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September 22, 2019

The genome of Artemisia annua provides insight into the evolution of Asteraceae family and artemisinin biosynthesis.

Artemisia annua, commonly known as sweet wormwood or Qinghao, is a shrub native to China and has long been used for medicinal purposes. A. annua is now cultivated globally as the only natural source of a potent anti-malarial compound, artemisinin. Here, we report a high-quality draft assembly of the 1.74-gigabase genome of A. annua, which is highly heterozygous, rich in repetitive sequences, and contains 63 226 protein-coding genes, one of the largest numbers among the sequenced plant species. We found that, as one of a few sequenced genomes in the Asteraceae, the A. annua genome contains a large number of genes specific to this large angiosperm clade. Notably, the expansion and functional diversification of genes encoding enzymes involved in terpene biosynthesis are consistent with the evolution of the artemisinin biosynthetic pathway. We further revealed by transcriptome profiling that A. annua has evolved the sophisticated transcriptional regulatory networks underlying artemisinin biosynthesis. Based on comprehensive genomic and transcriptomic analyses we generated transgenic A. annua lines producing high levels of artemisinin, which are now ready for large-scale production and thereby will help meet the challenge of increasing global demand of artemisinin. Copyright © 2018 The Author. Published by Elsevier Inc. All rights reserved.

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September 22, 2019

Sequence analysis of IncA/C and IncI1 plasmids isolated from multidrug-resistant Salmonella Newport using Single-Molecule Real-Time Sequencing.

Multidrug-resistant (MDR) plasmids play an important role in disseminating antimicrobial resistance genes. To elucidate the antimicrobial resistance gene compositions in A/C incompatibility complex (IncA/C) plasmids carried by animal-derived MDR Salmonella Newport, and to investigate the spread mechanism of IncA/C plasmids, this study characterizes the complete nucleotide sequences of IncA/C plasmids by comparative analysis. Complete nucleotide sequencing of plasmids and chromosomes of six MDR Salmonella Newport strains was performed using PacBio RSII. Open reading frames were assigned using prokaryotic genome annotation pipeline (PGAP). To understand genomic diversity and evolutionary relationships among Salmonella Newport IncA/C plasmids, we included three complete IncA/C plasmid sequences with similar backbones from Salmonella Newport and Escherichia coli: pSN254, pAM04528, and peH4H, and additional 200 draft chromosomes. With the exception of canine isolate CVM22462, which contained an additional IncI1 plasmid, each of the six MDR Salmonella Newport strains contained only the IncA/C plasmid. These IncA/C plasmids (including references) ranged in size from 80.1 (pCVM21538) to 176.5?kb (pSN254) and carried various resistance genes. Resistance genes floR, tetA, tetR, strA, strB, sul, and mer were identified in all IncA/C plasmids. Additionally, blaCMY-2 and sugE were present in all IncA/C plasmids, excepting pCVM21538. Plasmid pCVM22462 was capable of being transferred by conjugation. The IncI1 plasmid pCVM22462b in CVM22462 carried blaCMY-2 and sugE. Our data showed that MDR Salmonella Newport strains carrying similar IncA/C plasmids clustered together in the phylogenetic tree using chromosome sequences and the IncA/C plasmids from animal-derived Salmonella Newport contained diverse resistance genes. In the current study, we analyzed genomic diversities and phylogenetic relationships among MDR Salmonella Newport using complete plasmids and chromosome sequences and provided possible spread mechanism of IncA/C plasmids in Salmonella Newport Lineage II.

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September 22, 2019

The presence of colistin resistance gene mcr-1 and -3 in ESBL producing Escherichia coli isolated from food in Ho Chi Minh City, Vietnam.

Colistin is indicated for the treatment of multidrug-resistant gram-negative bacterial infections. However, the spread of colistin-resistant bacteria harbouring an mcr gene has become a serious concern. This study investigated local foods in Vietnam for contamination with colistin-resistant bacteria. A total of 261 extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Escherichia coli isolates from 330 meat and seafood products were analysed for colistin susceptibility and the presence of mcr genes. Approximately, 24% (62/261) of ESBL- or AmpC-producing E. coli isolates showed colistin resistance; 97% (60/62) of colistin-resistant isolates harboured mcr-1, whereas 3% (2/62) harboured mcr-3. As the result of plasmid analysis of two strains, both plasmids harbouring mcr-3 revealed that plasmid replicon type was IncFII. Sequencing analysis indicated that an insertion sequence was present near mcr-3, suggesting that IncFII plasmids harbouring mcr-3 could be transferred to other bacterial species by horizontal transfer of the plasmid or transfer with some insertion sequence. In conclusion, ESBL-producing E. coli and AmpC-producing E. coli have acquired colistin resistance because 24% of such isolates show colistin resistance and 3% of the colistin-resistant strains harbour mcr-3. We reported the present of the mcr-3-carrying ESBL-producing E. coli isolated from pork in Vietnam.

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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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September 22, 2019

Mycobacterial biomaterials and resources for researchers.

There are many resources available to mycobacterial researchers, including culture collections around the world that distribute biomaterials to the general scientific community, genomic and clinical databases, and powerful bioinformatics tools. However, many of these resources may be unknown to the research community. This review article aims to summarize and publicize many of these resources, thus strengthening the quality and reproducibility of mycobacterial research by providing the scientific community access to authenticated and quality-controlled biomaterials and a wealth of information, analytical tools and research opportunities.

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September 22, 2019

Multiplex assessment of protein variant abundance by massively parallel sequencing.

Determining the pathogenicity of genetic variants is a critical challenge, and functional assessment is often the only option. Experimentally characterizing millions of possible missense variants in thousands of clinically important genes requires generalizable, scalable assays. We describe variant abundance by massively parallel sequencing (VAMP-seq), which measures the effects of thousands of missense variants of a protein on intracellular abundance simultaneously. We apply VAMP-seq to quantify the abundance of 7,801 single-amino-acid variants of PTEN and TPMT, proteins in which functional variants are clinically actionable. We identify 1,138 PTEN and 777 TPMT variants that result in low protein abundance, and may be pathogenic or alter drug metabolism, respectively. We observe selection for low-abundance PTEN variants in cancer, and show that p.Pro38Ser, which accounts for ~10% of PTEN missense variants in melanoma, functions via a dominant-negative mechanism. Finally, we demonstrate that VAMP-seq is applicable to other genes, highlighting its generalizability.

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September 22, 2019

Computational comparison of availability in CTL/gag epitopes among patients with acute and chronic HIV-1 infection.

Recent studies indicate that there is selection bias for transmission of viral polymorphisms associated with higher viral fitness. Furthermore, after transmission and before a specific immune response is mounted in the recipient, the virus undergoes a number of reversions which allow an increase in their replicative capacity. These aspects, and others, affect the viral population characteristic of early acute infection.160 singlegag-gene amplifications were obtained by limiting-dilution RT-PCR from plasma samples of 8 ARV-naïve patients with early acute infection (<30?days, 22?days average) and 8 ARV-naive patients with approximately a year of infection (10 amplicons per patient). Sanger sequencing and NGS SMRT technology (Pacific Biosciences) were implemented to sequence the amplicons. Phylogenetic analysis was performed by using MEGA 6.06. HLA-I (A and B) typing was performed by SSOP-PCR method. The chromatograms were analyzed with Sequencher 4.10. Epitopes and immune-proteosomal cleavages prediction was performed with CBS prediction server for the 30 HLA-A and -B alleles most prevalent in our population with peptide lengths from 8 to 14 mer. Cytotoxic response prediction was performed by using IEDB Analysis Resource.After implementing epitope prediction analysis, we identified a total number of 325 possible viral epitopes present in two or more acute or chronic patients. 60.3% (n?=?196) of them were present only in acute infection (prevalent acute epitopes) while 39.7% (n?=?129) were present only in chronic infection (prevalent chronic epitopes). Within p24, the difference was equally dramatic with 59.4% (79/133) being acute epitopes (p?

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September 22, 2019

Diversity and evolution of the emerging Pandoraviridae family.

With DNA genomes reaching 2.5?Mb packed in particles of bacterium-like shape and dimension, the first two Acanthamoeba-infecting pandoraviruses remained up to now the most complex viruses since their discovery in 2013. Our isolation of three new strains from distant locations and environments is now used to perform the first comparative genomics analysis of the emerging worldwide-distributed Pandoraviridae family. Thorough annotation of the genomes combining transcriptomic, proteomic, and bioinformatic analyses reveals many non-coding transcripts and significantly reduces the former set of predicted protein-coding genes. Here we show that the pandoraviruses exhibit an open pan-genome, the enormous size of which is not adequately explained by gene duplications or horizontal transfers. As most of the strain-specific genes have no extant homolog and exhibit statistical features comparable to intergenic regions, we suggest that de novo gene creation could contribute to the evolution of the giant pandoravirus genomes.

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September 22, 2019

The genome of Rhizophagus clarus HR1 reveals a common genetic basis for auxotrophy among arbuscular mycorrhizal fungi.

Mycorrhizal symbiosis is one of the most fundamental types of mutualistic plant-microbe interaction. Among the many classes of mycorrhizae, the arbuscular mycorrhizae have the most general symbiotic style and the longest history. However, the genomes of arbuscular mycorrhizal (AM) fungi are not well characterized due to difficulties in cultivation and genetic analysis. In this study, we sequenced the genome of the AM fungus Rhizophagus clarus HR1, compared the sequence with the genome sequence of the model species R. irregularis, and checked for missing genes that encode enzymes in metabolic pathways related to their obligate biotrophy.In the genome of R. clarus, we confirmed the absence of cytosolic fatty acid synthase (FAS), whereas all mitochondrial FAS components were present. A KEGG pathway map identified the absence of genes encoding enzymes for several other metabolic pathways in the two AM fungi, including thiamine biosynthesis and the conversion of vitamin B6 derivatives. We also found that a large proportion of the genes encoding glucose-producing polysaccharide hydrolases, that are present even in ectomycorrhizal fungi, also appear to be absent in AM fungi.In this study, we found several new genes that are absent from the genomes of AM fungi in addition to the genes previously identified as missing. Missing genes for enzymes in primary metabolic pathways imply that AM fungi may have a higher dependency on host plants than other biotrophic fungi. These missing metabolic pathways provide a genetic basis to explore the physiological characteristics and auxotrophy of AM fungi.

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September 22, 2019

First report of the occurrence and whole-genome characterization of Edwardsiella tarda in the false killer whale (Pseudorca crassidens).

Although several Edwardsiella tarda infections have been reported, its pathogenic role in marine mammals has not been investigated at the genome level. We investigated the genome of E. tarda strain KC-Pc-HB1, isolated from the false killer whale (Pseudorca crassidens) found bycaught in South Korea. The obtained genome was similar to that of human pathogenic E. tarda strains, but distinct from other Edwardsiella species. Although type III and VI secretion systems, which are essential for the virulence of other Edwardsiella species, were absent, several virulence-related genes involved in the pathogenesis of E. tarda were found in the genome. These results provide important insights into the E. tarda infecting marine mammals and give valuable information on potential virulence factors in this pathogen.

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September 22, 2019

The impact of Staphylococcus aureus genomic variation on clinical phenotype of children with acute hematogenous osteomyelitis.

Children with acute hematogenous osteomyelitis (AHO) have a broad spectrum of illness ranging from mild to severe. The purpose of this study is to evaluate the impact of genomic variation of Staphylococcus aureus on clinical phenotype of affected children and determine which virulence genes correlate with severity of illness.De novo whole genome sequencing was conducted for a strain of Community Acquired Methicillin Resistant Staphylococcus aureus (CA-MRSA), using PacBio Hierarchical Genome Assembly Process (HGAP) from 6 Single Molecule Real Time (SMRT) Cells, as a reference for DNA library assembly of 71 Staphylococcus aureus isolates from children with AHO. Virulence gene annotation was based on exhaustive literature review and genomic data in NCBI for Staphylococcus aureus. Clinical phenotype was assessed using a validated severity score. Kruskal-Wallis rank sum test determined association between clinical severity and virulence gene presence using False Discovery Rate (FDR), significance <0.01.PacBio produced an assembled genome of 2,898,306 bp and 2054 Open Reading Frames (ORFs). Annotation confirmed 201 virulence genes. Statistical analysis of gene presence by clinical severity found 40 genes significantly associated with severity of illness (FDR =0.009). MRSA isolates encoded a significantly greater number of virulence genes than did MSSA (p < 0.0001). Phylogenetic analysis by maximum likelihood (PAML) demonstrated the relatedness of genomic distance to clinical phenotype.The Staphylococcus aureus genome contains virulence genes which are significantly associated with severity of illness in children with osteomyelitis. This study introduces a novel reference strain and detailed annotation of Staphylococcus aureus virulence genes. While this study does not address bacterial gene expression, a platform is created for future transcriptome investigations to elucidate the complex mechanisms involved in childhood osteomyelitis.

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September 22, 2019

Footprints of parasitism in the genome of the parasitic flowering plant Cuscuta campestris.

A parasitic lifestyle, where plants procure some or all of their nutrients from other living plants, has evolved independently in many dicotyledonous plant families and is a major threat for agriculture globally. Nevertheless, no genome sequence of a parasitic plant has been reported to date. Here we describe the genome sequence of the parasitic field dodder, Cuscuta campestris. The genome contains signatures of a fairly recent whole-genome duplication and lacks genes for pathways superfluous to a parasitic lifestyle. Specifically, genes needed for high photosynthetic activity are lost, explaining the low photosynthesis rates displayed by the parasite. Moreover, several genes involved in nutrient uptake processes from the soil are lost. On the other hand, evidence for horizontal gene transfer by way of genomic DNA integration from the parasite’s hosts is found. We conclude that the parasitic lifestyle has left characteristic footprints in the C. campestris genome.

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September 22, 2019

Landscape of the genome and host cell response of Mycobacterium shigaense reveals pathogenic features.

A systems approach was used to explore the genome and transcriptome of Mycobacterium shigaense, a new opportunistic pathogen isolated from a patient with a skin infection, and the host response transcriptome was assessed using a macrophage infection model. The M. shigaense genome comprises 5,207,883?bp, with 67.2% G+C content and 5098 predicted coding genes. Evolutionarily, the bacterium belongs to a cluster in the phylogenetic tree along with three target opportunistic pathogenic strains, namely, M. avium, M. triplex and M. simiae. Potential virulence genes are indeed expressed by M. shigaense under culture conditions. Phenotypically, M. shigaense had similar infection and replication capacities in a macrophage model as the opportunistic species compared to M. tuberculosis. M. shigaense activated NF-?B, TNF, cytokines and chemokines in the host innate immune-related signaling pathways and elicited an early response shared with pathogenic bacilli except M. tuberculosis. M. shigaense upregulated specific host response genes such as TLR7, CCL4 and CXCL5. We performed an integrated and comparative analysis of M. shigaense. Multigroup comparison indicated certain differences with typical pathogenic bacilli in terms of gene features and the macrophage response.

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September 22, 2019

Comparative genomics of Campylobacter concisus: Analysis of clinical strains reveals genome diversity and pathogenic potential.

In recent years, an increasing number of Campylobacter species have been associated with human gastrointestinal (GI) diseases including gastroenteritis, inflammatory bowel disease, and colorectal cancer. Campylobacter concisus, an oral commensal historically linked to gingivitis and periodontitis, has been increasingly detected in the lower GI tract. In the present study, we generated robust genome sequence data from C. concisus strains and undertook a comprehensive pangenome assessment to identify C. concisus virulence properties and to explain potential adaptations acquired while residing in specific ecological niche(s) of the GI tract. Genomes of 53 new C. concisus strains were sequenced, assembled, and annotated including 36 strains from gastroenteritis patients, 13 strains from Crohn’s disease patients and four strains from colitis patients (three collagenous colitis and one lymphocytic colitis). When compared with previous published sequences, strains clustered into two main groups/genomospecies (GS) with phylogenetic clustering explained neither by disease phenotype nor sample location. Paired oral/faecal isolates, from the same patient, indicated that there are few genetic differences between oral and gut isolates which suggests that gut isolates most likely reflect oral strain relocation. Type IV and VI secretion systems genes, genes known to be important for pathogenicity in the Campylobacter genus, were present in the genomes assemblies, with 82% containing Type VI secretion system genes. Our findings indicate that C. concisus strains are genetically diverse, and the variability in bacterial secretion system content may play an important role in their virulence potential.

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September 22, 2019

Sea cucumber genome provides insights into saponin biosynthesis and aestivation regulation.

Echinoderms exhibit several fascinating evolutionary innovations that are rarely seen in the animal kingdom, but how these animals attained such features is not well understood. Here we report the sequencing and analysis of the genome and extensive transcriptomes of the sea cucumber Apostichopus japonicus, a species from a special echinoderm group with extraordinary potential for saponin synthesis, aestivation and organ regeneration. The sea cucumber does not possess a reorganized Hox cluster as previously assumed for all echinoderms, and the spatial expression of Hox7 and Hox11/13b potentially guides the embryo-to-larva axial transformation. Contrary to the typical production of lanosterol in animal cholesterol synthesis, the oxidosqualene cyclase of sea cucumber produces parkeol for saponin synthesis and has “plant-like” motifs suggestive of convergent evolution. The transcriptional factors Klf2 and Egr1 are identified as key regulators of aestivation, probably exerting their effects through a clock gene-controlled process. Intestinal hypometabolism during aestivation is driven by the DNA hypermethylation of various metabolic gene pathways, whereas the transcriptional network of intestine regeneration involves diverse signaling pathways, including Wnt, Hippo and FGF. Decoding the sea cucumber genome provides a new avenue for an in-depth understanding of the extraordinary features of sea cucumbers and other echinoderms.

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