Reproductive seasonality and maturity of male Neophocaena phocaenoides asiaeorientalis in captivity: A case study based on the hormone evidence

To provide hormone evidence on reproductive seasonality and maturity of the Yangtze finless porpoise, the authors monitored the monthly variations of serum reproductive hormones of a male Yangtze finless porpoise in captivity via radioimmunoassay from 1997 to 2003. It was demonstrated that the immature animal had a serum T level of 28–101?ng?dL^?1 (65?±?52?ng?dL^?1), reached puberty with the serum T concentration ranging between 250 and 590?ng?dL^?1 (390?±?130?ng?dL^?1), and attained reproductive maturity when the serum T level went beyond 1120?±?180?ng?dL^?1 in the breeding season. The body length growth of the animal showed a pattern similar to the serum T variations during its adolescent period from the age of 4 to 6 years. Moreover, the serum T concentration of the male porpoise exhibited significant seasonal variations, indicating that its breeding season may start as early as March and end as late as September.     

Serum concentrations of gonadotropins and steroid hormones of Neophocaena phocaenoides asiaeorientalis in middle and lower regions of the Yangtze River

As a relatively isolated and unique freshwater population, the Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) is the most endangered subpopulation of this species. The objective of this study was to improve our understanding of their reproductive endocrinology by measuring (radioimmunoassay) serum concentrations of FSH, LH, estradiol (E(2)), progesterone (P(4)) and testosterone (T(2)) in free-ranging animals. Blood samples were collected from 66 Yangtze finless porpoises (41 males and 25 females) captured in the middle and lower reaches of the Yangtze River. Based on significant variation of serum T(2) concentrations in males with body length (BL) >138 cm, we inferred that they were mature; in these animals, there were significant seasonal variations in serum T(2) concentrations, with the highest concentrations in March and April (502.0+/-319.8 ng/dL, mean+/-S.D.) and the lowest in December (79.4+/-83.2 ng/dL). Serum T(2) concentrations positively correlated with serum concentrations of LH and weakly correlated with serum concentrations of E(2), whereas there was a significant negative correlation between serum LH and FSH concentrations in males >138 cm. The smallest apparently pregnant female porpoise had a BL of 130 cm. Serum P(4) concentrations ranged from 13.2 to 112.4 ng/mL (43.9+/-28.3 ng/mL) in pregnant females, and fluctuated under 1.0 ng/mL in non-pregnant females with BL>130 cm. Serum LH concentrations were significantly higher in non-pregnant females >130 cm versus those females <130 cm. To our knowledge, this is the first study of endocrine-related maturity and seasonal breeding characteristics of the Yangtze finless porpoise.     

Oestradiol and the preovulatory surges of luteinising hormone and follicle stimulating hormone in ewes during the breeding season and transition into anoestrus

This study was designed to see if giving exogenous oestradiol, during the follicular phase of the oestrous cycle of intact ewes, during the breeding season or transition into anoestrus, would alter the occurrence, timing or magnitude of the preovulatory surge of secretion of luteinising hormone (LH) or follicle stimulating hormone (FSH). During the breeding season and the time of transition, separate groups of ewes were infused (intravenously) with either saline (30 ml h⁻¹; n = 6) or oestradiol in saline (n = 6) for 30 h. Infusion started 12 h after removal of progestin-containing intravaginal sponges that had been in place for 12 days. The initial dose of oestradiol was 0.02 μg h⁻¹; this was doubled every 4 h for 20 h, followed by every 5 h up to 30 h, to reach a maximum of 1.5 μg h⁻¹. Following progestin removal during the breeding season, peak serum concentrations of oestradiol in control ewes were 10.31 ± 1.04 pg ml⁻¹, at 49.60 ± 3.40 h after progestin removal. There was no obvious peak during transition, but at a time after progestin removal equivalent to the time of the oestradiol peak in ewes at mid breeding season, oestradiol concentrations were 6.70 ± 1.14 pg ml⁻¹ in ewes in transition (P < 0.05). In oestradiol treated ewes, peak serum oestradiol concentrations (24.8 ± 2.1 pg ml⁻¹) and time to peak (41.00 ± 0.05 h) did not differ between seasons (P > 0.05). During the breeding season, all six control ewes and four of six ewes given oestradiol showed oestrus with LH and FSH surges. The two ewes not showing oestrus did not respond to oestrus synchronisation and had persistently high serum concentrations of progesterone. During transition, three of six control ewes showed oestrus but only two had LH and FSH surges; all oestradiol treated ewes showed oestrus and gonadotrophin surges (P < 0.05). The timing and magnitude of LH and FSH surges did not vary with treatment or season. In blood samples collected every 12 min for 6 h, from 12 h after the start of oestradiol infusion, mean serum concentrations of LH and LH pulse frequency were lower in control ewes during transition than during mid breeding season (P < 0.05). Oestradiol treatment resulted in lower mean serum concentrations of LH in season and lower LH pulse frequency in transition (P < 0.05). We concluded that enhancing the height of the preovulatory peak in serum concentrations of oestradiol during the breeding season did not alter the timing or the magnitude of the preovulatory surge of LH and FSH secretion and that at transition into anoestrus, oestradiol can induce oestrus and the surge release of LH and FSH as effectively as during the breeding season.     

长江江豚脐带永生化成纤维细胞系建立及细胞生长特性研究

取长江江豚(Neophocaena phocaenoids asiaeorientalis)产后胎盘脐静脉,经组织块培养,差异贴壁法纯化,构建长江江豚的原代细胞系;经外源癌基因SV40 T antigens(猿猴病毒T抗原)转染构建稳定脐带细胞系,并对长江江豚的永生化后的成纤维细胞的细胞形态、转染效率、生长曲线和活率等...     

长江江豚繁殖生物学研究概述

长江江豚,即江豚扬子亚种,是仅分布于长江及附属湖泊中的唯一江豚淡水亚种,也是3个江豚亚种中最濒危的一个亚种。本文概述了30 年来国内外关于长江江豚繁殖生物学方面的研究成果,主要从繁殖生态学、组织解剖学、生理学以及行为学等4 个方面阐述了长江江豚繁殖生物学研究现状,并对其中一些相互矛盾和有争议的问题进行了分析。探讨了今后研究中应注意的重点问题,力图为长江江豚的繁殖生物学研究指明方向,为早日实现长江江豚的人工繁殖奠定基础。
     

Physiological responses to capture and handling of free-ranging male Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis)

We captured free-ranging male Yangtze finless porpoises over three seasons and assayed leukocytes and serum biochemistry to investigate physiological responses to the capture and handlings. Serum thyroid hormones (THs) declined sharply in those porpoises compared with hormone variation in a captive male finless porpoise. Hypernatremia and hypokalemia were also significant in the free-ranging animals suggesting that conservation of serum sodium might be acutely vital for this freshwater subspecies. The animals captured in spring showed more significant neutrophilia and eosinopenia than those captured in autumn suggesting that they may be more affected by capture during the breeding season. Furthermore, physical examination of porpoises when out of the water was apparently stressful, particularly when they were kept out of the water for longer periods. However, an increase in circulating THs may be an adaptive response to accommodate these short-term stresses.     

繁殖期不同时段赤腹松鼠巢域的变化

繁殖期巢域的研究对了解繁殖期间动物的社群关系、种群动态、繁殖策略等有重要作用。2009 年3 ～12月和2010 年3 ～9 月对洪雅县赤腹松鼠的巢域面积变化进行观察。使用最小凸多边形(Minimum convex polygon,MCP)法计算赤腹松鼠各时期的巢域面积,结果显示: (1)整个繁殖期,雄性赤腹松鼠的巢域面积为1.34 ±0. 34 hm² ,显著大于雌性的巢域面积(0.60 ± 0. 08 hm² ); (2)雄性赤腹松鼠在求偶交配期主动增加其巢域面积,可能会增加雄性与雌性的遇见率,从而增加与雌性的交配数量;雌性赤腹松鼠在求偶交配期不主动增加巢域面积,而是具有较为稳定的活动范围。在妊娠育幼期雌性的巢域面积会减小,这可能是雌性为提高后代的成活率,在增加能量和降低捕食风险两者间做出的权衡;(3)整个繁殖期,雌性赤腹松鼠间无巢域重叠现象,而雄性间存在巢域重叠。两性之间仅在求偶交配期存在巢域重叠现象,因此,雌性赤腹松鼠在繁殖期有较强的领域性。     

Topical delivery of retinyl ascorbate: 4. Comparative anti-oxidant activity towards DPPH

The free radical scavenging properties of retinyl ascorbate (RA-AsA) were determined by monitoring the decomposition of 2,2-diphenyl-1-picrylhydrazyl (DPPH) as a function of time and in comparison with ascorbic acid (AsA), ascorbic acid palmitate (AsA-Pal), retinoic acid (RA), retinol (ROL) and retinol palmitate (Rol-Pal). The rate constant of RA-AsA (mean₃±SD) was 4.9±0.3 M^-1 s^-1, and indicated greater potency as an antioxidant compared to the rest of the test compounds (AsA 3.4±0.4 M^-1 s^-1, AsA-Pal, 2.9±0.2 M^-1 s^-1, RA 1.4±0.3 M^-1 s^-1, ROL 1.3±0.1 M^-1 s^-1, Rol-Pal exhibited insignificant activity). The decomposition rate constant of DPPH, 5±0.6 × 10^-8 M^-1 s^-1, in ethanol and BHA, 154±3 M^-1 s^-1 were both used as control. The compound RA-2-carboxy-2-hydroxy-ethanoate was isolated by prep-TLC and was identified, by ¹³C and ¹HNMR spectroscopy, as the major by-product from the reaction of RA-AsA with DPPH, which was also found to be potent antioxidant, 2.1±0.2 M^-1 s^-1. This suggests that oxidation of AsA moiety did not lead to the production of erythrulose species, which could cause deleterious modifications of cellular proteins.     

不同流态中长江江豚的栖息活动比较

长江江豚(Neophocaena asiaeorientalis)呈斑块状分布在长江干流,对栖息地有明确的选择性.流态是栖息地水文环境的核心内容.为研究长江江豚对不同流态的选择性,本文根据1993-2021年对长江下游15次样线考察记录到的412次,合计1 117头江豚个体的栖息活动进行了统计分析.结果表明:全年长江江...     

The potential role of microzooplankton in a northwestern Australian pelagic food web

The role of microzooplankton in waters adjacent to Australia's North West Cape (21°49'S 114°14'E) was studied during the austral summers 1997/1998 and 1998/1999. We estimated microzooplankton abundance and biomass at a shallow (∼20 m) shelf station and at a shelf break station (∼80 m). Microzooplankton were placed into six categories: four ciliate groups (strombidiids, strobilidiids, tintinnids, “other ciliates”), dinoflagellates, and sarcodines. Total microzooplankton abundances ranged between 0.14×10³ l^-1 and 3.4×10³ l^-1. The most abundant groups were the dinoflagellates (mean 459±73 standard error l^-1) and strombidiids (mean 334±42 standard error l^-1). Total microzooplankton biomass ranged between 0.03 and 1.70 µg C l^-1 (mean 0.33±0.05 standard error l^-1). Redundancy analysis indicated differences in microzooplankton community composition between stations and sampling years but no differences with sampling depth. The microzooplankton community showed considerable variability between adjacent sampling dates, reinforcing the conclusion of earlier studies that this area is a dynamic environment. Ciliate production on the shelf was estimated to be 1.05 µg C l^-1day^-1 (∼20 mg C m^-2day^-1) and 0.79 µg C l^-1day^-1(∼70 mg C m^-2day^-1) at the shelf break. Ciliate production near North West Cape was two- to six-fold higher than the rate of secondary production by juvenile copepods. Despite this, ciliate grazing appears to account for only ∼5% of primary production and ciliates do not appear to be a major conduit between primary producers and higher trophic levels in these waters.     

Characteristics of microbial fermentation and potential digestibility of fiber in the hindgut of dugongs (Dugong dugon)

The number of microorganisms in the hindgut of dugongs (Dugong dugon) were estimated and their in vitro volatile fatty acid (VFA) production and degradation of eelgrass measured. Scanning electron microscopy showed that some rod bacteria attached to the surface of plant tissue degraded and eroded the cell walls. Number of starch-, lactate-, cellobiose-, pectin-, xylan- and cellulose-utilizing bacteria, sulfate-reducing bacteria and methane-producing bacteria were estimated at 10⁹ ∼ 10¹⁰ colony forming units g^-1. Microorganisms degraded the cellulose and noncellulolytic components of the eelgrass, and about 47.3% of dry matter was degraded after 36 h in vitro incubation. The total VFA concentration was 10.5 mmol dL^-1 at 36 h incubation, which included 55.7 mol% acetate, 18.0 mol% n-butyrate and 15.1 mol% propionate. The gas composition of in vitro fermentation was 68.4% carbon dioxide, 22.2% methane and 9.4% hydrogen.     

Beta-naphthoflavone inhibits the induction of hepatic oestrogen-dependent proteins by 17alpha-ethynylestradiol in mosquitofish (Gambusia holbrooki)

The interactive effects of an aryl hydrocarbon receptor (AhR) agonist and of a xenoestrogen on biomarker responses were studied in the liver of male mosquitofish (Gambusia holbrooki). Hepatic 7-ethoxyresorufin O-deethylase (EROD) enzymatic activity was measured as a biomarker of exposure to the model AhR agonist beta-naphthoflavone (bNF). Hepatic proteins indicating the exposure of males to the synthetic oestrogen 17alpha-ethynylestradiol (EE2) were monitored by Western blot analysis using immunoserum prepared for this study. After a semi-static exposure only to waterborne EE2, Western blot analysis of liver homogenate revealed the induction of two protein bands (a double band at 205 kDa and a single band at 125 kDa). The interaction between bNF and EE2 was investigated by analysing, on the one hand, EROD activity and, on the other hand, immunoreactivity corresponding to the two oestrogen-dependent protein bands in the liver of fish exposed to different concentrations of bNF for 2 days, then to the same concentrations of bNF plus 0.1 µg l^-1 EE2 for 5 days. EE2 changed neither the basal activity of EROD nor its rate of induction with 1.0 and 4.0 µg l^-1 bNF. On the other hand, the induction of oestrogen-dependent proteins with 0.1 µg l^-1 EE2 was inhibited by exposure to 4.0 µg l^-1 bNF. These results together with literature data suggest that field monitoring of xenoestrogen contamination through the analysis of oestrogen-dependent protein in male fish as a biomarker should take into account the possible negative interference of AhR agonists.     

B-Mode ultrasonographic evaluation of the testis in relation to serum testosterone concentration in male Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) during the breeding season

The use of ultrasonography as a noninvasive tool for assessing the reproductive status of the male Yangtze finless porpoise (YFP; Neophocaena phocaenoides asiaeorientalis) was validated by correlating ultrasonographically determined testicular volume (TV) and testicular parenchyma pixel intensity (PI) with serum testosterone (T) concentration. The testes of 13 free-ranging male YFPs from the Tian-e-Zhou Reserve and three captive animals from the Baiji Dolphinarium (Wuhan, China) were examined ultrasonographically during April 2008. Testis volume was determined using Lambert's formula for an ellipsoid. Testicular parenchyma PI was evaluated by analyzing testicular ultrasonograms using pixel analysis software (Image J). Serum T concentrations were determined using a single-antibody radioimmunoassay. The TV, PI, and serum T concentration were low and similar in animals with body length <133 cm, highest in those with body length ≥142 cm, and highly variable in those with body length from 133 to 141 cm. Both TV and PI were closely correlated with serum T concentration (r = 0.91 and r = 0.85, respectively; P < 0.01), indicating a consistent association between structural and functional development of the testis. In conclusion, we inferred that puberty onset in male YFPs occurred when TV was >150 cm³ and PI was >60 during the breeding season and that testicular ultrasonography and pixel analysis was an efficient, noninvasive, real-time tool to evaluate testicular function of live male YFPs.     

北部湾印太江豚密度估算及适宜栖息地预测

目前我国印太江豚 (Neophocaena phocaenoides) 的基础信息十分有限,2018—2020年对北部湾离岸较近的广西壮族自治区合浦儒艮国家级自然保护区及离岸较远的涠洲岛水域进行了截线抽样法船只考察。应用DISTANCE估算出儒艮国家级自然保护区印太江豚密度约为每平方千米0.273 (0.133 ~ 0.561) 头,数量约88头 (95%CI：43 ~ 181);涠洲岛水域印太江豚密度约为每平方千米0.100 (0.048 ~ 0.210) 头,数量约137头 (95%CI：65 ~ 286)。利用最大熵模型 (MaxEnt) 分析北部湾水域印太江豚的适宜栖息地分布及面积。MaxEnt模型训练集和测试集AUC值分别为0.980和0.927,模型拟合度较好,结果表明离岸距离和海洋深度是影响印太江豚栖息地分布的主要因子,印太江豚的适宜栖息地在北部湾东北部近岸区和海南西侧,适宜度超过0.5的栖息地面积达14 630.62 km²。2018—2020年印太江豚适宜栖息地面积较2013年大幅度下降,0.5适宜度的栖息地面积减少了40%。参考印太江豚2个群体的平均密度和北部湾适宜栖息地面积,粗略估计北部湾水域印太江豚约1 463 ~ 3 994头。由此可见,北部湾水域内印太江豚密度比较高,应引起足够的重视。建议首先进行区域性海洋环境的整体保护,重点关注鱼类资源的恢复及对人类活动尤其渔船的管控,并尽快建立新的自然保护区。     

长江江豚自然保护区建设管理存在的问题及调整建议

长江江豚(Neophocaena asiaeorientalis)是生活在中国长江的珍稀濒危鲸类动物, 自然栖息地破坏是造成其种群衰退的重要原因。目前中国已经在长江中下游流域建立了8个长江江豚自然保护区。然而, 早期保护区的功能区划随经济发展和生态保护的现实矛盾日渐突出, 按照现行法规对保护区难以实行有效管理, 迫切需要进行优化调整。文章基于国家政策、长江江豚生物学特点及分布规律, 兼顾航运等重大经济发展需求, 提出了长江豚类自然保护区调整的原则和建议: (1)将长江江豚喜好的近岸水域及一定范围的重要江滩和洲滩湿地, 及江心洲汊江一侧水域划入核心保护区; (2)将中间主航道水域以及长江大堤至核心区边界之间的水域划入一般控制区; (3)兼顾历史遗留问题, 细化不同区域的管控措施。该调整方案一方面使保护区功能区划更加符合长江江豚的分布特征和偏好生境, 有利于对其自然种群和重要栖息地的有效保护。另一方面也有助于理顺自然保护区管理上存在的法律障碍, 为长江江豚自然保护区建设、规划和管理提供新的思路和模式。     

Establishment and characterization of fibroblast cell lines from the skin of the Yangtze finless porpoise

The Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis), as the sole freshwater subspecies of N. phocaenoides, is endemic to the Yangtze River and its adjacent lakes. Its population has declined significantly over recent decades. In this study, we established a skin-derived finite fibroblast cell line of the Yangtze finless porpoise, named YFP-SF1, using primary cell culture methods, and an immortalized cell line, T-YFP-SF1, through co-transfection (GFP and SV40 T antigens) techniques. YFP-SF1 proliferated continuously with a minimum population doubling time of 31 h and exhibited age-dependent changes in growth rate. T-YFP-SF1 cells exhibited fibroblast morphology and were characterized by a shorter doubling time, higher attachment efficiencies, colony formation at a low seeding density, and growth in low serum concentrations. Anchorage independence and foci formation in the cell monolayer were observed from passage 36. The chromosome number of YFP-SF1 and T-YFP-SF1 remained stable at 2n = 44 in the early passages, and the viability of thawed cells remained above 90% after cryopreservation in liquid nitrogen. Taken together, we have established fibroblast cell lines of Yangtze finless porpoise for the first time, which might assist as an in vitro model for this endangered mammal.     

Seasonal changes in the local distribution of Yangtze finless porpoises related to fish presence

The Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis) is an endangered freshwater porpoise subspecies unique to the Yangtze River basin. Seasonal variations in local distribution of the animal, as well as fish presence, sand dredging, ship navigation, and bridges were examined as potential factors affecting the occurrence of the animals. Passive acoustic surveys were performed regularly from May 2007 to August 2010, near the conjunction of the Yangtze River and Poyang Lake. The distribution of the porpoises was seasonally site‐specific. In May and August, the animals were detected more often at river junctions than in the lake, but vice versa from November to February. The rate of the porpoise detection was significantly higher in areas of fish presence than in areas of absence. The number of porpoises detected did not differ significantly between the sand dredging operation and the prohibition period (in 2008), although the number of vessels obviously declined in 2008. Ship traffic and bridges also did not appear to affect the presence of porpoises. These results showed the relative importance of the various environmental factors, which is important for conservation of not only Yangtze finless porpoise but also endangered isolated cetaceans.     

长江东流河道整治对长江江豚种群数量的影响

长江江豚是生活在长江中下游的濒危水生哺乳动物,通常活动在江心洲和浅水缓滩附近。航道整治改变了鱼类栖息活动的水文环境,也给豚类生存带来负面影响。本文根据长江东流河道整治施工前和施工后各3 次野外考察资料,分析了3 种流态中江豚的数量与群结构。结果表明,该工程实施前江豚主要栖息活动在边滩的分离区,其次在洲头的分流区。工程实施后,整治江段的江豚种群数量年下降率达8. 9% 。江豚在分流区活动已消失,在分离区集群规模较小,且在流态之间移动增大。本文研究结果也提示长江江豚就地保护难度越来越大,从长江干流中把长江江豚迁入故道中是一件刻不容缓的工作。     

Nitrogen and Phosphorus for Growth of Oil-Degrading Microorganisms in Seawater

Seawater was supplemented with NH₄⁺ and P to determine concentrations of N and P adequate for supporting exponential growth of bacteria utilizing crude oil, and to determine maximum rates of N and P uptake. Oil-degrading microorganisms were obtained by enrichment culture of indigenous oil-utilizing microorganisms in seawater. NH₄⁺ at a concentration of 5.5 µM was limiting to growth of bacteria on crude oil. Exponential growth occurred at concentrations higher than 30 µM NH₄⁺. The P concentration of 0.13 µM was limiting to growth of bacteria on crude oil. Exponential growth occurred at 1.8 |J,M P. The maximum NH4+ consumption rate was 426 ± 30 |J,g NH₄⁺ L^-1 hr^-1, and the maximum uptake rate of P was 48±4 µg P L^-1 hr^-1. Uptake of N and P with time showed zero-order kinetics, likely due to substrate solubility limitations. The uptake ratio of N:P was approximately 7:1 on a weight basis. Natural concentrations of N and P in marine and estuarine systems after hydrocarbon spillage initially may not limit oil biodegradation but may become limiting if adequate flux does not occur to replenish N and P depleted by microbial consumption.     

Hydrogen peroxide degradation by immobilized cells of alkaliphilic Bacillus halodurans

Whole cells of Bacillus halodurans LBK 261 were used as a source of catalase for degradation of hydrogen peroxide. The organism, B. halodurans grown at 55°C and pH 10, yielded a maximum catalase activity of 275 U g^-1 (wet wt.) cells. The catalase in the whole cells was active over a broad range of pH with a maximum at pH 8-9. The enzyme was optimally active at 55°C, but had low stability above 40°C. The whole cell biocatalyst exhibited a K_m of 6.6 mM for H₂O₂ and V_max of 707 mM H₂O₂ min^-1 g^-1 wet wt. cells, and showed saturation kinetics at 50 mM H₂O₂. The cells were entrapped in calcium alginate and used for H₂O₂ degradation at pH 9 in batch and continuous mode. In the batch process, the immobilized preparation containing 1.5 g (wet wt.) cells could be recycled at least four times for complete degradation of the peroxide in 50 mL solution at 25°C. An excess of immobilized biocatalyst could be used in a continuous stirred tank reactor for an average of 9 days at temperatures upto 55°C, and in a packed bed reactor (PBR) for 5 days before the beads started to deform.