Relevance of gut microbiome research in food safety assessment.

The gut microbiome is indispensable for the host physiological functioning. Yet, the impact of non-nutritious dietary compounds on the human gut microbiota and the role of the gut microbes in their metabolism and potential adverse biological effects have been overlooked. Identifying potential hazards and benefits would contribute to protecting and harnessing the gut microbiome's role in supporting human health. We discuss the evidence on the potential detrimental impact of certain food additives and microplastics on the gut microbiome and human health, with a focus on underlying mechanisms and causality. We provide recommendations for the incorporation of gut microbiome science in food risk assessment and identify the knowledge and tools needed to fill these gaps. The incorporation of gut microbiome endpoints to safety assessments, together with well-established toxicity and mutagenicity studies, might better inform the risk assessment of certain contaminants in food, and/or food additives.

Ensiling sorghum with unsalable pumpkin improves feed digestibility with minimal influence on the rumen microbial population using the rumen simulation technique.

This study aimed to determine the influence of sorghum ensiled with unsalable pumpkin at 20 or 40% dry matter (DM) basis on rumen fermentation characteristics and rumen microbial communities using the rumen simulation technique (RUSITEC). The experiment used a completely randomised design including silages comprising (1) 100% sorghum; (2) 80% sorghum + 20% DM pumpkin; or (3) 60% sorghum + 40% DM pumpkin. Each RUSITEC run (n = 2) was 15 d long, including 6 d of adaptation and 9 d of sampling. Dry matter digestibility (DMD) was measured on d 8 and 10-13. Gas production was measured daily, whereas methane and volatile fatty acids (VFA) production were measured from d 7-15. Solid-associated microbes (SAM) were collected on d 5, 10 and 15, whereas liquid-associated microbes (LAM) were collected after 15-d incubation. The V4 region of the 16S rRNA gene and the ITS1 region were sequenced to identify archaeal, bacterial and fungal communities. Ensiling 40% DM pumpkin with sorghum increased DMD and decreased the ratio of acetate to propionate (P ≤ 0.01). Both bacterial SAM and LAM communities were dominated by Megasphaera, and had the highest relative abundance (P = 0.03) with 40% DM pumpkin after 5 d incubation in the SAM community, while species of the Aspergillus genus dominated fungal SAM and LAM communities with 20 or 40% DM unsalable pumpkin. Therefore, ensiling up to 40% DM unsalable pumpkin with sorghum produces a high-quality ruminant feed with minimal influence on the rumen microbial population. KEY POINTS: • Including 40% DM unsalable pumpkin decreased acetate:propionate • Ensiling unsalable pumpkin with sorghum increases digestibility in a RUSITEC • Rumen microbial communities were slightly influenced by unsalable pumpkin inclusion.

Ozone Decreased Enteric Methane Production by 20% in an in vitro Rumen Fermentation System.

Ozone (O3) is volatile, highly oxidative, and has theoretical potential to reduce ruminant enteric methanogenesis by interactions between archaea and bacteria, and substrate and oxygen. The effects of O3 on the rumen microbiota, fermentation parameters, and CH4 emissions were studied through in vitro fermentation using a RUSITEC apparatus with O3 dissolved in the salivary buffer. The substrate consisted of maize silage or grain concentrates, and the treatments were (1) control (no O3) and (2) O3 at 0.07 ± 0.022 mg/L in the buffer. A 4-day adaptation period followed by a 6-day experimental period was used for measuring gas production and composition, as well as fermentation characteristics, which included ruminal volatile fatty acids (VFA) and liquid- and solid-associated microbial communities. Ozone treatment decreased total gas production by 15.4%, most notably CH4 production by 20.4%, and CH4 gas concentration by 5.8%, without compromising dry matter digestibility (DMD) of either maize silage or grain concentrates. There were no significant effects of O3 treatment on VFA production or pH. Ozone treatment reduced the relative abundance of methanogens, particularly Methanomicrobium. This study demonstrates the potential use of O3 as a method to reduce ruminant enteric methanogenesis.

Influence of probiotics on biofilm formation and diversity of bacteria colonising crop sorghum ensiled with unsalable vegetables.

The objective of this study was to characterise in situ digestion kinetics and bacterial colonisation of crop sorghum ensiled with unsalable carrot or pumpkin at 0, 20 or 40% dry matter (DM). Silages with or without the application of a commercial probiotic were incubated in situ for 0, 3, 6, 9, 24 and 48 h. Calculation of in situ digestion kinetics was conducted for DM, organic matter and neutral detergent fibre (aNDF). The V4 region of the 16S rRNA gene was sequenced to determine the composition and diversity of bacteria colonising the silage. Organic matter and DM digestion kinetics indicated that greater vegetable inclusion increased (P < 0.05) the soluble fraction and effective degradability. Bacterial richness at 48 h incubation was greater (P = 0.02) in 20% carrot and 40% pumpkin treatments, compared with the control. An effect of level × probiotic was observed with increased Shannon diversity (P = 0.01) for 40% carrot and 20% pumpkin probiotic treatments, respectively. Primary colonising bacteria were members of the Prevotella genus, dominating after 3 and 6 h of incubation. The abundance of Prevotella increased by 4.1% at 3 h (P < 0.01) and by 4.7% at 9 h incubation with probiotics, compared with the control. Secondary biofilm colonisers included members of Treponema, Saccharofermentans, Fibrobacter, Ruminobacter and Anaerosporobacter genera, dominant from 9 h incubation onward. This study demonstrated that including unsalable vegetables at 20 or 40% DM increases the soluble fraction and effective degradability of sorghum silage during in situ digestion and increases diversity of bacteria colonising ensiled vegetables within the rumen. KEY POINTS: • Ensiling unsalable vegetables is a viable strategy to reduce food waste. • Ensiled vegetables increased in situ soluble fraction and effective degradability. • Bacterial richness at 48 h incubation improved with 20% carrot or 40% pumpkin. • Diversity of colonising rumen bacteria increased with carrot or pumpkin inclusion.

Crop Sorghum Ensiled With Unsalable Vegetables Increases Silage Microbial Diversity.

Ensiling vegetables with forage crops is a suggested method of waste diversion and can be directly utilized as a livestock feed. Carrot or pumpkin, ensiled at 0, 20, or 40% dry matter (DM) with crop sorghum, and with or without a second-generation silage inoculant were assessed for nutritive composition, organic acid profiles, aerobic stability and in vitro rumen fermentation characteristics. The study was a completely randomized design, with the fixed effects consisting of vegetable type (carrot vs. pumpkin), level (i.e., the level of vegetables), inoculant (inoculant or non-inoculant) and the interactions, and mini-silos within treatment as the random effect. The experimental unit for sorghum treatments represented by each mini-silo (5 kg capacity). Silage was sampled after 70-days ensiling for nutrient composition, 14-day aerobic stability, organic acid profiles and microbial diversity. After 24 h in vitro incubation, rumen fermentation parameters were assessed, measuring gas and methane (CH4) production, in vitro digestibility and volatile fatty acid concentrations. Sorghum ensiled with carrot or pumpkin at 20% or 40% DM increased crude fat (P ≤ 0.01) and decreased (P ≤ 0.01) silage surface temperature upon aerobic exposure compared to the control. Bacterial communities analyzed through 16S rRNA gene sequencing linearly increased (P ≤ 0.01) in diversity across both vegetables when the vegetable proportion was increased in the silage; dominated by Lactobacillus species. ITS analysis of the fungal microbiota upon silage opening and after 14 days (aerobic stability) identified increased (P ≤ 0.03) fungal diversity with increasing vegetable proportions, predominantly populated by Fusarium denticulatum, Issatchenkia orientalis, Kazachstania humilis, and Monascus purpureus. Upon assessment in vitro, there was an increase (P ≤ 0.04) in in vitro digestibility and some CH4 parameters (% CH4, and mg CH4/g DM), with no effect (P ≥ 0.17) on remaining CH4 parameters (mL CH4/g DM, mg CH4/g digested DM), gas production or pH. However, increasing vegetable amount decreased percentage of acetic acid and increased percentage of propionic acid of the total VFA, decreasing A:P ratio and total VFA concentration as a result (P ≤ 0.01). The results from this study indicate including carrot or pumpkin at 20 or 40% DM in a sorghum silage can produce a highly digestible, microbially diverse and energy-rich livestock feed.

Microbial characterization and fermentative characteristics of crop maize ensiled with unsalable vegetables.

Incorporation of carrot or pumpkin at 0, 20 or 40% dry matter (DM-basis) with crop maize, with or without a silage inoculant was evaluated after 70 days ensiling for microbial community diversity, nutrient composition, and aerobic stability. Inclusion of carrots or pumpkin had a strong effect on the silage bacterial community structure but not the fungal community. Bacterial microbial richness was also reduced (P = 0.01) by increasing vegetable proportion. Inverse Simpson's diversity increased (P = 0.04) by 18.3% with carrot maize silage as opposed to pumpkin maize silage at 20 or 40% DM. After 70 d ensiling, silage bacterial microbiota was dominated by Lactobacillus spp. and the fungal microbiota by Candida tropicalis, Kazachstania humilis and Fusarium denticulatum. After 14 d aerobic exposure, fungal diversity was not influenced (P ≥ 0.13) by vegetable type or proportion of inclusion in the silage. Inoculation of vegetable silage lowered silage surface temperatures on day-7 (P = 0.03) and day-14 (P ≤ 0.01) of aerobic stability analysis. Our findings suggest that ensiling unsalable vegetables with crop maize can successfully replace forage at 20 or 40% DM to produce a high-quality livestock feed.

Effects of Hardwood Biochar on Methane Production, Fermentation Characteristics, and the Rumen Microbiota Using Rumen Simulation.

Biochar is a novel carbonized feed additive sourced from pyrolyzed biomass. This compound is known to adsorb gasses and carbon, participate in biological redox reactions and provide habitat biofilms for desirable microbiota proliferation. Therefore, biochar holds potential to modify rumen fermentation characteristics and reduce enteric CH4 emissions. The objective of this study was to investigate the effect of hardwood biochar supplementation on fermentation parameters, methane (CH4) production and the ruminal archaeal, bacterial, and fungal microbiota using the in vitro RUSITEC (rumen simulation technique) system. Treatments consisted of a control diet (oaten pasture: maize silage: concentrate, 35:35:30 w/w) and hardwood biochar included at 400 or 800 mg per day (3.6 and 7.2% of substrate DM, respectively), over a 15-day period. Biochar supplementation had no effect (P ≥ 0.37) on pH, effluent (mL/d), total gas (mL/d), dry matter (DM) digestibility or CH4 production (mg/d). The addition of 800 mg biochar per day had the tendency (P = 0.10) to lower the % of CH4 released in fermentation compared to 400 mg/d biochar treatment. However, no effect (P ≥ 0.44) was seen on total VFA, acetate, propionate, butyric, branched-chain VFA, valerate and caproate production and the ratio of acetate to propionate. No effect (P > 0.05) was observed on bacterial, archaeal or fungal community structure. However, biochar supplementation at 800 mg/d decreased the abundance of one Methanomethylophilaceae OTU (19.8-fold, P = 0.046) and one Lactobacillus spp. OTU (31.7-fold, P < 0.01), in comparison to control treatments. Two fungal OTUs classified as Vishniacozyma victoriae (5.4 × 107 increase) and Sporobolomyces ruberrimus (5.4 × 107-fold increase) were more abundant in the 800 mg/d biochar samples. In conclusion, hardwood biochar had no effects on ruminal fermentation characteristics and may potentially lower the concentration of enteric CH4 when included at higher dosages by manipulating ruminal microbiota abundances.