Identification of Signatures of Positive Selection That Have Shaped the Genomic Landscape of South African Pig Populations.

South Africa boasts a diverse range of pig populations, encompassing intensively raised commercial breeds, as well as indigenous and village pigs reared under low-input production systems. The aim of this study was to investigate how natural and artificial selection have shaped the genomic landscape of South African pig populations sampled from different genetic backgrounds and production systems. For this purpose, the integrated haplotype score (iHS), as well as cross population extended haplotype homozygosity (XP-EHH) and Lewontin and Krakauer's extension of the Fst statistic based on haplotype information (HapFLK) were utilised. Our results revealed several population-specific signatures of selection associated with the different production systems. The importance of natural selection in village populations was highlighted, as the majority of genomic regions under selection were identified in these populations. Regions under natural and artificial selection causing the distinct genetic footprints of these populations also allow for the identification of genes and pathways that may influence production and adaptation. In the context of intensively raised commercial pig breeds (Large White, Kolbroek, and Windsnyer), the identified regions included quantitative loci (QTLs) associated with economically important traits. For example, meat and carcass QTLs were prevalent in all the populations, showing the potential of village and indigenous populations' ability to be managed and improved for such traits. Results of this study therefore increase our understanding of the intricate interplay between selection pressures, genomic adaptations, and desirable traits within South African pig populations.

Chromosome-Level Assemblies for the Pine Pitch Canker Pathogen Fusarium circinatum.

The pine pitch canker pathogen, Fusarium circinatum, is globally regarded as one of the most important threats to commercial pine-based forestry. Although genome sequences of this fungus are available, these remain highly fragmented or structurally ill-defined. Our overall goal was to provide high-quality assemblies for two notable strains of F. circinatum, and to characterize these in terms of coding content, repetitiveness and the position of telomeres and centromeres. For this purpose, we used Oxford Nanopore Technologies MinION long-read sequences, as well as Illumina short sequence reads. By leveraging the genomic synteny inherent to F. circinatum and its close relatives, these sequence reads were assembled to chromosome level, where contiguous sequences mostly spanned from telomere to telomere. Comparative analyses unveiled remarkable variability in the twelfth and smallest chromosome, which is known to be dispensable. It presented a striking length polymorphism, with one strain lacking substantial portions from the chromosome's distal and proximal regions. These regions, characterized by a lower gene density, G+C content and an increased prevalence of repetitive elements, contrast starkly with the syntenic segments of the chromosome, as well as with the core chromosomes. We propose that these unusual regions might have arisen or expanded due to the presence of transposable elements. A comparison of the overall chromosome structure revealed that centromeric elements often underpin intrachromosomal differences between F. circinatum strains, especially at chromosomal breakpoints. This suggests a potential role for centromeres in shaping the chromosomal architecture of F. circinatum and its relatives. The publicly available genome data generated here, together with the detailed metadata provided, represent essential resources for future studies of this important plant pathogen.

Identification and Characterization of a QTL for Growth of Fusarium circinatum on Pine-Based Medium.

Fusarium circinatum is an economically important pathogen of pine and resides in the Fusarium fujikuroi species complex. Here we investigated the molecular processes underlying growth in F. circinatum by exploring the association between growth and the nutritional environment provided by the pine host. For this purpose, we subjected a mapping population consisting of F. circinatum X F. temperatum hybrid progeny to an analysis of growth rate on a pine-tissue derived medium. These data, together with the available genetic linkage map for F. circinatum, were then used to identify Quantitative Trait Loci (QTLs) associated with growth. The single significant QTL identified was then characterized using the available genome sequences for the hybrid progeny's parental isolates. This revealed that the QTL localized to two non-homologous regions in the F. circinatum and F. temperatum genomes. For one of these, the F. circinatum parent contained a two-gene deletion relative to the F. temperatum parent. For the other region, the two parental isolates encoded different protein products. Analysis of repeats, G+C content, and repeat-induced point (RIP) mutations further suggested a retrotransposon origin for the two-gene deletion in F. circinatum. Nevertheless, subsequent genome and PCR-based analyses showed that both regions were similarly polymorphic within a collection of diverse F. circinatum. However, we observed no clear correlation between the respective polymorphism patterns and growth rate in culture. These findings support the notion that growth is a complex multilocus trait and raise the possibility that the identified QTL contains multiple small-effect QTLs, of which some might be dependent on the genetic backgrounds. This study improved our current knowledge of the genetic determinants of vegetative growth in F. circinatum and provided an important foundation for determining the genes and processes underpinning its ability to colonize its host environment.

Phenolic degradation by catechol dioxygenases is associated with pathogenic fungi with a necrotrophic lifestyle in the Ceratocystidaceae.

Fungal species of the Ceratocystidaceae grow on their host plants using a variety of different lifestyles, from saprophytic to highly pathogenic. Although many genomes of fungi in the Ceratocystidaceae are publicly available, it is not known how the genes that encode catechol dioxygenases (CDOs), enzymes involved in the degradation of phenolic plant defense compounds, differ among members of the Ceratocystidaceae. The aim of this study was therefore to identify and characterize the genes encoding CDOs in the genomes of Ceratocystidaceae representatives. We found that genes encoding CDOs are more abundant in pathogenic necrotrophic species of the Ceratocystidaceae and less abundant in saprophytic species. The loss of the CDO genes and the associated 3-oxoadipate catabolic pathway appears to have occurred in a lineage-specific manner. Taken together, this study revealed a positive association between CDO gene copy number and fungal lifestyle in Ceratocystidaceae representatives.

IMA genome - F14 : Draft genome sequences of Penicillium roqueforti, Fusarium sororula, Chrysoporthe puriensis, and Chalaropsis populi.

Draft genomes of Penicillium roqueforti, Fusarium sororula, Chalaropsis populi, and Chrysoporthe puriensis are presented. Penicillium roqueforti is a model fungus for genetics, physiological and metabolic studies, as well as for biotechnological applications. Fusarium sororula and Chrysoporthe puriensis are important tree pathogens, and Chalaropsis populi is a soil-borne root-pathogen. The genome sequences presented here thus contribute towards a better understanding of both the pathogenicity and biotechnological potential of these species.

Genome comparisons suggest an association between Ceratocystis host adaptations and effector clusters in unique transposable element families.

Ceratocystis fimbriata is a host specific fungal pathogen of sweet potato (Ipomoea batatas). The closely related species, C. manginecans, is an important pathogen of trees (e.g. Acacia mangium and Mangifera indica) but has never been isolated from tuber crops. The genetic factors that determine the host range and host specificity of these species have not been determined. The aim of this study was to compare the genomes of C. fimbriata and C. manginecans in order to identify species-specific genetic differences that could be associated with host specificity. This included whole-genome alignments as well as comparisons of gene content and transposable elements (TEs). The genomes of the two species were found to be very similar, sharing similar catalogues of CAZymes, peptidases and lipases. However, the genomes of the two species also varied, harbouring species-specific genes (e.g. small secreted effectors, nutrient processing proteins and stress response proteins). A portion of the TEs identified (17%) had a unique distribution in each species. Transposable elements appeared to have played a prominent role in the divergence of the two species because they were strongly associated with chromosomal translocations and inversions as well as with unique genomic regions containing species-specific genes. Two large effector clusters, with unique TEs in each species, were identified. These effectors displayed non-synonymous mutations and deletions, conserved within a species, and could serve as mutational hot-spots for the development of host specificity in the two species.

Phylogenomic incongruence in Ceratocystis: a clue to speciation?

BACKGROUND: The taxonomic history of Ceratocystis, a genus in the Ceratocystidaceae, has been beset with questions and debate. This is due to many of the commonly used species recognition concepts (e.g., morphological and biological species concepts) providing different bases for interpretation of taxonomic boundaries. Species delineation in Ceratocystis primarily relied on genealogical concordance phylogenetic species recognition (GCPSR) using multiple standard molecular markers. RESULTS: Questions have arisen regarding the utility of these markers e.g., ITS, BT and TEF1-α due to evidence of intragenomic variation in the ITS, as well as genealogical incongruence, especially for isolates residing in a group referred to as the Latin-American clade (LAC) of the species. This study applied a phylogenomics approach to investigate the extent of phylogenetic incongruence in Ceratocystis. Phylogenomic analyses of a total of 1121 shared BUSCO genes revealed widespread incongruence within Ceratocystis, particularly within the LAC, which was typified by three equally represented topologies. Comparative analyses of the individual gene trees revealed evolutionary patterns indicative of hybridization. The maximum likelihood phylogenetic tree generated from the concatenated dataset comprised of 1069 shared BUSCO genes provided improved phylogenetic resolution suggesting the need for multiple gene markers in the phylogeny of Ceratocystis. CONCLUSION: The incongruence observed among single gene phylogenies in this study call into question the utility of single or a few molecular markers for species delineation. Although this study provides evidence of interspecific hybridization, the role of hybridization as the source of discordance will require further research because the results could also be explained by high levels of shared ancestral polymorphism in this recently diverged lineage. This study also highlights the utility of BUSCO genes as a set of multiple orthologous genes for phylogenomic studies.

The novel Huntiella omanensis mating gene, MAT1-2-7, is essential for ascomatal maturation.

Sexual reproduction is a highly conserved feature of the eukaryotes, yet sexual compatibility is determined by a wide variety of mechanisms. In ascomycete fungi, sexual development is controlled by genes at the mating type (MAT) locus that confer either MAT1-1 or MAT1-2 mating identity. Although the locus harbours, at minimum, a single gene, the individual MAT loci of certain species, including Huntiella omanensis, encode for two or more genes. The MAT1-2 idiomorph of H. omanensis is made up of MAT1-2-1, a primary MAT gene that is highly conserved in the Pezizomycotina and possesses a well-characterized DNA binding motif, the HMG-box domain. The idiomorph also harbours a novel secondary MAT gene, named MAT1-2-7, with no recognizable functional domains. In this study, we developed a transformation and CRISPR-Cas9-based genome editing protocol to characterize the MAT1-2-7 gene with respect to its function in mating. We have shown that MAT1-2-7 is essential for sexual reproduction and that isolates carrying the truncated MAT1-2-7 gene are incapable of ascomatal maturation and further sexual development. MAT1-2-7 was also shown to influence the vegetative radial growth rate of H. omanensis, illustrating the pleiotropic effects often associated with MAT genes.

Breed Ancestry, Divergence, Admixture, and Selection Patterns of the Simbra Crossbreed.

In this study, we evaluated an admixed South African Simbra crossbred population, as well as the Brahman (Indicine) and Simmental (Taurine) ancestor populations to understand their genetic architecture and detect genomic regions showing signatures of selection. Animals were genotyped using the Illumina BovineLD v2 BeadChip (7K). Genomic structure analysis confirmed that the South African Simbra cattle have an admixed genome, composed of 5/8 Taurine and 3/8 Indicine, ensuring that the Simbra genome maintains favorable traits from both breeds. Genomic regions that have been targeted by selection were detected using the linkage disequilibrium-based methods iHS and Rsb. These analyses identified 10 candidate regions that are potentially under strong positive selection, containing genes implicated in cattle health and production (e.g., TRIM63, KCNA10, NCAM1, SMIM5, MIER3, and SLC24A4). These adaptive alleles likely contribute to the biological and cellular functions determining phenotype in the Simbra hybrid cattle breed. Our data suggested that these alleles were introgressed from the breed's original indicine and taurine ancestors. The Simbra breed thus possesses derived parental alleles that combine the superior traits of the founder Brahman and Simmental breeds. These regions and genes might represent good targets for ad-hoc physiological studies, selection of breeding material and eventually even gene editing, for improved traits in modern cattle breeds. This study represents an important step toward developing and improving strategies for selection and population breeding to ultimately contribute meaningfully to the beef production industry.

QTL mapping of mycelial growth and aggressiveness to distinct hosts in Ceratocystis pathogens.

Some species of Ceratocystis display strong host specificity, such as C. fimbriata sensu stricto that is restricted to sweet potato (Ipomoea batatas) as host. In contrast, the closely related C. manginecans, infects Acacia mangium and Mangifera indica but is not pathogenic to I. batatas. Despite the economic importance of these fungi, knowledge regarding the genetic factors that influence their pathogenicity and host specificity is limited. A recent inheritance study, based on an interspecific cross between C. fimbriata and C. manginecans and the resultant 70 F1 progeny, confirmed that traits such as mycelial growth rate, spore production and aggressiveness on A. mangium and I. batatas are regulated by multiple genes. In the present study, a quantitative trait locus (QTL) analysis was performed to determine the genomic loci associated with these traits. All 70 progeny isolates were genotyped with SNP markers and a linkage map was constructed. The map contained 467 SNPs, distributed across nine linkage groups, with a total length of 1203 cm. Using the progeny genotypes and phenotypes, one QTL was identified on the linkage map for mycelial growth rate, one for aggressiveness to A. mangium and two for aggressiveness to I. batatas (P < 0.05). Two candidate genes, likely associated with mycelial growth rate, were identified in the QTL region. The three QTLs associated with aggressiveness to different hosts contained candidate genes involved in protein processing, detoxification and regions with effector genes and high transposable element density. The results provide a foundation for studies considering the function of genes regulating various quantitative traits in Ceratocystis.

It's All in the Genes: The Regulatory Pathways of Sexual Reproduction in Filamentous Ascomycetes.

Sexual reproduction in filamentous ascomycete fungi results in the production of highly specialized sexual tissues, which arise from relatively simple, vegetative mycelia. This conversion takes place after the recognition of and response to a variety of exogenous and endogenous cues, and relies on very strictly regulated gene, protein, and metabolite pathways. This makes studying sexual development in fungi an interesting tool in which to study gene-gene, gene-protein, and protein-metabolite interactions. This review provides an overview of some of the most important genes involved in this process; from those involved in the conversion of mycelia into sexually-competent tissue, to those involved in the development of the ascomata, the asci, and ultimately, the ascospores.

Distribution and Evolution of Nonribosomal Peptide Synthetase Gene Clusters in the Ceratocystidaceae.

In filamentous fungi, genes in secondary metabolite biosynthetic pathways are generally clustered. In the case of those pathways involved in nonribosomal peptide production, a nonribosomal peptide synthetase (NRPS) gene is commonly found as a main element of the cluster. Large multifunctional enzymes are encoded by members of this gene family that produce a broad spectrum of bioactive compounds. In this research, we applied genome-based identification of nonribosomal peptide biosynthetic gene clusters in the family Ceratocystidaceae. For this purpose, we used the whole genome sequences of species from the genera Ceratocystis,Davidsoniella,Thielaviopsis, Endoconidiophora,Bretziella, Huntiella, and Ambrosiella. To identify and characterize the clusters, different bioinformatics and phylogenetic approaches, as well as PCR-based methods were used. In all genomes studied, two highly conserved NRPS genes (one monomodular and one multimodular) were identified and their potential products were predicted to be siderophores. Expression analysis of two Huntiella species (H. moniliformis and H. omanensis) confirmed the accuracy of the annotations and proved that the genes in both clusters are expressed. Furthermore, a phylogenetic analysis showed that both NRPS genes of the Ceratocystidaceae formed distinct and well supported clades in their respective phylograms, where they grouped with other known NRPSs involved in siderophore production. Overall, these findings improve our understanding of the diversity and evolution of NRPS biosynthetic pathways in the family Ceratocystidaceae.

Genomic analysis of the aggressive tree pathogen Ceratocystis albifundus.

The overall goal of this study was to determine whether the genome of an important plant pathogen in Africa, Ceratocystis albifundus, is structured into subgenomic compartments, and if so, to establish how these compartments are distributed across the genome. For this purpose, the publicly available genome of C. albifundus was complemented with the genome sequences for four additional isolates using the Illumina HiSeq platform. In addition, a reference genome for one of the individuals was assembled using both PacBio and Illumina HiSeq technologies. Our results showed a high degree of synteny between the five genomes, although several regions lacked detectable long-range synteny. These regions were associated with the presence of accessory genes, lower genetic similarity, variation in read-map depth, as well as transposable elements and genes associated with host-pathogen interactions (e.g. effectors and CAZymes). Such patterns are regarded as hallmarks of accelerated evolution, particularly of accessory subgenomic compartments in fungal pathogens. Our findings thus showed that the genome of C. albifundus is made-up of core and accessory subgenomic compartments, which is an important step towards characterizing its pangenome. This study also highlights the value of comparative genomics for understanding mechanisms that may underly and influence the biology and evolution of pathogens.

The synergistic effect of concatenation in phylogenomics: the case in Pantoea.

With the increased availability of genome sequences for bacteria, it has become routine practice to construct genome-based phylogenies. These phylogenies have formed the basis for various taxonomic decisions, especially for resolving problematic relationships between taxa. Despite the popularity of concatenating shared genes to obtain well-supported phylogenies, various issues regarding this combined-evidence approach have been raised. These include the introduction of phylogenetic error into datasets, as well as incongruence due to organism-level evolutionary processes, particularly horizontal gene transfer and incomplete lineage sorting. Because of the huge effect that this could have on phylogenies, we evaluated the impact of phylogenetic conflict caused by organism-level evolutionary processes on the established species phylogeny for Pantoea, a member of the Enterobacterales. We explored the presence and distribution of phylogenetic conflict at the gene partition and nucleotide levels, by identifying putative inter-lineage recombination events that might have contributed to such conflict. Furthermore, we determined whether smaller, randomly constructed datasets had sufficient signal to reconstruct the current species tree hypothesis or if they would be overshadowed by phylogenetic incongruence. We found that no individual gene tree was fully congruent with the species phylogeny of Pantoea, although many of the expected nodes were supported by various individual genes across the genome. Evidence of recombination was found across all lineages within Pantoea, and provides support for organism-level evolutionary processes as a potential source of phylogenetic conflict. The phylogenetic signal from at least 70 random genes recovered robust, well-supported phylogenies for the backbone and most species relationships of Pantoea, and was unaffected by phylogenetic conflict within the dataset. Furthermore, despite providing limited resolution among taxa at the level of single gene trees, concatenated analyses of genes that were identified as having no signal resulted in a phylogeny that resembled the species phylogeny of Pantoea. This distribution of signal and noise across the genome presents the ideal situation for phylogenetic inference, as the topology from a ≥70-gene concatenated species phylogeny is not driven by single genes, and our data suggests that this finding may also hold true for smaller datasets. We thus argue that, by using a concatenation-based approach in phylogenomics, one can obtain robust phylogenies due to the synergistic effect of the combined signal obtained from multiple genes.

IMA Genome-F 11: Draft genome sequences of Fusarium xylarioides, Teratosphaeria gauchensis and T. zuluensis and genome annotation for Ceratocystis fimbriata.

Draft genomes of the fungal species Fusarium xylarioides, Teratosphaeria gauchensis and T. zuluensis are presented. In addition an annotation of the genome of Ceratocystis fimbriata is presented. Overall these genomes provide a valuable resource for understanding the molecular processes underlying pathogenicity and potential management strategies of these economically important fungi.

Ceratocystidaceae exhibit high levels of recombination at the mating-type (MAT) locus.

Mating is central to many fungal life cycles and is controlled by genes at the mating-type (MAT) locus. These genes determine whether the fungus will be self-sterile (heterothallic) or self-fertile (homothallic). Species in the ascomycete family Ceratocystidaceae have different mating strategies, making them interesting to consider with regards to their MAT loci. The aim of this study was to compare the composition of the MAT locus flanking regions in 11 species of Ceratocystidaceae representing four genera. Genome assemblies for each species were examined to identify the MAT locus and determine the structure of the flanking regions. Large contigs containing the MAT locus were then functionally annotated and analysed for the presence of transposable elements. Genes typically flanking the MAT locus in sordariomycetes were found to be highly conserved in the Ceratocystidaceae. The different genera in the Ceratocystidaceae displayed little synteny outside of the immediate MAT locus flanking genes. Even though species ofCeratocystis did not show much synteny outside of the immediate MAT locus flanking genes, species of Huntiella and Endoconidiophora were comparatively syntenic. Due to the high number of transposable elements present in Ceratocystis MAT flanking regions, we hypothesise that Ceratocystis species may have undergone recombination in this region.

Diversity and evolution of polyketide biosynthesis gene clusters in the Ceratocystidaceae.

Polyketides are secondary metabolites with diverse biological activities. Polyketide synthases (PKS) are often encoded from genes clustered in the same genomic region. Functional analyses and genomic studies show that most fungi are capable of producing a repertoire of polyketides. We considered the potential of Ceratocystidaceae for producing polyketides using a comparative genomics approach. Our aims were to identify the putative polyketide biosynthesis gene clusters, to characterize them and predict the types of polyketide compounds they might produce. We used sequences from nineteen species in the genera, Ceratocystis, Endoconidiophora, Davidsoniella, Huntiella, Thielaviopsis and Bretziella, to identify and characterize PKS gene clusters, by employing a range of bioinformatics and phylogenetic tools. We showed that the genomes contained putative clusters containing a non-reducing type I PKS and a type III PKS. Phylogenetic analyses suggested that these genes were already present in the ancestor of the Ceratocystidaceae. By contrast, the various reducing type I PKS-containing clusters identified in these genomes appeared to have distinct evolutionary origins. Although one of the identified clusters potentially allows for the production of melanin, their functional characterization will undoubtedly reveal many novel and important compounds implicated in the biology of the Ceratocystidaceae.

IMA Genome-F 9: Draft genome sequence of Annulohypoxylon stygium, Aspergillus mulundensis, Berkeleyomyces basicola (syn. Thielaviopsis basicola), Ceratocystis smalleyi, two Cercospora beticola strains, Coleophoma cylindrospora, Fusarium fracticaudum, Phialophora cf. hyalina, and Morchella septimelata.

Draft genomes of the species Annulohypoxylon stygium, Aspergillus mulundensis, Berkeleyomyces basicola (syn. Thielaviopsis basicola), Ceratocystis smalleyi, two Cercospora beticola strains, Coleophoma cylindrospora, Fusarium fracticaudum, Phialophora cf. hyalina and Morchella septimelata are presented. Both mating types (MAT1-1 and MAT1-2) of Cercospora beticola are included. Two strains of Coleophoma cylindrospora that produce sulfated homotyrosine echinocandin variants, FR209602, FR220897 and FR220899 are presented. The sequencing of Aspergillus mulundensis, Coleophoma cylindrospora and Phialophora cf. hyalina has enabled mapping of the gene clusters encoding the chemical diversity from the echinocandin pathways, providing data that reveals the complexity of secondary metabolism in these different species. Overall these genomes provide a valuable resource for understanding the molecular processes underlying pathogenicity (in some cases), biology and toxin production of these economically important fungi.