Omaha's Henry Doorly Research Articles

Not Her

Not Her Cindy Veach (bio) I don’t see the days. —Abdullah, age 9 The watch helps me remember our history. —Ahmed, age 17 —TIME It feels wrong to sit in the waiting area of my hair salon reading an article about Syrian refugees at the same time a woman in foils goes on about why it's condescending for a waiter to address a woman old enough to cover her roots as dear or sweetie. Wrong too that their words, paired alongside pictures of items they carried, rest on a retro coffee table. And if the words are the why then isn't it equally wrong that the flat screen in the corner is chattering about a drunk woman who broke into Omaha's Henry Doorly Zoo to pet the poor three-legged Malaysian tiger? It's dangerous to be adrift in a sea of tigers. When the bigger ship rammed the crowded dinghy it capsized. A refugee named Omar couldn't find his wife or children in the chop. All he could find were plastic water bottles which allowed him to float [End Page 90] the rest of the way. Tell me, how is it okay for a colorist to be mixing my personalized formula, as noted on a 3 x 5 index card with my name on it, at the same time I'm reading this? And this— Greek authorities erroneously reported they'd found Omar's wife—washed up and bloated. In other words, the things they carried were nothing but failed amulets. And isn't it outrageous that the blonde "Live at Five" anchor is telling me that the drunk woman will lose fingers? They told Omar his dead wife was wearing boots. I'm reading his exact words—She was barefoot in the boat that's how I knew it was not her— when my colorist chirps, "We're ready for you, dear." [End Page 91] Cindy Veach Cindy Veach's poems have appeared in Prairie Schooner, Poet Lore, Michigan Quarterly Review, The Journal, Chicago Review, and elsewhere. Her debut poetry collection, Gloved against Blood, is forthcoming from CavanKerry Press. She manages fundraising programs for nonprofit organizations and lives in Manchester, Massachusetts. Copyright © 2017 Cindy Veach

Systematics ofDiplazium laffanianum(Athyriaceae), a Fern Species Endemic to Bermuda

Diplazium laffanianum was last collected from its natural habitat in Bermuda in 1905. Now the species is considered extinct in the wild as it exists only in greenhouses, tissue cultures, and as a few young reintroduced sporophytes. For the last 13 years, a collaborative team of researchers from the Government of Bermuda and the Lab for Rare & Endangered Plants at Omaha's Henry Doorly Zoo in Nebraska, USA has been working to bring this species back from the brink of extinction. In this project, we developed propagation methods for D. laffanianum, and we studied molecular and morphological characters to determine the phylogenetic placement of D. laffanianum. We sequenced plastid DNA from D. laffanianum using high-throughput sequencing, downloaded six plastid markers (atpA, atpB, matK, rbcL, rps rps4-trnS, and trnL intron trnL-F) from GenBank for 21 Diplazium and outgroup taxa, and inferred a phylogeny. We examined 185 collections of five Diplazium species closely related to D. laffanianum, and we scored 15 morphological characters to test whether D. laffanianum is a distinct species. The phylogeny placed D. laffanianum in the D. cristatum group, suggesting a Neotropical ancestor, and two morphological characters distinguished D. laffanianum from close relatives. Our results provide evidence for the phylogenetic placement of D. laffanianum, and we are actively propagating and reintroducing individuals to the wild, aiding conservation of the species.

Discrimination of individual tigers (Panthera tigris) from long distance roars

This paper investigates the extent of tiger (Panthera tigris) vocal individuality through both qualitative and quantitative approaches using long distance roars from six individual tigers at Omaha's Henry Doorly Zoo in Omaha, NE. The framework for comparison across individuals includes statistical and discriminant function analysis across whole vocalization measures and statistical pattern classification using a hidden Markov model (HMM) with frame-based spectral features comprised of Greenwood frequency cepstral coefficients. Individual discrimination accuracy is evaluated as a function of spectral model complexity, represented by the number of mixtures in the underlying Gaussian mixture model (GMM), and temporal model complexity, represented by the number of sequential states in the HMM. Results indicate that the temporal pattern of the vocalization is the most significant factor in accurate discrimination. Overall baseline discrimination accuracy for this data set is about 70% using high level features without complex spectral or temporal models. Accuracy increases to about 80% when more complex spectral models (multiple mixture GMMs) are incorporated, and increases to a final accuracy of 90% when more detailed temporal models (10-state HMMs) are used. Classification accuracy is stable across a relatively wide range of configurations in terms of spectral and temporal model resolution.

116 DEVELOPMENT OF ASSISTED REPRODUCTION TECHNOLOGIES FOR THE ENDANGERED MISSISSIPPI GOPHER FROG (RANA SEVOSA) AND SPERM TRANSFER FOR IN VITRO FERTILIZATION

Species-specific differences in breeding strategies and physiology have limited the application of assisted reproductive technologies (ART) for critically endangered amphibians in captive assurance colonies. In 2006, the Memphis Zoo (MZ) initiated a program to develop ART for the critically endangered Mississippi gopher frog after natural breeding failed. Standard gamete collection and IVF developed by MZ for reproducing endangered toads such as the Wyoming or boreal toad were applied to the gopher frog with little success, especially hormonal therapy for sperm production. Using the leopard frog as a model species for Ranids, we tested the time and dose dependence of a luteinizing hormone releasing hormone analogue (LHRHa) and hCG on sperm quantity and quality. Initial findings from the leopard frog study were critical in designing the study on gopher frogs. Our objectives were to (1) compare 2 different hormones administered intraperitoneal (500 IU hCG vs 15 μg LHRHa) or their combination on spermiation in gopher frogs; (2) develop in vivo oocyte maturation and ovulation protocols using LHRHa (15 μg) and hCG (500 IU); and (3) transfer this technology to another institution as proof of principle. In gopher frogs, 100 and 83% of the males produced sperm in response to the LHRHa and the combination treatment, respectively, whereas only 16% responded to hCG alone. Sperm concentration peaked at 1 h post-administration for all treatments, with the LHRH/hCG cocktail treatment producing the highest concentration of sperm (mean = 4.6 × 106 ± 1.2 × 106 sperm mL–1, n = 6). No differences in motility were observed between treatments (P > 0.05). For females, a series of priming hormones of hCG and LHRHa were given several months before an ovulatory hormone regimen resulting in ovulation by 100% of the females (n = 6), whereas animals not primed failed to ovulate (n = 4). These 3 separate priming and IVF trials conducted between 2008 and 2010 resulted in each female laying ∼2000 eggs, with an average fertilization rate of 76% for inseminated eggs and hundreds of tadpoles produced. These IVF tadpoles represent the first captive reproduction of gopher frogs and highlight how ART can be applied to conservation and genetic management of threatened species. Subsequently, we tested our IVF protocols on gopher frogs at Omaha's Henry Doorly Zoo using fresh (collected on site) and chilled, shipped sperm from MZ. We collected 6169 eggs from 9 hormone-primed females with all animals ovulating. A portion of the total eggs ovulated were inseminated, resulting in 2401 fertilized eggs (38.9% of total eggs collected) across 18 different male–female pairings leading to viable tadpoles. In addition, sperm transferred overnight from the MZ produced 202/441 fertilized eggs (46%). The transfer of this technology and production of endangered amphibians using chilled, shipped sperm from live animals is a conservation milestone that can be applied to other captive breeding programs.

120 THE DEVELOPMENTAL COMPETENCE OF TADPOLES PRODUCED IN VITRO FROM THE ENDANGERED DUSKY GOPHER FROG (RANA SEVOSA) USING EXOGENOUS HORMONE TREATMENT

Dusky gopher frogs once existed throughout the states of Mississippi, Alabama and Louisiana. Presently, the USA Fish and Wildlife Service estimates that there are less than 100 frogs left in the wild, with almost all of these residing in a single pond in Mississippi, making the dusky gopher frog America's most endangered frog species. Their habitat has been threatened by residential and forestry development, as well as from fire suppression and the decline of gopher tortoises, whose burrows the frogs use for shelter. The USA Fish and Wildlife Service brought the first dusky gopher frogs into captivity in 2001 and they have been kept at the Memphis Zoo since 2003 and at Omaha's Henry Doorly Zoo since 2004. Natural breeding attempts in captivity have been unsuccessful despite efforts to artificially mimic what is known to be the appropriate seasonal environmental (e.g. light, humidity, temperature) and social cues (e.g. calls from wild frogs during the breeding season) that stimulate reproduction in situ. Also, there was a concern about the incidences of abnormalities (e.g. spindly legs) from inadequate husbandry conditions (e.g. nutritional deficiencies) or diseases in tadpoles and froglets produced ex situ (e.g. dermomycoides). The objectives of this study were (1) to examine the developmental competence of tadpoles produced by exogenous hormone treatment for IVF to complete normal metamorphosis into viable froglets ex situ and (2) to determine the incidence of developmental abnormalities or dermomycoides in the froglets produced ex situ. Males and females were first separated by the identification of black male nuptial pads used during amplexus. Males and females were stimulated to spermiate and ovulate, respectively, using a variation of a standard amphibian hormonal stimulation protocol (Kouba et al. 2012 Reprod. Fertil. Dev. 24). As a result, more than 1460 viable tadpoles (61% of fertilized eggs) were produced from 18 segregated genetic pairings. To date, 602 (41%) of the tadpoles have completed normal metamorphosis into viable froglets (over 6 times the known existing population in situ), 51% were either culled to examine for dermomycoides or died for a variety of known (accidental) and unknown reasons (abnormal appendage development accounting for less than 7%) and 8% have yet to metamorphose after 9 months. The incidence of dermomycoides in the culled individuals was 100%; however, a study being conducted concurrently in their native habitat has confirmed that the organism does exist in situ with the dusky gopher frogs, which act as natural carriers. Efforts are currently in place to establish reintroduction programs for the dusky gopher froglets produced ex situ to increase the dwindling wild population now that tadpoles produced by IVF have been shown to be developmentally competent. In conclusion, IVF can be used to produce tadpoles of the dusky gopher frog with a low incidence of abnormalities, but the tadpoles were found to be carriers of the organism dermomycoides similar to their counterparts in the wild.

Comparison of Carfentanil-Xylazine and Thiafentanil- Medetomidine in Electroejaculation of Captive Gaur (Bos gaurus)

Carfentanil citrate and thiafentanil oxalate have been used successfully to immobilize captive and free-ranging ungulates. The objective of this study was to compare the efficacy and certain physiologic parameters of protocols by using the 2 opioids in gaur (Bos gaurus). Eight adult gaur bulls were immobilized for electroejaculation at Omaha's Henry Doorly Zoo (Omaha, Nebraska, USA). All the animals were immobilized twice, by using each of the following protocols one time: 10 mg carfentanil combined with 100 mg xylazine (CX), reversed with 1,000 mg naltrexone and 24 mg yohimbine; and 12 mg thiafentanil combined with 20 mg medetomidine (TM), reversed with 120 mg naltrexone and 100 mg atipamezole. Immobilization drugs were delivered intramuscularly into the shoulder area via pole syringe. Electroejaculation was carried out by a standardized protocol to duplicate procedural stimulation on each animal. Induction and recovery times, initial rectal temperature, heart rate, respiratory rate, anesthetic depth, oxygen saturation, indirect blood pressure, and arterial blood gases were recorded at the time of initial handling, before ejaculation, and after ejaculation. Antagonists were administered 1/4 i.v. and 3/4 s.q. Both protocols require a small volume of drug for a large ungulate, provide smooth induction, and adequate anesthesia. Both protocols produced a significant hypoxemia, although the animals on CX showed slightly better blood gas values (based on lower partial pressure of carbon dioxide) and numerically lower blood pressure values. Animals on TM had better muscle relaxation and smoother recoveries, with no renarcotization noted. The results of the present study indicate the TM and CX protocols used for immobilizing gaur result in similar quality ejaculates that can be used for fertility examination as well as for assisted reproduction such as artificial insemination. Additional immobilizations need to take place to further compare these 2 combinations in this species.

Vocal power and pressure–flow relationships in excised tiger larynges

Despite the functional importance of loud, low-pitched vocalizations in big cats of the genus Panthera, little is known about the physics and physiology of the mechanisms producing such calls. We investigated laryngeal sound production in the laboratory using an excised-larynx setup combined with sound-level measurements and pressure-flow instrumentation. The larynges of five tigers (three Siberian or Amur, one generic non-pedigreed tiger with Bengal ancestry and one Sumatran), which had died of natural causes, were provided by Omaha's Henry Doorly Zoo over a five-year period. Anatomical investigation indicated the presence of both a rigid cartilaginous plate in the arytenoid portion of the glottis, and a vocal fold fused with a ventricular fold. Both of these features have been confusingly termed 'vocal pads' in the previous literature. We successfully induced phonation in all of these larynges. Our results showed that aerodynamic power in the glottis was of the order of 1.0 W for all specimens, acoustic power radiated (without a vocal tract) was of the order of 0.1 mW, and fundamental frequency ranged between 20 and 100 Hz when a lung pressure in the range of 0-2.0 kPa was applied. The mean glottal airflow increased to the order of 1.0 l s(-1) per 1.0 kPa of pressure, which is predictable from scaling human and canine larynges by glottal length and vibrational amplitude. Phonation threshold pressure was remarkably low, on the order of 0.3 kPa, which is lower than for human and canine larynges phonated without a vocal tract. Our results indicate that a vocal fold length approximately three times greater than that of humans is predictive of the low fundamental frequency, and the extraordinarily flat and broad medial surface of the vocal folds is predictive of the low phonation threshold pressure.

DERMATITIS IN CAPTIVE WYOMING TOADS (BUFO BAXTERI) ASSOCIATED WITH FUSARIUM SPP.

From May 2007 to June 2008, 30 of 49 Wyoming toads (Bufo baxteri) kept at Omaha's Henry Doorly Zoo (Nebraska, USA) died showing clinical signs of ventral erythema, inappetance, lethargy, and delayed righting reflex. Treatment with antifungals and antibiotics was unsuccessful in all cases. Histopathologic analyses revealed dermatitis as the primary problem in 20 of 21 toads in which skin was examined. Fungal dermatitis was present in 17 toads, with hyphae approximately 1-3 μm in diameter, and parallel cell walls and frequent septations. In 14 animals, the fungal dermatitis was the main pathologic lesion. Several species of bacteria were associated with all cases. A few animals tested positive for Ranavirus using polymerase chain reaction. Fusarium sp. was consistently cultured from skin, feces, kidneys, and from powdered food provided to crickets. Four isolates were identified as Fusarium proliferatum, Fusarium oxysporum, Fusarium solani, and Fusarium verticillioides, which suggested a secondary role of fungi. A specific underlying cause of disease could not be found, although the roles of humidity and Ranavirus infection are discussed, along with the well-known susceptibility of bufonids to fungal dermatitis.

Collaboration for amphibian conservation: the establishment of the Johannesburg Zoo Amphibian Conservation Center in South Africa with assistance from Omaha's Henry Doorly Zoo, USA

Amphibian populations around the world are rapidly declining in number as their habitats become increasingly unsuitable for survival. A call from the World Association of Zoos and Aquariums prompted Johannesburg Zoo to set up a centre for amphibian breeding and research. It soon became apparent that assistance would be required to learn the skills necessary to house and reproduce South African amphibian species successfully. Omaha's Henry Doorly Zoo, Nebraska, USA, was approached to assist in developing these skills at the Johannesburg Zoo. The result of this strong collaborative effort was the establishment of the Johannesburg Zoo Amphibian Conservation Center.

A training program for noninvasive semen collection in captive western lowland gorillas (Gorilla gorilla gorilla)

Staff at Omaha's Henry Doorly Zoo devised a training program for noninvasive semen collection from three male gorillas. Trained behaviors, shaped using food rewards and verbal praise, enabled the trainer to perform a cursory physical examination and collect >270 semen samples during the study period. Ejaculate volume, sperm motility, and progressive forward motion were recorded. Two of the gorillas regularly produced semen samples that were cryopreserved. Frozen semen from one gorilla was successfully used in an assisted reproductive procedure. The training program may be used as a guideline for other institutions to create similar programs. Zoo Biol 17:143–151, 1998. © 1998 Wiley-Liss, Inc.