Biology subjects

Lanclos, V. C.

Publications and source records attributed to Lanclos, V. C..

12 recordsLinked to original sources

bioRxiv2025

Bioplastic Production Potential of Azospira suillum PS: Growth-Associated PHB Production Under Aerobic and Anaerobic Conditions

Plastics are indispensable in modern society, yet less than 10% of the 380 million tons produced annually are recycled. This accumulation of non-degradable waste has intensified the search for sustainable alternatives. Polyhydroxyalkanoates (PHAs), particularly polyhydroxybutyrate (PHB), are biodegradable polymers produced by microorganisms. Here, we investigate the capacity of Azospira suillum PS, a model facultative perchlorate-reducing bacterium, to synthesize PHB. We show that unlike most organisms, PHB production is growth-associated both aerobically, and anaerobically with perchlorate as an electron acceptor under moderate nitrogen deplete conditions. Genomic analysis revealed four phaC homologs, with one, phaC4, co-localized with phaB and phaR, suggesting a conserved operon. While three of the phaC genes were successfully deleted, phaC4 could not be disrupted, indicating an essential role for growth. Redox profiling revealed a positive correlation between PHB accumulation and NADPH/NADP+ ratios, supporting the role of cytoplasmic reducing equivalents in PHB synthesis. This study presents the first demonstration of PHA production under perchlorate-respiring conditions and one of the few documented cases of growth-associated PHA synthesis. It offers new insights into the genetic and metabolic basis of PHB production in A. suillum PS, underscoring its potential as a platform organism for sustainable, growth-coupled bioplastic production.

microbiology↗

bioRxiv2025

Lithosyntrophy: Obligate syntrophy in a phosphite-oxidizing, methanogenic culture

The anaerobic conversion of organic matter to methane and carbon dioxide typically relies on obligate syntrophic interactions between bacteria and methanogenic archaea, where interspecies hydrogen (H2) transfer enables thermodynamically constrained reactions to proceed near equilibrium. Syntrophs couple the oxidation of fermentation products such as fatty acids and alcohols to the reduction of protons to form H2. These reactions can only proceed if low H2 concentrations are maintained by H2-consuming syntrophic partners. Here, we describe "lithosyntrophy," a novel mode of syntrophic interaction in which electrons that drive hydrogenotrophic methanogenesis originate from an inorganic compound rather than from the canonical organic substrates. Candidatus Phosphitivorax anaerolimi strain Phox-21 oxidizes phosphite (HPO32-, oxidation state +3) to phosphate coupled to hydrogenogenesis in an obligate energetic dependency on a hydrogenotrophic methanogen, Methanoculleus sp. Physiology experiments, thermodynamic calculations, genomic annotation, and metaproteomics analysis collectively revealed a mechanism for syntrophic phosphite oxidation in Phox-21, which requires phosphite, acetate, and CO2 as co-substrates. In this pathway, electrons derived from phosphite drive H2 production via an electron-confurcating hydrogenase. Unlike previously characterized acetogenic phosphite oxidizers that grow without exogenous acetate, Phox-21 requires acetate to regenerate AMP, a cofactor required by the phosphite dehydrogenase, PtdF. Lithosyntrophic phosphite oxidizers may play important roles both in transferring reducing equivalents as well as biologically available phosphorus to other members of their surrounding microbial communities. We infer that lithosyntrophic DPO emerged before acetoclastic methanogenesis and was a major sink for acetate in the Archaean when phosphite was more abundant. Significance statementDissimilatory phosphite-oxidizing microorganisms (DPOM) use phosphite as an energy source, producing phosphate. While the two previously isolated DPOM couple phosphite oxidation to carbon fixation via the Wood-Ljungdahl pathway, Candidatus Phosphitivorax anaerolimi strain Phox-21 instead performs lithosyntrophic metabolism, coupling DPO to proton reduction in obligate partnership with a hydrogenotrophic methanogen. This establishes a novel link between phosphorus and carbon redox cycles: like organosyntrophy, DPO can fuel methanogenesis. Lithosyntrophy expands our understanding of syntrophic interactions and suggests similar processes may occur with other inorganic substrates in anoxic ecosystems. We propose lithosyntrophic DPO predates acetoclastic methanogenesis, serving as an early acetate sink during the Archaean. Following Earths oxygenation, phosphite depletion likely reduced competition for acetate, enabling the later evolution of acetoclastic methanogenesis.

microbiology↗

bioRxiv2025

Culture-supported ecophysiology of the SAR116 clade demonstrates metabolic and spatial niche partitioning

Marine SAR116 bacterioplankton are ubiquitous in surface waters across global oceans and form their own order, Puniceispirillales, within the Alphaproteobacteria. To date no comparative physiology among diverse SAR116 isolates has been performed to capture the functional diversity within the clade, and further, diversity through the lens of metabolic potential and environmental preferences via clade-wide pangenomics remains poorly constrained. Using high-throughput dilution-to-extinction cultivation, we isolated and genome sequenced five new and diverse SAR116 isolates from the northern Gulf of Mexico. Here we present a comparative physiological analysis of these SAR116 isolates, along with a pangenomic investigation of the SAR116 clade using a combination of metagenome-assembled genomes (MAGs, n=258), single-amplified genomes (SAGs, n=84), previously existing (n=2), and new isolate genomes (n=5), totaling 349 SAR116 genomes. Phylogenomic investigation supported the division of SAR116 into three distinct subclades, each with additional structure totalling 15 monophyletic groups. Our SAR116 isolates belonged to three groups within subclade I representing distinct genera with different morphologies and varied phenotypic responses to salinity and temperature. Overall, SAR116 genomes encoded differences in vitamin and amino acid synthesis, trace metal transport, and osmolyte synthesis and transport. They also had genetic potential for diverse sulfur oxidation metabolisms, placing SAR116 at the confluence of the organic and inorganic sulfur pools. SAR116 subclades showed distinct patterns in habitat preferences across open ocean, coastal, and estuarine environments, and three of our isolates represented the most abundant coastal and estuarine subclade. This investigation provides the most comprehensive exploration of SAR116 to date anchored by new culture genomes and physiology.

microbiology↗

bioRxiv2024

Polyhydroxybutyrate production in freshwater SAR11 (LD12)

SAR11 bacteria (order Pelagibacterales) are oligotrophs and often the most abundant bacterioplankton in aquatic environments. A subset of sequenced SAR11 genomes, predominantly in the brackish and freshwater SAR11 subclades, contain homologs of pha genes, which in other organisms confer the ability to store carbon and energy via polyhydroxyalkanoate (PHA) polymers. Here, we investigated the relevance of PHA production to SAR11 biology. Phylogenetics showed that Pha proteins occurred on a long branch and provided evidence for origin at the common ancestor of the brackish IIIa and freshwater LD12 subclades, followed by horizontal transfer within SAR11. Using the LD12 representative "Candidatus Fonsibacter ubiquis" strain LSUCC0530, we found that a large minority of LSUCC0530 cells contained a single Nile red-staining granule, confirmed that the cells produced polyhydroxybutyrate (PHB), and estimated the total PHB content in the cells. We heterologously expressed the LSUCC0530 phaCAB locus in Escherichia coli, finding it to be functional and the likely origin of the PHB. We also determined that, irrespective of changes to carbon, nitrogen, and phosphorus concentrations, a similar fraction of LSUCC0530 cells generated PHB granules and expression of the phaCAB locus remained constant. We suggest that PHB synthesis in LSUCC0530 may be constitutively active due to the slow growth dynamics and minimal regulation that characterize SAR11 bacteria. This is the first characterization of polymer storage in SAR11, providing new insights into the likely fitness advantage for cells harboring this metabolism.

microbiology↗

bioRxiv2024

New isolates refine the ecophysiology of the Roseobacter CHAB-I-5 lineage

The CHAB-I-5 cluster is a pelagic lineage that can comprise a significant proportion of all roseobacters in surface oceans and have predicted roles in biogeochemical cycling via heterotrophy, aerobic anoxygenic photosynthesis (AAnP), CO oxidation, DMSP degradation, and other metabolisms. Though cultures of CHAB-I-5 have been reported, none have been explored and the best known representative, strain SB2, was lost from culture after obtaining the genome sequence. We have isolated two new CHAB-I-5 representatives, strains US3C007 and FZCC0083, and assembled complete, circularized genomes with 98.7% and 92.5% average nucleotide identities with the SB2 genome. Comparison of these three with 49 other unique CHAB-I-5 metagenome-assembled and single-cell genomes indicated that the cluster represents a genus with two species, and we identified subtle differences in genomic content between the two species subclusters. Metagenomic recruitment from over fourteen hundred samples expanded their known global distribution and highlighted both isolated strains as representative members of the clade. FZCC0083 grew over twice as fast as US3C007 and over a wider range of temperatures. The axenic culture of US3C007 occurs as pleomorphic cells with most exhibiting a coccobacillus/vibrioid shape. We propose the name Thalassovivens spotae, gen nov., sp. nov. for the type strain US3C007T.

microbiology↗

bioRxiv2023

The AEGEAN-169 clade of bacterioplankton is synonymous with SAR11 subclade V (HIMB59) and metabolically distinct

Bacterioplankton of the SAR11 clade are the most abundant marine microorganisms and consist of numerous subclades spanning Order level divergence (Pelagibacterales). The assignment of the earliest diverging subclade V (a.k.a. HIMB59) to the Pelagibacterales is highly controversial, with multiple recent phylogenetic studies placing them completely separate from SAR11. Other than through phylogenomics, subclade V has not received detailed examination due to limited genomes from this group. Here, we assessed the ecogenomic characteristics of subclade V to better understand the role of this group in comparison to the Pelagibacterales. We used a new isolate genome, recently released single amplified genomes (SAGs) and metagenome-assembled genomes (MAGs), and previously established SAR11 genomes to perform a comprehensive comparative genomics analysis. We paired this analysis with recruitment of metagenomes spanning open ocean, coastal, and brackish systems. Phylogenomics, average amino acid identity, and 16S rRNA gene phylogeny indicate that SAR11 subclade V is synonymous with the ubiquitous AEGEAN-169 clade, and support the contention that this group represents a taxonomic Family. AEGEAN-169 shared many bulk genome qualities with SAR11, such as streamlining and low GC content, but genomes were generally larger. AEGEAN-169 had overlapping distributions with SAR11 but was metabolically distinct from SAR11 in its potential to transport and utilize a broader range of sugars as well as in transport of trace metals and thiamin. Thus, regardless of the ultimate phylogenetic placement of AEGEAN-169, these organisms have distinct metabolic capacities that likely allow them to differentiate their niche from canonical SAR11 taxa. IMPORTANCEOne goal of marine microbiologists is to uncover the roles various microorganisms are playing in biogeochemical cycles. Success in this endeavor relies on differentiating groups of microbes and circumscribing their relationships. An early-diverging group (subclade V) of the most abundant bacterioplankton, SAR11, has recently been proposed as a separate lineage that does not share a most recent common ancestor. But beyond phylogenetics, little has been done to evaluate how these organisms compare with SAR11. Our work leverages dozens of new genomes to demonstrate the similarities and differences between subclade V and SAR11. In our analysis, we also establish that subclade V is synonymous with a group of bacteria established from 16S rRNA gene sequences, AEGEAN-169. Subclade V/AEGEAN-169 has clear metabolic distinctions from SAR11 and their shared traits point to remarkable convergent evolution if they do not share a most recent common ancestor.

microbiology↗

bioRxiv2022

Ecophysiology and genomics of the brackish water adapted SAR11 subclade IIIa

The Order Pelagibacterales (SAR11) is the most abundant group of heterotrophic bacterioplankton in global oceans and comprises multiple subclades with unique spatiotemporal distributions. Subclade IIIa is the primary SAR11 group in brackish waters and shares a common ancestor with the dominant freshwater IIIb (LD12) subclade. Despite its dominance in brackish environments, subclade IIIa lacks systematic genomic or ecological studies. Here, we combine closed genomes from new IIIa isolates, new IIIa MAGS from San Francisco Bay (SFB), and 466 high-quality publicly available SAR11 genomes for the most comprehensive pangenomic study of subclade IIIa to date. Subclade IIIa represents a taxonomic family containing three genera (denoted as subgroups IIIa.1, IIIa.2, and IIIa.3) that had distinct ecological distributions related to salinity. The expansion of taxon selection within subclade IIIa also established previously noted metabolic differentiation in subclade IIIa compared to other SAR11 subclades such as glycine/serine prototrophy, mosaic glyoxylate shunt presence, and polyhydroxyalkanoate synthesis potential. Our analysis further shows metabolic flexibility among subgroups within IIIa. Additionally, we find that subclade IIIa.3 bridges the marine and freshwater clades based on its potential for compatible solute transport, iron utilization, and bicarbonate management potential. Pure culture experimentation validated differential salinity ranges in IIIa.1 and IIIa.3 and provided the first IIIa cell size and volume data. This study is an important step forward for understanding the genomic, ecological, and physiological differentiation of subclade IIIa and the overall evolutionary history of SAR11.

microbiology↗

bioRxiv2021

Ecophysiology of the cosmopolitan OM252 bacterioplankton (Gammaproteobacteria)

Among the thousands of species that comprise marine bacterioplankton communities, most remain functionally obscure. One key cosmopolitan group in this understudied majority is the OM252 clade of Gammaproteobacteria. Although frequently found in sequence data and even previously cultured, the diversity, metabolic potential, physiology, and distribution of this clade has not been thoroughly investigated. Here we examined these features of OM252 bacterioplankton using a newly isolated strain and genomes from publicly available databases. We demonstrated that this group constitutes a globally distributed novel genus (Candidatus Halomarinus), sister to Litoricola, comprising two subclades and multiple distinct species. OM252 organisms have small genomes (median 2.21 Mbp) and are predicted obligate aerobes capable of alternating between chemoorganoheterotrophic and chemolithotrophic growth using reduced sulfur compounds as electron donors, with subclade I genomes encoding the Calvin-Benson-Bassham cycle for carbon fixation. One representative strain of subclade I, LSUCC0096, had extensive halotolerance but a mesophilic temperature range for growth, with a maximum of 0.36 doublings/hr at 35{degrees}C. Cells were curved rod/spirillum-shaped, ~1.5 x 0.2 m. Growth on thiosulfate as the sole electron donor under autotrophic conditions was roughly one third that of heterotrophic growth, even though calculations indicated similar Gibbs energies for both catabolisms. These phenotypic data show that some Ca. Halomarinus organisms can switch between serving as carbon sources or sinks and indicate the likely anabolic cost of lithoautotrophic growth. Our results thus provide new hypotheses about the roles of these organisms in global biogeochemical cycling of carbon and sulfur. ImportanceMarine microbial communities are teeming with understudied taxa due to the sheer numbers of species in any given sample of seawater. One group, the OM252 clade of Gammaproteobacteria, has been identified in gene surveys from myriad locations, and one isolated organism has even been genome sequenced (HIMB30). However, further study of these organisms has not occurred. Using another isolated representative (strain LSUCC0096) and publicly available genome sequences from metagenomic and single-cell genomic datasets, we examined the diversity within the OM252 clade, the distribution of these taxa in the worlds oceans, reconstructed the predicted metabolism of the group, and quantified growth dynamics in LSUCC0096. Our results generate new knowledge about the previously enigmatic OM252 clade and point towards the importance of facultative chemolithoautotrophy for supporting some clades of ostensibly "heterotrophic" taxa.

microbiology↗

bioRxiv2021

Mechanisms Driving Genome Reduction of a Novel Roseobacter Lineage Showing Vitamin B12 Auxotrophy

Members of the marine Roseobacter group are key players in the global carbon and sulfur cycles. While over 300 species have been described, only 2% possess reduced genomes (mostly 3-3.5 Mbp) compared to an average roseobacter (>4 Mbp). These taxonomic minorities are phylogenetically diverse but form a Pelagic Roseobacter Cluster (PRC) at the genome content level. Here, we cultivated eight isolates constituting a novel Roseobacter lineage which we named CHUG. Metagenomic and metatranscriptomic read recruitment analyses showed that CHUG members were globally distributed and active in marine environments. CHUG members possess some of the smallest genomes (~2.52 Mb) among all known roseobacters, but they do not exhibit canonical features of genome streamlining like higher coding density or fewer paralogues and pseudogenes compared to their sister lineages. While CHUG members are clustered with traditional PRC members at the genome content level, they show important differences. Unlike other PRC members, neither the relative abundances of CHUG members nor their gene expression levels are correlated with chlorophyll a concentration across the global samples. Moreover, CHUG members cannot synthesize vitamin B12, a key metabolite made by most roseobacters but not by many phytoplankton species and thus thought to mediate the roseobacter-phytoplankton interactions. This combination of features is evidence for the hypothesis that CHUG members may have evolved a free-living lifestyle decoupled from phytoplankton. This ecological transition was accompanied by the loss of signature genes involved in roseobacter-phytoplankton symbiosis, suggesting that relaxation of purifying selection is likely an important driver of genome reduction in CHUG.

microbiology↗

bioRxiv2020

Expanding the diversity of bacterioplankton isolates and modeling isolation efficacy with large scale dilution-to-extinction cultivation

Cultivated bacterioplankton representatives from diverse lineages and locations are essential for microbiology, but the large majority of taxa either remain uncultivated or lack isolates from diverse geographic locales. We paired large scale dilution-to-extinction (DTE) cultivation with microbial community analysis and modeling to expand the phylogenetic and geographic diversity of cultivated bacterioplankton and to evaluate DTE cultivation success. Here, we report results from 17 DTE experiments totaling 7,820 individual incubations over three years, yielding 328 repeatably transferable isolates. Comparison of isolates to microbial community data of source waters indicated that we successfully isolated 5% of the observed bacterioplankton community throughout the study. 43% and 26% of our isolates matched operational taxonomic units and amplicon single nucleotide variants, respectively, within the top 50 most abundant taxa. Isolates included those from previously uncultivated clades such as SAR11 LD12 and Actinobacteria acIV, as well as geographically novel members from other ecologically important groups like SAR11 subclade IIIa, SAR116, and others; providing the first isolates in eight putatively new genera and seven putatively new species. Using a newly developed DTE cultivation model, we evaluated taxon viability by comparing relative abundance with cultivation success. The model i) revealed the minimum attempts required for successful isolation of taxa amenable to growth on our media, and ii) identified possible subpopulation viability variation in abundant taxa such as SAR11 that likely impacts cultivation success. By incorporating viability in experimental design, we can now statistically constrain the effort necessary for successful cultivation of specific taxa on a defined medium. ImportanceEven before the coining of the term "great plate count anomaly" in the 1980s, scientists had noted the discrepancy between the number of microorganisms observed under the microscope and the number of colonies that grew on traditional agar media. New cultivation approaches have reduced this disparity, resulting in the isolation of some of the "most wanted" bacterial lineages. Nevertheless, the vast majority of microorganisms remain uncultured, hampering progress towards answering fundamental biological questions about many important microorganisms. Furthermore, few studies have evaluated the underlying factors influencing cultivation success, limiting our ability to improve cultivation efficacy. Our work details the use of dilution-to-extinction (DTE) cultivation to expand the phylogenetic and geographic diversity of available axenic cultures. We also provide a new model of the DTE approach that uses cultivation results and natural abundance information to predict taxon-specific viability and iteratively constrain DTE experimental design to improve cultivation success.

microbiology↗

bioRxiv2020

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 organisms 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 for which the results have been known for decades. We present a course-based undergraduate research experience (CURE) curriculum to involve students in characterization of previously untested, ecologically relevant bacterioplankton cultures to identify the carbon substrates used for growth, as well as the temperature and salinity ranges conducive to growth. Students use these results to connect their organisms 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 147 students during the fall semester of 2018.

scientific communication and education↗

bioRxiv2016

Insights on the importance of salinity from the first cultured freshwater SAR11 (LD12) representative

Evolutionary transitions between fresh and salt water happen infrequently among bacterioplankton. Within the ubiquitous and highly abundant heterotrophic Alphaproteobacteria order Pelagibacterales (SAR11), most members live in marine habitats, but the LD12 subclade has evolved as a unique freshwater lineage. LD12 cells occur as some of the most dominant freshwater bacterioplankton, yet this group has remained elusive to cultivation, hampering a more thorough understanding of its biology. Here, we report the first successful isolation of an LD12 representative, strain LSUCC0530, using high throughput dilution to extinction cultivation methods, and its complete genome sequence. Growth experiments corroborate ecological data suggesting active populations of LD12 in brackish water up to salinities of ~5. LSUCC0530 has the smallest closed genome thus far reported for a SAR11 strain (1.16 Mbp). The genome affirms many previous metabolic predictions from cultivation-independent analyses, like a complete Embden-Meyerhof-Parnas glycolysis pathway, but also provides novel insights, such as the first isocitrate dehydrogenase in LD12, a likely homologous recombination of malate synthase from outside of the SAR11 clade, and analogous substitutions of ion transporters with others that occur throughout the rest of the SAR11 clade. Growth data support metagenomic recruitment results suggesting temperature-based ecotype diversification within LD12. Key gene losses for osmolyte uptake provide a succinct hypothesis for the evolutionary transition of LD12 from salt to fresh water. For strain LSUCC0530, we propose the provisional nomenclature Candidatus Fonsibacter ubiquis.

microbiology↗