An Agar Plate Method for Rapid Screening of Rhizobium for Tolerance to Acid-Aluminum Stress

Abstract

An agar plate method has been developed for the rapid, preliminary screening of large numbers of rhizobia for tolerance to acidity and aluminum (acid-Al). Media containing pH indicators are used, and distinguish true tolerance from cases where growth depends on a previous pH shift. Strains which were highly tolerant or sensitive to acid Al on agar behaved similarly in defined liquid media. Strains that formed dry, pinpoint colonies were more sensitive to acid-AI than those that formed large, gummy colonies.Additional Index Words. acidity, aluminum, cowpea rhizobia, pH indicators, Rhizobium japonicum.Ayanaba, A., S. Asanuma, and D.N. Munns. 1983. An agar plate method for rapid screening of Rhizobium for tolerance to acid-aluminum stress. Soil Sci. Soc. Am. J. 47:256-258.METHODS have been developed recently for the preselection of Rhizobium strains tolerant of the soil acidity factors: aluminum, phosphate, and acidity (Date and Halliday, 1978;Keyser and Munns, 1979) and so of possible value as inoculant strains in the tropics. In these studies test strains are added to the stress media, incubated, and then examined for turbidity. In more refined tests, incubated cultures are monitored for cell multiplication prior to detectable pH changes.Screening for tolerance of acidity and Al can be vitiated by shifts in pH, which rhizobia sometimes induce depending on the buffering and particular composition of the medium (Date and Halliday, 1978). In agar media, local neutralization at the point of inoculation could become specially important, especially with large inocula. We report on an agar plate method that permits rapid screening of large numbers of rhizobia for tolerance to

Residual effects of successive exposure of soybean Bradyrhizobium strains to aluminium on solid defined medium

“…Both studies followed the methods described by Ayanaba et al (1983). In the present studies, it was found that the addition of Al decreased the pH of the media, similar to the studies of Ayanaba et al (1983). It was found, for example, that increases of Al from 0 mM Al to 500 mM Al decreased pH from 5.50 to 5.0±0.04.…”

“…The most likely hypothesis to explain the differences between those results and ours is the difference in the final pH of the defined medium. Both studies followed the methods described by Ayanaba et al (1983). In the present studies, it was found that the addition of Al decreased the pH of the media, similar to the studies of Ayanaba et al (1983).…”

“…It was found, for example, that increases of Al from 0 mM Al to 500 mM Al decreased pH from 5.50 to 5.0±0.04. In the present study, the pH was not adjusted to the initial pH (pH 5.5) as proposed by Ayanaba et al (1983). In the study by Octive (1990) the pH of the medium was returned to the original pH after addition of the different concentrations of aluminium.…”

“…A loopful of each culture was aseptically transferred, streaked on sterile Petri dishes containing 20 mL of solid defined medium (SDM) and incubated at 26°C for colony development. Three single colonies from each strain were selected randomly and grown in LDM to be used as inoculum (Figure 1) 50,100,150,200,250,300,350,400,450, 500 and 550 mM) were prepared and kept dry for two days (Ayanaba et al, 1983). Where required Al was added as a filter-sterilized solution of 50 mM AlK(SO 4 ) 2 .12H 2 O.…”

Preparation of pH 3.0 agar plate, enumeration of acid-tolerant, and Al-resistant microorganisms in acid soils

“…To alleviate these constraints selection of crops and production of transgenic plants that are Al-tolerant have been attempted recently. For the source of genes to produce transgenic plants, although Al-resistant microorganisms could be used, studies on Al-resistant microorganisms and genes have been carried out only in Rhizobia (Ayanba et al 1983;Johnson and Wood 1983;Flis et al 1993).…”