Where is lactobacillus bulgaricus found in nature

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Song, Y. Sci Rep 6, Download citation. Received : 26 October Accepted : 18 February Published : 04 March Anyone you share the following link with will be able to read this content:. Sorry, a shareable link is not currently available for this article. Provided by the Springer Nature SharedIt content-sharing initiative. Brazilian Journal of Microbiology BMC Genomics Applied Microbiology and Biotechnology Scientific Reports By submitting a comment you agree to abide by our Terms and Community Guidelines.

If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate. Advanced search. Skip to main content Thank you for visiting nature. Download PDF. Subjects Applied microbiology Bacterial evolution. Abstract Lactobacillus delbrueckii subsp. Ovals represent the core proteomes of L. The overall core of the 5 ssp.

A, 65 proteins are present in all ssp. When comparing the proteins from the core proteome of the ssp. The latter proteins are mainly involved in carbohydrate and amino-acid metabolism see below. The 8 ssp. For 24 among the 65 ssp. Likewise, for 5 of the 25 ssp. These observations suggest that at least part of the subspecies specificity results from the differential loss of ancestral genes.

A second in silico comparison was made using KEGG [ 12 ] functional annotations as a starting point. For this purpose, proteins from the predicted proteomes of strains ssp. Matching maps were compared, and for a selection of differences between the two strains the comparison was extended to the 10 strains used in this study using homology search based analyses.

Not unexpectedly, the results of this analysis to a great extent corroborate the results of the core genomes analysis for as far as genes with known functions are concerned, and in addition put a certain number of differences in a metabolic pathway context. A first impression of the global metabolic capacities of the two strains obtained using Ipath [ 14 ] suggests that these capacities are more extensive for the ssp.

Metabolic capacities of L. Metabolic pathway analysis was performed using KEGG [ 12 , 13 ]. Graphs were generated using ipath [ 14 ]. An overview of the experimentally determined carbohydrate metabolism potential of four ssp. However, a high level of variability was observed within the ssp. Because of this intra ssp. With only few exceptions, the observed fermentation profiles could be predicted on the basis of in silico metabolic pathway analyses using existing and newly established genome sequences Additional file 6 : Table S5.

The results of these in silico analyses also suggested that the ssp. This prediction was experimentally confirmed by growth on M17 medium containing starch as the only carbon source and a starch degradation assay results not shown. Apart from the difference in carbohydrate metabolism potential between the two ssp. This observation suggests that the ssp.

The results of the in silico analysis also indicate that for the uptake of carbohydrates L. A noticeable exception is the uptake of the milk sugar lactose in the ssp. The comparative genomics approach using KEGG pathways led to some particularly interesting observations with regard to the uptake and metabolism of the milk sugar lactose, a key feature of dairy lactic acid bacteria.

While for the ssp. Using this system, the spp. Lactose transport and metabolism pathways in L. PEP, phosphoenolpyruvate; Pyr, pyruvate. The first signs of a loss of this system are also found in the ssp.

It is interesting to note that several lactobacilli of the acidophilus group L. This family 42 protein is not present in L. As earlier reported [ 5 ], the lac repressor is inactivated in the ssp. Inactivation of this repressor and the loss of the galactose metabolic pathway in the spp. Furthermore, the ancestral PTS-lactose system still visible in the ssp. A second key to the comprehension of L. Earlier studies of the L. This view is confirmed in the other L.

All strains of the ssp. Neither of the two ssp. Both ssp. This number is not different, however, from the number found in Lactobacillus gasseri , a closely related Lactobacillus acidophilus group member from the gastro-intestinal tract not shown. The only significant adaptation of the proteolytic system of both L. This protease is not found in other lactobacilli of the acidophilus group except Lactobacillus equicursoris , the closest known relative of L.

In spite of this, the analysis of the ssp. For the latter enzymes, L. This vision is corroborated by the analysis of the other ssp. Ornithine can then be used to import more arginine through an arginine-ornithine antiporter encoded by arc D , or be decarboxylated by ornithine decarboxylase for which the ssp. In one of the ssp. Both the arginine deiminase pathway and glutamate decarboxylase can be found in other lactobacilli of the acidophilus group, and the above mentioned observations suggest that L.

Lactobacillus delbrueckii is one of the economically most important bacteria in dairy industries, where the ssp. The classical procedure of distinguishing the two ssp. While the ssp. The in silico analysis of the first complete genome sequence of an L. Here, we extend this observation to four other L. For the ssp. In this study we extend these genome-scale observations for the first time to the ssp.

Intriguingly, while for any of 14 mono- and di-saccharides studied at least one and on an average 4 out of 8 strains lost the capacity to ferment it, all L.

A second major adaptation to the milk environment concerns the loss of amino acid biosynthesis pathways, earlier observed in L. Like in the case of carbohydrate metabolism, this adaptation appears to be less advanced in the ssp. Explained by the ample availability of amino acids in the form of milk proteins, this adaptation is coherent with the presence of the major cell wall bound protease PrtB responsible for the first step in the degradation of milk proteins in both ssp.

The gene encoding this proteinase is not found in closely related lactobacilli. From these examples, the general picture emerges that both L. This conclusion is in line with an earlier study by Germond et al. The common ancestor would have evolved in an environment where various carbohydrates from plant origin constituted the main carbon sources, as can be deduced from the presence of remnants of genes involved in their fermentation in the various strains analyzed.

Interestingly, our analyses indicate that while the ssp. Whereas an antiporter is the system of choice in the lactose-rich milk environment, a PTS system, which excels in conditions where the substrate concentration is low, points to an ancestral environment where lactose was present but in low concentrations.

Together with the indication that the ancestral environment contained carbohydrates from plant origin, this may suggest that the ancestor evolved in the mammalian digestive tract, an environment where both conditions can be met. To our knowledge, this is the first report on the isolation and characterization of L. Yogurt is a fermented milk product obtained by fermentation of milk by the action of symbiotic starter cultures of Lactobacillus delbrueckii ssp.

However, the popularity of Bulgarian yogurt, currently known simply as yogurt, and consumption of it throughout the world, could be attributed to the Nobel Prize winner Ilya Metchnikoff. He observed the long life-span of Bulgarian peasants who consumed the traditional fermented milk, and introduced the probiotic concept for the first time.

He suggested that lactobacilli might counteract the putrefactive effects of the gastrointestinal metabolism Metchnikoff, There are some opinions concerning the natural habitat or distribution of yogurt-starter bacteria. Yogurt was indicated as the only known habitat of L.

After maintaining at this temperature, a dense milk coagulum is obtained and used to prepare home-made yogurt. This practice, however, is rarely applied at present. The origin of this traditional custom to prepare yogurt starter is not known precisely. According to Markoff , until more than years ago the shepherds from the Rodopi region of Bulgaria prepared yogurt starter by squashing the roots of Ononis spinosa and mixed the juice with sheep's milk.

Also Berberis vulgaris or Paliurus aculeatus were used for the same purpose Katrandjiev, Girginoff pointed Matricaria chamomilla , Prunus spinosa and Alfalfa as possible sources for isolation of yogurt-starter bacteria. Stefanova also mentioned that L. Although all these authors indicated plants as possible sources for isolation of yogurt-starter bacteria by examining the historical aspects of the custom to prepare home-made yogurt in Bulgaria, none of them presented any results of purposive isolation of L.

The preliminary investigation by Michaylova suggested the presence of mannose and glucose on the surface of Cornus mas and Prunus spinosa. The aims of this study were, first, to investigate the possibility for isolation of L. This paper reports, for the first time, the successful isolation of L.

Based on the results, we speculate that at least one possible origin of the yogurt-starter strains is plants in Bulgaria. A part of the results in this paper was reported previously Michaylova et al.

In the period from September to September , plant samples were collected at four regions in Bulgaria Fig. In each of the four regions, samples were taken from several sites that were away from human habitation to avoid potential contamination from home-made or commercial yogurt. Samples were collected from native plants height range up to 2 m that looked healthy. The target plant was Cornus mas and it was collected from each site. To get information on the relationship between the plant species and the existence of yogurt-starter bacteria, the other plant materials were also collected in an area about 20 m adjacent to the target plant.

Plant materials were carefully handled to avoid contamination, and each sample was immediately put in a glass tube containing 10 mL of sterile SM medium, which was then firmly plugged. Sampling areas in Bulgaria and detection of yogurt-starter bacteria. Each frozen sample was thawed and diluted with sterile 0. The following characteristics of the bacterial isolates were checked using standard protocols: Gram-staining, cell morphology, catalase activity, production of gas from glucose, halotolerance 6.

Twenty strains of L. Two industrial strains, L. B 26 and LBB. Image analysis of the band patterns, and calculations of the molecular weight of the DNA fragments based on markers, the density of the fragments and the matching among the lanes were carried out using rlfp scan software Scanalytics, Inc.

The generated matrix was subjected to clustering by the unweighted pair-group method with arithmetic means. Each L. The culture used for the measurement of acidity after 40 h of incubation was also used to obtain the whey by filtration through a paper filter No. The method described by Church was used to measure the proteolytic activity of L. The proteolytic activities were expressed as leucine equivalents, according to a standard curve obtained using leucine in a concentration range of 0—10 mM.

The strains of S. The cell suspension 0. The specific activity was defined as the number of units per absorbance of the cell suspension at nm. The method for yogurt preparation was based on the laboratory-scale manufacturing process for yogurt at the Food Research and Development Center, Meiji Dairies Corporation.

A yogurt mix containing 3. The concentration of WPI was fixed at 0. One percent of the yogurt-starter culture was inoculated to the mix. After mixing, c. Curd-tension, viscosity and particle size were measured using a curd-meter ME; I. Lactobacillus bulgaricus and S. To isolate yogurt-starter LAB from plants in Bulgaria, we analyzed plant samples collected from four regions Fig.

Nine hundred and eighty six single colonies from the test tubes on MRS or M17 agar plates were isolated after incubation for 48 h. All the bacterial cultures were preliminarily examined by simple taxonomic tests of Gram-staining, cell morphology, acid production and catalase test.

Sampling periods: a, September ; b, May ; c, March ; d, September The remaining bacterial isolates containing 70 rod-shaped bacteria hereafter referred to as rods and coccal bacteria cocci were judged as LAB. Rods were isolated from Calendula officinalis, Cornus mas , Galanthus nivalis , Prunus spinosa and other unidentified plant species.

Cocci were isolated from Calendula officinalis, Capsella bursa-pastoris, Chrysanthemum, Cichorium intybus, Colchicum, Cornus mas, Dianthus, Galanthus nivalis, Hedera, Nerium oleander, Plantago lanceolata, Prunus spinosa, Rosa, Tropaeolum and other unidentified plant species. Both rods and cocci were simultaneously isolated from Calendula officinalis, Cornus mas, Galanthus nivalis and Prunus spinosa. The phenotypes of the selected 70 rods and cocci were characterized using the standard methods.

All of them had no catalase activity and produced no gas from glucose. These characteristics, as well as the results obtained using the API system data not shown , identified them as L. The remaining seven rods utilized glucose, galactose, mannose and lactose and produced dl -lactate and were classified as Lactobacillus helveticus. These characteristics, as well as the results obtained using the API system data not shown , identified them as S.

The remaining eight cocci were tentatively identified as Lactococcus lactis based on their characteristics data not shown. These results show that L. The results of detection of yogurt-starter bacteria from plants are summarized in Table 1. Twenty representative strains of L. The selection process was designed so as to include the most diverse strains isolated from the plant samples.

Moreover, all 20 strains identified as S. These findings verified the identification of the 20 selected strains from plants in Bulgaria as L. To gain further insight into the relationships among the 20 selected strains of L.

The industrial yogurt-starter strains, L. Cluster analysis of the PFGE patterns of the bacterial isolates. The results from the industrial strains, L. The 20 selected strains of L. On the other hand, the 20 selected strains of S. Strains with the same PFGE pattern were frequently isolated from different plant species within one region, but were rarely isolated from two or more different regions for both L.

It was examined whether each selected strain of L. The acidity and pH of SMY medium, and the kinematic viscosity of the whey, were similar for all the strains of L. Moreover, all the selected strains of L. These values were consistent with our data so far obtained for L.

Properties of Lactobacillus delbrueckii ssp. Similarly, the acidity and pH of SMY medium and the kinematic viscosity of the whey did not significantly differ among all the strains of S. Furthermore, urease activity was detected in all strains of S. Properties of Streptococcus thermophilus isolated from plants in Bulgaria.

Based on these results, it was concluded that the 20 strains of L. To evaluate the quality of yogurt prepared with the selected strains, we prepared set-type yogurt using one of the 26 combinations of starter strains as shown in Table 5.