Gut microbiota-derived butyrate selectively interferes with growth of carbapenem-resistant Escherichia coli based on their resistance mechanism.

We investigated consequences of resistance acquisition in Escherichia coli clinical isolates during anaerobic (continuous culture) growth and examined their sensitivity to butyrate, a hallmark metabolite of healthy gut microbiota. Strains were stratified based on carrying either a carbapenemase (CARB) or displaying porin malfunctioning (POR). POR displayed markedly altered growth efficiencies, lower membrane stability and increased sensitivity to butyrate compared with CARB. Major differences in global gene expression between the two groups during anaerobic growth were revealed involving increased expression of alternative substrate influx routes, the stringent response and iron acquisition together with lower expression of various stress response systems in POR. Longitudinal analyses during butyrate wash-in showed common responses for all strains as well as specific features of POR that displayed strong initial "overshoot" reactions affecting various stress responses that balanced out over time. Results were partly reproduced in a mutant strain verifying porin deficiencies as the major underlying mechanism for results observed in clinical isolates. Furthermore, direct competition experiments confirmed butyrate as key for amplifying fitness disadvantages based on porin malfunctioning. Results provide new (molecular) insights into ecological consequences of resistance acquisition and can assist in developing measures to prevent colonization and infection based on the underlying resistance mechanism.

Role of Carbon, Nitrogen, Phosphate and Sulfur Metabolism in Secondary Metabolism Precursor Supply in Streptomyces spp.

The natural soil environment of Streptomyces is characterized by variations in the availability of nitrogen, carbon, phosphate and sulfur, leading to complex primary and secondary metabolisms. Their remarkable ability to adapt to fluctuating nutrient conditions is possible through the utilization of a large amount of substrates by diverse intracellular and extracellular enzymes. Thus, Streptomyces fulfill an important ecological role in soil environments, metabolizing the remains of other organisms. In order to survive under changing conditions in their natural habitats, they have the possibility to fall back on specialized enzymes to utilize diverse nutrients and supply compounds from primary metabolism as precursors for secondary metabolite production. We aimed to summarize the knowledge on the C-, N-, P- and S-metabolisms in the genus Streptomyces as a source of building blocks for the production of antibiotics and other relevant compounds.

Anoxygenic photosynthesis with emphasis on green sulfur bacteria and a perspective for hydrogen sulfide detoxification of anoxic environments.

The bacterial light-dependent energy metabolism can be divided into two types: oxygenic and anoxygenic photosynthesis. Bacterial oxygenic photosynthesis is similar to plants and is characteristic for cyanobacteria. Bacterial anoxygenic photosynthesis is performed by anoxygenic phototrophs, especially green sulfur bacteria (GSB; family Chlorobiaceae) and purple sulfur bacteria (PSB; family Chromatiaceae). In anoxygenic photosynthesis, hydrogen sulfide (H2S) is used as the main electron donor, which differs from plants or cyanobacteria where water is the main source of electrons. This review mainly focuses on the microbiology of GSB, which may be found in water or soil ecosystems where H2S is abundant. GSB oxidize H2S to elemental sulfur. GSB possess special structures-chlorosomes-wherein photosynthetic pigments are located. Chlorosomes are vesicles that are surrounded by a lipid monolayer that serve as light-collecting antennas. The carbon source of GSB is carbon dioxide, which is assimilated through the reverse tricarboxylic acid cycle. Our review provides a thorough introduction to the comparative eco-physiology of GSB and discusses selected application possibilities of anoxygenic phototrophs in the fields of environmental management, bioremediation, and biotechnology.

Dormancy-like Phenotype of Aggregatibacter actinomycetemcomitans: Survival during Famine.

Microbes frequently experience nutrient deprivations in the natural environment and may enter dormancy. Aggregatibacter actinomycetemcomitans is known to establish long-term infections in humans. This study examined the dormancy-like phenotype of an A. actinomycetemcomitans strain D7S-1 and its isogenic smooth-colony mutant D7SS. A tissue culture medium RPMI-1640 was nutrient-deficient (ND) and unable to support A. actinomycetemcomitans growth. RPMI-1640 amended with bases was nutrient-limited (NL) and supported limited growth of A. actinomycetemcomitans less than the nutrient-enriched (NE) laboratory medium did. Strain D7S-1, after an initial 2-log reduction in viability, maintained viability from day 4 to day 15 in the NL medium. Strain D7SS, after 1-log reduction in viability, maintained viability from day 3 to day 5. In contrast, bacteria in the NE medium were either non-recoverable (D7S-1; >6-log reduction) or continued to lose viability (D7SS; 3-log reduction) on day 5 and beyond. Scanning and transmission electron microscopy showed that A. actinomycetemcomitans in the NL medium formed robust biofilms similar to those in the NE medium but with evidence of stress. A. actinomycetemcomitans in the ND medium revealed scant biofilms and extensive cellular damage. We concluded that A. actinomycetemcomitans grown in the NL medium exhibited a dormancy-like phenotype characterized by minimum growth, prolonged viability, and distinct cellular morphology.

Corrigendum: Phenotypic and transcriptional characterization of F. tularensis LVS during transition into a viable but non-culturable state.

[This corrects the article DOI: 10.3389/fmicb.2024.1347488.].

Bacterial extracellular vesicles: Vital contributors to physiology from bacteria to host.

Bacterial extracellular vesicles (bEVs) represent spherical particles with diameters ranging from 20 to 400 nm filled with multiple parental bacteria-derived components, including proteins, nucleic acids, lipids, and other biomolecules. The production of bEVs facilitates bacteria interacting with their environment and exerting biological functions. It is increasingly evident that the bEVs play integral roles in both bacterial and host physiology, contributing to environmental adaptations to functioning as health promoters for their hosts. This review highlights the current state of knowledge on the composition, biogenesis, and diversity of bEVs and the mechanisms by which different bEVs elicit effects on bacterial physiology and host health. We posit that an in-depth exploration of the mechanistic aspects of bEVs activity is essential to elucidate their health-promoting effects on the host and may facilitate the translation of bEVs into applications as novel natural biological nanomaterials.

Tools and methods for high-throughput single-cell imaging with the mother machine.

Despite much progress, image processing remains a significant bottleneck for high-throughput analysis of microscopy data. One popular platform for single-cell time-lapse imaging is the mother machine, which enables long-term tracking of microbial cells under precisely controlled growth conditions. While several mother machine image analysis pipelines have been developed in the past several years, adoption by a non-expert audience remains a challenge. To fill this gap, we implemented our own software, MM3, as a plugin for the multidimensional image viewer napari. napari-MM3 is a complete and modular image analysis pipeline for mother machine data, which takes advantage of the high-level interactivity of napari. Here, we give an overview of napari-MM3 and test it against several well-designed and widely used image analysis pipelines, including BACMMAN and DeLTA. Researchers often analyze mother machine data with custom scripts using varied image analysis methods, but a quantitative comparison of the output of different pipelines has been lacking. To this end, we show that key single-cell physiological parameter correlations and distributions are robust to the choice of analysis method. However, we also find that small changes in thresholding parameters can systematically alter parameters extracted from single-cell imaging experiments. Moreover, we explicitly show that in deep learning-based segmentation, 'what you put is what you get' (WYPIWYG) - that is, pixel-level variation in training data for cell segmentation can propagate to the model output and bias spatial and temporal measurements. Finally, while the primary purpose of this work is to introduce the image analysis software that we have developed over the last decade in our lab, we also provide information for those who want to implement mother machine-based high-throughput imaging and analysis methods in their research.

The distinct cell physiology of Bradyrhizobium at the population and cellular level.

The α-Proteobacteria belonging to Bradyrhizobium genus are microorganisms of extreme slow growth. Despite their extended use as inoculants in soybean production, their physiology remains poorly characterized. In this work, we produced quantitative data on four different isolates: B. diazoefficens USDA110, B. diazoefficiens USDA122, B. japonicum E109 and B. japonicum USDA6 which are representative of specific genomic profiles. Notably, we found conserved physiological traits conserved in all the studied isolates: (i) the lag and initial exponential growth phases display cell aggregation; (ii) the increase in specific nutrient concentration such as yeast extract and gluconate hinders growth; (iii) cell size does not correlate with culture age; and (iv) cell cycle presents polar growth. Meanwhile, fitness, cell size and in vitro growth widely vary across isolates correlating to ribosomal RNA operon number. In summary, this study provides novel empirical data that enriches the comprehension of the Bradyrhizobium (slow) growth dynamics and cell cycle.

Phenotypic and transcriptional characterization of F. tularensis LVS during transition into a viable but non-culturable state.

Francisella tularensis is a gram-negative, intracellular pathogen which can cause serious, potentially fatal, illness in humans. Species of F. tularensis are found across the Northern Hemisphere and can infect a broad range of host species, including humans. Factors affecting the persistence of F. tularensis in the environment and its epidemiology are not well understood, however, the ability of F. tularensis to enter a viable but non-culturable state (VBNC) may be important. A broad range of bacteria, including many pathogens, have been observed to enter the VBNC state in response to stressful environmental conditions, such as nutrient limitation, osmotic or oxidative stress or low temperature. To investigate the transition into the VBNC state for F. tularensis, we analyzed the attenuated live vaccine strain, F. tularensis LVS grown under standard laboratory conditions. We found that F. tularensis LVS rapidly and spontaneously enters a VBNC state in broth culture at 37°C and that this transition coincides with morphological differentiation of the cells. The VBNC bacteria retained an ability to interact with both murine macrophages and human erythrocytes in in vitro assays and were insensitive to treatment with gentamicin. Finally, we present the first transcriptomic analysis of VBNC F. tularensis, which revealed clear differences in gene expression, and we identify sets of differentially regulated genes which are specific to the VBNC state. Identification of these VBNC specific genes will pave the way for future research aimed at dissecting the molecular mechanisms driving entry into the VBNC state.

Better off breathing: an explanation for the seemingly detrimental impact of aerobic respiration on Zymomonas mobilis.

The bacterium Zymomonas mobilis is best known for fermentatively producing more ethanol than yeast. However, Z. mobilis has also puzzled researchers for decades with the counterintuitive observation that disrupting aerobic respiration benefits aerobic growth, implying that fermentation remains favorable. Retention of detrimental respiration genes seemed to defy natural selection. New findings by Felczak et al. help clarify the importance of respiration for Z. mobilis and the factors that led to the confusing prior results (M. M. Felczak, M. P. Bernard, and M. A. TerAvest, 2023, mBio 14:e02043-23, https://doi.org/10.1128/mbio.02043-23). The team overcame redundancy from multiple genome copies to delete what turned out to be a key terminal oxidase. Unlike previous studies, wherein mutants exhibited low respiration rates and had improved aerobic growth, this mutant was incapable of respiration and had poor aerobic growth. Thus, respiration is important but surprisingly exceeds what is optimal under lab conditions. Respiration likely protects against toxic effects of oxygen, ensuring retention of respiration genes in the Z. mobilis genome.

Tradeoffs in bacterial physiology determine the efficiency of antibiotic killing.

Antibiotic effectiveness depends on a variety of factors. While many mechanistic details of antibiotic action are known, the connection between death rate and bacterial physiology is poorly understood. A common observation is that death rate in antibiotics rises linearly with growth rate; however, it remains unclear how other factors, such as environmental conditions and whole-cell physiological properties, affect bactericidal activity. To address this, we developed a high-throughput assay to precisely measure antibiotic-mediated death. We found that death rate is linear in growth rate, but the slope depends on environmental conditions. Growth under stress lowers death rate compared to nonstressed environments with similar growth rate. To understand stress's role, we developed a mathematical model of bacterial death based on resource allocation that includes a stress-response sector; we identify this sector using RNA-seq. Our model accurately predicts the minimal inhibitory concentration (MIC) with zero free parameters across a wide range of growth conditions. The model also quantitatively predicts death and MIC when sectors are experimentally modulated using cyclic adenosine monophosphate (cAMP), including protection from death at very low cAMP levels. The present study shows that different conditions with equal growth rate can have different death rates and establishes a quantitative relation between growth, death, and MIC that suggests approaches to improve antibiotic efficacy.

A novel synthetic inhibitor of polyamine utilization in Streptomyces coelicolor.

In this work, we present the first inhibitor of GlnA2Sc, a gamma-glutamylpolyamine synthetase, which allows Streptomyces coelicolor to detoxify high concentrations of polyamines and to utilize them as a carbon or nitrogen source. GlnA2 belongs to the class of glutamine synthetase-like (GS-like) enzymes that catalyze the glutamylation of different nitrogen-containing compounds. Whereas a number of inhibitors for GS are known, none of them are known to inhibit GlnA2. In this work, PPU268, an inhibitor for GlnA2 is presented that is structurally derived from the prototypic GS inhibitor-methionine sulfoximine (MSO). It combines two features: the binding mechanism of MSO and the amine substrate specificity of GlnA2Sc. This inhibitor is a novel compound to block the polyamine utilization in bacteria resulting in the inability to detoxify polyamines. This may offer a possibility to develop novel therapeutic strategies to combat actinobacterial human pathogens that encounter polyamines in the course of the infection processes.

System-wide optimization of an orthogonal translation system with enhanced biological tolerance.

Over the past two decades, synthetic biological systems have revolutionized the study of cellular physiology. The ability to site-specifically incorporate biologically relevant non-standard amino acids using orthogonal translation systems (OTSs) has proven particularly useful, providing unparalleled access to cellular mechanisms modulated by post-translational modifications, such as protein phosphorylation. However, despite significant advances in OTS design and function, the systems-level biology of OTS development and utilization remains underexplored. In this study, we employ a phosphoserine OTS (pSerOTS) as a model to systematically investigate global interactions between OTS components and the cellular environment, aiming to improve OTS performance. Based on this analysis, we design OTS variants to enhance orthogonality by minimizing host process interactions and reducing stress response activation. Our findings advance understanding of system-wide OTS:host interactions, enabling informed design practices that circumvent deleterious interactions with host physiology while improving OTS performance and stability. Furthermore, our study emphasizes the importance of establishing a pipeline for systematically profiling OTS:host interactions to enhance orthogonality and mitigate mechanisms underlying OTS-mediated host toxicity.

Resource allocation accounts for the large variability of rate-yield phenotypes across bacterial strains.

Different strains of a microorganism growing in the same environment display a wide variety of growth rates and growth yields. We developed a coarse-grained model to test the hypothesis that different resource allocation strategies, corresponding to different compositions of the proteome, can account for the observed rate-yield variability. The model predictions were verified by means of a database of hundreds of published rate-yield and uptake-secretion phenotypes of Escherichia coli strains grown in standard laboratory conditions. We found a very good quantitative agreement between the range of predicted and observed growth rates, growth yields, and glucose uptake and acetate secretion rates. These results support the hypothesis that resource allocation is a major explanatory factor of the observed variability of growth rates and growth yields across different bacterial strains. An interesting prediction of our model, supported by the experimental data, is that high growth rates are not necessarily accompanied by low growth yields. The resource allocation strategies enabling high-rate, high-yield growth of E. coli lead to a higher saturation of enzymes and ribosomes, and thus to a more efficient utilization of proteomic resources. Our model thus contributes to a fundamental understanding of the quantitative relationship between rate and yield in E. coli and other microorganisms. It may also be useful for the rapid screening of strains in metabolic engineering and synthetic biology.

Extending Validity of the Bacterial Cell Cycle Model through Thymine Limitation: A Personal View.

The contemporary view of bacterial physiology was established in 1958 at the "Copenhagen School", culminating a decade later in a detailed description of the cell cycle based on four parameters. This model has been subsequently supported by numerous studies, nicknamed BCD (The Bacterial Cell-Cycle Dogma). It readily explains, quantitatively, the coupling between chromosome replication and cell division, size and DNA content. An important derivative is the number of replication positions n, the ratio between the time C to complete a round of replication and the cell mass doubling time τ; the former is constant at any temperature and the latter is determined by the medium composition. Changes in cell width W are highly correlated to n through the equation for so-called nucleoid complexity NC (=(2n - 1)/(ln2 × n)), the amount of DNA per terC (i.e., chromosome) in genome equivalents. The narrow range of potential n can be dramatically extended using the method of thymine limitation of thymine-requiring mutants, which allows a more rigorous testing of the hypothesis that the nucleoid structure is the primary source of the signal that determines W during cell division. How this putative signal is relayed from the nucleoid to the divisome is still highly enigmatic. The aim of this Opinion article is to suggest the possibility of a new signaling function for nucleoid DNA.

Escherichia coli Aggregates Mediated by Native or Synthetic Adhesins Exhibit Both Core and Adhesin-Specific Transcriptional Responses.

Bacteria can rapidly tune their physiology and metabolism to adapt to environmental fluctuations. In particular, they can adapt their lifestyle to the close proximity of other bacteria or the presence of different surfaces. However, whether these interactions trigger transcriptomic responses is poorly understood. We used a specific setup of E. coli strains expressing native or synthetic adhesins mediating bacterial aggregation to study the transcriptomic changes of aggregated compared to nonaggregated bacteria. Our results show that, following aggregation, bacteria exhibit a core response independent of the adhesin type, with differential expression of 56.9% of the coding genome, including genes involved in stress response and anaerobic lifestyle. Moreover, when aggregates were formed via a naturally expressed E. coli adhesin (antigen 43), the transcriptomic response of the bacteria was more exaggerated than that of aggregates formed via a synthetic adhesin. This suggests that the response to aggregation induced by native E. coli adhesins could have been finely tuned during bacterial evolution. Our study therefore provides insights into the effect of self-interaction in bacteria and allows a better understanding of why bacterial aggregates exhibit increased stress tolerance. IMPORTANCE The formation of bacterial aggregates has an important role in both clinical and ecological contexts. Although these structures have been previously shown to be more resistant to stressful conditions, the genetic basis of this stress tolerance associated with the aggregate lifestyle is poorly understood. Surface sensing mediated by different adhesins can result in various changes in bacterial physiology. However, whether adhesin-adhesin interactions, as well as the type of adhesin mediating aggregation, affect bacterial cell physiology is unknown. By sequencing the transcriptomes of aggregated and nonaggregated cells expressing native or synthetic adhesins, we characterized the effects of aggregation and adhesin type on E. coli physiology.

Actively replicating gut bacteria identified by 5-ethynyl-2'-deoxyuridine (EdU) click chemistry and cell sorting.

The composition of the intestinal bacterial community is well described, but recent research suggests that the metabolism of these bacteria plays a larger role in health than which species are present. One fundamental aspect of gut bacterial metabolism that remains understudied is bacterial replication. Indeed, there exist few techniques which can identify actively replicating gut bacteria. In this study, we aimed to address this gap by adapting 5-ethynyl-2'-deoxyuridine (EdU) click chemistry (EdU-click), a metabolic labeling method, coupled with fluorescence-activated cell sorting and sequencing (FACS-Seq) to characterize replicating gut bacteria. We first used EdU-click with human gut bacterial isolates and show that many of them are amenable to this technique. We then optimized EdU-click and FACS-Seq for murine fecal bacteria and reveal that Prevotella UCG-001 and Ileibacterium are enriched in the replicating fraction. Finally, we labeled the actively replicating murine gut bacteria during exposure to cell wall-specific antibiotics in vitro. We show that regardless of the antibiotic used, the actively replicating bacteria largely consist of Ileibacterium, suggesting the resistance of this taxon to perturbations. Overall, we demonstrate how combining EdU-click and FACSeq can identify the actively replicating gut bacteria and their link with the composition of the whole community in both homeostatic and perturbed conditions. This technique will be instrumental in elucidating in situ bacterial replication dynamics in a variety of other ecological states, including colonization and species invasion, as well as for investigating the relationship between the replication and abundance of bacteria in complex communities.

The bacteria that live in our guts are known to influence our intestinal and overall health. Though we know a lot about which kinds of bacteria are in our guts, we still don't know much about which bacteria are actually alive and growing. This is important to know, because bacteria that are growing, or replicating, are more likely to impact our health than bacteria which are not replicating. Our research group aimed to address this issue by developing a new technique that can identify which gut bacteria are actively replicating. We first tested this technique on specific gut bacteria, and then we made sure the technique worked when it was used on the gut bacteria of mice. By using this technique, we identified several types of mouse gut bacteria that were actively replicating. We also demonstrated one possible application of this technique by using it to identify mouse gut bacteria that were able to replicate after they were grown with antibiotics. Overall, we have introduced a new technique to identify replicating gut bacteria and show how it can be used to increase our knowledge on which bacteria are growing in the gut. This technique will help us identify which bacteria may be more important to our health due to their active growth.

Stress Response Mechanisms of Salmonella Enteritidis to Sodium Hypochlorite at the Proteomic Level.

Salmonella Enteritidis (S. Enteritidis) can adapt to sublethal sodium hypochlorite conditions, which subsequently triggers stress resistance mechanisms in this pathogen. Hence, the current work aimed to reveal the underlying stress adaptation mechanisms in S. Enteritidis by phenotypic, proteomic, and physiological analyses. It was found that 130 ppm sodium hypochlorite resulted in a moderate inhibitory effect on bacterial growth and an increased accumulation of intracellular reactive oxygen species. In response to this sublethal treatment, a total of 492 proteins in S. Enteritidis showed significant differential abundance (p < 0.05; fold change >2.0 or <0.5), including 225 more abundant proteins and 267 less abundant proteins, as revealed by the tandem-mass-tags-based quantitative proteomics technology. Functional characterization further revealed that proteins related to flagellar assembly, two-component system, and phosphotransferase system were in less abundance, while those associated with ABC transporters were generally in more abundance. Specifically, the repression of flagellar-assembly-related proteins led to diminished swimming motility, which served as a potential energy conservation strategy. Moreover, altered abundance of lipid-metabolism-related proteins resulted in reduced cell membrane fluidity, which provided a survival advantage to S. Enteritidis. Taken together, these results indicate that S. Enteritidis employs multiple adaptation pathways to cope with sodium hypochlorite stress.

A CURE for Physiological Characterization of Bacterioplankton in Liquid Culture.

Bacterial characterization is an important aspect of microbiology that includes experimentally determining growth rates, environmental conditions conducive to growth, and the types of energy sources microorganisms can use. Researchers use this information to help understand and predict an organism's ecological distribution and environmental functions. Microbiology students generally conduct bacterial characterization experiments in their coursework; however, they are frequently restricted to model organisms without ecological relevance and already well-studied physiologies. We present a course-based undergraduate research experience (CURE) curriculum to involve students in characterization of previously untested, ecologically relevant aquatic free-living bacteria (bacterioplankton) cultures to identify the usable nutrient substrates, as well as the temperature and salinity ranges conducive to growth. Students use these results to connect their organism's physiology to the isolation environment. This curriculum also exposes students to advanced microbiology methods such as flow cytometry for measuring cell concentrations, teaches them to use the programming language R for data plotting, and emphasizes scientific communication through writing, speaking, poster creation/presentation, and social media. This CURE is an attractive introduction to scientific research and was successfully tested with 187 students in three semesters at two different universities. Students generated reproducible growth data for multiple strains across these different deployments, demonstrating the utility of the curriculum for research support.

Growth Substrate and Prophage Induction Collectively Influence Metabolite and Lipid Profiles in a Marine Bacterium.

Bacterial growth substrates influence a variety of biological functions, including the biosynthesis and regulation of lipid intermediates. The extent of this rewiring is not well understood nor has it been considered in the context of virally infected cells. Here, we used a one-host-two-temperate phage model system to probe the combined influence of growth substrate and phage infection on host carbon and lipid metabolism. Using untargeted metabolomics and lipidomics, we reported the detection of a suite of metabolites and lipid classes for two Sulfitobacter lysogens provided with three growth substrates of differing complexity and nutrient composition (yeast extract/tryptone [complex], glutamate and acetate). The growth medium led to dramatic differences in the detectable intracellular metabolites, with only 15% of 175 measured metabolites showing overlap across the three growth substrates. Between-strain differences were most evident in the cultures grown on acetate, followed by glutamate then complex medium. Lipid distribution profiles were also distinct between cultures grown on different substrates as well as between the two lysogens grown in the same medium. Five phospholipids, three aminolipid, and one class of unknown lipid-like features were identified. Most (≥94%) of these 75 lipids were quantifiable in all samples. Metabolite and lipid profiles were strongly determined by growth medium composition and modestly by strain type. Because fluctuations in availability and form of carbon substrates and nutrients, as well as virus pressure, are common features of natural systems, the influence of these intersecting factors will undoubtedly be imprinted in the metabolome and lipidome of resident bacteria. IMPORTANCE Community-level metabolomics approaches are increasingly used to characterize natural microbial populations. These approaches typically depend upon temporal snapshots from which the status and function of communities are often inferred. Such inferences are typically drawn from lab-based studies of select model organisms raised under limited growth conditions. To better interpret community-level data, the extent to which ecologically relevant bacteria demonstrate metabolic flexibility requires elucidation. Herein, we used an environmentally relevant model heterotrophic marine bacterium to assess the relationship between growth determinants and metabolome. We also aimed to assess the contribution of phage activity to the host metabolome. Striking differences in primary metabolite and lipid profiles appeared to be driven primarily by growth regime and, secondarily, by phage type. These findings demonstrated the malleable nature of metabolomes and lipidomes and lay the foundation for future studies that relate cellular composition with function in complex environmental microbial communities.