Single-strain Starter Cultures Research Articles

Starter culture-related changes in free amino acids, biogenic amines profile, and antioxidant properties of fermented red beetroot grown in Poland

Fermentation of two red beet cultivars (Wodan and Alto) with single-strain starter cultures consisting of selected strains of lactic acid bacteria (LAB) of plant origin (Weissella cibaria KKP2058, Levilactobacillus brevis ZF165) and a multi-strain culture (containing W. cibaria KKP2058, L. brevis ZF165, Lactiplantibacillus plantarum KKP1822, Limosilactobacillus fermentum KKP1820, and Leuconostoc mesenteroides JEIIF) was performed to evaluate their impact on betalains, free amino acids, formation of biogenic amines, and antioxidative properties of the juice formed. Next-generation sequencing data analysis used to identify the microbial community revealed that the starter cultures promoted the dominance of the genus Lactobacillus, and decreased the proportion of spoilage bacteria compared to spontaneously fermented juices. Generally, the fermentation process significantly influenced the amount of the analyzed compounds, leading in most cases to their reduction. The observed changes in the studied parameters depended on the starter culture used, indicating different metabolic activities of the LAB strains towards bioactive compounds. The use of multi-strain starter cultures allowed to largely prevent the reduction of betacyanins and histamine formation.

Effect of proteolytic starter culture isolated from Chinese Dong fermented pork (Nanx Wudl) on microbiological, biochemical and organoleptic attributes in dry fermented sausages

The effect of a proteolytic starter culture isolated from Nanx Wudl, on microbiological, biochemical and organoleptic attributes of dry fermented sausages was investigated during processing. Based on preliminary screening, the combination of Staphylococcus xylosus SX16 and Lactobacillus plantarum CMRC6, showing excellent proteolytic activity in vitro, was selected as the multi-strain starter (starter LS). For comparison, the single-strain starter culture of L. plantarum CMRC6 (starter LB) and non-inoculated control were also tested. During fermentation, lactic acid bacteria and staphylococci dominated the microbiota and suppressed the Enterobacteriaceae growth in LS-inoculated sausages. The addition of LS starter accelerated acidification and proteolysis during ripening, improving the contents of total free amino acids and several essential free amino acids (Phe, Ile and Leu). Volatile compounds analysis revealed that LS-fermented sausage showed the highest abundance of 3-methyl-1-butanol, probably due to the inoculated S. xylosus. The inoculation of LS starter improved the sensory properties of sausages, especially the flavor attribute. Therefore, S. xylosus SX16 and L. plantarum CMRC6 are promising candidates for inclusion as multi-strain starters in the manufacture of gourmet fermented dry sausage.

Utilization of indigenously isolated single strain starter cultures for the production of sourdough bread

<p>Sourdoughs were prepared with <em>Saccharomyces cerevisiae</em> (T<sub>0</sub>) and indigenously isolated starter cultures i.e<em> Lactobacillus brevis (</em>T<sub>1</sub>)<em>, Lactobacillus fermentum</em> (T<sub>2</sub>) and<em> Lactobacillus plantarum </em>(T<sub>3</sub>). Breads were prepared from all sourdoughs samples in triplicate and analyzed for pH, Total Titratable Acidity (TTA), loaf volume, microbial characteristics (total plate count and fungal count) and sensory profile (internal and external) in triplicate. The breads prepared from <em>Saccharomyces cerevisiae</em> (T<sub>0</sub>) exhibited the highest pH with the lowest TTA while T<sub>1 </sub>showed the lowest pH with the highest TTA. The T<sub>0</sub> breads got the highest values for loaf volume followed by T<sub>1</sub>. The breads produced with the addition of hetero-fermentative starter cultures (T<sub>1 </sub>and T<sub>2</sub>) showed resistance against the growth of the contaminating microorganisms. In the sensory evaluation, the breads produced with<em> </em>T<sub>1 </sub>ranked the best for color (crust and crumb), taste, aroma, texture and overall acceptability by the panelists. </p>

Two efficient nitrite-reducing Lactobacillus strains isolated from traditional fermented pork (Nanx Wudl) as competitive starter cultures for Chinese fermented dry sausage

Lactic acid bacteria isolated from traditional Dong pork product (Nanx Wudl) were investigated for their potential as starter cultures for Chinese fermented dry sausages. Based on preliminary screening, Lactobacillus plantarum CMRC6 and Lactobacillus sakei CMRC15, both showing excellent nitrite-reducing capacity, were used as single-strain starter cultures. For comparison, a commercial composite starter was also tested. In CMRC6 and CMRC15-inoculated sausages, lactic acid bacteria dominated the microflora and improved the microbiological safety by suppression of Enterobacteriaceae growth. Nitrite content of all inoculated sausages declined rapidly during ripening compared to non-inoculated. Texture profiles analysis showed inoculated sausages had more pronounced textural development during ripening. Sensory evaluation indicated CMRC6 and CMRC15-fermented sausages had comparable or more desirable organoleptic characteristics than sausage made with commercial starters. Therefore, CMRC6 and CMRC15 are promising candidates as multi-functional starter cultures for microbiological safety and residual nitrite control in gourmet Chinese dry sausage production.

Evaluation of a single strain starter culture, a selected inoculum enrichment, and natural microflora in the processing of Tonda di Cagliari natural table olives: Impact on chemical, microbiological, sensory and texture quality

The processing of natural table olives is still empirical and far to be controlled. Taking natural fermentation (NF) as control, the performances of a single strain LAB starter culture (SSL) and a selected inoculum enrichment (SIE) were compared. The results showed that the SIE was more efficient in controlling spoiling microflora (Enterobacteriaceae spp.) compared to SSL and NF. Indeed, Enterobacteriaceae in SIE samples were not detectable starting from 10 days, while in SSL and NF samples starting from 30 days. Both starters rapidly acidified the brine, lowering the pH to safety levels (<4.0) after 12 days, while NF samples reached pH 4.3 after 45 days, and these values were kept constant up to 150 days. Moreover, both starters showed debittering activity, as stated through sensory analyses and HPLC analysis of phenols. Compared to NF samples, SIE and SSL extracts retained more hydroxytyrosol at the end of the process, and this could partially account for their higher scavenging activity. Texture profile analysis (TPA) showed that olives processed with SIE inoculum were firmer and more elastic, compared to SSL inoculated olives, thus resulting more similar to NF samples.

Prevalence and impact of single-strain starter cultures of lactic acid bacteria on metabolite formation in sourdough

Flavour of type II sourdoughs is influenced by the ingredients, processing conditions, and starter culture composition. It is, however, not fully clear to what extent different sourdough lactic acid bacteria (LAB) contribute to flavour. Therefore, two types of flour (rye and wheat) and different LAB starter culture strains were used to prepare sourdoughs, thereby leaving the yeast microbiota uncontrolled. All LAB starter culture strains tested were shown to be prevalent and to acidify the flour/water mixture to pH values between 3.1 and 3.9 after 24h of fermentation. Multiple aldehydes, alcohols, ketones, and carboxylic acids were produced by the sourdough-associated microbiota throughout the fermentation period. Based on the organoleptic evaluation of breads produced with these sourdoughs, five LAB strains were selected to perform prolonged wheat and rye fermentations as to their capacity to result in an acidic (Lactobacillus fermentum IMDO 130101, Lactobacillus plantarum IMDO 130201, and Lactobacillus crustorum LMG 23699), buttermilk-like (Lactobacillus amylovorus DCE 471), or fruity flavour (Lactobacillus sakei CG1). Upon prolonged fermentation, higher metabolite concentrations were produced. For instance, L. sakei CG1 produced the highest amounts of 3-methyl-1-butanol, which was further converted into 3-methylbutyl acetate. The latter compound resulted in a fruity banana flavour after 48h of fermentation, probably due to yeast interference. Rye fermentations resulted in sourdoughs richer in volatiles than wheat, including 3-methyl-1-butanol, 2-phenylethanol, and ethyl acetate.

The origin of single strain starter culture usage for commercial Cheddar cheesemaking

The technology of using single strain starter cultures for commercial Cheddar cheesemaking is commonly believed to have originated in New Zealand in about 1934. However, W. S. Sutton of the New South Wales Department of Agriculture isolated pure strains of Streptococcus cremoris and, along with A. B. Shelton, conducted cheesemaking trials at the Hawkesbury Agricultural College, now the University of Western Sydney, from at least September 1930. Pure strains were being used in factories by October 1930 and, by July 1932, at least six factories were using pure single cultures. The technology was taken to New Zealand from New South Wales by Dr Moir.

Use of response surface methodology to investigate the effects of processing conditions on sourdough wheat bread quality

Response surface methodology was used to investigate the influence of three factors, sourdough fermentation time, proof time and amount of yeast addition on the quality of sourdough wheat bread. Each predictor variable was tested at five levels. Sourdough fermentation times were 5, 11, 20, 29 and 35 h, yeast addition rates were 0.05, 0.75, 1.77, 2.80 and 3.50% (flour weight basis) and proof times were 16, 40, 75, 110 and 134 min. The performance of two different starter culture types, a mixed strain starter culture called Bocker Reinzucht–Sauerteig Weizen and a single strain starter culture of Lactobacillus brevis, was compared by separately completing the experimental design for each. Independently non-acidified control bread was prepared. A range of loaf quality parameters was determined including pH, total titratable acidity, loaf height, specific volume, crumb mean cell area and crumb hardness. Overall breads with better specific volume values were achieved with the use of sourdough relative to the control. Results indicated that maximum loaf specific volume was achieved when L. brevis sourdough was used particularly when it was used in conjunction with a high rate of yeast. Given a lower rate of yeast addition however, the mixed strain starter culture yielded better bread.

Growth reduction of Listeria spp. caused by undefined industrial red smear cheese cultures and bacteriocin-producing Brevibacterium lines as evaluated in situ on soft cheese.

The undefined microbial floras derived from the surface of ripe cheese which are used for the ripening of commercial red smear cheeses have a strong impact on the growth of Listeria spp. In some cases, these microbial consortia inhibit Listeria almost completely. From such undefined industrial cheese-ripening floras, linocin M18-producing (lin+) (N. Valdés-Stauber and S. Scherer, Appl. Environ. Microbiol. 60:3809-3814, 1994) and -nonproducing Brevibacterium linens strains were isolated and used as single-strain starter cultures on model red smear cheeses to evaluate their potential inhibitory effects on Listeria strains in situ. On cheeses ripened with lin+ strains, a growth reduction of L. ivanovii and L. monocytogenes of 1 to 2 log units was observed compared to cheeses ripened with lin strains. Linocin M18 activity was detected in cheeses ripened with lin+ strains but was not found in those ripened with lin strains. We suggest that production of linocin M18 contributes to the growth reduction of Listeria observed on model red smear cheeses but is unsufficient to explain the almost complete inhibition of Listeria caused by some undefined microbial floras derived from the surface of ripe cheeses.

A Strategy for Rotation of Different Bacteriophage Defenses in a Lactococcal Single-Strain Starter Culture System

A new strategy for starter culture rotations was developed for a series of phage-resistant clones genetically derived from a single strain of Lactococcus lactis subsp. lactis. Phage-resistant derivatives carrying different defense systems were constructed via conjugation with various plasmids encoding abortive infection (Abi/Hsp) and/or restriction and modification (R/M) systems of different specificity. The plasmids included pTR2030 (Hsp R/M), pTN20 (Abi R/M), pTRK11 (R/M), and pTRK68 (R/M). Selected phage-resistant transconjugants or transformants were evaluated in different rotation sequences through cycles of the Heap-Lawrence starter culture activity test in milk contaminated with phage and whey from the previous cycle. When used in consecutive sequence, derivative strains carrying the R/M systems encoded by pTN20, pTRK11, and pTRK68 retarded phage development when the initial levels of phage contamination were below 10 PFU/ml but not when levels were increased to 10 PFU/ml. Use of a derivative bearing pTR2030 (Hsp R/M) at the beginning of the rotation prevented phage development, even when the initial levels of phage contamination were high (10 PFU/ml). Alternating the type and specificity of R/M and Abi defenses through the rotation prevented phage proliferation and in some cases eliminated contaminating phages. A model rotation sequence for the phage defense rotation strategy was developed and performed successfully over nine cycles of the Heap-Lawrence starter culture activity test in the presence of high-titer commercial phage composites. This phage defense rotation strategy is designed to protect a highly specialized Lactococcus strain from phage attack during continuous and extended use in the dairy industry.

Effect of cell-free milk culture liquid of Enterococcus faecalis subsp. liquefaciens on growth, acid production and proteolysis of some commercial starter cultures in milk

Inclusion of 50 μl/ml cell-free culture liquid of Enterococcus faecalis var. liquefaciens in skim milk intended for culturing separately 20 multi- and single-strain starters did not increase the acid production by the starters and neither the cell growth nor the proteolysis was stimulated. On the contrary, both growth and acid production were variably inhibited, with the reduction in acid production not always resulting from growth inhibition. The inhibition ranged from 0 to 99% for growth and 0.5 to 62% for acid production and was maximum by heating the cell-free culture liquid at 100°C for 5 min. Growth of the single-strain starter cultures was affected by the heated but not the unheated culture liquid in general. The crude inhibitor proved to be non-neutralizable, non-dialyzable, sensitive to proteolysis and resistant to heat (100°C for 30 min) at 0.59% acidity, implicating bacteriocin-like agent as a participant in the antimicrobial activity. Enterococcin positive strains of Enterococcus faecalis are, therefore, unsuitable for inclusion in the multi-strain starter cultures in cheese making even though these variants might have a good ability to grow at low temperatures and exhibit excellent fermentative and proteolytic characteristics in addition to high salt-tolerance. Multiplication of these strains in raw milk might also result in failure of the growth and acid production by the starter cultures.

Use of Galactose-Fermenting Streptococcus thermophilus in the Manufacture of Swiss, Mozzarella, and Short-Method Cheddar Cheese

Galactose-fermenting (galactose-positive) strains of Streptococcus thermophilus, alone and combined with galactose-positive and galactose-negative strains of Lactobacillus bulgaricus, were used as starter cultures in the manufacture of Swiss and Mozzarella cheese and were paired with Streptococcus lactis (also galactose-positive) in short-method Cheddar cheese manufacture. Experimental Swiss cheese made with the galactose-positive Streptococcus thermophilus starter alone contained a large amount of galactose (ca. 26 to 28 µmol/g of curd) 28h after hooping compared with control Swiss (< 2 µmol/g) made with a nongalactose fermenting strain of Streptococcus thermophilus and a galactose-positive strain of Lactobacillus bulgaricus. Mozzarella and short-method Cheddar made with only galactose-positive Streptococcus thermophilus also contained large amounts of galactose. Swiss cheese made with a galactose-positive strain of Streptococcus thermophilus and a galactose-negative strain of Lactobacillus bulgaricus had little galactose remaining after 28h, indicating that the Lactobacillus had a stimulatory effect on galactose metabolism in Streptococcus thermophilus. These results indicate that galactose-fermenting Streptococcus thermophilus may have limited potential when used as single strain starter cultures in Swiss cheese, but may be useful when combined with galactose-positive Lactobacillus in the manufacture of Mozzarella cheese.

Production of Staphylococcal Enterotoxin A in Cheddar and Colby Cheeses1,2,3

Enterotoxin A production by Staphylococcus aureus in Cheddar and Colby cheeses was determined by inoculating milk with different initial S. aureus populations and making cheese using normal and inhibited starter cultures. Detectable amounts of enterotoxin A were observed more frequently in Colby than in Cheddar cheese, both made with normal starter. With initial S. aureus inocula of at least 35,000 to 250,000 per milliliter of milk, enterotoxin A was detected when at least 15million S. aureus per gram in Colby and 28million in Cheddar developed. Starter failure induced by lactic bacteriophage during the early stages of cheese-making (draining or prior), resulted in extensive growth of S. aureus and detectable enterotoxin A, even with initial S. aureus populations of 1 to 2 thousand per milliliter of milk. Under these conditions, minimal S. aureus populations associated with enterotoxin A were about 3 to 5million pergram. Partial failure of mixed-type multiple-strain starter culture or late failure (beyond the milling in Cheddar cheese) of single strain starter culture also resulted in enterotoxin A without extensive growth of S. aureus, 3 to 11million per gram. Enterotoxin A persisted for over 3 years in Cheddar cheese made with normal or inhibited starter.

STUDIES WITH A BACTERIOPHAGE INHIBITORY MEDIUM: I. INHIBITION OF PHAGE AND GROWTH OF SINGLE STRAIN LACTIC STREPTOCOCCI AND LEUCONOSTOC

SummaryA phage resistant starter culture medium was examined for its ability to inhibit bacteriophages under several conditions. The medium also was tested for its ability to support growth of single strain starter culture organisms and for its effect on viability of starter bacteria during storage at 2 and −20 C. When prepared according to manufacturer's directions, the medium prevented phage proliferation and eliminated phages from infected cultures within three transfers. Preparation of the medium with water of 500 ppm hardness did not affect its phage-inhibiting properties. Single strains of Streptococcus lactis, Streptococcus cremoris and Streptococcus diacetilactis grew well in the medium though there was variation between strains; Leuconostoc species were not able to grow well. Storage survival of starter organisms in the phage resistant medium also revealed considerable variation between strains; Leuconostoc sp. and S. diacetilactis survived poorly when stored in the medium, especially at 2 C, while S. lactis and S. cremoris survived well when held at −20 C.

Some observations on slow and fast acid-producing variants of strains of Streptococcus cremoris and Str. lactis used as cheese starters

Improved growth of Streptococcus cremoris 924 was obtained in the presence of CO2.When fast and slow acid-producing variants of Str. cremoris 924 were mixed and the mixture subcultured daily the resulting cultures were all active acid producers, irrespective of the initial ratio of fast to slow cells. A balance is established between the two types of cell; probably this will vary with the cultural conditions and vary between strains of Str. cremoris and Str. lactis for any particular conditions of maintenance.Providing the strains are correctly selected, single-strain starter cultures can be maintained by daily subculture in litmus milk, apparently indefinitely. Activity varies with the present method within wide limits and this variation seems to be a function of the culture.Freeze drying is a satisfactory method of maintaining active single-strain starters.The author wishes to thank Miss Shelagh Gibbs for technical assistance, Miss H. R. Chapman for making the cheese, and Messrs. Cow and Gate, Ltd., for supplying the dried milk powder for these experiments.

SEASONAL VARIATION IN CHEESE STARTER ACTIVITY

Summary:Depression of activity of certain single strain starter cultures in H.T.S.T. pasteurized milk during the October‐April season was observed, but the effect was removed by momentary boiling. The addition of a small percentage of such milk to autoclaved milk reduced the activity of these cultures. These ‘inhibited’cultures were unable to grow in modified Niven's medium even when additional known growth factors were added. Using a whey agar medium, active ‘uninhibited’colonies may be picked and a ‘failing culture’revived.An hypothesis postulating an inhibitor‐growth factor antagonism is advanced, to explain the findings.