The steroid metabolome of pregnancy, insights into the maintenance of pregnancy and evolution of reproductive traits.

Modes of mammalian reproduction are diverse and not always conserved among related species. Progesterone is universally required to supports pregnancy but sites of synthesis and metabolic pathways vary widely. The steroid metabolome of mid-to late gestation was characterized, focusing on 5α-reduced pregnanes in species representing the Perissodactyla, Cetartiodactyla and Carnivora using mass spectrometry. Metabolomes and steroidogenic enzyme ortholog sequences were used in heirarchial analyses. Steroid metabolite profiles were similar within orders, whales within cetartiodactyls for instance, but with notable exceptions such as rhinoceros clustering with goats, and tapirs with pigs. Steroidogenic enzyme sequence clustering reflected expected evolutionary relationships but once again with exceptions. Human sequences (expected outgroups) clustered with perissodactyl CYP11A1, CYP17A1 and SRD5A1 gene orthologues, forming outgroups only for HSD17B1 and SRD5A2. Spotted hyena CYP19A1 clustered within the Perissodactyla, between rhinoceros and equid orthologues, whereas CYP17A1 clustered within the Carnivora. This variability highlights the random adoption of divergent physiological strategies as pregnancy evolved among genetically similar species.

Wagyu-Angus cross improves meat tenderness compared to Angus cattle but unaffected by mild protein restriction during late gestation.

For the spring-calving beef herds, late gestation coincides with winter and early spring, when cows are dependent on feed supplements with low quality hay, which is hard to meet their nutrient requirements. However, the effects of deficiencies of metabolizable protein intake during late gestation on offspring beef quality are unclear. Wagyu cattle have excellent marbling, and insemination with Wagyu vs Angus semen is a practical option for beef producers to improve beef quality. To test, Angus cows (621 ±â€¯73 kg) were selected and randomly separated into two groups. Each group was inseminated with either Angus or Wagyu semen. During the last 90 days of gestation, cows in each group were further separated and received either a low protein diet (85% of the NRC metabolizable protein requirement), which was a low quality hay-based diet common in northwestern region of the U.S., or an adequate protein diet (108% NRC requirement). All progeny was managed together and harvested at a final BW of 576.5 ±â€¯16.6 kg. Wagyu-sired offspring had higher marbling scores and quality grades than Angus (P < 0.01). Protein supplementation did not affect Slice Shear Force (SSF) in either breeds (P = 0.60). However, Wagyu-sired cattle had lower SSF than Angus-sired (P < 0.01). In addition, Wagyu-sired cattle had higher intramuscular fat (P < 0.05) and total collagen content (P < 0.05), but Angus-sired had greater mature collagen cross-links, as shown by higher contents of Pyridinoline (P < 0.01) and Ehrlich Chromogen (P < 0.01). Consistently, the mRNA expression of enzymes catalyzing collagen cross-linking was higher in Angus-sired offspring, including Plod 1 (P < 0.05), Plod 2 (P = 0.08), and P4Hα 2 (P < 0.01). In conclusion, Wagyu-sired cattle had greater tenderness and marbling score compared to Angus-sired, which was associated with lower collagen cross-links. Feeding mature grass hay-based diet with relatively low protein content during late gestation had no major effect on beef quality of subsequent cattle.

Double ovulation and occurrence of twinning in alpacas (Vicugna pacos).

Twin births are rare in alpacas despite the high incidence of double ovulation and are undesirable because they contribute to early and late pregnancy loss, abortion, and birth of nonviable neonates. The objective of the present study was to determine the incidence and outcome of twin pregnancy in double-ovulating alpacas by retrospective and prospective analysis. Data from double-ovulating females (N = 41) presented for pregnancy diagnosis were analyzed to determine pregnancy status at three stages after mating (14-16, 25-30, and 45-53 days). In a prospective study, adult reproductively sound alpacas (n = 21) were examined by ultrasonography to determine the incidence of multiple ovulations. A subset of those alpacas (n = 10) were euthanized either at 9 days (n = 5) or 14 days (n = 5) after a single mating and administration of GnRH to determine presence and number of embryos. A total of 31 cycles were included in the study to determine the incidence of multiple ovulations. In the retrospective study, twin pregnancies were identified between 25 and 30 days in 47.1% of double-ovulating females. There were more twins from bilateral ovulations (62.5%) than from unilateral ovulations (37.5%). Twin pregnancies were either reduced to a singleton (62.5%) or lost completely (37.5%). One set of twins remained viable until Day 52. In the prospective study, double ovulation occurred in 18.8% of the cycles. Two embryos were collected from two of the four double-ovulating females. In conclusion, twin conception is not rare in alpacas. A high rate of spontaneous reduction of twin pregnancies occurs before Day 45. However, a significant proportion of twin pregnancy may result in loss of both embryos. Determination of the number of ovulations (CL) at the time of early pregnancy diagnosis (14-16 days) should be an integral part of any pregnancy evaluation.

Bighorn sheep × domestic sheep hybrids survive Mannheimia haemolytica challenge in the absence of vaccination.

Bighorn sheep (BHS, Ovis canadensis) are much more susceptible than domestic sheep (DS, Ovis aries) to pneumonia caused by leukotoxin (Lkt)-producing members of the Family Pasteurellaceae, particularly Mannheimia haemolytica and Bibersteinia trehalosi. Leukotoxin is widely accepted as the critical virulence factor of these bacteria since Lkt-negative mutants do not cause death of BHS. Typically, DS carry Lkt-positive M. haemolytica and/or B. trehalosi as commensal bacteria in their nasopharynx. In contrast, most BHS do not carry Lkt-positive M. haemolytica or B. trehalosi, or carry Lkt-negative strains in their nasopharynx. In previous studies, we demonstrated that unimmunized DS resist M. haemolytica challenge while BHS succumb to it. We hypothesized that Lkt-neutralizing antibodies, induced by Lkt-positive M. haemolytica and/or B. trehalosi innately carried by DS in their nasopharynx, render them less susceptible to infection by these bacteria. In this study we developed BHS×DS F1 hybrids by artificial insemination of domestic ewes with BHS semen. F1 hybrids were fertile, and produced F2 hybrids and back-crosses. The F1, F2, and back-crosses were raised together with domestic ewes. All these animals acquired Lkt-positive M. haemolytica and/or B. trehalosi, and developed high titers of Lkt-neutralizing antibodies in the absence of vaccination. Furthermore, all of these animals resisted challenge with lethal dose of M. haemolytica. These results suggest that lack of previous exposure to Lkt is at least partially responsible for fatal pneumonia in BHS when they acquire Lkt-positive M. haemolytica and/or B. trehalosi from DS when the two species commingle.

Effect of reproductive tract scoring on reproductive efficiency in beef heifers bred by timed insemination and natural service versus only natural service.

The objective was to determine the effects of reproductive tract score (RTS) on reproductive performance in beef heifers bred by timed artificial insemination followed by natural service (AI-NS) or by natural service only (NSO). Angus cross beef heifers (n = 2660) in the AI-NS group were artificially inseminated at a fixed time (5- or 7-day CO-Synch + controlled internal drug release protocol) once, then exposed to bulls 2 weeks later (bull-to-heifer ratio = 1:40-1:50) for the reminder of the 85-day breeding season. Angus cross beef heifers (n = 1381) in NSO group were submitted to bulls (bull-to-heifer ratio = 1:20-1:25) for the entire 85-day breeding season. Heifers were reproductive tract scored from 1 (prepubertal) to 5 (cyclic) 4 weeks before, and were body condition scored (BCS) from 1 (emaciated) to 9 (obese) at the beginning of breeding season. Pregnancy diagnosis was performed 70 days after AI for AI-NS group and 2 months after the end of breeding season for both groups. Heifers in both groups were well managed and of similar age (14.9 ± 0.4 [AI-NS] and 14.7 ± 0.8 [NSO] months). Pregnancy rates (PRs) and number of days to become pregnant were calculated using PROC GLIMMIX and PROC LIFETEST procedures of SAS. Adjusting for BCS (P = 0.07), expressed estrus (P < 0.05), year (P < 0.05), and BCS by year interaction (P < 0.05), the AI-PR was greater for heifers in AI-NS group with higher RTS (P < 0.0001; 40.7%, 48.3%, 57.6%, and 64.6% for RTS of 2 or less, 3, 4, and 5, respectively). Controlling for BCS (P < 0.05), year (P < 0.05) and the breeding season pregnancy rates (BS-PRs) were greater for heifers in the AI-NS group with higher RTS (P < 0.01; 81.2%, 86.5%, 90.4%, and 95.2% for RTS of 2 or less, 3, 4, and 5, respectively). Similarly, adjusting for BCS, year (P < 0.05), the BS-PR was greater for heifers in NSO group with higher RTS (P < 0.01; 79.7%, 84.3%, 88.4%, and 90.2% for RTS of 2 or less, 3, 4, and 5, respectively). Heifers with higher RTS in both groups became pregnant earlier in the breeding season compared with heifers with lower RTS (log-rank statistics: P < 0.0001). Heifers in the AI-NS group become pregnant at a faster rate compared with those in the NSO group (P < 0.01). The BS-PR for heifers with RTS 5 was different between AI-NS and NSO groups (P < 0.0001). In conclusion, the RTS influenced both the number of beef heifers that became pregnant during the breeding season and the time at which they become pregnant. Furthermore, irrespective of RTS, heifers bred by NSO required more time to become pregnant than their counterparts in herds that used timed AI. The application of RTS system is reliant on the use of synchronization protocol. The application of RTS for selection may plausibly remove precocious females with lower RTS. On the contrary, application of RTS would help select heifers that will become pregnant earlier in breeding season.

Effects of vasectomy on seminal plasma alkaline phosphatase in male alpacas (Vicugña pacos).

Azoospermia is a common finding in male alpacas which present for infertility. The challenge is to differentiate azoospermia of testicular origin from non-testicular origin. In several species, alkaline phosphatase (AP) concentrations in seminal plasma have been used as a diagnostic marker of contributions of the testis and epididymis to the ejaculate. The purpose of this study was to determine whether AP assay could differentiate testicular from non-testicular azoospermia in male alpacas. An experimental model of bilateral outflow obstruction (pre-scrotal vasectomy) was used in 22 male alpacas, aged 2-9 years. No reproductive history was available. Animals were submitted for electroejaculation (EE) under general anaesthesia and vasectomy performed. Five weeks later, animals were submitted for EE. Vasectomy was not successful in one animal, which was removed from analysis. AP levels were compared in seminal plasma in the pre- and post-vasectomy samples. The mean ± SEM concentration of AP in pre-vasectomy seminal plasma was 504.29 ± 166.45 U/l (range 10-2910); the post-vasectomy levels were 252.48 ± 81.77 U/l (range 0-1640; p = 0.06). In 71.4% of animals, AP levels decreased, varying from 18% to 100% reduction. Results of this study suggest that AP is not produced exclusively by the testis and epididymis in alpacas and that AP assay is not a valid diagnostic test for determination of origin of azoospermia; the gold standard for diagnosis of origin of azoospermia remains testicular biopsy.

Chronology of early embryonic development and embryo uterine migration in alpacas.

The objectives were to: (1) describe the chronology of early embryonic development from ovulation to entry into the uterus; and (2) to determine the timing of embryo migration to the left uterine horn when ovulation occurred from the right ovary. The experiment was conducted in Peru. Females (n = 132) were randomly assigned to 15 experimental groups. All females were mated to an intact male, given 50 µg GnRH im (Cystorelin) and ovulation time determined by transrectal ultrasonography, conducted every 6 hours, starting 24 hours postmating. Animals were slaughtered at a specific intervals postovulation and reproductive tracts were recovered and subjected to oviductal and uterine flushing for females slaughtered between 1 and 6 days postovulation (dpo; Day 0 = ovulation) and uterine flushing for females slaughtered from 7 to 15 dpo for recovery of oocytes/embryos. Season of mating did not influence the interval from mating to ovulation (winter: 29 ± 6 hours vs. summer: 30 ± 6 hours; P = 0.49). Ovulation rates for females mated during winter and summer were 92% versus 100%, respectively (P = 0.05). Fertilization rates for winter and summer mated females were 72% and 82% (P = 0.29). Unfertilized ova were not retained in the uterine tube. All embryos collected were in the uterine tube ipsilateral to the side of ovulation between 1 and 5 dpo. Embryos reached the uterus on 6 dpo. Embryos began to elongate on 9 dpo; at this time, 83% of embryos derived from right-ovary ovulations were collected from the left uterine horn. Embryos occupied the entire uterine cavity by 10 dpo. In conclusion, we characterized early embryo development and location of embryo during its early developmental stages in alpaca. This was apparently the first report regarding chronology of embryo development and migration to the left horn in alpaca which merits further investigation regarding its role in maternal recognition of pregnancy.

Presynchronization with GnRH 7 days prior to resynchronization with CO-Synch did not improve pregnancy rate in lactating dairy cows.

The objective was to determine the effect of presynchronization with GnRH 7 d prior to the initiation of resynchronization with CO-Synch on pregnancy/AI (P/AI) of resynchronization in lactating dairy cows, and the effect of GnRH on P/AI from previous breeding. All parity Holstein cows (n = 3287) from four dairy farms were enrolled. Cows not detected in estrus by 28 ± 3 d (Day -7) after a previous breeding were assigned to receive either GnRH (100 µg, im; n = 1636) or no GnRH (Control; n = 1651). Cows not detected in estrus during the 7 d after GnRH underwent pregnancy diagnosis (35 ± 3 d after previous breeding, Day 0); non-pregnant cows (n = 1232) in the Control (n = 645) and GnRH (n = 587) groups were resynchronized with a CO-Synch protocol. Briefly, cows received 100 µg GnRH on Day 0, 25 mg PGF(2α) on Day 7, and 72 h later (Day 10) were given 100 µg GnRH and concurrently inseminated. Serum progesterone concentrations (n = 55 cows) were elevated in 47.3, 70.9, and 74.5% of cows on Days -7, 0, and 7, respectively. The proportion of cows with high progesterone concentrations on Day -7 and Day 0 were 44.1% and 88.2% (P < 0.003), and 55.2% and 33.2% (P > 0.1), for GnRH and Control groups, respectively. Accounting for significant variables such as locations (P < 0.0001) and parity categories (P < 0.05), the P/AI (35 ± 3 d after AI) for resynchronization was not different between GnRH and Control groups [26.7% (95% CI: 23.2, 30.5; (157/587) vs 28.4% (95% CI: 25.0, 31.9; (183/645); P > 0.1]. There were no significant location by treatment or parity by treatment interactions. Accounting for significant variables such as location (P < 0.0001) and parity categories (P < 0.001), the P/AI was not different between GnRH and Control groups for the previous service [60.2%; 95% CI: 57.9, 62.6; (986/1636) vs 59.1%; 95% CI: 56.7, 61.5; (976/1651); P > 0.1)]. There were no significant location by treatment or parity by treatment interactions. In conclusion, more cows presynchronized with GnRH 7 d prior to resynchronization with CO-Synch had elevated progesterone concentrations at initiation of resynchronization than those not presynchronized. The GnRH treatment 7 d prior to resynchronization with CO-Synch, when given 28 ± 3 d after a previous breeding, did not improve P/AI in lactating dairy cows; furthermore, compared to the control, it did not significantly affect pregnancy rate from the previous breeding.

In vitro and in vivo developmental competence of dromedary (Camelus dromedarius) oocytes following in vitro fertilization or parthenogenetic activation.

Parthenogenetic activation of the oocyte represents an important step in the somatic cloning. The aim of the present study was to evaluate the effectiveness (in term of in vitro development) of different methods of parthenogenetic activation of dromedary oocytes. Selected cumulus-oocytes-complexes (n=1264) collected by follicular aspiration from ovaries obtained postmortem were matured in vitro (IVM) for 30 h then divided randomly into seven groups and submitted to artificial activation. Two groups were preactivated with 25 microM of calcium ionophore (CaI) for 20 min then incubated for 4h with either 2mM 6-dimethylaminopurine (6-DMAP) (group 1, n=202) or with 10 microg/mL cycloheximide (CHX) (group 2, n=194). Group 3 (n=172) and group 4 (n=184), oocytes were pretreated with 5 microM ionomycin (Iono) for 5 min then incubated with either 2mM 6-DMAP or 10 microg/mL cycloheximide for 4h, respectively. Group 5 (n=161) and group 6 (n=155) oocytes were preactivated with electrical stimulation (ES) then activated with either 2mM 6-DMAP or 10 microg/mL cycloheximide for 4h, respectively. Group 7 (n=196) oocytes were submitted to in vitro fertilization (IVF) and served as a control. All groups containing oocytes were cultured in vitro following activation or IVF, at 38.5 degrees C under 5% CO(2) in air with >95% humidity. The in vitro development rates of dromedary oocytes exposed to 6-DMAP after CaI (61%), ES (74%) and the IVF group (71%) were similar and significantly greater (P<0.05) than other treatments (10% for group 2, 47% for group 3, 27% for group 4 and 41% for group 6). The blastocyst developmental rate was better (P<0.05) in parthenotes following activation with Iono/6-DMAP (21%) compared to activation with Iono/CHX (12%). However, all were less than that achieved in the IVF group (35%). We conclude that parthenogenetic activation of camel oocytes with 6-DMAP is more effective than activation with CHX for all pre-treatments tested (CaI, Iono or ES). The viability of activated (n=15) or IVF (n=10) hatched-dromedary embryos was examined by transfer to synchronized recipients. An embryonic vesicle was seen by ultrasonography at 15 days post transfer in four females (CaI/6-DMAP: 1/5; 20%, IVF: 3/10; 30%). The only pseudopregnancy obtained with an activated embryo resorbed at 25 days. One of the females receiving the IVF produced embryos aborted at 2 months and the other two females carried to term and gave birth to healthy calves (one female and one male). This study shows that artificial activation of dromedary oocytes with CaI/6-DMAP or ES/6-DMAP is more effective than other treatments in terms of in vitro embryo development. This provides efficient activation conditions which may lead to the development of the somatic cell nuclear transfer procedure in dromedary.

Reproductive emergencies in camelids.

Emergencies in theriogenology practice go beyond just saving the life of the patient, but also preserving its reproductive abilities. Camelid emergency medicine is a relatively new field. This paper discusses the most common reproductive emergencies, their diagnosis, treatment, and prognosis in male and female camelids. The conclusions drawn are based primarily on clinical observations by the authors over the last 25 years. Special consideration is given to peculiarities of the species, particularly in the choice of obstetrical manipulations and therapies.

Quality and developmental ability of dromedary (Camelus dromedarius) embryos obtained by IVM/IVF, in vivo matured/IVF or in vivo matured/fertilized oocytes.

The effect of source of cumulus-oocytes-complexes (COCs), maturation and fertilization conditions on developmental competence of dromedary embryos was examined. Thirty-six adult females were superovulated with equine Chorionic Gonadotropin (eCG) injection (3500 IU, IM) and divided in three groups of 12 females each. Group 1 provided 138 COC's collected from follicles >or= 5 mm 10 days after stimulation prior hCG treatment and matured in vitro for 30 h. Group 2 provided 120 in vivo matured oocytes which were aspirated from their follicles 20 h after hCG (3000 IU, IV) given on day 10 follow eCG injection. Group 3 provided 65 in vivo matured/fertilized oocytes. Females in Group 3 received hCG on day 10 following eCG treatment and then were mated 24 h later. Fertilized oocytes were collected from the oviducts of females 48-h post-mating. Quality of the oocytes was assessed after in vitro maturation (IVM), in vitro fertilization (IVF) and in vitro culture (IVC) of COCs. All cultures were performed in three replicates (n = 3) at 38.5 degrees C, under 5% CO(2) and high humidity (>95%). Only COCs with cumulus and homogenous (dark) cytoplasm were used. Nuclear maturation rate for Groups 1 and 2 was determined by epifluorescence microscopy in a sample of COCs (n = 30) denuded, fixed and stained with Hoechst 33342. To study the viability of obtained embryos, hatched blastocysts from each group were transferred to recipients followed by pregnancy diagnosis using ultrasonography at 15, 60 and 90 days. The percentage of COCs reaching metaphase II (MII) after 30 h of maturation was slightly but not significantly higher for in vivo matured oocytes (28/30; 93%) than those in vitro matured (25/30; 84%). The total rate of cleavage (2 cells to blastocyst stage) was not different for the three groups. However, significantly (p < 0.05) more blastocyst and hatched blastocysts were obtained from in vivo matured and in vivo fertilized oocytes (Group 3; 52% and 73%) than from in vitro fertilized oocytes whether they were matured in vitro (Group 1; 35% and 32%) or in vivo (Group 2; 32% and 45%). Pregnancy rates were not significantly different amongst all groups for the three first months following embryo transfer. All pregnancies were lost after day 90 follow transfer except for in vivo matured and in vivo matured/fertilized groups. Only in vivo matured/in vitro fertilized and in vivo matured/fertilized produced embryos continued normal development until term and resulted in the birth of normal and healthy live calves. Six claves (29%; 6/21) were born from Group 3 and one (8%; 1/13) calf was born from Group 2. This study shows that the IVC system used is able to support camel embryo development. However, developmental competence and viability of dromedary embryos may be directly related to the intrinsic quality (cytoplasmic maturation) of oocytes.

Current knowledge and future challenges in camelid reproduction.

Reproductive biology research on camelids offers some interesting peculiarities and challenges to scientists and animal production specialists. The objective of this paper is to review camelid reproduction, advances in reproductive physiology and reproductive biotechnologies in camelids and discuss some areas for further research. In the female, the focus has been on understanding follicular dynamics. This has allowed development of synchronization and superovulation strategies to support embryo transfer technologies which are now commonly used in camels. Some advances have been achieved in preservation of embryos by vitrification. Fertilization, early embryo development and embryo signaling for maternal recognition of pregnancy are still not fully understood. New information on the interaction of the developing embryo and the endometrium may shed some light on this signaling as well as the mechanism of prevention of luteolysis. The presence of a seminal ovulation-inducing factor (OIF) was confirmed in llamas and alpacas. Chronology of oocytes maturation has been described. In vitro production of embryos has been achieved resulting in successful pregnancies and births in the dromedary. These techniques offer a new tool for the production and study of interspecies/cross-species embryos and their effect on pregnancy. Male reproductive function remains poorly studied. Semen preservation and artificial insemination still present many challenges and are not used in production at the moment. The involvement of climatic and nutritional conditions as well as the role of leptin in the regulation of reproductive function need to be evaluated.

Effect of follicular size on in vitro developmental competence of oocytes and viability of embryos after transfer in the dromedary (Camelus dromedarius).

The aim of this work was to determine the effect of follicle size on camel oocyte quality as measured by developmental competence in vitro and in vivo. Ovaries from a local slaughterhouse were dissected to obtain two classes of follicle size: small (3-6 mm) and large (>6 mm) follicles. Quality of the oocytes was assessed after in vitro maturation (IVM), in vitro fertilization (IVF) and in vitro culture (IVC) of cumulus oocyte complexes (COCs). All cultures were done in four replicates at 38.5 degrees C, under 5% CO(2) and high humidity (>95%). Only COCs with cumulus and homogenous (dark) cytoplasm were used. The COCs were matured for 28 h in TCM-199 medium supplemented with 10% heat-treated fetal calf serum (FCS), 10 ng/mL EGF, and 250 microM cysteamine. Nuclear maturation rate for each class of follicle size was determined by contrast phase microscopy in a sample of COCs (n=30) denuded, fixed and stained with aceto-orcein. In vitro fertilization was performed using fresh semen (0.5 x 10(6)spermatozoa/mL in modified TALP-solution). Fertilized oocytes were cultured in mKSOMaa, under 5% O(2) and 90% N(2). The percentage of COCs reaching metaphase II (MII) after 28 h of maturation was 87% (26/30) and 73% (22/30) for oocytes originating from large and small follicles, respectively (P>0.1). The rate of total cleavage (two cells to blastocyst stage) was greater (P<0.05) for oocytes originating from large follicles (72%; 116/162) than for those derived from small follicles (59%; 140/237). The percentage of fertilized oocytes reaching the blastocyst stage was 35% (57/162) and 20% (48/237) for oocytes collected from large and small follicles, respectively (P<0.05). The viability of in vitro-produced hatched blastocyst from the two groups (15 from 3 to 6mm follicle size and 22 from follicles >6 mm) was assessed by transfer to synchronized recipients. None of the hatched blastocysts from small follicles resulted in a pregnancy whereas 68% (15/22) of the transferred hatched embryos from large follicles developed into a 25-day pregnancy. Of the resulting 15 pregnancies, 53% (n=8) aborted (five between 2 and 4 months and three between 5 and 7 months of pregnancy). The remaining seven pregnant females gave birth to normal healthy offsprings (four females and three males). The present study shows that dromedary oocytes developmental competence is acquired late during the final phase of follicular development and this developmental ability translates into greater pregnancy rates after transfer of in vitro produced hatched blastocysts.

Infectious causes of reproductive loss in camelids.

Reproductive losses in camelids are due to infertility, pregnancy loss, udder diseases and neonatal mortality caused by a variety of infectious diseases. Uterine infection and abortion represent the major complaint in camelid veterinary practice. The major infectious organisms in endometritis and metritis are E. coli and Streptococcus equi subspecies zooepidemicus. Abortion rates due to infectious diseases vary from 10% to more than 70% in some areas. Leptospirosis, toxoplasmosis and chlamydiosis have been diagnosed as the major causes of abortion in llamas and alpacas. In camels, brucellosis and trypanosomiasis represent the major causes of infectious abortion in the Middle East and Africa. Mastitis is rare in South American camelids. The prevalence of subclinical udder infection in camels can reach very high proportions in dairy camels. Udder infections are primarily due to Streptococcus agalactiae and Staphylococcus aureus. Neonatal mortality is primarily due to diarrhea following failure of passive transfer and exposure to E. coli, rotavirus, coronavirus, Coccidia and Salmonella. This paper reviews the etio-pathogenesis of these causes of reproductive losses, as well as the major risk factors and strategies to prevent their occurrence.

Biosecurity and biocontainment in alpaca operations.

Biosecurity on South American camelid operations involves both external and internal measures to prevent the introduction and spread of disease. External biosecurity involves practices and techniques directed at the prevention of entry of new diseases into a group of animals. Internal biosecurity or biocontainment, involves practices and techniques that are directed at the prevention or spread of disease within an existing group of animals. External biosecurity is particularly important in North America camelid operations due to the extensive movement of animals for breeding or show purposes. Internal biosecurity typically involves this the prevention and treatment of failure of passive transfer, maintenance of proper nutrition and housing, and the implementation of an appropriate vaccination program for endemic or relevant diseases. Attention to appropriate cleaning and disinfection procedures related to housing, feeding, and treatment equipment is important for the maintenance of both internal and external biosecurity practices. This paper discusses various risk factors associated with the control of infectious disease in the context of external and internal biosecurity measures in camelids operations.

Update on reproductive biotechnologies in small ruminants and camelids.

Recent advances in reproductive biotechnologies in small ruminants include improvement of methods for in vitro production of embryos and attempts at spermatogonial stem cell transplantation. In vitro production of embryos by IVM/IVF, intra-cytoplasmic sperm injection (ICSI), or nuclear transfer (NT) has been made possible by improvements in oocyte collection and maturation techniques, and early embryo culture systems. However, in vitro embryo production still is not very efficient due to several limiting factors affecting the outcome of each step of the process. This paper discusses factors affecting in vitro embryo production in small ruminants and camelids, as well as preliminary results with the technique of spermatogonial stem cell transplantation.

In Vitro and in vivo developmental competence of dromedary (Camelus dromedarius) embryos produced in vitro using two culture systems (mKSOMaa and oviductal cells).

Development competence and pregnancy rate of in vitro-produced (IVP) dromedary embryos were studied in two culture systems: (i) semi-defined modified medium (mKSOMaa) and (ii) co-culture using camel epithelial oviducal cells. Five hundred and three cumulus-oocytes complexes (COCs) were selected, allowed to mature, fertilized and cultured in vitro (38.5 degrees C; 5% CO2, maximum humidity > 95%, with concentration of oxygen of 5% for semi-defined medium and 20% for co-culture cells). Maturation was accomplished by incubation in TCM-199 medium supplemented with 10% heat-treated foetal calf serum (FCS), 10 ng/ml epidermal growth factor, 1 microg/ml follicle-stimulating hormone, 1 microg/ml oestradiol and 500 microM cysteamine for 30 h. In vitro fertilization (IVF) was performed using fresh semen (0.5 x 10(6) spermatozoa/ml in modified TALP solution). Fertilized COCs were denuded by vortexing, then cultured in either mKSOMaa (10% heat-treated FCS was added 24 h post-IVF), under 5% O2 and 90% N2 (group 1; n = 249) or with dromedary epithelial oviducal cell monolayers in TCM-199 with 10% heat-treated FCS under 20% O2 (group 2; n = 254). The rate of cleavage was significant higher (p < 0.05) for group 1 (63%, 156/249) than for group 2 (51%, 130/254). No significant difference was found between the two groups in the rate of development to blastocyst (21% vs 16.5%) and their hatchability (21% vs 14%). Pregnancy rates were similar for the first 60 days. However, all pregnancies were lost after 60 days with the exception of two of six (33%) from recipients of hatched blastocysts from group 1. We conclude that both systems support in vitro production of dromedary embryos by in vitro maturation (IVM)/IVF of oocytes. However, embryos obtained by culture in the semi-defined medium (mKSOMaa) appear to have a better in vivo development ability.

Changes in spermatogenesis and endocrine function in the ram testis due to irradiation and active immunization against luteinizing hormone-releasing hormone.

Spermatogonial stem cell transplantation is a technique that has potential in livestock to enhance genetic gain and generate transgenic offspring through the male germ line. A means for depletion of endogenous germ cells in a recipient's seminiferous tubules is necessary for this technology to be applied. The objectives of this study were to evaluate several methods for depletion of endogenous germ cells in the testes of adult rams and to evaluate ultrasound-guided injections into the rete testes as a means for infusing a suspension into the seminiferous tubules. Sixteen adult rams were randomly divided into 4 treatment groups (n = 4 per group). Treatments consisted of active immunization against LHRH (IMM), localized testicular irradiation (IR), LHRH immunization + irradiation (IMM+IR), and untreated control. Serial bleedings were conducted pretreatment and monthly after treatment for 4 mo, at which time all rams were castrated. Both IMM and IMM+IR rams received exogenous gonadotropin in the form of Perganol weekly for 8 wk before castration to bypass the immunization. All rams also received an ultrasound-guided injection of PBS containing 0.4% trypan blue into the rete testis of one testicle before castration. Rams receiving IMM and IMM+IR treatments had higher (P < 0.05) average percentages of seminiferous tubule cross sections with depleted germ cells compared with controls. Serum testosterone was decreased (P < 0.05) in IMM and IMM+IR rams 1 mo after treatment and throughout the remainder of the study compared with controls and IR rams, which were not different from each other. Serum inhibin concentration was unchanged in all rams following treatment indicating that Sertoli cell function was unaltered. A greater (P < 0.05) average percentage of the total testicular area could be filled with the trypan blue solution by rete testis injection in IMM and IMM+IR rams. These data demonstrate the depletion of endogenous germ cells in adult ram testes without alteration of Sertoli cell viability and function that have potential as methods for preparing recipient animals for germ cell transplantation.

Retrospective study on an unusual form of ovario-bursal pathology in the camel (Camelus dromedarius).

A peculiar form of ovarian bursa pathology, called hydrobursitis, is described in the dromedary camel. This malformation is characterized by a collection of fluid within the ovarian bursa with encapsulation of the ovary. Retrospective study of all diagnosed cases in our laboratory shows that this malformation is responsible for reduced reproductive performance due to abortion, infertility and embryonic death. Hydrobursitis was diagnosed in 33 of 355 animals examined. The incidence of this lesion was higher in animals that aborted (9/24) or were barren for more than 2 years (13/37). Diagnosis by palpation per rectum and ultrasonography, is described. Gross pathological and biochemical studies on specimens collected at surgery or in the slaughterhouse showed that the fluid is hemorrhagic and that it could be a mixture of blood and follicular fluid. Bacteriological and cytological studies were inconclusive. The origin of this lesion is discussed in light of genetic, pathological and management factors. Loss of embryos in affected animals could be due to the effect of increased tension on the uterus caused by the accumulation of fluid which ranged in volume from 250 mL to 4240 mL. The condition was treated in several females by surgical ablation. Embryo transfer was used in females that were bilaterally affected and underwent a bilateral ovariectomy. Pregnancy and calving rates were 52.5% and 38.1% for unilaterally ovariectomized females bred after surgery and 33.3% and 22.2% for bilaterally ovariectomized females used as embryo recipients after surgery. These results show that involvement of the uterus in this malformation is very limited and that surgery can be considered for valuable animals if they are unilaterally affected.

Septic orchitis in an alpaca.

An adult, intact male alpaca was presented with an acute onset of unilateral scrotal swelling. Following complete physical and ultrasonographic examination, the most likely differential diagnoses were orchitis, hematoma, and testicular torsion. The animal was castrated and histopathologic evaluation revealed unilateral orchitis. Streptococcus equi subsp. zooepidemicus was cultured.