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Comparison of fecal and cecal microbiotas reveals qualitative similarities but quantitative differences.

Stanley D, Geier MS, Chen H, Hughes RJ, Moore RJ - BMC Microbiol. (2015)

Bottom Line: In contrast, fecal sampling does not require sacrifice and thus allows the same bird to be sampled repeatedly over time.It was found that 88.55% of all operational taxonomic units (OTUs), containing 99.25% of all sequences, were shared between the two sample types, with OTUs unique for each sample type found to be very rare.The microbial populations present within the paired ceca of individual birds were also compared and shown to be similar.

View Article: PubMed Central - PubMed

Affiliation: Central Queensland University, School of Medical and Applied Sciences, Bruce Highway, Rockhampton, QLD, 4702, Australia. d.stanley@cqu.edu.au.

ABSTRACT

Background: The majority of chicken microbiota studies have used the ceca as a sampling site due to the specific role of ceca in chicken productivity, health and wellbeing. However, sampling from ceca and other gastrointestinal tract sections requires the bird to be sacrificed. In contrast, fecal sampling does not require sacrifice and thus allows the same bird to be sampled repeatedly over time. This is a more meaningful and preferred way of sampling as the same animals can be monitored and tracked for temporal studies. The commonly used practice of selecting a subset of birds at each time-point for sacrifice and sampling introduces added variability due to the known animal to animal variation in microbiota.

Results: Cecal samples and fecal samples via cloacal swab were collected from 163 birds across 3 replicate trials. DNA was extracted and 16S rRNA gene sequences amplified and pyrosequenced to determine and compare the phylogenetic profile of the microbiota within each sample. The fecal and cecal samples were investigated to determine to what extent the microbiota found in fecal samples represented the microbiota of the ceca. It was found that 88.55% of all operational taxonomic units (OTUs), containing 99.25% of all sequences, were shared between the two sample types, with OTUs unique for each sample type found to be very rare. There was a positive correlation between cecal and fecal abundance in the shared sequences, however the two communities differed significantly in community structure, represented as either alpha or beta diversity. The microbial populations present within the paired ceca of individual birds were also compared and shown to be similar.

Conclusions: Fecal sample analysis captures a large percentage of the microbial diversity present in the ceca. However, the qualitative similarities in OTU presence are not a good representation of the proportions of OTUs within the microbiota from each sampling site. The fecal microbiota is qualitatively similar to cecal microbiota but quantitatively different. Fecal samples can be effectively used to detect some shifts and responses of cecal microbiota.

No MeSH data available.


Correlation plots for the different minimal cecal (A) and fecal (B) abundances. Note that each shared OTU was plotted for all 163 birds and may be highly abundant in many of the birds, thus total percentages are not expected to add up to 100. For example, there were 11 birds that had OTUs with cecal abundance higher than 50% vs 43 birds that contained fecal OTUs with abundance over 50%.
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Fig5: Correlation plots for the different minimal cecal (A) and fecal (B) abundances. Note that each shared OTU was plotted for all 163 birds and may be highly abundant in many of the birds, thus total percentages are not expected to add up to 100. For example, there were 11 birds that had OTUs with cecal abundance higher than 50% vs 43 birds that contained fecal OTUs with abundance over 50%.

Mentions: The main question we aimed to answer in this study was whether fecal samples could provide a reliable snapshot of cecal community structure. The high number of sequences and samples across the three trials provides sufficient statistical power to attempt to make these predictions. The three trials were inspected separately and as a whole set for correlations between fecal and cecal abundance of all shared OTUs present in both cecal and fecal samples across all of the birds. All 3 trials showed positive correlation between cecal and fecal abundances (Figure 4). To inspect if the level of correlation is influenced by abundance, i.e., if more abundant OTUs show better correlation, we inspected correlations for subsets of data with different cecal and separately fecal minimal abundance for all trials. Although we expected that more abundant OTUs would have higher cecal-fecal correlation, we found the opposite trend for both cecal and fecal abundances (Figures 4,5,6). Rare OTUs seem to be of similar low abundance in both cecal and fecal while higher abundance taxa tended to differ more. This is especially notable in the fecal dominant OTUs. OTUs comprising more than 50% of cecal sequences in one bird (Figure 5A) had a wide range of fecal abundances. The number of birds with cecal microbiota dominated by more than 50% was much lower than in fecal samples. Figure 6 simplifies the message from detailed Figure 5A and B; shared OTUs present in lower abundance correlated better, while correlation for more abundant OTUs was very low.Figure 4


Comparison of fecal and cecal microbiotas reveals qualitative similarities but quantitative differences.

Stanley D, Geier MS, Chen H, Hughes RJ, Moore RJ - BMC Microbiol. (2015)

Correlation plots for the different minimal cecal (A) and fecal (B) abundances. Note that each shared OTU was plotted for all 163 birds and may be highly abundant in many of the birds, thus total percentages are not expected to add up to 100. For example, there were 11 birds that had OTUs with cecal abundance higher than 50% vs 43 birds that contained fecal OTUs with abundance over 50%.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4403768&req=5

Fig5: Correlation plots for the different minimal cecal (A) and fecal (B) abundances. Note that each shared OTU was plotted for all 163 birds and may be highly abundant in many of the birds, thus total percentages are not expected to add up to 100. For example, there were 11 birds that had OTUs with cecal abundance higher than 50% vs 43 birds that contained fecal OTUs with abundance over 50%.
Mentions: The main question we aimed to answer in this study was whether fecal samples could provide a reliable snapshot of cecal community structure. The high number of sequences and samples across the three trials provides sufficient statistical power to attempt to make these predictions. The three trials were inspected separately and as a whole set for correlations between fecal and cecal abundance of all shared OTUs present in both cecal and fecal samples across all of the birds. All 3 trials showed positive correlation between cecal and fecal abundances (Figure 4). To inspect if the level of correlation is influenced by abundance, i.e., if more abundant OTUs show better correlation, we inspected correlations for subsets of data with different cecal and separately fecal minimal abundance for all trials. Although we expected that more abundant OTUs would have higher cecal-fecal correlation, we found the opposite trend for both cecal and fecal abundances (Figures 4,5,6). Rare OTUs seem to be of similar low abundance in both cecal and fecal while higher abundance taxa tended to differ more. This is especially notable in the fecal dominant OTUs. OTUs comprising more than 50% of cecal sequences in one bird (Figure 5A) had a wide range of fecal abundances. The number of birds with cecal microbiota dominated by more than 50% was much lower than in fecal samples. Figure 6 simplifies the message from detailed Figure 5A and B; shared OTUs present in lower abundance correlated better, while correlation for more abundant OTUs was very low.Figure 4

Bottom Line: In contrast, fecal sampling does not require sacrifice and thus allows the same bird to be sampled repeatedly over time.It was found that 88.55% of all operational taxonomic units (OTUs), containing 99.25% of all sequences, were shared between the two sample types, with OTUs unique for each sample type found to be very rare.The microbial populations present within the paired ceca of individual birds were also compared and shown to be similar.

View Article: PubMed Central - PubMed

Affiliation: Central Queensland University, School of Medical and Applied Sciences, Bruce Highway, Rockhampton, QLD, 4702, Australia. d.stanley@cqu.edu.au.

ABSTRACT

Background: The majority of chicken microbiota studies have used the ceca as a sampling site due to the specific role of ceca in chicken productivity, health and wellbeing. However, sampling from ceca and other gastrointestinal tract sections requires the bird to be sacrificed. In contrast, fecal sampling does not require sacrifice and thus allows the same bird to be sampled repeatedly over time. This is a more meaningful and preferred way of sampling as the same animals can be monitored and tracked for temporal studies. The commonly used practice of selecting a subset of birds at each time-point for sacrifice and sampling introduces added variability due to the known animal to animal variation in microbiota.

Results: Cecal samples and fecal samples via cloacal swab were collected from 163 birds across 3 replicate trials. DNA was extracted and 16S rRNA gene sequences amplified and pyrosequenced to determine and compare the phylogenetic profile of the microbiota within each sample. The fecal and cecal samples were investigated to determine to what extent the microbiota found in fecal samples represented the microbiota of the ceca. It was found that 88.55% of all operational taxonomic units (OTUs), containing 99.25% of all sequences, were shared between the two sample types, with OTUs unique for each sample type found to be very rare. There was a positive correlation between cecal and fecal abundance in the shared sequences, however the two communities differed significantly in community structure, represented as either alpha or beta diversity. The microbial populations present within the paired ceca of individual birds were also compared and shown to be similar.

Conclusions: Fecal sample analysis captures a large percentage of the microbial diversity present in the ceca. However, the qualitative similarities in OTU presence are not a good representation of the proportions of OTUs within the microbiota from each sampling site. The fecal microbiota is qualitatively similar to cecal microbiota but quantitatively different. Fecal samples can be effectively used to detect some shifts and responses of cecal microbiota.

No MeSH data available.