Bacteriophages are as ubiquitous as their bacterial hosts and often more abundant. Understanding how bacteriophages control their bacterial host populations requires a number of different approaches. ...Bacteriophages can control bacterial populations through lysis, drive evolution of bacterial immunity systems through infection, provide a conduit for horizontal gene transfer and alter host metabolism by carriage of auxiliary metabolic genes. Understanding and quantifying how bacteriophages drive these processes, requires both technological developments to take measurements in situ, and laboratory-based studies to understand mechanisms. Technological advances have allowed quantification of the number of infected cells in situ, revealing far-lower levels than expected. Understanding how observations in laboratory conditions relate to what occurs in the environment, and experimental confirmation of the predicted function of phage genes from observations in environmental omics data, remains challenging.
•Multiomic approaches have increased our understanding of phage diversity in nature.•Lab studies reveal novel biology within phage-host interactions.•The challenge remains to link lab studies with diversity to understand how phages control bacterial communities (i.e. functional ecology).
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.
Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus are the most numerous photosynthetic organisms on our planet 1, 2. With a global population size of 3.6 × 1027 3, they are ...responsible for approximately 10% of global primary production 3, 4. Viruses that infect Prochlorococcus and Synechococcus (cyanophages) can be readily isolated from ocean waters 5–7 and frequently outnumber their cyanobacterial hosts 8. Ultimately, cyanophage-induced lysis of infected cells results in the release of fixed carbon into the dissolved organic matter pool 9. What is less well known is the functioning of photosynthesis during the relatively long latent periods of many cyanophages 10, 11. Remarkably, the genomes of many cyanophage isolates contain genes involved in photosynthetic electron transport (PET) 12–18 as well as central carbon metabolism 14, 15, 19, 20, suggesting that cyanophages may play an active role in photosynthesis. However, cyanophage-encoded gene products are hypothesized to maintain or even supplement PET for energy generation while sacrificing wasteful CO2 fixation during infection 17, 18, 20. Yet this paradigm has not been rigorously tested. Here, we measured the ability of viral-infected Synechococcus cells to fix CO2 as well as maintain PET. We compared two cyanophage isolates that share different complements of PET and central carbon metabolism genes. We demonstrate cyanophage-dependent inhibition of CO2 fixation early in the infection cycle. In contrast, PET is maintained throughout infection. Our data suggest a generalized strategy among marine cyanophages to redirect photosynthesis to support phage development, which has important implications for estimates of global primary production.
•Viruses of abundant marine cyanobacteria shut down CO2 fixation during infection•The strength of the shutdown is dependent on the viral gene content•Viruses encoding genes that actively shut down CO2 fixation are less productive
Puxty et al. show that marine cyanobacterial viruses modify host photosynthesis to maximize energy production but inhibit CO2 fixation. These phototrophs are responsible for ∼10% of global CO2 fixation, and up to 60% of cells are infected at any time. Therefore, this phenomenon has implications for the assessment of primary production of the biosphere.
This article summarises the activities of the Bacterial Viruses Subcommittee of the International Committee on Taxonomy of Viruses for the period of March 2021−March 2022. We provide an overview of ...the new taxa proposed in 2021, approved by the Executive Committee, and ratified by vote in 2022. Significant changes to the taxonomy of bacterial viruses were introduced: the paraphyletic morphological families
Podoviridae
,
Siphoviridae
, and
Myoviridae
as well as the order
Caudovirales
were abolished, and a binomial system of nomenclature for species was established. In addition, one order, 22 families, 30 subfamilies, 321 genera, and 862 species were newly created, promoted, or moved.
The Anderson phage typing scheme has been successfully used worldwide for epidemiological surveillance of Salmonella enterica serovar Typhimurium. Although the scheme is being replaced by whole ...genome sequence subtyping methods, it can provide a valuable model system for study of phage-host interaction. The phage typing scheme distinguishes more than 300 definitive types of Salmonella Typhimurium based on their patterns of lysis to a unique collection of 30 specific Salmonella phages. In this study, we sequenced the genomes of 28 Anderson typing phages of Salmonella Typhimurium to begin to characterize the genetic determinants that are responsible for the differences in these phage type profiles. Genomic analysis of typing phages reveals that Anderson phages can be classified into three different groups, the P22-like, ES18-like and SETP3-like clusters. Most Anderson phages are short tailed P22-like viruses (genus Lederbergvirus); but phages STMP8 and STMP18 are very closely related to the lambdoid long tailed phage ES18, and phages STMP12 and STMP13 are related to the long noncontractile tailed, virulent phage SETP3. Most of these typing phages have complex genome relationships, but interestingly, two phage pairs STMP5 and STMP16 as well as STMP12 and STMP13 differ by a single nucleotide. The former affects a P22-like protein involved in DNA passage through the periplasm during its injection, and the latter affects a gene whose function is unknown. Using the Anderson phage typing scheme would provide insights into phage biology and the development of phage therapy for the treatment of antibiotic resistant bacterial infections.
Bacterioplankton plays an essential role in aquatic ecosystems, and cyanobacteria are an influential part of the microbiome in many water bodies. In freshwaters used for recreational activities or ...drinking water, toxic cyanobacteria cause concerns due to the risk of intoxication with cyanotoxins, such as microcystins. In this study, we aimed to unmask relationships between toxicity, cyanobacterial community composition, and environmental factors. At the same time, we assessed the correlation of a genetic marker with microcystin concentration and aimed to identify the main microcystin producer. We used Illumina MiSeq sequencing to study the bacterioplankton in two recreational lakes in South Germany. We quantified a microcystin biosynthesis gene (
) using qPCR and linked this information with microcystin concentration to assess toxicity. Microcystin biosynthesis gene (
)-clone libraries were used to determine the origin of microcystin biosynthesis genes. Bloom toxicity did not alter the bacterial community composition, which was highly dynamic at the lowest taxonomic level for some phyla such as Cyanobacteria. At the OTU level, we found distinctly different degrees of temporal variation between major bacteria phyla. Cyanobacteria and Bacteroidetes showed drastic temporal changes in their community compositions, while the composition of Actinobacteria remained rather stable in both lakes. The bacterial community composition of Alpha- and Beta-proteobacteria remained stable over time in Lake Klostersee, but it showed temporal variations in Lake Bergknappweiher. The presence of potential microcystin degraders and potential algicidal bacteria amongst prevalent Bacteroidetes and Alphaproteobacteria implied a role of those co-occurring heterotrophic bacteria in cyanobacterial bloom dynamics. Comparison of both lakes studied revealed a large shared microbiome, which was shaped toward the lake specific community composition by environmental factors. Microcystin variants detected were microcystin-LR, -RR, and -YR. The maximum microcystin concentrations measured was 6.7 μg/L, a value still acceptable for recreational waters but not drinking water. Microcystin concentration correlated positively with total phosphorus and
copy number. We identified low abundant
sp. as the only microcystin producer in both lakes. Therefore, risk assessment efforts need to take into account the fact that non-dominant species may cause toxicity of the blooms observed.
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.
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.
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.
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. Video Abstract.
Antimicrobial resistance (AMR) is a major problem globally. The main bacterial organisms associated with urinary tract infection (UTI) associated sepsis are
and
along with
species. These all have AMR ...strains known as ESBL (Extended Spectrum Beta-Lactamase), which are featured on the WHO priority pathogens list as "critical" for research. Bacteriophages (phages), as viruses that can infect and kill bacteria, could provide an effective tool to tackle these AMR strains. There is currently no "gold standard" for developing a phage cocktail. Here we describe a novel approach to develop an effective phage cocktail against a set of ESBL-producing
and
largely isolated from patients in United Kingdom hospitals. By comparing different measures of phage efficacy, we show which are the most robust, and suggest an efficient screening cascade that could be used to develop phage cocktails to target other AMR bacterial species. A target panel of 38 ESBL-producing clinical strains isolated from urine samples was collated and used to test phage efficacy. After an initial screening of 68 phages, six were identified and tested against these 38 strains to determine their clinical coverage and killing efficiency. To achieve this, we assessed four different methods to assess phage virulence across these bacterial isolates. These were the Direct Spot Test (DST), the Efficiency of Plating (EOP) assay, the planktonic killing assay (PKA) and the biofilm assay. The final ESBL cocktail of six phages could effectively kill 23/38 strains (61%), for
13/19 (68%) and for
10/19 (53%) based on the PKA data. The ESBL
collection had six isolates from the prevalent UTI-associated ST131 sequence type, five of which were targeted effectively by the final cocktail. Of the four methods used to assess phage virulence, the data suggests that PKAs are as effective as the much more time-consuming EOPs and data for the two assays correlates well. This suggests that planktonic killing is a good proxy to determine which phages should be used in a cocktail. This assay when combined with the virulence index also allows "phage synergy" to inform cocktail design.
Marine picocyanobacteria of the genera
and
, the two most abundant phototrophs on Earth, thrive in oligotrophic oceanic regions. While it is well known that specific lineages are exquisitely adapted ...to prevailing in situ light and temperature regimes, much less is known of the molecular machinery required to facilitate occupancy of these low-nutrient environments. Here, we describe a hitherto unknown alkaline phosphatase, Psip1, that has a substantially higher affinity for phosphomonoesters than other well-known phosphatases like PhoA, PhoX, or PhoD and is restricted to clade III
and a subset of high light I-adapted
strains, suggesting niche specificity. We demonstrate that Psip1 has undergone convergent evolution with PhoX, requiring both iron and calcium for activity and likely possessing identical key residues around the active site, despite generally very low sequence homology. Interrogation of metagenomes and transcriptomes from TARA oceans and an Atlantic Meridional transect shows that
is abundant and highly expressed in picocyanobacterial populations from the Mediterranean Sea and north Atlantic gyre, regions well recognized to be phosphorus (P)-deplete. Together, this identifies
as an important oligotrophy-specific gene for P recycling in these organisms. Furthermore,
is not restricted to picocyanobacteria and is abundant and highly transcribed in some α-proteobacteria and eukaryotic algae, suggesting that such a high-affinity phosphatase is important across the microbial taxonomic world to occupy low-P environments.
A common cause of human food poisoning is through ingestion of pork products contaminated with
spp. Worryingly multi-drug resistant (MDR)
strains have been isolated from pigs, which motivates the ...need for alternative antimicrobials. In this study isolation and characterization of 21 lytic
phages is described. All 21 phages, labeled as SPFM phages were shown to efficiently infect MDR
strains isolated from United Kingdom pigs and phages SPFM1, SPFM3, SPFM10, SPFM14, SPFM15, SPFM17, and SPFM19 could lyse 100% of strains tested. The phage genome sizes range from 233 to 242 Kb, which qualifies them as jumbo phages. All SPFM phage genomes are approximately 95% similar to each other by average nucleotide identity, they encode between 258-307 coding sequences and share 188 core genes. Phylogenetic analysis shows these phages are most similar to phages of the genus
and to further characterize phages within the genus, genes under positive selection were identified. Several of the genes under evolutionary selection pressure were predicted to encode for proteins that interact with bacteria. We describe the phenotypic and genetic characterization of this novel
phage set. As the phages efficiently kill MDR
strains, they may offer a promising alternative to antibiotics.
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