The Promises and Pitfalls of Machine Learning for Detecting Viruses in Aquatic Metagenomes

Tools allowing for the identification of viral sequences in host-associated and environmental metagenomes allows for a better understanding of the genetics and ecology of viruses and their hosts. Recently, new approaches using machine learning methods to distinguish viral from bacterial signal using k-mer sequence signatures were published for identifying viral contigs in metagenomes. The promise of these content-based approaches is the ability to discover new viruses, with no or few known relatives. In this perspective paper, we examine the use of the content-based machine learning tool VirFinder for the identification of viral sequences in aquatic metagenomes and explore the possibility of using ecosystem-focused models targeted to marine metagenomes. We discuss the impact of the training set composition on the tool performance and the current limitation for the retrieval of low abundance viral sequences in metagenomes. We identify potential biases that could arise from machine learning approaches for viral hunting in real-world datasets and suggest possible avenues to overcome them.National Science Foundation [1640775]Open access journal.This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Contrôle redox de la sécrétion protéique chez Saccharomyces cerevisiae

The endoplasmic reticulum (ER) is the first intracellular compartment of the protein secretion pathway. Protein maturation in this compartment involves protein folding and post-traductionnal modification including formation of disulfide bonds. The formation of disulfide bonds is operated by a highly conserved redox relay made of the thiol oxidase Ero1 and the protein disulfide isomerase (PDI). Ero1p catalyzes disulfide bond formation and relays them by thiol-disulfide exchange to PDI, which in turn oxidizes substrates. Isomerization and terminal reduction of non-native disulfide bonds both rely on a reduction system that remains to be formally identified. Studies however suggest the importance of reduced glutathione in this reducing system. GSH is small redox tripeptide exclusively synthesized in the cytosol. In this study we (i) describe the main parameters of glutathione traffic across the ER membrane (ii) identify the main actors involved in the transport and (iii) analyze the physiological impact of a modification of the ER glutathione redox state.We established a system to monitor the fluxes of glutathione from the cytosol to the ER in S. cerevisiae. To artificially increase fluxes of glutathione, we used a cell over-expressing the GSH plasma membrane transporter HGT1, which when grown in presence of glutathione import high levels of this compound. Consequently, we monitored the intracellular relocation of imported GSH by following GSH fluxes using two specific redox probes. Our data indicate that:(i) GSH is transported into the ER by facilitated diffusion along a concentration gradient. GSSG can also be imported into the ER. Similarly, stress conditions that stimulate GSH synthesis, such as heat shoc, arsenite treatment, also triggered a GSH import in the ER.(ii) GSH import in the ER is achieved by the translocon Sec61, and is regulated by the lumenal chaperone Kar2.(iii) A rapid reduction of glutathione ER redox state leads to the activation of a non-apoptotic programmed cell death pathway, usually observed during high ER stress.Les protéines destinées à la sécrétion ou adressées à la membrane transitent par le réticulum endoplasmique (RE) où elles acquièrent leur conformation native et subissent des modifications post-traductionnelles comme la formation de ponts disulfures. Dans ce compartiment, la formation de ponts disulfures repose sur l’activité de l’oxydase Ero1 et de la Protein Disulfure Isomerase (PDI). Ero1 catalyse la formation de ponts disulfures et les transmet à la PDI qui à son tour oxyde les substrats. L’isomérisation ou la réduction terminale des ponts disulfures non-natifs repose sur un système de réduction dans le RE encore non élucidé. Des études suggèrent l’importance du glutathion réduit (GSH) dans ce système de réduction. Le GSH est un tripeptide redox exclusivement synthétisé dans le cytosol. Notre étude s’attache à (i) décrire les flux de glutathion entre RE et cytosol et (ii) identifier les acteurs de ce transport (iii) comprendre l’impact d’une modification de l’homéostasie redox du glutathion sur la physiologie du RE.Nous avons établi un système permettant d’étudier les flux de glutathion entre cytosol et RE. Afin de démasquer ces flux intracellulaires, nous avons utilisé une souche de S. cerevisiae surexprimant le transporteur plasmatique du glutathion, HGT1. Ce système permet de modifier rapidement et drastiquement la concentration cytosolique de glutathion. Les flux intracellulaires engendrés sont ensuite suivis grâce à des sondes redox spécifiques du glutathion adressées dans le RE ou le cytoplasme.(i) Nos résultats suggèrent que le GSH et le GSSG sont importés dans le RE depuis le cytosol. Le GSH est transporté selon un gradient de concentration via un système de transport de diffusion facilité. Ces flux sont également observés lors de stress stimulant la synthèse de GSH (stress thermique, arsenite…).(ii) Le transport de GSH dans le lumen est assuré par le translocon Sec61, et une régulation de cet import par la chaperone luminale Kar2 est observée.(iii) une réduction rapide de l’état redox du glutathion dans le RE conduit à une mort cellulaire programmée non apoptotique, également observée lors d’autre stress RE (traitement tunicamycine)

Complete Genome Sequence of Cyanobacterial Siphovirus KBS2A

Abstract We present the genome of a cyanosiphovirus (KBS2A) that infects a marine Synechococcus sp. (strain WH7803). Unique to this genome, relative to other sequenced cyanosiphoviruses, is the absence of elements associated with integration into the host chromosome, suggesting this virus may not be able to establish a lysogenic relationship. Go to: GENOME ANNOUNCEMENT As obligate parasites, viruses can regulate their host population dynamics but also influence the structure and productivity of microbial communities (1, 2). Synechococcus species are an abundant and ecologically important group of Cyanobacteria found in freshwater and marine ecosystems worldwide. Virus-cyanobacterium interactions may have important implications for global biogeochemical cycles. The most commonly isolated cyanophages are myoviruses and podoviruses (3, 4). Siphoviruses are a third group of viruses that infect cyanobacteria, but they have received less attention (5). The genomes of 5 cyanosiphoviruses have recently become available: that of P-SS2, a siphovirus infecting Prochlorococcus (MIT9313) (6), followed by the cyanosiphoviruses S-CBS1, S-CBS2, S-CBS3, and S-CBS4, isolated from the Chesapeake Bay Estuary, all infecting Synechococcus populations (5). Here, we present the complete genome of cyanosiphophage (KBS2A, originally named KBS-S-2A), a virus that infects Synechococcus sp. strain WH7803. The virus was isolated by plaque assay from the Chesapeake Bay by plating on Synechococcus sp. WH7803. Purified virus DNA was submitted to the Broad Institute as part of the Marine Phage Sequencing Project, where it was sequenced to ~30-fold coverage using 454 pyrosequencing. Translated open reading frames (ORFs) were compared with known protein sequences using the BLASTp program. ORF annotation was aided by the use of PSI-BLAST, HHpred, gene size, and domain conservation. The genome size of KBS2A is 40,658 bp. In total, 64 ORFs have been predicted in this genome; of these, 43 have homologues in databases, and among them, 33 have been assigned to a putative function. For most (88%) predicted ORFs with homologues, homology has been found with the other cyanosiphovirus genomes. We compared the genomic arrangements of the 6 sequenced cyanosiphoviruses using dot plot and global gene homology and found no common genomic organization, suggesting strong mosaicism in the cyanosiphoviruses. In cyanophages, cyanobacterium-related proteins can be found and are often associated with photosynthesis and transcriptional regulation (6). In previously sequenced cyanosiphovirus genomes (5, 6), numerous viral genes (6 to 40 per genome) possess homology with host genes. In the case of the KBS2A genome, only 3 ORFs (coding for RNA polymerase sigma factor RpoD, HNH endonuclease, and a putative DNA polymerase) show such homology, implying less exchange (and potentially interaction) with the host genome. The first annotated cyanosiphovirus genome (that of P-SS2) showed the presence of genes identified as encoding an integrase and excisionase, which are enzymes that allow for phage integration into the host’s genome (6). Moreover, the annotation of cyanosiphoviruses S-CBS1 and S-CBS3 led to the discovery of a prophage-like structure in two sequenced Synechococcus elongatus strains (5). In phage genomes, tRNA genes serve as indicators of potential phage integration by site-specific recombination (7, 8), although recent models have offered alternative suggestions for the role of these genes (9). Sequences of this nature can, however, be found in the P-SS2 and S-CBS4 genomes. No such features (tRNAs, integrases, etc.) were found in the genome of KBS2A, suggesting that this siphovirus might be an exclusively lytic phage rather than a temperate phage. Nucleotide sequence accession number. The complete sequence of the Synechococcus phage KBS2A genome can be accessed under the GenBank accession no. HQ634187. doi: 10.1128/genomeA.00472-1

Estimation of benthic macrofauna consumption by water birds in the bay of Saint-Brieuc (France)

The biomass and productivity of benthic invertebrates were studied in the bay of Saint-Brieuc in connection with the consumption by the nine most numerous wader and duck species present on the site. This consumption is about 1.9 g ash free dry weight/m2/year, it is similar to that obtained in the bay of Mont-Saint-Michel but very lower to results from Wadden Sea. Eutrophication generally induces an increase in overall productivity of the intertidal coastal ecosystem, as it has been observed in the German part of the Wadden Sea since the 1950s. Consequently, the bird populations grow very significantly in eutrophic systems. But this is not the case in the bay of Saint-Brieuc where macrozoobentic biomass produced and consumed by birds is comparable to what was observed in the bay of Mont-Saint-Michel, an oligotrophic marine site characterized by relatively low biomass and productivity. In the bay of Saint-Brieuc excessive inputs of nutrients induce an important development of Ulva armocicana, responsible for the spectacular green tides which greatly impact this bay each summer. The proliferation of Ulva brings the nutrient flows to a standstill and by the way regulates the production of microphytoplankton and so, indirectly, the abundance of higher trophic levelsLa biomasse et la production de la macrofaune benthique de la baie de Saint-Brieuc ont été analysées, en relation avec la consommation par les neuf principales espèces de limicoles et de canards présentes sur le site. Cette consommation est estimée à 1,9 g AFDW/m2/an, résultat similaire à celui obtenu en baie du Mont-Saint Michel, mais très inférieur à ceux observés en mer des Wadden. L'eutrophisation induit généralement une augmentation de la productivité globale des écosystèmes littoraux intertidaux. C'est par exemple ce qui a été observé dans la partie allemande de la mer des Wadden depuis les années 50. En conséquence, les effectifs d'oiseaux peuvent croître de façon très marquée dans les systèmes eutrophes. Or ce n'est pas le cas en baie de Saint-Brieuc où les biomasses macrozoobenthiques produites et consommées par les oiseaux sont comparables à ce qui est observé en baie du Mont-Saint-Michel, site océanique oligotrophe caractérisé par une biomasse et une productivité relativement faibles. En baie de Saint-Brieuc, les apports excessifs de sels nutritifs induisent d'important développements d'Ulva armocicana, responsables des spectaculaires marées vertes qui impactent très fortement la baie chaque été. La prolifération d'ulves bloque les flux nutritifs et régule la production de microphytoplancton et, indirectement, l'abondance des échelons trophiques supérieurs

Quantifier les besoins énergétiques des limicoles.

National audienceLa plupart des limicoles de l'hémisphère nord nichent dans la toundra circumpolaire et passent la période de non-reproduction sur les escales migratoires et les sites d'hivernage en domaine tempéré à tropical. Pendant cette longue période internuptiale, si quelques limicoles peuvent exploiter des habitats continentaux humides, la majorité dépend des domaines intertidaux de substrat meuble ou dur, et plus particulièrement des grands sites estuariens. Au sein du compartiment benthique d'un estuaire européen classique, si on exclut les pinnipèdes, on trouve schématiquement trois grands échelons trophiques : les oiseaux et les poissons, le macrozoobenthos et les algues (microalgues et macroalgues). Cette chaîne trophique courte s'explique en grande partie par la grande diversité d'oiseaux s'alimentant en domaine intertidal. Puisque les limicoles comptent parmi les principaux prédateurs du compartiment benthique des systèmes estuariens, l'évaluation de l'énergie qu'ils prélèvent par rapport à la ressource disponible est essentielle dans l'étude des réseaux trophiques intertidaux. Dans ce chapitre, nous proposons une courte revue des méthodes classiquement utilisées pour estimer l'énergie prélevée par une communauté de limicoles en fonction de la ressource et de l'espace benthiques disponibles au sein d'un domaine intertidal donné. Nous détaillerons successivement comment quantifier les besoins énergétiques des limicoles, puis comment quantifier la valeur énergétique du macrobenthos et de sa fraction récoltable par les limicoles, et enfin prendre en compte la superficie intertidale réellement disponible pour les limicoles

BENTHIC MACROFAUNA CONSUMPTION BY WATER BIRDS

The diversity and abundance of birds present in intertidal coastal ecosystems are closely related to the biomass of benthic invertebrates. The assessment of energy consumed compared with the available resource is one of the fundamental aspects of intertidal foodweb studies. The feeding of birds on benthic invertebrates was studied in the bay of Saint-Brieuc, a 2900ha tidal bay located on the Côtes d'Armor coast (Brittany). The consumption of the nine most numerous wader and duck species present on the site was valued at 1,9gAFDW/m²/yr. This is comparable with results observed in the Mont Saint-Michel bay but much lower than results from the Wadden sea. In order to further the understanding of the predator-prey system, the taking into account of local hypsometric singularities is relevant. Thus it is possible to determine an "average available foraging area" which takes into account the foreshore exposure frequency according to tidal conditions, which represents the real usage of the foreshore by birds. In the Bay of Saint-Brieuc foreshore, the available average surface is about 1115ha, i.e. a consumption of 4,9g/m²/yr. The use of this space by birds depends on the type, density and the accessibility of prey, the sediment characteristics, and the presence of events which can cause a disturbance. The distribution of the benthic macrofauna and spatial distribution of four wader species were studied to map the main feeding areas and the benthos they host. This data set will allow the study of potential feeding habitat compared with currently exploited habitat, and to analyse the compatibility of a complex benthic resources/birds/human activities system

Libra: scalable k-mer-based tool for massive all-vs-all metagenome comparisons

Background Shotgun metagenomics provides powerful insights into microbial community biodiversity and function. Yet, inferences from metagenomic studies are often limited by dataset size and complexity and are restricted by the availability and completeness of existing databases. De novo comparative metagenomics enables the comparison of metagenomes based on their total genetic content. Results We developed a tool called Libra that performs an all-vs-all comparison of metagenomes for precise clustering based on their k-mer content. Libra uses a scalable Hadoop framework for massive metagenome comparisons, Cosine Similarity for calculating the distance using sequence composition and abundance while normalizing for sequencing depth, and a web-based implementation in iMicrobe (http://imicrobe.us) that uses the CyVerse advanced cyberinfrastructure to promote broad use of the tool by the scientific community. Conclusions A comparison of Libra to equivalent tools using both simulated and real metagenomic datasets, ranging from 80 million to 4.2 billion reads, reveals that methods commonly implemented to reduce compute time for large datasets, such as data reduction, read count normalization, and presence/absence distance metrics, greatly diminish the resolution of large-scale comparative analyses. In contrast, Libra uses all of the reads to calculate k-mer abundance in a Hadoop architecture that can scale to any size dataset to enable global-scale analyses and link microbial signatures to biological processes.National Science Foundation [1640775]Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

The Promises and Pitfalls of Machine Learning for Detecting Viruses in Aquatic Metagenomes

Tools allowing for the identification of viral sequences in host-associated and environmental metagenomes allows for a better understanding of the genetics and ecology of viruses and their hosts. Recently, new approaches using machine learning methods to distinguish viral from bacterial signal using k-mer sequence signatures were published for identifying viral contigs in metagenomes. The promise of these content-based approaches is the ability to discover new viruses, with no or few known relatives. In this perspective paper, we examine the use of the content-based machine learning tool VirFinder for the identification of viral sequences in aquatic metagenomes and explore the possibility of using ecosystem-focused models targeted to marine metagenomes. We discuss the impact of the training set composition on the tool performance and the current limitation for the retrieval of low abundance viral sequences in metagenomes. We identify potential biases that could arise from machine learning approaches for viral hunting in real-world datasets and suggest possible avenues to overcome them

Bacteroides abundance drives birth mode dependent infant gut microbiota developmental trajectories

Background and aimsBirth mode and other early life factors affect a newborn's microbial colonization with potential long-term health effects. Individual variations in early life gut microbiota development, especially their effects on the functional repertoire of microbiota, are still poorly characterized. This study aims to provide new insights into the gut microbiome developmental trajectories during the first year of life. MethodsOur study comprised 78 term infants sampled at 3 weeks, 3 months, 6 months, and 12 months (n = 280 total samples), and their mothers were sampled in late pregnancy (n = 50). Fecal DNA was subjected to shotgun metagenomic sequencing. Infant samples were studied for taxonomic and functional maturation, and maternal microbiota was used as a reference. Hierarchical clustering on taxonomic profiles was used to identify the main microbiota developmental trajectories in the infants, and their associations with perinatal and postnatal factors were assessed. ResultsIn line with previous studies, infant microbiota composition showed increased alpha diversity and decreased beta diversity by age, converging toward an adult-like profile. However, we did not observe an increase in functional alpha diversity, which was stable and comparable with the mother samples throughout all the sampling points. Using a de novo clustering approach, two main infant microbiota clusters driven by Bacteroidaceae and Clostridiaceae emerged at each time point. The clusters were associated with birth mode and their functions differed mainly in terms of biosynthetic and carbohydrate degradation pathways, some of which consistently differed between the clusters for all the time points. The longitudinal analysis indicated three main microbiota developmental trajectories, with the majority of the infants retaining their characteristic cluster until 1 year. As many as 40% of vaginally delivered infants were grouped with infants delivered by C-section due to their clear and persistent depletion in Bacteroides. Intrapartum antibiotics, any perinatal or postnatal factors, maternal microbiota composition, or other maternal factors did not explain the depletion in Bacteroides in the subset of vaginally born infants. ConclusionOur study provides an enhanced understanding of the compositional and functional early life gut microbiota trajectories, opening avenues for investigating elusive causes that influence non-typical microbiota development.Peer reviewe

Ecosystem quality and natural heritage preservation: the case ofthe littoral eutrophication and the wintering of Brent Geese Branta b. bernicla in the bay ofSaint-Brieuc (France)

The national nature reserve of Saint-Brieuc Bay, located in the north of Brittany, is a site of international importance for the wintering of the Black-bellied Brent goose Branta bernicla bernicla although its preferred food, eelgrass, is completely absent. Local monitoring points out that the green seaweeds Ulva armocicana represents 90% of the food eaten by the Brent geese. These algae are responsible for the spectacular green tides which greatly impact this bay each summer. The 400 odd tons consumed each winter by the Brent geese are insufficient to curb this phenomenon. However, the resorption policy of the nitrogenized surpluses arriving in the bay in order to fight against the aestival green tides could, in the long term, reduce this food resource and force geese to find a substitution food which, locally, could be a grass growing on salt-marshes (Puccinellia maritima) or winter cereals growing in arable lands bordering the bay. To avoid possible crop, the national nature reserve could have to increase the surface of Puccinellia maritima meadows by mowing or by converting to pasture on the climacic areas with Halimione portulacoides as is the practice on the Atlantic coast, in Aiguillon Bay. This will have to be done with caution, the areas with sea-purslanes being important feeding grounds for young fish during the high tides. It is quite singular to see that, in this bay, a wintering area of international interest for a water bird developed thanks to serious environmental perturbation and that this ornithological interest could be disrupted by current curative policies of water quality restoration, should no compensatory measures come to balance for the hoped reduction of the green tideLa réserve naturelle nationale de la baie de Saint-Brieuc, située dans le nord de la Bretagne, est un site d'hivernage d'importance internationale de la Bernache cravant à ventre sombre Branta bernicla bernicla bien que sa nourriture préférée, les zostères, en soit totalement absente. Les suivis réalisés ont permis de mettre en évidence que les Ulves Ulva armocicana, représentent 90 % de la nourriture ingérée localement par les bernaches. Ces ulves sont responsables des spectaculaires marées vertes qui impactent très fortement la baie chaque été. Les quelque 400 tonnes consommées chaque hiver par les bernaches ne permettent pas d'endiguer ce phénomène. Toutefois, la politique de résorption des excédents azotés arrivant en baie afin de lutter contre la prolifération estivale des ulves pourrait, à terme, réduire cette ressource alimentaire et amener les bernaches à devoir trouver une nourriture de substitution qui, dans la baie, pourrait être une graminée poussant sur les prés-salés (Puccinellia maritima) ou les céréales d'hiver semées dans les champs bordant la baie. Pour éviter d'éventuels dégâts dans les cultures, la réserve pourrait être amenée à agrandir les prairies salées à Puccinellia maritima existantes, soit par la fauche, soit par le pâturage sur les zones climaciques à Obiones Halimione portulacoides comme cela se pratique en baie de l'Aiguillon. Ceci devra se faire avec prudence, les zones à Obiones étant d'importantes zones de nourrissage pour les jeunes poissons fréquentant les prés-salés lors des marées hautes. Il est tout à fait singulier de voir que, dans cette baie, un hivernage d'intérêt international d'une espèce d'oiseau d'eau s'est développé grâce à un grave déséquilibre du milieu et pourrait être remis en cause par les mesures curatives en cours de mise en œuvre si aucune mesure ne vient compenser la diminution espérée des stocks d'ulves