Impact of disease in dairy cows on ceftiofur pharmacokinetics, withdrawal times and emergence of antimicrobial resistance

Producing safe and wholesome food is a priority of animal agriculture in order to maintain consumer confidence in the products produced by the animal agriculture industries. Antimicrobial residues and antimicrobial resistant organisms are two of the most important concerns of consumers regarding food safety. Withholding periods are in place to minimize the risk of antimicrobial residues being present in food. However, these withholding periods are established on healthy animals, not clinically ill animals. Additionally, there is no such withholding period in place to minimize the risk of transporting antimicrobial resistant organisms through food following treatment, although it has been proposed. The first objective of this dissertation was to investigate the impact of clinical disease on the pharmacokinetics of ceftiofur. The second objective was to investigate changes in the fecal microbiota following drug therapy. The results of this work demonstrate the volume of distribution is increased in diseased animals and area under the concentration curve is decreased. Additionally, the mean elimination half-life can be significantly different. In both studies involving clinical disease, at least one animal in each group had an elimination half-life that was nearly twice as long as the mean of the control group. No animals were found to have violative drug residues present in tissues following observation of the labeled withholding time. Fecal Escherichia coli populations transiently decreased following therapy with ceftiofur and ceftiofur resistant populations were significantly different than untreated controls. There was a tendency towards a significantly higher ceftiofur resistant E. coli population in diseased animals treated with ceftiofur versus healthy animals treated controls. By 14 days following therapy, total and resistant E. coli populations returned to pre-treatment levels. E. coli isolates that were resistant to ceftiofur were also cross-resistant to ampicillin and ceftriaxone. Additionally, 64.3% of the resistant isolates were also resistant to tetracycline. The ÃÂ-lactamase gene blaCTX-M was most commonly found in ceftiofur resistant isolates but mechanism for ceftiofur resistance was not identified in 55.8% of the ceftiofur resistant isolates. Isolates that were determined to be phenotypically ceftiofur cross-resistant were resistant to 4.72 while isolates that were ceftiofur sensitive were cross-resistant to 1.1 antimicrobials. The current study suggests that changes in bacterial populations following clinical disease are not different from those of healthy cows treated with ceftiofur. Observation of the established drug withholding period following treatment with ceftiofur crystalline free acid minimizes the risk of transferring fecal isolates harboring antimicrobial resistance to the public

A Survey of Dairy Farm Treatment Practices on Midwest Dairy Farms

Judicious antimicrobial use and antimicrobial stewardship have become buzzwords in production animal agriculture over the last few years. While these words are becoming expectations in the industry, very little is understood about their true meaning and the level of implementation of judicious use practices on dairy farms. We conducted an investigation on 85 dairy farms in the Midwest to document drug use practices on these farms. Our results indicate that most farms are doing an adequate job of implementing judicious practices, but there is room for improvement to meet expectations of regulatory officials and consumers

Utilization of Liquid Chromatography/Mass Spectrometry to Detect Drug Residues in Milk: Applications for Research and Commercial Dairying

Prevention of drug residues in milk is a daily endeavor on dairy farms. There is increasing scrutiny from the public and government when it comes to drug residues in milk. Drug residues can result from simple human errors, disease processes not allowing for normal clearance of a drug, or malicious activity. The testing methodologies used to detect drug residues have become more sensitive with many tests available that can detect drug levels below ten parts per billion (ppb)

Topical Flunixin Meglumine Effects on Pain Associated Biomarkers after Dehorning

Twenty-four calves were dehorned and treated with either topical flunixin meglumine formulated for systemic absorption or a placebo. Biomarkers associated with pain were evaluated for up to 72 hour after the dehorning procedure. Plasma cortisol concentrations, 90 minutes post-dehorning, and mechanical nociception threshold at the control site were the only tested biomarkers where a significant difference was demonstrated. No other differences of biomarkers between the two dehorned groups were observed for any time points. Although this product is easy to dose and dispense, its effects on pain biomarkers appears to be negligible

A Highly Effective Protocol for the Rapid and Consistent Induction of Digital Dermatitis in Holstein Calves

Bovine Digital Dermatitis (DD) is a leading cause of lameness in dairy cattle. DD is reportedly increasing in prevalence in beef cattle feedlots of the US. The exact etiologic agent(s) responsible for the disease have yet to be determined. Multiple studies have demonstrated the presence of a variety of Treponema spp. within lesions. Attempts to reproduce clinically relevant disease using pure cultures of these organisms has failed to result in lesions that mirror the morphology and severity of naturally occurring lesions. This manuscript details the systematic development of an experimental protocol that reliably induces digital dermatitis lesions on a large enough scale to allow experimental evaluation of treatment and prevention measures. In total, 21 protocols from five experiments were evaluated on their effectiveness in inducing DD lesions in 126 Holstein calves (504 feet). The protocols varied in the type and concentration of inoculum, frequency of inoculation, duration the feet were wrapped, and type of experimental controls need to validate a successful induction. Knowledge gained in the first four experiments resulted in a final protocol capable of inducing DD lesions in 42 of 44 (95%) feet over a 28 day period. All induced lesions were macroscopically and microscopically identified as clinical DD lesions by individuals blinded to protocols. Lesions were also located at the site of inoculation in the palmer aspect of the interdigital space, and induced clinically measurable lameness in a significant portion of the calves. Collectively these results validate the model and provide a rapid and reliable means of inducing DD in large groups of calves

Deep Sequencing Analysis Reveals the Temporal Microbiota Changes Associated with the Development of Bovine Digital Dermatitis

Bovine digital dermatitis (DD) is a leading cause of lameness in dairy cattle throughout the world. Despite 35 years of research, the definitive etiologic agent associated with the disease process is still unknown. Previous studies have demonstrated that multiple bacterial species are associated with lesions, with spirochetes being the most reliably identified organism. This study details the deep sequencing-based metagenomic evaluation of 48 staged DD biopsy specimens collected during a 3-year longitudinal study of disease progression. Over 175 million sequences were evaluated by utilizing both shotgun and 16S metagenomic techniques. Based on the shotgun sequencing results, there was no evidence of a fungal or DNA viral etiology. The bacterial microbiota of biopsy specimens progresses through a systematic series of changes that correlate with the novel morphological lesion scoring system developed as part of this project. This scoring system was validated, as the microbiota of each stage was statistically significantly different from those of other stages (P\u3c 0.001). The microbiota of control biopsy specimens were the most diverse and became less diverse as lesions developed. Although Treponema spp. predominated in the advanced lesions, they were in relatively low abundance in the newly described early lesions that are associated with the initiation of the disease process. The consortium of Treponema spp. identified at the onset of disease changes considerably as the lesions progress through the morphological stages identified. The results of this study support the hypothesis that DD is a polybacterial disease process and provide unique insights into the temporal changes in bacterial populations throughout lesion development

Bacterial Causes of Digital Dermatitis (DD) in Dairy Cattle

Bovine digital dermatitis (DD) is a leading cause of lameness in dairy cattle throughout the world. Despite 35 years of research, the definitive cause of the disease process is still unknown. Previous studies have demonstrated that multiple bacterial species are associated with lesions, with spirochetes being the most reliably identified organism. This study utilized total DNA sequencing of 48 staged DD biopsy specimens collected during a 3-year longitudinal study of disease progression in dairy cattle. Over 175 million sequences were obtained and used to identify the bacterial species that were present in the biopsies. There was no evidence of a fungal or DNA viral causes. The bacterial communities present in the biopsy specimens was found to progress through a systematic series of changes that correlate with the novel visual lesion scoring system developed as part of this project. Although Treponema spp. predominated in the advanced lesions, they were in relatively low abundance in the newly described early lesions that are associated with the initiation of the disease process. The results of this study support the hypothesis that DD is a polybacterial disease process and provide unique insights into the temporal changes in bacterial populations throughout lesion development. These insights are expected to be critical in the development of new treatment strategies and the potential development of effective vaccines. In addition, the development and validation of a lesion scoring system will assist with determining the prognosis of a lesion

Altered Plasma Pharmacokinetics of Ceftiofur Hydrochloride in Cows Affected with Severe Clinical Mastitis

Ceftiofur is the most commonly used antimicrobial in lactating dairy cows. Recently, there has been an increase in the number of violative residues of ceftiofur in the tissues of cull dairy cows. This was the first project in a series of projects we will be completing aimed at characterizing the pharmacokinetics of ceftiofur in disease challenged animals. The results of this study indicate that diseased animals have lower plasma concentrations and altered pharmacokinetics compared to healthy animals. Future work will investigate the influence of altered pharmacokinetics on the presence of violative residues

The genetic architecture of complete blood counts in lactating Holstein dairy cows

Complete blood counts (CBCs) measure the abundance of individual immune cells, red blood cells, and related measures such as platelets in circulating blood. These measures can indicate the health status of an animal; thus, baseline circulating levels in a healthy animal may be related to the productive life, resilience, and production efficiency of cattle. The objective of this study is to determine the heritability of CBC traits and identify genomic regions that are associated with CBC measurements in lactating Holstein dairy cattle. The heritability of CBCs was estimated using a Bayes C0 model. The study population consisted of 388 cows with genotypes at roughly 75,000 markers and 16 different CBC phenotypes taken at one to three time points (n = 33, 131, and 224 for 1, 2, and 3 time points, respectively). Heritabilities ranged from 0.00 ± 0.00 (red cell distribution width) to 0.68 ± 0.06 (lymphocytes). A total of 96 different 1-Mb windows were identified that explained more than 1% of the genetic variance for at least one CBC trait, with 10 windows explaining more than 1% of the genetic variance for two or more traits. Multiple genes in the identified regions have functions related to immune response, cell differentiation, anemia, and disease. Positional candidate genes include RAD52 motif-containing protein 1 (RDM1), which is correlated with the degree of immune infiltration of immune cells, and C-X-C motif chemokine ligand 12 (CXCL12), which is critically involved in neutrophil bone marrow storage and release regulation and enhances neutrophil migration. Since animal health directly impacts feed intake, understanding the genetics of CBCs may be useful in identifying more disease-resilient and feed-efficient dairy cattle. Identification of genes responsible for variation in CBCs will also help identify the variability in how dairy cattle defend against illness and injury