Lactic Acid Bacteria In Cheese Research Articles

Bacteriocins against biogenic amine-accumulating lactic acid bacteria in cheese: Nisin A shows the broadest antimicrobial spectrum and prevents the formation of biofilms

Cheese is a food in which toxic concentrations of biogenic amines (BA) may be reached, mainly as a consequence of the decarboxylation of determined amino acids by certain lactic acid bacteria (LAB). To maintain the food safety of cheese, environmentally friendly strategies are needed that specifically prevent the growth of BA-producing LAB and the accumulation of BA. The bacteriocins produced by LAB are natural compounds with great potential as food biopreservatives. This work examines the antimicrobial potential of 7 bacteriocin-containing, cell-free supernatants (CFS: coagulin A-CFS, enterocin A-CFS, enterocin P-CFS, lacticin 481-CFS, nisin A-CFS, nisin Z-CFS and plantaricin A-CFS) produced by LAB against 48 strains of the LAB species largely responsible for the accumulation of the most important BA in cheese, i.e., histamine, tyramine and putrescine. Susceptibility to the different CFS was strain-dependent. The histamine-producing species with the broadest sensitivity spectrum were Lentilactobacillus parabuchneri (the species mainly responsible for the accumulation of histamine in cheese) and Pediococcus parvulus. The tyramine-producing species with the broadest sensitivity spectrum was Enterococcus faecium, while Enterococcus faecalis and Enterococcus hirae were among the most sensitive putrescine producers. Nisin A-CFS was active against 31 of the 48 BA-producing strains (the broadest antimicrobial spectrum recorded). Moreover, commercial nisin A prevented biofilm formation by 67% of the BA-producing, biofilm-forming LAB strains. These findings underscore the potential of bacteriocins in the control of BA-producing LAB, and support the use of nisin A as a food grade biopreservative for keeping BA-producing LAB in check and reducing BA accumulation in cheese.

Impact of LAB from Serpa PDO Cheese in Cheese Models: Towards the Development of an Autochthonous Starter Culture.

Serpa is a protected designation of origin (PDO) cheese produced with raw ewes' milk and coagulated with Cynara cardunculus. Legislation does not allow for milk pasteurization and starter culture inoculation. Although natural Serpa's rich microbiota allows for the development of a unique organoleptic profile, it also suggests high heterogeneity. This raises issues in the final sensory and safety properties, leading to several losses in the sector. A possible solution to overcoming these issues is the development of an autochthonous starter culture. In the present work, some Serpa cheese Lactic acid bacteria (LAB)-isolated microorganisms, previously selected based on their safety, technological and protective performance, were tested in laboratory-scale cheeses. Their acidification, proteolysis (protein and peptide profile, nitrogen fractions, free amino acids (FAA)), and volatiles generation (volatile fatty acids (VFA) and esters) potential was investigated. Significant differences were found in all parameters analyzed, showing a considerable strain effect. Successive statistical analyses were performed to compare cheese models and Serpa PDO cheese. The strains L. plantarum PL1 and PL2 and the PL1 and L. paracasei PC mix were selected as the most promising, resulting in a closer lipolytic and proteolytic profile of Serpa PDO cheese. In future work, these inocula will be produced at a pilot scale and tested at the cheese level to validate their application.

Microbial and chemical composition of Cheddar cheese supplemented with prebiotics from pasteurized milk to aging

Microbial and chemical properties of cheese is crucial in the dairy industry to understand their effects on cheese quality. Microorganisms within this fat, protein, and water matrix are largely responsible for physiochemical characteristics and associated quality. Prebiotics can be used as an energy source for lactic acid bacteria in cheese by altering the microbial community and provide the potential for value-added foods, with a more stable probiotic population. This research focuses on the addition of fructooligosaccharides (FOS) or inulin to the Cheddar cheese-making process to evaluate the effects on microbial and physicochemical composition changes. Laboratory-scale Cheddar cheese produced in 2 replicates was supplemented with 0 (control), 0.5, 1.0, and 2.0% (wt/wt) of FOS or inulin using 18 L of commercially pasteurized milk. A total of 210 samples (15 samples per replicate of each treatment) were collected from cheese-making procedure and aging period. Analysis for each sample were performed for quantitative analysis of chemical and microbial composition. The prevalence of lactic acid bacteria (log cfu/g) in Cheddar cheese supplemented with FOS (6.34 ± 0.11 and 8.99 ± 0.46; ± standard deviation) or inulin (6.02 ± 0.79 and 9.08 ± 1.00) was significantly higher than the control (5.84 ± 0.27 and 8.48 ± 0.06) in whey and curd, respectively. Fructooligosaccharides supplemented cheeses showed similar chemical properties to the control cheese, whereas inulin-supplemented cheeses exhibited a significantly higher moisture content than FOS and the control groups. Streptococcus and Lactococcus were predominant in all cheeses and 2% inulin and 2% FOS-supplemented cheeses possessed significant amounts of nonstarter lactic acid bacteria found to be an unidentified group of Lactobacillaceae, which emerged after 90 d of aging. In conclusion, this study demonstrates that prebiotic supplementation of Cheddar cheese results in differing microbial and chemical characteristics.

Symbiotic goat milk ice cream with umbu fortified with autochthonous goat cheese lactic acid bacteria

Autochthonous lactic acid bacteria (LAB) from goat cheese were selected for the preparation of symbiotic goat milk ice cream (GMIC) with the inclusion of umbu fruit (Spondias tuberosa). Its sensory attributes, microbiological and physical-chemical characteristics and its antioxidant capacity were measured in the GMIC produced with autochthonous isolates (GMIC 2) and with commercial cultures (GMIC 1). Three isolates from goat cheese identified as Lactobacillus plantarum, Lactobacillus rhamnosus and Pediococcus acidilactici were selected for their physiological, metabolic, protective, safety and functional characteristics for the production of GMIC 2. GMIC 2 showed greater viability of the microorganisms over time compared to GMIC 1. The selection of autochthonous cultures (AC) significantly influenced (P < 0.01) the protein content in the water phase, melting rate and antioxidant capacity of GMIC 2. The GMIC 2 was designated as tastier (P = 0.0167), and with the greater preference and purchase intention (P < 0.001) among the tasters. The combination of probiotic cultures and umbu fruit sensorially favoured both GMIC. However, the AC have distinct properties that certify the authenticity of the umbu-flavoured GMIC from the Caatinga biome of the northeastern semi-arid region and have potential applications in the local industry.

Viability of Staphylococcus aureus and expression of its toxins (SEC and TSST-1) in cheeses using Lactobacillus rhamnosus D1 or Weissella paramesenteroides GIR16L4 or both as starter cultures

Staphylococcus aureus is one of the main causative agents of food poisoning. This bacterium is an important component of cheese microbiota and plays an important role in foodborne diseases. Another important component of the microbiota is the lactic acid bacterium, which actively participates in processes that define the physicochemical, sensorial, and microbiological features of cheese. Of the various microbiological interactions in cheese, the interaction between lactic acid bacteria and Staph. aureus is most relevant. To this end, we evaluated the viability of Staph. aureus strains and the expression of their enterotoxins in cheeses produced experimentally, using Weissella paramesenteroides GIR16L4 or Lactobacillus rhamnosus D1 or both as starter cultures. Over 7 d, we observed that the presence of lactic acid bacteria did not impair Staph. aureus growth. However, via qPCR we observed a change in the gene expression of staphylococcal enterotoxins, suggesting that molecular communication exists between Staph. aureus strains and lactic acid bacteria in cheese.

Investigation of the Lactic Acid Bacteria in Kazak Cheese and Their Contributions to Cheese Fermentation.

Kazak cheese is a traditional dairy product fermented by lactic acid bacteria (LAB) in Xinjiang. To investigate the LAB in Kazak cheese and their contributions to cheese fermentation, four representative LAB, Streptococcus thermophilus B8, Lactobacillus helveticus B6, Weissella confusa B14, and Lactobacillus rhamnosus B10, were isolated from Kazak cheese and subsequently used to ferment cheeses, which were named StC, LhC, WcC, and LrC, respectively. The result showed that most of the physical and chemical indicators had no significant difference, except for moisture and fat. W. confusa B14 was beneficial to the production of amino acids, whereas S. thermophilus B8 promoted the formation of organic acids and contributed to formation ideal texture property. Furthermore, the four cheeses all possessed a strong fruity aroma, with brandy, sweet, herbaceous, pungent, and fatty aromas being the most prominent in WcC. This is because L. helveticus B6 produced a high concentration of hexanal, nonanal, octanal, 3-methylbutanoic acid, ethyl acetate, ethyl butanoate, isoamyl acetate, and ethyl hexanoate in LhC. Research on the fermentation mechanism of LAB in cheese will provide a theoretical basis for the quality control and industrial production of Kazak cheese.

Peptidoglycan Hydrolases of Probiotic Pediococcus acidilactici NCDC 252: Isolation, Physicochemical and In Silico Characterization

Pediococcus acidilactici NCDC 252 is a Gram-positive lactic acid bacteria (LAB) that is being studied to develop it as probiotic and starter culture for dairy and meat industry. Peptidoglycan hydrolases (PGHs) are also gaining interest because of their antibacterial activity. NCDC 252 genome was searched for PGH encoded genes and product of these genes were in silico characterized for different parameters. Extracellular PGHs of NCDC 252 were precipitated and characterized. PGHs were screened turbidometrically with Bacillus cereus and Staphylococcus albus as substrates. Spectrum of PGH(s) was studied in renaturing SDS-PAGE by varying substrates and renaturing conditions. PGH(s) band intensity decreased in the presence of NaCl whereas some additional secondary lytic bands appeared in the presence of EDTA. Autolysis of LAB in cheese and other fermented food products is associated with release of oligopeptides by proteolysis and other enzyme activities. Autolytic phenotype of P. acidilactici was evaluated and characterized. Lytic activity studies revealed 45–60% activity against Micrococcus lysodeikticus and B. cereus greater than for P. acidilactici. PGH of P. acidilactici exhibited broad growth inhibiting spectrum of antibacterial activity against studied Gram positive and Gram negative bacteria and thus can be promising in controlling pathogenic bacteria. This also confers probiotic attribute to the studied strain.

Major technological differences between an industrial-type and five artisan-type Greek PDO Galotyri market cheeses as revealed by great variations in their lactic acid microbiota

Galotyri is a Greek PDO acid-curd, traditionally ripened, soft spread cheese produced from boiled (>80–90 ℃), naturally acidified ewes’ or ewes’/goats’ milk in the regions of Epirus and Thessaly. In this survey study, the microbiology of an industrial Galotyri cheese variety (11 batches) was compared with five artisan-type Galotyri cheese varieties (1–2 batches each), with emphasis on the species of lactic acid bacteria (LAB) prevailing in the final product. All market cheese samples distributed either in packaged or bulk form, were purchased from retail outlets at Ioannina, Epirus during two sampling periods, 2003–2006 and 2014–2019. A total of 366 (189 industrial and 177 artisan) cheese LAB isolates were identified by key biochemical criteria. Strains of Streptococcus thermophilus and/or Lactobacillus delbrueckii subsp. bulgaricus were isolated nearly as a pure symbiotic starter culture (97.4%) from all industrial batches, labeled as a fresh cheese combining the nutritional properties of yogurt. Conversely, the artisan-type cheeses were ripened and more diversified in terms of LAB ecology. They contained strains of <i>Lactococcus lactis</i> (19.8%), <i>Lb. plantarum</i> (16.9%), <i>S. thermophilus</i> (14.7%), <i>Leuconostoc mesenteroides</i> group (11.3%), <i>Pediococcus inopinatus</i> (7.3%), <i>Enterococcus faecalis</i> (8.5%), <i>E. faecium</i> (6.2%), <i>E. durans</i> (5.1%), <i>Lb. rhamnosus</i> (3.4%), <i>Lb. gasseri</i> (3.4%), <i>Lb. bulgaricus</i> (2.8%) and <i>Lb. acidipiscis</i> (0.6%); the prevalence of each species was cheese product batch-dependent. In conclusion: (i) the industrial Galotyri cheese technology results in a fresh yogurt-like cheese of limited (thermophilic) LAB diversity; (ii) the artisan-type Galotyri cheese technologies may vary significantly, but share a decisive ripening step that favors the predominance of mesophilic LAB species in the final market products; (iii) a regulatory distinction between fresh and ripened varieties of commercial PDO-certified Galotyri cheeses is required.

EFEITO DA ADIÇÃO DO FERMENTO NATURAL SOBRE A CONTAGEM DE BACTÉRIAS LÁTICAS EM QUEIJO MINAS ARTESANAL DO SERRO

Os queijos Minas artesanais possuem grande importância econômica, social, ambiental e política para o estado de Minas Gerais, sendo o responsável pelo sustento direto e indireto de mais de 35 mil famílias. O fermento natural é utilizado pela grande maioria de produtores na forma de soro fermento ou porções de queijos fabricados anteriormente. Todavia o seu uso é indiscriminado o que contribui para a descaracterização do queijo. Até o presente momento não existem estudos envolvendo a utilização do fermento natural na fabricação de queijos Minas artesanais. O objetivo deste estudo foi avaliar o efeito da adição de fermento natural sobre as contagens de bactérias láticas do queijo Minas artesanal do Serro ao longo de 60 dias de maturação. Foram analisados queijos fabricados com e sem fermento natural de três diferentes produtores nos tempos: 8, 20, 40 e 60 dias de maturação, em três repetições. Os resultados mostraram que não houve diferença na contagem de bactérias láticas nos queijos fabricados com e sem fermento natural após oito dias de fabricação. Embora não tenha sido encontrada diferença nas contagens de bactérias láticas dos queijos, certamente há diferença qualitativa na microbiota presente dos queijos provenientes dos dois tratamentos, uma vez que a concentração de sal do fermento natural seleciona certos grupos de bactérias láticas. Estudos de caracterização da microbiota devem ser feitos para que o uso do fermento natural tenha base científica.

0550 Effect of rate of cooling and ripening temperatures on non-starter lactic acid bacteria in cheese

0550 Effect of rate of cooling and ripening temperatures on non-starter lactic acid bacteria in cheese

Evaluation of a differential medium for the preliminary identification of members of the Lactobacillus plantarum and Lactobacillus casei groups

A medium originally developed for differential enumeration of probiotic species in fermented milk (mMRS-BPB, Lee and Lee 2008) was evaluated for its ability to correctly discriminate members of the Lactobacillus plantarum (L. plantarum) and L. casei groups from other species. The medium was tested on 461 strains of lactic acid bacteria (LAB) belonging to eight genera and thirty-five species. Colony morphology was relatively consistent for L. plantarum, L. paraplantarum, L. pentosus, L. paracasei, L. casei, and L. rhamnosus, but, when used alone, was not always sufficient to discriminate these species from other species potentially present in cheese. A procedure based on tree classification was developed to obtain preliminary identification on the basis of colony morphology, cell morphology, and CO2 production from glucose. By combining results of the tree classification procedure and heuristic rules, correct preliminary identification at the species or group level could be obtained in 74.4 % of cases overall, and the percentage of correct identifications was as high as 88-100 % for members of the L. plantarum and L. casei groups. When species belonging to groups that can be easily discriminated by rapid molecular methods were combined, the decision tree allowed to correct identification at the group level for the 95 % of the strains. Logistic regression was used to evaluate the effect of strain, operator, light source, and incubation temperature. Although all factors significantly affected one or more of the characters used for identification, the classification procedure proved to be quite robust. It may be difficult to use mMRS-BPB in the differential enumeration of LAB in cheese, except when species composition is relatively simple; however, it can be used as a simple tool to guide molecular identification in studies focused on the isolation of new strains from cheese.

Application of ARISA to assess the influence of salt content and cation type on microbiological diversity of Cheddar cheese

The structure and dynamics of microbial populations play a significant role during cheese manufacture and ripening. Therefore, fast and accurate methods for identification and characterization of the microbial populations are of fundamental importance to the cheese industry. In this study, we investigate the application of the automated ribosomal intergenic spacer analysis (ARISA) for the assessment of the microbial dynamics in cheeses differing in salt cation level and type. We developed a database of the observed and theoretical length of the 16S-23S intergenic spacer of common lactic acid bacteria (LAB) found in cheese and used the database to describe the structure and dynamics of microbial populations during ripening. Salt content and cation concentration did not significantly influence the overall bacteria structure, except that lower salt levels resulted in enhanced starter survival. Presence of nonstarter LAB was detected by ARISA and denaturing gradient gel electrophoresis (DGGE) after 3months for all the cheeses analysed. ARISA used as fingerprinting method, proved to be a rapid and inexpensive technique for the discrimination of LAB in cheese and demonstrated higher resolution and performance in comparison with DGGE. Microbial communities play important roles during cheese making and ripening, hence rapid inexpensive methods to characterize this microbiota are of great interest to both academic and industrial scientists. The application of automated ribosomal intergenic spacer analysis (ARISA) was used to examine the microbial ecology of Cheddar cheese differing in salt level and type. ARISA is well suited to the analysis of the microbial ecology of cheese during ripening. Additionally, the results confirm that salt concentration influences starter culture survival in the cheese matrix, while significant differences were not observed in the nonstarter lactic acid bacteria.

Research Regarding the Use of Wheat Biodiversity for Obtaining some Cereal-Based Fermented Mashes

Cereal-based fermented beverages are an opportunity to include probiotics and fiber in human nutrition. Nowadays, increasing the interest for rediscovering the benefits, on scientific basis, of traditional fermented beverages based on cereals. Fermented mashes based on different cereal grist, namely Braga in Romanian, is a healthy and nutritious beverage, a source for new functional products with beneficial effects on human health. Physical-chemical and sensorial characteristics of these beverages depend on the ingredients and method of preparation. In this article was analyzed the influence of the five varieties of wheat on the main physical-chemical and sensorial characteristics of the fermented mashes obtained from grounded debranned millet (Panicum miliaceum), different species of wheat (Triticum aestivum - two variety, Triticum durum - one varieties, Triticum spelta - one variety and Triticum turgidum - one variety) and well modified Pilsner barley malt. There have been two series of determinations: first series with ground millet with the disc mill set to 0.1 mm and a second series with ground millet with the disc mill set to 0.4 mm disc gap. For both series, the other grains used were milled to 0.4 mm disc gap. The mashes content is 1:12 = grounded mix (millet : wheat : barley malt = 1.33:1:1): water. After boiling, reconstruction of the initially content with water, filtering and adding of 7% w/w white crystalline sugar those two series of five sweet mashes was inoculated with a mixed culture (0,01% w/w - dry wheat beer yeast and 0,04% w/w - dry cheese lactic acid bacteria), incubated at 30 ºC for 15 h in the thermostat and then stored at 6ºC for 63 h. At beginning and follow-up of fermentation the mashes were analyzed (pH, volatile and total acidity, total and soluble dry substances, alcohol concentration and dynamic viscosity). At the end of fermentation, it was done the sensory evaluation (colour, sweet, sour, taste, texture and stability). All determinations have made in triplicate and data are shown as mean ± SD. The best results have obtained for mashes containing T. Spelta wheat, followed by mashes with T. durum variety. Highest dynamic viscosity and total acidity was recorded for mashes obtained with Triticum turgidum variety of wheat.

Influence of Acidification Method on Composition, Texture, Psychrotrophs, and Lactic Acid Bacteria in Minas Frescal Cheese

The suitability of carbon dioxide (CO2) addition to milk for the manufacture of rennet-coagulated Minas Frescal cheeses was evaluated. The objectives of this work were to evaluate the influence of milk carbonation on Minas Frescal textural, microbiological, and physicochemical parameters as a method of acidification. A control (pH 6.8), two CO2-treated, and two lactic acid-treated (pH 6.4 and 6.0) cheeses were produced and evaluated. Milk preservation promoted the multiplication of lactic acid bacteria in cheeses produced by both processes. The use of lower pH milk showed higher effectiveness over psychrotrophs, proteolytic, and lipolytic psychrotroph counts on the first days of storage, extending the lag phase. From 12 days on this pH, inhibition effect over psychrotrophs was severely reduced, especially in CO2-treated cheeses, where a difference of 1.52 log (day 4) was reduced to 0.23 log (day 12) when comparing treatments of pH 6.0 to 6.4. Cheeses produced with CO2 showed higher hardness, regardless of pH. Moisture and pH decreased with time due to syneresis and the production of lactic acid, respectively. Significant differences were found in fat, protein, and ash contents. Cheeses treated with CO2 showed higher protein/fat proportions. According to the results of this work, CO2-treated thermized milk can be satisfactorily used in the manufacture of Minas Frescal cheese, showing psychrotroph inhibition equivalent to that with added lactic acid and improved hardness.

Níveis de células somáticas sobre a proteólise do queijo Mussarela

Objetivou-se, com a realização deste trabalho, avaliar o efeito do nível de células somáticas sobre a microbiota e a proteólise do queijo Mussarela, durante o período de armazenamento. Foram selecionadas vacas com contagem de células somáticas ≤200mil células/mL; de &gt; 200 a ≤400mil células/mL; de &gt;400mil células/mL a ≤750mil células/mL e &gt;750mil células/mL e que não tinham recebido tratamento com antimicrobianos nos dias que antecederam a coleta da matéria-prima. Os queijos produzidos foram avaliados após 1; 15 e 30 dias de armazenamento para a contagem de coliformes a 35ºC, coliformes a 45ºC, psicrotróficos e bactérias ácido lácticas. Paralelamente, foram determinados os índices de extensão e profundidade da proteólise. O experimento completo foi repetido quatro vezes e o delineamento experimental foi em blocos aleatórios. Na análise estatística, utilizou-se a análise de variância seguida do teste de Tukey, considerando p&lt;0,05 como probabilidade mínima aceitável para diferença entre as médias. O leite com elevada contagem de células somáticas apresentou concentração menor de proteína e maior de nitrogênio não proteico. Observou-se diminuição das bactérias ácido lácticas no queijo elaborado com leite composto de células somáticas &gt;750mil células/mL. Não obstante, ocorreu um aumento significativo na extensão e profundidade da proteólise durante o período de armazenamento, resultados observados nos queijos fabricados com o leite com células somáticas &gt;400mil células/mL. Portanto, para se produzir um queijo Mussarela de boa qualidade torna-se necessário o controle da matéria prima, e esta deve apresentar células somáticas inferiores a 400mil células/mL.

Growth and survival of cheese ripening bacteria on milk fat globule membrane isolated from bovine milk and its monosaccharides

The growth and survival of cheese lactic acid bacteria (LAB) on milk fat globule membrane (MFGM) isolated from bovine milk and on monosaccharides found in MFGM glycoconjugates were studied. The LAB studied were selected lactobacilli strains isolated from Norwegian semi-hard cheese, a starter Lactococcus sp. and the potential cheese contaminant Enterococcus hirae. The bacteria were able to grow on MFGM isolated from bovine milk and to utilize several of the monosaccharides found in the MFGM. Only one strain of lactobacilli was able to utilize N-acetyl-d-galactosamine for growth. Growing lactobacilli on the acylated aminosugars N-acetyl-d-glucosamine or N-acetyl-d-galactosamine induced a decrease in the number of culturable bacteria at an earlier stage than when grown on any of the other monosaccharides. The potential cheese contaminant Ec. hirae showed superior growth and survival abilities compared with the other lactic acid bacteria when grown on the MFGM media and on several of the MFGM monosaccharides.

Basic Physical-Chemical and Sensorial Evaluation of Some Fermented Mashes Produced from Wheat, Husked Millet, Barley Malt and Oat

Fermented mashes based on different cereal grist are a healthy and nutritious beverage, a source for new functional products with beneficial effects on human health. These mashes are a thick consistency and sour-sweet taste, different colors depending on the ingredients and method of preparation. In this study were determined the main physical and chemical characteristics (pH, volatile and total acidity, total and soluble dry substances, alcohol concentration, dynamic viscosity) and was made sensorial analysis of some boiled aqueous mashes obtained from wheat, husked millet, barley malt and oat. Mashes were fermented at 30°C with a mixed culture of mesophilic cheese lactic acid bacteria and wheat beer yeast for 23h, followed by a cold maturation at 4°C for 42h. This study allows exploring the potential of different cereals to produce cereal-based beverages. The best results were obtained for mashes containing a larger amount of barley malt, followed by mashes with oat. Highest dynamic viscosities were recorded for mashes obtained only from wheat and millet. All determinations were made in triplicate and data were shown as mean ± SD.

Characterization of non-starter lactic acid bacteria in traditionally produced home-made Radan cheese during ripening

Two hundred thirteen non-starter lactic acid bacteria isolated from Radan cheese during ripening were identified with both a classical biochemical test and rep-PCR with (GTG)5 primer. For most isolates, which belong to the Lactococcus lactis subsp. lactis, Leuconostoc mesenteroides, Lactobacillus plantarum, Lactobacillus paraplantarum and Enterococcus faecium, a phenotypic identification was in good agreement with rep-PCR identification. Lactococeus lactis subsp. lactis, Enterococcus faecium and subspecies from the Lenconostoc mesenteroides group were the dominant population of lactic acid bacteria in cheese until 10 days of ripening and only one Streptococcus thermophilus strain was isolated from the 5-day-old cheese sample. As ripening progressed, Lactobacillus plantarum became the predominant species together with the group of heterofermentative species of lactobacilli that could not be precisely identified with rep-PCR.

Randomly-amplified microsatellite polymorphism for preliminary typing of lactic acid bacteria from Bryndza Cheese

A high-throughput, medium-discrimination method for preliminary typing and selecting non-identical isolates of lactic acid bacteria in cheeses was developed. RAMP, a PCR with one microsatellite-targeted and one random primer, was used for preliminary typing of 1119 isolates of lactic acid bacteria from Slovak Bryndza cheese. A total of 59 genotypes were identified based on RAMP profiles consisting of 12-23 DNA fragments of 150-3000 bp. For example, 18, 17, 13 and 7 different RAMP-types were identified in Lactobacillus brevis, L. plantarum, L. paracasei and L. fermentum, respectively. The method facilitated well reproducible, medium-discrimination typing of Lactobacillus spp. and Pediococcus spp. at a subspecies level and proved to be suitable for preliminary typing of lactic acid bacteria isolated from cheese.

Key enzymes involved in methionine catabolism by cheese lactic acid bacteria

Cheese microbiota and their enzymatic conversion of l-methionine to volatile sulphur compounds (VSCs) play an important role in aroma formation during cheese ripening. Here, lactic acid bacteria (LAB) strains isolated from raw goats' milk cheeses were screened for the major enzymes critical to the formation of VSCs from l-methionine. A large natural biodiversity in enzyme capabilities and high inter- and intra-species variability was found among the LAB isolates investigated. From those isolates tested, lactococci displayed higher C–S lyase specificities towards the sulphur-containing compounds examined than did Lactobacillus and Leuconostoc, in some cases generating higher levels of VSCs than B. linens, known to be an efficient producer of methanethiol (MTL) and related VSCs. Moreover, these differences in C–S lyase activities (determined spectrophotometrically by measuring the formation of free thiol groups) were shown to correspond with the enzymatic potential of the isolates as determined by visualization of enzymatic activities. This technique could therefore prove valuable for the detection and preliminary characterization of C–S lyase activities among LAB isolates. Lactococci were also found to possess higher aminotransferase activities than lactobacilli and leuconostocs, while glutamate dehydrogenase activities were observed to be highest among Leuconostoc and Lactobacillus spp. Meanwhile, α-keto acid decarboxylase activities were highly variable and were measurable in only a limited number of isolates, mainly lactobacilli. From these data, combining indigenous isolates showing high VSCs-producing capabilities with those that facilitate the completion of the metabolic pathway responsible for degrading l-methionine into volatile compounds may provide an efficient approach to enhance cheese aroma development.