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Cordyceps militaris is a filamentous fungus used for both medicinal and culinary purposes. It exhibits a wide range of pharmacological activities due to its valuable contents of cordycepin, polysaccharides, carotenoids, terpenoids and other metabolites. However, C. militaris strains are highly susceptible to irreversible degradation in agricultural production, which is often manifested as a prolonged color change period and a significant decrease in the production of secondary metabolites. UDP-glycosyltransferases are an important enzyme family that participates in the synthesis of terpenoids by performing the glycosylation of key residues of enzymes or molecules. However, few studies have focused on its effect on the regulation of metabolite production in C. militaris. Therefore, in this study, we performed transcriptome analysis across four different developmental stages of C. militaris to target the putative glycosyltransferase gene CmUGT1, which plays important roles in metabolite production. We further constructed and screened a CmUGT1-overexpressing strain by Agrobacterium tumefaciens-mediated infestation of C. militaris spores. The major metabolite production of the wild-type and CmUGT1-overexpressing C. militaris strains was determined after short-term shake-flask cultivation of mycelia. The results showed that the yields of carotenoids and polysaccharides in the mycelia of the CmUGT1-overexpressing strains were 3.8 and 3.4 times greater than those in the mycelia of the wild type, respectively (p

ObjectiveTo explore the effects of probiotic fermentation products of germinated grains on cognitive and sleep improvement in mice with sleep deprivation induced by chlorophenylalanine (PCPA), and to provide theoretical and experimental basis for the development of natural products to alleviate insomnia.MethodsELISA and high-performance liquid chromatography (HPLC) were used to determine the contents of γ-aminobutyric acid and L-theanine in fermentation products. Open Field Test was used to analyze the changes of emotional behavior between groups before and after intervention. ELISA was used to analyze the changes of hypothalamic serotonin, GABA, glutamate, and serum interleukin 6. 16S rRNA sequencing was used to analyze the changes of intestinal flora before and after the intervention of compound fermentation products. LC–MS/MS was used to analyze the changes of intestinal SCFAs before and after the intervention.ResultsThe content of GABA and L-theanine in 7 L fermentation products was 12.555 μmol/L (1.295 mg/L) and 0.471 mg/mL by ELISA. Compared with the PCPA-induced Model group, the sleep duration of the KEY group was statistically significant (p < 0.0001). Compared with the PCPA-induced Model group, the number of crossing the central lattice in the KEY group was significantly increased, and the number of grooming was significantly reduced (all p

Exploring the bacterial community in the S. glauca rhizosphere was of great value for understanding how this species adapted to the saline-alkali environment and for the rational development and use of saline-alkali soils. In this study, high-throughput sequencing technology was used to investigate the diversity characteristics and distribution patterns of soil bacterial communities in the rhizosphere of S.glauca-dominated communities in the Hetao Irrigation Distract, Inner Mongolia, China. The relationships among bacterial characteristics, soil physicochemical properties and vegetation in four sampling sites were analyzed. The soil bacterial communities in the rhizosphere of S. glauca-dominated communities were mainly composed of 16 phyla (i.e., Proteobacteria, Actinobacteria, Bacteroidetes, Gemmatimonadetes, Chloroflexi, Acidobacteria, Firmicutes, Planctomycetes, Deinococcus-Thermus, Verrucomicrobia, Saccharibacteria, Cyanobacteria, Nitrospirae, JL-ETNP-Z39, Parcubacteria and Chlorobi), and these populations accounted for more than 99% of the total bacterial community. At the genus level, the main bacterial communities comprised Halomonas, Nitriliruptor, Euzebya and Pelagibius, which accounted for 15.70% of the total bacterial community. An alpha diversity analysis indicated that the richness and diversity of rhizosphere soil bacteria differed significantly among the sampling sites, and the bacterial richness and diversity indices of severe saline-alkali land were higher than those of light and moderate saline-alkali land. The principal component analysis (PCA) and linear discriminant analysis effect size (LEfSe) showed significant differences in the species composition of the rhizosphere soil bacterial community among different sampling sites. A correlation analysis showed that the number of bacterial species exhibited the highest correlation with the soil water content (SWC). The richness and evenness indices were significantly correlated with the SWC and SO42–, K+ and Mg2+ concentrations. The electrical conductivity (EC), soluble ions (Na+, CO32– + HCO3–, K+, Ca2+, Mg2+, and SO42+), SWC and vegetation coverage (VC) were the main drivers affecting the changes in its community structure. The bacterial community in the rhizosphere of S. glauca enhanced the adaptability of S. glauca to saline-alkali environment by participating in the cycling process of nutrient elements, the decomposition of organic matter and the production of plant growth regulating substances. These results provided a theoretical reference for further study on the relationship among rhizosphere soil microorganisms and salt tolerance in halophytes.

IntroductionAs a symbiotic probiotic for the host, Clostridium butyricum (CB) has the potential to strengthen the body’s immune system and improve intestinal health. However, the probiotic mechanism of CB is not completely understood. The Clostridium butyricum CBX 2021 strain isolated by our team from a health pig independently exhibits strong butyric acid production ability and stress resistance. Therefore, this study comprehensively investigated the efficacy of CBX 2021 in pigs and its mechanism of improving pig health.MethodsIn this study, we systematically revealed the probiotic effect and potential mechanism of the strain by using various methods such as microbiome, metabolites and transcriptome through animal experiments in vivo and cell experiments in vitro.ResultsOur in vivo study showed that CBX 2021 improved growth indicators such as daily weight gain in weaned piglets and also reduced diarrhea rates. Meanwhile, CBX 2021 significantly increased immunoglobulin levels in piglets, reduced contents of inflammatory factors and improved the intestinal barrier. Subsequently, 16S rRNA sequencing showed that CBX 2021 treatment implanted more butyric acid-producing bacteria (such as Faecalibacterium) in piglets and reduced the number of potentially pathogenic bacteria (like Rikenellaceae RC9_gut_group). With significant changes in the microbial community, CBX 2021 improved tryptophan metabolism and several alkaloids synthesis in piglets. Further in vitro experiments showed that CBX 2021 adhesion directly promoted the proliferation of a porcine intestinal epithelial cell line (IPEC-J2). Moreover, transcriptome analysis revealed that bacterial adhesion increased the expression of intracellular G protein-coupled receptors, inhibited the Notch signaling pathway, and led to a decrease in intracellular pro-inflammatory molecules.DiscussionThese results suggest that CBX 2021 may accelerate piglet growth by optimizing the intestinal microbiota, improving metabolic function and enhancing intestinal health.

Akkermansia muciniphila (A. muciniphila), an intestinal symbiont residing in the mucosal layer, shows promise as a probiotic. Our previous study found that the abundance of A. muciniphila was significantly higher in Ningxiang suckling piglets compared to other breeds, suggesting that early breast milk may play a crucial role. This study examines A. muciniphila’s ability to utilize Ningxiang pig milk oligosaccharides. We discovered that A. muciniphila can thrive on both Ningxiang pig colostrum and purified pig milk oligosaccharides. Genetic analysis has shown that A. muciniphila harbors essential glycan-degrading enzymes, enabling it to effectively break down a broad spectrum of oligosaccharides. Our findings demonstrate that A. muciniphila can degrade pig milk oligosaccharides structures such as 3′-FL, 3′-SL, LNT, and LNnT, producing short-chain fatty acids in the process. The hydrolysis of these host-derived glycan structures enhances A. muciniphila’s symbiotic interactions with other beneficial gut bacteria, contributing to a dynamic microbial ecological network. The capability of A. muciniphila to utilize pig milk oligosaccharides allows it to establish itself in the intestines of newborn piglets, effectively colonizing the mucosal layer early in life. This early colonization is key in supporting both mucosal and metabolic health, which is critical for enhancing piglet survival during lactation.

Endophytic bacteria have a complex coevolutionary relationship with their host macroalgae. Dioecious macroalgae are important producers in marine ecosystems, but there is still a lack of research on how sex influences their endophytic bacteria. In this study, the endophytic bacterial communities in male and female S. thunbergii and their reproductive tissues (receptacles) were compared using culture methods and high-throughput sequencing. The endophytic bacterial communities detected by the two methods were different. Among the 78 isolated strains, the dominant phylum, genus, and species were Bacillota, Alkalihalobacillus, and Alkalihalobacillus algicola, respectively, in the algal bodies, while in the receptacles, they were Bacillota, Vibrio, and Vibrio alginolyticus. However, 24 phyla and 349 genera of endophytic bacteria were identified by high-throughput sequencing, and the dominant phylum and genus were Pseudomonadota and Sva0996_ Marine_ Group, respectively, in both the algal body and the receptacles. The two methods showed similar compositions of endophytic bacterial communities between the samples of different sexes, but the relative abundances of dominant and specific taxa were different. The high-throughput sequencing results showed more clearly that the sex of the host alga had an effect on its endophyte community assembly and a greater effect on the endophytic bacterial community in the receptacles. Moreover, most specific bacteria and predicted functional genes that differed between the samples from the males and females were related to metabolism, suggesting that metabolic differences are the main causes of sex differences in the endophytic bacterial community. Our research is the first to show that host sex contributes to the composition of endophytic bacterial communities in dioecious marine macroalgae. The results enrich the database of endophytic bacteria of dioecious marine macroalgae and pave the way for better understanding the assembly mechanism of the endophytic bacterial community of algae.

Antibiotic resistance in Enterococcus faecium, Enterococcus faecalis, and Staphylococcus aureus remains a major public health concern worldwide. Furthermore, these microbes frequently co-exist in biofilm-associated infections, largely nullifying antibiotic-based therapy. Therefore, it is imperative to develop an efficient therapeutic strategy for combating infections caused by polymicrobial biofilms. In this study, we investigated the antibacterial and antibiofilm activity of the bacteriophage endolysin Ply113 in vitro. Ply113 exhibited high and rapid lytic activity against E. faecium, E. faecalis, and S. aureus, including vancomycin-resistant Enterococcus and methicillin-resistant S. aureus isolates. Transmission electron microscopy revealed that Ply113 treatment led to the detachment of bacterial cell walls and considerable cell lysis. Ply113 maintained stable lytic activity over a temperature range of 4–45°C, over a pH range of 5.0–8.0, and in the presence of 0–400 mM NaCl. Ply113 treatment effectively eliminated the mono-species biofilms formed by E. faecium, E. faecalis, and S. aureus in a dose-dependent manner. Ply113 was also able to eliminate the dual-species biofilms of E. faecium–S. aureus and E. faecalis–S. aureus. Additionally, Ply113 exerted potent antibacterial efficacy in vivo, distinctly decreasing the bacterial loads in a murine peritoneal septicemia model. Our findings suggest that the bacteriophage endolysin Ply113 is a promising antimicrobial agent for the treatment of polymicrobial infections.

Regulated cell death (RCD) is a strategy employed by host cells to defend invasions of pathogens, such as viruses and bacteria. Ferroptosis is a type of RCD characterized by excessive accumulation of iron and lipid peroxidation. While ferroptosis is primarily considered as a mechanism associated with tumorigenesis, emerging evidence begin to suggest that it may play essential role during virus infections. Recent studies illustrated that activation of ferroptosis could either induce or prohibit various types of RCDs to facilitate virus replication or evade host surveillance. More experimental evidence has demonstrated how viruses regulate ferroptosis to influence replication, transmission, and pathogenesis. This review summarizes ferroptosis-related metabolism, including iron metabolism, lipid peroxidation, and antioxidant metabolism. Furthermore, we discuss the interplay between viral infections and host ferroptosis process, with a focus on the mechanism of how viruses exploit ferroptosis for its own replication. Understanding how ferroptosis impacts virus infection can offer valuable insights into the development of effective therapeutic strategies to combat virus infections.

Heavy metal contamination from coal mining calls for advanced bioremediation, i.e., using sulfate-reducing bacteria (SRB) technology. Yet, the interaction of SRB with native soil microbiota during metal sequestration, especially in the presence of plants, remains ambiguous. In this study, we assessed the metal sequestration capabilities, ecological network interactions, and enzymatic functions in soils treated with a predominant SRB consortium, mainly Desulfovibrio (14 OTUs, 42.15%) and Desulfobulbus (7 OTUs, 42.27%), alongside Acacia dealbata (AD) and Pisum sativum (PS) plants. The SRB consortium notably enhanced the immobilization of metals such as Zn, Cu, As, and Pb in soil, with the conversion of metals to residual forms rising from 23.47 to 75.98%. Plant inclusion introduced variability, potentially due to changes in root exudates under metal stress. While AD flourished, PS demonstrated significant enhancement in conjunction with SRB, despite initial challenges. Comprehensive microbial analyses revealed the pivotal role of SRB in influencing microbial networking, underpinning critical ecological links. This interplay between plants and SRB not only enhanced microbial diversity but also enriched soil nutrients. Further, enzymatic assessments, highlighting enzymes like NADH:ubiquinone reductase and non-specific serine/threonine protein kinase, reinforced contribution of SRB to energy metabolism and environmental resilience of the entire soil microbial community. Overall, this research underscores the potential of SRB-driven bioremediation in revitalizing soils affected by coal mining.

Smallpox is an infectious disease caused by the variola virus, and it has a high mortality rate. Historically it has broken out in many countries and it was a great threat to human health. Smallpox was declared eradicated in 1980, and Many countries stopped nation-wide smallpox vaccinations at that time. In recent years the potential threat of bioterrorism using smallpox has led to resumed research on the treatment and prevention of smallpox. Effective ways of preventing and treating smallpox infection have been reported, including vaccination, chemical drugs, neutralizing antibodies, and clinical symptomatic therapies. Antibody treatments include anti-sera, murine monoclonal antibodies, and engineered humanized or human antibodies. Engineered antibodies are homologous, safe, and effective. The development of humanized and genetically engineered antibodies against variola virus via molecular biology and bioinformatics is therefore a potentially fruitful prospect with respect to field application. Natural smallpox virus is inaccessible, therefore most research about prevention and/or treatment of smallpox were done using vaccinia virus, which is much safer and highly homologous to smallpox. Herein we summarize vaccinia virus epitope information reported to date, and discuss neutralizing antibodies with potential value for field application.

The widespread and continuous cultivation of tobacco has led to soil degradation and reduced crop yields and quality. Green manure is an essential organic fertilizer that alleviates obstacles to continuous cultivation. However, the plant–soil microecological effects of green manure on flue-cured tobacco cultivation remain unclear. Thus, a positioning trail including two treatments, chemical fertilizer application only (treatment NPK) and chemical fertilizer application with turning ryegrass (treatment NPKG) was conducted, and the effect of ryegrass returning on the soil physicochemical properties, soil microbiome, crop yield, and quality of flue-cured tobacco in continuous cropping soil were investigated. Results showed that returning ryegrass to the field increased the thickness of soil humus layer from 13 cm to 15 cm, reduced the humus layer soil bulk density to 1.29 cm3/g. Ryegrass tilled and returned to the field increased soil organic matter content by 6.89–7.92%, increased rhizosphere soil available phosphorus content by 2.22–17.96%, and converted the soil non-exchangeable potassium into potassium that was available for plant absorption and utilization. Ryegrass tilling and returning to the field increased the potassium content of middle leaves of flue-cured tobacco by 7.69–10.07%, the increased potassium content in flue-cured tobacco was accompanied by increased total sugar, reducing sugar, and the ratio of reducing sugar to nicotine, which facilitated the harmonization of the chemical composition of cured tobacco leaves. Moreover, the increased number of markedly improved operational taxonomic units enhanced the complexity of the soil bacterial community and its compactness after ryegrass tillage and their return to the field. The available potassium, available phosphorus, total potassium content, pH, and sampling period of the rhizosphere soil had considerable effects on the rhizosphere microbial. Ryegrass tilling and returning to the field changed the soil microbiome, which increased the abundance of bulk soil Proteobacteria, rhizosphere soil Fibrobacterota, and microbes with anti-pathogen activity (Lysobacteria, Sphingomonas, Chaetomium, and Minimedusa); and reduced the abundance of pathogenic fungi Neocosmospore genus in the soil. In brief, ryegrass returned to the field, improved soil microecology and restored soil nutrients, and established a new dynamic balance of soil ecology, thereby improving the quality of cultivated land and the quality of flue-cured tobacco.

The broad-spectrum antimicrobial activity of Elsholtzia ciliate essential oil (ECO) has been previously reported, but its effectiveness against halitosis-causing bacteria such as Fusobacterium nucleatum and Porphyromonas gingivalis is not well understood. In this study, we investigated the bacteriostatic activity of ECO against planktonic cells and biofilms of F. nucleatum and P. gingivalis, as well as its ability to inhibit bacterial metabolism and production of volatile sulfur compounds (VSCs) at sub-lethal concentrations. Our findings revealed that ECO exhibited comparable activities to chlorhexidine against these oral bacteria. Treatment with ECO significantly reduced the production of VSCs, including hydrogen sulfide, dimethyl disulfide, and methanethiol, which are major contributors to bad breath. As the major chemical components of ECO, carvacrol, p-cymene, and phellandrene, were demonstrated in vitro inhibitory effects on F. nucleatum and P. gingivalis, and their combined use showed synergistic and additive effects, suggesting that the overall activity of ECO is derived from the cumulative or synergistic effect of multiple active components. ECO was found to have a destructive effect on the bacterial cell membrane by examining the cell morphology and permeability. Furthermore, the application of ECO induced significant changes in the bacterial composition of saliva-derived biofilm, resulting in the elimination of bacterial species that contribute to halitosis, including Fusobacterium, Porphyromonas, and Prevotella. These results provide experimental evidence for the potential clinical applications of ECOs in the prevention and treatment of halitosis.

Coral-associated microbial communities play a vital role in underpinning the health and resilience of reef ecosystems. Previous studies have demonstrated that the microbial communities of corals are affected by multiple factors, mainly focusing on host species and geolocation. However, up-to-date, insight into how the coral microbiota is structured by vast geographic distance with rich taxa is deficient. In the present study, the coral microbiota in six stony coral species collected from the coastal area of three countries, including United States of America (USA), Australia and Fiji, was used for analysis. It was found that the geographic influence on the coral microbiota was stronger than the coral host influence, even though both were significant. Interestingly, the contribution of the deterministic process to bacterial community composition increased as geographical distance grew. A total of 65 differentially abundant features of functions in coral microbial communities were identified to be associated with three geolocations. While in the same coastal area of USA, the similar relationship of coral microbiota was consistent with the phylogenetic relationship of coral hosts. In contrast to the phylum Proteobacteria, which was most abundant in other coral species in USA, Cyanobacteria was the most abundant phylum in Orbicella faveolata. The above findings may help to better understand the multiple natural driving forces shaping the coral microbial community to contribute to defining the healthy baseline of the coral microbiome.

The emergence and spread of carbapenemase genes, colistin resistance genes mcr-1, and tigecycline resistance gene tet(X) represent a significant threat to clinical therapy and public health. In this study, we investigated the presence of carbapenemase genes, mcr-1, and tet(X) in 298 Escherichia coli strains obtained from a teaching hospital in China. In total, eight (2.68%), six (2.01%), and one (0.34%) E. coli isolates carried blaNDM, mcr-1, and tet(X4), respectively. The blaNDM gene was located on IncX3 (n = 4), F2:A-:B- (n = 3), and F2:A1:B1 (n = 1) plasmids, with high similarity to multiple plasmids belonging to the same incompatibility type from Enterobacteriaceae. Six MCR-producing strains contained mcr-1-carrying IncI2 plasmids, organized similarly to other mcr-1-bearing IncI2 plasmids from animals in China. The blaCTX−M−55/64/132/199 gene located within a typical transposition unit (ISEcp1-blaCTX−M-orf477Δ) was inserted near dnaJ to generate 5-bp direct repeats in four mcr-1-positive plasmids. The tet(X) and another four resistance genes [aadA2, tet(A), floR, and Δlnu(F)] were co-located on an IncX1 plasmid, highly similar to other tet(X4)-carrying IncX1 plasmids from Escherichia and Klebsiella of animal or food origin, except that the conjugative transfer region of IncX1 plasmids was absent in our plasmid. Although a low prevalence of blaNDM, mcr-1, and tet(X) was observed in E. coli from patients in this study, their dissemination associated with some successful pandemic plasmids is of great concern. The continued surveillance of these crucial resistance genes in patients should be strengthened.

ObjectiveIncreasing evidence suggests that gut microbiota is involved in the occurrence and progression of urinary system diseases such as clear cell renal cell carcinoma (ccRCC). However, the mechanism of how alteration of gut metagenome promotes ccRCC remains unclear. Here we aim to elucidate the association of specific gut bacteria and their metabolites with ccRCC.MethodsIn a pilot case-control study among 30 ccRCC patients (RCC group) and 30 healthy controls (Control group), 16S ribosomal RNA (rRNA) gene sequencing were analyzed from fecal samples collected prior to surgery or hospitalization. Alpha diversity and beta diversity analysis of the gut microbiota were performed, and differential taxa were identified by multivariate statistics. Meanwhile, serum metabolism was measured by UHPLC-MS, and differential genes were identified based on the TCGA database.ResultsAlpha diversity found there were no significant microbial diversity differences of gut microbiota between the RCC group and the Control group. However, beta diversity analysis showed that the overall structures of the two groups were significantly separated (p = 0.008). Random Forests revealed the relative abundances of 20 species differed significantly between the RCC group and the Control group, among which nine species were enriched in the RCC group such as Desulfovibrionaceae, and 11 species were less abundant such as four kinds of Lactobacillus. Concomitantly, serum level of taurine, which was considered to be consumed by Desulfovibrionaceae and released by Lactobacillus, has decreased in the RCC group. In addition, macrophage-related genes such as Gabbr1 was upregulated in ccRCC patients.ConclusionReduction of protective bacteria, proliferation of sulfide-degrading bacteria Desulfovibrionaceae, reduction of taurine, and enrichment of macrophage related genes might be the risk predictors of ccRCC.

IntroductionThe human respiratory tract is considered to be a polymicrobial niche, and an imbalance in the microorganism composition is normally associated with several respiratory diseases. In addition to the well-studied bacteriome, the existence of fungal species in the respiratory tract has drawn increasing attention and has been suggested to have a significant clinical impact. However, the understanding of the respiratory fungal microbiota (mycobiome) in pulmonary diseases is still insufficient.MethodsIn this study, we investigated the fungal community composition of oropharynx swab (OS) samples from patients with five kinds of pulmonary disease, including interstitial lung disease (ILD), bacterial pneumonia (BP), fungal pneumonia (FP), asthma (AS) and lung cancer (LC), and compared them with healthy controls (HCs), based on high-throughput sequencing of the amplified fungal internal transcribed spacer (ITS) region.ResultsThe results showed significant differences in fungal composition and abundance between disease groups and HCs. Malassezia was the most significant genus, which was much more abundant in pulmonary diseases than in the control. In addition, many common taxa were shared among different disease groups, but differences in taxa abundance and specific species in distinct disease groups were also observed. Based on linear discriminant analysis effect size (LefSe), each group had its characteristic species. Furthermore, some species showed a significant correlation with the patient clinical characteristics.DiscussionOur study deepened our understanding of the respiratory tract mycobiome in some diseases that are less studied and identified the commonalities and differences among different kinds of pulmonary disease. These results would provide the solid basis for further investigation of the association between the mycobiome and pathogenicity of pulmonary diseases.

During the co-evolution of viruses and their hosts, viruses have developed various strategies for overcoming host immunological defenses so that they can proliferate efficiently. Porcine reproductive and respiratory syndrome virus (PRRSV), a significant virus to the swine industry across the world, typically establishes prolonged infection via diverse and complicated mechanisms, which is one of the biggest obstacles for controlling the associated disease, porcine reproductive and respiratory syndrome (PRRS). In this review, we summarize the latest research on how PRRSV circumvents host antiviral responses from both the innate and adaptive immune systems and how this virus utilizes other evasion mechanisms, such as the manipulation of host apoptosis and microRNA. A thorough understanding of the exact mechanisms of PRRSV immune evasion will help with the development of novel antiviral strategies against PRRSV.

Alfalfa is harvested two or three times a year in central and western Inner Mongolia, China. However, the variations in bacterial communities as affected by wilting and ensiling, and the ensiling characteristics of alfalfa among the different cuttings, are not fully understood. To enable a more complete evaluation, alfalfa was harvested three times a year. At each time of cutting, alfalfa was harvested at early bloom, wilted for 6 h, and then ensiled in polyethylene bags for 60 days. The bacterial communities and nutritional components of fresh alfalfa(F), wilted alfalfa(W) and ensiled alfalfa(S), and the fermentation quality and functional profile of bacterial communities of the three cuttings alfalfa silage, were then analyzed. Functional characteristics of silage bacterial communities were evaluated according to the Kyoto Encyclopedia of Genes and Genomes. The results showed that all nutritional components, fermentation quality, bacterial communities, carbohydrate, amino acid metabolism and key enzymes of bacterial communities were influenced by cutting time. The species richness of F increased from the first cutting to the third cutting; it was not changed by wilting, but was decreased by ensiling. At phylum level, Proteobacteria were more predominant than other bacteria, followed by Firmicutes (0.063–21.39%) in F and W in the first and second cuttings. Firmicutes (96.66–99.79%) were more predominant than other bacteria, followed by Proteobacteria (0.13–3.19%) in S in the first and second cuttings. Proteobacteria, however, predominated over all other bacteria in F, W, or S in the third cutting. The third-cutting silage showed the highest levels of dry matter, pH and butyric acid (p

Escherichia coli cysteine desulfurase (CD), IscS, modifies basal metabolism by transferring sulphur (S) from L-cysteine to numerous cellular pathways, whereas NFS1, a human CD, is active only in the formation of the [Acp]2:[ISD11]2:[NFS1]2 complex. Despite the accumulation of red-coloured IscS in E. coli cells as a result of the deficiency of accessible iron, as revealed in our previous studies, the mechanism of the potential enzymatic reaction remains unclear. In this study, the N-terminus of IscS was fused with the C-terminus of NFS1, which was reported to be almost fully active as IscS and exhibits a pyridoxal 5′-phosphate (PLP) absorption peak at 395 nm. Moreover, SUMO-EH-IscS exhibited significant growth recovery and NADH-dehydrogenase I activity in the iscS mutant cells. Furthermore, through in vitro and in vivo experiments combined with high-performance liquid chromatography and ultra-performance liquid chromatography–tandem mass spectrometry, it was shown that the new absorption peaks of the IscS H104Q, IscS Q183E, IscS K206A, and IscS K206A&C328S variants at 340 and 350 nm may correspond to the enzyme reaction intermediates, Cys-ketimine and Cys-aldimine, respectively. However, after mutation of the conserved active-site residues, additional absorption peaks at 420 and 430 nm were associated with PLP migration in the active-site pocket. Additionally, the corresponding absorption peaks of Cys-quinonoid, Ala-ketimine, and Ala-aldimine intermediates in IscS were 510, 325, and 345 nm, respectively, as determined by site-directed mutagenesis and substrate/product-binding analyses during the CD reaction process. Notably, red IscS formed in vitro by incubating IscS variants (Q183E and K206A) with excess L-alanine and sulphide under aerobic conditions produced an absorption peak similar to the wild-type IscS, at 510 nm. Interestingly, site-directed mutation of IscS with hydrogen bonds to PLP at Asp180 and Gln183 resulted in a loss of enzymatic activity followed by an absorption peak consistent with NFS1 (420 nm). Furthermore, mutations at Asp180 or Lys206 inhibited the reaction of IscS in vitro with L-cysteine (substrate) and L-alanine (product). These results suggest that the conserved active site residues (His104, Asp180, and Gln183) and their hydrogen bond with PLP in the N-terminus of IscS play a key role in determining whether the L-cysteine substrate can enter the active-site pocket and regulate the enzymatic reaction process. Therefore, our findings provide a framework for evaluating the roles of conserved active-site residues, motifs, and domains in CDs.

The present study was conducted to evaluate the effects of Clostridium butyricum (CB) and fructooligosaccharide (FOS) singly or combined, on performance, egg quality, amino acid digestibility, jejunal morphology, immune function and antioxidant capacity in peak-phase laying hens. A total of 288 Hy-Line Brown laying hens (30 weeks of age) were randomly assigned to 4 dietary groups that included basal diet, basal diet +0.02% of CB (zlc-17: 1 × 109 CFU/g) (PRO), basal diet +0.6% FOS (PRE), and basal diet +0.02% CB + 0.6% FOS (SYN) for 12 weeks. Each treatment had 6 replicates with 12 birds each. The results demonstrated that probiotics (PRO), prebiotics (PRE) and synbiotics (SYN) (p ≤ 0.05), respectively, exerted a positive effect on the performance and physiological response of the birds. There were significant increases in egg production rate, egg weight, egg mass, daily feed intake and reduced number of damaged eggs. and zero mortality rate due to dietary PRO, PRE and SYN (p ≤ 0.05) respectively. Also, feed conversion was improved by PRO (p ≤ 0.05). In addition, egg quality assessment showed that; eggshell quality was increased by PRO (p ≤ 0.05) and albumen indices (Haugh unit, thick albumen content, and albumen height) were enhanced by PRO, PRE and SYN (p ≤ 0.05). Further analysis showed that PRO, PRE and SYN (p ≤ 0.05), reduced heterophil to lymphocyte ratio, increased antioxidant enzymes and immunoglobulin concentration. Although spleen index was higher for PRO (p ≤ 0.05) group. The significant increase in villi height, villi width, villi height to crypt depth ratio and reduced crypt depth were obvious for PRO, PRE, and SYN (p ≤ 0.05). Furthermore, improved nutrient absorption and retention evidenced by increased digestibility of crude protein and amino acids, were notable for PRO, PRE, and SYN (p ≤ 0.05) group. Collectively, our findings revealed that dietary CB and FOS alone, or combined, enhanced productive performance, egg quality, amino acid digestibility, jejunal morphology, and physiological response in peak-phase laying hens. Our results would provide direction on nutritional strategies for gut enhancers and better physiological response of peak laying hens.

IntroductionSoil fumigant dazomet is a broad-spectrum nematicide and fungicide that can kill non-target microbes. Fungicides or organic fertilizers are often added after fumigation to improve the recovery of soil microbes. However, the effect of adding microbial organic fertilizers (MOF) after fumigation on the structure and function of rhizosphere soil microbial communities of crops is unclear.MethodsTherefore, we investigated the effects of adding Junweinong and Junlisu MOFs after dazomet fumigation on the structure and function of rhizosphere microbial communities and its relationship with soil properties and enzyme activities.Results and discussionThe results showed that the addition of these two MOFs after dazomet fumigation significantly reduced the rhizosphere soil available phosphorus, available potassium, organic matter content, and urease, alkaline phosphatase, and catalase activities, but increased the soil pH compared with the fumigation treatment. The application of MOFs after fumigation resulted in significant enrichment of bacteria such as Gaiella, norank_f_Vicinamibacteraceae, and Flavisolibacter and fungi such as Peroneutypa, Olpidium, and Microascus in the rhizosphere soil of the crop and increased the relative abundance of functional genes of 13 kinds of amino acids metabolism, pyruvate metabolism, TCA cycle, and pentose phosphate pathway as well as endophytic and epiphytic functional groups in the rhizosphere soil. In particular, NH4+-N, pH, and AK had the greatest effect on rhizosphere microorganisms. Overall, the addition of MOFs after fumigation promoted crop root nutrient uptake, enhanced rhizosphere soil microbial metabolism, allowed more beneficial communities to colonize the roots, and promoted soil microbiological health.

Salami is a kind of fermented meat product with rich nutrition and unique flavor. Because it is rich in fat, it is easy to oxidize to produce bad flavor. Compared with the way of adding artificial or natural antioxidants to reduce the degree of sausage oxidation, the antioxidant characteristics of developing the starter itself deserve more attention. In this study, firstly the antioxidant activities of 5 strains of yeast were measured in vitro, and then the mixture of yeast and Lactobacillus rhamnosus YL-1 was applied to fermented sausage model. The effect of the starter in the sausage model was investigated through physicochemical parameters, degree of fat oxidation, free fatty acid content, and though volatile flavor compound analysis, sensory evaluation and various indexes after storage were observed. Metagenomics was used to explore metabolic pathways, functional genes and key enzymes related to lipid oxidizing substances in sausage in yeast. The results showed that Wickerhamomyces anomalus Y12-3 and Y12-4 had strong tolerance to H2O2, and had higher SOD and CAT enzyme activities. The addition of yeast effectively reduced the material value of peroxidation value and active thiobarbiturate in salami. In flavor analysis, the content of flavor compounds associated with lipid oxidation, such as hexanal, heptanal, nonanal and (E)-2-decenal were significantly lower with the use of Debaryomyces hansenii Y4-1 and Y12-3. Meanwhile, the possible pathways of yeast metabolism of flavor substances related to lipid oxidation (mainly aldehydes) were discussed with the help of metagenomic techniques. According to the results of metagenomics, fatty acid degradation (ko00071) metabolic pathway was related to the degradation of aldehydes through aldehyde dehydrogenase, which was the potential key enzyme.

The roles of the microbe-gut-brain axis in metabolic homeostasis, development, and health are well-known. The hypothalamus integrates the higher nerve center system and functions to regulate energy balance, feeding, biological rhythms and mood. However, how the hypothalamus is affected by gut microbes in mammals is unclear. This study demonstrated differences in hypothalamic gene expression between the germ-free (GF) pigs and pigs colonized with gut microbiota (CG) by whole-transcriptome analysis. A total of 938 mRNAs, 385 lncRNAs and 42 miRNAs were identified to be differentially expressed between the two groups of pigs. An mRNA-miRNA-lncRNA competing endogenous RNA network was constructed, and miR-22-3p, miR-24-3p, miR-136-3p, miR-143-3p, and miR-545-3p located in the net hub. Gene function and pathway enrichment analysis showed the altered mRNAs were mainly related to developmental regulation, mitochondrial function, the nervous system, cell signaling and neurodegenerative diseases. Notably, the remarkable upregulation of multiple genes in oxidative phosphorylation enhanced the GF pigs’ hypothalamic energy expenditure. Additionally, the reduction in ATP content and the increase in carnitine palmitoyl transterase-1 (CPT1) protein level also confirmed this fact. Furthermore, the hypothalamic cell apoptosis rate in the CG piglets was significantly higher than that in the GF piglets. This may be due to the elevated concentrations of pro-inflammatory factors produced by gut bacteria. The obtained results collectively suggest that the colonization of gut microbes has a significant impact on hypothalamic function and health.

Long-interspersed element 1 (LINE-1) is an autonomous non-LTR retrotransposon. Its replication can cause mutation and rearrangement of host genomic DNA, which may result in serious genetic diseases. Host cells therefore developed defense strategies to restrict LINE-1 mobilization. In this study, we reported that CCHC-type zinc-finger protein ZCCHC3 can repress LINE-1 retrotransposition, and this activity is closely related to its zinc-finger domain. Further studies show that ZCCHC3 can post-transcriptionally diminish the LINE-1 RNA level. The association of ZCCHC3 with both LINE-1 RNA and ORF1 suggests that ZCCHC3 interacts with LINE-1 RNP and consequently causes its RNA degradation. These data demonstrate collectively that ZCCHC3 contributes to the cellular control of LINE-1 replication.

This study focused on evaluating the influence of Clostridium butyricum and Brevibacillus strains on egg production, egg quality, immune response and antioxidant function, apparent fecal amino acid digestibility, and jejunal morphology when supplemented as probiotics in the diets of laying hens in the peak phase. A total of 288 healthy 30-week-old Hy-Line Brown laying hens were arbitrarily assigned to four dietary groups, which included control diet and control diet supplemented with 0.02% C. butyricum zlc-17, C. butyricum lwc-13, or Brevibacillus zlb-z1, for 84 days. The results showed that dietary C. butyricum and Brevibacillus sp. exerted a positively significant influence (P ≤ 0.05) compared to the control group on the performance, egg quality, and physiological response of the birds. The diets could reduce mortality rate and enhance (P ≤ 0.05) egg weight and egg mass, egg production rate, and feed efficiency. Further analysis suggested that the probiotic strains can enhance (P ≤ 0.05) eggshell quality, Haugh unit, thick albumen content, and albumen height. Also, probiotics enhanced (P ≤ 0.05) the antioxidant status via increased antioxidant enzymes and jejunal morphology as evidenced by increased villi surface area (VSA), the ratio of villi height to crypt depth, villi width, and villi height, and a significant reduction in crypt depth. Besides, nutrient absorption and retention were enhanced, as apparent fecal amino acid digestibility of key essential amino acids was substantially improved in the diet-based group. The concentrations of immunoglobulin M and A (IgM and IgA) increased significantly (P ≤ 0.05) in the probiotics group and the same effect was notable for complement proteins (C3) and immune organ (Spleen). Conclusively, the supplementation of Clostridium butyricum zlc-17 in comparison to Clostridium butyricum lwc-13 and Brevibacillus zlb-z1 strains significantly (P ≤ 0.05) promoted the antioxidant status, modulated the intestinal structure, enhanced amino acid digestibility, and regulated the immunity index of the laying hens, which finally improves the laying performance and egg quality of the laying hens.

Biofilm communities play a major role in explaining the temporal variation of biogeochemical conditions in freshwater ecosystems, and yet we know little about how these complex microbial communities change over time (aka succession), and from different initial conditions, in comparison to other stream communities. This has resulted in limited knowledge on how biofilm community structure and microbial colonization vary over relevant time scales to become mature biofilms capable of significant alteration of the freshwater environment in which they live. Here, we monitored successional trajectories of biofilm communities from summer and winter in a headwater stream and evaluated their structural state over time by DNA high-throughput sequencing. Significant differences in biofilm composition were observed when microbial colonization started in the summer vs. winter seasons, with higher percentage of algae (Bacillariophyta) and Bacteroidetes in winter-initiated samples but higher abundance of Proteobacteria (e.g., Rhizobiales, Rhodobacterales, Sphingomonadales, and Burkholderiales), Actinobacteria, and Chloroflexi in summer-initiated samples. Interestingly, results showed that despite seasonal effects on early biofilm succession, biofilm community structures converged after 70 days, suggesting the existence of a stable, mature community in the stream that is independent of the environmental conditions during biofilm colonization. Overall, our results show that algae are important in the early development of biofilm communities during winter, while heterotrophic bacteria play a more critical role during summer colonization and development of biofilms.

Stover mulching as a conservation and sustainable agricultural practice is beneficial for maintaining soil nitrogen (N) requirements and plant health. The microbial functional guilds of the root and rhizosphere are important factors in the soil nitrogen cycle. However, it is unclear how the frequency and amount of stover mulching influence microbial functional guilds in the root and rhizosphere. Therefore, we investigated the responses of the microbial functional guilds in the endosphere and rhizosphere to maize stover mulching amounts (0, 1/3, 2/3, and total stover mulching every year) and frequencies (once every 3 years and twice every 3 years) under 10-year no-till management. The bacterial functional guilds of nitrogen fixation, nitrification, and anaerobic nitrate oxidation displayed the significantly correlation with C/N, total nitrogen, NO3−, and NH4+. The fungal functional guilds of plant pathogens and saprotrophs showed significantly correlations with C/N, total nitrogen, and NO3−. Moreover, we found that bacterial guilds play a pivotal role in maintaining N requirements at the jointing stage, whereas root endophytic fungal guilds play a more important role than bacterial guilds in regulating plant health at the mature stage. The frequency and amount of stover mulching had significant effects on the microbial functional guilds in the root and rhizosphere. Our data suggest that stover mulch application twice every 3 years is the optimal mulching frequency because it yielded the lowest abundance of nitrifying and anaerobic nitrate-oxidising bacteria and the highest abundance of nitrogen-fixing bacteria at the jointing stage, as well as the lowest abundance of fungal plant pathogens in roots at the mature stage.

To better understand the ensiling characteristics of sorghum stalk, the dynamic changes of fermentation parameters, nutrient components and bacterial community of sorghum stalk silage were analyzed by intermittently sampling on day 0, 1, 3, 7, 14, 28, and 56 of ensiling duration. The results showed that high-moisture sorghum stalk was well preserved during ensiling fermentation, with the DM loss of 4.10% and the little difference between the nutrients of sorghum stalk before and after ensiling. The pH value of silage declined to its lowest value of 4.32 by Day 7 of ensiling, and other fermentation parameters kept steady since Day 28 of ensiling. The amplicon sequencing analysis revealed that the alpha diversity parameters of silage bacterial community including Shannon index, observed features, Pielou evenness and Faith PD gradually declined (P < 0.01) with ensiling duration. Principal coordinate analysis (PCoA) revealed that bacterial profiles of raw material would experience a succession becoming a quite different community during ensiling fermentation. Taxonomic classification revealed a total of 10 and 173 bacterial taxa at the phylum and genus level, respectively, as being detected with relative abundances higher than 0.01% and in at least half samples. LEfSe analysis revealed that 26 bacterial taxa were affected by sampling timepoint (P < 0.05 and LDA score > 4). When focusing on the dynamic trend of silage bacterial taxa, lactic acid bacteria successfully dominated in the bacterial community on Day 1 of ensiling, and the bacterial community almost came to a plateau by Day 28 of ensiling, with Lactobacillus and Leuconostoc as the dominant genera. In a word, the succession of fermentation parameters, nutrient components and bacterial community indicate a successful dominance establishment of LAB and a fast advent of fermentation plateau, suggesting that high-moisture sorghum stalk can be ensiled directly, but the pH of mature silage is a little high.

Francisella tularensis is a dangerous pathogen that causes an extremely contagious zoonosis in humans named tularemia. Given its low-dose morbidity, the potential to be fatal, and aerosol spread, it is regarded as a severe threat to public health. The US Centers for Disease Control and Prevention (CDC) has classified it as a category A potential agent for bioterrorism and a Tier 1 Select Agent. Herein, we combined recombinase polymerase amplification (RPA) with CRISPR/Cas12a system to select the F. tularensis target gene (TUL4), creating a two-pronged rapid and ultrasensitive diagnostic method for detecting F. tularensis. The real-time RPA (RT-RPA) assay detected F. tularensis within 10 min at a sensitivity of 5 copies/reaction, F. tularensis genomic DNA of 5 fg, and F. tularensis of 2 × 102 CFU/ml; the RPA-CRISPR/Cas12a assay detects F. tularensis within 40 min at a sensitivity of 0.5 copies/reaction, F. tularensis genomic DNA of 1 fg, and F. tularensis of 2 CFU/ml. Furthermore, the evaluation of specificity showed that both assays were highly specific to F. tularensis. More importantly, in a test of prepared simulated blood and sewage samples, the RT-RPA assay results were consistent with RT-PCR assay results, and the RPA-CRISPR/Cas12a assay could detect a minute amount of F. tularensis genomic DNA (2.5 fg). There was no nonspecific detection with blood samples and sewage samples, giving the tests a high practical application value. For example, in on-site and epidemic areas, the RT-RPA was used for rapid screening and the RPA-CRISPR/Cas12a assay was used for more accurate diagnosis.

Bovine herpesvirus 1 (BoHV-1) is an alphaherpesvirus that causes infectious bovine rhinotracheitis and infectious pustular vulvovaginitis in cattle. Ιnterferon-gamma (IFN-γ) is a pleiotropic cytokine with antiviral activity that modulates the innate and adaptive immune responses. In this study, we prepared high-purity bovine interferon gamma (BoIFN-γ) dimer protein using prokaryotic expression system and affinity chromatography. We subsequently investigated the effect of BoIFN-γ on BoHV-1 infection in Madin-Darby bovine kidney (MDBK) cells. The results showed that BoIFN-γ pre-treament not only decreased the production of BoHV-1 but also reduced the cytopathic effect of the virus. Differential gene expression profiles of BoHV-1 infected MDBK cells were then analysed through high-throughput RNA sequencing. The data showed that BoIFN-γ pre-treatment reduced lipid metabolism disorder and DNA damage caused by BoHV-1 infection. Furthermore, BoIFN-γ treatment upregulated the transcription of interferon regulatory transcription factors (IRF1 and GBP5) and interferon-stimulated genes (ISGs) of MDBK cells. Additionally, BoIFN-γ promotes expression of cellular protein involved in complement activation and coagulation cascades response as well as antigen processing and presentation process, while BoHV-1 infection dramatically downregulates transcription of these immune components including C3, C1r, C1s, PLAT, ITGB2, PROCR, BoLA, CD74, B2M, PA28, BoLA-DRA, and TAPBP. Collectively, our findings revealed that BoIFN-γ pre-treatment can improve host resistance to BoHV-1 infection and regulate transcription or expression of host protein associated with cellular metabolism and innate immune response. This provides insights into the development of prophylactic agents for prevention and control of BoHV-1 infection.

This study aimed to investigate the prevalence and characterization of tet(X4) in Escherichia coli isolates from a pig farm in Shanghai, China, and to elucidate tet(X4) dissemination mechanism in this swine farm. Forty-nine (80.33%) E. coli strains were isolated from 61 samples from a pig farm and were screened for the presence of tet(X). Among them, six (12.24%) strains were positive for tet(X4) and exhibited resistance to tigecycline (MIC ≥ 16 mg/L). They were further sequenced by Illumina Hiseq. Six tet(X4)-positive strains belonged to ST761 with identical resistance genes, resistance profiles, plasmid replicons, and cgMLST type except that additional ColE10 plasmid was present in isolate SH21PTE35. Isolate SH21PTE31, as a representative ST761 E. coli strain, was further sequenced using Nanopore MinION. The tet(X4) in SH21PTE31 was located on IncFIA18/IncFIB(K)/IncX1 hybrid plasmid pYUSHP31-1, highly similar to other tet(X4)-carrying IncFIA18/IncFIB(K)/IncX1 plasmids from ST761 E. coli and other E. coli lineages in China. These IncFIA18/IncFIB(K)/IncX1 plasmids shared closely related multidrug resistance regions, and could reorganize, acquire or lose resistance modules mediated by mobile elements such as ISCR2 and IS26. Phylogenetic analysis were performed including all tet(X4)-positive isolates obtained in this pig farm combined with 43 tet(X4)-positive E. coli from pigs, cow, pork, wastewater, and patients with the same ST from NCBI. The 50 tet(X4)-carrying E. coli ST761 isolates from different areas in China shared a close phylogenetic relationship (0-49 SNPs). In conclusion, clonal transmission of tet(X4)-positive E. coli ST761 has occurred in this swine farm. E. coli ST761 has the potential to become a high-risk clone for tet(X4) dissemination in China.

BackgroundLeishmaniasis being a local disease, as kala-azar this particular form is a visceral form. It is transmitted by sandflies, and is a parasitic disease involving the reticuloendothelial system of mononuclear macrophages. Due to its poor prognosis and high fatality rate, the fatality rate of patients without effective treatment can exceed 95%. Thereby, early diagnosis and treatment can significantly improve its prognosis. The metagenomic next-generation sequencing (mNGS) has the advantage of being able to find pathogens that cannot be detected by traditional methods. More importantly, it can conduct nucleic acid detection of pathogens covering a wide range in a short time. For infectious diseases like kala-azar, which is clinically complicated and difficult, mNGS detection provides a basis for accurate etiological diagnosis.Case ReportWe report 2 cases of kala-azar in West China Hospital, Chengdu, China. The first case is a 47-year-old male patient who had recurrent fever for 4 months, accompanied by reduction of red blood cell, white blood cell, and blood platelet. He was detected by mNGS and clinically diagnosed as kala-azar (Leishmania detection), finally died of multiple organ failure. The second patient was a 15-year-old male who had fever for more than 10 days. He was detected by mNGS and clinically diagnosed as kala-azar (Leishmania detection). He recovered and discharged quickly after treatment with sodium stibogluconate.ConclusionEfforts should be made to improve early etiological diagnosis in order to improve patient prognosis. mNGS detection is beneficial to the diagnosis and treatment of infectious diseases with unknown causes in the early stage of emergency treatment.

Tryptophan (Trp) can modify the gut microbiota. However, there is no information about the effect of Trp on intestinal microbiota after lipopolysaccharide (LPS) challenge. This study aimed to investigate the effect of Trp on intestinal barrier function, inflammation, antioxidant status, and microbiota in LPS-challenged piglets. A total of 18 weaned castrated piglets were randomly divided into three treatments with 6 replicate per treatment, namely, (i) non-challenged control (CON); (ii) LPS-challenged control (LPS-CON); and (iii) LPS + 0.2% Trp (LPS-Trp). After feeding with control or 0.2% tryptophan-supplemented diets for 35 days, pigs were intraperitoneally injected with LPS (100 μg/kg body weight) or saline. At 4 h post-challenge, all pigs were slaughtered, and colonic samples were collected. The samples were analyzed for gut microbiota, fatty acids, antioxidant parameters, and the expression of mRNA and protein. The community bar chart showed that Trp supplementation to LPS-challenged pigs increased the relative abundance of Anaerostipes (P < 0.05) and tended to increase the relative abundance of V9D2013_group (P = 0.09), while decreased the relative abundance of Corynebacterium (P < 0.05) and unclassified_c__Bacteroidia (P < 0.01). Gas chromatography showed that Trp increased the concentrations of acetate, propionate, butyrate, and isovalerate in the colonic digesta (P < 0.05). Trp reduced the mRNA level of pro-inflammatory cytokines (P < 0.01), and increased mRNA level of aryl hydrocarbon receptor, cytochrome P450 (CYP) 1A1 and CYP1B1 (P < 0.05). Correlation analysis results showed that acetate, propionate, and butyrate concentrations were positively correlated with mRNA level of occludin and CYP1B1 (P < 0.05), and were negatively correlated with pro-inflammatory cytokines gene expression (P < 0.05). Isovalerate concentration was positively correlated with catalase activity (P < 0.05), and was negatively correlated with pro-inflammatory cytokines gene expression (P < 0.05). Furthermore, Trp enhanced the antioxidant activities (P < 0.01), and increased mRNA and protein expressions of claudin-1, occludin, and zonula occludens-1 (P < 0.01) after LPS challenge. These results suggest that Trp enhanced intestinal health by a modulated intestinal microbiota composition, improved the short chain fatty acids synthesis, reduced inflammation, increased antioxidant capacity, and improved intestinal barrier function.

The community structures of epiphytic bacteria on the surface of macroalgae are closely related to their host algae, but there is a lack of research on the differences between the epiphytic bacterial communities of male and female algae and their reproductive tissues. In this study, high-throughput sequencing was used to compare epiphytic bacterial community structures on the intertidal macroalgae Sargassum thunbergii and their receptacles between male and female samples. The epiphytic bacteria on the male and female algal bodies and receptacles had similar community structures with a large number of shared bacteria, but the samples clearly clustered separately, and the abundances of dominant taxa, specific bacteria, and indicator species differed, indicating that epiphytic bacterial communities differed significantly between the male and female S. thunbergii and their receptacles. In addition, the abundance of many predicted functional genes was significantly different between epiphytic bacteria on male and female algal bodies and receptacles, especially metabolism-related genes, and the abundances of predicted functional genes of epiphytic bacteria were significantly higher on both types of male samples than on female samples. Our study confirmed that the sex of the host algae influenced the epiphytic bacterial community structures on algae and algal reproductive tissues, and this role was mainly related to the host metabolism. The results reveal the role of host plant sex in the formation of epiphytic bacterial communities. These findings are helpful for obtaining an in-depth understanding of the construction mechanism of algae-associated bacteria.

Plant growth performance under a stressful environment, notably in the agriculture field, is directly correlated with the rapid growth of the human population, which triggers the pressure on crop productivity. Plants perceived many stresses owing to degraded land, which induces low plant productivity and, therefore, becomes a foremost concern for the future to face a situation of food scarcity. Land degradation is a very notable environmental issue at the local, regional, and global levels for agriculture. Land degradation generates global problems such as drought desertification, heavy metal contamination, and soil salinity, which pose challenges to achieving many UN Sustainable Development goals. The plant itself has a varied algorithm for the mitigation of stresses arising due to degraded land; the rhizospheric system of the plant has diverse modes and efficient mechanisms to cope with stress by numerous root-associated microbes. The suitable root-associated microbes and components of root exudate interplay against stress and build adaptation against stress-mediated mechanisms. The problem of iron-deficient soil is rising owing to increasing degraded land across the globe, which hampers plant growth productivity. Therefore, in the context to tackle these issues, the present review aims to identify plant-stress status owing to iron-deficient soil and its probable eco-friendly solution. Siderophores are well-recognized iron-chelating agents produced by numerous microbes and are associated with the rhizosphere. These siderophore-producing microbes are eco-friendly and sustainable agents, which may be managing plant stresses in the degraded land. The review also focuses on the molecular mechanisms of siderophores and their chemistry, cross-talk between plant root and siderophores-producing microbes to combat plant stress, and the utilization of siderophores in plant growth on degraded land.

Enterococcus faecalis is a common gram-positive non-spore-forming bacterium in nature and is found in the upper respiratory tract, intestine, and mouth of healthy people. E. faecalis is also one of the common pathogens causing nosocomial infections and is resistant to several antibiotics commonly used in practice. Thus, treating drug-resistant E. faecalis with antibiotics is challenging, and new approaches are needed. In this study, we isolated a bacteriophage named EFap02 that targets E. faecalis strain EFa02 from sewage at Southwest Hospital. Phage EFap02 belongs to the Siphoviridae family with a long tail of approximately 210 nm, and EFap02 can tolerate a strong acid and alkali environment and high temperature. Its receptor was identified as the capsular polysaccharide. Phage-resistant mutants had loss-of-function mutations in glycosyltransferase (gtr2), which is responsible for capsular polysaccharide biosynthesis, and this caused the loss of capsular polysaccharide and interruption of phage adsorption. Although phage-resistant mutants against EFap02 can be selected, such mutants are impaired in biofilm formation due to the loss of capsular polysaccharide, which compromises its virulence. Therefore, this study provided a detailed description of the E. faecalis EFap02 phage with the potential for treating E. faecalis infection.

Aiming to reveal the variation in bacteria community under oxygen depletion formed every summer in water column of central Bohai Sea, a time-scenario sampling from June to August in 2018 at a 20-day interval along one inshore–offshore transect was settled. Water samples were collected at the surface, middle, and bottom layer and then analyzed by high-throughput sequencing targeting both 16S rRNA and nosZ genes. Compared to the surface and middle water, oxygen depletion occurred at bottom layer in August. In top two layers, Cyanobacteria dominated the bacterial community, whereas heterotrophic bacteria became dominant in bottom water of Bohai Sea. Based on the time scenario, distinct community separation was observed before (June and July) and after (August) oxygen depletion (p = 0.003). Vertically, strict stratification of nosZ gene was stably formed along 3 sampling layers. As a response to oxygen depletion, the diversity indices of both total bacteria (16S rRNA) and nosZ gene-encoded denitrification bacteria all increased, which indicated the intense potential of nitrogen lose when oxygen depleted. Dissolved oxygen (DO) was the key impacting factor on the community composition of total bacteria in June, whereas nutrients together with DO play the important roles in August for both total and denitrifying bacteria. The biotic impact was revealed further by strong correlations which showed between Cyanobacteria and heterotrophic bacteria in June from co-occurrence network analysis, which became weak in August when DO was depleted. This study discovered the variation in bacteria community in oxygen-depleted water with further effort to understand the potential role of denitrifying bacteria under oxygen depletion in Bohai Sea for the first time, which provided insights into the microbial response to the world-wide expanding oxygen depletion and their contributions in the ocean nitrogen cycling.

Human cytomegalovirus (HCMV) is a β-herpesvirus whose genome consists of double stranded linear DNA. HCMV genome can generate non-coding RNAs (ncRNAs) through transcription in its host cells. Besides that, HCMV infection also changes the ncRNAs expression profile of the host cells. ncRNAs play a key role in maintaining the normal physiological activity of cells, and the disorder of ncRNAs expression has numerous adverse effects on cells. However, until now, the relationship between ncRNAs and HCMV-induced adverse effects are not summarized in detail. This review aims to give a systematic summary of the role of HCMV infection in ncRNAs expression while providing insights into the molecular mechanism of unnormal cellular events caused by ncRNAs disorder. ncRNAs disorder induced by HCMV infection is highly associated with cell proliferation, apoptosis, tumorigenesis, and immune regulation, as well as the development of cardiovascular diseases, and the potential role of biomarker. We summarize the studies on HCMV associated ncRNAs disorder and suggest innovative strategies for eliminating the adverse effects caused by HCMV infection.

Non-Saccharomyces (NS) yeasts with high β-glucosidase activity play a vital role in improving the aroma complexity of wines by releasing aroma compounds from glycosidic precursors during fermentation. In this study, the effect of sequential inoculation fermentation of Meyerozyma guilliermondii NM218 and Hanseniaspora uvarum BF345 with two Saccharomyces cerevisiae strains [Vintage Red™ (VR) and Aroma White™ (AW)] on volatile compounds and sensory characteristics of wines was investigated. Prior to winemaking trials, the sequential inoculation times of the two NS yeasts were evaluated in synthetic must, based on changes in strain population and enzyme activity. The intervals for inoculation of NM218 and BF345 with the S. cerevisiae strains were 48 and 24 h, respectively. In the main experiment, sequential inoculation fermentations of the two strains with S. cerevisiae were carried out in Cabernet Sauvignon (CS) and Chardonnay (CH) grape must. The oenological parameters, volatile composition, and sensory characteristics of the final wines were assessed. No clear differences were observed in the oenological parameters of the sequentially fermented CH wines compared with the control, except for residual sugar and alcohol. However, in CS wines, the total acid contents were significantly lower in the wines fermented by sequential inoculation compared to the control. Both NM218 and BF345 improved the aroma complexity of wines by increasing esters and terpenes when inoculated with S. cerevisiae strains compared to inoculation with S. cerevisiae strains alone. NM218 resulted in a more positive effect on CS wine aroma, with higher levels of citronellol and trans-nerolidol. BF345 significantly enhanced the floral and fruity aromas of CH wine by producing higher concentrations of geranyl acetone, β-damascenone, trans-nerolidol, and nerol. Both NM218 and BF345 yeasts could potentially be used to improve wine aroma and overall quality, especially wine floral and fruity aromas, when used in sequential inoculation with S. cerevisiae.

The potentially pathogenic species of the genus Vibrio pose a threat to both humans and animals, creating medical burdens and economic losses to the mariculture industry. Improvements in surveillance and diagnosis are needed to successfully manage vibriosis outbreaks. Matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) can provide rapid diagnosis and has been widely used in the identification of Vibrio spp. The main weakness of this technology is the limited number of strains and species of Vibrio in the existing commercial database. Here, we develop a new in-house database named PVBase containing 790 main spectra projections (MSP) of ten Vibrio species that come from various regions of China and include abundant clinical and environmental strains. PVBase was validated through a blind test of 65 Vibrio strains. The identification accuracy and scoring of Vibrio strains was greatly improved through the addition of PVBase. Identification accuracy increased from 73.4 to 100%. The number of strains with identification scores above 2.2 increased from 53.1% to 96.9% and 53.1% of strains had an identification score above 2.59. Moreover, perfect discrimination was obtained when using all of the MSPs created for the Vibrio species, even for very closely related species such as V. cholerae, V. albensis, and V. mimicus or V. alginolyticus, V. parahaemolyticus, and V. harveyi. In addition, we used phyloproteomic analysis to study whether there are differences in protein fingerprints of different regions or pathogenic strains. We found that MSP characteristics of Vibrio species were not related to their region or source. With the construction of PVBase, the identification efficiency of potentially pathogenic Vibrio species has been greatly improved, which is an important advance for epidemic prevention and control, and aquaculture disease detection.

The widespread emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) with limited therapeutic options has become a global concern. In this study, a K. pneumoniae strain called KP2e was recovered from a human case of fatal septic shock in a Chinese hospital. Polymerase chain reaction and sequencing, antimicrobial susceptibility testing, conjugation experiments, S1 nuclease-pulsed field gel electrophoresis/southern blot, whole genome sequencing and comparative genomics were performed to investigate the phenotypic and molecular characteristics of this isolate. KP2e possessed the NDM-6-encoding gene and exhibited resistance to almost all β-lactams except for monobactam. This strain belonged to sequence type 4024, the complete genome of which was composed of one chromosome and three plasmids. Furthermore, blaNDM–6 coexisted on two self-transmissible plasmids, which were assigned to types IncFIB and IncN. A structure of IS26-composite transposon capturing an identical Tn125 remnant (ΔISAba125-blaNDM–6-bleMBL-trpF-dsbC-cutA-groES-ΔgroEL) was identified in the two plasmids, and this conserved blaNDM-surrounding genetic context was similar to that of few IncN plasmids found in other regions of China. Our research appears to be the first description of a clinical strain that emerged co-harbouring dual blaNDM-carrying plasmids, and the first report of NDM-6-positive CRKP in China. These findings demonstrated that IncN is a key medium in the evolution and expanding dissemination of blaNDM genes among various species, which indicates that close monitoring and rapid detection of blaNDM-harbouring plasmids is necessary.

Extracellular DNA (eDNA) is a critical component in the extracellular matrix (ECM) of bacterial biofilms, while little is known about the mechanisms underlying how eDNA integrates into the ECM through potential macromolecular interactions. Myxococcus xanthus biofilm was employed as a suitable model for the investigation due to the co-distribution of eDNA and exopolysaccharides (EPS) owing to their direct interactions in the ECM. DNA is able to combine with M. xanthus EPS to form a macromolecular conjugate, which is dominated by the electrostatic forces participating in the polymer-polymer interactions. Without intercalation binding, DNA-EPS interactions exhibit a certain degree of reversibility. Acting as a strong extracellular framework during biofilm formation process, the eDNA-EPS complex not only facilitates the initial cell adhesion and subsequent establishment of ECM architecture, but also renders cells within biofilms stress resistances that are relevant to the survival of M. xanthus in some hostile environments. Furthermore, the EPS protects the conjugated DNA from the degradation by nucleic acid hydrolases, which leads to the continuous and stable existence of eDNA in the native ECM of M. xanthus biofilms. These results will shed light on developing prevention and treatment strategies against biofilm-related risks.

We characterized a negative sense single-stranded RNA mycovirus, Fusarium oxysporum mymonavirus 1 (FoMyV1), isolated from the phytopathogenic fungus Fusarium oxysporum. The genome of FoMyV1 is 10,114 nt, including five open reading frames (ORFs1–5) that are non-overlapping and linearly arranged. The largest, ORF5, encodes a large polypeptide L containing a conserved regions corresponding to Mononegavirales RNA-dependent RNA polymerase and mRNA-capping enzyme region V; the putative functions of the remaining four ORFs are unknown. The L protein encoded by ORF5 shared a high amino acid identity of 65% with that of Hubei rhabdo-like virus 4, a mymonavirus that associated with arthropods. However, the L protein of FoMyV1 also showed amino acid similarity (27–36%) with proteins of mynonaviruses that infect the phytopathogenic fungi Sclerotinia sclerotiorum and Botrytis cineaea. Phylogenetic analysis based on L protein showed that FoMyV1 is clustered with the members of the genus Hubramonavirus in the family Mymonaviridae. Moreover, we found that FoMyV1 could successfully transfer by hyphal anastomosis to a virus-free strain. FoMyV1 reduced the vegetative growth and conidium production of its fungal host but did not alter its virulence. To the best of our knowledge, this is not only the first mymonavirus described in the species F. oxysporum, but also the first Hubramonavirus species found to infect a fungus. However, the incidence of FoMyV1 infections in the tested F. oxysporum strains was only 1%.

Bacillus thuringiensis, a gram-positive bacteria, has three insecticidal proteins: Vip (vegetative insecticidal protein), Cry (crystal), and Sip (secreted insecticidal protein). Of the three, Sip proteins have insecticidal activity against larvae of Coleoptera. However, the Sip1Aa protein has little solubility in the supernatant because of inclusion bodies. This makes it more difficult to study, and thus research on Sip proteins is limited, which hinders the study of their mechanistic functions and insecticidal mechanisms. This highlights the importance of further investigation of the Sip1Aa protein. Disulfide bonds play an important role in the stability and function of proteins. Here, we successfully constructed mutant proteins with high insecticidal activity. The tertiary structure of the Sip1Aa protein was analyzed with homologous modeling and bioinformatics to predict the conserved domain of the protein. Cysteine was used to replace amino acids via site-directed mutagenesis. We successfully constructed Sip149-251, Sip153-248, Sip158-243, and Sip178-314 mutant proteins with higher solubility than Sip1Aa. Sip153-248 and Sip158-243 were the most stable compared to Sip1Aa, followed by Sip149-251 and Sip178-314. The insecticidal activity of Sip153-248 (Sip158-243) was 2.76 (2.26) times higher than that of Sip1Aa. The insecticidal activity of Sip149-251 and Sip178-314 did not differ significantly from that of Sip1Aa. Basic structural properties, physicochemical properties, and the spatial structure of the mutation site of Sip1Aa and the mutant proteins were analyzed. These results provide a molecular basis for using Sip1Aa to control Coleopteran insects and contribute to the study of the Sip1Aa insecticidal mechanism.

We investigated the effect of photoperiod on ileal morphology, barrier function, short-chain fatty acid (SCFA) contents, microbial flora, melatonin expression, and synthesis in laying ducks. After adaption, a total of 180 Jinding laying ducks (252 days old) were randomly divided into three treatments, receiving 12L (hours of light):12D (hours of darkness), 16L:8D, or 20L:4D. Each treatment had six replicates with 10 birds each. The formal experiment lasted 58 days. Compared with 12L:12D, the significantly higher values of villus height and goblet cell percentage (GCP) were observed in 16L:8D treatment, accompanied with the higher mRNA relative expression of zonula occludens-1, zonula occludens-2, zonula occludens-3, claudin-1, occludin, and mucin 2 (P < 0.05). Besides, significantly higher values of acetate and propionate, butyrate and total SCFA concentrations were simultaneously observed in ileal chyme of 16L:8D treatment (P < 0.05). For the ileal microbial community, the results of principal coordinate analysis (PCoA) visually presented that three photoperiod groups were mainly scattered into three clusters, indicating that the microbiota composition in different photoperiod treatments were quite dissimilar. Lower values of Shannon indicators were observed in the 20L:4D treatment (P < 0.05), meaning that the microbiota α-diversity decreased in the 20-h photoperiod. The relative abundance of Actinobacteria, Fusobacteria, and Proteobacteria at phylum level and Fusobacterium, Clostridium_sensu_stricto_1, and Pectobacterium at genus level kept an appropriate balance in the 16L:8D photoperiod. Melatonin level in serum decreased with the increasing photoperiods at 6:00 and 12:00, which was consistent with melatonin receptor expressions in the hypothalamus and ileal tissue. Meanwhile, the adenosine 3′,5′-cyclic phosphate (cAMP) contents were significantly downregulated in the pineal gland (P < 0.05), in response to the increase in photoperiod. In conclusion, an appropriate photoperiod could improve ileal morphology, barrier function, SCFA profile, and microbial flora, which may be attributed to the appropriate regulation of the circadian rhythm through melatonin as well as its receptor expression, and 16 h could be an adequate photoperiod for laying ducks.

Laribacter hongkongensis is a new emerging foodborne pathogen that causes community-acquired gastroenteritis and traveler’s diarrhea. However, the genetic features of L. hongkongensis have not yet been properly understood. A total of 45 aquatic animal-associated L. hongkongensis strains isolated from intestinal specimens of frogs and grass carps were subjected to whole-genome sequencing (WGS), along with the genome data of 4 reported human clinical strains, the analysis of virulence genes, carbohydrate-active enzymes, and antimicrobial resistance (AMR) determinants were carried out for comprehensively understanding of this new foodborne pathogen. Human clinical strains were genetically more related to some strains from frogs inferred from phylogenetic trees. The distribution of virulence genes and carbohydrate-active enzymes exhibited different patterns among strains of different sources, reflecting their adaption to different host environments and indicating different potentials to infect humans. Thirty-two AMR genes were detected, susceptibility to 18 clinical used antibiotics including aminoglycoside, chloramphenicol, trimethoprim, and sulfa was checked to evaluate the availability of clinical medicines. Resistance to Rifampicin, Cefazolin, ceftazidime, Ampicillin, and ceftriaxone is prevalent in most strains, resistance to tetracycline, trimethoprim-sulfamethoxazole, ciprofloxacin, and levofloxacin are aggregated in nearly half of frog-derived strains, suggesting that drug resistance of frog-derived strains is more serious, and clinical treatment for L. hongkongensis infection should be more cautious.

The rice blast pathogen, Magnaporthe oryzae, spreads through spores and invades rice through appressoria. Melanin is necessary for an appressorium to penetrate plant cells, but there are many unknown aspects of its role in fungal conidiation. In this study, we confirmed that melanin promotes spore production in M. oryzae, and that this effect is related to the background melanin content of wild-type strains. In the wild-type 70-15 strain with low melanin content of aerial hyphae, increased melanin synthesis promoted sporulation. In contrast, increased melanin synthesis in the wild-type Guy11 strain, which has higher melanin content, did not promote sporulation. The transcription factor Cnf1 (conidial production negative regulatory factor 1), which negatively regulates melanin synthesis, has opposite effects in conidiophore differentiation of Guy11 and 70-15. Deletion of CNF1 did not abolish the defects of Δcos1 and Δhox2 (where COS1/conidiophore stalk-less 1 or HOX2/homeodomain protein 2 was deleted) in conidiation, while increased the conidiation of Δgcc1 and Δgcf3 (where GCC1/growth, conidiation and cell wall regulatory factor 1, or GCF3/growth and conidiation regulatory factor 3 was deleted). Pig1 (pigment of Magnaporthe 1) regulates the melanin synthesis of hyphae but not of conidiophores, spores, or appressoria. Deletion of the same gene in different wild-type strains can lead to different phenotypes, partly because of differences in melanin content between fungal strains. Overall, this study reveals the functional diversity and complexity of melanin in different M. oryzae strains.

Marine picocyanobacteria Synechococcus exhibit highly diverse pigment types (PTs) and hence possess great advantage to utilize different spectrum of light effectively and to occupy a wide range of light niches. In this study, we explored the diversity of Synechococcus PTs in the eastern Indian Ocean (EIO), surface water of Strait of Malacca (SSM), and coastal waters of Sri Lanka (SSL). All the detected PTs were phycourobilin (PUB) containing PT 3 and showed distinct distribution patterns. Low PUB PT 3a and partial chromatic acclimater PT 3eA dominated in coastal and shallow waters (SSM and SSL). In contrast, high PUB and chromatic acclimaters PT 3dA and PT 3c/3dB were mainly distributed in open ocean (EIO). PT 3dA and PT 3c/3dB occurred at similar depths of the lower euphotic layers but showed distinct distribution pattern that are partially exclusive, indicating that they compete with each other for the same light niche. Interestingly, the newly described PT 3f was detected with high relative abundances at all stations and particularly dominated in the upper euphotic layer in EIO, which was confirmed with PT-specific quantitative polymerase chain reaction (qPCR). The relative abundance of PT 3f was negatively correlated with nutrient level, implying that PT 3f is adapted to oligotrophic waters. Pronounced niche partition of different PTs was observed in the upper and lower layers of euphotic zone in EIO and SSM/SSL. Light, nutrients, and strong stratification may play important roles in the niche partition of different PTs. Further analysis about ecologically significant taxonomic units revealed high diversity within each PT at different locations, which provided insights for understanding specific PT with wide range of niches.

This study aimed to investigate the prevalence of fosfomycin fosA7 in Salmonella enterica isolates from food animals and retail meat products in China and the impact of fosA7 on bacterial fitness. A total of 360 Salmonella isolates collected from 11 provinces and cities in China were detected for fosA7. All fosA7-positive Salmonella isolates were determined minimum inhibitory concentrations (MICs) and sequenced by Illumina Hiseq. The fosA7 gene of S. Derby isolate HA2-WA5 was knocked out. The full length of fosA7 was cloned into vector pBR322 and then transformed into various hosts. MICs of fosfomycin, growth curves, stability, and fitness of fosA7 were evaluated. The fosA7 gene was identified in S. Derby (ST40, n = 30) and S. Reading (ST1628, n = 5). MICs to fosfomycin of 35 fosA7-positive isolates were 1 to 32 mg/L. All fosA7 were located on chromosomes of Salmonella. The deletion of fosA7 in HA2-WA5 decreased fosfomycin MIC by 16-fold and slightly affected its fitness. The acquisition of plasmid-borne fosA7 enhanced MICs of fosfomycin in Salmonella (1,024-fold) and Escherichia coli (16-fold). The recombinant plasmid pBR322-fosA7 was stable in Salmonella Typhimurium, S. Pullorum, S. Derby, and E. coli, except for Salmonella Enteritidis, and barely affected on the growth of them but significantly increased biological fitness in Salmonella. The spread of specific Salmonella serovars such as S. Derby ST40 will facilitate the dissemination of fosA7. fosA7 can confer high-level fosfomycin resistance and enhance bacterial fitness in Salmonella if transferred on plasmids; thus, it has the potential to be a reservoir of the mobilized fosfomycin resistance gene.

Morganella morganii, which is often regarded as a human commensal organism, can be an opportunistic pathogen, causing a variety of clinical infections with serious morbidity and mortality. An efficient and convenient method for subtyping and identifying M. morganii strains in epidemiological surveillance and control is urgently needed. Serotyping based on bacterial surface polysaccharide antigens (O-antigen or K-antigens) is a standard subtyping method for many gram-negative bacteria. Here, through whole genome sequencing and comparative genomics analysis of 27 strains, we developed a molecular serotyping scheme based on the genetic variation of O-antigen gene clusters (O-AGC) in M. morganii, and 11 distinct O-AGC types were identified. A conventional serotyping scheme was also developed by the production of antisera and agglutination experiments, which was shown to be perfectly consistent with the molecular serotyping scheme, confirming that the variation in M. morganii O-AGC correlated with phenotypic O-antigen diversification. Furthermore, a microsphere-based suspension array (MSA) with high specificity was developed based on the specific genes within each O-AGC type. The sensitivity of MSA was determined to be 0.1 ng of genomic DNA and 103 CFU of pure culture. We further analyzed 104 M. morganii genomes available in GenBank, and an additional six novel O-AGC types were identified, indicating that the extension of this molecular serotyping scheme is convenient. Our work provides an important tool for the detection and epidemiological surveillance of M. morganii, and this method has the potential to be widely utilized, especially for bacterial genera/species without an efficient typing approach.