Biosynthesis Of Essential Amino Acids Research Articles

Dynamic response of essential amino acid biosynthesis in Buchnera aphidicola to supplement sub-optimal host nutrition

The endosymbiotic bacterium Buchnera aphidicola allows its host Acyrthosiphon pisum to utilise a nutritionally limited phloem sap diet without significant mortality by providing essential amino acids (EAAs), which it biosynthesises de novo via complex pathways consisting of multiple enzymes. Previous studies have reported how non-essential amino acids (NEAAs) provided by the host are utilised by B. aphidicola, along with how genes within the biosynthetic pathways respond to amino acid deficiency. Although the effect on B. aphidicola gene expression upon the removal of a single EAA and multiple NEAAs from the A. pisum diet has been reported, little is known about the effects of the complete simultaneous removal of multiple EAAs, especially branched-chain amino acids (BCAAs). To investigate this, A. pisum was provided with amino acid deficient diets ilv- (lacking isoleucine, leucine, valine) or thra- (lacking threonine, methionine, lysine). Due to their involvement in the production of several amino acids, the expression of genes ilvC, ilvD (both involved in isoleucine, leucine and valine biosynthesis) and thrA (involved in threonine, methionine and lysine biosynthesis) was analysed and the expression of trpC (involved in tryptophan biosynthesis) was used as a control. Survival was reduced significantly when A. pisum was reared on ilv- or thra- (P < 0.001 and P = 0.000 respectively) compared to optimal artificial diet and was significantly lower on ilv- (P < 0.001) than thra-. This is likely attributed to the EAAs absent from ilv- being required at higher concentrations for aphid growth, than those EAAs absent from thra-. Expression of ilvC and ilvD were upregulated 2.49- and 2.08-fold (respectively) and thrA expression increased 2.35- and 2.12-fold when A. pisum was reared on ilv- and thra- (respectively). The surprisingly large upregulation of thrA when reared on ilv- is likely due to threonine being an intermediate in isoleucine biosynthesis. Expression of trpC was not affected by rearing on either of the two amino acid deficient diets. To our knowledge this study has shown, for the first time, how genes within the biosynthetic pathways of an endosymbiont respond to the simultaneous complete omission of multiple EAAs as well as all three BCAAs (leucine, isoleucine, valine), from the host diet.

Functional genomic characterization reveals the probiotic tendency and safety assessment of Exiguobacterium acetylicum G1–33 isolated from the gut of the hybrid grouper (Epinephelus fuscoguttatus♀ × E. lanceolatus♂)

Probiotics are commonly used as nutritional supplements and antibiotic substitutes and thus are of great commercial interest. The present study was performed to demonstrate the probiotic potential of Exiguobacterium acetylicum G1–33 isolated from the gut of the hybrid grouper (Epinephelus fuscoguttatus♀ × E. lanceolatus♂) by integrating genomics and phenotypic traits. In silico genome analysis confirmed its identity as E. acetylicum. Functional annotation of the genome revealed the existence of molecular markers responsible for the functional properties of potential probiotics, such as resistance to the host’s harsh gastrointestinal environment, adhesion and colonization of the intestinal mucosa, competition and inhibition of pathogens, responses to oxidative stress, and biosynthesis of essential amino acids and vitamins. The genomic analysis was validated by in vitro experiments. Effective toxin-encoding genes were not identified in the G1–33 genome, and the safety of the strain was confirmed by in vitro experiments. These results indicate that E. acetylicum G1–33 exhibits excellent probiotic properties and can become a probiotic with high application value in aquaculture.

A genome-centric view of the role of the Acropora kenti microbiome in coral health and resilience

Microbial diversity has been extensively explored in reef-building corals. However, the functional roles of coral-associated microorganisms remain poorly elucidated. Here, we recover 191 bacterial and 10 archaeal metagenome-assembled genomes (MAGs) from the coral Acropora kenti (formerly A. tenuis) and adjacent seawater, to identify microbial functions and metabolic interactions within the holobiont. We show that 82 MAGs were specific to the A. kenti holobiont, including members of the Pseudomonadota, Bacteroidota, and Desulfobacterota. A. kenti-specific MAGs displayed significant differences in their genomic features and functional potential relative to seawater-specific MAGs, with a higher prevalence of genes involved in host immune system evasion, nitrogen and carbon fixation, and synthesis of five essential B-vitamins. We find a diversity of A. kenti-specific MAGs encode the biosynthesis of essential amino acids, such as tryptophan, histidine, and lysine, which cannot be de novo synthesised by the host or Symbiodiniaceae. Across a water quality gradient spanning 2° of latitude, A. kenti microbial community composition is correlated to increased temperature and dissolved inorganic nitrogen, with corresponding enrichment in molecular chaperones, nitrate reductases, and a heat-shock protein. We reveal mechanisms of A. kenti-microbiome-symbiosis on the Great Barrier Reef, highlighting the interactions underpinning the health of this keystone holobiont.

Metabolism, digestion, and horizontal transfer: potential roles and interaction of symbiotic bacteria in the ladybird beetle Novius pumilus and their prey Icerya aegyptiaca.

In this study, we first time sequenced and analyzed the 16S rRNA gene data of predator ladybird beetles Novius pumilus and globally distributed invasive pest Icerya aegyptiaca at different stages, and combined data with bacterial genome sequences in N. pumilus to explored the taxonomic distribution, alpha and beta diversity, differentially abundant bacteria, co-occurrence network, and putative functions of their microbial community. Our finding revealed that Candidatus Walczuchella, which exhibited a higher abundance in I. aegyptiaca, possessed several genes in essential amino acid biosynthesis and seemed to perform roles in providing nutrients to the host, similar to other obligate symbionts in scale insects. Lactococcus, Serratia, and Pseudomonas, more abundant in N. pumilus, were predicted to have genes related to hydrocarbon, fatty acids, and chitin degradation, which may assist their hosts in digesting the wax shell covering the scale insects. Notably, our result showed that Lactococcus had relatively higher abundances in adults and eggs compared to other stages in N. pumilus, indicating potential vertical transmission. Additionally, we found that Arsenophonus, known to influence sex ratios in whitefly and wasp, may also function in I. aegyptiaca, probably by influencing nutrient metabolism as it similarly had many genes corresponding to vitamin B and essential amino acid biosynthesis. Also, we observed a potential horizontal transfer of Arsenophonus between the scale insect and its predator, with a relatively high abundance in the ladybirds compared to other bacteria from the scale insects.IMPORTANCEThe composition and dynamic changes of microbiome in different developmental stages of ladybird beetles Novius pumilus with its prey Icerya aegyptiaca were detected. We found that Candidatus Walczuchella, abundant in I. aegyptiaca, probably provide nutrients to their host based on their amino acid biosynthesis-related genes. Abundant symbionts in N. pumilus, including Lactococcus, Serratia, and Pseudophonus, may help the host digest the scale insects with their hydrocarbon, fatty acid, and chitin degrading-related genes. A key endosymbiont Arsenophonus may play potential roles in the nutrient metabolisms and sex determination in I. aegyptiaca, and is possibly transferred from the scale insect to the predator.

In silico prediction of candidate gene targets for the management of African cassava whitefly (Bemisia tabaci, SSA1-SG1), a key vector of viruses causing cassava brown streak disease.

Whiteflies (Bemisia tabaci sensu lato) have a wide host range and are globally important agricultural pests. In Sub-Saharan Africa, they vector viruses that cause two ongoing disease epidemics: cassava brown streak disease and cassava mosaic virus disease. These two diseases threaten food security for more than 800 million people in Sub-Saharan Africa. Efforts are ongoing to identify target genes for the development of novel management options against the whitefly populations that vector these devastating viral diseases affecting cassava production in Sub-Saharan Africa. This study aimed to identify genes that mediate osmoregulation and symbiosis functions within cassava whitefly gut and bacteriocytes and evaluate their potential as key gene targets for novel whitefly control strategies. The gene expression profiles of dissected guts, bacteriocytes and whole bodies were compared by RNAseq analysis to identify genes with significantly enriched expression in the gut and bacteriocytes. Phylogenetic analyses identified three candidate osmoregulation gene targets: two α-glucosidases, SUC 1 and SUC 2 with predicted function in sugar transformations that reduce osmotic pressure in the gut; and a water-specific aquaporin (AQP1) mediating water cycling from the distal to the proximal end of the gut. Expression of the genes in the gut was enriched 23.67-, 26.54- and 22.30-fold, respectively. Genome-wide metabolic reconstruction coupled with constraint-based modeling revealed four genes (argH, lysA, BCAT & dapB) within the bacteriocytes as potential targets for the management of cassava whiteflies. These genes were selected based on their role and essentiality within the different essential amino acid biosynthesis pathways. A demonstration of candidate osmoregulation and symbiosis gene targets in other species of the Bemisia tabaci species complex that are orthologs of the empirically validated osmoregulation genes highlights the latter as promising gene targets for the control of cassava whitefly pests by in planta RNA interference.

Taxonomic and functional shifts of gut microbiome in immunoglobulin A vasculitis children and their mothers.

To examine the gut microbiota characteristics in children with immunoglobulin A vasculitis and their interrelationships with the host, while evaluate the vertical inheritance of microbiota in the development and progression of IgA vasculitis. This study investigated the gut microbiome of 127 IgA vasculitis mother-child pairs and 62 matched healthy mother-child pairs, and compared the gut microbial composition of different groups. The pathway enrichment analysis evaluated potential gut microbiome-mediated pathways involved in the pathophysiology of IgA vasculitis. The Spearman correlation analysis illustrated the relationships between clinical variables and bacterial biomarkers. This study identified distinct intestinal microbiome in IgA vasculitis children compared to healthy children, and further pointed out the association in gut microbiota between IgA vasculitis children's and their mother's. The relative abundance of Megamonas and Lactobacillus in IgAV children was positively correlated with that in their mothers. The pathway enrichment analysis found microbial biosynthesis of vitamins and essential amino acids was upregulated in children with IgA vasculitis. Correlation analysis showed bacterial biomarkers were correlated with indicators of blood coagulation. Children with IgA vasculitis have unique bacterial biomarkers and may affect coagulation function, and their gut microbiome was closely associated with that of their mothers. The observed association in gut microbiota between IgA vasculitis children and their mothers suggested a potential intergenerational influence of the maternal microbiota on the development or progression of IgA vasculitis in children.

A mitochondrial-targeted activity-based sensing probe for ratiometric imaging of formaldehyde reveals key regulators of the mitochondrial one-carbon pool.

Formaldehyde (FA) is both a highly reactive environmental genotoxin and an endogenously produced metabolite that functions as a signaling molecule and one-carbon (1C) store to regulate 1C metabolism and epigenetics in the cell. Owing to its signal-stress duality, cells have evolved multiple clearance mechanisms to maintain FA homeostasis, acting to avoid the established genotoxicity of FA while also redirecting FA-derived carbon units into the biosynthesis of essential nucleobases and amino acids. The highly compartmentalized nature of FA exposure, production, and regulation motivates the development of chemical tools that enable monitoring of transient FA fluxes with subcellular resolution. Here we report a mitochondrial-targeted, activity-based sensing probe for ratiometric FA detection, MitoRFAP-2, and apply this reagent to monitor endogenous mitochondrial sources and sinks of this 1C unit. We establish the utility of subcellular localization by showing that MitoRFAP-2 is sensitive enough to detect changes in mitochondrial FA pools with genetic and pharmacological modulation of enzymes involved in 1C and amino acid metabolism, including the pervasive, less active genetic mutant aldehyde dehydrogenase 2*2 (ALDH2*2), where previous, non-targeted versions of FA sensors are not. Finally, we used MitoRFAP-2 to comparatively profile basal levels of FA across a panel of breast cancer cell lines, finding that FA-dependent fluorescence correlates with expression levels of enzymes involved in 1C metabolism. By showcasing the ability of MitoRFAP-2 to identify new information on mitochondrial FA homeostasis, this work provides a starting point for the design of a broader range of chemical probes for detecting physiologically important aldehydes with subcellular resolution and a useful reagent for further studies of 1C biology.

Nutrient supplementation by genome-eroded Burkholderia symbionts of scale insects

Hemipterans are known as hosts to bacterial or fungal symbionts that supplement their unbalanced diet with essential nutrients. Among them, scale insects (Coccomorpha) are characterized by a particularly large diversity of symbiotic systems. Here, using microscopic and genomic approaches, we functionally characterized the symbionts of two scale insects belonging to the Eriococcidae family, Acanthococcus aceris and Gossyparia spuria. These species host Burkholderia bacteria that are localized in the cytoplasm of the fat body cells. Metagenome sequencing revealed very similar and highly reduced genomes (<900KBp) with a low GC content (~38%), making them the smallest and most AT-biased Burkholderia genomes yet sequenced. In their eroded genomes, both symbionts retain biosynthetic pathways for the essential amino acids leucine, isoleucine, valine, threonine, lysine, arginine, histidine, phenylalanine, and precursors for the semi-essential amino acid tyrosine, as well as the cobalamin-dependent methionine synthase MetH. A tryptophan biosynthesis pathway is conserved in the symbiont of G. spuria, but appeared pseudogenized in A. aceris, suggesting differential availability of tryptophan in the two host species’ diets. In addition to the pathways for essential amino acid biosynthesis, both symbionts maintain biosynthetic pathways for multiple cofactors, including riboflavin, cobalamin, thiamine, and folate. The localization of Burkholderia symbionts and their genome traits indicate that the symbiosis between Burkholderia and eriococcids is younger than other hemipteran symbioses, but is functionally convergent. Our results add to the emerging picture of dynamic symbiont replacements in sap-sucking Hemiptera and highlight Burkholderia as widespread and versatile intra- and extracellular symbionts of animals, plants, and fungi.

Comparative analysis of muscle profiles in silky fowl and white Leghorn Chicken: Insights from multi-omics and experimental approaches

Silky fowl is a highly valued breed of chicken with both culinary and medicinal significance. We conducted a comprehensive and integrated proteomic, transcriptomic, and metabolomic analysis to compare silky fowl meat (SFM) with Leghorn chicken meat (LCM). In SFM, 262 differentially expressed proteins (48 up-regulated, 214 down-regulated), 2577 differentially expressed genes (949 up-regulated, 1628 down-regulated), and 198 differentially abundant metabolites (52 up-regulated, 146 down-regulated) were identified. Our studies underscore the notable involvement of SFM in essential amino acid biosynthesis, accompanied by the up-regulation of genes associated with melanogenesis. Several genes exhibited consistent expression patterns with the results derived from real-time quantitative polymerase chain reaction (RT-qPCR). SFM contained higher levels of flavor-enhancing factors (glutathione, glycine, and serine) relative to LCM. Moreover, it harbored the presence of pharmacologically significant metabolites alongside up-regulated genes related to vitamin E metabolism. These multi-omics differences provide a comprehensive framework that lays the groundwork for future investigations into the biological, nutritional, and pharmacological applications of silky fowl.

Crystal Structure of Aspartate Semialdehyde Dehydrogenase from Porphyromonas gingivalis

Aspartate semialdehyde dehydrogenase (ASADH) catalyzes the biosynthesis of several essential amino acids, including lysine, methionine, and threonine, and bacterial cell components. Thus, ASADH is a crucial target for developing new antimicrobial agents that can potentially disrupt the biosynthesis of essential amino acids, thereby inhibiting the growth of pathogens. Herein, the crystal structures of ASADH obtained from Porphyromonas gingivalis (PgASADH, UniProtKB code A0A1R4DY25) were determined in apo- and adenosine-2′-5′-diphosphate (2′,5′-ADP)-bound complex forms at a resolution of 1.73 Å. The apo- and 2′,5′-ADP-complexed crystals of PgASADH belonged to the space groups of I212121 and C2221, respectively. Analytical size-exclusion chromatography showed a stable PgASADH dimer in a solution. Clustering analysis and structural comparison studies performed on PgASADH and previously known ASADHs revealed that ASADHs, including PgASADH, can be classified into three types depending on sequential and structural differences at the α-helical subdomain region. These findings provide valuable insights into developing structure-based species-specific new antibacterial drugs against the oral pathogen P. gingivalis.

The unique gut microbiome of giant pandas involved in protein metabolism contributes to the host’s dietary adaption to bamboo

BackgroundThe gut microbiota of the giant panda (Ailuropoda melanoleuca), a global symbol of conservation, are believed to be involved in the host’s dietary switch to a fibrous bamboo diet. However, their exact roles are still largely unknown.ResultsIn this study, we first comprehensively analyzed a large number of gut metagenomes giant pandas (n = 322), including 98 pandas sequenced in this study with deep sequencing (Illumina) and third-generation sequencing (nanopore). We reconstructed 408 metagenome-assembled genomes (MAGs), and 148 of which (36.27%) were near complete. The most abundant MAG was classified as Streptococcus alactolyticus. A pairwise comparison of the metagenomes and meta-transcriptomes in 14 feces revealed genes involved in carbohydrate metabolism were lower, but those involved in protein metabolism were greater in abundance and expression in giant pandas compared to those in herbivores and omnivores. Of note, S. alactolyticus was positively correlated to the KEGG modules of essential amino-acid biosynthesis. After being isolated from pandas and gavaged to mice, S. alactolyticus significantly increased the relative abundance of essential amino acids in mice jejunum.ConclusionsThe study highlights the unique protein metabolic profiles in the giant panda’s gut microbiome. The findings suggest that S. alactolyticus is an important player in the gut microbiota that contributes to the giant panda’s dietary adaptation by more involvement in protein rather than carbohydrate metabolism.4zh5Ny6QmwZifdgaSCFmHgVideo

Metagenome-assembled genomes for biomarkers of bio-functionalities in Laal dahi, an Indian ethnic fermented milk product

Laal dahi is a sweetened and soft pudding-like fermented milk product of the Eastern regions of India, which has not been studied for its microbial community structures and health promoting functionality in terms of ‘omics’ approaches. We applied metagenomic and metagenomes-assembled genomes (MAGs) tools to decipher the biomarkers for genes encoding for different health promoting functionalities in laal dahi. Abundance of bacterial domains was observed with negligible presence of eukaryotes and viruses. Bacillota was the most abundant phylum with different bacterial species viz., Enterococcus italicus, Lactococcus raffinolactis, Lactobacillus helveticus, Bifidobacterium mongoliense, Hafnia alvei, Lactococcus lactis, Acetobacter okinawensis, Streptococcus thermophilus, Thermus thermophilus, Leuconostoc citreum, Leuconostoc pseudomesenteroides, Acetobacter orientalis, Lactobacillus gallinarum, Lactococcus chungangensis and Lactobacillus delbrueckii. Comparison of laal dahi microbiome with that of similar fermented milk products was also carried out after retrieving the metagenomic datasets from public databases. Significant abundance of Lb. helveticus, E. italicus, Lc. raffinolactis and Lc. lactis in laal dahi. Interestingly, Bifidobacterium mongoliense, Lb. gallinarum, Lc. chungangensis and Acetobacter okinawensis were only detected in laal dahi but Streptococcus infantarius, Lacticaseibacillus rhamnosus and Lb. johnsonii were absent. Reconstruction of putative single environment-specific genomes from metagenomes in addition to subsampling of the abundant species resulted in five high-quality MAGs identified as Lactobacillus delbrueckii, Lactobacillus helveticus, Lactococcus chungangensis, Lactococcus lactis and Streptococcus thermophilus. All MAGs showed the presence of various genes with several putative functions corresponding to different probiotic and prebiotic functions, short-chain fatty acids production, immunomodulation, antitumor genes, essential amino acid and vitamin biosynthesis. Genes for γ-Aminobutyric acid (GABA) production were only detected in MAG of Lactococcus lactis. Gene clusters for secondary metabolites (antimicrobial peptides) were detected in all MAGs except Lc. chungangensis. Additionally, detection of clustered regularly interspaced short palindromic repeats (CRISPR)-associated (Cas) elements was observed only in Lactobacillus delbrueckii and Streptococcus thermophilus. Annotation of several genes with potential health beneficial properties in all five MAGs may support the need to explore the culturability of these MAGs for future use in controlled fermentation of functional dairy products.

Citrus pomace co-fermentation improved its protein and amino acids by Bacillus amyloliquefaciens and Candida utilis

High-value utilization of citrus pomace (CP) is of great significance to the sustainable development of the citrus industry. Co-fermentation of CP was investigated using the autochthonous probiotic bacteria, Bacillus amyloliquefaciens BF2 and Candida utilis GIM 2.9. Crude protein, soluble protein, and small peptide content of CP increased by 40.01%, 923.53%, and 626.67% after fermentation, respectively. Essential amino acids increased from 28.51% to 47.04% of total free amino acids. Leucine level reached 4.04 mg/g DW, increasing by 1293.10%, and produced histidine, valine, cysteine, and tyrosine. Scanning electron microscopy results showed that BF2 and GIM 2.9 could effectively degrade and utilize the CP fibers. Enzymolysis co-fermentation (ECF) system secreted higher levels of non-starch polysaccharide enzymes and proteases than co-fermentation (CF) system. Acid protease activity was positively correlated with soluble protein and free amino acid contents. BF2 possessed 63 essential amino acid-biosynthesis genes, nearly twice as many as that of non-essential amino acids. Eight key enzyme genes were identified for leucine biosynthesis in the BF2 genome, suggesting that BF2 played a key role in leucine biosynthesis. Our results provide a potential method for the high-value utilization of CP as a protein feed additive or dietary supplement.

Alteration of gut microbiota in wild-borne long-tailed macaques after 1-year being housed in hygienic captivity

The wild-born long-tailed macaques (Macaca fascicularis) were recently recruited and used as breeders for the National Primate Research Center of Thailand, Chulalongkorn University (NPRCT-CU), and changes in their in-depth gut microbiota profiles were investigated. The Oxford Nanopore Technology (ONT) was used to explore full-length 16S rDNA sequences of gut microbiota in animals once captured in their natural habitat and 1-year following translocation and housing in a hygienic environment at NPRCT-CU. Our findings show that the gut microbiota of macaques after 1 year of hygienic housing and programmed diets feeding was altered and reshaped. The prevalent gut bacteria such as Prevotella copri and Faecalibacterium prausnitzii were enriched after translocation, causing the lower alpha diversity. The correlation analysis revealed that Prevotella copri, Phascolarctobacterium succinatutens, and Prevotella stercorea, showed a positive correlation with each other. Significantly enriched pathways in the macaques after translocation included biosynthesis of essential amino acids, fatty acids, polyamine and butanoate. The effects of microbiota change could help macaques to harvest the energy from programmed diets and adapt their gut metabolism. The novel probiotics and microbiota engineering approach could be further developed based on the current findings and should be helpful for captive animal health care management.

Co-diversification of an intestinal Mycoplasma and its salmonid host

Understanding the evolutionary relationships between a host and its intestinal resident bacteria can transform how we understand adaptive phenotypic traits. The interplay between hosts and their resident bacteria inevitably affects the intestinal environment and, thereby, the living conditions of both the host and the microbiota. Thereby this co-existence likely influences the fitness of both bacteria and host. Whether this co-existence leads to evolutionary co-diversification in animals is largely unexplored, mainly due to the complexity of the environment and microbial communities and the often low host selection. We present the gut metagenome from wild Atlantic salmon (Salmo salar), a new wild organism model with an intestinal microbiota of low complexity and a well-described population structure, making it well-suited for investigating co-evolution. Our data reveal a strong host selection of a core gut microbiota dominated by a single Mycoplasma species. We found a clear co-diversification between the population structure of Atlantic salmon and nucleotide variability of the intestinal Mycoplasma populations conforming to expectations from co-evolution between host and resident bacteria. Our results show that the stable microbiota of Atlantic salmon has evolved with its salmonid host populations while potentially providing adaptive traits to the salmon host populations, including defence mechanisms, biosynthesis of essential amino acids, and metabolism of B vitamins. We highlight Atlantic salmon as a novel model for studying co-evolution between vertebrate hosts and their resident bacteria.

The Role of luxS in Campylobacter jejuni Beyond Intercellular Signaling.

The full role of the luxS gene in the biological processes, such as essential amino acid synthesis, nitrogen and pyruvate metabolism, and flagellar assembly, of Campylobacter jejuni has not been clearly described to date. Therefore, in this study, we used a comprehensive approach at the cellular and molecular levels, including transcriptomics and proteomics, to investigate the key role of the luxS gene and compared C. jejuni 11168ΔluxS (luxS mutant) and C. jejuni NCTC 11168 (wild type) strains. Transcriptomic analysis of the luxS mutant grown under optimal conditions revealed upregulation of luxS mutant metabolic pathways when normalized to wild type, including oxidative phosphorylation, carbon metabolism, citrate cycle, biosynthesis of secondary metabolites, and biosynthesis of various essential amino acids. Interestingly, induction of these metabolic pathways was also confirmed by proteomic analysis, indicating their important role in energy production and the growth of C. jejuni. In addition, genes important for the stress response of C. jejuni, including nutrient starvation and oxidative stress, were upregulated. This was also evident in the better survival of the luxS mutant under starvation conditions than the wild type. At the molecular level, we confirmed that metabolic pathways were upregulated under optimal conditions in the luxS mutant, including those important for the biosynthesis of several essential amino acids. This also modulated the utilization of various carbon and nitrogen sources, as determined by Biolog phenotype microarray analysis. In summary, transcriptomic and proteomic analysis revealed key biological differences in tricarboxylic acid (TCA) cycle, pyruvate, nitrogen, and thiamine metabolism as well as lipopolysaccharide biosynthesis in the luxS mutant. IMPORTANCE Campylobacter jejuni is the world's leading foodborne bacterial pathogen of gastrointestinal disease in humans. C. jejuni is a fastidious but widespread organism and the most frequently reported zoonotic pathogen in the European Union since 2005. This led us to believe that C. jejuni, which is highly sensitive to stress factors (starvation and oxygen concentration) and has a low growth rate, benefits significantly from the luxS gene. The role of this gene in the life cycle of C. jejuni is well known, and the expression of luxS regulates many phenotypes, including motility, biofilm formation, host colonization, virulence, autoagglutination, cellular adherence and invasion, oxidative stress, and chemotaxis. Surprisingly, this study confirmed for the first time that the deletion of the luxS gene strongly affects the central metabolic pathway of C. jejuni, which improves its survival, showing its role beyond the intercellular signaling system.

Cellular and potential molecular mechanisms underlying transovarial transmission of the obligate symbiont Sulcia in cicadas.

Vertical transmission of symbionts in insects is critical to persistence of symbioses across host generations. The key time point and related cellular/molecular mechanisms underlying the transmission in most insects remain unclear. Here, we reveal that in the bacteriome-endosymbiont system of the cicada Meimuna mongolica, the obligate symbiont Candidatus Sulcia muelleri (hereafter Sulcia) proliferates and migrates to the ovaries mainly after the adult emergence of cicadas. Sulcia cells swell to approximately twice their previous size with the outer membrane changed to be more irregular during this process. Almost all the Sulcia genes involved in biosynthesis of essential amino acids, heat shock protein, energy metabolism, DNA replication and repair and protein export were highly expressed in all life stages of cicadas. Among which, genes involved in DNA replication and synthesis of leucine and arginine were upregulated in the newly emerged adults relative to fifth-instar nymphs. Signal transduction is the pronounced function exhibited in both Sulcia and the cicada bacteriomes in newly emerged adults. The results suggest host sensing of arginine and leucine integrate Sulcia's output of host-EAAs into mTORC1 signalling. This study highlights the importance of signalling pathways in regulating the host/symbiont interaction and symbiont transmission in sap-feeding auchenorrhynchous insects.

Dietary Supplementation of a Commercial Prebiotic, Probiotic and Their Combination Affected Growth Performance and Transient Intestinal Microbiota of Red Drum (Sciaenops ocellatus L.).

In the present study, the potential synergism between beneficial lactic acid bacteria (Pediococcus acidilactici) contained in a probiotic and a mixture of fermentable complex carbohydrates and autolyzed brewer's yeast (or prebiotic) were explored in red drum. Four experimental diets were formulated from practical ingredients, and the basal diet was supplemented with either probiotic, prebiotic, or both supplements. Red drum juveniles (~5.5 g) were offered the four experimental diets for 56 days, and at the end of the feeding trial fish fed diets supplemented with probiotic had significantly better weight gain than those fed the non-supplemented diets, and higher protein content in their whole-body composition. Transient intestinal microbiome alpha and beta diversity were significantly affected by the dietary treatments. Interestingly, a higher relative abundance of the lactic acid genus Pediococcus was observed for fish fed diets supplemented with the prebiotic. A higher relative abundance was also observed for the predicted functions of the microbial metagenome, and many of these pathways involved the biosynthesis of essential amino acids, vitamins, and nucleotides. Even though no potential synergistic effect was observed, the individual inclusion of these prebiotic and probiotic supplements positively affected the intestinal health and growth performance of red drum, respectively.

De novo metatranscriptomic exploration of gene function in the millipede holobiont

Invertebrate–microbial associations are widespread in the biosphere and are often related to the function of novel genes, fitness advantages, and even speciation events. Despite ~ 13,000 species of millipedes identified across the world, millipedes and their gut microbiota are markedly understudied compared to other arthropods. Exploring the contribution of individual host-associated microbes is often challenging as many are uncultivable. In this study, we conducted metatranscriptomic profiling of different body segments of a millipede at the holobiont level. This is the first reported transcriptome assembly of a tropical millipede Telodeinopus aoutii (Demange, 1971), as well as the first study on any Myriapoda holobiont. High-throughput RNA sequencing revealed that Telodeinopus aoutii contained > 90% of the core Arthropoda genes. Proteobacteria, Bacteroidetes, Firmicutes, and Euryarchaeota represented dominant and functionally active phyla in the millipede gut, among which 97% of Bacteroidetes and 98% of Firmicutes were present exclusively in the hindgut. A total of 37,831 predicted protein-coding genes of millipede holobiont belonged to six enzyme classes. Around 35% of these proteins were produced by microbiota in the hindgut and 21% by the host in the midgut. Our results indicated that although major metabolic pathways operate at the holobiont level, the involvement of some host and microbial genes are mutually exclusive and microbes predominantly contribute to essential amino acid biosynthesis, short-chain fatty acid metabolism, and fermentation.

Resurrecting essential amino acid biosynthesis in mammalian cells.

Major genomic deletions in independent eukaryotic lineages have led to repeated ancestral loss of biosynthesis pathways for nine of the twenty canonical amino acids. While the evolutionary forces driving these polyphyletic deletion events are not well understood, the consequence is that extant metazoans are unable to produce nine essential amino acids (EAAs). Previous studies have highlighted that EAA biosynthesis tends to be more energetically costly, raising the possibility that these pathways were lost from organisms with access to abundant EAAs. It is unclear whether present-day metazoans can reaccept these pathways to resurrect biosynthetic capabilities that were lost long ago or whether evolution has rendered EAA pathways incompatible with metazoan metabolism. Here, we report progress on a large-scale synthetic genomics effort to reestablish EAA biosynthetic functionality in mammalian cells. We designed codon-optimized biosynthesis pathways based on genes mined from Escherichia coli. These pathways were de novo synthesized in 3 kilobase chunks, assembled in yeasto and genomically integrated into a Chinese hamster ovary (CHO) cell line. One synthetic pathway produced valine at a sufficient level for cell viability and proliferation. 13C-tracing verified de novo biosynthesis of valine and further revealed build-up of pathway intermediate 2,3-dihydroxy-3-isovalerate. Increasing the dosage of downstream ilvD boosted pathway performance and allowed for long-term propagation of second-generation cells in valine-free medium at 3.2 days per doubling. This work demonstrates that mammalian metabolism is amenable to restoration of ancient core pathways, paving a path for genome-scale efforts to synthetically restore metabolic functions to the metazoan lineage.