1. bookVolume 65 (2016): Issue 3 (August 2016)
Journal Details
First Published
04 Mar 1952
Publication timeframe
4 times per year
access type Open Access

Characterization of Rhizobial Bacteria Nodulating Astragalus corrugatus and Hippocrepis areolata in Tunisian Arid Soils

Published Online: 26 Aug 2016
Volume & Issue: Volume 65 (2016) - Issue 3 (August 2016)
Page range: 331 - 339
Received: 13 Jun 2015
Accepted: 11 Feb 2016
Journal Details
First Published
04 Mar 1952
Publication timeframe
4 times per year

Fifty seven bacterial isolates from root nodules of two spontaneous legumes (Astragalus corrugatus and Hippocrepis areolata) growing in the arid areas of Tunisia were characterized by phenotypic features, 16S rDNA PCR-RFLP and 16S rRNA gene sequencing. Phenotypically, our results indicate that A. corrugatus and H. areolata isolates showed heterogenic responses to the different phenotypic features. All isolates were acid producers, fast growers and all of them used different compounds as sole carbon and nitrogen source. The majority of isolate grew at pHs between 6 and 9, at temperatures up to 40°C and tolerated 3% NaCl concentrations. Phylogenetically, the new isolates were affiliated to four genera Sinorhizobium, Rhizobium, Mesorhizobium and Agrobacterium. About 73% of the isolates were species within the genera Sinorhizobium and Rhizobium. The isolates which failed to nodulate their host plants of origin were associated to Agrobacterium genus (three isolates).


Ardley J.K., M.A. Parker, S.E. De Meyer, R.D. Trengove, G.W. O’Hara, W.G. Reeve, R.J. Yates, M.J. Dilworth, A. Willems and J.G. Howieson. 2012. Microvirga lupini sp. nov., Microvirga lotononidis sp. nov., and Microvirga zambiensis sp. nov. areAlphaproteobacterial root nodule bacteria that specifically nodulate and fix nitrogen with geographically and taxonomically separate legume hosts. Int. J. Syst. Evol. Microbiol. 62: 2579–2588. Search in Google Scholar

Ben Romdhane S., H. Nasr, R. Samba-Mbaye, M. Neyra and M. Ghorbal. 2005. Diversity of Acacia tortilis rhizobia revealed by PCR/ RFLP on crushed root nodules in Tunisia. Ann. Microbiol. 55: 249–258. Search in Google Scholar

Benhizia Y., H. Benhizia, A. Benguedouar, R. Muresu, A. Giaco- mini and A. Squartini. 2004. Gammaproteobacteria can nodulate legumes of the genus Hedysarum. Syst. Appl. Microbiol. 27: 462–468.10.1078/0723202041438527Search in Google Scholar

Chen W.M., S. Laevens, T.M. Lee, T. Coenye, P. De Vos, M. Mer- geay and P. Vandamme. 2001. Ralstonia taiwanensis sp. nov., isolated from root nodules of Mimosa species and sputum of a cystic fibrosis patient. Int. J. Syst. Evol. Microbiol. 51: 1729–1735. Search in Google Scholar

Chen W.M., E.K. James, T. Coenye, J.H. Chou, E. Barrios, S.M. De Faria, G.N. Elliott, S.Y. Sheu, J.I. Sprent and P. Van- damme. 2006. Burkholderia mimosarum sp. nov., isolated from root nodules of Mimosa spp. from Taiwan and South America. Int. J. Syst. Evol. Microbiol. 56: 1847–1851. Search in Google Scholar

Chen W.M., S.M. De Faria, J.H. Chou, E.K. James, G.N. Elliott, J.I. Sprent, C. Bontemps, J.P. Young and P. Vandamme. 2008. Burkholderia sabiae sp. nov., isolated from root nodules of Mimosa caesalpiniifolia. Int. J. Syst. Evol. Microbiol. 58: 2174–2179. Search in Google Scholar

de Lajudie P., A. Willems, G. Nick, S.H. Mohamed, U. Torck, R. Coopman, A. Filali-Maltouf, K. Kersters, B. Dreyfus, K. Lind- ström and others. 1999. Agrobacterium bv. 1 strains isolated from nodules of tropical legumes. Syst. Appl. Microbiol. 22: 119–132.10.1016/S0723-2020(99)80035-6 Search in Google Scholar

Elkan G.H. 1992. Taxonomy of the rhizobia. Can. J. Microbiol. 38: 446–450.10.1139/m92-075 Search in Google Scholar

Fterich A., M. Mahdhi, M.A. Caviedes, E. Pajuelo, R. Rivas, I.D. Rodriguez-Llorente and M. Mars. 2011. Characterization of root-nodulating bacteria associated to Prosopis farcta growing in the arid regions of Tunisia. Arch. Microbiol. 193: 385–397.10.1007/s00203-011-0683-z Search in Google Scholar

Gao J.L., Z.D. Terefework, W.X. Chen and K. Lindström. 2001. Genetic diversity of rhizobia isolated from Astragalus adsurgens growing in different geographical regions of China. J. Biotechnol. 91: 155–168.10.1016/S0168-1656(01)00337-6 Search in Google Scholar

Gao J.L., S.L. Turner, F.L. Kan E.T Wang, Z.Y. Tan, Y.H. Qiu, J. Gu, Z. Terefework, J.P.W. Young, K. Lindström and others. 2004. Mesorhizobium septentrionale sp. nov. and M. temperatum sp. nov., isolated from Astragalus adsurgens growing in the northern regions of China. Int. J. Syst. Evol. Microbiol. 54: 2003–2012. Search in Google Scholar

Gnat S., M. Wójcik, S. Wdowiak-Wróbel, M. Kalita, A. Ptaszyńska and W. Małek. 2014. Phenotypic characterization of Astragalus glycyphyllos symbionts and their phylogeny based on the 16S rDNA sequences and RFLP of 16S rRNA gene. Antonie Van Leeuwenhoek. 105: 1033–1048.10.1007/s10482-014-0163-y401983124710996 Search in Google Scholar

Graham P.H., M.J. Sadowsky, H.H. Keiser, Y.M. Barnet, R.S. Bradley, J.E. Cooper, D.J. De Ley, B.D.W. Jarvis, E.B. Roslycky, B.W. Strijdom and others. 1991. Proposed minimal standards for the description of new genera and species of root and stem-nodu- lating bacteria. Int. J. Syst. Bacteriol. 41: 582–587.10.1099/00207713-41-4-582 Search in Google Scholar

Guerrouj K., E. Pe’rez-Valera, R. Chahboune, H. Abdelmoumen, E.J. Bedmar M.M. El Idrissi. 2013. Identification of the rhizobial symbiont of Astragalus glombiformis in Eastern Morocco as Meso- rhizobium camelthorni. Antonie van Leeuwenhoek. 104: 187–198.10.1007/s10482-013-9936-y23673873 Search in Google Scholar

Gurtler V., V.A. Wilson and B.C. Mayall. 1991. Classification of medically important clostridia using restriction endonuclease site differences of PCR-amplified 16S rDNA. J. Gen. Microbiol. 137: 2673–2679.10.1099/00221287-137-11-26731783911 Search in Google Scholar

Jaftha J.B., B.W. Strijdom and P.L. Steyn. 2002 Characterization of pigmented methylotrophic bacteria which nodulate Lotononis bainesii. Syst. Appl. Microbiol. 25: 440–449.10.1078/0723-2020-0012412421082 Search in Google Scholar

Jourand M., E. Giraud, G. Béna, A. Sy, A. Willems, M. Gillis, B. Dreyfus and P. De Lajudie. 2004. Methylobacterium nodulans sp. nov., for a group of aerobic, facultatively methylotrophic, legume root-nodule forming and nitrogen-fixing bacteria. Int. J. Syst. Evol. Microbiol. 54: 2269–2273. Search in Google Scholar

Karanja, N.K. and M. Wood. 1988. Selecting Rhizobium phaseoli strains for use with beans (phaseolus vulgaris L.) in Kenya: tolerance of high temperature and antibiotic resistance. Plant Soil. 112: 15–22.10.1007/BF02181747Search in Google Scholar

Kimura M. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J. Mol. Evol. 16: 111–120.10.1007/BF017315817463489 Search in Google Scholar

Klonowska A., C. Chaintreuil, P. Tisseyre1, L. Miche, R. Melko- nian, M. Ducousso, G. Laguerre, B. Brunel and L. Moulin. 2012. Biodiversity of Mimosa pudica rhizobial symbionts (Cupriavidus taiwanensis, Rhizobium mesoamericanum) in New Caledonia and their adaptation to heavy metal-rich soils. FEMS Microbiol. Ecol. 81: 618–635. Search in Google Scholar

Kumar S., K. Tamura, I.B. Jakobsen and M. Nei. 2001. MEGA2: Molecular Evolutionary Genetics Analysis software. Bioinformatics. 17: 1244–1245.10.1093/bioinformatics/17.12.124411751241 Search in Google Scholar

Lin D.X., E.T. Wang, H. Tang, T.X. Han, Y.R. He, S.H. Guan and W.X. Chen. 2008. Shinella kummerowiae sp. nov., a symbiotic bacterium isolated from root nodules of the herbal legume Kummerowia stipulacea. Int. J. Syst. Evol. Microbiol. 58: 1409–1413. Search in Google Scholar

Liu J., E.T. Wang and W.X Chen. 2005. Diverse rhizobia associated with woody legumes Wisteria sinensis, Cercis. Syst. Appl. Microbiol. 28: 465–477. Search in Google Scholar

Mahdhi M., A. Nzoué, F. Gueye, C. Merabet, P. de Lajudie and M. Mars. 2007. Phenotypic and genotypic diversity of Genista saha- rae microsymbionts from the infra-arid region of Tunisia. Lett. Appl. Microbiol. 54: 604–609.10.1111/j.1472-765X.2007.02233.x17916130 Search in Google Scholar

Mahdhi M., P. de Lajudie and M. Mars. 2008. Phylogenetic and symbiotic characterization of rhizobial bacteria nodulating Argy- rolobium uniflorum in Tunisian arid soils. Can. J. Microbiol. 54: 209–217.10.1139/W07-131 Search in Google Scholar

Mahdhi M., A. Fterich, M. Rejili, I.D. Rodriguez-Llorente and M. Mars. 2012. Legume-Nodulating Bacteria (LNB) from three pasture legumes (Vicia sativa, Trigonella maritima and Hedysarum spinosissimum) in Tunisia. Ann. Microbiol. 62: 61–68. Search in Google Scholar

Małek W. and E. Sajnaga. 1999. Current taxonomy of the rhizobia. Acta. Microbiol. Pol. 48: 109–122. Search in Google Scholar

Mantelin S., M. Fischer-Le Saux, F. Zakhia, G. Béna, S. Bonneau, H. Jeder, P. De Lajudie and J.C. Cleyret-Marel. 2006. Emended description of the genus Phyllobacterium and description of four novel species associated with plant roots: Phyllobacterium bourgo- gense sp. nov., Phyllobacterium ifriqiyense sp. nov., Phyllobacterium leguminum sp. nov. and Phyllobacterium brassicacearum sp. nov. Int. J. Syst. Evol. Microbiol. 56: 827–839. Search in Google Scholar

Mhamdi R., G. Laguerre, M.E. Aouani, M. Mars and N. Amarger. 2002. Different species and symbiotic genotypes of field rhizobia can nodulate Phaseolus vulgaris in Tunisian soils. FEMS Microbiol. Ecol. 41: 77–84.10.1111/j.1574-6941.2002.tb00968.x19709241 Search in Google Scholar

Mhamdi R., M. Mrabet, G. Laguerre, R. Tiwari and M.E. Aouani. 2005. Colonization of Phaseolus vulgaris nodules by Agrobacteriumlike strains. Can. J. Microbiol. 51: 105–111.10.1139/w04-12016091768 Search in Google Scholar

Mohamed S.H., A. Smouni, M. Neyra, D. Kharchaf and A. Filali- Maltouf. 2000. Phenotypic characteristics of root-nodulating bacteria isolated from Acacia spp. grown in Libya. Plant Soil. 224: 171–183.10.1023/A:1004838218642Search in Google Scholar

Moulin L., A. Munive, B. Dreyfus and C. Boivin-Masson. 2001. Nodulation of legumes by members of the beta-subclass of Proteo- bacteria. Nature 411: 948–950.10.1038/3508207011418858 Search in Google Scholar

Mrabet M., B. Mnasri, S. Ben Romdhane, G. Laguerre, M.E. Aouani and R. Mhamdi. 2006. Agrobacterium strains isolated from root nodules of common bean specifically reduce nodulation by Rhizobium gallicum. FEMS Microbiol. Ecol. 56: 304–309.10.1111/j.1574-6941.2006.00069.x16629759 Search in Google Scholar

Muresu R., E. Polone, L. Sulas, B. Baldan, A. Tondello, G. Delogu, P. Cappuccinelli, S. Alberghini, Y. Benhizia, H. Benhizia and others. 2008. Coexistence of predominantly nonculturable rhizo- bia with diverse, endophytic bacterial taxa within nodules of wild legumes. FEMS Microbiol. Ecol. 63: 383–400. Search in Google Scholar

Rejili M., M.J. Lorite, M. Mahdhi, J.S. Pinilla, A. Ferchichi and M. Mars. 2009. Genetic diversity of rhizobial populations recovered from three Lotus species cultivated in the infra-arid Tunisian Soils. Progress in Natural Science 19: 1079–1087.10.1016/j.pnsc.2009.02.003 Search in Google Scholar

Rejii M., M. Mahdhi, A. Fterich, S. Dhaoui, I. Guefrachi, R. Abdeddayem and M. Mars. 2012. Symbiotic nitrogen fixation of wild legumes in Tunisia: Soil fertility dynamics, field nodulation and nodules effectiveness. Agriculture, Ecosystems and Environment 157: 60–69. Search in Google Scholar

Rejii M., M. Mahdhi, J.A. Domínguez-Núñez and M. Mars. 2013. The phenotypic, phylogenetic and symbiotic characterization of rhi- zobia nodulating Lotus sp. in Tunisian arid soils. Ann. Microbiol. 64: 355–362. Search in Google Scholar

Rivas R., A. Willems, N.S. Subba-Rao, P.F. Mateos, F.B. Dazzo, R.M. Kroppenstedt, E. Martinez-Molina, M. Gillis and E. Velázquez. 2003. Description of Devosia neptuniae sp. nov. that nodulates and fixes nitrogen in symbiosis with Neptunia natans, an aquatic legume from India. Syst. Appl. Microbiol. 26: 47–53. Search in Google Scholar

Somasegaran P. and H.J. Hoben. 1994. Handbook of rhizobia, p. 450. In: Methods in legume-rhizobium technology. Springer, New York.10.1007/978-1-4613-8375-8 Search in Google Scholar

Trujillo E.M., A. Willems, A. Abril, A.M. Planchuelo, R. Rivas, D. Ludena, P.F. Mateos, E. Martinez-Molina and E. Velazquez. 2005. Nodulation of Lupinus albus by strains of Ochrobactrum lupine sp. nov. Appl. Environ. Microbiol. 71: 1318–1327.10.1128/AEM.71.3.1318-1327.2005106514515746334 Search in Google Scholar

Valverde A., E. Velázquez, F. Fernández-Santos, N. Vizcaíno, R. Rivas, P.F. Mateos, E. Martínez-Molina, J.M. Igual and A. Willems. 2005. Phyllobacterium trifolii sp. nov., nodulating Trifolium and Lupinus in Spanish soils. Int. J. Syst. Evol. Microbiol. 55: 1985–1989.Search in Google Scholar

Vandamme P., B. Pot, M. Gillis, P. de Vos, K. Kersters and J. Swing. 1996. Polyphasic taxonomy, a consensus approach to bacterial systematics. Microbiol. Rev. 60:407–438.10.1128/mr.60.2.407-438.19962394508801440 Search in Google Scholar

Vincent J.M. 1970. A manual for the practical study of root nodule bacteria. IBP Handbook, No. 15. Blackwell, Oxford Search in Google Scholar

Wei G.H., Z.Y. Tan, M.E. Zhu, E.T. Wang, S.Z. Han and W.X. Chen. 2003. Characterization of rhizobia isolated from legume species within the genera Astragalus and Lespedeza grown in the Loess Plateau of China and description of Rhizobium loessense sp. nov. Int. J. Syst. Evol. Microbiol. 53: 1575–1583.10.1099/ijs.0.02031-013130051 Search in Google Scholar

Wei G.H., Z.X. Zhang, C. Chen, W.M. Chen and W.T. Ju. 2008. Phenotypic and genetic diversity of rhizobia isolated from nodules of the legume genera Astragalus, Lespedeza and Hedysarum in northwestern China. Microbiol. Res. 163: 651–662. Search in Google Scholar

Weisburg W.G., S.M. Barns, D.A. Pelletior and D.J. Lane. 1991. 16S ribosomal DNA amplification for phylogenetic study. J. Bacte- riol. 173: 697–703.10.1128/jb.173.2.697-703.19912070611987160 Search in Google Scholar

Zakhia F., H. Jeder, O. Domergue, A. Willems, J.C. Cleyet-Marel, M. Gillis, B. Dreyfus and P. De Lajudie. 2004. Characterisation of legume nodulating bacteria (LNB) in arid regions of Tunisia. Syst. Appl. Microbiol. 27: 380–395.10.1078/0723-2020-0027315214644 Search in Google Scholar

Zakhia F., H. Jeder, A. Willems, M. Gillis, B. Dreyfus and P. De Lajudie. 2006. Diverse bacteria associated with root nodules of spontaneous legumes in Tunisia and first report for nifH-like gene within the genera Microbacterium and Starkeya. Microbial Ecol. 51: 375–393.10.1007/s00248-006-9025-016598639 Search in Google Scholar

Zhang X.X., S.L. Turner, X.W. Gao, H.J. Yang, F. Debelle, G.P. Yang, J. Denarie, J.P.W. Young and F.D. Li. 2000. The common nodulation genes of Astragalus sinicus rhizobia are conserved despite chromosomal diversity. Appl. Environ. Microbiol. 66: 2988–2995.10.1128/AEM.66.7.2988-2995.20009210110877796 Search in Google Scholar

Zhao C.T., E.T. Wang, W.F. Chen and W.X. Chen. 2008. Diverse genomic species and evidences of symbiotic gene lateral transfer detected among the rhizobia associated with Astragalus species grown in the temperate regions of China FEMS Microbiol. Lett. 286: 263–273.10.1111/j.1574-6968.2008.01282.x18657113 Search in Google Scholar

Zhao C.T., E.T. Wang, Y.M. Zhang, W.F. Chen, X.H. Sui, W.X. Chen, H.C. Liu and X.X. Zhang. 2012. Mesorhizobium silam- urunense sp. nov., isolated from root nodules of Astragalus species Int. J. Syst. Evol. Microbiol. 62: 2180–2186. Search in Google Scholar

Zheng W.T., Y. Li, R. Wang, X.H. Sui, X.X. Zhang, J.J. Zhang, E.T. Wang and W.X. Chen. 2013. Mesorhizobium qingshengii sp. nov., isolated from effective nodules of Astragalus sinicus Int. J. Syst. Evol. Microbiol. 63: 2002–2007. Search in Google Scholar

Zurdo-Piñero J.L., R. Rivas, M.E. Trujillo, J.A. Vizcaíno Carrasco, A. Chamber Palomares, P.F. Mateos, E. Martínez-Molina and E. Velázquez. 2007. Ochrobactrum cytisi sp. nov., isolated from nodules of Cytisus scoparius in Spain. Int. J. Syst. Evol. Microbiol. 57: 784–788. Search in Google Scholar

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