Ramiro Isaza

EPIDEMIOLOGY AND DIAGNOSIS OF MYCOBACTERIUM TUBERCULOSIS IN CAPTIVE ASIAN ELEPHANTS (ELEPHAS MAXIMUS)

Abstract The deaths of two Asian elephants (Elephas maximus) in August 1996 led the United States Department of Agriculture to require the testing and treatment of elephants for tuberculosis. From August 1996 to September 1999, Mycobacterium tuberculosis infection was confirmed by culture in 12 of 118 elephants in six herds. Eight diagnoses were made antemortem on the basis of isolation of M. tuberculosis by culture of trunk wash samples; the remainder (including the initial two) were diagnosed postmortem. We present the case histories, epidemiologic characteristics, diagnostic test results, and therapeutic plans from these six herds. The intradermal tuberculin test, enzyme-linked immunosorbent assay serology, the blood tuberculosis test, and nucleic acid amplification and culture are compared as methods to diagnose M. tuberculosis infection in elephants.

CONCEPTS AND ISSUES WITH INTERSPECIES SCALING IN ZOOLOGICAL PHARMACOLOGY

Abstract Zoologic medicine practitioners take approved agents (veterinary or human) and extrapolate their use to nonapproved species. The decision on dose, duration, and interval is often made with limited species-specific pharmacokinetic information. Because of the monetary value of these animals or their status as endangered species, this method of “trial and error” for therapeutic dosage selection is inappropriate. In zoologic medicine, various methods have been used in an attempt to extrapolate or predict safe and effective dosage regimens. The simplest and typical method of extrapolating a dosage to a nondomestic species is to use a mg/kg dose established for another domestic species or humans. However, this calculation results in a linear increase in the amount of drug administered as body weight increases. Although common, this method tends to overdose large animals and underdose small animals. The second method is similar, except that it takes the approved dose in a specific species and makes an additional assumption that links the dosage to a physiologic function or anatomic feature. Examples are the use of basal metabolic rate or body-surface area as the basis for dosage extrapolation. Allometric scaling of pharmacokinetic parameters is the final method of dosage extrapolation between species. This is commonly used in the pharmaceutical industry to establish the first dosage in human drug investigations. Adaptation of this method for zoologic medicine may enhance our ability to estimate therapeutic dosages for nondomestic species. This review discusses and compares these three methods for dosage selection and provides examples of extrapolation from the literature.

Proliferative osteoarthritis and osteoarthrosis in 15 snakes.

Abstract Fifteen snakes representing seven species with segmental, proliferative osteoarthritis and osteoarthrosis of the spine were presented for examination. All the snakes were captive, privately owned, and fed domestic rodents. Physical examination, radiography, blood culture, bone culture, necropsy, and histopathology were performed on each snake. All the snakes had similar physical examination, radiologic, and necropsy findings. There were three histologic types of lesions: active bacterial osteoarthritis, predominantly noninflammatory osteoarthrosis with multifocal inflammation suggestive of chronic bacterial osteoarthritis, and noninflammatory lesions consistent with osteoarthrosis without evidence of inflammation or bacteria. These findings suggest that all these snakes represent a single disease process, bacterial infection of the vertebrae. The different histologic lesions observed in these snakes may be a continuum of lesions, from acute to chronic. Gram-negative bacteria were isolated from the blood or bone lesions of 8 of the 15 snakes. In six of these eight snakes, Salmonella species were isolated. Gram-positive bacteria (Streptococcus sp.) were isolated from two other snakes. Blood and bone culture results were well correlated, so blood culture may be effective for detecting active bacterial osteoarthritis.

Simultaneous extraction and quantitation of fentanyl and norfentanyl from primate plasma with LC/MS detection.

The quantitation of both fentanyl and its desalkyl metabolite, norfentanyl, in plasma using LC/MS has not been previously described. The detection and quantitation of fentanyl and norfentanyl was achieved using LC/MS detection. The liquid-liquid extraction used toluene as the organic phase. Chromatography was carried out using a Zirchrom-PBD (50 mm x 2.1 mm, 3 microm) column with a mobile phase of acetonitrile-ammonium acetate (10 mM), citrate (0.1 mM, pH 4.4) (45:55, v/v) with a flow rate of 0.3 ml/min. Mass spectroscopy detection was performed using ESI in the positive mode. The LOQ for fentanyl was 25 pg/ml and norfentanyl was 50 pg/ml. For the concentrations of 75, 250, and 750 pg/ml, respectively, fentanyl had inter-day precisions of 6.6, 7.2, and 6.6% with accuracies of 4.0, 5.1, and 5.1% and intra-day precisions of 1.6, 1.9, and 1.9% with accuracies of 11.6, 9.4, and 8.4%, and norfentanyl had inter-day precisions of 7.4, 0.3, and 0.7% with accuracies of 9.1, 8.8, and 12.3% and intra-day precisions of 5.3, 1.4, and 0.1% with accuracies of 10.9, 8.9, and 12.8%. The recoveries of fentanyl were 85, 92, and 75% and of norfentanyl were 40, 49, and 46% at the 75, 250, and 750 pg/ml concentrations, respectively.

Halicephalobus gingivalis (Nematoda) infection in a Grevy's zebra (Equus grevyi).

Abstract A 6-yr-old female Grevys zebra (Equus grevyi) with a disseminated rhabditiform nematode infection is described. Antemortem clinical signs were limited to blindness and abnormal behavior believed to be caused by a recurrent nematode-induced uveitis. Histologic examination of the kidneys, heart, eyes, uterus, and lymph nodes revealed granulomas containing multiple sections of rhabditiform nematodes. Most of the recovered nematodes were larval stages with only a few adult females noted. The adults measured 243–297 µm × 11–16 µm (x̄ = 269 × 14 µm). The distinctive rhabditiform esophagi had corpus:isthmus:bulb proportions of 19:11:5. On the basis of adult morphology, the nematode was identified as Halicephalobus gingivalis. This is the first report of this parasite in a zebra and indicates that this parasitic granulomatous disease should be considered in zebras with neurologic disease.

USE OF FAMCICLOVIR FOR THE TREATMENT OF ENDOTHELIOTROPHIC HERPESVIRUS INFECTIONS IN ASIAN ELEPHANTS (ELEPHAS MAXIMUS)

Abstract Two juvenile Asian elephants (Elephas maximus) presented with an acute onset of facial edema and lethargy. Examination of the oral cavity of each animal revealed cyanosis of the tip and distal margins of the tongue suggestive of endothelial inclusion body disease (EIBD) of elephants. Whole-blood samples were obtained, and polymerase chain reaction tests confirmed the presence of elephant herpesvirus. The animals were administered famciclovir (Famvir, SmithKline Beecham Pharmaceuticals, Philadelphia, Pennsylvania 19101, USA), a potent human anti-herpesvirus drug, in the course of their disease, and recovery followed a treatment regime of 3–4 wk. These are the first known cases of elephants surviving EIBD.

Evaluation of a multiple-antigen enzyme-linked immunosorbent assay for detection of Mycobacterium tuberculosis infection in captive elephants.

Abstract Mycobacterium tuberculosis has become an important agent of disease in the captive elephant population of the United States, although current detection methods appear to be inadequate for effective disease management. This investigation sought to validate a multiple-antigen enzyme-linked immunosorbent assay (ELISA) for screening of M. tuberculosis infection in captive elephants and to document the elephants serologic response over time using a cross-sectional observational study design. Serum samples were collected from 51 Asian elephants (Elephas maximus) and 26 African elephants (Loxodonta africana) from 16 zoos and circuses throughout the United States. Infection status of each animal was determined by mycobacterial culture of trunk washes. Reactivity of each serum sample against six antigens was determined, and the linear combination of antigens that accurately predicted the infection status of the greatest number of animals was determined by discriminant analysis. The resulting classification functions were used to calculate the percentage of animals that were correctly classified (i.e., specificity and sensitivity). Of the 77 elephants sampled, 47 fit the criteria for inclusion in discriminant analysis. Of these, seven Asian elephants were considered infected; 25 Asian elephants and 15 African elephants were considered noninfected. The remaining elephants had been exposed to one or more infected animals. The specificity and sensitivity of the multiple-antigen ELISA were both 100% (91.9–100% and 54.4–100%, respectively) with 95% confidence intervals. Mycobacterium bovis culture filtrate showed the highest individual antigen specificity (95%; 83.0–100%) and sensitivity (100%; 54.4–100%). Serum samples from 34 elephants were analyzed over time by the response to the culture filtrate antigen; four of these elephants were culture positive and had been used to calculate the discriminant function. Limitations such as sample size, compromised ability to ascertain each animals true infection status, and absence of known-infected African elephants suggest that much additional research needs to be conducted regarding the use of this ELISA. However, the results indicate that this multiple-antigen ELISA would be a valuable screening test for detecting M. tuberculosis infection in elephant herds.

PRELIMINARY SINGLE-DOSE PHARMACOKINETICS OF MARBOFLOXACIN IN BALL PYTHONS (PYTHON REGIUS)

Abstract Pharmacokinetics of marbofloxacin in two male and four female adult ball pythons (Python regius) was determined after i.v. and p.o. administration of a single dose. Using a crossover design, each snake was given a single 10 mg/kg dose of marbofloxacin i.v. and p.o. Blood samples were collected prior to and 0.5, 1, 1.5, 3, 6, 12, and 24 hr after marbofloxacin administration. Marbofloxacin was quantitated by use of liquid chromatography-mass spectrometry. Following p.o. administration, marbofloxacin had a peak plasma concentration (Cmax) of 9.40 μg/ml and a time to Cmax (Tmax) of 9.0 hr. Based on the plasma pharmacokinetics generated in this study and pending any further studies to evaluate potential toxicity and multi-dose pharmacokinetics, we suggest a dosage for marbofloxacin in ball pythons of 10 mg/kg p.o. at least every 48 hr, depending on the sensitivity of the pathogen and as a basis for further research.

Moxidectin plasma concentrations following topical administration to llamas (Lama glama) and alpacas (Lama pacos)

Producers and veterinarians commonly administer pharmaceuticals labeled for cattle, sheep, or goats to llamas and alpacas, yet little is known about the safety and efficacy of this extra-label usage. There are many species-related factors, such as bioavailability and metabolism, that could lead to variation in the pharmacokinetic parameters between species and possibly decrease the activity of a compound. This experiment investigated the absorption and disposition of topical (pour-on) moxidectin in llamas and alpacas. Seven llamas and seven alpacas were obtained from commercial sources and were not treated with an avermectin/milbemycin agent within 60 days prior to the start of the study. The animals were housed outdoors with water and hay available ad libitum. All animals received moxidectin at a dose of 500g/kg after being clipped along the dorsal midline. Serial blood samples were collected post dose. Samples were analyzed for moxidectin using a validated liquid chromatography/mass spectroscopy assay. Moxidectin administration was well tolerated with no adverse effects noted in either species following topical administration. Median Cmax values were highly variable in the llamas and alpacas with two alpacas having no detectable plasma concentrations of moxidectin. Due to the limited absorption of this compound in both species, appropriate pharmacokinetic parameters could not be determined. In these two species of South American camelids, moxidectin was not well absorbed following topical administration.

Drug Delivery to Captive Asian Elephants - Treating Goliath

Captive Asian elephants have been maintained in captivity by humans for over 4000 years. Despite this association, there is little published literature on the treatment of elephant diseases or methods of drug administration to these animals. Elephants in captivity are generally healthy and require few therapeutic interventions over the course of their lifetime. However, when they become acutely ill, treatment becomes a serious issue. The successful and consistent administration of therapeutics to elephants is formidable in an animal that presents significant limitations in drug delivery options. The single most important factor in administering drugs to an elephant is the animals cooperation in accepting the medication. Working around elephants can be very dangerous and this is magnified when working around sick or injured animals where the elephant is subject to increased stress, pain, and unusual situations associated with treatment. The large body size of the Asian elephant produces a separate set of issues. In this paper, methods of drug administration and their associated limitations will be reviewed. Considerations of medicating such large animals can serve to highlight the problems and principles of treatment that are inherent in these species.