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1.
Nutrients ; 16(8)2024 Apr 19.
Article in English | MEDLINE | ID: mdl-38674902

ABSTRACT

Fermented foods have long been known to have immunomodulatory capabilities, and fermentates derived from the lactic acid bacteria of dairy products can modulate the immune system. We have used skimmed milk powder to generate novel fermentates using Lb. helveticus strains SC234 and SC232 and we demonstrate here that these fermentates can enhance key immune mechanisms that are critical to the immune response to viruses. We show that our novel fermentates, SC234 and SC232, can positively impact on cytokine and chemokine secretion, nitric oxide (NO) production, cell surface marker expression, and phagocytosis in macrophage models. We demonstrate that the fermentates SC234 and SC232 increase the secretion of cytokines IL-1ß, IL-6, TNF-α, IL-27, and IL-10; promote an M1 pro-inflammatory phenotype for viral immunity via NO induction; decrease chemokine expression of Monocyte Chemoattractant Protein (MCP); increase cell surface marker expression; and enhance phagocytosis in comparison to their starting material. These data suggest that these novel fermentates have potential as novel functional food ingredients for the treatment, management, and control of viral infection.


Subject(s)
Cytokines , Fermentation , Nitric Oxide , Phagocytosis , Cytokines/metabolism , Animals , Nitric Oxide/metabolism , Macrophages/immunology , Macrophages/metabolism , Mice , RAW 264.7 Cells , Virus Diseases/immunology
2.
MethodsX ; 11: 102393, 2023 Dec.
Article in English | MEDLINE | ID: mdl-37846356

ABSTRACT

Ex vivo colon model experiments are frequently employed as a means to assess the gut microbiome modulating potential of different foods, food ingredients and dietary supplements. A number of useful models already exist; however, they tend to be relatively low in terms of throughput (3-4 samples per experiment) with a long experiment duration of one to a number of weeks. Therefore, a need for a high-throughput system with a short duration time is required to enable screening of large numbers of samples. Therefore, we report here on the development of a system based on the Applikon micro-Matrix bioreactor which has the capacity to run 24 samples with an experiment duration of 48 h. However, Escherichia coli blooms are a common problem encountered in this model. Here, we describe the factors that contribute to such blooms and provide approaches to address them, providing:•Step by step optimisation of processes involved in conducting ex vivo distal colon experiments using the micro-Matrix bioreactor fermentation platform•Recommended steps for users on how to attenuate E. coli blooms in such ex vivo colon model experiments.

3.
Foods ; 12(18)2023 Sep 19.
Article in English | MEDLINE | ID: mdl-37761191

ABSTRACT

High-throughput DNA sequencing (HTS) was used to study the microbial diversity of commercial traditional Izmir Tulum (IT) and Izmir Brined Tulum (IBT) cheeses from Izmir, Türkiye. Simultaneously, cultivation-dependent methods were used to isolate, identify and characterize bacterial strains displaying probiotic potential. At the phylum level, Firmicutes dominated the microbiota of both cheese types comprising >98% of the population. Thirty genera were observed, with Streptococcus being the most abundant genus and with Streptococcus thermophilus and S. infantarius subsp. infantarius being the most abundant species. Genera, including Bifidobacterium and Chryseobacterium, not previously associated with IT and IBT, were detected. IT cheeses displayed higher operational taxonomic units (OTUs; Richness) and diversity index (Simpson) than IBT cheeses; however, the difference between the diversity of the microbiota of IT and IBT cheese samples was not significant. Three Lacticaseibacillus paracasei strains isolated from IBT cheeses exhibited probiotic characteristics, which included capacity to survive under in vitro simulated gastrointestinal conditions, resistance to bile salts and potential to adhere to HT-29 human intestinal cells. These findings demonstrate that Tulum cheeses harbor bacterial genera not previously reported in this cheese and that some strains display probiotic characteristics.

4.
Foods ; 12(13)2023 Jun 30.
Article in English | MEDLINE | ID: mdl-37444302

ABSTRACT

In recent years, chlorate has become a residue of concern internationally, due to the risk that it poses to thyroid gland function. However, little is known about its occurrence in dairy products of Irish origin. To address this, a study was conducted in which samples of milk (n = 317), cream (n = 199), butter (n = 178), cheese (n = 144) and yoghurt (n = 440) were collected from grocery stores in the Republic of Ireland. Sampling was conducted across spring, summer, autumn and winter of 2021. Samples from multiple manufacturers of each respective dairy product were procured and analysed for chlorate using UPLC-MS/MS. Chlorate was detected in milk, cream, natural, blueberry, strawberry and raspberry yoghurts. Mean chlorate levels detected in these products were 0.0088, 0.0057, 0.055, 0.067, 0.077 and 0.095 mg kg-1, respectively. Chlorate was undetected in butter and cheese (<0.01 mg kg-1). All products sampled, except yoghurt, were found to be compliant with the EU limit for chlorate in milk (0.10 mg kg-1). Some manufacturers produced product with greater incidence and levels of chlorate. Chlorate levels from samples tested at different times of the year did not differ significantly, with the exception of strawberry and raspberry yoghurts which had higher chlorate levels in the winter period.

5.
Eur J Nutr ; 62(4): 1755-1765, 2023 Jun.
Article in English | MEDLINE | ID: mdl-36820883

ABSTRACT

PURPOSE: Studies show that dairy fat consumed in the form of cheese reduce LDL-cholesterol concentration (LDL-c) compared to butter and mechanistic suggestions include the calcium content of cheese leading to enhanced faecal fat excretion. The aim of this study was to test the effect of varying the calcium content within a cheese, on faecal fat excretion as a primary outcome, and blood lipid markers, fasting glucose and calcium excretion as secondary outcomes. METHODS: 7 healthy males (BMI 18-25) participated in this randomized, cross-over control intervention, of 3 × 2 week periods. Diets contained 240 g/day cheese; a High Calcium Cheese (HCC) diet, a Reduced Calcium Cheese (RCC) diet, and a control arm: Reduced Calcium Cheese + CaCO3 Supplement (RCC + Supp) diet. Diets differed in calcium content and form but were otherwise controlled for energy and key macronutrients. Blood and 5-day faecal samples were collected. RESULTS: There was no significant difference in faecal fat excretion (g/day) between the diets (P = 0.066). Percent fat of faecel excretion was higher after RCC + Supp (P = 0.016). None of the individual fatty acids were different. Fasting LDL-c was significantly lower following the HCC diet vs. the other arms (P = 0.002). Faecal Ca was different across all diets (P = 0.001), lowest after RCC, and greatest after RCC + Supp. No differences were observed for fasting blood parameters or changes in anthropometry. CONCLUSION: Varying the calcium content within a cheese matrix significantly affected fasting LDL-c values. Results did not support higher faecal fat excretion as an underlying mechanism, but the high attrition rate was a limitation. Trial registerer Trial Registered at ISRCTN.org, registration number ISRCTN11663659 on 12.07.2022. Retrospectively registered.


Subject(s)
Carcinoma, Renal Cell , Cheese , Kidney Neoplasms , Humans , Male , Blood Glucose , Calcium , Calcium, Dietary , Cholesterol, HDL , Cholesterol, LDL , Cross-Over Studies , Dietary Fats
6.
MethodsX ; 9: 101922, 2022.
Article in English | MEDLINE | ID: mdl-36438855

ABSTRACT

Starter Lactic Acid Bacteria (LAB) are responsible for converting lactose to lactic acid during cheese manufacturing and, as a result, play a critical role in defining the attributes of the final product. There is great interest in isolating novel starter LAB strains to provide alternatives to existing industry cultures or to help enhance the quality and safety of cheeses traditionally made without starter cultures addition [1]. The Fast-Slow Differential Agar (FSDA) medium was developed in 1984 and still remains the standard to rapidly differentiate fast and slow milk-coagulating lactic streptococci and thus avoid screening a large number of isolates for acid production capacity [2]. However, we found that FSDA was unable to selectively isolate fast acid-producing strains from young, traditional, starter-free Izmir Brined Tulum cheeses, due to the presence of a diverse microbiome including Non-Starter LAB and spoilage Gram-negative microbiota [1, 3]. Here, we describe a modified FSDA (mFSDA) with increased selectivity and recovery efficiency towards lactic streptococci, which was successfully used to rapidly isolate potential starters from Tulum cheeses [1] and could similarly outperform FSDA in raw milk cheeses and other varieties containing high levels of "background" microbiota. The main differences between FSDA and mFSDA media consist in the presence of nalidixic acid, ascorbic acid and yeast extract in mFSDA. These targeted additions provide mFSDA with a two-prong selectivity that (I) suppresses unwanted microbiota, and (II) increases the recovery efficiency of lactic streptocci adept to using milk nutrients. Specifically:•Nalidixic acid is an antibiotic that primarily inhibit Gram-negative bacteria [4].•Ascorbic acid and yeast extract stimulate the growth of lactic streptococci [5] and were added to complement skim milk in creating an environment favoring the growth of lactose-positive, casein peptides-utilizing LAB.•The pH indicator bromocresol purple enabled the chromogenic discrimination between LAB with different acid production capability.

7.
Food Res Int ; 160: 111707, 2022 10.
Article in English | MEDLINE | ID: mdl-36076456

ABSTRACT

Izmir Brined Tulum (IBT) Cheese is a traditional semi hard cheese produced in the Aegean region of Türkiye. Lactic acid bacteria (LAB) isolates from IBT cheese samples taken during manufacture and from mature IBT cheeses were investigated for their acid producing capability with the aim of detecting LAB strains responsible for acid production in IBT cheese. Forty two out of 216 isolates decreased the pH of milk to 5.0 or below in 18 h at 37 °C or 42 °C. 16S rRNA Sanger sequencing revealed the presence of LAB species that had not been detected in IBT cheese previously and, indeed, were identified for the first time as the primary acid producers. The majority of these acid producing isolates were identified as putative Streptococcus lutetiensis/Streptococcus infantarius subsp. infantarius (Sii). Further analysis by sequencing the groES/groEL genes of these isolates established that they were Sii. The remaining isolates from cheese samples taken during manufacture were identified as Streptococcus macedonicus, S. thermophilus, Lactococcus lactis subsp. lactis, Lactobacillus delbrueckii subsp. sunkii and L. delbrueckii subsp. indicus and, from mature cheeses, as Enterococcus faecalis and L. delbrueckii subsp. sunkii. Pulsed-field gel electrophoresis (PFGE) results revealed a large genetic diversity amongst the Sii isolates recovered from the IBT cheeses. It was also established that the Sii strains exhibited efficient and consistent acidification ability equivalent to S. thermophilus. Whole-genome sequencing (WGS) and comparative genome analysis of the representative Sii AYB210 strain provided further insights. More specifically, the genome of AYB210 differed from the previously sequenced African dairy isolate Sii CJ18 and the human isolate ATCC®BAA-102™. Modifications in the lactose operon, which may be an indicator of dairy adaptation, were identified and a high number of CRISPR spacers and putative bacteriocin, virulence factor and antibiotic resistance genes were also detected.


Subject(s)
Cheese , Lactobacillales , Lactococcus lactis , Animals , Cheese/microbiology , Humans , Lactobacillales/genetics , Lactobacillus , Lactococcus lactis/genetics , Milk/microbiology , RNA, Ribosomal, 16S/genetics , Streptococcus
8.
Nutrients ; 14(7)2022 Apr 06.
Article in English | MEDLINE | ID: mdl-35406140

ABSTRACT

Fermented foods have been a part of human diet for almost 10,000 years, and their level of diversity in the 21st century is substantial. The health benefits of fermented foods have been intensively investigated; identification of bioactive peptides and microbial metabolites in fermented foods that can positively affect human health has consolidated this interest. Each fermented food typically hosts a distinct population of microorganisms. Once ingested, nutrients and microorganisms from fermented foods may survive to interact with the gut microbiome, which can now be resolved at the species and strain level by metagenomics. Transient or long-term colonization of the gut by fermented food strains or impacts of fermented foods on indigenous gut microbes can therefore be determined. This review considers the primary food fermentation pathways and microorganisms involved, the potential health benefits, and the ability of these foodstuffs to impact the gut microbiome once ingested either through compounds produced during the fermentation process or through interactions with microorganisms from the fermented food that are capable of surviving in the gastro-intestinal transit. This review clearly shows that fermented foods can affect the gut microbiome in both the short and long term, and should be considered an important element of the human diet.


Subject(s)
Fermented Foods , Gastrointestinal Microbiome , Diet , Fermentation , Humans
9.
Nutrients ; 12(6)2020 Jun 04.
Article in English | MEDLINE | ID: mdl-32512787

ABSTRACT

Consuming fermented foods has been reported to result in improvements in a range of health parameters. These positive effects can be exerted by a combination of the live microorganisms that the fermented foods contain, as well as the bioactive components released into the foods as by-products of the fermentation process. In many instances, and particularly in dairy fermented foods, the microorganisms involved in the fermentation process belong to the lactic acid group of bacteria (LAB). An alternative approach to making some of the health benefits that have been attributed to fermented foods available is through the production of 'fermentates'. The term 'fermentate' generally relates to a powdered preparation, derived from a fermented product and which can contain the fermenting microorganisms, components of these microorganisms, culture supernatants, fermented substrates, and a range of metabolites and bioactive components with potential health benefits. Here, we provide a brief overview of a selection of in vitro and in vivo studies and patents exclusively reporting the health benefits of LAB 'fermentates'. Typically, in such studies, the potential health benefits have been attributed to the bioactive metabolites present in the crude fermentates and/or culture supernatants rather than the direct effects of the LAB strain(s) involved.


Subject(s)
Fermented Foods , Food Microbiology , Lactobacillales , Nutritional Physiological Phenomena/physiology , Probiotics , Angiotensin-Converting Enzyme Inhibitors , Antioxidants , Cardiovascular Diseases/prevention & control , Cognition , Cultured Milk Products/microbiology , Diabetes Mellitus, Type 2/prevention & control , Fermentation , Fermented Foods/microbiology , Foodborne Diseases/prevention & control , Gastrointestinal Microbiome , Humans , Immunomodulation
10.
Am J Clin Nutr ; 108(4): 667-674, 2018 10 01.
Article in English | MEDLINE | ID: mdl-30107488

ABSTRACT

Background: Dairy fat consumed as cheese has different effects on blood lipids than that consumed as butter. It is unknown whether the effect is specific to fat interaction with other cheese nutrients (calcium, casein proteins), or to the cheese matrix itself. Objective: We aimed to test the effect of 6 wk daily consumption of ∼40 g dairy fat, eaten within macronutrient-matched food matrices, on markers of metabolic health, in overweight adults aged ≥50 y. Design: The study was a 6-wk randomized parallel intervention; 164 volunteers (75 men) received ∼40 g of dairy fat/d, in 1 of 4 treatments: (A) 120 g full-fat Irish cheddar cheese (FFCC) (n = 46); (B) 120 g reduced-fat Irish cheddar cheese + butter (21 g) (RFC + B) (n = 45); (C) butter (49 g), calcium caseinate powder (30 g), and Ca supplement (CaCO3) (500 mg) (BCC) (n = 42); or (D) 120 g FFCC, for 6 wk (as per A) (n = 31). Group D first completed a 6-wk "run-in" period, where they excluded all dietary cheese before commencing the intervention. Results: There was no difference in anthropometry, fasting glucose, or insulin between the groups at pre- or postintervention. However, a stepwise-matrix effect was observed between the groups for total cholesterol (TC) (P = 0.033) and LDL cholesterol (P = 0.026), with significantly lower postintervention TC (mean ± SD) (5.23 ± 0.88 mmol/L) and LDL cholesterol (2.97 ± 0.67 mmol/L) when all of the fat was contained within the cheese matrix (Group A), compared with Group C when it was not (TC: 5.57 ± 0.86 mmol/L; LDL cholesterol: 3.43 ± 0.78 mmol/L). Conclusion: Dairy fat, eaten in the form of cheese, appears to differently affect blood lipids compared with the same constituents eaten in different matrices, with significantly lower total cholesterol observed when all nutrients are consumed within a cheese matrix This trial was registered at ISRCTN as ISRCTN86731958.


Subject(s)
Calcium/pharmacology , Cheese , Cholesterol/blood , Diet , Dietary Fats/pharmacology , Dietary Proteins/pharmacology , Feeding Behavior , Aged , Blood Glucose/metabolism , Butter , Caseins/pharmacology , Cheese/analysis , Cholesterol, LDL/blood , Female , Humans , Insulin/metabolism , Male , Middle Aged
11.
Food Chem ; 265: 9-17, 2018 Nov 01.
Article in English | MEDLINE | ID: mdl-29884400

ABSTRACT

Appetite can be effectively reduced by targeting the production, secretion, circulation time or receptor of the enteric satiety hormone glucagon-like peptide-1 (GLP-1). The objective of this study was to investigate the potency of Irish Cheddar cheeses to modulate GLP-1 levels. Nine out of ten water-soluble extracts (WSEs) of representative Irish Cheddar cheeses, post 6 months ripening, significantly (p < 0.05) stimulated active GLP-1 secretion from the mouse enteroendocrine cell line STC-1. This secretion was associated with protein content and ripening time of cheese. C-57BL/6 mice (n = 9/10), who received the most potent sample, C2-WSE-8 M, had a significantly reduced cumulative food intake at 6 h compared to control (p < 0.05), but not overall treatment × time effect over a 7 h period. Simulated in vitro gastrointestinal digestion, that models the upper human gut, indicated loss of GLP-1 stimulating activity once C2-WSE-8M entered the intestinal phase, suggesting efficacy of C2-WSE-8M will depend on protection during gut transit.


Subject(s)
Cheese , Glucagon-Like Peptide 1/metabolism , Intestinal Mucosa/metabolism , Intestines/drug effects , Animals , Digestion , Eating , Male , Mice , Mice, Inbred C57BL
12.
J Dairy Sci ; 100(8): 6053-6073, 2017 Aug.
Article in English | MEDLINE | ID: mdl-28624283

ABSTRACT

The purpose of this study was to investigate the effects of pasture-based versus indoor total mixed ration (TMR) feeding systems on the chemical composition, quality characteristics, and sensory properties of full-fat Cheddar cheeses. Fifty-four multiparous and primiparous Friesian cows were divided into 3 groups (n = 18) for an entire lactation. Group 1 was housed indoors and fed a TMR diet of grass silage, maize silage, and concentrates; group 2 was maintained outdoors on perennial ryegrass only pasture (GRS); and group 3 was maintained outdoors on perennial ryegrass/white clover pasture (CLV). Full-fat Cheddar cheeses were manufactured in triplicate at pilot scale from each feeding system in September 2015 and were examined over a 270-d ripening period at 8°C. Pasture-derived feeding systems were shown to produce Cheddar cheeses yellower in color than that of TMR, which was positively correlated with increased cheese ß-carotene content. Feeding system had a significant effect on the fatty acid composition of the cheeses. The nutritional composition of Cheddar cheese was improved through pasture-based feeding systems, with significantly lower thrombogenicity index scores and a greater than 2-fold increase in the concentration of vaccenic acid and the bioactive conjugated linoleic acid C18:2 cis-9,trans-11, whereas TMR-derived cheeses had significantly higher palmitic acid content. Fatty acid profiling of cheeses coupled with multivariate analysis showed clear separation of Cheddar cheeses derived from pasture-based diets (GRS or CLV) from that of a TMR system. Such alterations in the fatty acid profile resulted in pasture-derived cheeses having reduced hardness scores at room temperature. Feeding system and ripening time had a significant effect on the volatile profile of the Cheddar cheeses. Pasture-derived Cheddar cheeses had significantly higher concentrations of the hydrocarbon toluene, whereas TMR-derived cheese had significantly higher concentration of 2,3-butanediol. Ripening period resulted in significant alterations to cheese volatile profiles, with increases in acid-, alcohol-, aldehyde-, ester-, and terpene-based volatile compounds. This study has demonstrated the benefits of pasture-derived feeding systems for production of Cheddar cheeses with enhanced nutritional and rheological quality compared with a TMR feeding system.


Subject(s)
Animal Feed , Animal Nutritional Physiological Phenomena , Cheese/analysis , Milk/chemistry , Animals , Butylene Glycols , Cattle , Diet/veterinary , Female , Poaceae , Taste
13.
mSystems ; 1(3)2016.
Article in English | MEDLINE | ID: mdl-27822529

ABSTRACT

A DNA sequencing-based strategy was applied to study the microbiology of Continental-type cheeses with a pink discoloration defect. The basis for this phenomenon has remained elusive, despite decades of research. The bacterial composition of cheese containing the defect was compared to that of control cheese using 16S rRNA gene and shotgun metagenomic sequencing as well as quantitative PCR (qPCR). Throughout, it was apparent that Thermus, a carotenoid-producing genus, was present at higher levels in defect-associated cheeses than in control cheeses. Prompted by this finding and data confirming the pink discoloration to be associated with the presence of a carotenoid, a culture-based approach was employed, and Thermus thermophilus was successfully cultured from defect-containing cheeses. The link between Thermus and the pinking phenomenon was then established through the cheese defect equivalent of Koch's postulates when the defect was recreated by the reintroduction of a T. thermophilus isolate to a test cheese during the manufacturing process. IMPORTANCE Pink discoloration in cheese is a defect affecting many cheeses throughout the world, leading to significant financial loss for the dairy industry. Despite decades of research, the cause of this defect has remained elusive. The advent of high-throughput, next-generation sequencing has revolutionized the field of food microbiology and, with respect to this study, provided a means of testing a possible microbial basis for this defect. In this study, a combined 16S rRNA, whole-genome sequencing, and quantitative PCR approach was taken. This resulted in the identification of Thermus, a carotenoid-producing thermophile, in defect-associated cheeses and the recreation of the problem in cheeses to which Thermus was added. This finding has the potential to lead to new strategies to eliminate this defect, and our method represents an approach that can be employed to investigate the role of microbes in other food defects of unknown origin.

15.
Int J Food Microbiol ; 197: 77-87, 2015 Mar 16.
Article in English | MEDLINE | ID: mdl-25574847

ABSTRACT

Sporeforming bacteria are a significant concern for the international dairy industry. Spores present in milk survive heat treatments and can persist during downstream processing. If they are present in sufficient numbers in dairy products they can cause spoilage or lead to illness as a result of toxin production. While many reviews have highlighted the threat posed by spores of aerobic bacteria to the dairy industry, few have focused on problems caused by the array of different species of anaerobic sporeformers (Clostridium and related genera) that can be found in milk. This is despite of the fact that members of these bacteria are found throughout the dairy farm environment, and can be toxigenic, neurotoxigenic or spoilage bacteria. This makes the possible presence of Clostridium and related spores in bulk tank milk (BTM) important from both a financial and a public health perspective. In this review dairy associated anaerobic sporeformers are assessed from a number of perspectives. This includes the taxonomy of this group of bacteria, the important subgroup of this genus the "sulphite reducing clostridia" (SRC), how these bacteria are detected in milk products, the epidemiological data regarding pathogenic species and strains within the SRC group as well as the influence of farming practices on the presence of SRC in BTM.


Subject(s)
Clostridium/physiology , Dairy Products/microbiology , Food Microbiology , Milk/microbiology , Animals , Clostridium/classification , Clostridium Infections/epidemiology , Clostridium Infections/microbiology , Clostridium Infections/transmission , Dairying/standards , Foodborne Diseases/epidemiology , Foodborne Diseases/microbiology , Humans , Public Health/standards
16.
Front Microbiol ; 6: 1418, 2015.
Article in English | MEDLINE | ID: mdl-26733963

ABSTRACT

Milk produced in udder cells is sterile but due to its high nutrient content, it can be a good growth substrate for contaminating bacteria. The quality of milk is monitored via somatic cell counts and total bacterial counts, with prescribed regulatory limits to ensure quality and safety. Bacterial contaminants can cause disease, or spoilage of milk and its secondary products. Aerobic spore-forming bacteria, such as those from the genera Sporosarcina, Paenisporosarcina, Brevibacillus, Paenibacillus, Geobacillus and Bacillus, are a particular concern in this regard as they are able to survive industrial pasteurization and form biofilms within pipes and stainless steel equipment. These single or multiple-species biofilms become a reservoir of spoilage microorganisms and a cycle of contamination can be initiated. Indeed, previous studies have highlighted that these microorganisms are highly prevalent in dead ends, corners, cracks, crevices, gaskets, valves and the joints of stainless steel equipment used in the dairy manufacturing plants. Hence, adequate monitoring and control measures are essential to prevent spoilage and ensure consumer safety. Common controlling approaches include specific cleaning-in-place processes, chemical and biological biocides and other novel methods. In this review, we highlight the problems caused by these microorganisms, and discuss issues relating to their prevalence, monitoring thereof and control with respect to the dairy industry.

17.
FEMS Microbiol Rev ; 37(5): 664-98, 2013 Sep.
Article in English | MEDLINE | ID: mdl-23808865

ABSTRACT

Here, we review what is known about the microorganisms present in raw milk, including milk from cows, sheep, goats and humans. Milk, due to its high nutritional content, can support a rich microbiota. These microorganisms enter milk from a variety of sources and, once in milk, can play a number of roles, such as facilitating dairy fermentations (e.g. Lactococcus, Lactobacillus, Streptococcus, Propionibacterium and fungal populations), causing spoilage (e.g. Pseudomonas, Clostridium, Bacillus and other spore-forming or thermoduric microorganisms), promoting health (e.g. lactobacilli and bifidobacteria) or causing disease (e.g. Listeria, Salmonella, Escherichia coli, Campylobacter and mycotoxin-producing fungi). There is also concern that the presence of antibiotic residues in milk leads to the development of resistance, particularly among pathogenic bacteria. Here, we comprehensively review these topics, while comparing the approaches, both culture-dependent and culture-independent, which can be taken to investigate the microbial composition of milk.


Subject(s)
Microbiota , Milk, Human/microbiology , Milk/microbiology , Animals , Food Safety , Food Storage , Humans
18.
J Dairy Sci ; 96(8): 4928-37, 2013 Aug.
Article in English | MEDLINE | ID: mdl-23746589

ABSTRACT

The microbial composition of raw and pasteurized milk is assessed on a daily basis. However, many such tests are culture-dependent, and, thus, bacteria that are present at subdominant levels, or that cannot be easily grown in the laboratory, may be overlooked. To address this potential bias, we have used several culture-independent techniques, including flow cytometry, real-time quantitative PCR, and high-throughput DNA sequencing, to assess the microbial population of milk from a selection of commercial milk producers, pre- and postpasteurization. The combination of techniques employed reveals the presence of a previously unrecognized and diverse bacterial population in unpasteurized cow milk. Most notably, the use of high-throughput DNA sequencing resulted in several bacterial genera being identified in milk samples for the first time. These included Bacteroides, Faecalibacterium, Prevotella, and Catenibacterium. Our culture-independent analyses also indicate that the bacterial population of pasteurized milk is more diverse than previously appreciated, and that nonthermoduric bacteria within these populations are likely to be in a damaged, nonculturable form. It is thus apparent that the application of state-of-the-art approaches can provide a detailed insight into the bacterial composition of milk and could potentially be employed in the future to investigate the factors that influence the composition of these populations.


Subject(s)
Milk/microbiology , Pasteurization/standards , Animals , Cattle , Flow Cytometry/methods , High-Throughput Nucleotide Sequencing/methods , Milk/chemistry , Real-Time Polymerase Chain Reaction/methods
19.
J Biosci Bioeng ; 115(3): 246-52, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23182500

ABSTRACT

To understand high amount of production and detailed processing of antihypertensive peptides, Val-Pro-Pro (VPP) and Ile-Pro-Pro (IPP), in Lactobacillus helveticus CM4 fermented milk, whole genome sequence of the CM4 strain was completed and compared to previously reported whole genome sequence of L. helveticus DPC4571. It revealed 2,028,493 bp of DNA sequence and encoding of 2174 open reading frames in the whole genome sequence with the highest homology to the genome sequence of L. helveticus DPC 4571. Comparative analysis focused on proteolytic enzymes between CM4 and DPC4571 strains revealed existence of 23 kinds of identical intracellular peptidase genes in both strains but no prtY type proteinase gene in DPC4571. Immunoblotting analysis with an antibody raised against the PrtY proteinase showed existence of the 45 kDa PrtY protein in CM4 but not in DPC4571 in the cell extracts. The cell wall-associated proteinase activity was higher in the CM4 than that in the DPC4571 throughout all fermentation period, and the amounts of VPP and IPP in CM4 and DPC4571 fermented milk were correlated with the proteinase activity on the cell wall. Moreover, slight difference of the ß-casein hydrolysates by cell wall-associated extracellular proteinases between CM4 and DPC4571 cells was detected by a MALDI-TOF/TOF analysis. These results suggest that the extracellular proteinase activity might affect on the productivity of VPP and IPP in L. helveticus fermented milk and some peptidases might play important role in following precise processing to release VPP and IPP.


Subject(s)
Antihypertensive Agents/metabolism , Lactobacillus helveticus/enzymology , Oligopeptides/metabolism , Peptide Hydrolases/metabolism , Amino Acid Sequence , Animals , Cell Wall/enzymology , DNA, Bacterial/chemistry , Fermentation , Genome, Bacterial , Lactobacillus helveticus/genetics , Lactobacillus helveticus/metabolism , Milk/metabolism , Molecular Sequence Data , Open Reading Frames , Peptide Hydrolases/genetics , Peptides/chemistry , Sequence Homology, Nucleic Acid
20.
Appl Environ Microbiol ; 78(16): 5717-23, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22685131

ABSTRACT

Here, high-throughput sequencing was employed to reveal the highly diverse bacterial populations present in 62 Irish artisanal cheeses and, in some cases, associated cheese rinds. Using this approach, we revealed the presence of several genera not previously associated with cheese, including Faecalibacterium, Prevotella, and Helcococcus and, for the first time, detected the presence of Arthrobacter and Brachybacterium in goats' milk cheese. Our analysis confirmed many previously observed patterns, such as the dominance of typical cheese bacteria, the fact that the microbiota of raw and pasteurized milk cheeses differ, and that the level of cheese maturation has a significant influence on Lactobacillus populations. It was also noted that cheeses containing adjunct ingredients had lower proportions of Lactococcus species. It is thus apparent that high-throughput sequencing-based investigations can provide valuable insights into the microbial populations of artisanal foods.


Subject(s)
Bacteria/classification , Bacteria/genetics , Biota , Cheese/microbiology , Animals , Cattle , Goats , High-Throughput Nucleotide Sequencing , Ireland , Milk/microbiology , Sheep
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