. The assays were performed under controlled conditions with leaflets detached from 6-year-old plants. The development of 20 C. heveae individuals on each of the clones GT 1, PB 235 and RRIM 600 was analysed. This experiment was performed four times during periods when C. heveae was abundant in the field: (P1) November–December 2005, (P2) January–February, (P3) March–April and (P4) May–June 2006. Accordingly, the leaflets used in each assay represented the physiological condition of the host plant during each period. This approach allowed us to evaluate the seasonal suitability of rubber tree clones to C. heveae. We observed seasonal differences in the suitability of rubber tree clones to mite attack. The mites reared on the PB235 had a shorter development period, the highest egg production and highest survivorship. This evidence showed that the PB 235 was the most suitable of those tested. We also observed that the leaflets used in the assays during periods P2 and P3 were the most favourable for the development of C. heveae. This finding emphasises the seasonal suitability of rubber tree leaflets. On the other hand, GT 1 showed higher resistance against C. heveae than did RRIM 600 and PB 235, primarily during the period from November to February. This result indicated that use of the GT 1 clone to control the mite might represent an alternative for growers.     

Expression of the sucrose transporter 3 (<Emphasis Type="Italic">HbSUT3</Emphasis>) in rubber tree and its relation to latex yield

Sucrose is not only a precursor of rubber biosynthesis, but it also participates in other latex regeneration between repeated tapping. Sucrose transporter (HbSUT) genes play key roles in sinks and sources of the rubber biosynthesis pathway. HbSUT3 was dominant in expression in latex among the SUT member genes. Therefore, the expression level of HbSUT3 in latex is a potential indicator distinguishing between high- and low-yielding clones. The aim of this research was to assess the potential of this gene in selective breeding to improve latex yield, from the correlation of HbSUT3 expression with latex yield. Four high-yielding clones were sampled in this study and compared with the common RRIM 600 clone as the baseline, paired by the field. Among the putative full-length cDNAs of Hevea sucrose transporters available in the NCBI database, only HbSUT3 was detected. The HbSUT3 gene was overexpressed in the four rubber clones relative to the control, and the NK1 clone had the highest expression level. The expression level of HbSUT3 correlated positively with latex yield but negatively with the sucrose content of latex. Gene expression analysis of rubber seedlings indicated that the bark had higher expression of this gene than the leaves, and the levels correlated with latex yields of these clones. These data provided new candidate selection criteria for use in the early selection of high-yielding rubber clones, necessary for rapid cycle breeding programs.     

Novel bacterial endophytes from Hevea brasiliensis as biocontrol agent against Phytophthora leaf fall disease

Bacterial endophytes offer control against many diseases of crop plants as potential biocontrol agents. Antagonistic bacterial endophytes acting against Phytophthora meadii have been screened from leaf, petiole and root tissues of Hevea brasiliensis. Six bacterial endophytes could exhibit more than 50 % inhibition of P. meadii, among which EIL-2, from disease-free zones showed a maximum of 62.5 % inhibition. The isolate EIL-2 was characterized as Alcaligenes sp. and the other isolates were identified as Pseudomonas aeruginosa. 16S rDNA sequence analysis showed that there existed genetic variation among the five isolates of P. aeruginosa from different tissues of the plant indicating the tissue type adaptation of the isolates. Dual culture technique with endophyte EIL-2 completely arrested the growth of P. meadii when inoculated prior to pathogen. The bioassay with EIL-2 in H. brasiliensis clones, RRII 105 showed 43 % reduction of lesion size on infected leaves whereas in RRIM 600 it was only 30 %.     

云南保存橡胶树魏克汉种质资源亲子系谱分析

对“农业部景洪橡胶树种质资源圃”（云南景洪）保存的290份魏克汉种质资源进行了亲子系谱分析。结果表明,无性系GT1、PR107、PB86、RRIM600、云研277-5、PB5/51、IAN873的后代为210份,占72.41%。从表型看,魏克汉种质资源群体变异丰富,实际上遗传基础很窄。本文以GT1、PR107、PB86、RRIM600、云研277-5、PB5/51、IAN873为主线建立了亲子系谱图,并对部分种质资源的性状进行了分析,获得一些性状的传递遗传规律,供育种工作者参考。     

The Role of Lipids and Proteins in the Mechanism of Latex Vessel Plugging in Hevea brasiliensis

The phospholipid content of the bottom fraction of latex as well as the neutral lipids of rubber particles were determined in thirty-one clonal mother trees of RRII clones of Hevea brasiliensis Muell. Arg. Ratios between cationic proteins and anionic proteins in the B-sera, which is the sera contained in the lutoid particles in latex, obtained from clones RRIM-501, PB 6/9, RRII-105 and Tjir-l were determined by electrophoresis. The influence of these factors on plugging index (a measure of the magnitude of latex vessel plugging) was investigated. Lutoid instability, as indicated by bursting index, is negatively ocrrelated with the phospholipid content of the bottom fraction of latex. The neutral lipid content of rubber particles is positively correlated with the colloidal stability of latex. The latex vessel plugging during latex flow is found to be negatively correlated with both the lutoid stability and the neutral lipids in the rubber particle. A high ratio of cationic and anionic proteins in B-serum may also enhance the process of plugging.     

Changes in leaf physiology caused by Calacarus heveae (Acari, Eriophyidae) on rubber tree

The influence of Calacarus heveae Feres on physiological processes was evaluated in two rubber tree clones. Experiments were conducted in a greenhouse with 5-month-old potted seedlings of RRIM 600 and GT 1 clones, that were either infested with C. heveae or not (non-infested control). The level of photosynthetic pigments, net photosynthetic rate, stomatal conductance, transpiration rate, changes in relative humidity between leaf surface and ambient air (Δw) and intercellular CO(2) concentration (Ci CO(2)) were evaluated. Infested plants showed significant reductions in the rate of transpiration, the rate of photosynthesis, stomatal conductance and Δw. RRIM 600 seedlings showed more pronounced physiological damage than GT 1 seedlings, indicating a lower physiological tolerance of the former clone to the mite. However, carotenoid levels were reduced only in GT 1 seedlings. Photosynthesis was probably reduced due to a decrease in stomatal opening, as indicated by reductions in transpiration rate and stomatal conductance and by the absence of differences in chlorophyll levels between treatments. Our results indicate that populations of C. heveae reduce the productivity of rubber trees. Thus, farmers must to be aware to control this mite pest in rubber tree plantations.     

Molecular and Pathogenic Variation Identified Among Isolates of <Emphasis Type="Italic">Corynespora cassiicola</Emphasis>

Pathogenicity tests and inter simple sequence repeat (ISSR) molecular fingerprinting markers were utilized to analyze 24 Corynespora cassiicola isolates obtained from a lot of Hevea clones grown in most rubber nurseries and a few plantations in China. The C. cassiicola isolates were collected from Hainan and Yunnan provinces, China, from 2006 to 2008. The assay of 24 C. cassiicola isolates on detached leaves of four different Hevea rubber clones (genotypes PR 107, Dafeng 95, RRIM 600, and Reyan 7-33-97) indicated that 23 of the isolates were susceptible to RRIM 600, and were therefore considered race 1 except for CC-023. ISSR analysis grouped 24 C. cassiicola isolates into four clusters (A, B, C, and D). Unweighted pair-group method with arithmetic averaging (UPGMA) analysis based on Nei and Li's coefficient (calculated from the binary matrix data of 103 DNA fragments generated from 16 ISSR primers) indicated that cluster A included 19 isolates from Hainan and Yunnan (this cluster was further divided into two sub clusters (I, II), sub cluster II contained isolate CC-023); clusters B and C comprised of 1 isolates from Hainan, respectively; while cluster D encompassed 3 isolates from Hainan and Yunnan. Pathogenicity tests and ISSR analysis showed that there was no correlation between race structure, the geographical origin of the pathogen and their ISSR clusters because 23 of the isolates belonging to four distinct clusters were considered race 1 except for isolate CC-023. However, most of the isolates with different pathogenicity levels shared the same clades, and furthermore, the ISSR clusters and cology color had an exact correlation. These results should facilitate the development of rubber clones with enhanced resistance against all genetic clusters of C. cassiicola.     

In vitro photoautotrophic acclimatization,direct transplantation and ex vitro adaptation of rubber tree (<Emphasis Type="Italic">Hevea brasiliensis</Emphasis>)

We investigated the effect of carbon dioxide (CO₂)-ambient (350 µmol CO₂ mol^?1) and CO₂-enriched (1500 µmol CO₂ mol^?1) conditions of in vitro photoautotrophic system on two cultivars, ‘RRIM600’ and ‘RRIT413’ of rubber tree (Hevea brasiliensis) in an acclimatization process of 45 days. Survival percentage of in vitro rubber tree plantlets derived from somatic embryos under ambient CO₂ was better than those under CO₂-enriched conditions, especially in cv. ‘RRIT413’. Subsequently, the survival rate of ex vitro transplanted plantlets was similar to the in vitro plantlets and abnormal morphological characters such as light-green leaves (SPAD), small leaves in cv. ‘RRIT413’ acclimatized under CO₂-enriched conditions were demonstrated 30 days after the plantlets were transferred into the soil. Maximum quantum yield of PSII, photon yield of PSII, stomatal conductance and transpiration rate in cv. ‘RRIT413’ acclimatized under CO₂-enriched conditions were sharply declined by 39.0, 50.6, 47.1 and 45.8%, respectively as compared to those acclimatized under ambient CO₂ conditions. In contrast, the in vitro acclimatized plantlets of cv. ‘RRIM600’ were un-responsive under both ambient- and enriched-CO₂ conditions. In conclusion, genotypic dependent in response to CO₂ enriched conditions in in-vitro acclimatization of rubber tree plantlets was evidently demonstrated as a key result to regulate plant growth and development in ex vitro environments. Interestingly, soluble sugar contents (sucrose, glucose and fructose) were increased after transplanting the plantlets of cv. ‘RRIM600’ acclimatized under CO₂-enriched condition into the soil and thus, can be considered as an adaptive indicator of ex vitro adaptation.     

低NPK胁迫对橡胶树实生苗新生叶蓬NPK含量的影响

以橡胶树3个品种（GT1、RRIM600、PR107）实生苗为材料,研究正常施肥到低NPK胁迫和恢复正常施肥过程中叶片NPK含量的变化。结果表明：橡胶树实生苗新生叶蓬叶片NPK含量与NPK亏缺与否和胁迫处理的时间密切相关,且胁迫后的缺素状况可通过适时恢复正常施肥得以缓解或改善;叶片N、P、K含量出现降增的时间及幅度有所不同,其中N素的响应较快,P素次之,K素最为缓慢;在低NPK胁迫情况下,N、P、K素间存在不同的互作现象;不同品种叶片养分含量的变化趋势基本一致,但耐受低NPK的胁迫能力及恢复能力有所不同,RRIM600较为敏感,PR107次之,GT1较不敏感。     

Xylem embolism and stomatal regulation in two rubber clones (<Emphasis Type="Italic">Hevea brasiliensis</Emphasis> Muell. Arg.)

Vulnerability to water-stress-induced embolism of stems, petioles, and leaf midribs was evaluated for two rubber clones (RRIM600 and RRIT251). The xylem conduits were relatively vulnerable to cavitation with 50% of embolism measured for xylem pressures between –1 and –2 MPa. This feature can be related to the tropical-humid origin of the species. A distinct basipetal gradient of vulnerability was found, leaf midribs being the least vulnerable. Substantial variation in vulnerability to cavitation was found between the two clones only at the petiole level. A correlation was found between the stomatal behavior and the development of cavitation. Stomata were nearly closed when the xylem pressure reached the point of xylem dysfunction. Stomata may thus contribute to controlling the risk of cavitation. However, for one clone a poor correlation was found between stomatal regulation and petiole vulnerability. This was consistent with a high degree of embolism measured in the petioles after a soil drought event. Therefore, xylem cavitation might represent a promising criterion to evaluate the performance of rubber clones under drought conditions.     

橡胶树两个品种的核型分析

热研7-33-97（Hevea brasiliensis,热研7-33-97）和RRIM600（Hevea brasiliensis,RRIM600）都是大规模推广种植的优良橡胶树品种,对其核型研究在橡胶树育种中有着重要的意义。采用去壁低渗方法对热研7-33-97和橡胶栽培品系RRIM600的染色体数目与核型进行了研究。结果表明：热研7-33-97的染色体核型公式为2n=36=34m（4sat）＋2sm,核型为2B型。RRIM600的染色体核型公式为2n=36=28m（4sat）＋8sm,核型为2B型。热研7-33-97和RRIM600的核型在进化上属于比较原始的类型。该研究可为橡胶树育种、种质资源鉴定及基因定位提供细胞学基础。     

Yield components in Hevea brasiliensis– theoretical considerations

Abstract A theoretical analysis of yield components of Hevea brasiliensis is attempted in this paper. The effect of the major yield components, i.e. initial flow rate per unit length of tapping cut, length of the cut, percentage rubber content and plugging index on rubber yield is represented by the formula Variation in yield within and between clones can be ascribed to variation to any one of the above components. The importance of high growth rate for maintaining high yield throughout the life cycle of the tree is theoretically elucidated. While the present contention of a theoretical maximum yield of 9.5 t ha^?1 with a stand of 350 trees is questioned, the theoretical possibility of attaining that yield by increasing the stand per ha to 600 is analysed.     

AFLP Fingerprinting Analysis of Elite Hevea brasiliensis Germplasm

There are more than 6 000 clones of Hevea brasiliensis Mull. Ary germplasm in the germplasm garden of Chinese National Key Biotechnology Laboratory for Tropical Crops and some of them are elite germplasm demonstrated by production and previous studies. AFLP (amplified fragment length polymorphism) fingerprinting analysis was performed on 25 clones (15 of Wickham clones and 10 of Amazon wild clones which possess phenotypes with high-yielding/low-yielding, cold-tolerance/cold-sensitivity, oidium-resistance/oidium-sensitivity, tapping panel dryness (TPD)/healthy) respectively through a 377 DNA sequencer (P. E. Corp.) and PAGE electrophoresis results were analyzed by using GeneScanTMand GenotypeTM Analysis software (P.E.Corp.). The fragment profiles of different clones were obtained. Five hundred and eighteen fragments were generated by two primer combinations screened from 64 primer combinations and 511 fragments appeared to be polymorphic (98.6%). Genetic distance ranged from 0.25 to 0.81 between clones and ranged from 0.07 to 0.17 within RRIM600 clone. A specific 320 bp fragment of the oidium-resistant clones was found through genotype analysis. These results showed that AFLP fingerprints were highly reproducible and powerful and can be widely used in germplasm identification and genetic diversity analysis of Hevea brasiliensis. In addition, based on the AFLP data, cluster analysis was performed. Cluster results showed that all the clones studied were almost clustered into a group one by one.      

AFLP在橡胶树优异种质研究中的应用

采用377DNA测序仪（P.E.Corp.)用AFLP技术对25种分别具有高产／低产、抗白粉病／感白粉病、抗寒／不抗寒、死皮／不死皮性状的橡胶树（Hevea brasiliensis Mull.Ary)无性系（其中Wickham种质15种,Amazon野生橡胶树种质10种）进行了指纹图谱分析,从64对引物组合中选出2对引物组合构建了所有被研究种质的指纹图谱。2对引物共扩增出518条带,多态性比率超过98．6％。遗传多样性分析表明,橡胶树种质间遗传距离为0．25－0．81,RRIM600种质内遗传距离为0．07－0．17。通过基因分型分析获得一条大小为320bp的抗白粉病种质特有的片段。根据以上的AFLP数据进行了聚类分析,结果表明所有被研究种质几乎是依次聚成一个大类。     

Susceptibility of rubber trees to Loranthaceae in Gabon

Abstract

In Gabon, two major species of Loranthaceae, Phragmanthera capitata (Sprengel) Balle and Globimetula braunii (Engl.) Danser to a lesser extent, parasitize Hevea brasiliensis (Müll.) Arg. The behavior of different clones of rubber trees facing the Loranthaceae parasitism has been studied in the large industrial plantation of Mitzic and in three neighboring smallholding plantations. The clones PR 261, RRIM 600 and PB 260, although never totally resistant, are far less susceptible to Loranthaceae than the clones AVROS 2037, PB 217, PB 235 and GT 1. The parasite development kinetics was more rapid on the most susceptible clones. Phellem thickness, cortical parenchyma thickness and tannin cell density were independent of susceptibility. The number of tree ramifications was significantly different between the clones. Interestingly, on the least susceptible trees, being also the most ramified, the parasite showed the slowest development.