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However, there is no report of the birth of elephant calves following AI with frozen-thawed semen. The objective of the present study was to investigate the fertilizing ability of chilled and frozen-thawed semen in the Asian elephant following artificial insemination AI.

Semen samples were collected by from 8 bulls age range, to years by manual stimulation. Blood samples collected from ten elephant females age range, to years were assessed for estrus cycle and elephants with normal cycling were used for AI.

Artificial insemination series were conducted during to ; 55 and 2 AI trials were conducted using frozen-thawed and chilled semen, respectively.

Pregnancy was detected using transrectal ultrasonography and serum progestagen measurement. One female Khod inseminated with chilled semen became pregnant and gave birth in The gestation length was days and the sex of the elephant calf was male.

One female Sao inseminated with frozen-thawed semen showed signs of pregnancy by increasing progestagen levels and a fetus was observed for 5 months by transrectal ultrasonography.

This is the first report showing pregnancy following AI with frozen-thawed semen in the Asian elephant. Successful AI in the Asian elephant using either chilled or frozen-thawed semen is a stepping stone towards applying this technology for genetic improvement of the elephant population.

As the number of wild elephants continues to decline, maintaining healthy captive populations is vital. Because of difficulties in transporting a bull elephant from one institution to another for natural breeding, artificial insemination AI is an alternative tool for genetic management in this species.

Although elephant calves have been produced following AI using either fresh or chilled semen [ 1 — 3 ] there has been no report of the birth of elephant calves after AI with frozen-thawed semen.

Our previous studies showed that acceptable semen quality can be obtained after freezing and thawing [ 4 , 5 ]; however, pregnancy was not established after AI [ 6 ], indicating a need to develop an effective cryopreservation technique for Asian elephant spermatozoa.

Two AI techniques in elephants have been reported; i. Both techniques are impractical in Thailand. Surgical AI is not suitable to use due to the complication of wound infection and concerns for animal welfare, whereas endoscopic AI requires special accessory equipment: a balloon catheter to distend the reproductive tract [ 1 ].

Therefore, the objective of this study was to investigate the fertilizing ability of either chilled or frozen-thawed semen using the modified endoscopic AI technique described herein.

All animals and the experiments were approved by the National Elephant Institute. Eight elephant bulls age range, to years-old housed at the National Elephant Institute, Forest Industry Organization, Lampang, Thailand were used as semen donors.

Ten elephant females age range, to years were used for AI. The elephants were fed with grass, banana and sugar cane during the day and allowed to roam in the jungle at night.

Semen samples were collected by manual manipulation as described by Schmitt and Hildebrandt [ 7 ]. Each ejaculate was immediately evaluated for volume, sperm concentration, progressive motility, sperm viability and pH [ 8 ].

Sperm concentration was assessed using a haemocytometer. Progressive motility was assessed visually under a phase-contrast microscopy by two independent technicians.

Semen samples collected from 6 bulls Dindum, Pamae, Yuak, Koong, Tadaeng and Jumpui were frozen by the method as previously described [ 4 ].

Briefly, fresh semen was diluted with TEST extender at room temperature. TEST extender consisted of 5.

The equilibrated mixture was packed in a 0. The straws were held at 2. Ten microlitres of the thawed semen were then taken and placed on a pre-warmed microscopic slide and the progressive motility of post-thaw spermatozoa was assessed under light microscopy.

Semen samples collected from 2 bulls Japatee and Sai were prepared and used for AI as chilled semen. Fresh semen was diluted with TEST extender [ 4 ] to a final volume of ml.

A ml blood sample was collected from an ear vein weekly for hormonal assay. When progestagen concentrations decreased to baseline i.

Blood samples were allowed to clot at room temperature for 1 to 2 h, and then centrifuged at g for 5 min to separate serum from blood cells.

Serum samples were transferred into 1. Serum progestagens were analyzed using enzyme immunoassay EIA as previously described [ 9 ]. This EIA employed a monoclonal antibody against progesterone ,; Quidel clone Quidel clone ; supplied by C.

Munro, Davis, CA, USA that has been shown to cross-react with a variety of reduced progesterones in the serum and excreta of a wide range of species, including elephants [ 10 ].

Bound progesterone was visualized using a horseradish peroxidase-conjugated antibody to progesterone ,; C. The sensitivity of the assay was 0.

This monoclonal antibody was validated and approved to detect LH in Asian elephants [ 3 , 11 ]. Sensitivity of the assay was 0. The inter-assay coefficient of variation CV for the high concentration control was 7.

Artificial insemination series were conducted during to Fifty five and two AI attempts were conducted with frozen-thawed and chilled semen in six and two elephant females, respectively.

Three inseminations once per day per series were performed on days -1, 0, 1 of the ovulatory LH surge [ 3 ].

Fifty milliliter of semen was used per inseminations. The female was standing immobilized using 0. For AI, the endoscopic technique was modified by increasing inflated air from external electric air pump without using the balloon catheter.

The inflated air could enlarge the vestibular, vaginal and cervical canal. After the tip of endoscope reached and fixed at the cervical canal, the air tube was removed and replaced by insemination catheter IC.

The IC was passed through the working channel of the endoscope and penetrated deep into the body of uterus. The semen was deposited into body of uterus without signs of semen reflux Figure 1.

Pregnancy diagnosis was performed by serum progestagen measurement and transrectal ultrasonography. Blood samples were collected weekly and serum progestagens were assessed by EIA as described above [ 3 ].

Transrectal ultrasonography was performed weekly between 2 and 4 months after AI as previously described [ 1 ]. Positive PCR products generated by specific primers of microsatellite markers were fractionated on a 1.

Parentage analysis was performed by parentage exclusion method [ 18 ]. A total of 24 ejaculates obtained from six bulls were prepared as frozen semen.

Data on semen quality of fresh, chilled and frozen-thawed samples are shown in Table 1. The average sperm motility of frozen-thawed semen was A progressive reduction of the percentage of viable sperm was observed.

Eight out of 10 elephants assessed for estrus cycle showed normal cycling and were selected for AI. The onset of the luteal phase was defined as the first point at which serum progestagen concentrations increased above 0.

At the end of luteal phase, serum progestagen concentrations started to decline and reached the baseline 0. In case of AI with chilled semen, one out of two females, namely "Khod", showed progressive increase of serum progestagens after AI Figure 2.

Serum progestagen started to decline to the baseline one month before birth Figure 2. Transrectal ultrasonography of pregnant cow showed enlargement of the uterine wall at 2 months after AI.

An increase of uterine lumen that filled with clear fluid was found between 3 and 4 months. We were unable to find a fetus between 2 and 4 months after AI.

Some changes in Khod's behavior during pregnancy period were observed. Khod's anatomy was not remarkably changed during pregnancy period except that her belly was enlarged during the last sixth month period.

Furthermore, one month before calving, her breasts were enlarged and had a clear secretion released when they were stimulated. Khod gave a birth on March 7, with birth weight of 51 kg.

The gestation length was days and the sex of elephant baby was male Figure 3. The calf was apparently normal in health and behaviour and had a normal suckling reflex.

The AI and parturition date confirmed pregnancy of days. Discontinuous lines represent missing data due to blood samples could not be collected.

Transrectal ultrasonography showed enlargement of the uterine body and a fetus was observed during 5 months of pregnancy Figure 4B. The pregnant cow is an ongoing pregnancy.

Increase and remain of serum progestagens at high level more than 20 weeks after AI indicated pregnancy. B Sonogram of elephant conception at day post ovulation.

The white arrow indicated embryo with the diameter of 43 mm. The results revealed the Medelian inheritance of all markers.

They indicated that the AI calf is the son of Japatee and Khod. The present study demonstrated that AI with a modified endoscopic technique using chilled semen was successfully used to produce a elephant calf.

The modified endoscopic technique using frozen-thawed semen demonstrated establishment of pregnancy. To our knowledge this is the first paper reporting establishment of pregnancy with AI using frozen-thawed semen in the Asian elephant.

This suggests that AI may present an alternative approach to traditional breeding for genetic and management in this species.

The elephant is an important animal and is recognized as the symbol of Thailand. Although the elephant is highly valued and plays an important economic role in our country the population of captive and wild elephants has decreased gradually.

It is well accepted that a major cause for the decline of captive elephants in Thailand is the loss of opportunity for propagation caused by the practice of separating male and female elephants and distributing them to either elephant camps or zoos for tourism purposes.

In addition to natural breeding, AI with either chilled or frozen-thawed semen will provide genetic diversity.

Moreover, ovLH surge, which induced ovulation, induction protocol was developed [ 19 ]. This novel method is an important advance for AI [ 20 ], and is useful for reducing the labor and number of samples needed for identifying the anLH surge to time the assisted breeding 3 weeks later [ 3 ].

Although the present study successfully produced one elephant calf out of 2 AI trials using chilled semen the success rate of AI with frozen-thawed semen is extremely low one successful pregnancy out of 55 AI trials.

The reason for the low success rate of AI with frozen-thawed semen samples is not clear. It might be due to the low sperm quality following freezing and thawing compared to chilled and fresh semen.

High quality of post-thaw sperm is a prerequisite for the success rate after AI. During freezing and thawing of elephant semen changes in the traditional sperm parameters, including sperm motility, viability, acrosome integrity and mitochondrial activity have been detected [ 4 , 5 ].

Furthermore, ultrastructural investigations using transmission electron microscopy TEM confirmed that the freezing and thawing procedure caused structural damage to elephant sperm, especially in the plasma membrane, acrosome and mitochondria.

Advanced Search. They are simply called breeding parties. The breeding parties are for Indian women, usually very upscale and immerses in Western circles, who want white babies but don't want a white husband, or at least don't want to divorce their Indian husbands.

If they have the consent or at least tolerance of their husbands and families, then they're set up with a breeding party. The goal here is for the Indian woman to get pregnant by a white man, and that man has to be tall, blue eyed, handsome, intelligent, usually a businessman who's in India for a time.

To insure that she gets pregnant, she has sex with multiple white. To simplify things and to make her feel comfortable, the sex and breeding is done in a group setting with other Indian women there to be bred also.

The women should be ovulating at that time. Most of the women get pregnant the at the breeding party, some don't and return again. One woman I heard of went again to a party, even though she was already pregnant from the first party she didn't tell anyone though.

She loved it so much the first time she had to do it again! Rishi Posts : Join date : Like Dislike. But it is a fact that Indian women who have children through white men show off their white skinned blue eyed infants.

In fact the Indian grand parents are much prouder than their daughters themselves. I know a Tamil brahmin lady whose daughter married a white American man.

When they went to India, her sister took her aside and asked her "when are you getting us a white baby? All of us are eagerly waiting for that,".

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For example, National Zoo reproductive physiologist Janine Brown discovered that elephants have a double LH surge, which turned out to be vital for the proper timing of the artificial insemination.

An elephant birth would help bolster the decreasing population of Asian elephants in North America, and it would be an important step toward creating a multi-generational herd at the National Zoo.

The Zoo is currently expanding its elephant exhibit to accommodate such a social grouping. National Zoo scientists have been studying Asian elephants in the wild for nearly 40 years, in an effort to help prevent them from becoming extinct.

Fewer than 30, Asian elephants remain in the wild. There also are 15, domesticated elephants found in Asian range countries, many of which live in substandard conditions in logging camps, temples, tourist resorts and other facilities.

Note to editors: Digital photos of Shanthi are available from the National Zoo's public affairs office.

January 19, News Release. One female Sao inseminated with frozen-thawed semen showed signs of pregnancy by increasing progestagen levels and a fetus was observed for 5 months by transrectal ultrasonography.

This is the first report showing pregnancy following AI with frozen-thawed semen in the Asian elephant. Successful AI in the Asian elephant using either chilled or frozen-thawed semen is a stepping stone towards applying this technology for genetic improvement of the elephant population.

As the number of wild elephants continues to decline, maintaining healthy captive populations is vital.

Because of difficulties in transporting a bull elephant from one institution to another for natural breeding, artificial insemination AI is an alternative tool for genetic management in this species.

Although elephant calves have been produced following AI using either fresh or chilled semen [ 1 — 3 ] there has been no report of the birth of elephant calves after AI with frozen-thawed semen.

Our previous studies showed that acceptable semen quality can be obtained after freezing and thawing [ 4 , 5 ]; however, pregnancy was not established after AI [ 6 ], indicating a need to develop an effective cryopreservation technique for Asian elephant spermatozoa.

Two AI techniques in elephants have been reported; i. Both techniques are impractical in Thailand. Surgical AI is not suitable to use due to the complication of wound infection and concerns for animal welfare, whereas endoscopic AI requires special accessory equipment: a balloon catheter to distend the reproductive tract [ 1 ].

Therefore, the objective of this study was to investigate the fertilizing ability of either chilled or frozen-thawed semen using the modified endoscopic AI technique described herein.

All animals and the experiments were approved by the National Elephant Institute. Eight elephant bulls age range, to years-old housed at the National Elephant Institute, Forest Industry Organization, Lampang, Thailand were used as semen donors.

Ten elephant females age range, to years were used for AI. The elephants were fed with grass, banana and sugar cane during the day and allowed to roam in the jungle at night.

Semen samples were collected by manual manipulation as described by Schmitt and Hildebrandt [ 7 ]. Each ejaculate was immediately evaluated for volume, sperm concentration, progressive motility, sperm viability and pH [ 8 ].

Sperm concentration was assessed using a haemocytometer. Progressive motility was assessed visually under a phase-contrast microscopy by two independent technicians.

Semen samples collected from 6 bulls Dindum, Pamae, Yuak, Koong, Tadaeng and Jumpui were frozen by the method as previously described [ 4 ].

Briefly, fresh semen was diluted with TEST extender at room temperature. TEST extender consisted of 5.

The equilibrated mixture was packed in a 0. The straws were held at 2. Ten microlitres of the thawed semen were then taken and placed on a pre-warmed microscopic slide and the progressive motility of post-thaw spermatozoa was assessed under light microscopy.

Semen samples collected from 2 bulls Japatee and Sai were prepared and used for AI as chilled semen. Fresh semen was diluted with TEST extender [ 4 ] to a final volume of ml.

A ml blood sample was collected from an ear vein weekly for hormonal assay. When progestagen concentrations decreased to baseline i. Blood samples were allowed to clot at room temperature for 1 to 2 h, and then centrifuged at g for 5 min to separate serum from blood cells.

Serum samples were transferred into 1. Serum progestagens were analyzed using enzyme immunoassay EIA as previously described [ 9 ].

This EIA employed a monoclonal antibody against progesterone ,; Quidel clone Quidel clone ; supplied by C.

Munro, Davis, CA, USA that has been shown to cross-react with a variety of reduced progesterones in the serum and excreta of a wide range of species, including elephants [ 10 ].

Bound progesterone was visualized using a horseradish peroxidase-conjugated antibody to progesterone ,; C. The sensitivity of the assay was 0.

This monoclonal antibody was validated and approved to detect LH in Asian elephants [ 3 , 11 ]. Sensitivity of the assay was 0.

The inter-assay coefficient of variation CV for the high concentration control was 7. Artificial insemination series were conducted during to Fifty five and two AI attempts were conducted with frozen-thawed and chilled semen in six and two elephant females, respectively.

Three inseminations once per day per series were performed on days -1, 0, 1 of the ovulatory LH surge [ 3 ].

Fifty milliliter of semen was used per inseminations. The female was standing immobilized using 0. For AI, the endoscopic technique was modified by increasing inflated air from external electric air pump without using the balloon catheter.

The inflated air could enlarge the vestibular, vaginal and cervical canal. After the tip of endoscope reached and fixed at the cervical canal, the air tube was removed and replaced by insemination catheter IC.

The IC was passed through the working channel of the endoscope and penetrated deep into the body of uterus. The semen was deposited into body of uterus without signs of semen reflux Figure 1.

Pregnancy diagnosis was performed by serum progestagen measurement and transrectal ultrasonography. Blood samples were collected weekly and serum progestagens were assessed by EIA as described above [ 3 ].

Transrectal ultrasonography was performed weekly between 2 and 4 months after AI as previously described [ 1 ]. Positive PCR products generated by specific primers of microsatellite markers were fractionated on a 1.

Parentage analysis was performed by parentage exclusion method [ 18 ]. A total of 24 ejaculates obtained from six bulls were prepared as frozen semen.

Data on semen quality of fresh, chilled and frozen-thawed samples are shown in Table 1. The average sperm motility of frozen-thawed semen was A progressive reduction of the percentage of viable sperm was observed.

Eight out of 10 elephants assessed for estrus cycle showed normal cycling and were selected for AI.

The onset of the luteal phase was defined as the first point at which serum progestagen concentrations increased above 0. At the end of luteal phase, serum progestagen concentrations started to decline and reached the baseline 0.

In case of AI with chilled semen, one out of two females, namely "Khod", showed progressive increase of serum progestagens after AI Figure 2. Serum progestagen started to decline to the baseline one month before birth Figure 2.

Transrectal ultrasonography of pregnant cow showed enlargement of the uterine wall at 2 months after AI. An increase of uterine lumen that filled with clear fluid was found between 3 and 4 months.

We were unable to find a fetus between 2 and 4 months after AI. Some changes in Khod's behavior during pregnancy period were observed.

Khod's anatomy was not remarkably changed during pregnancy period except that her belly was enlarged during the last sixth month period. Furthermore, one month before calving, her breasts were enlarged and had a clear secretion released when they were stimulated.

Khod gave a birth on March 7, with birth weight of 51 kg. In , Shanthi gave birth to Kandula. He was the fifth elephant in the world conceived by artificial insemination.

Through past artificial insemination procedures performed with Shanthi, zoo scientists were able to develop a greater understanding of elephant reproduction.

Zoo reproductive physiologist Janine Brown discovered that elephants have what is called a double luteinizing hormone surge, which turned out to be vital for the proper timing of the artificial insemination.

This discovery was very useful, as artificially inseminating an elephant is a challenging and difficult medical procedure.

The artificial insemination must be timed properly: Elephants have two surges of luteinizing hormone in about a three-week period.

Using blood samples, scientists are able to detect when the first surge occurs.

Hellsangel Posts : Join date : Ten elephant females age range, to years were used for AI. January 19, All authors Erotic finder and approved the final manuscript. High quality of post-thaw 18 ebony xxx is a prerequisite for the success rate after AI. Connie carter porn tube insemination Anon-v download insemination series were conducted during to Note Mandingo anal sex editors: Digital photos of Shanthi are Woman fingers herself from the National Zoo's public affairs office. This might be a key point of success and failure in previous AI August ames p*** in Spanking on bare butt elephants unpublished data. Int J Androl. He was the fifth elephant in the world conceived by artificial insemination. Using blood samples, scientists are able Premiumhentai com detect when the first surge occurs. Jessica.jaymes progestagen concentrations decreased Jodie gasson porn baseline i. Sensitivity of the assay was 0. For example, National Zoo reproductive physiologist Janine Brown discovered that elephants have a double LH surge, which turned out to be vital for the proper timing of the artificial insemination. Figure 1. Amateur Rio lee katt dylan, Videos. Besamung p 2 Min Hossawald - 20,6k Sichten. Asiatisch-kaukasische Babys machen. Asian Insemination p Free homemade mobile porn Sek Hossawald - 13,9k Sichten. Besamung Live Cams. We use cookies to optimize site functionality and give you the best possible experience. Nattalliex Alle Kategorien. Japanese bbw milf creampie and impregnate and enema p 5 min Mbs - Furry hemtai Danger Videos. Teens 46, Videos.

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