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An investigation of the phenotypic characteristics of 19 rhizobial strains, isolated from root nodules of different plant legumes grown in the soil of agriculture farms in Ismailia governorate, transpired. Most isolates were creamy or white opaque, mucoid, with a convex elevation, translucent, and smooth margined. Microscopic investigation revealed that all bacterial isolates were rod-shaped and had no positive affinity for Gram-stain. Identifying rhizobial cultures from any bacterial contaminants employed confirmatory tests based on prepared special media, including YMA supplement with Congo red, glucose peptone agar, Kit-lactose agar, and Hoffer's alkaline test. Based on an infectivity test, all isolates proved their ability to reinfect their host. These rhizobial isolates, classified into two categories, included fast and slow-growing rhizobia according to their growth in the YEM medium containing bromothymol blue (BTB). Meanwhile, the assessment of the genetic diversity among these isolates proceeded using ISSR and RAPD markers, which ISSR marker proved a more powerful tool in discriminating among the tested isolates than the RAPD marker. The cluster analysis, with the RAPD marker, classified the isolates into two main groups. The first group included the isolates (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 13), while the second group contained the isolates (12, 14, 15, 16, 17, 18, and 19). Moreover, using ISSR markers also showed a cluster of two main groups with diverse categories; the first cluster included isolates 1 to 11, and the second group contained isolates 12 to 19. [ABSTRACT FROM AUTHOR]

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Legume root nodules harbor rhizobia and other non-nodulating endophytes known as nodule-associated bacteria (NAB) whose role in the legume symbiosis is still unknown. We analysed the genetic diversity of 34 NAB isolates obtained from the root nodules of faba bean grown under various soil conditions in Egypt using 16S rRNA and concatenated sequences of three housekeeping genes. All isolates were identified as members of the family Enterobacteriaceae belonging to the genera Klebsiella , Enterobacter and Raoultella. We identified nine enterobacterial genospecies, most of which have not been previously reported as NAB. All isolated strains harbored nifH gene sequences and most of them possessed plant growth-promoting (PGP) traits. Upon co-inoculation with an N2 fixing rhizobium (Rlv NGB-FR128), two strains (Enterobacter sichanensis NGB-FR97 and Klebsiella variicola NGB-FR116) significantly increased nodulation, growth and N-uptake of faba bean plants over the single treatments or the uninoculated control. The presence of these enterobacteria in nodules was significantly affected by the host plant genotype, symbiotic rhizobium genotype and endophyte genotype, indicating that the nodule colonization process is regulated by plant–microbe–microbe interactions. This study emphasizes the importance of nodule-associated enterobacteria and suggests their potential role in improving the effectiveness of rhizobial inoculants. [ABSTRACT FROM AUTHOR]

In the last few decades, there has been a growing interest in environmentally friendly sustainable agricultural practices, thus increasing the role of biofertilizers such as rhizobia, which can decrease the need for chemical fertilizers, reduce adverse environmental effects, and help to save money. Therefore, information on the distribution and genetic variation of native rhizobial isolates would aid in selecting novel rhizobial strains that could be developed and used as biofertilizers in legume production. This research was conducted to characterize 24 rhizobial isolates from five legumes on morphological, biochemical, and molecular aspects and determine the phylogenetic relationships among them. Rhizobial isolates were obtained from five Egyptian legumes: faba bean, lentil, pea, clover, and soybean. Morphological characterization classified the isolates into fast and slow growers. Biochemical characterization using API 20E and API 20NE systems showed a large diversity, which may reflect their adaptation in different environments. Moreover, molecular detection of the 16S rRNA gene enabled to characterize 19 of them to the species level. Rhizobial isolates from pea, faba bean, clover, and lentil were identified as Rhizobium leguminosarum and those from soybean were identified as Bradyrhizobium japonicum. These data reflected a narrow diversity of rhizobial species in Egypt. A phylogenetic analysis of the 19 isolates confirmed that B. japonicum isolates were divergent from all other isolates. Furthermore, the phylogram revealed that each group of isolates that originated from the root nodule of a certain legume formed a separate subcluster. The obtained data suggested a narrow range of interspecies variations, which is consistent with the idea of the presence of biovars among the species. [ABSTRACT FROM AUTHOR]

In the present work we analyzed the taxonomic status of several Rhizobium strains isolated from Trifolium alexandrinum L. nodules in Egypt. The 16S rRNA genes of these strains were identical to those of Rhizobium bangladeshense BLR175 T and Rhizobium binae BLR195 T . However, the analyses of recA and atpD genes split the strains into two clusters. Cluster II strains are identified as R. bangladeshense with >98% similarity values in both genes. The cluster I strains are phylogenetically related to Rhizobium etli CFN42 T and R. bangladeshense BLR175 T , but with less than 94% similarity values in recA and atpD genes. DNA–DNA hybridization analysis showed 42% and 48% average relatedness between the strain 1010 T from cluster I with respect to R. bangladeshense BLR175 T and R. etli CFN42 T , respectively. Phenotypic characteristics of cluster I strains also differed from those of their closest related Rhizobium species. Analysis of the nodC gene showed that the strains belong to two groups within the symbiovar trifolii which was identified in Egypt linked to the species R. bangladeshense . Based on the genotypic and phenotypic characteristics, the group I strains belong to a new species for which the name Rhizobium aegyptiacum sp. nov. (sv. trifolii) is proposed, with strain 1010 T being designated as the type strain (= USDA 7124 T = LMG 29296 T = CECT 9098 T ). [ABSTRACT FROM AUTHOR]

Abstract: Twenty rhizobial strains isolated from the root nodules of soybean (Glycine max L.) were collected from nine governorates representing different agro-climatic and soil conditions in Egypt. The strains were characterized using a polyphasic approach, including nodulation pattern, phenotypic characterization, 16S rDNA sequencing, nifH and nodA symbiotic genes sequencing, and rep-PCR fingerprinting. Symbiotic properties assay revealed that all local rhizobial strains showed a wide spectrum of prolific nodulation and a marked increase in plant growth parameters compared to the un-inoculated control. Complete sequencing of 16S rRNA demonstrated that, native soybean nodulating rhizobia are phylogenetically related to Bradyrhizobium, Ensifer and Rhizobium (syn. Agrobacterium) genera. Study of tolerance ability to environmental stresses revealed that local strains survived in a wide pH ranges (pH 5–11) and a few of them tolerated high acidic conditions (pH 4). Agrobacterium strains were identified as the highest salt-tolerant and were survived under 6% NaCl, however Ensifer strains were the uppermost heat-tolerant and can grow at 42 °C. Agrobacterium strains have been shown to harbor nifH and nodA genes similar to those in other fast growing soybean symbionts and were largely distinct from symbiotic genes of slow growing bradyrhizobia. The symbiotic effectiveness stability of Agrobacterium strains to nodulate soybean roots was confirmed using plant nodulation assay. [Copyright &y& Elsevier]

Abstract: Snap bean fields in 12 of the 25 governorates of Egypt were surveyed to determine the distribution and taxonomy of snap bean-nodulating rhizobia. Nodulation rates in the field were very low, indicating that Egyptian soils do not have sufficient numbers of snap bean-compatible Rhizobium spp. A total of 87 rhizobial isolates were assayed on the most commonly grown cultivars in order to identify the most effective strains. The five most effective isolates (R11, R13, R28, R49 and R52) were fast-growing and utilized a wide range of carbon and nitrogen sources. A phylogenetic assignment of these strains by analysis of the 16S ribosomal RNA gene suggested that all fell within the Rhizobium etli–Rhizobium leguminosarum group. Strains R11, R49 and R52 all clustered with other identified R. etli strains, while strains R13 and R28 were more distinct. The distinctness of R13 and R28 was supported by physiological characteristics, such as their ability to utilize citrate, erythritol, dulcitol and lactate. Strains R13 and R28 also yielded the highest plant nitrogen content of all isolates. The highly effective strains isolated in this study, in particular strains R13 and R28, are promising candidates for improving crop yields. The data also suggested that these two strains represented a novel sub-group within the R. etli–R. leguminosarum group. As snap bean is a crop of great economic value to Egypt, the identification of highly effective rhizobial strains adapted to Egyptian soils, such as strains R13 and R28, is of great interest. [Copyright &y& Elsevier]

Berseem clover (T. alexandrinum) is the main forage legume crop used as animal feed in Egypt. Here, eighty rhizobial isolates were isolated from root nodules of berseem clover grown in different regions in Egypt and were grouped by RFLP-16S rRNA ribotyping. Representative isolates were characterized using phylogenetic analyses of the 16S rRNA, rpoB , glnA , pgi, and nodC genes. We also investigated the performance of these isolates using phenotypic tests and nitrogen fixation efficiency assays. The majority of strains (<90%) were closely related to Rhizobium aegyptiacum and Rhizobium aethiopicum and of the remaining strains, six belonged to the Rhizobium leguminosarum genospecies complex and only one strain was assigned to Agrobacterium fabacearum. Despite their heterogeneous chromosomal background, most of the strains shared nodC gene alleles corresponding to symbiovar trifolii. Some of the strains closely affiliated to R. aegyptiacum and R. aethiopicum had superior nodulation and nitrogen fixation capabilities in berseem clover, compared to the commercial inoculant (Okadein®) and N-added treatments. R. leguminosarum strain NGB-CR 17 that harbored a nodC allele typical of symbiovar viciae, was also able to form an effective symbiosis with clover. Two strains with nodC alleles of symbiovar trifolii, R. aegyptiacum strains NGB-CR 129 and 136, were capable of forming effective nodules in Phaseolus vulgaris in axenic greenhouse conditions. This adds the symbiovar trifolii which is well-established in the Egyptian soils to the list of symbiovars that form nodules in P. vulgaris. [ABSTRACT FROM AUTHOR]

A total of 38 bacterial strains were isolated from root nodules of Lotus , Vicia , Trifolium , Lathyrus , Astragalus , and Hippocrepis plants grown-wild at different geographical locations in Egypt. Analysis of the almost complete sequence of the 16S rRNA gene from the bacterial strains showed that seventeen of them isolated from all the six plant species were members of the genus Rhizobium , six from Lotus , Vicia and Hippocrepis were Ensifer , and the remaining six strains from Lotus and Vicia were Neorhizobium. Sequencing of the symbiotic nodC gene from eight selected rhizobial strains confirmed their affiliation with rhizobial species. These results suggest the existence of high rhizobial diversity in wild-grown Egyptian legumes. Nodules from Lotus , Trifolium , and Astragalus also contained bacterial species of the genera Achromobacter , Ancylobacter , Bacillus , Cronobacter , Curtobacterium , Flavobacterium , Paenibacillus and Pseudomonas. These results provide information about the diversity of rhizobia associated with wild-grown legumes in Egypt, and enlarged the rhizobial spectrum of Lotus , Vicia , Trifolium , Lathyrus , Hippocrepis and Astragalus , which supported to the idea that rhizobial promiscuity could be an adaption strategy to diverse environments. [ABSTRACT FROM AUTHOR]