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.

Ultrastructure of the Spermatozoa of the Yangtze Finless Porpoise (Neophocaena phocaenoides asiaeorientalis)

P>Semen sample was collected from two captive adult Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) during physical examination. One individual was aged about 9 years with body length 143 cm (total length) and body weight 46.1 kg in 2003. The age of the other was unknown and its body length was 147 cm and body weight was 43 kg in 2004. Ultrastructure of their spermatozoa was examined using scanning and transmission electron microscope. The sperm concentration was 4.17 x 10(9) spermatozoa per ml by the cytometer. The approximate dimensions of the spermatozoa were as follows: head length, 3.366 +/- 0.140 mu m (mean +/- SE, n = 15); head width, 1.896 +/- 0.099 mu m (n = 15); and neck length, 1.004 +/- 0.074 mu m (n = 10). The tail included midpiece, principal piece and terminal piece. The length of the midpiece was 1.882 +/- 0.077 mu m (n = 9). There is no apparent boundary between the principal piece and the terminal piece, so the length of the principal piece and the terminal piece was 44.612 +/- 3.485 mu m (n = 5). Total length of the spermatozoa was 53.314 +/- 4.580 mu m (n = 10). The acrosome covered approximately 45.8% of the anterior portion of the head.

Development and characterization of polymorphic microsatellite loci in the endangered Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis)

The finless porpoise (Neophocaena phocaenoides) is one of the smallest cetacean species widely distributed in the shallow coastal waters of the Indo-Pacific Oceans. The population size of the Yangtze subspecies (N. p. asiaeorientalis) has sharply decreased in the last two decades and access to objective data on its population structure and genetic diversity would be of great assistance for their proper management. Here we report on the isolation of nine polymorphic microsatellite using the "Fast Isolation by AFLP of Sequences Containing repeats" (FIASCO) protocol. Polymorphism was assessed using 30 porpoise individuals randomly sampled in the Yangtze River. The number of alleles per locus varies from 2 to 9, with an average value of 5.56, whereas the ranges of observed and expected heterozygosi ties were 0.300-0.633 (mean 0.496) and 0.473-0.804 (mean 0.659), respectively.

Estimation of the detection probability for Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) with a passive acoustic method

Yangtze finless porpoises were surveyed by using simultaneous visual and acoustical methods from 6 November to 13 December 2006. Two research vessels towed stereo acoustic data loggers, which were used to store the intensity and sound source direction of the high frequency sonar signals produced by finless porpoises at detection ranges up to 300 m on each side of the vessel. Simple stereo beam forming allowed the separation of distinct biosonar sound source, which enabled us to count the number of vocalizing porpoises. Acoustically, 204 porpoises were detected from one vessel and 199 from the other vessel in the same section of the Yangtze River. Visually, 163 and 162 porpoises were detected from two vessels within 300 m of the vessel track. The calculated detection probability using acoustic method was approximately twice that for visual detection for each vessel. The difference in detection probabilities between the two methods was caused by the large number of single individuals that were missed by visual observers. However, the sizes of large groups were underestimated by using the acoustic methods. Acoustic and visual observations complemented each other in the accurate detection of porpoises. The use of simple, relatively inexpensive acoustic monitoring systems should enhance population surveys of free-ranging, echolocating odontocetes. (C) 2008 Acoustical Society of America.

The ontogeny of echolocation in a Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis)

Acoustic and concurrent behavioral data from one neonatal male Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) in captivity were presented. The calf click train was first recorded at 22 days postnatal, and the frequency of hydrophone-exploration behavior with head scanning motions in conjunction with emissions of click trains by the calf increased gradually with age. The echolocation clicks in the first recorded click train were indistinguishable from those of adults. Calf echolocation trains were found to decrease in maximum click-repetition rate, duration, and number of clicks per train with age while the. minimum click-repetition rate remained more consistent. (c) 2007 Acoustical Society of America.

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. (c) 2006 Elsevier Inc. All rights reserved.

Respiratory pattern of captive Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis)

Cetacean respiration usually happen in bouts. The most widely applied quantitative method used to analyze the structure of these bouts is the log(e)-survivorship analysis, based on the assumption that the respiratory intervals are distributed as negative exponentials. However, for the data collected from three captive Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis), we failed to obtain a convergent result with the application of log,survivorship analysis. However, the two-Gaussian model, which was recently proposed to analyze the feeding behavior of cows, was successfully fitted to the data. According to the fitting results, the overall respiratory pattern of the captive Yangtze finless porpoises can be described as a dive with a mean duration of around 30-40 s, followed by two or three ventilations with a mean interval of approximately 9 s. The average intra-bout intervals during both active and inactive periods are constant at 7.7-9.9 s for all individuals. However, when shifting from active to inactive states, the adult male and female decrease their mean numbers of respirations per bout and average length of inter-bout respiratory intervals, while the estimates of both parameters increase for the juvenile female. It was pointed out that the two-Gaussian model might be more adequate for cetacean respiratory-bout structure analyses than the log(e)-survivorship technique.

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 28-101ngdL -1 (6552ngdL -1 ), reached puberty with the serum T concentration ranging between 250 and 590ngdL -1 (390130ngdL -1 ), and attained reproductive maturity when the serum T level went beyond 1120180ngdL -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.

Mercury concentrations in Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) from Eastern Dongting Lake, China

Mercury concentrations in some key tissues (liver, kidney, small intestine, stomach, blubber and brain) of five Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis), accidentally died in the Eastern Dongting Lake of China between 1998 and 2004, were investigated using Atomic Fluorescent Spectrometry (AFS). Total mercury (T-Hg) concentrations ranged between 0.17 to 181 mu g g wet weight. and varied significantly in different tissues. Liver is the most preferable target organ for mercury accumulation. although the highest T-Hg concentrations were not found exclusively in liver, but also in kidney and small intestine in some individuals. Moreover, it was demonstrated that mercury concentrations increased positively with age of the animals. However, the highest T-Hg concentrations in liver and kidney were both found in a 2-month-old calf, which implied that the transfer efficiency of mercury from mothers to babies in Yangtze finless porpoise is remarkable. And what is more important, it seems that the Yangtze finless porpoise in Eastern Donating Lake had much higher T-Hg levels than those reported for other Phocoenidae species. To be noticed, the T-Hg was accumulated tremendously from aquatic environment to the Yangtze finless porpoise's liver, reaching a bioaccumulation factor (BAF) of 4.3x10(5) in the Eastern Donating Lake ecosystem.

Diurnal changes of behavior and respiration of Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) in captivity

A preliminary study was carried out to investigate diurnal changes of behavior of three, one adult mate, one adult female, and one juvenile female, Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) in captivity. The respiration and behavior of the porpoises were recorded for 222 hr across 42 days. Behavioral data were recorded for eight general categories: aerial display and fast swimming, begging for fish, playing, nonsexual socializing, sexual behavior, resting, rubbing, and miscellaneous (i.e., other behaviors not included in the above categories). Each behavioral category was scored using one-zero sampling with 10-min intervals. The adult male showed shorter mean respiratory intervals at night (19:00-7:00 h), whereas the mean respiratory intervals of the females were shorter during the day (7:00-19:00 h). Begging for fish of all individuals, playing of the juvenile female, nonsexual socializing, and miscellaneous behavior of the adult female and resting of the male were observed more easily in the day, and aerial display and fast swimming of the adults and resting of the females were observed more easily at night. No significant diurnal difference was found, however, in the remaining categories of each individual. Each of the three porpoises therefore showed a distinct diurnal pattern, but none was obviously more active in the daytime than during the nighttime. Results suggest that daytime-only feeding schedules may be insufficient to meet the energetic needs of marine mammals that show a 24-hr activity cycle, and that nighttime feeding may be a worthwhile addition to husbandry routines.

Ex situ conservation status of an endangered Yangtze finless porpoise population (Neophocaena phocaenoides asiaeorientalis) as measured from microsatellites and mtDNA diversity

The Yangtze finless porpoise, Neophocaena phocaenoides asiaeorientalis, is all endangered small cetacean that occurs only in the middle and lower reaches of the Yangtze River of China. The establishment of a breeding population of the porpoise in Tian-e-Zhou Baiji National Natural Reserve represents the first attempt at ex situ conservation efforts for a cetacean species. With the goal of effective protection, management, and monitoring of this preserved population, we examined its genetic diversity using 930 bp of mtDNA control region sequences and 13 polymorphic microsatellite loci. A very low level of genetic variation (h = 0.6010 +/- 0.0029 s.d.; pi = 0.0007 +/- 0.0000002 s.d.) in the mtDNA control region sequences and a moderate genetic diversity (Ho = 0.5740 +/- 0.2575 s.d.) in the microsatellites were detected in the population. It is necessary to introduce more individuals with representative genetic variations into the reserve ill order to foml a larger and healthier group structure for long-term survival of the population. Successful establishment of the Yangtze finless porpoise population in the Reserve also provides a useful model for an ex situ conservation programme for other rare and endangered species. (c) 2005 International Council for the Exploration ofthe Sea. Published by Elsevier Ltd. All rights reserved.

Population genetic structure of the Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis): Implications for management and conservation

Understanding the population genetic structure is a prerequisite for conservation of a species. The degree of genetic variability characteristic of the mitochondrial DNA control region has been widely exploited in studies of population genetic structure and can be useful in identifying meaningful population subdivisions. To estimate the genetic profile of the Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis), an endangered freshwater population endemic to China, the complete mtDNA control region was examined in 39 individuals belonging to seven different stocks inhabiting the middle and lower reaches of the Yangtze River. Very low genetic diversity was found (nucleotide diversity 0.0011 +/- 0.0002 and haplotypic diversity 0.65 +/- 0.05). The mtDNA genetic pattern of the Yangtze population appears to indicate a founder event in its evolutionary history and to support the marine origin for this population. Analyses by F-st and Phi(st) yielded statistically significant population genetic structure (F-st = 0.44, P < 0.05; phi(st) = 0.36, P < 0.05). These results may have significant implications for the management and conservation of the Yangtze finless porpoise in the future.

Parentage determination of an isolated Yangtze finless porpoise population Neophocaena phocaenoides asiaeorientalis in the Yangtze Tian-e-Zhou Baiji National Natural Reserve based on molecular data

Reproductive behaviors are poorly known for the Yangtze finless porpoise Neophocaena phocaenoides asiaeorientalis. In this study, the parentage of an isolated Yangtze finless porpoise population inhabiting the Yangtze Tian-e-Zhou Baiji National Natural Reserve is determined by analysis of microsatellite loci and mitochondrial DNA (mtDNA) control region sequences, and the porpoise's reproductive behaviors are studied. Overall 4 full parentage assignments and additional 3 single parentage assignments were determined for the population of 23 individuals. The analysis shows that their estimated reproductive cycle is shorter than that reported previously and there probably exists an overlapping between gestation and lactation period. The Study also shows that the female does not show fidelity to a particular male for breeding and vice versa, the oldest males did not monopolize mating and the dominance rank could not be so strict for the porpoise society. Moreover, the porpoise's mating pattern and relatedness among candidate parents are discussed here. These results provide important information for making guidelines of management and conservation for this protected population.

Evoked-potential audiogram of the Yangtze finless porpoise Neophocaena phocaenoides asiaeorientalis (L)

Evoked-potential audiograms were obtained in two (one male and one female) Yangtze finless porpoises, Neophocaena phocaenoides asiaseorientalis. Sinusoidal amplitude-modulated 20-ms tone bursts were used as probes with recording envelope-following evoked potentials. A frequency range of 8 to 152 kHz was investigated. The range of greatest sensitivity covered frequencies from 45 to 139 kHz, and the lowest thresholds of 47.2 and 48.5 dB re: 1 μ Pa were found at a frequency of 54 kHz in the two subjects, respectively. At lower frequencies, threshold increased with a rate of around 14 dB/octave, and threshold steeply increased at 152 kHz. © 2005 Acoustical Society of America.

Observations on behavior and ecology of the Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) group at Tian-e-Zhou Oxbow of the Yangtze River

The authors made 39 surveys (a total of 161 days) in the Tian-e-Zhou Oxbow of the Yangtze River, China, for observing 13 Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) captured from the main stream of the Yangtze River and 7 juveniles born in the oxbow from January 1997 to July 2000. The animals were usually divided into several "core" groups and moved around in shallow, muddy-bottom areas with the largest individual in the lead. Each core group was composed of 2-3 animals (either 2 adults, 1 adult and 1 juvenile, 2 adults and 1 juvenile, or 2 adults and 1 calf). Newly-released animals joined the other animals first, and then reorganized their own groups one or two days later. Average breath interval was 34.4 s (+/- s.d. 4.39) for individuals in the group. The animals mated from May through June and gave birth during the second and last ten days of April of the next year. The gestation period was estimated as 310 - 320 days. Calves over 5 months old began to eat small fish. The distance of calves swimming apart from their suspected mothers increased each month. These findings will help in the management of the reserve to protect this unique freshwater porpoise.