Randomized controlled trial comparison of analgesic drugs for control of pain associated with induced lameness in lactating dairy cattle.

Both the economic loss and welfare implications of lameness affect the dairy industry. Currently no analgesic drugs are approved to alleviate lameness-associated pain in lactating dairy cattle in the United States. In this randomized controlled trial, 48 lactating Holsteins were enrolled to evaluate the effect of oral meloxicam and i.v. flunixin meglumine on induced lameness. Cows were allocated to 1 of 4 treatment groups (n = 12 per group): lameness and flunixin meglumine (LAME + FLU); lameness and meloxicam (LAME + MEL); lameness and placebo (LAME + PLBO); or sham induction and placebo (SHAM + PLBO). Six hours before treatment, arthritis-synovitis was induced in the distal interphalangeal joint with 20 mg of amphotericin B, whereas SHAM cows were given an intra-articular injection of an equal volume (4 mL) of isotonic saline. Cows in LAME + FLU received 2.2 mg/kg flunixin meglumine i.v. and whey protein placebo orally; LAME + MEL were administered 1 mg/kg meloxicam orally and 2 mL/45 kg sterile saline placebo i.v.; LAME + PLBO were administered 2 mL/45 kg sterile saline placebo i.v. and whey protein placebo orally; and SHAM + PLBO received 2 mL/45 kg sterile saline placebo i.v. and whey protein placebo orally. The initial treatment of MEL, FLU, or PLBO was identified as time 0 h and followed by a second dose 24 h later with data collection for 120 h. The methods used to assess analgesic efficacy were electronic pressure mat, visual lameness assessment, visual analog score, plasma cortisol concentration, plasma substance P concentration, mechanical nociception threshold, and infrared thermography imaging. Linear mixed effect modeling was the primary method of statistical analysis. Visual lameness scoring indicated a lower proportion of the FLU + LAME group was lame at the T2 h and T8 h time points in comparison to the positive controls, whereas MEL therapy resulted in a lower proportion of lame cows at the T8 h time point. Cortisol area under the effect curve was lower following FLU therapy compared with LAME + PBLO for the 0-2 h (LSM difference = 35.1 ng·h/mL, 95% CI: 6.8, 63.3 ng·h/mL), 2-8 h (LSM difference = 120.6 ng·h/mL, 95% CI: 77.2, 164.0 ng·h/mL), and 0-24 h (LSM difference = 226.0 ng·h/mL, 95% CI: 103.3, 348.8 ng·h/mL) time intervals. Following MEL therapy, cortisol area under the effect curve was lower than LAME + PLBO for both the 2 to 8 h (LSM difference = 93.6 ng·h/mL, 95% CI: 50.2, 137.0 ng·h/mL) and 0 to 24 h time intervals (LSM difference = 187.6 ng·h/mL, 95% CI: 64.9, 310.4 ng·h/mL). Analysis of data from other assessment modalities failed to discern biologically relevant differences between treatment groups. We conclude that meaningful differences were evident for visual lameness assessment and cortisol from MEL and FLU treatment versus the positive control. Further clinical research is needed toward development of a model that will create reproducible events that are more pronounced in severity and duration of lameness which can be validated as a substitute for naturally occurring lameness cases.

Impacts of chronic and increasing lipopolysaccharide exposure on production and reproductive parameters in lactating Holstein dairy cows.

Lipopolysaccharide (LPS) administration causes immunoactivation, which negatively affects production and fertility, but experimental exposure via an acute bolus is unlikely to resemble natural infections. Thus, the objectives were to characterize effects of chronic endotoxemia on production parameters and follicular development in estrous-synchronized lactating cows. Eleven Holstein cows (169 ± 20 d in milk; 681 ± 16 kg of body weight) were acclimated to their environmental surroundings for 3 d and then enrolled in 2 experimental periods (P). During P1 (3 d) cows consumed feed ad libitum and baseline samples were obtained. During P2 (7 d), cows were assigned to continuous infusion of either (1) saline-infused and pair-fed (CON-PF; 40 mL/h of saline i.v.; n = 5) or (2) LPS infused and ad libitum fed (LPS-AL; Escherichia coli O55:B5; 0.017, 0.020, 0.026, 0.036, 0.055, 0.088, and 0.148 µg/kg of body weight/h i.v. on d 1 to 7, respectively; n = 6). Controls were pair-fed to the LPS-AL group to eliminate confounding effects of dissimilar nutrient intake. Infusing LPS temporally caused mild hyperthermia on d 1 to 3 (+0.49°C) relative to baseline. Dry matter intake of LPS-AL cows decreased (28%) on d 1 of P2, then progressively returned to baseline. Relative to baseline, milk yield from LPS-AL cows was decreased on d 1 of P2 (12%). No treatment differences were observed in milk yield during P2. Follicular growth, dominant follicle size, serum progesterone (P4), and follicular P4 and 17ß-estradiol concentrations were similar between treatments. Serum 17ß-estradiol tended to increase (115%) and serum amyloid A and LPS-binding protein were increased (118 and 40%, respectively) in LPS-AL relative to CON-PF cows. Compared with CON-PF, neutrophils in LPS-AL cows were initially increased (45%), then gradually decreased. In contrast, monocytes were initially decreased (40%) and progressively increased with time in the LPS-AL cows. Hepatic mRNA abundance of cytochrome P450 family 2 subfamily C (CYP2C) or CYP3A was not affected by LPS, nor was there a treatment effect on toll-like receptor 4 or LBP; however, acyloxyacyl hydrolase and RELA subunit of nuclear factor kappa B tended to be increased in LPS-AL cows. These data suggest lactating dairy cows become tolerant to chronic and exponentially increasing LPS infusion in terms of production and reproductive parameters.

Efficacy of vaccination with a Klebsiella pneumoniae siderophore receptor protein vaccine for reduction of Klebsiella mastitis in lactating cattle.

Clinical mastitis caused by Klebsiella spp. is an emerging problem in the US dairy industry and results in a high degree of financial losses to dairy workers. This study was conducted as a randomized, blinded, and placebo-controlled efficacy study of a Klebsiella pneumoniae siderophore receptor protein (SRP) vaccine (Kleb-SRP), with a total of 569 cows and heifers enrolled. The study was designed to look at vaccine effect on Klebsiella mastitis; however, the SRP in Klebsiella are highly conserved across coliform bacteria, which means that the vaccine has potential for cross-protection against all coliforms. Cows were paired based on parity, days in milk at enrollment, and somatic cell count. Within pairs, individuals were randomized to receive either Kleb-SRP or a placebo formulation. Following vaccination, the incidence of Klebsiella spp. and total coliform mastitis from natural exposure were compared to determine the efficacy of the vaccine. When analyzing all cows, the reduction of mastitis risk was not significant, though milk production increased 0.31 kg/d and somatic cell counts were reduced by 20.1%. When administered before calving, the vaccine reduced the risk of Klebsiella and total coliform mastitis by 76.9 and 47.5% respectively; however, we observed no significant effect when administered after calving. The vaccine, when administered before calving, also increased milk production by an average of 1.74 kg/d and reduced somatic cell counts by 64.8%. When administered after calving, we noted a slight decrease in daily milk production (0.39 kg) but no significant effect on somatic cell counts. All cows in the study (including vaccinates and placebo) received multiple doses of a commercially available licensed Escherichia coli bacterin. It should be noted that this herd was chosen because of the high number of clinical Klebsiella clinical mastitis cases this herd experienced before the trial and the extreme environmental challenge that was present from bedding with dried manure solids. The data from this study demonstrate efficacy of the Kleb-SRP vaccine against Klebsiella mastitis alone and coliform mastitis in general (including all coliforms) when administered before the initiation of a lactation cycle.

Effect of chromium on bioenergetics and leukocyte dynamics following immunoactivation in lactating Holstein cows.

Activated immune cells are insulin sensitive and utilize copious amounts of glucose. Because chromium (Cr) increases insulin sensitivity and may be immunomodulatory, our objective was to evaluate the effect of supplemental Cr (KemTrace Cr propionate, 20 g/d; Kemin Industries Inc., Des Moines, IA) on immune system glucose utilization and immune system dynamics following an intravenous endotoxin challenge in lactating Holstein cows. Twenty cows (320 ± 18 d in milk) were randomly assigned to 1 of 4 treatments: (1) pair-fed (PF) control (PF-CON; 5 mL of saline; n = 5), (2) PF and Cr supplemented (PF-Cr; 5 mL of saline; n = 5), (3) lipopolysaccharide (LPS)-euglycemic clamp and control supplemented (LPS-CON; 0.375 µg/kg of body weight LPS; n = 5), and (4) LPS-euglycemic clamp and Cr supplemented (LPS-Cr; 0.375 µg/kg of body weight LPS; n = 5). The experiment was conducted serially in 3 periods (P). During P1 (3 d), cows received their respective dietary treatments and baseline values were obtained. At the initiation of P2 (2 d), either a 12-h LPS-euglycemic clamp was conducted or cows were PF to their respective dietary counterparts. During P3 (3 d), cows consumed feed ad libitum and continued to receive their respective dietary treatment. During P2, LPS administration decreased dry matter intake (DMI; 40%) similarly among diets, and by experimental design the pattern and magnitude of reduced DMI were similar in the PF cohorts. During P3, LPS-Cr cows tended to have decreased DMI (6%) relative to LPS-CON cows. Relative to controls, milk yield from LPS-challenged cows decreased (58%) during P2 and LPS-Cr cows produced less (16%) milk than LPS-CON cows. During P3, milk yield progressively increased similarly in LPS-administered cows, but overall milk yield remained decreased (24%) compared with PF controls. There were no dietary treatment differences in milk yield during P3. Circulating insulin increased 9- and 15-fold in LPS-administered cows at 6 and 12 h postbolus, respectively, compared with PF controls. Compared with LPS-CON cows, circulating insulin in LPS-Cr cows was decreased (48%) at 6 h postbolus. Relative to PF cows, circulating LPS binding protein and serum amyloid A from LPS-administered cows increased 2- and 5-fold, respectively. Compared with PF cows, blood neutrophil counts in LPS-infused cows initially decreased, then gradually increased 163%. Between 18 and 48 h postbolus, the number of neutrophils was increased (12%) in LPS-Cr versus LPS-CON cows. The 12-h total glucose deficit was 220 and 1,777 g for the PF and LPS treatments, respectively, but glucose utilization following immune activation was not influenced by Cr. In summary, supplemental Cr reduced the insulin response and increased circulating neutrophils following an LPS challenge but did not appear to alter the immune system's glucose requirement following acute and intense activation.

Comparison of milk and plasma pharmacokinetics of meloxicam in postpartum versus mid-lactation Holstein cows.

The objective of this study reported here was determine whether differences occurred in meloxicam pharmacokinetics between postpartum cows and mid-lactation cows. Preliminary data from a separate study (P. J. Gorden, unpublished data) in postpartum cows demonstrated elevated plasma and milk concentration profiles compared to previously published data (Malreddy, Coetzee, KuKanich, & Gehring, ). Two different groups were enrolled, each with 10 cows. The treatment group (TRT) was postpartum cows treated with meloxicam, and the positive control (PC) group was cows in mid-lactation treated with meloxicam. Plasma and milk meloxicam concentrations between the TRT and PC group were compared. Significant differences in meloxicam concentration in plasma were determined at all time points from 8 hr to 120 hr post-treatment. In milk, there was a treatment (p = .003), time (p < .001), and treatment by time interaction (p < .001). Significant differences in milk meloxicam concentration were determined at all time points from 8 hr to 96 hr post-treatment, except for the 16-hr time point. The time needed for meloxicam to no longer be detected in milk of the TRT group was longer compared to the PC group, indicating that a longer milk withdrawal is needed. These data suggest higher bioavailability as the underlying mechanism. Further research is needed to determine the mechanisms underlying differences this outcome.

A study to examine the relationship between metritis severity and depletion of oxytetracycline in plasma and milk after intrauterine infusion.

Metritis is a frequent problem in postpartum dairy cows. Intrauterine therapy with the antimicrobial oxytetracycline (OTC) is often used, although this therapy has not been shown to be superior to systemic therapy. The objectives of this study were to (1) determine the plasma and milk concentrations of OTC following intrauterine infusion in postpartum dairy cows with varying degrees of metritis severity; (2) determine the depletion time of OTC in an attempt to provide veterinarians withdrawal guidelines, should they use this therapy; and (3) correlate metritis severity scores with OTC concentrations in plasma and milk. Our hypothesis was that cows with more severe metritis would have higher OTC concentrations in milk following intrauterine therapy. Thirty-two cows were selected to participate in the study after farm personnel had determined that they had metritis based on evaluation of vaginal discharge between 4 and 14 DIM, in accordance with the farm's treatment protocols. Metritis scores (1-4) were assigned based on a published scheme: 1 represented yellow-to-orange thick discharge or translucent mucus with no fetid smell; 2 represented blood-tinged vaginal mucus, slightly watery, with little or no fetid smell; 3 represented red to red/brown watery discharge with moderate fetid smell; and 4 represented red to red/brown watery discharge containing pieces of placenta and an intense fetid smell. Trial cows received a single treatment of 4g of OTC (approximately 6.7mg/kg) via intrauterine infusion. Blood samples were collected over 96h, and milk samples were collected before intrauterine therapy and 3 times a day for 4 d following infusion. Following treatment, OTC rapidly diffused to plasma and subsequently to milk. Maximum OTC concentrations in plasma and milk occurred within the first 24h following intrauterine infusion, and 25 of the 32 cows had detectable OTC concentrations in milk at 4 d after intrauterine infusion. Cows with clinical metritis (metritis severity scores of 3 or 4) at the initiation of treatment were significantly and positively correlated with higher milk OTC concentrations at the second [time (T)9 h; r=0.43], fourth (T25 h; r=0.42), and fifth milking following treatment (T33 h; r=0.38) compared with cows with normal vaginal discharge. We also observed a positive correlation between initial metritis score and milk maximum concentration (r=0.36) and milk area under the concentration curve (r=0.36). Given that intrauterine administration of OTC is an extra-label therapy, dairy producers should consult with their veterinarian to ensure that milk is being tested at or below the established tolerance for OTC. This will ensure that violative drug residues do not enter the human food supply.