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April 21, 2020

Characterization of vanM carrying clinical Enterococcus isolates and diversity of the suppressed vanM gene cluster.

Here we report the prevalence of the suppressed vanM gene cluster as a reservoir of vancomycin resistance genes. Among 1284 clinical isolates of enterococci from four hospitals in Hangzhou, China, 55 isolates of Enterococcus faecium and one isolate of Enterococcus faecalis were screened positive for the vanM genotype. Antimicrobial susceptibility testing showed that 55 of the 56 vanM-positive isolates were susceptible to vancomycin and teicoplanin. Most of them (54/56) belonged to the main epidemic lineage CC17, mostly the ST78 type. The vanM gene clusters in the 55 vancomycin-susceptible isolates showed sequence diversity owing to different insertion locations of IS1216E. The vanM transposons could be classified into five types and they all carried two or more IS1216E elements, leading to complete or partial deletions of vanR, vanS, or vanX. Quantitative reverse transcription polymerase chain reaction showed that the expression level of vanM was significantly lower in the vancomycin-susceptible isolates than in the vancomycin-resistant isolate. Considering the prevalence of the vanM genotype and the potential for conversion to a resistant phenotype, vanM might act as an important determinant of glycopeptide resistance in the future. It is essential to strengthen the surveillance of vanM-containing enterococci to control the dissemination of vancomycin resistance. Copyright © 2018. Published by Elsevier B.V.


April 21, 2020

Rapid and Focused Maturation of a VRC01-Class HIV Broadly Neutralizing Antibody Lineage Involves Both Binding and Accommodation of the N276-Glycan.

The VH1-2 restricted VRC01-class of antibodies targeting the HIV envelope CD4 binding site are a major focus of HIV vaccine strategies. However, a detailed analysis of VRC01-class antibody development has been limited by the rare nature of these responses during natural infection and the lack of longitudinal sampling of such responses. To inform vaccine strategies, we mapped the development of a VRC01-class antibody lineage (PCIN63) in the subtype C infected IAVI Protocol C neutralizer PC063. PCIN63 monoclonal antibodies had the hallmark VRC01-class features and demonstrated neutralization breadth similar to the prototype VRC01 antibody, but were 2- to 3-fold less mutated. Maturation occurred rapidly within ~24 months of emergence of the lineage and somatic hypermutations accumulated at key contact residues. This longitudinal study of broadly neutralizing VRC01-class antibody lineage reveals early binding to the N276-glycan during affinity maturation, which may have implications for vaccine design.Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.


April 21, 2020

Epidemiologic and genomic insights on mcr-1-harbouring Salmonella from diarrhoeal outpatients in Shanghai, China, 2006-2016.

Colistin resistance mediated by mcr-1-harbouring plasmids is an emerging threat in Enterobacteriaceae, like Salmonella. Based on its major contribution to the diarrhoea burden, the epidemic state and threat of mcr-1-harbouring Salmonella in community-acquired infections should be estimated.This retrospective study analysed the mcr-1 gene incidence in Salmonella strains collected from a surveillance on diarrhoeal outpatients in Shanghai Municipality, China, 2006-2016. Molecular characteristics of the mcr-1-positive strains and their plasmids were determined by genome sequencing. The transfer abilities of these plasmids were measured with various conjugation strains, species, and serotypes.Among the 12,053 Salmonella isolates, 37 mcr-1-harbouring strains, in which 35 were serovar Typhimurium, were detected first in 2012 and with increasing frequency after 2015. Most patients infected with mcr-1-harbouring strains were aged <5?years. All strains, including fluoroquinolone-resistant and/or extended-spectrum ß-lactamase-producing strains, were multi-drug resistant. S. Typhimurium had higher mcr-1 plasmid acquisition ability compared with other common serovars. Phylogeny based on the genomes combined with complete plasmid sequences revealed some clusters, suggesting the presence of mcr-1-harbouring Salmonella outbreaks in the community. Most mcr-1-positive strains were clustered together with the pork strains, strongly suggesting pork consumption as a main infection source.The mcr-1-harbouring Salmonella prevalence in community-acquired diarrhoea displays a rapid increase trend, and the ESBL-mcr-1-harbouring Salmonella poses a threat for children. These findings highlight the necessary and significance of prohibiting colistin use in animals and continuous monitoring of mcr-1-harbouring Salmonella.Copyright © 2019. Published by Elsevier B.V.


April 21, 2020

The role of genomic structural variation in the genetic improvement of polyploid crops

Many of our major crop species are polyploids, containing more than one genome or set of chromosomes. Polyploid crops present unique challenges, including difficulties in genome assembly, in discriminating between multiple gene and sequence copies, and in genetic mapping, hindering use of genomic data for genetics and breeding. Polyploid genomes may also be more prone to containing structural variation, such as loss of gene copies or sequences (presence–absence variation) and the presence of genes or sequences in multiple copies (copy-number variation). Although the two main types of genomic structural variation commonly identified are presence–absence variation and copy-number variation, we propose that homeologous exchanges constitute a third major form of genomic structural variation in polyploids. Homeologous exchanges involve the replacement of one genomic segment by a similar copy from another genome or ancestrally duplicated region, and are known to be extremely common in polyploids. Detecting all kinds of genomic structural variation is challenging, but recent advances such as optical mapping and long-read sequencing offer potential strategies to help identify structural variants even in complex polyploid genomes. All three major types of genomic structural variation (presence–absence, copy-number, and homeologous exchange) are now known to influence phenotypes in crop plants, with examples of flowering time, frost tolerance, and adaptive and agronomic traits. In this review, we summarize the challenges of genome analysis in polyploid crops, describe the various types of genomic structural variation and the genomics technologies and data that can be used to detect them, and collate information produced to date related to the impact of genomic structural variation on crop phenotypes. We highlight the importance of genomic structural variation for the future genetic improvement of polyploid crops.


April 21, 2020

Real time monitoring of Aeromonas salmonicida evolution in response to successive antibiotic therapies in a commercial fish farm.

Our ability to predict evolutionary trajectories of pathogens in response to antibiotic pressure is one of the promising leverage to fight against the present antibiotic resistance worldwide crisis. Yet, few studies tackled this question in situ at the outbreak level, due to the difficulty to link a given pathogenic clone evolution with its precise antibiotic exposure over time. In this study, we monitored the real-time evolution of an Aeromonas salmonicida clone in response to successive antibiotic and vaccine therapies in a commercial fish farm. The clone was responsible for a four-year outbreak of furunculosis within a Recirculating Aquaculture System Salmo salar farm in China, and we reconstructed the precise tempo of mobile genetic elements (MGEs) acquisition events during this period. The resistance profile provided by the acquired MGEs closely mirrored the antibiotics used to treat the outbreak, and we evidenced that two subclonal groups developed similar resistances although unrelated MGE acquisitions. Finally, we also demonstrated the efficiency of vaccination in outbreak management and its positive effect on antibiotic resistance prevalence. Our study provides unprecedented knowledge critical to understand evolutionary trajectories of resistant pathogens outside the laboratory. © 2019 Society for Applied Microbiology and John Wiley & Sons Ltd.


April 21, 2020

Substantial Heritable Variation in Recombination Rate on Multiple Scales in Honeybees and Bumblebees.

Meiotic recombination shuffles genetic variation and promotes correct segregation of chromosomes. Rates of recombination vary on several scales, both within genomes and between individuals, and this variation is affected by both genetic and environmental factors. Social insects have extremely high rates of recombination, although the evolutionary causes of this are not known. Here, we estimate rates of crossovers and gene conversions in 22 colonies of the honeybee, Apis mellifera, and 9 colonies of the bumblebee, Bombus terrestris, using direct sequencing of 299 haploid drone offspring. We confirm that both species have extremely elevated crossover rates, with higher rates measured in the highly eusocial honeybee than the primitively social bumblebee. There are also significant differences in recombination rate between subspecies of honeybee. There is substantial variation in genome-wide recombination rate between individuals of both A. mellifera and B. terrestris and the distribution of these rates overlap between species. A large proportion of interindividual variation in recombination rate is heritable, which indicates the presence of variation in trans-acting factors that influence recombination genome-wide. We infer that levels of crossover interference are significantly lower in honeybees compared to bumblebees, which may be one mechanism that contributes to higher recombination rates in honeybees. We also find a significant increase in recombination rate with distance from the centromere, mirrored by methylation differences. We detect a strong transmission bias due to GC-biased gene conversion associated with noncrossover gene conversions. Our results shed light on the mechanistic causes of extreme rates of recombination in social insects and the genetic architecture of recombination rate variation. Copyright © 2019 by the Genetics Society of America.


April 21, 2020

Insights into the evolution and drug susceptibility of Babesia duncani from the sequence of its mitochondrial and apicoplast genomes.

Babesia microti and Babesia duncani are the main causative agents of human babesiosis in the United States. While significant knowledge about B. microti has been gained over the past few years, nothing is known about B. duncani biology, pathogenesis, mode of transmission or sensitivity to currently recommended therapies. Studies in immunocompetent wild type mice and hamsters have shown that unlike B. microti, infection with B. duncani results in severe pathology and ultimately death. The parasite factors involved in B. duncani virulence remain unknown. Here we report the first known completed sequence and annotation of the apicoplast and mitochondrial genomes of B. duncani. We found that the apicoplast genome of this parasite consists of a 34?kb monocistronic circular molecule encoding functions that are important for apicoplast gene transcription as well as translation and maturation of the organelle’s proteins. The mitochondrial genome of B. duncani consists of a 5.9?kb monocistronic linear molecule with two inverted repeats of 48?bp at both ends. Using the conserved cytochrome b (Cytb) and cytochrome c oxidase subunit I (coxI) proteins encoded by the mitochondrial genome, phylogenetic analysis revealed that B. duncani defines a new lineage among apicomplexan parasites distinct from B. microti, Babesia bovis, Theileria spp. and Plasmodium spp. Annotation of the apicoplast and mitochondrial genomes of B. duncani identified targets for development of effective therapies. Our studies set the stage for evaluation of the efficacy of these drugs alone or in combination against B. duncani in culture as well as in animal models.Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.


April 21, 2020

Competition between mobile genetic elements drives optimization of a phage-encoded CRISPR-Cas system: insights from a natural arms race.

CRISPR-Cas systems function as adaptive immune systems by acquiring nucleotide sequences called spacers that mediate sequence-specific defence against competitors. Uniquely, the phage ICP1 encodes a Type I-F CRISPR-Cas system that is deployed to target and overcome PLE, a mobile genetic element with anti-phage activity in Vibrio cholerae. Here, we exploit the arms race between ICP1 and PLE to examine spacer acquisition and interference under laboratory conditions to reconcile findings from wild populations. Natural ICP1 isolates encode multiple spacers directed against PLE, but we find that single spacers do not interfere equally with PLE mobilization. High-throughput sequencing to assay spacer acquisition reveals that ICP1 can also acquire spacers that target the V. cholerae chromosome. We find that targeting the V. cholerae chromosome proximal to PLE is sufficient to block PLE and is dependent on Cas2-3 helicase activity. We propose a model in which indirect chromosomal spacers are able to circumvent PLE by Cas2-3-mediated processive degradation of the V. cholerae chromosome before PLE mobilization. Generally, laboratory-acquired spacers are much more diverse than the subset of spacers maintained by ICP1 in nature, showing how evolutionary pressures can constrain CRISPR-Cas targeting in ways that are often not appreciated through in vitro analyses. This article is part of a discussion meeting issue ‘The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems’.


April 21, 2020

Reduced frequency of HIV superinfection in a high-risk cohort in Zambia.

Rates of HIV-1 superinfection, re-infection with a genetically distinct virus despite HIV-1 specific immune responses, vary in different risk populations. We previously found the rates of superinfection were similar to primary HIV infection (PHI) in a Zambian heterosexual transmission cohort. Here, we conduct a similar analysis of 47 HIV-positive Zambians from an acute infection cohort with more frequent follow-up, all infected by non-spousal partners. We identified only one case of superinfection in the first two years, significantly fewer than in our previous study, which was likely due to increased counseling during acute infection and an overall population-wide decline in factors associated with HIV transmission. The predominant virus detected after superinfection was a recombinant of the transmitted founder (TF) and the superinfecting strain. The superinfected individual mounted a neutralizing antibody response to the primary TF virus, which remained TF-specific over time and even after superinfection, did not neutralize the superinfecting variant.Copyright © 2019. Published by Elsevier Inc.


April 21, 2020

Development of CRISPR-Cas systems for genome editing and beyond

The development of clustered regularly interspaced short-palindromic repeat (CRISPR)-Cas systems for genome editing has transformed the way life science research is conducted and holds enormous potential for the treatment of disease as well as for many aspects of biotech- nology. Here, I provide a personal perspective on the development of CRISPR-Cas9 for genome editing within the broader context of the field and discuss our work to discover novel Cas effectors and develop them into additional molecular tools. The initial demonstra- tion of Cas9-mediated genome editing launched the development of many other technologies, enabled new lines of biological inquiry, and motivated a deeper examination of natural CRISPR-Cas systems, including the discovery of new types of CRISPR-Cas systems. These new discoveries in turn spurred further technological developments. I review these exciting discoveries and technologies as well as provide an overview of the broad array of applications of these technologies in basic research and in the improvement of human health. It is clear that we are only just beginning to unravel the potential within microbial diversity, and it is quite likely that we will continue to discover other exciting phenomena, some of which it may be possible to repurpose as molecular technologies. The transformation of mysterious natural phenomena to powerful tools, however, takes a collective effort to discover, characterize, and engineer them, and it has been a privilege to join the numerous researchers who have contributed to this transformation of CRISPR-Cas systems.


April 21, 2020

Wild relatives of maize

Crop domestication changed the course of human evolution, and domestication of maize (Zea mays L. subspecies mays), today the world’s most important crop, enabled civilizations to flourish and has played a major role in shaping the world we know today. Archaeological and ethnobotanical research help us understand the development of the cultures and the movements of the peoples who carried maize to new areas where it continued to adapt. Ancient remains of maize cobs and kernels have been found in the place of domestication, the Balsas River Valley (~9,000 years before present era), and the cultivation center, the Tehuacan Valley (~5,000 years before present era), and have been used to study the process of domestication. Paleogenomic data showed that some of the genes controlling the stem and inflorescence architecture were comparable to modern maize, while other genes controlling ear shattering and starch biosynthesis retain high levels of variability, similar to those found in the wild relative teosinte. These results indicate that the domestication process was both gradual and complex, where different genetic loci were selected at different points in time, and that the transformation of teosinte to maize was completed in the last 5,000 years. Mesoamerican native cultures domesticated teosinte and developed maize from a 6 cm long, popping-kernel ear to what we now recognize as modern maize with its wide variety in ear size, kernel texture, color, size, and adequacy for diverse uses and also invented nixtamalization, a process key to maximizing its nutrition. Used directly for human and animal consumption, processed food products, bioenergy, and many cultural applications, it is now grown on six of the world’s seven continents. The study of its evolution and domestication from the wild grass teosinte helps us understand the nature of genetic diversity of maize and its wild relatives and gene expression. Genetic barriers to direct use of teosinte or Tripsacum in maize breeding have challenged our ability to identify valuable genes and traits, let alone incorporate them into elite, modern varieties. Genomic information and newer genetic technologies will facilitate the use of wild relatives in crop improvement; hence it is more important than ever to ensure their conservation and availability, fundamental to future food security. In situ conservation efforts dedicated to preserving remnant populations of wild relatives in Mexico are key to safeguarding the genetic diversity of maize and its genepool, as well as enabling these species to continue to adapt to dynamic climate and environmental changes. Genebank ex situ efforts are crucial to securely maintain collected wild relative resources and to provide them for gene discovery and other research efforts.


April 21, 2020

Genome and plasmid diversity of Extended-Spectrum ß-Lactamase-producing Escherichia coli ST131 – tracking phylogenetic trajectories with Bayesian inference.

Clonal lineages of ESBL (Extended-Spectrum ß-Lactamase)-producing E. coli belonging to sequence type 131 (ST131) have disseminated globally during the last 30 years, leading to an increased prevalence of resistance to fluoroquinolones and extended-spectrum cephalosporins in clinical isolates of E. coli. We aimed to study if Swedish ESBL-producing ST131 isolates originated from single or multiple introductions to the population by assessing the amount of genetic variation, on chromosomal and plasmid level, between Swedish and international E. coli ST131. Bayesian inference of Swedish E. coli ST131 isolates (n?=?29), sequenced using PacBio RSII, together with an international ST131 dataset showed that the Swedish isolates were part of the international ST131 A, C1 and C2 clades. Highly conserved plasmids were identified in three clusters although they were separated by several years, which indicates a strong co-evolution between some ST131 lineages and specific plasmids. In conclusion, the tight clonal relationship observed within the ST131 clades, together with highly conserved plasmids, challenges investigation of strain transmission events. A combination of few SNPs on a genome-wide scale and an epidemiological temporospatial link, are needed to track the spread of the ST131 subclones.


April 21, 2020

Differences in resource use lead to coexistence of seed-transmitted microbial populations.

Seeds are involved in the vertical transmission of microorganisms in plants and act as reservoirs for the plant microbiome. They could serve as carriers of pathogens, making the study of microbial interactions on seeds important in the emergence of plant diseases. We studied the influence of biological disturbances caused by seed transmission of two phytopathogenic agents, Alternaria brassicicola Abra43 (Abra43) and Xanthomonas campestris pv. campestris 8004 (Xcc8004), on the structure and function of radish seed microbial assemblages, as well as the nutritional overlap between Xcc8004 and the seed microbiome, to find seed microbial residents capable of outcompeting this pathogen. According to taxonomic and functional inference performed on metagenomics reads, no shift in structure and function of the seed microbiome was observed following Abra43 and Xcc8004 transmission. This lack of impact derives from a limited overlap in nutritional resources between Xcc8004 and the major bacterial populations of radish seeds. However, two native seed-associated bacterial strains belonging to Stenotrophomonas rhizophila displayed a high overlap with Xcc8004 regarding the use of resources; they might therefore limit its transmission. The strategy we used may serve as a foundation for the selection of seed indigenous bacterial strains that could limit seed transmission of pathogens.


April 21, 2020

Whole genome sequencing identifies bacterial factors affecting transmission of multidrug-resistant tuberculosis in a high-prevalence setting.

Whole genome sequencing (WGS) can elucidate Mycobacterium tuberculosis (Mtb) transmission patterns but more data is needed to guide its use in high-burden settings. In a household-based TB transmissibility study in Peru, we identified a large MIRU-VNTR Mtb cluster (148 isolates) with a range of resistance phenotypes, and studied host and bacterial factors contributing to its spread. WGS was performed on 61 of the 148 isolates. We compared transmission link inference using epidemiological or genomic data and estimated the dates of emergence of the cluster and antimicrobial drug resistance (DR) acquisition events by generating a time-calibrated phylogeny. Using a set of 12,032 public Mtb genomes, we determined bacterial factors characterizing this cluster and under positive selection in other Mtb lineages. Four of the 61 isolates were distantly related and the remaining 57 isolates diverged ca. 1968 (95%HPD: 1945-1985). Isoniazid resistance arose once and rifampin resistance emerged subsequently at least three times. Emergence of other DR types occurred as recently as within the last year of sampling. We identified five cluster-defining SNPs potentially contributing to transmissibility. In conclusion, clusters (as defined by MIRU-VNTR typing) may be circulating for decades in a high-burden setting. WGS allows for an enhanced understanding of transmission, drug resistance, and bacterial fitness factors.


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