Spheroplast fusion of Acetobacter aceti and its application to the breeding of strains for vinegar production.

Abstract

Spheroplasts of auxotrophic mutants derived from Acetobacter aceti subsp. aceti No. 1023 were efficiently prepared by treatment with lysozyme, using sucrose as an osmotic stabilizer, and regenerated on an agar plate containing sorbitol and gelatin. In addition, spheroplast fusion between the several auxotrophic mutants was achieved in the presence of polyethylene glycol and CaCl2. The frequency of fusion was found to be about 5 x 10~5. Spheroplast fusion between A. aceti subsp. aceti No. 2 with the ability to grow at high temperature and A. aceti subsp. xylinum NBI1002 with high resistance to acetic acid was also achieved by the same method, with a frequency of 6.0 x 10~6. The fusants showed various degrees of resistance to acetic acid and ability to grow at high temperature. One of the fusants, named No. 116, could produce acetic acid from ethanol continuously under conditions under which both parent strains were unable to grow. This suggests that spheroplast fusion is applicable to the breeding of strains for vinegar production. The genus Acetobacter is industrially important for vinegar production. To improve strains of Acetobacter genetically, recombinant DNAtechniques are considered to be useful, and host-vector systems and an efficient transformation method for Acetobacter have been developed.1~4) However, cell fusion may also be applicable for such a purpose, especially

A thermotolerant and high acetic acid-producing bacterium Acetobacter sp. I14-2

“…In addition, the production rate at 35 or 37 °C was higher than that of a protoplast fusant strain, no. 116, which possessed a high resistance to acetic acid and ability to grow at high temperatures (Fukaya et al . 1989).…”

Development of Thermotolerant Acetic Acid Bacteria Useful for Vinegar Fermentation at Higher Temperatures

“…However, with respect to acetic acid productivity, the results obtained here, 5.06 to 4.42 g/(liter' h), are more than 2 to 3 times higher than those with mesophilic strains. According to the results by Fukaya et al,14) acetic acid productivity with A. aceti NBI 1002 was recorded to be 1.06g/(liter·h) under con· tinuous vinegar fermentation in a jar fermentor. In another example of continuous vinegar fermentation, it was reported to be 1.…”

“…and 0.7 g of acetic acid per 100 ml of tap water. This medium was almost the same as that used by Fukaya et al 14 ) During the cultivation, the ethanol concentration in the medium was always kept at the level of 1-2% by controlling the feeding rate.…”

Aerobic submerged fermentation by acetic acid bacteria for vinegar production: Process and biotechnological aspects

“…Over the past few years, there have been many studies attempting to understand the mechanisms of acetic acid resistance, and also for strain development for high acidity vinegar production [76][77][78] (Table 2). Fukaya and co-workers [78] developed a spheroplast fusion between A. aceti subsp. aceti and A. aceti subsp.…”