Probable enterotoxin-associated toxic shock syndrome caused by Staphylococcus epidermidis.

BACKGROUND: We describe a case of a toxic shock-like syndrome in a child, which was associated with Staphylococcus epidermidis instead of Staphylococcus aureus or Streptococcus pyogenes, the usual causes of toxic shock syndrome. CASE PRESENTATION: The patient was an 8-year-old boy who developed a toxic shock syndrome-like illness, including fever, hypotension, and rash. The Staphylococcus epidermidis isolate was cultured from urine, but this organism was unavailable for toxin testing. Multiple blood cultures were negative. Instead, a highly novel assay was used on acute plasma from the patient which demonstrated the presence of the genes for superantigens, staphylococcal enterotoxins A, C, D, and E. Superantigens are the known causes of toxic shock syndrome. CONCLUSIONS: Our study suggests strongly that Staphylococcus epidermidis was causing the TSS symptoms through the known Staphylococcus aureus superantigens. It is unknown how many other such patients exist; this should be explored. Of great importance is that PCR performed directly on blood plasma in the absence of microbial isolation could be used to demonstrate superantigen genes.

Inhibition of Candida albicans and Staphylococcus epidermidis mixed biofilm formation in a catheter disk model system treated with EtOH-EDTA solution.

Microbial colonization and the formation of biofilms on catheter surfaces pose a great risk for medical-related infections. We aimed (a) to evaluate polymicrobial biofilm formation of Candida albicans and Staphylococcus epidermidis and (b) to investigate the inhibition and effects of ethanol (EtOH) and EtOH-EDTA solutions on biofilms. Catheter disks were made and used as a substrate for biofilm formation. Varying concentrations of EtOH and EtOH-EDTA solutions were compared in deterring biofilm formation. The EtOH-EDTA solutions were further tested to remove mature and preformed biofilms. Compared to their monospecies counterparts, biofilm concentration significantly increases when C. albicans is co-cultured with S. epidermidis. Moreover, all treatments with EtOH-EDTA solution significantly lowered biofilm formation compared to EtOH alone (P ≤ 0.05). Lastly, biofilm was dramatically reduced when treated with 20%, 30%, 40%, and 50% EtOH-EDTA solutions (P ≤ 0.05). Our findings suggest that biofilms become more resilient to treatment when formed by multiple organisms. Nonetheless, treatment with EtOH-EDTA is effective against these polymicrobial biofilms.

Photocatalytic Inactivation of Co-Culture of E. coli and S. epidermidis Using APTES-Modified TiO2.

The presented work shows the antibacterial activity of TiO2 photocatalysts modified by 3-aminopropyltriethoxysilane (APTES). The APTES-functionalized TiO2 samples were obtained by the solvothermal process followed by calcination. The antibacterial activity of APTES/TiO2 samples was evaluated with two species of bacteria, Escherichia coli and Staphylococcus epidermidis, under artificial solar light (ASL) irradiation. The used bacteria are model organisms characterized by negative zeta potential (approx. -44.2 mV for E. coli and -42.3 mV for S. epidermidis). For the first time, the antibacterial properties of APTES-functionalized TiO2 were evaluated against mono- and co-cultured bacteria. The high antibacterial properties characterized the obtained APTES-modified nanomaterials. The best antibacterial properties were presented in the TiO2-4 h-120 °C-300 mM-Ar-300 °C sample (modified with 300 mM of APTES and calcined at 300 °C). The improvement of the antibacterial properties was attributed to a positive value of zeta potential, high surface area, and porous volume.

Emerging resistance in Staphylococcus epidermidis during dalbavancin exposure: a case report and in vitro analysis of isolates from prosthetic joint infections.

BACKGROUND: Dalbavancin, a semisynthetic lipoglycopeptide with exceptionally long half-life and Gram-positive spectrum, is an attractive option for infections requiring prolonged therapy, including prosthetic joint infections (PJIs). OBJECTIVES: To investigate the prevalence of reduced susceptibility to dalbavancin in a strain collection of Staphylococcus epidermidis from PJIs, and to investigate genomic variation in isolates with reduced susceptibility selected during growth under dalbavancin exposure. METHODS: MIC determination was performed on S. epidermidis isolates from a strain collection (n = 64) and from one patient with emerging resistance during treatment (n = 4). These isolates were subsequently cultured on dalbavancin-containing agar and evaluated at 48 h; MIC determination was repeated if phenotypical heterogeneity was detected during growth. Population analysis profile (PAP-AUC) was performed in isolates where a  ≥ 2-fold increase in MIC was detected, together with corresponding parental isolates (n = 21). Finally, WGS was performed. RESULTS: All strains grew at 48 h on agar containing 0.125 mg/L dalbavancin. PAP-AUC demonstrated significant differences between parental and derived strains in four of the eight analysed groups. An amino acid change in the walK gene coinciding with emergence of phenotypic resistance was detected in the patient isolates, whereas no alterations were found in this region in the in vitro derived strains. CONCLUSIONS: Exposure to dalbavancin may lead to reduced susceptibility to dalbavancin through either selection of pre-existing subpopulations, epigenetic changes or spontaneous mutations during antibiotic exposure. Source control combined with adequate antibiotic concentrations may be important to prevent emerging reduced susceptibility during dalbavancin treatment.

Stability and Resistance to Proteolysis of Enterotoxins SEC and SEL Produced by Staphylococcus epidermidis and Staphylococcus aureus.

The only staphylococcal enterotoxins produced by Staphylococcus epidermidis include SECepi and SELepi, whereas Staphylococcus aureus produces orthologous SECs and SEL having different sequences. We compared S. epidermidis and S. aureus SECs and SELs in terms of resistance to proteolysis and both, thermal and chemical stability. We show that SECepi and SELepi produced by S. epidermidis have similar resistance to proteolysis if compared with their respective orthologues produced by S. aureus. Studied S. epidermidis and S. aureus SEC variants incubated with pepsin at pH 2.0 were found to be more resistant to proteolysis than SELs. SELs turned out to be more resistant than SECs to proteolysis with trypsin at pH 8.0. SECepi was found to be more resistant to thermal denaturation if compared with its S. aureus orthologues. The S. epidermidis and S. aureus SEC variants were found to have higher thermal stability than SELs. Our data indicate that, due to their high stability, the enterotoxins SECepi and SELepi produced in food by S. epidermidis may pose a food safety risk comparable with that posed by S. aureus enterotoxins.

Molecular Detection and Characterization of the Staphylococcus epidermidis and Staphylococcus haemolyticus Isolated from Hospitalized Patients and Healthcare Workers in Iran.

Background: The present study is aimed at surveying the antibiotics resistance profile, biofilm formation ability, staphylococcal cassette chromosome mec (SCCmec) types, and molecular epidemiology of Staphylococcus epidermidis and Staphylococcus haemolyticus isolated from hospitalized patients and healthcare workers in four teaching hospitals in Iran. Methods: In total, 43 Staphylococcus epidermidis and 12 Staphylococcus haemolyticus were isolated from hospitalized patients, and 19 Staphylococcus epidermidis and 7 Staphylococcus haemolyticus isolated from healthcare workers were included in the present study. The antimicrobial resistance profile of isolates was determined using the disk diffusion method. Moreover, the resistance of isolates to methicillin was identified using the cefoxitin disk diffusion test. The microtiter-plate test was used for quantifying biofilm formation. Moreover, the frequency of icaA and icaD genes was determined using PCR assay. The molecular epidemiology of methicillin-resistant isolates was determined using SCCmec typing and pulsed-field gel electrophoresis methods. Results: Among all coagulase-negative staphylococci isolates, the highest resistance rate (81.5%) was seen for cefoxitin and cotrimoxazole. All of the isolates were susceptible to linezolid. Out of the 66 mecA-positive isolates, the most common SCCmec type was the type I (n = 23; 34.8%) followed by type IV (n = 13; 19.7%). Using pulsed-field gel electrophoresis (PFGE) assay, 27 PFGE types including 14 common types and 13 singletons were obtained among 51 methicillin-resistant S. epidermidis (MRSE) isolates. Moreover, among 12 methicillin-resistant S. haemolyticus (MRSH) isolates, 8 PFGE types were detected, of which 5 PFGE types were singletons. Conclusion: The high rate of resistance to antibiotics as well as the possibility of cross-infection shows the importance of a pattern shift in the management and controlling programs of coagulase-negative staphylococci, especially in healthcare centers. Clinical trial registration. The present study is not a clinical trial study. Thus, a registration number is not required.

Effect of Antibiotic Exposure on Staphylococcus epidermidis Responsible for Catheter-Related Bacteremia.

Coagulase-negative staphylococci (CoNS) and especially Staphylococcus epidermidis are responsible for health care infections, notably in the presence of foreign material (e.g., venous or central-line catheters). Catheter-related bacteremia (CRB) increases health care costs and mortality. The aim of our study was to evaluate the impact of 15 days of antibiotic exposure (ceftobiprole, daptomycin, linezolid and vancomycin) at sub-inhibitory concentration on the resistance, fitness and genome evolution of 36 clinical strains of S. epidermidis responsible for CRB. Resistance was evaluated by antibiogram, the ability to adapt metabolism by the Biofilm Ring test® and the in vivo nematode virulence model. The impact of antibiotic exposure was determined by whole-genome sequencing (WGS) and biofilm formation experiments. We observed that S. epidermidis strains presented a wide variety of virulence potential and biofilm formation. After antibiotic exposure, S. epidermidis strains adapted their fitness with an increase in biofilm formation. Antibiotic exposure also affected genes involved in resistance and was responsible for cross-resistance between vancomycin, daptomycin and ceftobiprole. Our data confirmed that antibiotic exposure modified bacterial pathogenicity and the emergence of resistant bacteria.

Undesirable consequences of the metallic nanoparticles action on the properties and functioning of Escherichia coli, Bacillus cereus and Staphylococcus epidermidis membranes.

Controversial and inconsistent findings on the toxicity of metallic nanoparticles (NPs) against many bacteria are common in recorded studies; therefore, further advanced experimental work is needed to elucidate the mechanisms underlying nanotoxicity. This study deciphered the direct effects of Ag-NPs, Cu-NPs, ZnO-NPs and TiO2-NPs on membrane permeability, cytoplasmic leakage, ATP level, ATPase activity and fatty acid profiling of Escherichia coli, Bacillus cereus and Staphylococcus epidermidis as model microorganisms. A multifaceted analysis of all collected results indicated the different influences of individual NPs on the measured parameters depending on their type and concentration. Predominantly, membrane permeability was correlated with increased cytoplasmic leakage, reduced total ATP levels and ATPase activity. The established fatty acid profiles were unique and concerned various changes in the percentages of hydroxyl, cyclopropane, branched and unsaturated fatty acids. Decisively, E. coli was more susceptible to changes in measured parameters than B. cereus and S. epidermidis. Also, it was established that ZnO-NPs and Cu-NPs had a major differentiating impact on studied parameters. Additionally, bacterial cell imaging using scanning electron microscopy elucidated different NPs distributions on the cell surface. The presented results are believed to provide novel, valuable and accumulated knowledge in the understanding of NPs action on bacterial membranes.

Antibacterial Activity on Orthopedic Clinical Isolates and Cytotoxicity of the Antimicrobial Peptide Dadapin-1.

In orthopedic surgery, biomaterial-associated infections represent a complication of serious concern. Most promising strategies to prevent these infections currently rely on the use of anti-infective biomaterials. Desirably, in anti-infective biomaterials, the antibacterial properties should be achieved by doping, grafting, or coating the material surfaces with molecules that are alternative to conventional antibiotics and exhibit a potent and highly specific activity against bacteria, without altering the biocompatibility. Antimicrobial peptides (AMPs) are among the most interesting candidate molecules for this biomaterial functionalization. Here, the potential expressed by the recently discovered peptide Dadapin-1 was explored by assaying its MIC, MBIC and MBC on clinical strains of relevant bacterial species isolated from orthopedic infections and by assessing its cytotoxicity on the human osteoblast-like MG63 cells. When appropriately tested in diluted Mueller Hinton Broth II (MHB II), Dadapin-1 exhibited significant antibacterial properties. MIC values were in the range of 3.1-6.2 µM for the gram-positive bacteria Staphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus warneri, and 12.4-24.9 µM for the gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa. Interestingly, the peptide was found non-cytotoxic, with an IC50 exceeding the highest concentration tested of 179 µM. Overall, Dadapin-1 expresses considerable potential for future application in the production of anti-infective biomaterials.

Antibiotic resistance and drug modification: Synthesis, characterization and bioactivity of newly modified potent pleuromutilin derivatives with a substituted piperazine moiety.

Antibiotic-resistant bacteria pose a major global public health concern, owing to the lack of effective antibacterial drugs. Consequently, the discovery and development of innovative antibacterial drug classes with unique mechanisms of action are urgently needed. In this study, we designed, synthesised, and tested a series of novel pleuromutilin derivatives with piperazine linker substituted by amino acids moieties to determine their antibacterial properties. Most synthesized compounds exhibited potent activities against Staphylococcus aureus (S. aureus), methicillin-resistant S. aureus (MRSA), and methicillin-resistant Staphylococcus epidermidis. Compound 6l, the most potent antibacterial agent created in this study, displayed a rapid bactericidal activity against MRSA, Klebsiella pneumoniae and S. aureus cfr N12. Moreover, pharmacokinetics study of compound 6l exhibited good PK performance with a low in vivo clearance (CL = 1965 mL/h/kg) and a suitable half-life (T1/2 = 11.614 ± 5.123 h). Molecular docking experiments revealed the binding model of compound 6l to the unmethylated A2503 of peptidyl transferase centre of 23S rRNA. Interaction pattern of 6l with cfr-mediated ribosomes revealed by molecular dynamics. Moreover in vivo mouse systemic infection experiments with compound 6l revealed its effectiveness against MRSA and S. aureus cfr N12 with the ED50 of 11.08 mg/kg and 14.63 mg/kg body weight, respectively.

Adhesion of Yeast and Bacteria to Oral Surfaces.

Colonization of surfaces in the human body by microorganisms is an early, essential, step in the initiation of infectious disease. We have developed in vitro assays to investigate interactions between yeast or bacterial cells and human tissues, fluids, or prostheses. Such assays can be used to identify the adhesins, ligands, and receptors involved in these interactions, for example, by determining which components of the microbe or human tissue/fluid interfere with adherence in the assay. The assays can also be applied to find ways of preventing adhesion, and subsequent disease, by investigating the effects of different conditions and added compounds on adherence in the in vitro assays. Here we describe assays for measuring adhesion of the oral yeast Candida albicans, a common commensal and opportunistic pathogen, or the bacterium Staphylococcus epidermidis, which is not normally pathogenic but is known to form biofilms on medical prostheses. The assays described belong to two approaches to investigating adhesion and biofilm formation: (i) retention at a fixed time point following liquid washes, and (ii) retention against a continuous flow of medium.

Attachment and optimization of Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa biofilms to a 3D printed lattice.

A lattice was designed and fabricated using three-dimensional (3D) printing that allows for the facile transfer of biofilms formed from either Staphylococcus aureus, Staphylococcus epidermidis, or Pseudomonas aeruginosa into a fresh cell culture flask. To enhance biofilm production onto the filaments, three protein-based treatments were compared: fetal bovine serum (FBS), bovine serum albumin (BSA), and fibrinogen (Fb). Protein treatments included either supplementing the growth broths or pre-coating the lattice prior to immersion into the broth. S. aureus and P. aeruginosa biofilms were observed on all tested filaments that contained the supplement Fb. S. epidermidis required BSA to form biofilm. Ultimately, polycarbonate (PC) was chosen as the optimal material for lattice creation since it can be autoclaved without warping key design features. In addition, this 3D printed design may facilitate biofilm transfer from the bacterial culture to different cell culture platforms.

Reduction of Staphylococcus epidermidis in the mammary tumor microbiota induces antitumor immunity and decreases breast cancer aggressiveness.

The mammary gland hosts a microbiota, which differs between malignant versus normal tissue. We found that aerosolized antibiotics decrease murine mammary tumor growth and strongly limit lung metastasis. Oral absorbable antibiotics also reduced mammary tumors. In ampicillin-treated nodules, the immune microenvironment consisted of an M1 profile and improved T cell/macrophage infiltration. In these tumors, we noted an under-representation of microbial recognition and complement pathways, supported by TLR2/TLR7 protein and C3-fragment deposition reduction. By 16S rRNA gene profiling, we observed increased Staphylococcus levels in untreated tumors, among which we isolated Staphylococcus epidermidis, which had potent inflammatory activity and increased Tregs. Conversely, oral ampicillin lowered Staphylococcus epidermidis in mammary tumors and expanded bacteria promoting an M1 phenotype and reducing MDSCs and tumor growth. Ampicillin/paclitaxel combination improved the chemotherapeutic efficacy. Notably, an Amp-like signature, based on genes differentially expressed in murine tumors, identified breast cancer patients with better prognosis and high immune infiltration that correlated with a bacteria response signature. This study highlights the significant influence of mammary tumor microbiota on local immune status and the relevance of its treatment with antibiotics, in combination with breast cancer therapies.

Mutation of gdpS gene induces a viable but non-culturable state in Staphylococcus epidermidis and changes in the global transcriptional profile.

BACKGROUND: In the genome of staphylococci, only the gdpS gene encodes the conserved GGDEF domain, which is the characteristic of diguanylate cyclases. In our previous study, we have demonstrated that the gdpS gene can modulate biofilm formation by positively regulating the expression of ica operon in Staphylococcus epidermidis. Moreover, this regulation seems to be independent of the c-di-GMP signaling pathway and the protein-coding function of this gene. Therefore, the biological function of the gdpS gene remains to be further investigated. RESULTS: In the present study, it was observed that mutation of the gdpS gene induced S. epidermidis to enter into a presumed viable but nonculturable state (VBNC) after cryopreservation with glycerol. Similarly, when moved from liquid to solid culture medium, the gdpS mutant strain also exhibited a VBNC state. Compared with the wild-type strain, the gdpS mutant strain autolyzed more quickly during storage at 4℃, indicating its increased susceptibility to low temperature. Transcriptional profiling analysis showed that the gdpS mutation affected the transcription of 188 genes (92 genes were upregulated and 96 genes were downregulated). Specifically, genes responsible for glycerol metabolism were most markedly upregulated and most of the altered genes in the mutant strain are those involved in nitrogen metabolism. In addition, the most significantly downregulated genes included the betB gene, whose product catalyzes the synthesis of glycine betaine and confers tolerance to cold. CONCLUSION: The preliminary results suggest that the gdpS gene may participate in VBNC formation of S. epidermidis in face of adverse environmental factors, which is probably achieved by regulating expression of energy metabolism genes. Besides, the gdpS gene is critical for S. epidermidis to survive low temperature, and the underlying mechanism may be partly explained by its influence on expression of betB gene.

Transcriptional adaptation of staphylococci during colonization of the authentic human environment: An overview of transcriptomic changes and their relationship to physiological conditions.

Staphylococci are commensals of human skin and mucous membranes, but some species can also cause serious infections. Host niches during both colonization and infection differ greatly and are characterized by specific environmental conditions (pH, temperature, oxygen, nutrient availability, and microbiota) that can affect gene expression and virulence of microbes. To successfully occupy extremely different habitats at different anatomical sites, Staphylococci are equipped with a variety of regulatory elements that allow specific adaptation to the changing environments. Not surprisingly, gene expression in vivo can be significantly different from the expression pattern observed in vitro. Niche specific stimuli that influence the bacterial ability to either cause infection or maintain colonization are only partially understood. Here, we describe habitat specific conditions and discuss the available literature analyzing staphylococcal gene expression, focusing on Staphylococcus aureus and S. epidermidis during colonization of the nose and skin.

CATASAN Is a New Anti-Biofilm Agent Produced by the Marine Antarctic Bacterium Psychrobacter sp. TAE2020.

The development of new approaches to prevent microbial surface adhesion and biofilm formation is an emerging need following the growing understanding of the impact of biofilm-related infections on human health. Staphylococcus epidermidis, with its ability to form biofilm and colonize biomaterials, represents the most frequent causative agent involved in infections of medical devices. In the research of new anti-biofilm agents against S. epidermidis biofilm, Antarctic marine bacteria represent an untapped reservoir of biodiversity. In the present study, the attention was focused on Psychrobacter sp. TAE2020, an Antarctic marine bacterium that produces molecules able to impair the initial attachment of S. epidermidis strains to the polystyrene surface. The setup of suitable purification protocols allowed the identification by NMR spectroscopy and LC-MS/MS analysis of a protein-polysaccharide complex named CATASAN. This complex proved to be a very effective anti-biofilm agent. Indeed, it not only interferes with cell surface attachment, but also prevents biofilm formation and affects the mature biofilm matrix structure of S. epidermidis. Moreover, CATASAN is endowed with a good emulsification activity in a wide range of pH and temperature. Therefore, its use can be easily extended to different biotechnological applications.

Critical roles for 'housekeeping' nucleases in type III CRISPR-Cas immunity.

CRISPR-Cas systems are a family of adaptive immune systems that use small CRISPR RNAs (crRNAs) and CRISPR-associated (Cas) nucleases to protect prokaryotes from invading plasmids and viruses (i.e., phages). Type III systems launch a multilayered immune response that relies upon both Cas and non-Cas cellular nucleases, and although the functions of Cas components have been well described, the identities and roles of non-Cas participants remain poorly understood. Previously, we showed that the type III-A CRISPR-Cas system in Staphylococcus epidermidis employs two degradosome-associated nucleases, PNPase and RNase J2, to promote crRNA maturation and eliminate invading nucleic acids (Chou-Zheng and Hatoum-Aslan, 2019). Here, we identify RNase R as a third 'housekeeping' nuclease critical for immunity. We show that RNase R works in concert with PNPase to complete crRNA maturation and identify specific interactions with Csm5, a member of the type III effector complex, which facilitate nuclease recruitment/stimulation. Furthermore, we demonstrate that RNase R and PNPase are required to maintain robust anti-plasmid immunity, particularly when targeted transcripts are sparse. Altogether, our findings expand the known repertoire of accessory nucleases required for type III immunity and highlight the remarkable capacity of these systems to interface with diverse cellular pathways to ensure successful defense.

Anti-biofilm activity of Pseudomonas fluorescens culture supernatants on biofilm formation of Staphylococcus epidermidis 1457.

OBJECTIVE: Staphylococcus epidermidis is a skin colonizer and a major cause of nosocomial infections that can lead to sepsis. It causes opportunistic infections by forming biofilms on medical devices, which are hard to control with conventional antibiotics. In an attempt to develop its biofilm inhibitors, the culture supernatant (CS) of Pseudomonas fluorescens was assessed. This study examined the effect of the CS on S. epidermidis 1457 biofilm formation, the characteristics of inhibitors in the CS, and the differential gene expression of S. epidermidis when treated with the CS. RESULTS: P. fluorescens CS specifically targeted the maturation stage of S. epidermidis biofilm formation while not affecting planktonic growth. RT-qPCR analysis revealed that P. fluorescens CS significantly downregulated S. epidermidis ica genes and upregulated an ica repressor, tcaR. This indicates that the CS reduced polysaccharide intercellular adhesin synthesis, a major component of the S. epidermidis 1457 biofilm matrix. Further studies are required to elucidate the specific inhibitory components in the CS and their mechanism of action. Our results indicate that inhibitors in the P. fluorescens CS may have a significant value for inhibiting S. epidermidis biofilm. Combinations of specific inhibitors from the CS and antibiotics against staphylococci may provide an effective measure to control S. epidermidis biofilm formation while avoiding antibiotic resistance and compensating the attenuated effectiveness of antibiotics on biofilms.

A model for microbial interactions and metabolomic alterations in Candida glabrata-Staphylococcus epidermidis dual-species biofilms.

The fungus Candida glabrata and the bacterium Staphylococcus epidermidis are important biofilm-forming microorganisms responsible of nosocomial infections in patients. In addition to causing single-species disease, these microorganisms are also involved in polymicrobial infections leading to an increased antimicrobial resistance. To expand knowledge about polymicrobial biofilms, in this study we investigate the formation of single- and dual-species biofilms of these two opportunistic pathogens employing several complementary approaches. First, biofilm biomass, biofilm metabolic activity and the microbial composition in single- and dual-species biofilms were assessed and compared. Then, the expression of three genes of C. glabrata and three genes of S. epidermidis positively related to the process of biofilm formation was evaluated. Although S. epidermidis is a stronger biofilm producer than C. glabrata, both biological and genetic data indicate that S. epidermidis growth is inhibited by C. glabrata which dominates the dual-species biofilms. To better understand the mechanisms of the interactions between the two microorganisms, a broad GC-MS metabolomic dataset of extracellular metabolites for planktonic, single- and dual-species biofilm cultures of C. glabrata and S. epidermidis was collected. As demonstrated by Partial Least Squares Discriminant Analysis (PLS-DA) of GC-MS metabolomic data, planktonic cultures, single- and dual-species biofilms can be sharply differentiated from each other by the nature and levels of an assortment of primary and secondary metabolites secreted in the culture medium. However, according to our data, 2-phenylethanol (secreted by C. glabrata) and the synergistically combined antifungal activity of 3-phenyllactic acid and of the cyclic dipeptide cyclo-(l-Pro-l-Trp) (secreted by S. epidermidis) play a major role in the race of the two microorganisms for predominance and survival.

The nitric oxide synthase gene negatively regulates biofilm formation in Staphylococcus epidermidis.

Staphylococcus epidermidis (S. epidermidis) is a clinically important conditioned pathogen that can cause a troublesome chronic implant-related infection once a biofilm is formed. The nitric oxide synthase (NOS) gene, which is responsible for endogenous nitric oxide synthesis, has already been found in the genome of S. epidermidis; however, the specific mechanisms associated with the effects of NOS on S. epidermidis pathogenicity are still unknown. The purpose of the current study was to investigate whether the NOS gene has an impact on biofilm formation in S. epidermidis. Bioinformatics analysis of the NOS gene was performed, and homologous recombination was subsequently employed to delete this gene. The effects of the NOS gene on biofilm formation of S. epidermidis and its underlying mechanisms were analyzed by bacterial growth assays, biofilm semiquantitative determination, Triton X-100-induced autolysis assays, and bacterial biofilm dispersal assays. Additionally, the transcription levels of fbe, aap, icaA, icaR and sigB, which are related to biofilm formation, were further investigated by qRT-PCR following NOS deletion. Phylogenetic analysis revealed that the NOS gene was conserved between bacterial species originating from different genera. The NOS deletion strain of S. epidermidis 1457 and its counterpart were successfully constructed. Disruption of the NOS gene resulted in significantly enhanced biofilm formation, slightly retarded bacterial growth, a markedly decreased autolysis rate, and drastically weakened bacterial biofilm dispersal. Our data showed that the fbe, aap and icaA genes were significantly upregulated, while the icaR and sigB genes were significantly downregulated, compared with the wild strain. Therefore, these data strongly suggested that the NOS gene can negatively regulate biofilm formation in S. epidermidis by affecting biofilm aggregation and dispersal.