Bronchoalveolar lavage for diagnosis of tuberculosis infection in elephants.

Tuberculosis (TB) has been known to affect elephants for thousands of years. It was put into spotlight when few circus elephants were diagnosed carrying Mycobacterium (M.) tuberculosis. Because of the zoonotic risk and high susceptibility to M. tuberculosis, periodic testing was enacted since, in captive breeding programmes. Presently, trunk wash is the recommended diagnostic procedure for TB. Trunk wash, however, puts the operator at risk, has low sensitivity, and is prone to contamination. Here, bronchoalveolar lavage is described for the first time for TB diagnosis in elephants. Bronchial, trunk and mouth fluids were investigated using bacterial culture, M. tuberculosis complex (MTC)-specific real-time quantitative PCR (qPCR) and mycobacterial genus-specific qPCR for overall presence of mycobacteria or mycobacterial DNA including bacteria or DNA of closely related genera, respectively, in 14 elephants. Neither bacteria of the MTC nor their DNA were identified in any of the elephants. Yet, 25% of the cultures grew non-tuberculous mycobacteria (NTM) or closely related bacterial species. Furthermore, 85% of the samples contained DNA of NTM or closely related bacterial genera. This finding might explain continued false-positive results from various serological tests. From a zoonotic point of view, bronchoalveolar lavage is safer for the testing personal, has higher probability of capturing MTC and, through PCR, identifies DNA NTM in elephants. Yet, necessary endoscopic equipment, animal sedation and access to a TB reference laboratory might pose challenging requirements in remote conditions in some elephant range countries.

Cryopreservation of captive roe deer (Capreolus capreolus) semen.

To address the need to preserve current genetic diversity before it is lost forever; further studies to adapt assisted reproductive technologies to various endangered species are needed, among other things. Roe deer (Capreolus capreolus), an over abundant wild deer, can serve as model species to develop or improve sperm cryopreservation of threatened or endangered deer species. The aim of this study was to compare the ability of three diluents (Berliner Cryomedium [BC]; Tris, citric acid, glucose [TCG]; TES, Tris, glucose) to support chilling, cryopreservation (with 5% glycerol; G) and postthaw incubation (at 22 °C and 37 °C) of roe deer spermatozoa collected by electroejaculation. Berliner Cryomedium was the diluent that better preserved roe deer spermatozoa during refrigeration, able to maintain motility for at least 14 days, longer than the other extenders. BC + G was the extender of choice for cryopreservation, showing higher viability compared with TCG + G (66.7 ± 3.4 vs. 54.5 ± 6.5; P < 0.05) and higher level of acrosome integrity compared with TES, Tris, glucose + G (79.4 ± 3.4 vs. 67.9 ± 5.0; P < 0.05). Maintaining the samples at 22 °C after thawing presented higher values in various parameters compared with 37 °C. The knowledge gained through this study can potentially act as a preliminary step toward development of new protocols to help increase the reproductive success of biologically similar, yet endangered, wild species.

Characterization of fetal growth by repeated ultrasound measurements in the wild guinea pig (Cavia aperea).

Fetal growth during pregnancy has previously been studied in the domesticated guinea pig (Cavia aperea f. porcellus) after dissecting pregnant females, but there are no studies describing the fetal growth in their wild progenitor, the wild guinea pig (C aperea). In this study, 50 pregnancies of wild guinea pig sows were investigated using modern ultrasound technique. The two most common fetal growth parameters (biparietal diameter [BPD] and crown-rump-length [CRL]) and uterine position were measured. Data revealed similar fetal growth patterns in the wild guinea pig and domesticated guinea pig in the investigated gestation period, although they differ in reproductive milestones such as gestation length (average duration of pregnancy 68 days), average birth weight, and litter mass. In this study, pregnancy lasted on average 60.2 days with a variance of less than a day (0.96 days). The measured fetal growth parameters are strongly correlated with each (R = 0.91; P < 0.001) other and with gestational age (BPD regression equation y = 0.04x - 0.29; P < 0.001 and CRL regression equation y = 0.17x - 2.21; P < 0.01). Furthermore, fetuses in the most frequent uterine positions did not differ in their growth parameters and were not influenced by the mother ID. Our results imply that ultrasound measurement of a single fetal growth parameter is sufficient to reliably estimate gestational age in the wild guinea pig.

Enriching the captive elephant population genetic pool through artificial insemination with frozen-thawed semen collected in the wild.

The first successful AI in an elephant was reported in 1998, using fresh semen. Since then almost 40 calves have been produced through AI in both Asian and African elephants worldwide. Following these successes, with the objective of enriching the captive population with genetic material from the wild, we evaluated the possibility of using frozen-thawed semen collected from wild bulls for AI in captivity. Semen, collected from a 36-yr-old wild African savanna elephant (Loxodonta africana) in South Africa was frozen using the directional freezing technique. This frozen-thawed semen was used for four inseminations over two consecutive days, two before and two after ovulation, in a 26-yr-old female African savanna elephant in Austria. Insemination dose of 1200 × 10(6) cells per AI with 61% motility resulted in pregnancy, which was confirmed through ultrasound examination 75, 110 and 141 days after the AI procedure. This represents the first successful AI using wild bull frozen-thawed semen in elephants. The incorporation of AI with frozen-thawed semen into the assisted reproduction toolbox opens the way to preserve and transport semen between distant individuals in captivity or, as was done in this study, between wild and captive populations, without the need to transport stressed or potentially disease-carrying animals or to remove animals from the wild. In addition, cryopreserved spermatozoa, in combination with AI, are useful methods to extend the reproductive lifespan of individuals beyond their biological lifespan and an important tool for genetic diversity management and phenotype selection in these endangered mammals.

Estrus induction in white rhinoceros (Ceratotherium simum).

The estrous cycle length in the white rhinoceros (Ceratotherium simum) is either 4 or 10 wk. The cause(s) for this variation as well as the poor fertility rate in captivity remains under debate in this species. Most captive adult white rhinoceros undergo long anovulatory periods without luteal activity which are considered a major reason for their low reproductive rate. In this study, the synthetic progestin chlormadinone acetate (CMA) was tested in combination with hCG or the GnRH analogue deslorelin for its efficiency to induce ovulation in fourteen females without luteal activity and in three, regular cycling females. HCG (N = 12), injectable GnRH analogue (N = 8) and GnRH analogue implants (N = 15) were given to induce ovulation after CMA treatment. Treatment success was determined using both transrectal ultrasonography and progesterone metabolite EIA analysis. A preovulatory sized follicle (3.5 ± 0.1 cm) or a corpus luteum (5.1 ± 0.7) was present on the ovary one day after induction in 93.1% of the treatments. Despite this high rate of ovarian response, ovulation rate differed between the study groups. The ovulation rate for hCG, injectable GnRH analogue and GnRH analogue implants was 66.7%, 62.5% and 93.3%, respectively. Ovulation rate in cyclic females treated with GnRH implants was 100% (6/6) compared with 89% (8/9) in females without luteal activity receiving the same treatment. The length of the estrous cycle when induced with hCG was 4 wk (85.7%). The estrous cycle when induced with GnRH analogue was predominantly 10 wk long. Two females without luteal activity treated with GnRH became pregnant. In conclusion, CMA in combination with GnRH analogue implants was highly effective to induce ovulation in white rhinoceroses and thus can contribute to efforts aimed at increasing natural mating and reproductive rates in the captive white rhinoceros population.

Life cycle of feline Corpora lutea: histological and intraluteal hormone analysis.

The corpus luteum (CL) is a transient hormone gland on the ovary that produces progesterone (P4) for the maintenance of pregnancy. It develops from residual follicular granulosa and theca cells after ovulation. Very little is known about the cellular and hormonal processes within CLs obtained from pregnant and pseudopregnant felids. Therefore, our aim was to review the luteal function in feline CLs of different reproductive stages in conjunction with our data obtained in domestic cats and Eurasian lynxes. Corpus luteum function in lynxes is of particular interest, as a post-partum luteal activity was suggested based on repeated ultrasonography and endocrine examinations. Histology of CL from pregnant and pseudopregnant domestic cats clearly reflects the luteal function. The formation of the CL after ovulation is characterized by transforming of theca and granulosa cells into steroidogenic luteal cells and is accompanied by increased intraluteal and circulating P4 levels. Luteal regression is steadily progressive; the first signs (coarsed vacuolization, increased proportion of non-steroidogenic cells) are visible already in CL from the second trimester of pregnancy.

Collection and preservation of pygmy hippopotamus (Choeropsis liberiensis) semen.

Knowledge about the reproduction of the endangered pygmy hippopotamus is almost non-existent. This study takes the first step toward changing this by devising a protocol for the collection, evaluation, and short-term preservation of semen of this endangered species. Semen was collected successfully from seven bulls by electroejaculation, using a specially designed rectal probe. Mean +/- SEM values of native sperm parameters from combined best fractions were: motility-80.0 +/- 4.1%, concentration-2421 +/- 1530 x 10(6) cells/mL, total collected cell number-759 +/- 261 x 10(6) cells, intact acrosome-87.8 +/- 1.2%, intact morphology-52.7 +/- 4.3%, and, for some, hypoosmotic swelling test-79.3 +/- 4.4% and seminal plasma osmolarity-297.5 +/- 3.3 mOsm. Seven different extenders were tested for sperm storage under chilling conditions: Berliner Cryomedium (BC), Biladyl, modification of Kenney modified Tyrode's medium (KMT), MES medium, Androhep((R)), boar M III() extender and Human Sperm Refrigeration Medium. While differences between males were apparent, the BC was consistently superior to all other extenders in sperm motility and facilitated storage for 7 d with up to 30% motility and some motility even after 3 weeks. With this knowledge in hand, the obvious two directions for future research are to conduct artificial insemination and to develop a technique for sperm cryopreservation.

Seminoma in a southern black rhinoceros (Diceros bicornis minor): diagnosis, surgical management and effect on fertility.

A testicular mass was identified by ultrasonography performed during a routine reproductive evaluation of an adult male southern black rhinoceros (Diceros bicornis minor). Histological examination of a testicular biopsy supported a presumptive diagnosis of testicular neoplasia. Hemi-castration was performed to excise the affected testis and a pathological diagnosis of a seminoma was made. Assessment of semen suggested reduced fertility as a consequence of the neoplastic process, but hemi-castration prevented further growth and metastasis of the tumour and ensured the animal's breeding potential. This is the second documented case of a seminoma in a rhinoceros species and the first case in a black rhinoceros.

Ovarian superstimulation, transrectal ultrasound-guided oocyte recovery, and IVF in rhinoceros.

Numerous reports on reproductive pathology in all rhinoceros species illustrate the abundance of female infertility in captive populations. In infertile rhinoceroses, oocyte collection and embryo production could represent the best remaining option for these animals to reproduce and to contribute to the genetic pool. We report here on superstimulation, repeated oocyte recovery, and attempted in vitro fertilization (IVF) in white and black rhinoceroses. Four anestrous rhinoceroses (two white, two black) with unknown follicular status were treated with gonadotropin-releasing hormone analogue, deslorelin acetate, for 6 to 7 d. Number and size of follicles in superstimulated females was significantly higher and larger compared with those in nonstimulated anestrous females (n=9). Ovum pick-up was achieved by transrectal ultrasound-guided follicle aspiration. Up to 15 follicles were aspirated per ovary. During six ovum pick-ups, a total of 29 cumulus-oocyte complexes (COCs) were harvested with a range of 2 to 9 COCs per collection. No postsurgical complications were noted on the rhinoceros ovaries using this minimally invasive approach. Various in vitro maturation (IVM) and IVF protocols were tested on the collected COCs. Despite the low total number of COCs available for IVM and IVF in this study, we can report the first rhinoceros embryo ever produced in vitro. The production of a 4-cell embryo demonstrated the potential of transrectal ultrasound-guided oocyte recovery as a valuable tool for in vitro production of rhinoceros embryos from otherwise infertile females.

Non cat-like ovarian cycle in the Eurasian and the Iberian lynx - ultrasonographical and endocrinological analysis.

The Iberian lynx is considered the most endangered felid species. Therefore, an ex situ conservation program was initiated to protect this species from extinction. Additional knowledge on lynx reproduction biology and reliable methods for reproductive monitoring are important for developing a captive breeding program. The aim of this study in lynx was to implement transrectal ultrasonography to visualize ovarian structures (follicles, corpora lutea) and to assess ovarian activity in addition to analysis of serum progesterone and oestradiol. Because of limited access to Iberian lynxes, the less-endangered Eurasian lynx and bobcat were also studied in this comparative study. Recent endocrinological studies based on faecal and urinary progesterone and oestrogen metabolites revealed that steroid profiles in both these species were alike and did not follow the typical pattern of other felids. Pregnancy diagnosis was not possible, since progesterone concentrations did not differ between pregnant and pseudopregnant animals. Progesterone was also detected after parturition as well as after weaning until the onset of a new oestrous cycle. In the present study, the presence of corpora lutea during the non-breeding season was confirmed by ultrasonography and by elevated serum levels of progesterone averaging 3.56 +/- 1.3 ng/ml in Eurasian and 6.1 +/- 0.26 ng/ml in Iberian lynx, respectively. The ultrasonographical findings on the ovarian structures suggest strongly that corpora lutea developed after ovulation stay active until November and regress before the onset of the next oestrus.

Seasonal profiles of ovarian activity in Iberian lynx (Lynx pardinus) based on urinary hormone metabolite analyses.

The Iberian Lynx Ex-Situ Conservation Programme is an essential part of a co-ordinated action plan to conserve the most endangered felid species of the world. Successful captive breeding demands reliable methods for reproduction monitoring including reliable non-invasive pregnancy diagnosis. During a 3-year study, urine samples from six captive Iberian lynx females were obtained (one non-pregnant, one pseudo-pregnant and 11 pregnant cycles). Progesterone, pregnanediol and oestradiol were determined in urinary extracts and relevant urinary oestrogen metabolites were characterized by high-performance liquid chromatography (HPLC). Urinary progestins did not follow the typical pregnancy-related course of felids. In the lynx, we failed to demonstrate an urinary progestin elevation during pregnancy. In contrast, urinary oestrogens increased from 3.8 +/- 0.6 to 8.6 +/- 0.5 ng/mg creatinine (p < 0.001) during the pregnancy. A comparison of pseudo-pregnant with pregnant cycles revealed a further increase of oestrogens caused by implantation (p < 0.05). In one female, which refused to mate, no difference was estimated between oestrogens levels during the breeding and non-breeding seasons. Almost 10-fold higher oestrogen concentrations were measured in urines of females that shared enclosures with males. HPLC analysis of oestrogens in urine samples collected from Iberian lynx during the pregnancy revealed that lynx urine is composed of two polar oestrogen metabolites in addition to oestrone and minor amounts of oestradiol. Oestrone was detectable in all urinary extracts (8-12% of metabolites), whereas oestradiol was elevated only during late pregnancy (18%). Thus, seasonal luteal activity in Iberian lynx can be monitored by urinary oestrogens. The increase of urinary oestradiol during late pregnancy might indicate an oestradiol secretion by the lynx placenta.

Pregnancy monitoring in dogs and cats using 3D and 4D ultrasonography.

Three-dimensional (3D)/four-dimensional (4D) volume ultrasound is an established method in human medicine that offers various options for analysing and presenting ultrasound volume data. However, the successful application of the different 3D/4D imaging modalities in pregnant dogs and cats has not yet been reported in the literature. The main reasons for this are: (1) the high costs of 3D/4D ultrasound systems, (2) operation difficulties due to high breathing frequency in non-sedated animals and (3) the missing specific knowledge in veterinary medicine concerning how to perform high-quality volume scans. Automatically acquired ultrasound volume data sets were generated with two different ultrasound systems: the portable Voluson i and the stationary Voluson Expert 730. Different 3D/4D imaging modalities were tested in regard of their practicability in pregnancy monitoring in dogs and cats. Nine different volume imaging modalities were applied using the saved files. For the presentation of the static 3D volume data sets, we used the multiplanar, niche, surface, transparency, glass body, inversion, volume calculation and tomographic ultrasound imaging modes. For the dynamic 4D data, the surface and glass body modes were applied. By changing the human standard settings to the requirements of small animal anatomy, it was found that 3D/4D ultrasound has great potential for the characterization of pregnancy in queens and bitches. The 3D/4D technology offered advanced information about pregnancy status and birth prediction and improved the diagnostic confidence. By using standardized examination protocols, 3D/4D ultrasound will allow a reduction in examination time by generating even more relevant information. These benefits, combined with possible future cost reduction of commercial ultrasound systems, might lead to frequent utilization in routine pregnancy diagnostic and birth management in small animal practice.

Frozen-thawed rhinoceros sperm exhibit DNA damage shortly after thawing when assessed by the sperm chromatin dispersion assay.

This study reports on the successful validation (via in situ nick translation and neutral comet assay) of the equine Sperm-Halomax kit as an appropriate methodology for the assessment of sperm DNA fragmentation in three species of rhinoceros. Rhinoceros sperm nuclei with fragmented DNA (validated using in situ nick translation) were evident as large halos with dispersed DNA fragments, whereas those with nonfragmented DNA displayed small halos of nondispersed DNA within the microgel. There was a high correlation (r) of 0.974 (R(2) value=0.949; P<0.01; n=16) between the respective assessments of the Sperm Chromatin Dispersion test (SCDt) and the neutral comet assay. Application of the SCDt to determine the DNA fragmentation dynamics of rhinoceros (n=6) sperm frozen in liquid nitrogen vapor and incubated postthaw at 37 degrees C for up to 48 h to mimic in vitro conditions in the female reproductive tract, revealed an increase (P=0.001) in DNA damage, as soon as 4h after the start of incubation. Linear regression equations were calculated for all six rhinoceroses over the first 6h of incubation and revealed individual animal variation. Freshly collected and incubated (37 degrees C) rhinoceros (n=3) sperm had no increase in the basal level of DNA fragmentation for up to 48 h, indicating that the cryopreservation of rhinoceros sperm in liquid nitrogen vapor, as used in this study, appeared to result in freeze-thaw DNA damage.

First successful artificial insemination with frozen-thawed semen in rhinoceros.

The first successful artificial insemination (AI) in a rhinoceros was reported in 2007 using fresh semen. Following that success, we decided to evaluate the possibility of using frozen-thawed semen for artificial insemination. Semen, collected from a 35-36 year old Southern white rhinoceros (Ceratotherium simum simum) in the UK was frozen using the directional freezing technique. This frozen semen was used in two intrauterine AI attempts on a 30 years old female rhinoceros in Hungary. The first attempt, conducted 30 days postpartum with an insemination dose of approximately 135 x 10(6) motile cells, failed. The second attempt, conducted two estrus cycles later with an insemination dose of approximately 500 x 10(6) motile cells, resulted in pregnancy and the birth of a healthy offspring. This represents the first successful AI using frozen-thawed semen in a rhinoceros, putting it among very few wildlife species in which AI with frozen-thawed semen resulted in a live birth. The incorporation of AI with frozen-thawed semen into the assisted reproduction toolbox opens the way to preserve and transport semen between distant individuals in captivity or between wild and captive populations, without the need to transport stressed or potentially disease carrying animals. In addition, cryopreserved spermatozoa, in combination with AI, are useful methods to extend the reproductive lifespan of individuals beyond their biological lifespan and an important tool for managing genetic diversity in these endangered mammals.

MALDI-TOF "fingerprint" phospholipid mass spectra allow the differentiation between ruminantia and feloideae spermatozoa.

The detailed comparative analysis of sperm lipids could essentially contribute to a better understanding of membrane function in the context of fertilization and, moreover, of sperm preservation. The application of sensitive analytical methods is particularly necessary for endangered species as the available amount of spermatozoa (and, accordingly, extractable lipids) is strongly limited. It will be shown that matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a fast, simple and sensitive method for the determination of the phospholipid composition of spermatozoa from several ruminantia (cattle, roe deer, Klipspringer) and feloideae species (domestic cat, Siberian tiger, fosa). Characteristic "fingerprints" are obtained from the positive ion spectra that allow the differentiation between both animal groups. In contrast to the lipid extracts of ruminantia spermatozoa which predominantly contain ether lipids including essential amounts of plasmalogens, the more complex phospholipid composition of feloideae spermatozoa is clearly dominated by diacyl phospholipids and contains only marginal amounts of plasmalogens. It will also be shown that the lipid compositions of ejaculated, electroejaculated and cauda epididymal spermatozoa of the same species are very similar and give comparable data. Therefore, the analysis of ejaculated spermatozoa is not an absolute must.

Pregnancy diagnosis in urine of Iberian lynx (Lynx pardinus).

Diagnosis of pregnancies is an important management tool for the Iberian lynx Conservation Breeding Program, a program geared to recover the world's most endangered felid. Non-invasive methods such as fecal hormone analyses are not applicable to the lynx, since fecal progestin does not follow the typical pregnancy pattern of felids. Therefore, we aimed to test whether urine can be used as an alternative substance for pregnancy diagnosis in the Iberian lynx. Progesterone immunoreactive metabolites were determined in urine samples of pregnant and non-pregnant females before and during breeding season. Additionally, we used the Witness Relaxin test to determine relaxin in blood and urine. No differences were found in progestin concentrations determined in urine samples collected from pregnant and non-pregnant animals between day 1 and 65 following mating. Although the Witness Relaxin test was positive in serum samples collected from animals between day 32 and 56 of pregnancy, it failed in both fresh and frozen urine samples collected from the same stage of pregnancy. A weak relaxin reaction in urine samples collected from animals between day 29 and 46 of pregnancy was detectable after urines were concentrated by ultrafiltration (>50x). Concentrated samples obtained from non-pregnant and early pregnant animals yielded negative test results. In conclusion, the Witness Relaxin test can be applied for pregnancy diagnosis in Iberian lynx in both serum and concentrated urine samples obtained during the second half of pregnancy. A positive relaxin test indicates an ongoing pregnancy, whereas negative tests must be judged carefully as hormone concentrations might be below detection thresholds.

Non-invasive monitoring of hormones: a tool to improve reproduction in captive breeding of the Eurasian lynx.

The survival of many critical endangered mammal species is often depending on successful captive breeding programmes which include the future option of reintroduction to the wild. Breeding in captivity also demands the application of modern assisted reproductive techniques to ensure maximal biodiversity, but knowledge on reproductive physiology is often limited. Therefore, non-invasive monitoring of urinary and faecal hormones has become an important tool for reproductive management. To exemplify the importance of non-invasive hormone monitoring, we choose the Eurasian lynx as a model for the world's most endangered felid species, the Iberian lynx. We analysed faecal samples of pregnant and pseudo-pregnant female Eurasian lynxes during a 3-year study period. Compared to pre-mating levels faecal progesterone metabolite profiles revealed a tendency towards higher levels in pregnant and pseudo-pregnant females with no difference between both categories. Oestrogen levels raised in both pregnant and pseudo-pregnant females with a tendency to be more elevated and prolonged in pregnant females. Surprisingly both E2 and P4 metabolites were highly correlated (r(2) =0.8131, p < 0.0001) showing a postpartum increase both in pregnant and pseudo-pregnant females. The results from the Eurasian lynx revealed that the measurement of faecal progesterone metabolites led to profiles dissimilar to profiles shown in other felid species, but similar to those from faecal gestagen metabolite analysis in the Iberian lynx. To identify faecal gestagen and oestrogen metabolites a radio-metabolism study was performed. Using the progesterone immunoassay two major progesterone metabolites were detected demonstrating that the assay indeed tracks the relevant metabolites. The oestrogen assay measured authentic 17beta-oestradiol and oestrone, and their conjugates. The analysis of the faecal metabolite composition in samples from early and late pregnancy and lactation particularly revealed a distinct shift in the relation between 17beta-oestradiol and oestrone that changed in favour of oestrone. This might indicate different hormone sources during and after pregnancy (corpus luteum, placenta). We hypothesize, that placental steroid analysis in combination with other highly sophisticated analytical techniques, like liquid chromatography mass spectrometry or urinary relaxin analysis may led to analytical options to confirm pregnancy and to differentiate this from pseudo-pregnancy in lynx species.

Obstetrics in elephants.

Obstetrics, one of the oldest fields in veterinary medicine, is well described and practiced in domestic and exotic animals. However, when providing care during elephant birth or dystocia, veterinary intervention options differ greatly from any domestic species, and are far more limited due to the dimensions and specific anatomy of the elephant reproductive tract. In addition, aging of captive elephant populations and advanced age of primiparous females make active birth management increasingly important. Intrauterine infection, uterine inertia and urogenital tract pathologies are emerging as major causes for dystocia, often leading to foetal and dam death. This paper reviews the current knowledge on elephant birth and the factors associated with dystocia. It then summarises recommendations for birth and dystocia management. As Caesarean section, the most common ultima ratio in domestic animal obstetrics, is lethal and therefore not an option in the elephant, non-invasive medical treatment, induction of the Fergusson reflex or the conscious decision to leave a retained foetus until it is expelled voluntarily, are key elements in elephant obstetrics. Surgical strategies such as episiotomy and foetotomy are sometimes inevitable in order to try to save the life of the dam, however, these interventions result in chronic post-surgical complications or even fatal outcome. Limited reliable data on serum calcium concentrations, and pharmacokinetics and effect of exogenous oestrogen, oxytocin, and prostaglandins during birth provide the scope of future research, necessary to advance scientific knowledge on obstetrics in elephants.

Early embryo development in the elephant assessed by serial ultrasound examinations.

The elephant has an extraordinary long pregnancy, lasting 21 months. However, knowledge on embryo development is limited. To date, only single morphological observations of elephant embryo development associated with placentation are available, all lacking correlation to gestational age. The present study describes morphological characteristics of early embryo development in the elephant with exact biometric staging. Six pregnancies in five Asian and one African elephants with known conception dates were followed by 2D and 3D ultrasound, covering the embryonic period from ovulation to day 116 post-ovulation. The embryonic vesicle was earliest observed was on day 50 p.o. The proper embryo was not detected until day 62 p.o. Embryonic heartbeat was first observed on day 71 p.o. The allantois, which became visible as a single sacculation on day 71 p.o. was subdivided in four compartments on day 76 p.o. By day 95 p.o., head, rump, front and hind legs were clearly distinguished. Between days 95 and 103 p.o. the choriovitelline placenta was replaced by the chorioallantoic placenta. A physiological midgut herniation was transiently present between days 95 and 116 p.o. On the basis of the late appearance of the embryonic vesicle, delayed implantation in the elephant is discussed. The study provides a coherent description of elephant embryonic development, formation of the extraembryonic organs and their role in placenta formation, all of which are of interest for both comparative evolutionary studies and the improvement of assisted reproduction techniques.

Assisted reproduction in female rhinoceros and elephants--current status and future perspective.

Over the last few decades, rhinoceroses and elephants became important icons in the saga of wildlife conservation. Recent surveys estimate the wild Asian (Elephas maximus) and African (Loxodonta africana) elephant populations to be, at most, 50 250 and 637 600 respectively. For the five rhinoceros species, black (Diceros bicornis), white (Ceratotherium simum), Indian (Rhinoceros unicornis), Javan (Rhinoceros sondaicus) and Sumatran rhinoceros (Dicerorhinus Sumatrensis), the population estimates of 3610, 11 330, 2400, 60 and 300, respectively, are of even greater concern. Protected against habitat loss, poaching and left undisturbed, rhinoceros and elephants reproduce well in the wild. But small and decreasing populations make successful captive management of these taxa increasingly important. In captivity, however, most populations face possible 'extinction' because of historically poor reproductive performance. From the first descriptions of the reproductive anatomy and the oestrous cycle (Laws 1969; Kassam and Lasley 1981; Balke et al. 1988a,b; Plotka et al. 1988; Godfrey et al. 1991) to the present use of advanced assisted reproduction technologies, researchers have strive to understand the function and dysfunction of the reproductive biology of these charismatic species. This paper reviewed the current knowledge on rhinoceros and elephant reproduction biology, reproductive cycle, gestation, dystocia, reproductive pathology, oestrous induction and artificial insemination, sperm sexing, IVF and contraception, and how this knowledge is or might be used to aid species conservation for maximal reproductive efficiency and enhancement of genetic management.