To establish the molecular bases for development of a microbiological system approaching excretive fermentation of useful lipids, a mutant strain that accumulates lipids in the medium was isolated from the laboratory yeast Saccharomyces cerevisiae. Following the mutagenesis to strain YP1, a long chain fatty acid utilizer with ethylmethane sulfonate, the mutant strain, STG1, was selected from about 80,000 colonies. The analysis of extracellular lipids and the monitoring of leakage of intracellular proteins indicated that strain STG1 secreted lipids containing triacylglycerols into the extracellular space without cell lysis. Genetic studies clarified that this mutation was recessive and was complemented by wild-type genomic DNA fragments. STG1 was considered to be a good tool for elucidation of the molecular mechanism for transmembrane lipid transport.
Gluconacetobacter liquefaciens and Gluconacetobacter xylinus share very similar phenotypic characteristics. They are differentiated by the production of a reddish-brown water-soluble pigment of the former and cellulose production of the latter. However, the loss of the two distinguishing features questions the separate standings of two species. The DNA base composition and the DNA relatedness of strains of the two species, including other established species of acetic acid bacteria, were determined. G. liquefaciens strains had the higher guanine-plus-cytosine content (G+C content) in DNA, ranging from 63.5 to 66.9 mol%, and G. xylinus had the lower range, from 59.4 to 63.2 mol%, DNA hybridization revealed a low level of DNA similarity between the two species. G. liquefaciens strains produced 2,5-diketogluconic acid and pyrones from glucose, and G. xylinus strains produced 5-ketogluconic acid. From these results, it is unequivocal that G. liquefaciens is a distinct species from G. xylinus.
An automated spectrophotometric (AS) method was used to evaluate the growth-promoting ability of yeast extracts (YE) on cultures of Lactobacillus acidophilus and Lactococcus lactis subsp. cremoris. The AS data were compared to that obtained from classical shake flask fermentations and from 250 ml bioreactors equipped with pH control. In assays involving the evaluation of 26 different commercial YE, maximum growth rate (μmax) values determined with the AS unit ranged from 0.25 to 0.45 h−1 for Lb. acidophilus and from 0.10 to 0.40 h−1 for Lc. cremoris. Good correlations were obtained between AS data and manual sampling from the shake flasks or the bioreactors for μmax, as well as maximum optical density (ODmax). The AS method is thus useful as a screening tool for the selection of YE lots in media formulation. Species reacted differently to the 26 YE, but less variation was observed between strains of the same species. This suggests that a producer of various lactococci or lactobacilli can expect a relatively constant response to a given YE lot between strains of the same species. However, it should not be assumed that the YE having the best growth-promoting properties for Lb. acidophilus will also be the best media supplements for the growth of Lc. cremoris.
Sixty-four strains of acetic acid bacteria were isolated from Indonesian sources such as fruits, flowers, and fermented foods by the enrichment culture at pH 3.5. Forty-five strains were routinely identified as Acetobacter strains because of their oxidation of acetate and lactate to carbon dioxide and water and their Q-9 isoprenolog, corresponding to 70% of all the 64 acetic acid bacteria isolated. Eight isolates were identified as Gluconacetobacter strains because of their oxidation of acetate and lactate and their Q-10 isoprenolog, occupying 13% of all the isolates. The remaining 11 isolates, accommodated in the genus Gluconobacter because of no oxidation of acetate and lactate and because of their Q-10 isoprenolog, accounted for 17% of all the isolates. They were divided into two groups based on DNA base compositions. One comprised the seven isolates, which had high G+C contents of DNA ranging from 60.3 to 63.5 mol% and of which DNAs hybridized with that of the type strain of Gluconobacter oxydans at values of 64–94% of DNA relatedness. The other comprised the remaining four isolates, which had low G+C contents of DNA ranging from 57.5 to 57.7 mol% and of which DNAs hybridized with that of the type strain of Gluconobacter frateurii at values of 63–77% of DNA relatedness. The high values of DNA relatedness, 84 to 96%, were obtained between the type strains of Gluconobacter cerinus and Gluconobacter asaii.
We examined the identity of Aspergillus penicillioides, the typical xerophilic and strictly anamorphic species, using an integrated analysis of the genotypic and phenotypic characters. Our experimental methods on two genotypic characters, i.e., DNA base composition using the HPLC method and DNA relatedness using the nitrocellulose filter hybridization technique between A. flavus, A. oryzae, and their close relations revealed a good agreement with the values by buoyant density (for DNA base composition) and spectrophotometric determination (for DNA relatedness) reported by Kurtzman et al. in 1986. On the basis of these comparisons, we examined DNA base composition and DNA relatedness of six selected strains of A. penicillioides, including IFO 8155 (originally described as A. vitricola), one strain of A. restrictus, and the respective strains from Eurotium amstelodami, E. repens, and E. rubrum. As a result, five strains within A. penicillioides, including the neotype strain NRRL 4548, had G+C contents of 46 to 49 mol%, whereas IFO 8155 had 50 mol%. A. restrictus had 52 mol%, and three Eurotium species ranged from 46 to 49 mol%. The DNA relatedness between A. penicillioides (five strains), except for IFO 8155, exhibited values greater than 70%, but the DNA complementarity between four strains and IFO 8155 in A. penicillioides revealed values of less than 40%. DNA relatedness values between three species of Eurotium were 65 to 72%. We determined 18S, 5.8S, and ITS rDNA sequences as other genotypic characters from A. penicillioides (six strains), A. restrictus, and related teleomorphic species of Eurotium. In three phylogenetic trees inferred from these sequences, five strains of A. penicillioides, including the neotype strain, were closely related to each other, whereas IFO 8155 was distantly related and grouped with other xerophilic species. Our results have suggested that A. penicillioides typified by NRRL 4548 and A. penicillioides IFO 8155 (ex holotype of A. vitricola) are not conspecific. The enzyme patterns as a genotypic character and general morphology and conidial ornamentation types as phenotypic characters supported this conclusion. Therefore the name A. vitricola Ohtsuki, typified by the holotype strain IFO 8155, should be revived. Evolutionary affinities among Aspergillus species and related teleomorphs, including the xerophilic taxa, are discussed.
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