[Objective] Listeria monocytogenes (Lm) is a ubiquitous foodborne pathogen causing listeriosis.Lm can grow at low temperatures and thus may cause safety problems of refrigerated food and threaten the public health. The growth ofLm at low temperatures involves the inhibition of flagellar gene expression, which restricts flagellar biosynthesis. MogR is a transcriptional repressor which represses the expression of flagellar genes during intracellular infection and during extracellular growth ofLm at 37 ℃, resulting in no biosynthesis of flagella. Whereas MogR is deprived of repression function and the bacteria produce flagella during growth at 20–30 ℃. Our studies demonstrated thatLm significantly reduced the flagellar production at 4 ℃, but the molecular mechanism of which remained unclear. This study aims to reveal the relationship between the reduction of flagella and MogR repression at 4 ℃. [Methods] We constructed themogR-deleted mutant ΔmogR and the flagellin geneflaA-deleted mutant ΔflaA (as the control strain with no flagella), and their complementary strains cΔmogR and cΔflaA with theLm strain ATCC 19115 as the parental strain. Then, we analyzed the swarming motility, flagellar biosynthesis, and transcriptional levels of flagellar genes in above five strains at 4 ℃, 28 ℃, and 37 ℃, respectively. The growth curves of these strains were determined at 4 ℃, 28 ℃, and 37 ℃, respectively. [Results] Compared with the parental strain, ΔmogR showed significantly increased in motility, flagellar biosynthesis, and transcriptional levels of flagellar genes (P < 0.01, 0.001, and 0.001, respectively) at 4 ℃. The growth of ΔmogR markedly decreased compared with the parental strain (P < 0.05) at 4 ℃. The data of motility, flagellar biosynthesis, and transcriptional levels of flagellar genes in cΔmogR had no significant differences compared with the parental strain. [Conclusion] The reduction in flagellar biosynthesis was associated with the repression function of MogR inLm at 4 ℃. The reduction in flagellar biosynthesis was of benefit toLm proliferation at low temperatures. This study enriched our understanding of the mechanism ofLm growth at low temperatures.

Pathogenic mycoplasma has the ability to invade host cells, which is a key mechanism to their pathogenicity. Functional proteins that mediate the entry of mycoplasma may serve as potential drug or vaccine targets. [Objective] To clone and express the LRR5 protein encoded byMBOVPG45_0564 inMycoplasmabovis and to explore its role in the invasion of host cells byM.bovis. [Methods] The NCBI database was used for the homology analysis ofMBOVPG45_0564, and the structure of LRR5 protein was predicted by the Discovery Studio Client system. After prokaryotic expression of LRR5 protein, the mouse polyclonal antibody was prepared, and the subcellular localization of LRR5 protein was observed by immunoelectron microscopy. The invasion ofM.bovis into embryonic bovine lung (EBL) cells after LRR5 antibody blocking was observed by plate counting and laser confocal microscopy. After conjugation of LRR5 protein to the surface of fluorescent microspheres, the entry of the microspheres into EBL cells was observed by laser confocal microscopy and a high-content live-cell imaging system. [Results] MBOVPG45_0564 was a conserved sequence inM.bovis and encoded LRR5, a membrane-associated protein with a typical crescent-like spatial conformation. Multiple repeating leucine motifs were assembled in a supercoiled manner to form a solenoid protein structural unit. After LRR5 antibody blocking, the invasion rate ofM.bovis to EBL cells reduced (P<0.05), and the fluorescent microspheres conjugated with LRR5 protein could successfully enter EBL cells. [Conclusion] The LRR5 protein encoded byMBOVPG45_0564 is localized on theM.bovis membrane and plays a role in the invasion ofM.bovis into host cells.

[Objective] To analyze the polysaccharide hydrolysis activity and genomic characteristics of a Gram-negative bacterial strain FZY0027 isolated from intertidal seawater. [Methods] The strain FZY0027 was identified based on the morphological characteristics, 16S rRNA gene sequence, and the whole genome sequence determined by Illumina NovaSeq and Oxford Nanopore PromethION. Bioinformatics tools such as dbCAN, EasyCGTree, BRIG, and Easyfig were used to compare the strain FZY0027 withSaccharophagus degradans 2-40^T. The 3, 5-dinitrosalicylic acid (DNS) method was employed to measure the polysaccharide hydrolysis activity of strain FZY0027. [Results] The 16S rRNA gene sequence showed the similarity of 99.9% between strain FZY0027 andS.degradans 2-40^T, and thus strain FZY0027 was preliminarily identified asS.degradans FZY0027. The highest levels of reducing sugars (2.28, 1.75, and 1.10 mg/mL, respectively) were produced by FZY0027 through the hydrolysis of starch, xylan, and mannose. The genome of strain FZY0027 was 5 178 381 bp, encoding a total of 4 156 genes, with the G+C content of 45.8%. The average nucleotide identity (ANI), average amino acid identity (AAI), and digital DNA-DNA hybridization (dDDH) values between strain FZY0027 andS.degradans 2-40^T were 96.5%, 96.7%, and 70.0%, respectively. A total of 303 genes were annotated in the Carbohydrate-Active Enzyme database, and there was a significant difference in the number (137 and 130, respectively) of genes encoding glycoside hydrolases (GHs) between strain FZY0027 andS.degradans 2-40^T. Strain FZY0027 carried multiple genes involved in the hydrolysis of starch and xylan, which was corresponding to its strong ability to hydrolyse starch and xylan. However, compared withS.degradans 2-40^T, strain FZY0027 could only hydrolyse a few polysaccharides under the experimental conditions in this study, which suggested that this strain may require specific culture conditions to fully exert its polysaccharide hydrolysis ability. [Conclusion] Strain FZY0027 is a versatile polysaccharide-hydrolyzing bacterium with the potential for bioresource utilization.

Persulfidation plays a role in protein functioning and signaling, maintaining the physiological and metabolic balance of cells, protecting cells from oxidative stress, and regulating sulfur homeostasis. This article summarized the internal relationship of hydrogen sulfide, reactive sulfur species, and cysteine metabolism, expounded the mechanism of sulfur homeostasis regulation, and introduced the role of persulfidation in microbial sulfur homeostasis, providing new thoughts for the future research.

Equine coronavirus (ECoV) is an emerging virus attacking the gastrointestinal tract in horses, and the infected adult horses mainly present fever, abdominal pain, and diarrhea. In 1975, ECoV infection first appeared in the United States, and since then it has been prevalent in many countries and regions. Only one recombinant strain of ECoV was isolated from the small intestine of a donkey experiencing diarrhea in Shandong Province, China. [Objective] Understanding the genetic composition, genetic relationship, and biological characteristics of ECoV strains in China can lay a foundation for unveiling the epidemic status and genetic evolution trend of ECoV and provide materials for the research and development of products for the prevention and control of ECoV. [Methods] RT-PCR was employed to detect the stool samples from a horse experiencing diarrhea in Huangpi District, Wuhan City, Hubei Province. The virus was isolated from the positive samples and verified by indirect immunofluorescence assay (IFA) with monoclonal antibodies targeting the S1 protein of ECoV. Based on the whole genome sequencing results of the isolate, the phylogenetic analysis and sequence alignments of the whole genome, N gene, and NS2 gene were performed. [Results] An ECoV strain was successfully isolated and named ECoV-JL. Transmission electron microscopy (TEM) showed that the isolated virus particles were spheroidal and had a capsule membrane and a typical spiroid structure of coronaviruses. The tissue culture infectious dose 50% (TCID₅₀) of ECoV-JL reached a peak of 10^6.16 TCID₅₀/mL 72 h post infection in HRT-18 cells. ECoV-JL strains could infect three human cell lines: HRT-18 (human ileocecal cancer cells), Caco-2 (human colorectal adenocarcinoma cells), and Huh7 (human liver cancer cells). The genome sequence of ECoV-JL and the ECoV genome sequences in GenBank showed the similarity within the range of 97.9%–99.0%. ECoV-JL was in a separate branch of the phylogenetic tree and far related to other strains, which indicated that ECoV-JL might be derived from recombination mutations. The NS2 gene presented more mutations, and the difference in NS2 gene was the main reason for the poor homology between ECoV-JL and other strains. [Conclusion] We isolated and identified an ECoV strain from the stool samples of horses with diarrhea and named it ECOV-JL. The study about the biological characteristics and phylogenetic relationship of this strain reflected the characteristics of the epidemic strains in Hubei Province, providing a clue for the epidemic status and evolution trend of ECoV in China.

Fusarium head blight (FHB) caused byFusarium graminearum is one of the major diseases limiting wheat production. Biocontrol has been considered an effective and sustainable method for the control of crop diseases. [Objective] To screen out endophytic strains with inhibitory effects onF.graminearum from wheat grains and evaluate their biocontrol potential, providing strain resources and theoretical support for the development and utilization of biocontrol agents against FHB in wheat. [Methods] Plate confrontation, spore germination, and cell-free supernatant (CFS) inhibition experiments were carried out to screen out the endophytic strains with antagonistic activity againstF.graminearum from wheat grains. Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) were employed to observe the conidial morphology, membrane integrity, and mycelial reactive oxygen species ofF.graminearum treated with the CFS. Pot experiments were performed to verify the biocontrol effects of the endophytic bacteria on FHB. Illumina HiSeq was used for whole genome sequencing. [Results] A highly efficient endophytic strain JB7 inhibiting the growth ofF.graminearum was isolated from wheat grains. The CFS of strain JB7 in the decline stage showed the inhibition rate of 85.23% on the spore germination ofF.graminearum. Moreover, it led to concavity on spore surface, cell membrane damage, leakage of nucleic acids and proteins, and accumulation of reactive oxygen species in the mycelia ofF.graminearum. The CFS of strain JB7 significantly increased the content of soluble protein and malondialdehyde. Strain JB7 could produce protease, cellulase, glucanase, and siderophores. Pot experiments showed that strain JB7 decreased the disease index of FHB (P < 0.05) in wheat. The strain was identified asBacillus methylotrophic JB7 by whole genome sequencing, and it carried 12 gene clusters for the synthesis of secondary metabolites with antimicrobial functions. [Conclusion] Strain JB7 could inhibit the growth ofF.graminearum and demonstrated a strong control effect on FHB, serving as a candidate strain for the biocontrol of FHB in wheat.

[Objective] To investigate the community structure, diversity, and plant-growth-promoting functions of endophytic bacteria in the protocorms ofEpidendrum sp. and mine the core bacterial groups associated with the seed germination. [Methods] We collected five samples including three types of protocorms (germinated on MS1 medium, pine bark, and rotten oak leaves) and two types of substrates (pine bark and rotten oak leaves). High-throughput sequencing of the 16S rRNA gene was employed to compare the diversity and community composition of bacterial communities under different germination conditions. The conventional method was used to isolate the endophytic bacteria from the protocorms germinated on pine bark and rotten oak leaves (i.e., from symbiotic seed germination), and the growth-promoting potential of the isolates was evaluated. [Results] A total of 2 735 operational taxonomic units (OTUs) were obtained from the five samples, belonging to 876 genera, 453 families of 41 phyla, among whichProteobacteria andActinobacteria were the dominant phyla. The results of principal coordinates analysis showed that there were differences in the bacterial community structure between the protocorms ofEpidendrum sp. and the substrates, and the community structure of endophytic bacteria in the protocorms germinated on the MS1 medium was closest to that on pine bark. Functional prediction indicated that the endophytic bacteria in the protocorms germinated on rotten oak leaves had higher nitrogen-fixing ability than that in other types of protocorms. Nineteen isolates were obtained from protocorms geminated on pine bark and rotten oak leaves, belonging to 16 species of 12 genera.Tumebacillus flagellatus,Bradyrhizobium cenepequi, andWeizmannia ginsengihumi were the common species in the protocorms germinated on pine bark and rotten oak leaves.Pseudomonas koreensis andW.ginsengihumi had the potential of solubilizing phosphorus and producing indole-3-acetic acid (IAA) and siderophores. [Conclusion] TheEpidendrum sp. protocorms germinated in different environments harbor rich endophytic bacteria. The endophytic bacteria isolated from the protocorms from symbiotic seed germination had plant growth-promoting functions, such as fixing nitrogen, solubilizing phosphorus, and producing IAA and siderophores. This study provides a scientific basis for mining the microbial resources related to seed germination of orchids and studying the interactions between orchids and microorganisms.

[Objective] Streptococcus thermophilus is widely used in the dairy industry as a common starter for fermented dairy products such as yogurt and cheese. Most strains ofS.thermophilus are galactose-negative (Gal⁻) and unable to metabolize galactose and excrete it extracellularly, which results in an increase in the galactose content in fermented milk.S.thermophilus can be treated by chemical mutagenesis to metabolize galactose and then used to develop the fermented milk products with low galactose content. [Methods] We used nitrosoguanidine (NTG) to induce the mutation ofS.thermophilus IMAU80846. Furthermore, we measured the activities of β-galactosidase (β-Gal), galactokinase (GalK), pyruvate kinase (PK), and glucokinase (GK) in the wild-type and mutant strains ofS.thermophilus IMAU80846 and analyzed the amino acid sequences encoding these enzymes.S.thermophilus IMAU80846Y that could metabolize galactose was obtained. We performed whole genome sequencing of the mutant strain and measured the content of lactic acid, lactose, galactose, and glucose in the fermented milk products produced with the wild-type strain and the mutant strain. We then compounded the wild-type strain and the mutant strain withLactobacillus delbrueckii subsp.bulgaricus IMAU20450, respectively, and characterized the two groups of fermented milk products during fermentation and storage. Finally, we prepared a fermented milk product with low galactose content. [Results] S.thermophiles IMAU80846Y had higher activities of β-Gal and GalK and lower activities of PK and GK than the wild-type strain. The amino acid sequences and whole genome sequences showed that the mutant strain had mutations in the genes involved in carbohydrate metabolism. The HPLC results showed that the fermented milk produced with the mutant strain had lower content of lactose and galactose and higher content of lactic acid and glucose than that produced with the wild-type strain. Compared with the compound group with the wild-type strain, the compound group with the mutant strain improved the titration acidity, viable cell count, viscosity, and water holding capacity of fermented milk. [Conclusion] The mutagenesis with NTG changed the ability ofS.thermophilus IMAU80846 to metabolize galactose, and the mutant strain could be used to produce the fermented milk with low galactose content.

Atopic dermatitis (AD) is a highly prevalent allergic skin disease characterised by recurrent attacks and severe itching. The pathogenesis of AD involves a variety of factors including genetic susceptibility, epidermal barrier dysfunction, microbiome dysbiosis, immune imbalance, and the environment, while the available therapeutic drugs have severe side effects and limited efficacy. Studies have demonstrated that gut microbiota, particularly probiotics, play a role in AD. Probiotics can alleviate AD symptoms by inhibiting pathogens, enhancing barriers, improving the intestinal environment, and balancing the Th1/Th2 immune response, among other mechanisms. In this review, we summarized the skin and intestinal microecological characteristics of AD patients and systematically elucidated the mechanisms of probiotics in alleviating AD from the pathogenesis and influencing factors of AD, aiming to provide theoretical support for probiotics in the treatment of AD and related allergic skin diseases.

[Objective] Pseudoalteromonas, an endemic and dominant genus in the ocean, possesses multiple methyl-accepting chemotaxis proteins (MCPs), the functions of which remain unclear. [Methods] The chemotaxis ofPseudoalteromonas arabiensis N1230-9 isolated from surface seawater of the Pacific Ocean towards 23 carbon sources was analyzed by a plate-based assay for swimming motility. Two sCache domain-containing genes (woc28264 andwoc27036) encoding MCPs were deleted by homologous recombination. The chemotaxis of the two mutants to 10 carbon sources was then analyzed. [Results] Strain N1230-9 showed obvious chemotaxis to 10 carbon sources: trehalose, maltose, sucrose, N-acetylglucosamine, L-malic acid, sodium acetate, sodium propionate, sodium pyruvate, citric acid, and succinic acid. WOC28264 was a specific chemotactic receptor for L-malic acid and sucrose, while WOC27036 was a specific chemotactic receptor for citric acid and succinic acid. Both WOC28264 and WOC27036 were chemotactic receptors for N-acetylglucosamine and trehalose. [Conclusion] WOC28264 and WOC27036 have overlapping carbon source effectors.