Experimental design considerations in microbiota/inflammation studies

There is now convincing evidence that many inflammatory diseases are precipitated, or at least exacerbated, by unfavourable interactions of the host with the resident microbiota. The role of gut microbiota in the genesis and progression of diseases such as inflammatory bowel disease, obesity, metabolic syndrome and diabetes have been studied both in human and in animal, mainly rodent, models of disease. The intrinsic variation in microbiota composition, both within one host over time and within a group of similarly treated hosts, presents particular challenges in experimental design. This review highlights factors that need to be taken into consideration when designing animal trials to investigate the gastrointestinal tract microbiota in the context of inflammation studies. These include the origin and history of the animals, the husbandry of the animals before and during experiments, details of sampling, sample processing, sequence data acquisition and bioinformatic analysis. Because of the intrinsic variability in microbiota composition, it is likely that the number of animals required to allow meaningful statistical comparisons across groups will be higher than researchers have generally used for purely immune-based analyses

Experimental design considerations in microbiota/inflammation studies

There is now convincing evidence that many inflammatory diseases are precipitated, or at least exacerbated, by unfavourable interactions of the host with the resident microbiota. The role of gut microbiota in the genesis and progression of diseases such as inflammatory bowel disease, obesity, metabolic syndrome and diabetes have been studied both in human and in animal, mainly rodent, models of disease. The intrinsic variation in microbiota composition, both within one host over time and within a group of similarly treated hosts, presents particular challenges in experimental design. This review highlights factors that need to be taken into consideration when designing animal trials to investigate the gastrointestinal tract microbiota in the context of inflammation studies. These include the origin and history of the animals, the husbandry of the animals before and during experiments, details of sampling, sample processing, sequence data acquisition and bioinformatic analysis. Because of the intrinsic variability in microbiota composition, it is likely that the number of animals required to allow meaningful statistical comparisons across groups will be higher than researchers have generally used for purely immune-based analyses

An insight into intestinal mucosal microbiota disruption after stroke

Recent work from our laboratory has provided evidence that indicates selective bacterial translocation from the host gut microbiota to peripheral tissues (i.e. lung) plays a key role in the development of post-stroke infections. Despite this, it is currently unknown whether mucosal bacteria that live on and interact closely with the host intestinal epithelium contribute in regulating bacterial translocation after stroke. Here, we found that the microbial communities within the mucosa of gastrointestinal tract (GIT) were significantly different between sham-operated and post-stroke mice at 24 h following surgery. The differences in microbiota composition were substantial in all sections of the GIT and were significant, even at the phylum level. The main characteristics of the stroke-induced shift in mucosal microbiota composition were an increased abundance of Akkermansia muciniphila and an excessive abundance of clostridial species. Furthermore, we analysed the predicted functional potential of the altered mucosal microbiota induced by stroke using PICRUSt and revealed significant increases in functions associated with infectious diseases, membrane transport and xenobiotic degradation. Our findings revealed stroke induces far-reaching and robust changes to the intestinal mucosal microbiota. A better understanding of the precise molecular events leading up to stroke-induced mucosal microbiota changes may represent novel therapy targets to improve patient outcomes. © 2018 The Author(s)

The time-course of broiler intestinal microbiota development after administration of cecal contents to incubating eggs

Background. The microbial populations that inhabit the gastrointestinal tract (GIT) are known to influence the health and growth performance of the host. Clean hatcheries and machine-based incubation practices in the commercial poultry industry can lead to the acquisition of aberrant microbiota in the GIT of chickens and a very high level of bird-to-bird variation. The lack of microbial profile flock uniformity presents challenges for harnessing and manipulating intestinal bacteria to better serve the host. Methods. Cecal contents from high or low performing chickens were used to inoculate the surface of eggs prior to hatching and then the initial gut colonisation was monitored and subsequent changes in gut microbiota composition were followed over time. Two different cecal treatment groups were compared to an untreated control group (n=32). Bacterial communities were characterised using high-throughput 16S rRNA gene sequencing techniques. Results. Cecal microbiota transfer via egg surface application did not transfer the performance profile of the donors to the recipient birds. One of the cecal inoculations provided a more uniform gut microbiota, but this was not reproduced in the second group with a different inoculum. Development of the intestinal community was reproducible in all three groups with some genera like Lactobacillus showing no change, others like Faecalibacterium increased in abundance slowly and steadily over time and others like Enterobacter were abundant only in the first days of life. Discussion. The cecal treatment reduced bird-to-bird variation in microbiota compo-sition. Although the high FCR performance of donor birds was not transferred with the cecal microbiota, all three groups, including the control, performed better than standard for the breed. The pattern of microbiota development was similar in all three flocks, indicating that the normal processes of microbiota acquisition largely swamped any effect of the cecal material applied to eggs. © 2017 Donaldson et al

Bacteria within the gastrointestinal tract microbiota correlated with improved growth and feed conversion: Challenges presented for the identification of performance enhancing probiotic bacteria

Identification of bacteria associated with desirable productivity outcomes in animals may offer a direct approach to the identification of probiotic bacteria for use in animal production. We performed three controlled chicken trials (n = 96) to investigate caecal microbiota differences between the best and poorest performing birds using four performance measures; feed conversion ratio (FCR), utilization of energy from the feed measured as apparent metabolisable energy, gain rate (GR), and amount of feed eaten (FE). The shifts in microbiota composition associated with the performance measures were very different between the three trials. Analysis of the caecal microbiota revealed that the high and low FCR birds had significant differences in the abundance of some bacteria as demonstrated by shifts in microbiota alpha and beta diversity. Trials 1 and 2 showed significant overall community shifts, however, the microbial changes driving the difference between good and poor performers were very different. Lachnospiraceae, Ruminococcaceae, and Erysipelotrichaceae families and genera Ruminococcus, Faecalibacterium and multiple lineages of genus Clostridium (from families Lachnospiraceae, Ruminococcaceae, and Erysipelotrichaceae) were highly abundant in good FCR birds in Trial 1. Different microbiota was associated with FCR in Trial 2; Catabacteriaceae and unknown Clostridiales family members were increased in good FCR and genera Clostridium (from family Clostridiaceae) and Lactobacillus were associated with poor FCR. Trial 3 had only mild microbiota differences associated with all four performance measures. Overall, the genus Lactobacillus was correlated with feed intake which resulted in poor FCR performance. The genus Faecalibacterium correlated with improved FCR, increased GR and reduced FE. There was overlap in phylotypes correlated with improved FCR and GR, while different microbial cohorts appeared to be correlated with FE. Even under controlled conditions different cohorts of birds developed distinctly different microbiotas. Within the different trial groups the abundance of certain bacterial groups correlated with productivity outcomes. However, with different underlying microbiotas there were different bacteria correlated with performance. The challenge will be to identify probiotic bacteria that can reliably deliver favorable outcomes from diverse microbiotas. © 2016 Stanley, Hughes, Geier and Moore

At-hatch administration of probiotic to chickens can introduce beneficial changes in gut microbiota

This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Recent advances in culture-free microbiological techniques bring new understanding of the role of intestinal microbiota in heath and performance. Intestinal microbial communities in chickens assume a near-stable state within the week which leaves a very small window for permanent microbiota remodelling. It is the first colonisers that determine the fate of microbial community in humans and birds alike, and after the microbiota has matured there are very small odds for permanent modification as stable community resists change. In this study we inoculated broiler chicks immediately post hatch, with 3 species of Lactobacillus, identified by sequencing of 16S rRNA and pheS genes as L. ingluviei, L. agilis and L. reuteri. The strains were isolated from the gut of healthy chickens as reproducibly persistent Lactobacillus strains among multiple flocks. Birds inoculated with the probiotic mix reached significantly higher weight by 28 days of age. Although each strain was able to colonise when administered alone, administering the probiotic mix at-hatch resulted in colonisation by only L. ingluviei. High initial abundance of L. ingluviei was slowly reducing, however, the effects of at-hatch administration of the Lactobacillus mix on modifying microbiota development and structure remained persistent. There was a tendency of promotion of beneficial and reduction in pathogenic taxa in the probiotic administered group. © 2018 Baldwin et al

The synthesis and characterisation of highly stable and reproducible selenium nanoparticles

This paper describes a simple and reproducible solution phase synthesis approach for selenium nanoparticles by reducing selenium tetrachloride in the presence of ascorbic acid. An optimization study with poly (sodium 4-styrenesulfonate) produced stable and spherical narrowly size distributed nanoparticles (46 nm) which are considered highly monodisperse. The presence of selenium nanoparticles was confirmed by UV-visible spectroscopy for surface plasmon resonance (262 nm), elemental dispersive spectroscopy (11 KeV and 12.5 KeV) and size ranges characterized by dynamic light scattering (PDI = 0.04, the ​size range of optimized nanoparticles = 35 nm to 75 nm), and visualized using scanning and transmission electron microscopy

Nanoparticles in feed: Progress and prospects in poultry research

© 2016 Elsevier LtdThe global poultry industry has greatly expanded due to an increase in demand for chicken meat and eggs. Growth of the industry was followed by growth in research which resulted in improved growth rate, feed efficiency, health status, and reduced carriage of pathogens. However, major research focus was improvement in productivity. It is possible to manipulate feed formulations to improve the feed conversion ratio (FCR), which results in a lower feed requirement to achieve market weight. Feed additives, containing vitamins and minerals, are commonly added to typical diets to support rapid growth and favourable FCR. Nanoparticles can be added to feed and provide an excellent platform to incorporate in various compounds, such as vaccines and nutrient supplements, due to large surface area to volume ratio and high absorption in the body. Nanoparticles can enable direct transportation of compounds to targeted organs or systems while avoiding fast degradability often seen with antibiotics and can encourage multiple health benefits. Silver, currently the most common nanoparticle investigated for use in chicken feed, has been shown to improve the microbiota of chickens. However, the positive results are tempered by the finding that silver nanoparticles have relatively high toxicity in birds. The question therefore arises as to whether other nanoparticle forms of essential metals and natural compounds can be safely delivered to provide positive impacts on health and productivity without the toxic side effects that can be seen with silver nanoparticles. Here, we review the current state of nanoparticle use as a poultry feed supplement - the successes and pitfalls of nano-feed as reported by researchers across the world

Understanding the mechanisms of zinc bacitracin and avilamycin on animal production: Linking gut microbiota and growth performance in chickens

© 2017, Springer-Verlag Berlin Heidelberg. Unravelling the mechanisms of how antibiotics influence growth performance through changes in gut microbiota can lead to the identification of highly productive microbiota in animal production. Here we investigated the effect of zinc bacitracin and avilamycin on growth performance and caecal microbiota in chickens and analysed associations between individual bacteria and growth performance. Two trials were undertaken; each used 96 individually caged 15-day-old Cobb broilers. Trial 1 had a control group (n = 48) and a zinc bacitracin (50 ppm) treatment group (n = 48). Trial 2 had a control group (n = 48) and an avilamycin (15 ppm) treatment group (n = 48). Chicken growth performance was evaluated over a 10-day period, and caecal microbiota was characterised by sequencing of bacterial 16S rRNA gene amplicons. Avilamycin produced no effect on growth performance and exhibited little significant disturbance of the microbiota structure. However, zinc bacitracin reduced the feed conversion ratio (FCR) in treated birds, changed the composition and increased the diversity of their caecal microbiota by reducing dominant species. Avilamycin only produced minor reductions in the abundance of two microbial taxa, whereas zinc bacitracin produced relatively large shifts in a number of taxa, primarily Lactobacillus species. Also, a number of phylotypes closely related to lactobacilli species were positively or negatively correlated with FCR values, suggesting contrasting effects of Lactobacillus spp. on chicken growth performance. By harnessing such bacteria, it may be possible to develop high-productivity strategies in poultry that rely on the use of probiotics and less on in-feed antibiotics

Sorghum and wheat differentially affect caecal microbiota and associated performance characteristics of meat chickens

This study compared the effects of wheat-and sorghum-based diets on broiler chickens. The growth performance and caecal microbial community of chickens were measured and correlations between productivity and specific gut microbes were observed. Cobb broilers 15 days of age were individually caged and two dietary treatments were used, one with a wheat-based diet (n D 48) and another one with a sorghum-based diet (n D 48). Growth performance measurements were taken over a 10 day period and samples for microbiota analysis were taken at the end of that period. Caecal microbiota was characterised by sequencing of 16S bacterial rRNA gene amplicons. Overall, the results indicated that a sorghum-based diet produced higher apparent metabolisable energy (AME) and body-weight gain (BWG) values in chickens, compared to a wheatbased diet. Nevertheless, sorghum-fed birds had higher feed conversion ratio (FCR) values than wheat-fed birds, possibly because of some anti-nutritional factors in sorghum. Further analyses showed that caecal microbial community was significantly associated with AME values, but microbiota composition differed between dietary treatments. A number of bacteria were individually correlated with growth performance measurements. Numerous OTUs assigned to strains of Lactobacillus crispatus and Lachnospiraceae, which were prevalent in sorghum-fed chickens, were correlated with high AME and BWG values, respectively. Additionally, a number of OTUs assigned to Clostridiales that were prevalent in wheat-fed chickens were correlated with low FCR values. Overall, these results suggest that between-diet variations in growth performance were partly associated with changes in the caecal microbiota. © 2017 Crisol-Martínez et al