Arthrospira platensis Extract: A Non-Invasive Strategy to Obtain Adjunct Attenuated Cultures

This study aims at proposing the use of Arthrospira platensis, commonly known as Spirulina, extract as a non-invasive method to attenuate the growth rate of non-starter adjunct cultures, thus preventing the over-acidification that may occur during cheese manufacturing. A preliminary screening using four different concentrations (0.20%, 0.30%, 0.50%, and 0.70%) of A. platensis extract and four starter and three non-starter lactic acid bacteria strains was performed by impedometric analysis. This allowed us to select one starter and one non-starter strain to be used in the in vitro simulation of a co-culture in milk with the best antimicrobial concentration (0.3%). The growth dynamics of the two selected strains, starter Lactococcus lactis 1426 and non-sarter Lacticaseibacillus rhamnosus 1473, co-cultured for 120 h was monitored by three different approaches: (i) plate counting on M17, for the enumeration of lactococci, and MRS for lactobacilli; (ii) fluorescence microscopic counting of viable and non-viable coccoid Lactococcus lactis 1426 and rod-shaped Lacticaseibacillus rhamnosus 1473 cells; (iii) the overall estimation of co-culture growth behavior by impedometric parameters Lag, Rate, and yEnd. All the data obtained from the in vitro simulation were in agreement, revealing that a slowdown of non-starter growth occurred, while the starter strain was not affected, or slightly stimulated, from the antimicrobial presence. In particular, the growth of Lb. rhamnosus 1473 was delayed without adversely compromise the cells’ integrity, connected with metabolic functions, showing a great potential for use in cheese production

Study on lipid fractions of Streptococcus thermophilus by TLC, GC and GC/MS techniques

In this research the lipid fraction of two strains of Streptococcus thermophilus was studied. Lipids were extracted by applying Folch method and fractioned by thin layer chromatography (TLC). Fatty acid composition was determined both before TLC, on the total fat extracted, and after TLC on diacylglycerol and apolar fractions. Gas chromatographic analysis was performed by using both flame ionization (FID) and mass spectrometer (MS) detector. The main difference between the two strains was the presence of short and medium chain fatty acids in food-born S. thermophilus. Moreover one of the most important bacterial fatty acids, C19 cyclopropane, was detected only in diacylglycerols, which, as reported in literature, are formed transiently as intermediates in the biosynthesis of glycerophospholipids

Phenotypic Diversity of Lactobacillus casei Group Isolates as a Selection Criterion for Use as Secondary Adjunct Starters

Autochthonous lactic acid bacteria (LAB) play a key role in the development of cheese flavor. As the pasteurization treatment on raw milk causes the elimination of LAB, secondary starter cultures are used in cheese manufacture to obtain cheeses with improved and standardized flavors. In this work, strains of the L. casei group isolated from traditional Italian cheeses were screened for their phenotypic features of technological interest for use as secondary starters. Their milk acidifying performance and the production of volatile compounds when grown in milk were evaluated. Simultaneously, the acetoin metabolic pathway presence was screened in the strains and assessed for its transcriptional activation. The results showed that the analyzed strains, despite belonging to taxonomically-related species, vary greatly according to the measured phenotypes. Four strains among the fourteen screened could be potentially used as adjunct cultures for cheese-making processes. The strain that showed the highest production of acetoin upregulated the aspartate pathway. An increased knowledge of volatile compounds’ production and acidifying properties of LAB strains isolated from traditional dairy products might guide the selection of strains for industrial applications

Can the development and autolysis of lactic acid bacteria influence the cheese volatile fraction? The case of Grana Padano

In this study, the relationship between the dynamics of the growth and lysis of lactic acid bacteria in Grana Padano cheese and the formation of the volatile flavor compounds during cheese ripening was investigated. The microbial dynamics of Grana Padano cheeses thatwere produced in two different dairies were followed during ripening. The total and cultivable lactic microflora, community composition as determined by length heterogeneity- PCR (LH-PCR), and extent of bacterial lysis using an intracellular enzymatic activity assaywere compared among cheeses after 2, 6 and 13 months of ripening in two dairies. The evolution of whole and lysed microbiota was different between the two dairies. In dairy 2, the number of total cells was higher than that in dairy 1 in all samples, and the number of cells that lysed during ripening was lower. In addition, at the beginning of ripening (2 months), the community structure of the cheese from dairy 2 was more complex and was composed of starter lactic acid bacteria (Lactobacillus helveticus and Lactobacillus delbrueckii) and NSLAB, possibly arising fromraw milk, including Lactobacillus rhamnosus/Lactobacillus casei and Pediococcus acidilactici. On the other hand, the cheese from dairy 1 that ripened for 2 months was mainly composed of the SLAB L. helveticus and L. delbrueckii. An evaluation of the free-DNA fraction through LH-PCR identified those species that had a high degree of lysis. Data on the dynamics of bacterial growth and lysis were evaluated with respect to the volatile profile and the organic acid content of the two cheeses after 13 months of ripening, producing very different results. Cheese from dairy 1 showed a higher content of free fatty acids, particularly those deriving from milk fat lipolysis, benzaldehyde and organic acids, such as pGlu and citric. In contrast, cheese from dairy 2 had a greater amount of ketones, alcohols, hydrocarbons, acetic acid and propionic acid. Based on these results, we can conclude that in the first cheese, the intracellular enzymes thatwere released fromlysiswere mainly involved in aromaformation,whereas in the second cheese, the greater complexity of volatile compounds may be associated with its more complex microbial composition caused from SLAB lysis and NSLAB (mainly L. rhamnosus/L. casei) growth during ripening

Izdvajanje i identifikacija mikroflore iz kravljeg kolostruma tradicionalnim i molekularnim metodama

Colostrum is the first milk produced by the mammary gland during the early postpartum period. In addition to nutrients, antimicrobial and growth factors, bovine colostrum contains valuable microflora, including members of the genera Lactobacillus and Bifidobacterium, which have been used widely in probiotic food. The aim of this work is to identify and quantify the microflora of colostrum samples from two Holstein dairy cows on the first three postpartum days through culture-dependent 16S rRNA gene sequencing and length heterogeneity polymerase chain reaction (LH-PCR) techniques, using an adapted method to extract the total DNA. Twenty-nine strains isolated in synthetic media were identified by their 16S rRNA gene sequences, revealing two potential probiotic strains (Lactobacillus casei and Bifidobacterium pseudolongum). Colostrum samples were subjected to LH-PCR analysis to obtain fingerprint profiles of the microflora. Lactobacillus casei was the main species present in the samples. The probiotic potential of these bacteria for use in fermented and functional foods remains to be evaluated.Kolostrum je prvo mlijeko, koje proizvodi mliječna žlijezda tijekom ranog postpartalnog razdoblja. Osim što sadrži hranjiva, antimikrobne tvari i hormone rasta, kravlji kolostrum sadržava dragocjenu mikrofloru, uključujući sojeve rodova Lactobacillus i Bifidobacterium što se koriste u proizvodnji probiotičke hrane. Svrha je ovoga rada bila identificirati i kvantificirati mikrofloru u uzorcima kolostruma dvaju Holstein mlječnih krava, uzetih tijekom prva tri dana postpartalnog razdoblja. Mikroorganizmi su određeni sekvenciranjem 16S rRNK gena i LH-PCR metodom kojom se ekstrahira ukupna DNK. U sintetičkoj je podlozi izolirano dvadeset i devet sojeva, identificiranih pomoću sekvencija 16S rRNK gena, te su izdvojena dva potencijalno probiotička soja (Lactobacillus casei i Bifidobacterium pseudolongum). Uzorci su kolostruma analizirani LH-PCR metodom kako bi se utvrdio DNK profil mikroflore, te je utvrđeno da u uzorcima prevladava vrsta Lactobacillus casei. Probiotička svojstva ove bakterije za proizvodnju fermentiranih i probiotičkih proizvoda tek treba istražiti

How the Fewest Become the Greatest. L. casei’s Impact on Long Ripened Cheeses

Members of the Lactobacillus casei group, including species classified currently as L. casei, L. paracasei, and L. rhamnosus, are among the most frequently found species in raw milk, hard cooked, long-ripened cheeses. Starting from very low numbers in raw milk, they become dominant in the cheese during ripening, selected by physical and chemical changes produced by cheese making and ripening. Their presence at different stages of cheese making and ripening is crucial in defining product features. For these reasons, the scientific community has been more and more interested in studying these “tiny but mighty microbes” and their implications during cheese making and ripening. The present paper reviews the current literature on the effect of L. casei in cheeses, with particular reference to the case of Parmigiano Reggiano and Grana Padano, two of the most famous PDO (Protected Designation of Origin) Italian cheeses. Recent advances regarding the selection of new wild strains able to persist until the end of ripening and carrying out slow but crucial activities resulting in specific aromatic features, are also presented

Izdvajanje i identifikacija mikroflore iz kravljeg kolostruma tradicionalnim i molekularnim metodama

Colostrum is the first milk produced by the mammary gland during the early postpartum period. In addition to nutrients, antimicrobial and growth factors, bovine colostrum contains valuable microflora, including members of the genera Lactobacillus and Bifidobacterium, which have been used widely in probiotic food. The aim of this work is to identify and quantify the microflora of colostrum samples from two Holstein dairy cows on the first three postpartum days through culture-dependent 16S rRNA gene sequencing and length heterogeneity polymerase chain reaction (LH-PCR) techniques, using an adapted method to extract the total DNA. Twenty-nine strains isolated in synthetic media were identified by their 16S rRNA gene sequences, revealing two potential probiotic strains (Lactobacillus casei and Bifidobacterium pseudolongum). Colostrum samples were subjected to LH-PCR analysis to obtain fingerprint profiles of the microflora. Lactobacillus casei was the main species present in the samples. The probiotic potential of these bacteria for use in fermented and functional foods remains to be evaluated.Kolostrum je prvo mlijeko, koje proizvodi mliječna žlijezda tijekom ranog postpartalnog razdoblja. Osim što sadrži hranjiva, antimikrobne tvari i hormone rasta, kravlji kolostrum sadržava dragocjenu mikrofloru, uključujući sojeve rodova Lactobacillus i Bifidobacterium što se koriste u proizvodnji probiotičke hrane. Svrha je ovoga rada bila identificirati i kvantificirati mikrofloru u uzorcima kolostruma dvaju Holstein mlječnih krava, uzetih tijekom prva tri dana postpartalnog razdoblja. Mikroorganizmi su određeni sekvenciranjem 16S rRNK gena i LH-PCR metodom kojom se ekstrahira ukupna DNK. U sintetičkoj je podlozi izolirano dvadeset i devet sojeva, identificiranih pomoću sekvencija 16S rRNK gena, te su izdvojena dva potencijalno probiotička soja (Lactobacillus casei i Bifidobacterium pseudolongum). Uzorci su kolostruma analizirani LH-PCR metodom kako bi se utvrdio DNK profil mikroflore, te je utvrđeno da u uzorcima prevladava vrsta Lactobacillus casei. Probiotička svojstva ove bakterije za proizvodnju fermentiranih i probiotičkih proizvoda tek treba istražiti

Lactobacillus paracasei 4341 as adjunct culture to enhance flavor in short ripened Caciotta-type cheese

Caciotta is the name used to define a type of Italian semi-hard cheese Caciotta-type cheese. Due to the short ripening time, pasteurization is necessary to eliminate the potential pathogenic bacteria, which may be present in raw milk, causing also the reduction of ripened cheese flavor. The purpose of this research was to evaluate the effect of a selected wild Lactobacillus paracasei strain experimentally used as adjunct culture to enhance the flavour formation in a short-ripened caciotta-type cheese. An integrated polyphasic approach was used to compare the experimental and control Caciotta produced in a company located in Emilia Romagna region (Italy). It was demonstrated how the L. paracasei 4341 was able to develop in curd and cheese interacting with the acidifying commercial starter. The main acidifying starter species, were differently affected by the presence of the adjunct culture. Streptococcus thermophilus shown comparable behavior in all cheese-making step of control and experimental Caciotta, while Lactobacillus delbrueckii subsp bulgaricus, growth was slowed down by the presence of the adjunct culture during the whole ripening time. The higher amount of volatile compounds and organic acids due to the adjunct L. paracasei 4341 lead to a clear differentiation of the experimental Caciotta respect to the control, in terms of aromatic profile, color, texture and sensorial perception