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1.
Food Microbiol ; 121: 104487, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38637064

RESUMO

Streptococcus thermophilus is a bacterium widely used in the production of yogurts and cheeses, where it efficiently ferments lactose, the saccharide naturally present in milk. It is also employed as a starter in dairy- or plant-based fermented foods that contain saccharides other than lactose (e.g., sucrose, glucose). However, little is known about how saccharide use is regulated, in particular when saccharides are mixed. Here, we determine the effect of the 5 sugars that S. thermophilus is able to use, at different concentration and when they are mixed on the promoter activities of the C-metabolism genes. Using a transcriptional fusion approach, we discovered that lactose and glucose modulated the activity of the lacS and scrA promoters in a concentration-dependent manner. When mixed with lactose, glucose also repressed the two promoter activities; when mixed with sucrose, lactose still repressed scrA promoter activity. We determined that catabolite control protein A (CcpA) played a key role in these dynamics. We also showed that promoter activity was linked with glycolytic flux, which varied depending on saccharide type and concentration. Overall, this study identified key mechanisms in carbohydrate metabolism - autoregulation and partial hierarchical control - and demonstrated that they are partly mediated by CcpA.


Assuntos
Glucose , Lactose , Lactose/metabolismo , Glucose/metabolismo , Metabolismo dos Carboidratos , Glicólise , Streptococcus thermophilus/genética , Streptococcus thermophilus/metabolismo , Sacarose/metabolismo
2.
Food Microbiol ; 121: 104521, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38637083

RESUMO

Natural whey starters (NWS) are cultures with undefined multiple-strains species commonly used to speed up the fermentation process of cheeses. The aim of this study was to explore the diversity and the viability of Comté cheese NWS microbiota. Culture-dependent methods, i.e. plate counting and genotypic characterization, and culture-independent methods, i.e. qPCR, viability-qPCR, fluorescence microscopy and DNA metabarcoding, were combined to analyze thirty-six NWS collected in six Comté cheese factories at two seasons. Our results highlighted that NWS were dominated by Streptococcus thermophilus (ST) and thermophilic lactobacilli. These species showed a diversity of strains based on Rep-PCR. The dominance of Lactobacillus helveticus (LH) over Lactobacillus delbrueckii (LD) varied depending on the factory and the season. This highlighted two types of NWS: the type-ST/LD (LD > LH) and the type-ST/LH (LD < LH). The microbial composition varied depending on cheese factory. One factory was distinguished by its level of culturable microbial groups (ST, enterococci and yeast) and its fungi diversity. The approaches used to estimate the viability showed that most NWS cells were viable. Further investigations are needed to understand the microbial diversity of these NWS.


Assuntos
Queijo , Lactobacillus delbrueckii , Lactobacillus helveticus , Soro do Leite , Queijo/microbiologia , Microbiologia de Alimentos , Proteínas do Soro do Leite/análise , Streptococcus thermophilus/genética
3.
Antonie Van Leeuwenhoek ; 117(1): 63, 2024 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-38561518

RESUMO

Phage resistance is crucial for lactic acid bacteria in the dairy industry. However, identifying all phages affecting these bacteria is challenging. CRISPR-Cas systems offer a resistance mechanism developed by bacteria and archaea against phages and plasmids. In this study, 11 S. thermophilus strains from traditional yogurts underwent analysis using next-generation sequencing (NGS) and bioinformatics tools. Initial characterization involved molecular ribotyping. Bioinformatics analysis of the NGS raw data revealed that all 11 strains possessed at least one CRISPR type. A total of 21 CRISPR loci were identified, belonging to CRISPR types II-A, II-C, and III-A, including 13 Type II-A, 1 Type III-C, and 7 Type III-A CRISPR types. By analyzing spacer sequences in S. thermophilus bacterial genomes and matching them with phage/plasmid genomes, notable strains emerged. SY9 showed prominence with 132 phage matches and 30 plasmid matches, followed by SY12 with 35 phage matches and 25 plasmid matches, and SY18 with 49 phage matches and 13 plasmid matches. These findings indicate the potential of S. thermophilus strains in phage/plasmid resistance for selecting starter cultures, ultimately improving the quality and quantity of dairy products. Nevertheless, further research is required to validate these results and explore the practical applications of this approach.


Assuntos
Bacteriófagos , Streptococcus thermophilus , Streptococcus thermophilus/genética , Sistemas CRISPR-Cas , Iogurte , Bacteriófagos/genética , Plasmídeos/genética
4.
Molecules ; 29(7)2024 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-38611831

RESUMO

In addition to traditional use in fermented dairy products, S. thermophilus also exhibits anti-inflammatory properties both in live and heat-inactivated form. Recent studies have highlighted that some hydrolysates from surface proteins of S. thermophilus could be responsible partially for overall anti-inflammatory activity of this bacterium. It was hypothesized that anti-inflammatory activity could also be attributed to peptides resulting from the digestion of intracellular proteins of S. thermophilus. Therefore, total intracellular proteins (TIP) from two phenotypically different strains, LMD-9 and CNRZ-21N, were recovered by sonication followed by ammonium sulphate precipitation. The molecular masses of the TIP of both strains were very close to each other as observed by SDS-PAGE. The TIP were fractionated by size exclusion fast protein liquid chromatography to obtain a 3-10 kDa intracellular protein (IP) fraction, which was then hydrolysed with pancreatic enzyme preparation, Corolase PP. The hydrolysed IP fraction from each strain exhibited anti-inflammatory activity by modulating pro-inflammatory mediators, particularly IL-1ß in LPS-stimulated THP-1 macrophages. However, a decrease in IL-8 secretion was only observed with hydrolysed IP fraction from CNRZ-21N, indicating that strain could be an important parameter in obtaining active hydrolysates. Results showed that peptides from the 3-10 kDa IP fraction of S. thermophilus could therefore be considered as postbiotics with potential beneficial effects on human health. Thus, it can be used as a promising bioactive ingredient for the development of functional foods to prevent low-grade inflammation.


Assuntos
Proteínas de Membrana , Streptococcus thermophilus , Humanos , Sulfato de Amônio , Interleucina-1beta , Macrófagos
5.
Nutrients ; 16(6)2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38542758

RESUMO

Research on regulating brain functions with probiotics and postbiotics through the gut-brain axis is attracting attention, offering the possibility of adjuvant therapy for Alzheimer's disease (AD). Three-month-old male APP/PS1 mice were gavaged with live and heat-inactivated S. thermophilus MN-002 for three months. This study demonstrated that live and heat-inactivated S. thermophilus MN-002 improved cognitive dysfunctions in APP/PS1 mice, especially in spatial memory. However, the main effects of live S. thermophilus MN-002 directly altered the intestinal microbiota composition and increased serum IL-1ß and IL-6. Heat-inactivated S. thermophilus MN-002 increased colonic propionic acid levels and enhanced the hippocampus's antioxidant capacity. Furthermore, the changes were more obvious in the high-dose group, such as astrogliosis in the hippocampus. These results indicate that different forms and doses of the same strain, S. thermophilus MN-002, can partly improve cognitive functions in AD model mice via the gut-brain axis.


Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Camundongos , Masculino , Animais , Precursor de Proteína beta-Amiloide/genética , Camundongos Transgênicos , Streptococcus thermophilus , Eixo Encéfalo-Intestino , Temperatura Alta , Doença de Alzheimer/tratamento farmacológico , Modelos Animais de Doenças , Peptídeos beta-Amiloides/uso terapêutico
6.
Molecules ; 29(6)2024 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-38542894

RESUMO

The lactic acid bacteria Streptococcus thermophilus and Lactobacillus helveticus are commonly used as starter cultures in dairy product production. This study aimed to investigate the characteristics of fermented milk using different ratios of these strains and analyze the changes in volatile compounds during fermentation and storage. A 10:1 ratio of Streptococcus thermophilus CICC 6063 to Lactobacillus helveticus CICC 6064 showed optimal fermentation time (4.2 h), viable cell count (9.64 log10 colony-forming units/mL), and sensory evaluation score (79.1 points). In total, 56 volatile compounds were identified and quantified by solid-phase microextraction and gas chromatography-mass spectrometry (SPME-GC-MS), including aldehydes, ketones, acids, alcohols, esters, and others. Among these, according to VIP analysis, 2,3-butanedione, acetoin, 2,3-pentanedione, hexanoic acid, acetic acid, acetaldehyde, and butanoic acid were identified as discriminatory volatile metabolites for distinguishing between different time points. Throughout the fermentation and storage process, the levels of 2,3-pentanedione and acetoin exhibited synergistic dynamics. These findings enhance our understanding of the chemical and molecular characteristics of milk fermented with Streptococcus thermophilus and Lactobacillus helveticus, providing a basis for improving the flavor and odor of dairy products during fermentation and storage.


Assuntos
Lactobacillus delbrueckii , Lactobacillus helveticus , Pentanonas , Animais , Leite/química , Streptococcus thermophilus/metabolismo , Fermentação , Acetoína/análise , Lactobacillus delbrueckii/metabolismo , Cetonas/análise
7.
J Food Sci ; 89(4): 1976-1987, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38454630

RESUMO

Seafood is highly perishable and has a short shelf-life. This study investigated the effect of chitosan and alginate (CH-SA) coating combined with the cell-free supernatant of Streptococcus thermophilus FUA329 (CFS) as a preservative on the quailty of white shrimp (Litopenaeus vannamei) refrigerated at 4° for 0, 3, 6, 9, 12, 15 days. Freshly shrimps were randomly divided into four groups: the CFS group (400 mL); the CH-SA group (1% chitosan/1% alginate); the CFS-CH-SA group (1% chitosan/1% alginate with 400 mL CFS) are treatment groups, and the control group (400 mL sterile water). The CFS-CH-SA coating effectively suppressed microbial growth total viable count and chemical accumulation (pH, total volatile basic nitrogen, thiobarbituric acid reactive substance) compared with the control. Additionally, the CFS-CH-SA coating improved the texture and sensory characteristics of shrimp during storage. The coated shrimp exhibited significantly reduced water loss (p < 0.05). The combination of CH-SA coating with CFS treatment can extend the shelf life of shrimp. PRACTICAL APPLICATION: Recently, edible films have received more consideration as a promising method to enhance the shelf life of seafood. The presence of Lactic acid bacteria metabolites in edible films reduces spoilage and improves consumer health. Our findings encourage the application of edible coating incorporated with cell-free supernatant of Streptococcus thermophilus FUA 329 to design multifubctional foods and preserve the qualities of shrimp.


Assuntos
Quitosana , Conservação de Alimentos , Conservação de Alimentos/métodos , Alginatos , Quitosana/farmacologia , Quitosana/química , Streptococcus thermophilus , Expectativa de Vida , Água
8.
Int J Food Microbiol ; 416: 110684, 2024 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-38513545

RESUMO

Urease operon is highly conserved within the species Streptococcus thermophilus and urease-negative strains are rare in nature. S. thermophilus MIMO1, isolated from commercial yogurt, was previously characterized as urease-positive Ni-dependent strain. Beside a mutation in ureQ, coding for a nickel ABC transporter permease, the strain MIMO1 showed a mutation in ureE gene which code for a metallochaperone involved in Ni delivery to the urease catalytic site. The single base mutation in ureE determined a substitution of Asp29 with Asn29 in the metallochaperone in a conserved protein region not involved in the catalytic activity. With the aim to investigate the role Asp29vs Asn29 substitution in UreE on the urease activity of S. thermophilus, ureE gene of the reference strain DSM 20617T (ureEDSM20617) was replaced by ureE gene of strain MIMO1 (ureEMIMO1) to obtain the recombinant ES3. In-gel detection of urease activity revealed that the substitution of Asp29 with Asn29 in UreE resulted in a higher stability of the enzyme complexes. Moreover, the recombinant ES3 showed higher level of urease activity compared to the wildtype without any detectable increase in the expression level of ureC gene, thus highlighting the role of UreE not only in Ni assembly but also on the level of urease activity. During the growth in milk, the recombinant ES3 showed an anticipated urease activity compared to the wildtype, and analogous milk fermentation performance. The overall data obtained by comparing urease-positive and urease-negative strains/mutants confirmed that urease activity strongly impacts on the milk fermentation process and specifically on the yield of the homolactic fermentation.


Assuntos
Streptococcus thermophilus , Urease , Animais , Urease/genética , Streptococcus thermophilus/metabolismo , Metalochaperonas/metabolismo , Proteínas de Transporte/genética , Níquel/metabolismo , Hidrólise , Leite/metabolismo , Ureia , Fermentação , Proteínas de Bactérias/genética
9.
J Microbiol Methods ; 220: 106924, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38548070

RESUMO

The CRISPRi system using dCas9Sth1 from Streptococcus thermophilus developed for Mycobacterium tuberculosis and M. smegmatis was modified to allow gene knock-out in M. abscessus. Efficacy of the knock-out system was evaluated by applying deletions and insertions to the mps1 gene. A comparative genomic analysis of mutants and wild type validated the target specificity.


Assuntos
Mycobacterium abscessus , Mycobacterium tuberculosis , Mycobacterium abscessus/genética , Sistemas CRISPR-Cas , Streptococcus thermophilus/genética , Mycobacterium tuberculosis/genética
10.
Arch Microbiol ; 206(3): 121, 2024 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-38400998

RESUMO

Yogurt, a globally consumed fermented dairy product, is recognized for its taste and potential health benefits attributed to probiotic bacteria, particularly Streptococcus thermophilus. In this study, we employed Multilocus Sequence Typing (MLST) to investigate the genetic diversity and phylogenetic relationships of 13 S. thermophilus isolates from traditional Turkish yogurt samples. We also assessed potential correlations between genetic traits and geographic origins. The isolates were identified as S. thermophilus using VITEK® MALDI-TOF MS, ribotyping, and 16S rRNA analysis methods. MLST analysis revealed 13 different sequence types (STs), with seven new STs for Turkey. The most prevalent STs were ST/83 (n = 3), ST/135 (n = 2), and ST/134 (n = 2). eBURST analysis showed that these isolates mainly were singletons (n = 7) defined as sequence types (STs) that cannot be assigned to any group and differ at two or more alleles from every other ST in the sample. This information suggests that the isolates under study were genetically distinct from the other isolates in the dataset, highlighting their unique genetic profiles within the population. Genetic diversity analysis of ten housekeeping genes revealed polymorphism, with some genes showing higher allelic variation than others. Tajima's D values suggested that selection pressures differed among these genes, with some being more conserved, likely due to their vital functions. Phylogenetic analysis revealed distinct genetic diversity between Turkish isolates and European and Asian counterparts. These findings demonstrate the genetic diversity of S. thermophilus isolates in Turkish yogurt and highlight their unique evolutionary patterns. This research contributes to our understanding of local microbial diversity associated with yogurt production in Turkey and holds the potential for identifyic strains with enhanced functional attributes.


Assuntos
Streptococcus thermophilus , Iogurte , Tipagem de Sequências Multilocus/métodos , Streptococcus thermophilus/genética , Filogenia , RNA Ribossômico 16S/genética , Turquia , Polimorfismo Genético , Variação Genética
11.
J Agric Food Chem ; 72(6): 3008-3016, 2024 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-38301119

RESUMO

Streptococcus thermophilus FUA329 converts ellagic acid (EA) to urolithin A (Uro-A), which is not autonomously converted by the gut microbiota to produce highly bioavailable and multibiologically active Uro-A in urolithin metabotype 0 (UM-0) populations. We consider that Streptococcus thermophilus FUA329 has the potential to be developed as a probiotic. Therefore, we utilized S. thermophilus FUA329 for in vitro cofermentation with gut microbiota. The results revealed that strain FUA329 increased the production of EA-converted Uro-A during in vitro cofermentation with the human gut microbiota of different urolithin metabotypes (UMs), with a significant increase in the production of Uro-A in the experimental group of UM-0. In addition, changes in the in vitro cofermentation microbial community were determined using high-throughput sequencing. Strain FUA329 modulated the structure and composition of the gut microbiota in different UMs, thereby significantly increasing the abundance of beneficial microbiota in the gut microbiota while decreasing the abundance of harmful microbiota. Of greatest interest was the significant increase in the abundance of Actinobacteria phylum after the cofermentation of strain FUA329 with UM-0 gut microbiota, which might be related to the significant increase in the production of Uro-A.


Assuntos
Microbioma Gastrointestinal , Microbiota , Humanos , Streptococcus thermophilus , Cumarínicos/química , Ácido Elágico
12.
Int J Mol Sci ; 25(4)2024 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-38397005

RESUMO

Gamma-aminobutyric acid (GABA)-producing lactic acid bacteria (LAB) can be used as starters in the development of GABA-enriched functional fermented foods. In this work, four GABA-producing strains each of Lactococcus lactis and Streptococcus thermophilus species were isolated from cow's milk, and their phenotypic, technological, and safety profiles determined. Genome analysis provided genetic support for the majority of the analyzed traits, namely, GABA production, growth in milk, and the absence of genes of concern. The operon harboring the glutamate decarboxylase gene (gadB) was chromosomally encoded in all strains and showed the same gene content and gene order as those reported, respectively, for L. lactis and S. thermophilus. In the latter species, the operon was flanked (as in most strains of this species) by complete or truncated copies of insertion sequences (IS), suggesting recent acquisition through horizontal gene transfer. The genomes of three L. lactis and two S. thermophilus strains showed a gene encoding a caseinolytic proteinase (PrtP in L. lactis and PrtS in S. thermophilus). Of these, all but one grew in milk, forming a coagulum of good appearance and an appealing acidic flavor and taste. They also produced GABA in milk supplemented with monosodium glutamate. Two L. lactis strains were identified as belonging to the biovar. diacetylactis, utilized citrate from milk, and produced significant amounts of acetoin. None of the strains showed any noticeable antibiotic resistance, nor did their genomes harbor transferable antibiotic resistance genes or genes involved in toxicity, virulence, or pathogenicity. Altogether these results suggest that all eight strains may be considered candidates for use as starters or components of mixed LAB cultures for the manufacture of GABA-enriched fermented dairy products.


Assuntos
Queijo , Lactobacillales , Lactococcus lactis , Animais , Leite/microbiologia , Lactococcus lactis/genética , Streptococcus thermophilus/genética , Ácido gama-Aminobutírico , Genômica , Fermentação , Queijo/microbiologia
13.
PLoS One ; 19(2): e0293378, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38386624

RESUMO

This study evaluated 15 lactic acid bacteria with a focus on their ability to degrade inosine and hypo-xanthine-which are the intermediates in purine metabolism-for the management of hyperuricemia and gout. After a preliminary screening based on HPLC, Lactiplantibacillus plantarum CR1 and Lactiplantibacillus pentosus GZ1 were found to have the highest nucleoside degrading rates, and they were therefore selected for further characterization. S. thermophilus IDCC 2201, which possessed the hpt gene encoding hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and exhibited purine degradation, was also selected for further characterization. These three selected strains were examined in terms of their probiotic effect on lowering serum uric acid in a Sprague-Dawley (SD) rat model of potassium oxonate (PO)-induced hyperuricemia. Among these three strains, the level of serum uric acid was most reduced by S. thermophilus IDCC 2201 (p < 0.05). Further, analysis of the microbiome showed that administration of S. thermophlilus IDCC 2201 led to a significant difference in gut microbiota composition compared to that in the group administered with PO-induced hyperuricemia. Moreover, intestinal short-chain fatty acids (SCFAs) were found to be significantly increased. Altogether, the results of this work indicate that S. thermophilus IDCC 2201 lowers uric acid levels by degrading purine-nucleosides and also restores intestinal flora and SCFAs, ultimately suggesting that S. thermophilus IDCC 2201 is a promising candidate for use as an adjuvant treatment in patients with hyperuricemia.


Assuntos
Hiperuricemia , Nucleosídeos de Purina , Ratos , Animais , Humanos , Nucleosídeos de Purina/metabolismo , Ácido Úrico , Hiperuricemia/metabolismo , Nucleosídeos , Streptococcus thermophilus , Ratos Sprague-Dawley , Xantina
14.
Food Funct ; 15(3): 1431-1442, 2024 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-38224462

RESUMO

Probiotic intervention, already showing promise in the treatment of various psychiatric disorders like depression, emerges as a potential therapy for anorexia nervosa (AN) with minimal side effects. In this study, we established an activity-based anorexia (ABA) model to probe the pathogenesis of AN and assess the impact of probiotics on ABA mice. ABA resulted in a compensatory increase in duodenal ghrelin levels, impairing the regulation of feeding and the brain reward system. Intervention with Streptococcus salivarius subsp. thermophilus CCFM1312 ameliorated these ABA-induced effects, and the activation of neurons in the nucleus tractus solitarius (NTS) was observed following probiotic administration, revealing the advantageous role of probiotics in AN through the vagus nerve. Furthermore, our metabolomics analysis of cecal contents unveiled that S. salivarius subsp. thermophilus CCFM1312 modulated gut microbiota metabolism and thereby regulated intestinal ghrelin levels.


Assuntos
Probióticos , Resiliência Psicológica , Streptococcus salivarius , Humanos , Animais , Camundongos , Grelina , Anorexia , Streptococcus thermophilus
15.
Arch Microbiol ; 206(2): 82, 2024 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-38294545

RESUMO

Probiotics are beneficial microorganisms, mostly lactic acid bacteria (LAB), that offer health benefits to the host when consumed in adequate amounts. This study assessed the probiotic efficacy and safety of LAB strains isolated from Laban, a traditional fermented milk product. Seven primarily selected Gram-positive, catalase-negative, non-spore-forming isolates were examined for their antimicrobial activity against the bacterial pathogens Bacillus cereus, Salmonella typhi, Staphylococcus aureus, and Vibrio cholera, and the fungal pathogen Candida albicans. Two isolates, identified as Pediococcus pentosaceus L1 and Streptococcus thermophilus L3, which showed antimicrobial activity against all pathogens, were further evaluated for their probiotic competence. The selected isolates demonstrated strong resistance to low pH, bile salts, and phenol, indicating their potential for gastric endurance. They also exhibited high cell surface hydrophobicity to various hydrocarbons, autoaggregation, and coaggregation properties, demonstrating strong adhesion abilities. In addition, both isolates showed strong antioxidant activity and were non-hemolytic. Although the isolates had some resistance to certain antibiotics, they were generally susceptible to commonly used antibiotics. The two LAB strains also exhibited promising technological properties, such as milk coagulation and exopolysaccharide production, indicating their potential to enhance the quality of dairy products. The results suggest that the LAB strains isolated from Laban have strong potential as probiotics, and due to their food origin, they are highly likely to exhibit maximal efficacy in food and pharmaceutical products for human consumption.


Assuntos
Produtos Fermentados do Leite , Probióticos , Humanos , Pediococcus pentosaceus , Streptococcus thermophilus , Antibacterianos/farmacologia
16.
Biotechnol Bioeng ; 121(2): 749-756, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37994543

RESUMO

Streptococcus thermophilus has been extensively used in industrial milk fermentation. However, lack of efficient genetic manipulation approaches greatly hampered the industrial application of this species. Here, we repurposed the endogenous CRISPR1 and CRISPR3 systems, both belong to type II-A CRISPR-Cas9, by delivering a self-targeting CRISPR array with DNA repair template into S. thermophilus LMD-9. We achieved 785-bp deletion in lacZ gene by repurposing CRISPR1 and CRISPR3 systems with efficiencies of 35% and 59%, respectively, when 1-kb DNA repair template was provided. While providing with 1.5-kb repair template, the editing efficiency for deletion in lacZ gene reached 90% using CRISPR3 systems. Diverse editing outcomes encompassing a stop code insertion and single nucleotide variation within lacZ, as well as a 234-bp DNA fragment insertion upstream of ster_0903, were generated with high efficiencies of 75%-100% using the CRISPR3 system. Harnessing the customized endogenous CRISPR3 system to target six genes of eps gene cluster, we obtained six single-gene knockout mutants with efficiencies of 29%-80%, and proved that the epsA, epsE, and epsG were the key genes affecting exopolysaccharides biosynthesis in S. thermophilus LMD-9. Altogether, repurposing the native type II-A CRISPR-Cas9 can be served as a toolkit for precise genome engineering in S. thermophilus for biotechnological applications.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Sistemas CRISPR-Cas/genética , Streptococcus thermophilus/genética , DNA
17.
J Dairy Sci ; 107(1): 123-140, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37641256

RESUMO

This study aimed to investigate the symbiosis between Streptococcus thermophilus CICC 6038 and Lactobacillus delbrueckii ssp. bulgaricus CICC 6047. In addition, the effect of their different inoculum ratios was determined, and comparison experiments of fermentation characteristics and storage stability of milk fermented by their monocultures and cocultures at optimal inoculum ratio were performed. We found the time to obtain pH 4.6 and ΔpH during storage varied among 6 inoculum ratios (1:1, 2:1, 10:1, 19:1, 50:1, 100:1). By the statistical model to evaluate the optimal ratio, the ratio of 19:1 was selected, which exhibited high acidification rate and low postacidification with pH values remaining between 4.2 and 4.4 after a 50-d storage. Among the 3 groups included in our analyses (i.e., the monocultures of S. thermophilus CICC 6038 [St] and Lb. bulgaricus CICC 6047 [Lb] and their cocultures [St+Lb] at 19:1), the coculture group showed higher acidification activity, improved rheological properties, richer typical volatile compounds, more desirable sensor quality after the fermentation process than the other 2 groups. However, the continuous accumulation of acetic acid during storage showed that acetic acid was more highly correlated with postacidification than d-lactic acid for the Lb group and St+Lb group. Our study emphasized the importance of selecting an appropriate bacterial consortium at the optimal inoculum ratio to achieve favorable fermentation performance and enhanced postacidification stability during storage.


Assuntos
Lactobacillus delbrueckii , Iogurte , Animais , Iogurte/microbiologia , Streptococcus thermophilus , Fermentação , Acetatos
18.
J Sci Food Agric ; 104(3): 1458-1469, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37814322

RESUMO

BACKGROUND: Streptococcus thermophilus is an important strain widely used in dairy fermentation, with distinct urea metabolism characteristics compared to other lactic acid bacteria. The conversion of urea by S. thermophilus has been shown to affect the flavor and acidification characteristics of milk. Additionally, urea metabolism has been found to significantly increase the number of cells and reduce cell damage under acidic pH conditions, resulting in higher activity. However, the physiological role of urea metabolism in S. thermophilus has not been fully evaluated. A deep understanding of this metabolic feature is of great significance for its production and application. Genome-scale metabolic network models (GEMs) are effective tools for investigating the metabolic network of organisms using computational biology methods. Constructing an organism-specific GEM can assist us in comprehending its characteristic metabolism at a systemic level. RESULTS: In the present study, we reconstructed a high-quality GEM of S. thermophilus S-3 (iCH492), which contains 492 genes, 608 metabolites and 642 reactions. Growth phenotyping experiments were employed to validate the model both qualitatively and quantitatively, yielding satisfactory predictive accuracy (95.83%), sensitivity (93.33%) and specificity (100%). Subsequently, a systematic evaluation of urea metabolism in S. thermophilus was performed using iCH492. The results showed that urea metabolism reduces intracellular hydrogen ions and creates membrane potential by producing and transporting ammonium ions. This activation of glycolytic fluxes and ATP synthase produces more ATP for biomass synthesis. The regulation of fluxes of reactions involving NAD(P)H by urea metabolism improves redox balance. CONCLUSION: Model iCH492 represents the most comprehensive knowledge-base of S. thermophilus to date, serving as a potent tool. The evaluation of urea metabolism led to novel insights regarding the role of urease. © 2023 Society of Chemical Industry.


Assuntos
Redes e Vias Metabólicas , Streptococcus thermophilus , Animais , Streptococcus thermophilus/genética , Streptococcus thermophilus/metabolismo , Fermentação , Leite/química , Ureia/metabolismo , Trifosfato de Adenosina/análise
19.
Int J Food Microbiol ; 411: 110549, 2024 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-38157636

RESUMO

Bouhezza is a traditional Algerian cheese produced and ripened in goatskin bags called Djeld. The aim of this study was to characterize the microbial ecosystem from Djeld (fresh and dried Djeld for making Bouhezza cheese) and the changes introduced by Lben microflora during its preparation and to identify its role in cheesemaking and its safety. Two replicates of fresh and dried skin bags (FS and DS) were sampled and analyzed before and after contact with Lben. The microbiological results showed no pathogens. Skins observed before the addition of Lben were less populated 2.86 and 3.20 log CFU cm-2 than skins examined after the addition of Lben (approximately 6.0 log CFU cm-2), suggesting a potential role of Lben in releasing some microorganisms into the skin during its time in the Djeld. However, an increase in mesophilic lactic acid bacteria and yeasts was observed in Lben after different periods of interaction with the skin. PCR-TTGE revealed the predominance of lactic acid bacteria (Lactiplantibacillus plantarum, Limosilactobacillus fermentum, Staphylococcus equorum subsp. linens, Lactococcus cremoris, Streptococcus thermophilus) and a few high-GC-content bacteria (Lacticaseibacillus paracasei, Brevibacterium casei). Transfer of several microbial species was observed between the goatskin bag biofilm and Lben during the overnight interaction. Bands corresponding to Lacticaseibacillus paracasei, Brevibacterium casei, and Lactobacillus delbrueckii subsp. lactis were detected in the fresh skin profile and in Lben after contact with the fresh skin. Lacticaseibacillus paracasei was found in dried skin and Lben after contact with dry skin. Lactobacillus helveticus and Enterococcus faecalis appeared in the Lben profile and persisted in Lben and the biofilm-covered dry skin after interaction. These results demonstrate an exchange of specific microbial populations between goatskin bag biofilm and Lben during the traditional preparation method, suggesting that the diversity of goatskin biofilm contributes to the microbial diversity of Lben used in the production of Bouhezza cheese.


Assuntos
Queijo , Lactobacillales , Animais , Ecossistema , Lactobacillus , Streptococcus thermophilus , Queijo/microbiologia , Contagem de Colônia Microbiana , Leite/microbiologia , Microbiologia de Alimentos
20.
Ultrason Sonochem ; 102: 106726, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38113583

RESUMO

In this study, the effects of ultrasonicated Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Lactiplantibacillus plantarum AF1 (100 W, 30 kHz, 3 min) on the safety and bioactive properties of stirred yoghurt during storage (4 °C for 21 days) were investigated. The results showed that sonicated cultures were more effective in reducing pathogens than untreated ones. The highest antioxidant activity (DPPH and ABTS), α-glucosidase and α-amylase inhibition capacity were found in yoghurt containing sonicated probiotic + sonicated yoghurt starter cultures (P + Y + ). The highest amount of peptides (12.4 mg/g) was found in P + Y + yoghurts at the end of the storage time. There were not significant differences between the exopolysaccharide content of P + Y+ (17.30 mg/L) and P + Y- (17.20 mg/L) yoghurts, although it was significantly (P ≤ 0.05) higher than the other samples. The use of ultrasonicated cultures could enhance the safety of stirred yoghurt and improve its functional and bioactive properties.


Assuntos
Lactobacillus delbrueckii , Lactobacillus plantarum , Lactobacillus delbrueckii/fisiologia , Streptococcus thermophilus , Iogurte , Fermentação
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