Bifidobacterium longum CCFM1077 Attenuates Hyperlipidemia by Modulating the Gut Microbiota Composition and Fecal Metabolites:A Randomized,Double-Blind,Placebo-Controlled Clinical Trial

An increasing number of studies have indicated that gut microbiota and its metabolites are crucial in the development of hyperlipidemia.Bifidobacterium longum(B.longum)CCFM1077 has been shown to have lipid-lowering effects in animals.This study aimed to evaluate the potential of B.longum CCFM1077 in lowering the lipid levels in patients with hyperlipidemia and investigate the effect of this bacterium on serum lipid abnormalities,gut microbiota,and fecal metabolites in these patients.This study was a six-week,randomized,double-blind,and placebo-controlled pilot clinical trial.Subjects with hyperlipidemia(N=62)were randomly assigned to receive placebo(N=31)or B.longum CCFM1077(1×1010colony-forming units(CFUs)per day;N=31).Serum lipid levels including total cholesterol(TC),lowdensity lipoprotein cholesterol(LDL-C),total triglyceride(TG),and high-density lipoprotein cholesterol(HDL-C)were examined at the baseline and interventio nal endpoints.Changes in the gut microbiota composition and diversity were measured based on 16S ribosomal RNA(rRNA)sequencing of the V3-V4region at the end of the intervention period.Non-targeted metabolomics of the feces was performed using ultra-performance liquid chromatography(UPLC)-Q-Exactive Orbitrap/mass spectrometer.Oral administration of B.longum CCFM1077 for six weeks significantly decreased the serum levels of TC(p<0.01)and LDL-C(p<0.01)in patients with hyperlipidemia.B.longum CCFM1077 treatment markedly increased gut microbiota diversity and the relative abundance of anti-obesity-related genera,including Lactobacillus,Butyricicoccus,Bifidobacterium,and Blautia,whereas it decreased the relative abundance of obesity-related genera,including Alistipes,Megamonas,and Catenibacterium.Additionally,some key metabolites(bile acids(BAs),biotin,and caffeine)and their corresponding metabolic pathways(primary BA biosynthesis,and taurine and hypotaurine,biotin,purine,and caffeine metabolisms)were enriched by B.longum CCFM1077,and thus it may lower lipid levels.B.longum CCFM1077 is a probiotic strain with the potential to lower serum TC and LDL-C levels patients with hyperlipidemia.The underlying mechanism may be related to the increased abundance of anti-obesity-related genera and fecal metabolites.These findings provide a foundation for future clinical applications of lipid-lowering probiotics in managing individuals with hyperlipidemia.

Fecal metabolomics reveals the positive effect of ethanol extract of propolis on T2DM mice

A large number of studies have shown that propolis has positive effects in the treatment of type 2 diabetes mellitus(T2DM).However,there are have only been a few reports that are based on an ultra-high performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS)analysis of the fecal metabolomics of ethanol extract of propolis(EEP)in the treatment of T2DM.The present investigation was designed to screen potential biomarkers of T2DM by the metabonomic method and to explain the possible anti-diabetes mechanism of EEP according to the changes in the biomarkers.The results showed that EEP improved the body weight(BW)of T2DM mice,lowered blood sugar levels,and significantly restored blood biochemical indicators related to T2DM,such as fasting insulin(FINS),homeostasis model assessment of insulin resistance(HOMA-IR),aspartate transaminase(AST),and alanine aminotransferase(ALT).Liver pathology showed that EEP reversed liver damage caused by T2DM.Metabolomics data identified 27 potential biomarkers in fecal samples.EEP effectively regulated the dysfunction in the metabolic pathways of glycerophospholipids,sphingolipids,riboflavins,and sterol lipids caused by T2DM.In summary,our research results revealed positive effects of EEP in the treatment of T2DM and provided potential candidate markers for further research and in the clinical treatment of T2DM.

Effects of dietary forage to concentrate ratio and wildrye length on nutrient intake, digestibility, plasma metabolites, ruminal fermentation and fecal microflora of male Chinese Holstein calves

Twenty-eight male, weaned Chinese Holstein calves（（156.8±33.4） kg） were used to investigate the effects of dietary forage to concentrate ratio（F：C） and forage length on nutrient digestibility, plasma metabolites, ruminal fermentation, and fecal microflora. Animals were randomly allocated to four treatments in a 2×2 factorial arrangement： whole-length forage（WL） with low F：C（50：50）; WL with high F：C（65：35）; short-length forage（SL） with high F：C（65：35）; and SL with low F：C（50：50）. Chinese wildrye was used as the only forage source in this trial. The grass in the SL treatments was chopped using a chaff cutter to achieve small particle size（-50% particles 〉19 mm）. Dry matter intake（DMI） and organic matter（OM） intake was increased by increasing both F：C（P〈0.01） and forage length（FL）（P〈0.05）, while acid detergent fiber（ADF） and neutral detergent fiber（NDF） intakes were only increased by increasing the F：C（P〈0.01）. The digestibility of NDF was increased as the FL increased（P〈0.01）, and it was also affected by interaction between F：C and FL（P〈0.05）. Cholesterol（CHO）（P〈0.01）, leptin（LP）（P〈0.05）, and growth hormone（GH）（P〈0.01） concentrations in plasma were increased as dietary F：C increased. A significant increase in plasma triglyceride（TG）（P〈0.01）, insulin（INS）（P〈0.05）, and GH（P〈0.01） levels was observed with decreasing dietary FL. Ruminal p H values of calves fed with low F：C diets were significantly lower than those in high F：C treatment（P〈0.05）. Increasing the F：C enhanced ruminal acetic acid（P〈0.05） and acetic acid/propionic acid（P〈0.01）. Fecal Lactobacillus content was significantly higher, while Escherichia coli and Salmonella contents were significantly lower in WL and high F：C groups（P〈0.05）. Lower fecal scores（higher diarrhea rate） were observed in calves fed with SL hay compared to WL hay（P〈0.05）. Denatured gradient gel electrophoresis（DGGE） bands and richness index（S） were significantly affected by the interaction between F：C and FL（P〈0.05）, under high F：C, band numbers and richness index from WL group were higher than that from SL group（P〈0.05）, whereas there were no differences between WL andSL groups under low F：C（P〉0.05）. Microflora similarity was 50–73% among the different treatments. It is concluded that the WL with high F：C（65：35） diet is suitable for weaned calves.

Gut microbiota links with cognitive impairment in amyotrophic lateral sclerosis: A multi-omics study

Recently, cognitive impairments(CI) and behavioral abnormalities in patients with amyotrophic lateral sclerosis(ALS) have been reported. However, the underlying mechanisms have been poorly understood. In the current study, we explored the role of gut microbiota in CI of ALS patients. We collected fecal samples from 35ALS patients and 35 healthy controls. The cognitive function of the ALS patients was evaluated using the Edinburgh Cognitive and Behavioral ALS Screen. We analyzed these samples by using 16S rRNA gene sequencing as well as both untargeted and targeted(bile acids) metabolite mapping between patients with CI and patients with normal cognition(CN). We found altered gut microbial communities and a lower ratio of Firmicutes/Bacteroidetes in the CI group, compared with the CN group. In addition, the untargeted metabolite mapping revealed that 26 and 17 metabolites significantly increased and decreased, respectively, in the CI group, compared with the CN group. These metabolites were mapped to the metabolic pathways associated with bile acids. We further found that cholic acid and chenodeoxycholic acid were significantly lower in the CI group than in the CN group. In conclusion, we found that the gut microbiota and its metabolome profile differed between ALS patients with and without CI and that the altered bile acid profile in fecal samples was significantly associated with CI in ALS patients. These results need to be replicated in larger studies in the future.

Monitoring the effects of land transformation on African clawless otters(Aonyx capensis)using fecal glucocorticoid metabolite concentrations as a measure of stress

In a time of increasing environmental change caused by anthropogenic disturbance,there is a greater need to understand animal adaptations to manmade environments.In this regard,the measurement of stress-related endocrine markers provides a useful tool to examine the impact of environmental challenges and the physiological consequences for wildlife occupying such space.The aims of the present study were to validate fecal glucocorticoid metabolite(fGCM)concentrations as a measure of stress using samples from a male African clawless otter(Aonyx capensis;n=1)and to compare fGCM concentrations of otters occurring in a transformed and in 2 natural areas in South Africa.From the 5 different enzyme-immunoassays(EIA)tested,a cortisol and oxoetiocholanolone(measuring 11,17 dioxoandrostanes)EIA revealed the highest response(74%and 48%increase,respectively)30 and 24 hours after a stress event(translocation of a captive individual as part of its rehabilitation prior to release),respectively.For both EIAs,fGCM concentrations were comparable for samples collected up to 3 h post-defecation.Using the cortisol EIA for subsequent analyses,fGCM concentrations of animals from the transformed area(n=20;mean[±SD]:0.468[±0.539]µg/g dry weight[DW])were significantly higher(P=0.013)than those from otters in the natural areas(n=17;0.242[±0.226]µg/g DW).These preliminary results suggest that African clawless otters may have increased adrenocortical activity that could be due to conditions linked to living in a transformed environment.

Effect of intensity and duration of anthropic noises on European mink locomotor activity and fecal cortisol metabolite levels

Human activities involving noise emission can affect wild animals.European mink was exposed to road noise and human voice playbacks to analyze how sound intensity level and duration of both noises altered the time that individuals were active and if their fecal cortisol metabolite(FCM)levels varied.A Hierarchical Analysis Cluster was performed to establish 2 mink groups with respect to both noise source type:short duration/low intensity(SL)and long duration/high intensity(LH).We performed general linear mixed models to evaluate the variation in locomotor activity duration(s)and FCM(nanogram per gram)levels,respectively.The results showed both road noise and human voices decreased locomotor activity duration in SL more sharply compared with LH,and human voices were the triggers that induced the most pronounced response to both exposure conditions.FCM(ng/g)levels increased in SL compared with LH during road noise while the opposite happened during human voices.Differences based on sex and age of individuals were observed.In conclusion,noise characteristics given by the sound type determined the variations in locomotor activity duration while noise exposure level determined the variations in FCM(ng/g)levels.Attention should be paid to noisy activities(e.g.,recreational activities for visitors in protected natural areas)and loud groups of people to conserve wildlife,especially noise sensitive species.

Validation of a radioimmunoassay-based fecal corticosteroid assay for Richardson＇s ground squirrels Urocitellus richardsonii and behavioural correlates of stress

We validated a radioimmunoassay-based method quantifying fecal glucocorticoid metabolites （FGMs） from captive male and female Richardson＇s ground squirrels Urocitellus richardsonii. Blood samples were drawn to explore the correlation between plasma cortisol and FGM concentrations. We also injected groups of squirrels with normal saline （CTL; control）, adre- nocorticotropic hormone （ACTH; stimulating adrenal activity）, or dexamethasone （DEX; suppressing adrenal activity）. Potential correlations between stress and behaviour were explored through quantification of fecal pellet production and the intervention necessary to elicit defecation, as well as the behaviour of subjects in the context of handling. Changes in plasma cortisol concen- tration between capture （baseline）, and following handling （stress-induced） were also quantified for free-living squirrels. While glucocorticoid concentrations recovered from feces during our captive-animal study were not well correlated with plasma eortisol concentrations, and uncorrelated with defecation or behaviour, FGM concentrations did reflect the activation of the hypothalamic- pituitary-adrenal （HPA） axis. FGM concentrations increased significantly during initial captivity, but declined to baseline level as individuals acclimated to the novel environment. Injection of subjects with ACTH increased FGMs above baseline, confirming activation of the HPA axis. Plasma cortisol concentrations increased significantly with induced stress, indicating that capture and handling activated the glucocorticoid stress response even among previously handled, free-living subjects. Our findings validate a non-invasive tool that will afford new insight into the physiological processes underlying social, reproductive and antipredator behaviour of Richardson＇s ground squirrels [Current Zoology 60 （5）： 591-601, 2014 ].

Endocrine correlates of gender and throat coloration in the southern ground-hornbill(Bucorvus leadbeateri)

The southern ground-hornbill(SGH)is a cooperatively breeding bird endemic to eastern and southern Africa,but is endangered in its southern distributional range.The national conservation restoration program harvests redundant chicks for captive breeding and reintroduction;with sexing and social grouping of the species evaluated by throat-skin coloration,with adult males displaying a completely red color compared to dark blue within the red observed in adult females.However,recent findings indicate that dominant and subordinate adult males exhibit patches of blue throat-skin.To optimize SGH management practices,it is vital to determine the role of red and blue coloration,as well as the possible drivers thereof.As a prerequisite,an enzyme immunoassay for monitoring fecal androgen metabolite(fAM)concentrations in SGH was established.Following this,fresh fecal samples were collected from 78 SGH,of various demographics and origin,across 12 captive institutions,to determine whether fAM concentrations differ between blue(B),partially blue(sB),and fully red(R)throat-skin colored SGH.Furthermore,fAM concentrations were compared between males housed in different social groups of different age and sex classes.Individual median fAM concentrations of B,sB,and R adult males did not differ significantly but were considerably higher in B and sB males compared to R males.Social dynamics within captivity,for example,dominance,played no role as a driver of male gonadal activity or throat skin coloration.The results of the study indicate that androgens and apparent social dynamics are not primary determinants of throat coloration in male SGH.

Stressful living in lower-quality habitats?Body mass,feeding behavior and physiological stress levels in wild wood mouse populations

Wild populations are continuously subjected to changes in environmental factors that pose different challenges.Body condition and hormones have been commonly used as health indicators due to their potential correlation with fitness.In the present study,we analyzed whether habitats of different quality influenced body mass,food intake and physiological stress levels in wild wood mice(Apodemus sylvaticus).Field work was seasonally carried out in Holm oak woods and pine forests in central Spain.A total of 93 wood mice from 4 different populations(2 per habitat type)were live-trapped.From each captured individual we noted body mass and food intake,measured as the amount of bait remaining in each trap.The physiological stress levels were measured non-inva-sively in collected fresh feces by quantifying fecal corticosterone metabolites(FCM)with a 5a-pregnane-3ß,11ß,21-triol-20-one enzyme immunoassay.Wood mice abundances decreased from spring to summer,were higher in Holm oak woods than in pine forests and also resulted in different age-class distribution between both habitats.Individuals inhabiting pine forests showed a lower body mass and increased food intake,probably because of the comparatively lower food quality and availability in this habitat.Furthermore,these individuals showed increased physiological stress levels,likely due to the lower quality habitat in relation to both food and vegetation cover availability.Overall,besides affecting local wood mouse abundance,our study underscores the effect of habitat quality on body mass,food intake and the endocrine stress response.Considering the wood mouse’s piv-otal position in ecosystems,these results could help in the understanding of environmental traits hampering the viability of wild populations.

Immune challenge of mating effort:steroid hormone profile,dark ventral patch and parasite burden in relation to intrasexual competition in male Iberian red deer

Testosterone secretion may regulate the reproductive effort and the development of sexual traits,but it may also involve costs at the immunological and metabolic levels.However,the evidence for this trade-off in wild populations is scarce.Cortisol also plays an important role in mediating the reproductive and immune functions.In this study,we analyzed whether the endoparasite burden relates to hormonal levels(fecal testosterone and cortisol metabolites)and/or morphological sexual traits(size of the dark ventral patch,a trait that indicates reproductive effort in males)in male Iberian red deer.For this purpose,we sampled male red deer harvested during hunting actions in 2 types of populations in south western Spain that differed in structure,affecting the level of male–male competition for mates.We used coprological analyses to estimate the parasite burden mainly of gastrointestinal and bronchopulmonary nematodes and of protozoa,and assessed testosterone and cortisol metabolite levels from fecal pellets.We found a positive relationship of host parasitation with both testosterone levels and the size of the dark ventral patch,but these relationships depended on the intensity of male–male competition in the population,being only found under the high-competition scenario.These results are discussed under the hypothesis of the testosterone immunocompetence handicap,suggesting a cost at the immunological level,and,therefore,higher susceptibility to parasite infection in males that make a greater reproductive effort.However,this effect seems to be modulated by the social environment(male–male competition)that might lead to different optima in testosterone production and sexual trait development.

Complex relationships between physiological stress and endoparasite infections in natural populations

Short-term elevation of glucocorticoids(GCs)is one of the major physiological mechanisms by which vertebrates cope with challenging environmental or social factors(stressors).However,when exposure to stressors occurs repeatedly or over a prolonged period of time,animals may experience chronic elevation of GCs,which reduces the immune response efficiency and can lead to higher intensity of parasitic infection.Here,we used invasive gray squirrels Sciurus carolinensis in troduced in Norther n Italy and their 2 most prevale nt gastrointesti nal parasites,the n ematode Strongyloides robustus and coccidia of the genus Eimeria,as a model to investigate relati on ships among macroparasite infection and concentrations of fecal glucocorticoid metabolites(FGMs),an integrated measure of circulating GCs.Our results revealed an association of FGMs with infection by St.robustus,but not with coccidia.Individuals with higher FGMs appear to be responsible for the greatest St.robustus egg shedding within gray squirrel populations,thus possibly acting as superspreaders.However,FGMs were negatively associated with adult St.robustus,suggesting that the abundance of adults of this nematode species does not induce elevation in FGMs,but is only affected by it through immun e-mediated effects on its fecun dity.Fin ally,the relati on ship betwee n St.robustus(both eggs and adult parasites)and FGMs was not linear,suggesting that only high levels of physiological stress in fluence parasite inf ection.Our fin dings highlight that the direction and magnitude of the stress-infection relationship may depend not only on the specific hostparasite system,but also on the different life stages of the same parasite.