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1. Pyruvate Production by Escherichia coli by Use of Pyruvate Dehydrogenase Variants

   https://doi.org/10.1128/AEM.00487-21  

   Moxley, W. Chris ; Eiteman, Mark A. ( June 2021 , Applied and Environmental Microbiology)

   Atomi, Haruyuki (Ed.)

   ABSTRACT Altering metabolic flux at a key branch point in metabolism has commonly been accomplished through gene knockouts or by modulating gene expression. An alternative approach to direct metabolic flux preferentially toward a product is decreasing the activity of a key enzyme through protein engineering. In Escherichia coli , pyruvate can accumulate from glucose when carbon flux through the pyruvate dehydrogenase complex is suppressed. Based on this principle, 16 chromosomally expressed AceE variants were constructed in E. coli C and compared for growth rate and pyruvate accumulation using glucose as the sole carbon source. To prevent conversion of pyruvate to other products, the strains also contained deletions in two nonessential pathways: lactate dehydrogenase ( ldhA ) and pyruvate oxidase ( poxB ). The effect of deleting phosphoenolpyruvate synthase ( ppsA ) on pyruvate assimilation was also examined. The best pyruvate-accumulating strains were examined in controlled batch and continuous processes. In a nitrogen-limited chemostat process at steady-state growth rates of 0.15 to 0.28 h −1 , an engineered strain expressing the AceE[H106V] variant accumulated pyruvate at a yield of 0.59 to 0.66 g pyruvate/g glucose with a specific productivity of 0.78 to 0.92 g pyruvate/g cells·h. These results provide proof of concept that pyruvate dehydrogenase complex variants can effectively shift carbon flux away from central carbon metabolism to allow pyruvate accumulation. This approach can potentially be applied to other key enzymes in metabolism to direct carbon toward a biochemical product. IMPORTANCE Microbial production of biochemicals from renewable resources has become an efficient and cost-effective alternative to traditional chemical synthesis methods. Metabolic engineering tools are important for optimizing a process to perform at an economically feasible level. This study describes an additional tool to modify central metabolism and direct metabolic flux to a product. We have shown that variants of the pyruvate dehydrogenase complex can direct metabolic flux away from cell growth to increase pyruvate production in Escherichia coli . This approach could be paired with existing strategies to optimize metabolism and create industrially relevant and economically feasible processes. 

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2. Cryptic community structure and metabolic interactions among the heritable facultative symbionts of the pea aphid

   https://doi.org/10.1111/jeb.14216  

   Peng, Linyao ; Hoban, Jessica ; Joffe, Jonah ; Smith, Andrew H. ; Carpenter, Melissa ; Marcelis, Tracy ; Patel, Vilas ; Lynn‐Bell, Nicole ; Oliver, Kerry M. ; Russell, Jacob A. ( September 2023 , Journal of Evolutionary Biology)

   Most insects harbour influential, yet non‐essential heritable microbes in their hemocoel. Communities of these symbionts exhibit low diversity. But their frequent multi‐species nature raises intriguing questions on roles for symbiont–symbiont synergies in host adaptation, and on the stability of the symbiont communities, themselves. In this study, we build on knowledge of species‐defined symbiont community structure across US populations of the pea aphid,Acyrthosiphon pisum. Through extensive symbiont genotyping, we show that pea aphids' microbiomes can be more precisely defined at the symbiont strain level, with strain variability shaping five out of nine previously reported co‐infection trends. Field data provide a mixture of evidence for synergistic fitness effects and symbiont hitchhiking, revealing causes and consequences of these co‐infection trends. To test whether within‐host metabolic interactions predict common versus rare strain‐defined communities, we leveraged the high relatedness of our dominant, community‐defined symbiont strains vs. 12 pea aphid‐derived Gammaproteobacteria with sequenced genomes. Genomic inference, using metabolic complementarity indices, revealed high potential for cooperation among one pair of symbionts—Serratia symbioticaandRickettsiella viridis. Applying the expansion network algorithm, through additional use of pea aphid and obligateBuchnerasymbiont genomes,SerratiaandRickettsiellaemerged as the only symbiont community requiring both parties to expand holobiont metabolism. Through their joint expansion of the biotin biosynthesis pathway, these symbionts may span missing gaps, creating a multi‐party mutualism within their nutrient‐limited, phloem‐feeding hosts. Recent, complementary gene inactivation, within the biotin pathways ofSerratiaandRickettsiella, raises further questions on the origins of mutualisms and host–symbiont interdependencies.

    

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3. The reduced genome of a heritable symbiont from an ectoparasitic feather feeding louse

   https://doi.org/10.1186/s12862-021-01840-7  

   Alickovic, Leila ; Johnson, Kevin P. ; Boyd, Bret M. ( December 2021 , BMC Ecology and Evolution)

   Abstract Background Feather feeding lice are abundant and diverse ectoparasites that complete their entire life cycle on an avian host. The principal or sole source of nutrition for these lice is feathers. Feathers appear to lack four amino acids that the lice would require to complete development and reproduce. Several insect groups have acquired heritable and intracellular bacteria that can synthesize metabolites absent in an insect’s diet, allowing insects to feed exclusively on nutrient-poor resources. Multiple species of feather feeding lice have been shown to harbor heritable and intracellular bacteria. We expected that these bacteria augment the louse’s diet with amino acids and facilitated the evolution of these diverse and specialized parasites. Heritable symbionts of insects often have small genomes that contain a minimal set of genes needed to maintain essential cell functions and synthesize metabolites absent in the host insect’s diet. Therefore, we expected the genome of a bacterial endosymbiont in feather lice would be small, but encode pathways for biosynthesis of amino acids. Results We sequenced the genome of a bacterial symbiont from a feather feeding louse ( Columbicola wolffhuegeli ) that parasitizes the Pied Imperial Pigeon ( Ducula bicolor ) and used its genome to predict metabolism of amino acids based on the presence or absence of genes. We found that this bacterial symbiont has a small genome, similar to the genomes of heritable symbionts described in other insect groups. However, we failed to identify many of the genes that we expected would support metabolism of amino acids in the symbiont genome. We also evaluated other gene pathways and features of the highly reduced genome of this symbiotic bacterium. Conclusions Based on the data collected in this study, it does not appear that this bacterial symbiont can synthesize amino acids needed to complement the diet of a feather feeding louse. Our results raise additional questions about the biology of feather chewing lice and the roles of symbiotic bacteria in evolution of diverse avian parasites. 

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4. Droplet superpropulsion in an energetically constrained insect

   https://doi.org/10.1038/s41467-023-36376-5  

   Challita, Elio J. ; Sehgal, Prateek ; Krugner, Rodrigo ; Bhamla, M. Saad ( February 2023 , Nature Communications)

   Food consumption and waste elimination are vital functions for living systems. Although how feeding impacts animal form and function has been studied for more than a century since Darwin, how its obligate partner, excretion, controls and constrains animal behavior, size, and energetics remains largely unexplored. Here we study millimeter-scale sharpshooter insects (Cicadellidae) that feed exclusively on a plant’s xylem sap, a nutrient-deficit source (95% water). To eliminate their high-volume excreta, these insects exploit droplet superpropulsion, a phenomenon in which an elastic projectile can achieve higher velocity than the underlying actuator through temporal tuning. We combine coupled-oscillator models, computational fluid dynamics, and biophysical experiments to show that these insects temporally tune the frequency of their anal stylus to the Rayleigh frequency of their surface tension-dominated elastic drops as a single-shot resonance mechanism. Our model predicts that for these tiny insects, the superpropulsion of droplets is energetically cheaper than forming jets, enabling them to survive on an extreme energy-constrained xylem-sap diet. The principles and limits of superpropulsion outlined here can inform designs of energy-efficient self-cleaning structures and soft engines to generate ballistic motions.

    

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5. Chromosome‐level genome assembly of the aster leafhopper ( Macrosteles quadrilineatus ) reveals the role of environment and microbial symbiosis in shaping pest insect genome evolution

   https://doi.org/10.1111/1755-0998.13919  

   Vasquez, Yumary M. ; Li, Zheng ; Xue, Allen Z. ; Bennett, Gordon M. ( April 2024 , Molecular Ecology Resources)

   Leafhoppers comprise over 20,000 plant‐sap feeding species, many of which are important agricultural pests. Most species rely on two ancestral bacterial symbionts,SulciaandNasuia, for essential nutrition lacking in their phloem and xylem plant sap diets. To understand how pest leafhopper genomes evolve and are shaped by microbial symbioses, we completed a chromosomal‐level assembly of the aster leafhopper's genome (ALF;Macrosteles quadrilineatus). We compared ALF's genome to three other pest leafhoppers,Nephotettix cincticeps,Homalodisca vitripennis, andEmpoasca onukii, which have distinct ecologies and symbiotic relationships. Despite diverging ~155 million years ago, leafhoppers have high levels of chromosomal synteny and gene family conservation. Conserved genes include those involved in plant chemical detoxification, resistance to various insecticides, and defence against environmental stress. Positive selection acting upon these genes further points to ongoing adaptive evolution in response to agricultural environments. In relation to leafhoppers' general dependence on symbionts, species that retain the ancestral symbiont,Sulcia, displayed gene enrichment of metabolic processes in their genomes. Leafhoppers with bothSulciaand its ancient partner,Nasuia, showed genomic enrichment in genes related to microbial population regulation and immune responses. Finally, horizontally transferred genes (HTGs) associated with symbiont support ofSulciaandNasuiaare only observed in leafhoppers that maintain symbionts. In contrast, HTGs involved in non‐symbiotic functions are conserved across all species. The high‐quality ALF genome provides deep insights into how host ecology and symbioses shape genome evolution and a wealth of genetic resources for pest control targets.

    

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