Metagenomic approaches to assess bacteriophages in various environmental niches

Bacteriophages are ubiquitous and numerous parasites of bacteria and play a critical evolutionary role in virtually every ecosystem, yet our understanding of the extent of the diversity and role of phages remains inadequate for many ecological niches, particularly in cases in which the host is unculturable. During the past 15 years, the emergence of the field of viral metagenomics has drastically enhanced our ability to analyse the so-called viral ‘dark matter’ of the biosphere. Here, we review the evolution of viral metagenomic methodologies, as well as providing an overview of some of the most significant applications and findings in this field of research

The Pragmatic School of Thought in Open Science Practice: A Case Study of Multi-stakeholder Participation in Shaping the Future of Internet Governance

The internet is a disruptive technology that continues to define our modern world. However, numerous ethical challenges remain for internet governance going forward, e.g. surveillance capitalism, terrorism and radicalisation. The ‘pragmatic’ school of thought in open science advocates for collaboration between diverse stakeholder groups (e.g. citizens, academics, practitioners, policymakers) to ensure an informed, and positive imprint for change. However, our understanding of how open science can be used for assimilating knowledge on complex socio-political issues remains nascent. To address this gap, we present findings from ‘We, the Internet’, a global consultation project which utilised open science practices such as stakeholder-led evaluations and open access publications to engage stakeholders in dialogue around the future of internet governance. Our findings discuss emergent themes on the future of internet governance, and highlight the potential of open science to mobilise groups and combat public scepticism in policy-making

GoferBot: A Visual Guided Human-Robot Collaborative Assembly System

The current transformation towards smart manufacturing has led to a growing demand for human-robot collaboration (HRC) in the manufacturing process. Perceiving and understanding the human co-worker's behaviour introduces challenges for collaborative robots to efficiently and effectively perform tasks in unstructured and dynamic environments. Integrating recent data-driven machine vision capabilities into HRC systems is a logical next step in addressing these challenges. However, in these cases, off-the-shelf components struggle due to generalisation limitations. Real-world evaluation is required in order to fully appreciate the maturity and robustness of these approaches. Furthermore, understanding the pure-vision aspects is a crucial first step before combining multiple modalities in order to understand the limitations. In this paper, we propose GoferBot, a novel vision-based semantic HRC system for a real-world assembly task. It is composed of a visual servoing module that reaches and grasps assembly parts in an unstructured multi-instance and dynamic environment, an action recognition module that performs human action prediction for implicit communication, and a visual handover module that uses the perceptual understanding of human behaviour to produce an intuitive and efficient collaborative assembly experience. GoferBot is a novel assembly system that seamlessly integrates all sub-modules by utilising implicit semantic information purely from visual perception

Functional and structural dissection of the tape measure protein of lactococcal phage TP901-1

The tail tape measure protein (TMP) of tailed bacteriophages (also called phages) dictates the tail length and facilitates DNA transit to the cell cytoplasm during infection. Here, a thorough mutational analysis of the TMP from lactococcal phage TP901-1 (TMPTP901-1) was undertaken. We generated 56 mutants aimed at defining TMPTP901-1 domains that are essential for tail assembly and successful infection. Through analysis of the derived mutants, we determined that TP901-1 infectivity requires the N-terminal 154 aa residues, the C-terminal 60 residues and the first predicted hydrophobic region of TMPTP901-1 as a minimum. Furthermore, the role of TMPTP901-1 in tail length determination was visualized by electron microscopic imaging of TMP-deletion mutants. The inverse linear correlation between the extent of TMPTP901-1-encoding gene deletions and tail length of the corresponding virion provides an estimate of TMPTP901-1 regions interacting with the connector or involved in initiator complex formation. This study represents the most thorough characterisation of a TMP from a Gram-positive host-infecting phage and provides essential advances to understanding its role in virion assembly, morphology and infection

Using the Rosat Catalogue to find Counterparts for Unidentified Objects in the 1st Fermi/LAT Catalogue

There are a total of 1451 gamma-ray emitting objects in the Fermi Large Area Telescope First Source Catalogue. The point source location accuracy of typically a few arcminutes has allowed the counterparts for many of these sources to be found at other wavelengths, but even so there are 630 which are described as having no plausible counterpart at 80% confidence. In order to help identify the unknown objects, we have cross-correlated the positions of these sources with the Rosat All Sky Survey Bright Source Catalogue. In this way, for Fermi sources which have a possible counterpart in soft X-rays, we can use the, much smaller, Rosat error box to search for identifications. We find a strong correlation between the two samples and calculate that there are about 60 sources with a Rosat counterpart. Using the Rosat error boxes we provide tentative associations for half of them, demonstrate that the majority of these are either blazars or blazar candidates and give evidence that most belong to the BL Lac class. Given that they are X-ray selected and most are high synchrotron peaked objects, which indicates the presence of high energy electrons, these sources are also good candidates for TeV emission, and therefore good probes of the extragalactic background light.Comment: 9 pages, 1 figure; Accepted for publication in MNRA

Polyploidy breaks speciation barriers in Australian burrowing frogs Neobatrachus

Polyploidy has played an important role in evolution across the tree of life but it is still unclear how polyploid lineages may persist after their initial formation. While both common and well-studied in plants, polyploidy is rare in animals and generally less understood. The Australian burrowing frog genus Neobatrachus is comprised of six diploid and three polyploid species and offers a powerful animal polyploid model system. We generated exome-capture sequence data from 87 individuals representing all nine species of Neobatrachus to investigate species-level relationships, the origin and inheritance mode of polyploid species, and the population genomic effects of polyploidy on genus-wide demography. We describe rapid speciation of diploid Neobatrachus species and show that the three independently originated polyploid species have tetrasomic or mixed inheritance. We document higher genetic diversity in tetraploids, resulting from widespread gene flow between the tetraploids, asymmetric inter-ploidy gene flow directed from sympatric diploids to tetraploids, and isolation of diploid species from each other. We also constructed models of ecologically suitable areas for each species to investigate the impact of climate on differing ploidy levels. These models suggest substantial change in suitable areas compared to past climate, which correspond to population genomic estimates of demographic histories. We propose that Neobatrachus diploids may be suffering the early genomic impacts of climate-induced habitat loss, while tetraploids appear to be avoiding this fate, possibly due to widespread gene flow. Finally, we demonstrate that Neobatrachus is an attractive model to study the effects of ploidy on the evolution of adaptation in animals

Structure and Assembly of TP901-1 Virion Unveiled by Mutagenesis

International audienceBacteriophages of the Siphoviridae family represent the most abundant viral morphology in the biosphere, yet many molecular aspects of their virion structure, assembly and associated functions remain to be unveiled. In this study, we present a comprehensive mutational and molecular analysis of the temperate Lactococcus lactis-infecting phage TP901-1. Fourteen mutations located within the structural module of TP901-1 were created; twelve mutations were designed to prevent full length translation of putative proteins by nonsense mutations, while two additional mutations caused aberrant protein production. Electron microscopy and Western blot analysis of mutant virion preparations, as well as in vitro assembly of phage mutant combinations, revealed the essential nature of many of the corresponding gene products and provided information on their biological function(s). Based on the information obtained, we propose a functional and assembly model of the TP901-1 Siphoviridae virion

Biocidal inactivation of Lactococcus lactis bacteriophages: efficacy and targets of commonly used sanitizers

Lactococcus lactis strains, being intensely used in the dairy industry, are particularly vulnerable to members of the so-called 936 group of phages. Sanitization and disinfection using purpose-made biocidal solutions is a critical step in controlling phage contamination in such dairy processing plants. The susceptibility of 36 936 group phages to biocidal treatments was examined using 14 biocides and commercially available sanitizers. The targets of a number of these biocides were investigated by means of electron microscopic and proteomic analyses. The results from this study highlight significant variations in phage resistance to biocides among 936 phages. Furthermore, rather than possessing resistance to specific biocides or biocide types, biocide-resistant phages tend to possess a broad tolerance to multiple classes of antimicrobial compounds

Ubiquitous carbohydrate binding modules decorate 936 lactococcal siphophage virions

With the availability of an increasing number of 3D structures of bacteriophage components, combined with powerful in silico predictive tools, it has become possible to decipher the structural assembly and functionality of phage adhesion devices. In the current study, we examined 113 members of the 936 group of lactococcal siphophages, and identified a number of Carbohydrate Binding Modules (CBMs) in the neck passage structure and major tail protein, on top of evolved Dit proteins, as recently reported by us. The binding ability of such CBM-containing proteins was assessed through the construction of green fluorescent protein fusion proteins and subsequent binding assays. Two CBMs, one from the phage tail and another from the neck, demonstrated definite binding to their phage-specific host. Bioinformatic analysis of the structural proteins of 936 phages reveals that they incorporate binding modules which exhibit structural homology to those found in other lactococcal phage groups and beyond, indicating that phages utilize common structural “bricks” to enhance host binding capabilities. The omnipresence of CBMs in Siphophages supports their beneficial role in the infection process, as they can be combined in various ways to form appendages with different shapes and functionalities, ensuring their success in host detection in their respective ecological niches

Needle in a Whey-Stack: PhRACS as a Discovery Tool for Unknown Phage-Host Combinations

The field of metagenomics has rapidly expanded to become the go-to method for complex microbial community analyses. However, there is currently no straightforward route from metagenomics to traditional culture-based methods of strain isolation, particularly in (bacterio)phage biology, leading to an investigative bottleneck. Here, we describe a method that exploits specific phage receptor binding protein (RBP)-host cell surface receptor interaction enabling isolation of phagehost combinations from an environmental sample. The method was successfully applied to two complex sample types-a dairy-derived whey sample and an infant fecal sample, enabling retrieval of specific and culturable phage hosts.IMPORTANCE PhRACS aims to bridge the current divide between in silico genetic analyses (i.e., phageomic studies) and traditional culture-based methodology. Through the labeling of specific bacterial hosts with fluorescently tagged recombinant phage receptor binding proteins and the isolation of tagged cells using flow cytometry, PhRACS allows the full potential of phageomic data to be realized in the wet laboratory