رسائل ماجيستير
English
2011
Improvement of Some Traits of Rhizobia Specific to Peanut (Arachis hypogaea)
أشرف بكري عبد الرازق, ابراهيم حلمي عباس, سهام محمد شاش, فيروز حسن عبد المجيد , محمود عبد المقصود ناصف
كلية العلوم-جامعة بنها · مصر
Legumes, including peanut are essential components for sustaining productivity in many agricultural systems. A hallmark benefit of legumes use in many agricultural systems is their ability to develop root nodules and to fix N2in symbiosis with compatible rhizobia. Peanut (Arachis hypogaea) is one of the most important oil crops in Egypt especially in newly reclaimed soils. Its economically importance make the cultivated areas expanded yearly. Soil born fungal peanut diseases especially infections with Aspergillus flavuswhich resulted in aflatoxin contamination is a major problem affecting peanut production in Egypt. Chemical fungicides are commonly used to protect seeds and seedlings from fungal diseases However, these fungicides adversely affect rhizobial inoculants efficiency as well as its hazardous effect on environment and human and animal health. Biological control can be used as a powerful alternative solution for chemical control of such fungal diseases. Rhizobacteria which have the ability to antagonize and inhibit these fungal pathogens are considered effective bio-agents. In addition to their antagonistic activity, rhizobacteria are used as powerful plant growth promoting bacteria. This study aimed at improvement of peanut rhizobial inoculants through acquiring new antagonistic traits using conjugation between efficient peanut nodulated rhizobia and effective rhizobacteria. The results of this study can be summarized as follows:1-The symbiotic effectiveness of all tested rhizobial isolates/ strains was evaluated with different peanut cultivars grown in Egypt under greenhouse conditions. The pot experiments involved two Bradyrhizobiumstrains (USDA 3456 and USDA
-97 -3339), four local isolates (ARC 601, ARC 617, isolate 1 and isolate 2) and six peanut cultivars (Early bunch, Gregory, NC, Giza 5, Giza 6and Ismailia 1). 2-The pot experiment revealed results showed that all tested rhizobial strains/ isolates exhibited prolific nodulation patterns with the local peanut cultivars (Giza 5, Giza 6 and Ismailia 1),. Rhizobial treatments showed total nodules dry weight ranged from 75.0 to 252.5 mg/plant, whereas the un-inoculated control did not form any nodules. The obtained results showed significant increases in plants shoot dry weight and shoot N-content with a range of 4.8-60.5% and 5.9-99.8%, respectively compared to the un-inoculated control3-All tested rhizobial strains /isolates had a positive effect with the foreign peanut cultivars (Gregory, NC and Early bunch). All tested rhizobia displayed different nodulation patterns with dry weight of nodules ranged from 104.9 to 235.3 mg/ plant compared to the un-inoculated control which did not form any nodules. The obtained results revealed significant increases in plants shoot dry weight and shoot N-content with a range of 8-62 % and 22 -123%, respectively compared to the un-inoculated control4-All bradyrhizobial treatments showed significant differences compared to the un-inoculated control in respect to peanut plants growth parameters and their shoot N2contents. Also, the results clarified that the Bradyrhizobium strain USDA 3456 was the highly efficient strain with all tested peanut cultivars than other tested rhizobial strains/ isolates.
-98 -5-The antagonistic activity of five isolates of Serratia marcescen ssp. (SER4, EG10, GT, BF9 and BEF91) and three isolates of Pseudomona ssp. (P. putida, P. aeruginosa and P. fluorescens) were tested in-vitro, by assaying their ability to inhibit the mycelial growth of Aspergillus flavus fungus.6-The results demonstrated that GT isolate of S. marcescenssp. and P. putidasp. exhibited the highly antagonistic effect against A. flavus; they had the largest inhibition zones compared to control. On the otherhand, other tested isolates of S. marcescenssp. and Pseudomonassp. did not show any antagonistic effect against fungal growth.7-The conjugation process between the highly efficient N2fixing bradyrhizobia, USDA 3456 as a recipient and the most antagonistic rhizobacteria, S. marcescens(GT) and P. putida as donors was carried out.8-The three bacterial strains/ isolates used in conjugation were genetically marked by four antibiotics (ampicillin, chloramphenicol, neomycin sulphate and streptomycin). The results demonstrated that all bacterial strains/ isolates were sensitive to ampicillin and resistance to neomycin sulphate. The results also clarified that bradyrhizobial strain USDA 3456 was sensitive to streptomycin and resistant to chloramphenicol. On the contrary, Serratia sp. (GT) and P. putida sp. were resistant to streptomycin and sensitive to chloramphenicol.9-Two bradyrhizobial transconjugants, DiM 71 (conjugation between Bradyrhizobium USDA 3456 and S. marcescensGT) and DiM 73 (conjugation between Bradyrhizobium USDA 3456 and P. putida) were obtained.
-99 -10-The symbiotic effectiveness and antagonistic activity of the DiM 71 and DiM 73 were evaluated. The obtained results showed that both transconjugants acquired the antagonistic traits as well as maintained their N2fixation effectiveness. The results also clarified that, there was no significant difference between the parent Bradyrhizobiumstrain USDA 3456 and the new transconjugants in respect to their symbiotic effectiveness. The results also revealed that the obtained transconjugants exhibited highly antagonistic effect against peanut fungal pathogen, A. flavus in-vitro.11-One-dimensional protein analysis of all tested rhizobial isolates/ strains was carried out using SDS-PAGE. The electrophoretic analysis showed a high level similarity among all tested bradyrhizobia however; several discriminating protein bands could differentiate between the tested rhizobia. Strain USDA 3456 along with isolate 1, ARC 601 and 617 were ranked in a separate SDS-PAGE cluster, while strain USDA 3339 and isolate 2 were grouped in another cluster.12-The conjugation process was tracked on proteomic level using SDS-PAGE. The electrophoretic analysis of the new transconjugants (DiM 71 and DiM 73) expressed different proteins of their related parents (Bradyrhizobium USDA 3456, S. marcescens GT and P. putida sp.), individually. However, intensive level of similarity was found between the new transconjugants (DiM 71 and DiM 73) and their donor Bradyrhizobium strain, USDA 3456