Streptococcus thermophilus strains: Multifunctional lactic acid bacteria

Streptococcus thermophilus has “Generally Recognized as Safe” status and is extensively used for the manufacture of several important fermented dairy foods, including yoghurt and some cheese varieties (Swiss, Limburger, Brick). S. thermophilus also has a number of functional activities such as production of extracellular polysaccharides, bacteriocins and vitamins. In addition, it also has potential as a probiotic, as demonstrated by various health effects, transient survival, and moderate adherence in the gastrointestinal tract. Therefore, the natural diversity among S. thermophilus strains with respect to their capacity to produce different metabolites has the potential to be exploited beyond fermentation for lactic acid production.     

Variability and molecular typing of Streptococcus thermophilus strains displaying different proteolytic and acidifying properties

Proteolytic and acidifying properties of Streptococcus thermophilus strains isolated from yoghurt or cheeses were evaluated. Among 30 strains tested, 12 exhibited cell envelope-associated proteinase activity (PrtS⁺), three displayed a slight PrtS activity (PrtS^+/?) and 15 were PrtS^?, despite the presence of the corresponding gene (prtS) in eight of them. Sequencing of the prtS gene in four PrtS^? and one PrtS⁺ strains revealed that the absence of PrtS activity in the PrtS^? strain probably results from an alteration of the prtS regulation. The strains displaying the highest acidifying capacities were all PrtS⁺. All but one PrtS⁺ strains were phylogenetically close, as shown by the sequencing of their rDNA internal transcribed spacer (ITS) 16S-23S. More specifically, the high proteolytic and acidifying capacities are associated with the presence of a type II-ITS.     

The effect of heteropolysaccharide‐producing strains of Streptococcus thermophilus on the texture and organoleptic properties of low‐fat yoghurt

Heteropolysaccharide‐producing Streptococcus thermophilus strains (LY03, ST 111, or 5077) were compared with nonropy S. thermophilus strains (NR or 5137), in respect of texture development of low‐fat yoghurt produced at 42°C. Whether milk solids were added or not, lower syneresis and improved texture and sensory properties were obtained in yoghurts produced with S. thermophilus LY03 or S. thermophilus 5077 compared with nonropy strains. Texture effects, lower syneresis and improved sensory properties because of the use S. thermophilus ST 111 were only noticeable in the presence of milk solids. Milk solids had no significant effect on the textural parameters of nonropy strains.     

Effects of exopolysaccharide-producing strains of Streptococcus thermophilus on technological and rheological properties of set-type yoghurt

This study investigated the effects of two Streptococcus thermophilus strains, ST 285 and ST 1275, on selected technological and rheological characteristics of set-type yoghurt. The strains were selected for their capability to produce distinctly different exopolysaccharides (EPS) and were thus coded as capsular (ST 285) or ropy-capsular (ST 1275). The culture performance and physico-chemical properties of yoghurt were assessed in relation to different fermentation temperatures (30, 37 or 42 °C) and prolonged storage (up to 30 days) at low temperature (4 °C). ST 1275 showed faster growth and acidification rates, resulting in yoghurt with lower syneresis and higher-flow behaviour index, than ST 285. EPS production appeared to be growth associated with the maximum given at growth temperatures of 37 and 42 °C for ST 285 and ST 1275, respectively; however, EPS concentration declined considerably during storage. Prolonged cold storage increased several rheological characteristics of yoghurt including G′, consistency index and hysteresis loop area. A weak correlation between EPS concentration and textural properties of yoghurt was observed.     

Interactions between EPS-producing Streptococcus thermophilus strains in mixed yoghurt cultures

Mixed cultures of different EPS-producing Streptococcus thermophilus strains in combination with a Lactobacillus delbrueckii subsp. bulgaricus strain with negligible EPS-production were used for yoghurt production. The yoghurt texture was characterised with respect to sensory, rheological and microstructural properties and the EPS-concentrations were determined. The cultures resulted in yoghurts with highly different texture properties, and positive interactions between certain Streptococcus thermophilus strains were observed. The underlying properties of yoghurt texture are multidimensional, but a number of microstructural characteristics were apparent in the yoghurts with the highest mouth thickness, creaminess and viscosity. A strong protein network, not too dense and with medium size pores containing EPS, seems associated with these properties. The presence of capsular polysaccharides (CPS) also appeared to be beneficial as did a combination of EPS types, which were distributed differently in the protein network (in serum pores, respectively in association with protein). Obviously, a certain concentration of EPS must be present to provide for these effects on yoghurt texture, but other factors than concentration per se seem more important.     

A comparison of beta-galactosidase specific activities in strains of Streptococcus thermophilus

Six Streptococcus thermophilus strains were examined for growth, acid production, and beta-galactosidase activity per milligram protein (specific activity), so that strain comparisons could be made. A wide range in activity was observed. Activity depended on growth time in M17 broth and, for most strains, continued to increase after cells had reached stationary phase. Maximum activity was at 16 h and ranged from 0 to 58 units/mg protein. Strain ST exhibited no beta-galactosidase activity but had trace phospho-beta-galactosidase activity (.8 units/mg protein after 2 h of growth). Strains 3641 and TS2B exhibited slower growth rates and lower beta-galactosidase activities in milk as compared to M17 broth. Further, strain 3641 exhibited 10 times the activity of strain TS2B (2.86 vs. .24 units) after 4 h of growth in milk.     

Selection of Streptococcus thermophilus strains able to produce exopolysaccharides in milk

A collection of 139 Streptococcus thermophilus strains was genetically screened by PCR targeting some genes involved in exopolysaccharide (EPS) biosynthesis. Twenty‐nine strains were found to be PCR positive and were grown in skim milk to highlight their ropy character, that is the ability to efficiently thicken the milk cultures and minimise syneresis. Three strains producing a skim milk culture with apparent viscosities of 275 ± 14, 340 ± 21 and 510 ± 42 mPa.s, significantly higher than the average value (135 ± 87.3 mPa.s), were selected. EPS from milk cultures of these three strains was extracted and quantified from 84.4 to 209.2 mg/L.     

二株嗜热链球菌益生特性的体外研究

对嗜热链球菌ST1和嗜热链球菌KS1的部分益生性及功能特性进行了研究。考察了ST1和KS1对酸和胆盐的耐受性,结果表明,ST1和KS1在pH值为3.0条件下培养3h的存活率分别为67.17%和67.49%,在pH值为4.0或质量分数为0.3%胆盐环境中仍可以生长,表现出较高的存活力。通过抑菌实验表明ST1和KS1可以抑制常见的致病菌金黄色葡萄球菌、大肠杆菌、鼠伤寒沙门氏菌和福氏志贺氏菌。在对抗生素敏感实验中,ST1对其中的3种抗生素有耐药性,而KS1对6种抗生素有耐药性。研究还发现ST1和KS1还具有较高的降胆固醇的能力,胆固醇的去除率分别为65.14%和74.49%。     

Streptococcus thermophilus bacteriophages

At least 345 bacteriophages infecting Streptococcus thermophilus starter cultures have been isolated; general characteristics include high thermal resistance, short latent periods and large burst size. Phages with such characteristics are primed to thrive in the cheese making environment, lysing bacterial cultures and generating low-quality fermented products. All S. thermophilus phages isolated to date are members of the Siphoviridae family and the Caudovirales order and appear to constitute a polythetic phage species comprising two large groups, cos- and pac-types, based on the mode of DNA packaging. Comparative analyses have shown that S. thermophilus phage genomes are similarly organized into distinct modular regions and allow the detection of a core genome region. Several PCR-based techniques have been designed to detect them in cheese whey and milk samples. Similar S. thermophilus phages are globally distributed and endemic in specific dairy environments. The biogeography of S. thermophilus phages reinforces their current classification.     

Characterization of Streptococcus thermophilus strains that undergo lysis under unfavourable environmental conditions

The autolysis of starter lactic acid bacteria appears as a promising way to enhance the flavour of fermented dairy products. The present work was aimed at investigating the autolysis phenomenon in Streptococcus thermophilus, a thermophilic lactic acid bacteria involved in the starters used for the production of yoghurts, Italian and Swiss-type cheeses. Out of 146 strains screened for their aptitude to spontaneously lyse at the end of growth in M17 medium containing lactose in limited concentration, six strains, among which is the type strain CNRZ 1358, were found to be highly autolytic. These autolytic strains are characterized by a typical bell-shaped growth curve. Lysis of the type strain, which was studied as the model, was triggered under unfavourable environmental conditions, such as lactose depletion and NaCl or organic solvents addition. The lysogenic character of this strain was evidenced. Taken together, our results indicate that the autolytic phenotype in S. thermophilus is linked to the lysogenic character but does not result from the massive prophage induction under stressing conditions.     

Differentiation of Streptococcus thermophilus strains in commercial Direct Vat Set yoghurt starter

Based on inhibition of acid production, 100 individual colonies of Streptococcus thermophilus isolated from commercial Direct Vat Set yoghurt starter DVS-M were typed into 2 groups of sensitive and insensitive to phage φ001, which was isolated from the lysis culture of the turbidity test. The ratio between the sensitive and insensitive isolates in DVS-M was calculated at 3:2. Whole-cell protein profile (WCPP) and randomly amplified polymorphic DNA (RAPD) analysis showed that both WCPP and RAPD patterns between sensitive and insensitive isolates were distinct, whereas isolates with the same sensitivity or insensitivity shared common WCPP and RAPD patterns. It suggested that all the sensitive isolates belong to one S. thermophilus strain and all the insensitive isolates belong to another one. This study revealed that DVS-M yoghurt starter consists of 2 S. thermophilus strains with different phage sensitivity.     

Selection and properties of Streptococcus thermophilus mutants deficient in urease

Natural variations of the urea content of milk have a detrimental effect on the regularity of acidification by Streptococcus thermophilus strains used in dairy processes. The aim of the present study was to select urease-deficient mutants of S. thermophilus and to investigate their properties. Using an improved screening medium on agar plates, mutants were selected from 4 different parent strains after mutagen treatment and by spontaneous mutation. Most mutants were stable and had a phage sensitivity profile similar to that of their parent strain. Some of them contained detrimental secondary mutations, as their acidifying activity was lower than that of the parent strain cultivated in the presence of the urease inhibitor flurofamide. The proportion of this type of mutant was much lower among spontaneous mutants than among mutants selected after mutagen treatment. Utilization of urease-deficient mutants in dairy processes may have several advantages, such as an increase in acidification, an improved regularity of acidification, and a lower production of ammonia in whey.     

Streptococcus thermophilus strains of plant origin as dairy starters: Isolation and characterisation

Streptococcus thermophilus strains have been isolated mainly from dairy environments. To prospect for new strains of S. thermophilus, isolation was made from different plant sources. In this study, 74 plant isolates were characterised as S. thermophilus by a polyphasic approach and by 16S rRNA sequencing. The isolates were further evaluated for their physiological and biochemical properties. Plant isolates exhibited good acid production and varying proteolytic activity. All the isolates showed acetaldehyde and capsular exopolysaccharide (EPS) production and few revealed diacetyl production. In contrast to industrial strains, six isolates were able to ferment galactose and 25 were found to have no urease activity. Both the plant isolates and reference dairy cultures were found to possess similar physiological and biochemical properties and can be considered for developing new starters.     

Selection and properties of alpha-acetolactate decarboxylase-deficient spontaneous mutants of Streptococcus thermophilus

Many lactic acid bacteria produce diacetyl, which is a desirable aroma compound in some fermented dairy products. Strains or mutants of Lactococcus lactis subsp. lactis biovar. diacetylactis that are deficient in alpha-acetolactate decarboxylase are used in some food processes for their ability to produce large amounts of diacetyl. However, up until now, the use of alpha-acetolactate decarboxylase-deficient mutants of Streptococcus thermophilus for increased diacetyl production has not been evaluated. The objective of the present study was to devise a procedure for selecting spontaneous alpha-acetolactate decarboxylase-deficient mutants of S. thermophilus. We observed that in a chemically defined medium (CDM) containing alpha-ketobutyrate plus leucine, or alpha-ketobutyrate plus leucine plus isoleucine, the alpha-acetolactate decarboxylase-deficient mutant TIL865, obtained by directed mutagenesis, grew faster than its parent strain. This property was used for selecting spontaneous alpha-acetolactate decarboxylase-deficient mutants on agar plates. The resulting mutants were able to grow in milk, and their acidifying activity was slightly lower than that of the parent strain. Under partial anaerobic or aerobic conditions, they produced approximately three times more diacetyl than the parent strain. Such spontaneous mutants may be useful for increasing the diacetyl content of fermented milks whose production involves S. thermophilus strains.     

Interactions between Lactococcus lactis and Streptococcus thermophilus strains in Cheddar cheese processing conditions

The growth of pure and mixed cultures of Lactococcus lactis and Streptococcus thermophilus under simulated Cheddar cheese manufacture was examined. Cell-free wheys (CFW) of the cultures were prepared for analysis by automated spectrophotometry (AS). The maximal growth rate of the lactococci in S. thermophilus R0083 CFW was 13% higher than that noted in their own CFW and three lactococci also gave higher biomass levels (OD_max). During simulated Cheddar cheese fermentations with four paired cultures, one L. lactis strain grew 20% less when paired with S. thermophilus R0083, and an increase in colony forming units (cfu) was found with one other lactococcal strain. Viable counts of S. thermophilus in mixed cultures varied by less than 0.1 log cfu mL^?1. The AS data on OD_max in CFW were useful in predicting the evolution of cfu in the fermented mixed cultures. As a function of strain, the presence of S. thermophilus in a Cheddar fermentation process can enable extended growth of the lactococci.     

奶豆腐来源植物乳杆菌的益生特性评价

以奶豆腐来源的3株植物乳杆菌(Lactobacillus plantarum)为研究对象,分析了菌株的耐酸耐胆盐能力、表面疏水性和体外抗氧化能力,旨在评价菌株的益生特性。结果表明:植物乳杆菌对强酸和高胆盐具有较好的耐受能力,能保持较高的活性;供试菌株均对二甲苯的疏水性作用较强,而对三氯甲烷的疏水作用较弱;3株菌株均具有较好抗氧化活性。     

Nisin抗性嗜热链球菌的筛选

采用两倍稀释法在脱脂乳培养基和乳清培养基中测定了nisin对嗜热链球菌9的最小抑菌浓度(MIC)。在脱脂乳培养基中逐渐增加nisin的浓度，驯化嗜热链球菌9，直到其抗性达到300IU／ml。经生产性能鉴定和遗传性状实验，获得两株适宜做酸奶发酵剂的nisin抗性嗜热链球菌9.31和9．4。     

Exopolysaccharide-producing Streptococcus thermophilus strains as functional starter cultures in the production of fermented milks

Relationships between exopolysaccharide (EPS) production (amount, molecular mass and sugar composition of the EPS) by different Streptococcus thermophilus strains as a functional starter culture, and textural characteristics (viscosity) of fermented milk and yoghurt have been studied. Five interesting heteropolysaccharide-producing strains have been tested. Both S. thermophilus LY03 and S. thermophilus CH101 produced the highest amounts of EPS and also displayed the highest apparent viscosities in fermented milk. S. thermophilus ST 111 and S. thermophilus STD differed considerably in EPS yields, but not in apparent viscosities of fermented milk. In addition, S. thermophilus ST 111 displayed a high variability in EPS amounts when cultivated in milk. In milk medium, S. thermophilus LY03 produced two heteropolysaccharides, a high-molecular-mass (HMM) EPS and a low-molecular-mass (LMM) EPS of the same composition (Gal/Glu/GalNAc=3.4:1.4:1.0). S. thermophilus ST 111 produced only a HMM-EPS (Gal/Rha=2.5:1.0), while S. thermophilus CH 101 (Gal/Glu=1.0:1.0), S. thermophilus ST 113 (Gal/Glu/Rha/GalNAc=1.7:3.9:1.5:1.0) and S. thermophilus STD (Gal/Glu/Rha/GalNAc=3.5:6.2:1.2:1.0) produced only LMM-EPS. Both HMM-EPS and LMM-EPS solutions (S. thermophilus LY03) demonstrated a pseudoplastic character; HMM-EPS solutions of 0.2% (m/v) displayed a high consistency as well. Although its production of high EPS amounts, S. thermophilus LY03 resulted in relatively thin yoghurts, so that texture values did not directly correlate with EPS production capacity. Once structure/function relationships are known, one can determine the molecular properties of the isolated and purified EPS (molecular size, structural characteristics) from candidate strains to predict their potential in texture formation. For a final selection of interesting EPS-producing starter strains one should test the EPS production under yoghurt manufacturing conditions.     

Rheological properties of fermented milk produced by a single exopolysaccharide producing Streptococcus thermophilus strain in the presence of added calcium and sucrose

The effect of calcium supplementation, presence of exopolysaccharides (EPS) and sucrose on the texture of low-fat fermented milk was examined. Reconstituted skim milk (14%) was acidified by 2% glucono-δ-lactone (GDL) or a single strain, EPS-producing Streptococcus thermophilus , and incubated at 42°C. The levels of calcium and sucrose addition ranged from 0 to 9 m m and 0 to 45 m m respectively. Compared with acidified milk (GDL), fermented milk containing EPS showed different rheological properties in the presence of added calcium and sucrose. Calcium addition reduced gel characteristics: G', K, and the second maximum of derivative of shear rate-shear stress; while sucrose addition also increased them but subsequently reduced syneresis. Added calcium or sucrose increased gel hysteresis and reduced apparent yield stress.