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April 21, 2020

Genome sequence of Jatropha curcas L., a non-edible biodiesel plant, provides a resource to improve seed-related traits.

Jatropha curcas (physic nut), a non-edible oilseed crop, represents one of the most promising alternative energy sources due to its high seed oil content, rapid growth and adaptability to various environments. We report ~339 Mbp draft whole genome sequence of J. curcas var. Chai Nat using both the PacBio and Illumina sequencing platforms. We identified and categorized differentially expressed genes related to biosynthesis of lipid and toxic compound among four stages of seed development. Triacylglycerol (TAG), the major component of seed storage oil, is mainly synthesized by phospholipid:diacylglycerol acyltransferase in Jatropha, and continuous high expression of homologs of oleosin over seed development contributes to accumulation of high level of oil in kernels by preventing the breakdown of TAG. A physical cluster of genes for diterpenoid biosynthetic enzymes, including casbene synthases highly responsible for a toxic compound, phorbol ester, in seed cake, was syntenically highly conserved between Jatropha and castor bean. Transcriptomic analysis of female and male flowers revealed the up-regulation of a dozen family of TFs in female flower. Additionally, we constructed a robust species tree enabling estimation of divergence times among nine Jatropha species and five commercial crops in Malpighiales order. Our results will help researchers and breeders increase energy efficiency of this important oil seed crop by improving yield and oil content, and eliminating toxic compound in seed cake for animal feed. © 2018 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

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April 21, 2020

The Genome of Cucurbita argyrosperma (Silver-Seed Gourd) Reveals Faster Rates of Protein-Coding Gene and Long Noncoding RNA Turnover and Neofunctionalization within Cucurbita.

Whole-genome duplications are an important source of evolutionary novelties that change the mode and tempo at which genetic elements evolve within a genome. The Cucurbita genus experienced a whole-genome duplication around 30 million years ago, although the evolutionary dynamics of the coding and noncoding genes in this genus have not yet been scrutinized. Here, we analyzed the genomes of four Cucurbita species, including a newly assembled genome of Cucurbita argyrosperma, and compared the gene contents of these species with those of five other members of the Cucurbitaceae family to assess the evolutionary dynamics of protein-coding and long intergenic noncoding RNA (lincRNA) genes after the genome duplication. We report that Cucurbita genomes have a higher protein-coding gene birth-death rate compared with the genomes of the other members of the Cucurbitaceae family. C. argyrosperma gene families associated with pollination and transmembrane transport had significantly faster evolutionary rates. lincRNA families showed high levels of gene turnover throughout the phylogeny, and 67.7% of the lincRNA families in Cucurbita showed evidence of birth from the neofunctionalization of previously existing protein-coding genes. Collectively, our results suggest that the whole-genome duplication in Cucurbita resulted in faster rates of gene family evolution through the neofunctionalization of duplicated genes. Copyright © 2019 The Author. Published by Elsevier Inc. All rights reserved.

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April 21, 2020

Endogenous pararetrovirus sequences are widely present in Citrinae genomes.

Endogenous pararetroviruses (EPRVs) are characterized in several plant genomes and their biological effects have been reported. In this study, hundreds of EPRV segments were identified in six Citrinae genomes. A total of 1034 EPRV segments were identified in the genomes of sweet orange, 2036 in pummelo, 598 in clementine mandarin, 752 in Ichang papeda, 2060 in citron and 245 in atalantia. Genomic analysis indicated that EPRV segments tend to cluster as hot spots in the genomes, particularly on chromosome 2 and 5. Large numbers of simple repeats and transposable elements were identified in the 2-kb flanking regions of the EPRV segments. Comparative genomic analysis and PCR experiments showed that there are highly conserved EPRV segments and species-specific EPRV segments between the Citrinae genomes. Phylogenetic analysis suggested that the integration events of EPRVs could initiate in a common progenitor of Citrinae species and repeatedly occur during the Citrinae divergence.Copyright © 2018 Elsevier B.V. All rights reserved.

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April 21, 2020

Genomic variation and strain-specific functional adaptation in the human gut microbiome during early life.

The human gut microbiome matures towards the adult composition during the first years of life and is implicated in early immune development. Here, we investigate the effects of microbial genomic diversity on gut microbiome development using integrated early childhood data sets collected in the DIABIMMUNE study in Finland, Estonia and Russian Karelia. We show that gut microbial diversity is associated with household location and linear growth of children. Single nucleotide polymorphism- and metagenomic assembly-based strain tracking revealed large and highly dynamic microbial pangenomes, especially in the genus Bacteroides, in which we identified evidence of variability deriving from Bacteroides-targeting bacteriophages. Our analyses revealed functional consequences of strain diversity; only 10% of Finnish infants harboured Bifidobacterium longum subsp. infantis, a subspecies specialized in human milk metabolism, whereas Russian infants commonly maintained a probiotic Bifidobacterium bifidum strain in infancy. Groups of bacteria contributing to diverse, characterized metabolic pathways converged to highly subject-specific configurations over the first two years of life. This longitudinal study extends the current view of early gut microbial community assembly based on strain-level genomic variation.

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April 21, 2020

Genome sequence and genetic transformation of a widely distributed and cultivated poplar.

Populus alba is widely distributed and cultivated in Europe and Asia. This species has been used for diverse studies. In this study, we assembled a de novo genome sequence of P. alba var. pyramidalis (= P. bolleana) and confirmed its high transformation efficiency and short transformation time by experiments. Through a process of hybrid genome assembly, a total of 464 M of the genome was assembled. Annotation analyses predicted 37 901 protein-coding genes. This genome is highly collinear to that of P. trichocarpa, with most genes having orthologs in the two species. We found a marked expansion of gene families related to histone and the hormone auxin but loss of disease resistance genes in P. alba if compared with the closely related P. trichocarpa. The genome sequence presented here represents a valuable resource for further molecular functional analyses of this species as a new tree model, poplar breeding practices and comparative genomic analyses across different poplars. © 2018 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

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April 21, 2020

Patterns of non-ARD variation in more than 300 full-length HLA-DPB1 alleles.

Our understanding of sequence variation in the HLA-DPB1 gene is largely restricted to the hypervariable antigen recognition domain (ARD) encoded by exon 2. Here, we employed a redundant sequencing strategy combining long-read and short-read data to accurately phase and characterise in full length the majority of common and well-documented (CWD) DPB1 alleles as well as alleles with an observed frequency of at least 0.0006% in our predominantly European sample set. We generated 664 DPB1 sequences, comprising 279 distinct allelic variants. This allows us to present the, to date, most comprehensive analysis of the nature and extent of DPB1 sequence variation. The full-length sequence analysis revealed the existence of two highly diverged allele clades. These clades correlate with the rs9277534 A???G variant, a known expression marker located in the 3′-UTR. The two clades are fully differentiated by 174 fixed polymorphisms throughout a 3.6?kb stretch at the 3′-end of DPB1. The region upstream of this differentiation zone is characterised by increasingly shared variation between the clades. The low-expression A clade comprises 59% of the distinct allelic sequences including the three by far most frequent DPB1 alleles, DPB1*04:01, DPB1*02:01 and DPB1*04:02. Alleles in the A clade show reduced nucleotide diversity with an excess of rare variants when compared to the high-expression G clade. This pattern is consistent with a scenario of recent proliferation of A-clade alleles. The full-length characterisation of all but the most rare DPB1 alleles will benefit the application of NGS for DPB1 genotyping and provides a helpful framework for a deeper understanding of high- and low-expression alleles and their implications in the context of unrelated haematopoietic stem-cell transplantation.Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

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April 21, 2020

Secretion of an Argonaute protein by a parasitic nematode and the evolution of its siRNA guides.

Extracellular RNA has been proposed to mediate communication between cells and organisms however relatively little is understood regarding how specific sequences are selected for export. Here, we describe a specific Argonaute protein (exWAGO) that is secreted in extracellular vesicles (EVs) released by the gastrointestinal nematode Heligmosomoides bakeri, at multiple copies per EV. Phylogenetic and gene expression analyses demonstrate exWAGO orthologues are highly conserved and abundantly expressed in related parasites but highly diverged in free-living genus Caenorhabditis. We show that the most abundant small RNAs released from the nematode parasite are not microRNAs as previously thought, but rather secondary small interfering RNAs (siRNAs) that are produced by RNA-dependent RNA Polymerases. The siRNAs that are released in EVs have distinct evolutionary properties compared to those resident in free-living or parasitic nematodes. Immunoprecipitation of exWAGO demonstrates that it specifically associates with siRNAs from transposons and newly evolved repetitive elements that are packaged in EVs and released into the host environment. Together this work demonstrates molecular and evolutionary selectivity in the small RNA sequences that are released in EVs into the host environment and identifies a novel Argonaute protein as the mediator of this. © The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.

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April 21, 2020

The CF Canada-Sick Kids Program in individual CF therapy: A resource for the advancement of personalized medicine in CF.

Therapies targeting certain CFTR mutants have been approved, yet variations in clinical response highlight the need for in-vitro and genetic tools that predict patient-specific clinical outcomes. Toward this goal, the CF Canada-Sick Kids Program in Individual CF Therapy (CFIT) is generating a “first of its kind”, comprehensive resource containing patient-specific cell cultures and data from 100 CF individuals that will enable modeling of therapeutic responses.The CFIT program is generating: 1) nasal cells from drug naïve patients suitable for culture and the study of drug responses in vitro, 2) matched gene expression data obtained by sequencing the RNA from the primary nasal tissue, 3) whole genome sequencing of blood derived DNA from each of the 100 participants, 4) induced pluripotent stem cells (iPSCs) generated from each participant’s blood sample, 5) CRISPR-edited isogenic control iPSC lines and 6) prospective clinical data from patients treated with CF modulators.To date, we have recruited 57 of 100 individuals to CFIT, most of whom are homozygous for F508del (to assess in-vitro: in-vivo correlations with respect to ORKAMBI response) or heterozygous for F508del and a minimal function mutation. In addition, several donors are homozygous for rare nonsense and missense mutations. Nasal epithelial cell cultures and matched iPSC lines are available for many of these donors.This accessible resource will enable development of tools that predict individual outcomes to current and emerging modulators targeting F508del-CFTR and facilitate therapy discovery for rare CF causing mutations.Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

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April 21, 2020

Full-length transcriptome analysis of Litopenaeus vannamei reveals transcript variants involved in the innate immune system.

To better understand the immune system of shrimp, this study combined PacBio isoform sequencing (Iso-Seq) and Illumina paired-end short reads sequencing methods to discover full-length immune-related molecules of the Pacific white shrimp, Litopenaeus vannamei. A total of 72,648 nonredundant full-length transcripts (unigenes) were generated with an average length of 2545 bp from five main tissues, including the hepatopancreas, cardiac stomach, heart, muscle, and pyloric stomach. These unigenes exhibited a high annotation rate (62,164, 85.57%) when compared against NR, NT, Swiss-Prot, Pfam, GO, KEGG and COG databases. A total of 7544 putative long noncoding RNAs (lncRNAs) were detected and 1164 nonredundant full-length transcripts (449 UniTransModels) participated in the alternative splicing (AS) events. Importantly, a total of 5279 nonredundant full-length unigenes were successfully identified, which were involved in the innate immune system, including 9 immune-related processes, 19 immune-related pathways and 10 other immune-related systems. We also found wide transcript variants, which increased the number and function complexity of immune molecules; for example, toll-like receptors (TLRs) and interferon regulatory factors (IRFs). The 480 differentially expressed genes (DEGs) were significantly higher or tissue-specific expression patterns in the hepatopancreas compared with that in other four tested tissues (FDR <0.05). Furthermore, the expression levels of six selected immune-related DEGs and putative IRFs were validated using real-time PCR technology, substantiating the reliability of the PacBio Iso-seq results. In conclusion, our results provide new genetic resources of long-read full-length transcripts data and information for identifying immune-related genes, which are an invaluable transcriptomic resource as genomic reference, especially for further exploration of the innate immune and defense mechanisms of shrimp. Copyright © 2019 Elsevier Ltd. All rights reserved.

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April 21, 2020

Current advances in HIV vaccine preclinical studies using Macaque models.

The macaque simian or simian/human immunodeficiency virus (SIV/SHIV) challenge model has been widely used to inform and guide human vaccine trials. Substantial advances have been made recently in the application of repeated-low-dose challenge (RLD) approach to assess SIV/SHIV vaccine efficacies (VE). Some candidate HIV vaccines have shown protective effects in preclinical studies using the macaque SIV/SHIV model but the model’s true predictive value for screening potential HIV vaccine candidates needs to be evaluated further. Here, we review key parameters used in the RLD approach and discuss their relevance for evaluating VE to improve preclinical studies of candidate HIV vaccines.Crown Copyright © 2019. Published by Elsevier Ltd. All rights reserved.

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April 21, 2020

Analyses of four new Caulobacter Phicbkviruses indicate independent lineages.

Bacteriophages with genomes larger than 200 kbp are considered giant phages, and the giant Phicbkviruses are the most frequently isolated Caulobacter crescentus phages. In this study, we compare six bacteriophage genomes that differ from the genomes of the majority of Phicbkviruses. Four of these genomes are much larger than those of the rest of the Phicbkviruses, with genome sizes that are more than 250 kbp. A comparison of 16 Phicbkvirus genomes identified a ‘core genome’ of 69 genes that is present in all of these Phicbkvirus genomes, as well as shared accessory genes and genes that are unique for each phage. Most of the core genes are clustered into the regions coding for structural proteins or those involved in DNA replication. A phylogenetic analysis indicated that these 16 CaulobacterPhicbkvirus genomes are related, but they represent four distinct branches of the Phicbkvirus genomic tree with distantly related branches sharing little nucleotide homology. In contrast, pairwise comparisons within each branch of the phylogenetic tree showed that more than 80?% of the entire genome is shared among phages within a group. This conservation of the genomes within each branch indicates that horizontal gene transfer events between the groups are rare. Therefore, the Phicbkvirus genus consists of at least four different phylogenetic branches that are evolving independently from one another. One of these branches contains a 27-gene inversion relative to the other three branches. Also, an analysis of the tRNA genes showed that they are relatively mobile within the Phicbkvirus genus.

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April 21, 2020

Fudania jinshanensis gen. nov., sp. nov., isolated from faeces of the Tibetan antelope (Pantholops hodgsonii) in China.

Two hitherto unknown bacteria (strains 313T and 352) were recovered from the faeces of Tibetan antelopes on the Tibet-Qinghai Plateau, PR China. Cells were rod-shaped and Gram-stain-positive. The optimal growth conditions were at 37?°C and pH 7. The isolates were closely related to Actinotignum sanguinis (92.6?% 16S rRNA gene sequence similarity), Arcanobacterium haemolyticum (92.5?%), Actinotignum schaalii (92.4?%), Actinobaculum massiliense (92.2?%) and Flaviflexus huanghaiensis (91.6?%). Phylogenetic analyses showed that strains 313T and 352 clustered independently in the vicinity of the genera Actinotignum, Actinobaculum and Flaviflexus, but could not be classified clearly as a member of any of these genera. Phylogenomic analysis also indicated that strains 313T and 352 formed an independent branch in the family Actinomycetaceae. The major cellular fatty acids of the strains were C16?:?0 and C18?:?1?9c. The polar lipids comprised diphosphatidylglycerol, phosphatidylinositol mannoside, phosphatidylglycerol, phosphatidylinositol and five unidentified components. The peptidoglycan contained lysine, alanine and glutamic acid. The respiratory quinone was absent. The whole-cell sugars included glucose and rhamnose. The DNA G+C?content of strain 313T was 60.6?mol%. Based on the low 16S rRNA gene sequence similarities, its taxonomic position in the phylogenetic and phylogenomic trees and its unique lipid pattern, we propose that strains 313T and 352 represent members of a novel species in a new genus, for which the name Fudania jinshanensis gen. nov., sp. nov. is proposed. The type strain is 313T (=CGMCC 4.7453T=DSM 106216T).

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April 21, 2020

Genetic and biochemical characterization of FRI-3, a novel variant of the Ambler class A carbapenemase FRI-1.

To characterize a new variant of the FRI class A carbapenemase isolated from an MDR clinical Enterobacter cloacae isolate.A carbapenem-resistant E. cloacae was isolated from a rectal swab from a patient in an ICU in Southern Germany. Various phenotypic tests confirmed production of a putative class A carbapenemase. The new bla gene variant, blaFRI-3, and its genetic environment were characterized by WGS. Biochemical characterization was performed by heterologous expression in Escherichia coli TOP10 and by purification of the enzyme with subsequent determination of its kinetic parameters.PCR and sequencing carried out for different class A carbapenemase genes confirmed the presence of a novel variant of blaFRI-1. The novel variant was named FRI-3 and exhibited 91%, 96% and 92% amino acid identity to FRI-1, FRI-2 and FRI-4, respectively. E. coli TOP10 expressing blaFRI-3 showed increased resistance to almost all ß-lactams. Comparing the catalytic behaviour of FRI-3 and FRI-1, it was shown that FRI-3 had the same substrate spectrum, but basically hydrolysed ß-lactams less efficiently than FRI-1. WGS data revealed that blaFRI-3 was located on a 111?kb plasmid.The biochemical characterization of FRI-3 illustrates that even a few differences in the amino acid sequence can lead to altered catalytic activities of ß-lactamases belonging to the same family. © The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.

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April 21, 2020

Vaccine-induced protection from homologous tier 2 SHIV challenge in nonhuman primates depends on serum-neutralizing antibody titers.

Passive administration of HIV neutralizing antibodies (nAbs) can protect macaques from hard-to-neutralize (tier 2) chimeric simian-human immunodeficiency virus (SHIV) challenge. However, conditions for nAb-mediated protection after vaccination have not been established. Here, we selected groups of 6 rhesus macaques with either high or low serum nAb titers from a total of 78 animals immunized with recombinant native-like (SOSIP) Env trimers. Repeat intrarectal challenge with homologous tier 2 SHIVBG505 led to rapid infection in unimmunized and low-titer animals. High-titer animals, however, demonstrated protection that was gradually lost as nAb titers waned over time. An autologous serum ID50 nAb titer of ~1:500 afforded more than 90% protection from medium-dose SHIV infection. In contrast, antibody-dependent cellular cytotoxicity and T cell activity did not correlate with protection. Therefore, Env protein-based vaccination strategies can protect against hard-to-neutralize SHIV challenge in rhesus macaques by inducing tier 2 nAbs, provided appropriate neutralizing titers can be reached and maintained. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

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April 21, 2020

Biomimetic hydroxyapatite nanocrystals are an active carrier for Salmonella bacteriophages.

The use of bacteriophages represents a valid alternative to conventional antimicrobial treatments, overcoming the widespread bacterial antibiotic resistance phenomenon. In this work, we evaluated whether biomimetic hydroxyapatite (HA) nanocrystals are able to enhance some properties of bacteriophages. The final goal of this study was to demonstrate that biomimetic HA nanocrystals can be used for bacteriophage delivery in the context of bacterial infections, and contribute – at the same time – to enhance some of the biological properties of the same bacteriophages such as stability, preservation, antimicrobial activity, and so on.Phage isolation and characterization were carried out by using Mitomycin C and following double-layer agar technique. The biomimetic HA water suspension was synthesized in order to obtain nanocrystals with plate-like morphology and nanometric dimensions. The interaction of phages with the HA was investigated by dynamic light scattering and Zeta potential analyses. The cytotoxicity and intracellular killing activities of the phage-HA complex were evaluated in human hepatocellular carcinoma HepG2 cells. The bacterial inhibition capacity of the complex was assessed on chicken minced meat samples infected with Salmonella Rissen.Our data highlighted that the biomimetic HA nanocrystal-bacteriophage complex was more stable and more effective than phages alone in all tested experimental conditions.Our results evidenced the important contribution of biomimetic HA nanocrystals: they act as an excellent carrier for bacteriophage delivery and enhance its biological characteristics. This study confirmed the significant role of the mineral HA when it is complexed with biological entities like bacteriophages, as it has been shown for molecules such as lactoferrin.

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