Frequent nonhomologous replacement of replicative helicase loaders by viruses in Vibrionaceae.

Several microbial genomes lack textbook-defined essential genes. If an essential gene is absent from a genome, then an evolutionarily independent gene of unknown function complements its function. Here, we identified frequent nonhomologous replacement of an essential component of DNA replication initiation, a replicative helicase loader gene, in Vibrionaceae. Our analysis of Vibrionaceae genomes revealed two genes with unknown function, named vdhL1 and vdhL2, that were substantially enriched in genomes without the known helicase-loader genes. These genes showed no sequence similarities to genes with known function but encoded proteins structurally similar with a viral helicase loader. Analyses of genomic syntenies and coevolution with helicase genes suggested that vdhL1/2 encodes a helicase loader. The in vitro assay showed that Vibrio harveyi VdhL1 and Vibrio ezurae VdhL2 promote the helicase activity of DnaB. Furthermore, molecular phylogenetics suggested that vdhL1/2 were derived from phages and replaced an intrinsic helicase loader gene of Vibrionaceae over 20 times. This high replacement frequency implies the host's advantage in acquiring a viral helicase loader gene.

Taxonomic difference in marine bloom-forming phytoplanktonic species affects the dynamics of both bloom-responding prokaryotes and prokaryotic viruses.

The production of dissolved organic matter during phytoplankton blooms and consumption by heterotrophic prokaryotes promote marine carbon biogeochemical cycling. Although prokaryotic viruses presumably affect this process, their dynamics during blooms are not fully understood. Here, we investigated the effects of taxonomic difference in bloom-forming phytoplankton on prokaryotes and their viruses. We analyzed the dynamics of coastal prokaryotic communities and viruses under the addition of dissolved intracellular fractions from taxonomically distinct phytoplankton, the diatom Chaetoceros sp. (CIF) and the raphidophycean alga Heterosigma akashiwo (HIF), using microcosm experiments. Ribosomal RNA gene amplicon and viral metagenomic analyses revealed that particular prokaryotes and prokaryotic viruses specifically increased in either CIF or HIF, indicating that taxonomic difference in bloom-forming phytoplankton promotes distinct dynamics of not only the prokaryotic community but also prokaryotic viruses. Furthermore, combining our microcosm experiments with publicly available environmental data mining, we identified both known and novel possible host-virus pairs. In particular, the growth of prokaryotes associating with phytoplanktonic organic matter, such as Bacteroidetes (Polaribacter and NS9 marine group), Vibrio spp., and Rhodobacteriales (Nereida and Planktomarina), was accompanied by an increase in viruses predicted to infect Bacteroidetes, Vibrio, and Rhodobacteriales, respectively. Collectively, our findings suggest that changes in bloom-forming species can be followed by an increase in a specific group of prokaryotes and their viruses and that elucidating these tripartite relationships among specific phytoplankton, prokaryotes, and prokaryotic viruses improves our understanding of coastal biogeochemical cycling in blooms.IMPORTANCEThe primary production during marine phytoplankton bloom and the consumption of the produced organic matter by heterotrophic prokaryotes significantly contribute to coastal biogeochemical cycles. While the activities of those heterotrophic prokaryotes are presumably affected by viral infection, the dynamics of their viruses during blooms are not fully understood. In this study, we experimentally demonstrated that intracellular fractions of taxonomically distinct bloom-forming phytoplankton species, the diatom Chaetoceros sp. and the raphidophycean alga Heterosigma akashiwo, promoted the growth of taxonomically different prokaryotes and prokaryotic viruses. Based on their dynamics and predicted hosts of those viruses, we succeeded in detecting already-known and novel possible host-virus pairs associating with either phytoplankton species. Altogether, we propose that the succession of bloom-forming phytoplankton would change the composition of the abundant prokaryotes, resulting in an increase in their viruses. These changes in viral composition, depending on bloom-forming species, would alter the dynamics and metabolism of prokaryotes, affecting biogeochemical cycling in blooms.

Virus-prokaryote infection pairs associated with prokaryotic production in a freshwater lake.

Viruses infect and kill prokaryotic populations in a density- or frequency-dependent manner and affect carbon cycling. However, the effects of the stratification transition, including the stratified and de-stratified periods, on the changes in prokaryotic and viral communities and their interactions remain unclear. We conducted a monthly survey of the surface and deep layers of a large and deep freshwater lake (Lake Biwa, Japan) for a year and analyzed the prokaryotic production and prokaryotic and viral community composition. Our analysis revealed that, in the surface layer, 19 prokaryotic species, accounting for approximately 40% of the total prokaryotic abundance, could potentially contribute to the majority of prokaryotic production, which is the highest during the summer and is suppressed by viruses. This suggests that a small fraction of prokaryotes and phages were the key infection pairs during the peak period of prokaryotic activity in the freshwater lake. We also found that approximately 50% of the dominant prokaryotic and viral species in the deep layer were present throughout the study period. This suggests that the "kill the winner" model could explain the viral impact on prokaryotes in the surface layer, but other dynamics may be at play in the deep layer. Furthermore, we found that annual vertical mixing could result in a similar rate of community change between the surface and deep layers. These findings may be valuable in understanding how communities and the interaction among them change when freshwater lake stratification is affected by global warming in the future.IMPORTANCEViral infection associated with prokaryotic production occurs in a density- or frequency-dependent manner and regulates the prokaryotic community. Stratification transition and annual vertical mixing in freshwater lakes are known to affect the prokaryotic community and the interaction between prokaryotes and viruses. By pairing measurements of virome analysis and prokaryotic production of a 1-year survey of the depths of surface and deep layers, we revealed (i) the prokaryotic infection pairs associated with prokaryotic production and (ii) the reset in prokaryotic and viral communities through annual vertical mixing in a freshwater lake. Our results provide a basis for future work into changes in stratification that may impact the biogeochemical cycling in freshwater lakes.

A rumen virosphere with implications of contribution to fermentation and methane production, and endemism in cattle breeds and individuals.

Viruses have a potential to modify the ruminal digestion via infection and cell lysis of prokaryotes, suggesting that viruses are related to animal performance and methane production. This study aimed to elucidate the genome-based diversity of rumen viral communities and the differences in virus structure between individuals and cattle breeds and to understand how viruses influence on the rumen. To these ends, a metagenomic sequencing of virus-like particles in the rumen of 22 Japanese cattle, including Japanese Black (JB, n = 8), Japanese Shorthorn (n = 2), and Japanese Black sires × Holstein dams crossbred steers (F1, n = 12) was conducted. Additionally, the rumen viromes of six JB and six F1 that were fed identical diets and kept in a single barn were compared. A total of 8,232 non-redundant viral genomes (≥5-kb length and ≥50% completeness), including 982 complete genomes, were constructed, and rumen virome exhibited lysogenic signatures. Furthermore, putative hosts of 1,223 viral genomes were predicted using tRNA and clustered regularly interspaced short palindromic repeat (CRISPR)-spacer matching. The genomes included 1 and 10 putative novel complete genomes associated with Fibrobacter and Ruminococcus, respectively, which are the main rumen cellulose-degrading bacteria. Additionally, the hosts of 22 viral genomes, including 2 complete genomes, were predicted as methanogens, such as Methanobrevibacter and Methanomethylophilus. Most rumen viruses were highly rumen and individual specific and related to rumen-specific prokaryotes. Furthermore, the rumen viral community structure was significantly different between JB and F1 steers, indicating that cattle breed is one of the factors influencing the rumen virome composition.IMPORTANCEHere, we investigated the individual and breed differences of the rumen viral community in Japanese cattle. In the process, we reconstructed putative novel complete viral genomes related to rumen fiber-degrading bacteria and methanogen. The finding strongly suggests that rumen viruses contribute to cellulose and hemicellulose digestion and methanogenesis. Notably, this study also found that rumen viruses are highly rumen and individual specific, suggesting that rumen viruses may not be transmitted through environmental exposure. More importantly, we revealed differences of viral communities between JB and F1 cattle, indicating that cattle breed is a factor that influences the establishment of rumen virome. These results suggest the possibility of rumen virus transmission from mother to offspring and its potential to influence beef production traits. These rumen viral genomes and findings provide new insights into the characterizations of the rumen viruses.

Population-level prokaryotic community structures associated with ferromanganese nodules in the Clarion-Clipperton Zone (Pacific Ocean) revealed by 16S rRNA gene amplicon sequencing.

Although deep-sea ferromanganese nodules are a potential resource for exploitation, their formation mechanisms remain unclear. Several nodule-associated prokaryotic species have been identified by amplicon sequencing of 16S rRNA genes and are assumed to contribute to nodule formation. However, the recent development of amplicon sequence variant (ASV)-level monitoring revealed that closely related prokaryotic populations within an operational taxonomic unit often exhibit distinct ecological properties. Thus, conventional species-level monitoring might have overlooked nodule-specific populations when distinct populations of the same species were present in surrounding environments. Herein, we examined the prokaryotic community diversity of nodules and surrounding environments at the Clarion-Clipperton Zone in Japanese licensed areas by 16S rRNA gene amplicon sequencing with ASV-level resolution for three cruises from 2017 to 2019. Prokaryotic community composition and diversity were distinct by habitat type: nodule, nodule-surface mud, sediment, bottom water and water column. Most ASVs (~80%) were habitat-specific. We identified 178 nodule-associated ASVs and 41 ASVs associated with nodule-surface mud via linear discriminant effect size analysis. Moreover, several ASVs, such as members of SAR324 and Woeseia, were highly specific to nodules. These nodule-specific ASVs are promising targets for future investigation of the nodule formation process.

Prevalence of Viral Frequency-Dependent Infection in Coastal Marine Prokaryotes Revealed Using Monthly Time Series Virome Analysis.

Viruses infecting marine prokaryotes have a large impact on the diversity and dynamics of their hosts. Model systems suggest that viral infection is frequency dependent and constrained by the virus-host encounter rate. However, it is unclear whether frequency-dependent infection is pervasive among the abundant prokaryotic populations with different temporal dynamics. To address this question, we performed a comparison of prokaryotic and viral communities using 16S rRNA amplicon and virome sequencing based on samples collected monthly for 2 years at a Japanese coastal site, Osaka Bay. Concurrent seasonal shifts observed in prokaryotic and viral community dynamics indicated that the abundance of viruses correlated with that of their predicted host phyla (or classes). Cooccurrence network analysis between abundant prokaryotes and viruses revealed 6,423 cooccurring pairs, suggesting a tight coupling of host and viral abundances and their "one-to-many" correspondence. Although stable dominant species, such as SAR11, showed few cooccurring viruses, a fast succession of their viruses suggests that viruses infecting these populations changed continuously. Our results suggest that frequency-dependent viral infection prevails in coastal marine prokaryotes regardless of host taxa and temporal dynamics. IMPORTANCE There is little room for doubt that viral infection is prevalent among abundant marine prokaryotes regardless of their taxa or growth strategy. However, comprehensive evaluations of viral infections in natural prokaryotic communities are still technically difficult. In this study, we examined viral infection in abundant prokaryotes by monitoring the monthly dynamics of prokaryotic and viral communities at a eutrophic coastal site, Osaka Bay. We compared the community dynamics of viruses with those of their putative hosts based on genome-based in silico host prediction. We observed frequent cooccurrence among the predicted virus-host pairs, suggesting that viral infection is prevalent in abundant prokaryotes regardless of their taxa or temporal dynamics. This likely indicates that frequent lysis of the abundant prokaryotes via viral infection has a considerable contribution to the biogeochemical cycling and maintenance of prokaryotic community diversity.

Year-round dynamics of amplicon sequence variant communities differ among eukaryotes, Imitervirales and prokaryotes in a coastal ecosystem.

Coastal microbial communities are affected by seasonal environmental change, biotic interactions and fluctuating nutrient availability. We investigated the seasonal dynamics of communities of eukaryotes, a major group of double-stranded DNA viruses that infect eukaryotes (order Imitervirales; phylum Nucleocytoviricota), and prokaryotes in the Uranouchi Inlet, Kochi, Japan. We performed metabarcoding using ribosomal RNA genes and viral polB genes as markers in 43 seawater samples collected over 20 months. Eukaryotes, prokaryotes and Imitervirales communities characterized by the compositions of amplicon sequence variants (ASVs) showed synchronic seasonal cycles. However, the community dynamics showed intriguing differences in several aspects, such as the recovery rate after a year. We also showed that the differences in community dynamics were at least partially explained by differences in recurrence/persistence levels of individual ASVs among eukaryotes, prokaryotes and Imitervirales. Prokaryotic ASVs were the most persistent, followed by eukaryotic ASVs and Imitervirales ASVs, which were the least persistent. We argue that the differences in the specificity of interactions (virus-eukaryote vs prokaryote-eukaryote) as well as the niche breadth of community members were at the origin of the distinct community dynamics among eukaryotes, their viruses and prokaryotes.

Taxonomic and functional characterization of the rumen microbiome of Japanese Black cattle revealed by 16S rRNA gene amplicon and metagenome shotgun sequencing.

This study aimed to determine the taxonomic and functional characteristics of the Japanese Black (JB) steer rumen microbiome. The rumen microbiomes of six JB steers (age 14.7 ± 1.44 months) and six JB sires × Holstein dams crossbred (F1) steers (age 11.1 ± 0.39 months), fed the same diet, were evaluated. Based on 16S rRNA gene sequencing, the beta diversity revealed differences in microbial community structures between the JB and F1 rumen. Shotgun sequencing showed that Fibrobacter succinogenes and two Ruminococcus spp., which are related to cellulose degradation were relatively more abundant in the JB steer rumen than in the F1 rumen. Furthermore, the 16S rRNA gene copy number of F. succinogenes was significantly higher in the JB steer rumen than in the F1 rumen according to quantitative real-time polymerase chain reaction analysis. Genes encoding the enzymes that accelerate cellulose degradation and those associated with hemicellulose degradation were enriched in the JB steer rumen. Although Prevotella spp. were predominant both in the JB and F1 rumen, the genes encoding carbohydrate-active enzymes of Prevotella spp. may differ between JB and F1.

Calcium salts of long-chain fatty acids from linseed oil decrease methane production by altering the rumen microbiome in vitro.

Calcium salts of long-chain fatty acids (CSFA) from linseed oil have the potential to reduce methane (CH4) production from ruminants; however, there is little information on the effect of supplementary CSFA on rumen microbiome as well as CH4 production. The aim of the present study was to evaluate the effects of supplementary CSFA on ruminal fermentation, digestibility, CH4 production, and rumen microbiome in vitro. We compared five treatments: three CSFA concentrations-0% (CON), 2.25% (FAL) and 4.50% (FAH) on a dry matter (DM) basis-15 mM of fumarate (FUM), and 20 mg/kg DM of monensin (MON). The results showed that the proportions of propionate in FAL, FAH, FUM, and MON were increased, compared with CON (P < 0.05). Although DM and neutral detergent fiber expressed exclusive of residual ash (NDFom) digestibility decreased in FAL and FAH compared to those in CON (P < 0.05), DM digestibility-adjusted CH4 production in FAL and FAH was reduced by 38.2% and 63.0%, respectively, compared with that in CON (P < 0.05). The genera Ruminobacter, Succinivibrio, Succiniclasticum, Streptococcus, Selenomonas.1, and Megasphaera, which are related to propionate production, were increased (P < 0.05), while Methanobrevibacter and protozoa counts, which are associated with CH4 production, were decreased in FAH, compared with CON (P < 0.05). The results suggested that the inclusion of CSFA significantly changed the rumen microbiome, leading to the acceleration of propionate production and the reduction of CH4 production. In conclusion, although further in vivo study is needed to evaluate the reduction effect on rumen CH4 production, CSFA may be a promising candidate for reduction of CH4 emission from ruminants.

Predetermined clockwork microbial worlds: Current understanding of aquatic microbial diel response from model systems to complex environments.

In the photic zone of aquatic ecosystems, microorganisms with different metabolisms and their viruses form complex interactions and food webs. Within these interactions, phototrophic microorganisms such as eukaryotic microalgae and cyanobacteria interact directly with sunlight, and thereby generate circadian rhythms. Diel cycling originally generated in microbial phototrophs is directly transmitted toward heterotrophic microorganisms utilizing the photosynthetic products as they are excreted or exuded. Such diel cycling seems to be indirectly propagated toward heterotrophs as a result of complex biotic interactions. For example, cell death of phototrophic microorganisms induced by viral lysis and protistan grazing provides additional resources of dissolved organic matter to the microbial community, and so generates diel cycling in other heterotrophs with different nutrient dependencies. Likewise, differences in the diel transmitting pathway via complex interactions among heterotrophs, and between heterotrophs and their viruses, may also generate higher variation and time lag diel rhythms in different heterotrophic taxa. Thus, sunlight and photosynthesis not only contribute energy and carbon supply, but also directly or indirectly control diel cycling of the microbial community through complex interactions in the photic zone of aquatic ecosystems.

Differential Responses of a Coastal Prokaryotic Community to Phytoplanktonic Organic Matter Derived from Cellular Components and Exudates.

The phytoplanktonic production and prokaryotic consumption of organic matter significantly contribute to marine carbon cycling. Organic matter released from phytoplankton via three processes (exudation of living cells, cell disruption through grazing, and viral lysis) shows distinct chemical properties. We herein investigated the effects of phytoplanktonic whole-cell fractions (WF) (representing cell disruption by grazing) and extracellular fractions (EF) (representing exudates) prepared from Heterosigma akashiwo, a bloom-forming Raphidophyceae, on prokaryotic communities using culture-based experiments. We analyzed prokaryotic community changes for two weeks. The shift in cell abundance by both treatments showed similar dynamics, reaching the first peak (~4.1×106| |cells| |mL-1) on day 3 and second peak (~1.1×106| |cells| |mL-1) on day 13. We classified the sequences obtained into operational taxonomic units (OTUs). A Bray-Curtis dissimilarity analysis revealed that the OTU-level community structure changed distinctively with the two treatments. Ten and 13 OTUs were specifically abundant in the WF and EF treatments, respectively. These OTUs were assigned as heterotrophic bacteria mainly belonging to the Alteromonadales (Gammaproteobacteria) and Bacteroidetes clades and showed successive dynamics following the addition of organic matter. We also analyzed the dynamics of these OTUs in the ocean using publicly available metagenomic data from a natural coastal bloom in Monterey Bay, USA. At least two WF treatment OTUs showed co-occurrence with H. akashiwo, indicating that the blooms of H. akashiwo also affect these OTUs in the ocean. The present results strongly suggest that the thriving and dead cells of uninfected phytoplankton differentially influence the marine prokaryotic community.

In silico Prediction of Virus-Host Interactions for Marine Bacteroidetes With the Use of Metagenome-Assembled Genomes.

Bacteroidetes is one of the most abundant heterotrophic bacterial taxa in the ocean and play crucial roles in recycling phytoplankton-derived organic matter. Viruses of Bacteroidetes are also expected to have an important role in the regulation of host communities. However, knowledge on marine Bacteroidetes viruses is biased toward cultured viruses from a few species, mainly fish pathogens or Bacteroidetes not abundant in marine environments. In this study, we investigated the recently reported 1,811 marine viral genomes to identify putative Bacteroidetes viruses using various in silico host prediction techniques. Notably, we used microbial metagenome-assembled genomes (MAGs) to augment the marine Bacteroidetes reference genomic data. The examined viral genomes and MAGs were derived from simultaneously collected samples. Using nucleotide sequence similarity-based host prediction methods, we detected 31 putative Bacteroidetes viral genomes. The MAG-based method substantially enhanced the predictions (26 viruses) when compared with the method that is solely based on the reference genomes from NCBI RefSeq (7 viruses). Previously unrecognized genus-level groups of Bacteroidetes viruses were detected only by the MAG-based method. We also developed a host prediction method based on the proportion of Bacteroidetes homologs in viral genomes, which detected 321 putative Bacteroidetes virus genomes including 81 that were newly recognized as Bacteroidetes virus genomes. The majority of putative Bacteroidetes viruses were detected based on the proportion of Bacteroidetes homologs in both RefSeq and MAGs; however, some were detected in only one of the two datasets. Putative Bacteroidetes virus lineages included not only relatives of known viruses but also those phylogenetically distant from the cultured viruses, such as marine Far-T4 like viruses known to be widespread in aquatic environments. Our MAG and protein homology-based host prediction approaches enhanced the existing knowledge on the diversity of Bacteroidetes viruses and their potential interaction with their hosts in marine environments.

An Optimized Metabarcoding Method for Mimiviridae.

Mimiviridae is a group of viruses with large genomes and virions. Ecological relevance of Mimiviridae in marine environments has been increasingly recognized through the discoveries of novel isolates and metagenomic studies. To facilitate ecological profiling of Mimiviridae, we previously proposed a meta-barcoding approach based on 82 degenerate primer pairs (i.e., MEGAPRIMER) targeting the DNA polymerase gene of Mimiviridae. The method detected a larger number of operational taxonomic units (OTUs) in environmental samples than previous methods. However, it required large quantities of DNA and was laborious due to the use of individual primer pairs. Here, we examined coastal seawater samples using varying PCR conditions and purification protocols to streamline the MEGAPRIMER method. Mixing primer pairs in "cocktails" reduced the required amount of environmental DNA by 90%, while reproducing the results obtained by the original protocol. We compared the results obtained by the meta-barcoding approach with quantifications using qPCR for selected OTUs. This revealed possible amplification biases among different OTUs, but the frequency profiles for individual OTUs across multiple samples were similar to those obtained by qPCR. We anticipate that the newly developed MEGAPRIMER protocols will be useful for ecological investigation of Mimiviridae in a larger set of environmental samples.

Diel cycling of the cosmopolitan abundant Pelagibacter virus 37-F6: one of the most abundant viruses on earth.

The spatiotemporal dynamics for marine viral populations has only recently been explored. However, nothing is known about temporal activities of the uncultured Pelagibacter virus vSAG 37-F6, which was discovered by single-virus genomics as potentially the most abundant marine virus. Here, we investigate the diel cycling of 37-F6 virus and the putative SAR11 host using coastal and oceanic transcriptomic and viromic time-series data from Osaka Bay and North Pacific Subtropical Gyre. Virus 37-F6 and relatives displayed diel cycling of transcriptional activities synchronized with its putative host. In both virus and host, the lowest transcription rates were observed at 14:00-15:00, coinciding roughly with maximum solar irradiance, while higher transcriptional rates were detected during the night/early morning and afternoon. Diel abundance of free viruses of 37-F6 in seawater roughly mirrored the transcriptional activities of both virus and host. In Osaka Bay, among viral relatives (genus level), virus 37-F6 specifically showed the highest ratio of transcriptional activity to virome abundance, a proxy for viral transcriptional activity relative to free viral particle abundance. This high ratio suggests high infection rate efficiencies in vSAG 37-F6 virus compared to viral relatives. Thus, time-series data revealed temporal transcript activities in one of the most abundant viruses in Earth.

Cooccurrence of Broad- and Narrow-Host-Range Viruses Infecting the Bloom-Forming Toxic Cyanobacterium Microcystis aeruginosa.

Viruses play important roles in regulating the abundance and composition of bacterial populations in aquatic ecosystems. The bloom-forming toxic cyanobacterium Microcystis aeruginosa is predicted to interact with diverse cyanoviruses, resulting in Microcystis population diversification. However, current knowledge of the genomes from these viruses and their infection programs is limited to those of Microcystis virus Ma-LMM01. Here, we performed a time series sampling at a small pond in Japan during a Microcystis bloom and then investigated the genomic information and transcriptional dynamics of Microcystis-interacting viruses using metagenomic and metatranscriptomic approaches. We identified 15 viral genomic fragments classified into three groups, groups I (including Ma-LMM01), II (high abundance and transcriptional activity), and III (new lineages). According to the phylogenetic distribution of Microcystis strains possessing spacers against each viral group, the group II-original viruses interacted with all three phylogenetically distinct Microcystis population types (phylotypes), whereas the groups I and III-original viruses interacted with only one or two phylotypes, indicating the cooccurrence of broad- (group II) and narrow (groups I and III)-host-range viruses in the bloom. These viral fragments showed the highest transcriptional levels during daytime regardless of their genomic differences. Interestingly, M. aeruginosa expressed antiviral defense genes in the environment, unlike what was seen with an Ma-LMM01 infection in a previous culture experiment. Given that broad-host-range viruses often induce antiviral responses within alternative hosts, our findings suggest that such antiviral responses might inhibit viral multiplication, mainly that of broad-host-range viruses like those in group II.IMPORTANCE The bloom-forming toxic cyanobacterium Microcystis aeruginosa is thought to have diversified its population through the interactions between host and viruses in antiviral defense systems. However, current knowledge of viral genomes and infection programs is limited to those of Microcystis virus Ma-LMM01, which was a narrow host range in which it can escape from the highly abundant host defense systems. Our metagenomic approaches unveiled the cooccurrence of narrow- and broad-host-range Microcystis viruses, which included fifteen viral genomic fragments from Microcystis blooms that were classified into three groups. Interestingly, Microcystis antiviral defense genes were expressed against viral infection in the environment, unlike what was seen in a culture experiment with Ma-LMM01. Given that viruses with a broad host range often induce antiviral responses within alternative hosts, our findings suggest that antiviral responses inhibit viral reproduction, especially that of broad-range viruses like those in group II. This paper augments our understanding of the interactions between M. aeruginosa and its viruses and fills an important knowledge gap.

Systematic identification of synthetic lethal mutations with reduced-genome Escherichia coli: synthetic genetic interactions among yoaA, xthA and holC related to survival from MMS exposure.

Reduced-genome Escherichia coli strains lacking up to 38.9% of the parental chromosome have been constructed by combining large-scale chromosome deletion mutations. Functionally redundant genes involved in essential processes can be systematically identified using these reduced-genome strains. One large-scale chromosome deletion mutation could be introduced into the wild-type strain but not into the largest reduced-genome strain, suggesting a synthetic lethal interaction between genes removed by the deletion and those already absent in the reduced-genome strain. Thus, introduction of the deletion mutation into a series of reduced-genome mutants could allow the identification of other chromosome deletion mutations responsible for the synthetic lethal phenotype. We identified a synthetic lethality caused by disruption of nfo and xthA, two genes encoding apurinic/apyrimidinic (AP) endonucleases involved in the DNA base excision repair pathway, and two other large-scale chromosome deletions. We constructed temperature-sensitive mutants harboring quadruple-deletion mutations in the affected genes/chromosome regions. Using these mutants, we identified two multi-copy suppressors: holC, encoding the chi subunit of DNA polymerase III, and yoaA, encoding a putative DNA helicase. Addition of the yoaA disruption increased the methyl methanesulfonate (MMS) sensitivity of xthA single-deletion or xthA nfo double-deletion mutants. This increased MMS sensitivity was not suppressed by the presence of multi-copy holC. These results indicate that yoaA is involved in MMS sensitivity and suggest that YoaA functions together with HolC.