Semen collection methods in alpacas.

South American camelids, particularly alpacas, are gaining in popularity outside their native lands. Reproductive biotechnologies, such as artificial insemination or embryo transfer with cryopreserved embryos, are more complicated in these species than many others due to differences in their reproductive physiology compared to the more commonly encountered domestic livestock. This article reviews the methods currently available for obtaining semen or spermatozoa from alpacas and describes some of the problems associated with handling the viscous seminal plasma characteristic of camelids. Possibilities and limitations of reproductive biotechnologies in this species are discussed, and future developments are outlined, especially some new techniques which are currently being developed for use with camel semen and may be transferable to alpacas.

Comparison of differents methods of sperm selection of llama raw semen.

The objective of this study was to compare the efficiency of different sperm selection methods applied to the same llama ejaculate. Four treatments were compared: two variants of the swim up technique (with and without seminal plasma), and two different colloids, Androcoll-E-Large and Percoll(®). Using electroejaculation, 21 semen samples were obtained from 7 llama males (n=7, r=3). The ejaculates were incubated in a solution of 0.1% collagenase, to decrease thread formation, and then split into 4 aliquots: one aliquot was layered over a column of Androcoll-E-Large (SLC) and the second over a column of Percoll (45%). The third aliquot was deposited in a tube with culture medium and was incubated at a 45° angle for 30min at 37°C (SU1). The last aliquot was centrifuged to separate the spermatozoa and seminal plasma. The sperm pellet obtained was resuspended, and transferred to a tube with culture medium which was incubated at an angle of 45° for 30min at 37°C (SU2). Both aliquots SLC and P showed higher proportions of progressive motility and plasma membrane functionality (p≤0.05) than raw semen. There were no significant differences (p>0.05) in sperm viability and in normal spermatozoa between raw semen and treatments. Nevertheless, only SLC did not have a significant increase of bent tails. In conclusion SLC centrifugation would be the method of choice for selecting llama spermatozoa.

Changes in bull sperm kinematics after single layer centrifugation.

The objective of this study was to evaluate bull sperm kinematics after centrifugation through a single layer of a colloid [Single Layer Centrifugation (SLC)]. Ejaculates from 20 bulls were extended and stored at 4-6°C for 24 h during transport to the laboratory for SLC through Androcoll-B, followed by measurement of sperm kinematics in all samples. Total motility (86% and 88% for uncentrifuged and SLC samples, respectively) and progressive motility (84% for both the groups) were similar (p > 0.05). In contrast, straightness (STR) (0.65 vs 0.69), linearity (LIN) (0.32 vs 0.35) and beat cross frequency (BCF) (22.3 vs 23.6 Hz) were significantly higher in the SLC-selected samples than in the uncentrifuged samples, whereas velocity of the average path (VAP) (95 vs 90 µm/s), curvilinear velocity (VCL) (192 vs 180 µm/s), amplitude of lateral head deviation (ALH) (7 µm vs 6.5 µm) and hypermotility (49% vs 38%) were significantly decreased. The kinematics of the samples with the poorest motility was improved most by SLC. In conclusion, even though SLC had no direct effect on total and progressive motility, it appeared to have a positive influence on several other kinematic parameters that may be important for fertilization after artificial insemination.

CDC-42 controls early cell polarity and spindle orientation in C. elegans.

BACKGROUND: Generation of asymmetry in the one-cell embryo of C. elegans establishes the anterior--posterior axis (A-P), and is necessary for the proper identity of early blastomeres. Conserved PAR proteins are asymmetrically distributed and are required for the generation of this early asymmetry. The small G protein Cdc42 is a key regulator of polarity in other systems, and recently it has been shown to interact with the mammalian homolog of PAR-6. The function of Cdc42 in C. elegans had not yet been investigated, however. RESULTS: Here, we show that C. elegans cdc-42 plays an essential role in the polarity of the one-cell embryo and the proper localization of PAR proteins. Inhibition of cdc-42 using RNA interference results in embryos with a phenotype that is nearly identical to par-3, par-6, and pkc-3 mutants, and asymmetric localization of these and other PAR proteins is lost. We further show that C. elegans CDC-42 physically interacts with PAR-6 in a yeast two-hybrid system, consistent with data on the interaction of human homologs. CONCLUSIONS: Our results show that CDC-42 acts in concert with the PAR proteins to control the polarity of the C. elegans embryo, and provide evidence that the interaction of CDC-42 and the PAR-3/PAR-6/PKC-3 complex has been evolutionarily conserved as a functional unit.

Scarred patterns in surface waves.

Surface wave patterns are investigated experimentally in a system geometry that has become a paradigm of quantum chaos: the stadium billiard. Linear waves in bounded geometries for which classical ray trajectories are chaotic are known to give rise to scarred patterns. Here, we utilize parametrically forced surface waves (Faraday waves), which become progressively nonlinear beyond the wave instability threshold, to investigate the subtle interplay between boundaries and nonlinearity. Only a subset (three main types) of the computed linear modes of the stadium are observed in a systematic scan. These correspond to modes in which the wave amplitudes are strongly enhanced along paths corresponding to certain periodic ray orbits. Many other modes are found to be suppressed, in general agreement with a prediction by Agam and Altshuler based on boundary dissipation and the Lyapunov exponent of the associated orbit. Spatially asymmetric or disordered (but time-independent) patterns are also found even near onset. As the driving acceleration is increased, the time-independent scarred patterns persist, but in some cases transitions between modes are noted. The onset of spatiotemporal chaos at higher forcing amplitude often involves a nonperiodic oscillation between spatially ordered and disordered states. We characterize this phenomenon using the concept of pattern entropy. The rate of change of the patterns is found to be reduced as the state passes temporarily near the ordered configurations of lower entropy. We also report complex but highly symmetric (time-independent) patterns far above onset in the regime that is normally chaotic.

Inducible immunity to Trichomonas vaginalis in a mouse model of vaginal infection.

We studied the protective effect of subcutaneous immunization with Trichomonas vaginalis in a mouse model of vaginal infection. BALB/c mice were immunized with various doses of T. vaginalis (4.5 x 10(5), 9 x 10(6), and 1 x 10(8) organisms per ml) suspended in Freund's complete adjuvant 56 days prior to vaginal infection and were given booster injections of the same doses of T. vaginalis in Freund's incomplete adjuvant 4 weeks later. Control mice were immunized and given booster injections of phosphate-buffered saline suspended in Freund's complete and incomplete adjuvants. The mice were tail bled and vaginal washes were performed at weekly intervals for 4 weeks to determine the isolation of T. vaginalis and the serum and vaginal antibody reactivity. Mice which had been immunized and given booster immunizations had significantly fewer intravaginal infections and had increased serum and vaginal antibody responses compared with those of control mice (P < 0.01). Mice that were vaginally infected, treated with metronidazole, and then reinfected vaginally did not develop protective immunity. Subcutaneous immunization with whole T. vaginalis organisms appears to confer protection against intravaginal challenge with T. vaginalis, protection which is not achieved as a result of prior vaginal infection.