Ratio of electron donor to acceptor influences metabolic specialization and denitrification dynamics in Pseudomonas aeruginosa in a mixed carbon medium

© The Author(s), 2021. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Zhang, I. H., Mullen, S., Ciccarese, D., Dumit, D., Martocello, D. E., Toyofuku, M., Nomura, N., Smriga, S., & Babbin, A. R. Ratio of electron donor to acceptor influences metabolic specialization and denitrification dynamics in Pseudomonas aeruginosa in a mixed carbon medium. Frontiers in Microbiology, 12, (2021): 711073, https://doi.org/10.3389/fmicb.2021.711073.Denitrifying microbes sequentially reduce nitrate (NO3–) to nitrite (NO2–), NO, N2O, and N2 through enzymes encoded by nar, nir, nor, and nos. Some denitrifiers maintain the whole four-gene pathway, but others possess partial pathways. Partial denitrifiers may evolve through metabolic specialization whereas complete denitrifiers may adapt toward greater metabolic flexibility in nitrogen oxide (NOx–) utilization. Both exist within natural environments, but we lack an understanding of selective pressures driving the evolution toward each lifestyle. Here we investigate differences in growth rate, growth yield, denitrification dynamics, and the extent of intermediate metabolite accumulation under varying nutrient conditions between the model complete denitrifier Pseudomonas aeruginosa and a community of engineered specialists with deletions in the denitrification genes nar or nir. Our results in a mixed carbon medium indicate a growth rate vs. yield tradeoff between complete and partial denitrifiers, which varies with total nutrient availability and ratios of organic carbon to NOx–. We found that the cultures of both complete and partial denitrifiers accumulated nitrite and that the metabolic lifestyle coupled with nutrient conditions are responsible for the extent of nitrite accumulation.Funding for this work was provided by Simons Foundation award 622065 and an MIT Environmental Solutions Initiative seed grant to AB. Additional support was received by the MIT Ferry Fund

Microbial Ecology of Four Coral Atolls in the Northern Line Islands

Microbes are key players in both healthy and degraded coral reefs. A combination of metagenomics, microscopy, culturing, and water chemistry were used to characterize microbial communities on four coral atolls in the Northern Line Islands, central Pacific. Kingman, a small uninhabited atoll which lies most northerly in the chain, had microbial and water chemistry characteristic of an open ocean ecosystem. On this atoll the microbial community was equally divided between autotrophs (mostly Prochlorococcus spp.) and heterotrophs. In contrast, Kiritimati, a large and populated (∼5500 people) atoll, which is most southerly in the chain, had microbial and water chemistry characteristic of a near-shore environment. On Kiritimati, there were 10 times more microbial cells and virus-like particles in the water column and these microbes were dominated by heterotrophs, including a large percentage of potential pathogens. Culturable Vibrios were common only on Kiritimati. The benthic community on Kiritimati had the highest prevalence of coral disease and lowest coral cover. The middle atolls, Palmyra and Tabuaeran, had intermediate densities of microbes and viruses and higher percentages of autotrophic microbes than either Kingman or Kiritimati. The differences in microbial communities across atolls could reflect variation in 1) oceaonographic and/or hydrographic conditions or 2) human impacts associated with land-use and fishing. The fact that historically Kingman and Kiritimati did not differ strongly in their fish or benthic communities (both had large numbers of sharks and high coral cover) suggest an anthropogenic component in the differences in the microbial communities. Kingman is one of the world's most pristine coral reefs, and this dataset should serve as a baseline for future studies of coral reef microbes. Obtaining the microbial data set, from atolls is particularly important given the association of microbes in the ongoing degradation of coral reef ecosystems worldwide

Ecological significance of bacteria associated with coral reef fish feces

Bacteria may play important roles in the biogeochemical cycling of coral reef fish feces and in the interactions between fishes and corals. This interaction potential was observed in a study of milkfish (Chanos chanos) aquaculture farms in the Philippines. Effluents from suspended fish pens created steep gradients of particulate organic carbon and other water characteristics that extended into nearby coral reefs. Highly similar bacterial phylotypes co-occurred in milkfish feces and in corals indicating the potential for transport of fecal particles and interaction with coral. In a separate study at Palmyra Atoll, bacteria abundances ranged 1̃0⁹ to 10¹¹ g⁻¹ dry wt among feces of parrotfish (Chlorurus sordidus), snapper (Lutjanus bohar), and surgeonfish (Acanthurus nigricans). Bacteria in parrotfish feces grew at a rate of 2̃ x 10⁸ cells g⁻¹ dry wt feces h⁻¹ . To improve our ability to observe growing marine bacteria, I tested a method for using the thymidine analogue 5-ethynyl-2'-deoxyuridine (EdU), which becomes incorporated during DNA synthesis and can be detected using c̀lick' chemistry combined with epifluorescence microscopy. The percentage of EdU-labeled bacteria ranged from 6̃% to 18% during a time course incubation of natural seawater assemblages. Additionally, cell specific signal intensities could be quantified, demonstrating the method's potential for determining individual cell growth rate. Other studies addressed phylotype composition of feces-associated assemblages. Analyses of feces-derived 16S rRNA gene clones revealed that Vibrionaceae dominated parrotfish and snapper feces. Many of these genes clustered phylogenetically to cultured Vibrio spp. and Photobacterium spp. Other Vibrionaceae- like sequences comprised a distinct phylogenetic group that may represent 'feces-specific' taxa. PCR primers specific to this f̀ish feces vibrio-like' group (FFV-L) were used to screen 'aged' parrotfish feces. FFV-L could be detected in feces collected over several days, indicating that feces may permit persistence of FFV-L in reefs. In addition to FFV-L, other bacteria phylotypes consistently occurred in aged feces, as determined via denaturing gradient gel electrophoresis (DGGE) analyses. To test the responses of coral-associated bacteria during short-term interactions with feces, both in situ and aquaria experiments were performed whereby corals were challenged with acute feces doses. The results reinforce the potential for bacteria transfer between feces and corals. Meanwhile, acute challenges with parrotfish feces did not impair the overall health of three coral specie

Denitrifying bacteria respond to and shape microscale gradients within particulate matrices

Heterotrophic denitrification enables facultative anaerobes to continue growing even when limited by oxygen (O₂) availability. Particles in particular provide physical matrices characterized by reduced O₂ permeability even in well-oxygenated bulk conditions, creating microenvironments where microbial denitrifiers may proliferate. Whereas numerical particle models generally describe denitrification as a function of radius, here we provide evidence for heterogeneity of intraparticle denitrification activity due to local interactions within and among microcolonies. Pseudomonas aeruginosa cells and microcolonies act to metabolically shade each other, fostering anaerobic processes just microns from O₂-saturated bulk water. Even within well-oxygenated fluid, suboxia and denitrification reproducibly developed and migrated along sharp 10 to 100 µm gradients, driven by the balance of oxidant diffusion and local respiration. Moreover, metabolic differentiation among densely packed cells is dictated by the diffusional supply of O₂, leading to distinct bimodality in the distribution of nitrate and nitrite reductase expression. The initial seeding density controls the speed at which anoxia develops, and even particles seeded with few bacteria remain capable of becoming anoxic. Our empirical results capture the dynamics of denitrifier gene expression in direct association with O₂ concentrations over microscale physical matrices, providing observations of the co-occurrence and spatial arrangement of aerobic and anaerobic processes.Simons Foundation (Award 622065

Chemotaxis toward phytoplankton drives organic matter partitioning among marine bacteria

The microenvironment surrounding individual phytoplankton cells is often rich in dissolved organic matter (DOM), which can attract bacteria by chemotaxis. These “phycospheres” may be prominent sources of resource heterogeneity in the ocean, affecting the growth of bacterial populations and the fate of DOM. However, these effects remain poorly quantified due to a lack of quantitative ecological frameworks. Here, we used video microscopy to dissect with unprecedented resolution the chemotactic accumulation of marine bacteria around individual Chaetoceros affinis diatoms undergoing lysis. The observed spatiotemporal distribution of bacteria was used in a resource utilization model to map the conditions under which competition between different bacterial groups favors chemotaxis. The model predicts that chemotactic, copiotrophic populations outcompete nonmotile, oligotrophic populations during diatom blooms and bloom collapse conditions, resulting in an increase in the ratio of motile to nonmotile cells and in the succession of populations. Partitioning of DOM between the two populations is strongly dependent on the overall concentration of bacteria and the diffusivity of different DOM substances, and within each population, the growth benefit from phycospheres is experienced by only a small fraction of cells. By informing a DOM utilization model with highly resolved behavioral data, the hybrid approach used here represents a new path toward the elusive goal of predicting the consequences of microscale interactions in the ocean.National Science Foundation (U.S.) (Ocean Sciences Postdoctoral Fellowship)Gordon and Betty Moore Foundation (Marine Microbial Initiative Investigator Award GBMF 3783

Intermittent turbulence in flowing bacterial suspensions

Dense suspensions of motile bacteria, possibly including the human gut microbiome, exhibit collective dynamics akin to those observed in classic, high Reynolds number turbulence with important implications for chemical and biological transport, yet this analogy has remained primarily qualitative. Here, we present experiments in which a dense suspension of Bacillus subtilis bacteria was flowed through microchannels and the velocity statistics of the flowing suspension were quantified using a recently developed velocimetry technique coupled with vortex identification methods. Observations revealed a robust intermittency phenomenon, whereby the average velocity profile of the suspension fluctuated between a plug-like flow and a parabolic flow profile. This intermittency is a hallmark of the onset of classic turbulence and Lagrangian tracking revealed that it here originates from the presence of transient vortices in the active, collective motion of the bacteria locally reinforcing the externally imposed flow. These results link together two entirely different manifestations of turbulence and show the potential of the microfluidic approach to mimic the environment characteristic of certain niches of the human microbiome