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July 7, 2019

Divergent and convergent modes of interaction between wheat and Puccinia graminis f. sp. tritici isolates revealed by the comparative gene co-expression network and genome analyses.

Two opposing evolutionary constraints exert pressure on plant pathogens: one to diversify virulence factors in order to evade plant defenses, and the other to retain virulence factors critical for maintaining a compatible interaction with the plant host. To better understand how the diversified arsenals of fungal genes promote interaction with the same compatible wheat line, we performed a comparative genomic analysis of two North American isolates of Puccinia graminis f. sp. tritici (Pgt).The patterns of inter-isolate divergence in the secreted candidate effector genes were compared with the levels of conservation and divergence of plant-pathogen gene co-expression networks (GCN) developed for each isolate. Comprative genomic analyses revealed substantial level of interisolate divergence in effector gene complement and sequence divergence. Gene Ontology (GO) analyses of the conserved and unique parts of the isolate-specific GCNs identified a number of conserved host pathways targeted by both isolates. Interestingly, the degree of inter-isolate sub-network conservation varied widely for the different host pathways and was positively associated with the proportion of conserved effector candidates associated with each sub-network. While different Pgt isolates tended to exploit similar wheat pathways for infection, the mode of plant-pathogen interaction varied for different pathways with some pathways being associated with the conserved set of effectors and others being linked with the diverged or isolate-specific effectors.Our data suggest that at the intra-species level pathogen populations likely maintain divergent sets of effectors capable of targeting the same plant host pathways. This functional redundancy may play an important role in the dynamic of the “arms-race” between host and pathogen serving as the basis for diverse virulence strategies and creating conditions where mutations in certain effector groups will not have a major effect on the pathogen’s ability to infect the host.

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July 7, 2019

Benzalkonium tolerance genes and outcome in Listeria monocytogenes meningitis.

Listeria monocytogenes is a food-borne pathogen that can cause meningitis. The listerial genotype ST6 has been linked to increasing rates of unfavourable outcome over time. We investigated listerial genetic variation and the relation with clinical outcome in meningitis.We sequenced 96 isolates from adults with listerial meningitis included in two prospective nationwide cohort studies by whole genome sequencing, and evaluated associations between bacterial genetic variation and clinical outcome. We validated these results by screening listerial genotypes of 445 cerebrospinal fluid and blood isolates from patients over a 30-year period from the Dutch national surveillance cohort.We identified a bacteriophage, phiLMST6 co-occurring with a novel plasmid, pLMST6, in ST6 isolates to be associated with unfavourable outcome in patients (p 2.83e-05). The plasmid carries a benzalkonium chloride tolerance gene, emrC, conferring decreased susceptibility to disinfectants used in the food-processing industry. Isolates harbouring emrC were growth inhibited at higher levels of benzalkonium chloride (median 60 mg/L versus 15 mg/L; p <0.001), and had higher MICs for amoxicillin and gentamicin compared with isolates without emrC (both p <0.001). Transformation of pLMST6 into naive strains led to benzalkonium chloride tolerance and higher MICs for gentamicin.These results show that a novel plasmid, carrying the efflux transporter emrC, is associated with increased incidence of ST6 listerial meningitis in the Netherlands. Suggesting increased disease severity, our findings warrant consideration of disinfectants used in the food-processing industry that select for resistance mechanisms and may, inadvertently, lead to increased risk of poor disease outcome. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

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July 7, 2019

Geographical structure of endosymbiotic bacteria hosted by Bathymodiolus mussels at eastern Pacific hydrothermal vents.

Chemolithoautotrophic primary production sustains dense invertebrate communities at deep-sea hydrothermal vents and hydrocarbon seeps. Symbiotic bacteria that oxidize dissolved sulfur, methane, and hydrogen gases nourish bathymodiolin mussels that thrive in these environments worldwide. The mussel symbionts are newly acquired in each generation via infection by free-living forms. This study examined geographical subdivision of the thiotrophic endosymbionts hosted by Bathymodiolus mussels living along the eastern Pacific hydrothermal vents. High-throughput sequencing data of 16S ribosomal RNA encoding gene and fragments of six protein-coding genes of symbionts were examined in the samples collected from nine vent localities at the East Pacific Rise, Galápagos Rift, and Pacific-Antarctic Ridge.Both of the parapatric sister-species, B. thermophilus and B. antarcticus, hosted the same numerically dominant phylotype of thiotrophic Gammaproteobacteria. However, sequences from six protein-coding genes revealed highly divergent symbiont lineages living north and south of the Easter Microplate and hosted by these two Bathymodiolus mussel species. High heterogeneity of symbiont haplotypes among host individuals sampled from the same location suggested that stochasticity associated with initial infections was amplified as symbionts proliferated within the host individuals. The mussel species presently contact one another and hybridize along the Easter Microplate, but the northern and southern symbionts appear to be completely isolated. Vicariance associated with orogeny of the Easter Microplate region, 2.5-5.3 million years ago, may have initiated isolation of the symbiont and host populations. Estimates of synonymous substitution rates for the protein-coding bacterial genes examined in this study were 0.77-1.62%/nucleotide/million years.Our present study reports the most comprehensive population genetic analyses of the chemosynthetic endosymbiotic bacteria based on high-throughput genetic data and extensive geographical sampling to date, and demonstrates the role of the geographical features, the Easter Microplate and geographical distance, in the intraspecific divergence of this bacterial species along the mid-ocean ridge axes in the eastern Pacific. Altogether, our results provide insights into extrinsic and intrinsic factors affecting the dispersal and evolution of chemosynthetic symbiotic partners in the hydrothermal vents along the eastern Pacific Ocean.

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July 7, 2019

HALC: High throughput algorithm for long read error correction.

The third generation PacBio SMRT long reads can effectively address the read length issue of the second generation sequencing technology, but contain approximately 15% sequencing errors. Several error correction algorithms have been designed to efficiently reduce the error rate to 1%, but they discard large amounts of uncorrected bases and thus lead to low throughput. This loss of bases could limit the completeness of downstream assemblies and the accuracy of analysis.Here, we introduce HALC, a high throughput algorithm for long read error correction. HALC aligns the long reads to short read contigs from the same species with a relatively low identity requirement so that a long read region can be aligned to at least one contig region, including its true genome region’s repeats in the contigs sufficiently similar to it (similar repeat based alignment approach). It then constructs a contig graph and, for each long read, references the other long reads’ alignments to find the most accurate alignment and correct it with the aligned contig regions (long read support based validation approach). Even though some long read regions without the true genome regions in the contigs are corrected with their repeats, this approach makes it possible to further refine these long read regions with the initial insufficient short reads and correct the uncorrected regions in between. In our performance tests on E. coli, A. thaliana and Maylandia zebra data sets, HALC was able to obtain 6.7-41.1% higher throughput than the existing algorithms while maintaining comparable accuracy. The HALC corrected long reads can thus result in 11.4-60.7% longer assembled contigs than the existing algorithms.The HALC software can be downloaded for free from this site: https://github.com/lanl001/halc .

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July 7, 2019

Determination of nucleopolyhedrovirus’ taxonomic position

To date , over 78 genomes of nucleopolyhedroviruses (NPVs) have been sequenced and deposited in NCBI. How to define a new virus from the infected larvae in the field is usually the first question. Two NPV strains, which were isolated from casuarina moth (L. xylina) and golden birdwing larvae (Troides aeacus), respectively, displayed the same question. Due to the identity of polyhedrin (polh) sequences of these two isolates to that of Lymantria dispar MNPV and Bombyx mori NPV, they are named LdMNPV-like virus and TraeNPV, provisionally. To further clarify the relationships of LdMNPV-like virus and TraeNPV to closely related NPVs, Kimura 2-parameter (K-2-P) analysis was performed. Apparently, the results of K-2-P analysis that showed LdMNPV-like virus is an LdMNPV isolate, while TraeNPV had an ambiguous relationship to BmNPV. Otherwise, MaviNPV, which is a mini-AcMNPV, also exhibited a different story by K-2-P analysis. Since K-2-P analysis could not cover all species determination issues, therefore, TraeNPV needs to be sequenced for defining its taxonomic position. For this purpose, different genomic sequencing technologies and bioinformatic analysis approaches will be discussed. We anticipated that these applications will help to exam nucleotide information of unknown species and give an insight and facilitate to this issue.

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July 7, 2019

Multiple genome sequences of exopolysaccharide-producing, brewery-associated Lactobacillus brevis strains.

Lactobacillus brevis represents one of the most relevant beer-spoiling bacteria. Besides strains causing turbidity and off flavors upon growth and metabolite formation, this species also comprises strains that produce exopolysaccharides (EPSs), which increase the viscosity of beer. Here, we report the complete genome sequences of three EPS-producing, brewery-associated L. brevis strains. Copyright © 2017 Fraunhofer et al.

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July 7, 2019

Coping with living in the soil: the genome of the parthenogenetic springtail Folsomia candida.

Folsomia candida is a model in soil biology, belonging to the family of Isotomidae, subclass Collembola. It reproduces parthenogenetically in the presence of Wolbachia, and exhibits remarkable physiological adaptations to stress. To better understand these features and adaptations to life in the soil, we studied its genome in the context of its parthenogenetic lifestyle.We applied Pacific Bioscience sequencing and assembly to generate a reference genome for F. candida of 221.7 Mbp, comprising only 162 scaffolds. The complete genome of its endosymbiont Wolbachia, was also assembled and turned out to be the largest strain identified so far. Substantial gene family expansions and lineage-specific gene clusters were linked to stress response. A large number of genes (809) were acquired by horizontal gene transfer. A substantial fraction of these genes are involved in lignocellulose degradation. Also, the presence of genes involved in antibiotic biosynthesis was confirmed. Intra-genomic rearrangements of collinear gene clusters were observed, of which 11 were organized as palindromes. The Hox gene cluster of F. candida showed major rearrangements compared to arthropod consensus cluster, resulting in a disorganized cluster.The expansion of stress response gene families suggests that stress defense was important to facilitate colonization of soils. The large number of HGT genes related to lignocellulose degradation could be beneficial to unlock carbohydrate sources in soil, especially those contained in decaying plant and fungal organic matter. Intra- as well as inter-scaffold duplications of gene clusters may be a consequence of its parthenogenetic lifestyle. This high quality genome will be instrumental for evolutionary biologists investigating deep phylogenetic lineages among arthropods and will provide the basis for a more mechanistic understanding in soil ecology and ecotoxicology.

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July 7, 2019

Characterization and genome comparisons of three Achromobacter phages of the family Siphoviridae.

In this study, we present the characterization and genomic data of three Achromobacter phages belonging to the family Siphoviridae. Phages 83-24, JWX and JWF were isolated from sewage samples in Paris and Braunschweig, respectively, and infect Achromobacter xylosoxidans, an emerging nosocomial pathogen in cystic fibrosis patients. Analysis of morphology and growth parameters revealed that phages 83-24 and JWX have similar properties, both have nearly the same head and tail measurements, and both have a burst size between 85 and 100 pfu/cell. In regard to morphological properties, JWF had a much longer and more flexible tail compared to other phages. The linear double-stranded DNAs of all three phages are terminally redundant and not circularly permutated. The complete nucleotide sequences consist of 81,541 bp for JWF, 49,714 bp for JWX and 48,216 bp for 83-24. Analysis of the genome sequences showed again that phages JWX and 83-24 are quite similar. Comparison to the GenBank database via BLASTN revealed partial similarities to Roseobacter phage RDJL phi1 and Burkholderia phage BcepGomr. In contrast, BLASTN analysis of the genome sequence of phage JWF revealed only few similarities to non-annotated prophage regions in different strains of Burkholderia and Mesorhizobium.

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July 7, 2019

Starvation and recovery in the deep-sea methanotroph Methyloprofundus sedimenti.

In the deep ocean, the conversion of methane into derived carbon and energy drives the establishment of diverse faunal communities. Yet specific biological mechanisms underlying the introduction of methane-derived carbon into the food web remain poorly described, due to a lack of cultured representative deep-sea methanotrophic prokaryotes. Here, the response of the deep-sea aerobic methanotroph Methyloprofundus sedimenti to methane starvation and recovery was characterized. By combining lipid analysis, RNA analysis, and electron cryotomography, it was shown that M. sedimenti undergoes discrete cellular shifts in response to methane starvation, including changes in headgroup-specific fatty acid saturation levels, and reductions in cytoplasmic storage granules. Methane starvation is associated with a significant increase in the abundance of gene transcripts pertinent to methane oxidation. Methane reintroduction to starved cells stimulates a rapid, transient extracellular accumulation of methanol, revealing a way in which methane-derived carbon may be routed to community members. This study provides new understanding of methanotrophic responses to methane starvation and recovery, and lays the initial groundwork to develop Methyloprofundus as a model chemosynthesizing bacterium from the deep sea.© 2016 John Wiley & Sons Ltd.

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July 7, 2019

Butterfly genomics: insights from the genome of Melitaea cinxia

The first lepidopteran genome (Bombyx mori) was published in 2004. Ten years later the genome of Melitaea cinxia came out as the third butterfly genome published, and the first eukaryotic genome sequenced in Finland. Owing to Ilkka Hanski, the M. cinxia system in the Åland Islands has become a famous model for metapopulation biology. More than 20 years of research on this system provides a strong ecological basis upon which a genetic framework could be built. Genetic knowledge is an essential addition for understanding eco-evolutionary dynamics and the genetic basis of variability in life history traits. Here we review the process of the M. cinxia genome project, its implications for lepidopteran genome evolution, and describe how the genome has been used for gene expression studies to identify genetic consequences of habitat fragmentation. Finally, we introduce some future possibilities and challenges for genomic research in M. cinxia and other Lepidoptera.

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July 7, 2019

2015 epidemic of severe Streptococcus agalactiae sequence type 283 infections in Singapore associated with the consumption of raw freshwater fish: a detailed analysis of clinical, epidemiological, and bacterial sequencing data.

Streptococcus agalactiae (group B Streptococcus [GBS]) has not been described as a foodborne pathogen. However, in 2015, a large outbreak of severe invasive sequence type (ST) 283 GBS infections in adults epidemiologically linked to the consumption of raw freshwater fish occurred in Singapore. We attempted to determine the scale of the outbreak, define the clinical spectrum of disease, and link the outbreak to contaminated fish.Time-series analysis was performed on microbiology laboratory data. Food handlers and fishmongers were screened for enteric carriage of GBS. A retrospective cohort study was conducted to assess differences in demographic and clinical characteristics of patients with invasive ST283 and non-ST283 infections. Whole-genome sequencing was performed on human and fish ST283 isolates from Singapore, Thailand, and Hong Kong.The outbreak was estimated to have started in late January 2015. Within the study cohort of 408 patients, ST283 accounted for 35.8% of cases. Patients with ST283 infection were younger and had fewer comorbidities but were more likely to develop meningoencephalitis, septic arthritis, and spinal infection. Of 82 food handlers and fishmongers screened, none carried ST283. Culture of 43 fish samples yielded 13 ST283-positive samples. Phylogenomic analysis of 161 ST283 isolates from humans and fish revealed they formed a tight clade distinguished by 93 single-nucleotide polymorphisms.ST283 is a zoonotic GBS clone associated with farmed freshwater fish, capable of causing severe disease in humans. It caused a large foodborne outbreak in Singapore and poses both a regional and potentially more widespread threat.

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July 7, 2019

Heterogeneous resistance to quizartinib in acute myeloid leukemia revealed by single-cell analysis.

Genomic studies have revealed significant branching heterogeneity in cancer. Studies of resistance to tyrosine kinase inhibitor therapy have not fully reflected this heterogeneity because resistance in individual patients has been ascribed to largely mutually exclusive on-target or off-target mechanisms in which tumors either retain dependency on the target oncogene or subvert it through a parallel pathway. Using targeted sequencing from single cells and colonies from patient samples, we demonstrate tremendous clonal diversity in the majority of acute myeloid leukemia (AML) patients with activating FLT3 internal tandem duplication mutations at the time of acquired resistance to the FLT3 inhibitor quizartinib. These findings establish that clinical resistance to quizartinib is highly complex and reflects the underlying clonal heterogeneity of AML.© 2017 by The American Society of Hematology.

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July 7, 2019

Complete genome sequence of Microbulbifer sp. CCB-MM1, a halophile isolated from Matang Mangrove Forest, Malaysia.

Microbulbifer sp. CCB-MM1 is a halophile isolated from estuarine sediment of Matang Mangrove Forest, Malaysia. Based on 16S rRNA gene sequence analysis, strain CCB-MM1 is a potentially new species of genus Microbulbifer. Here we describe its features and present its complete genome sequence with annotation. The genome sequence is 3.86 Mb in size with GC content of 58.85%, harbouring 3313 protein coding genes and 92 RNA genes. A total of 71 genes associated with carbohydrate active enzymes were found using dbCAN. Ectoine biosynthetic genes, ectABC operon and ask_ect were detected using antiSMASH 3.0. Cell shape determination genes, mreBCD operon, rodA and rodZ were annotated, congruent with the rod-coccus cell cycle of the strain CCB-MM1. In addition, putative mreBCD operon regulatory gene, bolA was detected, which might be associated with the regulation of rod-coccus cell cycle observed from the strain.

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July 7, 2019

Maize defective kernel mutant generated by insertion of a Ds element in a gene encoding a highly conserved TTI2 cochaperone.

We have used the newly engineered transposable element Dsg to tag a gene that gives rise to a defective kernel (dek) phenotype. Dsg requires the autonomous element Ac for transposition. Upon excision, it leaves a short DNA footprint that can create in-frame and frameshift insertions in coding sequences. Therefore, we could create alleles of the tagged gene that confirmed causation of the dek phenotype by the Dsg insertion. The mutation, designated dek38-Dsg, is embryonic lethal, has a defective basal endosperm transfer (BETL) layer, and results in a smaller seed with highly underdeveloped endosperm. The maize dek38 gene encodes a TTI2 (Tel2-interacting protein 2) molecular cochaperone. In yeast and mammals, TTI2 associates with two other cochaperones, TEL2 (Telomere maintenance 2) and TTI1 (Tel2-interacting protein 1), to form the triple T complex that regulates DNA damage response. Therefore, we cloned the maize Tel2 and Tti1 homologs and showed that TEL2 can interact with both TTI1 and TTI2 in yeast two-hybrid assays. The three proteins regulate the cellular levels of phosphatidylinositol 3-kinase-related kinases (PIKKs) and localize to the cytoplasm and the nucleus, consistent with known subcellular locations of PIKKs. dek38-Dsg displays reduced pollen transmission, indicating TTI2’s importance in male reproductive cell development.

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July 7, 2019

Discovering and sequencing new plant viral genomes by next-generation sequencing: description of a practical pipeline

Small-scale sequencing has improved substantially in recent decades, culminating in the development of next-generation sequencing (NGS) technologies. Modern NGS methods have helped the discovery of many new plant viruses. Nevertheless, there is still a need to establish solid assembly pipelines targeting small genomes characterised by low identities to known viral sequences. Here, we describe and discuss the fundamental steps required for discovering and sequencing new plant viral genomes by NGS. A practical pipeline and standard alternative tools used in NGS analysis are presented.

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