Spatiotemporal patterns of the pregnancy microbiome and links to reproductive disorders.

The microbiome of females undergoes extensive remodeling during pregnancy, which is likely to have an impact on the health of both mothers and offspring. Nevertheless, large-scale integrated investigations characterizing microbiome dynamics across key body habitats are lacking. Here, we performed an extensive meta-analysis that compiles and analyzes microbiome profiles from  >10,000 samples across the gut, vagina, and oral cavity of pregnant women from diverse geographical regions. We have unveiled unexpected variations in the taxonomic, functional, and ecological characteristics of microbial communities throughout the course of pregnancy. The gut microbiota showed distinct trajectories between Western and non-Western populations. The vagina microbiota exhibited fluctuating transitions at the genus level across gestation, while the oral microbiota remained relatively stable. We also identified distinctive microbial signatures associated with prevalent pregnancy-related disorders, including opposite variations in the oral and gut microbiota of patients with gestational diabetes and disrupted microbial networks in preterm birth. This study establishes a comprehensive atlas of the pregnancy microbiome by integrating multidimensional datasets and offers foundational insights into the intricate interplay between microbes and host factors that underlie reproductive health.

MAMI: a comprehensive database of mother-infant microbiome and probiotic resources.

Extensive evidence has demonstrated that the human microbiome and probiotics confer great impacts on human health, particularly during critical developmental stages such as pregnancy and infancy when microbial communities undergo remarkable changes and maturation. However, a major challenge in understanding the microbial community structure and interactions between mothers and infants lies in the current lack of comprehensive microbiome databases specifically focused on maternal and infant health. To address this gap, we have developed an extensive database called MAMI (Microbiome Atlas of Mothers and Infants) that archives data on the maternal and neonatal microbiome, as well as abundant resources on edible probiotic strains. By leveraging this resource, we can gain profound insights into the dynamics of microbial communities, contributing to lifelong wellness for both mothers and infants through precise modulation of the developing microbiota. The functionalities incorporated into MAMI provide a unique perspective on the study of the mother-infant microbiome, which not only advance microbiome-based scientific research but also enhance clinical practice. MAMI is publicly available at https://bioinfo.biols.ac.cn/mami/.

PandaGUT provides new insights into bacterial diversity, function, and resistome landscapes with implications for conservation.

BACKGROUND: The gut microbiota play important roles in host adaptation and evolution, but are understudied in natural population of wild mammals. To address host adaptive evolution and improve conservation efforts of threatened mammals from a metagenomic perspective, we established a high-quality gut microbiome catalog of the giant panda (pandaGUT) to resolve the microbiome diversity, functional, and resistome landscapes using approximately 7 Tbp of long- and short-read sequencing data from 439 stool samples. RESULTS: The pandaGUT catalog comprises 820 metagenome-assembled genomes, including 40 complete closed genomes, and 64.5% of which belong to species that have not been previously reported, greatly expanding the coverage of most prokaryotic lineages. The catalog contains 2.37 million unique genes, with 74.8% possessing complete open read frames, facilitating future mining of microbial functional potential. We identified three microbial enterotypes across wild and captive panda populations characterized by Clostridium, Pseudomonas, and Escherichia, respectively. We found that wild pandas exhibited host genetic-specific microbial structures and functions, suggesting host-gut microbiota phylosymbiosis, while the captive cohorts encoded more multi-drug resistance genes. CONCLUSIONS: Our study provides largely untapped resources for biochemical and biotechnological applications as well as potential intervention avenues via the rational manipulation of microbial diversity and reducing antibiotic usage for future conservation management of wildlife. Video Abstract.

Gut microbiota dysbiosis associated with different types of demyelinating optic neuritis in patients.

BACKGROUND: Demyelinating optic neuritis (DON) causes rapid vision loss in young and middle-aged people. The limited efficacy of treatment and the toxic side effects of drugs significantly affect the quality of life of patients with DON. Therefore, DON pathogenesis has always been a research hotspot in terms of prevention and treatment. Studies have suggested that gut microbiota imbalances may be involved in autoimmune disease development via the modulation of multiple inflammatory cytokines and anti-inflammatory metabolites. Therefore, this study aims to explore gut microbiota differences between healthy controls (HCs) and patients with DON. METHODS: A total of 54 patients with DON and 41 HCs were recruited. Fecal and blood samples were collected before and after intravenous methylprednisolone pulse (IVMP) treatment. The Shannon index, gut microbiota structure, and differential bacteria were evaluated and compared. RESULTS: The Shannon diversity index was decreased in patients with DON (p < 0.001) but was higher after IVMP treatment (p < 0.05). In patients with DON, Blautia, Escherichia-Shigella, and Ruminococcus showed higher abundances, whereas Bacteroides, Faecalibacterium, Roseburia, Parabacteroides, Romboutsia, and Alistipes showed lower abundances compared to that in the HCs. After IVMP treatment, the Shannon index of the myelin oligodendrocyte glycoprotein-immunoglobulin G (+) (MOG-IgG (+)) and both aquaporin-4 (AQP4)-IgG (-) and MOG-IgG (-) groups increased (p < 0.05). Bacteroides was negatively correlated with interleukin (IL)-21, IL-17E, and tumor necrosis factor-α levels (p < 0.05, r = -0.54; p < 0.05, r= -0.50; p < 0.05, r =-0.55, respectively). Escherichia was positively correlated with macrophage inflammatory protein-3α (p < 0.05, r = 0.51). Alistipes was negatively correlated with soluble CD40 ligand (p < 0.05, r = -0.52). CONCLUSION: The gut microbiota differed significantly between patients with DON and HCs; however, IVMP treatment may restore gut microbiota diversity and structure in patients with DON. Moreover, gut microbiota changes may play a role in DON pathogenesis.

A novel peptide protects against diet-induced obesity by suppressing appetite and modulating the gut microbiota.

OBJECTIVE: The obesity epidemic and its metabolic complications continue to be a major global public health threat with limited effective treatments, especially drugs that can be taken orally. Peptides are a promising class of molecules that have gained increased interest for their applications in medicine and biotechnology. In this study, we focused on looking for peptides that can be administrated orally to treat obesity and exploring its mechanisms. DESIGN: Here, a 9-amino-acid peptide named D3 was designed and administered orally to germ-free (GF) mice and wild-type (WT) mice, rats and macaques. The effects of D3 on body weight and other basal metabolic parameters were evaluated. The effects of D3 on gut microbiota were evaluated using 16S rRNA amplicon sequencing. To identify and confirm the mechanisms of D3, transcriptome analysis of ileum and molecular approaches on three animal models were performed. RESULTS: A significant body weight reduction was observed both in WT (12%) and GF (9%) mice treated with D3. D3 ameliorated leptin resistance and upregulated the expression of uroguanylin (UGN), which suppresses appetite via the UGN-GUCY2C endocrine axis. Similar effects were also found in diet-induced obese rat and macaque models. Furthermore, the abundance of intestinal Akkermansia muciniphila increased about 100 times through the IFNγ-Irgm1 axis after D3 treatment, which may further inhibit fat absorption by downregulating Cd36. CONCLUSION: Our results indicated that D3 is a novel drug candidate for counteracting diet-induced obesity as a non-toxic and bioactive peptide. Targeting the UGN-GUCY2C endocrine axis may represent a therapeutic strategy for the treatment of obesity.

Changes to gut amino acid transporters and microbiome associated with increased E/I ratio in Chd8+/- mouse model of ASD-like behavior.

Autism spectrum disorder (ASD), a group of neurodevelopmental disorders characterized by social communication deficits and stereotyped behaviors, may be associated with changes to the gut microbiota. However, how gut commensal bacteria modulate brain function in ASD remains unclear. Here, we used chromodomain helicase DNA-binding protein 8 (CHD8) haploinsufficient mice as a model of ASD to elucidate the pathways through which the host and gut microbiota interact with each other. We found that increased levels of amino acid transporters in the intestines of the mouse model of ASD contribute to the high level of serum glutamine and the increased excitation/inhibition (E/I) ratio in the brain. In addition, elevated α-defensin levels in the haploinsufficient mice resulted in dysregulation of the gut microbiota characterized by a reduced abundance of Bacteroides. Furthermore, supplementation with Bacteroides uniformis improved the ASD-like behaviors and restored the E/I ratio in the brain by decreasing intestinal amino acid transport and the serum glutamine levels. Our study demonstrates associations between changes in the gut microbiota and amino acid transporters, and ASD-like behavioral and electrophysiology phenotypes, in a mouse model.

Aliidiomarina halalkaliphila sp. nov., a haloalkaliphilic bacterium isolated from a soda lake in Inner Mongolia Autonomous Region, China.

A haloalkaliphilic strain (IM 1326T) was isolated from brine sampled at a soda lake in the Inner Mongolia Autonomous Region, China. Cells of the strain were rod-shaped and motile. Strain IM 1326T was able to grow at 4-42 °C (optimum, 37 °C) with 0-13.0 % (w/v) NaCl concentrations (optimum at 4.0-6.0 %) and at pH 7.5-11.0 (optimum at 9.0-10.0). The 16S rRNA gene phylogenetic analysis revealed that the isolate belongs to the genus Aliidiomarina and is closely related to the type strains of Aliidiomarina sanyensis (95.8 % sequence similarity), Aliidiomarina shirensis (95.7 %), Aliidiomarina iranensis (95.4 %) and Aliidiomarina haloalkalitolerans (95.3 %). The whole genome of strain IM 1326T was sequenced, and the genomic DNA G+C content was 49.7 mol%. Average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values between the isolate and the related Aliidiomarina species were 68.1-84.9 %, 76-78 % and 18.4-20.4 %, respectively. The respiratory quinone was ubiquinone-8. The polar lipid profile included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and one unidentified aminophospholipid. The predominant cellular fatty acids were summed feature 9 (10-methyl-C16 : 0/iso-C17 : 1 ω9c, 22.2 %), iso-C15 : 0 (16.1 %) and iso-C17 : 0 (13.1 %). Based on the results of phylogenetic analysis, genome relatedness, and the physiological and chemotaxonomic properties of the isolate, strain IM 1326T is considered to represent a novel species of the genus Aliidiomarina, for which the name Aliidiomarina halalkaliphila sp. nov. is proposed (type strain IM 1326T=CGMCC 1.17056T=JCM 34227T).

Maternal and neonatal viromes indicate the risk of offspring's gastrointestinal tract exposure to pathogenic viruses of vaginal origin during delivery.

A cumulative effect of enterovirus and gluten intake on the risk of celiac disease autoimmunity in infants highlights the significance of viral exposure in early life on the health of children. However, pathogenic viruses may be transmitted to the offspring in an earlier period, raising the possibility that women whose vaginas are inhabited by such viruses may have had their babies infected as early as the time of delivery. A high-resolution intergenerational virome atlas was obtained by metagenomic sequencing and virome analysis on 486 samples from six body sites of 99 mother-neonate pairs. We found that neonates had less diverse oral and enteric viruses than mothers. Vaginally delivered newborns seconds after birth had a more similar oral virome and more viruses of vaginal origin than cesarean-section (C-section) newborns (56.9% vs. 5.8%). Such viruses include both Lactobacillus phage and potentially pathogenic viruses, such as herpesvirus, vaccinia virus, and hepacivirus, illustrating a relatively high variety of the pioneer viral taxa at the time of delivery and a delivery-dependent mother-to-neonate transmission along the vaginal-oral-intestinal route. Neonates are exposed to vaginal viruses as they pass through the reproductive tract, and viruses of vaginal origin may threaten their health. These findings challenge the conventional notion that vaginal delivery is definitely better than cesarean delivery from the perspective of microbial transmission. Screening for vaginal virome before delivery is a worthwhile step to advocate in normal labor to eliminate the risk of intergenerational transmission of pathogenic viruses to offspring.

Highly integrated adaptive mechanisms in Spiribacter halalkaliphilus, a bacterium abundant in Chinese soda-saline lakes.

Soda-saline lakes are polyextreme environments inhabited by many haloalkaliphiles, including one of the most abundant Spiribacter species. However, its mechanisms of adaptation are not ecophysiologically characterized. Based on a large-scale cultivation strategy, we obtained a representative isolate of this Spiribacter species whose relative abundance was the highest (up to 15.63%) in a wide range of salinities in the soda-saline lakes in Inner Mongolia, China. This species is a chemoorganoheterotrophic haloalkaliphile. It has a small and streamlined genome and utilizes a wide variety of compatible solutes to resist osmotic pressure and multiple monovalent cation/proton antiporters for pH homeostasis. In addition to growth enhancement by light under microaerobic conditions, cell growth, organic substrate consumption and polyhydroxybutyrate biosynthesis were also improved by inorganic sulfide. Both quantitative RT-PCR and enzymatic assays verified that sulfide:quinone oxidoreductase was upregulated during this process. Metatranscriptomic analysis indicated that all genes related to environmental adaptation were transcribed in natural environments. Overall, this study has identified a novel abundant haloalkaliphile with multiple and highly integrated adaptive strategies and found that inorganic sulfide was able to improve the adaptation of a heterotroph to polyextreme environments.

Multi-omics reveals the positive leverage of plant secondary metabolites on the gut microbiota in a non-model mammal.

BACKGROUND: Flavonoids are important plant secondary metabolites (PSMs) that have been widely used for their health-promoting effects. However, little is known about overall flavonoid metabolism and the interactive effects between flavonoids and the gut microbiota. The flavonoid-rich bamboo and the giant panda provide an ideal system to bridge this gap. RESULTS: Here, integrating metabolomic and metagenomic approaches, and in vitro culture experiment, we identified 97 flavonoids in bamboo and most of them have not been identified previously; the utilization of more than 70% flavonoid monomers was attributed to gut microbiota; the variation of flavonoid in bamboo leaves and shoots shaped the seasonal microbial fluctuation. The greater the flavonoid content in the diet was, the lower microbial diversity and virulence factor, but the more cellulose-degrading species. CONCLUSIONS: Our study shows an unprecedented landscape of beneficial PSMs in a non-model mammal and reveals that PSMs remodel the gut microbiota conferring host adaptation to diet transition in an ecological context, providing a novel insight into host-microbe interaction. Video abstract.

Optimising PHBV biopolymer production in haloarchaea via CRISPRi-mediated redirection of carbon flux.

The haloarchaeon Haloferax mediterranei is a potential strain for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) production, yet the production yield and cost are the major obstacles hindering the use of this archaeal strain. Leveraging the endogenous type I-B CRISPR-Cas system in H. mediterranei, we develop a CRISPR-based interference (CRISPRi) approach that allows to regulate the metabolic pathways related to PHBV synthesis, thereby enhancing PHBV production. Our CRISPRi approach can downregulate the gene expression in a range of 25% to 98% depending upon the target region. Importantly, plasmid-mediated CRISPRi downregulation on the citrate synthase genes (citZ and gltA) improves the PHBV accumulation by 76.4% (from 1.78 to 3.14 g/L). When crRNA cassette integrated into chromosome, this further shortens the PHBV fermentation period and enhances PHA productivity by 165%. Our transcriptome analysis shows that repression of citrate synthase genes redirects metabolic flux from the central metabolic pathways to PHBV synthesis pathway. These findings demonstrate that the CRISPRi-based gene regulation is a transformative toolkit for fine-tuning the endogenous metabolic pathways in the archaeal system, which can be applied to not only the biopolymer production but also many other applications.

Comprehensive profiling of circular RNAs with nanopore sequencing and CIRI-long.

Reconstructing the sequence of circular RNAs (circRNAs) from short RNA sequencing reads has proved challenging given the similarity of circRNAs and their corresponding linear messenger RNAs. Previous sequencing methods were unable to achieve high-throughput detection of full-length circRNAs. Here we describe a protocol for enrichment and full-length sequencing of circRNA isoforms using nanopore technology. Circular reverse transcription and size selection achieves a 20-fold higher enrichment of circRNAs from total RNA compared to previous methods. We developed an algorithm, called circRNA identifier using long-read sequencing data (CIRI-long), to reconstruct the sequence of circRNAs. The workflow was validated with simulated data and by comparison to Illumina sequencing as well as quantitative real-time RT-PCR. We used CIRI-long to analyze adult mouse brain samples and systematically profile circRNAs, including mitochondria-derived and transcriptional read-through circRNAs. We identified a new type of intronic self-ligated circRNA that exhibits special splicing and expression patterns. Our method takes advantage of nanopore long reads and enables unbiased reconstruction of full-length circRNA sequences.

Natronorubrum halalkaliphilum sp. nov., a haloalkaliphilic archaeon isolated from soda lake in Inner Mongolia Autonomous Region, China.

A haloalkaliphilic strain JWXQ-INN-674T was isolated from the water sample of a soda lake in Inner Mongolia Autonomous Region, China. Cells of the strain were coccoid, motile, and strictly aerobic. The strain was able to grow in presence of 2.6-5.4 M NaCl (optimum concentration is 3.4 M) at 30-50 °C (optimum temperature is 42 °C) and pH 7-9.5 (optimum pH is 9.0). The 16S rRNA gene sequence of strain JWXQ-INN-674T showed 95.3-96.6% similarity to members of the genus Natronorubrum of the family Natrialbaceae. The whole genome sequencing of strain JWXQ-INN-674T revealed a genome size of 4.56 M bp and a DNA G + C content of 62.5 mol%. Genome relatedness of strain JWXQ-INN-674T and other species in the genus Natronorubrum was analyzed by average nucleotide identity and digital DNA-DNA hybridization with the values of 76.8-90.6 and 23.1-39.3%, respectively. The strain possessed the polar lipids phosphatidylglycerol and methylated phosphatidylglycerol phosphate lipid. No glycolipids were detected. Based on phylogenetic analysis, phenotypic characteristics, chemotaxonomic properties and genome relatedness, the isolate was proposed as the type strain of a novel species of genus Natronorubrum, Natronorubrum halalkaliphilum sp. nov. (type strain JWXQ-INN-674T = CGMCC 1.17283T = JCM 34245T).

Halalkalirubrum salinum gen. nov., sp. nov., a halophilic archaeon isolated from a saline lake.

A novel extremely halophilic archaeon, strain N1521T, was isolated from a saline lake in Tibet, China. Cells of the strain were pleomorphic and Gram-stain-negative. It produced red pigments. Growth was observed at 4-42 °C (optimum, 37 °C), pH 7.0-10.5 (optimum, 8.0-9.5), NaCl 11%-25% (optimum, 15%) and in the presence of 0-0.1 M MgCl2 (optimum, 0.05 M) in aerobic conditions. The minimum NaCl concentration that prevented cell lysis was 2% (w/v). The major polar lipids of strain N1521T were phosphatidylglycerol sulfate, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol and an unidentified glycolipid. The DNA G + C content was 58.37 mol%. According to 16S rRNA gene sequence comparisons, strain N1521T revealed the highest sequence similarity to Haloprofundus halophilus NK23T (91.38%) and Halogranum amylolyticum TNN58T (91.00%), and low sequence similarities (< 91%) with other genera in the order Haloferacales. Phylogenetic analysis based on the 16S rRNA gene and rpoB' gene sequence showed that strain N1521T was distinct from the members of the order Haloferacales. The digital DNA-DNA hybridization, average nucleotide identity and average amino acid identity values calculated from whole genome-sequence comparison between strain N1521T and the members of the order Haloferacales were in the ranges of 15.1-18.2%, 68.8-73.0%, and 58.4-63.9%, respectively. Phylogenetic tree reconstructions based on the whole-genome sequences revealed that strain N1521T was closer to the members of the family Halorubraceae. Based on the data obtained, strain N1521T is thus considered to represent a novel species of a new genus within the family Halorubraceae, for which the name Halalkalirubrum salinum gen. nov., sp. nov. is proposed. The type strain is N1521T (= CGMCC 1.16693 = JCM 33785).

Salinadaptatus halalkaliphilus gen. nov., sp. nov., a haloalkaliphilic archaeon isolated from salt pond in Inner Mongolia Autonomous Region, China.

A haloalkaliphilic strain XQ-INN 246T was isolated from the sediment of a salt pond in Inner Mongolia Autonomous Region, China. Cells of the strain were rods, motile and strictly aerobic. The strain was able to grow in the presence of 2.6-5.3 M NaCl (optimum concentration is 4.4 M) at 30-50 °C (optimum temperature is 42 °C) and pH 7.0-10.0 (optimum pH is 8.0-8.5). The whole genome sequencing of strain XQ-INN 246T revealed a genome size of 4.52 Mbp and a DNA G+C content of 62.06 mol%. Phylogenetic tree based on 16S rRNA gene sequences and concatenated amino acid sequences of 122 single-copy conserved proteins revealed a robust lineage of the strain XQ-INN 246T with members of related genera of the family Natrialbaceae. The strain possessed the polar lipids of phosphatidylglycerol and phosphatidylglycerol phosphate methyl ester. No glycolipids were detected. Based on phylogenetic analysis, phenotypic characteristics, chemotaxonomic properties and genome relatedness, the isolate was proposed as the type strain of a novel species of a new genus within the family Natrialbaceae, for which the name Salinadaptatus halalkaliphilus gen. nov., sp. nov. is proposed. The type strain is XQ-INN 246T (=CGMCC 1.16692T=JCM 33751T).

Unusual Phosphoenolpyruvate (PEP) Synthetase-Like Protein Crucial to Enhancement of Polyhydroxyalkanoate Accumulation in Haloferax mediterranei Revealed by Dissection of PEP-Pyruvate Interconversion Mechanism.

Phosphoenolpyruvate (PEP)/pyruvate interconversion is a major metabolic point in glycolysis and gluconeogenesis and is catalyzed by various sets of enzymes in different Archaea groups. In this study, we report the key enzymes that catalyze the anabolic and catabolic directions of the PEP/pyruvate interconversion in Haloferax mediterranei The in silico analysis showed the presence of a potassium-dependent pyruvate kinase (PYKHm [HFX_0773]) and two phosphoenol pyruvate synthetase (PPS) candidates (PPSHm [HFX_0782] and a PPS homolog protein named PPS-like [HFX_2676]) in this strain. Expression of the pykHm gene and ppsHm was induced by glycerol and pyruvate, respectively; whereas the pps-like gene was not induced at all. Similarly, genetic analysis and enzyme activities of purified proteins showed that PYKHm catalyzed the conversion from PEP to pyruvate and that PPSHm catalyzed the reverse reaction, while PPS-like protein displayed no function in PEP/pyruvate interconversion. Interestingly, knockout of the pps-like gene led to a 70.46% increase in poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) production. The transcriptome sequencing (RNA-Seq) and quantitative reverse transcription-PCR (qRT-PCR) results showed that many genes responsible for PHBV monomer supply and for PHBV synthesis were upregulated in a pps-like gene deletion strain and thereby improved PHBV accumulation. Additionally, our phylogenetic evidence suggested that PPS-like protein diverged from PPS enzyme and evolved as a distinct protein with novel function in haloarchaea. Our findings attempt to fill the gaps in central metabolism of Archaea by providing comprehensive information about key enzymes involved in the haloarchaeal PEP/pyruvate interconversion, and we also report a high-yielding PHBV strain with great future potentials.IMPORTANCEArchaea, the third domain of life, have evolved diversified metabolic pathways to cope with their extreme habitats. Phosphoenol pyruvate (PEP)/pyruvate interconversion during carbohydrate metabolism is one such important metabolic process that is highly differentiated among Archaea However, this process is still uncharacterized in the haloarchaeal group. Haloferax mediterranei is a well-studied haloarchaeon that has the ability to produce polyhydroxyalkanoates (PHAs) under unbalanced nutritional conditions. In this study, we identified the key enzymes involved in this interconversion and discussed their differences with their counterparts from other members of the Archaea and Bacteria domains. Notably, we found a novel protein, phosphoenolpyruvate synthetase-like (PPS-like), which exhibited high homology to PPS enzyme. However, PPS-like protein has evolved some distinct sequence features and functions, and strikingly the corresponding gene deletion helped to enhance poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) synthesis significantly. Overall, we have filled the gap in knowledge about PEP/pyruvate interconversion in haloarchaea and reported an efficient strategy for improving PHBV production in H. mediterranei.

Engineering Haloferax mediterranei as an Efficient Platform for High Level Production of Lycopene.

Lycopene attracts increasing interests in the pharmaceutical, food, and cosmetic industries due to its anti-oxidative and anti-cancer properties. Compared with other lycopene production methods, such as chemical synthesis or direct extraction from plants, the biosynthesis approach using microbes is more economical and sustainable. In this work, we engineered Haloferax mediterranei, a halophilic archaeon, as a new lycopene producer. H. mediterranei has the de novo synthetic pathway for lycopene but cannot accumulate this compound. To address this issue, we reinforced the lycopene synthesis pathway, blocked its flux to other carotenoids and disrupted its competitive pathways. The reaction from geranylgeranyl-PP to phytoene catalyzed by phytoene synthase (CrtB) was identified as the rate-limiting step in H. mediterranei. Insertion of a strong promoter PphaR immediately upstream of the crtB gene, or overexpression of the heterologous CrtB and phytoene desaturase (CrtI) led to a higher yield of lycopene. In addition, blocking bacterioruberin biosynthesis increased the purity and yield of lycopene. Knock-out of the key genes, responsible for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) biosynthesis, diverted more carbon flux into lycopene synthesis, and thus further enhanced lycopene production. The metabolic engineered H. mediterranei strain produced lycopene at 119.25 ± 0.55 mg per gram of dry cell weight in shake flask fermentation. The obtained yield was superior compared to the lycopene production observed in most of the engineered Escherichia coli or yeast even when they were cultivated in pilot scale bioreactors. Collectively, this work offers insights into the mechanism involved in carotenoid biosynthesis in haloarchaea and demonstrates the potential of using haloarchaea for the production of lycopene or other carotenoids.