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African painted dogs (Lycaon pictus, APD) are highly endangered, with fewer than 7000 remaining in nature. Captive breeding programs can preserve a genetically diverse population and provide a source of individuals for reintroductions. However, most programs are initiated from few founders and suffer from low genetic diversity and inbreeding. The aims of this study were to use molecular markers to assess genetic variation, inbreeding, and relatedness among APDs in the North American captive population, to use these data to realign studbook records, and to compare these data to wild populations and to the European captive population to facilitate the development of a global management plan. We sequenced mitochondrial and major histocompatibility (MHC) class II loci and genotyped 14 microsatellite loci from 109 APDs from 34 institutions in North America. We identified three likely studbook errors and resolved ten cases of uncertain paternity. Overall, microsatellite heterozygosity was higher than reported in Europe, but effective population size estimates were lower. Mitochondrial sequence variation was extremely limited, and there were fewer MHC haplotypes than in Europe or the wild. Although the population did not show evidence of significant inbreeding overall, several individuals shared high relatedness values, which should be incorporated into future breeding programs.

Dissertação de Mestrado Integrado em Medicina Veterinária The genetic diversity and population structure of the California sea lion (Zalophus californianus) were assessed based on the pedigree information registered in the European studbook, currently under the management of the Lisbon Zoo, as an EEP programme. Records collected from 1930 until 2020 were used for this analysis, including a total of 1998 individuals, 477 of which represent the current living population. To our knowledge, this is the most comprehensive and up-to-date analysis for this species, assessing the evolution of genetic diversity, inbreeding and relationships, and the genetic contribution of founders, using conventional pedigree analysis procedures and additional software tools commonly used in domestic species, such as ENDOG. The average equivalent complete generations were 1.54 (±1.20) per animal for the overall population. However, there has been an increase in pedigree depth throughout the years, with the reference population reaching 3.37 (±0.76) complete generations, where all animals had maternal grandparents known and nearly 85% to 90% had paternal grandparents known. The whole population had a generation interval of 11.52 (±4.70) years, however the mean ages at which individuals had their progeny has been increasing, particularly in the last decade. Regarding the number of offspring for each parent, dams had an average of 4.05 (±3.16) pups and sires 7.97 (±8.57) pups. The overall mean inbreeding was 2.03% (±5.92), though throughout the years, this has been increasing, with the reference population reaching 3.34% (±6.08). The rate of increase in inbreeding was around 1.6% per generation. The number of inbred animals has also been growing, representing 16.87% and 52.94% of the total and reference populations, respectively. However, the average inbreeding coefficient, of the inbred individuals has shown a significant decrease, with the value for the reference population being nearly half that of the overall population. The mean coancestry among individuals from the same institution of birth, was 0.10, and from different institutions was 0.01, providing the opportunity to maintain inbreeding under control by planned exchange of breeders between institutions. The number of founders contributing to 50% of the gene pool was 36 and 12, for the total and reference populations, respectively, with an effective number of founders of 96 and 34 in the two populations. The effective population size, for the whole population was 36. It is important to maintain a sound program aimed at the genetic management of the population, including management of selection and mating decisions, to avoid further losses of genetic diversity. RESUMO - Diversidade genética do Leão-marinho Californiano avaliada através da análise do pedigree - A diversidade genética e a estrutura da população de leões-marinhos Californianos (Zalophus californianus) foi avaliada com base nos registos de pedigree do studbook Europeu, sob a gestão do Jardim Zoológico de Lisboa, como um programa EEP. Os registos, recolhidos desde 1930 até 2020, que foram utilizados para esta análise, incluem um total de 1998 indivíduos, 477 dos quais representam a população viva atual. Para nosso conhecimento, esta é a análise mais compreensiva e atualizada desta espécie, que avalia a evolução da diversidade genética, consanguinidade, parentescos, e contribuição genética de fundadores, usando métodos convencionais de análise de pedigree e ferramentas de software adicional, tipicamente usadas para espécies domésticas, como o ENDOG. A média de gerações completas equivalentes foi de 1.54 (±1.20) por animal, para a população total. No entanto, tem havido um aumento na profundidade do pedigree, ao longo dos anos, com a população de referência a alcançar as 3.37 (±0.76) gerações completas, e onde todos os animais tinham avós maternos conhecidos e 85% a 90% tinham avós paternos conhecidos. A população total teve um intervalo de geração de 11.52 (±4.70) anos, no entanto a média das idades às quais os indivíduos tiveram filhos tem vindo a aumentar, particularmente na última década. Em relação ao número de filhos por progenitor, para as fêmeas foi 4.05 (±3.16) crias e para os machos 7.97 (±8.57) crias. A consanguinidade média geral foi de 2.03% (±5.92), embora ao longo dos anos, tenha vindo a aumentar, com a população de referência a alcançar um valor de 3.34% (±6.08). A taxa de aumento da consanguinidade foi de 1.6% por geração. O número de animais consanguíneos tem vindo a aumentar, representando 16.87% e 52.94% das populações total e de referência, respetivamente. No entanto, a média do coeficiente de consanguinidade, dos consanguíneos, mostra uma diminuição significativa, com o valor da população de referência a ser metade do da população total. O relacionamento médio, nos indivíduos da mesma instituição de nascimento foi de 0.10 e de diferentes instituições de 0.01, proporcionando a oportunidade de manter a consanguinidade sob controle por meio da troca planeada entre as instituições. O número de fundadores que contribuíram para 50% do pool genético foi de 36 e 12, para as populações totais e de referência, respetivamente, com um número efetivo de fundadores de 96 e 34, nas duas populações. O tamanho efetivo da população total foi de 36. É importante manter um programa apontado para a gestão genética da população, incluindo o maneio da seleção e decisões de acasalamento, para evitar maiores perdas de diversidade genética. N/A

Dissertação de Mestrado apresentada no ISPA – Instituto Universitário para obtenção de grau de Mestre na especialidade de Biologia Marinha e Conservação Com o tempo, o homem tem vindo a tomar cada vez mais consciência da grande importância da proteção, preservação e recuperação do meio ambiente, assim como das espécies ameaçadas. Aliadas à missão de conservação estão várias entidades que, diariamente, trabalham com intuito de manter populações saudáveis e autossustentáveis. Este estudo apresenta os resultados da análise demográfica e genética da população do Leão-marinho-da-califórnia [Zalophus californianus (Lesson, 1828)], espécie gerida no âmbito de Programa ex situ da EAZA (EEP). Realizou-se um questionário que foi enviado a diversas instituições, com o objetivo de criar um Plano de Gestão a Longo Prazo para a espécie, através de um bom controlo do tamanho da sua população. Os dados desta população encontram-se registados no seu Studbook desde 1991, dados estes que são essenciais para monitorizar e gerir o estabelecimento e o crescimento de populações mantidas em cativeiro, facilitando o desenvolvimento de uma população viável e ajudando a maximizar a diversidade genética e a estabilidade demográfica. Através da atualização e correção de erros do Studbook foi possível aumentar a percentagem de pedigree conhecido. Para a análise demográfica e genética da população utilizou-se o software PMx. Os resultados mostram que a população do EEP se encontra em crescimento, demostrando parâmetros demográficos positivos, tais como: taxa instantânea de crescimento (r =0,039); taxa de crescimento populacional (λ = 1,040) e; taxa líquida de reprodução (Ro = 1,605). Relativamente aos parâmetros genéticos, a diversidade genética da população é alta e tem potencial para aumentar, o que indica um bom estado genético. Over time, man has become increasingly aware of the great importance of protecting, preserving, and recovering the environment, as well as threatened species. Allied to the conservation mission are several entities that work daily in order to maintain healthy and self-sustainable populations. This study presents the results of the demographic and genetic analysis of the population of the California Sea Lion [Zalophus californianus (Lesson, 1828)], a species managed under the EAZA Ex situ Programme (EEP). A questionnaire was prepared and sent to several institutions, with the aim of creating a long-term management plan for the species, through good control over the size of its population. This population's data has been recorded in a Studbook since 1991. This data is essential to monitor and manage the establishment and growth of populations kept in captivity, facilitating the development of a viable population, and helping to maximize genetic diversity and demographic stability. By updating and correcting Studbook errors, it was possible to increase the percentage of pedigree known. For the demographic and genetic analysis of the population, the PMx software was used. The results show that the population of the EEP is growing, showing positive demographic parameters, such as: instantaneous growth rate (r = 0.039); population growth rate (λ = 1,040); net reproductive rate (Ro = 1.605). Regarding genetic parameters, the genetic diversity of the population is high and has the potential to increase, which indicates a good genetic status.

Osoitan väitöskirjassani nykyään seurattavien kotieläinten jalostusperiaatteiden synnyn suomalaisen hevosrodun perustamisvaiheiden kautta. Puhtaan rodun jalostusteorialla oli käänteentekevä merkitys kotieläinten jalostukseen Euroopassa 1800­-luvulla. Eläinten jalostus ei ollut ajatuksena uusi, mutta uusi teoria pysäytti eri eläinkantojen sekalaiset risteytykset, tieteellistytti jalostuksen ja nosti eläinten luokittelun perustaksi rodun käsitteen sen modernissa merkityksessä. Keskipisteeseen nousivat modernin eläinjalostuksen käsitteet, rotu ja sen kantakirja. Koska tutkin suomalaista hevosjalostusta kotieläinjalostusteorioiden kansainvälisessä murroskohdassa, sitä ei voi nähdä vain kansallisena liikkeenä. Tutkimukseni yksi tärkeä näkökulma on jalostuksen eurooppalainen oppihistoria ja tieteellistyminen. Sitä kautta analysoin, miten kansainväliset eläinjalostusvaikutteet ja kansalliset hevostarpeet yhdistyivät suomalaisessa hevosjalostuksessa. Tutkin väitöskirjassani, miten kansainvälisestä kantakirjainnovaatiosta muodostui hevosjalostuksen perusta Suomessa vuosina 1893–1907. Tutkimukseni osat ovat suomalaisen hevosen kantakirjan kolmivaiheisen institutionalisoitumisprosessin mukaan innovaation tulo Suomeen (1891–1895), järjestelmän rakentuminen (1893–1899) ja aatteen valtakunnallinen läpimurto (1900–1907). Aikarajaus alkaa Suomen ensimmäisen kantakirjahevosen valitsemisesta paikalliseen Uudenmaan ja Hämeen läänien maanviljelysseuran kantakirjaan ja päättyy suomalaisen hevosrodun valtakunnallisen valtion kantakirjan läpimurtoon. Tähän väliin mahtuvassa suomalaisen hevosrodun syntyprosessissa on yhteneväisyyksiä innovaatioteorioihin, joista olen seurannut Everett M. Rogersin teoriaa innovaatiota koskevasta päätöksenteosta. Tutkimukseni paljastaa kantakirjan hallinnoinnista muodostuneen kilpajuoksun kansalaisyhteiskunnan ja valtionhallinnon välillä. Kantakirjasta kilpailivat Uudenmaan ja Hämeen läänien maanviljelysseura, senaatin maataloushallinto ja hevosjalostuksen ympärille perustettu Hippos­-yhdistysliike. Yksilön merkitys näkyi siinä, että eri tahojen kantakirjat henkilöityivät vahvasti yksittäisiin toimijoihin. Tutkimukseni useat tasot selittävät siten lähdeaineiston laajuuden. Päälähteet ovat maataloushallinnon, seurojen ja virkamiesten arkistot sekä muu viranomaisaineisto ja aikalaiskirjoitukset. Tutkimukseni osoittaa, että kantakirjakirjainnovaatio tuotiin Suomen hevosjalostuksen perustaksi kansainvälisen kotieläinjalostusteorian ja kansallisten hevostarpeiden yhteisvaikutuksesta. Ylirajaisen kotieläinjalostusliikkeen rantautumisessa suomalaiseen hevosjalostukseen oli omat erityispiirteensä. Valtion vahva puuttuminen kantakirjan hallinnointiin heti innovaation käytäntöön viennin alusta alkaen oli yleisesti yhdistyksiä harvinaisempaa. Vielä erikoisempaa oli, että kantakirja kohdistettiin yksin kotimaiseen, alkuperäiseen hevostyyppiin. Yleisesti osa maatiaiseläimistä ehti hävitä, ennen kuin ne otettiin järjestelmällisen jalostuksen piiriin. Kotieläinjalostuksen kansainvälisessä modernisaatiossa harvinaislaatuisesti hevosjalostuksen perustaksi nousi Suomessa yhden rodun – maatiaishevosen – valtakunnallinen kantakirja. Valinnan keskeisenä perusteena oli hevosvoiman käyttötarve, monipuolisen työhevosen ensisijaisuus. Maa-­ ja metsätalouden sekä liikenteen teknologisen järjestelmän tärkeä osa, maatiaishevonen oli joutunut kovan muutospaineen alle etenkin maatalouden hevosvetoisten koneiden kehityksen seurauksena. Teoreettisen viitekehyksen tutkimukselle antaa tätä kautta myös Thomas P. Hughesin teknologisen järjestelmän käsite. Yhtenäisen eläinrodun muodostaminen on pitkä prosessi – jossa käytäntö ja ideologia eivät aina ole ristiriidattomia –, mutta kantakirjan perustamisen seurauksena suomalaista maatiaishevosta voitiin alkaa jalostaa rotuna oman rotumääritelmän mukaisesti. Suomenhevosen – kuten suomenkarjan – olemassaolon nykypäivän suojeltuna alkuperäisrotuna mahdollisti siten eurooppalaisen puhtaan rodun jalostusteorian sovellus maatiaiseläimiin 1800­-luvun lopulla. Yhtenä maatiaiseläinjalostuksen edelläkävijänä Suomi toi kotieläinjalostuksen kansainväliseen modernisaatioon oman erityislaatuisuutensa. Suomessa hevonen puolestaan oli kotieläimistä yhtenä keskeisenä esikuvana yleiselle jalostuksen modernisaatiolle, joka myöhemmin laajeni koskemaan kotieläinjalostusta kokonaisuudessaan. In my study, I discuss the origin principles of domestic animal breeding and how these are reflected in modern animal breeding. I show this via the stages of establishment of a horse breed now known as the Finnhorse. The purebred breeding theory revolutionized breeding of domestic animals in Europe in the 1800s. Animal breeding was not new as an idea, but the new theory stopped the production of miscellaneous crosses; breeding became scientific and the concept of breed in its modern sense became the basis for the classification of animals. The concepts of modern animal breeding, the breed and the studbook rose to the core. As I analyze Finnish horse breeding at the international turning point of domestic animal breeding theories, it cannot only be seen as a national movement. One of the main aspects of my study is the history of European doctrine of breeding and the scientification of breeding. Through this, I analyze how international influences of animal breeding and national requirements for horses merged in Finnish horse breeding. In my study, I analyze how international studbook innovation became the basis for horse breeding in Finland between 1893 and 1907. The three­stage process of institutionalization of the Finnish horse studbook is the basis of my study. The stages are: the arrival of innovation to Finland (1891–1895), the structural development of the system (1893–1899), and the wider breakthrough of the idea on a national level (1900–1907). The study begins with how the first horse was chosen for the local studbook, which was maintained by a regional agricultural society, called Uudenmaan ja Hämeen läänien maanviljelysseura. The study ends with the breakthrough of a national studbook of the Finnish horse breed maintained by the state. There are convergences in the establishment of the Finnish horse breed and the innovation theories in which I have followed the theory of decision­making concerning innovation by Everett M. Rogers. My study reveals that there was a scramble for the administrative rights of the studbook between the civil society and the government. The regional agricultural society, Uudenmaan ja Hämeen läänien maanviljelysseura, the agricultural administration of the Senate and horse breeding associations, collectively known as Hippos, were all competing for the ownership of the studbook. In all quarters, the studbooks were strongly linked to individuals. The extent of the source materials hence explains the multiple levels in my study. The main sources of the study are the archives of the agricultural administration, society archives, public officials’ archives, other official records and the writings of contemporaries. My study shows that studbook innovation became the basis of Finnish horse breeding and this was a combined result of the arrival of the international animal breeding theory and the need for horses on a practical level – i.e. for agriculture, forestry, transport and industry. This was something of a national requirement. The arrival of the cross­regional movement of domestic animal breeding in Finnish horse breeding had its own characteristics. Right from the beginning, the state was strongly interventional in the administration of the studbook, which was less common – something that was usually initially left to the associations to implement. It was even more peculiar that the studbook concentrated on the national native type of horse alone. In general, many of the native types of animal had already died out before they were taken into account within a systematic breeding program. In Finland, the nationwide studbook for one breed – the native type of horse – became the basis for horse breeding, which was rare internationally within the modernization of domestic animal breeding at the time. The requirement for the utilization of horsepower, for producing an all­round work horse, was the main reason why the Finnish native horse was selected first for a systematic breeding program. An important part of the technological system of agriculture, forestry and transport, the native type of horse had been put under a lot pressure for a change, in particular as a result of the advances made in the development of horse­drawn agricultural machinery. The concept of technological system by Thomas P. Hughes hence provides a theoretical framework for this study too. It is a long process to create a uniform animal breed – the practice and ideology do not always exist without contradictions. However, as a result of the founding of the studbook, it was possible to begin breeding the Finnish native horse within its own breed definition. As a result, had the European purebred breeding theory not been applied to the native types of animal at the end of the 19th century, breeds such as the Finnhorse and Finncattle would not exist as protected native breeds today. As one of the pioneers in native breed breeding, Finland introduced its own specific characteristics to the international modernization of domestic animal breeding. In Finland, the horse for its part served as one of the central models in the general modernization of breeding, which later expanded to involve domestic animal breeding as a whole.

The aim of this work was to analyze genetic parameters such as the inbreeding coe cient (F), relatedness coe cient (AR) and partial inbreeding coe cient (Fij) of the whole PRE population, and the ancestors which account for 50% of the total genetic variability of the current population, from genealogical information. The average F of the whole PRE population (328,706 animals) has decreased from 8.45% to 7.51% in the least 20 years. The Fij was estimated for the whole PRE population, resulting in a database of 58,772,533 records containing one record for each Fij that each animal receives from a certain common ancestor (CA). A total of 10,244 CAs contributed to the Fij with an average of 5370 descendants, with each descendant having an average of 170 CAs. Over the generations, the number of CAs has increased, while the proportion of Fij by each one has decreased. In addition, the contributions of the more influential ancestors have changed. The increased census, the limited use of artificial insemination and our increased knowledge about inbreeding depression and the animals’ breeding values allow breeders to select horses more for their functionality and conformation than for their pedigree reputation, which is the cause of all these changes.

Ameriški quarter konj je pasma, ki se je začela razvijati v 17. stoletju. Vsak konj te pasme mora biti registriran pri American Quarter Horse Association, kot tudi v državi, v kateri se nahaja. Z Uredbo (EU) 2016/1012 … (2016) je potrebna prenova rejskih programov in vzpostavitev rodovniške knjige za vse pasme, ki se nahajajo v državi. Poleg tega mora vsakega konja pri prodaji spremljati zootehniški dokument. Prvi ameriški quarter konj je bil v Slovenijo pripeljan jeseni leta 1995. To je bil Future Enjoy, ki ima v Sloveniji registriranih 13 potomcev. Do julija 2019 je bilo v Sloveniji skotenih 452 ameriških quarter konj, skupno registriranih pa kar 821. Za sprejem v rodovniško knjigo je bilo pregledanih 38 od 115 kobil, ki izpolnjujejo osnovne pogoje. V letu 2019 je licenciranih 6 žrebcev pasme ameriški quarter konj. Linearno oceno, ki je potrebna za razvrščanje v pododdelke rodovniške knjige, je od leta 2015 naprej prejelo 73 ameriških quarter konj. Predlagamo naslednjo obliko rodovniške knjige: tri glavne rodovniške knjige (Knjiga žrebcev, Knjiga Kobil in Knjiga kastratov in steriliziranih kobil), ki so naprej deljene na Osnovno knjigo, Splošno knjigo, Knjigo I in Knjigo II, poleg glavne knjige pa predlagamo še dodatni del. The American Quarter Horse is a breed developed in the 17th century. Every horse of this breed has to be registered with the American Quarter Horse Association and also in the country in which the horse resides. Changing of the EU legislation (Regulation (EU) 2016/1012 …, 2016) requires the revision of breeding programs and establishment of Stud Books for all horse breeds located in member country. Also, every horse must be accompanied by zootechnical certificate in trade. The first American Quarter Horse arrived in Slovenia in the Autumn of 1995. The horse was Future Enjoy, which had 13 registered offsprings in Slovenia. Altogether, the 452 American Quarter Horses were born by July 2019 and there were 821 registered American Quarter Horses in Slovenia. Thirty-eight of 115 mares which qualify for acceptance in the Stud Book have already been assessed. In 2019, there are six licensed American Quarter Horse breeding stallions. From 2015 to 2018, 73 horses were linearly scored, which is necessary for classification of horses into the subdivisions of the Stud Book. We suggest three major books (Stallions’ Book, Mares’ Book and Geldings’ and Sterilized Mares’ Book) and Appendix Book. The major books should have subdivisions named Basic Book, General Book, Book I and Book II.

In many birds and mammals, the size and sex composition of litters can have important downstream effects for individual offspring. Primates are model organisms for questions of cooperation and conflict, but the factors shaping interactions among same-age siblings have been less-studied in primates because most species bear single young. However, callitrichines (marmosets, tamarins, and lion tamarins) frequently bear litters of two or more, thereby providing the opportunity to ask whether variation in the size and sex composition of litters affects development, survival, and reproduction. To investigate these questions, we compiled a large dataset of 9 species of callitrichines (n= 27,080 individuals; Callithrix geoffroyi, Callithrix jacchus, Cebuella pygmaea, Saguinus imperator, Saguinus oedipus, Leontopithecus chrysomelas, Leontopithecus chrysopygus, Leontopithecus rosalia, and Callimico goeldii) from zoo and laboratory populations spanning 80 years (1938 to 2018). Through this comparative approach, we found several lines of evidence that litter size and sex composition may impact fitness. Singletons have higher survivorship than litter-born peers and they significantly outperform litter-born individuals on two measures of reproductive performance. Further, for some species, individuals born in a mixed-sex litter outperform isosexually-born individuals (i.e., those born in all-male or all-female litters), suggesting that same-sex competition may limit reproductive performance. We also document several interesting demographic trends. All but one species (Cebuella pygmaea) has a male-biased birth sex ratio (BSR) with higher survivorship from birth to sexual maturity among females (although this was significant in only two species). Isosexual litters occurred at the expected frequency (with one exception: Cebuella pygmaea), unlike other animals, where isosexual litters are typically over-represented. Taken together, our results indicate a modest negative effect of same-age sibling competition on reproductive output in captive callitrichines. This study also serves to illustrate the value of zoo and laboratory records for biological inquiry.

Traditionally, studbooks have been used as the major tool for the scientific management of ex-situ populations organised in breeding programmes. Well documented pedigree information enables managers to sufficiently monitor population size, demographic stability and the level of genetic diversity. However, breeding programes can only maintain the genetic information which is brought into captivity by wild caught founder animals. In most studbooks little is known about those individuals. This is also true for the red panda (Ailurus fulgens), a charismatic but threatened flagship species. Conservation breeding of this remarkable mammal species started in the early 1960s and is nowadays coordinated through a Global Species Management Plan (GSMP). In Europe the Endangered Species Programme (EEP) forms the biggest regional breeding group of red pandas (subspecies A. f. fulgens). Little is known about the true origin of the 23 founders of the EEP and how their unknown relationships may affect their living descendants now and in the future. Studbook data on 1350 individuals were analysed for different assumptions about founder relationships and compared with each other. We provide studbook-based evidence that the annual change in genetic parameters (gene diversity and mean inbreeding) are not affected in long-term by the original founder relationship within strongly intermixed zoo populations managed for mean kinship. Our results point out the importance of good genetic management in the early years of breeding programmes. These early years are crucial in implementing knowledge of founder relationships in the studbook, since a population becomes genetically equalised within the first decades and new information obtained later has no significant effects on subsequent genetic trends.

El objetivo principal de ésta tesis fue realizar un análisis demográfico, genético y reproductivo de la población de jaguares (Panthera onca) cautivos en Colombia. En el análisis demográfico se realizó la construcción del Studbook Regional para Jaguares (Panthera onca) (n=38), se obtuvo el censo de los animales que han estado presentes en las colecciones zoológicas de Colombia desde el año de 1970 hasta el año 2009. Se estableció la distribución de los ejemplares vivos en la actualidad por clases de edades. Se obtuvieron las características demográficas para las poblaciones actuales y poblaciones modelos de jaguares machos y hembras; también se construyó la tabla de vida para machos y hembras. En el análisis genético, a partir de los registros, se calcularon las estadísticas poblacionales. Se encontró que en la población se encuentran 2 nuevos posibles fundadores potenciales (n=2). Se realizaron las proyecciones reproductivas para un tiempo de 5, 10, 15 y 20 años con el fin de incrementar la pobalción de 25 individuos a 40. Se realizó la selección de apareamientos empleando el valor de la media de parentesco MK, y la ubicación geográfica. Se realizó una propuesta de meta poblacional para los jaguares en Colombia de conservar el 90% de la diversidad genética durante un periodo de 100 años, teniendo en cuenta las siguientes variables: Tiempo generacional (T =12), Tasa de crecimiento poblacional (λ= 1.02), Tamaño poblacional actual (N =25), Número efectivo poblacional actual (Ne =6.5), Relación Ne/N (0.26), Diversidad genética actual (GD =0.78), Población máxima permitida (n=40), Población requerida para alcanzar metas (n=37), FGE reclutado por cada nuevo fundador (FGE =0.4). El plan de manejo poblacional para el jaguar en parques zoológicos de Colombia fue propuesto estableciendo metas de manejo poblacional para los siguientes 100 años. También se realizó un análisis genético de la población de jaguares cautivos en parques zoológicos de Colombia mediante el análisis tanto de registros del Studbook Regional para Jaguares como de muestras de ADN obtenidas de 20 animales vivos (n=20), con el cual se construyó el pedigrí para la población empleando 9 microsatélites. Las paternidades y maternidades fueron asignadas por el programa CERVUS y las relaciones de parentesco fueron obtenidas por el programa KINSHIP. El análisis del Studbook se realizó por medio del programa PM2000 en donde se obtuvo el valor de la diversidad genética (GD) actual de la población (GD=0.7832), el valor de diversidad genética potencial (GDp=0.9113), valor génico (GV=0.7846), el valor del equivalente del genoma fundador actual (FGE=2.31) y potencial (FGEp=5.64), el valor del genoma fundador sobreviviente actual (FGS=3.14) y potencial (FGSp=5.64), el coeficiente medio de consanguinidad (F=0.0179), el número efectivo poblacional (Ne=5.26), al igual que la relación (Ne/N =0.2667) y la media de parentesco MK (Mean KINSHIP) para cada individuo. En el análisis de los microsatélites se calcularon las frecuencias alélicas por cada microsatélite y se encontró un promedio de alelos por locus de 5.67. El promedio de la heterocigocidad observada general fue de 0,687 y el de la heterocigocidad esperada 0,684. Se encontró que todos los locus se encuentran en equilibrio Hardy–Weinberg. Para esta población se encontró un exceso de heterocigotos en los locus FCA075, FCA043, FCA736, FCA220 y FCA391 y exceso de homocigotos solo para los locus FCA 441, FCA 008, FCA 224 mediante los estimados de Fis de Weir y Cockerham's (W and C), y Robertson y Hill's (R and H). El promedio de Fis para la población fue (Fis W and C= -0,0157) y (Fis R and H=-0,0109) indicando que la población se encuentra en exogamia. No se encontraron diferencias significativas para la prueba de desequilibrio por ligamiento. Los análisis de verosimilitud obtenidos en CERVUS resultaron con asignaciones de la maternidad y de paternidad. Los asignamientos se realizaron según el logaritmo natural de la tasa de verosimilitud (LOD). Se obtuvieron las probabilidades de relación y parentesco asignadas por KINSHIP de las simulaciones y se encontraron relaciones significativas (p0.05) en 6 parejas de jaguares, (p0.01) para 3 parejas y (p0.001) para 11 parejas. Con este estudio se logró establecer la relación entre individuos y se logró construir el pedigrí de la población de jaguares cautivos en parques zoológicos de Colombia como base fundamental para el establecimiento de un plan de conservación ex situ para el Jaguar en parques zoológicos de Colombia. Por último se realizó la caracterización reproductiva de un grupo de jaguares cautivos (n=6) en las instalaciones de la Fundación Zoológico Santa Cruz en Cundinamarca, Colombia, mediante un análisis longitudinal no invasivo de estradiol y progesterona en hembras y testosterona en machos. En un periodo de un 1 año dividido en cuatro épocas climatológicas las hembras confinadas en solitario fueron evaluadas con el objetivo de caracterizar la actividad ovárica y determinar la presentación de ovulaciones espontáneas. En los machos la actividad testicular. En las hembras se realizó un segundo seguimiento después de la aplicación de un protocolo convencional de gonadotropinas. La hormonas fueron extraídas de muestras de materia fecal de tres hembras (n=3) y dos machos (n=2). La medición de estradiol se realizó por medio de radioinmunoanálisis (RIA) y la medición de progesterona y testosterona por medio de ensayo por inmunoabsorción ligado a enzimas (ELISA). El modelo lineal de regresión de efectos mixtos encontró diferencias significativas en las concentraciones de estadiol en las diferentes estaciones (Chi square = 15.97, DF = 3, p 0.01). Las comparaciones de medias estacionales se realizó mediante el método de la diferencia significativa honesta de Tukey (DSH), revelando diferencias significativas entre las estaciones: Sequía 1 vs. Lluvias 2 (p 0.01), Lluvias 1 vs. Lluvias 2 (p 0.05), y Sequía 2 vs. Lluvias 2 (p 0.05). En las tres jaguares hembras se encontraron elevaciones de progesterona compatibles con ovulaciones expontáneas y el modelo lineal de regresión de efectos mixtos también encontró diferencias significativas en las concentraciones de progesterona en las diferentes estaciones (Chi square = 28.56, DF = 3, p 0.01). El DSH mostró diferencias significativas solo entre las estaciones: Sequía 2 vs.Lluvias 2 (p 0.01). La estación con el nivel promedio más bajo fue lluvias 2 (Octubre, Noviembre, y Diciembre). Durante esta estación periodos de anestro fueron registrados entre 31 y 83 días. Las tres hembra mostraron picos de estradiol después de la administración de gonadotropina coriónica equina (eCG). El análisis longitudinal no invasivo para los machos (Macho 1 y Macho 2) no mostró diferencias significativas en el curso del año de análisis. El análisis seminal de los tres machos adultos (n = 3) fue realizado por medio de electroeyaculación bajo anestesia general (Macho 1 and 2) y por lavado epididmal post mortem (Macho 3). La concentración media de espermatozoides encontrada fue 5.7 x 106 ± 1.1 x 106 espermatozoides/ml. La motilidad progresiva promedio fue de 80 %. El porcentaje de espermatozoides normales obtenidos por electro eyaculación + desviación estandar (DS) fue 80 ± 2.8%, y las anormalidades más frecuentemente encontradas fueron defectos de cabeza (7 ± 1.4%). El fluido obtenido en el lavado epididimal mostró 35 ± 1.4% de espermatozoides normales, y la anormalidad más frecuente fueron las gotas citoplasmáticas distales (39 ± 11.3%). Abstract. The objective of this thesis was to analyze the demographic, genetic and reproductive status of captive jaguars in Colombia. In the demographic analysis the Regional Studbook for Jaguars (Panthera onca) (n=38) was constructed using the software SPARKS. The studbook data were analyzed by the software population management 2000 (PM2000). The census of the animals that had been present in the zoological collections of Colombia from the year of 1970 until the year 2009 was obtained. The distribution of the living animals by age classes was established and the demographic characteristics were obtained for the current population and model population of male and female jaguars, as well as the life table for males and females. The reproductive projections were obtained for 5, 10, 15 and 20 years in order to increase the captive population from 25 individuals to 40. The selection of matings was based on the Mean KINSHIP (MK) value, and the geographical location. It is presented the population goals for the jaguars in Colombia to conserve 90% of the genetic diversity during a 100 year period, for the variables: Generational Time (T =12), populational growth rate (λ = 1.02), current populational size (N =25), current populational effective Number (Ne =6.5), Relationship (Ne/N =0.26), current genetic Diversity (GD =0.78), allowed maximum Population (n=40), required Population to reach goals (n=37), FGE recruited by each new founder (FGE =0.4). The population management plan for the Jaguar (Panthera onca) in Colombia was proposed for the Colombian Zoological Parks establishing population management goals for the next 100 years. Also, a genetic analysis for the captive population of jaguars (Panthera onca) was done in zoological parks of Colombia based upon the Colombian Regional Studbook for Jaguars and from DNA obtained form 20 living animals (n=20) in order to establish the pedigree for the captive population using 9 microsatellites. The assignments for paternities and maternities were done with for the program CERVUS and the relationship between animals were established with the KINSHIP program. The analysis of the Studbook was done with the PM2000 program generating the following values: genetic diversity for the population (GD=0.7832), potential genetic diversity (GD=0.9113), genic value (GV=0.7846), current founder genome equivalent (FGE=2.31) and potential p(FGE=5.64), current and potential founder genome survivor (FGS=3.14; pFGS=5.64 respectively). Other values calculated were, mean coefficient of consanguinity (F=0.0179), population effective number (Ne=5.26) and the Mean KINSHIP (MK) for each individual. With the microsatellites analysis, the allelic frequencies were calculated for each locus with an average of alleles per loci of 5.67. The averages of the observed and expected heterozigocity were 0.687 and 0.684 respectively. It was found that all loci are in Hardy-Weinberg equilibrium. For this population an excess of heterozygous was found for the locus FCA075, FCA043, FCA736, FCA220 and FCA391 and an excess of homozygous for the locus FCA 441, FCA 008, FCA 224 by Fis of Weir and Cockerham's (W and C), and Robertson and Hill's (R and H). The average of Fis for the population was (Fis W and C = -0.0157) and (Fis R and H=-0.0109) indicating that the population is in exogamy. No significant differences were found for the linkage disequilibrium test. The likelihood analyses obtained in CERVUS assigned maternities and paternities. The assignments were done according to the value Pair Top LOD. The relationship probabilities were obtained by KINSHIP. In the simulations, a significant relationship (p 0.05) were found in 6 pairs of jaguars, (p 0.01) in three pairs and (p 0.001) for 11 pairs. With this study the relationships between animals was established and the pedigree of the captive population of jaguars was built in order to develop an ex situ conservation plan for the species in the Colombian zoological parks. In the last part of the study a reproductive analysis of a captive group of jaguars (n = 6) at the Santacruz Zoological Foundation in Cundinamarca, Colombia, was conducted by performing a longitudinal, noninvasive, hormonal analysis of estradiol and progestogens in females and of androgens in males. During four seasons, female jaguars confined in solitary were evaluated for ovarian activity and spontaneous ovulation, male jaguars for testicular activity. A second hormonal follow-up was conducted in the females after administration of gonadotropins. Hormones were extracted from fecal samples of three females (n = 3) and two males (n = 2). Estradiol measurements were obtained by radioimmunoassay (RIA) and progestogens by enzyme immunoassay (EIA). The linear mixed-effect regression (LMER) showed that there was a significant effect of seasons in the concentrations of estradiol (Chi square = 15.97, DF = 3, p 0.01). Post hoc comparisons of all pairs of seasonal means were conducted according to Tukey’s honest significant difference (HSD), revealing significant differences between seasons: Dry 1 vs. Rains 2 (p 0.01), Rains 1 vs. Rains 2 (p 0.05), and Dry 2 vs. Rains 2 (p 0.05). Elevations of progestogens compatible with spontaneous ovulation occurred in three jaguars and the LMER showed that there was also a significant effect of seasons (Chi square = 28.56, DF = 3, p 0.01). Post hoc comparisons showed significant differences only between seasons: Dry 2 vs. Rains 2 (p 0.01). The season with the lowest average concentration was Rains 2 (October, November, and December). During this season, periods of anestrous were registered that lasted between 31 and 83 days. The three females presented estradiol peaks after the administration of eCG. A noninvasive longitudinal analysis for androgens was also made (Males 1 and 2) over the course of one year, and no significant differences were found between the different seasons. A seminal analysis of three adult male jaguars (Panthera onca) (n = 3) was also performed after electro-ejaculation under general anesthesia (Male 1 and 2) and by a postmortem epididymal wash (Male 3). The mean concentration of spermatozoids was 5.7 x 106 ± 1.1 x 106 spermatozoa/ml. The progressive motility + SD averaged 80 %. The percentage of normal spermatozoids obtained by electro-ejaculation was 80 ± 2.8%, and the abnormalities found more frequently were head defects (7 ± 1.4%). The seminal fluid obtained by epididymal flush contained 35 ± 1.4% normal spermatozoids, and the most frequent abnormalities found corresponded to distal cytoplasmic droplets (39 ± 11.3%). Doctorado

Trabajo Fin de Master en Medicina Veterinaria, Universidad de Lisboa, Between 1970 and 1975, 72 (36♂.36♀) Saharawi dorcas gazelles (Gazella dorcas neglecta) arrived in Finca Experimental la Hoya, Almería, southeast Spain. They were the founding individuals of a captive breeding programme, now managed as an European Endangered species Programme, currently with 236 (104.132) living individuals distributed among 12 institutions. Previous demographic and genetic assessment of this population was published in 2002 (Abáigar, 2002). An updated analysis was commended, ultimately motivated by the recent establishment of in situ reintroductions in Senegal. Besides the inclusion of the last 14 years of studbook data, this is the first thorough study to use the software programmes Population Modelling, ENDOG and SplitsTree4 for this taxon. The census history shows an overall increasing population, consistent with the incorporation of more institutions in the programme, although a tendency towards a stationary population is observed since 2010. Such tendency is corroborated by the current demographic parameters of annual growth rate (1.058), instantaneous rate of change (0.056) and net reproductive rate (1.283). Age-specific life table parameters show higher mortality rate, fecundity rate and reproductive value in males in almost all age-classes in contrast with higher survival rate and life expectancy in females. Sex and age structure of the global living population is close to pyramidal shape, as would occur in demographically stable populations. Plotting of annual gene diversity and mean inbreeding coefficient shows a positive impact of the incorporation of eight wild-caught individuals in the programme in the 90s. Current genetic parameters are the result of proper reproductive management of threatened species, and are in accordance with standardised targets when planning cooperative metapopulation breeding.

Genealogical records of animals (studbook) are created to avoid reproduction between closely related individuals, which could cause inbreeding, particularly for such endangered species as the Panthera onca (Linnaeus, 1758). Jaguar is the largest felid in the Americas and is considered an important ecological key species. In Mexico, wild jaguar populations have been significantly reduced in recent decades, and population decline typically accompany decreases in genetic variation. There is no current census of captive jaguars in Mexico, and zoos do not follow a standardized protocol in breeding programs based on genetic studies. Here, we emphasise the importance of maintaining an adequate level of genetic variation and propose the implementation of standardised studbooks for jaguars in Mexico, mainly to avoid inbreeding. In addition, achieving the aims of studbook registration would provide a population genetic characterisation that could serve as a basis for ex situ conservation programmes.

Since 1829 the African Cheetah Acinonyx jubatus has been exhibited in over 373 zoological facilities. As at 31 December 1994 the international captive population was 1218 animals of which 880 (72%) were captive-bred and 338 (28%) were wild-caught. The steady increase in the captive population is a result of captive breeding, co-operative captive-management programmes and importation from the wild. Of facilities holding Cheetah 26%. (96) have bred the species, 15% of which have bred continuously producing 63%> (n= 1580) of all cubs born in captivity. Although the number of facilities breeding Cheetah has increased, in 1994 only 10%) of them reported successful reproduction. The Ne has increased gradually and in 1994 was equivalent to 17% of the captive population. Of 1564 animals that have been imported, c. 20% (n= 308) have reproduced and in 1994 155 have living descendants in the captive population. Except for a few East African Cheetah Acinonyx jubatus raineyi all of the imported animals are the southern African subspecies Acinonyx jubatus jubatus. There has been an increase in the number of subspecific hybrids in the captive population and between 1990 and 1994 28 hybrids produced 24%, (190) of cubs. The captive population is not yet self-sustaining and is maintained by the importation of wild-caught animals. Continued progress can be achieved by implication of a co-ordinated global management programme.

The aims of this study were to verify the distribution of births of captive maned wolves Chrysocyon brachyurus and the causes of their deaths during the period from 1980 to 1998, based on the registry of births and deaths in the International Studbook for Maned Wolves. To determine birth distribution and average litter size, 361 parturitions were analyzed for the 1989-98 period. To analyze causes of mortality, the animals were divided into four groups: 1. pups born in captivity that died prior to one year of age; 2. animals born in captivity that died at more than one year of age; 3. animals captured in the wild that died at any age; and 4. all animals that died during the 1980-98 period. In group 1, the main causes of mortality were parental incompetence (67%), infectious diseases, (9%) and digestive system disorders (5%). The average mortality rate for pups was 56%. Parental incompetence was responsible for 95% of pup deaths during the first week of life. In group 2, the main causes were euthanasia (18%) and disorders of the genitourinary (10%) and digestive systems (8%). Euthanasia was implemented due to senility, congenital disorders, degenerative diseases, and trauma. In group 3, the main causes were digestive system disorders (12%), infectious diseases (10%), and lesions or accidents (10%). The main causes of mortality of maned wolves in captivity (group 4) were parental incompetence (38%), infectious diseases (9%), and digestive system disorders (7%). O objetivo deste estudo foi verificar a distribuição de nascimentos e as principais causas de mortalidade de lobos-guarás cativos, com base nos registros de nascimento e óbito no International Studbook for Maned Wolf, no período de 1980-98. Foram analisadas 361 parições, no período de 1989-98, a fim de verificar a distribuição de nascimentos e o tamanho médio das ninhadas. Os animais foram distribuídos em quatro grupos para análise das causas de mortalidade: 1. filhotes nascidos em cativeiro que morreram com menos de um ano de idade; 2. espécimes nascidos em cativeiro que morreram com mais de um ano de idade; 3. espécimes capturados na natureza que morreram com qualquer idade; e 4. lobos-guarás mortos no período de 1980-98. No grupo 1, as principais causas de mortalidade foram incompetência parental (67%), doenças infecciosas (9%) e alterações do sistema digestivo (5%). A mortalidade média de filhotes foi de 56%. A incompetência parental foi responsável por 95% das mortes dos filhotes na primeira semana de vida. No grupo 2, as principais causas foram eutanásia (18%), alterações gênito-urinárias (10%) e alterações do sistema digestivo (8%). As eutanásias ocorreram devido à senilidade, alterações congênitas, doenças degenerativas e traumas. No grupo 3, as principais causas foram alterações no aparelho digestivo (12%), doenças infecciosas (10%) e lesões e acidentes (10%). As principais causas de mortalidade de lobos-guarás em cativeiro, no período de 1980-98, foram incompetência parental (38%), doenças infecciosas (9%) e alterações do sistema digestivo (7%).

Financiadora de Estudos e Projetos Species kept in captivity are of great importance for in situ conservation of natural populations. In order to fulfill this role, they need to be properly managed with the aim to avoid the consequences of a captive environment, such as: loss of genetic diversity, inbreeding depression, accumulation of deleterious alleles and genetic adaptation to captivity (the first three due to the small number of breeding stock). Thus, with the aim to test founder effects in captive ―populations‖, we used as study model the spectacled bears from Brazilian zoos. This ―population‖ consists of twenty-one bears found in eight zoos along the country, which are originated from just twelve founding individuals. To test a possible founder effect, we analyzed twelve heterologous microsatellite loci for all the specimens of the species in Brazil. Results for the number of alleles and allelic richness obtained through the rarefaction method showed that spectacled bears in captivity have reduced levels of allelic richness in relation to three natural populations (Colombia, Ecuador and Venezuela) studied by other authors; suggesting that bears from Brazilian zoos suffered the consequences of a founding effect, as they have only a fraction of the gene pool of the ancestral population. From the information present in the studbook was possible to verify that there are two families of spectacled bears, and according to the individuals´s multilocus genotypes, we found alleles that are present in a same family and absent in the other. Regarding the kinship, there was a discrepancy in the results with the molecular estimators through the softwares ML-Relate and Coancestry and the information given by the institutions. The information obtained in this work are fundamental to the ex situ conservation program in Brazil with the objective of maintain as high as possible the genetic diversity for future reintroduction plans of the species in nature. Espécies mantidas em cativeiro são de grande importância na conservação in situ de populações naturais. A fim de que possam desempenhar este papel é necessário que sejam devidamente manejadas, para que as consequências que o cativeiro traz sejam evitadas, como por exemplo: perda de diversidade genética, depressão endogâmica, acúmulo de alelos deletérios e adaptação genética ao cativeiro (as três primeiras devido ao pequeno número de espécimes dos plantéis). No Brasil, uma espécie de mamífero que vem sendo criada em cativeiro desde 1970 e que passou por um efeito fundador, é o urso de óculos (Tremarctos ornatus). O plantel existente hoje totaliza o número de vinte e um ursos, encontrados em oito zoológicos do país, o qual se originou de apenas doze indivíduos fundadores. Para avaliar as consequências de um conhecido efeito fundador, analisamos doze locos heterólogos de microssatélites para todos os exemplares da espécie nos zoos do país, através de amostragem não-invasiva. Resultados referentes ao número de alelos e à riqueza alélica obtida por meio do método de rarefação mostraram que os ursos de óculos de cativeiro possuem valores reduzidos de riqueza alélica em relação a três populações naturais (Colômbia, Equador e Venezuela) estudadas por outros autores; sugerindo que os ursos dos zoológicos brasileiros sofreram as consequências de um efeito fundador, pois possuem apenas uma fração do pool gênico da população ancestral. A partir das informações presentes no studbook foi possível verificar que existem duas famílias de ursos de óculos, e de acordo com os genótipos multilocos dos indivíduos, encontramos alelos que estão presentes em uma mesma família e ausentes na outra. Em relação ao parentesco, houve uma discrepância nos resultados encontrados com os estimadores moleculares através dos softwares ML-Relate e Coancestry e o histórico cedido pelas instituições. As informações obtidas em nosso trabalho, portanto, são fundamentais para que os ursos encontrados nos zoológicos do Brasil possam participar na formação de casais em futuros programas de reprodução ex situ com o propósito de serem utilizados em planos de reintrodução da espécie na natureza.

The last representatives of the Barbary lion\ud (Panthera leo leo), once numerous in North Africa but\ud exterminated from the wild by the 1940s, are believed to be\ud the captive lions descended from the Moroccan Royal\ud Collection, numbering less than 90 animals in zoos\ud worldwide. The genetic fitness of these captive “Royal\ud Lions” may now be under threat since, although most zoos\ud have avoided hybridisation with animals of other origin, no\ud formal breeding programme currently exists and several\ud institutions have halted breeding activities. This situation\ud has arisen since the distinctiveness of Barbary lions and the\ud representative status of Royal Lions remain inconclusive\ud and definitive molecular studies have yet to be completed.\ud Previously, in the 1970s, morphological and phenotypic\ud traits were used to match Royal Lions and the historic\ud Barbary lion and an ex situ breeding programme was\ud initiated involving a number of selected “founder” animals.\ud This paper outlines the status of the descendent population\ud within zoos in Morocco and Europe, including all known\ud pure-bred descendents from the Royal Palace collection.\ud Founder representation is shown to be greater across\ud European collections than the Moroccan collection. Breeding\ud exchanges are recommended between institutions in\ud order to improve genetic diversity and maintain the genetic\ud health of the population and a studbook for European zoo\ud animals has been developed to support this action. This\ud analysis serves as a benchmark for guiding effective\ud maintenance of the captive population, thereby allowing\ud time to clarify the conservation value of Royal Lions and\ud their relevance to North African ecology.

The African cheetah has been bred in North American zoological facilities since 1956. The captive population has since grown to around 200 animals because of a combined increase in importation plus captive births. From 1982 to 1986, the captive birth rate declined by 50%, primarily because of a low frequency of breeding individuals in the population. The 1986 population had an effective breeding size of 28.1 in a total population of over 193 cheetahs. The incidence of infant mortality has been high (36.7%) relative to other zoo-bred species, perhaps as a consequence of the previously observed genetic impoverishment of the species. The combination of low fecundity, high infant mortality, and population dynamics indicates that the North American captive cheetah population is neither a self-sustaining nor a theoretically “viable population” as defined by Soule et al. [ZOO BIOLOGY 5:101–114, 1986]. Possible recommendations for improving captive cheetah propagation are discussed.

日本国内で1931年～2012年までに飼育されたチーターAcinonyx jubatusの繁殖傾向と生存状況を調査するために,国際血統登録「International Cheetah Studbook 2012」とチーター国内血統登録を用いて分析を行った。16施設(現在飼育施設9施設)で548頭が飼育され,そのうち314頭は,雄36頭,雌42頭から産まれた。これらの繁殖個体の繁殖時の年齢について,野生個体,海外個体,国内個体間で統計的な有意差が確認された(p, We investigated the reproductive trend and survival of cheetah Acinonyx jubatus, reared between 1931 and 2012 in Japan. We analyzed the "International Cheetah Studbook 2012" and the "Cheetah Internal Studbook" for the history of cheetahs reared in Japan, population movement, breeding changes, age at death, rearing and breeding results at each animal-breeding facility, and breeding results when females were reared at the same place and for the same period. A total of 548 animals were bred in 16 facilities (9 facilities at present), and 314 of them were produced in Japan. The offspring were born from 36 males and 42 females. A significant difference (p