Abstract Background Bacteriocins are peptides that demonstrate antimicrobial activity and are produced by bacteria. Bacillus velezensis is considered a reliable producer of bacteriocins, with a strong antimicrobial spectrum and antimicrobial activity. In this study, B. velezensis G7 isolated from mangrove plant roots was used as the experimental strain. Whole-genome sequencing (WGS) analysis was performed to annotate functional genes. Results The results demonstrated that B. velezensis G7 has a genome size of 3,894,836 bp, a GC content of 46.67%, and predicts 3,715 CDSs, 10 rRNA genes, and 83 tRNA genes. Gene annotation revealed pathways related to bacteriocin synthesis and immunity. Identified 8 bacteriocin and secondary metabolite biosynthetic gene clusters in B. velezensis G7, including Sactipeptides, LCI, Amylocyclicin, ComX4, Mersacidin, Lanthipeptide, Bacilysin, and NRPS. The crude protein from B. velezensis G7 shows broad-spectrum antibacterial activity, with the strongest inhibition against Listeria monocytogenes and the weakest against Staphylococcus aureus . Conclusions The genome of B. velezensis G7 was analyzed, and bacteriocin-related genes were identified. The results suggest that B. velezensis G7 has the potential to produce bacteriocins, thereby providing a theoretical basis for future studies on bacteriocin production in B. velezensis . In addition, these findings may facilitate further advances in bacteriocin research and bioinformatics analysis.

The global rise of antimicrobial resistance has intensified the demand for novel antimicrobial agents with broad-spectrum efficacy and unique mechanisms of action. Herein, a marine-derived strain, Bacillus velezensis (B. velezensis) AM12, exhibiting clear inhibitory activity against eight major foodborne pathogens, was isolated from coastal seawater near Zhanjiang, China. Whole-genome sequencing revealed a 3,992,266 bp circular chromosome with a GC content of 46

Bacteriocins are peptides produced by bacteria that have antibacterial effects on other bacterial species. They have emerged as novel antimicrobial alternatives or feed additives in the field of veterinary medicine. Heterologous expression systems have been demonstrated to be capable of high-level expression of bacteriocins when bacteriocins are used in commercial and industrial contexts. The bacteriocin amylocyclicin was expressed in Escherichia coli BL21 (DE3), and the induction conditions were optimized by a single-factor method. The composition of the induction medium was optimized using response surface methodology (RSM). The induction conditions that yielded optimal results were an isopropylthio-β-galactopyranoside (IPTG) concentration of 0.5 mM, an induction temperature of 28 ℃, and an induction time of 6 h. The optimal sucrose, ethanol, and NaCl additions were 0.804 g/100 mL, 1.547

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) and its robust biofilm-forming capability pose severe threats to public health, livestock production, and food safety, and underscores the urgent need for novel antibacterial and anti-biofilm agents. In this study, we identified and characterized a novel bacteriocin, PFB252, derived from Bacillus velezensis through a multistep purification process involving acid precipitation, TA-GF75 gel column chromatography, Tiderose Q HP anion-exchange chromatography (TRUKING, Changsha, China), and reversed-phase HPLC. PFB252 exhibited remarkable thermal stability, pH tolerance, and resistance to enzymatic degradation, and demonstrated potent antibacterial activity against MRSA. At subinhibitory concentrations (1/32× minimum inhibitory concentration [MIC] and 1/16× MIC), PFB252 significantly disrupted biofilm formation and impaired the metabolic viability of embedded bacteria, and it drastically reduced extracellular polysaccharide, the key component of the biofilm matrix. Transcriptional analysis further revealed that PFB252 at subinhibitory concentrations downregulated critical biofilm-associated genes. PFB252 exhibited strong antimicrobial efficacy in dairy applications and could reduce MRSA counts in milk from 103 to <10 cfu/mL within 4 d at MIC and maintain suppression in cheese below 102 cfu/g over 7 d. These properties highlight PFB252's potential as a natural biopreservative for combating MRSA in food systems and offer a promising solution for food safety applications.

Guangdong Province is an important area of poultry breeding in China. Zhanjiang city is located in the western part of Guangdong Province, where there are many broiler farms. To investigate antimicrobial resistance (AMR) and the presence of resistance genes in Escherichia coli from broiler farms, a total of 220 samples were collected from soil and feces at eight broiler farms. Subsequently, 220 strains of E. coli were isolated for drug resistance analysis and detection of AMR genes. The results revealed that the isolated E. coli strains exhibited high prevalence of multidrug resistance to 12 antimicrobial drugs including amoxicillin, tetracycline, cotrimoxazole and sulfisoxazole. Among the isolated strains, 95% of the isolates were resistant to more than three antimicrobial agents; notably, thirty-nine strains showed multidrug resistance to ten tested drugs, while four strain exhibits multidrug resistance to as many as fifteen antibacterial drugs. Additionally, seven AMR genes such as blaTEM and sul2 were detected in over half (≥50%) of the isolated E. coli samples; thirteen AMR genes had relatively low detection prevalence (≤30%). Correlation analysis indicated a strong association between certain AMR genes (blaTEM, pexA, aadA1, blaAIM, ant(3")-I, sul2, sul3, tet(D)) and AMR (≥50%). In conclusion, E.coli strains obtained from soil and fecal samples in broiler farms exhibited multidrug resistant phenotypes along with carrying various AMR genes. This provides a reference for the scientific control of E. coli multidrug resistance in this area.

Listeria monocytogenes is a significant zoonotic pathogen capable of forming biofilms on food and other materials, representing a considerable risk to human health and animal husbandry. The use of bacteriocins as potential new antibacterial and antibiofilm reagents has attracted considerable interest. This study aimed to determine the inhibitory effects of bacteriocin PCM7−4 on L. monocytogenes biofilm formation. In this study, bacteriocin PCM7−4 of SICs (1/16 × MIC, 1/8 × MIC) significantly inhibited the formation of L. monocytogenes biofilm. Bacteriocin PCM7−4 of SICs significantly reduced the production of bacterial extracellular polysaccharides, and could decrease the bacterial motility, meanwhile, PCM7−4 significantly reduced the number and viability of bacteria within the biofilm. RT-qPCR results showed that bacteriocin PCM7−4 significantly reduced the expression of flagella, community sensing and virulence factor genes associated with biofilm formation. The results demonstrated the considerable potential of bacteriocin PCM7−4 as a therapeutic agent for the prevention and treatment of L. monocytogenes biofilms.

Bacteriocins are defined as proteins that are produced by bacteria and that have antibacterial effects on various pathogenic bacteria. This study explored the effects of the crude extract of bacteriocin produced by Bacillus velezensis G02 (PRJNA1303554) on the intestinal microbiota of normal mice. Observation through liver and kidney tissue sections showed that the bacteriocin crude extract had no obvious adverse reactions on the internal organs of mice, and no mice died. A 16 S rDNA amplicon sequencing analysis revealed that the crude extract of the bacteriocin significantly changed the α-diversity (Shannon (P = 0.51), Simpson (P = 0.51), and Chao1 (P = 0.28)) and β-diversity (PCoA, (P = 0.1) NMDS, (P = 0.0132)) of the intestinal microbiota in normal mice, reducing the species abundance and increasing the evenness of their intestinal microbiota. In addition, the bacteriocin PG02 crude extract exhibited broad-spectrum antibacterial activity against both gram-positive and gram-negative bacteria. The purified bacteriocin was identified by LC-MS/MS in combination with sequence coverage analysis; its molecular weight was found to be 20.1 31 kDa, and it was speculated to be a putative novel bacteriocin named bacteriocin PG02. This study provides a reference for the microbiology field research and development of feed additive.

IntroductionBacteriocin P7 was extracted from the cell-free supernatant (CFS) of Bacillus velezensis G7, which is a strain isolated from mangrove plants.MethodsIn this study, the culture conditions of B. velezensis G7 were optimised using an orthogonal test. The (CFS) was subsequently purified by using TA-GF75 gel chromatography, Tiderose Q HP anion chromatography and reversed-phase high-performance liquid chromatography (RP-HPLC). Finally, the bacteriocin was identified by using LC-MS/MS.Results and discussionThe optimal culture conditions for B. velezensis G7 are 4.5 g/100 mL glucose, 1.5 g/100 mL yeast, and 1.2 g/100 mL MgSO4·7H2O. The stability of the CFS is affected by several factors, including heat, UV treatment and different storage conditions. High temperatures and long UV irradiation treatments significantly reduce the stability of CFS, which is more sensitive to strong acids, bases and enzymatic degradation. The minimum inhibitory concentration (MIC) of purified bacteriocin P7 against S. aureus was determined to be 30.352 μg/mL. On the basis of the results of the haemolytic activity assay, it was concluded that the use of bacteriocin P7 at concentrations equal to or below the 2 × MIC is safe. The addition of organic solvents and inorganic salts did not affect the bacteriocin P7, while the incorporation of SDS could enhance its antimicrobial efficacy. The bacteriocin was subjected to analysis by LC-MS/MS, which revealed that it was similar to the class I bacteriocin amyloliquecidin GF610. The findings of the present study indicate that the endophytic B. velezensis G7 from mangrove plant can produce bacteriocins, thereby providing a reference point for the expansion of novel bacteriocin sources.

Listeria monocytogenes, a pathogenic bacterium causing zoonotic diseases, necessitates the urgent search for novel anti-Listeria monocytogenes drugs due to the continuous emergence of drug-resistant bacteria. In this study, we isolated and identified a bacteriocin-producing strain CM7-4 from seawater as Bacillus velezensis through 16S rRNA sequence analysis. Moreover, we successfully purified a novel bacteriocin named PCM7-4 from Bacillus velezensis CM7-4. The molecular weight of PCM7-4 was determined to be 40,228.99 Da. Notably, PCM7-4 exhibited broad-spectrum antibacterial activity against both Gram-positive and Gram-negative bacteria with a minimum inhibitory concentration (MIC) of 5.625 mu g/mL against Listeria monocytogenes specifically. It demonstrated heat resistance and high stability within the pH range of 2-12 while being sensitive to proteinase K degradation without any observed hemolytic activity. Furthermore, SEM analysis revealed that PCM7-4 effectively inhibited biofilm formation and disrupted cell membranes in Listeria monocytogenes cells. Transcriptome analysis revealed that PCM7-4 exerts an impact on genes associated with crucial metabolic pathways, encompassing the biosynthesis of secondary metabolites, phosphotransferase systems (PTS), and starch/sucrose metabolism. These findings highlight the significant potential of bacteriocin PCM7-4 for the development of effective antimicrobial interventions targeting food-borne pathogenic bacteria.

To investigate the impact of florfenicol on soil microbial community structure and diversity, an indoor florfenicol exposure model was established. Soil samples were collected at different concentrations of florfenicol (0, 0.05, 0.5, 5, and 50 mg/kg) on days 0, 7, 30, and 60. High-throughput sequencing was employed to examine the changes in soil microbial community structure, diversity, and abundance. The analysis revealed a total of 31874 operational taxonomic units ( OTUs ) in the soil samples, averaging 9524 OTUs per sample. The number of soil microbial community OTUs declined with increasing florfenicol concentration. The microbial species richness and diversity showed a decreasing trend at day 7, while the treatment group with 50 mg/kg florfenicol at 60 days exhibited the lowest richness index. Examination of the soil microbial community structure identified 50 phyla and 1303 genera. At the phylum level, the abundance of Actinobacteria and Bacteroides decreased with increasing florfenicol concentration. Similarly, at the genus level, some of the dominant genera displayed a decline in abundance with the rise of florfenicol concentration. Cluster analysis demonstrated significant temporal and concentration variability. The results indicate differences in the composition, diversity, dominance, abundance, and evenness of soil bacterial communities among different groups. Florfenicol has a significant negative impact on the structure and diversity of soil microbial communities. The findings of this study provide a scientific basis for the rational use of florfenicol in livestock farming to maintain a healthy and stable soil microecological environment.

Introduction: The antibacterial protein PAG14 was extracted from a metabolite of Bacillus G14 isolated from mangrove plants.Methods: In this study, Pseudomonas aeruginosa, Pasteurell multocide, Enterobacter aerogenes, and Enterococcus faecalis were used as indicator bacteria to screen endophytes that exhibited antibacterial activity. The endophyte culture conditions were optimized to enhance productivity. Subsequently, the culture supernatant was salted using ammonium sulfate, followed by purification using dextran gel chromatography and ion exchange column techniques. Finally, the structures of antibacterial proteins were identified using mass spectrometry.Results and Discussion: The optimal culture conditions for Bacillus G14 were 2% mannitol, 0.5% fish peptone, 0.05% KH2PO4 + 0.05% K2HPO4 + 0.025% MnSO4·H2O. The antibacterial substances exhibited stability within the temperature range of 30–40°C and pH range of 5.0–7.0, while displaying sensitivity toward enzymes. The antibacterial activity decreased as the duration of UV irradiation increased. The antibacterial protein PAG14, isolated from the culture broth of Bacillus G14 through purification using dextran gel and ion-exchange columns, was identified as a class III bacteriocin using LC-MS/MS, similar to Lysozyme C. These findings serve as a theoretical foundation for the investigation and application of bacteriocins in food products.

为了从红树林植物根际土壤中分离出具有广谱抑菌作用的海洋细菌,试验以大肠杆菌、金黄色葡萄球菌、沙门氏菌、链球菌4种常见致病菌为指示菌,采用M10和P3培养基从红树植物根际土壤中分离细菌,以固体琼脂打孔法对分离的细菌进行抑菌活性筛选,并提取分离菌株的基因组DNA,PCR扩增16S rDNA基因并测序,同时采用邻接法构建系统发育树.结果表明:从红树植物土壤中分离得到654株细菌,其中107株细菌具有抑菌活性,占分离菌株的16.4％.菌株M36对大肠杆菌和链球菌抑菌活性最强,抑菌圈直径分别为13 mm、16 mm;菌株P19和P25对金黄色葡萄球的抑菌活性最强,抑菌圈直径均为16 mm;菌株P83对沙门氏菌抑菌活性最强,抑菌圈直径为13 mm;菌株M36和P31的抑菌谱最广.综合两种培养基的培养结果有抑菌活性的菌株被分成4个属,分别是肠杆菌属、芽孢杆菌属、葡萄球菌属、伯克霍尔德氏菌属,其中肠杆菌属为优势菌属,其次为芽孢杆菌属,占比分别为69.2％和15.9％.说明红树植物根际土壤中存在具有广谱抑菌活性的细菌.

Florfenicol has a significant therapeutic effect on animal bacterial diseases, most of metabolites enter the soil in the form of metabolites in feces or urine and pollute environment. Soil ecological model was established by experiment to investigate the effects of florfenicol residues in soil on 16S rDNA sequence diversity of phosphorus-solubilizing bacteria. Five different concentrations of florfenicol (0 mg center dot kg-1, 0.1 mg center dot kg-1, 1 mg center dot kg-1, 10 mg center dot kg-1 and 100 mg center dot kg-1) were used to collect soil samples on the 7 d, 21 d and 49 d after dosing. The effects of florfenicol on 16S rDNA sequence diversity of soil phosphorus-solubilizing bacteria were determined by amplifed ribosomal DNA restriction analysis (ARDRA) and enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) method. The results showed that the number of Operational Taxonomic Units (OTUs) types decreased with the increase of florfenicol concentration, and the percentage of OTUs number to bacteria were the lowest at 100 mg center dot kg-1 florfenicol concentration after 21d treating, which was 8.33%. The phosphorus-solubilizing bacteria were amplified by ERIC-PCR after 21d treating, the fingerprint type of ERIC-PCR decreased with the increase of drug concentration, and the diversity index of the drug group was significantly lower than that of the blank control group. This indicated that florfenicol had an effect on the dominance, richness and evenness of soil phosphorus-solubilizing bacteria community.

Introduction:The widespread use of antibiotics in animal agriculture has increased the resistance of Escherichia coli, and pathogenic E. coli often harbor complex virulence factors. Antimicrobial resistance in pathogenic bacteria can cause public health problems. Correlation analyses of the resistance, virulence, and serotype data from the pathogenic bacteria found on farms and in the surrounding environment can thus provide extremely valuable data to help improve public health management.Methods:In this investigation, we have assessed the drug resistance and virulence genes as well as the molecular typing characteristics of 30 E. coli strains isolated from duck farms in the Zhanjiang area of China. Polymerase chain reaction was used to detect the drug resistance and virulence genes as well as serotypes, and whole-genome sequencing was used to analyze the multilocus sequence typing.Results:The detection rates for the oqxA resistance gene and fimC virulence gene were highest (93.3%, respectively). There were no correlations between the drug resistance and virulence gene numbers in the same strain. The epidemic serotype was O81 (5/24), ST3856 was an epidemic sequence type, and strains I-9 and III-6 carried 11 virulence genes. The E. coli strains from the duck farms in the Zhanjiang area were thus found to have a broad drug resistance spectrum, various virulence genes, complex serotypes, and certain pathogenicity and genetic relationship.Discussion:Monitoring the spread of pathogenic bacteria and the provision of guidance regarding the use of antibiotics in the livestock and poultry industries will be required in the future in the Zhanjiang area.

Florfenicol is used worldwide for its low side effects and strong bactericidal effect. Florfenicol is physicochemically stable and can persist in natural water bodies and affect water denitrification. Indoor aquatic microcosm models were constructed and water samples were collected at different florfenicol concentrations (0.1, 1, 10, and 100 mg/L) on days 0, 7, 30, and 60 to extract the microbial genome DNA and determine the water properties. qPCR and amplicon sequencing were used to study the dynamic changes of nirS gene and nirS-type denitrifying communities structure, diversity and abundance, respectively. The results showed that higher florfenicol concentrations caused accumulation of nitrate and ammonium nitrogen in water. Florfenicol stress caused orders of magnitude changes in nirS gene abundance, showing a trend of increasing first and then decreasing. 100 mg/L florfenicol addition led to a sustained increase of nirS gene abundance in water bodies. The florfenicol addition altered denitrifying community structure and suppressed the richness and diversity index of denitrifying bacteria in water body. Over time, the richness and diversity index gradually recovered. Proteobacteria was always the dominant denitrifying phylum in water. The relative abundance of Pseudomonas and beta proteobacterium showed obvious positive correlation with nirS gene abundance and were the dominant genera under florfenicol stress. Our study provided a scientific basis for the rational use of florfenicol in aquaculture to maintain a healthy and stable microecological environment, and also provided a preliminary understanding of the response characteristics of water denitrifying microorganisms to florfenicol exposure.

从土壤中分离得到10株解磷菌,将菌株接种至LB液体培养基中,30℃培养24 h作为种子液.在无机磷液体培养基中加入氟苯尼考,使培养基含药浓度分别为0、0.1、1.0、10.0、50.0 μg/mL.按照1%的接种量接种种子液,30℃培养5 d,每24 h取2 mL培养液,利用钼锑抗分光光度法测定培养液中速效磷含量.结果表明,10株解磷菌在50.0 μg/mL药物浓度下,培养基中速效磷含量明显低于其他药物浓度组;大部分菌株在药物浓度0～50.0 μg/mL菌液中细菌数量变化不明显.说明氟苯尼考对解磷菌的解磷能力产生了抑制作用,但与细菌的生长繁殖并无明显相关性.

Florfenicol is one of the most widely used antibiotics in aquaculture and veterinary clinics because of its low side effects and strong bactericidal effect. A total of 45~60% of florfenicol is not absorbed by the animal body and accumulates in the aquatic environment through a variety of pathways, which affects denitrification. Indoor aquatic microcosm models were constructed and sediment samples were collected at different florfenicol concentrations (0.1, 1, 10, and 100 mg/L) on days 0, 7, 30, and 60 to extract the microbial genome DNA and determine the water properties. qPCR and amplicon sequencing were used to study the dynamic changes in the nirS gene and nirS-type denitrification community structure, diversity, and abundance, respectively. The results showed that high florfenicol stress influenced the sediment's physicochemical properties, reducing conductivity, alkaline dissolved nitrogen, and organic matter content. In addition, the abundance of nirS, a functional denitrification gene, increased obviously with increased florfenicol concentrations but decreased the diversity of nirS-type denitrification microorganisms. Proteobacteria was the dominant denitrifying phylum in the sediment. Our study provides a scientific basis for the rational use of florfenicol in aquaculture to maintain a healthy and stable microecological environment and also provides a preliminary understanding of the response characteristics of water denitrifying microorganisms to florfenicol exposure.

In order to explore the impact of antibiotics (enrofloxacin) on microbial community in aquatic environment, an indoor aquatic ecological model was built, and different concentrations of enrofloxacin (0.05, 0.5, 5, and 50 mg/L) were added in the aquatic ecological model. In addition, the water and sediment samples were collected on the 0, 7, 30, and 60 days, and the changes in microbial community were studied through 16S rDNA high-throughput sequencing. The results showed that when the concentration of enrofloxacin was 50 mg/L, the relative abundance of Actinomycetes was increased. In the water, the bacterial richness and diversity communities first decreased and then gradually recovered with the passage of time; On the 7th day, the diversity and richness index of species in the treatment groups with enrofloxacin at 5 and 50 mg/L decreased to the lowest; On the 30th day, the diversity and richness index of species began to rise; On the 60th day, the diversity index and richness index of water species began to increase, while the diversity index and richness index of sediment species decreased. In conclusion, the addition of enrofloxacin negatively affected the microbial community structure in an indoor aquatic ecological model, 50 mg/L enrofloxacin could increase the relative abundance of Actinomycetes, and decrease the diversity and richness index of water and sediment.

[目的]建立水体模型,探究氟苯尼考对水生细菌耐药性的影响.[方法]将水体样品分成Ⅰ、Ⅱ、Ⅲ、Ⅳ、Ⅴ组,每组3个重复,各组添加不同浓度的氟苯尼考.分离培养建模前以及建模后第14、28、56天水生细菌,利用微量肉汤二倍稀释法和KB药敏纸片法对分离的水生细菌分别进行氟苯尼考和17种抗菌药物的敏感性试验.[结果]第56天收集的Ⅴ组水生细菌对氟苯尼考的耐药率最高,达到25.00％;分离的氟苯尼考敏感菌和耐药菌对阿莫西林、链霉素和多西环素等8种药物的敏感性差异极显著(P<0.01).[结论]氟苯尼考在一定程度上增加了水生细菌的耐药性.

This study evaluates the effects of a broad-spectrum antibiotic (florfenicol) on antibiotic resistance genes (ARGs) and bacterial community structure in aquatic environments. We constructed an indoor aquatic microcosm model, adding different concentrations of florfenicol (0.1, 1, 10, 100 mg L−1), and water and sediment samples were collected after 0, 7, 30, and 60 days. qPCR and 16S rDNA amplicon sequencing were used to study the changes in the ARGs and bacterial community structure of the collected samples. The results show that the inclusion of florfenicol resulted in an increased abundance of the floR and optrA genes. Adding 100 mg L−1 florfenicol to the water increased the abundance of optrA gene copies with the maximum on the Day 7, and increased the abundance of floR gene copies with the maximum on Day 30. Adding 100 mg L−1 florfenicol to the sediment increased the abundance of floR and optrA genes by one order of magnitude on Day 60. Meanwhile, the average number of operational taxonomic units (OTUs) in the water samples was 257, and the average number of OTUs in sediment samples was 823. The bacterial community diversity and richness in sediments were higher than those in water. The difference between the maximal and minimal values of the Shannon diversity index in the water and sediment samples was 4.36 and 1.95, respectively. The effect of florfenicol on the bacterial community structure in water was much higher than that in sediment. At 30 days, the diversity index and richness index of the florfenicol treatment groups with 1 and 10 mg L−1 concentrations began to increase; at 60 days, the diversity and richness indices of the 100 mg L−1 florfenicol treatment group began to increase. The samples at the same sampling time in the sediments clustered closer together. The results of this study provide a scientific basis for guiding the rational use of florfenicol in aquaculture, maintaining a healthy and stable microecological environment in aquaculture, and provide theoretical data for environmental ecological risk assessment and safety management caused by microbial resistance under the abuse of florfenicol.