Whey and Buttermilk-Based Formulas Modulate Gut Microbiota in Mice with Antibiotic-Induced Dysbiosis

Scope

Diet is one of the main factors that modifies intestinal microbiota composition. The search for foods that can reverse situations of intestinal dysbiosis such as that induced by antibiotics is of great interest. Buttermilk and whey are the main by-products produced by the dairy industry containing bioactive compounds. The aim of this study is to investigate the ability of whey and buttermilk-based formulas supplemented with lactoferrin and milk fat globule membrane (MFGM) to modulate the effects of clindamycin on mouse intestinal microbiota.

Methods and results

Male C57BL/6 mice are treated with saline (control), clindamycin (Clin), a formula containing whey (F1) or buttermilk (F2), Clin+F1 or Clin+F2, and their fecal microbiota profiles are analyzed by sequencing of 16S rRNA gene using the MinION device. Clin induces alterations in both the composition and metabolic functions of the mice intestinal microbiota. The treatment with F1 or F2 reverses the effects of clindamycin, restoring the levels of Rikenellaceae and Lactobacillaceae families and certain pathways related to short-chain fatty acids production and tetrahydrofolate biosynthesis.

Conclusion

Whey and buttermilk supplemented with lactoferrin and MFGM may be a bioactive formula for functional foods to prevent or restore microbiota alterations induced by antibiotic administration.     

Infant formula supplemented with 1,3-olein-2-palmitin regulated the immunity,gut microbiota,and metabolites of mice colonized by feces from healthy infants

Infant formula is currently an important food to cope with insufficient breastfeeding. Although 1,3-olein-2-palmitin (OPO) has been used in infant formula, its effects on the immune system, gut microbiota, and metabolites for infants remain unclear. This study constructed a mouse model of colonizing healthy infant feces using antibiotic treatment and fecal microbial transplantation. Thus, the gap between the infant formula supplemented with OPO and human milk in mouse serum biochemistry, immune system, intestinal microbiota, short-chain fatty acid production, and metabolites was evaluated. Our results showed that regarding IL-9, IL-10 levels, fecal secretory IgA, and endotoxin, formula supplemented with OPO and human milk types had comparable levels. Additionally, OPO slightly increased the content of short-chain fatty acids. The 16S rRNA gene sequence analysis and metabonomics analysis demonstrated that feeding different foods affects the gut microbiota of mice; in particular, supplementing formula feeding with OPO enriched the abundance of bifidobacteria. Furthermore, feeding different foods leads to unique intestinal content of metabolites, and the gut microbiota regulates the metabolites' differences. Our results reveal a brand new perspective of OPO regarding gut microbiota and metabolites.     

饮食、肠道微生物与健康的关系研究进展

肠道微生物是膳食与人体健康的桥梁,与营养物质的吸收和代谢密切相关,是人体重要的“微生物器官”。同时,研究学者们也已经清楚地认识到这个重要的“微生物器官”与人体生理功能、健康和疾病关系密切,对人体的重要性不容忽视。为此,本文就近几年来有关膳食对肠道微生物结构以及代谢产物影响的研究进行综述,并介绍肠道微生物与人体生理功能及健康的关系,旨在为未来研究饮食、肠道微生物与健康的关系提供参考。     

Changes in the rumen bacterial community in response to sunflower oil and fish oil supplements in the diet of dairy sheep

Rumen microbial biohydrogenation of dietary unsaturated fatty acids has a major effect on the process of developing healthier dairy products. This study aimed to investigate in vivo the effect of diet supplementation with sunflower (SO) and fish (FO) oils on the rumen bacterial community in dairy sheep. First, 32 lactating ewes, divided in 8 lots of 4 animals each (2 lots per treatment), were fed a high-concentrate total mixed ration supplemented with 0, 2% SO, 1% FO, or 2% SO plus 1% FO. After 21 d, rumen fluid samples were taken from each lot for DNA extraction and fluorescence in situ hybridization (FISH) analysis. In a second experiment, 5 cannulated ewes were first fed the same TMR, with the exception of a higher forage level, and then changed to the same diet supplemented with 2% SO plus 1% FO. After 0, 3, and 10 d, rumen content samples were taken for DNA extraction and FISH analysis (fluid). Total bacteria and the Butyrivibrio group were studied in microbial DNA by terminal RFLP analysis (T-RFLP), and real-time PCR was used to quantify Butyrivibrio bacteria that produce vaccenic acid or stearic acid. In rumen fluid samples, total bacteria and clostridial clusters IX and XIV were analyzed by FISH. Dietary supplementation with SO plus FO seemed to induce important changes in the total bacteria and Butyrivibrio populations, and a high interindividual variation was observed, and the speed of the effect of the lipid supplementation depended on the individual microbial composition. Analysis by T-RFLP and FISH showed increases in cluster IX bacteria with SO plus FO supplementation, presumably Quinella-like microorganisms. The abundances of vaccenic acid- and stearic acid-producing Butyrivibrio relative to total bacteria, estimated by real time PCR, were low (0.28 and 0.18%, respectively, in rumen fluid, and 0.86 and 0.81% in rumen contents) and only that of SA-producing bacteria seemed to be reduced by diets containing FO, although differences were only significant in lactating ewes. The T-RFLP analysis showed a variable effect of lipid supplementation on different bacteria of the family Lachnospiraceae, which includes the cultured bacteria known to be actively involved in rumen biohydrogenation. These results suggest that the latter bacteria do not play a dominant role in this process, and therefore other as-yet-uncultivated microorganisms might be more relevant.     

Effects of a probiotic fermented milk beverage containing Lactobacillus casei strain Shirota on defecation frequency,intestinal microbiota,and the intestinal environment of healthy individuals with soft stools

The effects of drinking a fermented milk beverage that contains Lactobacillus casei strain Shirota (LcS) at 40 billion bacterial cells/bottle for 4 weeks (probiotics, 1 bottle/day) on defecation frequency, intestinal microbiota and the intestinal environment of healthy individuals with soft stools were evaluated. Thirty-four healthy adults who had soft stools were randomised into 2 groups, and the effects of a regular 4-week intake of probiotics were evaluated by a placebo-controlled, double-blind, parallel-group comparative design. Defecation frequency significantly decreased after the 4-week intake period compared with before the probiotic treatment. The stool quality significantly improved (hardened) compared to the placebo. Also, the water content of the stools was lower in the probiotic group than in the placebo group. Live LcS was recovered at 6.9 ± 1.3 and 7.2 ± 0.8 log₁₀ CFU per 1 g of stool after 2 and 4 weeks, respectively, of probiotic treatment. The number of bifidobacteria in the stools also increased significantly compared with the level before starting the probiotics. The organic acid levels (total, acetic acid, propionic acid, and butyric acid) significantly increased compared with the level before intake in both the probiotic and placebo groups, but they returned to the original levels after the end of the intake period. These results suggest that probiotic fermented milk beverage has an intestine-conditioning effect by improving the frequency of defecation and stool quality and increasing the intrinsic bifidobacteria in healthy individuals with soft stool.     

Rumen bacterial community evaluated by 454 pyrosequencing and terminal restriction fragment length polymorphism analyses in dairy sheep fed marine algae

Developing novel strategies to increase the content of bioactive unsaturated fatty acids (FA) in ruminant-derived products requires a deeper understanding of rumen biohydrogenation and bacteria involved in this process. Although high-throughput pyrosequencing may allow for a great coverage of bacterial diversity, it has hardly been used to investigate the microbiology of ruminal FA metabolism. In this experiment, 454 pyrosequencing and a molecular fingerprinting technique (terminal restriction fragment length polymorphism; T-RFLP) were used concurrently to assess the effect of diet supplementation with marine algae (MA) on the rumen bacterial community of dairy sheep. Eleven lactating ewes were divided in 2 lots and offered a total mixed ration based on alfalfa hay and concentrate (40:60), supplemented with 0 (control) or 8 (MA) g of MA/kg of dry matter. After 54 d on treatments, animals were slaughtered and samples of rumen content and fluid were collected separately for microbial analysis. Pyrosequencing yielded a greater coverage of bacterial diversity than T-RFLP and allowed the identification of low abundant populations. Conversely, both molecular approaches pointed to similar conclusions and showed that relevant changes due to MA addition were observed within the major ruminal phyla, namely Bacteroidetes, Firmicutes, and Proteobacteria. Decreases in the abundance of unclassified Bacteroidales, Porphyromonadaceae, and Ruminococcaceae and increases in as-yet uncultured species of the family Succinivibrionaceae, might be related to a potential role of these groups in different pathways of rumen FA metabolism. Diet supplementation with MA, however, had no effect on the relative abundance of Butyrivibrio and Pseudobutyrivibrio genera. In addition, results from both 454 pyrosequencing and T-RFLP indicate that the effect of MA was rather consistent in rumen content or fluid samples, despite inherent differences between these fractions in their bacterial composition.     

Effect of low inulin doses with different polymerisation degree on lipid metabolism,mineral absorption,and intestinal microbiota in rats with fat-supplemented diet

Inulin with different degree of polymerisation from chicory (Orafti Beneo^TMGR, GR) and artichoke (ArtBiochem Artinulin^TM, Art) as well as the mixture of these inulins with chicory oligofructose (Orafti Beneo^TMSynergy1, Syn; and Artinulin^TM plus Orafti Beneo^TMP95 oligofructose, Art+P95) were assayed on developing rats fed with fat-supplemented diet. The effect of these inulins was evaluated on seven haematological and 16 serobiochemical parameters, three minerals, feedstuff intake, growth rate, food utility index and intestinal microbiota upon consumption of a human (60 kg weight) equivalent dose of 0.82 g/day for 75 days. The low inulin dose assayed exerted beneficial effects on rats with fat-supplemented diet upon increase of iron absorption (Art), regulation of ALP (phosphatase alkaline) activity (Art, Syn and Art+P95), increase of growth rate without modification of the final weight (Syn1 and Art+P95), increase of HDL-cholesterol (GR, Syn1 and Art+P95) and reduction of enterobacteria (GR, Syn1 and Art+P95).     

茯苓多糖对营养性肥胖幼鼠的降脂和肠道菌群调节作用

该研究探讨了茯苓多糖对高脂饲料喂养的营养性肥胖幼鼠的降脂及肠道菌群调节作用。将幼鼠（3周龄）随机分成正常对照组、模型组、茯苓多糖低、中、高剂量组（50、100、200 mg/kg）,每组10只。正常对照组给予普通饲料喂养,其他各组均给予高脂饲料喂养构建营养性肥胖大鼠模型,造模成功后给予茯苓多糖灌胃,1次/d,4周后进行检测。结果显示,200 mg/kg茯苓多糖干预可显著减轻营养性肥胖大鼠体质量（16.31%）及肝脏中脂肪堆积（P<0.05）,降低脂肪系数（59.88%）、Lee’s指数（13.10%）及血清中TC（45.17%）、TG（45.76%）、和LDL-C（57.14%）水平,提高HDL-C（56.16%）水平,同时提高普雷沃菌属（Prevotella）（93.33%）、拟杆菌属（Bacteroides）（159.26%）、萨特氏菌属（Sutterella）（88.70%）相对丰度及肠道菌群多样性（P<0.05）,降低厚壁菌门（Firmicutes）/拟杆菌门（Bacteroidetes）比值（67.01%）及摩根氏菌属（Morganella）相对丰度（37.37%）,然而茯苓多糖对营养性肥胖大鼠日均摄食量无显著影响（P>0.05）。因此,茯苓多糖可通过降低血脂、调节肠道菌群结构和增加肠道菌群多样性改善高脂饮食诱导的大鼠营养性肥胖。     

Nicotine, cotinine, withdrawal, and craving patterns during smoking and nicotine nasal spray use: results from a pilot study with African American men.

Nicotine intake via smoking is highly variable. Individualized dosing of nicotine replacement therapy (NRT) may improve product efficacy, but a better understanding of the within-day and within-subject relationships between smoking, NRT use, nicotine and cotinine concentrations in blood, and cravings and withdrawal symptoms is needed to inform dosing algorithms. A pilot study was undertaken to collect data on these relationships and to assess the feasibility of the methods needed for this type of research, including a sophisticated statistical modeling technique (a two-part mixed-effects model with correlated random effects that accounts for clumping at zero). Because nicotine metabolism varies by gender and race, the sample was homogeneous with respect to these characteristics. In a within-subjects study, 27 African American adult male smokers carried a computerized cigarette dispenser for 1 week, capturing the time each cigarette was smoked. Subjects then entered an inpatient setting for 1 day of scheduled smoking (matched to data from the cigarette dispenser to create an ecologically valid schedule) and 4 days of ad libitum nicotine nasal spray use, while tobacco abstinent. Eight times per day, at 2-hour intervals, blood was drawn and ratings of cigarette cravings and withdrawal symptoms were obtained. On average, subjects used less than half of the manufacturer's recommended minimum daily dose of nicotine nasal spray. Large differences in nicotine and cotinine levels were observed between individuals. When predicting nicotine, cotinine, withdrawal, and cravings, we observed significant interactions between route of nicotine intake and a variety of independent variables.     

Characterization of fecal branched-chain fatty acid profiles and their associations with fecal microbiota in diarrheic and healthy dairy calves

Branched-chain fatty acids (BCFA) have recently been reported to play a role in human gut health during early life. However, little information is available on the fecal BCFA profiles in young ruminants and whether they are associated with the development of neonatal calf diarrhea. The objectives of this study were to (1) characterize BCFA profiles in feces collected from young calves, (2) compare the fecal BCFA composition between diarrheic and nondiarrheic dairy calves, and (3) explore the potential relationships between BCFA and microbiota in the feces. A total of 32 male Holstein dairy calves (13 ± 3 d old) with the same diet management were grouped as diarrheic (n = 16) or healthy (n = 16) based on fecal score (determined by liquid fecal consistency with some solid particles); diarrhea cases were defined as fecal score ≥2 for at least 2 d. Fecal samples were collected on the seventh day after calf arrival, and the fecal BCFA and microbial profiles were assessed using gas chromatograph and amplicon sequencing, respectively. In total, 7 BCFA were detected in the feces of all dairy calves; however, the concentrations of fecal BCFA differed between diarrheic and nondiarrheic calves. Specifically, the concentrations of iso-C16:0, iso-C17:0, anteiso-C17:0, and total even-chain BCFA were significantly higher in the feces of diarrheic calves. When the associations between BCFA and bacteria were studied, the relative abundance of Eggerthella was positively correlated with the concentrations of iso-C16:0 (ρ = 0.67), iso-17:0 (ρ = 0.77), anteiso-C17:0 (ρ = 0.73), and iso-C18:0 (ρ = 0.65), whereas the relative abundance of Subdoligranulum was positively correlated with the concentrations of iso-C14:0 (ρ = 0.62), iso-C15:0 (ρ = 0.78), and anteiso-C15:0 (ρ = 0.63). Use of random forest algorithm showed that BCFA such as anteiso-C15:0, iso-C16:0, iso-C17:0, iso-C18:0, and total even-chain BCFA could be used as biomarkers to differentiate diarrheic calves from healthy ones. Our findings generated fundamental knowledge on the potential roles of BCFA in neonatal calf gut health. Follow-up studies with larger animal populations are warranted to validate the feasibility of using BCFA as indicators of health status in neonatal calves.     

Milking system and premilking routines have a strong effect on the microbial community in bulk tank milk

In this study, we investigated the variation in the microbial community present in bulk tank milk samples and the potential effect of different farm management factors. Bulk tank milk samples were collected repeatedly over one year from 42 farms located in northern Sweden. Total and thermoresistant bacteria counts and 16S rRNA gene-based amplicon sequencing were used to characterize microbial community composition. The microbial community was in general heterogeneous both within and between different farms and the community composition in the bulk tank milk was commonly dominated by Pseudomonas, Acinetobacter, Streptococcus, unclassified Peptostreptococcaceae, and Staphylococcus. Principal component analysis including farm factor variables and microbial taxa data revealed that the microbial community in milk was affected by type of milking system. Milk from farms using an automatic (robot) milking system (AMS) and loose housing showed different microbial community composition compared with milk from tiestall farms. A discriminant analysis model revealed that this difference was dependent on several microbial taxa. Among farms using an automatic milking system, there were further differences in the microbial community composition depending on the brand of the milking robot used. On tiestall farms, routines for teat preparation and cleaning of the milking equipment affected the microbial community composition in milk. Total bacteria count (TBC) in milk differed between the farm types, and TBC were higher on AMS than tiestall farms (log 4.05 vs. log 3.79 TBC/mL for AMS and tiestalls, respectively). Among tiestall farms, milk from farms using a chemical agent in connection to teat preparation and a more frequent use of acid to clean the milking equipment had lower TBC in milk, than milk from farms using water for teat preparation and a less frequent use of acid to clean the milking equipment (log 3.68 vs. 4.02 TBC/mL). There were no significant differences in the number of thermoresistant bacteria between farm types. The evaluated factors explained only a small proportion of total variation in the microbiota data, however, despite this, the study highlights the effect of routines associated with teat preparation and cleaning of the milking equipment on raw milk microbiota, irrespective of type of milking system used.     

Is a vegan or a vegetarian diet associated with the microbiota composition in the gut? Results of a new cross-sectional study and systematic review

Abstract

It is assumed that diet influences the composition of gut microbiota, which in turn may affect human health status. This systematic review aimed to summarize associations of a vegan or vegetarian diet with the composition of microbiota. A literature search was conducted in PubMed and Embase for eligible human studies with vegan or vegetarian diets as an exposure and microbiota composition as an outcome in healthy adults. Furthermore, data from our cross-sectional study with vegan participants were included. Out of sixteen included studies, six investigated the association between gut microbiota composition in both vegans and in vegetarians, six in vegans and four studies in vegetarians compared to omnivores, respectively. Among 5 different phyla, 28 families, 96 genera and 177 species, Bacteroides, Bifidobacterium and Prevotella were the most reported genera, followed by the species Prevotella copri, Faecalibacterium prausnitzii and Escherichia coli in all diets. No consistent association between a vegan diet or vegetarian diet and microbiota composition compared to omnivores could be identified. Moreover, some studies revealed contradictory results. This result could be due to high microbial individuality, and/or differences in the applied approaches. Standardized methods with high taxonomical and functional resolutions are needed to clarify this issue.     

Growth performance,blood metabolites,ruminal fermentation,and bacterial community in preweaning dairy calves fed corn silage-included starter and total mixed ration

The objective of this study was to evaluate the effects of the inclusion of whole-plant corn silage (WPCS) in a starter or total mixed ration (TMR) on growth, blood metabolites, ruminal fermentation, and microbial community in preweaning dairy calves. A total of 45 healthy dairy calves were blocked by date of birth and randomly assigned to 1 of 3 treatments: 100% calf starter (CONS), a mix of 85% calf starter and 15% WPCS [dry matter (DM) basis; CSCS], or 100% WPCS-based lactation TMR (CTMR). Pasteurized normal milk was fed to all the animals under the same regimen. The experiment ran from when the calves were 2 d old to weaning at 63 d. Milk and feed intakes were recorded daily. Growth performance data and blood samples were collected on wk 3, 5, 7, and 9 of the experiment. Rumen fluid was sampled at 40 and 60 d. The 3 treatments had different particle size fractions. The CSCS group had greater medium fraction (<19 mm, >8 mm) and particles retained on 8-mm sieves than the other 2 groups, whereas the CTMR group had the greatest long (>19 mm) and fine (<4 mm) fractions and physically effective neutral detergent fiber (NDF) on 8- and 4-mm sieves, but had the smallest short fraction (<8 mm, >4 mm) and particles retained on 4-mm sieves. The 24-h in vitro digestibility of DM, crude protein (CP), NDF, and acid detergent fiber (ADF) were decreased in order by the CONS, CSCS, and CTMR groups. Compared with the CONS group, the digestibility of ether extract (EE) was lower in the CSCS and CTMR groups, whereas the digestibility of starch was similar among treatments. During the experimental period, the DM, CP, and metabolizable energy intakes from milk, solid feed, and total feed were not affected by treatments. The NDF, ADF, and EE intakes and potentially digestible intakes were greater in the CTMR group than in the other 2 groups. With the exception that body barrel was greater for calves fed CSCS, growth parameters and blood metabolites were similar among treatments. Compared with the CSCS group, the CTMR group had greater rumen pH and total volatile fatty acids, propionate, and isovalerate concentrations, but a lower acetate:propionate ratio. The CTMR group had greater relative abundances of some cellulolytic bacteria (Rikenellaceae RC9 gut group, Christensenellaceae R7, Ruminococcaceae NK4A214, Ruminococcaceae UCG, Ruminococcus, and Erysipelotrichaceae UCG) in the rumen, which may be beneficial for the early acquisition of specific adult-associated microorganisms. In summary, a WPCS-based lactation TMR, but not the WPCS-included starter, had the potential to be an alternative starter in preweaning calves without having significant adverse effects. These findings provide theoretical and practical implications for the rational application of TMR in the early life of dairy calves.