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The objective of this study was to investigate the effect and composition of a standardized natural citrus extract (SNCE) on both broiler chickens’ growth performances and intestinal microbiota. A total of 930 one-day-old males were randomly assigned to three dietary treatments: a control treatment (CTL) in which broiler chickens were fed with a standard diet and two citrus treatments in which broiler chickens were fed with the same standard diet supplemented with 250 ppm and 2,500 ppm of SNCE, respectively. Each dietary treatment was composed of 10 experimental units (pen) of 31 broiler chickens each. Growth performances such as feed consumption, body weight, and feed conversion ratio (FCR) were recorded weekly until day 42. Litter quality was also weekly recorded while mortality was daily recorded. One broiler chicken was randomly selected from each pen (10 chickens/group) and ceca samples were collected for microbiota analysis at day 7 and 42. Chromatographic methods were used to determine molecules that enter into the composition of the SNCE. Results from the characterization of SNCE allowed to identify pectic oligosaccharides (POS) as a major component of the SNCE. In addition, 35 secondary metabolites, including eriocitrin, hesperidin, and naringin, were identified. The experiment performed on broiler chickens showed that the final body weight of broiler chickens fed diets supplemented with SNCE was higher than those fed the CTL diets (P < 0.01). Broiler cecal microbiota was impacted by age (P < 0.01) but not by the dietary supplementation of SNCE. Results indicate that SNCE allowed enhancing chickens’ performances without any modulation of the cecal microbiota of broiler chickens. The characterization of SNCE allowed to identify compounds such as eriocitrin, naringin, hesperidin, and POS. Thus, opening new horizons for a better understanding of the observed effect on broiler chickens’ growth performances.

We investigated the effect of the Standardized Natural Citrus Extract (SNCE; Nor-Spice AB, Nor-Feed SAS, France) on the microbiota of the sows and on the weight gain of their piglets. Fifty sows were randomly divided into two groups: a control group (23 sows) with a standard diet and a SNCE group (27 sows) with a standard diet supplemented with 2,500 ppm of SNCE. Supplementation occurred 10 d before and 5 d after farrowing. Fecal samples from 16 sows (8 randomly selected sows of each dietary treatment) were collected for the fecal microbiota analysis 5 d after farrowing. The supplementation of SNCE increases the amount of cultivable Lactobacillus threefold in vitro. Microbial DNA was extracted from the fecal samples for sequencing of the 16S rRNA gene. The SNCE, which affected the microbiota as a discriminant analysis, was able to separate the microbial communities of the eight sows that received SNCE from the three control sows with 21 Operational Taxonomic Units (area under the ROC curve = 96%). SNCE also reduced the interval between farrowing and the first dejection of the sow and increased their feed intake (P-value < 0.05). Furthermore, feeding the sows with SNCE improved the weight gain of the piglets in the first week of life. These results show that SNCE supplementation allows to enhance zootechnical performances of peripartum’ sows, possibly due to the modulation of the microbiota transmitted to the piglets.

International audience; In suckling mammals, the onset of solid food ingestion is coincident with the maturation of the gut barrier. This ontogenic process is driven by the colonization of the intestine by the microbiota. However, the mechanisms underlying the microbial regulation of the intestinal development in early life are not fully understood. Here, we studied the co-maturation of the microbiota (composition and metabolic activity) and of the gut barrier at the suckling-to-weaning transition by using a combination of experiments in vivo (suckling rabbit model), ex vivo (Ussing chambers) and in vitro (epithelial cell lines and organoids). The microbiota composition, its metabolic activity, para-cellular epithelial permeability and the gene expression of key components of the gut barrier shifted sharply at the onset of solid food ingestion in vivo, despite milk was still predominant in the diet at that time. We found that cecal content sterile supernatant (i.e. containing a mixture of metabolites) obtained after the onset of solid food ingestion accelerated the formation of the epithelial barrier in Caco-2 cells in vitro and our results suggested that these effects were driven by the bacterial metabolite butyrate. Moreover, the treatment of organoids with cecal content sterile supernatant partially replicated in vitro the effects of solid food ingestion on the epithelial barrier in vivo. Altogether, our results show that the metabolites produced by the microbiota at the onset of solid food ingestion contribute to the maturation of the gut barrier at the suckling-to-weaning transition. Targeting the gut microbiota metabolic activity during this key developmental window might therefore be a promising strategy to promote intestinal homeostasis.

In pigs, the gut microbiota composition plays a major role in the process of digestion, but is influenced by many external factors, especially diet. To be used in breeding applications, genotype by diet interactions on microbiota composition have to be quantified, as well as their impact on genetic covariances with feed efficiency (FE) and digestive efficiency (DE) traits. This study aimed at determining the impact of an alternative diet on variance components of microbiota traits (genera and alpha diversity indices) and estimating genetic correlations between microbiota and efficiency traits for pigs fed a conventional (CO) or a high-fiber (HF) diet. Fecal microbes of 812 full-siblings fed a CO diet and 752 pigs fed the HF diet were characterized at 16 weeks of age by sequencing the V3-V4 region of the 16S rRNA gene. A total of 231 genera were identified. Digestibility coefficients of nitrogen, organic matter, and energy were predicted analyzing the same fecal samples with near infrared spectrometry. Daily feed intake, feed conversion ratio, residual feed intake and average daily gain (ADG) were also recorded. The 71 genera present in more than 20% of individuals were retained for genetic analyses. Heritability (h²) of microbiota traits were similar between diets (from null to 0.38 ± 0.12 in the CO diet and to 0.39 ± 0.12 in the HF diet). Only three out of the 24 genera and two alpha diversity indices with significant h² in both diets had genetic correlations across diets significantly different from 0.99 (P 0.05), indicating limited genetic by diet interactions for these traits. When both diets were analyzed jointly, 59 genera had h² significantly different from zero. Based on the genetic correlations between these genera and ADG, FE, and DE traits, three groups of genera could be identified. A group of 29 genera had abundances favorably correlated with DE and FE traits, 14 genera were unfavorably correlated with DE traits, and the last group of 16 genera had abundances with correlations close to zero with production traits. However, genera abundances favorably correlated with DE and FE traits were unfavorably correlated with ADG, and vice versa. Alpha diversity indices had correlation patterns similar to the first group. In the end, genetic by diet interactions on gut microbiota composition of growing pigs were limited in this study. Based on this study, microbiota-based traits could be used as proxies to improve FE and DE in growing pigs.The link between the composition of the gut microbiota, i.e the composition of microorganisms in the gut, in pigs and their feed efficiency, i.e. their ability to utilize nutrients, as well as their ability to digest were studied from a genetic point of view. A family structure of 1,564 pigs were studied and fed with two different diets. One of the full-sib was fed a conventional diet used in breeding farms and the other one an alternative diet containing raw materials, less expensive but with a higher content of dietary fibers more difficult to digest. This study has shown that some microbiota microorganisms were genetically correlated with feed and digestive efficiency performances, positively or negatively, depending on the microorganisms. In addition, the diversity of microorganisms in the animal’s gut was favorably correlated with the feed and digestive performances studied. Therefore, there is a genetic link between these performances and the composition of the animal’s gut microbiota. Thus, a potential genetic selection on some intestinal microorganisms or diversity of microorganisms would allow to improve these performances, and in particular when pigs are fed with diet more difficult to digest.

Background The objective of the present study was to investigate how variation in the faecal microbial composition is associated with variation in average daily gain (ADG), backfat thickness (BFT), daily feed intake (DFI), feed conversion ratio (FCR), and residual feed intake (RFI), using data from two experimental pig lines that were divergent for feed efficiency. Estimates of microbiability were obtained by a Bayesian approach using animal mixed models. Microbiome-wide association analyses (MWAS) were conducted by single-operational taxonomic units (OTU) regression and by back-solving solutions of best linear unbiased prediction using a microbiome covariance matrix. In addition, accuracy of microbiome predictions of phenotypes using the microbiome covariance matrix was evaluated. Results Estimates of heritability ranged from 0.31 ± 0.13 for FCR to 0.51 ± 0.10 for BFT. Estimates of microbiability were lower than those of heritability for all traits and were 0.11 ± 0.09 for RFI, 0.20 ± 0.11 for FCR, 0.04 ± 0.03 for DFI, 0.03 ± 0.03 for ADG, and 0.02 ± 0.03 for BFT. Bivariate analyses showed a high microbial correlation of 0.70 ± 0.34 between RFI and FCR. The two approaches used for MWAS showed similar results. Overall, eight OTU with significant or suggestive effects on the five traits were identified. They belonged to the genera and families that are mainly involved in producing short-chain fatty acids and digestive enzymes. Prediction accuracy of phenotypes using a full model including the genetic and microbiota components ranged from 0.60 ± 0.19 to 0.78 ± 0.05. Similar accuracies of predictions of the microbial component were observed using models that did or did not include an additive animal effect, suggesting no interaction with the genetic effect. Conclusions Our results showed substantial associations of the faecal microbiome with feed efficiency related traits but negligible effects with growth traits. Microbiome data incorporated as a covariance matrix can be used to predict phenotypes of animals that do not (yet) have phenotypic information. Connecting breeding environment between training sets and predicted populations could be necessary to obtain reliable microbiome predictions.

Chronic stress and the gut microbiota appear to comprise a feed-forward loop, which contributes to the development of depressive disorders. Evidence suggests that memory can also be impaired by either chronic stress or microbiota imbalance. However, it remains to be established whether these could be a part of an integrated loop model and be responsible for memory impairments. To shed light on this, we used a two-pronged approach in Japanese quail: first stress-induced alterations in gut microbiota were characterized, then we tested whether this altered microbiota could affect brain and memory function when transferred to a germ-free host. The cecal microbiota of chronically stressed quails was found to be significantly different from that of unstressed individuals with lower α and β diversities and increased Bacteroidetes abundance largely represented by the Alistipes genus, a well-known stress target in rodents and humans. The transfer of this altered microbiota into germ-free quails decreased their spatial and cue-based memory abilities as previously demonstrated in the stressed donors. The recipients also displayed increased anxiety-like behavior, reduced basal plasma corticosterone levels and differential gene expression in the brain. Furthermore, cecal microbiota transfer from a chronically stressed individual was sufficient to mimic the adverse impact of chronic stress on memory in recipient hosts and this action may be related to the Alistipes genus. Our results provide evidence of a feed-forward loop system linking the microbiota-gut-brain axis to stress and memory function and suggest that maintaining a healthy microbiota could help alleviate memory impairments linked to chronic stress. Availability of data and materials The datasets used during the current study are available from the corresponding author on reasonable request and they are available at https://doi.org/10.15454/JYITK4 .

Chronic stress and the gut microbiota appear to comprise a feed-forward loop, which contributes to the development of depressive disorders. Evidence suggests that memory can also be impaired by either chronic stress or microbiota imbalance. However, it remains to be established whether these could be a part of an integrated loop model and be responsible for memory impairments. To shed light on this, we used a two-pronged approach in Japanese quail: first stress-induced alterations in gut microbiota were characterized, then we tested whether this altered microbiota could affect brain and memory function when transferred to a germ-free host. The cecal microbiota of chronically stressed quails was found to be significantly different from that of unstressed individuals with lower α and β diversities and increased Bacteroidetes abundance largely represented by the Alistipes genus, a well-known stress target in rodents and humans. The transfer of this altered microbiota into germ-free quails decreased their spatial and cue-based memory abilities as previously demonstrated in the stressed donors. The recipients also displayed increased anxiety-like behavior, reduced basal plasma corticosterone levels and differential gene expression in the brain. Furthermore, cecal microbiota transfer from a chronically stressed individual was sufficient to mimic the adverse impact of chronic stress on memory in recipient hosts and this action may be related to the Alistipes genus. Our results provide evidence of a feed-forward loop system linking the microbiota-gut-brain axis to stress and memory function and suggest that maintaining a healthy microbiota could help alleviate memory impairments linked to chronic stress.

BackgroundBacterial consumption of viruses has never yet been reported, even though bacteria feed on almost anything. Viruses are omnipresent predators for all organisms, but have no acknowledged active biocontrol. The viral biomass undoubtedly reintegrates the trophic cycles, however the mechanisms of this phase still remain unknown.MethodsHere, we used stable isotope probing with 13C labelled T4 phages to monitor the increase of density of the bacterial DNA concomitant with the decrease of plaque forming units. We used 12C T4 phages as control.ResultsT4 phage disappearance in wastewater sludge was found to occur mainly through predation by Aeromonadacea. Phage consumption also favours significant in situ bacterial growth. Furthermore, an isolated strain of Aeromonas was observed to grow on T4 phages as sole source of carbon, nitrogen and phosphorus.Conclusionsbacterial species are capable of consuming bacteriophages in situ, which is likely a widespread and underestimated type of biocontrol.This assay is anticipated as a starting point for harnessing the bacterial potential in limiting the diffusion of harmful viruses within environments such as gut or water.

Understanding the transmission of antibiotic resistance genes (ARGs) is critical for human health. For this, it is necessary to identify which type of mobile genetic elements is able to spread them from animal reservoirs into human pathogens. Previous research suggests that in pig feces, ARGs may be encoded by bacteriophages. However, convincing proof for phage-encoded ARGs in pig viromes is still lacking, because of bacterial DNA contaminating issues. We collected 14 pig fecal samples and performed deep sequencing on both highly purified viral fractions and total microbiota, in order to investigate phage and prophage-encoded ARGs. We show that ARGs are absent from the genomes of active, virion-forming phages (below 0.02% of viral contigs from viromes), but present in three prophages, representing 0.02% of the viral contigs identified in the microbial dataset. However, the corresponding phages were not detected in the viromes, and their genetic maps suggest they might be defective. Furthermore, our dataset allows for the first time a comprehensive view of the interplay between prophages and viral particles.

International audience; The gut microbiota plays a key role in intestinal development at the suckling-to-weaning transition. The objective of this study was to analyze the production of metabolites by the gut microbiota in suckling and weaned piglets. We studied piglets raised in two separate maternity farms and weaned at postnatal day 21 in the same farm. The fecal metabolome (1H nuclear magnetic resonance) and the microbiota composition (16S rRNA gene amplicon sequencing) and its predicted functions (PICRUSt2) were analyzed in the same piglets during the suckling period (postnatal day 13) and 2 days after weaning (postnatal day 23). The relative concentrations of the bacterial metabolites methylamine, dimethylamine, cadaverine, tyramine, putrescine, 5-aminovalerate, succinate, and 3-(4-hydroxyphenylpropionate) were higher during the suckling period than after weaning. In contrast, the relative concentrations of the short-chain fatty acids acetate and propionate were higher after weaning than during the suckling period. The maternity of origin of piglets also influenced the level of some bacterial metabolites (propionate and isobutyrate). The fecal metabolome signatures observed in suckling and weaned piglets were associated with specific microbiota-predicted functionalities, structure, and diversity. Gut microbiota-derived metabolites, which are differentially abundant between suckling and weaned piglets (e.g., short-chain fatty acids and biogenic amines), are known to regulate gut health. Thus, identification of metabolome signatures in suckling and weaned piglets paves the way for the development of health-promoting nutritional strategies, targeting the production of bacterial metabolites in early life.

SUMMARYThe objective of this study was to analyze in piglets the impact of weaning on the production of metabolites by gut bacteria and to determine whether early life environment influences the functional maturation of the gut microbiota. Fecal metabolome and microbiome were analyzed in piglets raised in two separate maternity farms and mixed at weaning. In piglets from both maternity farms, the relative abundance ofLactobacillusand of the predicted function “Fucose degradation” decreased after weaning while the relative abundance ofRuminococcus 2and of the predicted function “Starch degradation” increased. In piglets from the first maternity farm, the relative concentration of biogenic amines and the relative abundance ofEscherichi-Shigelladecreased after weaning while the relative concentration of short chain fatty acids and the relative abundance ofChristensenellaceae R-7 groupandRuminococcaceae UCG-002increased. These changes were not observed at weaning in piglets from the second maternity farm probably because they already had high relative concentration of short chain fatty acids and higher relative abundance ofChristensenellaceae R-7 groupandRuminococcaceae UCG-002during the suckling period. In conclusion, the functional maturation of the microbiota at weaning is highly dependent on the maternal environment in piglets.ORIGINALITY - SIGNIFICANCE STATEMENTBacterial metabolites are key molecular intermediates between the gut microbiota and host cells. Our study in piglets reveals that the metabolic activity of the gut microbiota shifts at weaning, a key developmental period for intestinal and immune health. We also show that this functional maturation of the gut microbiota is strongly influenced by maternal environment. Thus, targeting early life environmental factors is a promising strategy to program health trough the production of beneficial bacterial metabolites at the suckling-to-weaning transition.

International audience; The objective of this study was to evaluate the impact of acute and repeated exposure to a diet naturally contaminated with mycotoxins (deoxynivalenol, DON) on the performance and fecal microbiota composition of finishing pigs. In total, 160 pigs raised in two successive batches were randomly distributed between four experimental groups. The control group received a standard finishing diet (uncontaminated) from 99 to 154 days of age. The experimental groups were fed a diet formulated with naturally DON-contaminated maize (3.02 mg DON/kg) for 7 days beginning on 113, 134, or both 113 and 134 days of age. Microbiota composition was analyzed using 16S rRNA sequencing applied to fecal samples collected at 99, 119, 140, and 154 days of age. DON exposure decreased daily feed intake (1.90 vs 2.64 kg/d following the first challenge period, 2.27 vs 3.27 kg/d following the second challenge period, P < 0.05). This resulted in a decrease in average daily gain measured over the entire experiment (0.94 vs 1.05 kg/d, P < 0.05). The DON challenge resulted in short-term modification of microbiota composition and bacterial functions, particularly those related to DON detoxification in the intestinal tract. After a 2-week recovery period, no difference in microbiota composition was observed between control and challenged pigs, suggesting resilience of the microbiota after perturbation caused by ingestion of a contaminated diet.; L’objectif de cette étude est d’évaluer l’impact d’expositions courtes et répétées à un aliment naturellement contaminé avec des mycotoxines et notamment de la déoxynivalénol (DON) sur les performances et la composition microbienne fécale de porcs en finition. Au total, 160 porcs issus de deux bandes successives ont été répartis aléatoirement entre quatre traitements expérimentaux. Le groupe témoin a reçu un aliment finition standard (non contaminé) entre 99 et 154 jours d’âge. Les groupes expérimentaux ont reçu un aliment formulé à partir de maïs naturellement contaminé par DON (3,02 mg DON/kg) pendant 7 jours à partir de 113, 134 et à partir à la fois de 113 et 134 jours. Une exposition à DON a entraîné une réduction de l’ingéré (1,90 vs2,64 kg/j suite à la première période d’exposition et 2,27 vs 3,27 kg/j suite à la deuxième période d’exposition, P < 0,05). Il en résulte une diminution du gain de poids mesuré sur l’ensemble de la période de test (0,94 kg/j vs 1,05 kg/j, P < 0,05). La composition du microbiote a été analysée par séquençage du gène de l’ARN 16S dans des fèces collectées à 99, 119, 140 et 154 jours d’âge. Une exposition à DON entraîne une modification temporaire de la composition microbienne et des fonctions bactériennes, notamment celles associées à la dégradation de DON dans le tube digestif. Après une période de récupération de 2 semaines, aucune différence n’est observée entre les animaux exposés et les témoins suggérant une forte résilience du microbiote suite à une perturbation liée à l’ingestion d’un aliment contaminé.

International audience; This study describes variability among farms of the response of microbiota to weaning and evaluates its relation to the robustness of piglets. Faecal samples and growth data were collected just before and seven days after weaning in 288 piglets from 16 commercial farms characterised by their pathogenic status and antimicrobial use. We observed in all farms the presence of four bacterial enterotypes (E1 to E4), differing notably in their abundance of Prevotella, Faecalibacterium, Roseburia, and Lachnospira, and corresponding to a gradual maturational shift from pre- to post-weaning microbiota. After weaning, of the 1 177 operational taxonomic units (OTU) contained in the dataset, 462 and 340 had their abundance affected (P < 0.05) by the use of antimicrobials and by the health status of the herd, respectively. Animals were then classified according to their relative average daily gain (rADG) from 26 to 48 days of age. The high rADG piglets had less Oscillospira (P < 0.05) and Campylobacter (P < 0.01), and more Prevotella (P < 0.01), Coprococcus (P < 0.01) and Lachnospira (P < 0.05) after weaning, which resembled the E4 enterotype. Thus, using animals from 16 farms, we identified four ubiquitous maturation enterotypes and found that piglets that reached enterotype E4 just after weaning might be more robust. Besides antimicrobial use, the general health status of the farm has a strong influence on the early post-weaning evolution of piglet microbiota.; Cette étude décrit la variabilité entre élevages de l’évolution du microbiote au moment du sevrage, et évalue la relation entre cette dernière et la robustesse des porcelets. Des échantillons fécaux et des enregistrements de croissance ont été collectés juste avant et sept jours après le sevrage pour 288 porcelets issus de 16 élevages commerciaux, caractérisés pour leur statut sanitaire et leur usage d’antibiotiques. Nous avons observé dans toutes les fermes la présence de quatre entérotypes bactériens (E1 à E4), différant notamment par leur abondance de Prevotella, Faecalibacterium, Roseburia, et Lachnospira, et correspondant à des stades de maturation graduelle pré- vers post-sevrage de la flore. Après le sevrage, des 1 177 unitéstaxonomiques (OTU) contenues dans le jeu de données, 462 et 340 voyaient leur abondance respectivement influencée par l’utilisation d’antibiotiques et par le statut sanitaire de l’élevage. Les animaux ont ensuite été classés selon leur gain de poids relatif (GMQr) calculé entre 26 et 48 jours d’âge. Les porcelets à haut GMQr avaient moins d’Oscillospira (P < 0,05) et Campylobacter (P < 0,01), et plus de Prevotella (P < 0,01), Coprococcus (P < 0,01) et Lachnospira (P < 0,05) après le sevrage, ce qui s’apparentait à l’entérotype E4. Ainsi, en utilisant des animaux provenant de 16 fermes, nous avons pu identifier quatreentérotypes maturationnels ubiquitaires, et supposé que les porcelets ayant atteint l’entérotype E4 juste après le sevrage pourraient être plus robustes. Hormis l’usage d’antibiotiques, le statut sanitaire global de l’élevage a une influence importante sur l’évolution précoce du microbiote après le sevrage.

Metabarcoding of the 16S rRNA gene is commonly used to characterize microbial communities, by estimating the relative abundance of microbes. Here, we present a method to retrieve the concentrations of the 16S rRNA gene per gram of any environmental sample using a synthetic standard in minuscule amounts (100 ppm to 1% of the 16S rRNA sequences) that is added to the sample before DNA extraction and quantified by two quantitative polymerase chain reaction (qPCR) reactions. This allows normalizing by the initial microbial density, taking into account the DNA recovery yield. We quantified the internal standard and the total load of 16S rRNA genes by qPCR. The qPCR for the latter uses the exact same primers as those used for Illumina sequencing of the V3‐V4 hypervariable regions of the 16S rRNA gene to increase accuracy. We are able to calculate the absolute concentration of the species per gram of sample, taking into account the DNA recovery yield. This is crucial for an accurate estimate as the yield varied between 40% and 84%. This method avoids sacrificing a high proportion of the sequencing effort to quantify the internal standard. If sacrificing a part of the sequencing effort to the internal standard is acceptable, we however recommend that the internal standard accounts for 30% of the environmental 16S rRNA genes to avoid the PCR bias associated with rare phylotypes. The method proposed here was tested on a feces sample but can be applied more broadly on any environmental sample. This method offers a real improvement of metabarcoding of microbial communities since it makes the method quantitative with limited efforts., Here, we present a spike‐and‐recovery method to get quantitative estimates from 16S rRNA surveys. The method relies on adding an artificial strand of DNA to the lysis buffer before the DNA extraction and measuring its recovery either by direct sequencing or by quantitative polymerase chain reaction (qPCR). The low detection limit achieved by qPCR allows to add minute amounts of the internal standard so that the sequencing effort is focused on the unknown sequences.

Antibiotic resistance of microbes thriving in the animal gut is a growing concern for public health as it may serve as a hidden reservoir for antibiotic resistance genes (ARGs). We compared 16 control piglets to 24 piglets fed for 3 weeks with S1 or S2 fecal suspensions from two sows that were not exposed to antibiotics for at least 6 months: the first suspension decreased the erythromycin resistance gene ermB and the aminoglycoside phosphotransferase gene conferring resistance to kanamycine (aphA3), while the second decreased the tetracycline resistance gene tetL, with an unexpected increase in ARGs. Using 16S RNA sequencing, we identified microbial species that are likely to carry ARGs, such as the lincosamide nucleotidyltransferase lnuB, the cephalosporinase cepA, and the tetracycline resistance genes tetG and tetM, as well as microbes that never co-exist with the tetracycline resistance gene tetQ, the erythromycin resistance gene ermG and aphA3. Since 73% of the microbes detected in the sows were not detected in the piglets at weaning, a neutral model was applied to estimate whether a microbial species is more important than chance would predict. This model confirmed that force-feeding modifies the dynamics of gut colonization. In conclusion, early inoculation of gut microbes is an interesting possibility to stimulate gut microbiota towards a desirable state in pig production, but more work is needed to be able to predict which communities should be used.

International audience; Mycotoxins are a major contaminant of pig feed and have negative effects on health and performance. The present study investigated the impact of single or repeated acute challenges with a diet naturally contaminated with deoxynivalenol (DON) and zearalenone (ZEN) on growth performances of finishing pigs and their fecal microbiota composition. A total of 160 pigs (castrated males and females) in two successive batches were randomly divided into four experimental groups of 40 pigs each. The control group received a control finisher diet from 99 to 154 days of age. Challenged groups were subjected to a 7-day acute challenge by being fed a DON- and ZEN-contaminated diet (3.02 mg DON/kg feed and 0.76 mg ZEN/kg feed) at 113 days (group DC), 134 days (group CD) or both 113 and 134 days (group DD). Microbiota composition was analyzed via 16S rRNA sequencing from fecal samples collected from the 80 females at 99, 119, 140 and 154 days. Challenged pigs (i.e. groups DC, CD and DD) reduced their average daily feed intake by 25% and 27% ( P < 0.001) and feed efficiency by 34% and 28% ( P < 0.05) during the first and second mycotoxin exposure, respectively. Microbiota composition was affected by mycotoxin exposure ( P = 0.07 during the first exposure and P = 0.01 during the second exposure). At the family level, mycotoxin exposure significantly ( P < 0.05) decreased the relative abundances of Ruminococcaceae, Streptococcaceae and Veillonellaceae and increased that of Erysipelotrichaceae at both 119 and 140 days of age. After the 7-day DON/ZEN challenge, the relative abundance of 6 to 148 operational taxonomic units (OTUs) differed among the treatment groups. However, none of these OTUs changed in all treatment groups. Using 27 functional pathways, pigs exposed to DON/ZEN challenges could be distinguished from control pigs using sparse partial least squares discriminant analysis, with a 15% misclassification rate. Regarding the functionality of these predictors, two pathways were involved in detoxifying mycotoxins: drug metabolism and xenobiotic metabolism by cytochrome P450. In challenged pigs, microbiota composition returned to the initial state within 3 weeks after the end of a single or repeated DON/ZEN challenge, highlighting the resilience of the gut microbiome. The feeding and growth performances of the pigs during challenge periods were significantly correlated with biological pathways related to health problems and modifications in host metabolism. To conclude, short-term DON/ZEN challenges resulted in transient modifications in the composition and functions of fecal microbiota.

The present study aimed at investigating the evolution of pigs’ fecal microbiota composition from post-weaning to finishing in a longitudinal analysis. The experiment was conducted on 160 Pietrain × (Large White × Landrace) castrated male and female pigs in two replicates. Feces were collected at 52, 99, 119, 140, and 154 days of age for further 16S rRNA sequencing to analyze the microbiota composition. Pig microbiota evolved strongly from 52 to 99 days of age with an increased abundance of Streptococcaceae and a decreased abundance of Lactobacillaceae. During the finishing stage, microbiota kept evolving at a slower rate. To link the community structure to the performances, the fecal samples were clustered into enterotypes sharing a similar bacterial composition. At 52 days, two enterotypes dominated either by Lactobacillus or by Prevotella–Sarcina were identified. They differed from the two enterotypes determined from 99 to 154 days which were dominated either by Lactobacillus or by Turicibacter–Clostridium sensu stricto. During this time period, 75% of the pigs switched enterotypes. The enterotypes were not related to differences in the overall growth or feeding performance. The enterotype definition was time-dependent and seemed to be related to the sex type at 99 days of age.

The interaction between the gut microbiota (GM) and the brain has led to the concept of the microbiota-gut-brain axis but data for birds remain scarce. We tested the hypothesis that colonization of germ-free chicks from a quail line selected for a high emotional reactivity (E+) with GM from a line with low emotional reactivity (E-) would reduce their emotional behaviour in comparison with germ-free chicks from an E+ line colonized with GM from the same E+ line. The GM composition analysis of both groups revealed a shift in terms of microbial diversity and richness between day 21 and day 35 and the GM of the two groups of quails were closer to each other at day 35 than at day 21 at a phylum level. Quails that received GM from the E- line expressed a lower emotional reactivity than quails colonized by GM from the E+ line in tonic immobility and novel environment tests carried out during the second week of age. This result was reversed in a second tonic immobility test and an open-field run 2 weeks later. These behavioural and GM modifications over time could be the consequence of the resilience of the GM to recover the equilibrium present in the E+ host, which is in part driven by the host genotype. This study shows for the first time that a GM transfer can influence emotional reactivity in Japanese quails, supporting the existence of a microbiota-gut-brain axis in this species of bird.

International audience; The interaction between the gut microbiota (GM) and the brain has led to the concept of the microbiota-gut-brain axis but data in birds remain scarce. We tested the hypothesis that colonization of germ-free chicks from a quail line selected for a high emotional reactivity (E+) with GM from a line with low emotional reactivity (E-) would reduce their emotional behaviour in comparison with germ-free chicks from E+ line colonized with GM from the same E+ line. The GM composition analysis of both groups revealed a shift in term of microbial diversity and richness between Day 21 and Day 35 and the GM of the two groups of quails were closer to each other at Day 35 than at Day 21 at a phylum level. Quails that received GM from the E- line expressed a lower emotional reactivity than the quails colonized by GM from the E+line in the tonic immobility and the novel environment tests proceeded during the second week of age. This result was reversed in a second tonic immobility test and an open-field run two weeks later. These behavioural and GM modifications over time could be the consequence of the resilience of the GM to recover its equilibrium present in the E+ host, which is in part driven by the host genotype. This study shows for the first time that a gut microbiota transfer can influence emotional reactivity in Japanese quails strengthening the existence of a microbiota-gut-brain axis in this species of bird.

International audience; The aim of this experiment was to evaluate the significance of neonatal environment on feed efficiency. For that purpose, rabbits from a line selected for residual feed intake (RFI) during 10 generations (G10 kits) were cross-fostered with non-selected control does (i.e., G0 line), and reciprocally. In parallel, sibs were fostered by mothers from their original line. Nine hundred animals were raised in individual (N = 456) or collective (N = 320) cages. Traits analysed in this study were body weight at 32 days and at 63 days, average daily gain (ADG), feed intake between weaning and 63 days (FI), feed conversion ratio (FCR) and RFI. The maternal environment offered by does from the line selected for RFI deteriorated the FCR of the kits, independently of their line of origin, during fattening (+0.08 +/- 0.02) compared to FCR of kits nursed by G0 does. The line, the type of housing and the batch were significant effects for all the measured traits: G10 kits were lighter than their G0 counterparts at 32 days (-82.9 +/- 9 g, p < 0.0001) and at 63 days (-161 +/- 16 g, p < 0.0001). They also had a lower ADG (-2.36 +/- 0.36 g/day, p < 0.0001), RFI (-521 +/- 24 g/day, p < 0.0001) and a lower FI (-855 +/- 31 g, p < 0.0001), resulting in a more desirable feed efficiency (FCR: -0.35 +/- 0.02). There was no significant difference in the contrast of G10 and G0 performances between collective and individual/digestive cages (p > 0.22): -2.35 g/day versus 2.94 g/day for ADG, -0.39 versus -0.40 for FCR, -577 g versus -565 g for RFI and -879 g versus -859 g for FI, respectively). Thus, no genotype-by-environment (housing) interaction is expected at the commercial level, that is, no re-ranking of the animals due to collective housing.

This study describes the response of piglet microbiota to weaning in various environments, and investigates whether microbiota composition is associated with the robustness of piglets. Faecal samples and growth data were collected just before and 7 days after weaning in 288 piglets from 16 commercial farms characterised by their pathogenic status and antimicrobial use. The effect of weaning on the most abundant microbial families of faecal microbiota was roughly the same in all farms and in agreement with previous findings. Four enterotypes, ubiquitous in all farms, were identified, for which the most discriminating genera werePrevotella,Faecalibacterium,Roseburia, andLachnospira.They corresponded to a gradual maturational shift from pre to post-weaning microbiota. Besides antimicrobial use, the pathogenic status of the farm had a major influence on the post-weaning microbial evolution of apparently healthy piglets. Regarding individual characteristics, piglets whose growth was less perturbed by weaning had moreBacteroidetes(P < 0.01) and lessProteobacteria (P < 0.001).In response to weaning, they showed a greater increase inPrevotella(P < 0.01),Coprococcus(P< 0.01) andLachnospira(P < 0.05) than piglets that grew more slowly. Thus, the microbiota of robust piglets shares common characteristics regardless the living environment of animals.

To get insights into selection criteria for feed efficiency, 2 rabbit lines have been created: the ConsoResidual line was selected for residual feed intake (RFI) with ad libitum feeding and the ADGrestrict line was selected for ADG under restricted feeding (-20% of voluntary intake). The first objective of this study was to evaluate, after 9 generations of selection, the direct and correlated responses to selection on production traits in the 2 lines for traits recorded during growth. Second, applying the 2 feeding conditions used for selection to both selected lines plus the control unselected line (generation 0, G0) in a 2 × 3 factorial trial, the line performances were compared and the gut microbiota of the lines was characterized. The correlated responses in feed conversion ratio (FCR) were remarkably equivalent in both selected lines (-2.74 genetic σ) but correlated responses in other traits were notably different. In the ConsoResidual line, selection for decreased RFI resulted in a small negative correlated response in BW at 63 d old (BW63) and in a null response in ADG. In the ADGrestrict line, on the contrary, the correlated response in BW63 was substantial (+1.59 σ). The 2 selected lines had a FCR reduced by 0.2 point compared with the G0 line, and the same difference was found in both feeding regimens ( < 0.001). Indeed, selection on ADG would lead to heavier animals with no significant reduction of feed costs, whereas selection on RFI leads to lower feed costs and no increase of animal BW under ad libitum feeding. Altogether, our results do not suggest any genotype × environment interaction in the response to feeding regimens. The intestinal microbial communities from efficient rabbits differed from their unselected counterparts in terms of fermentation end products and microbial phylotypes, suggesting a central role of these microbes in the better feed efficiency of the rabbits.

The present study aimed at describing the impact of a deoxynivalenol (DON) - contaminated diet on microbiota composition. A total of 147 (Large White x Landrace) x Piétrain pigs from two replicates were affected to four different experimental treatments. Pigs in the control group (CC) received a control diet from 99 to 154 days of age. Groups DC, CD, and DD were given the DON-contaminated diet (3.02 mg DON/kg feed) for 7 days at 113, at 134, and at 113 and 134 days of age, respectively. The DON-contaminated diet was formulated with a naturally contaminated corn. The body weight gain and the feed intake were daily recorded from day 99 to 154. Fecal samples were collected at the beginning of the experiment (d99), immediately after the end of the 7-days DON challenges (d119 and 140), and at the end of the experiment (d154) for further 16S rRNA sequencing. During challenge periods, ADFI was decreased by 26% to 32% (P < 0.05) and ADG by 40% to 60% (P < 0.05). Short-term DON challenges induced transient changes in microbiota composition. Two weeks after the end of the DON challenges, this composition went back to control state. Whatever the age, DON challenged pigs could be discriminated from controls in a sparse PLS discriminant analysis based on 88 operational taxonomic units or 27 functional pathways with 16% error-rate. The main predictors raised by this discrimination belonged to Streptococcaceae, Lachnospiraceae and Clostridiaceae families. In our experimental conditions, changes in microbiota composition observed during DON challenges were poorly correlated to changes in ADG and ADFI. This experiment revealed a transient modification of the microbiota composition following the exposition to DON-contaminated diet, with no long-term impact on pigs’ performance. This study is part of the Feed-a-Gene Project funded from the European Union’s H2020 Program (grant 633531).

The present study aimed at predicting feed efficiency (FE) based on fecal microbiota, using partial least square regression (PLSR), sparse PLSR, and random forest regression (RF). Fecal samples from 147 Pietrain x (Large White x Landrace) pigs reared in two consecutive batches were collected at 99 days of age. Daily live body weight and feed intake were individually measured in pigs fed ad libitum with a corn soybean diet. The relative abundances of operational taxonomic units (OTU) resulting from fecal 16S rRNA sequencing were used to build the prediction models of FE between 99 and 113 days. From these data, neither PLSR nor RF models have been validated on external datasets. An important over-fitting has been observed in PLSR. With this aim to test the ability of the methods to retrieve information, synthetic OTU were created to fit an artificial Pearson correlation with FE (r² = 0 to 0.9) and were added among the predictors in the dataset. Artificial OTU correlated above 0.37 with FE improved the prediction in sparse PLSR and RF, and reduced the over-fitting. The best predictions were achieved by sparse PLSR. The present study emphasized the ability of sparse PLSR and RF to build valid prediction models of a quantitative phenotype, based on fecal microbiota composition. Since no OTU was correlated above 0.30 with FE in the real dataset, the power of the prediction methods was not enough to extract useful information from the fecal microbiota. The functional redundancy of the microbiota could explain the lack of relevant information in the real dataset to predict pigs’ quantitative phenotype. These results suggest that the best strategy is to run sparse PLSR only if a correlation higher than 0.37 is observed. This study is part of the Feed-a-Gene Project funded from the European Union’s H2020 Program (grant 633531).

International audience; Gut microbiota is a key contributor to feed use in monogastric species, in particular via the digestion dietary fibres. Molecular techniques are now available to run large studies and decipher the potential of gut microbiota to improve livestock. Studies on human and mice are more advanced: different factors have been demonstrated to influence the gut microbiota composition and functions, including maternal transmission, environment (diet composition and quantity, humidity and heat), age and physiological status. Studies also evaluated if the host controls its gut microbiota. In pigs, chicken and rabbits, microbiota differences between animals of extreme phenotypes within populations, and between lines divergently selected for specific traits, are reported. Other studies reported that some microbiota abundancies are heritable. Linear mixed models have been used to evaluate its contribution to trait variability, or microbiability, with different data (full vs 16S sequencing of gut or faecal contents) and different variance matrices. They showed significant contribution to production traits, reaching more than 30%. However, confounding effects exist, such as the maternal inheritance of the microbiota and the genetic determinism by the host : many studies show reduction of the genetic additive variance, in addition to reduction of the residual variance with microbiability. Specific datasets, testing a given genetic in different environments or using cross-fostering, are used to better understand the relative contribution of each effect (host genetics and microbiota) to the trait variance, and propose solutions to livestock.

International audience; Microbial population in the gastrointestinal tract plays a central role in health and nutrient digestion. The objective of the present study was to investigate the relationships between microbiota and apparent digestibility coefficients with respect to age and diet. Pigs from Large-White, Duroc or Pietrain breeds were raised under the same housing conditions and fed alternately a low-fiber (LF) and a high-fiber diet (HF) during 4 successive 3-week periods. Data collection for digestibility measurements was achieved during the last week of each period. At the end of each period, fecal microbiota was collected for 16S rRNA gene sequencing. The microbiota remained stable across periods whereas digestibility of energy, crude proteins and cell wall components increased. The microbiota was resilient to diet effect and pigs fed the LF diet were discriminated to those fed the HF diet using 31 predicting OTUs with a mean classification error-rate of 3.9%. Clostridiaceae and Turicibacter were negatively correlated whereas Lactobacillus was positively correlated with protein and energy digestibility coefficients in the LF group. In addition, Lachnospiraceae and Prevotella were negatively correlated with cell wall component digestibility. In contrast, no significant correlation was found between microbiota composition and digestibility coefficients when pigs were fed the HF diet. Interestingly, it was also no longer possible to distinguish animals from different breeds once the animals were fed a HF diet, so that the microbiota could only trace the breed origin in the first period and in the LF group. In our experimental conditions, 3 weeks of adaptation to a new diet seems to be sufficient to observe resilience in growing pigs' microbiota. We demonstrated that fecal microbiota can be used to classify pigs according to their dietary treatment. Some bacteria are favorable or unfavorable to digestibility. This suggests that manipulations of bacterial populations can improve digestibility and feed efficiency.

Dietary fiber content and composition affect microbial composition and activity in the gut, which in turn influence energetic contribution of fermentation products to the metabolic energy supply in pigs. This may affect feed efficiency (FE) in pigs. The present study investigated the relationship between the fecal microbial composition and FE in individual growing-finishing pigs. In addition, the effects of diet composition and sex on the fecal microbiome were studied. Fecal samples were collected of 154 grower-finisher pigs (3-way crossbreeds) the day before slaughter. Pigs were either fed a diet based on corn/soybean meal (CS) or a diet based on wheat/barley/by-products (WB). Fecal microbiome was characterized by 16S ribosomal DNA sequencing, clustered by operational taxonomic unit (OTU), and results were subjected to a discriminant approach combined with principal component analysis to discriminate diets, sexes, and FE extreme groups (10 high and 10 low FE pigs for each diet by sex-combination). Pigs on different diets and males vs. females had a very distinct fecal microbiome, needing only 2 OTU for diet (P = 0.020) and 18 OTU for sex (P = 0.040) to separate the groups. The 2 most important OTU for diet, and the most important OTU for sex, were taxonomically classified as the same bacterium. In pigs fed the CS diet, there was no significant association between FE and fecal microbiota composition based on OTU (P > 0.05), but in pigs fed the WB diet differences in FE were associated with 17 OTU in males (P = 0.018) and to 7 OTU in females (P = 0.010), with 3 OTU in common for both sexes. In conclusion, our results showed a diet and sex-dependent relationship between FE and the fecal microbial composition at slaughter weight in grower-finisher pigs.

Limiting the post-weaning intake of the young rabbit is known to improve its resistance to digestive disorders, whereas a degradation of its housing hygiene is assumed to have a negative impact on its health. This study aims at providing insights into the mechanism of digestive health preservation regarding both host (growth and immune response) and its symbiotic digestive microbiota. A 2×2 factorial design from weaning (day 28) to day 64 was set up: ad libitum intake or restricted intake at 70% of ad libitum, and high v. low hygiene of housing (n=105 per group). At day 36 and day 45, 15 animals/group were subcutaneously immunized with ovalbumin (OVA) to assess their specific immune response. Blood was sampled at 36, 45, 57 and 64 days of age to determine total and anti-OVA immunoglobulin type G (IgG) and haptoglobin levels. The cecal bacterial community was explored (18 per group) by 454 pyrosequencing of genes coding for the 16S ribosomal RNA, whereas cecal pH, NH3 and volatile fatty acid (VFA) concentrations were measured to characterize fermentative activity. A 30% reduction in feed intake reduced the growth by only 17% (P

International audience; The addition of sodium chloride to freshwater or diluted minimal salt medium increases the adsorption of T2 phages on Escherichia coli. For the first time the adsorption in diluted minimal salt medium was measured by counting unadsorbed phages (i.e. free particles) using flow cytometry, allowing a gentle separation between adsorbed and unadsorbed phages. Flow cytometry was able to detect weakly adsorbed phage that remained undetected using classical centrifugation-based methods and this allowed us to show that increasing ionic strength enhances the phage adsorption to its bacterial host with an extremely low detection limit. A key result was that the adsorption in high ionic strength (i.e. 100 mM) reached 4.5 +/- 0.1 x 10(-5) mL/min which is 1400 fold higher than previously reported values.In order to understand the mechanism underpinning such a weak phage adsorption, the zeta potentials and the diffusion coefficient of the particles were measured by dynamic light scattering. The bacterial cells and the phages had zeta potentials between -60 mV and -10 mV and -30 mV and -10 mV, respectively. The diffusion coefficient of the phage was 2.8 +/- 0.4 x 10(-12) m(2) s(-1) corresponding to a hydrodynamic radius of 104 +/- 15 nm. However significant adsorption occurs in conditions where the DLVO theory predicts that minimal encounter, suggesting that forces other that electrostatic repulsion and Van der Waals interaction (e.g. potential impurities, particle shape and other biological characteristics) are likely to interplay.

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International audience; Changes in bacterial and fungal communities in chicken litter with high and low moisture content over a five week period during a single chicken grow out cycle in a poultry shed in subtropical Australia were investigated to study the association between specific microbes and odour production. 2. Microbial biomass, as indicated by DNA yields, was higher and community composition was more dynamic over time in moist compared with dry chicken litter. 3. Bacillus, Atopostipes and Aspergillus species increased in relative abundance in moist chicken litter samples over time reflecting the relatively high fitness and hence activity of these specific bacteria and this specific fungus in this environment.

National audience; The natural coprophagy behavior expressed by young rabbits can be amplified by including hard feces in the nest box. The objectives of our study were 1) to describe the feasibility of this practice under commercial conditions, and 2) to study the interests and limits of this practice on offsprings’ health until the slaughter age. The experiment was conducted on 363 litters in a professional unit that stop to use antibiotics for more than 10 years, during 5 consecutive batches from birth to 70 days of age. In the group F (feces added, n = 183litters), exogenous hard feces (5 pellets per day and nest) were manually added to the nest between days 2 and 18 of lactation. The exogenous hard feces were obtained from future reproductive females reared in the same unit. In the group T (control, n = 180 litters) no hard feces were added to the nest box, but the natural female behavior of dropping hard feces in the nest was accepted. Manually adding hard feces in the nest box did not improved the offspring mortality rate during lactation (T: 7,5 % vs. F: 7,2 %). A significant interaction between treatment and batch on the mortality rate after weaning was observed, with a favorable result for batch 2 (T: 20,4 % vs. F: 8,2 %). The difference observed between the results obtained here under professional conditions and previously in experimental conditions, shows that additional studies regarding the origin of feces, when starting the treatment and at which period of the day, are necessary to validate this practice on professional conditions.; Le comportement naturel de coprophagie exprimé par les jeunes lapereaux au nid peut être amplifié par l’ajout de fèces dures exogènes. L'objectif de notre étude est 1) d'évaluer la faisabilité de cette pratique en conditions d’élevage professionnel, et 2) d’étudier son intérêt et ses limites sur la santé des lapereaux jusqu’à l’âge de la vente. L’expérience a été réalisée sur 363 portées, dans un élevage professionnel n'utilisant pas d'antibiotiques depuis plus de 10 ans, et pendant 5 bandes consécutives, de la naissance à 70 jours d’âge. Dans le groupe F (fèces ajoutés, n = 183 portées), des fèces exogènes (5 crottes par jour et par nid) ont été ajoutés dans le nid entre 2 et 18 jours de lactation. Ces fèces exogènes provenaient de femelles futures reproductrices du même élevage. Dans le groupe T (Témoin, n = 180 portées), aucun apport de fèces exogène n’a été réalisé, mais le comportement naturel de dépôt des fèces dures dans les nids était respecté. L’ajout de fèces dures exogènes dans les nids n’a pas amélioré le taux de mortalité des lapereaux pendant l’allaitement (T: 7,5 % vs. F: 7,2 %). Une interaction significative entre les effets traitement et bande a été observée pour le taux de mortalité après sevrage, où une influence favorable pendant la bande 2 (T: 20,4 % vs. F: 8,2 %) a été constatée. Les écarts de résultats obtenus ici entre élevage professionnel et préalablement en conditions expérimentales, démontrent qu'il faut des études complémentaires sur l’origine des fèces, le moment du début du traitement et la période de la journée à laquelle les fèces exogènes sont déposées dans le nid, pour valider cette pratique.

Aims: To evaluate the importance of the bacterial composition on the resilience of the organic matter assimilation in the sea. Methods and Results: Chemostats were inoculated with coastal and offshore bacterial communities. Bacterial density and protein synthesis increased before stabilizing, and this response to confinement was more marked in the offshore chemostats. Before the toluene perturbation the community structure in the coastal chemostats remained complex whereas the offshore chemostats became dominated by Alteromonas sp. After the perturbation, bacterial protein synthesis was inhibited before peaking briefly at a level fivefold to that observed before the perturbation and then stabilizing at a level comparable to that before the perturbation. Alteromonas dominated both the coastal and the offshore communities immediately after the perturbation and the coastal communities did not recover their initial complexity. Conclusions: Cell lysis induced by the toluene perturbation favoured the growth of Alteromonas which could initiate growth rapidly in response to the nutrient pulse. Despite their different community structure in situ, the resilience of protein synthesis of coastal and offshore bacterial communities was dependent on Alteromonas, which dominated in the chemostats. Significance and Impact of the Study: Here we show that although Alteromonas sp. dominated in artificial offshore and coastal communities in chemostats, their response time to the shock was different. This suggests that future perturbation studies on resilience in the marine environment should take account of ecosystem history.