Cloacal Swabs Are Unreliable Sources for Estimating Lower Gastro-Intestinal Tract Microbiota Membership and Structure in Broiler Chickens.

Travis Williams,Giridhar Athrey

doi:10.3390/microorganisms8050718

Abstract

The gastrointestinal microbiota of chickens plays a central role in health and performance. Cloacal swabs, due to their proximity to the ceca (a vital site of functional activity), are an alternative, non-invasive method used for assaying microbial communities and might be a viable option for longitudinal studies. In this study, the microbiota of twenty paired cecal content and cloacal swab samples representing two dietary treatments was assessed using 16S rRNA V4 hypervariable region sequencing and was analyzed using the MOTHUR pipeline, Phyloseq, and Vegan packages. Paired t-test and Wilcoxon signed-rank tests showed significant differences in the Chao1 index (p-value <0.0001 and p-value <0.0001, respectively) but not in the Inverse Simpson species diversity estimator (p-value = 0.06763 and p-value = 0.06021, respectively) between the cecal content and cloacal swabs. β-diversity between the cloacal swabs and cecal microbiota also showed significant differences using PERMANOVA, HOMOVA, and weighted UniFrac testing (p-values < 0.001). Based on a paired sample analysis, this study provided evidence of the high inter-individual variation and randomness of cloacal microbiota, in contrast to cecal microbiota. Our findings indicated that cloacal swabs do not approximate the α or β diversity of cecal samples and are not suitable for longitudinal studies of gut microbiota.

Highlights

Chicken (Gallus gallus domesticus) is the source of the most consumed animal protein globally, at nearly twice the amount of pork and beef combined [1]
The cecal samples had an average of 22,656 reads (IQR 20,312–24,515 reads), whereas the cloacal swabs had an average of 13,900 reads (IQR 6902–20,366 reads)
We showed that cloacal swabs do not faithfully approximate either the α and β diversity of cecal samples, based on the paired samples

Summary

Introduction

Chicken (Gallus gallus domesticus) is the source of the most consumed animal protein globally, at nearly twice the amount of pork and beef combined [1]. The role of gut microbiota in improving performance [6,7,8], welfare [9], and health [4,10,11,12,13,14], is a topic of intense interest. The gut microbiota is studied intensively in chicken; an NCBI PubMed Central search for “Poultry Gut Microbiota” yielded 2586 research articles within the last five years. The ceca, a pair of blind sacs, are especially important as the site of functional activity relevant to microbial communities and species studied in performance and health [7,17]. A standard experimental method of microbiota analysis in poultry involves the invasive sampling of the ceca, following euthanasia, which prevents longitudinal studies of the same experimental animals

Methods

Results

Discussion

Conclusion