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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.


Alpha diversity is significantly different between fecal and cecal microbiota. Alpha diversity indicators of cecal (red) and fecal (blue) samples all showed significant (p < 1e−5) difference based on 1e5 Monte Carlo permutations. The indices plotted in the four panels are; A, Chao 1; B, Simpson; C, Shannon; and D, Equitability.
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Fig1: Alpha diversity is significantly different between fecal and cecal microbiota. Alpha diversity indicators of cecal (red) and fecal (blue) samples all showed significant (p < 1e−5) difference based on 1e5 Monte Carlo permutations. The indices plotted in the four panels are; A, Chao 1; B, Simpson; C, Shannon; and D, Equitability.

Mentions: A number of alpha diversity measures were inspected to compare within sample diversity of cecal and fecal samples. All of the diversity indicators showed statistically significant differences, all with P < 1e−5, the lowest possible p-value based on the 1e5 Monte Carlo permutations used (Qiime). Both non-phylogenetic estimator Chao1 (Figure 1A) and phylogenetic diversity (PD) indicator whole tree analysis showed higher diversity in cecal samples relative to fecal samples. Cecal richness, based on number of observed species, was significantly higher than richness in fecal communities. The singles and doubles estimator suggested more rare OTUs in cecal samples. Simpson’s evenness (Figure 1B) and Strong’s dominance index confirmed that fecal samples have more dominant OTUs. Shannon entropy (Figure 1C) and the Equitability indices (Figure 1D) showed cecal phylotypes as more evenly distributed within samples.Figure 1


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)

Alpha diversity is significantly different between fecal and cecal microbiota. Alpha diversity indicators of cecal (red) and fecal (blue) samples all showed significant (p < 1e−5) difference based on 1e5 Monte Carlo permutations. The indices plotted in the four panels are; A, Chao 1; B, Simpson; C, Shannon; and D, Equitability.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig1: Alpha diversity is significantly different between fecal and cecal microbiota. Alpha diversity indicators of cecal (red) and fecal (blue) samples all showed significant (p < 1e−5) difference based on 1e5 Monte Carlo permutations. The indices plotted in the four panels are; A, Chao 1; B, Simpson; C, Shannon; and D, Equitability.
Mentions: A number of alpha diversity measures were inspected to compare within sample diversity of cecal and fecal samples. All of the diversity indicators showed statistically significant differences, all with P < 1e−5, the lowest possible p-value based on the 1e5 Monte Carlo permutations used (Qiime). Both non-phylogenetic estimator Chao1 (Figure 1A) and phylogenetic diversity (PD) indicator whole tree analysis showed higher diversity in cecal samples relative to fecal samples. Cecal richness, based on number of observed species, was significantly higher than richness in fecal communities. The singles and doubles estimator suggested more rare OTUs in cecal samples. Simpson’s evenness (Figure 1B) and Strong’s dominance index confirmed that fecal samples have more dominant OTUs. Shannon entropy (Figure 1C) and the Equitability indices (Figure 1D) showed cecal phylotypes as more evenly distributed within samples.Figure 1

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.