抗虫耐除草剂转基因玉米花粉生活力研究

使用离体培养法检测不同玉米材料花粉萌发率并于显微镜下观察花粉粒直径,研究转Cry1Ab、EPSPS基因玉米对花粉生活力的影响。结果表明,刚离体花粉萌发活力最强,培养180 min后花粉萌发率可达89.4%,高温(36℃)条件下放置2 h花粉基本丧失萌发能力。与非转基因对照相比,转Cry1Ab、EPSPS基因玉米对花粉离体后的生存能力无显著影响,对花粉粒直径大小无影响。     

Quantification of Bacillus thuringiensis Cry1Ab protein in tissues of YieldGard (MON810) corn hybrids tested at multiple field locations in India

Transgenically expressed Bacillus thuringiensis insecticidal-protein Cry1Ab was quantified in target tissues of insect feeding of several YieldGard^® corn hybrids. The Cry1Ab protein is intended to protect corn plants from two economically important stem borers, Chilo partellus and Sesamia inferens. A total of seven YieldGard hybrids, all with MON810 event, were field-tested in a total of fourteen locations during the dry season (October–March) of 2005/2006 and wet season (May–October) of 2006. S. inferens and C. partellus oviposit on leaves of young corn plants, 15–60 days after emergence (DAE). The neonates initially feed by scraping the leaf lamina before migrating to bore into the stem. Thus high concentrations of Cry1Ab in whorl leaf and stem tissues would ensure effective control of the borers. The mean tissue Cry1Ab concentrations during the oviposition window of the borers (15–60 DAE), ranged from 50.05 to 21.01 ppm in whorl leaf, and between 9.26 and 3.47 ppm in stem tissue during the same period in the dry season of 2005/06. Similarly, Cry1Ab concentrations in whorl leaf and stem between 15 and 60 DAE during the wet season of 2006 ranged between 19.30 to 11.08 and 14.28 to 4.69 ppm, respectively. The baseline-sensitivity data of these insects to Cry1Ab in laboratory assays was determined. The concentrations of Cry1Ab in the target tissues as studied in seven YieldGard hybrids tested suggest effective management of the two borers. This paper also provides a summary of the expression of the Cry1Ab gene in various genetic backgrounds.     

Transgenic wheats with elevated levels of Dx5 and/or Dy10 high-molecular-weight glutenin subunits yield doughs with increased mixing strength and tolerance

To test the effects of independently increasing the in vivo levels of high-molecular-weight glutenin subunits (HMW-GS) Dx5 and Dy10 on wheat flour properties, we increased the copy numbers of their corresponding genes by genetic transformation. Thirteen transformants with increases in one or both subunits were chosen for biochemical and functional characterization by solvent fractionation, RP-HPLC, SDS-sedimentation, and micro-mixing. Increases in Dx5 and Dy10 contents ranged from 1.4- to 3.5-fold and 1.2- to 5.4-fold, respectively, and generally resulted in increased polymeric protein, increased mixing times and tolerances, and lower peak resistances. Increases in Dx5 content had larger effects on most parameters than comparable increases in Dy10. Flours with more than 2.6-times the native levels of Dx5 could not be mixed under standard 2-g mixograph conditions, while flours with 5.4 times the native levels of Dy10 could be mixed if sufficient time was allowed. Increases in Dx5 and Dy10 had additive effects on mixing behavior. These experiments demonstrate that dough mixing strength and tolerance can be increased by raising the levels of native HMW-GS Dx5 or Dy10, but that the effects of the two subunits are quantitatively and qualitatively different.     

Study on antagonistic effects of Talaromyces flavus on Verticillium albo-atrum, the causal agent of potato wilt disease

Talaromyces flavus a fungal antagonist, was isolated from soil samples collected from potato fields in Varamin and Karaj districts, Tehran province, Iran. Antagonistic effects of T. flavus isolates against Verticillium albo-atrum, the causal agent of potato wilt disease were investigated in the laboratory and greenhouse conditions. T. flavus colonies were recovered after three weeks from soil samples cultured on selective medium. Antagonistic effects of volatile and non-volatile extracts of T. flavus isolates on V. albo-atrum growth were investigated in the laboratory and five that caused higher growth inhibition of V. albo-atrum, were selected for greenhouse experiments. Infection index was compared in the greenhouse in a split plot trial with five isolates applied to soil, seed, or both arranged in a randomized complete block design with four replications. The minimum infection index was observed when seed were treated with T. flavus with the most effective isolate being Tf-Po-V-52. On seed, the minimum infection index was observed with Tf-Po-V-50. The most effective T. flavus isolate was also evaluated in a field experiment. Results indicated that infection index in seed treatment contained this isolate was 0.15 whereas that of control was 3.5. The overall results of this study showed that it may be possible to manage potato Verticillium wilt disease effectively by using T. flavus, a biocontrol fungus.     

Susceptibility of Helicoverpa armigera from different host plants in northern China to Bacillus thuringiensis toxin Cry1Ac

Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) is a serious pest of cotton and many other crops in northern China. To evaluate the contribution of alternative hosts as an effective refuge for transgenic cotton expressing the Bacillus thuringiensis (Bt) Cry1Ac toxin, the susceptibility to this toxin was measured in progeny derived from field-collected H. armigera larvae and pupae from different hosts in the Xiajin’s region of the Shandong Province in northern China. During 2008-2010, progeny from a total of 258,56,184 and 160 single-pair crosses derived from wheat (first-generation), Bt cotton (second-generation), Bt cotton (third-generation), and corn (third-generation) were screened on Cry1Ac diets, respectively. Based on relative average development rates (RADR) of H. armigera larvae in these F₁ tests, the second and third-generation moths emerging from Bt cotton fields were more tolerant to the Bt toxin than the first and third-generation moths emerging from wheat and corn each year. These results suggest that there is significant variation in susceptibility to Bt toxins among H. armigera populations derived from different host crops. Alternate crops, such as corn, that maintain Bt susceptible populations of H. armigera could be used as refugia to minimize the evolution of resistance to Bt cotton.     

Potato viruses in China

Potato (Solanum tuberosum L.) is globally the fourth largest staple crop after rice, wheat and maize. China is the largest potato producer worldwide, accounting for 26.3% and 22.2% of the global total area and yield, respectively. Viral diseases have been a limiting factor for sustainable production of potato in China. This article provides update and comprehensive information on the most important viruses, their effect on yield and their control in China. Meristem culture is currently the most widely used technique for production of virus-free potato plants and these plants are used as nuclear stock for the production of certified seed tubers. Although a significant increase in tuber yield has been achieved using certified seed potatoes, lack of application of a national seed potato production scheme has hindered progress. Conventional breeding using virus-resistant cultivars introduced from other countries as parents has already led to the production of Chinese cultivars. Use of virus-resistant wild Solanum species and genetic transformation techniques will provide new virus-resistant varieties in the future to further enhance control of potato viruses in China.     

Susceptibility of Cry1Ab-susceptible and -resistant sugarcane borer to transgenic corn plants containing single or pyramided Bacillus thuringiensis genes

Multiple independent trials were conducted to evaluate the performance of Cry1Ab-susceptible (Cry1Ab-SS), -heterozygous (Cry1Ab-RS), and -resistant (Cry1Ab-RR) genotypes of the sugarcane borer, Diatraea saccharalis (F.), on eight commercial hybrids and six experimental corn lines. The commercial varieties included two non-Bt and six Bt corn hybrids that expressed a single Bt protein (either Cry1Ab or Cry1F) targeting above-ground lepidopteran pests. The six experimental lines consisted of two non-Bt and four Bt corn lines, two expressing just the Cry1Ab protein and two containing the pyramided-genes Cry1A.105 and Cry2Ab2 (event MON 89034). Larval mortality on non-Bt corn leaf tissue ranged from 6 to 45% after 12 d across insect genotypes. The 12 d mortality of Cry1Ab-SS on leaf tissue of commercial Cry1Ab or Cry1F corn was 96-100%, whereas it was 80-96% for Cry1Ab-RS and 68-78% for Cry1Ab-RR. On intact plants, 39-64% of larvae survived on non-Bt corn plants after 21-25 d. Larval survivorship on intact plants of commercial Cry1Ab or Cry1F corn was 0-8.1% for Cry1Ab-SS, 1.3-34% for Cry1Ab-RS, and 19-51% for Cry1Ab-RR. Larvae of Cry1Ab-RR and -RS also caused significant plant injury to most of the commercial Bt corn hybrids, especially to the Cry1Ab corn. Cry1Ab resistance in D. saccharalis was incompletely dominant on commercial Bt corn hybrids. However, both experimental lines with pyramided genes of Cry1A.105 and Cry2Ab2 provided complete control of all three insect genotypes in both leaf tissue and intact plant tests. Results of this study suggest that MON 89034 should offer a means for Bt resistance management in D. saccharalis.     

Mycoparasitic Trichoderma viride as a biocontrol agent against Fusarium oxysporum f. sp. adzuki and Pythium arrhenomanes and as a growth promoter of soybean

Trichoderma viride was proved as an effective biocontrol agent against two fungal pathogens, Fusarium oxysporum f. sp. adzuki and Pythium arrhenomanes, infecting soybean. During an in vitro biocontrol test, Trichoderma showed mycoparasitism and destructive control against the tested fungal pathogens. Both the pathogens significantly influence the germination and P. arrhenomanes had a severe effect (only 5% germination). The root system of the soybean plant was poorly developed due to the infection and it exerted a negative influence on the nodulation and further growth phases of the plant. During pot assay along with biocontrol activity, Trichoderma showed growth promoting action on the soybean plant. Trichoderma enhanced growth of shoot and root systems and fruit yield after 12 weeks of growth. Pythium and Fusarium infected plants treated with Trichoderma had ∼194% and 141% more height than pathogens alone. The fruit yield treated with Trichoderma was ∼66 per plant whereas the yield was only 41 for a control plant. The plants infected with Pythium and Fusarium and treated with Trichoderma had fruit yields of 43 and 53 respectively and those were 5 and 1.6 times higher than plants infected with pathogens.     

Effects of the nematicide 1,3-dichloropropene on weed populations and stem canker disease severity in potatoes

The soil fumigant 1,3-dichloropropene (1,3-D) has been used in the UK for the control of potato cyst nematodes (PCN), Globodera pallida (Stone) and Globodera rostochiensis (Wollenweber), but its potential herbicidal activity has not been extensively investigated in this country. Field and glasshouse studies were therefore conducted to evaluate the potential of 1,3-D for the control of weeds in potatoes, and observations were made on the severity of potato stem canker, caused by the fungus Rhizoctonia solani Kühn [teleomorph: Thanatephorus cucumeris (Frank) Donk]. Autumn application of 1,3-D at 211.5 L active substance (a.s.) ha⁻¹ significantly suppressed the number of germinating weeds and the percentage of weed ground cover by 83% and 79%, respectively, relative to controls. There were also species-specific significant decreases (field pansy, Viola arvensis, in particular) in the number of weed seeds germinating in field soil in the glasshouse post-1,3-D treatment. The effect of 1,3-D declined in time and single (autumn or spring) or combined application produced a slight, but not significant, reduction in the number of weeds germinated on potato ridges relative to those recorded in untreated soil. The severity of stem canker on potato plants was not significantly reduced by 1,3-D but both mean number and weight of stems per plant were significantly increased compared with plants from untreated plots. These studies demonstrated that 1,3-D, in addition to giving PCN control, has efficacy against weeds; implications are the potential for reduced herbicide input in the crop rotation with accompanying economic and environmental benefits.     

Damage and survivorship of fall armyworm (Lepidoptera: Noctuidae) on transgenic field corn expressing Bacillus thuringiensis Cry proteins

Field corn, Zea mays L., plants expressing Cry1Ab and Cry1F insecticidal crystal (Cry) proteins of Bacillus thuringiensis (Bt) Berliner are planted on considerable acreage across the Southern region of the United States. The fall armyworm, Spodoptera frugiperda (J.E. Smith), is an economically important pest during the mid-to-late season on non-Bt and some commercial Bt corn hybrids. The objective of this study was to quantify foliar injury and survivorship of fall armyworm on transgenic corn lines expressing Cry1Ab or Cry1F Bt proteins. Corn lines/hybrids expressing Cry1Ab, Cry1F, and a conventional non-Bt cultivar were evaluated against artificial infestations of fall armyworm in field trials. Larvae (second instars) of fall armyworm were placed on corn plants (V8-V10 stages). Leaf injury ratings were recorded 14 d after infestation. Hybrids expressing Cry1F had significantly lower feeding injury ratings than non-Bt corn plants. Development and survivorship of fall armyworm on Bt corn lines/hybrids were also evaluated in no-choice laboratory assays by offering freshly harvested corn leaf tissue to third instars. Transgenic corn hybrids expressing Cry1Ab or Cry1F significantly reduced growth, development, and survivorship of fall armyworm compared to those offered non-Bt corn tissue. However, 25-76% of third instars offered Bt corn leaf tissues successfully pupated and emerged as adults. These results suggest Cry1Ab has limited effects on fall armyworm; whereas Cry1F demonstrated significant reductions in foliar injury and lower survivorship compared to that on non-Bt corn tissues. Although fall armyworm is not considered a primary target for insect resistance management by the U.S. Environmental Protection Agency, these levels of survivorship could impact selection pressures across the farmscape, especially when considering that transgenic Bt cotton cultivars express similar Cry (Cry1Ac or Cry1F) proteins.     

Comparative assessment of the field-susceptibility of Sesamia nonagrioides to the Cry1Ab toxin in areas with different adoption rates of Bt maize and in Bt-free areas

A three year (2003-2005) field study compared the susceptibility to the Cry1Ab toxin, expressed in Bt maize, of Mediterranean corn borer (MCB) Sesamia nonagrioides populations collected from areas with different adoption rates of Bt maize in Spain with Bt-free areas in Greece. Spain is the only European country where the cultivar Compa CB derived from the event Bt176 was commercially grown, from 1998 to 2005. The large decrease of the titer of the toxin in this cultivar at later growth stages represented the worst-case scenario for resistance development of MCB, since larvae of the second and third generations were exposed to sublethal concentrations of Cry1Ab toxin. Our data revealed that the variation in susceptibility to Cry1Ab for the MCB Spanish field populations analyzed in the three years was very low, with LC₅₀ values fluctuating between 12 and 30 ng Cry1Ab/cm², regardless of the region of origin, the type of maize (Bt or non-Bt) and the year. Furthermore, no significant differences were found when comparisons were made with a laboratory population (LC₅₀ values: 18-26 ng Cry1Ab/cm²) or with field populations from Greece (Bt-free areas), which displayed LC₅₀ values ranging between 22 and 27 ng Cry1Ab/cm². Standardizing bioassay protocols proved to be essential for obtaining comparable results. These findings suggest that resistant MCB populations did not evolve in those Spanish maize areas where Compa CB was largely cultivated for eight years, contradicting the expected rapid development of resistance under these unfavourable conditions. Additionally, our results can be used as baseline indices in post-market resistance monitoring programs if Bt maize is introduced in Greece. Further studies should continue, since the insights gained from a resistance monitoring program may help to enhance the durability of Bt maize.     

Management of the root-knot nematode Meloidogyne incognita on tomato with combinations of different biocontrol organisms

The nematicidal effect of Pseudomonas fluorescens, Paecilomyces lilacinus, Pichia guilliermondii and Calothrix parietina singly or in combination was tested against root-knot nematode, Meloidogyne incognita. Treatments with P. fluorescens and P. lilacinus caused mortality of M. incognita as 45% and 30% of juveniles after 48 h of exposures, respectively compared to water control in vitro. Under greenhouse conditions, all treatments reduced the disease severity and enhanced plant growth compared to untreated control. Application of P. fluorescens, P. lilacinus and P. guilliermondii Moh 10 was more effective compared to C. parietina. There was a negative interaction between C. parietina and either P. lilacinus or P. guilliermondii. Fresh and dry weight of shoots and roots of plants were significantly reduced as a result of infection with M. incognita, however application of biocontrol agents singly or in mix recovered this reduction. Moreover, they enhanced the growth parameters compared with the control. Our results proved that application of different biocontrol agents (P. fluorescens, P. lilacinus and P. guilliermondii) not only has a lethal effect on nematode, but also enhances the plant growth, supplying many nutritional elements and induction the systemic resistance in plants. Presence of C. parietina as a soil inhabitant cyanobacterium could antagonize biocontrol agents leading to the reduction of their practical efficiency in soil.     

Addressing current and future problems of parasitic weeds in rice

Significant areas of rain-fed rice in the Sahel, savannah and derived savannah zones of sub-Saharan Africa (SSA), Madagascar and other Indian Ocean Islands are infested by parasitic weeds. The affected area accommodates some of the poorest farmers of the world. Without appropriate management parasitic weeds in rice are expected to increase in importance in SSA due to their general invasive nature and their abilities to adapt to changing conditions such as imposed by predicted climate changes. The most important parasitic weeds in rice are: Striga hermonthica, Striga asiatica, Striga aspera and Rhamphicarpa fistulosa. The first two are primarily found in free-draining uplands while S. aspera is also found on hydromorphic soils and R. fistulosa is restricted to unimproved lowlands including inland valleys. As parasitic weeds are typical production constraints in subsistence rice production, targeting them would directly contribute to poverty alleviation and food security. This paper provides an overview of the problems caused by parasitic weeds in rice and discusses management options and opportunities for research for development.     

Genetic characterization of kernel polyphenol oxidases in wheat and related species

Polyphenol oxidase (PPO) activity causes undesirable darkening of raw Asian noodles and other wheat products. In this study we investigate the genetic origins and diversity of wheat kernel PPO. PPO was characterized via activity assays, antigenic staining, and Southern blots in Triticum aestivum, Triticum dicoccoides, Triticum durum, Triticum dicoccum, Triticum monococcum, Triticum urartu, Aegilops speltoides, and Aegilops tauschii. Among these species, PPO activity was well-correlated with antigenic staining intensity toward a wheat kernel-type PPO antibody. High PPO activity was observed in all three T. monococcum accessions (A^m genome), one Ae. speltoides accession, one T. durum accession, and two hexaploid wheat cultivars. Southern blots suggested the presence of two or more kernel-type PPO genes in diploid progenitors of the hexaploid A, B, and D genomes. Whole-kernel PPO activity was evaluated in disomic substitution lines derived from three T. dicoccoides accessions in the background of T. durum ‘Langdon’. PPO activity was primarily associated with chromosome 2A and to a much lower degree with chromosome 2B. DNA sequence comparisons showed that the intron associated with the high PPO allele on chromosome 2AL of hexaploid wheat had 94% nucleotide identity with the homeologous intron found in T. monococcum, a species with high kernel PPO activity. This implies that the ancestral PPO allele on the A genome is one of the high activity, and the low PPO allele found in hexaploid wheat represents a relatively recent genetic alteration. Results confirm the presence of multiple kernel-type PPO genes in the diploid and tetraploid progenitors and relatives of hexaploid wheat. However, it is likely that relatively few of the many kernel-type PPO genes present in wheat contribute substantially to kernel PPO activity. A single genetic locus on homeologous group 2 chromosomes may be the primary cause of high PPO activity in wheat kernels.     

Isolation and characterization of EMS-induced Dy10 and Ax1 high molecular weight glutenin subunit deficient mutant lines of elite hexaploid wheat (Triticum aestivum L.) cv. Summit

The mixing properties of the dough are critical in the production of bread and other food products derived from wheat. The high molecular weight glutenin subunits (HMW-GS) are major determinants of wheat dough processing qualities. The different alleles of the HMW-GS genes in hexaploid wheat vary in their effect on dough quality. To determine the contribution of the individual HMW-GS alleles, lines deficient in HMW-GS proteins were generated by chemical mutagenesis in the elite bread wheat Triticum aestivum cv. Summit. In this report we describe the identification and characterization of Dy10 and Ax1 deficient lines. Examination of the effect of Dy10 and Ax1 deficiency on dough rheological properties by mixography showed shorter mixing time to reach peak resistance, and weaker and less extensible doughs relative to the wild type control. This is the first time that the role of Dy10 in vivo has been examined apart from the Dx5 + Dy10 allelic pair combination.     

Effects of fungicide seed treatments on germination, population, and yield of maize grown from seed infected with fungal pathogens

Seedborne fungi can reduce survival, growth, and yield of maize (Zea mays L.). Laboratory, field, and growth chamber experiments were conducted to determine the effects of the seed treatment fungicides fludioxonil, mefenoxam, and azoxystrobin on germination, plant population, and grain yield of maize grown from low-quality hybrid seed infected with seedborne fungal pathogens. Study I used seed of four hybrids infected at 0-54% incidence with Fusarium spp., Stenocarpella maydis, Penicillium spp., Rhizopus spp., and/or Aspergillus spp. Study II used three seed lots for each of two hybrids infected at 7-37% incidence with S. maydis. Warm and cold germination for untreated seed varied among hybrids in both studies. Warm germination of the seed lot with the highest incidence of S. maydis in study II treated with azoxystrobin and fludioxonil was significantly greater (+7%) than the nontreated control. Plant population in study I was significantly affected by seed treatment, hybrid, and their interactions. Populations were greater (≥9%) for fludioxonil, fludioxonil + mefenoxam, and fludioxonil + mefenoxam + azoxystrobin treatments compared to controls. In growth chamber experiments with pasteurized soil, emergence (≥5%) and plant dry weight (≥14%) were both greater than controls only with the triple seed treatment. Plant populations in study II for all seed treatments except mefenoxam and azoxystrobin alone were greater (≥4%) than controls. Yield in study I was significantly affected by hybrid and seed treatment. Yield for one hybrid was higher (≥20%) than the control with all seed treatments except fludioxonil, whereas yield with another hybrid was consistently greater (≥26%) only with the triple seed treatment. Yield in study II was significantly affected by hybrid, seed treatment, and their interactions. Yield was greater (≥8%) than the controls for all seed treatments with one hybrid and with all (≥5%) except azoxystrobin for the other hybrid. Highest yields occurred with the triple seed treatment. Results indicate that fludioxonil and azoxystrobin can increase germination, population, and yield of maize grown from seed infected by S. maydis and other fungi.     

Comparative susceptibility of Ostrinia furnacalis, Ostrinia nubilalis and Diatraea saccharalis (Lepidoptera: Crambidae) to Bacillus thuringiensis Cry1 toxins

Transgenic corn hybrids that express toxins from Bacillus thuringiensis (Bt) are highly effective against the European corn borer, Ostrinia nubilalis (Hübner), and the closely related Asian corn borer, Ostrinia furnacalis (Guenée). Since the registration of Bt corn hybrids in the U.S. in 1996, there has been a great deal of information generated on O. nubilalis. However, relatively little information exists for O. furnacalis. To help determine whether the information generated for O. nubilalis can be leveraged for decisions regarding the use of transgenic Bt corn against O. furnacalis, experiments were designed to determine whether the pattern of sensitivity to various Bt Cry1 toxins is similar between the two species. Test insects included laboratory-reared O. furnacalis originating from Malaysia, a Bt-susceptible laboratory colony of O. nubilalis maintained at the University of Nebraska-Lincoln (UNL) and an out-group consisting of the sugarcane borer, Diatraea saccharalis (F.), from Louisiana which represents a different genus from the same family. O. furnacalis and O. nubilalis exhibited a similar pattern of susceptibility to all the Cry1 toxins and were highly susceptible to the range of Bt toxins tested including Cry1Aa, Cry1Ab, Cry1Ac and Cry1F. Both of the Ostrinia species were more tolerant to Cry1Ba compared with D. saccharalis, although sensitivity of O. furnacalis was intermediate and did not differ significantly from that of O. nubilalis and D. saccharalis. D. saccharalis was also susceptible to the range of toxins tested but unlike the two Ostrinia species, was more tolerant to Cry1F and more susceptible to Cry1Ba. These results indicate that both of the Ostrinia corn borer species are similar in sensitivity to the Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ba and Cry1F toxins, thus suggesting shared toxin receptors and mechanisms of toxicity for the two species.