The usefulness of near infrared spectroscopy (NIRS) to monitor urine estrogen concentrations was studied in order to determine optimal timing for breeding captive female giant pandas. NIR spectra of daily urine samples from a female giant panda (Ailuropoda melanoleuca) were acquired in the period between March 1st and 25th, 2007 (n ¼ 53). Estrone-3-glucuronide (E 1 G) concentrations in the samples were also measured by enzyme immunoassay (EIA). Transmittance spectra of all urine samples were obtained in the wavelength range from 1100 to 2432 nm (excluding the range from 1884 to 2012 nm) with sample thickness of 1 mm. Partial least square regression was applied to the spectra and good correlation was obtained between E 1 G concentration measured by EIA and predicted values by NIR (R 2 ¼ 0.94, SECV ¼ 10.04 ng ml À1 ). The results of both soft-independent modeling of class analogy (SIMCA) and moving principal component analysis (MPCA) could detect the time changes in E 1 G concentration as measured by EIA (the Pearson's correlation coefficients between E 1 G concentration and the interclass distances of SIMCA or the index of MPCA were r ¼ 0.64 and r ¼ 0.81 respectively, P < 0.01). As for MPCA index, the index sharply dropped on March 24th corresponding to the decrease of the E 1 G concentration indicating ovulation. Finally, artificial insemination was performed for 3 consecutive days including the peak day, March 24th, and the female became pregnant. These results indicated that NIRS and the following MPCA analysis of the respective urine spectral data could detect the changes of urinary hormones during estrous cycle at a nanogram level. The NIRS can find the optimal timing for breeding quicker and easier than EIA, so this technique can be useful for captive breeding of this threatened species.
Abstract. To detect estrus for reproductive management, and to determine the relationship between urinary estrogen and estrous behavior, in a female giant panda, we developed and evaluated a rapid enzyme immunoassay (EIA) system for urinary Estrone-3-glucuronide (E1G) using commercial reagents. The developed EIA system took only around 3 hours, including all procedures to obtain a result. It indicated good reproducibility (intra-assay CV of 5.16%, interassay CV of 15.4%) and sensitivity (lowest standard concentration was 0.0156 ng/ml) for measurement of the urinary concentrations of E1G in the giant panda. There was a positive correlation (r=0.934) with the data for estrone (E1) in the same samples, as measured by radioimmunoassay (RIA) performed in a commercial laboratory. The changes in the E1G concentrations were almost synchronous with the changes in E1 assayed by RIA in urine collected during 4 consecutive estrous seasons. The dynamics of urinary E1G measured by this system highly correlated with the occurrence of the presenting estrous behavior in the giant panda. The above results indicate that this assay system may be normally, rapidly and practically used for measurement of the urinary concentration of E1G in the giant panda. Key words: Giant panda (Ailuropoda melanoleuca), Enzyme immunoassay, Estrone-3-glucuronide, Estrous behavior (J. Reprod. Dev. 54: [281][282][283][284][285] 2008) he giant panda (Ailuropoda melanoleuca) is a critically endangered species. Its natural habitat is located and limited to Western China at elevations of 2,000 m to 4,000 m. The actual population is estimated to be no more than 1,500 individuals [1]. The giant panda is categorized as "Endangered" on the Red List of Threatened Species published by the International Union for Conservation of Nature and Natural Resources [2].Female giant pandas have a single annual estrus period and generally are in estrus for only a few days between February and May [3]. This characteristic constitutes a major limitation on production of new born in giant pandas.At the Kobe Municipal Oji Zoo, a collaborative project concerning reproduction of the giant panda has been conducted with China since 2000. Since the main purpose of this project is production of baby pandas, the primary work required is establishment of a rapid assay system to measure the sex steroid hormones to evaluate the giant panda's reproductive status and especially to detect estrus by measuring estrogen concentrations in the female for natural and artificial breeding.Because zoo and wild animals are not domesticated or are untrained, frequent sampling of blood is very difficult, and so urine or feces, which are easily collected without being invasive to the animals, are used to measure hormones. In Japan, another study of reproduction of the giant panda has been underway at Ueno Zoological Gardens, Tokyo, since 1972. They have succeeded in production of giant panda by artificial insemination (AI) [4]; however, they have also experienced unsuccessful reproduction in a fema...
In order to enable monitoring of the reproductive status of the female giant panda after observation of estrus behavior, we developed an enzyme immunoassay (EIA) system for urinary pregnanediol-3-glucuronide (PdG), a progesterone metabolite, using commercial reagents and examined the changes in the urinary concentration of PdG in a female giant panda that showed pseudopregnancy and suspicious pseudopregnancy in 6 consecutive years. The developed EIA system had good reproducibility (intra-and interassay CVs 6.1% and 16.3%, respectively), good parallelism between the standard curve and the dose response curve of serial diluted samples and positive correlation (r=0.836) with the data for PdG in the same samples measured by gas chromatography. Urinary PdG in the female panda showed two phases of increase. The first elevation was observed immediately after estrus with the levels of PdG below 100 ng/ Crmg, while the second phase was characterized by a drastic elevation above 100 ng/Crmg until the level began to decrease at the end of pseudopregnancy or suspicious pseudopregnancy. The length of the second phase had wider range than that of the first phase. In the present study, a new EIA assay system for urinary PdG in the female giant panda was developed, and we found that the length of the second phase is unstable in the pseudopregnant and suspicious pseudopregnant giant panda, in contrast with the unstable length of the first phase caused by delayed implantation in the pregnant giant panda.
An American greater flamingo(Phoenicopterus ruber) chick that hatched in Kobe Municipal Oji Zoo presented with a walking difficulty, as it grew, because of a twisted knock-knee leg. To remedy this, we developed a double-joint device orthosis made from polypropylene and fixed it from the tibiotarsus to the tarsometatarsus region in the chick. The chick began to walk normally and responded to the effects of corrective treatment. However, we also made a number of improvements to the joint bending to this orthosis because of the formation of a superficial ulcer in the intertarsal joint of the chick, continuing corrective treatment for 254 days. Results confirm twisted leg symptom improvements following the removal of the orthosis, and the chick continued to live with the other flamingos.
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