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We investigated lameness outbreaks at 3 commercial broiler farms in Arkansas. We isolated several distinct bacterial species from Bacterial Chondronecrosis with Osteomyelitis (BCO) lesions from these 3 farms. The results show that BCO-lameness pathogens on particular farms can differ significantly. We characterized genomes for isolates of the 2 most prevalent species, Escherichia coli and Staphylococcus aureus. Genomes assembled for E. coli isolates from all 3 farms were quite different between farms, and most similar to genomes from different geographical locations and hosts. The E. coli phylogenomics suggests frequent host shifts for this species. Genomes for S. aureus isolates from one farm were highly related to those from chicken isolates from Europe. Highly related isolates have also been characterized from chickens in the Arkansas area for at least a decade. Phylogenomics suggest that this S. aureus has been restricted to poultry for more than 40 y. Detailed analysis of genomes from 2 neighboring clades of S. aureus human and chicken isolates, identifies the acquisition of a specific pathogenicity island in the transition from human to chicken pathogen and that pathogenesis for this clade in chickens may depend on this mobile element. Investigation of the evolution of this chicken-restricted clade from 1980 in Ireland, Poland in 2008, Oklahoma in 2010 and Arkansas in 2019, reveals the acquisition of additional virulence determinants including pathogenicity islands. Isolate-specific genome characterizations will help further our understanding of the disease mechanisms of BCO-lameness, a significant animal welfare issue., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

This report demonstrates that high levels of lameness can be induced by a limited bacterial challenge in drinking water for birds raised on litter flooring, comparable with lameness induced by the gold standard for inducing lameness, growth on suspended wire flooring. The bacterium used in the challenge was cultured from lesions in birds induced for bacterial chondronecrosis with osteomyelitis (BCO) in the wire-flooring model so the epidemiology appears similar. The litter-flooring model could better approximate broiler operations. Furthermore, the work demonstrates that 2 commercial probiotics (GalliProTect and GalliProMax) can reduce lameness in the bacterial challenge litter-flooring model. Lameness attributable to BCO is one of the most significant animal welfare issues for broiler production. The wire-flooring and litter-flooring models afford alternatives for understanding the etiology, and epidemiology of BCO, and development of management strategies to reduce lameness. Probiotics afford a promising management strategy. The results suggest that the probiotic protection may extend beyond just intestinal health and intestinal barrier function., (Copyright © 2020. Published by Elsevier Inc.)

Staphylococcus agnetis has been previously associated with subclinical or clinically mild cases of mastitis in dairy cattle and is one of several staphylococcal species that have been isolated from the bones and blood of lame broilers. We reported that S. agnetis could be obtained frequently from bacterial chondronecrosis with osteomyelitis (BCO) lesions of lame broilers (A. Al-Rubaye et al., PLoS One 10:e0143336, 2015 [https://doi.org/10.1371/journal.pone.0143336]). A particular isolate, S. agnetis 908, can induce lameness in over 50% of exposed chickens, exceeding normal BCO incidences in broiler operations. We reported the assembly and annotation of the genome of isolate 908. To better understand the relationship between dairy cattle and broiler isolates, we assembled 11 additional genomes for S. agnetis isolates, an additional chicken BCO strain, and ten isolates from cattle milk, mammary gland secretions, or udder skin from the collection at the University of Missouri. To trace phylogenetic relationships, we constructed phylogenetic trees based on multilocus sequence typing and genome-to-genome distance comparisons. Chicken isolate 908 clustered with two of the cattle isolates, along with three isolates from chickens in Denmark and an isolate of S. agnetis we isolated from a BCO lesion on a commercial broiler farm in Arkansas. We used a number of BLAST tools to compare the chicken isolates to those from cattle and identified 98 coding sequences distinguishing isolate 908 from the cattle isolates. None of the identified genes explain the differences in host or tissue tropism. These analyses are critical to understanding how staphylococci colonize and infect different hosts and potentially how they can transition to alternative niches (bone versus dermis). IMPORTANCE has been recently recognized as associated with disease in dairy cattle and meat-type chickens. The infections appear to be limited in cattle and systemic in broilers. This report details the molecular relationships between cattle and chicken isolates in order to understand how this recently recognized species infects different hosts with different disease manifestations. The data show that the chicken and cattle isolates are very closely related, but the chicken isolates all cluster together, suggesting a single jump from cattle to chickens.Staphylococcus agnetis has been recently recognized as associated with disease in dairy cattle and meat-type chickens. The infections appear to be limited in cattle and systemic in broilers. This report details the molecular relationships between cattle and chicken isolates in order to understand how this recently recognized species infects different hosts with different disease manifestations. The data show that the chicken and cattle isolates are very closely related, but the chicken isolates all cluster together, suggesting a single jump from cattle to chickens., (Copyright © 2020 American Society for Microbiology.)

Background: Ascites syndrome is a hypertensive, multifactorial, multigene trait affecting meat-type chickens imposing significant economic losses on the broiler industry. A region containing the CPQ gene has been previously identified as significantly affecting ascites phenotype. The region was discovered through whole genome resequencing focused on chicken chromosome 2. The association was confirmed through further genotyping in multiple broiler populations., Results: The whole genome resequencing analyses have now been extended to the current chicken genome assembly. DNA samples were pooled according to gender and phenotype and the pools subjected to next generation sequencing. Loci were identified as clusters of single nucleotide polymorphisms where frequencies of the polymorphisms differed between resistant and susceptible chickens. The chickens are an unselected line descended from a commercial elite broiler line. Regions identified were specific to one or both genders. The data identify a total of 28 regions as potential quantitative trait loci for ascites. The genes from these regions have been associated with hypertensive-related traits in human association studies. One region on chicken chromosome 28 contains the LRRTM4 gene. Additional genotyping for the LRRTM4 region demonstrates an epistatic interaction with the CPQ region for ascites phenotype., Conclusions: The 28 regions identified were not previously identified in a multi-generational genome wide association study using 60k Single Nucleotide Polymorphism panels. This work demonstrates the utility of whole genome resequencing as a cost effective, direct, and efficient method for identifying specific gene regions affecting complex traits. The approach is applicable to any organism with a genome assembly and requires no a priori assumptions.

A quantitative trait locus on chromosome 9 was previously shown to be associated with ascites in multiple experimental and commercial populations. A study to evaluate the association of the QTL, based on variable number tandem repeat genotypes, with economically important traits was carried out on a commercial male elite line. Results indicated the highest fat and the lowest fillet mean were associated with the most resistant ascites genotype. All other traits measured for this genotype showed no trend towards positive or negatively impacting production values. The results suggest that a balanced approach could be undertaken in commercial broiler breeding operations to reduce ascites susceptibility in broiler populations without compromising overall genetic progress for traits of economic importance., (© 2017 Poultry Science Association Inc.)

Previously, we reported a genome wide association study (GWAS) that had shown association of a region between 11.8 and 13.6 Mbp on chromosome 9 with ascites phenotype in broilers. We had used microsatellite loci to demonstrate an association of particular genotypes for this region with ascites in experimental ascites lines and commercial broiler breeder lines. We identified two potential candidate genes, AGTR1 and UTS2D, within that chromosomal region for mediating the quantitative effect. We have now extended our analysis using SNPs for these genes to assess association with resistance or susceptibility to ascites in these same broiler lines. Surprisingly, in contrast to our previous GWAS and microsatellite data for this region, we find no association of the SNP genotypes or haplotypes in the region suggesting that the two genes might have limited association with the disease phenotype., (© 2016 Poultry Science Association Inc.)

Lameness in broiler chickens is a significant animal welfare and financial issue. Bacterial chondronecrosis with osteomyelitis (BCO) leading to lameness can be enhanced by rearing young broilers on wire flooring. Using the wire floor system, we identified Staphylococcus agnetis as the predominant isolate in BCO of the proximal tibiae and femora, and blood of lame broilers. Administration of S. agnetis isolates in water can induce lameness. We now report that the wire floor system increases bacterial translocation into the blood stream. We have also determined that approximately 10 5 CFU/mL is the minimum effective dose in the drinking water and that challenge at 10, 20, or 30 days of age produces similar incidences of lameness. BCO isolates of S. agnetis are much more effective than other Staphylococcus species and can overwhelm the protective effects of some commercial probiotics. Finally, we also demonstrated that the BCO lameness induced by administration of S. agnetis in the drinking water is transmissible to unchallenged broilers in the same pen., (© 2016 Poultry Science Association Inc.)

Bacterial chondronecrosis with osteomyelitis (BCO) is a common cause of lameness in commercial broiler chickens worldwide. BCO represents substantial production loss and welfare issues of chickens. The bacterial species or communities underlying BCO pathogenesis still remain to be fully characterized. To gain insights on blood microbiota in broilers and its potential association with BCO, blood samples collected from healthy (n = 240) and lame (n = 12) chickens were analyzed by deep sequencing of 16S RNA genes. The chicken blood microbiota were dominated by Proteobacteria (60.58% ± 0.65) followed by Bactroidetes (13.99% ± 0.29), Firmicutes (11.45% ± 0.51), Actinobacteria (10.21% ± 0.37) and Cyanobacteria (1.96% ± 0.21) that constituted 98.18% (± 0.22) of the whole phyla. The bacterial communities consist of 30-40 OTUs in the blood of broiler chickens, regardless of ages and other environmental or host conditions, and the blood microbiomes of BCO chickens were largely distinct from those of healthy chickens. In addition, Linear discriminant analysis (LDA) effect size (LEfSe) method revealed that Staphylococcus, Granulicatella, and Microbacterium were significantly enriched in BCO chickens as compared to healthy chickens. The results from this study have significant implications in understanding blood microbiota present in broiler chickens and its potential role in BCO pathogenesis.

Lameness in broiler chickens is a significant animal welfare and financial issue. Lameness can be enhanced by rearing young broilers on wire flooring. We have identified Staphylococcus agnetis as significantly involved in bacterial chondronecrosis with osteomyelitis (BCO) in proximal tibia and femorae, leading to lameness in broiler chickens in the wire floor system. Administration of S. agnetis in water induces lameness. Previously reported in some cases of cattle mastitis, this is the first report of this poorly described pathogen in chickens. We used long and short read next generation sequencing to assemble single finished contigs for the genome and a large plasmid from the chicken pathogen. Comparison of the S. agnetis genome to those of other pathogenic Staphylococci shows that S.agnetis contains a distinct repertoire of virulence determinants. Additionally, the S. agnetis genome has several regions that differ substantially from the genomes of other pathogenic Staphylococci. Comparison of our finished genome to a recent draft genome for a cattle mastitis isolate suggests that future investigations focus on the evolutionary epidemiology of this emerging pathogen of domestic animals.

A genome-wide SNP survey was used to identify chromosomal regions that showed linkage disequilibrium with respect to ascites susceptibility and ventricular hypertrophy in an F2 cross between previously described ascites-resistant and -susceptible lines. Variable number tandem repeats were used to obtain genotype data to further characterize these regions. A region on chromosome 9 (12 to 13 Mbp in 2011 assembly) shows association with ascites in the ascites lines and in several commercial broiler breeder lines with a significant sex effect. There are 2 candidate genes, AGTR1 (an angiotensin II type 1 receptor) and UTS2D (urotensin 2 domain containing), in this region that have been associated with hypertension and hypoxic response in mammals.

Staphylococcus agnetis is an emerging pathogen in chickens but has been most commonly isolated from sub-clinical mastitis in bovines. Previous whole-genome analyses for known virulence genes failed to identify determinants for the switch from mild ductal infections in cattle to severe infections in poultry. We now report identification of a family of 15 kbp, 17–19 gene mobile genetic elements (MGEs) specific to chicken osteomyelitis and dermatitis isolates of S. agnetis. These MGEs can be present in multiple copies per genome. The MGE has been vectored on a Staphylococcus phage that separately lysogenized two S. agnetis osteomyelitis strains. The S. agnetis genome from a broiler breeder case of ulcerative dermatitis contains 2 orthologs of this MGE, not associated with a prophage. BLASTn and phylogenetic analyses show that there are closely related intact MGEs found in genomes of S. aureus. The genome from a 1980s isolate from chickens in Ireland contains 3 copies of this MGE. More recent chicken isolates descended from that genome (Poland 2009, Oklahoma 2010, and Arkansas 2018) contain 2 to 4 related copies. Many of the genes of this MGE can be identified in disparate regions of the genomes of other chicken isolates of S. aureus. BLAST searches of the NCBI databases detect no similar MGEs outside of S. aureus and S. agnetis. These MGEs encode no proteins related to those produced by Staphylococcus aureus Pathogenicity Islands, which have been associated with the transition of S. aureus from human to chicken hosts. Other than mobilization functions, most of the genes in these new MGEs annotate as hypothetical proteins. The MGEs we describe appear to represent a new family of Chromosomal Islands (CIs) shared amongst S. agnetis and S. aureus. Further work is needed to understand the role of these CIs/MGEs in pathogenesis. Analysis of horizontal transfer of genetic elements between isolates and species of Staphylococci provides clues to evolution of host-pathogen interactions as well as revealing critical determinants for animal welfare and human diseases. [ABSTRACT FROM AUTHOR]