雌性小熊猫粪样中雌二醇与孕酮水平的变化与繁殖启动的关系

于1999年12月-2000年4月在成都动物园和成都大熊猫繁育研究基地,用放射免疫分析法测定了4只雌性小熊猫（Ailurus fulgens)粪样中雌二醇(E2)和孕酮(P)水平的变化,同时对其交配行为进行了观察。研究结果表明：雌性小熊猫进入发情期后,粪样中E2迅速升高,并多次出现峰值,表明小熊猫可能是季节性多发情动物。交配行为发生前E2出现峰值,P水平上升,说明E2和P是启动雌性小熊猫交配行为的重要因素;未伴随有P水平相对上升的E2峰值不能引起雌性小熊猫发生交配行为,该现象为暗发情。交配行为发生后P水平持续升高,据此推断小熊猫的排卵方式可能是诱导排卵,诱导因子是交配刺激。     

雄性小熊猫粪便中睾酮水平的变化与繁殖周期的关系

1999 年12 月至2000 年9 月, 使用放射免疫分析法, 测定了成都动物园和成都大熊猫繁育研究基地的4 只雄性小熊猫粪样中睾酮水平变化情况, 同时对其行为进行了观察。研究结果表明, 雄性粪样中睾酮水平在繁殖期(12 月～3 月) 显著高于非繁殖期(4 月～9 月) (P < 0.01) , 且睾酮水平变化与繁殖行为密切相关。在非繁殖期,雄性小熊猫的睾酮水平处于基础的较低水平时, 其繁殖行为如蹭阴、舔阴、舔标记物和咩叫等几乎没有发生或很少发生; 而在繁殖期, 雄性小熊猫的睾酮水平明显升高, 这些繁殖行为频繁发生。以上结果说明, 睾酮是调节雄性小熊猫繁殖行为季节性变化的生理因子。     

小熊猫妊娠期粪便中孕酮、雌二醇的水平变化

2000年3—8月在成都动物园和成都大熊猫繁育研究基地,用放射免疫分析法测定了6只雌性小熊猫粪样中孕酮（P）和雌二醇（E2）的水平变化,同时对其繁殖行为进行了观察。研究结果表明,交配后的雌体粪样中E2浓度下降至基础水平,P浓度在30d后上升至150ng／g以上;怀孕雌体在妊娠期粪样中P水平在200ng／g以上,P峰值可以达500ng／g以上;假妊娠者粪样中P水平一般不超过150ng／g,P峰值在200ng／g以下;雌体产前半个月孕酮迅速下降,E2缓慢上升,产后P保持在基础水平。这意味着可通过测定粪便中P水平进行早孕诊断、产期预测和真假妊娠判断。根据行为观察和粪样激素分析可知,雌兽妊娠期为123—128d（X=124,N=5）,产仔率为1．8／胎（N=5）。     

Influences of mating groups on the reproductive success of the Southern Sichuan Red Panda (Ailurus fulgens styani)

The reproduction of female red pandas under three different husbandry conditions (monogamy, polygamy, and polyandry) was studied at the Chengdu Research Base for Giant Panda Breeding, and Chengdu Zoological Park, China. We monitored the fecal steroid profiles of seven females by radioimmunoassay in order to understand how the different mating groupings affect the animals' reproductive success. Of the seven females studied, all gave birth except the one that was kept in the polyandrous group. Although fecal estradiol metabolites exhibited several peaks during estrus, only one coincided with mating behavior. Fecal progesterone metabolites did not display a sustained rise in the absence of mating behavior, even though estradiol displayed a peak of secretion. This suggests that the red panda may be an induced ovulator. During gestation, fecal progesterone metabolites were maintained at a comparatively high level, and were much higher than those of nonpregnant females. The female in the polyandrous group showed an abnormal estradiol metabolite profile, with a level that was four to five times higher than those of the females in the polygynous groupings. It was observed that this female always escaped and hid in the trees to avoid courtships with males, and did not give birth. This phenomenon suggests that polyandrous groupings may be unsuitable for successful reproduction in captivity. Zoo Biol 24:169–176, 2005. © 2005 Wiley‐Liss, Inc.     

圈养小熊猫育幼行为的初步观察

Nursing behaviors of the captive md pandas were quantitatively studied by focal sampling methods in Chengdu Research Base of Giant Panda Breeding from Jldy to August, 2000. The results indicated that frequencies of activity and rest were low in the first three days after birth, and became higher as time went by. The frequency of licking cub was higher in the first day after birth and deserting cub was only observed in the seventh day. Frequencies of some behaviors, such as rest, licking cub, cherishing cub,sniffing cub, returning to shed and deserting cub, differed sioaificantly in different phases of the first month after birth. Frequencies of licking cub, cherishing cub, sniffing cub and returning to shed were significantly higher in the pre-nursing period than in the midnursing and post-nursing periods. However, frequencies of rest and deserting cub were sioaificantly higher in the post-nursing period than in the per-nursing and mid-nursing periods. The degree of maternal behaviors can be inferred from degree that the mother exposes her babies to the envim~unent and the time that the mother leaves her shed.     

Microsatellite variability reveals the necessity for genetic input from wild giant pandas (Ailuropoda melanoleuca) into the captive population

Recent success in breeding giant pandas in captivity has encouraged panda conservationists to believe that the ex situ population is ready to serve as a source for supporting the wild population. In this study, we used 11 microsatellite DNA markers to assess the amount and distribution of genetic variability present in the two largest captive populations (Chengdu Research Base of Giant Panda Breeding, Sichuan Province and the China Research and Conservation Center for the Giant Panda at Wolong, Sichuan Province). The data were compared with those samples from wild pandas living in two key giant panda nature reserves (Baoxing Nature Reserve and Wanglang Nature Reserve). The results show that the captive populations have retained lower levels of allelic diversity and heterozygosity compared to isolated wild populations. However, low inbreeding coefficients indicate that captive populations are under careful genetic management. Excessive heterozygosity suggests that the two captive populations have experienced a genetic bottleneck, presumably caused by founder effects. Moreover, evidence of increased genetic divergence demonstrates restricted breeding options within facilities. Based on these results, we conclude that the genetic diversity in the captive populations is not optimal. Introduction of genetic materials from wild pandas and improved exchange of genetic materials among institutions will be necessary for the captive pandas to be representative of the wild populations.     

降低圈养大熊猫排粘液频率的措施初探

大熊猫(Ailuropoda melanoleuca)是中国特有的珍稀动物,在其整个消化道内膜上,都分布有丰富的粘液腺。在野外,这些粘液腺的分泌物包裹于大熊猫摄食的竹茎、竹叶和竹笋表面而排出体外;圈养条件下,常常见到这些分泌物积聚到一定数量而成团排出,这即被称为大熊猫的排粘液现象(王平     

圈养大熊猫群体间的基因流状况分析

圈养群体的遗传管理一个非常重要的手段就是实现不同群体间的基因交流.为了全面评估大熊猫圈养群体间的基因流状况,本研究以卧龙中国大熊猫保护中心的31只圈养大熊猫(简称卧龙群体)和成都大熊猫繁育研究基地与楼观台陕西省珍稀野生动物抢救饲养研究中心的37只圈养大熊猫(简称成都群体,其中楼观台1只)为研究对象,以7个大熊猫微卫星位点为分子标记,发现卧龙群体和成都群体间的遗传分化水平很低(Fst=0.041,P=0.001);尽管整个圈养群体的近交程度较低(Fis=0.026),但是成都群体的近交系数(Fis=0.045)远高于卧龙群体的近交系数(Fis=0.002);谱系分析、贝叶斯分析和系统进化法分析均显示,这两个群体间存在着基因流,但是这种基因流是单向的.此结果提示各个大熊猫饲养单位之间必须实现更多的合作,将大熊猫群体作为一个管理单元进行管理,从而实现更多的基因流.     

圈养小熊猫繁殖行为变化及繁殖行为对策

In order to investigate the change of reproductive behaviors and understand reproductive strategies of both male and female red pandas, one-year behavioral observation was conducted through the focal sampling method in the Chengdu Research Base of Giant Panda Breeding from December 1999 to November 2000. Our results indicated that reproductive behaviors showed significant differences between the estrous and non-estrous seasons. Frequencies of the rubbing anogenital, sniffing and licking marking were much higher in estrus than in non-estrus. Bleating only appeared in the estrus and can be regarded as an estrous indicator.The result also demonstrated that both male and females applied different reproductive behavioral strategies. Frequencies of activity, rubbing anogenital, licking and sniffing marking, and bleat were much higher in the male than in the female. However, those of resting and investigating were much lower in the male than in the female. This indicated that the male was more active than the female during the estrus and might imply that the male acts mainly as an estrous message sender, and the female as a message receiver in the estrous season.     

应用微卫星分型方法进行大熊猫父亲鉴定

1979年我们利用水稻花粉单倍体植株的 茎节、幼穗、叶鞘和叶片等组织进行离体培养, 诱导出二倍体植株。     

圈养大熊猫食谱组成与营养成分分析

为了探讨圈养大熊猫主食竹食谱组成及主要影响因素,并为圈养单位提供科学合理的食物营养配比和饲养管理建议。本研究通过对成都大熊猫繁育研究基地１０ 只成年大熊猫４ 年（２００９. ０３ －２０１３.０２）的饲养数据,３只成年大熊猫１ 年（２０１２. ０７ －２０１３. ０６）的觅食行为观察数据,以及投饲的３ 种竹笋和５ 种主食竹常规养分和总黄酮含量的统计分析。结果发现,基地圈养大熊猫偏好竹笋,喜食巴山木竹叶、白夹竹茎和苦竹茎,随机采食箬竹叶和刺竹茎;基地大熊猫根据投饲竹种类和季节的变化,形成了较为稳定的食谱组成;对投饲竹各部位成分比较分析发现,投饲竹养分含量的变化呈现出一定的规律性：其中粗蛋白、粗脂肪、蛋白质和能量比都以竹叶最高,竹枝次之,竹茎最低;干物质和粗纤维的含量则以竹茎最高,竹枝次之,竹叶最少;竹茎和竹枝呈现钙少磷多的现象,而竹叶则是钙多磷少;各投饲竹不同部位总黄酮含量以叶中最高,枝中其次,茎中最低。研究结果表明,圈养大熊猫主食竹食谱的选择遵循能量和营养学的规律,选择适口性强、营养价值高的食物假说。     

圈养小熊猫的昼夜活动节律

Daily activity rhytlms of three radio-collared captive red pandas were monitored individually at 15 minute intervals for 3 euntinuous days each month at Yele Nature Reserve, Xiangling Mountains, June- October 1995. Our data indicated that mean rate of activity [0.51 ) of captive red pandas was lower than that of wild red pandas. Three captive red pandas showed similar daily activity patterns, being least active during night and more active during daytime. Mean rate of activity during daylight (0.58)was higher than during nighttime (0.41). Daily mean durations of activity and rest were 12.21 hours and 11.79 hours, respectively. Active times of captive red pandas accounted for 50.6% of each 24 hours, of which 66.5% were recorded during daylight and 33.5% during night. Two active peaks appeared at07: 30-09: 15 and 17: 30- 19: 00. We recorded a mean of 2.17, 2.13 and 0.88 long, mid-length, and short resting bouts daily, which had mean durations of 3.47, 1.65 and 0.87 hour, respectively.Among these long rests, 46.1% occurred during daytime and 53.8% during nighttime.     

圈养大熊猫受配后的行为研究

1999～2002年,对成都大熊猫繁育研究基地的8只健康大熊猫20只次受配后行为进行了观察,以了解大熊猫配种后的行为变化,结果发现有以下4种情况：1)妊娠反应明显;2)妊娠反应不明显;3)假妊娠;4)无任何反应。并对这4种情况从行为上进行了探讨,明显的妊娠反应是怀孕产仔大熊猫的普遍规律,在产前27d左右明显减食、拒食,静卧安静休息时间在91％以上,有明显的临产反应,外阴、乳房发生明显的变化;不明显的妊娠反应是怀孕产仔大熊猫的特殊情况,有所减食,静卧安静休息时间无明显变化,也无明显的临产反应,有的外阴变化不大;假妊娠反应类似于明显的妊娠反应,出现在部分受配和未受配的大熊猫中,食欲变化有波动,到一定时间后食欲、活动量及外阴逐渐恢复;无任何反应是受配后一直无任何与妊娠有关的行为表现,食欲、活动情况如常。     

圈养大熊猫催产素水平与母性行为之间关系

Maintaining a good cub-rearing relationship is of great significance for the healthy development of newborn giant panda cubs. Oxytocin plays a key role in this cub-rearing relationship development during the breeding period. To investigate the relationship between oxytocin levels and maternal behavior, we sampled the maternal behavior of 6 adult female giant pandas (3 in lactation group, 3 in non-lactation group) at Chengdu Research Base of Giant Panda Breeding from June to September 2019 by using the focal sampling method, and tested the urine oxytocin level of each individuals by using enzyme-linked immunosorbent assay. The results showed that the oxytocin levels were significantly higher in the lactation group (278. 86 pg/mg ±44. 42 pg/mg) than in the non-lactation group. For the two types of breastfeeding groups, the level of oxytocin in the multiparous female pandas (185. 64 pg/mg ±44. 61 pg/mg) was significantly lower than that in the primiparous female pandas (465. 30 pg/mg ±82. 39 pg/mg). Compared with the primiparous female pandas, the multiparous female pandas had more embracing cub behavior (77. 45% ±1. 24%) and feeding behavior (15. 22% ±1. 62%), but less licking cub (14. 26% ±0. 91%) and position adjustment behavior (1. 69% ±0. 29%). Finally, we found the licking cub behavior was significantly positively correlated with the oxytocin level in the lactation group, and the embracing cub behavior was significantly negatively correlated with the oxytocin level in the lactation group. The results indicate that primiparous female pandas show more maternal behavior, less individual behavior and higher oxytocin levels. The results of this study suggest that the different reactions of oxytocin in the two types of giant panda females may help to improve the refined and differentiated breeding techniques for captive giant pandas in the future.     

圈养小熊猫遗传多样性与种群遗传结构分析

小熊猫是亚洲特有的珍稀濒危动物,目前受到栖息地减少、片断化和人类活动干扰等威胁。中国圈养小熊猫已经有60 多年历史,约55 个机构曾经饲养过小熊猫,现今圈养数量有400 多只,评估小熊猫圈养种群的遗传多样性和遗传结构对科学维持圈养种群和保存遗传种质资源意义重大。本研究利用19 个微卫星座位,对中国境内11 个小熊猫圈养种群的116 只个体进行了遗传多样性评估及遗传结构分析。结果显示11 个种群都具有较高的遗传多样性,平均基因丰富度3.505 ± 1.033 (北京)至4.026 ± 1.219 (冕宁),期望杂合度0.631 ± 0.225(黄山)至0.782 ±0.171 (温岭)。其中福州和无锡种群极显著偏离Hardy-Weinberg 平衡。整个圈养群体内各个种群遗传分化系数为0.055,呈显著分化,表明11 个种群遗传分化水平较高。Bayesian 遗传聚类分析将11 个种群聚为三个遗传簇,与野生种群的遗传聚类结果一致。结论:小熊猫圈养种群与野生种群相比,同样具有较高的遗传多样性。因此,圈养小熊猫遗传管理的重点不再是引进野生个体充实圈养种群,应制订科学的繁殖计划,避免近交,从而维持圈养种群的遗传多样性。     

圈养小熊猫几种病毒的PCR检测

本文采用巢式PCR/ RT-PCR 方法,对我国10 个动物园中无临床症状的圈养小熊猫的71 个肛拭子和61 个唾液拭子样品,进行犬瘟热病毒(CDV)、犬细小病毒(CPV)、犬冠状病毒(CCV)、犬腺病毒(CAV)和犬疱疹病毒(CHV)的检测,以评估我国圈养小熊猫是否面临这几种病毒的威胁。对阳性PCR 结果进行测序分析,并与GenBank 上的序列进行比较。结果,在肛拭子样品中检测到3 个CPV 和6 个CCV 阳性结果,经测序后,与GenBank 上序列的同源性分别达99% 和100% 。而在唾液拭子样品中没有检测到任何阳性结果,且CDV、CAV和CCV 的检测结果均为阴性。从阳性CPV 的肛拭子样品中分离到一株细小病毒毒株,表明圈养小熊猫已受到细小病毒和犬冠状病毒的感染,今后应加强这两种病毒的预防工作。本文所采用的PCR 方法检测病毒性疾病,能检测到微量的病毒模板,可对小熊猫病毒性感染进行早期诊断。     

Sixteen novel microsatellite loci developed for the giant panda (Ailuropoda melanoleuca)

Sixteen novel microsatellite DNA loci were developed from the giant panda (Ailuropoda melanoleuca) using a magnetic-bead capture method. A total of 115 alleles were obtained for these markers, ranging from 4 to 12 alleles per locus (average 7.188). These loci exhibited high levels of polymorphic information content and expected heterozygosity, 0.558–0.855 (average 0.729) and 0.628–0.885 (average 0.778), respectively. Therefore, the allelic polymorphism and heterozygosity show that the giant pandas raised in China Research and Conservation Center possess abundant genetic variation. In addition, if the three markers showing null alleles were excluded, the remaining 13 microsatellite loci still presented extremely low non-exclusion probabilities of parentage (0.002), paternity (0.000) and identity (0.000). As a result, this new suit of microsatellite markers would be a very informative tool for the genetic and conservation studies of giant pandas.     

紫色色杆菌感染小熊猫引起肺炎的临床调查

本文报道了福州大熊猫研究中心小熊猫感染紫色色杆菌引起肺炎的群发病例。7 只小熊猫中,出现以呼吸衰竭为主要临床症状的3 只小熊猫在发病3 d内全部死亡,病死率100% 。死亡的小熊猫解剖病变为气管内有白色泡沫样分泌物,肺有化脓性坏死并严重淤血;胸腔,心包积液且胸水浑浊;肝脏呈弥漫性空泡变性,淤血和散在局灶性坏死。通过流行病学调查,病理解剖观察,细菌分离培养和鉴定及动物回归试验,诊断为紫色色杆菌无色变种感染。根据GenBank 数据库提供的紫色色杆菌16S RNA 基因序列,设计一对引物(5’GAG CAAACA GGA TTA GAT ACC 3’;5’TTA CGG TTA CCT TGT TAC GAC 3’),获得目的基因片段739 bp,将核苷酸序列测定的结果与GenBank 数据库提供的7 株紫色色杆菌进行同源性比较。分离的菌株(FJ08A) 与CV09 株、ESBV4400株同源性均为98.8% ,与AY117554 株、EAV2 株、AJ871127 株、LMG3953 株、JS1 株同源性分别为98. 2% 、98.0% 、94.9%、93.1% 、92.8% 。依据调查结果和药物敏感试验,对其余4 只未发病的小熊猫采取肌
肉注射头孢哌酮钠和口服复方新诺明进行预防性治疗,效果良好,未有新病发生。结果表明:(1)紫色色杆菌是革兰氏阴性菌,致病性强,致死率高,腹腔注射分离的紫色色杆菌菌液导致接种小白鼠在2 ～3 d 内全部死亡。小熊猫感染后,发病迅速,致死率高,应引起重视;(2)紫色色杆菌是一种条件致病菌,外伤感染是主要致病因素。     

An ARMS-based technique for sex determination of red panda (Ailurus fulgens)

Molecular sexing is a key component in the investigation of wild populations. In this study, we developed a fast, accurate and reliable amplification refractory mutation system (ARMS) technique for sex determination of red panda based on the exon 4 of the ZFX/ZFY gene. The amplicons were distinguished simply by agarose gel electrophoresis, exhibiting one fragment in females (X: 300 bp) and two in males (X: 300 bp, Y: 166 bp). Robustness of this ARMS system was confirmed by testing both 43 captive red pandas using DNA samples with known-sex and 10 wild red pandas using faecal DNA samples with unknown sex.     

蜂桶寨自然保护区小熊猫觅食特征和营养对策

2002 年4 ～11 月,在蜂桶寨自然保护区对小熊猫的觅食特征和营养对策进行了研究,并采用Vanderloeg 和Scavia 选择指数Ei衡量小熊猫对竹笋和竹茎年龄的选择程度。结果表明,小熊猫食物由竹叶(70.5% )、竹笋(22.1% )、野果(7.2% )以及毛(0.2% )等组成。小熊猫几乎全以冷箭竹为食,未发现采食短锥玉山竹。春季食物主要由竹叶和竹笋组成,夏秋季主要由竹叶和果实组成,而冬季以竹叶为主。小熊猫对竹笋、竹叶的觅食具有很强的选择性。基径在3 mm 以上被采食竹笋,E_i的值为0.071 9 ～0. 094 4,竹高在10 ～70 cm 未被采食竹笋, E_i 的值为0.149 6 ～0.1989。对不同年龄竹茎和竹叶的选择在各个月份间存在变化,4 ～6 月一年生、二年生和多年生竹竹叶E_i分别为0. 036 2 、0.224 8 和0.487 6, 7 ～8 月三者E_i分别为0.0071、0.0027 和0. 0098,9～11 月特别喜食当年生老笋上的竹叶(E_i =0.552 1)。此外,小熊猫在6 ～ 8 月还特别喜食已展开(53.4% )和未展开的卷曲嫩叶(36.3% ),不喜食较老的竹叶(10.3% )。小熊猫的食性及其对食物的选择体现了在长期进化过程中觅食对策的优化,可最大化觅食过程中的物质或能量摄入。