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Occurrence, isolation and DNA identification of

Profile image of Christine DelormeChristine Delorme

2011, African Journal of Biotechnology

https://doi.org/10.5897/AJB11.2462
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Abstract

Streptococcus thermophilus isolates from traditional butter 'Smen', a fermented product from cow's and ewe's milk in arid area was subjected to taxonomical investigations. The identification procedure included phenotypic approaches, molecular characterization by using genus polymerase chain reaction (PCR) amplifications for sodA gene encoding the manganese-dependant superoxide dismutase A, and species-specific primers from gene encoding glucose kinase (glcK), gene encoding DNA polymerase III (dnaE) and gene encoding threonyl-tRNA synthetase (thrS) housekeeping genes in order to distinguish among reference, and wild strains of S. thermophilus and for their differentiation from Enterococcus spp. A total of 12 strains were tested by DNA identification analysis and these indigenous isolates were unambiguously characterised by their housekeeping gene profiles. Finally, four genotypes were recognised. To our knowledge, this is the first report on the isolation and molecular characterization of S. thermophilus strains from fermented milk transformed into home-production butter provided from Southern West Algeria. These results are of particular interest as they favour the selection of future cultures starters as S. thermophilus from traditional dairy products. It is, therefore, of great importance to preserve the genetic pool of the wild strains.

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2017

Streptococcus thermophilus is one of the commercially important dairy bacterium and considered as second most important starter culture in the manufacturing of dairy products (Yoghurt and cheese). To enhance the variety of these starter culture are not only essential to bring new strains with unique and desired characteristics, but also for the maintenance of these yoghurt starters which reduces with the overuse. In this review, different genes are used for the detection of S. thermophilus by molecular methods and as well strains producing exopolysaccharide are discussed. Molecular analysis including pulsed field gel electrophoresis and polymerase chain reaction are addressed. Since the dairy industry is keep in searching of new dairy starters for increasing the diversity of its product range, nowadays their keen attention is to explore new starter strains with high potentiality to give fermentation products

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Comparison of RAPD-PCR and PFGE analysis for the typing of Streptococcus thermophilus strains isolated from traditional Turkish yogurts
G. Candan Gultekin

Annals of Microbiology, 2015

The dairy industry is constantly developing new starters with specific traits. Therefore, strains from natural sources have huge potential for exploration and characterization. The aim of this work was to identify the most efficient, useful, and reliable option for typing 55 Streptococcus thermophilus isolates from traditional Turkish yogurts and four industrial strains. Then, we statistically evaluated whether our conclusions could be extended to the true population. For this purpose, the results of the rapid amplified polymorphic DNA polymerase chain reaction (RAPD-PCR; using the OPI-02 MOD, M13, and XD9 primers) and pulsed-field gel electrophoresis (PFGE; using the SmaI and ApaI enzymes) typing methods were compared. The discriminatory power, typeability, and reproducibility were analyzed. Additionally, the congruence between typing methods was quantified using the adjusted Rand index (AR), Wallace index (W), and expected Wallace coefficient. Both methods revealed high genetic diversity of the S. thermophilus strains, even in the same yogurt sample. The numerical combination of results for these primers or restriction enzymes increased the congruence between the methods and provided more complete information on the strains. The comparison of these two options showed that using SmaI with ApaI was more advisable and useful than using OPI-02 MOD with M13. Additionally, the first combination represented the best tool to discriminate S. thermophilus strains (Simpson's index of diversity [DI] of 0.999 [0.997-1.000]). This finding was statistically supported. The RAPD (OPI-02 MOD) typing result showed an ability to confidently predict the PFGE (SmaI, ApaI) type (AR = 0.782 [0.618-0.949], W = 0.946 [0.865-1.000]). This result had some statistical support and might represent an important application in the dairy industry for screening strains.

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Genome comparison and physiological characterization of eight Streptococcus thermophilus strains isolated from Italian dairy products
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Eight Streptococcus thermophilus strains of dairy origin isolated in Italy were chosen to investigate autochthonous bacterial diversity in this important technological species. In the present study a comparative analysis of all the 17 S. thermophilus genomes publicly available was performed to identify the core and the variable genes, which vary among strains from 196 to 265. Additionally, correlation between the isolation site and the genetic distance was investigated at genomic level. Results highlight that the phylogenetic reconstruction differs from the geographical strain distribution. Moreover, strain M17PTZA496 has a genome of 2.15 Mbp, notably larger than that of the others, determined by lateral gene transfer (including phage-mediated incorporation) and duplication events. Important technological characters, such as growth kinetics, bacteriocin production, acidification kinetics and surface adhesion capability were studied in all the Italian strains. Results indicate a wide range of variability in adhesion properties that significantly clustered strains into four groups. Genomic differences among strains in relation to these characters were identified but a clear correlation between genotype and phenotype was not always found since most of the genomic modifications arise from single nucleotide polymorphisms. This research represents a step forward in the identification of strains-specific functions in Streptococcus thermophilus and it

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Short communication: Technological and genotypic comparison between Streptococcus macedonicus and Streptococcus thermophilus strains coming from the same dairy environment
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The species Streptococcus thermophilus is widely used for the preparation of several dairy products, and its technological contribution is clear. On the other hand, although Streptococcus macedonicus was first described more than 10 yr ago and, despite the scientific interest around this issue, the exact role of Strep. macedonicus in cheese making has yet to be clarified. In this study, 121 strains belonging to both species and isolated from the same dairy environment were genetically characterized by random amplification of polymorphic DNA (RAPD)-PCR and compared for the main biochemical features of technological interest, such as acid production, galactose utilization, citrate metabolism, exopolysaccharide production, and lipolytic, ureolytic, exocellular proteolytic, and decarboxylasic activities. Analysis by RAPD-PCR highlighted a remarkable genotypic heterogeneity among strains in both species, and, at a similarity level of 78%, all the isolates and reference strains of Strep. thermophilus grouped together and were well separated from the strains of Strep. macedonicus, confirming that these 2 species are different microbial entities. Comparison between genetic and phenotypic or biotechnological data did not reveal any relationships.

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I IN ND DO O A AM ME ER RI IC CA AN N J JO OU UR RN NA AL L O OF F P PH HA AR RM MA AC CE EU UT TI IC CA AL L S SC CI IE EN NC CE ES S A REVIEW ON MOLECULAR DETECTION OF STREPTOCOCCUS THERMOPHILUS IN DAIRY PRODUCTS
Editor iajps

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Comparison of Streptococcus thermophilus strains by pulse field gel electrophoresis of genomic DNA
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FEMS Microbiology Letters, 1998

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Genome Sequences of Four Italian Streptococcus thermophilus Strains of Dairy Origin
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Genomic Analysis of Dairy Starter Culture Streptococcus thermophilus MTCC 5461
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Journal of Microbiology and Biotechnology, 2013

The lactic acid bacterium Streptococcus thermophilus is widely used as a starter culture for the production of dairy products. Whole-genome sequencing is expected to utilize the genetic basis behind the metabolic functioning of lactic acid bacterium (LAB), for development of their use in biotechnological and probiotic applications. We sequenced the whole genome of Streptococcus thermophilus MTCC 5461, the strain isolated from a curd source, by 454 GS-FLX titanium and Ion Torrent PGM. We performed comparative genome analysis using the local BLAST and RDP for 16S rDNA comparison and by the RAST server for functional comparison against the published genome sequence of Streptococcus thermophilus CNRZ 1066. The whole genome size of S. thermophilus MTCC 5461 is of 1.73 Mb size with a GC content of 39.3%. Streptococcal virulence-related genes are either inactivated or absent in the strain. The genome possesses coding sequences for features important for a probiotic organism such as adhesion, acid tolerance, bacteriocin production, and lactose utilization, which was found to be conserved among the strains MTCC 5461 and CNRZ 1066. Biochemical analysis revealed the utilization of 17 sugars by the bacterium, where the presence of genes encoding enzymes involved in metabolism for 16 of these 17 sugars were confirmed in the genome. This study supports the facts that the strain MTCC 5461 is nonpathogenic and harbors essential features that can be exploited for its probiotic potential.

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LWT - Food Science and Technology, 2014

Streptococcus thermophilus is a commonly used starter bacterium in dairy industry. It reduces the pH of milk rapidly and equilibrates the medium for the growth of Lactobacillus delbrueckii subsp. bulgaricus during yoghurt fermentation. Efforts to increase the diversity of artisanal yoghurt starters are not only important to bring new strains with novel and desirable characteristics, but also for the preservation of natural diversity which diminishes with the overuse and spread of industrial starters to natural resources. In the present study, 14 artisanal yoghurt samples were processed for the isolation of promising strains for yoghurt starter culture production and 66 strains were subsequently characterized. They were all identified as S. thermophilus using species-specific PCR and 16S rRNA gene sequencing. Genotypic diversity at the strain level was investigated by pulsed field gel electrophoresis (PFGE), and 22 homology groups were obtained. Further phenotypic characterization unearthed a significant phenotypic heterogeneity within homology groups, mostly with atypical novel character. Only 7 out of 66 strains showed S. thermophilus type-strain like phenotypic traits. Majority of the isolates were determined to be protease positive and fast milk acidifier to be used as yoghurt starter culture.

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References (41)

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Occurrence, isolation and DNA identification of Streptococcus thermophilus involved in Algerian traditional butter ‘Smen’
Christine Delorme

AFRICAN JOURNAL OF BIOTECHNOLOGY, 2011

Streptococcus thermophilus isolates from traditional butter 'Smen', a fermented product from cow's and ewe's milk in arid area was subjected to taxonomical investigations. The identification procedure included phenotypic approaches, molecular characterization by using genus polymerase chain reaction (PCR) amplifications for sodA gene encoding the manganese-dependant superoxide dismutase A, and species-specific primers from gene encoding glucose kinase (glcK), gene encoding DNA polymerase III (dnaE) and gene encoding threonyl-tRNA synthetase (thrS) housekeeping genes in order to distinguish among reference, and wild strains of S. thermophilus and for their differentiation from Enterococcus spp. A total of 12 strains were tested by DNA identification analysis and these indigenous isolates were unambiguously characterised by their housekeeping gene profiles. Finally, four genotypes were recognised. To our knowledge, this is the first report on the isolation and molecular characterization of S. thermophilus strains from fermented milk transformed into home-production butter provided from Southern West Algeria. These results are of particular interest as they favour the selection of future cultures starters as S. thermophilus from traditional dairy products. It is, therefore, of great importance to preserve the genetic pool of the wild strains.

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Genotyping of Streptococcus thermophilus strains isolated from traditional Egyptian dairy products by sequence analysis of the phosphoserine phosphatase (serB) gene with phenotypic characterizations of the strains
Mohamed Samir Darwish

diversity of Streptococcus thermophilus strains isolated from traditional Egyptian fermented dairy products and to characterize phenotypic traits of those strains related to their potential use in bioprocessing applications. Methods and Results: A novel, simplified approach was developed for genotyping Strep. thermophilus involving the analysis of nucleotide sequence variations within a housekeeping gene encoding the phosphoserine phosphatase (SerB). Using this method, it was possible to identify ten genotypes involving diverse serB alleles among 54 Strep. thermophilus isolates cultured from Egyptian dairy products. These isolates harboured five de novo serB alleles that have not been detected in other Strep. thermophilus strains, deposited in a multilocus sequence typing (MLST) database. To assess distinct genotypes of the organism with phenotypic traits relevant to their potential use in industry, Strep. thermophilus strains were all subjected to a series of phenotypic characterizations. The strains were found to exhibit phenotypic diversity in terms of their ability to ferment lactose and galactose, express urease activity, produce exopolysaccharides and develop acidity. Conclusions: The analysis of nucleotide sequence variations within the serB gene could serve as a suitable tool for probing diverse genotypes of Strep. thermophilus. Streptococcus thermophilus isolates associated with traditional Egyptian dairy products show high degree of genetic and phenotypic diversity. Significance and Impact of the Study: This study presents a novel, simplified procedure based on serB nucleotide sequencing for genotyping Strep. thermophilus. It also provides a pool of phenotypically diverse Strep. thermophilus cultures, from which certain strains could be selected for use in bioprocessing applications including the preparation of fermented dairy products.

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Genetic diversity and technological properties of Streptococcus thermophilus strains isolated from dairy products
Monica Gatti

Journal of Applied Microbiology, 2002

Aims: To evaluate the genetic diversity and the technological properties of 44 strains of Streptococcus thermophilus isolated from dairy products. Methods and Results: Strains were analysed for some relevant technological properties, i.e. exopolysaccharide (EPS) production, growth kinetic in skim milk medium, urease activity and galactose fermentation. The EPS production, determined by evaluating the colour of the colonies grown in ruthenium red milk agar, was observed in 50% of the analysed strains. Urease activity, determined by colorimetric and conductimetric methods, showed that 91% of the isolates, all except four, could hydrolyse urea. A conductimetric approach was also used for the evaluation of the overall metabolic behaviour in milk of Strep. thermophilus strains and the differences observed allowed grouping of the strains in seven different clusters. A total of 11 strains were able to produce acid in presence of galactose. Genetic diversity of Streptococcus thermophilus strains, evaluated by Random Amplified Polymorphic DNA fingerprinting (RAPD) and amplified epsC-D restriction analysis, allowed the identification of 21 different genotypes. Conclusions: Comparison between the genotypic and phenotypic data highlights an interesting correlation between some important technological properties and well-defined genotypes. Significance and Impact of the Study: The genetic and technological characterization carried out on several Strep. thermophilus strains of dairy origin should expand the knowledge on this important lactic acid bacteria species and lead to a simple, rapid, and reliable identification of strains on the basis of well-defined biotechnological properties.

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Genotypic and phenotypic heterogeneity of Streptococcus thermophilus strains isolated from dairy products
Monica Gatti

Journal of Applied Microbiology, 2001

Aims: Forty strains of Streptococcus thermophilus isolated from dairy products were identi®ed and typed by a polyphasic approach which included phenotypic and genotypic criteria. Methods and Results: Strains were identi®ed by sugar fermentation pattern and species-speci®c PCR. Phenotypic diversity was evaluated by a chemometric model taking into account some biochemical characteristics (e.g. acidifying and peptidase activities) of technological interest. Genotypic diversity was evidenced by PCR ®ngerprinting. The overall phenotypic and genotypic information was elaborated on a multivariate statistical basis by principal components analysis and cluster analysis, respectively. When acidifying and peptidase activities were considered, PCA indicated that most of the strains isolated from Pecorino Toscano cheese were separable from the others. Similarly, most of the starter culture strains tended to separate from the cheese isolates. Conclusions: A wide strain heterogeneity among Strep. thermophilus strains isolated from dairy products was observed. Signi®cance and Impact of the Study: A computerized analysis of genotypic and phenotypic information could be applied successfully to differentiate and characterize reliably and rapidly isolates occurring in different dairy products and to comprehend the technological role of speci®c Strep. thermophilus strains in dairy technology.

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Occurrence, isolation and DNA identification of involved in Algerian traditional butter ‘Smen’
Christine Delorme

African Journal of Biotechnology, 2011

Streptococcus thermophilus isolates from traditional butter 'Smen', a fermented product from cow's and ewe's milk in arid area was subjected to taxonomical investigations. The identification procedure included phenotypic approaches, molecular characterization by using genus polymerase chain reaction (PCR) amplifications for sodA gene encoding the manganese-dependant superoxide dismutase A, and species-specific primers from gene encoding glucose kinase (glcK), gene encoding DNA polymerase III (dnaE) and gene encoding threonyl-tRNA synthetase (thrS) housekeeping genes in order to distinguish among reference, and wild strains of S. thermophilus and for their differentiation from Enterococcus spp. A total of 12 strains were tested by DNA identification analysis and these indigenous isolates were unambiguously characterised by their housekeeping gene profiles. Finally, four genotypes were recognised. To our knowledge, this is the first report on the isolation and molecular characterization of S. thermophilus strains from fermented milk transformed into home-production butter provided from Southern West Algeria. These results are of particular interest as they favour the selection of future cultures starters as S. thermophilus from traditional dairy products. It is, therefore, of great importance to preserve the genetic pool of the wild strains.

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    Microorganisms

    In Algeria, Smen is a fermented butter produced in households using empirical methods. Smen fermentation is driven by autochthonous microorganisms; it improves butter shelf-life and yields highly fragrant products used as ingredients in traditional dishes as well as in traditional medicine. The present study is aimed at investigating microbial diversity and dynamics during Algerian Smen fermentation using both culture-dependent and culture-independent approaches, as well as by monitoring volatile organic compound production. To reach this goal, fifteen Smen samples (final products) produced in households from different regions in Algeria were collected and analyzed. In addition, microbial and volatile compound dynamics at the different stages of Smen manufacturing were investigated for one Smen preparation. The results showed that Smen is a microbiologically safe product, as all hygiene and safety criteria were respected. The dominant microorganisms identified by both techniques wer...

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