Pharmacokinetics of subcutaneous versus intramuscular administration of ceftiofur crystalline-free acid to bearded dragons (Pogona vitticeps)

Sarah M. Churgin Department of Surgical Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706.

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Kari E. Musgrave Department of Surgical Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706.

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Sherry K. Cox Department of Biomedical and Diagnostic Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996.

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Kurt K. Sladky Department of Surgical Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706.

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Abstract

Objective—To compare pharmacokinetics after a single IM or SC injection of ceftiofur crystalline-free acid (CCFA) to bearded dragons (Pogona vitticeps).

Animals—8 adult male bearded dragons.

Procedures—In a preliminary experiment, doses of 15 and 30 mg/kg, SC, were compared in 2 animals, and 30 mg/kg resulted in a more desirable pharmacokinetic profile. Then, in a randomized, complete crossover experimental design, each bearded dragon (n = 6) received a single dose of 30 mg of CCFA/kg IM or SC; the experiment was repeated after a 28-day washout period with the other route of administration. Blood samples were collected at 10 time points for 288 hours after injection. Plasma concentrations of ceftiofur and desfuroylceftiofur metabolites were measured via reverse-phase high-performance liquid chromatography. Data were analyzed with a noncompartmental model.

Results—No adverse effects were observed. Plasma concentrations greater than a target minimum inhibitory concentration of 1 μg/mL were achieved by 4 hours after administration by both routes. Mean plasma concentrations remained > 1 μg/mL for > 288 hours for both routes of administration.

Conclusions and Clinical Relevance—A single dose of CCFA (30 mg/kg) administered IM or SC to bearded dragons yielded plasma concentrations of ceftiofur and its metabolites > 1 μg/mL for > 288 hours. The SC route would be preferred because of less variability in plasma concentrations and greater ease of administration than the IM route. Future studies should include efficacy data as well as evaluation of the administration of multiple doses.

Abstract

Objective—To compare pharmacokinetics after a single IM or SC injection of ceftiofur crystalline-free acid (CCFA) to bearded dragons (Pogona vitticeps).

Animals—8 adult male bearded dragons.

Procedures—In a preliminary experiment, doses of 15 and 30 mg/kg, SC, were compared in 2 animals, and 30 mg/kg resulted in a more desirable pharmacokinetic profile. Then, in a randomized, complete crossover experimental design, each bearded dragon (n = 6) received a single dose of 30 mg of CCFA/kg IM or SC; the experiment was repeated after a 28-day washout period with the other route of administration. Blood samples were collected at 10 time points for 288 hours after injection. Plasma concentrations of ceftiofur and desfuroylceftiofur metabolites were measured via reverse-phase high-performance liquid chromatography. Data were analyzed with a noncompartmental model.

Results—No adverse effects were observed. Plasma concentrations greater than a target minimum inhibitory concentration of 1 μg/mL were achieved by 4 hours after administration by both routes. Mean plasma concentrations remained > 1 μg/mL for > 288 hours for both routes of administration.

Conclusions and Clinical Relevance—A single dose of CCFA (30 mg/kg) administered IM or SC to bearded dragons yielded plasma concentrations of ceftiofur and its metabolites > 1 μg/mL for > 288 hours. The SC route would be preferred because of less variability in plasma concentrations and greater ease of administration than the IM route. Future studies should include efficacy data as well as evaluation of the administration of multiple doses.

Ceftiofur is a third-generation, parenterally administered cephalosporin. Ceftiofur and its active metabolite, desfuroylceftiofur, have broad-spectrum bactericidal activity against a variety of gram-positive and gram-negative bacteria, including some anaerobes.1 There is a paucity of studies in which the activity of ceftiofur or its metabolites against bacterial isolates from reptiles has been evaluated.

Three major formulations of ceftiofur are available for use in veterinary medicine. There are 2 short-acting formulations, ceftiofur sodium and ceftiofur hydrochloride, and 1 long-acting formulation, CCFA. Ceftiofur sodium is licensed for use in the United States for various bacterial infections in cattle, swine, sheep, goats, horses, dogs, and 1-day-old chicks and poults, whereas ceftiofur hydrochloride is licensed in the United States for use in cattle and swine.2 In contrast, CCFA is licensed in the United States for use in cattle and swine as a single-injection antimicrobial and for use in horses as 2 injections administered 4 days apart.2 Ceftiofur crystalline-free acid differs from the other formulations of ceftiofur in that it is suspended in an aprylic-capric triglyceride and cottonseed oil–based carrier, which results in the sustained-release property.3

The pharmacokinetic profile of CCFA has been evaluated in a number of nondomestic species, including Asian elephants (Elephas maximus),4 helmeted guineafowl (Numida meleagris),5 American black ducks (Anas rubripes),6 ringneck doves (Streptopelia risoria),7 and ball pythons (Python regius).8 Dosages and results have differed among these studies, but CCFA has promise as a long-acting antimicrobial in these nondomestic species. For example, investigators found that IM administration of a dose of 10 mg/kg to guineafowl5 and ducks6 provided adequate plasma concentrations of ceftiofur and its metabolites for at least 72 hours. In elephants, a dose of 6.6 mg/kg, SC, provided adequate plasma concentrations for 7 to 10 days.4 Extended-duration pharmaceuticals in various nondomestic species could be of great benefit, particularly in animals that are difficult to handle or medicate.

Long-acting injectable antimicrobials are of particular interest in reptiles because of the difficulties associated with administration by other methods. Many reptiles do not eat on a daily basis, so administration of medicines in food is not practical. Direct oral administration is possible but may be difficult or impossible in those species (eg, large chelonians) that firmly resist opening their mouth, and there is a substantial risk of receiving a bite when handling any reptile. In the case of venomous reptiles, oral administration carries an even greater risk to handlers and is avoided whenever possible. Therefore, injectable antimicrobials are preferred for reptiles, but daily administration may be stressful for both handlers and animals.

Currently, one of the injectable antimicrobials most frequently used in reptile medicine is ceftazidime, which is typically prescribed at a dosing interval of 72 hours.9,10 This antimicrobial is effective against many bacterial pathogens in reptile species, and owners can be taught to administer the medication at home; however, in the experience of one of the authors (SMC), many practitioners do not routinely maintain a supply of ceftazidime at their clinic unless they have a large reptilian caseload. Additionally, although a 72-hour dosing interval is preferable to a shorter interval, that interval still requires frequent handling of dangerous reptiles, which is of particular concern when treating venomous species.

The objective of the study reported here was to perform a preliminary experiment to determine the pharmacokinetic profile of CCFA in bearded dragons (Pogona vitticeps). The dosage determined on the basis of the preliminary experiment was then used in a primary experiment to compare results for IM and SC administration. We hypothesized that SC administration of CCFA would provide a longer duration of plasma concentrations above a target MIC of 1 μg/mL, compared with results for the same dose administered IM. The target MIC of 1 μg/mL was selected on the basis that similar target values have been used in pharmacokinetic evaluations of ceftiofur in other exotic species.5,6,11

Materials and Methods

Animals—Eight bearded dragons were used in 2 experiments. Two bearded dragons were used in a preliminary experiment, and 6 bearded dragons were used in the subsequent primary experiment. Both experiments were approved by the University of Wisconsin School of Veterinary Medicine Institutional Animal Care and Use Committee.

The preliminary experiment included 2 client-owned adult male bearded dragons (body weight, 0.415 and 0.475 kg). Exact age of each of these animals was unknown because they had both been adopted as adults. Housing, diet, and husbandry were not standardized but were reported to be similar for both animals. Owner consent was obtained for inclusion of both of these bearded dragons.

The primary experiment included 6 adult male bearded dragons (mean ± SD body weight, 0.226 ± 0.021 kg); all 6 were purchased from a commercial supplier. Exact age of each of these animals was unknown, but they were estimated to be approximately 1 year old on the basis of size. Animals were maintained under controlled conditions at the University of Wisconsin School of Veterinary Medicine throughout the experiment. The bearded dragons were housed indoors at a constant temperature of 30°C throughout the experimental period, with a cycle of 12 hours of light and 12 hours of darkness; light was provided by full-spectrum UVA-UVB bulbs. All animals were fed the same diet, which consisted of dark leafy green vegetables mixed with a small amount of carrots and strawberries. Each bearded dragon was fed 1 live invertebrate (cricket or meal worm) daily. Physical examinations revealed all animals were apparently healthy before the start of the experimental period and after conclusion of the experiment, and all bearded dragons were subsequently adopted by owners.

Preliminary experiment—Both bearded dragons were examined, weighed, and administered a single dose of CCFAa (15 or 30 mg/kg). Injections were administered SC in the left scapular region with a 27-gauge needle. Blood samples (300 μL/sample) were collected from the caudal (tail) vein with a 27-gauge needle immediately before and 4, 12, 24, 48, 72, 120, 144, 192, and 288 hours after CCFA administration. An additional sample was obtained 28 days after CCFA administration to ensure that CCFA plasma concentrations had returned to baseline values. All blood samples were immediately placed into lithium heparin tubes. Samples were centrifuged (1,100 × g for 10 minutes) within 30 minutes after blood collection, and plasma was harvested and stored at −70°C until shipment for analysis. Plasma samples were shipped frozen on dry ice, and they were frozen when received by the analytic laboratory at the University of Tennessee College of Veterinary Medicine; samples were stored at −80°C until analysis.

Primary experiment—Bearded dragons used in the primary experiment were allowed an initial acclimatization period of ≥ 2 weeks before the start of the experiment. The primary experiment was conducted in 2 phases. In phase 1, each bearded dragon was examined, weighed, and assigned to a treatment group by use of a randomization procedure (computerized random numbers). Each animal then was administered a single dose of CCFAa (30 mg/kg) in the left biceps brachii muscle (IM) or the left scapular region (SC). Injections were administered with a 27-gauge needle. Blood samples (300 μL/sample) were collected from the caudal (tail) vein by use of a 27-gauge needle immediately before and 4, 12, 24, 48, 72, 120, 144, 192, and 288 hours after CCFA administration. After a 28-day washout period from the time of the initial injection, the experiment was repeated (phase 2) with the same dose of CCFA but the opposite route of administration for each animal. All blood samples were immediately placed into lithium heparin. Samples were centrifuged (1,100 × g for 10 minutes) within 30 minutes after blood collection, and plasma was harvested and stored at −70°C until shipment to the analytic laboratory for analysis. Frozen plasma was shipped to and stored at the analytic laboratory at the University of Tennessee College of Veterinary Medicine as described for the preliminary experiment.

Sample analysis—Maximum amount of time plasma samples from both experiments were stored frozen before analysis was approximately 6 weeks. Stability data gathered by one of the investigators (SKC; unpublished data) at the analytic laboratory indicated that there was no loss of drug after samples from guineafowl and wobbegong sharks were stored for a minimum of 6 weeks at −80°C, and all bearded dragon samples in the present study were analyzed within this time frame.

Analysis of ceftiofur concentrations in plasma samples was conducted with reverse-phase high-performance liquid chromatography. The system consisted of a separation moduleb and a UV detector.c Separation was attained on a 4.6 × 250-mm (diameter, 5 μm) C18 columnd with a 5-μm guard column.e The mobile phase was a mixture of 0.1% TFA in water and 0.1% TFA in acetonitrile. The mixture was pumped at a starting gradient of 90% TFA in water and 10% TFA in acetonitrile and was adjusted to 75% TFA in water and 25% TFA in acetonitrile over 25 minutes, then back to initial conditions over 3 minutes. The flow rate was 1.0 mL/min. Ceftiofur was quantified with UV detection at 265 nm.

Ceftiofur was extracted from plasma samples via a slightly modified derivation method12 that converted ceftiofur and all desfuroylceftiofur metabolites to DCA. Briefly, frozen plasma samples were thawed and mixed in a vortexer. An aliquot (100 μL) of each sample was transferred to a clean test tube, and 15 μL of internal standard (cefotaxime, 100 μg/mL) was added. Seven milliliters of 0.4% dithioerythritol in borate buffer was added, and tubes were placed in a 50°C water bath for 15 minutes. The tubes were removed and allowed to cool to room temperature (approx 23°C), and then 1.5 mL of iodoacetamide buffer was added. The solution was passed through a prewet hydrophilic-lipophilic balanced extraction column.f Samples were eluted with a solution of 5% glacial acetic acid in methanol, which was evaporated to dryness under a steady stream of nitrogen gas. Samples were reconstituted in 200 μL of mobile phase, and 50 μL was injected into the high-performance liquid chromatography system.

Standard curves and quality-control samples were prepared by spiking plasma obtained from ceftiofur naïve bearded dragons. The curve was linear for the concentration range of 0.1 to 100 μg/mL. Spiked standards and quality-control samples were treated the same as plasma samples obtained from the bearded dragons during the preliminary and primary experiments. Mean recovery was 99% for ceftiofur. Intra-assay variation ranged from 0.7% to 4.5%, and interassay variation ranged from 3.6% to 8.8%. The lower limit of quantification was 0.1 μg/mL.

Pharmacokinetic analysis—For each route of administration, plasma concentrations of DCA were analyzed by means of noncompartmental analysis with the aid of a computer program.g Mean and SD were calculated for the elimination rate constant, Tmax, Cmax, AUC, and mean residence time. Mean residence time and AUC were calculated from time 0 to the last measured concentration. The harmonic mean and pseudo-SD were calculated for terminal half-life.

Statistical analysis—Mean ± SD plasma DCA concentrations at each time point for each route of administration were calculated by the use of standard methods for normally distributed data.

Results

Animals—No adverse effects were observed in any bearded dragons during either of the experiments. There were no perceived changes in appetite or attitude despite the frequent handling and venipuncture. No reactions at injection sites were observed.

Preliminary experiment—Pharmacokinetic parameters were not calculated because the population size was small (n = 2) and diet, husbandry, and environmental temperature were not completely controlled. The bearded dragon that received CCFA at 15 mg/kg, SC, had a Cmax of 3.14 μg of DCA/mL with a Tmax of 12 hours, whereas the bearded dragon that received 30 mg/kg, SC, had a Cmax of 5.07 μg of DCA/mL with a Tmax of 12 hours. In the preliminary experiment, plasma concentrations of DCA were < 1 μg/mL by 192 hours for the 15 mg/kg dose, but they remained > 1 μg/mL until 288 hours after the 30 mg/kg dose. Plasma DCA concentrations were undetectable for both animals at 28 days after administration. Because no adverse effects were observed with either dose, the higher dose was selected for the primary experiment.

Primary experiment—Mean plasma DCA concentrations were determined for each time point for both IM and SC administration (Figure 1). Pharmacokinetic parameters were calculated for both IM and SC administration (Table 1). One bearded dragon had a measurable plasma DCA concentration (1.23 μg/mL) at the time of CCFA injection in phase 2 (ie, after the 28-day washout period). This was confirmed by a second analysis of the sample. This animal had received SC administration of CCFA in phase 1. To avoid bias, results for this bearded dragon were included in calculations for the SC administration but were excluded from all calculations for IM administration.

Figure 1—
Figure 1—

Mean ± SD plasma DCA concentration over time after IM (A) and SC (B) administration of CCFA (30 mg/kg) to healthy adult male bearded dragons (Pogona vitticeps). Values represent results for 6 bearded dragons after SC administration but only 5 bearded dragons after IM administration because 1 dragon had a measurable concentration of CCFA after the 28-day washout period following SC administration. Target MIC values of 1 μg/mL (dashed horizontal line) and 2 μg/mL (dotted horizontal line) are indicated.

Citation: American Journal of Veterinary Research 75, 5; 10.2460/ajvr.75.5.453

Table 1—

Values for noncompartmental pharmacokinetic parameters after administration of CCFA (30 mg/kg, IM or SC) to bearded dragons (Pogona vitticeps).

ParameterIMSC
Terminal half-life (h)70.8 ± 16.389.3 ± 46.0
Λz (1/h)0.0098 ± 0.00220.0078 ± 0.0035
Tm ax (h)18.7 ± 27.333.3 ± 27.3
Cmax (μg/mL)14.1 ± 5.910.4 ± 2.9
AUC0–∞ (h•μg/mL)1,891 ± 1,1911,701 ± 659
AUC0–last (h•μg/mL)1,699 ± 9681,441 ± 539
MRT (h)114.1 ± 28.9154.4 ± 42.9

Values reported are the mean and SD for 6 bearded dragons after SC administration but only 5 bearded dragons after IM administration because 1 dragon had a measurable concentration of CCFA after the 28-day washout period following SC administration. Values for terminal half-life are the harmonic mean and psuedo-SD.

AUC0–∞ = The AUC from time 0 to infinity. AUC0–last = The AUC from time 0 to the last measured concentration. Λz = Elimination rate constant. MRT = Mean residence time.

Mean plasma DCA concentrations exceeded the target MIC concentration of 1 μg/mL by 4 hours after CCFA administration in all bearded dragons for both routes of administration, and they remained above that target concentration for > 288 hours (Figure 1). Because a higher target MIC of 2 μg/mL was used in a previous pharmacokinetic study13 on ceftiofur in green iguanas, we also included a target concentration of 2 μg/mL in our evaluations. Mean plasma concentrations were maintained at > 2 μg/mL for > 192 hours but < 288 hours for both administration routes.

Although mean plasma DCA concentrations remained above the target MIC of 1 μg/mL for > 288 hours, some variation among individual bearded dragons was observed. The plasma concentration decreased to < 1 μg/mL between 192 and 288 hours after SC administration in 1 bearded dragon, between 192 and 288 hours after IM administration in 1 bearded dragon, and between 144 and 192 hours after IM administration in 1 bearded dragon. Additionally, as evidenced by differences in SD, there was less variation at all time points for SC administration than for IM administration. Thus, the SC route was considered to provide more consistent results.

Discussion

Analysis of the results of the present study confirmed that CCFA administration to bearded dragons provided plasma DCA concentrations > 1 μg/mL for > 288 hours (12 days) after both IM and SC administration in most animals. Although some individual variation was seen, SC administration provided less variation than did IM administration and thus would be preferred. No adverse reactions were observed after CCFA administration in any bearded dragon during the experiments. Furthermore, no problems were reported by owners during follow-up conversations.

Subjectively, the SC route of administration was technically easier for the investigators and less stressful for the bearded dragons than was the IM route. Given the small size of bearded dragons, it was challenging to administer a small volume of CCFA into the biceps brachii muscle, and that administration appeared to cause discomfort to the animals. Although larger muscle masses of the pelvic limbs or tail were available for CCFA administration, the investigators chose to avoid any possible rapid clearance by the reptilian renal portal system.14 Because of the ease of administration and subjectively less patient discomfort, we recommend SC rather than IM administration of CCFA to bearded dragons.

An important finding was the difference between results of the present study and those of a previous study8 conducted to evaluate CCFA pharmacokinetics in ball pythons. In that study,8 the only other study of CCFA administration to reptilians of which the authors are aware, investigators evaluated results for a single dose of 15 mg/kg, IM. In the ball pythons, mean ± SD Tmax was 2.17 ± 0.98 hours, and mean Cmax reached 7.096 ± 1.95 μg/mL. Plasma ceftiofur concentrations in the ball pythons remained above a target MIC of 0.1 μg/mL for at least 5 days and above a target MIC of 0.5 μg/mL for 20 hours.8 In contrast, in the present study of the administration of CCFA (30 mg/kg) to bearded dragons, mean ± SD Tmax was 21.6 ± 29.5 hours for IM administration, whereas mean Cmax reached 13.1 ± 6.0 μg/mL and mean plasma concentrations of DCA remained above a target MIC of 1 μg/mL for > 12 days. The disparate results obtained between the 2 studies are potentially attributable to a combination of several factors. First, there are substantial differences in methods between the 2 studies. In the study8 of ball pythons, only parent ceftiofur was measured in plasma samples, whereas in the study of bearded dragons reported here, the parent compound and all active metabolites were measured as DCA. This makes it difficult to directly compare results. Another factor is the difference in dose (15 vs 30 mg/kg). Finally, the results may have differed because of species-specific differences in ceftiofur metabolism. Drug dosages are often extrapolated from one reptile species or even taxonomic order to another, but even careful allometric scaling based on similar species can result in unpredictable pharmacokinetic profiles and drug toxicoses.15,16 Thus, the possibility of a species-specific difference in CCFA metabolism between ball pythons and bearded dragons must be considered and represents an area in which future research is needed.

Ceftiofur is a third-generation cephalosporin with broad-spectrum bactericidal activity against a variety of gram-positive, gram-negative, and anaerobic bacteria.1 In a study17 conducted to investigate antimicrobial susceptibility of 231 species of enteric bacteria isolated from turkey poults, ceftiofur was 1 of the 3 most active antimicrobials. Similarly, in a study18 of bacterial isolates obtained from ducks with air sacculitis and septicemia, ceftiofur was 1 of the 2 most active compounds. Major organisms of importance with moderate to high susceptibility to ceftiofur in birds include most Streptococcus spp, coagulase-positive Staphylococcus spp, and Escherichia coli as well as Enterobacter spp, Salmonella spp, Klebsiella spp, Serratia spp, and Proteus spp.6,17,18 For all of these organisms, the concentration of ceftiofur required to inhibit growth of 50% of isolates (ie, MIC50) is generally ≤ 1 μg/mL and the concentration required to inhibit growth of 90% of isolates (ie, MIC90) is ≤ 2 μg/mL for most. Important organisms considered resistant to ceftiofur include Pseudomonas spp and Enterococcus spp, for which isolates from birds have consistently had an MIC90 > 8 μg/mL.

A paucity of studies of the antimicrobial susceptibility of bacterial isolates from reptiles exist, and few have included ceftiofur as one of the antimicrobials evaluated. In 1 study,19 investigators evaluated the antimicrobial susceptibility of Devriesea agamarum, a common bacterial organism that causes dermatitis in lizards. By use of isolates from naturally infected lizards of several species, Devriesea organisms had an MIC90 of 0.12 μg/mL for ceftiofur sodium. Both experimentally infected bearded dragons and naturally infected Uromastyx lizards with skin infections attributable to Devriesea spp were successfully treated with ceftiofur sodium in that study.19 However, the use of long-acting CCFA in the lizards of that study19 was not evaluated. Aeromonas hydrophila isolated from a septicemic Nile crocodile (Crocodylus niloticus) during postmortem examination was susceptible to ceftiofur, but an MIC was not reported.20 Investigators in 2 large-scale studies21,22 of antimicrobial susceptibility for bacterial isolates from reptiles did not specifically test for susceptibility to ceftiofur.

Desfuroylceftiofur, an active metabolite of ceftiofur, has a similar (although not identical) susceptibility profile to that of the parent compound in vitro.1,23 The susceptibility profile for Staphylococcus organisms, in particular, differs between the 2 compounds (ceftiofur is considerably more active against Staphylococcus spp than is its metabolite desfuroylceftiofur).1,23 However, it is not known whether bearded dragons, or reptiles in general, have the same rapid metabolism of ceftiofur into desfuroylceftiofur as mammals. Indeed, a study24 conducted to compare the metabolism of the antimicrobial marbofloxacin in ball pythons and blue-and-gold macaws (Ara ararauna) revealed distinct differences in metabolite conversion between the 2 species as well as between the macaws and domestic chickens. Thus, until additional research into ceftiofur metabolism in reptiles is conducted, clinicians should be cautious when using CCFA to treat Staphylococcus infections in reptiles because it is unknown whether ceftiofur or desfuroylceftiofur will be the predominant antimicrobial compound present in the systemic circulation or in tissues.

Ceftiofur, similar to other β-lactam antimicrobials, is a time-dependent, bactericidal antimicrobial.25 The recommended dosing regimen for a time-dependent antimicrobial would be expected to maintain plasma drug concentrations above a target MIC throughout most or all of the dosing interval. There remains some question as to the exact portion of the dosing interval that must be above the MIC to ensure the most efficacious treatment. Even in critically ill humans with bacterial infections, there is a discrepancy in results when comparing continuous infusion and intermittent administration of β-lactam antimicrobials.26 Additionally, for a long-acting, time-dependent antimicrobial such as CCFA, it is not considered important to have concentrations that greatly exceed the target MIC because this will not improve the bactericidal effect.25 In the study reported here, plasma DCA concentrations after IM and SC administration of 30 mg/kg far exceeded the target MIC for a substantial portion of the 12-day period. Thus, it is possible that a lower dose (ie, between 15 and 30 mg/kg) may prolong the time the concentration exceeds the target MIC but not result in concentrations that greatly exceed the target MIC.

Not all target pathogens will have an MIC < 1 μg/mL for ceftiofur. Thus, when treating an infection with a higher MIC as determined by the results of antimicrobial susceptibility testing, the extended duration of high plasma concentrations of CCFA (> 1 to 2 μg/mL) detected in the present study would be advantageous. Further clinical trials that include the use of various dosing regimens would be required to determine the lowest effective, long-acting dose of CCFA in bearded dragons. However, the dose of CCFA (30 mg/kg) appeared to be safe and effective for the duration of the present study.

Ceftazidime is an antimicrobial commonly used for a variety of bacterial diseases in reptiles. Similar to CCFA, ceftazidime is a third-generation cephalosporin, but its spectrum differs from most others in this class because of high activity against Pseudomonas spp and low activity against gram-positive organisms.27 In 2 large-scale studies21,22 conducted to investigate common bacterial isolates from reptiles, Pseudomonas spp were frequently isolated from all orders of reptiles. Because Pseudomonas spp are considered resistant to ceftiofur, ceftazidime would be a superior antimicrobial choice to CCFA for use in patients from which Pseudomonas spp are isolated. These results highlight the need for bacterial culture and susceptibility testing so antimicrobial treatment can be tailored to the target microorganisms. If it is not possible to perform susceptibility testing or antimicrobial susceptibility results are not yet available, clinical judgment based on history and clinical signs should guide the initial treatment choice. In the experience of one of the authors (SMC), veterinary clinics with a low reptilian caseload are more likely to maintain a supply of CCFA than ceftazidime; thus, it may be an excellent first option. Additionally, for convenience and reduced animal stress, it appears that a single dose of CCFA may suffice for treatment of uncomplicated infections, in contrast to the more frequent dosing interval (every 72 hours) recommended for ceftazidime. This extended duration of efficacy makes CCFA a more desirable antimicrobial for use with aggressive or venomous animals, in situations in which handlers are unable to administer injections, or when an animal is unlikely to be available for follow-up evaluation.

The present study had several important limitations, including a small study population, lack of data on administration of multiple doses, lack of species-specific data on the effect of storage of frozen samples on stability of measured compounds, and lack of CBCs and biochemical analyses before and after treatment to ensure patient health and safety. The small study population and lack of data after administration of multiple doses of CCFA could be addressed in subsequent larger-scale studies. It was not possible to obtain species-specific data on stability during freezer storage during this study because of limitations in funding as well as logistical challenges associated with safely collecting an increased quantity of blood from these small animals. Because this stability testing was not performed, we extrapolated from data in other species (eg, guineafowl and wobbegong shark) and assumed that plasma ceftiofur concentrations would not be substantially altered during storage at −70° or −80°C for up to 6 weeks. However, because that was an untested assumption for plasma samples obtained from bearded dragons, it is possible that the results reported here may not accurately reflect plasma concentrations of ceftiofur in bearded dragons. Finally, the lack of CBCs and biochemical analyses before and after treatment is of particular importance because it is unknown whether any animals in this study had occult renal disease, which could have altered the metabolism of ceftiofur. However, in follow-up conversations with owners, all bearded dragons used in this study were alive and apparently healthy. Future studies of CCFA in reptiles should include evaluation of organ function if possible.

Analysis of the results of the present study suggested that a single dose of CCFA (30 mg/kg, SC) may be used to treat infections susceptible to ceftiofur in bearded dragons. One dose provided mean plasma concentrations of ceftiofur and its active metabolites, measured as DCA, at or above a target MIC of 1 μg/mL for > 288 hours. For chronic, complicated, or severe infections susceptible to ceftiofur, multiple doses of CCFA administered at an interval of 10 to 12 days may be clinically useful, but multiple-dose pharmacokinetic trials would be required before this recommendation can be made. Further research is needed to determine specific details regarding reptilian metabolism of ceftiofur and to investigate the activity of ceftiofur and its metabolites against reptile-specific bacterial isolates. Studies of CCFA pharmacokinetics in other reptile species as well as the use of CCFA in clinically ill reptiles are also needed.

ABBREVIATIONS

AUC

Area under the plasma concentration–time curve

CCFA

Ceftiofur crystalline-free acid

Cmax

Maximum plasma concentration

DCA

Desfuroylceftiofur acetamide

MIC

Minimum inhibitory concentration

TFA

Trifluoracetic acid

Tmax

Time to maximum plasma concentration

a.

Excede, Pharmacia and Upjohn Co, New York, NY.

b.

2695 Separation module, Waters, Milford, Mass.

c.

2487 Ultraviolet detector, Waters, Milford, Mass.

d.

Symmetry separation column, Waters, Milford, Mass.

e.

Symmetry guard column, Waters, Milford, Mass.

f.

Oasis HLB column, Waters, Milford, Mass.

g.

WinNONLIN, version 5.3, Pharsight Corp, Mountain View, Calif.

References

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