Department of Genetics

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Browsing Department of Genetics by browse.metadata.advisor "Bester-van der Merwe, Aletta Elizabeth"

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    The evolutionary history of the genus Seriola and the phylogeography and genetic diversity of S. lalandi (yellowtail) across its distribution range
    (Stellenbosch : Stellenbosch University, 2014-04) Swart, Belinda Louisa; Roodt-Wilding, Rouvay; Bester-van der Merwe, Aletta Elizabeth; Von der Heyden, Sophie; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: The genus Seriola includes several important commercial fish species, yet the phylogenetic relationships between species have not been fully investigated to date. This study reports the first molecular phylogeny for this genus based on two mitochondrial (Cytb and COI) and two nuclear gene (RAG1 and Rhod) fragments for all extant Seriola species (nine species, n = 27). The phylogenetic patterns resolved three main lineages: a ((S. fasciata and S. peruana), S. carpenteri) clade, a (S. dumerili and S. rivoliana) clade and a (S. lalandi and S. quinqueradiata) clade. The closure of the Tethys Sea (12 - 20 MYA) coincides with divergence of the ((S. fasciata and S. peruana), S. carpenteri) clade from the rest of the Seriola species; while the uplifting of the Isthmus of Panama (± 3 MYA) played an important role in speciation between S. fasciata and S. peruana. The climate and water temperature fluctuation in the Pliocene played important roles during the divergence of the remainder of the Seriola species. This study is also the first to describe the evolutionary history of the commercially important species Seriola lalandi across its distribution range. Global patterns of genetic variation within S. lalandi (n = 190) were examined using three genes fragments (mitochondrial DNA COI, Cytb and nuclear RAG1). Three distinct clades were identified, corresponding to three different geographic regions (North-western Pacific - Japan, North-eastern Pacific - USA, and the southern hemisphere clade). These groupings correspond with the previously identified subspecies of S. lalandi (North-western Pacific – S. lalandi aureovittata, North-eastern Pacific – S. lalandi dorsalis, and the southern hemisphere clade - S. lalandi lalandi). AMOVA results and pairwise FST values revealed significant population differentiation between these groups. The population subdivision between these clades in all probability is maintained by biogeographic or oceanographic barriers (such as the equator and East Pacific Barrier) that disrupt gene flow. The southern hemisphere clade comprised of samples from the southern Pacific (AUS, NZL and Chile) and the southern Atlantic (SA). No haplotypes were shared between these areas in the southern hemisphere. This southern hemisphere clade was further investigated with six microsatellite markers. The analyses revealed the South African populations as genetically distinct from populations of the South Pacific oceans (AMOVA, FCA and STRUCTURE results). In summary, the South African and southern Pacific grouping could be the result of recent vicariant events during the Pleistocene glacial / interglacial periods and / or contemporary oceanographic forces acting on these populations. Further population differentiation was found within the South African samples, but not in the South Pacific. In the southern Pacific clade this lack of population structure is the result of high gene flow (analysed with MIGRATE) between the sampling localities. This is the first report on the genetic structure of this commercial important species for South African populations. Five sampling localities from the west- to the east coast of South Africa were sampled (n = 201). The microsatellite analyses revealed two potentially genetically distinct groups. AMOVA, FST and FCA results suggest small but significant differentiation between populations from the west coast and from the south- and east coast, suggesting a potential genetic break in the Cape Point region (BARRIER). However, the program STRUCTURE showed a high level of admixture along the South African coast and the migration results (MIGRATE and BAYESASS) also suggest a high degree of gene flow between these regions.
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    Genetic diversity and mating systems in a mass-reared black soldier fly (Hermetia illucens) population
    (Stellenbosch : Stellenbosch University, 2021-03) Hoffmann, Lelanie; Rhode, Clint; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Black soldier flies are gaining popularity as an alternative source of protein in animal feed. They have a high feed conversion ratio and can be reared on biowaste, reducing the energy input required for mass-rearing. As the number of mass-reared colonies is increasing worldwide, the importance of genetic management in commercial populations is becoming clear. This study aimed to determine the effects of domestication and mating behaviour on the genetic diversity of a mass-reared black soldier fly population. Eight microsatellite markers were used to estimate genetic diversity in two temporally separated samples of a wild black soldier fly colony (Wild2015 and Wild2018) and three distinct generations of a mass-reared black soldier fly colony (F28, F48 and F52). Diversity estimates decreased significantly in the mass-reared colony over time, when compared to the two wild samples. The mass-reared colony also saw an increase in relatedness over time, with a relatedness coefficient as high as 0.430 in generation F48. These results indicate severe inbreeding in the mass-reared colony. Effective population sizes of between 22.6 and 59.0 in the mass-reared colony are also a cause for concern, as populations with low effective population sizes are more vulnerable to inbreeding depression and extinction. The high levels of genetic diversity observed in the two wild samples provide the potential for the wild colony to become a donor population, providing immigrants to introduce genetic diversity into the mass-reared colony. However, based on FST estimates, the two populations appear to be diverging from each other over time. Moderate differentiation was observed between Wild2015 and F28 (FST=0.062; p=0.000), while great differentiation was observed between Wild2018 and F52 (FST=0.161; p=0.000). To minimise the risk of outbreeding depression, the compatibility of wild individuals with the artificial environment would therefore need to be tested before immigrants are introduced into the mass-reared population. To study the mating behaviour of the black soldier fly, five mating pairs were randomly sampled in copula from generation F48 of the mass-reared colony. All candidate parents and 25 offspring from each clutch were genotyped and subjected to parentage analysis. Multiple paternity was detected in two of the five families, providing evidence for polyandry. This was a novel finding, as observation had previously led to the hypothesis that this species is monogamous. The occurrence of polyandrous mating provides evidence that adult flies can mate multiply despite being unable to replenish energy through feeding, thereby creating the potential for polygynous mating. Additionally, polyandrous mating has positive implications for the genetic management of commercial black soldier fly populations. However, these results are limited to mass-reared colonies, as the higher population densities found in captive populations increase the probability of remating. Finally, diversity estimates and inbreeding estimates were calculated for the candidate parents, offspring, and the population the parents were sourced from. Individual parent pairs showed increased levels of relatedness when compared to the source population, indicating positive assortative mating. As markers from random genomic regions were used for this study, the observed increase in relatedness may provide additional evidence for inbreeding in the mass-reared population. However, inbred populations show greater genome wide linkage disequilibrium, meaning that mate selection for desirable traits could potentially be detected in markers not directly related to traits of interest.
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    Genetic diversity and population genetic structure in the South African commercially important shark species, the common smoothhound (Mustelus mustelus)
    (Stellenbosch : Stellenbosch University, 2014-12) Maduna, Simo Njabulo; Bester-van der Merwe, Aletta Elizabeth; Roodt-Wilding, Rouvay; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Deciphering patterns of intraspecies population genetic structuring in commercially important shark species is essential for an integrated fisheries management approach to conservation of regional biodiversity. The common smoothhound shark, Mustelus mustelus, is an overexploited, commercially and recreationally important shark species in South Africa. Considering the vulnerable status of the common smoothhound shark and due to very limited available genetic information, this study aimed to develop molecular markers, assess patterns of genetic diversity and population connectivity along the South African coast using multilocus data generated from 12 microsatellite markers and the mitochondrial gene, NADH dehydrogenase subunit 4 (ND4). The cross-species amplification of microsatellites proved useful for genetic diversity and population genetic analysis of the common smoothhound shark. These microsatellites could aid in the molecular characterisation of other endemic and cosmopolitan species and provide valuable tools for the conservation of potentially threatened or exploited shark species. For the microsatellite data, moderate levels of genetic diversity based on the heterozygosity, allelic richness and haplotype diversity were found in a total of 144 individuals sampled across eight study populations. Estimates for pairwise population differentiation, F-statistics, AMOVA and factorial correspondence analysis (FCA) indicated significant genetic structure within and between west- and east coast populations. Additionally, Bayesian clustering analyses detected two putative ancestral gene pools, supporting the presence of a biogeographic barrier at the Cape Agulhas region and therefore genetic discontinuity between the Indian and Atlantic Ocean samples. On the contrary, mitochondrial data indicated that common smoothhound shark is genetically homogenous with substantial interoceanic gene flow. Such conflicting signals found between nuclear and mitochondrial DNA (mitonuclear discordance) can be attributed to a number of factors and could simply be due to the inherent differences in marker properties or an indication of sex biased dispersal. Despite an indication of an expanding common smoothhound shark population based on both marker types, a contemporary genetic bottleneck may have gone undetected as genetic divergence was very low in some of the study populations. Nonetheless, contemporary restriction to gene flow and historical demographics such as range expansion are proposed as the most likely forces explaining genetic structure in present-day common smoothhound sharks in South Africa. For future sustainable exploitation of common smoothhound shark, the possible existence of two genetically differentiated populations and observed asymmetric gene flow along the South African coast should be taken into consideration. It is also recommended that in the future further evaluations of finescale genetic structure and seasonal migration patterns in this commercially important species are conducted in order to allow integration of this knowledge into existing fisheries management practices.
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    Genetic diversity in a commercial black soldier fly, Hermetia illucens (Diptera: Stratiomydiae), population
    (Stellenbosch : Stellenbosch University, 2017-03) Badenhorst, Rozane; Rhode, Clint; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: The applications of black soldier fly (BSF), Hermetia illucens, as a first-generation biotechnology address two global problems - the pressure on food security arising from unsustainable food production practices which exhausting fisheries resources by fishmeal production; and the vast accumulation of organic waste. Understanding the phenotypic and genetic changes experienced when establishing colonies of H. illucens is important for successfully establishing new colonies and managing already existing mass-rearing facilities. The aim of this study was to assess the changes in genetic diversity coupled with the phenotypic changes associated with colony establishment. Commercially-relevant phenotypic parameters were measured; including pupae weight, percentage eclosion, post-mating longevity, hatchability and egg clutch sizes over six successive generations (F0-F5). Hatchability, clutch size and pupae weight increased from F1 to F4, but a decline was apparent from F3 to F4. Post-mating longevity dramatically decreased from F1 (100%) to F4 (33%) and the F5 generation did not develop beyond the pre-pupae stage resulting in colony-collapse. The phenotypic changes can be explained by a combination of environmental, physiological and genetic effects. The positive trends observed during earlier generations (F0-F1) may reflect the population reacting to relaxed natural selection. While, at the end of the study period (F3-F5), natural selection in captivity resulted in a decrease in phenotypic variables. The colony collapse in the final generation, despite constant environmental conditions, possibly reflects the deleterious effects of random genetic drift and inbreeding depression. For the assessment of the genetic processes which may have resulted in the observed phenotypic changes, ten microsatellite markers were developed and characterised. Marker utility was tested on 37 wild individuals using three multiplex-PCR reactions. The number of alleles for each locus ranged from 4 to 21. Polymorphism information content ranged from 0.52 to 0.90, while observed and expected heterozygosity ranged from 0.30-0.65 and 0.55-0.91, respectively. Furthermore, the markers are useful in individual identification (PID = 1.2x10-11and PIDsib = 1.1x10-4) and parentage analysis (P1=1; P2 = 1). The markers proved useful in the assessment of genetic diversity and were used to understand the genetic mechanisms underlying phenotypic trends previously described. The assessment of genetic diversity revealed significant population differentiation as described by FST estimates across the experimental generations (P < 0.01), accompanied by a significant loss of genetic diversity. The final generation indicated a significantly higher number of private alleles and significantly lower observed heterozygosity in comparison to preceding generations (P < 0.05). Population bottlenecks and inbreeding were evident in the estimation of effective population size (Ne) and relatedness. Phenotypic parameters which significantly correlated (Pearson’s r) to changes in genetic diversity were pupation (%), clutch size, pupae weight and female post-mating longevity (P < 0.05). Results indicate the effects of inbreeding depression and genetic drift were amplified by a decrease in effective population size and increased relatedness amongst individuals. In summation, the assessment of changes in phenotypic measurements and genetic diversity during the establishment of an H. illucens colony, gave valuable insight into the genetic processes which occur during colony establishment; and are applicable to colony establishment and the management of already existing colonies.
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    Mapping of dwarf growth habit traits in apple (Malus pumila Mill.) using molecular markers and transcriptomics approaches
    (Stellenbosch : Stellenbosch University, 2020-03) Mbulawa, Zama Thandekile Laureen; Bester-van der Merwe, Aletta Elizabeth; Kriel, Johan; Tobutt, Kenneth R.; Stellenbosch University. Faculty of Agrisciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Apple (Malus pumila Mill.) is one of the most important deciduous fruit crops worldwide. Apples are traditionally valued as an important dietary source of fibre and are high in antioxidants, contributing to human nutrition. In South Africa, the apple industry plays a vital role in the country's agricultural economy due to global exports. In recent years, more emphasis has been directed to dwarf trees, as they are well suited for profitable high-density orchards and sustainability of fruit production. However, dwarfism cannot always be linked to increased yield. At Bien Donné Research Farm of the Agricultural Research Council (ARC) Infruitec-Nietvoorbij’, several dwarf growth habits exist, which is related to a form of hybrid incompatibility, hybrid necrosis. One of them is associated with undesirable characteristics such as crinkled leaves and poor growth. Expression of hybrid necrosis in plants can lead to a significant reduction in productivity, due to the deleterious epistatic interactions between alleles that arose from divergent genetic backgrounds. Few, if any, genetic studies have thus far investigated crinkle dwarf growth traits in apple. This study aimed to examine the genetic basis underlying the crinkle dwarf phenotype by employing multidisciplinary approaches that included segregation pattern studies, assessment of self-incompatibility (hybrid incompatibility), molecular mapping and transcriptomic profiling of pooled samples of apical buds and young leaves from normal and from crinkle dwarf phenotypes. The genetics behind the crinkle dwarf trait was undertaken by studying the segregation patterns of the first filial generation (F1) apple progenies, where parental combinations were heterozygous. Segregation ratios of 9:7 and 3:1 were observed, for which crinkled dwarf phenotypes is expressed when one of the two genes is homozygous recessive (D-ee or ddE-). Additionally, the involvement of self-incompatibility (S) was investigated by identifying the parental S-genotypes using PCR based consensus and allele-specific primers of the apple S-RNAse gene. Eight parental S-genotypes were determined. Herein, the S-genotypes of Malling 1 (‘M.1’) (S3S9) and TSR1T187 (S7S24) were deduced for the first time. High-density SNP-based parental genetic linkage maps of ‘McIntosh’ and ‘M.1’ were constructed using the apple 20K Infinium SNP array. The crinkle dwarf trait was mapped on linkage group (LG) 8 in ‘McIntosh’ and on LG2 in ‘M.1’. In the consensus genetic map, crinkle dwarf trait also mapped on LG8. Additionally, the crinkle dwarf trait obtained for the parental genetic maps were validated using Kruskal-Wallis (KW) analysis. To gain deeper insights into the genes regulating crinkle dwarf phenotype, transcriptome profiles of pooled meristematic tissues of normal and crinkle dwarf phenotypes were generated using RNA-sequencing technology. A total of 921 significantly differentially expressed genes (DEGs), with 763 up-regulated and 158 down-regulated transcripts, were identified. Gene expression analyses revealed that defense signaling and stress-related genes were up-regulated during the expression of crinkle dwarf phenotype along with the activation of several antioxidant proteins/enzymes. The high expression of lactoperoxidase (Class III peroxidase) together with glutathione S-transferase suggests the involvement of reactive oxygen species (ROS). Genes typically encoding for pathogenesis-related proteins (chitinase and pectin), antioxidant enzymes, receptor-like protein (protein serine/threonine phosphatase), as well as alpha-linolenic acid, a precursor of the phytohormone jasmonic acid were all up-regulated during expression of crinkle dwarf phenotype. These findings support the notion that crinkle dwarf phenotype does indeed exhibit hybrid necrosis symptoms. Consequently, an autoimmune response might have been triggered by the allele incompatibilities, in this case between ‘McIntosh’ and ‘M.1’. Overall, the information generated in this study will aid in designing an in-house screening system for eliminating seedlings carrying crinkle dwarf genes from the ARC breeding material. In future, these findings will also aid in the design of crosses with predictable outcomes and in broadening a sustainable genetic base of the apple cultivars for high productivity orchards, while avoiding raising seedlings with dwarf growth habit associated with crinkled leaves.
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    Marine forensics : a molecular tool for trade monitoring and compliance in southern African fisheries, with focus on commercially exploited elasmobranch species
    (Stellenbosch : Stellenbosch University, 2019-12) Asbury, Tamaryn Anne; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Marine forensic science can be described as protecting fisheries resources, marine mammals and endangered species based on enforcement of the nation's laws. Species identification becomes challenging when morphological features (such as fins, scales and heads) are removed, or if confiscated samples are already in a processed state. The harvesting of elasmobranchs (sharks, rays and skates) is driven by the international shark meat and shark fin trade. In recent decades, a combination of increasing demand and economic globalisation has created a global market for elasmobranch products, especially the highly prized shark fins for Asian markets. In this study, marine forensics was assessed as a tool for complementing traditional identification methods – through the development of a mini-barcoding assay as well as investigating High Resolution Melting (HRM) analysis – for species identification, with focus on elasmobranch species occurring in the southern African region. Firstly, this involved the testing and optimisation of the standard barcoding region of the cytochrome oxidase c subunit I (COI) gene, and then using traditional barcoding primers as well as nested polymerase chain reaction (PCR) primers in a multiplex assay. Preliminary results (only a 22 % species identification success rate) indicated the limitations of using only the traditional COI primers and warranted the inclusion of alternate COI gene fragments for species identification in future related forensic cases. A mini-barcoding multiplex assay, comprising three primer sets, was optimised and applied to a wide range of forensic case studies involving confiscated shark fins, possibly for illegal trade. A significant number of CITES-listed and endangered species were identified when confiscated specimens from various regions in southern Africa were tested. Secondly, PCR amplification of a 16S ribosomal RNA (16S rRNA) gene fragment was optimised based on six southern African houndshark species and seven other commercially exploited species, including: hammerhead sharks Sphyrna lewini and Sphyrna zygaena, copper shark Carcharhinus brachyurus, dusky shark Carcharhinus obscurus, bull shark Carcharhinus leucas, blacktip shark Carcharhinus limbatus and blue shark Prionace glauca. High Resolution Melting analysis using the 16S rRNA gene region was investigated as a species identification method for these species. The HRM assay was successfully applied for the identification of seven commercially exploited shark species, including some of the top commercially important sharks and one endemic houndshark Scylliogaleus quecketti. Although further optimisation is required, this relatively fast and cost-effective approach will be a valuable tool for the initial screening of detained shipments, for possible illicit trade. Accordingly, this research presents species identification assays suitable for various shark related forensic case studies, and in future could be applied to identify most, if not all, elasmobranch species involved in trade regionally.
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    Medium-throughput SNP genotyping and linkage mapping in Haliotis midae
    (Stellenbosch : Stellenbosch University, 2012-12) Du Plessis, Jana; Roodt-Wilding, Rouvay; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Haliotis midae (locally also known as perlemoen) is the largest of five endemic species found along the coast of South Africa. It is the only species with commercial value contributing to the exploitation of these animals. Due to declines of natural stocks, farming practices were established during the early 1990s in order to supply the international demand. To facilitate efficient breeding methods and ensure the sustainability of these commercial populations, genetic management, which can be accomplished with the use of molecular markers such as single nucleotide polymorphisms (SNPs), is necessary. Single nucleotide polymorphisms have become the markers of choice in various applications in aquaculture genetics due to their abundance in genomes, reduction in developmental costs and increased throughput of genotyping assays. Identification of SNPs in non-model species such as H. midae can be achieved by in silico approaches. In silico methods are suitable for de novo SNP identification and are both cost- and time-efficient. It is based on the analysis of multiple alignments where mismatches may be reported as candidate SNPs. Various medium-throughput genotyping methods are available to confirm putative SNPs, but the ideal method depends on factors such as cost, accuracy and multiplexing capacity. Although SNP markers can have various applications within the aquaculture environment the focus for this current study was saturating the linkage map of H. midae with additional markers. This would assist in the identification of quantitative trait loci associated with economically important traits, which in turn could ultimately be employed for marker-assisted selection and improved molecular breeding programs. In order to identify in silico SNPs, sequenced transcriptome data from a previous study was used and subjected to a series of criteria: minor allele frequency 10%, minimum coverage 80, 60 bp flanking regions. Selected loci were genotyped using a 192-plex assay with the Illumina GoldenGate genotyping assay with the VeraCode technology on the BeadXpress platform, in individuals from six mapping families. A conversion rate of 69.35% and global success rate of 76.34% was achieved. Polymorphic loci were subjected to linkage analysis using JoinMap® v.4.1 to create sex-average and sex-specific maps and to saturate the current linkage map for H. midae. Along with previously developed markers, 54% of the newly developed SNPs could be successfully incorporated into the linkage map of H. midae. A total of 18 linkage groups were observed with an average marker spacing of 6.9 cM and genome coverage of 79.1%. Bioinformatic analyses and setting stringent criteria to identify SNPs from sequenced transcriptomic data proved to be an efficient way for SNP discovery in the current study. Genotyping of the identified loci with the GoldenGate genotyping assay demonstrated a high success rate; providing a genotyping assay adequate for species with little genomic information. The linkage map created in this study illustrated the utility of SNP markers in conjunction with microsatellite markers for linkage map construction and the adequate marker spacing obtained provides a step closer to quantitative trait loci mapping in this species.
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    Molecular aspects of exploited sharks in South Africa : multiple paternity and identification of novel molecular markers
    (Stellenbosch : Stellenbosch University, 2015-12) Rossouw, Charne; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of Science. Dept. of Genetics.
    ENGLISH ABSTRACT: Sharks have existed for the past 400 million years and play an important role in the oceanic ecosystem as they occupy the upper categories of the food web. Since the 1920s they have been fished on a commercial scale as the demand for shark related products increased. This placed considerable pressure on shark populations, resulting in a global decline of many populations and an increased risk of population extinction. As the genetic diversity of a population determines it’s resilience to changing environmental factors, including such information has become paramount for short and long-term management and conservation of individual species. This study therefore aimed to add to the growing body of biological and genetic data by investigating mating strategy through assessing the presence of multiple paternity (MP) in three commercially important shark species: the common smoothhound Mustelus mustelus, dusky shark Carcharhinus obscurus and the scalloped hammerhead shark Sphyrna lewini; and by identifying potential microsatellite and single nucleotide polymorphism (SNP) markers in two of the species, M. mustelus and C. obscurus, through the use of Next-Generation Sequencing (NGS) platforms. The first aim of this study was achieved by cross-amplifying microsatellite markers developed in closely related species to the study species. A total of 22 microsatellite markers were initially tested on four litters of M. mustelus and C. obscurus in order to determine the most informative markers for parentage analysis. Reduced marker panels of five to six microsatellites were selected and parentage analysis in GERUD and COLONY revealed the presence of MP in all three species. Mustelus mustelus had the highest frequency of MP (67%), followed by S. lewini (46%) and C. obscurus (35%). The second aim of this study entailed reduced genome sequencing of one M. mustelus and one C. obscurus individual using the HiSeq Illumina and Ion Proton platforms, respectively. For M. mustelus, 51,5 million reads with an average read length of 250bp were obtained, whereas C. obscurus yielded 27,6 million reads with an average read length of 213bp. Contigs were constructed for both species in order to search for perfect repeat motifs. In total, 2 700 and 1 255 microsatellite-containing regions were identified for M. mustelus and C. obscurus respectively. In order to search for SNP-containing regions, both sample species were aligned to previously assembled scaffolds of the ghost shark Callorhinchus milii, which served as a reference genome. After quality filtering, only 767 SNP-containing regions were identified for M. mustelus, whereas the identification of potential SNPs for C. obscurus was not successful. The insights gained into the mating strategies of M. mustelus, C. obscurus and S. lewini as well as the identification of potential species-specific molecular markers add to the growing body of information and genetic resources available for exploited species. In future, this information could be used for further molecular assessment of shark populations and a more intergrated approach to conservation and management of these already vulnerable sharks.
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    Molecular species identification and spatio-temporal assessment of genetic diversity in the smooth hammerhead shark Sphyrna zygaena in South Africa
    (Stellenbosch : Stellenbosch University, 2017-03) Kuguru, Gibbs; Bester-van der Merwe, Aletta Elizabeth; Rhode, Clint; Gennari, E.; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: The South African coast hosts a unique oceanographic regime with an enriched habitat able to support a diverse biota of chondrichthyans (sharks, skates, rays and chimaeras). Investigating these species and populations on a molecular level could aid in conserving this rich chondrichthyan biodiversity. As a precursor, a case study regarding the composition of species in three different South African fisheries was evaluated to determine the utility of the mitochondrial cytochrome oxidase subunit 1 (CO1) gene in delimiting species identity. Through this, a number of issues surrounding misidentification and cryptic speciation were recognized, and the efficacy of CO1 was tested and proved to be useful in identifying chondrichthyans affected in South African fisheries. One of these species, the smooth hammerhead shark (Sphyrna zygaena) displays a high degree of site fidelity to Mossel Bay as evidenced by the rise in the number of neonate and juvenile hammerheads during the summer season. This species is vulnerable as they are in danger of overfishing and the destruction of their natural habitat. With a low fecundity and a long generational time, recovery of near-depleted populations is prolonged. In this study mitochondrial sequence data and microsatellite markers were used to assess genetic diversity within and between S. zygaena sampling cohorts collected from Mossel Bay to the KwaZulu Natal coast. Additionally, kinship between the juvenile individuals was determined and parental genotypes were reconstructed from the neonate and juvenile smooth hammerhead sharks sampled in the Mossel Bay area. Significant population subdivision was evident between individuals sampled in the warm temperate south coast (Mosselbay) and the subtropical east coast (Algoa Bay and KwaZulu Natal), with asymmetric gene flow mainly from the south to the east coast. Highly significant population differentiation was seen between sampling years, indicative of differential temporal stocks utilizing Mossel Bay each year. Analysis of kinship revealed a high degree of sibling relationships within and between seasons, which is likely due to an overlap of some parental genotypes across seasons. The results obtained here can assist with decisions regarding the conservation of chondrichthyan biodiversity in South Africa while it is also recommended that genetic structure and temporal variation of S. zygaena populations be evaluated on a finer scale in the future.
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    Population dynamics of two endemic catshark species inferred from molecular and tag-recapture data
    (Stellenbosch : Stellenbosch University, 2023-03) Hubble, Emma Rebecca; Klein, Juliana Dunia; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of Agrisciences. Dept. of Genetics.
    ENGLISH ABSTRACT: The endemic catshark genus, Poroderma, consists of two closely related species, the pyjama catshark Poroderma africanum and the leopard catshark Poroderma pantherinum. They are found within the southern African oceans and are threatened by overexploitation due to being a bycatch product of commercial fisheries. In recent years, this previously understudied genus has gained more attention for their role as important meso-predator species within the South African marine environment. There is a lack of information regarding the movement patterns and population dynamics of these catsharks and the effective management and conservation of these sharks remains a huge challenge. This study aimed at addressing these knowledge gaps using a multifaceted approach; 1) molecular markers to infer the genetic variation within and between species, and 2) a long-term tag-recapture dataset from 1984 to 2020, to investigate the habitat usage, movement patterns and site fidelity of both species. Successful cross-amplification of microsatellite markers from the small-spotted catshark Scyliorhinus canicula was done to complement the de novo-developed Poroderma microsatellite markers. Traditional population genetics analyses (F-statistics and AMOVA) of nuclear data did not reveal any significant intraspecific genetic differentiation (P > 0.05) while multivariate (Discriminant Analysis of Principal Components) and Bayesian clustering analyses also indicated a high level of gene flow between populations i.e. very weak intraspecific genetic differentiation for both species. Nonetheless, all analyses indicated clear genetic differentiation between species, with a slightly higher level of genetic diversity and clinal structure observed in Poroderma pantherinum. Each species was assigned to distinct genetic clusters based on the nuclear data and there were no mitochondrial (NADH-2) haplotypes shared between the species. The presence of only two mutation events between the major haplotypes for each species highlighted the low sequence divergence that has previously been noted for these catsharks. Regarding their movement patterns, a high overall recapture rate was observed (10.36%) for both species and high site fidelity was confirmed for this genus since 72.52% of all tag-recaptures occurred in the same location as the tagging. The tag- recapture data indicated that several individuals from both species appeared to travel further distances (>30 km) than other endemic catsharks, which may result in the notable lack of genetic differentiation and population structure. Investigation into the utility of seasonal or no-take marine protected areas for these catsharks could be worthwhile as it would reduce the vulnerability of these species to the fishing trade and allow their unique habitats to recover from previous fishing pressure. Despite the lack of genetic differentiation, the high site fidelity means that there are specific locations that play a crucial role in the ecology and reproduction of these catsharks. Accordingly, the protection and management of these critical habitats could help prevent these species from becoming even more threatened.
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    Population genetics of Galeorhinus galeus, Carcharhinus brachyurus and Rhinobatos annulatus- implications for regional fisheries and elasmobranch conservation
    (Stellenbosch : Stellenbosch University, 2016-03) Bitalo, Daphne Nyachaki; Bester-van der Merwe, Aletta Elizabeth; Roodt-Wilding, Rouvay; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Elasmobranchs (sharks, skates and rays) are highly exploited world-wide and more vulnerable than most teleosts due to their life history traits (e.g. late age at maturity, low fecundity and slow growth). Most elasmobranchs are either targeted by commercial fisheries or unintentionally taken as bycatch in mixed-species fisheries. Among these, the tope shark Galeorhinus galeus, the copper shark Carcharhinus brachyurus and the southern African endemic lesser sandshark Rhinobatos annulatus, are targeted globally and locally in demersal, pelagic and recreational fisheries. Across the Southern Hemisphere, the International Union for the Conservation of Nature (IUCN) categorizes both the tope and copper sharks as “vulnerable” while the lesser sandshark as “data deficient” within its region of endemism. Information is urgently needed on their regional genetic structure and diversity to help delineate management units (MUs) for better fisheries monitoring and conserving local biodiversity. Regional and local population genetic structure of these species was assessed using previously optimised cross-species microsatellite panels and/or the mitochondrial NADH2 and NADH4 genes. Patterns of evolutionary and demographic history were inferred using coalescent and Bayesian statistical methods. For G. galeus, the data showed a lack of contemporary gene flow and deep historical divergence across the Southern Hemisphere. Two geographically distinct mitochondrial clades were recovered, one including the Atlantic and Indo-Pacific collections (ARG, SA and AUS) and one comprising the Pacific samples (NZ and CHI) as well as single divergent haplotype restricted to South Africa. Nuclear data also revealed large population subdivisions (FST = 0.050 to 0.333, P < 0.05) indicating very limited gene flow for tope sharks across ocean basins. On a local scale, F-statistics, multivariate and clustering analyses supported gene flow with substantial admixture along the South African coastline (FST = 0.016 to 0.048, P > 0.05), with some degree of genetic structure between the Atlantic and Indian Ocean samples. The east coast samples of Port Elizabeth were significantly differentiated from the rest (FST = 0.023 to 0.091, P > 0.05). For C. brachyurus, estimates of pairwise population differentiation were significant (average FST = 0.031, P = 0.000) indicating some degree of gene flow between sampling sites while the sub-structuring observed at Strandfontein indicated the existence of a possible distinct, more admixed group of individuals. Neither AMOVA (FCT = -0.011, P = 1.000) nor Bayesian clustering analyses indicated genetic discontinuity or significant population structure across the Atlantic/Indian boundary. Although the ND4 results also alluded to historical dispersal across this boundary, the population of Mossel Bay harboured four highly divergent haplotypes, indicating that this region might be a potential nursery site for C. brachyurus. The genetic diversity and genetic connectivity of R. annulatus was inferred using cross-amplified polymorphic microsatellite loci across the Agulhas bioregion that coincides with the warm temperate biogeographical province of South Africa. Significant genetic differentiation was observed over a small sampling range (FST = 0.016 to 0.094, P < 0.050) implying that the species might be highly structured throughout its entire geographical range. Overall effective population size for R. annulatus was very low (Ne = 106) and not in accordance to the abundance proposed for the species. As this is the first regional assessment for all three of these species, the findings of this study could have immediate implications for the regional management and conservation of commercial and recreational sharks.
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    Signatures of selection in natural and cultured Abalone (Haliotis midae) : a population genomics study
    (Stellenbosch : Stellenbosch University, 2013-03) Rhode, Clint; Roodt-Wilding, R.; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of Science. Dept. of Genetics.
    ENGLISH ABSTRACT: The South African abalone, Haliotis midae, commonly known as perlemoen, is an economically important gastropod mollusc. Historically, this species maintained a lucrative fisheries sector; however with increasingly lower landings there has now been a shift to aquaculture. Efforts to conserve natural populations and to improve abalone aquaculture production are thus running in parallel. Previous studies reported significant disparities in parental contributions in aquaculture populations that could explain the rapid divergence of commercial stocks from wild populations. Furthermore, subtle, but significant, population differentiation has also been reported for wild populations on the west-, south-, and east coast of the South African coastline. This study therefore aimed to investigate the evolutionary forces, in particularly selection, facilitating population divergence in wild and cultured H. midae populations using a population genomics approach. By using both microsatellite- and single nucleotide polymorphism (SNP) markers it was found that approximately 10% to 27% of the H. midae genome may be influenced by selection. When incorporating these loci into analyses of population differentiation (e.g. AMOVA, factorial correspondence analysis and estimates of genetic distance) there was a marked increase in genetic divergence between wild and cultured populations (especially when using microsatellite loci) and amongst populations from different geographic regions (particularly supported by the SNP loci). The differences in population clustering as highlighted by microsatellite- and SNP markers can most likely be attributed to the genomic distribution of the respective loci: The SNP markers were developed from EST sequences and therefore mostly represents protein structural variation; whereas the microsatellite markers, found to be putatively under selection, were mainly located in regulatory motifs. The results of this study therefore confirmed previous observations of divergence amongst wild- and cultured populations, but more importantly demonstrated that selection is an important factor driving this divergence. In wild populations selection probably facilitates adaptation to local environmental conditions, whilst amongst aquaculture population adaptation to captivity, husbandry practices and artificial selection may be important determinants. There is evidence for population bottlenecks in wild- and cultured populations; nonetheless long-term effective population sizes seem to be large. Amongst the wild populations, however, short-term population sizes appear to be small most likely due to differential spawning rates amongst reproductively active animals leading to temporal fluctuation in genetic diversity. The results indicate that contact between wild and cultured abalone should be minimised to prevent any adverse effects due to outbreeding depression. With regards to conservation, an emphasis on maintaining adaptive diversity of the wild stocks might be warranted. Continued genetic monitoring is advisable for both wild and cultured abalone populations as to optimally manage the abalone resource for both conservation and commercial viability and sustainability.
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    Unravelling the mystery of the shark genus Mustelus in southern Africa using a multidisciplinary approach
    (Stellenbosch : Stellenbosch University, 2017-12) Maduna, Simo Njabulo; Bester-van der Merwe, Aletta Elizabeth; Stellenbosch University. Faculty of Agrisciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Multidisciplinary approaches have previously offered some alternative innovative ways of addressing classical ecological questions while providing novel insights into behaviour and biology of elasmobranch species. The species-rich shark genus Mustelus, or smoothhounds (smoothhound sharks), is one of the most bio-economically important groups of elasmobranchs in the world’s oceans. Despite the commercial value of Mustelus, its systematics remains largely unresolved and the knowledge on the copulating and dispersal strategies of species of Mustelus is scarce. Here, a multidisciplinary approach – molecular, morphology and histology – with different methods of analysis on various spatial scales was used. First, this study investigated the evolutionary origin of the shark genus Mustelus in southern Africa using molecular phylogenetic and statistical biogeography approaches. Results gave strong support for a northern hemisphere origin of southern African Mustelus species, and that the radiation of Mustelus in this region was primarily driven by long-distance dispersal. The monophyly of expanded Mustelus indicated that southern African species of the genus arose from at least two separate colonisation events from the Northern Hemisphere. On a microevolutionary scale, a comparative population genetics approach was used to gain insight into spatial genetic structure and dispersal patterns in species of Mustelus (M. mustelus and M. palumbes) and other co-distributed demersal sharks (Galeorhinus galeus and Triakis megalopterus) characterised by assorted life histories, habitat preferences, and dispersal behaviour. Based on novel Next Generation Sequencing (NGS)-mined microsatellites, the null hypothesis of genetic homogeneity was rejected for all species investigated except for T. megalopterus. Most noteworthy is that the coalescent analysis of migration supported asymmetric gene flow from the Indian to the Atlantic Ocean, concordant with the Atlantic Ocean–Indian Ocean connection via Agulhas leakage proposed for many marine species along the South African coast. In terms of fisheries forensics, a dermal denticle identification key guide and two molecular assays (a microsatellite panel and High-resolution melting assay) were successfully developed for species identification of southern African Mustelus (M. mosis, M. mustelus and M. palumbes) and three other shark species (Galeorhinus galeus, Scylliogaleus quecketti and Triakis megalopterus) commonly confused with species of Mustelus in the region. Additionally, a SNP discovery and genotyping pipeline was optimised that could in future be used to obtain genome-wide data that will enable population genetic and demographic processes of the study species to be assessed more accurately. Lastly, evidence of sperm storage in female common smoothhound sharks was reported for the first time using a histological approach. The molecular analysis of a single common smoothhound litter also hinted at the within-species variation in the presence and frequency of multiple paternity previously reported for elasmobranchs. Overall, this study provides the most comprehensive set of conservation genetic resources for the common smoothhound shark to date. The results provide novel insights into the conservation biogeography, species identification and ecology of dispersal as well as mating behaviours in species of Mustelus. This will help inform existing and ongoing management and conservation efforts for smoothhound sharks occurring in southern Africa.