Diversity Symbiotic And Plant Growth Promoting Properties Of Rhizobia And Rhizobacteria Of Grass Pea (lathyrus Sativus L.) From Central Ethiopia Implication To The Selection And Use Of Microbial Inoculants In Low Input Agricu

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Grass pea (Lathyrus sativus L.) is widely cultivated for food and feed in some developingrncountries including Ethiopia. However, its alkaloid content did not attract attention to thernresearch community for its over-exaggerated neuro-lathyrism causing paralysis of limbs makingrnit as one of the most neglected orphan crops in the world. However, the crop is considered as anrninsurance crop by resource-poor farmers for it produces reliable yields when all other crops fail.rnThe increase in production in several areas in Ethiopia presupposes that it is an important sourcernof protein to the diet of the population and makes it the fifth most important pulse crops inrnEthiopia after faba bean, chickpea, field pea and haricot bean in terms of production. Thisrnindicates the necessity of looking beyond the prejudices held on the crop for long to expand thernfrontier of food security and soil fertility against the changing climate in the country. To this end,rna study was made to characterize and select rhizobial and rhizobacteria from grass pea fromrncentral Ethiopia using standard methods. Isolates were tested for their phenotypic, symbiotic,rngenetic, in vitro stress tolerance, variations in substrate utilization and PGP properties within therncontext of their taxonomic characteristics and selection of tolerant isolates for field applications.rnBased on their preliminary symbiotic effectiveness, stress tolerance (pH tolerance, intrinsicrnantibiotic resistance (IAR)), PGP (phosphate solubilization, indole acetic acid production, etc),rnand antagonistic properties (suppression of fungal pathogen, etc), four rhizobial and twornrhizobacteria isolates were selected and tested for single and co-inoculation trials of the crop onrnsoil culture under greenhouse conditions. The data showed that all but one rhizobium isolates (49rnisolates) were authenticated as root nodule bacteria (renodulating the host variety upon reinoculation)rnand 86% of the isolates accumulated more than 50% of the shoot dry weight (SDW)rn(0.273-1.148g/p in relation to N-fetilized control plants indicator of symbiotic effectivenessrn(SE%) in nitrogen fixation. The inoculated plants also showed variations in nodule numberrn(NN) ranging from 17.7-116/plant, Nodule dry weight (NDW) 0.011g-0.098/plant indicating arnseven-fold and ten-fold difference among the isolates. Among the highly symbiotic effectivernisolates (SDW of >80%), isolates AAUGR- 2, 5, 6, 9, 11, 14, 15, 19, 20, 24, 30 and 50 acquiredrna wide range of substrate utilization and ecological tolerance under in vitro-conditions and thernxvrnmajority of these isolates showed multiple plant growth promoting properties (MPGP) (4-8)rnranging from good potential of phosphate solubilization and suppression of fungal pathogens.rnThe PCR amplification of 16SRNA and nif-H genes showed that the majority of the isolatesrn(87%) showed 99% sequence homology with Rhizobium spp. obtained from NCBI gene datarnbase and a few isolates (AAUGR-2, AAUGR-24) were grouped into Ensifer meliloti andrnRhizobium leguminosarum var viceae indicating that grass pea rhizobia were more diverse thanrnthe hitherto established cross-nodulation grouping to Rhizobium leguminosarum var viceae. Allrnthe selected rhizobial isolates were highly significant (P=0.000) in their ability to solubilize TCPrnwith zones of solubilisation (solubilization index; SI) ranging from 1.24- 3.37 cm and therncapacity of producing organic acids within the range of 256- 418 μg/ml. The highest amount ofrnIAA (74.69±1.72 μg/ml) was produced by Rhizobium species (AAUGR-14) followed byrn74.18±1.95 μg/ml produced by AAUGR-30. Regarding the antagonistic properties, all isolatesrnproduced catalase and ammonia whereas, a number of isolates were positive for chitinase,rnprotease, cellulase, HCN and in vitro fungal inhibition assay. Among a total of 100 isolates ofrnrhizobacteria, 39% were phosphate solubilizers of which 22 isolates that showed SI of greaterrnthan 2.4 cm were selected for further screening process for multiple growth promoting propertiesrn(MPGP). The 16S rRNA gene sequence analysis of five rhizobacterial isolates, AAUGPR- 38,rn53, 73, 91 and 92 showed 94- 99 % identity to different rhizobacterial genera includingrnEnterobacter, Enterococcus, Kluyvera, pantoea and Serratia type. Under the circumstancesrnmost of them displayed 5-8 MPGP of which AAUGPR-53 identified as Enterococcus species,rnEnterococcus casseliflavus strain LAHHAB-24 and Enterococcus gallinarum strain F1 showedrnthe highest phosphate solubilization index (PSI) and IAA production efficiency of 4.81±0.02rn(cm) and 56.55±0.45 (μg/ml), respectively. Seventeen (77.3%) of the isolates showed in vitrornantifungal inhibition against Fusarium oxysporum f. sp. lentis with isolates AAUGPR- 92 and 91rnidentified as Enterococcus species, Enterococcus casseliflavus and Enterococcus gallinarumrnexhibiting a percent radial growth inhibition of 73 and 83%, respectively. In general, 46%-100%rnof the isolates showed variations in their ability to produce chitinase, protease, cellulase, HCNrnand NH3 associated with antagonistic properties of the isolates. With regard to the performancernof the selected inoculants to inoculation and coinoculation result, Rhizobium. sp. 14 performedrnxvirnbest in all single and co-inoculation trials with E. casseliflavus/gallinarum- 92. In general, thernmixed inoculants; Rhizobium sp. 6, R. sp. 9, R. sp. 11, R. sp. 14 and E. casseliflavus/gallinarum-rn53 & 92 displayed the highest performance in all parameters. In all cases, treatments increasedrnnodulation, growth and symbiotic effectiveness over the negative and positive controls. Thernmixed inoculants can be recommended as good candidates for plant growth improvement trialsrnand validation under field conditions to exploit the crop for its multipurpose use with higherrntolerance under extreme environmental factors, higher protein content and high yield under lowrnsoil fertility/input in the country.

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Diversity Symbiotic And Plant Growth Promoting Properties Of Rhizobia And Rhizobacteria Of Grass Pea (lathyrus Sativus L.) From Central Ethiopia Implication To The Selection And Use Of Microbial Inoculants In Low Input Agricu

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