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Lactobacillus kunkeei is an inhabitant of fructose-rich niches and is a potential member of the fructophilic lactic acid bacteria. In the present study, the phylogenetic and biochemical characteristics of the type strain and eight isolates of L. kunkeei, originating from wine, flowers and honey, were studied. The nine isolates, including the type strain, formed a well-defined phylogenetic subcluster based on the analysis of 16S rRNA gene sequences. The subcluster was not closely related to other subclusters in the Lactobacillus phylogenetic group. Biochemically, the eight new isolates showed typical fructophilic characteristics. The eight isolates grew poorly on glucose, but grew well on fructose. Good growth on glucose was only recorded in the presence of electron acceptors. The type strain of L. kunkeei differed from the other isolates only on the basis of poor growth on fructose. Although they belong to a group of obligately heterofermentative lactic acid bacteria, all nine isolates, including the type strain, produced almost equimolar amounts of lactic acid and acetic acid and very little ethanol from glucose. Eight of the isolates can thus be regarded as typical 'obligately' fructophilic lactic acid bacteria. Although the type strain of L. kunkeei was phenotypically slightly different from the other isolates, it possessed several important fructophilic characteristics. On the basis of the evidence gathered in this study, the type strain of L. kunkeei is recognized as a member of the 'obligately' fructophilic lactic acid bacteria.

A fructophilic lactic acid bacterium, designated strain F214-1 T , was isolated from a flower of Tropaeolum majus in South Africa. Based on phylogenetic analysis of 16S rRNA gene sequences, the strain formed a subcluster with Fructobacillus ficulneus and Fructobacillus pseudoficulneus and, based on recA gene sequences, the strain formed a subcluster with F. ficulneus. DNA-DNA hybridization studies showed that strain F214-1 T was phylogenetically distinct from its closest relatives. Acid was produced from the fermentation of D-glucose, D-fructose and D-mannitol only. D-Fructose was the preferred sole carbon and energy source and was fermented more rapidly than D-glucose. Growth of the strain on D-glucose under anaerobic conditions was very weak but external electron acceptors such as oxygen and pyruvate enhanced growth on D-glucose. Lactic acid and acetic acid were produced from D-glucose in equimolar amounts. Ethanol was produced at very low levels, despite the strain's obligately heterofermentative metabolism. Based on these data, strain F214-1 T represents a novel species of fructophilic bacteria in the genus Fructobacillus, for which the name Fructobacillus tropaeoli sp. nov. is proposed. and sodium azide (0.05; pH 6.8), was added to the crushed samples and the bags were incubated at 30 u C for 24 h. After incubation, each sample was inoculated into FYP broth by using a loop and incubated at 30 u C under aerobic conditions on an orbital shaker (120 r.p.m.) for 2 days. The culture was then streaked onto FYP agar plates containing (l 21) 5 g CaCO 3 and 12 g agar. The plates were incubated at 30 u C under aerobic conditions until visible colonies were observed. Colonies were selected based on differences in morphology, such as colony size and shape, and the clearance zone formed from hydrolysis of CaCO 3 by lactic acid, inoculated into FYP broth and GYP broth and incubated statically for 24 h at 30 u C. GYP broth differed from FYP broth by containing 10 g l 21 D-glucose instead of D-fructose.Strain F214-1 T grew well in FYP broth but poorly in GYP broth and was, therefore, regarded as a fructophilic lactic acid bacterium. The strain was stored at 280 u C in nutrient broth containing 20 % (v/v) glycerol. Strains F. durionis LMG 22556 T , F. ficulneus JCM 12225 T , F. fructosus NRIC 1058T and F. pseudoficulneus DSM 14568 T were cultured in FYP broth and used as reference strains.The 16S rRNA gene sequence of strain F214-1 T was determined as described previously (Endo & Okada, 2005). The closest recognized relatives of the isolate were determined by performing database searches and sequences of closely related species were retrieved from DDBJ. Multiple sequence alignments were carried out by using CLUSTAL_X, version 1.18 (Thompson et al., 1997). Distance matrices for the aligned sequences were calculated by using the two-parameter method of Kimura (1980) and phylogenetic trees were reconstructed by using the neighbourjoining, maximum-likelihood and maximum-parsimony methods in PHYLIP version 3.65 software as descr...

Three strains of fructophilic lactic acid bacteria were isolated from flowers in South Africa. The isolates formed a subcluster in the Lactobacillus buchneri phylogenetic group, closely related to Lactobacillus fructivorans, Lactobacillus homohiochii, Lactobacillus lindneri and Lactobacillus sanfranciscensis according to phylogenetic analysis based on the 16S rRNA gene sequences. Levels of DNA-DNA relatedness indicated that the three strains belonged to the same taxon and formed a genetically distinct group, well separated from their phylogenetic relatives. The three strains produced acids from only two of the 49 carbohydrates tested, i.e. D-glucose and Dfructose. D-Fructose was more rapidly fermented than D-glucose. Good growth was recorded on D-fructose or D-glucose in the presence of external electron acceptors. However, delayed growth was recorded on D-glucose without electron acceptors. The novel strains produced lactic acid, ethanol and acetic acid from D-glucose at a ratio of 1 : 0.8 : 0.2. These characteristics were distinct from other species of the genus Lactobacillus. Based on the data provided, the three isolates represent a fructophilic and novel species of the genus Lactobacillus, for which the name Lactobacillus florum sp. nov. is proposed. The type strain is F9-1 T (5JCM 16035 T 5DSM 22689 T 5NRIC 0771 T ).Fructophilic lactic acid bacteria (LAB) are a group of LAB described recently (Endo & Okada, 2008). These organisms are known to have unique characteristics, i.e. they grow well on D-fructose or D-glucose in the presence of external electron acceptors, but grow poorly on D-glucose without electron acceptors (Endo & Okada, 2008). Pyruvate and oxygen are used as electron acceptors. At the time of writing, only five species with these characteristics have been described, Fructobacillus fructosus, Fructobacillus durionis, Fructobacillus ficulneus and Fructobacillus pseudoficulneus (Endo & Okada, 2008) and Lactobacillus kunkeei (Endo et al., 2009).During a study of fructophilic LAB, three strains were isolated from flowers in South Africa (Endo et al., 2009). The isolates shared several common characteristics with known fructophilic LAB, but also had a few different characteristics. They formed a subcluster within the Lactobacillus buchneri cluster on phylogenetic analysis based on their 16S rRNA gene sequences and were closely related to Lactobacillus lindneri. Levels of DNA2DNA similarity and biochemical characteristics showed clear separation of the isolates from L. lindneri. This paper describes the taxonomic study of the three novel isolates and their classification as a novel species of the genus Lactobacillus. The three novel strains, F9-1 T , F9-2 and F17, grew well in FYP broth, but poorly in GYP broth and were thus regarded as fructophilic LAB. They were preserved at 280 u C in nutrient broth containing 20 % (v/v) glycerol.The 16S rRNA gene sequences of the three isolates were determined according to a previously described method (Endo & Okada, 2005). The closest recognized relatives of t...

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