Michele Miller

Tuberculosis in elephants: antibody responses to defined antigens of Mycobacterium tuberculosis, potential for early diagnosis, and monitoring of treatment.

ABSTRACT Tuberculosis (TB) in elephants is a re-emerging zoonotic disease caused primarily by Mycobacterium tuberculosis. Current diagnosis relies on trunk wash culture, the only officially recognized test, which has serious limitations. Innovative and efficient diagnostic methods are urgently needed. Rapid identification of infected animals is a crucial prerequisite for more effective control of TB, as early diagnosis allows timely initiation of chemotherapy. Serology has diagnostic potential, although key antigens have not been identified and optimal immunoassay formats are not established. To characterize the humoral responses in elephant TB, we tested 143 serum samples collected from 15 elephants over time. These included 48 samples from five culture-confirmed TB cases, of which four were in Asian elephants infected with M. tuberculosis and one was in an African elephant with Mycobacterium bovis. Multiantigen print immunoassay (MAPIA) employing a panel of 12 defined antigens was used to identify serologic correlates of active disease. ESAT-6 was the immunodominant antigen recognized in elephant TB. Serum immunoglobulin G antibodies to ESAT-6 and other proteins were detected up to 3.5 years prior to culture of M. tuberculosis from trunk washes. Antibody levels to certain antigens gradually decreased in response to antitubercular therapy, suggesting the possibility of treatment monitoring. In addition to MAPIA, serum samples were evaluated with a recently developed rapid test (RT) based on lateral flow technology (ElephantTB STAT-PAK). Similarly to MAPIA, infected elephants were identified using the RT up to 4 years prior to positive culture. These findings demonstrate the potential for TB surveillance and treatment monitoring using the RT and MAPIA, respectively.

Highly accurate antibody assays for early and rapid detection of tuberculosis in African and Asian elephants.

ABSTRACT Tuberculosis (TB) in elephants is a reemerging zoonotic disease caused primarily by Mycobacterium tuberculosis. Current methods for screening and diagnosis rely on trunk wash culture, which has serious limitations due to low test sensitivity, slow turnaround time, and variable sample quality. Innovative and more efficient diagnostic tools are urgently needed. We describe three novel serologic techniques, the ElephantTB Stat-Pak kit, multiantigen print immunoassay, and dual-path platform VetTB test, for rapid antibody detection in elephants. The study was performed with serum samples from 236 captive African and Asian elephants from 53 different locations in the United States and Europe. The elephants were divided into three groups based on disease status and history of exposure: (i) 26 animals with culture-confirmed TB due to M. tuberculosis or Mycobacterium bovis, (ii) 63 exposed elephants from known-infected herds that had never produced a culture-positive result from trunk wash samples, and (iii) 147 elephants without clinical symptoms suggestive of TB, with consistently negative trunk wash culture results, and with no history of potential exposure to TB in the past 5 years. Elephants with culture-confirmed TB and a proportion of exposed but trunk wash culture-negative elephants produced robust antibody responses to multiple antigens of M. tuberculosis, with seroconversions detectable years before TB-positive cultures were obtained from trunk wash specimens. ESAT-6 and CFP10 proteins were immunodominant antigens recognized by elephant antibodies during disease. The serologic assays demonstrated 100% sensitivity and 95 to 100% specificity. Rapid and accurate antibody tests to identify infected elephants will likely allow earlier and more efficient treatment, thus limiting transmission of infection to other susceptible animals and to humans.

Two cases of atypical mycobacteriosis caused by Mycobacterium szulgai associated with mortality in captive African elephants (Loxodonta africana).

Abstract Mycobacterium szulgai was associated with mortality in two captive African elephants (Loxodonta africana) housed at Lincoln Park Zoo. The first elephant presented with severe, acute lameness of the left rear limb. Despite extensive treatments, the animal collapsed and died 13 mo after initial presentation. Necropsy revealed osteomyelitis with loss of the femoral head and acetabulum and pulmonary granulomas with intralesional M. szulgai. The second elephant collapsed during transport to another institution with no premonitory clinical signs. This animal was euthanized because of prolonged recumbency. Granulomatous pneumonia with intralesional M. szulgai was found at necropsy. Two novel immunoassays performed on banked serum samples detected antibody responses to mycobacterial antigens in both infected elephants. It was not possible to determine when the infection was established or how the elephants were infected. When reviewing the epidemiology of this organism in humans, however, transmission between elephants seemed unlikely because human-to-human transmission of this organism has never been reported and a third elephant in the herd was not affected. In addition to Mycobacterium bovis and Mycobacterium tuberculosis, atypical mycobacterial organisms need to be considered potentially pathogenic in elephants.

IMPLICATIONS OF SIMIAN RETROVIRUSES FOR CAPTIVE PRIMATE POPULATION MANAGEMENT AND THE OCCUPATIONAL SAFETY OF PRIMATE HANDLERS

Abstract Nonhuman primates can be naturally infected with a plethora of viruses with zoonotic potential, including retroviruses. These simian viruses present risks to both captive nonhuman primate populations and persons exposed to nonhuman primates. Simian retroviruses, including simian immunodeficiency virus, simian type D retrovirus, simian T-lymphotropic virus, and gibbon ape leukemia virus, have been shown to cause clinical disease in nonhuman primates. In contrast, simian foamy virus, a retrovirus that is highly prevalent in most nonhuman primates, has not been associated with clinical disease in naturally infected primates. Although it has been shown that human retrovirus infections with human T-lymphotropic virus and human immunodeficiency virus originated through multiple independent introductions of simian retroviruses into human populations that then spread globally, little is known about the frequency of such zoonotic events. In this article, exogenous simian retroviruses are reviewed as a concern for zoo and wildlife veterinarians, primate handlers, other persons in direct contact with nonhuman primates, and other nonhuman primates in a collection. The health implications for individual animals as well as managed populations in zoos and research institutions are discussed, the cross-species transmission and zoonotic disease potential of simian retroviruses are described, and suggestions for working safely with nonhuman primates are provided.

One Health in the shrinking world: Experiences with tuberculosis at the human–livestock–wildlife interface

Tuberculosis (TB) is a global anthropozoonotic infection that has raised awareness of the impact of disease at the human-livestock-wildlife interface. There are examples of transmission from livestock resulting in establishment of reservoirs in wildlife populations, and exposures from interactions between humans and wildlife that have resulted in disease outbreaks. A One Health approach is crucial to managing and protecting the health of humans, livestock, wildlife and the environment. Although still in its infancy in many areas of the world, the use of transdisciplinary teams to address wildlife-human-livestock boundary diseases will broaden the scope of options for solutions. This paper reviews some less commonly known examples of threats and outcomes using lessons learned from tuberculosis.

Field Application of Serodiagnostics To Identify Elephants with Tuberculosis prior to Case Confirmation by Culture

ABSTRACT Three serologic methods for antibody detection in elephant tuberculosis (TB), the multiantigen print immunoassay (MAPIA), ElephantTB STAT-PAK kit, and DPP VetTB test, were evaluated using serial serum samples from 14 captive elephants infected with Mycobacterium tuberculosis in 5 countries. In all cases, serological testing was performed prior to the diagnosis of TB by mycobacterial culture of trunk wash or tissue samples collected at necropsy. All elephants produced antibody responses to M. tuberculosis antigens, with 13/14 recognizing ESAT-6 and/or CFP10 proteins. The findings supported the high serodiagnostic test accuracy in detecting infections months to years before M. tuberculosis could be isolated from elephants. The MAPIA and/or DPP VetTB assay demonstrated the potential for monitoring antimycobacterial therapy and predicting TB relapse in treated elephants when continuously used in the posttreatment period. History of exposure to TB and past treatment information should be taken into consideration for proper interpretation of the antibody test results. Data suggest that the more frequent trunk wash culture testing of seropositive elephants may enhance the efficiency of the TB diagnostic algorithm, leading to earlier treatment with improved outcomes.

USE OF BUTORPHANOL DURING IMMOBILIZATION OF FREE-RANGING WHITE RHINOCEROS (CERATOTHERIUM SIMUM)

Abstract:  Forty free-ranging white rhinoceros (Ceratotherium simum) were anesthetized with etorphine, azaperone, and hyaluronidase in Kruger National Park, South Africa, between February and August 2009. Eighteen rhinoceros received butorphanol in the dart combination, and 22 rhinoceros had butorphanol administered intravenously within 15 min of darting. Body position, blood gas values, heart rate, respiratory rate, and temperature were measured at two time points after darting, approximately 10 min apart (sample 1 mean collection time after darting, 9.4 ± 2.7 min; sample 2 mean collection time, 18.6 ± 2.8 min). A significant number of field-captured rhinoceros remained standing at the first sample period when butorphanol was administered in the dart. Higher median values for arterial partial pressure of oxygen (PaO2) in combination with lower arterial partial pressure of carbon dioxide (PaCO2) in standing versus recumbent rhinoceros suggested improved ventilation in this posture (P < 0.05). When the effect of time, body position, and age was controlled, median values for respiratory rate, lactate, and pH were better in rhinoceros that received butorphanol in the dart (P < 0.05). There was also a trend toward higher median values for SO2 and bicarbonate in rhinoceros receiving butorphanol in the dart. Intravenous administration of butorphanol resulted in significantly decreased median PaCO2 and heart rate in recumbent rhinoceros (P < 0.05) without changes in PaO2 between sample periods 1 and 2. However, rhinoceros remained hypoxemic during the short anesthetic procedure despite butorphanol administration. Preliminary results suggest that administration of butorphanol (either in the dart or intravenously) improves some metabolic parameters in free-ranging recumbent white rhinoceros without significantly affecting ventilation. It is hypothesized that this may be due to a lighter state of immobilization. Addition of butorphanol to the dart provides handling and physiologic advantages because the majority of rhinoceros remain standing.

Agreement between assays of cell-mediated immunity utilizing Mycobacterium bovis-specific antigens for the diagnosis of tuberculosis in African buffaloes (Syncerus caffer).

We assessed the use of Mycobacterium bovis-specific peptides for the diagnosis of tuberculosis in African buffaloes (Syncerus caffer) by evaluating the agreement between the single intradermal comparative tuberculin test (SICTT), the Bovigam(®) EC (BEC) assay, the Bovigam(®) HP (BHP) assay and two assays utilizing the QuantiFERON(®) TB-Gold (in tube) system employing 20 h (mQFT20 assay) and 30 h (mQFT30 assay) whole blood incubation periods. Of 84 buffaloes, 45% were SICTT-positive, 48% were BEC-positive, 50% were BHP-positive, 37% were mQFT20-positive and 43% were mQFT30-positive. Agreement between the BEC and BHP Bovigam(®) assays was high (κ=0.86, 95% CI 0.75-0.97) and these detected the most test-positive animals suggesting that they were the most sensitive assays. Interferon-gamma release was significantly greater in buffaloes that were test-positive for all tests than in animals with discordant but positive Bovigam(®) results. Agreement between the mQFT assays was equally high (κ=0.88, 95% CI 0.77-0.98); however, all buffaloes with discordant mQFT results (n=6) were mQFT30-positive/mQFT20-negative, including three confirmed M. bovis-infected animals, suggesting that the mQFT30 assay is the more sensitive of the two. Agreements between the two Bovigam(®) and two mQFT assays were moderate, suggesting that in its current format the mQFT assay is less sensitive than either the BEC or the BHP assays.

ANESTHETIC INDUCTION OF CAPTIVE TIGERS (PANTHERA TIGRIS) USING A MEDETOMIDINE-KETAMINE COMBINATION

Abstract Six adult female tigers (Panthera tigris) were anesthetized repeatedly for elective medical procedures using 3 mg medetomidine and 200 mg ketamine i.m. Inductions were rapid and smooth, although supplemental ketamine was needed for safe transport after induction in 6 of 17 procedures. Reversal of the medetomidine-induced sedation with 15 mg atipamezole i.m. 59–232 min after induction resulted in smooth, rapid recoveries.

Butorphanol with oxygen insufflation corrects etorphine-induced hypoxaemia in chemically immobilized white rhinoceros ( Ceratotherium simum )

BackgroundOpioid-induced immobilization is associated with severe respiratory depression in the white rhinoceros. We evaluated the efficacy of butorphanol and oxygen insufflation in alleviating opioid-induced respiratory depression in eight boma-managed rhinoceros.ResultsChemical immobilization with etorphine, azaperone and hyaluronidase, as per standard procedure for the white rhinoceros, caused severe respiratory depression with hypoxaemia (PaO2 = 27 ± 7 mmHg [mean ± SD]), hypercapnia (PaCO2 = 82 ± 6 mmHg) and acidosis (pH =7.26 ± 0.02) in the control trial at 5 min. Compared to pre-intervention values, butorphanol administration (without oxygen) improved the PaO2 (60 ± 3 mmHg, F(3,21) =151.9, p <0.001), PaCO2 (67 ± 4 mmHg, F(3,21) =22.57, p <0.001) and pH (7.31 ± 0.06, F(3,21) =27.60, p <0.001), while oxygen insufflation alone exacerbated the hypercapnia (123 ± 20 mmHg, F(3,21) =50.13, p <0.001) and acidosis (7.12 ± 0.07, F(3,21) =110.6, p <0.001). Surprisingly, butorphanol combined with oxygen fully corrected the opioid-induced hypoxaemia (PaO2 = 155 ± 53 mmHg) and reduced the hypercapnia over the whole immobilization period (p <0.05, areas under the curves) compared to the control trial. However, this intervention (butorphanol + oxygen) did not have any effect on the arterial pH.ConclusionsOxygen insufflation combined with a single intravenous dose of butorphanol improved the immobilization quality of boma-managed white rhinoceros by correcting the opioid-induced hypoxaemia, but did not completely reverse all components of respiratory depression. The efficacy of this intervention in reducing respiratory depression in field-captured animals remains to be determined.