The initial development of intestinal microbiota in poultry plays an important role in production performance, overall health and resistance against microbial infections. Multiplexed sequencing of 16S ribosomal RNA gene amplicons is often used in studies, such as feed intervention or antimicrobial drug trials, to determine corresponding effects on the composition of intestinal microbiota. However, considerable variation of intestinal microbiota composition has been observed both within and across studies. Such variation may in part be attributed to technical factors, such as sampling procedures, sample storage, DNA extraction, the choice of PCR primers and corresponding region to be sequenced, and the sequencing platforms used. Furthermore, part of this variation in microbiota composition may also be explained by different host characteristics and environmental factors. To facilitate the improvement of design, reproducibility and interpretation of poultry microbiota studies, we have reviewed the literature on confounding factors influencing the observed intestinal microbiota in chickens. First, it has been identified that host-related factors, such as age, sex, and breed, have a large effect on intestinal microbiota. The diversity of chicken intestinal microbiota tends to increase most during the first weeks of life, and corresponding colonization patterns seem to differ between layer- and meat-type chickens. Second, it has been found that environmental factors, such as biosecurity level, housing, litter, feed access and climate also have an effect on the composition of the intestinal microbiota. As microbiota studies have to deal with many of these unknown or hidden host and environmental variables, the choice of study designs can have a great impact on study outcomes and interpretation of the data. Providing details on a broad range of host and environmental factors in articles and sequence data repositories is highly recommended. This creates opportunities to combine data from different studies for meta-analysis, which will facilitate scientific breakthroughs toward nutritional and husbandry associated strategies to improve animal health and performance.
Keywords: 16S rRNA; confounding factors; gut health; gut microbiota; microbiome; poultry.