Fast neutron mutagenesis of soybean (Glycine soja L.) produces a supernodulating mutant containing a large deletion in linkage group H

We describe for the first time the application of fast neutron mutagenesis to the genetic dissection of root nodulation in legumes. We demonstrate the utility of chromosomal deletion mutations through production of a soybean supernodulation mutant FN37 that lacks the internal autoregulation of nodulation mechanism. After inoculation with microsymbiont Bradyrhizobium japonicum, FN37 forms at least 10 times more nodules than the wild type G. soja parent and has a phenotype identical to that of chemically induced allelic mutants nts382 and nts1007 (NTS-1 locus). Reciprocal grafting of shoots and roots confirmed systemic shoot control of the FN37 nodulation phenotype. RFLP/PCR marker pUTG132a and AFLP marker UQC-IS1 which are tightly linked to NTS-1 allowed the isolation of BAC contigs delineating both ends of the deletion. The genetic/physical distance ratio in the NTS-1 region is 279 kb/cM. The deletion is estimated to be about 460 kb based on the absence of markers and bacterial artificial chromosomes (BAC) ends as well as genetic and physical mapping. Deletion break points were determined physically and placed within flanking BAC contigs.

Generation of CTL responses using Kunjin replicon RNA

The Kunjin replicon was used to express a polytope that consisted of seven hepatitis C virus cytotoxic T lymphocyte epitopes and one influenza cytotoxic T lymphocyte epitope for vaccination studies. The self-replicating nature of, and expression from, the ribonucleic acid was confirmed in vitro . Initial vaccinations with one dose of Kun-Poly ribonucleic acid showed that an influenza-specific cytotoxic T lymphocyte response was elicited more efficiently by intradermal inoculation compared with intramuscular delivery. Two micrograms of ribonucleic acid delivered in the ear pinnae of mice was sufficient to elicit a detectable cytotoxic T lymphocyte response 10 days post-vaccination. Further vaccination studies showed that four of the seven hepatitis C virus cytotoxic T lymphocyte epitopes were able to elicit weak cytotoxic T lymphocyte responses whereas the influenza epitope was able to elicit strong, specific cytotoxic T lymphocyte responses following three doses of Kun-Poly ribonucleic acid. These studies vindicate the use of the Kunjin replicon as a vector to deliver encoded proteins for the development of cell-mediated immune responses.

Evaluation of potential biocontrol agents against Claviceps africana in vitro and in vivo

Undiluted culture filtrates of two commercial products of Trichoderma spp., Trichopel and Trichoflow, and two isolates of Penicillium citrinum completely inhibited the conidial germination of macroconidia of Claviceps africana , the cause of ergot or sugary disease of sorghum (Sorghum bicolor) in vitro . Similarly, Pseudomonas aeruginosa and Burkholderia cepacia completely inhibited macroconidial germination, with the former being more effective at high dilutions. In contrast, these bacterial isolates failed to inhibit infection in vivo in glasshouse tests with ergot-inoculated sorghum, but all fungal biocontrol agents (including an isolate of Epicoccum nigrum) reduced the severity of disease (percentage of infected spikelets per panicle), in some cases completely inhibiting the development of ergot. In a second glasshouse trial, optimum control was achieved when the biocontrol agents were applied 3-7 days before inoculation with conidia of C. africana .

DNA vaccine coding for the full-length infectious Kunjin virus RNA protects mice against the New York strain of West Nile virus

A plasmid DNA directing transcription of the infectious full-length RNA genome of Kunjin (KUN) virus in vivo from a mammalian expression promoter was used to vaccinate mice intramuscularly. The KUN viral cDNA encoded in the plasmid contained the mutation in the NS1 protein (Pro-250 to Leu) previously shown to attenuate KUN virus in weanling mice. KUN virus was isolated from the blood of immunized mice 3-4 days after DNA inoculation, demonstrating that infectious RNA was being transcribed in vivo; however, no symptoms of virus-induced disease were observed. By 19 days postimmunization, neutralizing antibody was detected in the serum of immunized animals. On challenge with lethal doses of the virulent New York strain of West Nile (WN) or wild-type KUN virus intracerebrally or intraperitoneally, mice immunized with as little as 0.1-1 mug of KUN plasmid DNA were solidly protected against disease. This finding correlated with neutralization data in vitro showing that serum from KUN DNA-immunized mice neutralized KUN and WN,viruses with similar efficiencies. The results demonstrate that delivery of an attenuated but replicating KUN virus via a plasmid DNA vector may provide an effective vaccination strategy against virulent strains of WN virus.

Early pregnancy factor treatment suppresses the inflammatory response and adhesion molecule expression in the spinal cord of SJL/J mice with experimental autoimmune encephalomyelitis and the delayed-type hypersensitivity reaction to trinitrochlorobenzene in normal BALB/c mice

Early pregnancy factor (EPF) is a secreted protein, present in serum during early pregnancy and essential for maintaining viability of the embryo. It is a homologue of chaperonin 10 (Cpn10) but, unlike Cpn10, it has an extracellular role. EPF has immunosuppressive and growth regulatory properties. Previously we have reported the preparation of recombinant EPF (rEPF) and shown that treatment with rEPF will suppress clinical signs of MBP-EAE in Lewis rats and PLP-EAE in SJL/J mice. In the present study, these findings have been extended to investigate possible mechanisms involved in the action of EPF. Following treatment of mice with rEPF from the day of inoculation, there were fewer infiltrating CD3+ and CD4+ cells in the parenchyma of the spinal cord during the onset of disease and after the initial episode, compared with mice treated with vehicle. Expression of the integrins LFA-1, VLA-4 and Mac-1 and of members of the immunoglobulin superfamily of adhesion molecules ICAM-1 and VCAM-1 was suppressed in the central nervous system (CNS) following rEPF treatment. The expression of PECAM-1 was not affected. To determine if rEPF suppressed T cell activation in the periphery, the delayed-type hypersensitivity (DTH) reaction of normal BALB/c mice to trinitrochlorobenzene (TNCB) following treatment with rEPF was studied. The results showed that treatment with rEPF suppressed the DTH reaction, demonstrating the ability of EPF to downregulate the cell-mediated immune response. These results indicate that suppression of immunological mechanisms by rEPF plays a major role in the reduction of clinical signs of disease in experimental autoimmune encephalomyelitis (EAE). (C) 2003 Elsevier Science B.V. All rights reserved.

Early pregnancy factor suppresses the infiltration of lymphocytes and macrophages in the spinal cord of rats during experimental autoimmune encephalomyelitis but has no effect on apoptosis

Early pregnancy factor (EPF) is a secreted protein with immunosuppressive and growth factor properties that has been shown to suppress acute experimental autoimmune encephalomyelitis (EAE) induced with myelin basic protein (MBP) in Lewis rats. EAE is associated with infiltration of the central nervous system (CNS) with inflammatory cells. Spontaneous recovery involves the loss of T lymphocytes from the CNS and the selective apoptosis of Vbeta8.2(+) cells. In the present study, T cell, macrophage (CD11b/c(+)) and B cell (CD45RA(+)) populations in spinal cord and popliteal lymph nodes (LN) of Lewis rats with EAE were quantitated and apoptosis was studied. Rats were treated with EPF or vehicle. Following treatment on day 14 after inoculation with MBP, neither 1 x 100 mug nor 2 x 100 mug doses of EPF affected the total number of cells infiltrating the spinal cord on day 15, although the higher dose caused a decrease in the number of CD5(+) and CD11b/c(+) cells. Treatment with 2 x 100 mug/day from days 10 to 14 decreased the total number of infiltrating cells, and the numbers of CD5(+), CD11b/c(+) and CD45RA(+) cells. Apoptosis was unaffected. No alteration on the number or type of inflammatory cells in the popliteal LN was observed after treatment on days 10-14. However, treatment with EPF from days 0 to 11 increased the total number of T and B cells and CD5(+) T cells found on day 12 in the LN. Similarly, there was an increase in the frequency of MBP-reactive cells in the LN as determined by limiting dilution analysis. These results suggest that EPF treatment reduces the numbers of lymphocytes and macrophages in the CNS, possibly through an effect on cell trafficking. (C) 2003 Elsevier B.V. All rights reserved.

Ovary colonization by Claviceps africana is related to ergot resistance in male-sterile sorghum lines

Ergot, caused by Claviceps africana, has emerged as a serious threat to sorghum hybrid seed production worldwide. In the absence of gene-for-gene-based qualitative resistance in commercial cultivars, varieties with high pollen production that can escape ergot infection are preferred. Recent demonstration of differences in ergot susceptibility among male-sterile lines has indicated the presence of partial resistance. Using chitin-specific fluorescin-isothiocyanate-conjugated wheat germ agglutin and callose-specific aniline blue, this study investigated the process of sorghum ovary colonization by C. africana. Conidia germinated within 24 h after inoculation (a.i.); the pathogen was established in the ovary by 79 h a.i., and at least half of the ovary was converted into sphacelial tissue by 120 h a.i. Changes in fungal cell wall chitin content and strategic callose deposition in the host tissue were associated with penetration and invasion of the ovary. The rate of ovary colonization differed in three male-sterile lines that also differed in ergot susceptibility. This work demonstrates a possible histological basis for partial resistance in male-sterile sorghum lines that could lay the foundation for variety improvement through further breeding and selection.

Distribution of Sugarcane mosaic virus in sugarcane plants

Variation in the concentration of virus in different parts of the plant has implications for virus-indexing programs. To allow more reliable detection of Sugarcane mosaic virus (SCMV), the distribution of the virus in sugarcane plants after artificial inoculation was studied using a reverse transcription polymerase chain reaction (RT-PCR) assay. Leaves of susceptible and moderately resistant sugarcane were mechanically inoculated with SCMV 6 weeks after planting. Weekly for 8 weeks after inoculation, plants were examined for mosaic symptoms and samples of leaves, roots and tillers were tested by RT-PCR to detect virus. SCMV moved from the point of inoculation to younger leaves, roots and tillers and eventually to leaves that emerged prior to inoculation. The pattern of SCMV distribution in moderately resistant and susceptible cultivars was not substantially different. However, the virus moved more slowly in the moderately resistant than in the susceptible cultivar. Young leaves proved to be the most suitable tissue for testing.

Effect of storage time and conditions on the cotyledon cell wall of the adzuki bean (Vigna angularis)

Two varieties of adzuki grown in Australia, Bloodwood and Erimo, were stored for up to 6 months at three temperatures (10, 20 and 30 degreesC), and two relative humidities (RH; 40 and 65%). The amount of cell wall material increased with time under all storage conditions. This increase was greatest at 30 degreesC and 40% RH. Storage time and conditions did not affect the total pectin levels in the cell wall. Erimo constantly exhibited a higher total pectin level than Bloodwood. The Bloodwood soluble pectin, Ca++ and Mg++ and Erimo Ca++ in the cell wall remained stable during storage, while the Erimo soluble pectin and Mg++ exhibited a slight decrease at 20 and 30 degreesC after 3 months of storage. (C) 2002 Elsevier Science Ltd. All rights reserved.

Influence of shapes of selected vegetable materials on drying kinetics during fluidized bed drying

Three different particular geometrical shapes of parallelepiped, cylinder and sphere were taken from cut green beans (length:diameter = 1:1, 2:1 and 3:1) and potatoes (aspect ratio = 1:1, 2:1 and 3:1) and peas, respectively. Their drying behaviour in a fluidised bed was studied at three different drying temperatures of 30, 40 and 50 degreesC (RH = 15%). Drying curves were constructed using non-dimensional moisture ratio (MR) and time and their behaviour was modelled using exponential (MR = exp(-kt)) and Page (MR = exp(-kt(n))) models. The effective diffusion coefficient of moisture transfer was determined by Fickian method using uni- and three-dimensional moisture movements. The diffusion coefficient was least affected by the size when the moisture movement was considered three-dimensional, whereas the drying temperature had a significative effect on diffusivity as expected. The drying constant and diffusivity coefficients were on the descending order for potato, beans and peas. The Arrhenius activation energy for the peas was also highest, indicating a strong barrier to moisture movement in peas as compared to beans and skinless cut potato pieces. (C) 2003 Elsevier Science Ltd. All rights reserved.

Formas de aplicação de inoculante e seus efeitos sobre a nodulação da soja

Atualmente, tem-se difundido a aplicação de inoculante no sulco de semeadura na cultura da soja, mas há poucas informações que dão suporte a essa prática e comprovam sua eficiência em diferentes ambientes manejados sob plantio direto. Este trabalho teve como objetivo avaliar a viabilidade da aplicação de inoculantes na cultura da soja, via semente e sulco de semeadura, em solo já cultivado ou não com soja. Foram realizados dois experimentos em campo a partir de dezembro de 2004 em Latossolo Vermelho-Amarelo, seguindo o mesmo método e tratamentos, porém em dois locais distintos, com ou sem cultivo anterior de soja. Foram testados oito tratamentos: (1) inoculação via semente (inoculante + fungicida + micronutriente); (2) sem inoculação (fungicida + micronutriente); (3) testemunha (semente pura, sem tratamento); (4) aplicação no sulco-dose 1 (dose do inoculante recomendada no sulco); (5) aplicação no sulco-dose 2 (duas vezes a dose recomendada no sulco); (6) aplicação no sulco-dose 3 (três vezes a dose recomendada no sulco); (7) sulco-dose 1 + inoculação via semente; e (8) adubação com N (200 kg ha-1 N). Foram avaliados massa de matéria seca de nódulos e número de nódulos totais e nódulos viáveis e não-viáveis aos 30 e 75 dias após emergência. A melhor nodulação foi obtida com aplicação de inoculante + fungicida + micronutriente via semente no solo ainda não cultivado. No solo previamente cultivado com soja, destacaram-se os tratamentos uma e duas vezes a dose do inoculante no sulco. Menores valores de massa seca de nódulos na soja foram obtidos no tratamento com adubação mineral. A aplicação via sulco do inoculante mostrou-se uma prática viável, em razão da semelhança dos resultados obtidos com a aplicação tradicional via semente.

Toxicidade de óleo de mamona a Helicoverpa zea e a Trichogramma pretiosum

O objetivo deste trabalho foi avaliar os efeitos do óleo de mamona sobre a broca-grande do tomateiro, Helicoverpa zea, e sobre o parasitoide de ovos Trichogramma pretiosum. Foi determinada a mortalidade larval da broca, que foi submetida ao óleo de mamona por ingestão e contato às concentrações 0,5, 1,0, 1,5, 2,0, 2,5 e 3,0% (v v-1). A ação do óleo de mamona sobre T. pretiosum foi determinada mediante testes de seletividade e suscetibilidade. As maiores mortalidades de larvas foram observadas no tratamento por contato com as menores concentrações (0,5 e 1,0%), enquanto com as concentrações intermediárias (1,5; 2,0 e 2,5%), não houve diferença entre as vias de aplicação. À maior concentração testada (3,0%), o tratamento por ingestão proporcionou mortalidade superior àquela obtida por contato. O número de ovos parasitados por T. pretiosum, no teste de seletividade, foi afetado negativamente pelo óleo de mamona, contudo, os demais parâmetros avaliados quanto à seletividade e à suscetibilidade não foram afetados. O óleo de mamona reduz a sobrevivência de larvas de H. zea tanto por ingestão quanto por contato. Além disso, não prejudica o desenvolvimento de T. pretiosum, desde que as pulverizações sejam realizadas após as liberações do parasitoide.

Crescimento, produção e rendimento dos clones da variedade ‘Conilon Vitória’ em condições de déficit hídrico e irrigado

O Estado do Espírito Santo é o maior produtor de grãos de café conilon do Brasil. Devido aos trabalhos na seleção de plantas produtivas, os avanços na produtividade desta cultura são mais pronunciados com a utilização da técnica da irrigação. A utilização da irrigação na cafeicultura possibilita a adoção de novas tecnologias de plantio, expansão da área produtiva, uniformização da produção diminuindo os grãos chochos e mal granados, além de mitigar os problemas da deficiência hídrica nos períodos críticos da cultura. Diante do exposto, objetivou-se com este estudo, avaliar o crescimento, a produção e o rendimento dos treze clones da variedade de café ‘Conilon Vitória’, em condições de déficit hídrico e irrigado. O experimento foi instalado no ano de 2010 no Instituto Federal do Espírito Santo, Campus Alegre, em esquema de parcelas subdivididas 3 x 13, sendo nas parcelas o manejo de irrigação em três níveis (irrigado, irrigado com déficit hídrico de 50% e sem irrigação) e nas subparcelas os clones da variedade ‘Conilon Vitória’ em treze níveis, em um delineamento em blocos casualizados, com três repetições. Cada parcela experimental foi constituída por cinco plantas úteis. O manejo da irrigação adotado foi via clima, onde a ETo foi estimada por meio da equação de Hargreaves e Samani (1985). Os dados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey para comparações entre o fator manejo e Scott-Knott para comparações entre o fator clone. O crescimento dos ramos das plantas foi diferenciado em relação ao manejo de irrigação aplicado. Plantas sem irrigação apresentaram maiores picos de crescimento. Plantas irrigadas apresentaram menores picos de crescimento dos ramos, porém, com maior período de crescimento. A produtividade média de plantas irrigadas foi cerca de 474,5% superior a plantas sem irrigação. A produtividade média de plantas irrigadas com déficit hídrico de 50% foi cerca de 329,6 % superior a plantas sem irrigação. O rendimento entre plantas irrigadas e irrigadas com déficit hídrico de 50% não diferiu. Já plantas sem irrigação apresentaram menor rendimento em todos os clones. A irrigação influenciou no tamanho dos grãos, plantas irrigadas apresentaram maiores percentagens de grãos retidos em peneiras 13 e superiores. Já plantas sem irrigação apresentaram maiores percentagens de grãos retidos na fundagem.

Crescimento, produção e rendimento dos clones da variedade ‘Conilon Vitória’ em condições de déficit hídrico e irrigado

O Estado do Espírito Santo é o maior produtor de grãos de café conilon do Brasil. Devido aos trabalhos na seleção de plantas produtivas, os avanços na produtividade desta cultura são mais pronunciados com a utilização da técnica da irrigação. A utilização da irrigação na cafeicultura possibilita a adoção de novas tecnologias de plantio, expansão da área produtiva, uniformização da produção diminuindo os grãos chochos e mal granados, além de mitigar os problemas da deficiência hídrica nos períodos críticos da cultura. Diante do exposto, objetivou-se com este estudo, avaliar o crescimento, a produção e o rendimento dos treze clones da variedade de café ‘Conilon Vitória’, em condições de déficit hídrico e irrigado. O experimento foi instalado no ano de 2010 no Instituto Federal do Espírito Santo, Campus Alegre, em esquema de parcelas subdivididas 3 x 13, sendo nas parcelas o manejo de irrigação em três níveis (irrigado, irrigado com déficit hídrico de 50% e sem irrigação) e nas subparcelas os clones da variedade ‘Conilon Vitória’ em treze níveis, em um delineamento em blocos casualizados, com três repetições. Cada parcela experimental foi constituída por cinco plantas úteis. O manejo da irrigação adotado foi via clima, onde a ETo foi estimada por meio da equação de Hargreaves e Samani (1985). Os dados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey para comparações entre o fator manejo e Scott-Knott para comparações entre o fator clone. O crescimento dos ramos das plantas foi diferenciado em relação ao manejo de irrigação aplicado. Plantas sem irrigação apresentaram maiores picos de crescimento. Plantas irrigadas apresentaram menores picos de crescimento dos ramos, porém, com maior período de crescimento. A produtividade média de plantas irrigadas foi cerca de 474,5% superior a plantas sem irrigação. A produtividade média de plantas irrigadas com déficit hídrico de 50% foi cerca de 329,6 % superior a plantas sem irrigação. O rendimento entre plantas irrigadas e irrigadas com déficit hídrico de 50% não diferiu. Já plantas sem irrigação apresentaram menor rendimento em todos os clones. A irrigação influenciou no tamanho dos grãos, plantas irrigadas apresentaram maiores percentagens de grãos retidos em peneiras 13 e superiores. Já plantas sem irrigação apresentaram maiores percentagens de grãos retidos na fundagem.

Damage level of the two-spotted spider mite Tetranychus urticae Koch (acari: tetranychidae) in soybeans

Among phytophagous spider mites, the two-spotted spider mite Tetranychus urticae Koch, 1836 is one of the most important agricultural pests, not only because of the damage it causes, but also because it has a wide host range, infesting many commercial crops such as leafy greens, cotton, beans, and soybeans, among others. This study was carried out in a greenhouse of the Faculdade de Ciências Agrárias (FCA) of the Universidade Federal da Grande Dourados (UFGD), located in the city of Dourados, state of Mato Grosso do Sul. The experiment was arranged in a randomized block design with 5 treatments and 4 replicates. The treatments consisted of 5 levels in percentage of chlorotic symptoms (indicating mite damage): 0%, 25%, 50%, 75%, and 100%. All of the characteristics evaluated, except for number of pods per plant, the number of seeds per plant, the total weight (productivity), and the weight of 1000 seeds, were significantly influenced by the different levels of chlorotic symptoms. The economic damage level for the two-spotted spider mite Tetranychus urticae, according to the equation y = 66.63-0.51 x, based on the price of US$ 11.00 per bag of soybeans and a control cost of US$ 16.00, would be 15.80% chlorotic symptoms. At a price of US$ 29.00 per bag with the same control cost, the economic damage level would be 13% of chlorotic symptoms.