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April 21, 2020

A Novel Bacteriophage Exclusion (BREX) System Encoded by the pglX Gene in Lactobacillus casei Zhang.

The bacteriophage exclusion (BREX) system is a novel prokaryotic defense system against bacteriophages. To our knowledge, no study has systematically characterized the function of the BREX system in lactic acid bacteria. Lactobacillus casei Zhang is a probiotic bacterium originating from koumiss. By using single-molecule real-time sequencing, we previously identified N6-methyladenine (m6A) signatures in the genome of L. casei Zhang and a putative methyltransferase (MTase), namely, pglX This work further analyzed the genomic locus near the pglX gene and identified it as a component of the BREX system. To decipher the biological role of pglX, an L. casei Zhang pglX mutant (?pglX) was constructed. Interestingly, m6A methylation of the 5′-ACRCAG-3′ motif was eliminated in the ?pglX mutant. The wild-type and mutant strains exhibited no significant difference in morphology or growth performance in de Man-Rogosa-Sharpe (MRS) medium. A significantly higher plasmid acquisition capacity was observed for the ?pglX mutant than for the wild type if the transformed plasmids contained pglX recognition sites (i.e., 5′-ACRCAG-3′). In contrast, no significant difference was observed in plasmid transformation efficiency between the two strains when plasmids lacking pglX recognition sites were tested. Moreover, the ?pglX mutant had a lower capacity to retain the plasmids than the wild type, suggesting a decrease in genetic stability. Since the Rebase database predicted that the L. casei PglX protein was bifunctional, as both an MTase and a restriction endonuclease, the PglX protein was heterologously expressed and purified but failed to show restriction endonuclease activity. Taken together, the results show that the L. casei Zhang pglX gene is a functional adenine MTase that belongs to the BREX system.IMPORTANCELactobacillus casei Zhang is a probiotic that confers beneficial effects on the host, and it is thus increasingly used in the dairy industry. The possession of an effective bacterial immune system that can defend against invasion of phages and exogenous DNA is a desirable feature for industrial bacterial strains. The bacteriophage exclusion (BREX) system is a recently described phage resistance system in prokaryotes. This work confirmed the function of the BREX system in L. casei and that the methyltransferase (pglX) is an indispensable part of the system. Overall, our study characterizes a BREX system component gene in lactic acid bacteria. Copyright © 2019 American Society for Microbiology.

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April 21, 2020

First Draft Genome Sequence of a Pearl Millet Blast Pathogen, Magnaporthe grisea Strain PMg_Dl, Obtained Using PacBio Single-Molecule Real-Time and Illumina NextSeq 500 Sequencing.

The first draft genome sequence of the pearl millet blast pathogen Magnaporthe grisea PMg_Dl from India is presented. The genome information of M. grisea will be useful to understand the Magnaporthe speciation, genetic diversity, environmental adaptation, and pathogenic and host range determinants.Copyright © 2019 Prakash et al.

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April 21, 2020

Draft Genome Sequences of Type VI Secretion System-Encoding Vibrio fischeri Strains FQ-A001 and ES401.

The type VI secretion system (T6SS) facilitates lethal competition between bacteria through direct contact. Comparative genomics has facilitated the study of these systems in Vibrio fischeri, which colonizes the squid host Euprymna scolopes Here, we report the draft genome sequences of two lethal V. fischeri strains that encode the T6SS, FQ-A001 and ES401.Copyright © 2019 Bultman et al.

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April 21, 2020

Plantibacter flavus, Curtobacterium herbarum, Paenibacillus taichungensis, and Rhizobium selenitireducens Endophytes Provide Host-Specific Growth Promotion of Arabidopsis thaliana, Basil, Lettuce, and Bok Choy Plants.

A collection of bacterial endophytes isolated from stem tissues of plants growing in soils highly contaminated with petroleum hydrocarbons were screened for plant growth-promoting capabilities. Twenty-seven endophytic isolates significantly improved the growth of Arabidopsis thaliana plants in comparison to that of uninoculated control plants. The five most beneficial isolates, one strain each of Curtobacterium herbarum, Paenibacillus taichungensis, and Rhizobium selenitireducens and two strains of Plantibacter flavus were further examined for growth promotion in Arabidopsis, lettuce, basil, and bok choy plants. Host-specific plant growth promotion was observed when plants were inoculated with the five bacterial strains. P. flavus strain M251 increased the total biomass and total root length of Arabidopsis plants by 4.7 and 5.8 times, respectively, over that of control plants and improved lettuce and basil root growth, while P. flavus strain M259 promoted Arabidopsis shoot and root growth, lettuce and basil root growth, and bok choy shoot growth. A genome comparison between P. flavus strains M251 and M259 showed that both genomes contain up to 70 actinobacterial putative plant-associated genes and genes involved in known plant-beneficial pathways, such as those for auxin and cytokinin biosynthesis and 1-aminocyclopropane-1-carboxylate deaminase production. This study provides evidence of direct plant growth promotion by Plantibacter flavusIMPORTANCE The discovery of new plant growth-promoting bacteria is necessary for the continued development of biofertilizers, which are environmentally friendly and cost-efficient alternatives to conventional chemical fertilizers. Biofertilizer effects on plant growth can be inconsistent due to the complexity of plant-microbe interactions, as the same bacteria can be beneficial to the growth of some plant species and neutral or detrimental to others. We examined a set of bacterial endophytes isolated from plants growing in a unique petroleum-contaminated environment to discover plant growth-promoting bacteria. We show that strains of Plantibacter flavus exhibit strain-specific plant growth-promoting effects on four different plant species.Copyright © 2019 American Society for Microbiology.

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April 21, 2020

Complete Genome Sequences of Two USA300-Related Community-Associated Methicillin-Resistant Staphylococcus aureus Clinical Isolates.

USA300 is a predominant community-associated methicillin-resistant Staphylococcus aureus strain causing significant morbidity and mortality in North America. We present the full annotated genome sequences of two methicillin-resistant Staphylococcus aureus isolates related to the USA300 pulsotype with the goal of studying the evolutionary relationships of this highly successful strain type.Copyright © 2019 McClure and Zhang.

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April 21, 2020

Resequencing the Genome of Malassezia restricta Strain KCTC 27527.

The draft genome sequence of Malassezia restricta KCTC 27527, a clinical isolate from a patient with dandruff, was previously reported. Using the PacBio Sequel platform, we completed and reannotated the genome of M. restricta KCTC 27527 for a better understanding of the genome of this fungus.Copyright © 2019 Cho et al.

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April 21, 2020

Sequence and Evolutionary Features for the Alternatively Spliced Exons of Eukaryotic Genes.

Alternative splicing of pre-mRNAs is a crucial mechanism for maintaining protein diversity in eukaryotes without requiring a considerable increase of genes in the number. Due to rapid advances in high-throughput sequencing technologies and computational algorithms, it is anticipated that alternative splicing events will be more intensively studied to address different kinds of biological questions. The occurrences of alternative splicing mean that all exons could be classified to be either constitutively or alternatively spliced depending on whether they are virtually included into all mature mRNAs. From an evolutionary point of view, therefore, the alternatively spliced exons would have been associated with distinctive biological characteristics in comparison with constitutively spliced exons. In this paper, we first outline the representative types of alternative splicing events and exon classification, and then review sequence and evolutionary features for the alternatively spliced exons. The main purpose is to facilitate understanding of the biological implications of alternative splicing in eukaryotes. This knowledge is also helpful to establish computational approaches for predicting the splicing pattern of exons.

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April 21, 2020

Transposable Elements Adaptive Role in Genome Plasticity, Pathogenicity and Evolution in Fungal Phytopathogens.

Transposable elements (TEs) are agents of genetic variability in phytopathogens as they are a source of adaptive evolution through genome diversification. Although many studies have uncovered information on TEs, the exact mechanism behind TE-induced changes within the genome remains poorly understood. Furthermore, convergent trends towards bigger genomes, emergence of novel genes and gain or loss of genes implicate a TE-regulated genome plasticity of fungal phytopathogens. TEs are able to alter gene expression by revamping the cis-regulatory elements or recruiting epigenetic control. Recent findings show that TEs recruit epigenetic control on the expression of effector genes as part of the coordinated infection strategy. In addition to genome plasticity and diversity, fungal pathogenicity is an area of economic concern. A survey of TE distribution suggests that their proximity to pathogenicity genes TEs may act as sites for emergence of novel pathogenicity factors via nucleotide changes and expansion or reduction of the gene family. Through a systematic survey of literature, we were able to conclude that the role of TEs in fungi is wide: ranging from genome plasticity, pathogenicity to adaptive behavior in evolution. This review also identifies the gaps in knowledge that requires further elucidation for a better understanding of TEs’ contribution to genome architecture and versatility.

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April 21, 2020

Genomic Diversity and Recombination among Xylella fastidiosa Subspecies.

Xylella fastidiosa is an economically important bacterial plant pathogen. With insights gained from 72 genomes, this study investigated differences among the three main subspecies, which have allopatric origins: X. fastidiosa subsp. fastidiosa, multiplex, and pauca The origin of recombinogenic X. fastidiosa subsp. morus and sandyi was also assessed. The evolutionary rate of the 622 genes of the species core genome was estimated at the scale of an X. fastidiosa subsp. pauca subclade (7.62?×?10-7 substitutions per site per year), which was subsequently used to estimate divergence time for the subspecies and introduction events. The study characterized genes present in the accessory genome of each of the three subspecies and investigated the core genome to detect genes potentially under positive selection. Recombination is recognized to be the major driver of diversity in X. fastidiosa, potentially facilitating shifts to novel plant hosts. The relative effect of recombination in comparison to point mutation was calculated (r/m?=?2.259). Evidence of recombination was uncovered in the core genome alignment; X. fastidiosa subsp. fastidiosa in the United States was less prone to recombination, with an average of 3.22 of the 622 core genes identified as recombining regions, whereas a specific clade of X. fastidiosa subsp. multiplex was found to have on average 9.60 recombining genes, 93.2% of which originated from X. fastidiosa subsp. fastidiosa Interestingly, for X. fastidiosa subsp. morus, which was initially thought to be the outcome of genome-wide recombination between X. fastidiosa subsp. fastidiosa and X. fastidiosa subsp. multiplex, intersubspecies homologous recombination levels reached 15.30% in the core genome. Finally, there is evidence of X. fastidiosa subsp. pauca strains from citrus containing genetic elements acquired from strains infecting coffee plants as well as genetic elements from both X. fastidiosa subsp. fastidiosa and X. fastidiosa subsp. multiplex In summary, our data provide new insights into the evolution and epidemiology of this plant pathogen.IMPORTANCEXylella fastidiosa is an important vector-borne plant pathogen. We used a set of 72 genomes that constitutes the largest assembled data set for this bacterial species so far to investigate genetic relationships and the impact of recombination on phylogenetic clades and to compare genome content at the subspecies level, and we used a molecular dating approach to infer the evolutionary rate of X. fastidiosa The results demonstrate that recombination is important in shaping the genomes of X. fastidiosa and that each of the main subspecies is under different selective pressures. We hope insights from this study will improve our understanding of X. fastidiosa evolution and biology.Copyright © 2019 American Society for Microbiology.

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April 21, 2020

A high-quality genome assembly from a single, field-collected spotted lanternfly (Lycorma delicatula) using the PacBio Sequel II system

Background A high-quality reference genome is an essential tool for applied and basic research on arthropods. Long-read sequencing technologies may be used to generate more complete and contiguous genome assemblies than alternate technologies; however, long-read methods have historically had greater input DNA requirements and higher costs than next-generation sequencing, which are barriers to their use on many samples. Here, we present a 2.3 Gb de novo genome assembly of a field-collected adult female spotted lanternfly (Lycorma delicatula) using a single Pacific Biosciences SMRT Cell. The spotted lanternfly is an invasive species recently discovered in the northeastern United States that threatens to damage economically important crop plants in the region. Results The DNA from 1 individual was used to make 1 standard, size-selected library with an average DNA fragment size of ~20 kb. The library was run on 1 Sequel II SMRT Cell 8M, generating a total of 132 Gb of long-read sequences, of which 82 Gb were from unique library molecules, representing ~36× coverage of the genome. The assembly had high contiguity (contig N50 length = 1.5 Mb), completeness, and sequence level accuracy as estimated by conserved gene set analysis (96.8% of conserved genes both complete and without frame shift errors). Furthermore, it was possible to segregate more than half of the diploid genome into the 2 separate haplotypes. The assembly also recovered 2 microbial symbiont genomes known to be associated with L. delicatula, each microbial genome being assembled into a single contig. Conclusions We demonstrate that field-collected arthropods can be used for the rapid generation of high-quality genome assemblies, an attractive approach for projects on emerging invasive species, disease vectors, or conservation efforts of endangered species.

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April 21, 2020

The Complete Genome of the Atypical Enteropathogenic Escherichia coli Archetype Isolate E110019 Highlights a Role for Plasmids in Dissemination of the Type III Secreted Effector EspT.

Enteropathogenic Escherichia coli (EPEC) is a leading cause of moderate to severe diarrhea among young children in developing countries, and EPEC isolates can be subdivided into two groups. Typical EPEC (tEPEC) bacteria are characterized by the presence of both the locus of enterocyte effacement (LEE) and the plasmid-encoded bundle-forming pilus (BFP), which are involved in adherence and translocation of type III effectors into the host cells. Atypical EPEC (aEPEC) bacteria also contain the LEE but lack the BFP. In the current report, we describe the complete genome of outbreak-associated aEPEC isolate E110019, which carries four plasmids. Comparative genomic analysis demonstrated that the type III secreted effector EspT gene, an autotransporter gene, a hemolysin gene, and putative fimbrial genes are all carried on plasmids. Further investigation of 65 espT-containing E. coli genomes demonstrated that different espT alleles are associated with multiple plasmids that differ in their overall gene content from the E110019 espT-containing plasmid. EspT has been previously described with respect to its role in the ability of E110019 to invade host cells. While other type III secreted effectors of E. coli have been identified on insertion elements and prophages of the chromosome, we demonstrated in the current study that the espT gene is located on multiple unique plasmids. These findings highlight a role of plasmids in dissemination of a unique E. coli type III secreted effector that is involved in host invasion and severe diarrheal illness.Copyright © 2019 American Society for Microbiology.

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April 21, 2020

Advantage of the F2:A1:B- IncF Pandemic Plasmid over IncC Plasmids in In Vitro Acquisition and Evolution of blaCTX-M Gene-Bearing Plasmids in Escherichia coli.

Despite a fitness cost imposed on bacterial hosts, large conjugative plasmids play a key role in the diffusion of resistance determinants, such as CTX-M extended-spectrum ß-lactamases. Among the large conjugative plasmids, IncF plasmids are the most predominant group, and an F2:A1:B- IncF-type plasmid encoding a CTX-M-15 variant was recently described as being strongly associated with the emerging worldwide Escherichia coli sequence type 131 (ST131)-O25b:H4 H30Rx/C2 sublineage. In this context, we investigated the fitness cost of narrow-range F-type plasmids, including the F2:A1:B- IncF-type CTX-M-15 plasmid, and of broad-range C-type plasmids in the K-12-like J53-2 E. coli strain. Although all plasmids imposed a significant fitness cost to the bacterial host immediately after conjugation, we show, using an experimental-evolution approach, that a negative impact on the fitness of the host strain was maintained throughout 1,120 generations with the IncC-IncR plasmid, regardless of the presence or absence of cefotaxime, in contrast to the F2:A1:B- IncF plasmid, whose cost was alleviated. Many chromosomal and plasmid rearrangements were detected after conjugation in transconjugants carrying the IncC plasmids but not in transconjugants carrying the F2:A1:B- IncF plasmid, except for insertion sequence (IS) mobilization from the fliM gene leading to the restoration of motility of the recipient strains. Only a few mutations occurred on the chromosome of each transconjugant throughout the experimental-evolution assay. Our findings indicate that the F2:A1:B- IncF CTX-M-15 plasmid is well adapted to the E. coli strain studied, contrary to the IncC-IncR CTX-M-15 plasmid, and that such plasmid-host adaptation could participate in the evolutionary success of the CTX-M-15-producing pandemic E. coli ST131-O25b:H4 lineage.Copyright © 2019 Mahérault et al.

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April 21, 2020

A Highly Unusual V1 Region of Env in an Elite Controller of HIV Infection.

HIV elite controllers represent a remarkable minority of patients who maintain normal CD4+ T-cell counts and low or undetectable viral loads for decades in the absence of antiretroviral therapy. To examine the possible contribution of virus attenuation to elite control, we obtained a primary HIV-1 isolate from an elite controller who had been infected for 19?years, the last 10 of which were in the absence of antiretroviral therapy. Full-length sequencing of this isolate revealed a highly unusual V1 domain in Envelope (Env). The V1 domain in this HIV-1 strain was 49 amino acids, placing it in the top 1% of lengths among the 6,112 Env sequences in the Los Alamos National Laboratory online database. Furthermore, it included two additional N-glycosylation sites and a pair of cysteines suggestive of an extra disulfide loop. Virus with this Env retained good infectivity and replicative capacity; however, analysis of recombinant viruses suggested that other sequences in Env were adapted to accommodate the unusual V1 domain. While the long V1 domain did not confer resistance to neutralization by monoclonal antibodies of the V1/V2-glycan-dependent class, it did confer resistance to neutralization by monoclonal antibodies of the V3-glycan-dependent class. Our findings support results in the literature that suggest a role for long V1 regions in shielding HIV-1 from recognition by V3-directed broadly neutralizing antibodies. In the case of the elite controller described here, it seems likely that selective pressures from the humoral immune system were responsible for driving the highly unusual polymorphisms present in this HIV-1 Envelope.IMPORTANCE Elite controllers have long provided an avenue for researchers to reveal mechanisms underlying control of HIV-1. While the role of host genetic factors in facilitating elite control is well known, the possibility of infection by attenuated strains of HIV-1 has been much less studied. Here we describe an unusual viral feature found in an elite controller of HIV-1 infection and demonstrate its role in conferring escape from monoclonal antibodies of the V3-glycan class. Our results suggest that extreme variation may be needed by HIV-1 to escape neutralization by some antibody specificities. Copyright © 2019 Silver et al.

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April 21, 2020

Salmonella Genomic Island 3 Is an Integrative and Conjugative Element and Contributes to Copper and Arsenic Tolerance of Salmonella enterica.

Salmonella genomic island 3 (SGI3) was first described as a chromosomal island in Salmonella 4,[5],12:i:-, a monophasic variant of Salmonella enterica subsp. enterica serovar Typhimurium. The SGI3 DNA sequence detected from Salmonella 4,[5],12:i:- isolated in Japan was identical to that of a previously reported one across entire length of 81?kb. SGI3 consists of 86 open reading frames, including a copper homeostasis and silver resistance island (CHASRI) and an arsenic tolerance operon, in addition to genes related to conjugative transfer and DNA replication or partitioning, suggesting that the island is a mobile genetic element. We successfully selected transconjugants that acquired SGI3 after filter-mating experiments using the S. enterica serovars Typhimurium, Heidelberg, Hadar, Newport, Cerro, and Thompson as recipients. Southern blot analysis using I-CeuI-digested genomic DNA demonstrated that SGI3 was integrated into a chromosomal fragment of the transconjugants. PCR and sequencing analysis demonstrated that SGI3 was inserted into the 3′ end of the tRNA genes pheV or pheR The length of the target site was 52 or 55?bp, and a 55-bp attI sequence indicating generation of the circular form of SGI3 was also detected. The transconjugants had a higher MIC against CuSO4 compared to the recipient strains under anaerobic conditions. Tolerance was defined by the cus gene cluster in the CHASRI. The transconjugants also had distinctly higher MICs against Na2HAsO4 compared to recipient strains under aerobic conditions. These findings clearly demonstrate that SGI3 is an integrative and conjugative element and contributes to the copper and arsenic tolerance of S. enterica.Copyright © 2019 American Society for Microbiology.

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April 21, 2020

Medusavirus, a Novel Large DNA Virus Discovered from Hot Spring Water.

Recent discoveries of new large DNA viruses reveal high diversity in their morphologies, genetic repertoires, and replication strategies. Here, we report the novel features of medusavirus, a large DNA virus newly isolated from hot spring water in Japan. Medusavirus, with a diameter of 260?nm, shows a T=277 icosahedral capsid with unique spherical-headed spikes on its surface. It has a 381-kb genome encoding 461 putative proteins, 86 of which have their closest homologs in Acanthamoeba, whereas 279 (61%) are orphan genes. The virus lacks the genes encoding DNA topoisomerase II and RNA polymerase, showing that DNA replication takes place in the host nucleus, whereas the progeny virions are assembled in the cytoplasm. Furthermore, the medusavirus genome harbored genes for all five types of histones (H1, H2A, H2B, H3, and H4) and one DNA polymerase, which are phylogenetically placed at the root of the eukaryotic clades. In contrast, the host amoeba encoded many medusavirus homologs, including the major capsid protein. These facts strongly suggested that amoebae are indeed the most promising natural hosts of medusavirus, and that lateral gene transfers have taken place repeatedly and bidirectionally between the virus and its host since the early stage of their coevolution. Medusavirus reflects the traces of direct evolutionary interactions between the virus and eukaryotic hosts, which may be caused by sharing the DNA replication compartment and by evolutionarily long lasting virus-host relationships. Based on its unique morphological characteristics and phylogenomic relationships with other known large DNA viruses, we propose that medusavirus represents a new family, MedusaviridaeIMPORTANCE We have isolated a new nucleocytoplasmic large DNA virus (NCLDV) from hot spring water in Japan, named medusavirus. This new NCLDV is phylogenetically placed at the root of the eukaryotic clades based on the phylogenies of several key genes, including that encoding DNA polymerase, and its genome surprisingly encodes the full set of histone homologs. Furthermore, its laboratory host, Acanthamoeba castellanii, encodes many medusavirus homologs in its genome, including the major capsid protein, suggesting that the amoeba is the genuine natural host from ancient times of this newly described virus and that lateral gene transfers have repeatedly occurred between the virus and amoeba. These results suggest that medusavirus is a unique NCLDV preserving ancient footprints of evolutionary interactions with its hosts, thus providing clues to elucidate the evolution of NCLDVs, eukaryotes, and virus-host interaction. Based on the dissimilarities with other known NCLDVs, we propose that medusavirus represents a new viral family, Medusaviridae.Copyright © 2019 Yoshikawa et al.

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