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Background The prediction of bacteriophage sequences in metagenomic datasets has become a topic of considerable interest, leading to the development of many novel bioinformatic tools. A comparative analysis of ten state-of-the-art phage identification tools was performed to inform their usage in microbiome research. Methods Artificial contigs generated from complete RefSeq genomes representing phages, plasmids, and chromosomes, and a previously sequenced mock community containing four phage species, were used to evaluate the precision, recall, and F1 scores of the tools. We also generated a dataset of randomly shuffled sequences to quantify false-positive calls. In addition, a set of previously simulated viromes was used to assess diversity bias in each tool’s output. Results VIBRANT and VirSorter2 achieved the highest F1 scores (0.93) in the RefSeq artificial contigs dataset, with several other tools also performing well. Kraken2 had the highest F1 score (0.86) in the mock community benchmark by a large margin (0.3 higher than DeepVirFinder in second place), mainly due to its high precision (0.96). Generally, k-mer-based tools performed better than reference similarity tools and gene-based methods. Several tools, most notably PPR-Meta, called a high number of false positives in the randomly shuffled sequences. When analysing the diversity of the genomes that each tool predicted from a virome set, most tools produced a viral genome set that had similar alpha- and beta-diversity patterns to the original population, with Seeker being a notable exception. Conclusions This study provides key metrics used to assess performance of phage detection tools, offers a framework for further comparison of additional viral discovery tools, and discusses optimal strategies for using these tools. We highlight that the choice of tool for identification of phages in metagenomic datasets, as well as their parameters, can bias the results and provide pointers for different use case scenarios. We have also made our benchmarking dataset available for download in order to facilitate future comparisons of phage identification tools.

Colonial New Zealand was built on the ideal of creating better lives for settlers. Emigrants came looking to escape the shackles of the class-system and poor conditions in Industrial Revolution period Britain. Colonial propaganda claimed that most emigrants achieved their aims, but the lives the colonists actually experienced upon reaching New Zealand remain relatively unexplored from a biosocial perspective. In this paper we present a pilot study of stable isotope results of bone collagen from seven adults interred in the St. John’s Cemetery (SJM), Milton, New Zealand (ca. AD 1860–1900). We interpret the diet at Milton and broadly compare our isotopic results with contemporaneous samples from Britain. We show that, like contemporary Britain, the diet of our studied individuals was focused on C3 crops and terrestrial meat sources. Despite higher ????15N values in contemporary UK populations (which can simplistically be interpreted as indicative of higher meat intake), consideration of different local baselines makes it likely that this New Zealand population had relatively similar levels of meat intake. Interestingly marine resources did not form an important part of the Milton diet, despite the site's proximity to the ocean, hinting at the possible stigmatisation of local resources and the development of a European New Zealand (pākehā) food identity.

The bacteriophage vB_AhM_PVN02 (PVN02), infecting Aeromonas hydrophila, was isolated from a striped catfish pond water sample in Can Tho City, Vietnam. The phage had high lytic activity with a latent period and burst size of approximately 20 min and 105 plaque-forming units per cell, respectively. Observation of the phage by transmission electron microscopy indicated that PVN02 belongs to the family Myoviridae. The genome of PVN02 is a double-stranded linear DNA with a length in 51,668 bp and a content of 52% GC. Among the 64 genes, 16 were predicted to encode proteins with predicted functions. No virulence or antibiotic resistance genes were found in the genome, suggesting it would be a useful biocontrol agent. Classification of the phage based on sequence comparisons, phylogenetic analysis, and gene-sharing networks was carried out, and it was found to be the first representative of a new species within a previously undefined genus in the family Myoviridae. This study confirmed that PVN02 is a novel lytic phage that could potentially be used as an agent to control Aeromonas hydrophila in striped catfish in the Mekong Delta, Vietnam.

Megaphages, bacteriophages harbouring extremely large genomes, have recently been found to be ubiquitous, being described from a variety of microbiomes ranging from the animal gut to soil and freshwater systems. However, no complete marine megaphage has been identified to date. Here, using both short and long read sequencing, we assembled >900 high-quality draft viral genomes from water in the English Channel. One of these genomes included a novel megaphage, Mar_Mega_1 at >650 Kb, making it one of the largest phage genomes assembled to date. Utilising phylogenetic and network approaches, we found this phage represents a new family of megaphages. Genomic analysis showed Mar_Mega_1 shares relatively few homologues with its closest relatives, but, as with other megaphages Mar_Mega_1 contained a variety of auxiliary metabolic genes responsible for carbon metabolism and nucleotide biosynthesis, including a NADP-dependent isocitrate dehydrogenase [Idh] and nicotinamide-nucleotide amidohydrolase [PncC], which have not previously been identified in megaphages. Mar_Mega_1 was abundant in a marine virome sample and related phages are widely prevalent in the oceans.

Fine-resolution sampling of pollen, non-pollen palynomorphs and microcharcoal as well as sedimentological data in a 6-m sediment core were used to reconstruct both natural conditions and human impacts in the late fifth and early fourth millennia cal bc in the environs of the Nebelivka megasite in Kirovograd Oblast, Central Ukraine. This 238-ha site, dating to the Middle (or BII) Phase of the Trypillia culture, represents one of the first low-density urban establishments in Europe. Despite what was believed to be a sizable population, local human impacts reconstructed from the multi-proxy palaeo-ecological record were moderate in character. There was no positive evidence to indicate a depositional hiatus in the P1 core and no sign of a major ecological impact at any stage in the high-resolution record. The palaeo-ecological record indicates modest settlement agglomeration with less permanent populations rather than permanent populations of tens of thousands of people.

The European Union (EU) is at a turning point. With the bipolar order of the Cold War fast becoming a distant memory, the European Union must quickly establish itself on the global stage before it loses the opportunity to do so. With Northeast Asia fast emerging as a new economic giant and political center for world affairs, the EU must reform its Common Foreign and Security Policy in order to develop a reputation and image as a global actor of soft power, based on its long-standing values and peaceful diplomacy. However, despite major reforms in the Lisbon Treaty (2009), the EU has not been able to form cohesion amongst its members, thus hindering its progress in achieving such international recognition. This paper therefore analyses the reforms taken place so far and suggests further reforms that will build a strong foundation for a united and cohesive foreign policy. It will then look at how the reformed framework will allow the EU to establish itself as a global actor in political affairs, in particular in Northeast Asia.

In this study, we aimed to investigate the spatio-temporal distribution patterns of the picocyanobacterium Synechococcus, an important contributor to primary production in many freshwater lakes. Our study sites were four lakes with different trophic states within the Osterseen Lake District in Southern Germany. Flow cytometry counts and next-generation sequencing were used from April to October 2015 to analyse the occurrence of Synechococcus and heterotrophic prokaryotes in relation to physical and chemical habitat properties. Synechococcus was identified as the main representative of the autotrophic picoplankton, but cell counts varied widely. The Synechococcus taxa identified by flow cytometry were confirmed by sequencing data, but the comparison of the flow cytometry counts and sequence data revealed discrepancies for cells in the exponential phase. The eutrophic Lake Schiffhuettensee was dominated by algae and had the highest abundance of heterotrophic prokaryotes. The presence of distinct operational taxonomic units of Synechococcus varied seasonally and was lake-specific, indicating local niche adaptation. Our study sheds light on the ecology of these important primary producers in freshwater systems. Furthermore, the discrepancy observed with the direct comparison of the widely used methods of next-generation sequencing and flow cytometry should serve as a caveat for future data analysis.

The anaerobic gut pathogen, Clostridioides difficile, forms adherent biofilms that may play an important role in recurrent C. difficile infections. The mechanisms underlying C. difficile community formation and inter-bacterial interactions are nevertheless poorly understood. C. difficile produces AI-2, a quorum sensing molecule that modulates biofilm formation across many bacterial species. We found that a strain defective in LuxS, the enzyme that mediates AI-2 production, is defective in biofilm development in vitro. Transcriptomic analyses of biofilms formed by wild type (WT) and luxS mutant (luxS) strains revealed a downregulation of prophage loci in the luxS mutant biofilms compared to the WT. Detection of phages and eDNA within biofilms may suggest that DNA release by phage-mediated cell lysis contributes to C. difficile biofilm formation. In order to understand if LuxS mediates C. difficile crosstalk with other gut species, C. difficile interactions with a common gut bacterium, Bacteroides fragilis, were studied. We demonstrate that C. difficile growth is significantly reduced when co-cultured with B. fragilis in mixed biofilms. Interestingly, the absence of C. difficile LuxS alleviates the B. fragilis-mediated growth inhibition. Dual species RNA-sequencing analyses from single and mixed biofilms revealed differential modulation of distinct metabolic pathways for C. difficile WT, luxS and B. fragilis upon co-culture, indicating that AI-2 may be involved in induction of selective metabolic responses in B. fragilis. Overall, our data suggest that C. difficile LuxS/AI-2 utilises different mechanisms to mediate formation of single and mixed species communities.

Cyanophage genomes frequently possess the psbA gene, encoding the D1 polypeptide of photosystem II. This protein is believed to maintain host photosynthetic capacity during infection and enhance phage fitness under high-light conditions. Although the first documented cyanophage-encoded psbA gene contained a group I intron, this feature has not been widely reported since, despite a plethora of new sequences becoming available. In this study, we show that in cyanophage S-PM2, this intron is spliced during the entire infection cycle. Furthermore, we report the widespread occurrence of psbA introns in marine metagenomic libraries, and with psbA often adjacent to a homing endonuclease (HE). Bioinformatic analysis of the intergenic region between psbA and the adjacent HE gene F-CphI in S-PM2 showed the presence of an antisense RNA (asRNA) connecting these two separate genetic elements. The asRNA is co-regulated with psbA and F-CphI, suggesting its involvement with their expression. Analysis of scaffolds from global ocean survey datasets shows this asRNA to be commonly associated with the 3′ end of cyanophage psbA genes, implying that this potential mechanism of regulating marine ‘viral’ photosynthesis is evolutionarily conserved. Although antisense transcription is commonly found in eukaryotic and increasingly also in prokaryotic organisms, there has been no indication for asRNAs in lytic phages so far. We propose that this asRNA also provides a means of preventing the formation of mobile group I introns within cyanophage psbA genes.\ud \ud

Tuberculosis (TB) was once a major killer in Europe, but it is unclear how the strains and patterns of infection at ‘peak TB' relate to what we see today. Here we describe 14 genome sequences of M. tuberculosis, representing 12 distinct genotypes, obtained from human remains from eighteenth-century Hungary using metagenomics. All our historic genotypes belong to M. tuberculosis Lineage 4. Bayesian phylogenetic dating, based on samples with well-documented dates, places the most recent common ancestor of this lineage in the late Roman period. We find that most bodies yielded more than one M. tuberculosis genotype and we document an intimate epidemiological link between infections in two long-dead individuals. Our results suggest that metagenomic approaches usefully inform detection and characterization of historical and contemporary infections., Tuberculosis was once a major killer in Europe. Here the authors use metagenomics to obtain genomic sequences of Mycobacterium tuberculosis from human remains from eighteenth-century Hungary, revealing mixed infections within individuals as well as presence of the same strain in two individuals.

A bacteriophage may protect itself and its host against a deadly effect of bright sunlight.