Osteomyelitis In Broilers Research Articles

Embryo lethality assay as a tool for assessing virulence of isolates from bacterial chondronecrosis with osteomyelitis in broilers

We used an embryo lethality assay (ELA) to assess virulence for different isolates from cases of bacterial chondronecrosis with osteomyelitis (BCO) in broilers. Lameness is among the most significant animal welfare issues in the poultry industry. Bacterial infections are a major cause of lameness and different bacterial species have been obtained from lame broilers. Reliable lab-based assays are required to assess relative virulence of bacteria obtained from lame broilers. ELA has been used to assess lethal dosage of Enterococcus faecalis and Enterococcus cecorum. We hypothesized that ELA could substitute for more laborious and costly assessments of BCO isolate pathogenicity using live birds. We evaluated 2 different levels of bacteria injected into eggs from layer and commercial broiler embryos. Significant findings include 1) Escherichia coli from neighboring farms operated by the same integrator had very different embryo lethality, 2) isolate Staphylococcus agnetis 908 had low virulence in ELA, even though this isolate can induce more than 50% BCO lameness, 3) Enterococcus cecorum 1415 also had low pathogenicity; even though it was recovered from severe bilateral tibial dyschondroplasia, 4) human and chicken BCO isolates of S. aureus had significant pathogenicity, 5) virulence for some isolates was highly variable possibly corresponding with quality of the embryos/fertile eggs used, and 6) ELA pathogenicity was much lower for our BCO isolates than previous reports which may reflect maternal environment. Overall, ELA virulence and BCO virulence are not always concordant indicating that ELA may not be an effective measure for assessing virulence with respect to BCO.

Chondronecrosis with osteomyelitis in broilers: further defining a bacterial challenge model using standard litter flooring and protection with probiotics

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.

Chondronecrosis with osteomyelitis in broilers: further defining lameness-inducing models with wire or litter flooring to evaluate protection with organic trace minerals

The feed additive Availa-ZMC was investigated for the ability to reduce lameness in broilers using 2 alternative models for inducing lameness. The mixture of organic trace minerals was effective in reducing lameness by 20% in the wire flooring model and 25% in the litter flooring model with the bacterial challenge. Lameness in both models is overwhelmingly attributable to bacterial chondronecrosis with osteomyelitis. The reduction in lameness was associated, at least in part, with enhanced intestinal barrier integrity mediated by elevated expression of tight junction proteins and stimulation of bactericidal killing of adherent peripheral blood monocytes obtained from the birds treated with Availa-ZMC. Lameness is a major animal welfare concern in broiler production. The wire flooring model and litter flooring model with the bacterial challenge are effective models for evaluation of management strategies for mitigating infectious causes of lameness.

RNA-seq reveals downregulated osteochondral genes potentially related to tibia bacterial chondronecrosis with osteomyelitis in broilers

BackgroundBacterial chondronecrosis with osteomyelitis (BCO) develops in the growth plate (GP) of the proximal femur and tibia and is initiated by damage to the less mineralized chondrocytes followed by colonization of opportunistic bacteria. This condition affects approximately 1% of all birds housed, being considered one of the major causes of lameness in fast growing broilers. Although several studies have been previously performed aiming to understand its pathogenesis, the molecular mechanisms involved with BCO remains to be elucidated. Therefore, this study aimed to generate a profile of global differential gene expression involved with BCO in the tibia of commercial broilers, through RNA sequencing analysis to identity genes and molecular pathways involved with BCO in chickens.ResultsOur data showed 192 differentially expressed (DE) genes: 63 upregulated and 129 downregulated in the GP of the tibia proximal epiphysis of BCO-affected broilers. Using all DE genes, six Biological Processes (BP) were associated with bone development (connective tissue development, cartilage development, skeletal system development, organ morphogenesis, system development and skeletal system morphogenesis). The analyses of the upregulated genes did not indicate any significant BP (FDR < 0.05). However, with the downregulated genes, the same BP were identified when using all DE genes in the analysis, with a total of 26 coding genes explaining BCO in the tibia: ACAN, ALDH1A2, CDH7, CHAD, CHADL, COL11A1, COMP, CSGALNACT1, CYR61, FRZB, GAL3ST1, HAPLN1, IHH, KIF26B, LECT1, LPPR1, PDE6B, RBP4A, SERINC5, SFRP1, SOX8, SOX9, TENM2, THBS1, UCHL1 and WFIKKN2. In addition, seven transcription factors were also associated to BCO: NFATC2, MAFB, HIF1A-ARNT, EWSR1-FLI1, NFIC, TCF3 and NF-KAPPAB.ConclusionsOur data show that osteochondral downregulated genes are potential molecular causes of BCO in broilers, and the bacterial process seems to be, in fact, a secondary condition. Sixteen genes responsible for bone and cartilage formation were downregulated in BCO-affected broilers being strong candidate genes to trigger this disorder.

Proximal femoral head transcriptome reveals novel candidate genes related to epiphysiolysis in broiler chickens

BackgroundThe proximal femoral head separation (FHS) or epiphysiolysis is a prevalent disorder affecting the chicken femur epiphysis, being considered a risk factor to infection which can cause bacterial chondronecrosis with osteomyelitis in broilers. To identify the genetic mechanisms involved in epiphysiolysis, differentially expressed (DE) genes in the femur of normal and FHS-affected broilers were identified using RNA-Seq technology. Femoral growth plate (GP) samples from 35-day-old commercial male broilers were collected from 4 healthy and 4 FHS-affected broilers. Sequencing was performed using an Illumina paired-end protocol. Differentially expressed genes were obtained using the edgeR package based on the False Discovery Rate (FDR < 0.05).ResultsApproximately 16 million reads/sample were generated with 2 × 100 bp paired-end reads. After data quality control, approximately 12 million reads/sample were mapped to the reference chicken genome (Galgal5). A total of 12,645 genes were expressed in the femur GP. Out of those, 314 were DE between groups, being 154 upregulated and 160 downregulated in FHS-affected broilers. In the functional analyses, several biological processes (BP) were overrepresented. Among them, those related to cell adhesion, extracellular matrix (ECM), bone development, blood circulation and lipid metabolism, which are more related to chicken growth, are possibly involved with the onset of FHS. On the other hand, BP associated to apoptosis or cell death and immune response, which were also found in our study, could be related to the consequence of the FHS.ConclusionsGenes with potential role in the epiphysiolysis were identified through the femur head transcriptome analysis, providing a better understanding of the mechanisms that regulate bone development in fast-growing chickens. In this study, we highlighted the importance of cell adhesion and extracellular matrix related genes in triggering FHS. Furthermore, we have shown new insights on the involvement of lipidemia and immune response/inflammation with FHS in broilers. Understanding the changes in the GP transcriptome might support breeding strategies to address poultry robustness and to obtain more resilient broilers.

Vertebral Osteomyelitis in Broilers: A Review

Locomotor diseases are still a challenge in modern poultry. Vertebral osteomyelitis (VO) is an emerging disease in broilers worldwide. The inflammatory process in the affected thoracic vertebra (T4) and subsequent spinal cord compression leads to clinical signs related to locomotor impairment, inadequate feeding and drinking, and increased mortality in the affected flocks. The pathogenesis of the disease is poorly understood and Enterococcus cecorum is the bacterium most frequently associated with the disease. However, other bacteria such as E. faecalis, E. durans, Escherichia coli and Staphylococcus aureus have been recently detected in cases of the disease, raising questions regarding its etiopathogenesis. As many questions about VO in broilers remain unanswered, knowledge on its prevention, control and treatment is limited. In this review, we compile and discuss the currently available information concerning VO in broilers and highlight important aspects of the disease.

Chondronecrosis with osteomyelitis in broilers: Further defining a bacterial challenge model using the wire flooring model

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 105 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.

The involvement of RUNX2 and SPARC genes in the bacterial chondronecrosis with osteomyelitis in broilers

Economic losses due to an increase of leg disorders in broilers have become a major concern of the poultry industry. Despite the efforts to reduce skeletal abnormalities in chickens, insufficient progress has been made. Bacterial chondronecrosis with osteomyelitis (BCO) is one of the main disorders that affect bone integrity in broilers. However, the genetic pathways and genes involved in most bone problems, including BCO, remains unclear. In this study, femoral samples from male broilers with 45 days of age affected or not with BCO were used to compare the relative expression with a reverse transcription real time PCR approach of 13 candidate genes: SPP1 (osteopontin), TNFRSF11B (osteoprotegerin), SPARC (osteonectin), CALB1 (calbidin 1), CALM (Calmodulin 2), IBSP (sialoprotein), COL1A2 (collagen, type I, α 2), BMP2 (bone morphogenetic protein 2), BMP3 (bone morphogenetic protein 3), RANKL (κ-B nuclear factor ligand), SMAD1 (SMAD family member 1), LEPR (leptin receptor) and RUNX2 (related transcription factor Runt 2). Differential expression test between affected and non-affected groups was performed using the REST software. The RUNX2 and SPARC genes were downregulated (P<0.05) in the affected group, with reduced expression of fourfold when compared with the non-affected group. This result indicates that the downregulation of RUNX2 and SPARC can contribute to an increased incidence of BCO in broilers.

Prophylactic administration of a combined prebiotic and probiotic, or therapeutic administration of enrofloxacin, to reduce the incidence of bacterial chondronecrosis with osteomyelitis in broilers

Bacteria entering the bloodstream via translocation from the gastrointestinal tract spread hematogenously and can trigger bacterial chondronecrosis with osteomyelitis (BCO) by infecting osteochondrotic microfractures in the epiphyseal-physeal cartilage of the proximal femora and tibiae. In experiment 1, broilers were fed control feed or the same feed containing BacPack 2X, which includes the prebiotic IMW50 (a mannan oligosaccharide beta-glucan yeast cell wall product) plus the probiotic Calsporin (Bacillus subtilis C-3102). Broilers reared on wire flooring consistently developed higher incidences of BCO than hatchmates reared on wood shavings litter (≥24 vs. ≤4%, respectively; P = 0.001). Adding BacPack 2X to the feed on d 1 through 56 delayed the age of onset and reduced the cumulative incidence of BCO on wire flooring when compared with broilers fed the control feed (24.0 vs. 40.7%, respectively; P = 0.003). In experiment 2, broilers reared on wire flooring received tap water on d 1 through 62 (control group) or therapeutic levels of the potent fluoroquinolone antimicrobial enrofloxacin in the water on d 35 through 54 (enrofloxacin group). During enrofloxacin administration, half as many birds developed BCO in the enrofloxacin group when compared with the control group (8.1 vs. 19.5%, respectively, on d 35 through 54; P = 0.001), whereas both groups had similar BCO incidences subsequent to withdrawing enrofloxacin on d 55 through 62 (14.8 vs. 18.2% for the enrofloxacin vs. control groups; P = 0.386). Cumulative lameness incidences for d 1 through 62 were higher for the control group than for the enrofloxacin group (39.0 vs. 25.8%, respectively; P = 0.003). These results demonstrate that wire flooring imposes a rigorous challenge that leads to high incidences of BCO that can be difficult to suppress, even with therapeutic doses of enrofloxacin. Prophylactically adding BacPack 2X to the feed reduced the incidence of BCO lameness by a proportion similar to that achieved with enrofloxacin, indicating that probiotics potentially can provide effective alternatives to antibiotics for reducing BCO lameness attributable to bacterial translocation and hematogenous distribution.

Bacterial chondronecrosis with osteomyelitis in broilers: Influence of sires and straight-run versus sex-separate rearing

Two experiments (E1, E2) were conducted to compare the influence of sires (sire A on dam C vs. sire B on dam C) and straight-run versus sex-separate rearing on the incidence of bacterial chondronecrosis with osteomyelitis (BCO) in broilers. Fertile eggs from commercial breeder flocks were incubated and hatched at the University of Arkansas Poultry Research Hatchery. Male and female chicks were reared together (straight-run) or separately (sex-separate) in 3 × 3 m pens on litter or flat wire flooring with 65 (E1) or 60 (E2) birds per pen. Necropsies revealed lesions that are pathognomonic for BCO in ≥98% of the birds that became lame. The SigmaStat Z-test was used to compare cumulative BCO incidences through 8 wk of age. For birds reared on litter, the incidences of BCO were low regardless of cross or sex (range: 1.7 to 5.1%; P ≥ 0.6). Within a cross and sex, rearing the broilers straight-run versus sex-separate on wire flooring did not significantly affect the incidence of BCO. Significant incidences of BCO did not develop until after d 40. Males from the sire A cross developed a higher incidence of BCO than males from the sire B cross in E1 (27 vs. 17%, respectively; P = 0.009) but not in E2 (28.5 vs. 22.6%, respectively; P = 0.141). In both experiments, males from the sire A cross developed higher incidences of BCO than females from the sire B cross (27 vs. 11.9%, in E1; 28.5 vs. 14.8%, in E2). With the sexes pooled, broilers from the sire A cross consistently developed higher incidences of BCO than broilers from the sire B cross (21.4 vs. 14.9%, P = 0.005 in E1; 26.5 vs. 18.7%, P = 0.003 in E2). High susceptibilities to both femoral head (all femoral head necrosis = 66 to 85% incidences) and tibial head (all tibial head necrosis = 81 to 96% incidences) BCO lesions were demonstrated in lame birds from both sexes and crosses. This study supports a sire influence on the susceptibility of broilers to BCO. Sire lines can be chosen to reduce BCO susceptibility when broilers are grown beyond 6 wk of age.

Evaluating portable wire-flooring models for inducing bacterial chondronecrosis with osteomyelitis in broilers

Rearing broilers on flat or sloping wire flooring is an effective method for consistently triggering lameness attributable to bacterial chondronecrosis with osteomyelitis (BCO). Portable obstacles known as speed bumps (SB) also consistently trigger modest incidences of BCO when they are installed between feed and water lines in litter flooring facilities. Two experiments were conducted to determine the most effective broiler age for introducing the SB into litter flooring pens, and to evaluate alternative configurations of the traditional SB with the expectation that amplified mechanical challenges to the legs of broilers should increase the incidence of BCO. Broiler chicks obtained from commercial hatcheries (lines B and D in experiment 1, lines A and B in experiment 2) were reared in floor pens with ad libitum feed and water and a 23L:1D photoperiod. In experiment 1, the 5 floor treatments included wood shavings litter only (L), flat wire only (W), or litter plus SB installed at 14, 28, or 42 d of age. Line B was more susceptible to lameness than line D (25.9 vs. 15.3% for all treatments combined; P = 0.001). Both lines developed low incidences of lameness on L (11 to 13%), intermediate incidences on SB regardless of day of installation (12 to 23%), and high incidences on W (21 to 39%). In experiment 2, broilers were reared with 7 floor treatments, including L, W, SB with a 50% slope (SB50%); SB50% with a limbo bar installed over the apex; SB with a 66% slope and limbo bar; SB50% with a nipple water line suspended over the apex; and a pagoda-top SB. All SB were inserted on d 28. Line B was more susceptible to lameness than line A (20.2 vs. 16.1% for all treatments combined; P < 0.05), and for both lines combined the lameness percentages averaged 7.7 (L), 29.2 (W), 17.3 (SB50%), 16.2 (SB50% with a limbo bar), 21.5 (SB with a 66% slope and limbo bar), 20.8 (SB50% with a nipple water line), and 11.5% (pagoda-top). These studies demonstrate the portable SB can be effectively used to experimentally trigger BCO in broilers.

Development of an experimental model of bacterial chondronecrosis with osteomyelitis in broilers following exposure to Staphylococcus aureus by aerosol, and inoculation with chicken anaemia and infectious bursal disease viruses

A series of experiments was designed in an attempt to reproduce bacterial chondronecrosis with osteomyelitis in broiler chickens using a natural route of infection. Birds in isolators were exposed to a suspension of Staphylococcus aureus by aerosol or exposed to S. aureus and subsequently inoculated with chicken anaemia virus (CAV) alone, or with CAV and infectious bursal disease virus (IBDV). Subsequently, S. aureus was recovered and bacterial chondronecrosis with osteomyelitis was diagnosed, by histology, in the proximal end of the femur and/or tibiotarsus of lame birds exposed to S. aureus with and without CAV and IBDV infections. Birds fed 60% of the recommended feed intake for the breed developed a lower incidence of S. aureus infection and/or bacterial chondronecrosis (P 0.05) than birds fed 100% of the recommended intake. A significantly lower incidence of S. aureus was recovered (P 0.05) in birds simultaneously exposed to S. aureus and inoculated with CAV and IBDV at day 21, than in birds exposed to S. aureus at day 10, and inoculated with CAV and IBDV at day 21. With the exception of birds exposed to S. aureus at 1 day old, a higher incidence of bacterial chondronecrosis was diagnosed in birds exposed to S. aureus and inoculated with CAV and IBDV than in birds exposed to S. aureus alone. It is hypothesised that inoculation with CAV and IBDV at day 21 enhanced the development of bacterial chondronecrosis in birds exposed to S. aureus at day 10 and fed 100% of the recommended feed intake or ad libitum.