Succession of the turkey gastrointestinal bacterial microbiome related to weight gain

Abstract

Because of concerns related to the use of antibiotics in animal agriculture, antibiotic-free alternatives are greatly needed to prevent disease and promote animal growth. One of the current challenges facing commercial turkey production in Minnesota is difficulty obtaining flock average weights typical of the industry standard, and this condition has been coined "Light Turkey Syndrome" or LTS. This condition has been identified in Minnesota turkey flocks for at least five years, and it has been observed that average flock body weights never approach their genetic potential. However, a single causative agent responsible for these weight reductions has not been identified despite numerous efforts to do so. The purpose of this study was to identify the bacterial community composition within the small intestines of heavy and light turkey flocks using 16S rRNA sequencing, and to identify possible correlations between microbiome and average flock weight. This study also sought to define the temporal succession of bacteria occurring in the turkey ileum. Based upon 2.7 million sequences across nine different turkey flocks, dominant operational taxonomic units (OTUs) were identified and compared between the flocks studied. OTUs that were associated with heavier weight flocks included those with similarity to Candidatus division Arthromitus and Clostridium bartlettii, while these flocks had decreased counts of several Lactobacillus species compared to lighter weight flocks. The core bacterial microbiome succession in commercial turkeys was also defined. Several defining markers of microbiome succession were identified, including the presence or abundance of Candidatus division Arthromitus, Lactobacillus aviarius, Lactobacillus ingluviei, Lactobacillus salivarius, and Clostridium bartlettii. Overall, the succession of the ileum bacterial microbiome in commercial turkeys proceeds in a predictable manner. Efforts to prevent disease and promote growth in the absence of antibiotics could involve target dominant bacteria identified in the turkey ileum that are associated with increased weight gain.

Keywords: 16S; Bacteria; Candidatus Arthromitus; Gastrointestinal; Lactobacillus; Microbiome; Poultry; Segmented filamentous bacteria; Succession; Turkeys.

Figure 1. Average flock weights for Light versus Heavy flocks in experiment #1.

To calculate the average sample flock weight, 40 birds from each flock were weighed or an average weight from the barn scale readings was used. Dashed line indicates genetic potential of bird type used.

Figure 2. Taxonomic classification of groups in this study.

Class-level taxonomic classification was based upon average proportional abundance of normalized samples. Designations in the X-axis are “CF”, individual birds from commercial flock (experiment #2); “RF”, individual birds from research flock (experiment #2); “Heavy”, pooled samples categorized as heavy weights (experiment #1); and “Light”, pooled samples categorized as lighter weights (experiment #1). “W”, age of birds in weeks.

Figure 3. Cladogram of operational taxonomic units (OTUs).

Cladogram illustrating relationships of OTUs in the entire study, classified using the Ribosomal Database Project and/or BLAST. The outer ring depicts relative log₁₀ abundances of OTUs in the entire dataset. Figure was generated using the Interactive Tree of Life (iTOL).

Figure 4. Dendrogram of operational taxonomic units (OTUs) with similarity to Lactobacillus.

Phylogenetic relationships were inferred using Maximum Likelihood analysis with a General Time Reversible Model using 1,000 bootstrap replicates in MEGA5. The dataset was generated from representative OTUs with similarity to Lactobacillus spp., and extracted Lactobacillus sequences from the NCBI database for the V3 hypervariable region of the 16S rRNA region.

Figure 5. Light microscopy of segmented filamentous bacteria in the turkey ileum.

Scaled line is approximately 10 microns.

Figure 6. Heatmap of average OTU abundance for each group studied.

Averages for multiple birds at each group and timepoint are depicted for the top 50 OTUs in this study. Heatmap is in log₁₀ normalized counts.

Figure 7. Heatmap of OTU abundance in individual birds.

Normalized counts for individual birds within each group and timepoint are depicted for the top 50 OTUs in this study. Heatmap is in log₁₀ normalized counts.

Figure 8. Principal coordinate analysis (PCoA) of samples of differing group and timepoint.

Dashed line indicates the approximate movement of turkeys from brooder to grow-out barns. Color is based on age of turkeys in weeks.

Figure 9. Cladogram depicting relationships between groups studied.

Cladogram was generated using two-way hierarchical clustering and Neighbor-Joining algorithm. Groups are designated as “CF”, commercial flock (experiment #2); “RF”, research flock (experiment #2); “Heavy”, flocks of heavier weights (experiment #1); and “Light”, flocks of lighter weights (experiment #1).

Figure 10. Temporal succession of bacteria in the turkey ileum.

Model is based upon OTUs identified as core in the dataset at each timepoint. Red-colored groups are present in weeks 1–2 and remain in the ileum through 7 weeks, green-colored groups emerge at 3–4 weeks, and blue-colored groups emerge at 5–7 weeks.