Comparative Genomics Unveils Functional Diversity, Pangenome Openness, and Underlying Biological Drivers among Bacillus subtilis Group.

The Bacillus subtilis group (Bs group), with Bacillus subtilis as its core species, holds significant research and economic value in various fields, including science, industrial production, food, and pharmaceuticals. However, most studies have been confined to comparative genomics analyses and exploration within individual genomes at the level of species, with few conducted within groups across different species. This study focused on Bacillus subtilis, the model of Gram-positive bacteria, and 14 other species with significant research value, employing comparative pangenomics as well as population enrichment analysis to ascertain the functional enrichment and diversity. Through the quantification of pangenome openness, this work revealed the underlying biological drivers and significant correlation between pangenome openness and various factors, including the distribution of toxin-antitoxin- and integrase-related genes, as well as the number of endonucleases, recombinases, repair system-related genes, prophages, integrases, and transfer mobile elements. Furthermore, the functional enrichment results indicated the potential for secondary metabolite, probiotic, and antibiotic exploration in Bacillus licheniformis, Bacillus paralicheniformis, and Bacillus spizizenii, respectively. In general, this work systematically exposed the quantification of pangenome openness, biological drivers, the pivotal role of genomic instability factors, and mobile elements, providing targeted exploration guidance for the Bs group.

Silencing Ditylenchus destructor cathepsin L-like cysteine protease has negative pleiotropic effect on nematode ontogenesis.

Ditylenchus destructor is a migratory plant-parasitic nematode that severely harms many agriculturally important crops. The control of this pest is difficult, thus efficient strategies for its management in agricultural production are urgently required. Cathepsin L-like cysteine protease (CPL) is one important protease that has been shown to participate in various physiological and pathological processes. Here we decided to characterize the CPL gene (Dd-cpl-1) from D. destructor. Analysis of Dd-cpl-1 gene showed that Dd-cpl-1 gene contains a signal peptide, an I29 inhibitor domain with ERFNIN and GNFD motifs, and a peptidase C1 domain with four conserved active residues, showing evolutionary conservation with other nematode CPLs. RT-qPCR revealed that Dd-cpl-1 gene displayed high expression in third-stage juveniles (J3s) and female adults. In situ hybridization analysis demonstrated that Dd-cpl-1 was expressed in the digestive system and reproductive organs. Silencing Dd-cpl-1 in 1-cell stage eggs of D. destructor by RNAi resulted in a severely delay in development or even in abortive morphogenesis during embryogenesis. The RNAi-mediated silencing of Dd-cpl-1 in J2s and J3s resulted in a developmental arrest phenotype in J3 stage. In addition, silencing Dd-cpl-1 gene expression in female adults led to a 57.43% decrease in egg production. Finally, Dd-cpl-1 RNAi-treated nematodes showed a significant reduction in host colonization and infection. Overall, our results indicate that Dd-CPL-1 plays multiple roles in D. destructor ontogenesis and could serve as a new potential target for controlling D. destructor.

Effect of simplified inoculum agent on performance and microbiome during cow manure-composting at industrial-scale.

A simplified inoculum agent, only comprising Bacillus subtilis and Aspergillus niger, was utilized for industrial-scale cow-manure composting to investigate its impact on composting performance and microbiome. Inoculants elevated the average and peak temperatures by up to 7 and 10 °C, respectively, during the thermophilic stage, reduced organic matter content, and raised germination index. Inoculation also extended the period of composting above 50 °C from 12 to 26 days. Sequencing unveiled significant shifts in microbial diversity, composition, and function. Aspergillus thrived during the mesophilic phase, potentially initiating composting, whereas Bacillus, Lysinibacillus, and Clostridium were enriched during the thermophilic stage. Metagenomic sequencing revealed an increased abundance of carbohydrate-active enzymes and glycometabolism-related genes responsible for lignocellulose degradation and heat generation after inoculation. These enriched microbes and functional genes contributed to organic matter degradation and temperature maintenance during thermophilic stage, expediting composting. This suggests the effectiveness of this simplified inoculum in industrial-level cow-manure composting.

Starter Culture Development and Innovation for Novel Fermented Foods.

Interest in fermented foods is increasing because fermented foods are promising solutions for more secure food systems with an increased proportion of minimally processed plant foods and a smaller environmental footprint. These developments also pertain to novel fermented food for which no traditional template exists, raising the question of how to develop starter cultures for such fermentations. This review establishes a framework that integrates traditional and scientific knowledge systems for the selection of suitable cultures. Safety considerations, the use of organisms in traditional food fermentations, and the link of phylogeny to metabolic properties provide criteria for culture selection. Such approaches can also select for microbial strains that have health benefits. A science-based approach to the development of novel fermented foods can substantially advance their value through more secure food systems, food products that provide health-promoting microbes, and the provision of foods that improve human health. Expected final online publication date for the Annual Review of Food Science and Technology, Volume 15 is April 2024. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Unzipped chromosome-level genomes reveal allopolyploid nematode origin pattern as unreduced gamete hybridization.

The formation and consequences of polyploidization in animals with clonal reproduction remain largely unknown. Clade I root-knot nematodes (RKNs), characterized by parthenogenesis and allopolyploidy, show a widespread geographical distribution and extensive agricultural destruction. Here, we generated 4 unzipped polyploid RKN genomes and identified a putative novel alternative telomeric element. Then we reconstructed 4 chromosome-level assemblies and resolved their genome structures as AAB for triploid and AABB for tetraploid. The phylogeny of subgenomes revealed polyploid RKN origin patterns as hybridization between haploid and unreduced gametes. We also observed extensive chromosomal fusions and homologous gene expression decrease after polyploidization, which might offset the disadvantages of clonal reproduction and increase fitness in polyploid RKNs. Our results reveal a rare pathway of polyploidization in parthenogenic polyploid animals and provide a large number of high-precision genetic resources that could be used for RKN prevention and control.

Sequence variety in the CC' loop of Siglec-8/9/3 determines the recognitions to sulfated oligosaccharides.

Siglecs are important lectins found in different types of immune cells and function as regulatory molecules by recognizing self-associated glycans and converting extracellular interactions into signals for inhibiting immune cell functions. Although many Siglecs have been found to show broad specificities and recognize different types of sulfated oligosaccharides, Siglec-8 and Siglec-9 displayed a high degree of specificity for sialyl N-acetyllactosamine (sLacNAc) with sulfations at O6-positions of the galactose (6'-sulfation) and N-acetylglucosamine (6-sulfation), respectively. Siglec-3 was recently discovered to bind sLacNAc both sulfations. In addition to a conserved arginine residue for binding to sialic acid residue, the sequence variety in the CC' loop may provide binding specificities to sulfated oligosaccharides in Siglecs. Thus, the present study employed molecular models to study the impact of different residues in the CC' loops of Siglec-8/9/3 to the recognitions of 6-sulfations in Gal and/or GlcNAc of sLacNAc. The negatively charged residues in the CC' loop of Siglec-9 formed unfavorable electrostatic repulsions with the 6-sulfate in Gal and resulted no recognitions, in contrast to the favorable interactions formed between the positively charged residues in the CC' loop of Siglec-8 and the 6-sulfate in Gal resulting strong specificity. A two-state binding model was proposed for Siglec-3 recognizing 6-sulfations in Gal and GlcNAc of sLacNAc, as the neutral residues in the CC' loop of Siglec-3 could not form strong favorable interactions to lock the 6-sulfate in Gal within a single binding pose or strong unfavorable interactions to repel the 6-sulfate in Gal. The oligosaccharide adopted two distinctive binding poses and oriented the sulfate groups to form interactions with residues in the CC' loop and G-strand. The present study provided a structural mechanism for the sequence variety in the CC' loop of Siglec-8/9/3 determining the recognitions to the sulfated oligosaccharides and offered insights into the binding specificities for Siglecs.

Metagenome-assembled genomes reveal carbohydrate degradation and element metabolism of microorganisms inhabiting Tengchong hot springs, China.

A hot spring is a distinctive aquatic environment that provides an excellent system to investigate microorganisms and their function in elemental cycling processes. Previous studies of terrestrial hot springs have been mostly focused on the microbial community, one special phylum or category, or genes involved in a particular metabolic step, while little is known about the overall functional metabolic profiles of microorganisms inhabiting the terrestrial hot springs. Here, we analyzed the microbial community structure and their functional genes based on metagenomic sequencing of six selected hot springs with different temperature and pH conditions. We sequenced a total of 11 samples from six hot springs and constructed 162 metagenome-assembled genomes (MAGs) with completeness above 70% and contamination lower than 10%. Crenarchaeota, Euryarchaeota and Aquificae were found to be the dominant phyla. Functional annotation revealed that bacteria encode versatile carbohydrate-active enzymes (CAZYmes) for the degradation of complex polysaccharides, while archaea tend to assimilate C1 compounds through carbon fixation. Under nitrogen-deficient conditions, there were correspondingly fewer genes involved in nitrogen metabolism, while abundant and diverse set of genes participating in sulfur metabolism, particularly those associated with sulfide oxidation and thiosulfate disproportionation. In summary, archaea and bacteria residing in the hot springs display distinct carbon metabolism fate, while sharing the common energy preference through sulfur metabolism. Overall, this research contributes to a better comprehension of biogeochemistry of terrestrial hot springs.

Translocation of gut microbes to epididymal white adipose tissue drives lipid metabolism disorder under heat stress.

Heat stress induces multi-organ damage and serious physiological dysfunction in mammals, and gut bacteria may translocate to extra-intestinal tissues under heat stress pathology. However, whether gut bacteria translocate to the key metabolic organs and impair function as a result of heat stress remains unknown. Using a heat stress-induced mouse model, heat stress inhibited epididymal white adipose tissue (eWAT) expansion and induced lipid metabolic disorder but did not damage other organs, such as the heart, liver, spleen, or muscle. Microbial profiling analysis revealed that heat stress shifted the bacterial community in the cecum and eWAT but not in the inguinal white adipose tissue, blood, heart, liver, spleen, or muscle. Notably, gut-vascular barrier function was impaired, and the levels of some bacteria, particularly Lactobacillus, were higher in the eWAT, as confirmed by catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) staining when mice were under heat stress. Moreover, integrated multi-omics analysis showed that the eWAT microbiota was associated with host lipid metabolism, and the expression of genes involved in the lipid metabolism in eWAT was upregulated under heat stress. A follow-up microbial supplementation study after introducing Lactobacillus plantarum to heat-stressed mice revealed that the probiotic ameliorated heat stress-induced loss of eWAT and dyslipidemia and reduced gut bacterial translocation to the eWAT by improving gut barrier function. Overall, our findings suggest that gut bacteria, particularly Lactobacillus spp., play a crucial role in heat stress-induced lipid metabolism disorder and that there is therapeutic potential for using probiotics, such as Lactobacillus plantarum.

Microbe-driven elemental cycling enables microbial adaptation to deep-sea ferromanganese nodule sediment fields.

BACKGROUND: Ferromanganese nodule-bearing deep-sea sediments cover vast areas of the ocean floor, representing a distinctive habitat in the abyss. These sediments harbor unique conditions characterized by high iron concentration and low degradable nutrient levels, which pose challenges to the survival and growth of most microorganisms. While the microbial diversity in ferromanganese nodule-associated sediments has been surveyed several times, little is known about the functional capacities of the communities adapted to these unique habitats. RESULTS: Seven sediment samples collected adjacent to ferromanganese nodules from the Clarion-Clipperton Fracture Zone (CCFZ) in the eastern Pacific Ocean were subjected to metagenomic analysis. As a result, 179 high-quality metagenome-assembled genomes (MAGs) were reconstructed and assigned to 21 bacterial phyla and 1 archaeal phylum, with 88.8% of the MAGs remaining unclassified at the species level. The main mechanisms of resistance to heavy metals for microorganisms in sediments included oxidation (Mn), reduction (Cr and Hg), efflux (Pb), synergy of reduction and efflux (As), and synergy of oxidation and efflux (Cu). Iron, which had the highest content among all metallic elements, may occur mainly as Fe(III) that potentially functioned as an electron acceptor. We found that microorganisms with a diverse array of CAZymes did not exhibit higher community abundance. Instead, microorganisms mainly obtained energy from oxidation of metal (e.g., Mn(II)) and sulfur compounds using oxygen or nitrate as an electron acceptor. Chemolithoautotrophic organisms (Thaumarchaeota and Nitrospirota phyla) were found to be potential manganese oxidizers. The functional profile analysis of the dominant microorganisms further indicated that utilization of inorganic nutrients by redox reactions (rather than organic nutrient metabolism) is a major adaptive strategy used by microorganisms to support their survival in the ferromanganese nodule sediments. CONCLUSIONS: This study provides a comprehensive metagenomic analysis of microbes inhabiting metal-rich ferromanganese nodule sediments. Our results reveal extensive redundancy across taxa for pathways of metal resistance and transformation, the highly diverse mechanisms used by microbes to obtain nutrition, and their participation in various element cycles in these unique environments. Video Abstract.

Population genomics and pathotypic evaluation of the bacterial leaf blight pathogen of rice reveals rapid evolutionary dynamics of a plant pathogen.

The Xanthomonas oryzae pv. oryzae (Xoo) is a bacterial pathogen causing bacterial blight disease in rice, resulting in significant yield reductions of up to 50% in rice production. Despite its serious threat to food production globally, knowledge of its population structure and virulence evolution is relatively limited. In this study, we employed whole-genome sequencing to explore the diversity and evolution of Xoo in the main rice-growing areas of China over the past 30 years. Using phylogenomic analysis, we revealed six lineages. CX-1 and CX-2 primarily contained Xoo isolates from South China, while CX-3 represented Xoo isolates from North China. Xoo isolates belonging to CX-5 and CX-6 were the most prevalent across all studied areas, persisting as dominant lineages for several decades. Recent sporadic disease outbreaks were primarily caused by Xoo isolates derived from the two major lineages, CX-5 and CX-6, although Xoo isolates from other lineages also contributed to these outbreaks. The lineage and sub-lineage distributions of Xoo isolates were strongly correlated with their geographical origin, which was found to be mainly determined by the planting of the two major rice subspecies, indica and japonica. Moreover, large-scale virulence testing was conducted to evaluate the diversity of pathogenicity for Xoo. We found rapid virulence evolution against rice, and its determinant factors included the genetic background of Xoo, rice resistance genes, and planting environment of rice. This study provides an excellent model for understanding the evolution and dynamics of plant pathogens in the context of their interactions with their hosts, which are shaped by a combination of geographical conditions and farming practices. The findings of this study may have important implications for the development of effective strategies for disease management and crop protection in rice production systems.

Postbiotics from Pichia kudriavzevii promote intestinal health performance through regulation of Limosilactobacillus reuteri in weaned piglets.

Postbiotics are attractive as alternatives to antibiotics for use against post-weaning diarrhea. However, their beneficial mechanisms are largely unknown. In the current study, we first demonstrated that supplementation with 0.5% Pichia kudriavzevii FZ12 postbiotics in the diet significantly reduced diarrhea incidence, promoted growth performance, improved gut health performance, and significantly enriched beneficial bacteria, particularly Lactobacillus spp., in the intestines of weaned piglets. Importantly, we identified a heat- and proteinase K-sensitive component, cytochrome c, of the postbiotics that significantly promoted the growth and biofilm formation of Limosilactobacillus reuteri FP13. We demonstrated the importance of P. kudriavzevii FZ12 postbiotics in improving the intestinal health of a model animal and revealed that cytochrome c is one of the important components of yeast postbiotics. These findings may provide new insights into microbe-postbiotics interplay that can be applied to guidelines for dietary modulation to alleviate weaning-induced diarrhea.

A phylogenomic analysis of Limosilactobacillus reuteri reveals ancient and stable evolutionary relationships with rodents and birds and zoonotic transmission to humans.

BACKGROUND: Gut microbes play crucial roles in the development and health of their animal hosts. However, the evolutionary relationships of gut microbes with vertebrate hosts, and the consequences that arise for the ecology and lifestyle of the microbes are still insufficiently understood. Specifically, the mechanisms by which strain-level diversity evolved, the degree by which lineages remain stably associated with hosts, and how their evolutionary history influences their ecological performance remain a critical gap in our understanding of vertebrate-microbe symbiosis. RESULTS: This study presents the characterization of an extended collection of strains of Limosilactobacillus reuteri and closely related species from a wide variety of hosts by phylogenomic and comparative genomic analyses combined with colonization experiments in mice to gain insight into the long-term evolutionary relationship of a bacterial symbiont with vertebrates. The phylogenetic analysis of L. reuteri revealed early-branching lineages that primarily consist of isolates from rodents (four lineages) and birds (one lineage), while lineages dominated by strains from herbivores, humans, pigs, and primates arose more recently and were less host specific. Strains from rodent lineages, despite their phylogenetic divergence, showed tight clustering in gene-content-based analyses. These L. reuteri strains but not those ones from non-rodent lineages efficiently colonize the forestomach epithelium of germ-free mice. The findings support a long-term evolutionary relationships of L. reuteri lineages with rodents and a stable host switch to birds. Associations of L. reuteri with other host species are likely more dynamic and transient. Interestingly, human isolates of L. reuteri cluster phylogenetically closely with strains from domesticated animals, such as chickens and herbivores, suggesting zoonotic transmissions. CONCLUSIONS: Overall, this study demonstrates that the evolutionary relationship of a vertebrate gut symbiont can be stable in particular hosts over time scales that allow major adaptations and specialization, but also emphasizes the diversity of symbiont lifestyles even within a single bacterial species. For L. reuteri, symbiont lifestyles ranged from autochthonous, likely based on vertical transmission and stably aligned to rodents and birds over evolutionary time, to allochthonous possibly reliant on zoonotic transmission in humans. Such information contributes to our ability to use these microbes in microbial-based therapeutics.

Enrofloxacin-induced transfer of multiple-antibiotic resistance genes and emergence of novel resistant bacteria in red swamp crayfish guts and pond sediments.

Antibiotic resistance genes (ARGs) can be transferred from environmental microbes to human pathogens, thus leading to bacterial infection treatment failures. The aquaculture polluted by over-used antibiotics is considered as a notorious reservoir of ARGs. However, the origin, diachronic changes, and mobility of ARGs under antibiotic exposure in aquaculture systems remain elusive. Our findings showed that enrofloxacin application also increased the relative abundance of various ARGs in addition to quinolone-resistance genes and induced ARG dissemination in crayfish gut and sediment bacteria. Further investigation indicated that the transposase-mediated recombination was the major driver of horizontal gene transfer (HGT) of ARGs under antibiotic stress. Notably, enrofloxacin application also induced the generation of some metagenome-assembled genomes (MAGs) carrying multiple ARGs, which were identified as novel species. Additionally, Enterobacteriaceae constituted a mobile ARG pool in aquaculture. Therefore, aquaculture provides potential wide environmental pathways for generation and spread of antibiotic resistance. Our findings of ARG temporal variations and dissemination pattern in aquaculture with artificial use of antibiotics are critical to the management of antibiotic resistance, which is of great ecosystem and health implications.

After the storm-Perspectives on the taxonomy of Lactobacillaceae.

In 2020, a taxonomic reorganization of the lactic acid bacteria reclassified over 300 species in 7 genera and 2 families into one family, the Lactobacillaceae, with 31 genera including 23 new genera to include organisms formerly classified as Lactobacillus species. This communication aims to provide a debrief on the taxonomic reorganization of lactobacilli to identify shortcomings in the proposed taxonomic framework, and to outline perspectives and opportunities provided by the current taxonomy of the Lactobacillaceae. The current taxonomy of lactobacilli not only necessitates becoming familiar with 23 new genus names but also provides substantial new opportunities in scientific discovery and regulatory approval of these organisms. First, description of new species in the Lactobacillaceae is facilitated and a solid framework for description of novel genera is provided. Second, the current taxonomy greatly enhances the resolution of genus-level sequencing approaches (e.g., 16S rRNA-based metagenomics) when identifying the composition and function of microbial communities. Third, the current taxonomy greatly facilitates the formulation of hypotheses linking phylogeny to metabolism and ecology of lactobacilli.

The Microbiome Structure of a Rice-Crayfish Integrated Breeding Model and Its Association with Crayfish Growth and Water Quality.

The rice-crayfish (RC) integrated breeding model is an important and special agricultural ecosystem that provides a unique ecological environment for exploring the microbial diversity, composition, and functional capacity. To date, little is known about the effect of the breeding model on microbiome assembly and breeding model-specific microbiome composition and the association of the microbiome with water quality and crayfish growth. In the present study, we assessed the taxonomic shifts in gut and water microbiomes and their associations with water quality and crayfish growth in the RC and crayfish monoculture (CM) breeding models across six time points of rice growth, including seedling (a), tillering and jointing (b), blooming (c), filling (d), fruiting (e), and rotting of rice residues (f). Dominant bacterial phyla, such as Proteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes, were detected in both gut and water microbiomes across breeding models. Notably, the diversity and structure of the gut and water microbiomes significantly (P < 0.001) differed between the RC and CM models, with higher microbial diversity being noted in the RC model than in the CM model. The taxa enriched in the RC model included Bacillus sp., Streptomyces sp., Lactobacillus sp., Prevotella sp., Rhodobacter sp., Bifidobacterium sp., Akkermansia sp., and Lactococcus sp., some of which are potentially beneficial to animals. However, opportunistic pathogens, such as Citrobacter sp. and Aeromonas sp., were depleted in the RC model. Furthermore, in the RC model, the enriched taxa that formed complex cooccurrence networks showed a significant positive correlation with water quality and crayfish growth, whereas the depleted taxa showed a significant negative correlation with water quality and crayfish growth. These results suggest that the RC model has a better microbiome composition and that RC model-specific microbes could play important roles in improving crayfish growth and water quality. IMPORTANCE The present study comprehensively compared two different breeding models in terms of their microbiome composition and the associations of the microbiomes with crayfish growth and water quality. RC model-specific microbiome composition was identified; these microbes were found to have a positive association with water quality and crayfish growth. These results provide valuable information for guiding microbial isolation and culture and for potentially harnessing the power of the microbiome to improve crayfish production and health and water quality.

Enhanced protein degradation by black soldier fly larvae (Hermetia illucens L.) and its gut microbes.

Black soldier fly larvae (BSFL) can convert a variety of organic wastes into biomass, and its gut microbiota are involved in this process. However, the role of gut microbes in the nutrient metabolism of BSFL is unclear. In this study, germ-free BSFL (GF) and gnotobiotic BSFL (GB) were evaluated in a high-protein artificial diet model. We used 16S rDNA sequencing, ITS1 sequencing, and network analysis to study gut microbiota in BSFL that degrade proteins. The protein reduction rate of the GB BSFL group was significantly higher (increased by 73.44%) than that of the GF BSFL group. The activity of gut proteinases, such as trypsin and peptidase, in the GB group was significantly higher than the GF group. The abundances of different gut microbes, including Pseudomonas spp., Orbus spp. and Campylobacter spp., were strongly correlated with amino acid metabolic pathways. Dysgonomonas spp. were strongly correlated with protein digestion and absorption. Issatchenkia spp. had a strong correlation with pepsin activity. Campylobacter spp., Pediococcus spp. and Lactobacillus spp. were strongly correlated with trypsin activity. Lactobacillus spp. and Bacillus spp. were strongly correlated with peptidase activity. Gut microbes such as Issatchenkia spp. may promote the gut proteolytic enzyme activity of BSFL and improve the degradation rate of proteins. BSFL protein digestion and absorption involves gut microbiota that have a variety of functions. In BSFL the core gut microbiota help complete protein degradation. These results demonstrate that core gut microbes in BSFL are important in protein degradation.

Deciphering Succession and Assembly Patterns of Microbial Communities in a Two-Stage Solid-State Fermentation System.

Although the importance of microbiota in the natural environment and in industrial production has been widely recognized, little is known about the formation and succession patterns of the microbial community, particularly secondary succession after disturbance. Here, we choose the Xiaoqu liquor brewing process as an experimental model in which sorghum grains were first aerobically saccharified and then anaerobically fermented after being stirred and acidified to explore multistage community succession patterns. We analyzed microbial composition, physicochemical factors, and metabolites of brewing grains inoculated with two different starters, pure starter and traditional starter, respectively. Two groups showed similar succession patterns where the saccharification microbiota was mainly derived from starters, while environmental microorganisms, mainly Lactobacillaceae and Saccharomyces, dominated the fermentation microbiota regardless of the original saccharification community composition. Species replacement shaped the bacterial community, while species replacement and loss both contributed to fungal community succession in both groups. Grain acidification and hypoxia led to the succession of bacterial and fungal communities during fermentation, respectively. Despite inoculation with starters containing different microorganisms, similar microbial communities during the fermentation stage of the two groups exhibited similar metabolite composition. However, higher abundance of Rhizopus in the saccharification of the pure starter group led to more alcohols, while higher abundance of Monascus and Saccharomycopsis in the traditional starter group promoted acid and ester metabolism. These results revealed the microbial succession patterns of two-stage liquor brewing and its influence on flavor metabolism, which could be used to regulate the microbial community in food fermentation to further promote the modernization of the fermented food industry. IMPORTANCE Revealing formation and assembly mechanisms of microbiota can help us to understand and further regulate its roles in the ecosystems. The Xiaoqu liquor brewing system is a tractable microbial ecosystem with low complexity. This two-stage microbial ecosystem can be used as an experimental model to analyze the multistage temporal succession pattern of microbial communities. Our results demonstrated the dynamic composition and succession pattern of a microbial community in the two-stage liquor brewing system. The results also revealed the microbial origins determining community composition, the ecological processes dominating microbial community succession patterns, the determinants affecting microbial community successions, and the effect of microbial community changes on metabolite synthesis. Overall, our study not only provides an insight into multistage succession patterns of microbial communities in liquor brewing systems but also provides reference for optimizing the quality of fermented products, which will be helpful to understand the succession patterns of microbial communities in other natural ecosystems.

Ecology and Function of the Transmissible Locus of Stress Tolerance in Escherichia coli and Plant-Associated Enterobacteriaceae.

The transmissible locus of stress tolerance (tLST) is a genomic island which confers resistance to heat and chlorine. In this study, we determined that the tLST is frequent in genomes of those Enterobacteriaceae that occur in association with plants as well as the intestines of humans and animals and are relevant as nosocomial pathogens, e.g., Klebsiella and Cronobacter species. The tLST is more frequent in environmental and clinical isolates of Klebsiella pneumoniae than in animal isolates, and heat and chlorine resistance of tLST-positive strains of K. pneumoniae matched the resistance of tLST-positive strains of Escherichia coli. The function of 13 tLST genes was determined by assessing the heat and chlorine resistance of E. coli MG1655 mutants. The deletion of sHsp20, clpKGI, sHspGI, pscA, pscB, and hdeDGI reduced both heat and chlorine resistance; deletion of kefB reduced only chlorine resistance. Genes coding for heat shock proteins sHsp20, clpKGI, and sHspGI decreased the oxidation of cytoplasmic proteins, while kefB decreased the oxidation of membrane lipids. The fitness cost of the tLST for E. coli MG1655 was assessed by pairwise competition experiments with isogenic tLST-positive or tLST-negative strains. The tLST imposes a fitness cost that is compensated for by frequent and lethal challenges with chlorine. All core genes need to be present to maintain the ecological advantage relative to the fitness cost. Taken together, core tLST genes are necessary to provide protection for E. coli against heat and chlorine stress, and the selective pressure for the tLST maintains core genes. IMPORTANCE The transmissible locus of stress tolerance (tLST) is a genomic island comprising 10 core genes that occurs in diverse Enterobacteriaceae and confers resistance to heat and chlorine. Experimentation described in the manuscript describes the physiological function of the core genes by characterization of the resistance of 13 single-knockout (KO) mutants and by characterization of protein and membrane oxidation in these strains after chlorine challenge. Results identify tLST resistance as a genomic island that is specific for those Enterobacteriaceae that occur in plant-associated habitats as well in the intestines of vertebrates. In addition, the ecological function of the genomic island was characterized by large-scale genomic analysis and competition experiments of wild-type and mutant strains. Results suggest that tLST-mediated resistance to chlorine may contribute to the persistence of nosocomial pathogens in hospitals.

Comparative Genomics of Exiguobacterium Reveals What Makes a Cosmopolitan Bacterium.

Although the strategies used by bacteria to adapt to specific environmental conditions are widely reported, fewer studies have addressed how microbes with a cosmopolitan distribution can survive in diverse ecosystems. Exiguobacterium is a versatile genus whose members are commonly found in various habitats. To better understand the mechanisms underlying the universality of Exiguobacterium, we collected 105 strains from diverse environments and performed large-scale metabolic and adaptive ability tests. We found that most Exiguobacterium members have the capacity to survive under wide ranges of temperature, salinity, and pH. According to phylogenetic and average nucleotide identity analyses, we identified 27 putative species and classified two genetic groups: groups I and II. Comparative genomic analysis revealed that the Exiguobacterium members utilize a variety of complex polysaccharides and proteins to support survival in diverse environments and also employ a number of chaperonins and transporters for this purpose. We observed that the group I species can be found in more diverse terrestrial environments and have a larger genome size than the group II species. Our analyses revealed that the expansion of transporter families drove genomic expansion in group I strains, and we identified 25 transporter families, many of which are involved in the transport of important substrates and resistance to environmental stresses and are enriched in group I strains. This study provides important insights into both the overall general genetic basis for the cosmopolitan distribution of a bacterial genus and the evolutionary and adaptive strategies of Exiguobacterium. IMPORTANCE The wide distribution characteristics of Exiguobacterium make it a valuable model for studying the adaptive strategies of bacteria that can survive in multiple habitats. In this study, we reveal that members of the Exiguobacterium genus have a cosmopolitan distribution and share an extensive adaptability that enables them to survive in various environments. The capacities shared by Exiguobacterium members, such as their diverse means of polysaccharide utilization and environmental-stress resistance, provide an important basis for their cosmopolitan distribution. Furthermore, the selective expansion of transporter families has been a main driving force for genomic evolution in Exiguobacterium. Our findings improve our understanding of the adaptive and evolutionary mechanisms of cosmopolitan bacteria and the vital genomic traits that can facilitate niche adaptation.

Exiguobacterium algae sp. nov. and Exiguobacterium qingdaonense sp. nov., two novel moderately halotolerant bacteria isolated from the coastal algae.

Two Gram-stain-positive, facultatively anaerobic, rod-shaped bacterial strains, S126T and S82T, were isolated from coastal algae of China. Strains S126T and S82T are halotolerant and could grow in the presence of 0-13% NaCl and 0-14% NaCl, respectively. The two strains shared 98.9% 16S rRNA gene sequence similarity with each other and 93.4-99.8% similarity with type strains of Exiguobacterium species. The major fatty acids (> 10%) of strains S126T and S82T were iso-C17:0, iso-C13:0, anteiso-C13:0 and iso-C15:0. The predominant quinones of strains S126T and S82T were MK-7 and MK-8. The polar lipid profiles of strain S126T and S82T contained diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The cell-wall peptidoglycans of both strains S126T and S82T were of the A3α L-Lys-Gly type. The average nucleotide identity (ANI) and average nucleotide index (AAI) between strains S126T and S82T and type strains of Exiguobacterium species were all below the thresholds to discriminate bacterial species, indicating that they constitute two novel species in the genus Exiguobacterium. Based on polyphasic taxonomy characterization and genomic aspects, the names Exiguobacterium algae sp. nov. and Exiguobacterium qingdaonense sp. nov. are proposed for the two novel species, with type strains being S126T (= CGMCC 1.17116T = KCTC 43079 T) and S82T (= CGMCC 1.17115T = KCTC 43078T), respectively.