Potential Prebiotic Properties of Whey Protein and Glycomacropeptide in Gut Microbiome.

Proteins in whey have prebiotic and antimicrobial properties. Whey protein comprises numerous bioactive proteins and peptides, including glycomacropeptide (GMP), a hydrophilic casein peptide that separates with the whey fraction during cheese making. GMP has traditionally been used as a protein source for individuals with phenylketonuria and also has prebiotic (supporting the growth of Bifidobacterium and lactic acid bacteria) and antimicrobial activities. GMP supplementation may help positively modulate the gut microbiome, help treat dysbiosis-related gastrointestinal disorders and improve overall health in consumers.

Correction to: Understanding the effects of dietary components on the gut microbiome and human health.

[This corrects the article DOI: 10.1007/s10068-020-00811-w.].

Effects of Whey Protein Supplementation on Inflammatory Marker Concentrations in Older Adults.

Although whey protein isolate (WPI) has been shown to be immunomodulatory, its ability to modulate production of a broad array of inflammatory markers has not previously been investigated in healthy adults. We investigated the effects of daily supplementation with 35 g of WPI for 3 weeks on inflammatory marker concentrations in the blood serum and feces of 14 older adult subjects (mean age: 59). Serum was analyzed using a multiplex assay to quantify the cytokines IFN-γ, IL-1ß, IL-1RA, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12p70, IL-13, IL-17A and TNF-α. Fecal samples were analyzed using an ELISA for the inflammatory markers calprotectin and lactoferrin. Our results yielded high inter-subject variability and a significant proportion of cytokine concentrations that were below our method's limit of quantification. We observed decreases in serum IL-12p70 in the washout phase compared with baseline, as well as the washout stage for fecal lactoferrin relative to the intervention stage. Serum IL-13 was also significantly reduced during the intervention and washout stages. Our data suggest that whey protein supplementation did not significantly alter most inflammatory markers measured but can alter concentrations of some inflammatory markers in healthy older adults. However, our study power of 35% suggests the number of participants was too low to draw strong conclusions from our data.

Fecal Methylmercury Correlates With Gut Microbiota Taxa in Pacific Walruses (Odobenus rosmarus divergens).

OBJECTIVES: Methylmercury metabolism was investigated in Pacific walruses (Odobenus rosmarus divergens) from St. Lawrence Island, Alaska, United States. METHODS: Total mercury and methylmercury concentrations were measured in fecal samples and paired colon samples (n = 16 walruses). Gut microbiota composition and diversity were determined using 16S rRNA gene sequencing. Associations between fecal and colon mercury and the 24 most prevalent gut microbiota taxa were investigated using linear models. RESULTS: In fecal samples, the median values for total mercury, methylmercury, and %methylmercury (of total mercury) were 200 ng/g, 4.7 ng/g, and 2.5%, respectively, while in colon samples, the median values for the same parameters were 28 ng/g, 7.8 ng/g, and 26%, respectively. In fecal samples, methylmercury was negatively correlated with one Bacteroides genus, while members of the Oscillospirales order were positively correlated with both methylmercury and %methylmercury (of total mercury). In colon samples, %methylmercury (of total mercury) was negatively correlated with members of two genera, Romboutsia and Paeniclostridium. CONCLUSIONS: Median %methylmercury (of total mercury) was 10 times higher in the colon compared to the fecal samples, suggesting that methylmercury was able to pass through the colon into systemic circulation. Fecal total mercury and/or methylmercury concentrations in walruses were comparable to some human studies despite differences in seafood consumption rates, suggesting that walruses excreted less mercury. There are no members (at this time) of the Oscillospirales order which are known to contain the genes to methylate mercury, suggesting the source of methylmercury in the gut was from diet and not in vivo methylation.

Shifts of microbiota during cheese production: impact on production and quality.

The high-throughput DNA sequencing (HTS) method is used to identify microbes in cheese and their potential functional properties. The technique can be applied to the microbiota of the cheese processing environment, raw milk, curd, whey, and starter cultures, and be used to improve the quality, safety, and other physicochemical properties of the final product. The HTS method is also utilized to study the microbiota shift of different types of cheeses during processing, as the composition and functional properties of the microbiome provide unique characteristics to different cheeses. Although there are several reviews that focused on microbiota of various types of cheeses, this review focuses on evaluating the microbiota shift of different types of cheese production and highlights key bacteria in each step of the processing as well as microbiota of various types of cheeses. KEY POINTS: • High-throughput sequencing can be applied to identify microbiota in cheese. • Microbiota in cheese is changed during making process and aging. • Starter culture plays an important role to establish microbiota in cheese.

Understanding the effects of dietary components on the gut microbiome and human health.

The gut microbiome is the complex microbial ecosystem found in the gastrointestinal tract of humans and animals. It plays a vital role in host development, physiology and metabolism, and has been implicated as a factor in brain function, behavior, mental health, and many disease states. While many factors, including host genetics and environmental factors, contribute to the composition of the gut microbiome, diet plays a large role. Microorganisms differ in their nutrient requirements, and alterations in host dietary composition can have strong impacts on the microbial inhabitants of the gastrointestinal tract. The health implications of these dietary and microbial changes are relevant as various global populations consume diets comprised of different macronutrient ratios, and many diets promote alterations to recommended macronutrient ratios to promote health. This review will outline the ways in which specific macro- and micronutrients impact the gut microbiome and host health.

Assessment of overall microbial community shift during Cheddar cheese production from raw milk to aging.

Cheese is a fermented dairy product that is made from animal milk and is considered to be a healthy food due to its available nutrients and potential probiotic characteristics. Since the microbes in the cheese matrix directly contribute to the quality and physicochemical properties of cheese, it is important to understand the microbial properties of cheese. In this study, Cheddar cheeses produced on three different dates at the Arbuthnot Dairy Center at Oregon State University were collected to determine the microbial community structure. A total of 773,821 sequencing reads and 271 amplicon sequence variants (ASVs) were acquired from 108 samples. Streptococcus and Lactococcus were observed as the most abundant ASVs in the cheese, which were used as the starter lactic acid bacteria (SLAB). Escherichia coli was detected in the raw milk; however, it was not detected after inoculating with SLAB. According to an alpha diversity analysis, SLAB inoculation decreased the microbial richness by inhibiting the growth of other bacteria present in the milk. A beta diversity analysis showed that microbial communities before the addition of SLAB clustered together, as did the samples from cheese making and aging. Non-starter lactic acid bacteria (NSLAB) were detected 15 weeks into aging for the June 6th and June 26th produced cheeses, and 17 weeks into aging for the cheese produced on April 26th. These NSLAB were identified as an unidentified group of Lactobacillaceae. This study characterizes the changes in the Cheddar cheese microbiome over the course of production from raw milk to a 6-month-aged final product. KEY POINTS: • 271 ASVs were acquired from cheese production from raw milk to 6-month aging. • Addition of SLAB changed the microbial diversity during Cheddar cheese making procedure. • NSLAB were detected more than 15 weeks after aging. Graphical Abstract.

Design of PCR assays to specifically detect and identify 37 Lactobacillus species in a single 96 well plate.

BACKGROUND: Lactobacillus species are used as probiotics and play an important role in fermented food production. However, use of 16S rRNA gene sequences as standard markers for the differentiation of Lactobacillus species offers a very limited scope, as several species of Lactobacillus share similar 16S rRNA gene sequences. In this study, we developed a rapid and accurate method based on comparative genomic analysis for the identification of 37 Lactobacillus species that are commonly used in probiotics and fermented foods. RESULTS: To select species-specific sequences or genes, a total of 180 Lactobacillus genome sequences were compared using Python scripts. In 14 out of 37 species, species-specific sequences could not be found due to the similarity of the 16S-23S rRNA gene. Selected unique genes were obtained using comparative genomic analysis and all genes were confirmed to be specific for 52,478,804 genomes via in silico analysis; they were found not to be strain-specific, but to exist in all strains of the same species. Species-specific primer pairs were designed from the selected 16S-23S rRNA gene sequences or unique genes of species. The specificity of the species-specific primer pairs was confirmed using reference strains, and the accuracy and efficiency of the polymerase chain reaction (PCR) with the standard curve were confirmed. The PCR method developed in this study is able to accurately differentiate species that were not distinguishable using the 16S rRNA gene alone. This PCR assays were designed to detect and identify 37 Lactobacillus species. The developed method was then applied in the monitoring of 19 probiotics and 12 dairy products. The applied tests confirmed that the species detected in 17 products matched those indicated on their labels, whereas the remaining products contained species other than those appearing on the label. CONCLUSIONS: The method developed in this study is able to rapidly and accurately distinguish different species of Lactobacillus, and can be used to monitor specific Lactobacillus species in foods such as probiotics and dairy products.

Microbial communities of a variety of cheeses and comparison between core and rind region of cheeses.

Understanding the microbial community of cheese is important in the dairy industry, as the microbiota contributes to the safety, quality, and physicochemical and sensory properties of cheese. In this study, the microbial compositions of different cheeses (Cheddar, provolone, and Swiss cheese) and cheese locations (core, rind, and mixed) collected from the Arbuthnot Dairy Center at Oregon State University were analyzed using 16S rRNA gene amplicon sequencing with the Illumina MiSeq platform (Illumina, San Diego, CA). A total of 225 operational taxonomic units were identified from the 4,675,187 sequencing reads generated. Streptococcus was observed to be the most abundant organism in provolone (72 to 85%) and Swiss (60 to 67%), whereas Lactococcus spp. were found to dominate Cheddar cheese (27 to 76%). Species richness varied significantly by cheese. According to alpha diversity analysis, porter-soaked Cheddar cheese exhibited the highest microbial richness, whereas smoked provolone cheese showed the lowest. Rind regions of each cheese changed color through smoking and soaking for the beverage process. In addition, the microbial diversity of the rind region was higher than the core region because smoking and soaking processes directly contacted the rind region of each cheese. The microbial communities of the samples clustered by cheese, indicated that, within a given type of cheese, microbial compositions were very similar. Moreover, 34 operational taxonomic units were identified as biomarkers for different types of cheese through the linear discriminant analysis effect size method. Last, both carbohydrate and AA metabolites comprised more than 40% of the total functional annotated genes from 9 varieties of cheese samples. This study provides insight into the microbial composition of different types of cheese, as well as various locations within a cheese, which is applicable to its safety and sensory quality.

Application of Whole-Genome Sequencing to Transposon Mutants of Salmonella Heidelberg.

Transposons are elements widely dispersed among organisms which are able to move and replicate fragments of genomes. The extensive variability in transposons present in most organisms requires extensive identification and interpretation of the resulting transposon mutants after transposon mutagenesis. However, much of this is reliant on utilizing randomness characteristics of transposon to identify essential genes for the organism of interest. Integration of the transposon mutant approach with commercialized next-generation sequencing (NGS) technology has helped to advance transposon identification by sequencing millions of reads generated from a single run on an NGS platform. Transposon sequencing is defined as a gene sequencing methodology that allows for the identification of nonessential genes and the determination of gene function using a random transposon insertional mutagenesis followed by massively parallel sequencing. The detailed protocol will be outlined in this chapter. The genomic DNA integrated with the transposons is sequenced using an NGS platform in order to determine the location of each mutation.