Invited review: Current state of genetic improvement in dairy sheep

Dairy sheep have been farmed traditionally in the Mediterranean basin in southern Europe, central Europe, eastern Europe, and in Near East countries. Currently, dairy sheep farming systems vary from extensive to intensive according to the economic relevance of the production chain and the specific environment and breed. Modern breeding programs were conceived in the 1960s. The most efficient selection scheme for local dairy sheep breeds is based on pyramidal management of the population with the breeders of nucleus flocks at the top, where pedigree and official milk recording, artificial insemination, controlled natural mating, and breeding value estimation are carried out to generate genetic progress. The genetic progress is then transferred to the commercial flocks through artificial insemination or natural-mating rams. Increasing milk yield is still the most profitable breeding objective for several breeds. Almost all milk is used for cheese production and, consequently, milk content traits are very important. Moreover, other traits are gaining interest for selection: machine milking ability and udder morphology, resistance to diseases (mastitis, internal parasites, scrapie), and traits related to the nutritional value of milk (fatty acid composition). Current breeding programs based on the traditional quantitative approach have achieved appreciable genetic gains for milk yield. In many cases, further selection goals such as milk composition, udder morphology, somatic cell count, and scrapie resistance have been implemented. However, the possibility of including other traits of selective interest is limited by high recording costs. Also, the organizational effort needed to apply the traditional quantitative approach limits the diffusion of current selection programs outside the European Mediterranean area. In this context, the application of selection schemes assisted by molecular information, to improve either traditional dairy traits or traits costly to record, seems to be attractive in dairy sheep. At the moment, the most effective strategy seems to be the strengthening of research projects aimed at finding causal mutations along the genes affecting traits of economic importance. However, genome-wide selection seems to be unfeasible in most dairy sheep breeds.     

CURRENT STATUS OF BOVINE SPONGIFORM ENCEPHALOPATHY-A REVIEW

Bovine spongiform encephalopathy (BSE) is a fatal brain disease of cattle which is presumed to have originated from feeding meat and bone meal (MBM) manufactured from rendered scrapie-infected sheep. More than 170,000 cases of BSE have been identified in UK cattle since it was specifically diagnosed in 1986. Most are in dairy herds. This paper reviews the background of the disease, changes in rendering in the United Kingdom which appear to have led to the development of BSE in cattle, characteristics of the disease, the disease-causing agent (prions), related prion diseases in other species, the impact of genetics on prion disease, prion strains, species barriers to transmission, risk of Creutzfeld-Jakob disease to humans, inactivating prion infectivity, and tallow and gelatin production and safety. Efforts at controlling the disease in the United Kingdom and preventing the disease in the United States are also discussed.     

Prion degradation in soil: possible role of microbial enzymes stimulated by the decomposition of buried carcasses

This study is part of a European project focused on understanding the biotic and abiotic mechanisms involved in the retention and dissemination of transmissible spongiform encephalopathies (TSE) infectivity in soil in order to propose practical recommendations to limit environmental contamination. A 1-year field experiment was conducted with lamb carcasses buried in a pasture soil at three depths (25, 45, and 105 cm). Microbial community response to carcasses was monitored through the potential proteolytic activity and substrate induced respiration (SIR). Soil above carcasses and control soil exhibited low proteolytic capacity, whatever the depth of burial. Contrastingly, in soil beneath the carcasses, proteolysis was stimulated. Decomposing carcasses also stimulated SIR, i.e., microbial biomass, suggesting that proteolytic populations specifically developed on lixiviates from animal tissues. Decomposition of soft tissues occurred within 2 months at subsurface while it lasted at least 1 year at deeper depth where proteolytic activities were season-dependent. The ability of soil proteases to degrade the beta form of prion protein was shown in vitro and conditions of burial relevant to minimize the risk of prion protein dissemination are discussed.     

朊蛋白疾病及其肉食品安全的研究进展

英国疯牛病造成了人类疾病传播的潜在危害,牛羊肉等肉食品的安全备受关注。为了避免疯牛病污染的肉品进入食物链,世界多个研究小组对各类肉食品开展了基于疯牛病传播因子安全方面的研究。疯牛病患病牛肉可以直接感染人类并导致发病,阳性病牛应彻底销毁并进行无害化处理,严禁进入食物链。染病的羊肉和鹿肉属于高度危险食品,也应做销毁处理,而猪肉、马肉、兔肉和水牛肉等则相对比较安全。同时,朊蛋白疾病的感染还与人类自身朊蛋白编码基因多态性有关。在严格检疫的同时,培育抗病品种的家畜以及食用不易感的肉类食品成为未来肉食品安全的一种发展趋势。本文总结了疯牛病、痒病等可传播海绵状脑病患病动物的肉用安全性,还分析了疯牛病可能突破种间屏障而造成跨物种传播的可能性,为肉食品安全及检疫提供参考。     

Novel sequence types (STs) of Staphylococcus aureus isolates causing clinical and subclinical mastitis in flocks of sheep in the northeast of Brazil

Staphylococcus aureus is one of the most important infectious mastitis causative agents in small ruminants. In order to know the distribution of Staph. aureus strains associated with infectious mastitis in flocks of sheep in the northeast of Brazil and establish whether these clones are related to the strains distributed internationally, this study analysed the genetic diversity of Staph. aureus isolates from cases of clinical and subclinical mastitis in ewes by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). In this research, 135 ewes with mastitis from 31 sheep flocks distributed in 15 districts were examined. Staph. aureus was isolated from sheep milk in 9 (29%) out of 31 herds located in 47% of the districts surveyed. MLST analysis allowed the identification of four STs (ST750, ST1728, ST1729 and ST1730). The last three with their respective novel alleles (glp-220; pta-182 and yqil-180) were recently reported in the Staph. aureus MLST database (http://www.mlst.net). Each novel allele showed only a nucleotide different from those already described. The occurrence of CC133 (ST750 and ST1729) in this study is in agreement with other reports that only a few clones of Staph. aureus seem to be responsible for most cases of mastitis in dairy farms and that some of these clones may have broad geographic distribution. However, the prevalence of CC5 (ST1728 and ST1730)--an important group related to cases of colonization or infection in humans--differs from previous studies by its widespread occurrence and may suggest human contamination followed by selective pressures of the allelic diversifications presented for these STs.     

Short communication: Investigation of Coxiella burnetii occurrence in dairy sheep flocks by bulk-tank milk analysis and antibody level determination

To estimate the prevalence of Coxiella burnetii in the dairy sheep population from the Basque Country (northern Spain), a study was carried out combining molecular and serological techniques. First, bulk-tank milk samples from 154 flocks belonging to the Latxa Breed Farmers Association were analyzed by PCR, with 22% of flocks testing positive for C. burnetii. Then, a selection of 34 flocks (7 PCR positive and 17 negative) was investigated for the presence of serum antibodies by ELISA test on 1,011 ewes (approximately 30 ewes per flock). A total of 8.9% of the animals were seropositive, 67.6% of the flocks had at least one seropositive animal, but only in 14.7% of them was seroprevalence greater than 25%. Older ewes showed a significantly greater prevalence (17.5%) compared with yearlings (7.5%) or replacement lambs (1.5%). A marginally significant association was found between seroprevalence and PCR detection of C. burnetii in bulk-tank milk. The widespread distribution of C. burnetii in the region advocates for the implementation of Q fever control strategies and highlights the potential risk of sheep as a reservoir and infection source for other domestic and wildlife species and the human population.     

A field study on fertility and purity of sex-sorted cattle sperm

The study assessed the fertility and purity of sexed semen used for inseminating Holstein-Friesian heifers in commercial dairy herds. Sex-sorted semen from 4 proven Holstein-Friesian bulls and available under commercial conditions was used on nulliparous Holstein heifers reared on 61 dairy farms of northern Italy. Data from 536 artificial inseminations with pregnancy diagnosis and 258 calvings were analyzed using the logistic regression procedure. The effects of year and season of insemination or calving, age at insemination or calving, heifer inbreeding, and the sperm dose used for insemination on the probability of a positive pregnancy diagnosis or of the birth of a female calf, respectively, were studied. The overall pregnancy rate for sexed semen was 51% and was affected by year of insemination and bull. Heifers inseminated with sexed semen from 2 bulls had lower pregnancy rates than heifers inseminated with sexed semen from other bulls. Purity of the sexed sperm, based on the proportion of female calves, was 87% and this percentage was not affected by explanatory variables included in the logistic regression. The results demonstrate that bulls differ in terms of fertility of their sexed semen. Careful selection of the insemination sires used for sorted semen is advisable for avoiding low fertility inseminations.     

Multiple typing for the epidemiological study of the contamination of broilers with Salmonella from the hatchery to the slaughterhouse

Eighteen Belgian broiler flocks were followed from the hatchery to the slaughterhouse by a multiple typing approach (sero-, geno-, and phage types) for the investigation of the transmission of Salmonella and its subtypes. For 12 of the 18 flocks, there was no correlation between the serotypes found preharvest and those isolated from the feces in the transport crates and on the carcasses in the slaughterhouse. Serotypes found in the crates were usually also found on the carcasses. In 5 of the 10 flocks with Salmonella-positive broilers, complex contamination patterns with the involvement of different serotypes, genotypes, or both were revealed. In two of these flocks (flocks 8 and 9), the Salmonella Enteritidis contamination of the broilers could be traced to the hatchery. In flock 9, evidence was found for the acquisition, during rearing, of a megaplasmid in the Salmonella Enteritidis strain. In the other three positive flocks (flocks 6, 7, and 10), the environment and movable material (e.g., footwear) played a determining role in the infection and shedding pattern of the broilers. For flocks 6 and 7, reared consecutively in the same broiler house, a persistent Salmonella Hadar geno/phage type predominated in the preharvest period, while another Salmonella Hadar geno/phage type was found in the house or the environment but never in the broilers. Only for the above-mentioned five flocks were the same strains that were found preharvest also recovered from the carcasses, although these strains were not predominant on the carcasses, with the exception of one flock (flock 10). In conclusion, it can be said that most of the time, Salmonella strains that contaminate Belgian broiler carcasses do not predominate in the preharvest environment.     

A 100-Year Review: Reproductive technologies in dairy science

Reproductive technology revolutionized dairy production during the past century. Artificial insemination was first successfully applied to cattle in the early 1900s. The next major developments involved semen extenders, invention of the electroejaculator, progeny testing, addition of antibiotics to semen during the 1930s and 1940s, and the major discovery of sperm cryopreservation with glycerol in 1949. The 1950s and 1960s were particularly productive with the development of protocols for the superovulation of cattle with both pregnant mare serum gonadotrophin/equine chorionic gonadotrophin and FSH, the first successful bovine embryo transfer, the discovery of sperm capacitation, the birth of rabbits after in vitro fertilization, and the development of insulated liquid nitrogen tanks. Improved semen extenders and the replacement of glass ampules with plastic semen straws followed. Some of the most noteworthy developments in the 1970s included the initial successes with in vitro culture of embryos, calves born after chromosomal sexing as embryos, embryo splitting resulting in the birth of twins, and development of computer-assisted semen analysis. The 1980s brought flow cytometric separation of X- and Y-bearing sperm, in vitro fertilization leading to the birth of live calves, clones produced by nuclear transfer from embryonic cells, and ovum pick-up via ultrasound-guided follicular aspiration. The 20th century ended with the birth of calves produced from AI with sexed semen, sheep and cattle clones produced by nuclear transfer from adult somatic cell nuclei, and the birth of transgenic cloned calves. The 21st century has seen the introduction of perhaps the most powerful biotechnology since the development of artificial insemination and cryopreservation. Quick, inexpensive genomic analysis via the use of single nucleotide polymorphism genotyping chips is revolutionizing the cattle breeding industry. Now, with the introduction of genome editing technology, the changes are becoming almost too rapid to fully digest.     

Genetic parameters of semen quality traits and genetic correlations with service sire nonreturn rate in Nordic Holstein bulls

Despite the importance of the quality of semen used in artificial insemination to the reproductive success of dairy herds, few studies have estimated the extent of genetic variability in semen quality traits. Even fewer studies have quantified the correlation between semen quality traits and male fertility. In this study, records of 100,058 ejaculates collected from 2,885 Nordic Holstein bulls were used to estimate genetic parameters for semen quality traits, including pre- and postcryopreservation semen concentration, sperm motility and viability, ejaculate volume, and number of doses per ejaculate. Additionally, summary data on nonreturn rate (NRR) obtained from insemination of some of the bulls (n = 2,142) to cows in different parities (heifers and parities 1–3 or more) were used to estimate correlations between the semen quality traits and service sire NRR. In the study, low to moderate heritability (0.06–0.45) was estimated for semen quality traits, indicating the possibility of improving these traits through selective breeding. The study also showed moderate to high genetic and phenotypic correlations between service sire NRR and some of the semen quality traits, including sperm viability pre- and postcryopreservation, motility postcryopreservation, and sperm concentration precryopreservation, indicating the predictive values of these traits for service sire NRR. The positive moderate to high genetic correlations between semen quality and service sire NRR traits also indicated that selection for semen quality traits might be favorable for improving service sire NRR.     

Sperm survival and transport in the female reproductive tract

Fertilization failure, mostly due to absence of sperm in the oviducts, is a major cause of reproductive inefficiency of farm animals. Sperm may be transported to the oviducts of cattle and sheep within a few minutes after mating or insemination, but these sperm probably fertilize few ova. Slower transport, with establishment of sperm populations in each segment of the reproductive tract, requires a few to several hours. In swine, sperm capable of fertilizing ova reach the oviducts in less than 1 h. Smooth muscle contractions of the reproductive tract, ciliary beats, fluid currents, and flagellar activity of sperm are primary mechanisms of sperm transport. Sperm become hyperactive in the oviducts in association with capacitation. Most sperm in an inseminate drain from the female reproductive tract within a few minutes or hours after insemination; remaining sperm are removed from the tract by slower drainage or phagocytosis. Sperm survival and transport in estrous ewes is reduced drastically by pastures with high estrogen content and by regulating estrus with progestogen or prostaglandin F2 alpha. The cervix is the initial site of inhibition of sperm transport in ewes, and endocrine imbalances probably are the basis of inhibition. Sperm transport problems generally are associated with immobilization and death of sperm in the uterus and anterior segments of the cervix within 2 h after mating. After gilts are inseminated with frozen-thawed semen, relatively few sperm are retained in the reproductive tract, apparently accounting for lowered fertilization rates. Sperm transport has been improved by adding to semen or administering to females such compounds as prostaglandin F2 alpha, oxytocin, estradiol, phenylephrine, or ergonovine. Estradiol, prostaglandin F2 alpha, phenylephrine, and ergonovine administered to rabbits at insemination each increased fertilization rates.     

Contamination of carcasses with Salmonella during poultry slaughter

Successively slaughtered poultry flocks were sampled for Salmonella to study the relationship between gastrointestinal colonization of the birds and contamination of the carcasses after slaughter. Samples from 56 broiler flocks and 16 spent layer and breeder flocks were collected in six slaughterhouses. Salmonella isolates were serotyped and further characterized by pulsed-field gel electrophoresis (PFGE). Although only 7 (13%) broiler flocks were colonized with Salmonella at slaughter, carcasses of 31 (55%) broiler flocks were contaminated after slaughter. Concerning the layer and breeder flocks, 11 (69%) flocks were colonized in the gastrointestinal tract, but after slaughter, carcasses of all flocks were contaminated. The Salmonella status determined at the farm did not always correlate to the status at slaughter. On the other hand, the slaughter of Salmonella-colonized flocks did not always result in the contamination of the carcasses with the same PFGE types isolated from the gastrointestinal tract. When only uncolonized flocks were slaughtered, the carcasses of flocks were on some occasions still contaminated with Salmonella. This indicates possible cross-contamination from the slaughter equipment or transport crates. These observations show that it is difficult to reach the benefits of logistic slaughter in commercial poultry slaughterhouses.     

Border disease virus seroprevalence correlates to antibodies in bulk-tank milk and reproductive performance of dairy sheep flocks

There is a great need to establish effective tools to control border disease virus (BDV) in European dairy sheep flocks. Hence, our main aim was to investigate the accuracy of analyzing anti-BDV antibodies in bulk-tank milk (BTM) in detecting the real BDV seroprevalence in dairy sheep flocks. Furthermore, the relevance of BDV to reproductive performance of dairy sheep flocks prompted us to search for the association between BDV seroprevalence and reproductive parameters. For these purposes, 34 flocks were selected based on different percentages of antibody inhibition (AIP) values in BTM as estimated by ELISA. Serum samples from 10 replacement lambs older than 6 mo, 10 ewes 1 to 2 yr old, and 10 ewes >2 yr old were collected and analyzed for the presence of anti-BDV antibodies by ELISA. A negative relationship between BDV AIP in BTM and within-flock seroprevalence was observed. Flocks with a high AIP (>80%) had an average of 2.5% seropositive animals; flocks with a moderate AIP (46-79%) had 11.4% seropositive animals; and finally, flocks with an AIP ≤45% showed a high flock seroprevalence (57.2%). Ten out of 34 flocks showed a high BDV seroprevalence in lambs, suggesting the presence of persistently infected animals in the flock. The observed AIP values in BTM from these likely BDV-infected flocks were indicative of a high seroprevalence. The analysis of reproductive-parameters data collected from these flocks showed no differences in fertility or prolificacy in relation to BDV circulation rates. Nonetheless, lamb mortality was significantly greater in flocks with low-moderate seroprevalence (10-30%), probably as a result of a first-time contact with BDV of previously naïve ewes. These findings suggest that testing of BTM samples may be useful in inferring the BDV seroprevalence in a flock.     

Impact of site of inseminate deposition and environmental factors that influence reproduction of dairy cattle

Uterine body and cornual inseminations (n = 2127) were evaluated over a 3-yr period in Holstein and Jersey cattle. For cornual insemination one-half of each semen dose was deposited approximately 2.5 cm into each uterine horn. Conception rate was lower for cervical insemination (39.4%) than uterine body (48.1%) or cornual (49.3%) inseminations. Pregnancy site distribution was equal for both insemination techniques but cervical insemination resulted in 60% right horn pregnancies. Primary housing, service number, inseminator, lactation number, and sire affected conception rate. Older cows were least fertile (31.4%), second service conception rate was lowest (42.6%), and barn housed cattle had a 39.7% conception rate. Days open was affected by primary housing, service number, sire, site of semen placement, and twinning. Twinning increased days open by 10 d. Optimum time for insemination of lactating cows was between 6 and 12 h after the initial observation of estrus. From this study we conclude that shallow cornual insemination is as effective as uterine body insemination, and conception rate is optimized when estrus is positively assessed.     

STRATEGIES FOR THE CONTROL OF BSE AND SCRAPIE

Transmissible Spongiform Encephalopathies, or Prion diseases, are fatal neurodegenerative diseases that affect both humans and animals. Examples include Creutzfeldt-Jakob disease (CJD) in humans, Bovine Spongiform Encephalopathy (BSE) in cattle and scrapie in sheep and goats. They are characterized by a long incubation period, ranging from months to years, and a variable length clinical course, both determined by the particular infection and the species involved. Some 170,000 cattle in the UK have been diagnosed with BSE and have been destroyed. In the rest of Europe there have also been cases of BSE, though the numbers involved are much lower. The recent potential link between BSE and a new variant form of CJD has highlighted the importance of controlling both BSE and scrapie in Ireland. Control of the disease in both cattle and sheep is complicated by the long incubation period, the lack of suitable tests and the general lack of detailed information in relation to the biology of the disease. Factors that must be considered in designing an effective control strategy include (1) better understanding of the epidemiology of the disease, (2) developing rapid and reliable tests for the disease, (3) introducing a suitable surveillance system for both BSE and scrapie, (4) minimizing the consequences for farmers that have BSE-infected animals, and (5) implementing and monitoring legislation to prohibit the spread of potentially infectious material. Each of these areas is discussed and it is proposed that the introduction of best available technology in both the diagnosis and surveillance of the national herd could form the basis for an effective program against animal prion diseases.     

Use of insemination data for joint evaluation of male and female fertility in predominantly seasonal-calving dairy herds

Conception in dairy cattle is influenced by the fertility of the cow and the bull and their interaction. Despite genetic selection for female fertility in many countries, selection for male fertility is largely not practiced. The primary objective of this study was to quantify variation in male and female fertility using insemination data from predominantly seasonal-calving herds. Nonreturn rate (NRR) was derived by coding each insemination as successful (1) or failed (0) based on a minimum of at least 25 d. The NRR was treated as a trait of the bull with semen (male fertility) and the cow that is mated (female fertility). The data (805,463 cows that mated to 5,776 bulls) were used to estimate parameters using either models that only included bulls with mating data or models that fitted the genetic and permanent environmental (PE) effects of bulls and cows simultaneously. We also evaluated whether fitting genetic and PE effects of bulls as one term is better for ranking bulls based on NRR compared with a model that ignored genetic effect. The age of cows that were mated, age of the bulls with semen data, season of mating, breed of cow that mated, inbreeding of cows and bulls, and days from calving to mating date were found to have a significant effect on NRR. Only about 3% of the total variance was explained by the random effects in the model, despite fitting the genetic and PE effects of the bull and cow. The 2 components of fertility (male and fertility) were not correlated. The heritability of male fertility was low (0.001 to 0.008), and that of female fertility was also low (~0.016). The highest heritability estimate for male fertility was obtained from the model that fitted the additive genetic relationship matrix and PE component of the bull as one term. When this model was used to calculate bull solutions, the difference between bulls with at least 100 inseminations was up to 19.2% units (−9.6 to 9.6%). Bull solutions from this model were compared with bull solutions that were predicted fitting bull effects ignoring pedigree. Bull solutions that were obtained considering pedigree had (1) the highest accuracy of prediction when early insemination was used to predict yet-to-be observed insemination data of bulls, and (2) improved model stability (i.e., a higher correlation between bull solutions from 2 randomly split herds) compared with the model which fitted bull with no pedigree. For practical purposes, the model that fitted genetic and PE effect as one term can provide more accurate semen fertility values for bulls than the model without genetic effect. To conclude, insemination data from predominantly seasonal-calving herds can be used to quantify variability between bulls for male fertility, which makes their ranking on NRR feasible. Potentially this information can be used for monitoring bulls and can supplement efforts to improve herd fertility by avoiding or minimizing the use of semen from subfertile bulls.     

Evaluation of bulk tank milk PCR and bulk tank milk modified ELISA tests for the detection of paratuberculosis at the herd level in goat and sheep dairies in Ontario,Canada

Early identification of dairy goat herds and dairy sheep flocks infected with Mycobacterium avium ssp. paratuberculosis is important for controlling this infection and minimizing economic losses. The objective of this study was to evaluate 2 bulk tank milk (BTM) paratuberculosis tests (PCR and modified ELISA) as potential herd-level tests. These tests were compared with the results obtained from testing 20 randomly selected lactating animals per farm (>2 yr) with an individual animal test (fecal culture, fecal PCR, serum ELISA, and milk ELISA). The study was conducted using 29 dairy goat herds and 21 dairy sheep flocks in Ontario, Canada, visited between October 2010 and August 2011. The sensitivity of the BTM PCR was poor in both the dairy goat herds (0.0%) and dairy sheep flocks (25.0%), but exhibited 100% specificity in both species. In comparison, the BTM modified ELISA demonstrated higher sensitivity. In goats, sensitivity ranged from 33.3 to 34.8% when fecal culture and PCR were the reference tests, respectively (specificities were both 100%), and 71.4 to 87.5% when the milk and serum ELISA, respectively, were the reference tests (specificities were 86.4 and 95.2%). The BTM modified ELISA in dairy sheep demonstrated comparable sensitivities, but lower specificities. When fecal culture and PCR were the reference test, sensitivities were 50.0 and 46.7%, respectively (specificities were 77.8 and 83.3%). The sensitivities when the milk and serum ELISA were the reference tests were 87.5 and 72.7%, respectively (specificities were 92.3 and 100%). Fecal PCR was the only individual animal test to identify significantly more farms as positive than the BTM PCR and modified ELISA test in both species. Therefore, whereas the BTM modified ELISA may provide an organization or control program with a high level of confidence that a BTM-positive farm is actually positive (high positive predictive value), if a producer wishes to increase the odds that a positive farm will test positive, so as not to miss an infection, then sampling and testing 20 animals with fecal PCR will better meet that objective.     

Adsorption of pathogenic prion protein to quartz sand

Management responses to prion diseases of cattle, deer, and elk create a significant need for safe and effective disposal of infected carcasses and other materials. Furthermore, soil may contribute to the horizontal transmission of sheep scrapie and cervid chronic wasting disease by serving as an environmental reservoirforthe infectious agent. As an initial step toward understanding prion mobility in porous materials such as soil and landfilled waste, the influence of pH and ionic strength (l) on pathogenic prion protein (PrPsc) properties (viz. aggregation state and zeta-potential) and adsorption to quartz sand was investigated. The apparent average isoelectric point of PrPsc aggregates was 4.6. PrPsc aggregate size was largest between pH 4 and 6, and increased with increasing l at pH 7. Adsorption to quartz sand was maximal near the apparent isoelectric point of PrPsc aggregates and decreased as pH either declined or increased. PrPsc adsorption increased as suspension l increased, and reached an apparent plateau at l approximately 0.1 M. While trends with pH and l in PrPsc attachment to quartz surfaces were consistent with predictions based on Born-DLVO theory, non-DLVO forces appeared to contribute to adsorption at pH 7 and 9 (l = 10 mM). Our findings suggest that disposal strategies that elevate pH (e.g., burial in lime or fly ash), may increase PrPsc mobility. Similarly, PrPsc mobility may increase as a landfill ages, due to increases in pH and decreases in l of the leachate.     

Semen-induced luteal phase and identification of a LH surge in the koala (Phascolarctos cinereus)

The koala ovulates in response to mating. The purpose of this study was to document the LH surge induced by copulation and to investigate the potential roles of mechanical stimulation of the urogenital sinus and deposition of semen in induction of the luteal phase. In experiment 1, serial blood samples from four koalas that underwent normal mating showed elevated concentrations of LH approximately 24-32 h post-coitus. There was no corresponding elevation in LH in koalas (n=4) that were exposed to the presence of a male but received no physical contact. In experiment 2, koalas on day 2 of oestrus were exposed to one of the following treatments (n=9 per group): artificial insemination with 1 ml 0.9% sterile saline (control group), insemination with 1 ml koala semen, stimulation of the urogenital sinus with a purpose built glass rod (designed to mimic the action of the penis during natural mating) and urogenital stimulation with the glass rod followed by insemination of 1 ml koala semen. Confirmation of a luteal phase was based on evidence of a prolonged return to oestrus, parturition and/or elevated progesterone concentrations. Insemination of saline (0/9) and urogenital stimulation (0/9) failed to induce a luteal phase. Insemination of semen without glass rod stimulation resulted in a luteal phase in 4/9 koalas, three of which gave birth. Insemination of semen in combination with urogenital stimulation produced a luteal phase in 7/9 koalas, four of which gave birth. Semen had a significant effect on induction of the koala luteal phase (P <0.001) but glass rod stimulation had no such effect (P=0.335). It was concluded that semen must be involved in the induction of a luteal phase in the koala. The results presented in this study will serve to improve optimal timing and induction of ovulation for artificial insemination in the koala.