Lactiplantibacillus pentosus
9D3, a prominent gamma-aminobutyric acid (GABA)-producing bacteria isolated from Thai pickled weed was characterized for its safety and probiotic properties
via
...whole-genome analysis and
in vitro
testing. The whole-genome sequence of
L. pentosus
9D3 was determined using a hybrid-sequencing approach, combining PacBio and Illumina technologies. A 3.81-Mbp genome of
L. pentosus
9D3 consisting of one 3.65-Mbp chromosome and six plasmids (1.9–71.9 Kbp) was identified with an estimated GC content of 46.09% and 3,456 predicted genes. The strain was confirmed to be
Lactiplantibacillus pentosus
according to the high average nucleotide identity value of >95% and digital DNA–DNA hybridization scores of >70% to the
L. pentosus
type strain. Comparative genome analysis with other
L. pentosus
strains showed that the GABA-producing capability was specific to the strain 9D3. Genes related to GABA biosynthesis and transport were identified on a plasmid, pLPE-70K, indicating the acquired nature of this property. The safety of
L. pentosus
9D3 was demonstrated through the lack of genes related to the production of toxins, biogenic amines, and antimicrobial drugs. Although the strain exhibited resistance to ampicillin and chloramphenicol, none of the antimicrobial resistance (AMR) genes were associated with mobile elements, i.e., plasmids and prophages. Therefore, the strain is considered to have low risk of transferring the AMR genes to other, potentially pathogenic bacteria. In addition,
L. pentosus
9D3 showed good survivability in the gastrointestinal tract environment and was able to adhere to the intestinal cell
in vitro.
Therefore,
L. pentosus
9D3 is concluded to be safe, with the potential to be used as a probiotic, exerting its health benefit through GABA production in the food system. The GABA-producing capability of the strain
in vivo
is the subject of further investigation.
Dissecting the cellular metabolism of
Cordyceps militaris
is important for the efficient production of bioactive compounds of a target with medicinal and industrial applications. However, the ...metabolic functions during developmental stages in this fungus at a system level are still unexplored. In this study, we aimed to reveal the metabolic functions and regulation of
C. militaris
TBRC6039 relevant to its developmental stages, including mycelial growth (MY) and fruiting body (FB) stages through integrative transcriptome analysis. The transcriptome analysis showed that 9,256 genes of
C. militaris
were expressed in both stages. Of them, 1,877 genes, residing primarily in the cell division cycle and amino acid, carbohydrate, and lipid metabolisms, exhibited significant differences in transcript levels between the MY and FB stages. Through integration with genome-scale networks analysis, the unique reporter metabolites (
e.g.
, α-D-glucose, β-D-glucose, D-galactose, triacylglycerol, and diacylglycerol) and key regulators (
e.g.
, AtfA, Atf2, and Yap1 transcription factors) were identified in
C. militaris
when grown at FB stage, linking to up-regulation of the metabolic genes involved in galactose and polysaccharide metabolisms, as well as glycerolipid and glycerophospholipid biosynthesis. Moreover, the high cordycepin content is related to the upregulated genes in lipid metabolism during the FB stage. Our findings suggest that the transcriptional regulation of these metabolic pathways played a crucial role in specific developmental stages of
C. militaris
. This study serves for cultivation process improvement for overproduction of valuable metabolites in
C. militaris
through an emerging systems and synthetic biology approach.
Display omitted
Cordyceps militaris is an entomopathogenic fungus which is often used in Asia as a traditional medicine developed from age-old wisdom. Presently, cordycepin from C. militaris is a ...great interest in medicinal applications. However, cellular growth of C. militaris and the association with cordycepin production remain poorly understood. To explore the metabolism of C. militaris as potential cell factories in medical and biotechnology applications, this study developed a high-quality genome-scale metabolic model of C. militaris, iNR1329, based on its genomic content and physiological data. The model included a total of 1329 genes, 1821 biochemical reactions, and 1171 metabolites among 4 different cellular compartments. Its in silico growth simulation results agreed well with experimental data on different carbon sources. iNR1329 was further used for optimizing the growth and cordycepin overproduction using a novel approach, POPCORN, for rational design of synthetic media. In addition to the high-quality GEM iNR1329, the presented POPCORN approach was successfully used to rationally design an optimal synthetic medium with C:N ratio of 8:1 for enhancing 3.5-fold increase in cordycepin production. This study thus provides a novel insight into C. militaris physiology and highlights a potential GEM-driven method for synthetic media design and metabolic engineering application. The iNR1329 and the POPCORN approach are available at the GitHub repository: https://github.com/sysbiomics/Cordyceps_militaris-GEM.
Cordyceps militaris is currently exploited for commercial production of specialty products as its biomass constituents are enriched in bioactive compounds, such as cordycepin. The rational process ...development is important for economically feasible production of high quality bioproducts. Light is an abiotic factor affecting the cultivation process of this entomopathogenic fungus, particularly in its carotenoid formation. To uncover the cell response to light exposure, this study aimed to systematically investigate the metabolic responses of C. militaris strain TBRC6039 using integrative genome-wide transcriptome and genome-scale metabolic network (GSMN)-driven analysis. The genome-wide transcriptome analysis showed 8747 expressed genes in the glucose and sucrose cultures grown under light-programming and dark conditions. Of them, 689 differentially expressed genes were significant in response to the light-programming exposure. Through integration with the GSMN-driven analysis using the improved network (iRT1467), the reporter metabolites, e.g., adenosine-5′-monophosphate (AMP) and 2-oxoglutarate, were identified when cultivated under the carotenoid-producing condition controlled by light-programming exposure, linking to up-regulations of the metabolic genes involved in glyoxalase system, as well as cordycepin and carotenoid biosynthesis. These results indicated that C. militaris had a metabolic control in acclimatization to light exposure through transcriptional co-regulation, which supported the cell growth and cordycepin production in addition to the accumulation of carotenoid as a photo-protective bio-pigment. This study provides a perspective in manipulating the metabolic fluxes towards the target metabolites through either genetic or physiological approaches.
KUB-AC5 displays the hallmark features of probiotic properties for food and feed industries. Optimization of cultivation condition for the industrial production is important to reach cell ...concentration and cost reduction. Considering the strain-specific growth physiology, metabolic capability, and essential nutrients of
KUB-AC5, the genome-scale metabolic model (GSMM) of
KUB-AC5 was developed. Hereby, the GSMM of
TN656 was successfully constructed which contained 656 genes, 831 metabolites, and 953 metabolic reactions. The
TN656 model could show a metabolic capability under various carbon sources and guide potentially 14 essential single nutrients (e.g., vitamin B complex and amino acids) and 2 essential double nutrients (pairwise glutamine-glutamate and asparagine-aspartate) for
KUB-AC5 growth through single and double omission analysis. Promisingly, the
TN656 model was further integrated with transcriptome data suggesting that putative metabolic routes as preferable paths e.g., sucrose uptake, nucleotide biosynthesis, urea cycle, and glutamine transporter for
KUB-AC5 growth. The developed GSMM offers a powerful tool for multi-level omics analysis, enabling probiotic strain optimization for biomass overproduction on an industrial scale.
The gut microbiome plays a major role in the maintenance of human health. Characterizing the taxonomy and metabolic functions of the human gut microbiome is necessary for enhancing health. Here, we ...analyzed the metagenomic sequencing, assembly and construction of a meta-gene catalogue of the human gut microbiome with the overall aim of investigating the taxonomy and metabolic functions of the gut microbiome in Thai adults. As a result, the integrative analysis of 16S rRNA gene and whole metagenome shotgun (WMGS) sequencing data revealed that the dominant gut bacterial families were
and
of the Firmicutes phylum. Consistently, across 3.8 million (M) genes annotated from 163.5 gigabases (Gb) of WMGS sequencing data, a significant number of genes associated with carbohydrate metabolism of the dominant bacterial families were identified. Further identification of bacterial community-wide metabolic functions promisingly highlighted the importance of
and
involvement in central carbon metabolism, sugar utilization and metabolism towards butyrate biosynthesis. This work presents an initial study of shotgun metagenomics in a Thai population-based cohort in a developing Southeast Asian country.
WJ11 is a lipid-producing strain with industrial potential. A holistic approach using gene manipulation and bioprocessing development has improved lipid production and the strain's economic ...viability. However, the systematic regulation of lipid accumulation and carotenoid biosynthesis in
remains unknown. To dissect the metabolic mechanism underlying lipid and carotenoid biosynthesis, transcriptome analysis and reporter metabolites identification were implemented between the wild-type (WJ11) and
WJ11 strains of
. As a result, transcriptome analysis revealed 10,287 expressed genes, with 657 differentially expressed genes (DEGs) primarily involved in amino acid, carbohydrate, and energy metabolism. Integration with a genome-scale metabolic model (GSMM) identified reporter metabolites in the
WJ11 strain, highlighting metabolic pathways crucial for amino acid, energy, and nitrogen metabolism. Notably, the downregulation of genes associated with carotenoid biosynthesis and acetyl-CoA generation suggests a coordinated relationship between the carotenoid and fatty acid biosynthesis pathways. Despite disruptions in the carotenoid pathway, lipid production remains stagnant due to reduced acetyl-CoA availability, emphasizing the intricate metabolic interplay. These findings provide insights into the coordinated relationship between carotenoid and fatty acid biosynthesis in
that are valuable in applied research to design optimized strains for producing desired bioproducts through emerging technology.
The genome-scale metabolic model (GSMM) of
provides a comprehensive basis of carbon assimilation for cell growth and metabolite production. However, the model with a simple mass balance concept shows ...limited capability to probe the metabolic responses of
under light exposure. This study, therefore, employed the transcriptome-integrated GSMM approach to extend the investigation of
's metabolism under light conditions. Through the gene inactivity moderated by metabolism and expression (GIMME) framework, the
PS1474-
GSMM model was furnished with the transcriptome data, thus providing a simulation that described reasonably well the metabolic responses underlying the phenotypic observation of
under the particular light conditions. The
PS1474-
GSMM obviously showed an improved prediction of metabolic fluxes in correlation with the expressed genes involved in the cordycepin and carotenoid biosynthetic pathways under the sucrose culturing conditions. Further analysis of reporter metabolites suggested that the central carbon, purine, and fatty acid metabolisms towards carotenoid biosynthesis were the predominant metabolic processes responsible in light conditions. This finding highlights the key responsive processes enabling the acclimatization of
metabolism in varying light conditions. This study provides a valuable perspective on manipulating metabolic genes and fluxes towards the target metabolite production of
.
Gut microbiome plays an essential role in host health, and there is interest in utilizing diet to modulate the composition and function of microbial communities. Copra meal hydrolysate (CMH) is ...commonly used as a natural additive to enhance health. However, the gut microbiome is largely unknown at species level and is associated with metabolic routes involving short-chain fatty acids (SCFAs). In this study, we aimed to analyze, using integrative metagenomics, the predominant species and metabolic routes involved in SCFAs production in the human gut microbiome after treatment with CMH. The effect of CMH treatment on the Thai gut microbiome was demonstrated using 16S rRNA genes with whole-metagenome shotgun (WMGS) sequencing technology. Accordingly, these results revealed that CMH has potentially beneficial effects on the gut microbiome. Twelve predominant bacterial species, as well as their potential metabolic routes, were involved in cooperative microbiome networks under sugar utilization (e.g., glucose, mannose, or xylose) and energy supply (e.g., NADH and ATP) in relation to SCFAs biosynthesis. These findings suggest that CMH may be used as a potential prebiotic diet for modulating and maintaining the gut microbiome. To our knowledge, this is the first study to reveal the predominant bacterial species and metabolic routes in the Thai gut microbiome after treatment with potential prebiotics.
Beyond comparative genomics, we identified 85 sugar transporter genes in
, clustering into nine subfamilies as sequence- and phylogenetic-based functional classification, presuming the versatile ...capability of the fungal growths on a range of sugars. Further analysis of the global gene expression patterns of
showed 123 genes were significantly expressed across the sucrose, glucose, and xylose cultures. The sugar transporters specific for pentose were then identified by gene-set enrichment analysis. Of them, the putative pentose transporter, CCM_06358 gene, was highest expressed in the xylose culture, and its functional role in xylose transport was discovered by the analysis of conserved structural motifs. In addition, a battery of molecular modeling methods, including homology modeling, transport pathway analysis, residue interaction network combined with molecular mechanics Poisson-Boltzmann surface area simulation (MM-PBSA), was implemented for probing the structure and function of the selected pentose transporter (CCM_06358) as a representative of sugar transportome in
. Considering the network bottlenecks and structural organizations, we further identified key amino acids (Phe38 and Trp441) and their interactions with other residues, contributing the xylose transport function, as verified by binding free energy calculation. The strategy used herein generated remarkably valuable biological information, which is applicable for the study of sugar transportome and the structure engineering of targeted transporter proteins that might link to the production of bioactive compounds derived from xylose metabolism, such as cordycepin.
PDF
