Testis tissue xenografting (TTX) provides a novel in vivo model for the study of testis function, and a previously-unavailable opportunity to produce spermatozoa in the grafts from immature donors of diverse species. The overall objectives of this thesis were to examine a number of factors that potentially affect the outcome of TTX, and to apply TTX using immature bison and deer donors as models for endangered ungulates. The objective of the first experiment was to examine the effects of recipient mouse strain, gender and gonadal status on the outcome of TTX. Eight small fragments of neonatal porcine testis tissue (~5 mg each) were grafted under the back skin of immunodeficient mice of different strains (SCID vs. nude), gender (male vs. female), and gonadal status (intact vs. gonadectomised), using a 2×2×2 factorial design (8 groups, n = 7 mice/group). The xenografts were recovered at 8 mo post-grafting and evaluated for gross and histological attributes. Gonadectomy of the recipients did not affect any of the measured outcomes of TTX (P > 0.05), and data were pooled into four groups based on recipient strain and gender. Overall, male recipient mice had grafts with higher mean (+SEM) recovery rate (97 ± 2.3% vs. 88 ± 2.4%, P = 0.004), weight (348 ± 26.3 vs. 104 ± 27.0 mg, P < 0.001), seminiferous tubular diameter (150 ± 3.3 vs. 108 ± 5.3 mg, P < 0.001), percentage of tubules containing spermatozoa (32 ± 3.2 vs. 6 ± 1.8%, P < 0.001), elongated spermatids (13 ± 1.4% vs. 4 ± 0.8%, P < 0.001), and round spermatids (10 ± 1.2% vs. 6 ± 1.1%, P = 0.006) than female mice. Overall, SCID mice had grafts with higher recovery rate (98 ± 2.4% vs. 87 ± 2.3%, P = 0.001), average weight (292 ± 27.0 vs. 160 ± 26.3 mg, P = 0.001), tubular density (44 ± 3.3 vs. 33 ± 2.1, P = 0.02), percentage of tubular cross-sections containing spermatocytes (27 ± 3.7% vs. 13 ± 2.3%, P = 0.003) than nude mice. Among the four groups of recipients, the grafts from male SCID mice had the highest weight (P < 0.05) and percentage of tubules containing spermatozoa (P < 0.05).<p> The objective of the second experiment was to evaluate the effect of using different numbers of donor testis tissue fragments on the outcome of TTX. Fragments of donor piglet testis tissue were grafted subcutaneously under the back skin of four groups of castrated male nude mice (n = 10/group). Each group of recipient mice received 2, 4, 8, or 16 fragments per mouse. Mice were sacrificed at 8 mo post-grafting, and xenografts were evaluated for physical growth and histological development. The relative weight of the vesicular gland (index) was also determined as a measure of bioactive androgen production by grafts in castrated recipient mice. The overall graft recovery rate was ~94% (range 86-98%) which did not differ among the groups (P > 0.05). The group of mice that received 16 testis tissue fragments had higher mean (+ SEM) graft weights (278 ± 39.4 vs. 106 ± 38.0, P = 0.02), total graft weight (2,443 ± 338.8 vs. 192 ± 76.2, P < 0.001), vesicular gland index (0.5 ± 0.06 vs. 0.1 ± 0.06, P = 0.007), and percentage of seminiferous tubules with round spermatids (11 ± 1.5 vs. 3 ± 1.3, P = 0.03) than the group of mice that received two testis tissue fragments. The objective of the third experiment was to assess the use to salvage testis tissue from neonatal/immature bison or deer donors using TTX into immunodeficient recipient mice as models for closely-related rare or endangered ungulates. Donor testis tissue fragments from two newborn bison calves (Bison bison bison) and a 2-mo-old white-tailed deer fawn (Odocoileus virginianus) were grafted under the back skin of gonadectomised nude mice (n = 15 and n = 7 for bison and deer groups, respectively, 8 testis fragments/mouse). To examine the potential effect of individual donors, we grafted four testis tissue fragments from one bison calf on one side of the recipient and four fragments from the second bison calf on the other side. Single grafts were surgically removed from representative recipient mice every 2 mo for up to 16- and 14 mo post-grafting, for bison and deer groups, respectively. The overall graft recovery rates were 69% and 63% for bison and deer groups, respectively. For bison grafts, a donor effect on efficiency of spermatogenesis was also observed. The weight of bison testis tissue xenografts increased (P < 0.02) ~4-fold by 2 mo and ~10-fold by 16 mo post-grafting, and gradual maturational changes were evident in the form of seminiferous tubule expansion starting at 2 mo, first appearance of spermatocytes at 6 mo, round spermatids at 12 mo, and elongated spermatids at 16 mo post-grafting. Testis tissue xenografts from donor white-tailed deer also showed a gradual development starting with tubular expansion by 2 mo and presence of spermatocytes by 6 mo post-grafting, round and elongated spermatids by 8 mo, followed by fully-formed spermatozoa by 12 mo post-grafting. The timing of complete spermatogenesis roughly corresponded to the reported timing of sexual maturation in these species.<p> Taken together, the findings in this thesis suggest that male SCID mice provide a more suitable recipient model for TTX with neonatal porcine testis tissue; recipient mice can be grafted with as many as 16 testis tissue fragments for optimal results; and that TTX is a feasible strategy for salvaging genetic materials from immature males of rare or endangered ungulates that die prematurely.

Testis tissue xenografting (TTX) provides a novel in vivo model for the study of testis function, and a previously-unavailable opportunity to produce spermatozoa in the grafts from immature donors of diverse species. The overall objectives of this thesis were to examine a number of factors that potentially affect the outcome of TTX, and to apply TTX using immature bison and deer donors as models for endangered ungulates. The objective of the first experiment was to examine the effects of recipient mouse strain, gender and gonadal status on the outcome of TTX. Eight small fragments of neonatal porcine testis tissue (~5 mg each) were grafted under the back skin of immunodeficient mice of different strains (SCID vs. nude), gender (male vs. female), and gonadal status (intact vs. gonadectomised), using a 2×2×2 factorial design (8 groups, n = 7 mice/group). The xenografts were recovered at 8 mo post-grafting and evaluated for gross and histological attributes. Gonadectomy of the recipients did not affect any of the measured outcomes of TTX (P > 0.05), and data were pooled into four groups based on recipient strain and gender. Overall, male recipient mice had grafts with higher mean (+SEM) recovery rate (97 ± 2.3% vs. 88 ± 2.4%, P = 0.004), weight (348 ± 26.3 vs. 104 ± 27.0 mg, P < 0.001), seminiferous tubular diameter (150 ± 3.3 vs. 108 ± 5.3 mg, P < 0.001), percentage of tubules containing spermatozoa (32 ± 3.2 vs. 6 ± 1.8%, P < 0.001), elongated spermatids (13 ± 1.4% vs. 4 ± 0.8%, P < 0.001), and round spermatids (10 ± 1.2% vs. 6 ± 1.1%, P = 0.006) than female mice. Overall, SCID mice had grafts with higher recovery rate (98 ± 2.4% vs. 87 ± 2.3%, P = 0.001), average weight (292 ± 27.0 vs. 160 ± 26.3 mg, P = 0.001), tubular density (44 ± 3.3 vs. 33 ± 2.1, P = 0.02), percentage of tubular cross-sections containing spermatocytes (27 ± 3.7% vs. 13 ± 2.3%, P = 0.003) than nude mice. Among the four groups of recipients, the grafts from male SCID mice had the highest weight (P < 0.05) and percentage of tubules containing spermatozoa (P < 0.05).<p> The objective of the second experiment was to evaluate the effect of using different numbers of donor testis tissue fragments on the outcome of TTX. Fragments of donor piglet testis tissue were grafted subcutaneously under the back skin of four groups of castrated male nude mice (n = 10/group). Each group of recipient mice received 2, 4, 8, or 16 fragments per mouse. Mice were sacrificed at 8 mo post-grafting, and xenografts were evaluated for physical growth and histological development. The relative weight of the vesicular gland (index) was also determined as a measure of bioactive androgen production by grafts in castrated recipient mice. The overall graft recovery rate was ~94% (range 86-98%) which did not differ among the groups (P > 0.05). The group of mice that received 16 testis tissue fragments had higher mean (+ SEM) graft weights (278 ± 39.4 vs. 106 ± 38.0, P = 0.02), total graft weight (2,443 ± 338.8 vs. 192 ± 76.2, P < 0.001), vesicular gland index (0.5 ± 0.06 vs. 0.1 ± 0.06, P = 0.007), and percentage of seminiferous tubules with round spermatids (11 ± 1.5 vs. 3 ± 1.3, P = 0.03) than the group of mice that received two testis tissue fragments. The objective of the third experiment was to assess the use to salvage testis tissue from neonatal/immature bison or deer donors using TTX into immunodeficient recipient mice as models for closely-related rare or endangered ungulates. Donor testis tissue fragments from two newborn bison calves (Bison bison bison) and a 2-mo-old white-tailed deer fawn (Odocoileus virginianus) were grafted under the back skin of gonadectomised nude mice (n = 15 and n = 7 for bison and deer groups, respectively, 8 testis fragments/mouse). To examine the potential effect of individual donors, we grafted four testis tissue fragments from one bison calf on one side of the recipient and four fragments from the second bison calf on the other side. Single grafts were surgically removed from representative recipient mice every 2 mo for up to 16- and 14 mo post-grafting, for bison and deer groups, respectively. The overall graft recovery rates were 69% and 63% for bison and deer groups, respectively. For bison grafts, a donor effect on efficiency of spermatogenesis was also observed. The weight of bison testis tissue xenografts increased (P < 0.02) ~4-fold by 2 mo and ~10-fold by 16 mo post-grafting, and gradual maturational changes were evident in the form of seminiferous tubule expansion starting at 2 mo, first appearance of spermatocytes at 6 mo, round spermatids at 12 mo, and elongated spermatids at 16 mo post-grafting. Testis tissue xenografts from donor white-tailed deer also showed a gradual development starting with tubular expansion by 2 mo and presence of spermatocytes by 6 mo post-grafting, round and elongated spermatids by 8 mo, followed by fully-formed spermatozoa by 12 mo post-grafting. The timing of complete spermatogenesis roughly corresponded to the reported timing of sexual maturation in these species.<p> Taken together, the findings in this thesis suggest that male SCID mice provide a more suitable recipient model for TTX with neonatal porcine testis tissue; recipient mice can be grafted with as many as 16 testis tissue fragments for optimal results; and that TTX is a feasible strategy for salvaging genetic materials from immature males of rare or endangered ungulates that die prematurely.

Submitted by Erika Demachki (erikademachki@gmail.com) on 2017-04-25T20:03:15Z No. of bitstreams: 2 Tese - Bruna Paula Alves da Silva - 2017.pdf: 1772056 bytes, checksum: 70764419b45a8c2ff0d3f4e7ed3fd5b4 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-04-26T12:54:20Z (GMT) No. of bitstreams: 2 Tese - Bruna Paula Alves da Silva - 2017.pdf: 1772056 bytes, checksum: 70764419b45a8c2ff0d3f4e7ed3fd5b4 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-04-26T12:54:20Z (GMT). No. of bitstreams: 2 Tese - Bruna Paula Alves da Silva - 2017.pdf: 1772056 bytes, checksum: 70764419b45a8c2ff0d3f4e7ed3fd5b4 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-03-29 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Fundação de Amparo à Pesquisa do Estado de Goiás - FAPEG / The brazilian bovine breeds Curraleiro Pé-Duro and Pantaneiro, inhabitants of Cerrado, Semi-árido and Pantanal are in the process of extinction, at risk of disappearing before their characteristics are adequately studied. The objective was perform the genealogical structure of breeds Curraleiro Pé-Duro and Pantaneiro, with a view to the conservation of animal genetic resources, in order to maintain the genetic variability within breeds, avoiding inbreeding, through proposals from matings based on genealogy. Samples of hair bulb from 1073 Curraleiro Pé-Duro cattle from nineteen herds and 290 Pantaneiros from four herds were analyzed. Amplification of 27 DNA microsatellites was performed by PCR. The average number of alleles per locus, allele frequency, heterozygosity expected and observed and polymorphic information content were calculated. For each animal an opinion of the affiliation control was issued. Large allelic variability was observed for the breeds, where the Curraleiro Pé-Duro presented an average of 8,70 alleles per loco and the Pantaneiro 6,70. The values of expected and observed heterozygosity were 0,713 and 0,653, respectively, for the Curraleiro Pé-Duro and 0,701 and 0,672 for Pantaneiro. The average value of polymorphic information content for the Curraleiro Pé-Duro was 0,671 and for the Pantaneiro 0,655. The probability of combined exclusion for the breeds was 0,99992. The mating lots were set at the minimum male:female ratio of 1:13 to the maximum of 1:26 for the Curraleiro Pé-Duro and 1:13 to 1:21 for the Pantaneiro, depending on the size of the herd. The paternity tests enabled the elaboration of reproductive programs for the herds, aiming to improve the genetic management of these populations, avoiding inbreeding. Both breeds showed great genetic variability inside of breeds, demonstrating that although they are in extinction have herds with a lot of genetic diversity. / As raças bovinas locais brasileiras Curraleiro Pé-Duro e Pantaneiro, habitantes do Cerrado, Semi-árido e do Pantanal encontram-se em processo de extinção, correndo o risco de desaparecerem antes que suas características sejam adequadamente estudadas. Objetivou-se realizar a estruturação genealógica de rebanhos das raças Curraleiro Pé-Duro e Pantaneiro, com vistas à conservação de recursos genéticos animais, visando manter a variabilidade genética dentro das raças, evitando-se a endogamia, por meio de propostas de acasalamentos baseadas na genealogia. Foram analisadas amostras de pelos de 1073 bovinos Curraleiro Pé-Duro de dezenove rebanhos e 290 Pantaneiros de quatro rebanhos. Realizou-se a amplificação de 27 microssatélites de DNA por PCR. Foi calculado o número médio de alelos por loco, frequência alélica, heterozigosidade esperada e observada e conteúdo de informação polimórfica. Para cada animal foi emitido parecer do controle de filiação. Observou-se grande variabilidade alélica para as raças, onde o Curraleiro Pé-Duro apresentou média de 8,70 alelos por loco e o Pantaneiro 6,70. Os valores médios de heterozigosidade esperada e observada foram 0,713 e 0,653, respectivamente, para o Curraleiro Pé-Duro e 0,701 e 0,672, para o Pantaneiro. O valor médio de conteúdo de informação polimórfica para o Curraleiro Pé-Duro foi 0,671 e para o Pantaneiro 0,655. A probabilidade de exclusão combinada para as raças foi 0,99992. Os lotes de acasalamentos foram montados na proporção macho:fêmea mínima de 1:13 à máxima de 1:26 para o Curraleiro Pé-Duro e de 1:13 à 1:21 para o Pantaneiro, dependendo do tamanho do rebanho. Os testes de paternidade possibilitaram a elaboração de programas reprodutivos para os rebanhos, com vistas a melhorar a gestão genética destas populações, evitando-se a endogamia. Ambas as raças apresentaram grande variabilidade genética intra-racial, demonstrando que apesar de estarem em extinção possuem rebanhos com muita diversidade genética.

The illegal trade in endangered animals in KwaZulu-Natal, with an emphasis on rhino poaching, is tactically addressed in this dissertation. The aim is to expose the nature and extent of these crimes; the victims, offenders and modus operandi involved; the adjudication of wildlife offences; the causes and consequences concerned; the relevant criminological theories to explain these crimes; and recommendations for prevention. This research intends to examine the contemporary pandemic of rhino poaching in KwaZulu-Natal, South Africa, and offer potential techniques for intervention. Furthermore, one of the main goals of the study is to reveal and enhance the extremely neglected field of conservation criminology. A general disregard by society for the environment, as well as the overall ineffectiveness and corruption of criminal justice and conservation authorities, comes to the fore. The purpose of the research is therefore to suggest possible prevention strategies in order to protect the rights of endangered species. / Criminology and Security Science / M.A. (Criminology)

The illegal trade in endangered animals in KwaZulu-Natal, with an emphasis on rhino poaching, is tactically addressed in this dissertation. The aim is to expose the nature and extent of these crimes; the victims, offenders and modus operandi involved; the adjudication of wildlife offences; the causes and consequences concerned; the relevant criminological theories to explain these crimes; and recommendations for prevention. This research intends to examine the contemporary pandemic of rhino poaching in KwaZulu-Natal, South Africa, and offer potential techniques for intervention. Furthermore, one of the main goals of the study is to reveal and enhance the extremely neglected field of conservation criminology. A general disregard by society for the environment, as well as the overall ineffectiveness and corruption of criminal justice and conservation authorities, comes to the fore. The purpose of the research is therefore to suggest possible prevention strategies in order to protect the rights of endangered species. / Criminology and Security Science / M.A. (Criminology)