Due to overpopulation of African elephants in South Africa and the consequent threat to biodiversity, the need for a method of population control has become evident. The potential use of the porcine zona pellucida (pZP) vaccine as a safe and effective means for population control is explored. While potential effects of pZP treatment on social behaviour of African elephants have been investigated, no examination of the influence of pZP vaccination on the endocrine correlates in treated females has been undertaken. The ovarian activity of freeranging, pZP-treated African elephant females was monitored non-invasively for one year duration by measuring faecal progestagen concentrations via enzyme immunoassay. Behavioural observations were recorded for comparison with progestagen concentrations and to determine any behavioural changes surrounding the pZP vaccine darting event. Each elephant under study showed progestagen concentrations rising above baseline at some period during the study indicating luteal functionality. Average progestagen concentrations were 1.61 ± 0.46 ìg/g. Within sampled females, 42.9% exhibited oestrous cycles within the range reported for African elephants, 14.3% had irregular cycles, and 42.9% did not appear to be cycling. Average oestrous cycle duration was 14.72 ± 0.85 weeks. Behavioural oestrous coincided with the onset of the luteal phase and a subsequent rise in progestagen concentrations. Focal sampling to determine activity budgets before and after the darting event revealed no significant change in behavioural activities. In the week following immunization, individual progestagen concentrations decreased significantly from overall average concentrations. Average progestagen concentrations positively correlated with rainfall and with herd dominance. No association between average individual progestagen concentrations or cyclicity status with age, lactation, or parity were detected. Earlier determination of efficacy was made indicating reproductive control was established 22 months post-treatment. Results indicate the presence of ovarian activity amongst pZP-treated female African elephants in two years following initial immunization. Further study should be aimed toward studying the long term effects of pZP vaccination on the reproductive function of female African elephants. / Dissertation (MSc)--University of Pretoria, 2010. / Production Animal Studies / unrestricted

The Bureau of Land Management (BLM) maintains a growing number of feral horses on public rangelands. With population growth rates as high as 22% annually, herds are exceeding their carrying capacity and millions of dollars are spent maintaining captured horses in holding facilities awaiting adoption. To manage the feral horse population, the BLM is seeking a contraceptive that is safe, can be remotely delivered, requires only a single administration and is effective for several years. Contraceptive strategies have been developed for feral horses that include hormone implants, chemical intrauterine devices, and immunocontraception. Porcine zona pellucida (pZP) immunocontraceptive vaccines have shown great potential for providing safe, long-term contraception in feral horses. ImmunoVaccine Technologies (Halifax, Nova Scotia, Canada) has developed a liposome encapsulated pZP formulation known as SpayVac™ (SpayVac), which after a single-dose provides multi-year contraceptive efficacy. In a continued effort to optimize the acceptability and efficacy of SpayVac, ImmunoVaccine Technologies developed alternative adjuvant preparations using either killed Mycobacterium butyricum (Modified Freund's Adjuvant; MFA) or a proprietary non-Mycobacterium based adjuvant (IVT) that are proposed to have less of the undesirable side-effects associated with Mycobacterium tuberculosis. Therefore, the objective of this research was to evaluate SpayVac in different adjuvant formulations for efficacy of contraception as measured by pZP titers and estrous cyclicity in treated mares. Domestic mares (n=28) were randomly assigned to four treatments (7 mares per treatment): adjuvant alone or saline (Control) or SpayVac vaccines in one of three adjuvant preparations: IVT or MFA in either an aqueous (MFA aq) or non-aqueous (MFA non-aq) suspension. Pre-immune blood samples were collected from each mare and mares were injected in the neck with a single injection of the Control or SpayVac. Subsequent blood samples were collected at weekly intervals for 26 weeks. Sera were analyzed for pZP titers and progesterone using ELISA. At the conclusion of the study, ovaries were recovered by ovariectomy (16 mares) or at necropsy (12 mares) for histologic analysis and collection of morphometric data and oocytes. Titers for pZP were greater (P<0.05) in IVT and MFA mares compared to Control mares and for MFA compared to IVT mares. Mares vaccinated with MFA aq had greater (P<0.05) pZP titers at 2 weeks post-injection compared to mares injected with IVT or MFA non-aq and at 3 weeks post-injection compared to mares injected with IVT. MFA non-aq mares had greater (P<0.05) pZP titers at 6 weeks post-injection compared to IVT mares and, although not significantly different, titers in MFA non-aq mares remained greater during weeks 8, 10, 14, 18 and 22 compared to IVT and MFA aq mares. Mean serum progesterone concentrations were greater (P<0.05) in Control compared to MFA non-aq mares. Mean ovarian weights, oocyte diameters, zona pellucida thicknesses and the number of horse sperm bound to oocytes recovered from vaccinated mares were greater (P<0.05) in Control mares compared to IVT and MFA mares. As judged by pZP titers and serum progesterone, these results suggest that SpayVac suspended in the MFA non-aqueous formulation exerted the greatest contraceptive effects in treated mares. This preparation of SpayVac may meet the criteria cited by the BLM for their most desirable immunocontraceptive. / Graduation date: 2012

L’efficacité de la chimiothérapie contre la croissance tumourale repose sur l'induction d'une mort des cellules tumourales dite immunogène. Les cellules tumourales mourantes agissent alors comme un vaccin thérapeutique en stimulant une réponse immunitaire anti-tumourale capable de contrôler, voire d'éliminer, les cellules cancéreuses résiduel. Les trois marqueurs principales de la mort cellulaire immunogène (MCI), sont (i) l'exposition pré-apoptotique de la calréticuline (CRT) à la surface des cellules, (ii) la sécrétion de l'ATP pendant l'apoptose qui dépend du processus d'autophagie, et (iii) le relargage post-apoptotique de la protéine non-histone de fixation à la chromatine HMGB1 (High Mobility Group B1). CRT, ATP et HMGB1 se lient respectivement à CD91, au récepteur purinergique P2RX7 (Purinergic Receptor P2X, ligand-gated ion channel 7) et au récepteur TLR4 (Toll-like Receptor 4) situés à la surface des cellules dendritiques. En retour, ces interactions initient respectivement la phagocytose des cellules mourantes, la production de l'interleukine-1β et la cross-présentation des antigènes tumouraux aux lymphocytes T.Notre laboratoire a précédemment démontré que la chimiothérapie adjuvante présente une efficacité réduite chez des patients atteints de cancers colorectaux et du sein portant un polymorphisme d'un seul nucléotide ou SNP (Single-Nucleotide Polymorphism) compromettent la fonction des gènes P2RX7 et TLR4, notamment rs3751143 (496Glu>Ala) pour le gène P2RX7 et rs4986790 (299Asp>Gly) pour le gène TLR4.Compte tenu des ces résultats que mettent en évidence la relation étroite entre l’efficacité de la chimiothérapie anticancéreuse et un système immunitaire opérationnel, nous avons décidé d'étudier l'effet de ces SNPs, soit sur la survie globale soit sur la survie sans événement, chez des patients atteints de cancer pulmonaire non à petites cellules ou NSCLC (Non-Small Cell Lung Carcinoma) et de cancers de la tête et du cou ou HNSCC (Head and Neck Squamous Cell Carcinoma). De plus, nous avons porté notre attention sur un autre SNP affectant le gène ATG16L1 (Autophagy-related 16-Like 1), notamment rs2241880 (300Thr>Ala) qui compromet l’activité d’un gène fondamental dans le processus d’autophagie.Dans les cancers NSCLC, les allèles mutants "perte de fonction" des gènes ATG16L1, P2RX7 et TLR4 n'affectent pas la survie globale, indépendamment du type de chimiothérapie administrée. Au contraire, les patientes atteintes de HNSCC, portant au moins un allèle "perte de fonction" d’ATG16L1 et TLR4, présentent un taux de survie sans récidive inferieure par rapport aux patients qui présentent un genotype sauvage. Ce travail décrit pour la première fois un biomarqueur prognostique pour ce type de cancer.De plus, nous avons pu définir, par génotypage à haut débit, une signature de SNPs prédictive de la réponse à la chimiothérapie neoadjuvantes à base d'anthracyclines et des taxanes chez les patients atteints de cancer du sein. En particulier, la combinaison des deux paramètres clinicopathologiques classiques (âge lors du diagnostic et récepteur aux œstrogènes) avec les génotypes rs1076669 du gène ECE1 (Endothelin Converting Enzyme 1; 341Thr>Ile) et rs2277413 du gène PZP (Pregnancy-Zone Protein; 813Val>Ala) a permis de définir trois grandes catégories des patients et leur probabilité respective d'atteindre la réponse complète pathologique après traitement.L'identification de nouveaux biomarqueurs associés à une absence/diminution de réponse à la chimiothérapie apparaît critique pour choisir des alternatives thérapeutiques appropriées et éviter les effets secondaires indésirables chez les non-répondeurs. / Successful chemotherapeutics can induce a type of tumour cell death that is immunogenic, implying that patient’s dying cancer cells function as a therapeutic vaccine and elicit an anti-tumour immune response that controls the residual disease. The three main hallmarks of immunogenic cell death (ICD) are the pre-apoptotic exposure of calreticulin (CRT) on the cell surface, the autophagy-dependent secretion of ATP during the blebbing phase of apoptosis and the post-apoptotic release of the chromatin-binding non-histone protein high mobility group B1 (HMGB1). CRT, ATP and HMGB1 interact with CD91, purinergic receptor P2X, ligand-gated ion channel, 7 (P2RX7) and Toll-like receptor 4 (TLR4), respectively, on the surface of dendritic cells (DCs), thus promoting the engulfment of dying cells, the production of IL-1β and the cross-presentation of tumour-associated antigens to T cells, respectively.Our laboratory has demonstrated that adjuvant chemotherapy exhibits a reduced efficacy in breast and colorectal cancer patients bearing single-nucleotide polymorphisms (SNPs) that compromise the function of P2RX7 or TLR4, such as rs3751143 in P2RX7 (Glu496Ala) and rs4986790 in TLR4 (Asp299Gly).Driven by these results, underpinning the intimate relationship between the success of anti-cancer chemotherapy and an operational immune system, we decided to investigate the effect of these SNPs on disease outcome among non-small cell lung carcinoma (NSCLC) and head and neck squamous cell carcinoma (HNSCC) patients. Additionally, we focused our attention on a SNP in autophagy related 16-like 1 (ATG16L1), namely rs2241880 (Thr300Ala), which compromises the activity of one pivotal autophagic gene. In NSCLC patients, loss-of-function ATG16L1, P2RX7 and TLR4 alleles do not affect overall survival, irrespective of the administration and type of chemotherapy. Conversely, HNSCC patients bearing at least one loss-of-function ATG16L1 or TLR4 allele exhibit a reduced disease-free survival when compared to their wild-type counterparts. This is the first report highlighting a putative prognostic biomarker for this malignancy. Furthermore, taking advantage of a high throughput genotyping study, we delineated a SNP-based signature that predicts the response of breast cancer patients to anthracycline- and taxane-based neoadjuvant chemotherapy. Particularly, the combination of two classical clinicophatological parameters (age at diagnosis and estrogen receptor) and genotype at the ECE1 (rs1076669 Thr341Ile) and PZP (rs2277413 Val813Ala) loci allowed us to define three broad categories with a correspondent probability of achieving pathological complete response. The identification of new biomarkers associated with a reduced/absent response to chemotherapy appears critical for selecting appropriate therapeutic alternatives, and avoiding undesired side effects among non-responders.

Mammal species often live in social groups, but the factors that promote group cohesion can be difficult to analyze due to the prevalence of strong group affiliations. Feral horses maintain stable harems of one or two males and several females, and harem stability is strongly related to individual fitness. Anecdotal evidence and an early study in the non-breeding season suggest that management of the Shackleford Banks island horses with immunocontraception reduces harem stability in the population, providing an opportunity to study the factors that influence harem stability. I investigated the effects of the immunocontraceptive PZP on harem stability during the breeding season and examined mare activity budgets and harassment rates to determine if these factors influence harem stability. I hypothesized that 1) immunocontraception would increase the rates at which mares changed harems during the breeding season 2) activity budgets of contracepted individuals would differ significantly from those of uncontracepted individuals, and 3) contracepted mares would experience greater levels of harassment associated with changing harems than uncontracepted mares. I found that the immunocontraceptive does increase harem changes during the breeding season. I also found that contracepted mares have different activity budgets than uncontracepted mares; as predicted, contracepted mares grazed less and moved more than uncontracepted controls. The factors that influence mare activity budgets included immunocontraception, harem stallion, number of individuals in the harem, number of mares in the harem and body condition of the mare, as well as some interactions between factors. I found that high harassment rates by both harem stallions and other mares in the harem are correlated with higher harem change rates and that contracepted mares are harassed more than uncontracepted mares. These results indicate that the immunocontraceptive does influence harem stability in this feral horse population, potentially through alterations in activity budgets and harassment rates.