![]() ![]() Overall, these results indicate that while stool and biopsy samples reveal different microbial communities in both membership and diversity, rectal swabs may provide a viable easy-to-collect alternative to stool while keeping in mind that the proportions of aerobic genera will be elevated.Īnother body site of interest that has been studied comprehensively is the human oral microbiome. However, it should be noted that they did find rectal swab microbial profiles, matched stool profiles closer than biopsies. Furthermore, they found numerous differences in levels 1 and 2 KEGG pathways through metagenomic shotgun sequencing (MGS) analysis. They found that 24 of 48 families were significantly different in relative abundance between swab and stool samples through 16S rRNA gene sequencing, and a higher proportion of aerobic bacteria could be found in rectal swabs. reported significant differences in both taxonomic makeup and functional pathway abundances between rectal swab and stool samples. has shown similar results however, a recent study by Jones et al. compared stool samples and rectal swabs from the same patients and found no significant difference in microbial abundances between these collection methods. In contrast, multiple studies have compared the observed microbial communities from rectal swabs and fecal samples and have found similar profiles. For example, biopsies often have higher levels of human DNA and do not account for impacts on community composition during the transit of stool in the colon. This is not surprising given the drastically different collection methods. In both cases, it was found that collection of samples by biopsy introduced strong biases toward the identification of specific microbes such as those that adhere to the mucosal wall of the colon. Work on the gut microbiome has compared colon biopsy samples to both stool and rectal swab samples. This has been highlighted in studies of the various areas of the human microbiome. The method of sample collection is particularly important to consider as different types of systematic bias can be introduced depending on the sampling environment. All these details should be noted during sample collection and accounted for during downstream analysis to ensure consistent results across studies. The way in which the sample is collected, how long the sample is stored, and what the sample is stored in can all impact the underlying observed microbial composition. Sample collection and storage are a vital part of an experiment and if not carefully planned can often introduce unaccounted bias. Throughout each section, we will highlight the various biases that are introduced during each step, and we will then review current approaches that have been taken to help alleviate those biases to improve our understanding of the human microbiome (Table 1). In this report, we will review each step in a typical marker gene and metagenomic shotgun sequencing microbiome study, starting with sample collection and ending with downstream bioinformatic analysis (Fig. While sometimes these biases can be subtle, they often result in significant impacts on biological conclusions. The introduction of these biases often results in distorted observations of the true underlying microbial composition contained within a sample. ![]() Throughout a typical microbiome study, there are numerous areas where biases are introduced. Microbiome studies are particularly at fault for elevated levels of these biases due to the high sensitivity of DNA sequencing instruments and the relatively unknown underlying microbial compositions within a sample. One possible reason for inconsistent results across studies is the high level of random and systemic bias that is introduced throughout sequenced-based human microbiome studies. However, results from different studies often do not match with previous findings. Investigation into these microbes has led to the development of both novel therapeutics and diagnostic tools. These communities and their cellular functions are known as the human microbiome, which has now been associated with multiple different influences on human health. Recent advances in DNA sequencing technology have allowed community-wide investigation into the microbes that live on and within the human body. ![]()
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