The link between broiler flock heterogeneity and cecal microbiome composition.

BACKGROUND: Despite low genetic variation of broilers and deployment of considerate management practices, there still exists considerable body weight (BW) heterogeneity within broiler flocks which adversely affects the commercial value. The purpose of this study was to investigate the role of the cecal microbiome in weight differences between animals. Understanding how the gut microbiome may contribute to flock heterogeneity helps to pave the road for identifying methods to improve flock uniformity and performance. RESULTS: Two hundred eighteen male broiler chicks were housed in the same pen, reared for 37 days, and at study end the 25 birds with highest BW (Big) and the 25 birds with lowest BW (Small) were selected for microbiome analysis. Cecal contents were analyzed by a hybrid metagenomic sequencing approach combining long and short read sequencing. We found that Big birds displayed higher microbial alpha diversity, higher microbiome uniformity (i.e. lower beta diversity within the group of Big birds), higher levels of SCFA-producing and health-associated bacterial taxa such as Lachnospiraceae, Faecalibacterium, Butyricicoccus and Christensenellales, and lower levels of Akkermansia muciniphila and Escherichia coli as compared to Small birds. CONCLUSION: Cecal microbiome characteristics could be linked to the size of broiler chickens. Differences in alpha diversity, beta diversity and taxa abundances all seem to be directly associated with growth differences observed in an otherwise similar broiler flock.

A Humanized Diet Profile May Facilitate Colonization and Immune Stimulation in Human Microbiota-Colonized Mice.

BACKGROUND: In spite of the importance of the use of gnotobiotic mice for human fecal transfer, colonization efficiency and immune stimulation after human microbiota inoculation in mice are poorly studied compared to mouse microbiota inoculation. We tested the colonization efficiency and immune responses in mice bred for one additional generation after inoculating the parent generation with either a human (HM) or a mouse microbiota (MM). Furthermore, we tested if colonization efficiency and immune stimulation could be improved in HM-colonized mice by dietary approaches: if these were fed a diet closer to the human diet either in its sources of animal fat and protein [the "animal source" (AS) diet] or in its proportions of macronutrients from the normal sources of a mouse diet [the "human profile" (HP) diet]. RESULTS: Although significantly lower in mice with a human microbiota (30-40% vs. 61-70%) the colonization efficiency was significantly higher in HM mice fed the HP diet (40%), and in MM mice fed AS (70%). The microbiota of mice fed HP was comparable to the microbiota of mice fed a standard rodent chow, while the microbiota of mice fed the animal source diet (AS) clustered separately. Mice inoculated with mouse fecal matter had significantly more CD4+ T cells and Cd4 expression and significantly fewer regulatory T cells (Tregs) and FoxP3 expression than human microbiota inoculated mice, but cell proportions differences were mostly apparent between mice fed the AS diet. Mice fed the HP diet had significantly higher expression of Cd8a. CONCLUSION: It is concluded that a diet with a humanized profile could support the establishment of a human microbiota in mice, which will, however, still elicit a lower colonization efficiency compared to mice inoculated with a mouse microbiota.

Human microbiota-transplanted C57BL/6 mice and offspring display reduced establishment of key bacteria and reduced immune stimulation compared to mouse microbiota-transplantation.

Transplantation of germ-free (GF) mice with microbiota from mice or humans stimulates the intestinal immune system in disparate ways. We transplanted a human microbiota into GF C57BL/6 mice and a murine C57BL/6 microbiota into GF C57BL/6 mice and Swiss-Webster (SW) mice. Mice were bred to produce an offspring generation. 56% of the Operational Taxonomic Units (OTUs) present in the human donor microbiota established in the recipient mice, whereas 81% of the C57BL/6 OTUs established in the recipient C57BL/6 and SW mice. Anti-inflammatory bacteria such as Faecalibacterium and Bifidobacterium from humans were not transferred to mice. Expression of immune-related intestinal genes was lower in human microbiota-mice and not different between parent and offspring generation. Expression of intestinal barrier-related genes was slightly higher in human microbiota-mice. Cytokines and chemokines measured in plasma were differentially present in human and mouse microbiota-mice. Minor differences in microbiota and gene expression were found between transplanted mice of different genetics. It is concluded that important immune-regulating bacteria are lost when transplanting microbiota from humans to C57BL/6 mice, and that the established human microbiota is a weak stimulator of the murine immune system. The results are important for study design considerations in microbiota transplantation studies involving immunological parameters.

Humanizing the gut microbiota of mice: Opportunities and challenges.

Mouse colonized with human fecal microbiota is an interesting model concept with pros and cons like any other model system. The concept provides an ecologically relevant context to study food component and drug metabolism, and is an invaluable tool for phenotype transfer studies to prove the role of the gut microbiota in health and disease. The major drawbacks are the difficulties with transferring certain components of the human microbiota to the recipient mice, and immunological abnormalities observed in these mice. There seem to be unexplored opportunities for trying to reduce these limitations, but careful evaluation of pros, cons and possible alternatives is still necessary.

Microbiota composition of simultaneously colonized mice housed under either a gnotobiotic isolator or individually ventilated cage regime.

Germ-free rodents colonized with microbiotas of interest are used for host-microbiota investigations and for testing microbiota-targeted therapeutic candidates. Traditionally, isolators are used for housing such gnotobiotic rodents due to optimal protection from the environment, but research groups focused on the microbiome are increasingly combining or substituting isolator housing with individually ventilated cage (IVC) systems. We compared the effect of housing systems on the gut microbiota composition of germ-free mice colonized with a complex microbiota and housed in either multiple IVC cages in an IVC facility or in multiple open-top cages in an isolator during three generations and five months. No increase in bacterial diversity as assessed by 16S rRNA gene sequencing was observed in the IVC cages, despite not applying completely aseptic cage changes. The donor bacterial community was equally represented in both housing systems. Time-dependent clustering between generations was observed in both systems, but was strongest in the IVC cages. Different relative abundance of a Rikenellaceae genus contributed to separate clustering of the isolator and IVC communities. Our data suggest that complex microbiotas are protected in IVC systems, but challenges related to temporal dynamics should be addressed.

Antibiotic-treated versus germ-free rodents for microbiota transplantation studies.

We recently investigated the applicability of antibiotic-treated recipient mice for transfer of different gut microbiota profiles. With this addendum we elaborate on perspectives and limitations of using antibiotics as an alternative to germ-free (GF) technology in microbial transplantation studies, and we speculate on the housing effect. It is possible to transfer host phenotypes via fecal transplantation to antibiotic-treated animals, but problems with reproducibility, baseline values, and antibiotic resistance genes should be considered. GF animals maintained in isolators still seem to be the best controlled models for long-term microbial transplantation, but antibiotic-treated recipients are also commonly utilized. We identify a need for systematic experiments investigating the stability of microbial transplantations by addressing 1) the recipient status as either GF, antibiotic-treated or specific pathogen free and 2) different levels of protected housing systems. In addition, the developmental effect of microbes on host physiological functions should be evaluated in the different scenarios.

Gastrointestinal microbiota and local inflammation during oxazolone-induced dermatitis in BALB/cA mice.

At present, laboratory animals are not standardized with regard to the gastrointestinal microbiota (GM), but differences in this feature may alter various parameters in animal models. We hypothesized that variation in the GM correlated with variation in clinical parameters of a murine oxazolone-induced skin inflammation model of atopic dermatitis. BALB/cA mice were sensitized with oxazolone over a 28-d period and variation in gastrointestinal microbiota in fecal and cecal samples was assessed by PCR-denaturing gradient gel electrophoresis. Clinical parameters included transepidermal water loss, ear thickness, inflammatory factors in ear tissue and plasma, and histopathologic evaluation. The fecal microbiota before induction of skin inflammation strongly correlated with the levels of some proinflammatory cytokines (IFNγ, IL1ß, IL12, and TNFα), the antiinflammatory cytokines IL4 and IL10, and the chemokine KC/GRO that were measured in ear samples at study termination. Cecal microbiota at termination correlated with ear thickness and transepidermal water loss. There was no correlation between cytokine responses and ear thickness or transepidermal water loss. In addition, GM changed during the study period in the oxazolone-treated mice, whereas this was not the case for the control mice. The current study shows that the GM of mice influences the development of oxazolone-induced skin inflammation and that the model itself likely induces a pathophysiologic response that alters the composition of the GM.