Nitrogen source influences the interactions of comammox bacteria with aerobic nitrifiers.

While the co-existence of comammox Nitrospira with canonical nitrifiers is well documented in diverse ecosystems, there is still a dearth of knowledge about the mechanisms underpinning their interactions. Understanding these interaction mechanisms is important as they may play a critical role in governing nitrogen biotransformation in natural and engineered ecosystems. In this study, we tested the ability of two environmentally relevant factors (nitrogen source and availability) to shape interactions between strict ammonia and nitrite-oxidizing bacteria and comammox Nitrospira in continuous flow column reactors. The composition of inorganic nitrogen species in reactors fed either ammonia or urea was similar during the lowest input nitrogen concentration (1 mg-N/L), but higher concentrations (2 and 4 mg-N/L) promoted significant differences in nitrogen species composition and nitrifier abundances. The abundance and diversity of comammox Nitrospira were dependent on both nitrogen source and input concentrations as multiple comammox Nitrospira populations were preferentially enriched in the urea-fed system. In contrast, their abundance was reduced in response to higher nitrogen concentrations in the ammonia-fed system. The preferential enrichment of comammox Nitrospira in the urea-fed system could be associated with their ureolytic activity calibrated to their ammonia oxidation rates, thus minimizing ammonia accumulation, which may be partially inhibitory. However, an increased abundance of comammox Nitrospira was not associated with a reduced abundance of nitrite oxidizers in the urea-fed system while a negative correlation was found between them in the ammonia-fed system, the latter dynamic likely emerging from reduced availability of nitrite to strict nitrite oxidizers at low ammonia concentrations. IMPORTANCE: Nitrification is an essential biological process in drinking water and wastewater treatment systems for treating nitrogen pollution. The discovery of comammox Nitrospira and their detection alongside canonical nitrifiers in these engineered ecosystems have made it necessary to understand the environmental conditions that regulate their abundance and activity relative to other better-studied nitrifiers. This study aimed to evaluate two important factors that could potentially influence the behavior of nitrifying bacteria and, therefore, impact nitrification processes. Column reactors fed with either ammonia or urea were systematically monitored to capture changes in nitrogen biotransformation and the nitrifying community as a function of influent nitrogen concentration, nitrogen source, and reactor depth. Our findings show that with increased ammonia availability, comammox Nitrospira decreased in abundance while nitrite oxidizers abundance increased. Yet, in systems with increasing urea availability, comammox Nitrospira abundance and diversity increased without an associated reduction in the abundance of canonical nitrifiers.

Rigorous science demands support of transgender scientists.

To build a just, equitable, and diverse academy, scientists and institutions must address systemic barriers that sex and gender minorities face. This Commentary summarizes (1) critical context informing the contemporary oppression of transgender people, (2) how this shapes extant research on sex and gender, and (3) actions to build an inclusive and rigorous academy for all.

Ammonia-oxidizing bacteria and archaea exhibit differential nitrogen source preferences.

Ammonia-oxidizing microorganisms (AOM) contribute to one of the largest nitrogen fluxes in the global nitrogen budget. Four distinct lineages of AOM: ammonia-oxidizing archaea (AOA), beta- and gamma-proteobacterial ammonia-oxidizing bacteria (ß-AOB and γ-AOB) and complete ammonia oxidizers (comammox), are thought to compete for ammonia as their primary nitrogen substrate. In addition, many AOM species can utilize urea as an alternative energy and nitrogen source through hydrolysis to ammonia. How the coordination of ammonia and urea metabolism in AOM influences their ecology remains poorly understood. Here we use stable isotope tracing, kinetics and transcriptomics experiments to show that representatives of the AOM lineages employ distinct regulatory strategies for ammonia or urea utilization, thereby minimizing direct substrate competition. The tested AOA and comammox species preferentially used ammonia over urea, while ß-AOB favoured urea utilization, repressed ammonia transport in the presence of urea and showed higher affinity for urea than for ammonia. Characterized γ-AOB co-utilized both substrates. These results reveal contrasting niche adaptation and coexistence patterns among the major AOM lineages.

Building a queer- and trans-inclusive microbiology conference.

Microbiology conferences can be powerful places to build collaborations and exchange ideas, but for queer and transgender (trans) scientists, they can also become sources of alienation and isolation. Many conference organizers would like to create welcoming and inclusive events but feel ill-equipped to make this vision a reality, and a historical lack of representation of queer and trans folks in microbiology means we rarely occupy these key leadership roles ourselves. Looking more broadly, queer and trans scientists are systematically marginalized across scientific fields, leading to disparities in career outcomes, professional networks, and opportunities, as well as the loss of unique scientific perspectives at all levels. For queer and trans folks with multiple, intersecting, marginalized identities, these barriers often become even more severe. Here, we draw from our experiences as early-career microbiologists to provide concrete, practical advice to help conference organizers across research communities design inclusive, safe, and welcoming conferences, where queer and trans scientists can flourish.

Thermophilic Dehalococcoidia with unusual traits shed light on an unexpected past.

Although the phylum Chloroflexota is ubiquitous, its biology and evolution are poorly understood due to limited cultivability. Here, we isolated two motile, thermophilic bacteria from hot spring sediments belonging to the genus Tepidiforma and class Dehalococcoidia within the phylum Chloroflexota. A combination of cryo-electron tomography, exometabolomics, and cultivation experiments using stable isotopes of carbon revealed three unusual traits: flagellar motility, a peptidoglycan-containing cell envelope, and heterotrophic activity on aromatics and plant-associated compounds. Outside of this genus, flagellar motility has not been observed in Chloroflexota, and peptidoglycan-containing cell envelopes have not been described in Dehalococcoidia. Although these traits are unusual among cultivated Chloroflexota and Dehalococcoidia, ancestral character state reconstructions showed flagellar motility and peptidoglycan-containing cell envelopes were ancestral within the Dehalococcoidia, and subsequently lost prior to a major adaptive radiation of Dehalococcoidia into marine environments. However, despite the predominantly vertical evolutionary histories of flagellar motility and peptidoglycan biosynthesis, the evolution of enzymes for degradation of aromatics and plant-associated compounds was predominantly horizontal and complex. Together, the presence of these unusual traits in Dehalococcoidia and their evolutionary histories raise new questions about the timing and selective forces driving their successful niche expansion into global oceans.

Ammonia-Oxidizing Bacteria Maintain Abundance but Lower amoA-Gene Expression during Cold Temperature Nitrification Failure in a Full-Scale Municipal Wastewater Treatment Plant.

In this study, we explore the relationship between community structure and transcriptional activity of ammonia-oxidizing bacteria during cold temperature nitrification failure in three parallel full-scale sequencing batch reactors (SBRs) treating municipal wastewater. In the three reactors, ammonia concentrations increased with declines in wastewater temperature below 15°C. We quantified and sequenced 16S rRNA and ammonia monooxygenase (amoA) gene fragments in DNA and RNA extracts from activated sludge samples collected from the SBRs during the warmer seasons (summer and fall) and when water temperatures were below 15°C (winter and spring). Taxonomic community composition of amoA genes and transcripts did not vary much between the warmer and colder seasons. However, we observed significant differences in amoA transcript copy numbers between fall (highest) and spring (lowest). Ammonia-oxidizing bacteria of the genus Nitrosomonas sp. could maintain their population abundance despite lowering their amoA gene expression during winter and spring. In spite of relatively low population abundance, an amoA amplicon sequence variant (ASV) cluster identified as most similar to the amoA gene of Nitrosospira briensis showed the highest amoA transcript-to-gene ratio throughout all four seasons, indicating that some nitrifiers remain active at wastewater temperatures below 15°C. Our results show that 16S rRNA and amoA gene copy numbers are limited predictors of cell activity. To optimize function and performance of mixed community bioprocesses, we need to collect high-resolution quantitative transcriptomic and potentially proteomic data to resolve the response of individual species to changes in environmental parameters in engineered systems. IMPORTANCE The diverse microbial community of activated sludge used in biological treatment systems exhibits dynamic seasonal shifts in community composition and activity. Many wastewater treatment plants in temperate/continental climates experience seasonal cold temperature nitrification failure. "Seasonal nitrification failure" is the discharge of elevated concentrations of ammonia (greater than 4 mg/liter) with treated wastewater during the winter (influent wastewater temperatures below 13°C). This study aims at expanding our understanding of how ammonia-oxidizing bacteria in activated sludge change in activity and growth across seasons. We quantified the ammonia monooxygenase (amoA) gene and transcript copy numbers using real-time PCR and sequenced the amoA amplicons to reveal community structure and activity changes of nitrifying microbial populations during seasonal nitrification failure in three full-scale sequencing batch reactors (SRBs) treating municipal wastewater. Relevant findings presented in this study contribute to explain seasonal nitrification performance variability in SRBs.

Seasonal Dynamics of the Activated Sludge Microbiome in Sequencing Batch Reactors, Assessed Using 16S rRNA Transcript Amplicon Sequencing.

Activated sludge is comprised of diverse microorganisms which remediate wastewater. Previous research has characterized activated sludge using 16S rRNA gene amplicon sequencing, which can help to address questions on the relative abundance of microorganisms. In this study, we used 16S rRNA transcript sequencing in order to characterize "active" populations (via protein synthesis potential) and gain a deeper understanding of microbial activity patterns within activated sludge. Seasonal abundances of individual populations in activated sludge change over time, yet a persistent group of core microorganisms remains throughout the year which are traditionally classified on presence or absence without monitoring of their activity or growth. The goal of this study was to further our understanding of how the activated sludge microbiome changes between seasons with respect to population abundance, activity, and growth. Triplicate sequencing batch reactors were sampled at 10-min intervals throughout reaction cycles during all four seasons. We quantified the gene and transcript copy numbers of 16S rRNA amplicons using real-time PCR and sequenced the products to reveal community abundance and activity changes. We identified 108 operational taxonomic units (OTUs) with stable abundance, activity, and growth throughout the year. Nonproliferating OTUs were commonly human health related, while OTUs that showed seasonal abundance changes have previously been identified as being associated with floc formation and bulking. We observed significant differences in 16S rRNA transcript copy numbers, particularly at lower temperatures in winter and spring. The study provides an analysis of the seasonal dynamics of microbial activity variations in activated sludge based on quantifying and sequencing 16S rRNA transcripts.IMPORTANCE Sequencing batch reactors are a common design for wastewater treatment plants, particularly in smaller municipalities, due to their low footprint and ease of operations. However, like for most treatment plants in temperate/continental climates, the microbial community involved in water treatment is highly seasonal and its biological processes can be sensitive to cold temperatures. The seasonality of these microbial communities has been explored primarily in conventional treatment plants and not in sequencing batch reactors. Furthermore, most studies often only address which organisms are present. However, the activated sludge microbial community is very diverse, and it is often hard to discern which organisms are active and which organisms are simply present. In this study, we applied additional sequencing techniques to also address the issues of which organisms are active and which organisms are growing. By addressing these issues, we gained new insights into seasonal microbial populations dynamics and activity patterns affecting wastewater treatment.

Composition and Dynamics of the Activated Sludge Microbiome during Seasonal Nitrification Failure.

Wastewater treatment plants in temperate climate zones frequently undergo seasonal nitrification failure in the winter month yet maintain removal efficiency for other contaminants. We tested the hypothesis that nitrification failure can be correlated to shifts in the nitrifying microbial community. We monitored three parallel, full-scale sequencing batch reactors over the course of a year with respect to reactor performance, microbial community composition via 16S rRNA gene amplicon sequencing, and functional gene abundance using qPCR. All reactors demonstrated similar changes to their core microbiome, and only subtle variations among seasonal and transient taxa. We observed a decrease in species richness during the winter, with a slow recovery of the activated sludge community during spring. Despite the change in nitrification performance, ammonia monooxygenase gene abundances remained constant throughout the year, as did the relative sequence abundance of Nitrosomonadacae. This suggests that nitrification failure at colder temperatures might result from different reaction kinetics of nitrifying taxa, or that other organisms with strong seasonal shifts in population abundance, e.g. an uncultured lineage of Saprospiraceae, affect plant performance in the winter. This research is a comprehensive analysis of the seasonal microbial community dynamics in triplicate full-scale sequencing batch reactors and ultimately strengthens our basic understanding of the microbial ecology of activated sludge communities by revealing seasonal succession patterns of individual taxa that correlate with nutrient removal efficiency.

Photobiological transformation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) using Rhodobacter sphaeroides.

Pump-and-treat strategies for groundwater containing explosives may be necessary when the contaminated water approaches sensitive receptors. This project investigated bacterial photosynthesis as a strategy for ex situ treatment, using light as the primary energy source to facilitate RDX transformation. The objective was to characterize the ability of photosynthetic Rhodobacter sphaeroides (strain ATCC(®) 17023 ™) to transform the high-energy explosive RDX. R. sphaeroides transformed 30 µM RDX within 40 h under light conditions; RDX was not fully transformed in the dark (non-photosynthetic conditions), suggesting that photosynthetic electron transfer was the primary mechanism. Experiments with RDX demonstrated that succinate and malate were the most effective electron donors for photosynthesis, but glycerol was also utilized as a photosynthetic electron donor. RDX was transformed irrespective of the presence of carbon dioxide. The electron shuttling compound anthraquinone-2,6-disulfonate (AQDS) increased transformation kinetics in the absence of CO2, when the cells had excess NADPH that needed to be re-oxidized because there was limited CO2 for carbon fixation. When CO2 was added, the cells generated more biomass, and AQDS had no stimulatory effect. End products indicated that RDX carbon became CO2, biomass, and a soluble, uncharacterized aqueous metabolite, determined using (14)C-labeled RDX. These data are the first to suggest that photobiological explosives transformation is possible and will provide a framework for which phototrophy can be used in environmental restoration of explosives contaminated water.