Seasonal Effects on Body Condition and Characteristics of the Estrous Cycle in Captive Asian Elephants (Elephas maximus) in Thailand: A Retrospective Study.

The aim of this study was to investigate the effects of season on the body condition score (BCS), the characteristics of the estrous cycle (luteal phase [LPL], follicular phase [FPL], estrous cycle [ECL] lengths, and the start of the luteal phase [SLP] and follicular phase [SFP]), and progesterone levels (baseline and peak) of eight captive Asian elephants (Elephas maximus) in Thailand. From 2014 to 2019, blood samples were collected weekly for serum progesterone enzyme immunoassays (EIAs). Estrous cycles (n = 70), including the luteal and follicular phases, and BCS (n = 70) were recorded. Based on the BCS, the LPL, FPL, and ECL were assigned to the following two groups: normal (BCS = 3.0-4.0, n = 38) and overweight (BCS = 4.5-5.0, n = 32). The findings demonstrated that there was no difference in LPL between the groups. However, in the normal group, the ECL was one week longer (14.9 ± 1.7 vs. 13.9 ± 1.7 weeks; p < 0.05), and the FPL also tended to be one week longer (7.2 ± 1.7 vs. 6.4 ± 1.5 weeks; p = 0.06) than in the overweight group. The mean progesterone level during the rainy, hot, and cool seasons was not statistically different. Based on the yearly averaged BCS from three seasons, the baseline and peak levels of progesterone were classified into the normal (n = 16) and overweight (n = 12) groups. Females with a normal BCS tended to exhibit higher progesterone peak levels (p = 0.08). The majority of peaks appeared during the rainy season (53.57%). The BCS was highest during the hot (4.47) and rainy (4.38) seasons, but not during the cool (4.12) season. The LPL, FPL, and ECL were not affected by the season in which the luteal phase occurred. On the other hand, the rainy season had a significant effect on the SFP, resulting in a longer LPL (p < 0.05) and ECL (p = 0.01); both were the longest during the rainy season. In conclusion, the effects of season on BCS may be related to characteristics of the estrous cycle and peak progesterone levels. Ultimately, these findings provide ground knowledge to assist elephant managers and owners in planning breeding activities using seasonal effects and BCS measurements in tropical climates.

Variability of gait characteristics in lameness elephant.

Lameness has a significant impact not only on the economy but also on elephant welfare. Several gait characteristics are altered to compensate for the discomfort. The traditional approach to detecting lameness has relied on mahout and veterinarian observation. The study aimed to determine how lameness affected the variability of an elephant's gait by using a three-dimensional inertial measurement unit (IMU) with Wi-Fi sensors. Five elephants with lameness, as determined by an experienced veterinarian and two, non-lamed elephants, aged between 58-80 years were included in the study. Gait biomechanics including limb segment motion, obtained from individually gyrometric- and accelero-based parameters and demonstrated as a graphical pattern showing changes in absolute rotation angle over time. The result revealed some character changes in gait kinematics parameters, but it was heterogeneity with an inconclusive pattern. The interlimb coordination could be a part of maintaining the actual locomotion pattern, or it could be a result of the mild degree of lameness for which all of the clients have compensated. This study introduces a new objective method for quantifying gait changes caused by lameness, additional research is required to make this objective more clinically applicable.

Dexmedetomidine Effectively Sedates Asian Elephants (Elephas maximus).

This study investigated the sedative effects of dexmedetomidine in Asian elephants. We hypothesized that 2 µg/kg dexmedetomidine would provide sufficient standing sedation. A crossover design study was performed in three Asian elephants. Each elephant was assigned to 1 of 3 treatment groups-1 (D1), 1.5 (D1.5) or 2 (D2) µg/kg dexmedetomidine (intramuscular injection, IM) with a two-week 'washout period' between doses. Elephants were monitored for 120 min. At 120 min (Ta), atipamezole was administered IM. Sedation and responsiveness scores were evaluated. Physiological parameters (pulse rate, respiratory rate, and %SpO2) and clinical observations were monitored during the study and for 3 days post drug administration. D2 provided the longest sedation (approximately 70 min), compared to D1 and D1.5. After Ta, each elephant's sedative stage lessened within 10-15 min without complications. No significant abnormal clinical observations were noted throughout and during the 3-days post study period. These data suggest that a single 2 µg/kg IM dexmedetomidine injection provides sufficient standing sedation for approximately 70 min in Asian elephants.

Development of Nonstructural Protein-Based Indirect ELISA to Identify Elephant Endotheliotropic Herpesvirus (EEHV) Infection in Asian Elephants (Elephas maximus).

Disease caused by elephant endotheliotropic herpesvirus (EEHV) infection is the most highly fatal hemorrhagic disease in Asian elephant calves worldwide. To date, adult elephants that have been infected with EEHV have predominantly displayed mild clinical signs, while they are believed to serve as EEHV shedders to other elephants. Hence, the diagnostic tools employed for monitoring EEHV-active infection are considered vitally important. In this study, partial EEHV-DNA polymerase (DNApol) nonstructural proteins (NSPs) were used to detect anti-EEHV antibodies through the use of an in-house indirect enzyme-linked immunosorbent assay (ELISA). The results were then compared to those obtained from a PCR test. In this study, a total of 175 serum samples were collected from Asian elephants living in elephant camps located in Chiang Mai and Lampang Provinces, Thailand. The elephants were aged between 2 and 80 years old. The overall percentages of positive samples by the PCR and EEHV-DNApol ELISA tests were 4% (21/175) and 12% (21/175), respectively. The ELISAs demonstrated values of 77.9% (95% posterior probability interval (PPI) = 52.5-95%) sensitivity and 87.7% (PPI = 82.5-91.9%) specificity, respectively. Accordingly, the sera obtained from the elephants exhibiting no clinical signs of EEHV infection, and those who were negative according to PCR tests, revealed a value of 14% seropositivity for EEHV-DNApol. Our results indicate that these asymptomatic, active EEHV-infected elephants could likely serve as a source of EEHV shedding within elephant herds. Consequently, the developed EEHV-DNApol NSPs-based ELISA test employed in the present study may be of use for routine monitoring and identification of EEHV shedders in elephant herds, and could be an alternative diagnostic tool for EEHV detection in Asian elephants.

Serum Insulin-like Growth Factor-1 Is a Biomarker of Testosterone Production and Intact Acrosome in Asian Elephants (Elephas maximus).

The objective of this study was to find relationships among serum IGF-1, serum testosterone, seminal plasma IGF-1 concentrations and semen parameters in Asian elephants (Elephas maximus). A total of 17 ejaculates (one to three ejaculates/bull) were collected from seven captive elephant bulls by performing rectal massage. Before each ejaculation, blood samples were obtained for serum IGF-1 and testosterone assays. Subsequently, the semen characteristics of each ejaculate were evaluated. Mean serum IGF-1 concentration of elephant bulls was estimated as 326.3 ± 114.6 ng/mL (median, 286.2 ng/mL; range, 167.4-542.7 ng/mL). An increase in serum IGF-1 concentration was found to correlate with the percentage of spermatozoa with intact acrosomes. In addition, IGF-1 concentration was positively correlated with testosterone level. However, seminal IGF-1 concentrations could not be detected. In conclusion, our findings suggest that serum IGF-1 concentration is likely a biomarker of normal testicular functions, particularly spermatogenesis in elephants. Moreover, this commercial IGF-1 ELISA is eligible for analyzing serum IGF-1 concentration in Asian elephants.

Clinical characteristics of elephant endotheliotropic herpesvirus (EEHV) cases in Asian elephants (Elephas maximus) in Thailand during 2006-2019.

BACKGROUND: Elephant endotheliotropic herpesvirus causes a hemorrhagic disease (EEHV-HD) that is a major cause of death in juvenile Asian elephants with EEHV1 and EEHV4 being the most prevalent. AIM: To perform a retrospective clinical data analysis. METHODS: Records of a total of 103 cases in Thailand confirmed by polymerase chain reaction (PCR) on blood and/or tissue samples. RESULTS: The severity of clinical signs varied among EEHV subtypes. EEHV1A was the most prevalent with 58%, followed by EEHV4 with 34%, EEHV1B with 5.8% and EEHV1&4 co-infection with 1.9%. Overall case fatality rate was 66%. When compared among subtypes, 100% case fatality rate was associated with EEHV1&4 co-infection, 83% with EEHV1B, 75% with EEHV1A, and the lowest at 40% for EEHV4. Calves 2- to 4-year old were in the highest age risk group and exhibited more severe clinical signs with the highest mortality. Majority of cases were found in weaned or trained claves and higher number of cases were observed in rainy season. A gender predilection could not be demonstrated. Severely affected elephants presented with thrombocytopenia, depletion of monocytes, lymphocytes and heterophils, a monocyte:heterophil (M:H) ratio lower than 2.37, hypoproteinemia (both albumin and globulin), severe grade of heterophil toxicity, and low red blood cell counts and pack cell volumes. Survival was not affected by antiviral drug treatment in the severely compromised animals. CONCLUSION: Early detection by laboratory testing and aggressive application of therapies comprising of supportive and anti-viral treatment can improve survival outcomes of this disease.

Serological study on brucellosis in captive elephants (Elephas maximus) and stray dogs in North Thailand.

BACKGROUND AND AIM: Brucellosis is considered as an important zoonotic disease caused by various strains of Brucella in numerous host species. Although brucellosis has been reported in almost animal species, the relevance of brucellosis infection and diagnostic technique in Asian elephant (Elephas maximus) has been limited. The present serological investigation aimed to investigate the antibody response to Brucella abortus in captive Asian elephants in North Thailand. Moreover, further serological survey was also conducted to detect the antibody response to Brucella canis in stray dogs cohabiting the same area as the elephant herd. MATERIALS AND METHODS: Serum samples were collected from 40 captive Asian elephants and submitted for serological analysis based on B. abortus antigen using Rose Bengal plate test (RBPT) in combination with ethylenediaminetetraacetic acid-tube agglutination test (EDTA-TAT) as a supplementary test and by commercial indirect enzyme-linked immunosorbent assay (iELISA). In addition, serum samples were also obtained from 16 stray dogs that live nearby the elephant-raising area and were tested using commercial Dot-ELISA based on B. canis antigen. RESULTS: Serological analysis in captive Asian elephants showed 100% seronegative (40/40) from all serological tests response to B. abortus. For stray dogs, 12.5% (2/16) had a low positive reaction response to B. canis. CONCLUSION: The serological survey for brucellosis in Asian elephant was adapted and applied using RBPT, EDTA-TAT, and iELISA in the present study. For future evaluation, we recommended the use of a combination of serological tests with validation together with comparing by direct detection such as bacterial isolation to provide an appropriate brucellosis surveillance program in Asian elephants. In addition, the surveillance of stray dogs or multispecies habitation should be kept into considerations.

Subclinical infection of captive Asian elephants (Elephas maximus) in Thailand with elephant endotheliotropic herpesvirus.

Elephant endotheliotropic herpesvirus (EEHV) infection is a conservation threat to the endangered Asian elephant (Elephas maximus), causing fatal hemorrhagic disease in juvenile elephants throughout the world, including Thailand. This study revealed a subclinical EEHV1 infection rate of 5.5% in healthy captive Asian elephants in Thailand (n = 362). The virus was detected in all age classes above one year old, in both sexes, and across the country - even in facilities with no history of hemorrhagic disease (EEHV HD). Subclinical EEHV infection in Thailand urgently requires proper health management.

Management factors affecting adrenal glucocorticoid activity of tourist camp elephants in Thailand and implications for elephant welfare.

Elephant camps are among the most popular destinations in Thailand for tourists from many countries. A wide range of management strategies are used by these camps, which can have varied impacts on health and welfare of elephants. The objectives of this study were to examine relationships between FGM (fecal glucocorticoid metabolite) concentrations and camp management factors (work routine, walking, restraint, rest area, foraging), and to other welfare indicators (stereotypic behaviors, body condition, foot health, and skin wounds). Data were obtained on 84 elephants (18 males and 66 females) from 15 elephant camps over a 1-year period. Elephants were examined every 3 months and assigned a body condition score, foot score, and wound score. Fecal samples were collected twice monthly for FGM analysis. Contrary to some beliefs, elephants in the observation only program where mahouts did not carry an ankus for protection had higher FGM concentrations compared to those at camps that offered riding with a saddle and shows. Elephants that were tethered in the forest at night had lower FGM concentrations compared to elephants that were kept in open areas inside the camps. There was an inverse relationship between FGM concentrations and occurrence of stereotypy, which was not anticipated. Thus, assessing adrenal activity via monitoring of FGM concentrations can provide important information on factors affecting the well-being of elephants. Results suggest that more naturalistic housing conditions and providing opportunities to exercise may be good for elephants under human care in Thailand, and that a no riding, no hook policy does not necessarily guarantee good welfare.

Survival analysis of confirmed elephant endotheliotropic herpes virus cases in Thailand from 2006 - 2018.

The elephant endotheliotropic herpesvirus (EEHV) has been a known cause of death of young elephants in Thailand for over a decade. In this study, we report on the demography, disease characteristics and mortality of 58 elephants with confirmed EEHV hemorrhagic disease between January 2006 and August 2018 using retrospective data subjected to survival analysis. Median age of EEHV presentation was 29 months, and the mortality rate was 68.97% with a median survival time of 36 h. Most EEHV cases occurred in the north of Thailand, the region where most of the country's captive elephants reside. The hazard ratio analysis identified application of medical procedures and antiviral medications as being significant factors correlated to the risk of death. Our results indicate a need to focus EEHV monitoring efforts on young elephants and to follow current protocols that advise starting treatments before clinical signs appear.

Evidence of high EEHV antibody seroprevalence and spatial variation among captive Asian elephants (Elephas maximus) in Thailand.

BACKGROUND: Elephant endotheliotropic herpesviruses (EEHV) can cause an acute highly fatal hemorrhagic disease in young Asian elephants (Elephas maximus), both ex situ and in situ. Amongst eight EEHV types described so far, type 1 (subtype 1A and 1B) is the predominant disease-associated type. Little is known about routes of infection and pathogenesis of EEHV, and knowledge of disease prevalence, especially in range countries, is limited. METHODS: A large cross-sectional serological survey was conducted in captive elephants (n = 994) throughout Thailand using an EEHV-1A glycoprotein B protein antigen specific antibody ELISA. RESULTS: Antibody seroprevalence was 42.3%, with 420 of 994 elephants testing positive. Associations between seropositivity and potential risk factors for EEHV infection were assessed and included: elephant age, sex, camp cluster size, management type (extensive versus intensive), sampling period (wet vs. dry season) and location of camp (region). Univariable regression analysis identified management system and region as risk factors for the presence of EEHV antibodies in elephants, with region being significant in the final multivariable regression model. Prevalence was highest in the North region of the country (49.4%). CONCLUSIONS: This study produced baseline serological data for captive elephants throughout Thailand, and showed a significant EEHV burden likely to be maintained in the captive population.

ELEPHANT ENDOTHELIOTROPIC HERPESVIRUS ASSOCIATED WITH CLOSTRIDIUM PERFRINGENS INFECTION IN TWO ASIAN ELEPHANT ( ELEPHAS MAXIMUS) CALVES.

Elephant endotheliotropic herpesvirus (EEHV) is an infection associated with fatal hemorrhagic disease in young Asian elephants ( Elephas maximus). This brief communication describes the postmortem evaluation of two Asian elephant calves diagnosed with EEHV4 and EEHV1A in conjunction with Clostridium perfringens infection. Case 1 was a 7-mo-old, male captive-born Asian elephant that developed diarrhea and died 2 days after clinical presentation. Examination of the heart, lungs, liver, and spleen revealed predominantly basophilic intranuclear inclusion bodies in the endothelial cells of the blood vessels. Case 2 was a 3-mo-old, female wild-born Asian elephant that showed signs of lethargy, anorexia, and convulsions and died 6 hr after clinical presentation. No intranuclear inclusion bodies were observed. The heart, lung, liver, and spleen of both calves tested positive for EEHV by polymerase chain reaction. Phylogenetic analysis identified EEHV4 and EEHV1A in Case 1 and 2, respectively. Additionally, liver, spleen, and hemorrhagic intestinal tissue samples tested positive for C. perfringens α, ß, and ε toxins. This is the first reported case to describe coinfection of EEHV and C. perfringens in Asian elephant calves.

Osteoarthritis in two marine mammals and 22 land mammals: learning from skeletal remains.

The occurrence of osteoarthritis (OA) in marine mammals is still questionable. Here we investigated the prevalence of OA in marine (dolphin and dugong) and terrestrial mammals (Asian elephant, Asiatic buffalo, camel, cat, cattle, deer, dog, domestic goat, horse, human, hyena, impala, lion, Malayan tapir, Assam macaque, mule, pig, rabbit, red kangaroo, sheep, tiger and waterbuck). Skeletal remains obtained from five institutes were used as subjects; a total of 45 different parts (locations) of bones were observed for OA lesions. The prevalence of OA was reported as number of OA lesions/total number of bones. Our results revealed that the presence of OA in marine species (dolphin and dugong) was 2.44% and 3.33%, respectively. In dolphins, the highest OA occurrence was on the left and right humeral trochlea, with 13.68% and 12.63%, respectively, while the highest number of OA lesions in dugongs was on the lumbar vertebrae (8.79%). No significant difference (P > 0.05) in the prevalence of OA between sexes in dolphins and dugongs was observed, but we found a significant difference (P < 0.05) in 24 bone locations of human bones, which had the highest OA prevalence (48.93%), followed by dogs (3.94%). In conclusion, OA can occur in marine mammals, similar to terrestrial mammals, even though their natural habitat is the ocean.

SUCCESSFUL TREATMENT OF A CLINICAL ELEPHANT ENDOTHELIOTROPIC HERPESVIRUS INFECTION: THE DYNAMICS OF VIRAL LOAD, GENOTYPE ANALYSIS, AND TREATMENT WITH ACYCLOVIR.

This article describes the treatment of clinical elephant endotheliotropic herpesvirus (EEHV) infection in a male Asian elephant ( Elephas maximus; approximately 3 yr old), the dynamics of viral load during the active infection, and genetic analysis of the virus. Treatment included injectable acyclovir (12 mg/kg iv, bid), antibiotic, vitamin, and fluids. Quantitative polymerase chain reaction was used to measure the viral levels in blood, which decreased continuously after initiation of intravenous acyclovir. Low levels of virus were detected in the blood for 2 wk, and the virus was undetectable after 1 mo. No complication was observed during the treatment period. This case report suggests that acyclovir, given parenterally, could potentially enhance survival of clinical EEHV-infected individuals.

Anatomy, histology and elemental profile of long bones and ribs of the Asian elephant (Elephas maximus).

This study evaluated the morphology and elemental composition of Asian elephant (Elephas maximus) bones (humerus, radius, ulna, femur, tibia, fibula and rib). Computerized tomography was used to image the intraosseous structure, compact bones were processed using histological techniques, and elemental profiling of compact bone was conducted using X-ray fluorescence. There was no clear evidence of an open marrow cavity in any of the bones; rather, dense trabecular bone was found in the bone interior. Compact bone contained double osteons in the radius, tibia and fibula. The osteon structure was comparatively large and similar in all bones, although the lacuna area was greater (P < 0.05) in the femur and ulna. Another finding was that nutrient foramina were clearly present in the humerus, ulna, femur, tibia and rib. Twenty elements were identified in elephant compact bone. Of these, ten differed significantly across the seven bones: Ca, Ti, V, Mn, Fe, Zr, Ag, Cd, Sn and Sb. Of particular interest was the finding of a significantly larger proportion of Fe in the humerus, radius, fibula and ribs, all bones without an open medullary cavity, which is traditionally associated with bone marrow for blood cell production. In conclusion, elephant bones present special characteristics, some of which may be important to hematopoiesis and bone strength for supporting a heavy body weight.

Osteocalcin and bone-specific alkaline phosphatase in Asian elephants (Elephas maximus) at different ages.

Bone turnover markers could offer a potential alternative means for the early diagnosis of metabolic bone disease in young growing elephants although the baseline of bone turnover markers in elephant is not well established. The aim of this study was to determine any relationship between the age of captive Asian elephants (Elephas maximus) and markers of bone formation. Serum samples from 24 female Asian elephants were collected to evaluate levels of two bone formation markers, namely, osteocalcin (OC) and bone-specific alkaline phosphatase (BAP). Both intact and N-terminal midfragment OC and BAP were negatively correlated with age. The findings demonstrate that younger elephants have a higher rate of bone turnover than older elephants. Use of these and additional bone markers could lead to the establishment of validated protocols for the monitoring of bone disease in elephants.

Risk factors for saddle-related skin lesions on elephants used in the tourism industry in Thailand.

BACKGROUND: Lesions related to working conditions and improper saddle design are a concern for a variety of working animals including elephants. The objectives of the present study were to determine the prevalence of cutaneous lesions in anatomic regions (i.e., neck, girth, back, tail) in contact with saddle-related equipment among elephants in Thailand working in the tourism industry, and to identify potential risk factors associated with these lesions. Data for this cross-sectional study were collected between May 2007 and July 2007 on 194 elephants from 18 tourism camps across Thailand. RESULTS: There was a high prevalence (64.4 %; 95 % CI 57.3 - 71.2) of active lesions, most often located on the back region. Using multilevel multivariable logistic regression modelling containing a random intercept for camp we identified the following risk factors: increasing elephant age, the use of rice sacks as padding material in contact with the skin, and the provision of a break for the elephants. Working hours had a quadratic relationship with the log odds of an active lesion where the probability of an active lesion initially increased with the number of working hours per day and then declined possibly reflecting a "healthy worker" bias where only animals without lesions continue to be able to work these longer hours. CONCLUSIONS: While we recognize that the cross-sectional nature of the study posed some inferential limitations, our results offer several potential intervention points for the prevention of these lesions. Specifically, we recommend the following until longitudinal studies can be conducted: increased monitoring of older elephants and the back region of all elephants, working less than 6 hours per day, and the avoidance of rice sacks as padding material in contact with skin.

Diet composition, food intake, apparent digestibility, and body condition score of the captive Asian elephant (Elephas maximus): a pilot study in two collections in Thailand.

Limited data are available regarding the nutrition and feeding of captive Asian elephants in range countries. In this study, feeding regimens of two collections in northern Thailand and their actual diets shaped by availability of forage and mahout preferences were assessed for nutritional quality. The composition of dietary intake, fecal output, and the dietary regimen were individually recorded for 5 days in 10 elephants. The proportion of forage in the diet represented 41 to 62% of the dry matter intake (DMI) in one collection whereas in the other collections it varied between 68 and 72%. Between 8.5 and 24% of the diet consisted of commercial pellets, and hulled rice represented up to 25% of the DMI in one collection. Sugar cane, corn cobs, and fruits such as bananas were eaten in smaller amounts. Body condition scores and weights were measured, which revealed that nine animals were in good condition. Representative samples of each food as well as fecal samples were analyzed for dry matter, crude protein, fat, crude fiber, gross energy, ash, calcium, and phosphorus. Diet adequacy was assessed by calculating the digestible nutrients in the rations and by comparing them to the recommendations from literature. The digestible energy (DE) intake varied between 0.6 and 1.4 megajoules (MJ) per kg(0.75) per day; therefore, higher than the estimated recommendations of 0.65 MJ per kg(0.75) per day for nine of the elephants. In all elephants the crude protein intake was less than the maintenance recommendations and ranged between 6.01 and 7.56% of the DMI. Calcium intake was low in one collection and there was an inverse calcium: phosphorus ratio, which was inadequate. The present study adds to the knowledge of captive elephant diets in Asia and is a starting point for further research, which is necessary to design optimum diet plans for captive Asian elephants in Thailand.

Mycobacterium tuberculosis infection of domesticated Asian elephants, Thailand.

Four Asian elephants were confirmed to be infected with Mycobacterium tuberculosis by bacterial culture, other diagnostic procedures, and sequencing of 16S-23S rDNA internal transcribed spacer region, 16S rRNA, and gyrase B gene sequences. Genotyping showed that the infectious agents originated from 4 sources in Thailand. To identify infections, a combination of diagnostic assays is essential.

Successful artificial insemination in the Asian elephant (Elephas maximus) using chilled and frozen-thawed semen.

BACKGROUND: Artificial insemination (AI) using frozen-thawed semen is well established and routinely used for breeding in various mammalian species. 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). METHODS: Semen samples were collected by from 8 bulls (age range, 12-to 42-years) by manual stimulation. Semen with high quality were either cooled to 4 degrees C or frozen in liquid nitrogen (-196 degrees C) before being used for AI. Blood samples collected from ten elephant females (age range, 12-to 52-years) were assessed for estrus cycle and elephants with normal cycling were used for AI. Artificial insemination series were conducted during 2003 to 2008; 55 and 2 AI trials were conducted using frozen-thawed and chilled semen, respectively. Pregnancy was detected using transrectal ultrasonography and serum progestagen measurement. RESULTS: One female (Khod) inseminated with chilled semen became pregnant and gave birth in 2007. The gestation length was 663 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. CONCLUSION: 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.