甘草提取物对Staphylococcus aureus的抑菌活性及作用机理

食源性病菌为食品安全带来了巨大挑战,甘草提取物作为食品添加剂对金黄色葡萄球菌(Staphylococcus aureus)具有良好的抑菌作用,可作为天然食品防腐剂的候选原料,但目前对其抑菌机理的研究还不深入,影响了其应用。为探究甘草提取物对S.aureus的抑菌机理,该研究通过生长曲线、氧化损伤实验、细胞膜壁分析、蛋白质分析和DNA分析,评价了甘草提取物对S.aureus的抑菌作用机制。结果表明,甘草提取物导致S.aureus核酸渗漏,说明其膜完整性被破坏;同时,甘草提取物降低了几种能量代谢酶:琥珀酸脱氢酶(succinate dehydrogenase,SDH)和总ATP酶的活力;此外,光谱和竞争分析表明,甘草提取物与DNA发生了静电结合和凹槽结合。总之,甘草提取物主要是通过对S.aureus细胞壁膜、蛋白质合成、细菌代谢活力和遗传物质发挥作用,从而抑制其生长。该研究为甘草提取物在食品防腐方面的应用提供了理论基础。

金黄色葡萄球菌(Staphylococcus aureus)液体增菌培养基的优化

为研发一种金黄色葡萄球菌(Staphylococcus aureus)选择性增菌液,选用针对革兰氏阴性菌及非葡萄球菌属菌株具有选择抑制作用的7.5%氯化钠肉汤作为基础培养基,探索氨基酸、丙酮酸钠、甘露醇对金黄色葡萄球菌的促生长能力,通过正交试验确定培养基的最优配方,并使用优化后的培养基与原始培养基作用于来源不同的四株金黄色葡萄球菌标准菌株,对其增菌效果和抑制大肠杆菌(Escherichia coli)的效果进行对比。结果表明,最佳培养基配方为在基础培养基上添加L-丙氨酸1.0 g/L、甘露醇6.0 g/L、丙酮酸钠1.2 g/L。菌株在优化后培养基上增菌浊度及转接至Baird-Parker的菌落数上均有促进效果,最终菌株的OD550 nm值有0.28~0.34的增长,转接Baird-Parker后的菌落数也有10~100倍的增长,且在促进目标菌生长的同时对大肠杆菌的抑制性也没有受到影响,表明该增菌培养基选择性良好。

Staphylococcus aureus and biofilms:transmission, threats, and promising strategies in animal husbandry

Staphylococcus aureus(S. aureus) is a common pathogenic bacterium in animal husbandry that can cause diseases such as mastitis, skin infections, arthritis, and other ailments. The formation of biofilms threatens and exacerbates S. aureus infection by allowing the bacteria to adhere to pathological areas and livestock product surfaces, thus triggering animal health crises and safety issues with livestock products. To solve this problem, in this review, we provide a brief overview of the harm caused by S. aureus and its biofilms on livestock and animal byproducts(meat and dairy products). We also describe the ways in which S. aureus spreads in animals and the threats it poses to the livestock industry. The processes and molecular mechanisms involved in biofilm formation are then explained. Finally, we discuss strategies for the removal and eradication of S. aureus and biofilms in animal husbandry, including the use of antimicrobial peptides, plant extracts, nanoparticles, phages, and antibodies. These strategies to reduce the spread of S. aureus in animal husbandry help maintain livestock health and improve productivity to ensure the ecologically sustainable development of animal husbandry and the safety of livestock products.

Structural insights on anti-biofilm mechanism of heated slightly acidic electrolyzed water technology against multi-resistant Staphylococcus aureus biofilm on food contact surface

Slightly acidic electrolyzed water(SAEW)has proven to be an efficient and novel sanitizer in food and agriculture field.This study assessed the efficacy of SAEW(30 mg/L)at 40℃on the inactivation of foodbome pathogens and detachment of multi-resistant Staphylococcus aureus(MRSA)biofilm.Furthermore.the underlying mechanism of MRS A biofilm under heated SAEW at 40℃treatment on metabolic profiles was investigated.The results showed that the heated SAEW at 40℃significantly effectively against foodbome pathogens of 1.96-7.56(lg(CFU/g))reduction in pork,chicken,spinach,and lettuce.The heated SAEW at 40℃treatment significantly reduced MRS A biofilm cells by 2.41(lg(CFU/cm^(2))).The synergistic effect of SAEW treatment showed intense anti-biofilm activity in decreasing cell density and impairing biofilm cell membranes.Global metabolic response of MRSA biofilms,treated by SAEW at 40℃,revealed the alterations of intracellular metabolites,including amino acids,organic acid,fatty acid,and lipid.Moreover,signaling pathways involved in amino acid metabolism,energy metabolism,nucleotide synthesis,carbohydrate metabolites,and lipid biosynthesis were functionally disrupted by the SAEW at 40℃treatment.As per our knowledge,this is the first research to uncover the potential mechanism of heated SAEW treatment against MRSA biofilm on food contact surface.

Elimination of methicillin‑resistant Staphylococcus aureus biofilms on titanium implants via photothermally‑triggered nitric oxide and immunotherapy for enhanced osseointegration

Background:Treatment of methicillin-resistant Staphylococcus aureus(MRSA)biofilm infections in implant placement surgery is limited by the lack of antimicrobial activity of titanium(Ti)implants.There is a need to explore more effective approaches for the treatment of MRSA biofilm infections.Methods:Herein,an interfacial functionalization strategy is proposed by the integration of mesoporous polydopamine nanoparticles(PDA),nitric oxide(NO)release donor sodium nitroprusside(SNP)and osteogenic growth peptide(OGP)onto Ti implants,denoted as Ti-PDA@SNP-OGP.The physical and chemical properties of Ti-PDA@SNP-OGP were assessed by scanning electron microscopy,X-ray photoelectron spectroscope,water contact angle,photothermal property and NO release behavior.The synergistic antibacterial effect and elimination of the MRSA biofilms were evaluated by 2′,7′-dichlorofluorescein diacetate probe,1-N-phenylnaphthylamine assay,adenosine triphosphate intensity,O-nitrophenyl-β-D-galactopyranoside hydrolysis activity,bicinchoninic acid leakage.Fluorescence staining,assays for alkaline phosphatase activity,collagen secretion and extracellular matrix mineralization,quantitative real‑time reverse transcription‑polymerase chain reaction,and enzyme-linked immunosorbent assay(ELISA)were used to evaluate the inflammatory response and osteogenic ability in bone marrow stromal cells(MSCs),RAW264.7 cells and their co-culture system.Giemsa staining,ELISA,micro-CT,hematoxylin and eosin,Masson's trichrome and immunohistochemistry staining were used to evaluate the eradication of MRSA biofilms,inhibition of inflammatory response,and promotion of osseointegration of Ti-PDA@SNP-OGP in vivo.Results:Ti-PDA@SNP-OGP displayed a synergistic photothermal and NO-dependent antibacterial effect against MRSA following near-infrared light(NIR)irradiation,and effectively eliminated the formed MRSA biofilms by inducing reactive oxygen species(ROS)-mediated oxidative stress,destroying bacterial membrane integrity and causing leakage of intracellular components(P<0.01).In vitro experiments revealed that Ti-PDA@SNP-OGP not only facilitated osteogenic differentiation of MSCs,but also promoted the polarization of pro-inflammatory M1 macrophages to the anti-inflammatory M2-phenotype(P<0.05 or P<0.01).The favorable osteo-immune microenvironment further facilitated osteogenesis of MSCs and the anti-inflammation of RAW264.7 cells via multiple paracrine signaling pathways(P<0.01).In vivo evaluation confirmed the aforementioned results and revealed that Ti-PDA@SNP-OGP induced ameliorative osseointegration in an MRSA-infected femoral defect implantation model(P<0.01).Conclusions:Ti-PDA@SNP-OGP is a promising multi-functional material for the high-efficient treatment of MRSA infections in implant replacement surgeries.

The ever-changing microenvironment of Staphylococcus aureus in cutaneous infections

Background:Staphylococcus aureus is responsible for the majority of skin and soft tissue infections,which are often diagnosed at a late stage,thereby impacting treatment efficacy.Our study was designed to reveal the physiological changes at different stages of infection by S.aureus through the combined analysis of variations in the skin microenvironment,providing insights for the diagnosis and treatment of S.aureus infections.Methods:We established a murine model of skin and soft tissue infection with S.aureus as the infectious agent to investigate the differences in the microenvironment at different stages of infection.By combining analysis of the host immune status and histological observations,we elucidate the progression of S.aureus infection in mice.Results:The results indicate that the infection process in mice can be divided into at least two stages:early infection(1–3 days post-i nfection)and late infection(5–7 days post-i nfection).During the early stage of infection,notable symptoms such as erythema and abundant exudate at the infection site were observed.Histological examination revealed infiltration of numerous neutrophils and bacterial clusters,accompanied by elevated levels of cytokines(IL-6,IL-10).There was a decrease in microbial alpha diversity within the microenvironment(Shannon,Faith's PD,Chao1,Observed species,Simpson,Pielou's E).In contrast,during the late stage of infection,a reduction or even absence of exudate was observed at the infected site,accompanied by the formation of scabs.Additionally,there was evidence of fibroblast proliferation and neovascularization.The levels of cytokines and microbial composition gradually returned to a healthy state.Conclusion:This study reveals synchrony between microbial composition and histological/immunological changes during S.aureus-i nduced SSTIs.

Eugenol targeting CrtM inhibits the biosynthesis of staphyloxanthin in Staphylococcus aureus

Staphylococcus aureus is a serious foodborne pathogen threatening food safety and public health.Especially the emergence of methicillin-resistant Staphylococcus aureus(MRSA)increased the difficulty of S.aureus treatment.Staphyloxanthin is a crucial virulence factor of S.aureus.Blocking staphyloxanthin production could help the host immune system counteract the invading S.aureus cells.In this study,we first screened for staphyloxanthin inhibitors using a virtual screening method.The outcome of the virtual screening method resulted in the identification of eugenol(300μg/mL),which significantly inhibits the staphyloxanthin production in S.aureus ATCC 29213,S.aureus Newman,MRSA ATCC 43300 and MRSA ATCC BAA1717by 84.2%,63.5%,68.1%,and 79.5%,respectively.The outcome of the growth curve assay,field-emission scanning electron,and confocal laser scanning microscopy analyses confirmed that eugenol at the test concentration did not affect the morphology and growth of S.aureus.Moreover,the survival rate of S.aureus ATCC 29213 and MRSA ATCC 43300 under H_(2)O_(2) pressure decreased to 51.9%and 45.5%in the presence of eugenol,respectively.The quantitative RT-PCR and molecular simulation studies revealed that eugenol targets staphyloxanthin biosynthesis by downregulating the transcription of the crtM gene and inhibiting the activity of the CrtM enzyme.Taken together,we first determined that eugenol was a prominent compound for staphyloxanthin inhibitor to combat S.aureus especially MRSA infections.

Electrochemical and colorimetric dual-signal detection of Staphylococcus aureus enterotoxin B based on AuPt bimetallic nanoparticles loaded Fe-N-C single atom nanocomposite

Sensitive detection of Staphylococcus aureus enterotoxin B(SEB)is of importance for preventing food poisoning from threatening human health.In this work,an electrochemical and colorimetric dual-signal detection assay of SEB was developed.The probe(Ab2/AuPt@Fe-N-C)was bound to SEB captured by Ab1,where the Ab2/AuPt@Fe-N-C triggered methylene blue degradation and resulted in the decrease of electrochemical signal.Furthermore,the probe catalyzed the oxidation of 3,3’,5,5’-tetramethyl biphenyl to generate a colorimetric absorbance at 652 nm.Once the target was captured and formed a sandwich-like complex,the color changed from colorless to blue.SEB detection by colorimetric and electrochemical methods showed a linear relationship in the concentration ranges of 0.0002-10.0000 and 0.0005-10.0000 ng/mL,with limits of detection of 0.0667 and 0.1670 pg/mL,respectively.The dual-signal biosensor was successfully used to detect SEB in milk and water samples,which has great potential in toxin detection in food and the environment.

Antimicrobial and synergistic effects of lemongrass and geranium essential oils against Streptococcus mutans,Staphylococcus aureus,and Candida spp.

BACKGROUND The oral cavity harbors more than 700 species of bacteria,which play crucial roles in the development of various oral diseases including caries,endodontic infection,periodontal infection,and diverse oral diseases.AIM To investigate the antimicrobial action of Cymbopogon Schoenanthus and Pelargonium graveolens essential oils against Streptococcus mutans,Staphylococcus aureus,Candida albicans,Ca.dubliniensis,and Ca.krusei.METHODS Minimum microbicidal concentration was determined following Clinical and Laboratory Standards Institute documents.The synergistic antimicrobial activity was evaluated using the Broth microdilution checkerboard method,and the antibiofilm activity was evaluated with the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay.Data were analyzed by one-way analysis of variance followed by the Tukey post-hoc test(P≤0.05).RESULTS C.schoenanthus and P.graveolens essential oils were as effective as 0.12%chlorhexidine against S.mutans and St.aureus monotypic biofilms after 24 h.After 24 h P.graveolens essential oil at 0.25%was more effective than the nystatin group,and C.schoenanthus essential oil at 0.25%was as effective as the nystatin group.CONCLUSION C.schoenanthus and P.graveolens essential oils are effective against S.mutans,St.aureus,Ca.albicans,Ca.dubliniensis,and Ca.krusei at different concentrations after 5 min and 24 h.

葡萄球菌Staphylococcus condimenti对广式腊肠风味的影响

采用固相微萃取方法结合气质联用仪,对Staphylococcus condimenti发酵广式腊肠中风味物质成分进行测定,分析接种分离菌株对广式腊肠风味的影响。从对照组和接种组(SC)中均鉴定出22种风味物质成分,主要为醇类、醛类、酮类和酯类物质,还有部分烃类和环类化合物。研究表明,Staphylococcus condiment可以加速脂肪和蛋白质的降解与氧化,有助于腊肠风味的改善。

Cloning and Prokaryotic Expression of FnBP Ligand Binding Gene of Staphylococcus aureus

[Objective] The study aimed to clone the FnBP ligand binding gene of Staphylococcus aureus and run prokaryotic expression by constructing a prokaryotic expression vector. [Method] The gene encoding FnBP ligand binding gene was amplified from S.aureus chromosomal DNA by PCR technique. After T-A cloning, plasmid pMD18- FnBP was constructed. pMD18- FnBP and pET28a(+)were digested by BamH Ⅰ and EcoR Ⅰ double enzymes, then the purified FnBP ligand binding gene was subcloned into the expression vector pET28a(+), and the prokaryotic expression vector pET28a-FnBP was thus constructed. The constructed plasmid pET28a-FnBP was transformed into Escherichia coli BL21(DE3) competent cells. The bacterium was induced by IPTG and the expressed products were analyzed by SDS-PAGE and Western blot. [Result] The gene fragment with the length of 370 bp was amplified by PCR approach. One approximately 30 kD exogenous protein was observed in SDS-PAGE analysis. Western blot analysis indicates the protein has antigenicity of S.aureus. [Conclusion] The FnBP ligand binding gene of S.aureus was successfully cloned and expressed in prokaryotic cells.

The Effect of Culture Condition on Type 5 Capsular Polysaccharide Production of Staphylococcus aureus from Diary Cattle

[Objective] The effect of different culture conditions on type 5 capsular polysaccharide production of Staphylococcus aureus from diary cattle was studied to provide simple way for CP production and preparation and laid foundation for carrying out new polysaccharide vaccine research. [Method] Staph-ylococcus aureus was isolated from milk sample of sick dairy cattle and capsular polysaccharide serotypes were identified. Type 5 capsular polysaccharide was cultured on BHI,solid columbia and mod110 culture media. Glucose and lactose were taken as carbon sources for every culture media in solid and liquid state. Therefore 9 different culture conditions were taken to study the effect of culture conditions on capsular polysaccharide production. [Result] Different culture conditions indicated that compared with columbia culture media, BHI culture media could decline capsular polysaccharide production and mod110 culture media could increase capsular polysaccharide production. While for same culture media, solid culture media was better for capsular polysaccharide production,meanwhile,taken lactose as carbon source could increase capsular polysaccharide production.

Inhibitory Effects of Bacillus subtilis on Staphylococcus aureus

[Objective] To produce drug resistance, seek non-toxic environmental so as to change the current biological drugs that did not excessive use of antibiotics. [Methods] A strain of Bacillus was purified and isolated from fresh and healthy in- testines of grass carps. Biochemical identification was carried out by conventional bacterial biochemical test method. Two pairs of primers were designed, 16S rRNA detection and sequencing analysis were carried out. Drug sensitive test was carried out by agar diffusion method. In vitro inhibition test on Staphylococcus aureus was carried out by Oxford cup method. [Results] The isolated bacterium had basically the same biochemical characters as Bacillus subtilis; and the homology reached 100%. Thus, the isolated bacterium was identified to be Bacillus subtilis. It was insensitive to amoxicillin, ampicillin, penicillin G and so on, but sensitive to amikacin, cefalexin, ciprofloxacin and cefradine. The inhibitory effects of Bacillus subtilis on Staphylococ- cus aureus were significant. The minimum inhibitory concentration （MIC） was 2.8×10^8×2^-5/ml and minimum bactericidal concentration （MBC） was 2.8×10^8×2^-2/ml. [Conclusions] The isolated Bacillus subtilis could be used to prevent and control diseases caused by Staphylococcus aureus, and reduce the abuse of antibiotics.

叶绿素镁钠盐对液态食品中Staphylococcus aureus的光动力杀菌研究

以叶绿素镁钠盐为光敏剂,研究了其对几种液体食品中金黄色葡萄球菌的光动力杀菌效果。结果表明,叶绿素镁钠盐对几种液态食品中的金黄色葡萄球菌具有很强的杀菌效果,其中以10-5mol/L的叶绿素镁钠盐杀菌效果最佳,10 min金黄色葡萄球菌减少了4.5个对数;随着光照时间的延长,液态食品中的金黄色葡萄球菌的致死率明显增加,特别是在起始10 min内;一般透光性好的食品包装材料对液态食品中金黄色葡萄球菌光动力杀菌效率没有影响,但液态食品的pH对其杀菌效率有一定的影响。对于不同的液态食品,在澄清或酸性的食品中叶绿色镁钠盐的光动力杀菌效果较好,但当食品中含固体颗粒或较浑浊时,其杀菌效率显著降低。

一株甲胺磷高效降解菌——巴氏葡萄球菌(Staphylococcus pasteuri)的筛选及其分子鉴定

从湘潭南天化工厂的甲胺磷废水和污泥中分离细菌样品,以甲胺磷为唯一碳源和能源,经过定向筛选,得到1株可高效降解甲胺磷的菌株HN003,气相色谱测定其对甲胺磷的平均降解效率在24h、48h和72h分别达到45.8%、88.5%和100%。对其进行常规生理生化测试,结果表明,菌株HN003革兰氏染色为阳性,在10-42℃、pH3.0-pH14.0范围内均能生长,并与巴氏葡萄球菌的表型特征非常相似。为了进一步确定HN003的分类学地位,测定了其16S rRNA基因序列(1541bp),分析了相关细菌相应序列的同源性,构建了分子系统发育树,结果表明,菌株HN003与巴氏葡萄球菌的亲缘关系最近。综合上述结果,菌株HN003可鉴定为巴氏葡萄球菌(Staphylococcus pasteuri)。

Antibacterial Activity of Herbal Preparations against Staphylococcus aureus and Streptococcus agalactiae in Cow Mastitis

[Objective] This study aimed to analyze the antibacterial activity of herbal preparations against Staphylococcus aureus and Streptococcus agalactiae in cow mastitis. [Method] The crude drug solutions of four different prescriptions for Zengrujianniusan were prepared through reflux extraction. Their antibacterial activity in vitro against Staphylococcus aureus and Streptococcus agalactiae in cow mastitis were investigated. [Result] All the four different prescriptions exhibited antibacterial activity against S. aureus and S. agalactiae. Among them, prescription Ⅲ was extremely sensitive, and had the best bactericidal effect. The other three prescriptions were highly sensitive. [Conclusion] This study provides a theoretical basis for the development of herbal preparations for the treatment of cow mastitis.

TMPD降解菌Staphylococcus pasteuri的鉴定

通过形态学观察、生理生化特征测定和16S rDNA序列同源性分析,对1株TMPD降解菌(LF-2)进行了鉴定。结果表明:LF-2的形态及主要生理生化特征与葡萄球菌属一致,而且与巴氏葡萄球菌(AJ717376)的16S rDNA序列具有99.80%的同源性;在系统发育树上构成一个分支。由此确定TMPD降解菌LF-2属于巴氏葡萄球菌(Staphylococcus pasteuri)。

从Staphylococcus simulans消除含有溶葡球菌酶基因质粒的研究

溶葡球菌酶是Staphylococcus simulans分泌的能分解葡萄球菌的酶,它的基因位于一个约40kb的质粒DNA上。为了探索用高拷贝质粒取代原有的质粒的可能性,本文首先进行了从该菌株中消除含有溶葡球菌酶基因质粒的实验研究,获得了相应的“消除”菌株。根据对目的菌株和原始菌株的比较分析,包括细胞蛋白质的SDS-聚丙烯酰胺凝胶电泳,Western blot分析,质粒DNA的琼脂糖胶电泳、Southern Blot分析,质粒DNA的限制性内切核酸酶酶切分析以及对溶葡球菌酶作用敏感性的分析,都表明该目的菌株确系Staphylococcus simulans的衍生菌株,只是清除了其中含有溶葡球菌酶基因的质粒。在此基础上,本文也进行了转化实验。

生物胺降解菌Staphylococcus xylosus JCM 2418的特性研究及应用

从鱼露中筛选获得一株具有生物胺降解能力的木糖葡萄球菌——Staphylococcus xylosus JCM 2418,对其安全性、发酵性能进行评价同时对生物胺降解相关酶展开分析,并将其接种至发酵食品中进行应用,监测发酵食品生物胺降解率以及风味的变化情况。结果表明,S.xylosus JCM 2418表现出弱耐药性,不具有溶血性,不产生生物膜;具有良好的耐盐性、蛋白酶和脂肪酶活性;生物胺降解相关酶为细胞膜上的胺氧化酶;应用在发酵食品中,可以有效控制生物胺的积累,同时改善发酵产品的风味,为提升发酵产品品质提供优良菌种。

Characterization of Staphylococcus aureus Isolated from Clinical Specimens by Matrix Assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry

Objective To develop a matrix assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS） approach to identify Staphylococcus aureus （5. aureus） and differentiate methicillin-resistant 5. aureus （MRSA） from methicillin-sensitive S. aureus （MSSA）. Methods A total of 100 5. aureus strains isolated from clinical specimens and farm workers were collected and analyzed by MALDI-TOF-MS. And data obtained were interpreted with biotyper software. Results Ninety-two strains were identified by MALDI-TOF-MS as 5. aureus at a level of secure genus and probable species, and 4 strains were identified at probable genus after their cultivation, spectral collection and data preprocessing. One strain was identified as 5. aureus with lower score. It was revealed that identification of 5. aureus by MALDI-TOF-MS was highly correlated with typing by biochemical and serological methods with an accuracy as high as 97%. The biotyper cluster analysis showed that 100 isolates were divided into 2 types at the distance level of 400. Higher peak intensity in the mass of both 3784 Da and 5700 Da was observed in MRSA, whereas that was absent from MSSA. Conclusion MALDI-TOF-MS is considered as a simple, high-throughput and accuracy for the identification of S from MSSA. rapid and highly reproducible technique with aureus and it can reliably differentiate MRSA