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Guarnieri, Michael T.

Publications and source records attributed to Guarnieri, Michael T..

7 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Cyanobacteria newly isolated from marine volcanic seeps display rapid sinking and robust, high-density growth

Cyanobacteria are photosynthetic organisms that play important roles in carbon cycling and are promising bioproduction chassis. Here, we isolate two novel cyanobacteria with 4.6Mbp genomes, UTEX 3221 and UTEX 3222, from a unique marine environment with naturally elevated CO₂. We describe complete genome sequences for both isolates and, focusing on UTEX 3222 due to its planktonic growth in liquid, characterize biotechnologically relevant growth and biomass characteristics. UTEX 3222 outpaces other fast-growing model strains on a solid medium. It can double every 2.35 hours in a liquid medium and grows to high density (>31 g/L biomass dry weight) in batch culture, nearly double that of Synechococcus sp. PCC 11901, whose high-density growth was recently reported. In addition, UTEX 3222 sinks readily, settling more quickly than other fast-growing strains, suggesting favorable economics of harvesting UTEX 3222 biomass. These traits may make UTEX 3222 a compelling choice for marine carbon dioxide removal (CDR) and photosynthetic bioproduction from CO₂. Overall, we find that bio-prospecting in environments with naturally elevated CO₂ may uncover novel CO₂-metabolizing organisms with unique characteristics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Polyphosphate kinase deletion increases laboratory productivity in cyanobacteria

Identification and manipulation of cellular energy regulation mechanisms may be a strategy to increase productivity in photosynthetic organisms. This work tests the hypothesis that polyphosphate synthesis and degradation play a role in energy management by storing or dissipating energy in the form of ATP. A polyphosphate kinase ( ppk ) knock-out strain unable to synthesize polyphosphate was generated in the cyanobacterium Synechocystis sp. PCC 6803. This mutant strain demonstrated higher ATP levels and faster growth than the wildtype strain in high-carbon conditions and had a growth defect under multiple stress conditions. In a strain that combined ppk deletion with heterologous expression of ethylene-forming enzyme, higher ethylene productivity was observed than in the wildtype background. These results support the role of polyphosphate synthesis and degradation as an energy regulation mechanism and suggest that such mechanisms may be effective targets in biocontainment design.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Risk Assessment of Industrial Microbes Using a Terrestrial Mesocosm Platform

Abstract Industrial microbes and bio-derived products have emerged as an integral component of the bioeconomy, with an array of agricultural, bioenergy, and biomedical applications. However, the rapid development of microbial biotechnology raises concerns related to environmental escape of laboratory microbes, detection and tracking thereof, and resultant impact upon native ecosystems. Indeed, though wild-type and genetically modified microbes are actively deployed in industrial bioprocesses, an understanding of microbial interactivity and impact upon the environment is severely lacking. In particular, the persistence and sustained ecosystem impact of industrial microbes following laboratory release or unintentional laboratory escape remains largely unexplored. Herein, we investigate the applicability of soil-sorghum mesocosms for the ecological risk assessment of the industrial microbe, Saccharomyces cerevisiae . We developed and applied a suite of diagnostic and bioinformatic analyses, including digital droplet PCR, microscopy, and phylogenomic analyses to assess the impacts of a terrestrial ecosystem perturbation event over a 30-day time course. The platform enables reproducible, high-sensitivity tracking of S. cerevisiae in a complex soil microbiome and analysis of the impact upon abiotic soil characteristics and soil microbiome population dynamics and diversity. The resultant data indicate that even though S. cerevisiae is relatively short-lived in the soil, a single perturbation event can have sustained impact upon mesocosm soil composition and underlying microbial populations in our system, underscoring the necessity for more comprehensive risk assessment and development of mitigation and biocontainment strategies in industrial bioprocesses.

09 BIOMASS FUELS↗
DOE OSTI2023

Metabolic engineering of Synechococcus elongatus 7942 for enhanced sucrose biosynthesis

The capability of cyanobacteria to produce sucrose from CO 2 and light has a remarkable societal and biotechnological impact since sucrose can serve as a carbon and energy source for a variety of heterotrophic organisms and can be converted into value-added products. However, most metabolic engineering efforts have focused on understanding local pathway alterations that drive sucrose biosynthesis and secretion in cyanobacteria rather than analyzing the global flux re-routing that occurs following induction of sucrose production by salt stress. Here, we investigated global metabolic flux alterations in a sucrose-secreting (cscB-overexpressing) strain relative to its wild-type Synechococcus elongatus 7942 parental strain. Here we used targeted metabolomics, 13 C metabolic flux analysis (MFA), and genome-scale modeling (GSM) as complementary approaches to elucidate differences in cellular resource allocation by quantifying metabolic profiles of three cyanobacterial cultures - wild-type S. elongatus 7942 without salt stress (WT), wild-type with salt stress (WT/NaCl), and the cscB-overexpressing strain with salt stress (cscB/NaCl) - all under photoautotrophic conditions. We quantified the substantial rewiring of metabolic fluxes in WT/NaCl and cscB/NaCl cultures relative to WT and identified a metabolic bottleneck limiting carbon fixation and sucrose biosynthesis. This bottleneck was subsequently mitigated through heterologous overexpression of glyceraldehyde-3-phosphate dehydrogenase in an engineered sucrose-secreting strain. Our study also demonstrates that combining 13 C-MFA and GSM is a useful strategy to both extend the coverage of MFA beyond central metabolism and to improve the accuracy of flux predictions provided by GSM.

09 BIOMASS FUELS↗
DOE OSTI2023

Heterologous expression of formate dehydrogenase enables photoformatotrophy in the emerging model microalga, Picochlorum renovo

Rising global greenhouse gas emissions and the impacts of resultant climate change necessitate development and deployment of carbon capture and conversion technologies. Amongst the myriad of bio-based conversion approaches under evaluation, a formate bio-economy has recently been proposed, wherein CO 2 -derived formate serves as a substrate for concurrent carbon and energy delivery to microbial systems. To date, this approach has been explored in chemolithotrophic and heterotrophic organisms via native or engineered formatotrophy. However, utilization of this concept in phototrophic organisms has yet to be reported. Herein, we have taken the first steps to establish formate utilization in Picochlorum renovo, a recently characterized eukaryotic microalga with facile genetic tools and promising applied biotechnology traits. Plastidial heterologous expression of a formate dehydrogenase (FDH) enabled P. renovo growth on formate as a carbon and energy source. Further, FDH expression enhanced cultivation capacity on ambient CO 2 , underscoring the potential for bypass of conventional CO 2 capture and concentration limitations. This work establishes a photoformatotrophic cultivation regime that leverages light energy-driven formate utilization. The resultant photosynthetic formate platform has widespread implications for applied phototrophic cultivation systems and the bio-economy at large.

09 BIOMASS FUELS↗