HELICOBACTER SCREENING OF GRAND CAYMAN BLUE IGUANA (CYCLURA LEWISI) AND NORTH ANTILLEAN SLIDER (TRACHEMYS DECUSSATA ANGUSTA) ON GRAND CAYMAN, CAYMAN ISLANDS.

The endemic Grand Cayman or blue iguana (Cyclura lewisi) is endangered. Beginning in 2015 significant morbidity and mortality occurred in captive and wild blue iguanas within Grand Cayman's Queen Elizabeth II Botanic Park (QEIIBP). Investigation identified a novel Helicobacter sp., provisionally named Helicobacter sp. Grand Cayman Blue Iguana 1 (GCBI1), as the cause. Invasive green iguanas (Iguana iguana) are believed to play a role in GCBI1 transmission to the blue iguana; however, the origin and transmission pathways have not been determined. To assess the likelihood of blue iguanas asymptomatically harboring GCBI1, in May 2022 population-level screening of captive blue iguanas at QEIIBP was conducted on half (n = 102) of the captive blue iguana population (n = 201) including half of each age class. Helicobacter sp. GCBI1 is closely related to a chelonian Helicobacter sp. and 10 sympatric wild north Antillean sliders (Trachemys decussata angusta) were sampled in October 2019. Combined choana/cloacal swabs were screened by a GCBI1-specific quantitative polymerase chain reaction (qPCR) assay. All samples were negative, suggesting that GCBI1 is not present asymptomatically in the captive blue iguana population or in north Antillean sliders. These results provide support for the hypothesis that GCBI1 is periodically introduced to captive and wild blue iguanas from another species or source.

HEMATOLOGIC AND BIOCHEMICAL VALUES OF FREE-RANGING HAWKSBILL SEA TURTLES (ERETMOCHELYS IMBRICATA) IN GLOVER'S REEF, BELIZE.

Blood samples were obtained from the dorsal cervical sinus of free-ranging hawksbill sea turtles (Eretmochelys imbricata) collected at Glover's Reef Marine Reserve, 42 km east of the coast of Belize, for hematology and plasma biochemistry analysis. Unknown sex, subadult turtles (N = 32) were sampled in 2013 (n = 22) and 2017 (n = 10). To provide a more robust data set, parameters that did not have statistically significant differences were pooled and treated as a single population. Eleven hematologic parameters were evaluated; of these, five were pooled. Twenty-three plasma biochemical parameters were evaluated; of these, 15 were pooled. The PCV observed in this study (mean 33.44%) was double that observed in two studies of juvenile hawksbills in Dubai (means 17% and 16%), whereas the total WBC count was half that observed in immature and adult hawksbills sampled in the Galápagos (mean 2.91 × 103 versus 5.3 × 103/µl). Total protein and albumin were lower than regionally similar, adult female hawksbills in Brazil (means 3.36 versus 5.45 g/dl and 0.93 versus 2.11 g/dl, respectively). Globulins were higher (mean 2.43 versus 1.06 and 0.5 g/dl), driving the albumin:globulin ratio lower than that observed in two studies of juvenile hawksbills in Dubai (0.4:1 versus 1.1:1 and 1:1, respectively). These findings represent a geographically distinct population from previous reports, highlight the variability in blood parameters from disparate populations, and reaffirm the vital importance of considering a multitude of variables when interpreting reptilian blood values. The similarities in the majority of values observed in 2013 and 2017 provide confidence in the stability of these parameters in this population.

GLOBAL RETROSPECTIVE REVIEW OF SEVERE ACUTE RESPIRATORY SYNDROME SARS COV-2 INFECTIONS IN NONDOMESTIC FELIDS: MARCH 2020-FEBRUARY 2021.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in nondomestic felids have been documented in North America, South America, Africa, Europe, and Asia. Between March 2020 and February 2021, at nine institutions across three continents, infection was confirmed in 16 tigers (Panthera tigris), 14 lions (Panthera leo), three snow leopards (Panthera uncia), one cougar (Puma concolor), and one Amur leopard cat (Prionailurus bengalensis euptilurus) ranging from 2 to 21 yr old (average, 10 yr). Infection was suspected in an additional 12 tigers, 4 lions, and 9 cougars. Clinical signs (in order of most to least common) included coughing, ocular and/or nasal discharge, wheezing, sneezing, decreased appetite, lethargy, diarrhea, and vomiting. Most felids recovered uneventfully, but one geriatric tiger with comorbidities developed severe dyspnea and neurologic signs necessitating euthanasia. Clinical signs lasted 1-19 d (average, 8 d); one tiger was asymptomatic. Infection was confirmed by various methods, including antigen tests and/or polymerase chain reaction (PCR) of nasal or oral swabs, tracheal wash, and feces, or virus isolation from feces or tracheal wash. Infection status and resolution were determined by testing nasal swabs from awake animals, fecal PCR, and observation of clinical signs. Shedding of fecal viral RNA was significantly longer than duration of clinical signs. Postinfection seropositivity was confirmed by four institutions including 11 felids (5 lions, 6 tigers). In most instances, asymptomatic or presymptomatic keepers were the presumed or confirmed source of infection, although in some instances the infection source remains uncertain. Almost all infections occurred despite using cloth facemasks and disposable gloves when in proximity to the felids and during food preparation. Although transmission may have occurred during momentary lapses in personal protective equipment compliance, it seems probable that cloth masks are insufficient at preventing transmission of SARS-CoV-2 from humans to nondomestic felids. Surgical or higher grade masks may be warranted when working with nondomestic felids.

ASSESSMENT OF MULTIANTIGEN PRINT IMMUNOASSAY AND RAPID LATERAL-FLOW TEST FOR THE DETECTION OF MYCOBACTERIUM BOVIS INFECTION IN MALAYAN TAPIR (TAPIRUS INDICUS).

A multiantigen print immunoassay (MAPIA) and rapid test (RT) developed and validated for detection of mycobacterial antibodies in elephants (Elephas maximus and Loxodonta africana) was assessed in Malayan tapir (Tapirus indicus). Retrospective analysis of banked serum from one Mycobacterium bovis infected and seven presumably uninfected tapir was performed by MAPIA and RT. A sample collected 2 mon prior to the death of a culture-confirmed M. bovis-infected tapir served as a positive control. Seroreactivity of this sample was demonstrated via both MAPIA and RT testing. Seven uninfected animals, including four without postmortem evidence of mycobacterial disease and three that remain healthy, were negative controls; none demonstrated seroreactivity to key antigens with either test. These results suggest that MAPIA and RT have potential utility for rapid detection of M. bovis infection in Malayan tapir.

HEMATOLOGY, PLASMA BIOCHEMISTRY, AND PLASMA PROTEIN ELECTROPHORESIS REFERENCE INTERVALS FOR BLUE IGUANAS (CYCLURA LEWISI) FROM GRAND CAYMAN ISLAND.

The blue iguana (Cyclura lewisi) is an endangered rock iguana species native to Grand Cayman, in the Cayman Islands. Health assessments were conducted on captive and free-roaming iguanas in 2001 and 2003-2014 and were performed in the summer wet season (June-July) of 2003-2004 and 2010-2014 and in the winter dry season (November-December) of 2001 and 2005-2009. Morphometric data were recorded from iguanas when blood samples were collected: 903 samples were collected and data from 890 samples from 775 iguanas were included. Samples were analyzed for hematology, plasma biochemistry, protein electrophoresis, mineral panels, 25-hydroxyvitamin D, and testosterone. Reference intervals were created for captive subadults, captive adults, and free-roaming adults when data were sufficient. Significant differences among these groups were described, as were differences on the basis of sex, season, and origin (captive vs free-roaming). In captive iguanas, most analytes were significantly different between subadults and adults, mature heterophils and copper were significantly higher in the dry season, zinc levels were significantly higher in the wet season, and cholesterol and triglycerides were significantly higher in adult females than adult males. Testosterone in adult males was significantly higher in the dry season. These results will aid in future health assessments and disease investigations in wild and captive populations of blue iguanas and are of comparative value for other Cyclura species that are free-roaming, captive, and, especially, in similar conservation release programs.

PHARMACOKINETICS AND CLINICAL SAFETY OF A SUSTAINED-RELEASE FORMULATION OF CEFTIOFUR CRYSTALLINE FREE ACID IN RINGNECK DOVES (STREPTOPELIA RISORIA) AFTER A SINGLE INTRAMUSCULAR INJECTION.

Ceftiofur crystalline free acid (CCFA) is a third-generation, oil-based, cephalosporin antimicrobial marketed as a once weekly treatment in cattle and swine, and as a two-time dose with 10-day duration in horses. Because handling and restraint times can be reduced, long-acting antibiotic preparations are particularly useful for treatment of nondomestic species. This study evaluated the pharmacokinetics of CCFA in ringneck doves (Streptopelia risoria). A single intramuscular (IM) injection of CCFA at 50 mg/kg was administered to each of 30 doves, and blood was collected from subsamples of 6 birds at predetermined sampling times (i.e., with a postinjection range of 0.5 to 192 hr). All ringneck doves were scheduled for euthanasia because of reasons unrelated to the study; this was performed at the conclusion of the study; and complete postmortem and histopathologic examinations were performed. Plasma concentrations of CCFA remained above the minimum inhibitory concentration (1.0 µg/ml; observed for most avian pathogenic bacteria) for 108 hr. No abnormalities were identified on individual birds before and after clinical pathology results (i.e., hematocrits and plasma biochemistry profiles), and only minimal gross and histopathologic changes such as mild tissue inflammation at the injection site were observed. Based on these results, one IM injection of CCFA at 50 mg/kg seems to be a potential option for treatment of ringneck doves.

CEFOVECIN PROTEIN BINDING AS A PREDICTOR FOR EXTENDED DURATION OF ACTION: A REVIEW OF CURRENT LITERATURE AND IN VITRO ANALYSIS IN MULTIPLE ZOOLOGICAL SPECIES.

Cefovecin is a third-generation cephalosporin antibiotic with an efficacy of 2 wk following a single injection in domestic dogs and cats. A high degree of plasma protein binding to cefovecin has been proposed as one of the mechanisms responsible for the long elimination half-life, but protein binding has not been evaluated extensively in nondomestic species. In this study, a review of the current literature was conducted, and pharmacokinetic data were compiled for species in which cefovecin has been evaluated thus far. Additionally, in vitro cefovecin protein binding was evaluated in plasma from 22 nondomestic species representing a broad range of taxa. Animals of the order Carnivora demonstrated protein-binding levels of >98%, which is supportive of the long elimination half-life seen in related species. Protein binding was highly variable in Artiodactyl and Perissodactyl species, with dolphins (Tursiops truncatus) and barasingha (Rucervus duvaucelii) displaying high protein binding (99.12% to >99%), but not gazelles (Eudorcas thomsonii) or equids (91.76-92.70%). Cefovecin was not highly bound in any reptiles or birds, corresponding to short half-lives reported for these taxa. These results suggest that a high percentage of plasma protein binding in vitro may predict in which species cefovecin may exhibit a long half-life and duration of action in vivo. These findings may aid in selecting species for cefovecin pharmacokinetic research and for empirical treatment of infections caused by susceptible bacteria.

SARS-COV-2 INFECTION AND LONGITUDINAL FECAL SCREENING IN MALAYAN TIGERS (PANTHERA TIGRIS JACKSONI), AMUR TIGERS (PANTHERA TIGRIS ALTAICA ), AND AFRICAN LIONS (PANTHERA LEO KRUGERI) AT THE BRONX ZOO, NEW YORK, USA.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) emerged as the cause of a global pandemic in 2019-2020. In March 2020, New York City became the epicenter in the United States for the pandemic. On 27 March 2020, a Malayan tiger (Panthera tigris jacksoni) at the Bronx Zoo in New York City developed a cough and wheezing with subsequent inappetence. Over the next week, an additional Malayan tiger and two Amur tigers (Panthera tigris altaica) in the same building and three lions (Panthera leo krugeri) in a separate building also became ill. The index case was anesthetized for diagnostic workup. Physical examination and bloodwork results were unremarkable. Thoracic radiography and ultrasonography revealed a bronchial pattern with peribronchial cuffing and mild lung consolidation with alveolar-interstitial syndrome, respectively. SARS-CoV-2 RNA was identified by real-time, reverse transcriptase PCR (rRT-PCR) on oropharyngeal and nasal swabs and tracheal wash fluid. Cytologic examination of tracheal wash fluid revealed necrosis, and viral RNA was detected in necrotic cells by in situ hybridization, confirming virus-associated tissue damage. SARS-CoV-2 was isolated from the tracheal wash fluid of the index case, as well as the feces from one Amur tiger and one lion. Fecal viral RNA shedding was confirmed in all seven clinical cases and an asymptomatic Amur tiger. Respiratory signs abated within 1-5 days for most animals, although they persisted intermittently for 16 days in the index case. Fecal RNA shedding persisted for as long as 35 days beyond cessation of respiratory signs. This case series describes the clinical presentation, diagnostic evaluation, and management of tigers and lions infected with SARS-CoV-2 and describes the duration of viral RNA fecal shedding in these cases. This report documents the first known natural transmission of SARS-CoV-2 from humans to nondomestic felids.

BURMESE ROOFED TURTLE (BATAGUR TRIVITTATA) DISEASE SCREENING IN MYANMAR.

The Burmese roofed turtle (Batagur trivittata), a critically endangered freshwater turtle, is endemic to Myanmar. Once thought to be extinct, remnant wild populations were discovered in 2001 and limited captive individuals identified in pagoda ponds or confiscated from fishers in Myanmar. These and their offspring are maintained in five facilities in Myanmar and form the basis of a conservation program (habitat protection, captive breeding, nest protection, egg collection, head-starting, and release). Prerelease health screenings were performed in 2014 and 2018 at Yadanabon Zoological Gardens, a head-starting facility in Limpha Village, and Lawkanandar Wildlife Park. One hundred forty-three turtles were assessed (37 male, 50 female, 56 juveniles [too young to determine sex]; two females were assessed in both years), age range of 1 to 12 y (one unknown age adult founder), and body mass range of 0.111 to 32.72 kg. Health evaluations both years included physical examination and combined choanal/cloacal swab samples for polymerase chain reaction testing of the potential chelonian pathogens intranuclear coccidia, Mycoplasma, Herpesvirus, Ranavirus, and Adenovirus (not all tests performed each year). In 2018, cloacal swabs from 30 and 20 turtles at the Yadanabon Zoological Gardens and Lawkanandar Wildlife Park, respectively, were cultured for Salmonella. All turtles were assessed as healthy based on normal physical examination findings, and all had negative test results. Prerelease health screening, such as performed in this study, is an important component of release, reintroduction, and translocation projects to prevent introduction of novel pathogens into naïve wild populations.

PARASITES OF THE BLUE IGUANA (CYCLURA LEWISI) FROM GRAND CAYMAN ISLAND.

Feces (n = 226; 2004-2015) from healthy captive and wild blue iguanas (Cyclura lewisi) from Grand Cayman, Cayman Islands, were examined for endoparasites. Parasites identified included Nyctotherus sp. and Entamoeba sp. cysts and trophozoites, trichomonad trophozoites, and oxyurid and trichostrongylid eggs. Endoparasites from postmortem examinations (n = 13) included adult and larval nematodes: Ozolaimus megatyphlon, Ozolaimus monhystera, Alaeuris travassosi, Atractis mega, and an unidentified species of Oswaldocruzia. Entamoeba spp. were more likely in captive juveniles of both sexes than captive or wild adults of either sex; Entamoeba spp. were more likely in captive adult females than captive adult males; trichomonad trophozoites were more likely in adult captive and wild iguanas of both sexes than in captive juveniles of either sex; and Nyctotherus spp. were more likely in juvenile captive males than captive adult males or females and more likely in adult wild males than captive juvenile males. Trichostrongylid eggs were more likely in adult wild females than adult captive females and more likely in captive and wild adults of both sexes than in captive juveniles of both sexes. Oxyurid eggs were more likely in adult captive and wild iguanas of both sexes than captive juveniles of either sex. Blue iguanas have a variety of endoparasites regardless of age, sex, or captive vs wild status, with no type found exclusively in either captive or wild populations. Ectoparasites from wild adults included adult ticks (Amblyomma torrei) and a single adult mite (Hirstiella trombidiformis). All are new host records for this species and Grand Cayman. Knowledge of parasite status of captive and wild populations is important to evaluate the relative risk of introduction of captive animals into wild populations.

PREVALENCE, INCIDENCE, AND IDENTIFICATION OF SALMONELLA ENTERICA FROM WILD AND CAPTIVE GRAND CAYMAN IGUANAS ( CYCLURA LEWISI).

From 2005 to 2017, a total of 334 fresh fecal samples was obtained from 236 captive and free-ranging Grand Cayman iguanas ( Cyclura lewisi) in a captive breeding and release program. One-hundred-sixteen samples were obtained from individual captive iguanas living in enclosures with natural substrate, 161 samples from captive iguanas living in elevated wire-bottom enclosures, and 57 samples from free-ranging wild iguanas. These samples were cultured to isolate subspecies of Salmonella enterica. as part of prerelease health evaluations, and to determine baseline health assessments of captive and wild populations of these iguanas. There was a 5.45% (95% confidence interval [CI]: 1.14-15.12) prevalence of S. enterica in iguanas housed on natural substrate ( n = 51), 3.85% (CI: 1.42-8.19) in iguanas housed in elevated wire-bottom enclosures ( n = 157), and 6.06% (CI: 0.74-20.23) in wild free-ranging iguanas ( n = 38). These results demonstrate no significant difference ( P = 0.73) in S. enterica prevalence among these housing conditions. The incidence of S. enterica from 2005 to 2017 in the population sampled was 4.19% (CI 3.10-5.29). Ten different serotypes of S. enterica were isolated from 14 iguanas. Salmonella enterica ser. Saintpaul was the most frequent isolate. Annual S. enterica prevalence was calculated for iguanas housed in different enclosure types, for free-roaming wild iguanas, and for all cultured iguanas. The highest yearly prevalence was 23.08% (CI: 5.04-53.81) in iguanas tested in 2007 ( n = 21). No Salmonella enterica were cultured in 2005, 2008, 2009, 2010, 2011, 2012, and 2015. These results suggest that the shedding of S. enterica was not significantly different between housing types or between captive versus wild iguanas and therefore that release of captive iguanas did not significantly affect the health of the wild population or their exposure to S. enterica.

LONG-TERM SURVEILLANCE OF LANGUR ALPHAHERPESVIRUS IN A ZOO POPULATION OF SILVERED LANGURS ( TRACHYPITHECUS CRISTATUS).

Langur alphaherpesvirus (HVL), a provisionally named alphaherpesvirus in the Simplexvirus genus, was first identified in 1991 at the Bronx Zoo in wild-origin silvered langurs ( Trachypithecus cristatus) and their descendants. HVL is closely related to B virus ( Macacine alphaherpesvirus 1) based on serologic and genetic data, but its natural history and zoonotic potential remain unknown. A cohort study was undertaken to describe the epidemiology, clinical impact, and potential management implications of this virus in a naturally infected, zoo-based population of silvered langurs. Opportunistic surveillance sampling from 1991 through 2015 resulted in 235 serum samples and 225 mucosal swabs from 75 individuals. A total of 43 individuals (57.3%) were seropositive for HVL within this period. Seroprevalence increased significantly with age, and indirect evidence suggested a peak in transmission at the onset of sexual maturity. These findings were similar to the behavior of other simplexviruses in their adapted hosts. Yearly cumulative incidence declined significantly through the study period, with zero or one new case detected each year from 2007 through 2015. The density of this population decreased within the study period for management reasons unrelated to HVL infection, and a change in age distribution or less-frequent contacts may have contributed to low transmission. In addition, clinical signs of simplexvirus infection were rare, and virus isolation was negative on all mucosal swabs, suggesting that viral shedding was infrequent. Yearly period seroprevalence remained relatively constant with a median of 45.8%, likely because of the extended survival of infected individuals within the population. Maintenance of a naturally occurring, novel virus with unknown zoonotic potential in a zoo population for over 25 yr highlights the importance of biosecurity and biosafety for management of silvered langurs and all primate species.

DISEASE SCREENING IN SOUTHERN RIVER TERRAPINS ( BATAGUR AFFINIS EDWARDMOLLI) IN CAMBODIA.

Southern River terrapins ( Batagur affinis) are among the most critically endangered turtles in the world. To augment the Cambodia population, a head-start program was established for the endemic subspecies Batagur affinis edwardmolli in 2006, and in 2015, prerelease health assessments were performed on 70 subadults (hatch years, 2006-2011). Combined choanal/cloacal swab samples ( n = 70) were collected and screened by polymerase chain reaction (PCR) for Mycoplasma, herpesvirus, and ranavirus. Cloacal samples ( n = 50) were also collected and cultured for Salmonella sp. Of 70 tested samples, six (8.6%) were positive for Mycoplasma, and all other PCR and culture test results were negative. Phylogenetic analysis of the 16S ribosomal RNA gene placed the Mycoplasma sp. from B. affinis edwardmolli in the chelonian Mycoplasma cluster that groups within the Mycoplasma pulmonis clade. This mollicute was not associated with clinical disease (defined as observable clinical abnormalities, such as depression, lethargy, respiratory signs, and anorexia) and is likely part of the endemic microbial flora of these terrapins.

Uropygial gland squamous cell carcinoma in chinstrap (Pygoscelis antarcticus) and gentoo (Pygoscelis papua) penguins at the Wildlife Conservation Society's Central Park Zoo.

Uropygial, or preen, glands are found in a variety of avian species including penguins. These glands have a multitude of functions and can develop a variety of conditions including impaction, rupture, adenitis, squamous metaplasia, and neoplasia of various types, with squamous cell carcinoma the most commonly reported. A case series of uropygial gland squamous cell carcinoma in five penguins at the Wildlife Conservation Society's Central Park Zoo is described. Most birds were aged (>10 yr) with a history of chronic, recurrent uropygial gland problems including impaction, rupture, abscess formation, or a combination of conditions. Before and after neoplasia diagnosis, these cases were managed conservatively, and palliative care was provided. Because many of these cases were preceded by chronic inflammation, it is possible this inflammation predisposed the uropygial gland to neoplastic transformation, and more aggressive treatment early in the disease process may therefore be warranted.

COMPARISON OF FRESH AND FROZEN FECAL SAMPLES FOR DETECTION OF ENTERIC SALMONELLA FROM CAPTIVE INDIAN STAR TORTOISES (GEOCHELONE ELEGANS).

The use of frozen fecal samples in enrichment media to detect Salmonella spp. strains was evaluated in Indian star tortoises (Geochelone elegans) to determine the utility of this test method for field collection. Fresh fecal samples were collected from 10 captive adult Indian star tortoises. Each sample was split, with one portion in enrichment media at room temperature for 1 or 2 days before submission to a reference laboratory for standard enteric culture. The other was placed in the same enrichment media and frozen at -20°C for 2 wk. Afterwards, it was transferred to a -80°C freezer for 1 mo before submission to the same reference laboratory. These freezer temperatures and holding times were selected to replicate typical field collection procedures. Salmonella enterica was isolated from all 10 fresh samples but from only six frozen samples. Statistically, results showed no significant difference between the two methods; however, this may be due to the limited sample size, and if so, utilization of frozen fecal samples to determine Salmonella spp. prevalence may underestimate the actual prevalence.

Dried blood spot analysis for elements of nutritional concern as demonstrated in studies of Galápagos land iguanas (Conolophus species).

BACKGROUND: Dried blood spot (DBS) technology is valuable in providing simple means of storing blood samples from wildlife with small blood volumes. Methods designed for heavy metal analysis on DBS become more useful if extended to elements of nutritional significance. PURPOSE: (1) Development of procedures for measuring Mn, Fe, Co, Cu, Zn, Se and Mo in DBS; (2) use the designed methods in health assessments of Galápagos land iguanas (Conolophus species). PROCEDURES: Elements were measured by inductively coupled plasma/mass spectrometry (ICP-MS) following acid digestion of whole blood or DBS from the same animal for direct comparison. Study animals comprised free-ranging iguanas from separate islands in the Galápagos archipelago. MAIN FINDINGS: DBS spikes (Mn, Fe, Co, Cu, Zn, Se and Mo) demonstrated accuracy to ∼100 ppb; reporting limits were set there except for Fe and Zn which were set at 1000 ppb. Plasma samples - generally preferable for nutritional element diagnostics - were submitted from Galápagos land iguanas along with DBS as part of a large-scale health assessment. In plasma versus DBS concentration comparisons, Fe, Cu, Se and Mn correlated well with R^2 values of 0.799, 0.818, 0.896 and 0.899, respectively, and slopes ranging 0.88 - 1.3. Co and Zn showed greater scatter. Mo had insufficient points above its reporting limit and offered advantages for toxicity assessments. Bland-Altman diagrams showed flat scatter between 2x standard deviation boundaries with no undue trends except for Mn which had few points above its reporting limit. Bias, defined as the average difference [DBS - plasma] divided by the average value, was relatively low throughout, with values of - 19.3 % (Fe), - 48.7 % (Co), - 19.6 % (Cu), - 6.9 % (Zn), - 21.4 % (Se) and + 40.7 % (Mn). Normal distribution assessment of iguana Cu, Zn, Se and Fe plasma values showed unanticipated divergences between two species. CONCLUSIONS: The DBS approach for nutritional element analysis offers a suitable methodology for determining crucial elements Mn, Fe, Co, Cu, Zn, Se, and Mo in veterinary samples. Analyses of samples from Conolophus species revealed interesting divergences particularly for Cu, Zn, Se and Fe, elements generally associated with defense against oxidative stress.

Efficacy of treatment and long-term follow-up of Batrachochytrium dendrobatidis PCR-positive anurans following itraconazole bath treatment.

All anuran specimens in the Wildlife Conservation Society's collections testing positive for Batrachochytrium dendrobatidis (Bd) were treated with itraconazole and then studied after treatment to assess the long-term effects of itraconazole and the drug's effectiveness in eliminating Bd carriers. Twenty-four individuals and eight colonies of 11 different species (75 total specimens) tested positive for Bd via polymerase chain reaction (PCR) on multicollection survey. All positive individuals and colonies were treated with a 0.01% itraconazole bath solution and retested for Bd via one of two PCR methodologies within 14 days of treatment completion, and all were negative for Bd. A total of 64 animals received secondary follow-up PCR testing at the time of death, 6-8 mo, or 12-15 mo post-treatment. Fourteen animals (14/64, 21.9%) were PCR positive for Bd on second follow-up. The highest percentage positive at second recheck were green-and-black poison dart frogs (Dendrobates auratus; 5/5 specimens, 100%), followed by red-eyed tree frogs (Agalychnis callidryas; 4/11, 36.4%), grey tree frogs (Hyla versicolor; 1/3, 33.3%), and green tree frogs (Hyla cinera; 3/11, 27.3%). Re-testing by PCR performed on 26/28 individuals that died during the study indicated 11/26 (42.3%) were positive (all via DNA extracted from formalin-fixed paraffin-embedded skin sections). However, there was no histologic evidence of chytridiomycosis in any of 27/28 individuals. The small number of deceased animals and effects of postmortem autolysis limited the ability to determine statistical trends in the pathology data, but none of the necropsied specimens showed evidence of itraconazole toxicity. Problems with itraconazole may be species dependent, and this report expands the list of species that can tolerate treatment. Although itraconazole is effective for clearance of most individuals infected with Bd, results of the study suggest that repeat itraconazole treatment and follow-up diagnostics may be required to ensure that subclinical infections are eliminated in amphibian collections.

Systemic Helicobacter infection and associated mortalities in endangered Grand Cayman blue iguanas (Cyclura lewisi) and introduced green iguanas (Iguana iguana).

The Blue Iguana Recovery Programme maintains a captive breeding and head-starting program for endangered Grand Cayman blue iguanas (Cyclura lewisi) on Grand Cayman, Cayman Islands. In May 2015, program staff encountered two lethargic wild Grand Cayman blue iguanas within the Queen Elizabeth II Botanic Park (QEIIBP). Spiral-shaped bacteria were identified on peripheral blood smears from both animals, which molecular diagnostics identified as a novel Helicobacter species (provisionary name Helicobacter sp. GCBI1). Between March 2015 and February 2017, 11 Grand Cayman blue iguanas were identified with the infection. Two of these were found dead and nine were treated; five of the nine treated animals survived the initial infection. Phylogenetic analysis of the 16S rRNA gene suggests Helicobacter sp. GCBI1 is most closely related to Helicobacter spp. in chelonians. We developed a Taqman qPCR assay specific for Helicobacter sp. GCBI1 to screen tissue and/or blood samples from clinical cases, fecal and cloacal samples from clinically healthy Grand Cayman blue iguanas, including previously infected and recovered iguanas, and iguanas housed adjacent to clinical cases. Fecal and/or cloacal swab samples were all negative, suggesting that Grand Cayman blue iguanas do not asymptomatically carry this organism nor shed this pathogen per cloaca post infection. Retrospective analysis of a 2014 mortality event affecting green iguanas (Iguana iguana) from a separate Grand Cayman location identified Helicobacter sp. GCBI1 in two of three cases. The source of infection and mode of transmission are yet to be confirmed. Analysis of rainfall data reveal that all infections occurred during a multi-year dry period, and most occurred shortly after the first rains at the end of seasonal drought. Additionally, further screening has identified Helicobacter sp. GCBI1 from choanal swabs of clinically normal green iguanas in the QEIIBP, suggesting they could be asymptomatic carriers and a potential source of the pathogen.

Evaluation of medetomidine-ketamine-butorphanol anesthesia with atipamezole-naltrexone antagonism in captive male guanacos (Lama guanicoe).

Seven captive adult male guanacos (Lama guanicoe) weighing 112.0 +/- 10.9 kg (mean +/- standard deviation) were anesthetized with a combination of medetomidine (90.0 +/- 8.8 microg/kg), ketamine (2.7 +/- 0.3 mg/kg), and butorphanol (0.3 +/- 0.03 mg/kg) administered intramuscularly to evaluate its anesthetic and cardiopulmonary effects. Inductions were smooth and rapid, with a mean time to initial effect of 3 +/- 1.5 min and a mean time to recumbency of 5.1 +/- 3.1 min. Anesthesia was predictable, smooth, and characterized by excellent muscle relaxation. Spontaneous ventilation was maintained throughout anesthesia in all animals. Marked bradycardia ranging from 24 to 52 beats/min was noted across all time points for all individuals. Median heart rates decreased during the procedures, but median heart rate, temperature, respiratory rate, and end-tidal carbon dioxide values over the 20 min monitoring period were not significantly different. Mean arterial partial pressure of oxygen (PaO2) on initial sampling was 65.9 +/- 14.8 mm Hg, with six of seven animals exhibiting hypoxemia (PaO2 <80 mm Hg). After oxygen supplementation for 20 min, mean PaO2 values showed statistically significant increases to a mean value of 127.7 +/- 32.4 mm Hg (P = 0.0014). Mean arterial partial pressure of carbon dioxide (PaCO2) showed a significant increase over the monitoring period (P = 0.0004), and mild hypoventilation (PaCO2 >45 mm Hg) was noted in four animals. Mean total duration of procedure time was 23.3 +/- 3.3 min. All guanacos received 0.45 +/- 0.04 mg/kg atipamezole and 2.7 +/- 0.25 mg/kg naltrexone administered intramuscularly for anesthetic antagonism. Recoveries were smooth and uncomplicated. Mean time to sternal recumbency after antagonist administration was 7.7 +/- 4.5 min, and time to successful standing was 12.9 +/- 5.0 min, with all animals standing on first attempt.

Use of a Mycobacterium tuberculosis rapid lateral flow test for assessment of non-specific tuberculin responses in silvered langurs (Trachypithecus cristatus ultima).

Nonspecific responses to intradermal tuberculin skin testing (TST) have been frequently documented in silvered langurs (Trachypithecus cristatus ultima) at the Wildlife Conservation Society's Bronx Zoo. A commercial rapid lateral flow immunoassay (RT) for the identification of Mycobacterium tuberculosis and M. bovis antibodies was employed to assess this test's correlation with TST results and tuberculosis status. Twenty-two silvered langurs were tested with both TST and RT. TST results were characterized into three groups as nonspecific tuberculin responses (n=9), responses to avian tuberculin only (n=6), or negative (n=7). Rapid lateral flow test results were negative in 20 of 22 individuals with equivocal results in two avian tuberculin responders. There was no difference in RT results between TST groups (P = 0.178). Rapid lateral flow test results were not in agreement with mammalian old tuberculin TST results (kappa = 0.08) but showed raw agreement with tuberculosis status (proportion of overall agreement = 0.909). Results suggest the RT assay may useful in the confirmation of nonspecific TST responders in silvered langurs.