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. 2021 Mar 31;9(4):721.
doi: 10.3390/microorganisms9040721.

Metagenomic Analysis of the Respiratory Microbiome of a Broiler Flock from Hatching to Processing

Kelly A Mulholland  1 , Monique G Robinson  1 , Sharon J Keeler  1 , Timothy J Johnson  2 , Bonnie W Weber  2 , Calvin L Keeler Jr  1
Affiliations

Metagenomic Analysis of the Respiratory Microbiome of a Broiler Flock from Hatching to Processing

Kelly A Mulholland et al. Microorganisms. .

Abstract

Elucidating the complex microbial interactions in biological environments requires the identification and characterization of not only the bacterial component but also the eukaryotic viruses, bacteriophage, and fungi. In a proof of concept experiment, next generation sequencing approaches, accompanied by the development of novel computational and bioinformatics tools, were utilized to examine the evolution of the microbial ecology of the avian trachea during the growth of a healthy commercial broiler flock. The flock was sampled weekly, beginning at placement and concluding at 49 days, the day before processing. Metagenomic sequencing of DNA and RNA was utilized to examine the bacteria, virus, bacteriophage, and fungal components during flock growth. The utility of using a metagenomic approach to study the avian respiratory virome was confirmed by detecting the dysbiosis in the avian respiratory virome of broiler chickens diagnosed with infection with infectious laryngotracheitis virus. This study provides the first comprehensive analysis of the ecology of the avian respiratory microbiome and demonstrates the feasibility for the use of this approach in future investigations of avian respiratory diseases.

Keywords: avian; bioinformatics; chicken; microbiome; respiratory; virome.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript. Or in the decision to publish the results.

Figures

Figure 1
Figure 1
Normalized relative abundance of detected DNA (A) and RNA (B) viral species at each time point. * No DNA viruses detected at Week 1.
Figure 2
Figure 2
Heat map with phylogenetic tree representing the detection frequency of eukaryotic viral families at each individual week. Color corresponds to the range of relative abundance of each week from 0 to 100%. The sum of each column, or week, is 100%.
Figure 3
Figure 3
Heat map with phylogenetic tree representing the detection intensity of each eukaryotic viral family from hatching to processing. Color intensity corresponds to the range of relative abundance of each family from 0 to 100%. The sum of each row, or viral family, is 100%.
Figure 4
Figure 4
Abundance of (A) virus, (B) bacteria, (C) bacteriophage and (D) fungi at Week 0, Week 1 and Week 7. Taxa represented at family (A,C) and phylum (B,D) level.
Figure 4
Figure 4
Abundance of (A) virus, (B) bacteria, (C) bacteriophage and (D) fungi at Week 0, Week 1 and Week 7. Taxa represented at family (A,C) and phylum (B,D) level.
Figure 5
Figure 5
Phylogenetic tree of (A) virus, (B) bacteria, (C) bacteriophage and (D) yeast and fungi. Node diameter indicates average abundance at species (A,C,D) and genera (B) level. Taxonomic levels range from phylum to genera (B), order to species (C) phylum to species (D). Viruses (A) are organized according to structural classification.
Figure 6
Figure 6
Microbial network of the complete healthy avian respiratory microbiome including detected RNA viruses, DNA viruses, yeast and fungi, bacteria, and bacteriophage. Taxa nodes are arranged by order. Node diameter correlates to taxa frequency.
Figure 7
Figure 7
Correlation matrix comparing bacteria and bacteriophage taxa at the family level. Node diameter corresponds to level of correlation. Node color corresponds to the Pearson correlation coefficient and ranges from -1 to 1 indicated by red and blue, respectively.
Figure 8
Figure 8
Changes in the eukaryotic viral microbiome associated with avian respiratory disease.
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