Dose determination and confirmation of a long-acting formulation of ceftiofur (ceftiofur crystalline free acid) administered subcutaneously for the treatment of bovine respiratory disease.

The objective of this work was to determine and confirm an effective dose of ceftiofur crystalline free acid sterile oil suspension (CCFA-SS, 100 mg ceftiofur equivalents (CE)/mL], a long-acting single-administration ceftiofur formulation, for the treatment of the bacterial component of bovine respiratory disease (BRD). Study 1 was a dose determination study that used an intratracheal Mannheimia haemolytica (Pasteurella haemolytica) challenge model to evaluate single-administration doses of CCFA-SS at 0.0, 1.1, 2.2, 3.3, 4.4 or 5.5 mg CE/kg body weight (BW) for the treatment of BRD. Data from this study were used to select doses for field testing in three multi-location clinical studies. In Study 2, the efficacy of a single administration dose of CCFA-SS at 4.4 mg CE/kg BW was compared with a negative control for the treatment of naturally occurring BRD in feedlot cattle. Treatments were administered when uniform clinical signs of BRD were present. Study 3 used a design similar to Study 2, and compared single-administration doses of CCFA-SS at 3.0 or 4.4 mg CE/kg BW with the positive-control tilmicosin (Micotil(R) 300 Injection, Elanco Animal Health) at 10 mg/kg BW. Study 4 compared the efficacy of single doses of CCFA-SS of 1.1-8.8 mg CE/kg BW with tilmicosin at 10 mg/kg BW. A total of 1176 cattle were included in these clinical studies. In Study 1, a dose of 4.55 mg CE/kg BW was determined to be effective. This was rounded to 4.4 mg CE/kg for field testing. In Study 2, a single dose of CCFA-SS at 4.4 mg CE/kg BW had a higher treatment success rate on day 14 (61%) than negative controls (26%, P < 0.01). However, in Study 3 this dose was judged to be at the beginning of an efficacious dose range for the treatment of BRD when compared with tilmicosin. In Study 4, day 28 treatment success rates were higher for CCFA-SS at 4.4-8.8 CE/kg BW than for tilmicosin (P=0.002) or the noneffective CCFA-SS dose of 1.1 mg CE/kg BW (P < 0.001). Based on decision criteria for Study 4, the effective dose was determined to be 4.4-5.5 mg CE/kg BW. These clinical studies demonstrated that a single dose of CCFA-SS (100 mg CE/mL) administered subcutaneously (s.c.) in the neck at 4.4-5.5 mg CE/kg BW is an effective treatment for BRD in feedlot cattle. However, this route of administration is no longer being considered for this formulation because of the ceftiofur residues that are present at the injection site for extended periods of time.

Comparison of plasma pharmacokinetics and bioequivalence of ceftiofur sodium in cattle after a single intramuscular or subcutaneous injection.

Ceftiofur sodium, a broad-spectrum cephalosporin, is active against gram-positive and gram-negative pathogens of veterinary importance. This study was designed to compare the bioequivalence of the sodium salt in cattle after a single intramuscular (i.m.) or subcutaneous dose (s.c.) of 2.2 mg ceftiofur equivalents/kg body weight. The criteria used to evaluate bioequivalence were (1) the area under the curve from time of injection to the limit of quantitation (LOQ) of the assay (AUC0-LOQ), and (2) time concentrations remained above 0.2 microg/mL (t>0.2). Twelve crossbred beef cattle were enrolled in a three-period, two-treatment crossover trial, with a minimum 2-week washout period between doses of 2.2 mg ceftiofur equivalents/kg. Blood samples were collected serially for up to 72 h post-injection. Plasma samples were then analyzed using a validated assay that measures ceftiofur, and all desfuroylceftiofur-related metabolites, by high-performance liquid chromatography (HPLC) as the stable derivative, desfuroylceftiofur acetamide. A maximum plasma concentration (Cmax) of 13.9+/-3.55 microg/mL was observed from 0. 67-2.0 h after i.m. administration, whereas a Cmax of 13.6+/-3.85 microg/mL was observed from 0.67-3.0 h after s.c. administration. The AUC0-LOQ was 108+/-35.0 microg. h/mL after i.m. dosing, compared with 105+/-29.8 microg. h/mL after s.c. dosing. The pre-established criterion for equivalence of the AUC0-LOQ for the i.m. and s.c. routes of administration was satisfied. The t>0.2 was 49.2+/-8.55 h after i.m. administration, compared with 47.0+/-9.40 h after s.c. administration. The pre-established criterion for equivalence of the t>0.2 for i.m. and s.c. administration was satisfied. The equivalence of AUC0-LOQ and t>0.2 for i.m. and s.c. administration of 2.2 mg ceftiofur equivalents (CE)/kg doses of ceftiofur sodium suggest similar therapeutic efficacy and systemic safety for the two routes of administration.

Efficacy of neomycin sulfate water medication on the control of mortality associated with colibacillosis in growing turkeys.

The objective of this investigation was to evaluate the efficacy, safety, and toxicity of neomycin sulfate (Neomix 325) water medication to control mortality associated with colibacillosis (Escherichia coli) in growing turkeys. One efficacy trial was conducted at five locations; each location included 2,880 sexed 21-d-old turkey poults that were naturally challenged with litter from turkey flocks that had colibacillosis. Between 5 and 7 d after challenge, and when mortality had reached 0.5%, poults were randomized within sex into three treatment groups of 0, 11, or 22 mg neomycin sulfate/kg body weight. In each location, each treatment was replicated 12 times with 40 poults per sex per replicate. All treatments were administered in the drinking water for 5 d. The pivotal decision criterion was mortality. Mortality was defined as 1) supported mortality (SM): positive microbial culture for E. coli and gross lesions, 2) diagnosed mortality (DM): diagnosed as associated with E. coli but not supported by lesions or positive microbiological cultures, 3) overall mortality (OM): mortality associated with E. coli or other microorganisms and miscellaneous reasons such as accidents (trampling or suffocations). Performance data (growth and feed utilization) also were measured and are reported without statistical analysis. Results from this efficacy study clearly demonstrated the effectiveness of neomycin sulfate against E. coli as measured by a reduction in mortality. In the target animal safety and toxicity study (done in conjunction with the efficacy study), neomycin sulfate in the drinking water at 66, 110, or 220 mg/kg per d for 15 d had no observable adverse effects on poult performance, as measured by feed or water consumption, body weight, gross pathology, or mortality.

Effects of age on the pharmacokinetics of single dose ceftiofur sodium administered intramuscularly or intravenously to cattle.

The effects of maturation on the intravenous (IV) and intramuscular (IM) pharmacokinetics of ceftiofur sodium following a dose of 2.2 mg ceftiofur equivalents/kg body weight were evaluated in 16 one-day-old Holstein bull calves (33-53 kg body weight initially; Group 1) and 14 six-month-old Holstein steers (217-276 kg body weight initially; Group 2). Group 1 calves were fed unmedicated milk replacer until 30 days of age and were then converted to the same roughag/concentrate diet as Group 2. Groups 1-IV and 2-IV received ceftiofur sodium IV, and Groups 1-IM and 2-IM received ceftiofur sodium IM. Group 1 calves were dosed at 7 days of age and at 1 and 3 months of age; group 2 calves were dosed at 6 and 9 months of age. Blood samples were obtained serially from each calf, and plasma samples were analysed using an HPLC assay that converts ceftiofur and all desfuroylceftiofur metabolites to desfuroylceftiofur acetamide. Cmax values were similar in all calves, and were no higher in younger calves than in older calves. Plasma concentrations remained above 0.150 microgram ceftiofur free acid equivalents/mliter for 72 h in 7-day-old calves, but were less than 0.150 microgram/mliter within 48 h following IV or IM injection for 6- and 9-month-old calves. Intramuscular bioavailability, assessed by comparing the model-derived area under the curve (AUCmod) from IM and IV injection at each age, appeared to be complete. After IV administration, the AUCmod in 7-day-old and 1-month-old calves (126.92 +/- 21.1 micrograms.h/mliter and 135.0 +/- 21.6 micrograms.h/mliter, respectively) was significantly larger than in 3-, 6- and 9-month-old calves (74.0 +/- 10.7 micrograms.h/mliter, 61.0 +/- 17.7 micrograms.h/mliter and 68.5 +/- 12.8 micrograms.h/mliter, respectively; P < 0.0001). The Vd(ss) decreased linearly within the first 3 months of life in cattle (0.345 +/- 0.0616 L/kg, 0.335 +/- 0.919 L/kg and 0.284 +/- 0.0490 L/kg, respectively; P = 0.031), indicative of the decreasing extracellular fluid volume in maturing cattle. The ClB was significantly smaller in 7-day-old and 1-month-old calves (0.0178 +/- 0.00325 L/h.kg and 0.0167 +/- 0.00310 L/h.kg, respectively) than in 3-, 6- and 9-month-old calves (0.0303 +/- 0.0046 L/h.kg, 0.0398 +/- 0.0149 L/h.kg and 0.0330 +/- 0.00552 L/h.kg, respectively; P < or = 0.001). This observation may be indicative of maturation of the metabolism and/or excretion processes for ceftiofur and desfuroylceftiofur metabolites. The approved dosage regimens for ceftiofur sodium of 1.1-2.2 mg/kg administered once daily for up to 5 consecutive days will provide plasma concentrations above the MIC for bovine respiratory disease pathogens for a longer period of time in neonatal calves than in older calves. Peak plasma concentrations of ceftiofur and desfuroylceftiofur metabolites were no higher in neonatal calves than in more mature cattle, highly suggestive that peak tissue concentrations would be no higher in neonatal calves than in more mature cattle.

Direct DNA extraction for PCR-mediated assays of soil organisms.

By using the rDNA of a plant wilt pathogen (Verticillium dahliae) as the target sequence, a direct method for the extraction of DNA from soil samples which can be used for PCR-mediated diagnostics without a need for further DNA purification has been developed. The soil organisms are disrupted by grinding in liquid nitrogen with the natural abrasives in soil, and losses due to degradation and adsorption are largely eliminated by the addition of skim milk powder. The DNA from disrupted cells is extracted with sodium dodecyl sulfate-phenol and collected by ethanol precipitation. After suitable dilution, this DNA extract can be assayed directly by PCR amplification technologies. The method is rapid, cost efficient, and when combined with suitable internal controls can be applied to the detection and quantification of specific soil organisms or pathogens on a large-scale basis.

The conduct of clinical trials and good clinical practices: the sponsor's perspective. Food and Drug Administration.

The Guideline, "Conduct of Clinical Investigations: Responsibilities of Clinical Investigators and Monitors for Investigational New Animal Drug Studies," which was issued by the Center for Veterinary Medicine (CVM) in October 1992 is intended to improve standards of clinical research. The purpose of clinical trials is to obtain data to support dose selection and to confirm the dose for regulatory approval of New Animal Drug Applications (NADA). The processes and procedures described in the Guideline should improve the "fitness for use" of data from clinical trials. Clinical investigators who desire to work with drug sponsors must understand the tenor of the Guideline and implement its recommendations when conducting pivotal clinical research. Protocols in which procedures are clearly described and responsibilities for monitors and investigators defined, in concert with a preplanned definitions of source data supported by appropriate quality control and quality assurance procedures, will ensure that the basis of approval for new animal drugs is defensible.

Dietary sodium bicarbonate as a treatment for exertional rhabdomyolysis in a horse.

A 3-year-old mare repeatedly had clinical signs of rhabdomyolysis on mild exertion. Serum creatine kinase and aspartate transaminase activities were high at rest. Responses to dietary sodium bicarbonate were tested through 7 alternating periods of supplementation of a basal ration of timothy hay and oats. Physical signs; venous blood pH and gases; blood glucose and lactate; serum electrolytes, enzymes, and creatinine; and urine pH were monitored before and after exercise. Dietary sodium bicarbonate raised resting venous blood pH and bicarbonate slightly and significantly increased urine pH from pH 7.46 to 8.2 (P less than 0.001). An exercise test included 5 minutes at the walk followed by 20 minutes at the trot. The exercise induced gait stiffness, muscle fasciculations, and muscle induration when the diet was not supplemented, but not when it was supplemented with sodium bicarbonate. Myoglobin was present in 16 of 21 urine samples after exercise during nonsupplemented periods, but only in 3 of 28 urine samples during supplemented periods (P less than 0.0001). Bicarbonate supplementation significantly decreased the responses of blood lactic acid, serum creatine kinase, and aspartate transaminase to exercise. Supplementation of the diet was associated with higher venous blood pH and bicarbonate ion concentrations throughout exercise. Dietary sodium bicarbonate apparently mitigated or prevented physical, chemical, and enzymatic characteristics of exertional rhabdomyolysis in this mare, possibly through its enhancement of buffering capacity in muscle tissue fluids.

Nature's Ballistic Missile.

The parasitic fungus Haptoglossa mirabilis infects its rotifer host by means of a gun-shaped attack cell. The anterior end of the cell is elongated to form a barrel; the wall at the mouth is invaginated deep into the cell to form a bore. A walled chamber at the base of the bore houses a complex, missile-like attack apparatus. The projectile is fired from the gun cell at high speed to accomplish initial penetration of the host.