Evaluation of the liver abscess microbiome and liver abscess prevalence in cattle reared for production of natural branded beef

Transcription

1 Evaluation of the liver abscess microbiome and liver abscess prevalence in cattle reared for production of natural branded beef K.L. Huebner, J.N. Martin C.J. Weissend, K.L. Holzer, M. Weinroth, Z. Abdo, J.L. Metcalf, I. Geornaras, J.K. Parker, P.S. Morley, K.E Belk

2 Background Liver abscesses are primary cause of liver condemnation at slaughter Abscess rate highly variable Tylosin fed to prevent abscesses at low inclusion dosages Traditional culture techniques: Polymicrobial, gram negative anaerobe Fusobacterium necrophorum, Trueperella pyogenes Salmonella spp. recently isolated

3 Liver Abscesses in Beef Cattle Severe abscesses have greatest effect on losses 2 Pathogenesis 2

4 Estimated: 108 bacteria per gram of soil of different species Only ~1% culturable 6 Amplification and sequencing of conserved regions of the 16S rrna gene enables genotyping of bacteria without having to culture them Limits organisms surveyed to bacteria and archaea Offers increased sampling of bacterial diversity per $ spent Appropriate if interested in bacterial diversity / ecology 16S rrna Sequencing

5 Objectives The objectives of this study were to evaluate the flora of liver abscesses in naturally raised finishing beef steers using 16S ribosomal RNA marker genes to define operational taxonomic units (OTUs), and secondly to compare these results to traditional cultural findings.

6 Materials and Methods Colorado beef steers Certified natural beef products (no antimicrobials or growth hormones) Liver abscess rates and grade recorded for 4,323 individual animals as part of another study. 33 liver abscess samples total collected at harvest representing 14 pens, 7 pen blocks of 2 pens each adjacent to each other (shared border) One to five liver abscesses per pen Bacterial community (microbiome) characterized by 16S rrna gene amplification and next generation sequencing

7 Liver abscess scores collected on individual animals at slaughter 5 Sterile extraction of purulent material Storage at 80 Celsius 2 pens made up one pen block. 7 pen blocks were sampled from a Colorado feedlot raised for natural branded beef products Sequence Data Bioinformatic analysis Trimming and quality filtering Open Reference OTU picking Sequence clustering (UCLUST) Taxonomy assignment (Greengenes) database Statistical analysis DNA Isolation DNA Quantification/Quality Control 16S Library Preparation 3 Illumina sequencing by synthesis 4

8 Results

9 Phylum level taxonomic summary sorted by adjacent pen blocks (PB) of cattle. 100% PB1 PB2 PB3 PB4 PB6 PB7 PB9 80% 60% Each pen block represents two pens of cattle housed adjacent to each other in the feedlot during the entire finishing period. 40% 20% 0% k Bacteria;p Tenericutes k Archaea;p Crenarchaeota k Bacteria;p Armatimonadetes k Bacteria;p Planctomycetes k Bacteria;p Nitrospirae k Bacteria;p Gemmatimonadetes

10 PCoA Plot based on unweighted Unifrac distances for bacteria of liver abscess microbiome plotted on total liver abscess rate gradient. Each pen block represents two pens of cattle housed adjacent to each other in the feedlot during the entire finishing period.

11 Summary Liver abscess prevalence 38.5% (95% CI ). Taxa phyla Fusobacterium, Bacteroides, and unassigned taxa were most represented. Actinobacteria (including Trueperella spp.) had lower relative abundance in the samples. There was a correlation between abscess rate and pen block microbiome, with shifts in the microbial community of adjacent pens accounting for 55 % of variation in the PCoA plot. OTU differences in the genus, Fusobacterium, are driving some of the differences in clustering by abscess rate and pen block.

12 Future Directions Liver abscesses are made up of polymicrobial communities 16s rrna analysis is an effective methodology that corroborates culture findings. Next generation sequencing methods are effective at capturing bacterial diversity and ecology in liver abscess samples of beef cattle. Future work will be directed at understanding how the liver abscess microbiome is related to changes in feedlot management, including diet, feed additives, antimicrobial use, and geographical differences in the soil microbiome.

13 References 1.Amachawadi & Nagaraja. Amer. Soc. Of Anim. Sci : Nagaraja and Lechtenberg. Vet Clin Food Anim Novogene Illumina Elanco Animal Health. Liver Check Service. 6. Handelsman et al. Chem & Biol. 1998

14 Acknowledgements Funding for this project was provided by the National Cattlemen s Beef Association. Thank you to our collaborators from JBS, Diamond V, West Texas A&M University, Iowa State University, and Diamond T. Also thank you to the CSU Animal Science graduate students.