Rhizobium strain identification in Arachis hypogaea nodules by enzyme-linked immunosorbent assay (ELISA)

Abstract

The technique of enzyme-linked immunosorbent assay (ELISA) was used for serological identification of peanut Rhizobium strains both in cell suspension of pure culture and in single root nodules of groundnut (Arachis hypogaea) plants. Antisera of three peanut Rhizobium strains were tested against eight different Rhizobium isolates. Three serogroups identified by agglutination and immunodiffusion tests were confirmed by ELISA. In this experiment ELISA was more sensitive by four to six orders of magnitude than the agglutination and immunodiffusion tests and enabled the detection of Rhizobium antigens in cell suspensions of 10(4)-10(5) cells per millilitre. The reactions of culture and nodule antigens were identical for all strains investigated. ELISA enabled the precise typing of rhizobial isolates in single small root nodules. The minimum fresh weight of nodule tissue necessary to perform the ELISA test was 0.4 mg crushed in 1 ml of phosphate-buffered saline (PBS). ELISA was also successfully used for strain identification in mixed inoculated plants. One of the strains in each pair formed most of the nodules examined.

Potential for inoculation of common bean by effective rhizobia in Tunisian soils

“…This result confirms those obtained by Graham (1981) and Amarger (1986) (Kishinevsky and Bar-Joseph, 1978;Ayanaba et al, 1986;Cleyet-Marel, 1987;Evans et al, 1996 (Cregan and Keyser, 1988) and white clover (Mytton, 1975 (Schuller et al, 1986;Heakman, 1987;Salez and Saint-Macary, 1987;Minchin et al, 1989) …”

“…This can be accomplished by a number of procedures. Serotyping is the most commonly used method for Rhizobium identification (Kishinevsky and Bar-Joseph, 1978;Ayanaba et al, 1986;Cleyet-Marel, 1987;Wolff et al, 1991). (Vincent, 1970 (Beck et al, 1993) (Somasegaran and Hoben, 1985) (Fe(II)EDTA instead of Fe-citrate).…”

A Modified, Highly Sensitive Enzyme-linked Immunosorbent Assay (ELISA) for Rhizobium meliloti Strain Identification

“…The results of the testing of different ELISA procedures showed that the presented final ELISA test procedure gave maximum ELISA values. When applying the designed ELISA test with enzyme conjugate derived by whole-cell immunization it was possible to detect and recognize strains of rhizobia in samples containing 1 x loJ cells ml- (Table l), a concentration lower than in earlier reports (Kishinevsky & Bar-Joseph, 1978;Berger et al, 1979;Morley & Jones, 1980). Strain specificity of the ELISA test was also obtained in nodule extracts containing 1 x lo5 cells ml-I ( Table 3).…”

The use of lacZ marker in enumeration of Azotobacter chroococcum in carrier based inoculants

“…Moreover, loss of antibiotic markers during growth makes this method unsuitable for strain identification. Serological techniques include agglutination, immunogel diffusion, immunofluorescence and ELISA, and have been used to identify Rhizobium japonicum grown in culture and from root nodules (Dudman, 1971; Kishinevsky and Bar Joseph, 1978; Olsen and Rice, 1989). However, cross reactive antigens present in other strains give faulty results and limit the use of these techniques (Vincent and Humphery, 1970).…”