Local adaptation with gene flow in a highly dispersive shark.

Adaptive divergence in response to environmental clines are expected to be common in species occupying heterogeneous environments. Despite numerous advances in techniques appropriate for non-model species, gene-environment association studies in elasmobranchs are still scarce. The bronze whaler or copper shark (Carcharhinus brachyurus) is a large coastal shark with a wide distribution and one of the most exploited elasmobranchs in southern Africa. Here, we assessed the distribution of neutral and adaptive genomic diversity in C. brachyurus across a highly heterogeneous environment in southern Africa based on genome-wide SNPs obtained through a restriction site-associated DNA method (3RAD). A combination of differentiation-based genome-scan (outflank) and genotype-environment analyses (redundancy analysis, latent factor mixed models) identified a total of 234 differentiation-based outlier and candidate SNPs associated with bioclimatic variables. Analysis of 26,299 putatively neutral SNPs revealed moderate and evenly distributed levels of genomic diversity across sites from the east coast of South Africa to Angola. Multivariate and clustering analyses demonstrated a high degree of gene flow with no significant population structuring among or within ocean basins. In contrast, the putatively adaptive SNPs demonstrated the presence of two clusters and deep divergence between Angola and all other individuals from Namibia and South Africa. These results provide evidence for adaptive divergence in response to a heterogeneous seascape in a large, mobile shark despite high levels of gene flow. These results are expected to inform management strategies and policy at the national and regional level for conservation of C. brachyurus populations.

A comprehensive phylogenomic study unveils evolutionary patterns and challenges in the mitochondrial genomes of Carcharhiniformes: A focus on Triakidae.

The complex evolutionary patterns in the mitochondrial genome (mitogenome) of the most species-rich shark order, the Carcharhiniformes (ground sharks) has led to challenges in the phylogenomic reconstruction of the families and genera belonging to the order, particularly the family Triakidae (houndsharks). The current state of Triakidae phylogeny remains controversial, with arguments for both monophyly and paraphyly within the family. We hypothesize that this variability is triggered by the selection of different a priori partitioning schemes to account for site and gene heterogeneity within the mitogenome. Here we used an extensive statistical framework to select the a priori partitioning scheme for inference of the mitochondrial phylogenomic relationships within Carcharhiniformes, tested site heterogeneous CAT + GTR + G4 models and incorporated the multi-species coalescent model (MSCM) into our analyses to account for the influence of gene tree discordance on species tree inference. We included five newly assembled houndshark mitogenomes to increase resolution of Triakidae. During the assembly procedure, we uncovered a 714 bp-duplication in the mitogenome of Galeorhinus galeus. Phylogenetic reconstruction confirmed monophyly within Triakidae and the existence of two distinct clades of the expanded Mustelus genus. The latter alludes to potential evolutionary reversal of reproductive mode from placental to aplacental, suggesting that reproductive mode has played a role in the trajectory of adaptive divergence. These new sequences have the potential to contribute to population genomic investigations, species phylogeography delineation, environmental DNA metabarcoding databases and, ultimately, improved conservation strategies for these ecologically and economically important species.

Addressing the complex phylogenetic relationship of the Gempylidae fishes using mitogenome data.

The Gempylidae (snake mackerels) family, belonging to the order Perciformes, consists of about 24 species described in 16 genera primarily distributed in tropical, subtropical, and temperate seas worldwide. Despite substantial research on this family utilizing morphological and molecular approaches, taxonomy categorization in this group has remained puzzling for decades prompting the need for further investigation into the underlying evolutionary history among the gempylids using molecular tools. In this study, we assembled eight complete novel mitochondrial genomes for five Gempylidae species (Neoepinnula minetomai, Neoepinnula orientalis, Rexea antefurcata, Rexea prometheoides, and Thyrsites atun) using Ion Torrent sequencing to supplement publicly available mitogenome data for gempylids. Using Bayesian inference and maximum-likelihood tree search methods, we investigated the evolutionary relationships of 17 Gempylidae species using mitogenome data. In addition, we estimated divergence times for extant gempylids. We identified two major clades that formed approximately 48.05 (35.89-52.04) million years ago: Gempylidae 1 (Thyrsites atun, Promethichthys prometheus, Nealotus tripes, Diplospinus multistriatus, Paradiplospinus antarcticus, Rexea antefurcata, Rexea nakamurai, Rexea prometheoides, Rexea solandri, Thyrsitoides marleyi, Gempylus serpens, and Nesiarchus nasutus) and Gempylidae 2 (Lepidocybium flavobrunneum, Ruvettus pretiosus, Neoepinnula minetomai, Neoepinnula orientalis, and Epinnula magistralis). The present study demonstrated the superior performance of complete mitogenome data compared with individual genes in phylogenetic reconstruction. By including T. atun individuals from different regions, we demonstrated the potential for the application of mitogenomes in species phylogeography.

Gene expression differentials driven by mass rearing and artificial selection in black soldier fly colonies.

The micro-evolutionary forces that shape genetic diversity during domestication have been assessed in many plant and animal systems. However, the impact of these processes on gene expression, and consequent functional adaptation to artificial environments, remains under-investigated. In this study, whole-transcriptome dynamics associated with the early stages of domestication of the black soldier fly (BSF), Hermetia illucens, were assessed. Differential gene expression (DGE) was evaluated in relation to (i) generational time within the cultured environment (F2 vs. F3), and (ii) two selection strategies [no artificial selective pressure (NS); and selection for greater larval mass (SEL)]. RNA-seq was conducted on 5th instar BSF larvae (n = 36), representing equal proportions of the NS (F2 = 9; F3 = 9) and SEL (F2 = 9; F3 = 9) groups. A multidimensional scaling plot revealed greater gene expression variability within the NS and F2 subgroups, while the SEL group clustered separately with lower levels of variation. Comparisons between generations revealed 898 differentially expressed genes (DEGs; FDR-corrected p < 0.05), while between selection strategies, 213 DEGs were observed (FDR-corrected p < 0.05). Enrichment analyses revealed that metabolic, developmental, and defence response processes were over-expressed in the comparison between F2 and F3 larvae, while metabolic processes were the main differentiating factor between NS and SEL lines. This illustrates the functional adaptations that occur in BSF colonies across generations due to mass rearing; as well as highlighting genic dynamics associated with artificial selection for production traits that might inform future selective breeding strategies.

Discerning the global phylogeographic distribution of Phyllosticta citricarpa by means of whole genome sequencing.

Phyllosticta citricarpa is a fungal pathogen causing citrus black spot (CBS). As a regulated pest in some countries, the presence of the pathogen limits the export of fruit and is therefore of agricultural and economic importance. In this study, we used high throughput sequencing data to infer the global phylogeographic distribution of this pathogen, including 71 isolates from eight countries, Argentina, Australia, Brazil, China, Cuba, Eswatini, South Africa and the United States of America. We assembled draft genomes and used a pairwise read mapping approach for the detection and enumeration of variants between isolates. We performed SSR marker discovery based on the assembled genome with the best assembly statistics, and generated genotype profiles for all isolates with 1987 SSR markers in silico. Furthermore, we identified 32,560 SNPs relative to a reference sequence followed by population genetic analyses based on the three datasets; pairwise variant counts, SSR genotypes and SNP genotypes. All three analysis approaches gave similar overall results. Possible pathways of dissemination among the populations from China, Australia, southern Africa and the Americas are postulated. The Chinese population is the most diverse, and is genetically the furthest removed from all other populations, and is therefore considered the closest to the origin of the pathogen. Isolates from Australia, Eswatini and the South African province Mpumalanga are closely associated and clustered together with those from Argentina and Brazil. The Eastern Cape, North West, and KwaZulu-Natal populations in South Africa grouped in another cluster, while isolates from Limpopo are distributed between the two aforementioned clusters. Southern African populations showed a close relationship to populations in North America, and could be a possible source of P. citricarpa populations that are now found in North America. This study represents the largest whole genome sequencing survey of P. citricarpa to date and provides a more comprehensive assessment of the population genetic diversity and connectivity of P. citricarpa from different geographic origins. This information could further assist in a better understanding of the epidemiology of the CBS pathogen, its long-distance dispersal and dissemination pathways, and can be used to refine phytosanitary regulations and management programmes for the disease.

Site fidelity and shallow genetic structure in the common smooth-hound shark Mustelus mustelus confirmed by tag-recapture and genetic data.

The common smooth-hound shark, Mustelus mustelus, is a widely distributed demersal shark under heavy exploitation from various fisheries throughout its distribution range. To assist in the development of appropriate management strategies, the authors evaluate stock structure, site fidelity and movement patterns along the species' distribution in southern Africa based on a combination of molecular and long-term tag-recapture data. Eight species-specific microsatellite markers (N = 73) and two mitochondrial genes, nicotinamide adenine dehydrogenase subunit 4 and control region (N = 45), did not reveal any significant genetic structure among neighbouring sites. Nonetheless, tagging data demonstrate a remarkable degree of site fidelity with 76% of sharks recaptured within 50 km of the original tagging location. On a larger geographic scale, dispersal is governed by oceanographic features as demonstrated by the lack of movements across the Benguela-Agulhas transition zone separating the South-East Atlantic Ocean (SEAO) and South-West Indian Ocean (SWIO) populations. Microsatellite data supported very shallow ocean-based structure (SEAO and SWIO) and historical southward gene flow following the Agulhas Current, corroborating the influence of this dynamic oceanographic system on gene flow. Moreover, no movements between Namibia and South Africa were observed, indicating that the Lüderitz upwelling formation off the Namibian coast acts as another barrier to dispersal and gene flow. Overall, these results show that dispersal and stock structure of M. mustelus are governed by a combination of behavioural traits and oceanographic features such as steep temperature gradients, currents and upwelling systems.

Patterns of Genetic Diversity and Mating Systems in a Mass-Reared Black Soldier Fly Colony.

The black soldier fly (BSF), Hermetia illucens, is a promising candidate for the emerging insect farming industry with favourable characteristics for both bioremediation and production of animal delivered nutritive and industrial compounds. The genetic management of commercial colonies will become increasingly important for the sustainability of the industry. However, r-selected life history traits of insects pose challenges to conventional animal husbandry and breeding approaches. In this study, the long-term genetic effects of mass-rearing were evaluated as well as mating systems in the species to establish factors that might influence genetic diversity, and by implication fitness and productivity in commercial colonies. Population genetic parameters, based on microsatellite markers, were estimated and compared amongst two temporal wild sampling populations and four generations (F28, F48, F52, and F62) of a mass-reared colony. Furthermore, genetic relationships amongst mate pairs were evaluated and parentage analysis was performed to determine the oc-currence of preferential mate choice and multiple paternity. The mass-reared colony showed a reduction in genetic diversity and evidence for inbreeding with significant successive generational genetic differentiation from the wild progenitor population. Population-level analysis also gave the first tentative evidence of positive assortative mating and genetic polyandry in BSF. The homoge-neity of the mass-reared colony seems to result from a dual action caused by small effective popu-lation size and increased homozygosity due to positive assortative mating. However, the high ge-netic diversity in the wild and a polyandrous mating system might suggest the possible restoration of diversity in mass-reared colonies through augmentation with the wild population.

Genetic Diversity and Mating Type Distribution of Pseudocercospora fijiensis on Banana in Uganda and Tanzania.

Black Sigatoka, caused by Pseudocercospora fijiensis, is a major foliar disease of banana and plantain worldwide. There are few available data regarding the genetic diversity and population structure of the pathogen in East Africa, which are needed to design effective and durable disease management strategies. We genotyped 319 single-spore isolates of P. fijiensis collected from seven regions in Uganda and Tanzania and five isolates from Nigeria using 16 simple sequence repeat markers and mating type-specific primers. Isolates from each country and region within the country were treated as populations and subpopulations, respectively. A total of 296 multilocus genotypes (MLGs) were recovered, representing a clonal fraction of 7%. Subpopulations had a moderate level of genetic diversity (Hexp = 0.12 to 0.31; mean, 0.29). Mating type distribution did not deviate from equilibrium (MAT1-1: MAT1-2, 1:1 ratio) in Uganda; however, in Tanzania the mating types were not in equilibrium (4:1 ratio). The index of association tests (IA and r̄d) showed that all populations were at linkage equilibrium (P > 0.05), thus supporting the hypothesis of random association of alleles. These findings are consistent with a pathogen that reproduces both clonally and sexually. Low and insignificant levels of population differentiation were detected, with 90% of the variation occurring among isolates within subpopulations. The high intrapopulation variation has implications in breeding for resistance to P. fijiensis because isolates differing in aggressiveness and virulence are likely to exist over small spatial scales. Diverse isolates will be required for resistance screening to ensure selection of banana cultivars with durable resistance to Sigatoka in East Africa.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Spatial and Temporal Genetic Analyses of Phyllosticta citricarpa in Two Lemon Orchards in South Africa Reveal a Role of Asexual Reproduction Within Sexually Reproducing Populations.

Citrus black spot (CBS), caused by Phyllosticta citricarpa, is a disease that affects citrus worldwide. In different regions of the world where both mating types occur, reports differ as to whether asexually produced pycnidiospores play an important role in the epidemiology of CBS and fruit infections. Therefore, we investigated the potential role of pycnidiospores in two lemon orchards in South Africa by using microsatellite-based analysis of fruit populations over time (two seasons) and space (distance). The two orchards were situated in the semiarid North West province (NW) and subtropical Mpumalanga province (MP). Each population contained both mating types in 1:1 ratios, and linkage disequilibrium analysis indicated a random mating population. A total of 109 and 94 multilocus genotypes (MLGs) were detected across the two seasons in the NW and MP orchards, respectively. Psex analyses indicated that most MLGs probably resulted from sexual reproduction, but there were six predominant MLGs in each orchard that were probably replicated via asexual reproduction. Each of the predominant MLGs was monomorphic for mating type. In the NW, five predominant and widespread MLGs caused 46 and 44% of the fruit infections in the two seasons, whereas in MP, three MLGs caused 34 and 48% of the infections. Asexual reproduction in both orchards was supported by low MLG evenness values in all populations. In both orchards, distance was not a reliable predictor of population genetic substructuring or season. Populations of P. citricarpa in the MP and NW orchards were significantly genetically differentiated from each other.

New polymorphic microsatellite loci revealed for the dusky shark Carcharhinus obscurus through Ion Proton double-digest RAD sequencing.

The non-model shark species, dusky shark Carcharhinus obscurus, is a bio-economically and recreationally important shark in many areas of its range. Despite of the fishery importance of C. obscurus few genetic resources are currently available for the species. Here, we report on the isolation of eight novel microsatellite loci from C. obscurus using a double-digest restriction site associated DNA (RAD) sequencing approach on the Ion Proton semiconductor platform (ddRADseq-ion). We characterised the loci in 26 individuals and all loci were polymorphic, exhibiting 5-10 alleles (average 6.6), and observed and expected heterozygosities of 0.385-0.962 and 0.479-0.847, respectively. We found that all pairs of loci were in linkage equilibrium and conformed to Hardy-Weinberg expectations. The loci reported in this study are only the second set of microsatellite loci ever characterized for C. obscurus and will be valuable for molecular ecology studies for this vulnerable species.

The complete mitochondrial genome and phylogenetic position of the leopard catshark, Poroderma pantherinum.

We present the first mitochondrial genome of a South African endemic catshark, Poroderma pantherinum. The complete mitogenome is 16,686 bp in length, comprising 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and one non-coding control region. Similar to other shark mitogenomes, it is AT rich (61.1%), with a GC content of 38.9%. Protein-coding genes used one of two start codons (ATG and GTG) and one stop codon (TAA/TA-/T-). Phylogenetic analysis of the leopard catshark and 34 carcharhinid species showed that it clusters with two other scyliorhinid species (Cephaloscyllium umbratile and Scyliorhinus canicula) with 100% support.

Characterization of the complete mitochondrial genome of the common smoothhound shark, Mustelus mustelus (Carcharhiniformes: Triakidae).

We present the complete mitochondrial genome of the common smoothhound , Mustelus mustelus,which is 16,755 bp long, contains 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and non-coding control region. All protein-coding genes begin with the ATG codon, except for the COI gene, which begins with GTG. Six protein-coding genes terminated with the TAA codon, and six with incomplete codons, T or TA. The phylogenetic reconstruction places M. mustelus within the genus Mustelus, with the closest relationship to the placental species, M. griseus. This mitogenome provides valuable information to further unravel the evolution of alternate reproductive modes within the genus.

Population genetics of Southern Hemisphere tope shark (Galeorhinus galeus): Intercontinental divergence and constrained gene flow at different geographical scales.

The tope shark (Galeorhinus galeus Linnaeus, 1758) is a temperate, coastal hound shark found in the Atlantic and Indo-Pacific oceans. In this study, the population structure of Galeorhinus galeus was determined across the entire Southern Hemisphere, where the species is heavily targeted by commercial fisheries, as well as locally, along the South African coastline. Analysis was conducted on a total of 185 samples using 19 microsatellite markers and a 671 bp fragment of the NADH dehydrogenase subunit 2 (ND2) gene. Across the Southern Hemisphere, three geographically distinct clades were recovered, including one from South America (Argentina, Chile), one from Africa (all the South African collections) and an Australia-New Zealand clade. Nuclear data revealed significant population subdivisions (FST = 0.192 to 0.376, p<0.05) indicating limited gene flow for tope sharks across ocean basins. Marked population connectivity was however evident across the Indian Ocean based on Bayesian clustering analysis. More locally in South Africa, F-statistics and multivariate analysis supported moderate to high gene flow across the Atlantic/Indian Ocean boundary (FST = 0.035 to 0.044, p<0.05), with exception of samples from Struisbaai and Port Elizabeth which differed significantly from the rest. Discriminant and Bayesian clustering analysis indicated admixture in all sampling populations, decreasing from west to east, corroborating possible restriction to gene flow across regional oceanographic barriers. Mitochondrial sequence data recovered seven haplotypes (h = 0.216, π = 0.001) for South Africa, with one major haplotype shared by 87% of the individuals and at least one private haplotype for each sampling location except Port Elizabeth. As with many other coastal shark species with cosmopolitan distribution, this study confirms the lack of both historical dispersal and inter-oceanic gene flow while also implicating contemporary factors such as oceanic currents and thermal fronts to drive local genetic structure of G. galeus on a smaller spatial scale.

Species identification and comparative population genetics of four coastal houndsharks based on novel NGS-mined microsatellites.

The common smooth-hound (Mustelus mustelus) is the topmost bio-economically and recreationally important shark species in southern Africa, western Africa, and Mediterranean Sea. Here, we used the Illumina HiSeq™ 2000 next-generation sequencing (NGS) technology to develop novel microsatellite markers for Mustelus mustelus. Two microsatellite multiplex panels were constructed from 11 polymorphic loci and characterized in two populations of Mustelus mustelus representative of its South African distribution. The markers were then tested for cross-species utility in Galeorhinus galeus, Mustelus palumbes, and Triakis megalopterus, three other demersal coastal sharks also subjected to recreational and/or commercial fishery pressures in South Africa. We assessed genetic diversity (NA, AR, HO, HE, and PIC) and differentiation (FST and Dest) for each species and also examined the potential use of these markers in species assignment. In each of the four species, all 11 microsatellites were variable with up to a mean NA of 8, AR up to 7.5, HE and PIC as high as 0.842. We were able to reject genetic homogeneity for all species investigated here except for T. megalopterus. We found that the panel of the microsatellite markers developed in this study could discriminate between the study species, particularly for those that are morphologically very similar. Our study provides molecular tools to address ecological and evolutionary questions vital to the conservation and management of these locally and globally exploited shark species.

Microsatellite cross-species amplification and utility in southern African elasmobranchs: A valuable resource for fisheries management and conservation.

BACKGROUND: Similarly to the rest of the world, southern Africa's diverse chondrichthyan fauna is currently experiencing high fishing pressures from direct and non-direct fisheries to satisfy market demands for shark products such as fins and meat. In this study, the development of microsatellite markers through cross-species amplification of primer sets previously developed for closely related species is reported as an alternative approach to de novo marker development. This included the design of four microsatellite multiplex assays and their cross-species utility in genetic diversity analysis of southern African elasmobranchs. As this study forms part of a larger project on the development of genetic resources for commercially important and endemic southern African species, Mustelus mustelus was used as a candidate species for testing these multiplex assays in down-stream applications. RESULTS: Thirty five microsatellite primer sets previously developed for five elasmobranch species were selected from literature for testing cross-species amplification in 16 elasmobranch species occurring in southern Africa. Cross-species amplification success rates ranged from 28.6%-71.4%. From the successfully amplified microsatellites, 22 loci were selected and evaluated for levels of polymorphism, and four multiplex assays comprising of the 22 microsatellites were successfully constructed, optimised and characterised in a panel of 87 Mustelus mustelus individuals. A total of 125 alleles were observed across all loci, with the number of alleles ranging from 3-12 alleles. Cross-species amplification of the four optimised multiplex assays was further tested on 11 commercially important and endemic southern African elasmobranch species. Percentage of polymorphism ranged from 31.8%-95.5% in these species with polymorphic information content decreasing exponentially with evolutionary distance from the source species. CONCLUSIONS: Cross-species amplification of the 35 microsatellites proved to be a time- and cost-effective approach to marker development in elasmobranchs and enabled the construction of four novel multiplex assays for characterising genetic diversity in a number of southern African elasmobranch species. This study successfully demonstrated the usefulness of these markers in down-stream applications such as genetic diversity assessment and species identification which could potentially aid in a more integrative, multidisciplinary approach to management and conservation of commercially important cosmopolitan and endemic elasmobranch species occurring in southern Africa.

Comparison of population genetic estimates amongst wild, F1 and F2 cultured abalone (Haliotis midae).

Haliotis midae is South Africa's most important aquaculture species. The reproduction cycle is currently not closed as many farms rely on wild-caught broodstock for seed production. However, there is an increasing interest in genetic improvement in commercial stocks, with a growing number of producers implementing selective breeding strategies. High throughput commercial production and mass spawning make it difficult to maintain breeding records; therefore, mostly mass selection is practised. The high fecundity and unequal parental contributions also often lead to increased levels of inbreeding. This study therefore aimed to assess the genetic effects of such breeding practices on commercial populations of H. midae. Using microsatellite loci, the genetic properties of a wild, an F1 and an F2 population were estimated and compared. Although there was no significant loss of genetic diversity amongst the cultured populations in comparison with the wild progenitor population, there was low-to-moderate genetic differentiation between populations. Relatedness amongst the F2 population was significant, and the rate of inbreeding was high. The effective population size for the F2 (±50) was also comparatively small with respect to the wild (∞) and F1 (±470) populations. These results suggest that farms need to give caution to breeding practices beyond the first (F1) generation and aim to increase effective population sizes and minimise inbreeding to ensure long-term genetic gain and productivity. This study also confirms the usefulness of population genetic analyses for commercial breeding and stock management in the absence of extensive pedigree records.

Detection of molecular signatures of selection at microsatellite loci in the South African abalone (Haliotis midae) using a population genomic approach.

Identifying genomic regions that may be under selection is important for elucidating the genetic architecture of complex phenotypes underlying adaptation to heterogeneous environments. A population genomic approach, using a classical neutrality test and various Fst-outlier detection methods was employed to evaluate genome-wide polymorphism data in order to identify loci that may be candidates for selection amongst six populations (three cultured and three wild) of the South African abalone, Haliotis midae. Approximately 9% of the genome-wide microsatellite markers were putatively subject to directional selection, whilst 6-18% of the genome is thought to be influenced by balancing selection. Genetic diversity estimates for candidate loci under directional selection was significantly reduced in comparison to candidate neutral loci, whilst candidate balancing selection loci demonstrated significantly higher levels of genetic diversity (Kruskal-Wallis test, P<0.05). Pairwise Fst estimates based on candidate directional selection loci also demonstrated increased levels of differentiation between study populations. Various candidate loci under selection showed significant inter-chromosomal linkage disequilibrium, suggesting possible gene-networks underling adaptive phenotypes. Furthermore, several loci had significant hits to known genes when performing BLAST searches to NCBI's non-redundant databases, whilst others are known to be derived from expressed sequences even though homology to a known gene could not be established. A number of loci also demonstrated relatively high similarity to transposable elements. The association of these loci to functional and genomically active sequences could in part explain the observed signatures of selection.