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

<p>Abstract</p> <p>Background</p> <p>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).</p> <p>Methods</p> <p>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°C or frozen in liquid nitrogen (-196°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.</p> <p>Results</p> <p>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.</p> <p>Conclusion</p> <p>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.</p

Circadian Rhythm of Salivary Immunoglobulin A and Associations with Cortisol as A Stress Biomarker in Captive Asian Elephants (Elephas maximus)

Salivary immunoglobulin A (sIgA) has been proposed as a potential indicator of welfare for various species, including Asian elephants, and may be related to adrenal cortisol responses. This study aimed to distinguish circadian rhythm effects on sIgA in male and female Asian elephants and compare patterns to those of salivary cortisol, information that could potentially have welfare implications. Subjects were captive elephants at an elephant camp in Chiang Mai province, Thailand (n = 5 males, 5 females). Salivette® kits were used to collect saliva from each elephant every 4 h from 06:00 to 22:00 h for 3 consecutive days (n = 15 samples/elephant). Enzyme immunoassays were used to quantify concentrations of IgA and cortisol in unextracted saliva. Circadian rhythm patterns were determined using a generalized least-squares method. Both sIgA and cortisol followed a circadian rhythm, although the patterns differed. sIgA displayed a daily quartic trend, whereas cortisol concentrations demonstrated a decreasing linear trend in concentrations throughout the day. There was no clear relationship between patterns of sIgA and salivary cortisol, implying that mechanisms of control and secretion differ. Results demonstrate for the first time that circadian rhythms affect sIgA, and concentrations follow a daily quartic pattern in Asian elephants, so standardizing time of collection is necessary

Effect of Tourist Activities on Fecal and Salivary Glucocorticoids and Immunoglobulin A in Female Captive Asian Elephants in Thailand

Asian elephants have been an important part of wildlife ecotourism in Thailand for over two decades. Elephants in tourist camps are exposed to a variety of management styles and daily activities that can potentially affect health and welfare. This study investigated relationships between a novel welfare biomarker, immunoglobulin A (IgA), and daily camp activities, and compared results to glucocorticoid (GC) measures. Often no-riding camps are portrayed as providing better welfare than camps that offer riding. Therefore, we predicted that elephants at no-riding camps would have lower GC and higher IgA concentrations, and a low GC/IgA ratio. Forty-four female elephants from six elephant camps were divided into three groups based on riding activities: saddle-riding, bareback-riding, and no-riding. Fecal and salivary samples were collected monthly for 1 year along with evaluations of body condition, foot health, and wounding. Camp environment and management varied among camps, although the major difference was in riding activities. Concentrations of GCs and IgA varied among the working groups, but not always consistently between sample matrices. Overall fecal glucocorticoid metabolite concentrations were lowest in the saddle-riding group. Only in one bareback-riding camp did the elephants exhibit a potentially positive welfare response with a low GC/IgA ratio over time. Other results varied between the two biomarkers, with considerable variability across camps, suggesting there is more to good welfare than whether elephants participate in riding or not. Several other human-induced stressors, like chaining, ankus use, and limited social opportunities are likely to be impacting well-being and should be considered to ensure management practices meet physical and psychological welfare needs

Characterizing the scent and chemical composition of Panthera leo marking fluid using solid-phase microextraction and multidimensional gas chromatography–mass spectrometry-olfactometry

Lions (Panthera leo) use chemical signaling to indicate health, reproductive status, and territorial ownership. To date, no study has reported on both scent and composition of marking fluid (MF) from P. leo. The objectives of this study were to: 1) develop a novel method for simultaneous chemical and scent identification of lion MF in its totality (urine + MF), 2) identify characteristic odorants responsible for the overall scent of MF as perceived by human panelists, and 3) compare the existing library of known odorous compounds characterized as eliciting behaviors in animals in order to understand potential functionality in lion behavior. Solid-phase microextraction and simultaneous chemical-sensory analyses with multidimensional gas-chromatography-mass spectrometry-olfactometry improved separating, isolating, and identifying mixed (MF, urine) compounds versus solvent-based extraction and chemical analyses. 2,5-Dimethylpyrazine, 4-methylphenol, and 3-methylcyclopentanone were isolated and identified as the compounds responsible for the characteristic odor of lion MF. Twenty-eight volatile organic compounds (VOCs) emitted from MF were identified, adding a new list of compounds previously unidentified in lion urine. New chemicals were identified in nine compound groups: ketones, aldehydes, amines, alcohols, aromatics, sulfur-containing compounds, phenyls, phenols, and volatile fatty acids. Twenty-three VOCs are known semiochemicals that are implicated in attraction, reproduction, and alarm-signaling behaviors in other species

Elephant reproduction: improvement of breeding efficiency and development of a breeding strategy

The efficiency of reproduction of the Asian elephant (Elephas maximus) has become of major concern. Captive breeding programs worldwide have met with limited success and few ex situ elephant populations are self-sustaining. The low birth rate and high mortality cause the captive population to decline rapidly. Self-sustaining populations in tourist elephant camps, timber elephant camps, circus or zoos would be essential to prevent a drain of wild populations and to reduce illegal wild capture to replace the captive population. To improve captive breeding management, we need to understand the reproductive physiology and, in particular, the estrous cycle of the Asian elephant. We investigated the female reproductive physiology in semi-captive elephants in Thailand, where the climate, ecology and nutrition are close to those of the wild population. For efficient breeding, adequate timing of copulation and insemination is essential. Hence, reproductive behaviors would be evaluated to identify the estrous period and to pinpoint the period of receptivity. This resulted in the development of an estrus detection method to improve the breeding efficiency. In addition, a GnRH-agonist administration protocol to induce a predictably timed ovulatory LH surge, which induced ovulation, was investigated to increase the success of both artificial and natural mating. Evaluation of the genetic relationship between breeding animals has been studied in order to maintain the genetic diversity. To this end, microsatellite markers were evaluated, which led to an optimal panel of markers for individual identification, paternal and maternal analysis and genetic diversity assessment. This panel has been proposed as the standard for studying genetic diversity across regions in Asia. Furthermore, genetic data of Asian elephants in Thailand and Dutch zoos were assessed. This information is most relevant for conservation programs and management. Pairing the right partners at the right time is one of the strategies to improve breeding efficiency. Reliable and informative pedigree data, generally available in zoo elephants but not in tourist and logging elephants, preferably combined with individual genetic analysis, is required to obtain insight in the genetic relatedness and to establish an effective breeding program. Breeding with least genetic relatedness partners, paired at the receptivity period of the female, would increase conception rate and prevent inbreeding. Furthermore, the proven bull with sufficient physical comfort for quality of life and female during the optimal reproductive stage of life (15-30 years) are recommended to achieve successful breeding. The results of these studies can be implemented in the captive breeding program in elephant camps and zoos to protect the sustainability of the Asian elephants. It is beneficial for both owners and conservationists to increase elephant numbers and maintain a viable captive population. Efficient breeding strategies, considering both the reproductive process and a broad genetic basis, will serve to increase and maintain the Asian elephant population for the future

Molecular characterisation and genetic variation of Elephant Endotheliotropic Herpesvirus infection in captive young Asian elephants in Thailand

Elephant Endotheliotropic Herpesvirus (EEHV) is emerging as a new threat for elephant conservation, since being identified as the cause of severe, often fatal, haemorrhagic disease in young Asian elephants. To describe positive cases and the molecular relatedness of virus detected in elephants in Thailand, we re-examined all available of EEHV samples occurring in young elephants in Thailand between 2006 and 2014 (n = 24). Results indicated 75% (18/24) of suspected cases were positive for EEHV by semi-nested PCR. Further gene analysis identified these positive cases as EEHV1A (72%, 13/18 cases), EEHV1B (11%, 2/18) and EEHV4 (17%, 3/18). This study is the first to phylogenetically analyse and provide an overview of most of the known EEHV cases that have occurred in Thailand. Positive individuals ranged in age from one to nine years, with no sex association detected, and occurred across geographical locations throughout the country. All individuals, except one, were captive-born. No history of direct contact among the cases was recorded, and this together with the fact that various subtype clusters of virus were found, implied that none of the positive cases were epidemiologically related. These results concur with the hypothesis that EEHV1 is likely to be an ancient endogenous pathogen in Asian elephants. It is recommended that active surveillance and routine monitoring for EEHV should be undertaken in all elephant range countries, to gain a better understanding of the epidemiology, transmission and prevention of this disease

Characteristics of gut microbiota in captive Asian elephants (Elephas maximus) from infant to elderly

Abstract Gut microbiota play an important role in the health and disease of Asian elephants, however, its characteristics at each stage of life have not been thoroughly investigated in maintaining and regulating health of elephants. This study, therefore, aimed to characterize the profiles of the gut microbiota of captive Asian elephants from infants to the elderly. Gut microbiota were identified by 16S rRNA sequencing from the feces of captive Asian elephants with varying age groups, including infant calves, suckling calves, weaned calves, subadult and adult elephants, and geriatric elephants. The diversity of the gut microbiota was lowest in infants, stable during adulthood, and slightly decreased in the geriatric period. The gut microbiota of the infant elephants was dominated by milk-fermenting taxa including genus Bifidobacterium of family Bifidobacteriaceae together with genus Akkermansia. The fiber-fermenting taxa such as Lachnospiraceae_NK3A20_group were found to be increased in suckling elephants in differential abundance analysis by Analysis of Compositions of Microbiomes with Bias Correction (ANCOM-BC). The gut microbiota profiles after weaning until the adult period has been uniform as indicated by no significant differences in beta diversity between groups. However, the composition of the gut microbiota was found to change again in geriatric elephants. Understanding of the composition of the gut microbiota of captive Asian elephants at various life stages could be beneficial for promoting good health throughout their lifespan, as well as ensuring the welfare of captive elephants

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