Ectosymbiotic Role of Food Bacteria for Paramecium: Bacterial Detoxification of Paramecia-Killing Toxin Contained in Wheat Grass Powder

Abstract

Bacterized plant infusion is a popular culture medium for Paramecium, using Klebsiella pneumoniae for the bacterium and Wheat Grass Powder (WGP) for the plant. It has been thought that WGP plays a role in the growth of bacteria, which in turn serve as the direct food for paramecia. However, we found that bacteria suspended in saline solution were unable to support the growth of paramecia. WGP including no bacteria was able to support neither the growth nor the survival of paramecia; instead, it killed paramecia. The killing effect of the WGP-derived substance(s), estimated to be of molecular weight less than 1,000, was abolished when bacteria were once grown in the WGP and then eliminated, suggesting that bacteria might change the toxic substance into an inactive form. This inactivation of the toxic substance may be caused either by metabolization inside of the bacteria or by neutralization by means of bacteria-derived substance outside of the bacteria. The second alternative is likely, because paramecia were able to survive and grow in the WGP medium containing a sufficient amount of dead bacteria killed by formalin or kanamycin. Dead bacteria killed by autoclaving were ineffective, probably because bacterial contents were lost. These findings revealed an ectosymbiotic role of bacteria; they confer benefits upon paramecia not only as food but also as machinery to detoxicate a plant toxin.

Catalase is the Bacteria‐Derived Detoxifying Substance against Paramecia‐ Killing Toxin in Wheat Grass Powder Infusion

“…Paramecium multimicronucleatum , syngen 3 stock YM25 (mating type IV), was used throughout this study. The 0.5% WGP infusion supplemented with stigmasterol (0.5 mg/liter) and Na2HPO4·12H2O (0.5 g/liter) was autoclaved, stored, and used as culture medium after inoculating with K. pneumoniae 2 d before use (Tokusumi and Takagi 2000). Usually WGP infusion was autoclaved under 1.2 atm for 20 min, but in some experiments to confirm the effect of autoclaving, the time was modified.…”

“…Solutions containing the bacteria‐derived protein were subjected to serial two‐fold dilution with synthetic medium for Blepharisma (SMB) containing 1.5 mM NaCl, 0.05 mM KC1, 0.4 mM CaCl2, 0.05 mM MgCl2, 0.05 mM MgSO4, and 2 mM Na‐phosphate buffer (pH 6.8). This SMB was modified from the original SMBIII, that contained EDTA (Miyake 1981) and had been used as physiological balanced solution for Paramecium (Shimomura and Takagi 1985; Tokusumi and Takagi 2000). The diluted solution (0.1 ml) was mixed with 1 % WGP infusion (0.1 ml) and incubated overnight.…”

“…The diluted solution (0.1 ml) was mixed with 1 % WGP infusion (0.1 ml) and incubated overnight. Five paramecia were introduced to the mixture in three‐well depression slides and incubated at 25°C for 12 h. To kill bacteria, all paramecia were pretreated with 50 μg/ml kanamycin, which was shown to be harmless to paramecia (Tokusumi and Takagi 2000). The experiment for each diluted solution was done with a set of 6 or 9 wells (i.e.…”

“…It has long been thought that the plant infusion promotes the growth of bacteria, which in turn are the direct food for paramecia (Sonneborn 1950, 1970). However, it is not that straightforward in light of a recent study showing that the WGP infusion contained a substance toxic to paramecia, and that the toxicity was neutralized when K. pneumoniae was once grown in the WGP infusion (Toku‐sumi and Takagi 2000). To date, the toxic substance in the WGP infusion, and detoxifying substance derived from K. pneumoniae , have remained unknown.…”

Variability of Autogamy-Maturation Pattern in Genetically Identical Populations of Paramecium tetraurelia

“…Cells were grown in 0.5% Wheat Grass Powder (Pines Int., Inc., Lawrence, KS) medium that had been inoculated with Klebsiella pneumoniae 2 days before use (Tokusumi and Takagi, 2000). All cultures were kept at 25 ° C.…”