Root colonization and systemic spreading of Azoarcus sp. strain BH72 in grasses

The invasive properties of Azoarcus sp. strain BH72, an endorhizospheric isolate of Kallar grass, on gnotobiotically grown seedlings of Oryza sativa IR36 and Leptochloa fusca (L.) Kunth were studied. Additionally, Azoarcus spp. were localized in roots of field-grown Kallar grass. To facilitate localization and to assure identity of bacteria, genetically engineered microorganisms expressing beta-glucuronidase were also used as inocula. beta-Glucuronidase staining indicated that the apical region of the root behind the meristem was the most intensively colonized. Light and electron microscopy showed that strain BH72 penetrated the rhizoplane preferentially in the zones of elongation and differentiation and colonized the root interior inter- and intracellularly. In addition to the root cortex, stelar tissue was also colonized; bacteria were found in the xylem. No evidence was obtained that Azoarcus spp. could reside in living plant cells; rather, plant cells were apparently destroyed after bacteria had penetrated the cell wall. A common pathogenicity test on tobacco leaves provided no evidence that representative strains of Azoarcus spp. are phytopathogenic. Compared with the control, inoculation with strain BH72 significantly promoted growth of rice seedlings. This effect was reversed when the plant medium was supplemented with malate (0.2 g/liter). N2 fixation was apparently not involved, because the same response was obtained with a nifK mutant of strain BH72, which has a Nif- phenotype. Also, Western blot (immunoblot) analysis of protein extracts from rice seedlings gave no indication that nitrogenase was present. PCR and Western immunoblotting, using primers specific for eubacteria and antibodies recognizing type-specific antigens, respectively, indicated that strain BH72 could colonize rice plants systemically, probably mediated by longitudinal spreading through vessels.

[1]  B. Hoste,et al.  Azoarcus gen. nov., Nitrogen-Fixing Proteobacteria Associated with Roots of Kallar Grass (Leptochloa fusca (L.) Kunth), and Description of Two Species, Azoarcus indigens sp. nov. and Azoarcus communis sp. nov. , 1993 .

[2]  F. Olivares,et al.  Endophytic Diazotrophs in Sugar Cane, Cereals and Tuber Plants , 1993 .

[3]  J. Vasse,et al.  Control of Infection in the Alfalfa- Rhizobium Meliloti Symbiosis , 1993 .

[4]  T. Hurek,et al.  Improved histochemical detection of GUS reporter gene expression by light microscopy , 1992 .

[5]  M. Montagu,et al.  Infection of Intact Roots of Kallar Grass and Rice Seedlings by Azoarcus , 1991 .

[6]  J. Lynch,et al.  Anatomy and community structure of the rhizosphere. , 1990 .

[7]  C. You,et al.  Non-nodular endorhizospheric nitrogen fixation in wetland rice , 1989 .

[8]  S. Perotto,et al.  Common components of the infection thread matrix and the intercellular space identified by immunocytochemical analysis of pea nodules and uninfected roots , 1989, The EMBO journal.

[9]  J. Baldani,et al.  Field inoculation of sorghum and rice with Azospirillum spp. and Herbaspirillum seropedicae , 1988 .

[10]  Y. Bashan,et al.  Interaction Between Azospirillum Brasilense Cd and Wheat Root Cells During Early Stages of Root Colonization , 1988 .

[11]  H. Antoun,et al.  Xylem-residing bacteria in alfalfa roots , 1987 .

[12]  K. A. Malik,et al.  Isolation and identification of a N2‐fixing zoogloea‐forming bacterium from kallar grass histoplane , 1987 .

[13]  T. Hurek,et al.  Cross-Reaction of Predominant Nitrogen-Fixing Bacteria with Enveloped, Round Bodies in the Root Interior of Kallar Grass , 1987, Applied and environmental microbiology.

[14]  T. Hurek,et al.  Root-Zone-Specific Oxygen Tolerance of Azospirillum spp. and Diazotrophic Rods Closely Associated with Kallar Grass , 1987, Applied and environmental microbiology.

[15]  T. Hurek,et al.  Close Association of Azospirillum and Diazotrophic Rods with Different Root Zones of Kallar Grass , 1986, Applied and environmental microbiology.

[16]  K. Kersters,et al.  Alcaligenes piechaudii, a New Species from Human Clinical Specimens and the Environment , 1986 .

[17]  R. C. Foster.  The Ultrastructure of the Rhizoplane and Rhizosphere , 1986 .

[18]  Y. Okon.  Azospirillum as a potential inoculant for agriculture , 1985 .

[19]  D. Gabrielson,et al.  Enumeration, location, and characterization of endophytic bacteria within sugar beet roots , 1985 .

[20]  J. Torrey,et al.  Early development of Rhizobium-induced root nodules of Parasponia rigida. II. Nodule morphogenesis and symbiotic development , 1985 .

[21]  D. M. Hinton,et al.  The distribution and ultrastructure of the endophyte of toxic tall fescue , 1985 .

[22]  J. Wang,et al.  Drosophila DNA topoisomerase I is associated with transcriptionally active regions of the genome. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[23]  J. Roth,et al.  Applications of immunocolloids in light microscopy. III. Demonstration of antigenic and lectin-binding sites in semithin resin sections. , 1984, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[24]  M. Kloss,et al.  Organic acids in the root exudates of diplachne fusca (linn.) beauv. , 1984 .

[25]  R. Hawkes,et al.  A dot-immunobinding assay for monoclonal and other antibodies. , 1982, Analytical biochemistry.

[26]  H. Noller,et al.  Gene organization and primary structure of a ribosomal RNA operon from Escherichia coli. , 1981, Journal of molecular biology.

[27]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[28]  J. Döbereiner,et al.  Light microscopy observations of tetrazolium-reducing bacteria in the endorhizosphere of maize and other grasses in Brazil. , 1978, Canadian journal of microbiology.

[29]  F. Dazzo,et al.  Cross-reactive antigens and lectin as determinants of symbiotic specificity in the Rhizobium-clover association. , 1975, Applied microbiology.

[30]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[31]  Z. Klement.  Rapid Detection of the Pathogenicity of Phytopathogenic Pseudomonads , 1963, Nature.

[32]  J. Marmur.  A procedure for the isolation of deoxyribonucleic acid from micro-organisms , 1961 .

[33]  K. Arashi,et al.  Studies on the Lysigenous Intercellular Space as the Ventilating System in the Culm of Rice and Some Other Graminaceous Plants. , 1955 .