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.     

Digesta retention and fibre digestion in brushtail possums, ringtail possums and rabbits

1. Mean retention times (MRTs) of fluid (marked with Co-EDTA), fine particles (mordanted with Yb) and large particles (mordanted with Cr) were measured in brushtail possums (Trichosurus vulpecula), ringtail possums (Pseudocheirus peregrinus) and laboratory rabbits fed semipurified diets. 2. In brushtail possums there were no significant differences in MRT among the three digesta markers. 3. In ringtail possums MRTs of the fluid and fine particle markers were approximately twice that of the large particle marker, indicative of selective retention of both fluid and fine particles in the caecum. 4. In the rabbit MRT of fine particles was also greater than that of large particles, again indicative of selective retention of fine particles in the caecum. 5. Fibre digestibility was greater in the rabbits than in the ringtail possums, and greater for neutral-detergent fibre (including agar) but less for acid-detergent fibre in the rabbits than in the brushtails. Differences in fibre digestibility between brushtails and rabbits were explained by differences in patterns of digesta flow. However, the higher digestibilities of fibre in the rabbits than in the ringtail possums could not be explained on a similar basis.     

Genetics of fibre production and fleece characteristics in small ruminants, Angora rabbit and South American camelids

This paper reviews genetics of fibre production and fleece characteristics in small ruminants, Angora rabbit and South American camelids with a special distinction between single-coated (SC) and double-coated (DC) species. Considering the biology of fibre production, there are variations in coat composition and structure, fibre growth pattern and fibre structure and quality between these two main kinds of fibre-producing animals. In SC species, all fibres are nearly similar in dimensions and are produced from individual follicles that have a very long period, essentially permanent, of active growth without a synchronous phase of rest between follicles. In contrast, in DC species the fleece comprises a coarse outer coat and a fine inner coat with variations of coat composition and structure, and fibre growth pattern according to the season with a well-defined duration of fibre growth. Genetic basis of hair growth pattern, coat composition and fibre structure are different between species. In small ruminants, these coat characters are additive and because of several genes whereas in rabbit, several autosomal recessive genes determine fibre growth, coat composition and structure. In alpaca, the fleece type (Suri or Huacaya) is determined by a single dominant gene. This paper also reviews genetic parameters of fibre production traits in Angora goat, Angora rabbit and alpaca in which many aspects of the genetic basis of fibre production are analogous. There are many traits controlling both fibre quality and fibre quantity, and most of these traits tend to be moderately to strongly inherited so that a rapid genetic progress in any traits is possible and indeed has been achieved. However, there are differences in breeding programmes. In DC Angora rabbit, selection for one single trait, the easy measurable total fleece weight has general beneficial effects on fleece quality. However, because of antagonistic relations between qualitative and quantitative traits in SC species, achieving this goal requires a multi-trait selection index approach. Gene mapping studies have recently identified several putative quantitative trait loci and major genes affecting fibre and fleece characteristics in sheep, goat and rabbit are reviewed. The whole genome sequence of sheep and rabbit will be available in the near future and the use of high-density single nucleotide polymorphism chip will allow fine mapping and dissection of the genetic basis of many production traits including fibre production and fleece characteristics. The application of these techniques will thus contribute to improving the efficiency, profitability and sustainability of small ruminant and rabbit fibre production.     

Excretion patterns of fluid and different sized particle passage markers in banteng (Bos javanicus) and pygmy hippopotamus (Hexaprotodon liberiensis): two functionally different foregut fermenters

Processing of ingesta particles plays a crucial role in the digestive physiology of herbivores. In the ruminant forestomach different sized particles are stratified into a small and a large particle fraction and only the latter is regurgitated and remasticated to smaller, easier-to-digest particles. In contrast, it has been suggested that in non-ruminating foregut fermenters, such as hippopotamuses, larger particles should be selectively excreted since they tend to be digested at a slower rate and hence can be considered intake-limiting bulk. In our study we determined the mean retention time (MRT) of fluids and different sized particles (2 mm and 10 mm) in six pygmy hippos (Hexaprotodon liberiensis) and six banteng (Bos javanicus) on a diet of fresh grass at two intake levels. We used cobalt ethylendiamintetraacetate (Co-EDTA) as fluid and chromium (Cr)-mordanted fibre (2 mm) and cerium (Ce)-mordanted fibre (10 mm) as particle markers, mixed in the food. Average total tract MRT for fluid, small and large particles at the high intake level was 32, 76 and 73 h in pygmy hippos and 25, 56 and 60 h in banteng, and at the low intake level 39, 109, and 105 h in pygmy hippos and 22, 51 and 58 h in banteng, respectively. In accordance with the prediction, large particles moved faster than, or as fast as the small particles, through the gut of pygmy hippos. In contrast, large particles were excreted slower than the small particles in the ruminant of this study, the banteng. Pygmy hippos had longer retention times than the banteng, which probably compensate for the less efficient particle size reduction. Although the results were not as distinct as expected, most likely due to the fact that ingestive mastication of the larger particle marker could not be prevented, they confirm our hypothesis of a functional difference in selective particle retention between ruminating and non-ruminating foregut fermenters.     

Sexual behavior in ewes and other domestic ruminants

Similarities as well as differences across species in the control of sexual behavior are helping to fully understand the subtle relations between physiology and eco-ethological constraints and how the brain integrates such information. We will illustrate this with sexual behavior in domestic ruminants and especially ewes. Females of these species like humans, but unlike rodents, have a long luteal phase. A prolonged exposure to progesterone (Pg) before the preovulatory estradiol rise is necessary for estrous behavior to be displayed. Estradiol action and receptor localization is very similar to that observed in other species. But not too surprisingly, the role of Pg is rather different with a priming effect not observed in rodents. However, as in rodents, Pg also has an inhibitory effect, is necessary for the display of proceptivity and is responsible for the timing of the different periovulatory events. These steroids act on the central nervous system in similar areas across mammalian species to regulate estrous behavior. Steroid fluctuations during the estrous cycle cause changes in catecholaminergic activity in the hypothalamus. Interestingly, these neurotransmitters seem to have very similar effects in ewes and rats as illustrated by the norepinephrine rise after male-female interactions observed in both species. Similar comparisons can be made regarding the action of some neuropeptides, including oxytocin and GnRH, and more integrative processes like sexual differentiation and modulation of reproduction by social interactions. Data on sheep, goats and cows will be compared with those of rodents.     

The maximum attainable body size of herbivorous mammals: morphophysiological constraints on foregut,and adaptations of hindgut fermenters

An oft-cited nutritional advantage of large body size is that larger animals have lower relative energy requirements and that, due to their increased gastrointestinal tract (GIT) capacity, they achieve longer ingesta passage rates, which allows them to use forage of lower quality. However, the fermentation of plant material cannot be optimized endlessly; there is a time when plant fibre is totally fermented, and another when energy losses due to methanogenic bacteria become punitive. Therefore, very large herbivores would need to evolve adaptations for a comparative acceleration of ingesta passage. To our knowledge, this phenomenon has not been emphasized in the literature to date. We propose that, among the extant herbivores, elephants, with their comparatively fast passage rate and low digestibility coefficients, are indicators of a trend that allowed even larger hindgut fermenting mammals to exist. The limited existing anatomical data on large hindgut fermenters suggests that both a relative shortening of the GIT, an increase in GIT diameter, and a reduced caecum might contribute to relatively faster ingesta passage; however, more anatomical data is needed to verify these hypotheses. The digestive physiology of large foregut fermenters presents a unique problem: ruminant-and nonruminant-forestomachs were designed to delay ingesta passage, and they limit food intake as a side effect. Therefore, with increasing body size and increasing absolute energy requirements, their relative capacity has to increase in order to compensate for this intake limitation. It seems that the foregut fermenting ungulates did not evolve species in which the intake-limiting effect of the foregut could be reduced, e.g. by special bypass structures, and hence this digestive model imposed an intrinsic body size limit. This limit will be lower the more the natural diet enhances the ingesta retention and hence the intake-limiting effect. Therefore, due to the mechanical characteristics of grass, grazing ruminants cannot become as big as the largest browsing ruminant. Ruminants are not absent from the very large body size classes because their digestive physiology offers no particular advantage, but because their digestive physiology itself intrinsically imposes a body size limit. We suggest that the decreasing ability for colonic water absorption in large grazing ruminants and the largest extant foregut fermenter, the hippopotamus, are an indication of this limit, and are the outcome of the competition of organs for the available space within the abdominal cavity. Our hypotheses are supported by the fossil record on extinct ruminant/tylopod species which did not, with the possible exception of the Sivatheriinae, surpass extant species in maximum body size. In contrast to foregut fermentation, the GIT design of hindgut fermenters allows adaptations for relative passage acceleration, which explains why very large extinct mammalian herbivores are thought to have been hindgut fermenters.     

Digesta retention in the gastro-intestinal tract of the orang utan (Pongo pygmaeus)

The diet of the orang utanPongo pygmaeus consists of fruit, leaves, communal insects, and bark, and contains appreciable amounts of non-starch polysaccharides. These complex carbohydrates require microbial fermentation before they can be used as an energy source by the orang utans. The gastrointestinal tract ofP. pygmaeus consists of a simple or unipartite stomach, a relatively long small intestine, and a complex haustrated caecum and colon. This morphology suggests that the capacious proximal colon is the principal site of digesta retention and fermentation of non-starch polysaccharides. We measured several parameters of digesta retention by giving three captive adultP. pygmaeus a pulse dose of inert markers specific for the solute and particulate phases of the digesta and collected their faeces at regular intervals over 192–338 hours. Transit times (times of first appearance of the markers in the faeces) and mean retention times (MRT) were long, consistent with a large complex gastro-intestinal tract. MRTs for the particulate marker were longer (p=0.032) than for the solute marker, indicative of selective retention of large particulate digesta. These results are consistent with the patterns of marker excretion in other mammals that use the digestive strategy of colon fermentation.     

Digesta retention time and recovery rates of ants and termites in Chinese pangolins (Manis pentadactyla)

Pangolins are myrmecophagous mammals whose biology and ecology remain poorly studied. Termite mandibles and ant head capsules are the two primary remains found in pangolin feces. Determining the retention time of insect cuticles is important for understanding the digestive physiology of pangolins, while determining the recovery rate of termites and ants in feces is required to estimate the number of these prey items that are consumed by pangolins. In this study, the authors conducted feeding trials with captive Chinese pangolins (Manis pentadactyla). Sixty grams of the fungus-growing termite Odontotermes formosanus (18,816 individuals) and 15–20 g of the yellow crazy ant Anoplolepis gracilipes (14,400–19,200 individuals) were fed to each pangolin. After feeding, pangolin feces were collected daily for 1 week. The authors also assessed the accuracy of using chromium (III) oxide powder (Cr₂O₃) as a proxy for determining gut passage time, as has been done in previous studies. The results showed that remaining termite mandibles and ant head capsules in feces peaked at 66 and 90 hr after feeding and their recovery rates were 0.35 ± 0.10 and 0.65 ± 0.04, respectively. In both feeding trials, the retention time of Cr₂O₃ was much shorter than that of the termite mandibles and ant head capsules, indicating that Cr₂O₃ is not an appropriate indicator for estimating food retention time of myrmecophagous animals. Our results revealed that the ant head capsules were preserved better in feces compared with the termite mandibles, suggesting that termites may be considerably underestimated in the feces of wild pangolins.     

Rubisco activity in guard cells compared with the solute requirement for stomatal opening

We investigated whether the reductive pentose phosphate path in guard cells of Pisum sativum had the capacity to contribute significantly to the production of osmotica during stomatal opening in the light. Amounts of ribulose 1,5-bisphophate carboxylase/oxygenase (Rubisco) were determined by the [¹⁴C]carboxyarabinitol bisphosphate assay. A guard cell contained about 1.2 and a mesophyll cell about 324 picograms of the enzyme; the ratio was 1:270. The specific activities of Rubisco in guard cells and in mesophyll cells were equal; there was no indication of a specific inhibitor of Rubisco in guard cells. Rubisco activity was 115 femtomol per guard-cell protoplast and hour. This value was different from zero with a probability of 0.99. After exposure of guard-cell protoplasts to ¹⁴CO₂ for 2 seconds in the light, about one-half of the radioactivity was in phosphorylated compounds and <10% in malate. Guard cells in epidermal strips produced a different labelling pattern; in the light, <10% of the label was in phosphorylated compounds and about 60% in malate. The rate of solute accumulation in intact guard cells was estimated to have been 900 femto-osmol per cell and hour. If Rubisco operated at full capacity in guard cells, and hexoses were produced as osmotica, solutes could be supplied at a rate of 19 femto-osmol per cell and hour, which would constitute 2% of the estimated requirement. The capacity of guard-cell Rubisco to meet the solute requirement for stomatal opening in leaves of Pisum sativum is insignificant.     

Salt requirements for membrane transport and solute retention in some moderate halophiles

Abstract Many species of bacteria isolated from saline environments require Na⁺ specifically for membrane transport. Transport occurs by a Na⁺ symport process energized by an electrochemical gradient of Na⁺ ions. The gradient at neutral pH appears to be produced by a primary electrogenic extrusion of protons coupled to a secondary, outwardly directed Na⁺ pump, a Na⁺/proton antiporter. At alkaline pH Vibrio alginolyticus may also produce the gradient by an energy-dependent primary extrusion of Na⁺ ions. Alteromonas haloplanktis and Vibrio costicola require salts in the medium to retain intracellular solutes. For A. haloplanktis the effects of the salts are primarily osmotic. For V. costicola , only NaCl is effective in retaining solutes and Na⁺ is required by this organism to maintain the membrane potential. In Escherichia coli a single substitution in the nucleotide sequence of the gene coding for the melibiose transport protein changed the cation specificity of the transport system. The possible ecological significance of this finding has been considered.     

Alcohol consumption patterns of shiftworkers compared with dayworkers

The detrimental effects of excessive alcohol consumption are well documented. There is some evidence that shiftworkers consume more alcohol than dayworkers as a sleep aid to compensate for sleep difficulties associated with work schedules. This study investigated drinking patterns between shiftworkers and dayworkers using the 2006 and 2007 waves from the Household Income and Labour Dynamics Survey. A subset of workers who were not in full-time study and had a single job were selected; participants who did not drink alcohol (n = 2090) were excluded. Using the 2001 Australian Government alcohol guidelines, alcohol consumption for risk of short-term harm (7+ standard drinks for men, 5+ for women) was investigated. The number of workers who drank alcohol "nearly every day" or "every day" was also examined. Some 13% of shiftworkers and 10% of those on standard schedules reported consuming alcohol at levels risky for short-term harm. Having a child less than 17 yrs (odds ratio [OR]?=?.39, 95% confidence interval [CI]?=?.22-.69), higher job demands (OR =?.71, 95% CI =?.58-.86), being female (OR =?.45, 95% CI=. 26-.79), and being older (OR =?.89, 95% CI =?.87-.92) significantly reduced, whereas being a shiftworker (OR = 2.10, 95% CI = 1.08-4.12) significantly increased, the odds of drinking alcohol in short-term risky levels. Nearly 10% of shiftworkers and 8% of those on standard schedules reported consuming alcohol in short-term risky levels at least weekly. Having a child less than 17 yrs (OR =?.40, 95% CI =?.22-.74), higher job demands (OR =?.69, 95% CI =?.56-.86), being female (OR =?.28, 95% CI =?.15-.53), and being older (OR =?.92, 95% CI =?.89-.94) were associated with a significant reduction in the odds of consuming alcohol at risky levels at least weekly. Being a shiftworker was not associated with a significant increase in the odds of consuming alcohol at such risky levels at least weekly, but a trend was evident (OR = 1.47, 95% CI =?.73-3.00). Some 13.5% of shiftworkers and 21% of those on standard schedules reported consuming alcohol in any amount "near daily" or "daily." Working more hours than preferred (OR = 1.80, 95% CI = 1.12-2.89) and being older (OR = 1.10, 95% CI = 1.07-1.13) were associated with a significant increase, and being female (OR =?.18, 95% CI =?.10-.33), and being a shiftworker (OR =?.20, 95% CI =?.09-.45) were associated with a significant decrease in the odds of consuming alcohol "daily" or "near daily." Overall, the results suggest that shiftworkers may be more likely to consume alcohol at levels considered to be risky for health in the short term. In contrast, they appear less likely to drink alcohol daily. This pattern is suggestive of "binge drinking" behavior.     

The fate of sulphadimethoxine in primates compared with other species

1. The metabolism of sulphadimethoxine (2,4-dimethoxy-6-sulphanilamidopyrimidine) was examined in nine species of primates and nine species of non-primates. 2. The main metabolite of the drug in the urine in man, rhesus monkey, baboon, squirrel monkey, capuchin, bushbaby, slow loris and tree shrew was sulphadimethoxine N(1)-glucuronide. In the green monkey, although the main metabolite was N(4)-acetylsulphadimethoxine, the N(1)-glucuronide was also a major metabolite. 3. In the dog, rat, mouse, guinea pig, Indian fruit bat and hen the N(1)-glucuronide was a minor metabolite in the urine, whereas in the cat, ferret and rabbit this glucuronide was not found in the urine. 4. All the species examined except the dog excreted some N(4)-acetylsulphadimethoxine, which was the major metabolite in the green monkey, rabbit and guinea pig. 5. In the tree shrew, a doubtful primate, N(1)-glucuronide formation was similar to that in the other primates. 6. It is suggested that the slow excretion of the drug by the rat may be due partly to strong binding of the drug to tissue proteins and that the strength of binding may vary with species. 7. In the rat the amount of N(1)-glucuronide found in the urine is not a true indication of the extent of this conjugation since much more of the conjugate was found in the bile (7% of the dose) than in the urine (1%). In the rabbit, no N(1)-glucuronide was found in the bile or urine, but a small amount of sulphadimethoxine N(4)-glucuronide was found in the bile of the rat (0.5% of dose) and rabbit (0.8%).     

An assessment of the behavioral toxicity of high-energy iron particles compared to other qualities of radiation

Conditioned taste aversion was used to evaluate the behavioral toxicity of exposure to high-energy iron particles (56Fe, 600 MeV/amu) in comparison to that of gamma photons (60Co), high-energy electrons, or fission neutrons. Exposure to high-energy iron particles (5-500 cGy) produced a dose-dependent taste aversion with a maximal effect achieved with a dose of 30 cGy. Gamma photons and electrons were the least effective stimuli for producing a conditioned taste aversion, with a maximal aversion obtained only after exposure to 500 cGy, while the effectiveness of fission neutrons was intermediate to that of photons and iron particles, and a maximal aversion was obtained with a dose of 100 cGy. In the second experiment, rats with lesions of the area postrema were exposed to iron particles (30 cGy), but failed to acquire a taste aversion. The results indicate that (1) high-energy iron particles are more toxic than other qualities of radiation and (2) similar mechanisms mediate the behavioral toxicity of gamma photons and high-energy iron particles.     

Liquid circulation and mixing in tower fermenters with recirculation loop

Liquid circulation and mixing time studies were carried out in a laboratory bubble column and modified bubble columns. Liquid circulation and mixing time are both strongly dependent on the geometrical configuration of the reactor. Internal recycle loop and external recycle loop shorten the mixing time compared to the bubble column.     

Inositolphosphoceramide metabolism in Trypanosoma cruzi as compared with other trypanosomatids

Chagas disease is caused by Trypanosoma cruzi and is endemic to North, Central and South American countries. Current therapy against this disease is only partially effective and produces adverse side effects. Studies on the metabolic pathways of T. cruzi, in particular those with no equivalent in mammalian cells, might identify targets for the development of new drugs. Ceramide is metabolized to inositolphosphoceramide (IPC) in T. cruzi and other kinetoplastid protists whereas in mammals it is mainly incorporated into sphingomyelin. In T. cruzi, in contrast to Trypanosoma brucei and Leishmania spp., IPC functions as lipid anchor constituent of glycoproteins and free glycosylinositolphospholipids (GIPLs). Inhibition of IPC and GIPLs biosynthesis impairs differentiation of trypomastigotes into the intracellular amastigote forms. The gene encoding IPC synthase in T. cruzi has been identified and the enzyme has been expressed in a cell-free system. The enzyme involved in IPC degradation and the remodelases responsible for the incorporation of ceramide into free GIPLs or into the glycosylphosphatidylinositols anchoring glycoproteins, and in fatty acid modifications of these molecules of T. cruzi have been understudied. Inositolphosphoceramide metabolism and remodeling could be exploited as targets for Chagas disease chemotherapy.