Inhibition of Rhizobium trifolii by Yeast Extracts or Glycine is Prevented by Calcium

Abstract

SUMMARYSeveral yeast water preparations and commercial yeast extracts varied in their toxicity to two strains of Rhizobium trifolii. Strains of R. japonicum and R. leguminosarum were also susceptible to yeast extract, while strains of R. meliloti were resistant.An interaction between glycine and monovalent cations was the main cause of yeast extract toxicity. Divalent cations, notably Ca2+, added to inhibitory media prevented the toxicity and enhanced growth with increased concentrations of yeast extract. I N T R O D U C T I O NYeast extract mannitol agar (YEMA) is the most commonly used medium for the culture of rhizobia, and is made from either yeast water preparations, or commercial yeast extract powders and pastes. Attempts to recover Rhizobium trifolii quantitatively by plate counts, gave anomalous results which were traced to inhibition by commercial yeast extracts.Toxicity of yeast extracts to other bacterial species has been reported occasionally but there have been very few references to inhibition of rhizobia. Jordan (1965) reported poor growth, with apparent bacteroid formation, of Rhizobium leguminosarum in mannitol broth containing 0.35% Difco yeast extract. On the other hand, many workers have described experiments with rhizobia in media containing 0.5 to I 76 (w/v) commercial yeast extracts, without comment on the growth yield. This paper reports an examination of the effects of a number of yeast extracts on the growth of Rhizobium trfolii. METHODSMedia. Unless otherwise stated, YEMA contained (g/l) : mannitol, 10; K2HP04, 0.5 ; MgS04.7H20, 0.2; NaCl, 0.1; Oxoid no. 3 agar, 15; CaCO,, 3 (Fred, Baldwin & McCoy, 1932). Commercial yeasts extracts or yeast water preparations were added at specified concentrations w/v for extracts and v/v for yeast water. Standard YEMA contained 0.3% Oxoid yeast extract ('L 20' Batch 118). All media were adjusted to pH 6.8; additives were sterilized separately. Chemicals were analytical reagent grade where possible.Yeast extract mannitol broth (YEMB) was YEMA without agar. The medium was boiled, cooled, while the CaCO, settled, then the supernatant liquid was decanted.Yeast extracts'. Six commercial yeast extract preparations were used : two Oxoid pastes (L 20), batches 118 and 123; three Oxoid powders (L 21), batches 2233, 2719 and 2878; one Difco powder, batch 493534.

Mutation of a Broadly Conserved Operon (RL3499-RL3502) from Rhizobium leguminosarum Biovar viciae Causes Defects in Cell Morphology and Envelope Integrity

“…Glycine toxicity in bacteria is a well-documented phenomenon and has been linked to a 20 to 40% decrease in the extent of cross-linking of the peptidoglycan and significant changes in cell morphology, including elongation, spheroplast formation, and rounding of rod-shaped cells (26). Yeast extract or glycine induces significant changes in cell morphology in some strains of Rhizobium spp., resulting in alteration of the typical rodshaped cells into enlarged, distorted, pleomorphic shapes and spheroplasts that sometimes resemble bacteroid cells (46,47). The RL3499-RL3502 mutants had similarly altered cell shape in the presence of glycine, providing further evidence for cell wall alterations in these mutants.…”

“…In some reports this sensitivity was attributed to the presence of the amino acid glycine (46). To determine whether our mutants were also sensitive to glycine, growth of the mutants was tested on VMM with 1 mM glycine.…”

“…Earlier research studying rhizobial growth has documented sensitivity to yeast extract and peptide sources such as hydrolyzed casein (46,47). In some reports this sensitivity was attributed to the presence of the amino acid glycine (46).…”

“…Alterations in the cell morphology of R. leguminosarum when grown in the presence of yeast extract and glycine have been previously observed (46,47). Therefore, the cell morphologies of the SM1 and nonpolar mutants were examined using light microscopy.…”

Efficacy of Micronutrients in Influencing Growth Behavior of Rhizobium of Pigeonpea (Cajanus cajan L. [Millsp.])

“…The common media used for culturing rhizobia is yeast extract mannitol media, which is made up of yeast extract powder or pastes 13 . Yeast extract mannitol media is found to be best suiting nutrient media for Rhizobium growth.…”