Superovulatory responses to eCG in llamas (Lama glama )

Llamas are copulation-induced single-ovulators, and multiple ovulation and embryo transfer (MOET) methods have not yet been developed for this species. Superovulatory responses to eCG given during an induced (Group A) or simulated (Group B) luteal phase were investigated using ultrasound to observe ovarian follicles and corpora lutea (CLs) and plasma progesterone was used to assess luteal function. Embryos were recovered nonsurgically. Group A (n = 19): donors were given 8 microg, im GnRH analogue (Day 0) to induce ovulation of a mature follicle, 1000 IU, im eCG (Day 7), and 250 microg PGF(2alpha) analogue (Day 9). Group B (n = 17): donors were given a subcutaneous progestagen implant (3 mg Norgestomet) at Days 0 to 7) and 1000 IU, im eCG (Day 5). When most (>65%) of the follicles in both Groups A and B had matured at 5 to 11 d post eCG, the donors were given 8 microg, im GnRH and mated once (n = 26) or twice within a 24-h interval (n = 10); embryos were recovered 6 to 9 d post ovulation. More follicles and corpora lutea were induced in Group B than in Group A, but a similar mean number of embryos were recovered (1.3 vs 1.6), and a similar proportion of donors yielded multiple embryos (35 vs 32%). The embryo recovery rate was similar for Groups A and B (39 and 37%), but it was higher (P < 0.001) with 2 (72%) rather than 1 (22%) mating, and it was negatively correlated with CL number (P < 0.05). Overall, 80% of the llamas had a precocious CL and elevated plasma progesterone concentrations when multiple follicles reached maturity. This was associated with increased subsequent superovulation and embryo recovery (P < 0.01). Peak plasma progesterone was positively correlated with the CL number (P < 0.05). From these results we conclude that superovulation may be achieved with eCG given during either an induced or a simulated luteal phase, that embryo recovery is improved following 2 matings rather than 1, and that MOET may indeed be feasible for use in the llama.     

Prediction of gestational age by ultrasonic fetometry in llamas (Lama glama) and alpacas (Lama pacos)

Fetal biparietal diameter (BPD) and thorax height (TH) were measured by ultrasound during intrauterine growth in pregnant llamas (Lama glama) and alpacas (Lama pacos). The goal was to establish representative curves that allows estimation of gestational age (GA) from real-time ultrasonic measurements of these fetal structures at any stage of gestation. Llamas and alpacas were mated under controlled conditions. Ultrasound exams were conducted to determine pregnancy status 1 month later. Measurements of fetal BPD and TH were conducted from the second month of pregnancy until term. Observation and assessment of fetal TH was difficult during the last 3 months of pregnancy, specially in llamas. Regression curves were calculated from the data as a function of GA, with the best fit represented by the following equations: llama GA=(BPD-0.002399)43.02293,r=0.98,P<0.001; llama GA=(TH-0.07137)46.94485, r=0.95,P<0.001; alpaca GA=(BPD-0.11376)47.23287, r=0.98,P<0.001; alpaca GA=(TH-0.36436)52.87663, r=0.96,P<0.001, where GA was measured in days and BPD and TH in centimeters. Results indicate that ultrasonic measurement of these fetal biometric variables constitute a valuable tool to estimate GA at any stage of pregnancy in these domestic South American camelids.     

Psoroptes sp. in two llamas (Lama glama) in Washington.

Psoroptes sp. mites were isolated from two llamas (Lama glama), a 4-mo-old male and its dam, in Washington. Mites were restricted to the ears and were responsible for head-shaking and incoordination. Infestations were treated successfully with a subcutaneous injection of ivermectin at 0.2 mg/kg of body weight and with 2 drops of ivermectin diluted in saline given topically in each ear. This is the first known report of Psoroptes sp. in llamas in the United States.     

Experimental infestation of llamas (Lama glama) with Boophilus microplus (Acari: Ixodidae)

Two llamas (Lama glama) were infested with approximately 20,000 Boophilus microplus larvae each and maintained under controlled conditions to collect the detached adult ticks. A total of 72 engorged and semi-engorged females were recovered with a mean weight of 163.0±70.6mg. Sixty-three (87.5%) of these ticks oviposited and 60 (95.2%) of the egg masses produced larvae. Six of the heaviest females laid 18,838 eggs from which 16,545 (87.8%) hatched. These results show that llamas are able to maintain a population of B. microplus under experimental conditions. Further studies are required to determine the role of llamas as alternative hosts for B. microplus under field conditions.     

Experimental infections of Eimeria alpacae and Eimeria punoensis in llamas (Lama glama).

Four llamas (Lama glama) ranging in age from 1.5 yr to 7 yr each were inoculated orally with 10,000 (n = 2) or 50,000 (n = 2) sporulated oocysts of Eimeria alpacae (25%) and Eimeria punoensis (75%). The prepatent period for E. aplacae was 16-18 days, and it was 10 days for E. punoensis. Patent periods for E. alpacae and E. punoensis were approximately 9 days and 24 days, respectively. Although large numbers of oocysts were present in feces, no clinical sign of coccidiosis was observed. Based on ths experiment, E. alpacae and E. punoensis at the numbers given are not likely pathogenic in healthy llamas older than 1 yr.     

First report of Neospora caninum infection in adult alpacas (Vicugna pacos) and llamas (Lama glama)

Neospora caninum is a cyst-forming coccidian that mainly affects bovines, although Neospora infection has also been described in other domestic and wild ruminant species. Serum samples from 78 alpacas (Vicugna pacos) and 73 llamas (Lama glama) at a unique dilution of 1:50 tested by indirect fluorescent antibody test (IFAT) were further analyzed serologically by IFAT and Western blot in both ruminant species to avoid cross-reactions with closely related coccidian parasites and to confirm the existence of N. caninum-specific antibodies. IFAT titers ranging between 1:50 and 1:800 were found. When using Western blot, N. caninum tachyzoite-specific immunodominant antigens with apparent molecular weights of 17-18, 34-35, 37, and 60-62 kDa were also recognized, although some sera with 1:50 IFAT titers proved not to have N. caninum-specific antibodies. As expected, higher IFAT titers were associated with higher anti-N. caninum reactivity in Western blot. This report documents for the first time the presence of N. caninum infection in adult alpacas and llamas from Peru.     

Clinical and serologic evaluation of two llamas (Lama glama) infected with Toxoplasma gondii during gestation

Two pregnant llamas (Lama glama) infected with Toxoplasma gondii and their offspring were evaluated clinically and serologically. Llama 1 was inoculated orally with 1,000 infective occysts of the P89 strain of T. gondii at 82 days of gestation (DOG). Llama 2 became naturally infected with T. gondii between 26 and 119 DOG. Both llamas remained clinically normal and delivered healthy offspring. Sera collected from both llamas during pregnancy and from their offspring before and after colostral ingestion were evaluated for antibodies to T. gondii by the modified agglutination test (MAT), latex agglutination test (LAT), indirect hemagglutination test (IHAT), and the Sabin-Feldman dye test (DT). In llama 1, MAT antibody titers were < 1:20, 1:320, 1:1,280, 1:640, and 1:80 at 82, 97, 109, 132, and 152 DOG, respectively. The MAT titers in naturally infected llama 2 were < 1:32, 1:320-1:640, and 1:1,280 at 26, 119-200, and 346 DOG, respectively. In both llamas, antibody titers in the DT were of similar magnitude as the MAT, but titers in the LAT and IHAT were inconsistent. Antibodies to T. gondii were not detected in precolostral sera obtained from offspring of both llamas suggesting there was no fetal T. gondii infection.     

Changes in testicular histology and sperm quality in llamas (Lama glama), following exposure to high ambient temperature

The aim of the study was to investigate whether a moderately elevated ambient temperature (29 degrees C, 4 weeks, 24hperday) has an effect on the spermatogenesis in male llamas (Lama glama) and to monitor the recrudescence of spermatogenesis. Thirteen llamas were used. Semen parameters were monitored in four of the llamas and six animals were castrated at different times after the heat treatment. An additional three llamas were used as control animals and were castrated without any treatment. Spermatogenesis was found to be severely impaired due to the high environmental temperature. Sperm concentration declined from 97.5 million to 10 million spermatozoa/ml. Sperm motility declined from 63.1% to 15.0% and the percentage of morphologically abnormal sperm cells increased from 26.3% up to 50.5%. The changes in sperm parameters corresponded to the histological analysis of the testes. We found an increase in destroyed tubules, where no stage of the spermatogenic cycle could be established from 1.8% up to 38.2%, and a reduction of the spermatogonial proliferation rate (Ki-67 histochemistry) represented by tubules with proliferating spermatogonia from 79.5% to 45.7% directly after the heat treatment. Apoptosis (TUNEL assay) showed no significant changes during the experiment. The recrudescence of spermatogenesis within 6 weeks after the heat treatment was found to be due to an increase of mitotic proliferation of spermatogonia and not due to a decrease in the apoptotic rate. Our data indicate that in llamas the thermoregulatory ability is not sufficient enough to prevent heat caused damage to the testis at longer acting ambient temperature of 29 degrees C.     

Suckling behavior of Llama (Lama glama glama) and Chapman's Zebra (Equus burchelli antiquorum) in captivity

Suckling behavior of one Llama foal and one Chapman's Zebra foal born in captivity is presented. Both animals suckled less frequently and spent more time grazing as they grew older. The suckling frequency of the Zebra foal was twice that of the Llama but on the average each Llama-suckle lasted twice as long. These differences are analyzed and evidence for a dual function of suckling behavior is discussed.     

Successful transfer of vitrified Ilama (Lama glama) embryos

The exploitation of the domestic animals species of South American camelids is of great social importance for the native people living in the High Andes. The reproductive physiology of these species is a unique challenge in the development of advanced breeding techniques. At present, the cryopreservation of embryos has not been developed and very few investigations have been conducted. The objective of the present work was to evaluate the in vivo survival of vitrified llama embryos after transfer to recipient females. Donors females were treated with a CIDR-estradiol benzoate-eCG regimen and were mated naturally 6 days after CIDR withdrawal. One ovulatory dose (8 microg) of GnRH was administered immediately after mating. A second mating was allowed 24 h later. Embryo recovery was performed nonsurgically between 8 and 8.5 days after the first mating. Twenty-two ova/embryos were recovered from 12 donor females. Hatched blastocysts were exposed to vitrification solution (20% glycerol + 20% ethylene glycol + 0.3 M sucrose + 0.375 M glucose + 3% polyethylene glycol (P/V)) in three steps, and after loading into 0.25 ml straws, were plunged into liquid nitrogen. For embryo transfer, recipients animals were ovulation-synchronized using GnRH administered at the same time as donors. A total of eight vitrified-warmed embryos and 12 fresh embryos were nonsurgically transferred to four and six recipient females, respectively (two embryo per recipient). The pregnancy rates were 50 and 33.3% for recipients that had received vitrified embryos and fresh embryos, respectively. The results demonstrated the effectiveness of this simple vitrification method for cryopreservation of llama embryos.     

Transvaginal collection and ultrastructure of llama (Lama glama) oocytes

Ultrasound-guided transvaginal follicle aspiration has been described as a noninvasive and repeatable procedure for oocyte collection in several species, but its use has not been described for any of the members of the family, Camelidae. A study was designed to determine the feasibility of an ultrasound-guided transvaginal approach for oocyte collection in llamas. Fifteen non-pregnant, adult female llamas (10 non-stimulated and 5 superstimulated) were examined by transrectal ultrasonography with a 7.5-MHz linear-array transducer to determine the number and diameter of follicles available for aspiration. After caudal epidural anesthesia was induced, the 7.5-MHz linear-array transducer was fastened to a long rigid handle and inserted intravaginally. The free hand was placed into the rectum to manipulate the ovaries, one at a time, in position against the vaginal wall over the face of the transducer. A 20-gauge, 55-cm-long, single-lumen needle was advanced through the vaginal fornix and into follicles > or = 3 mm in diameter. Follicular contents were aspirated using a regulated vacuum pump (flow rate = 33 mL/min; approximately 150 mm Hg) into a tube containing 3 mL of phosphate buffered saline and 0.2% BSA. Fluid was filtered (75 microm mesh), and oocytes were located and morphologically evaluated using a stereomicroscope. Overall, 134 follicles were aspirated, and 76 oocytes were collected (collection rate = 57%). Thirty-two oocytes (42%) were surrounded by multiple layers of compacted granulosa cells and had homogenous dark ooplasm; 13 oocytes (17%) were surrounded by the corona radiata layer only and had heavily granulated ooplasm; 9 oocytes (12%) were denuded and had homogenous dark ooplasm; and 22 oocytes (29%) were denuded and displayed signs of ooplasm degeneration. The ultrastructure of llama oocytes was similar to that of cattle except for conspicuous accumulation of large lipid droplets in the cytoplasm. Twenty-four hours after follicle aspiration, the ovaries were examined by transrectal ultrasonography and intrafollicular hematomas were detected in 3 llamas (9 of 48 follicles aspirated). Results demonstrate the potential utility of a transvaginal ultrasound-guided technique for oocyte collection and in vitro embryo production in llamas. Oocytes of llamas bear an ultrustructural resemblance to those of cattle, but are distinguished by a predominance of cytoplasmic lipid.     

Use of cloprostenol as an abortifacient in the llama (Lama glama)

As part of a larger project investigating the development and heritability of choanal atresia glama), it was necessary to develop a protocol for aborting llamas at various stages of gestation. Twenty-seven animals between 4 and 7 mo of gestation were successfully aborted a total of 53 times following two 250 microg intramuscular injections of cloprostenol at 24 h intervals. Abortion was induced once in 10 animals and multiple times (range 2 to 5) in 17 animals. Twenty-four animals (45.2%) aborted 3 d following the first injection, with 20 animals (37.7%) aborting 4 d post prostaglandin administration. Other animals aborted at 2 d (n=6, 11.3%), 5 d (n=2, 3.8%), and 7 d (n=1, 1.9%) following drug administration. Forty-nine (92.5%) of the abortions occurred following a single series of injections, while 4 animals (7.5%) aborted following a second series of injections. No confirmed pregnant animals failed to abort following the second series of cloprostenol injections. Conception rates in animals rebred 2 to 4 wk following an abortion were comparable to those of untreated animals in the research herd. Unlike the severe hypertension and death that has been reported following dinoprost tromethamine administration in the llama, no adverse reactions were observed in this study following cloprostenol administration. The results demonstrate that llamas can be safely and effectively aborted up to 7 mo of gestation (normal full term gestation = 342 +/- 10 days) without adverse effects on subsequent fertility.     

Melanins and melanosomes from llama (Lama glama L.)

Analysis of melanins and melanosomes in eight hair and skin samples taken of adult pigmented Argentine llamas (Lama glama L.) has been carried out. In each sample, eumelanins, pheomelanins and alkali-soluble melanins were identified. The total amount of melanins and the amount of eumelanins both decreased from black to reddish brown colour, while pheomelanins were found to be present in small quantities in each sample. Eumelanosomes were round and oval-shaped, displaying transverse striations clearly visible at low magnification. Dark brown samples revealed all four melanosomes stages. Stages I and II melanosomes appeared as large, asymmetrical vacuoles containing numerous microvesicles randomly scattered within an amorphous proteinaceous material (vesiculo-globular bodies). Stage III melanosomes had microgranular melanin deposits in the microvesicles and in the matrix. The fully melanized melanosomes (stage IV) were primarily round-shaped, showing an irregular outline and the electron-dense pigment was arranged to form large clusters. In light brown melanocytes, numerous melanosomes at different maturation stages could be found. Premelanosomes appeared ovoid, containing amorphous proteinaceous material and spotty and microgranular deposits. Mature melanosomes were fully melanized, homogeneously electron-dense, ovoid granules.     

Quantitative variation of melanins in llama (Lama glama L.)

The amount of melanin pigments was investigated in 80 Argentinean llama, representative of seven phenotypes and four different fleece colours, by means of spectrophotometric assays: SpEM (spectrophotometric eumelanin), SpPM (spectrophotometric pheomelanin), SpASM (spectrophotometric alkali soluble melanin), and SpTM (spectrophotometric total melanin). It was found that, although to a different extent, the quantitative variation of these pigments was affected both by phenotypes and fleece colours and, hence, it was possible to identify and to distinguish homogeneous groups on the basis of these two factors. In particular, SpEM revealed the most reliable parameter for a discrimination among these groups. Low concentrations of this pigment characterize red, wild, and red black phenotypes and distinguish them from black and tan and non-agouti black ones, both showing high concentrations; low concentrations also distinguish reddish brown fleece colour from very dark red and black ones.     

Synchronisation of ovarian follicular waves in the dromedary camel (Camelus dromedarius)

This study was designed to compare the efficacy of various treatments intended to synchronise follicular wave cycles in dromedary camels by removing the existing follicle of unknown size and replacing it with a follicle capable of ovulating at a known time. Camels were randomly assigned to one of five groups and treated with either (1) 5mg oestradiol benzoate (i.m.) and 100mg progesterone (i.m.; E/P, n=15), (2) 20 icrog GnRH analogue, buserelin (i.m.; GnRH, n=15), (3) 20 microg buserelin (i.m.) on Day 0 (T=0) and 500 microg prostaglandin on Day T+7 (GnRH/PG n=15), (4) transvaginal ultrasound-guided follicle ablation of all follicles > or =0.5 cm (ABL, n=15) or (5) 5 ml saline (i.m; Controls n=15). All camels were subsequently injected with 20 microg buserelin 14 days after the first treatment was given. The ovarian response was monitored daily by transrectal ultrasonography and the intervals from treatment to follicular wave emergence and also the day on which the new dominant follicle reached 1.3 cm was recorded. Amongst the treatment groups the mean interval from treatment to new follicle wave emergence and treatment to time taken for the new dominant follicle to reach 1.3 cm in diameter was shortest in the ABL group (2.3+/-0.5 days and 8.8+/-1.1 days respectively, P=0.044) and longest in the E/P group (6.4+/-0.8 days and 12.2+/-1.0 days respectively, P<0.001) whereas the GnRH and GnRH/PG groups were intermediate (3.0+/-0.5 days and 11.1+/-0.8 days GnRH; and 4.5+/-0.5 days and 10.7+/-0.7 days GnRH/PG). A total of 11/15 camels in both the GnRH and GnRH/PG groups had dominant follicles between 1.3 and 1.9 cm 14 days post treatment, of which 21 of the 22 follicles ovulated after GnRH injection on T+14. The ABL, E/P and control groups however, showed greater variability in follicle size with less camels having dominant follicles between 1.3 and 1.9 cm than the GnRH and GnRH/PG groups and more in the > or =2.0 cm or follicle regressing groups, therefore fewer of these camels ovulated (ABL n=7; E/P n=9; Control n=6) after GnRH injection on Day T+14. In conclusion, two GnRH injections 14 days apart or two GnRH injections 14 days apart and PG on Day 7 after the first GnRH were the most effective methods to synchronise ovulation rate in dromedary camels at a fixed time interval of 14 days after treatment.     

Collection method, season and individual variation on seminal characteristics in the llama (Lama glama)

The objective of the present study was to evaluate the effect of semen collection method (electroejaculation "EE" as compared with the artificial vagina "AV"), the season (summer versus winter) and the male used on macroscopic and microscopic characteristics of ejaculates in llamas. A total of 110 ejaculates were collected from six males and 92 of them were analyzed. Ejaculate volume, concentration, total sperm and the following sperm characteristics were studied: motility, membrane function (HOS test), membrane integrity (CFDA/PI fluorochromes) and morphology. A mixed linear model, that considered season and collection method as the fixed variables and the male as the random variable, was used for the statistical analysis. Variability was found between males (p相似文献   

Evaluation of DNA fragmentation in llama (Lama glama) sperm using the sperm chromatin dispersion test

The integrity of sperm chromatin is now viewed as an important factor in male fertility and in early embryonic development. The objectives of this study were: (1) adapt the simple and inexpensive sperm chromatin dispersion (SCD) test to evaluate DNA fragmentation in llama sperm and establish the halo patterns observed in this species, (2) determine an effective and reliable positive control for this technique and (3) evaluate correlation between the SCD test and the toluidine blue (TB) stain. To adapt the SCD test, three different mercaptoethanol (ME) concentrations were assayed (2.5%, 5% and 10% ME). To determine an effective positive control, three treatments (incubation at 100 °C for 30 min, incubation with 0.3 M NaOH for 30 min at room temperature and exposure to UV light for 2h) were assayed. The concentration selected to use in the SCD test was 5% ME, because it produced the largest halo while still conserving the structure of the core. Four DNA dispersion patterns were clearly observed: (I) nuclei with large DNA dispersion halos; (II) nuclei with medium halos; (III) nuclei with very small halos and (IV) nuclei with no halo. All treatments used as positive controls were effective in producing DNA fragmentation. A high correlation (r=0.84, P=0.03) was observed between spermatozoa without halos and TB positive cells. To conclude, SCD patterns in llama sperm have been established as well as a repeatable positive control for the assay. The SCD test and TB stain are simple and inexpensive techniques that can be used to evaluate DNA damage in llama sperm.     

Occurrence of microtubules during erythropoiesis in Llama, Lama glama

Studies on the ellipsoid erythroblasts of Llama, L a m gluma, L. during erythropoiesis showed the appearance of the marginal band composed of approximately 29 microtubules. As the maturation of erythrocytes goes on, the number of marginal band microtubules diminishes. No microtubules were found in mature erythrocytes of Llama.     

Endocrine changes during pregnancy, parturition and the early post-partum period in the llama (Lama glama)

Mean (+/- s.d.) pregnancy length for the 14 llamas in this study was 350 +/- 4.5 days. Plasma progesterone concentrations increased by 5 days after mating and remained elevated (greater than 2.0 ng/ml) throughout most of pregnancy. At about 2 weeks before parturition, plasma progesterone concentrations began to decline, dropped markedly during the final 24 h before parturition, and returned to basal concentrations (less than 0.5 ng/ml) by the day of parturition. The combined oestrone + oestradiol-17 beta and oestradiol-17 beta concentrations varied between 6 and 274 pg/ml and 4 and 114 pg/ml, respectively, during the first 9 months of pregnancy. Concentrations increased between 9 months after mating and the end of pregnancy with peak mean concentrations of 827 +/- 58 (s.e.m.) pg oestrone + oestradiol-17 beta/ml (range: 64-1658) and 196 +/- 10 pg oestradiol-17 beta/ml (31-294) during the last week of pregnancy. Concentrations then declined to 87 +/- 14 pg oestrone + oestradiol-17 beta/ml (7-488) and 25 +/- 5 pg oestradiol-17 beta/ml (2.5-142) during the first week post partum. Plasma cortisol concentrations varied between 2.6 and 51.9 ng/ml (14.0 +/- 0.5) from mating until 2 weeks before parturition when the concentrations began to decline. Only a slight increase in plasma cortisol concentrations was observed in association with parturition. Plasma triiodothyronine concentrations varied between 0.5 and 4.5 ng/ml (1.9 +/- 0.1) throughout pregnancy and the periparturient period.(ABSTRACT TRUNCATED AT 250 WORDS)