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September 22, 2019  |  

Microsatellite polymorphism in the endangered snail kite reveals a panmictic, low diversity population

Genetic structure and genetic diversity are key population characteristics that can inform conservation decisions, such as delineating management units or assessing potential risks for inbreeding depression. Evidence of genetic structuring or low genetic diversity in the critically endangered snail kite (Rostrhamus sociabilis plumbeus) would have implications for monitoring and planning decisions. Recent work on understanding connectivity across the snail kite range indicated that there is less dispersal between northern and southern parts of the current range, and that dispersal is shaped by individual habitat preference. We examine whether there is neutral genetic structure and the amount of genetic variation in the population by non-lethally sampling 235 nestlings from unique nests across the entire breeding range between 2013 and 2014. Data on 15 microsatellite revealed low diversity (e.g., Na?=?2.54, He?=?0.37) and range-wide panmixia based on AMOVA, Bayesian clustering, spatial autocorrelation, isolation by distance, and spatially explicit ordination analyses. Our results emphasize that long-term recovery goals and management strategies should be based on viewing snail kites as a single genetic population, despite evidence for non-random dispersal between wetlands over ecological time scales. These results also highlight the need to understand potential effects of low genetic diversity on population dynamics and viability of snail kites. More broadly, these results add to the growing evidence for potential discrepancies between dispersal and genetic patterns, emphasizing that care should be taken if using one to interpret the other, particularly for widely-ranging species.

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September 22, 2019  |  

Expansions of intronic TTTCA and TTTTA repeats in benign adult familial myoclonic epilepsy.

Epilepsy is a common neurological disorder, and mutations in genes encoding ion channels or neurotransmitter receptors are frequent causes of monogenic forms of epilepsy. Here we show that abnormal expansions of TTTCA and TTTTA repeats in intron 4 of SAMD12 cause benign adult familial myoclonic epilepsy (BAFME). Single-molecule, real-time sequencing of BAC clones and nanopore sequencing of genomic DNA identified two repeat configurations in SAMD12. Intriguingly, in two families with a clinical diagnosis of BAFME in which no repeat expansions in SAMD12 were observed, we identified similar expansions of TTTCA and TTTTA repeats in introns of TNRC6A and RAPGEF2, indicating that expansions of the same repeat motifs are involved in the pathogenesis of BAFME regardless of the genes in which the expanded repeats are located. This discovery that expansions of noncoding repeats lead to neuronal dysfunction responsible for myoclonic tremor and epilepsy extends the understanding of diseases with such repeat expansion.

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September 22, 2019  |  

Transposable element genomic fissuring in Pyrenophora teres is associated with genome expansion and dynamics of host-pathogen genetic interactions.

Pyrenophora teres, P. teres f. teres (PTT) and P. teres f. maculata (PTM) cause significant diseases in barley, but little is known about the large-scale genomic differences that may distinguish the two forms. Comprehensive genome assemblies were constructed from long DNA reads, optical and genetic maps. As repeat masking in fungal genomes influences the final gene annotations, an accurate and reproducible pipeline was developed to ensure comparability between isolates. The genomes of the two forms are highly collinear, each composed of 12 chromosomes. Genome evolution in P. teres is characterized by genome fissuring through the insertion and expansion of transposable elements (TEs), a process that isolates blocks of genic sequence. The phenomenon is particularly pronounced in PTT, which has a larger, more repetitive genome than PTM and more recent transposon activity measured by the frequency and size of genome fissures. PTT has a longer cultivated host association and, notably, a greater range of host-pathogen genetic interactions compared to other Pyrenophora spp., a property which associates better with genome size than pathogen lifestyle. The two forms possess similar complements of TE families with Tc1/Mariner and LINE-like Tad-1 elements more abundant in PTT. Tad-1 was only detectable as vestigial fragments in PTM and, within the forms, differences in genome sizes and the presence and absence of several TE families indicated recent lineage invasions. Gene differences between P. teres forms are mainly associated with gene-sparse regions near or within TE-rich regions, with many genes possessing characteristics of fungal effectors. Instances of gene interruption by transposons resulting in pseudogenization were detected in PTT. In addition, both forms have a large complement of secondary metabolite gene clusters indicating significant capacity to produce an array of different molecules. This study provides genomic resources for functional genetics to help dissect factors underlying the host-pathogen interactions.

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September 22, 2019  |  

Developing collaborative works for faster progress on fungal respiratory infections in cystic fibrosis.

Cystic fibrosis (CF) is the major genetic inherited disease in Caucasian populations. The respiratory tract of CF patients displays a sticky viscous mucus, which allows for the entrapment of airborne bacteria and fungal spores and provides a suitable environment for growth of microorganisms, including numerous yeast and filamentous fungal species. As a consequence, respiratory infections are the major cause of morbidity and mortality in this clinical context. Although bacteria remain the most common agents of these infections, fungal respiratory infections have emerged as an important cause of disease. Therefore, the International Society for Human and Animal Mycology (ISHAM) has launched a working group on Fungal respiratory infections in Cystic Fibrosis (Fri-CF) in October 2006, which was subsequently approved by the European Confederation of Medical Mycology (ECMM). Meetings of this working group, comprising both clinicians and mycologists involved in the follow-up of CF patients, as well as basic scientists interested in the fungal species involved, provided the opportunity to initiate collaborative works aimed to improve our knowledge on these infections to assist clinicians in patient management. The current review highlights the outcomes of some of these collaborative works in clinical surveillance, pathogenesis and treatment, giving special emphasis to standardization of culture procedures, improvement of species identification methods including the development of nonculture-based diagnostic methods, microbiome studies and identification of new biological markers, and the description of genotyping studies aiming to differentiate transient carriage and chronic colonization of the airways. The review also reports on the breakthrough in sequencing the genomes of the main Scedosporium species as basis for a better understanding of the pathogenic mechanisms of these fungi, and discusses treatment options of infections caused by multidrug resistant microorganisms, such as Scedosporium and Lomentospora species and members of the Rasamsonia argillacea species complex.

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September 22, 2019  |  

Gene presence-absence polymorphism in castrating anther-smut fungi: Recent gene Gains and Phylogeographic Structure.

Gene presence-absence polymorphisms segregating within species are a significant source of genetic variation but have been little investigated to date in natural populations. In plant pathogens, the gain or loss of genes encoding proteins interacting directly with the host, such as secreted proteins, probably plays an important role in coevolution and local adaptation. We investigated gene presence-absence polymorphism in populations of two closely related species of castrating anther-smut fungi, Microbotryum lychnidis-dioicae (MvSl) and M. silenes-dioicae (MvSd), from across Europe, on the basis of Illumina genome sequencing data and high-quality genome references. We observed presence-absence polymorphism for 186 autosomal genes (2% of all genes) in MvSl, and only 51 autosomal genes in MvSd. Distinct genes displayed presence-absence polymorphism in the two species. Genes displaying presence-absence polymorphism were frequently located in subtelomeric and centromeric regions and close to repetitive elements, and comparison with outgroups indicated that most were present in a single species, being recently acquired through duplications in multiple-gene families. Gene presence-absence polymorphism in MvSl showed a phylogeographic structure corresponding to clusters detected based on SNPs. In addition, gene absence alleles were rare within species and skewed toward low-frequency variants. These findings are consistent with a deleterious or neutral effect for most gene presence-absence polymorphism. Some of the observed gene loss and gain events may however be adaptive, as suggested by the putative functions of the corresponding encoded proteins (e.g., secreted proteins) or their localization within previously identified selective sweeps. The adaptive roles in plant and anther-smut fungi interactions of candidate genes however need to be experimentally tested in future studies.

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September 22, 2019  |  

Genomic architecture of haddock (Melanogrammus aeglefinus) shows expansions of innate immune genes and short tandem repeats.

Increased availability of genome assemblies for non-model organisms has resulted in invaluable biological and genomic insight into numerous vertebrates, including teleosts. Sequencing of the Atlantic cod (Gadus morhua) genome and the genomes of many of its relatives (Gadiformes) demonstrated a shared loss of the major histocompatibility complex (MHC) II genes 100 million years ago. An improved version of the Atlantic cod genome assembly shows an extreme density of tandem repeats compared to other vertebrate genome assemblies. Highly contiguous assemblies are therefore needed to further investigate the unusual immune system of the Gadiformes, and whether the high density of tandem repeats found in Atlantic cod is a shared trait in this group.Here, we have sequenced and assembled the genome of haddock (Melanogrammus aeglefinus) – a relative of Atlantic cod – using a combination of PacBio and Illumina reads. Comparative analyses reveal that the haddock genome contains an even higher density of tandem repeats outside and within protein coding sequences than Atlantic cod. Further, both species show an elevated number of tandem repeats in genes mainly involved in signal transduction compared to other teleosts. A characterization of the immune gene repertoire demonstrates a substantial expansion of MCHI in Atlantic cod compared to haddock. In contrast, the Toll-like receptors show a similar pattern of gene losses and expansions. For the NOD-like receptors (NLRs), another gene family associated with the innate immune system, we find a large expansion common to all teleosts, with possible lineage-specific expansions in zebrafish, stickleback and the codfishes.The generation of a highly contiguous genome assembly of haddock revealed that the high density of short tandem repeats as well as expanded immune gene families is not unique to Atlantic cod – but possibly a feature common to all, or most, codfishes. A shared expansion of NLR genes in teleosts suggests that the NLRs have a more substantial role in the innate immunity of teleosts than other vertebrates. Moreover, we find that high copy number genes combined with variable genome assembly qualities may impede complete characterization of these genes, i.e. the number of NLRs in different teleost species might be underestimates.

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September 22, 2019  |  

Genome-wide identification of simple sequence repeats and development of polymorphic SSR markers for genetic studies in tea plant (Camellia sinensis)

The tea plant (Camellia sinensis (L.) O. Kuntze) is one of the most popular non-alcoholic beverage crops worldwide. The availability of complete genome sequences for the Camellia sinensis var. ‘Shuchazao’ has provided the opportunity to identify all types of simple sequence repeat (SSR) markers by genome-wide scan. In this study, a total of 667,980 SSRs were identified in the ~?3.08 Gb genome, with an overall density of 216.88 SSRs/Mb. Dinucleotide repeats were predominant among microsatellites (72.25%), followed by trinucleotide repeats (15.35%), while the remaining SSRs accounted for less than 13%. The motif AG/CT (49.96%) and AT/TA (40.14%) were the most and the second most abundant among all identified SSR motifs, respectively; meanwhile, AAT/ATT (41.29%) and AAAT/ATTT (67.47%) were the most common among trinucleotides and tetranucleotides, respectively. A total of 300 primer pairs were designed to screen six tea cultivars for polymorphisms of SSR markers using the five selected repeat types of microsatellite sequences. The resulting 96 SSR markers that yielded polymorphic and unambiguous bands were further deployed on 47 tea cultivars for genetic diversity assessment, demonstrating high polymorphism of these SSR markers. Remarkably, the dendrogram revealed that the phylogenetic relationships among these tea cultivars are highly consistent with their genetic backgrounds or places of origin. The identified genome-wide SSRs and newly developed SSR markers will provide a powerful means for genetic researches in tea plant, including genetic diversity and evolutionary origin analysis, fingerprinting, QTL mapping, and marker-assisted selection for breeding.

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September 22, 2019  |  

Insights into platypus population structure and history from whole-genome sequencing.

The platypus is an egg-laying mammal which, alongside the echidna, occupies a unique place in the mammalian phylogenetic tree. Despite widespread interest in its unusual biology, little is known about its population structure or recent evolutionary history. To provide new insights into the dispersal and demographic history of this iconic species, we sequenced the genomes of 57 platypuses from across the whole species range in eastern mainland Australia and Tasmania. Using a highly improved reference genome, we called over 6.7?M SNPs, providing an informative genetic data set for population analyses. Our results show very strong population structure in the platypus, with our sampling locations corresponding to discrete groupings between which there is no evidence for recent gene flow. Genome-wide data allowed us to establish that 28 of the 57 sampled individuals had at least a third-degree relative among other samples from the same river, often taken at different times. Taking advantage of a sampled family quartet, we estimated the de novo mutation rate in the platypus at 7.0?×?10-9/bp/generation (95% CI 4.1?×?10-9-1.2?×?10-8/bp/generation). We estimated effective population sizes of ancestral populations and haplotype sharing between current groupings, and found evidence for bottlenecks and long-term population decline in multiple regions, and early divergence between populations in different regions. This study demonstrates the power of whole-genome sequencing for studying natural populations of an evolutionarily important species.

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September 22, 2019  |  

Characterization of phenotypic variation and genome aberrations observed among Phytophthora ramorum isolates from diverse hosts.

Accumulating evidence suggests that genome plasticity allows filamentous plant pathogens to adapt to changing environments. Recently, the generalist plant pathogen Phytophthora ramorum has been documented to undergo irreversible phenotypic alterations accompanied by chromosomal aberrations when infecting trunks of mature oak trees (genus Quercus). In contrast, genomes and phenotypes of the pathogen derived from the foliage of California bay (Umbellularia californica) are usually stable. We define this phenomenon as host-induced phenotypic diversification (HIPD). P. ramorum also causes a severe foliar blight in some ornamental plants such as Rhododendron spp. and Viburnum spp., and isolates from these hosts occasionally show phenotypes resembling those from oak trunks that carry chromosomal aberrations. The aim of this study was to investigate variations in phenotypes and genomes of P. ramorum isolates from non-oak hosts and substrates to determine whether HIPD changes may be equivalent to those among isolates from oaks.We analyzed genomes of diverse non-oak isolates including those taken from foliage of Rhododendron and other ornamental plants, as well as from natural host species, soil, and water. Isolates recovered from artificially inoculated oak logs were also examined. We identified diverse chromosomal aberrations including copy neutral loss of heterozygosity (cnLOH) and aneuploidy in isolates from non-oak hosts. Most identified aberrations in non-oak hosts were also common among oak isolates; however, trisomy, a frequent type of chromosomal aberration in oak isolates was not observed in isolates from Rhododendron.This work cross-examined phenotypic variation and chromosomal aberrations in P. ramorum isolates from oak and non-oak hosts and substrates. The results suggest that HIPD comparable to that occurring in oak hosts occurs in non-oak environments such as in Rhododendron leaves. Rhododendron leaves are more easily available than mature oak stems and thus can potentially serve as a model host for the investigation of HIPD, the newly described plant-pathogen interaction.

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September 22, 2019  |  

Knockout of rapC improves the bacillomycin D yield based on de novo genome sequencing of Bacillus amyloliquefaciens fmbJ.

Bacillus amyloliquefaciens, a Gram-positive and soil-dwelling bacterium, could produce secondary metabolites that suppress plant pathogens. In this study, we provided the whole genome sequence results of B. amyloliquefaciens fmbJ, which had one circular chromosome of 4?193?344 bp with 4249 genes, 87 tRNA genes, and 27 rRNA genes. In addition, fmbJ was found to contain several gene clusters of antimicrobial lipopeptides (bacillomycin D, surfactin, and fengycin), and bacillomycin D homologues were further comprehensively identified. To clarify the influence of rapC regulating the synthesis of lipopeptide on the yield of bacillomycin D, rapC gene in fmbJ was successfully deleted by the marker-free method. Finally, it was found that the deletion of rapC gene in fmbJ significantly improved bacillomycin D production from 240.7 ± 18.9 to 360.8 ± 30.7 mg/L, attributed to the increased the expression of bacillomycin D synthesis-related genes through enhancing the transcriptional level of comA, comP, and phrC. These results showed that the production of bacillomycin D in B. amyloliquefaciens fmbJ might be regulated by the RapC-PhrC system. The findings are expected to advance further agricultural application of Bacillus spp. as a promising source of natural bioactive compounds.

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September 22, 2019  |  

Whole-genome analysis of three yeast strains used for production of sherry-like wines revealed genetic traits specific to Flor yeasts.

Flor yeast strains represent a specialized group of Saccharomyces cerevisiae yeasts used for biological wine aging. We have sequenced the genomes of three flor strains originated from different geographic regions and used for production of sherry-like wines in Russia. According to the obtained phylogeny of 118 yeast strains, flor strains form very tight cluster adjacent to the main wine clade. SNP analysis versus available genomes of wine and flor strains revealed 2,270 genetic variants in 1,337 loci specific to flor strains. Gene ontology analysis in combination with gene content evaluation revealed a complex landscape of possibly adaptive genetic changes in flor yeast, related to genes associated with cell morphology, mitotic cell cycle, ion homeostasis, DNA repair, carbohydrate metabolism, lipid metabolism, and cell wall biogenesis. Pangenomic analysis discovered the presence of several well-known “non-reference” loci of potential industrial importance. Events of gene loss included deletions of asparaginase genes, maltose utilization locus, and FRE-FIT locus involved in iron transport. The latter in combination with a flor-yeast-specific mutation in the Aft1 transcription factor gene is likely to be responsible for the discovered phenotype of increased iron sensitivity and improved iron uptake of analyzed strains. Expansion of the coding region of the FLO11 flocullin gene and alteration of the balance between members of the FLO gene family are likely to positively affect the well-known propensity of flor strains for velum formation. Our study provides new insights in the nature of genetic variation in flor yeast strains and demonstrates that different adaptive properties of flor yeast strains could have evolved through different mechanisms of genetic variation.

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September 22, 2019  |  

Identification of a leucine-rich repeat receptor-like serine/threonine-protein kinase as a candidate gene for Rvi12 (Vb)-based apple scab resistance

Apple scab caused by Venturia inaequalis is the most important fungal disease of apples (Malus × domestica). Currently, the disease is controlled by up to 15 fungicide applications to the crop per year. Resistant apple cultivars will help promote the sustainable control of scab in commercial orchards. The breakdown of the Rvi6 (Vf) major-gene based resistance, the most used resistance gene in apple breeding, prompted the identification and characterization of new scab resistance genes. By using a large segregating population, the Rvi12 scab resistance gene was previously mapped to a genetic location flanked by molecular markers SNP_23.599 and SNP_24.482. Starting from these markers, utilizing chromosome walking of a Hansen’s baccata #2 (HB2) BAC-library; a single BAC clone spanning the Rvi12 interval was identified. Following Pacific Biosciences (PacBio) RS II sequencing and the use of the hierarchical genome assembly process (HGAP) assembly of the BAC clone sequence, the Rvi12 resistance locus was localized to a 62.3-kb genomic region. Gene prediction and in silico characterization identified a single candidate resistance gene. The gene, named here as Rvi12_Cd5, belongs to the LRR receptor-like serine/threonine-protein kinase family. In silico comparison of the resistance allele from HB2 and the susceptible allele from Golden Delicious (GD) identified the presence of an additional intron in the HB2 allele. Conserved domain analysis identified the presence of four additional LRR motifs in the susceptible allele compared to the resistance allele. The constitutive expression of Rvi12_Cd5 in HB2, together with its structural similarity to known resistance genes, makes it the most likely candidate for Rvi12 scab resistance in apple.

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September 22, 2019  |  

The complete chloroplast genome sequence of Actinidia arguta using the PacBio RS II platform.

Actinidia arguta is the most basal species in a phylogenetically and economically important genus in the family Actinidiaceae. To better understand the molecular basis of the Actinidia arguta chloroplast (cp), we sequenced the complete cp genome from A. arguta using Illumina and PacBio RS II sequencing technologies. The cp genome from A. arguta was 157,611 bp in length and composed of a pair of 24,232 bp inverted repeats (IRs) separated by a 20,463 bp small single copy region (SSC) and an 88,684 bp large single copy region (LSC). Overall, the cp genome contained 113 unique genes. The cp genomes from A. arguta and three other Actinidia species from GenBank were subjected to a comparative analysis. Indel mutation events and high frequencies of base substitution were identified, and the accD and ycf2 genes showed a high degree of variation within Actinidia. Forty-seven simple sequence repeats (SSRs) and 155 repetitive structures were identified, further demonstrating the rapid evolution in Actinidia. The cp genome analysis and the identification of variable loci provide vital information for understanding the evolution and function of the chloroplast and for characterizing Actinidia population genetics.

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September 22, 2019  |  

Whole genome sequence and comparative analysis of Borrelia burgdorferi MM1.

Lyme disease is caused by spirochaetes of the Borrelia burgdorferi sensu lato genospecies. Complete genome assemblies are available for fewer than ten strains of Borrelia burgdorferi sensu stricto, the primary cause of Lyme disease in North America. MM1 is a sensu stricto strain originally isolated in the midwestern United States. Aside from a small number of genes, the complete genome sequence of this strain has not been reported. Here we present the complete genome sequence of MM1 in relation to other sensu stricto strains and in terms of its Multi Locus Sequence Typing. Our results indicate that MM1 is a new sequence type which contains a conserved main chromosome and 15 plasmids. Our results include the first contiguous 28.5 kb assembly of lp28-8, a linear plasmid carrying the vls antigenic variation system, from a Borrelia burgdorferi sensu stricto strain.

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September 22, 2019  |  

Genetic diversity of Cryptosporidium hominis in a Bangladeshi community as revealed by whole genome sequencing.

We studied the genetic diversity of Cryptosporidium hominis infections in slum-dwelling infants from Dhaka over a 2-year period. Cryptosporidium hominis infections were common during the monsoon, and were genetically diverse as measured by gp60 genotyping and whole-genome resequencing. Recombination in the parasite was evidenced by the decay of linkage disequilibrium in the genome over <300 bp. Regions of the genome with high levels of polymorphism were also identified. Yet to be determined is if genomic diversity is responsible in part for the high rate of reinfection, seasonality, and varied clinical presentations of cryptosporidiosis in this population.

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