Co-transcriptional DNA and RNA Cleavage during Type III CRISPR-Cas Immunity.

Immune systems must recognize and destroy different pathogens that threaten the host. CRISPR-Cas immune systems protect prokaryotes from viral and plasmid infection utilizing small CRISPR RNAs that are complementary to the invader's genome and specify the targets of RNA-guided Cas nucleases. Type III CRISPR-Cas immunity requires target transcription, and whereas genetic studies demonstrated DNA targeting, in vitro data have shown crRNA-guided RNA cleavage. The molecular mechanism behind these disparate activities is not known. Here, we show that transcription across the targets of the Staphylococcus epidermidis type III-A CRISPR-Cas system results in the cleavage of the target DNA and its transcripts, mediated by independent active sites within the Cas10-Csm ribonucleoprotein effector complex. Immunity against plasmids and DNA viruses requires DNA, but not RNA, cleavage activity. Our studies reveal a highly versatile mechanism of CRISPR immunity that can defend microorganisms against diverse DNA and RNA invaders.

A CRISPR View of Cleavage.

Seminal studies showed that CRISPR-Cas systems provide adaptive immunity in prokaryotes and promising gene-editing tools from bacteria to humans. Yet, reports diverged on whether some CRISPR systems naturally target DNA or RNA. Here, Samai and colleagues unify the studies, showing that a single type III CRISPR-Cas system cleaves both DNA and RNA targets, independently.

Staphylococcal Periscope proteins Aap, SasG, and Pls project noncanonical legume-like lectin adhesin domains from the bacterial surface.

Staphylococcus aureus and Staphylococcus epidermidis are frequently associated with medical device infections that involve establishment of a bacterial biofilm on the device surface. Staphylococcal surface proteins Aap, SasG, and Pls are members of the Periscope Protein class and have been implicated in biofilm formation and host colonization; they comprise a repetitive region ("B region") and an N-terminal host colonization domain within the "A region," predicted to be a lectin domain. Repetitive E-G5 domains (as found in Aap, SasG, and Pls) form elongated "stalks" that would vary in length with repeat number, resulting in projection of the N-terminal A domain variable distances from the bacterial cell surface. Here, we present the structures of the lectin domains within A regions of SasG, Aap, and Pls and a structure of the Aap lectin domain attached to contiguous E-G5 repeats, suggesting the lectin domains will sit at the tip of the variable length rod. We demonstrate that these isolated domains (Aap, SasG) are sufficient to bind to human host desquamated nasal epithelial cells. Previously, proteolytic cleavage or a deletion within the A domain had been reported to induce biofilm formation; the structures suggest a potential link between these observations. Intriguingly, while the Aap, SasG, and Pls lectin domains bind a metal ion, they lack the nonproline cis peptide bond thought to be key for carbohydrate binding by the lectin fold. This suggestion of noncanonical ligand binding should be a key consideration when investigating the host cell interactions of these bacterial surface proteins.

Biosorption and transformation of cadmium and lead by Staphylococcus epidermidis AS-1 isolated from industrial effluent.

BACKGROUND: Rapid utilization of natural resources and other anthropogenic activities intruded heavy metals into the food chain and raised alarming concern for all life forms. The available methods proved insufficient in handling waste and pollutants due to the high cost and generation of toxic residues. Bioremediation strategies have offered sustainable solutions for toxic pollutants. In the current study, cadmium and lead (Cd and Pb respectively) tolerant strains have been isolated from industrial effluent and characterized for tolerance towards target pollutants. The strain was identified by 16s rRNA gene and further used for metal removal from the industrial effluents. RESULTS: Bacterial isolates were obtained from industrial discharge and evaluated for their tolerance towards Cd and Pb. AS-1 bacterial isolate exhibited maximum tolerance towards both the metals and hence was selected for further study. The isolate was identified as Staphylococcus epidermidis. ICP-MS and energy dispersive X-ray (EDX) analysis of biomass revealed that a significant proportion of cadmium (90.89%) and lead (94.87%) available in effluent were sequestered within bacterial biomass. Characteristic peaks at 2Ɵ (31.8637 and 45.6247 for cadmium) and (21.0397, 27.0127, 46.0537, 54.2707 and 75.6547 for lead) confirmed the crystalline nature of the sequestered metals. The selected strain was characterized on biochemical and molecular basis and was found to be Staphylococcus epidermidis. Based on 16 S rDNA sequence analysis, a phylogenetic dendrogram was created for the maximum likelihood of the bacterial strain. The sequence was deposited in the NCBI repository (accession number PP587422). CONCLUSION: The work has shown the possible way out of heavy metal pollution sustainably. To the best of the author's knowledge, this is the first report on the sequestration and reduction of cadmium and lead by a nonpathogenic strain of Staphylococcus epidermidis AS-1 that may be useful for alleviating heavy metal contamination.

Different modes of spacer acquisition by the Staphylococcus epidermidis type III-A CRISPR-Cas system.

CRISPR-Cas systems provide prokaryotic organisms with an adaptive defense mechanism that acquires immunological memories of infections. This is accomplished by integration of short fragments from the genome of invaders such as phages and plasmids, called 'spacers', into the CRISPR locus of the host. Depending on their genetic composition, CRISPR-Cas systems can be classified into six types, I-VI, however spacer acquisition has been extensively studied only in type I and II systems. Here, we used an inducible spacer acquisition assay to study this process in the type III-A CRISPR-Cas system of Staphylococcus epidermidis, in the absence of phage selection. Similarly to type I and II spacer acquisition, this type III system uses Cas1 and Cas2 to preferentially integrate spacers from the chromosomal terminus and free dsDNA ends produced after DNA breaks, in a manner that is enhanced by the AddAB DNA repair complex. Surprisingly, a different mode of spacer acquisition from rRNA and tRNA loci, which spans only the transcribed sequences of these genes and is not enhanced by AddAB, was also detected. Therefore, our findings reveal both common mechanistic principles that may be conserved in all CRISPR-Cas systems, as well as unique and intriguing features of type III spacer acquisition.

Electrogenic Staphylococcus epidermidis colonizes nasal cavities and alleviates IL-6 progression induced by the SARS2-CoV nucleocapsid protein.

AIM: Certain probiotic bacteria have been shown to possess an immunomodulatory effect and a protective effect on influenza infections. Using the Staphylococcus epidermidis K1 colonized mice model, we assessed the effect of nasal administration of glycerol or flavin mononucleotide (FMN) on the production of interleukin (IL)-6 mediated by the severe acute respiratory syndrome coronavirus 2 (SARS2-CoV) nucleocapsid protein (NPP). METHODS AND RESULTS: FMN, one of the key electron donors for the generation of electricity facilitated by S. epidermidis ATCC 12228, was detected in the glycerol fermentation medium. Compared to the S. epidermidis ATCC 12228, the S. epidermidis K1 isolate showed significant expression of the electron transfer genes, including pyruvate dehydrogenase (pdh), riboflavin kinase (rk), 1,4-dihydroxy-2-naphthoate octaprenyltransferase (menA), and type II NADH quinone oxidoreductase (ndh2). Institute of cancer research (ICR) mice were intranasally administered with S. epidermidis K1 with or without pretreatment with riboflavin kinase inhibitors, then nasally treated with glycerol or FMN before inoculating the NPP. Furthermore, J774A.1 macrophages were exposed to NPP serum and then treated with NPP of SARS2-CoV. The IL-6 levels in the bronchoalveolar lavage fluid (BALF) of mice and macrophages were quantified using a mouse IL-6 enzyme-linked immunosorbent assay kit. CONCLUSIONS: Here, we report that nasal administration of NPP strongly elevates IL-6 levels in both BALF and J774A.1 macrophages. It is worth noting that NPP-neutralizing antibodies can decrease IL-6 levels in macrophages. The nasal administration of glycerol or FMN to S. epidermidis K1-colonized mice results in a reduction of NPP-induced IL-6 production.

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.

The expression of glycosyltransferases sdgA and sdgB in Staphylococcus epidermidis depends on the conditions of biofilm formation.

The Staphylococcus aureus SdrG protein is glycosylated by SdgA and SdgB for protection against its degradation by the neutrophil cathepsin G. So far, there is no information about the role of Staphylococcus epidermidis SdgA or SdgB in biofilm-forming; therefore, the focus of this work was to determine the distribution and expression of the sdrG, sdgA and sdgB genes in S. epidermidis under in vitro and in vivo biofilm conditions. The frequencies of the sdrG, sdgA and sdgB genes were evaluated by PCR in a collection of 75 isolates. Isolates were grown in dynamic (non-biofilm-forming) or static (biofilm-forming) conditions. The expression of sdrG, sdgA and sdgB was determined by RT-qPCR in cells grown under dynamic conditions (CGDC), as well as in planktonic and sessile cells from a biofilm and cells adhered to a catheter implanted in Balb/c mice. The sdrG and sdgB genes were detected in 100% of isolates, while the sdgA gene was detected in 71% of the sample (p < 0.001). CGDC did not express sdrG, sdgA and sdgB mRNAs. Planktonic and sessile cells expressed sdrG and sdgB, and the same was observed in cells adhered to the catheter. In particular, one isolate, capable of inducing a biofilm under treatment with cathepsin G, expressed sdrG and sdgB in planktonic and sessile cells and cells adhering to the catheter. This suggests that bacteria require biofilm conditions as an important factor for the transcription of the sdgA, sdgB and sdrG genes.

The Transcription Factor SpoVG Is of Major Importance for Biofilm Formation of Staphylococcus epidermidis under In Vitro Conditions, but Dispensable for In Vivo Biofilm Formation.

Staphylococcus epidermidis is a common cause of device related infections on which pathogens form biofilms (i.e., multilayered cell populations embedded in an extracellular matrix). Here, we report that the transcription factor SpoVG is essential for the capacity of S. epidermidis to form such biofilms on artificial surfaces under in vitro conditions. Inactivation of spoVG in the polysaccharide intercellular adhesin (PIA) producing S. epidermidis strain 1457 yielded a mutant that, unlike its parental strain, failed to produce a clear biofilm in a microtiter plate-based static biofilm assay. A decreased biofilm formation capacity was also observed when 1457 ΔspoVG cells were co-cultured with polyurethane-based peripheral venous catheter fragments under dynamic conditions, while the cis-complemented 1457 ΔspoVG::spoVG derivative formed biofilms comparable to the levels seen with the wild-type. Transcriptional studies demonstrated that the deletion of spoVG significantly altered the expression of the intercellular adhesion (ica) locus by upregulating the transcription of the ica operon repressor icaR and down-regulating the transcription of icaADBC. Electrophoretic mobility shift assays (EMSA) revealed an interaction between SpoVG and the icaA-icaR intergenic region, suggesting SpoVG to promote biofilm formation of S. epidermidis by modulating ica expression. However, when mice were challenged with the 1457 ΔspoVG mutant in a foreign body infection model, only marginal differences in biomasses produced on the infected catheter fragments between the mutant and the parental strain were observed. These findings suggest that SpoVG is critical for the PIA-dependent biofilm formation of S. epidermis under in vitro conditions, but is largely dispensable for biofilm formation of this skin commensal under in vivo conditions.

Evaluation of the Effects of Ag, Cu, ZnO and TiO2 Nanoparticles on the Expression Level of Oxidative Stress-Related Genes and the Activity of Antioxidant Enzymes in Escherichia coli, Bacillus cereus and Staphylococcus epidermidis.

Although the molecular response of bacteria exposed to metal nanoparticles (NPs) is intensively studied, many phenomena related to their survival, metal uptake, gene expression and protein production are not fully understood. Therefore, this work aimed to study Ag-NPs, Cu-NPs, ZnO-NPs and TiO2-NPs-induced alterations in the expression level of selected oxidative stress-related genes in connection with the activity of antioxidant enzymes: catalase (CAT), peroxidase (PER) and superoxide dismutase (SOD) in Escherichia coli, Bacillus cereus and Staphylococcus epidermidis. The methodology used included: the extraction of total RNA and cDNA synthesis, the preparation of primers for selected housekeeping and oxidative stress genes, RT-qPCR reaction and the measurements of CAT, PER and SOD activities. It was established that the treatment of E. coli and S. epidermidis with NPs resulted mainly in the down-regulation of targeted genes, whilst the up-regulation of genes was confirmed in B. cereus. The greatest differences in the relative expression levels of tested genes occurred in B. cereus and S. epidermidis treated with TiO2-NPs, while in E. coli, they were observed under ZnO-NPs exposure. The changes found were mostly related to the expression of genes encoding proteins with PER and CAT-like activity. Among NPs, ZnO-NPs and Cu-NPs increased the activity of antioxidants in E. coli and B. cereus. In turn, TiO2-NPs had a major effect on enzymes activity in S. epidermidis. Considering all of the collected results for tested bacteria, it can be emphasised that the impact of NPs on the antioxidant system functioning was dependent on their type and concentration.

Regulation of cyclic oligoadenylate synthesis by the Staphylococcus epidermidis Cas10-Csm complex.

CRISPR-Cas systems are a class of adaptive immune systems in prokaryotes that use small CRISPR RNAs (crRNAs) in conjunction with CRISPR-associated (Cas) nucleases to recognize and degrade foreign nucleic acids. Recent studies have revealed that Type III CRISPR-Cas systems synthesize second messenger molecules previously unknown to exist in prokaryotes, cyclic oligoadenylates (cOA). These molecules activate the Csm6 nuclease to promote RNA degradation and may also coordinate additional cellular responses to foreign nucleic acids. Although cOA production has been reconstituted and characterized for a few bacterial and archaeal Type III systems, cOA generation and its regulation have not been explored for the Staphylococcus epidermidis Type III-A CRISPR-Cas system, a longstanding model for CRISPR-Cas function. Here, we demonstrate that this system performs Mg2+-dependent synthesis of 3-6 nt cOA. We show that activation of cOA synthesis is perturbed by single nucleotide mismatches between the crRNA and target RNA at discrete positions, and that synthesis is antagonized by Csm3-mediated target RNA cleavage. Altogether, our results establish the requirements for cOA production in a model Type III CRISPR-Cas system and suggest a natural mechanism to dampen immunity once the foreign RNA is destroyed.

The small non-coding RNA RsaE influences extracellular matrix composition in Staphylococcus epidermidis biofilm communities.

RsaE is a conserved small regulatory RNA (sRNA) which was previously reported to represent a riboregulator of central carbon flow and other metabolic pathways in Staphylococcus aureus and Bacillus subtilis. Here we show that RsaE contributes to extracellular (e)DNA release and biofilm-matrix switching towards polysaccharide intercellular adhesin (PIA) production in a hypervariable Staphylococcus epidermidis isolate. Transcriptome analysis through differential RNA sequencing (dRNA-seq) in combination with confocal laser scanning microscopy (CLSM) and reporter gene fusions demonstrate that S. epidermidis protein- and PIA-biofilm matrix producers differ with respect to RsaE and metabolic gene expression. RsaE is spatiotemporally expressed within S. epidermidis PIA-mediated biofilms, and its overexpression triggers a PIA biofilm phenotype as well as eDNA release in an S. epidermidis protein biofilm matrix-producing strain background. dRNA-seq and Northern blot analyses revealed RsaE to exist as a major full-length 100-nt transcript and a minor processed species lacking approximately 20 nucleotides at the 5'-end. RsaE processing results in expansion of the mRNA target spectrum. Thus, full-length RsaE interacts with S. epidermidis antiholin-encoding lrgA mRNA, facilitating bacterial lysis and eDNA release. Processed RsaE, however, interacts with the 5'-UTR of icaR and sucCD mRNAs, encoding the icaADBC biofilm operon repressor IcaR and succinyl-CoA synthetase of the tricarboxylic acid (TCA) cycle, respectively. RsaE augments PIA-mediated biofilm matrix production, most likely through activation of icaADBC operon expression via repression of icaR as well as by TCA cycle inhibition and re-programming of staphylococcal central carbon metabolism towards PIA precursor synthesis. Additionally, RsaE supports biofilm formation by mediating the release of eDNA as stabilizing biofilm matrix component. As RsaE itself is heterogeneously expressed within biofilms, we consider this sRNA to function as a factor favoring phenotypic heterogeneity and supporting division of labor in S. epidermidis biofilm communities.

Biofilm formation of Staphylococcus epidermidis imaged using atmospheric scanning electron microscopy.

Staphylococcus epidermidis are gram-positive bacteria that form a biofilm around implanted devices and develop an infection into a chronic state. Recently, it has been revealed that microvesicles have important roles in biofilm formation and intercellular communication among bacteria. However, biofilm formation of Staphylococcus epidermidis, and its relation to microvesicle secretion, is poorly understood because of the difficulty required to preserve the delicate water-rich morphology of biofilm for high-resolution observations. Here, we successfully imaged the microvesicles secreted from Staphylococcus epidermidis and the subsequent process of their integration into biofilm using liquid-phase imaging using atmospheric scanning electron microscopy (ASEM). In the biofilm, cells were connected by nanotube-like structures attached by microvesicles, and surrounded by extracellular polymeric substances. Cells cultured in the ASEM specimen holder were aldehyde-fixed and stained using positively charged nanogold labelling and/or using National Center for Microscopy and Imaging Research method. The samples immersed in aqueous radical scavenger glucose buffer were imaged by the inverted SEM of ASEM. Information regarding the morphologies of microvesicles, nanotube-like fibrils, and biofilm formed by Staphylococcus epidermidis is expected to be useful to elucidate the biological mechanism of biofilm formation and to develop a medicine against biofilms and their associated infections.

The Extracellular Vesicles from the Commensal Staphylococcus Epidermidis ATCC12228 Strain Regulate Skin Inflammation in the Imiquimod-Induced Psoriasis Murine Model.

Extracellular vesicles (EVs) are evaginations of the cytoplasmic membrane, containing nucleic acids, proteins, lipids, enzymes, and toxins. EVs participate in various bacterial physiological processes. Staphylococcus epidermidis interacts and communicates with the host skin. S. epidermidis' EVs may have an essential role in this communication mechanism, modulating the immunological environment. This work aimed to evaluate if S. epidermidis' EVs can modulate cytokine production by keratinocytes in vitro and in vivo using the imiquimod-induced psoriasis murine model. S. epidermidis' EVs were obtained from a commensal strain (ATC12228EVs) and a clinical isolated strain (983EVs). EVs from both origins induced IL-6 expression in HaCaT keratinocyte cultures; nevertheless, 983EVs promoted a higher expression of the pro-inflammatory cytokines VEGF-A, LL37, IL-8, and IL-17F than ATCC12228EVs. Moreover, in vivo imiquimod-induced psoriatic skin treated with ATCC12228EVs reduced the characteristic psoriatic skin features, such as acanthosis and cellular infiltrate, as well as VEGF-A, IL-6, KC, IL-23, IL-17F, IL-36γ, and IL-36R expression in a more efficient manner than 983EVs; however, in contrast, Foxp3 expression did not significantly change, and IL-36 receptor antagonist (IL-36Ra) was found to be increased. Our findings showed a distinctive immunological profile induction that is dependent on the clinical or commensal EV origin in a mice model of skin-like psoriasis. Characteristically, proteomics analysis showed differences in the EVs protein content, dependent on origin of the isolated EVs. Specifically, in ATCC12228EVs, we found the proteins glutamate dehydrogenase, ornithine carbamoyltransferase, arginine deiminase, carbamate kinase, catalase, superoxide dismutase, phenol-soluble ß1/ß2 modulin, and polyglycerol phosphate α-glucosyltransferase, which could be involved in the reduction of lesions in the murine imiquimod-induced psoriasis skin. Our results show that the commensal ATCC12228EVs have a greater protective/attenuating effect on the murine imiquimod-induced psoriasis by inducing IL-36Ra expression in comparison with EVs from a clinical isolate of S. epidermidis.

Insight into the Antibacterial Activity of Selected Metal Nanoparticles and Alterations within the Antioxidant Defence System in Escherichia coli, Bacillus cereus and Staphylococcus epidermidis.

The antimicrobial activity of nanoparticles (NPs) is a desirable feature of various products but can become problematic when NPs are released into different ecosystems, potentially endangering living microorganisms. Although there is an abundance of advanced studies on the toxicity and biological activity of NPs on microorganisms, the information regarding their detailed interactions with microbial cells and the induction of oxidative stress remains incomplete. Therefore, this work aimed to develop accurate oxidation stress profiles of Escherichia coli, Bacillus cereus and Staphylococcus epidermidis strains treated with commercial Ag-NPs, Cu-NPs, ZnO-NPs and TiO2-NPs. The methodology used included the following determinations: toxicological parameters, reactive oxygen species (ROS), antioxidant enzymes and dehydrogenases, reduced glutathione, oxidatively modified proteins and lipid peroxidation. The toxicological studies revealed that E. coli was most sensitive to NPs than B. cereus and S. epidermidis. Moreover, NPs induced the generation of specific ROS in bacterial cells, causing an increase in their concentration, which further resulted in alterations in the activity of the antioxidant defence system and protein oxidation. Significant changes in dehydrogenases activity and elevated lipid peroxidation indicated a negative effect of NPs on bacterial outer layers and respiratory activity. In general, NPs were characterised by very specific nano-bio effects, depending on their physicochemical properties and the species of microorganism.

Quebrachitol from Rhizophora mucronata inhibits biofilm formation and virulence production in Staphylococcus epidermidis by impairment of initial attachment and intercellular adhesion.

Staphylococcus epidermidis is well recognized nosocomial pathogen in clinical settings for their implants associated infections. Biofilm and virulence production executes a S. epidermidis pathogenesis against host. Hence, interfering of biofilm formation has become an auspicious to control the pathogenesis of S. epidermidis. The present study evaluates antibiofilm potential of Rhizophora mucronata against S. epidermidis biofilms. Rhizophora mucronata leaves extract significantly inhibited the biofilm formation and quebrachitol was identified as an active compound responsible for the biofilm inhibition. Quebrachitol significantly inhibited biofilm formation at concentration dependent manner without exhibit non-bactericidal property. And, quebrachitol reduced the biofilm building components such as exopolysaccharides, lipase and proteins production. Confocal laser scanning microscopic studies obtained quebrachitol surface independent biofilm efficacy against S. epidermidis. Notably, quebrachitol significantly reduced S. epidermidis adherence on biotic (coated with type I collagen and fibrinogen) and abiotic (hydrophobic and hydrophilic) surfaces. Addition of quebrachitol inhibits autolysis mediated initial attachment and accumulation associated aggregation process. Moreover, quebrachitol significantly reduced the hydrolases virulence production which supports S. epidermidis invasion into the host. Furthermore, gene expression analysis revealed the ability of quebrachitol to downregulate the virulence genes expression which are mainly involved in biofilm formation and virulence production. The results obtained from the present study suggest that quebrachitol as an ideal candidate for the therapeutic action against S. epidermidis pathogenesis.

Streptomyces-derived actinomycin D inhibits biofilm formation via downregulating ica locus and decreasing production of PIA in Staphylococcus epidermidis.

AIM: The objective of this study was to investigate the biofilm inhibitory activity of Streptomyces-derived actinomycin D against biofilm formation by Staphylococcus epidermidis. METHODS AND RESULTS: The microtitre plate method and microscopy were used to detect the biofilm formation of S. epidermidis. And an attempt was made to detect the effect of actinomycin D on important biofilm components, exopolysaccharides (EPS) in S. epidermidis using precolumn derivation HPLC. Also cell surface hydrophobicities of S. epidermidis were assessed to explore action mechanisms. The qPCR was performed to demonstrate the genetic mechanisms of biofilm formation by S. epidermidis. Unlike other antibiotics, actinomycin D (1·5 µg ml-1 ) from Streptomyces luteus significantly inhibited biofilm formation by S. epidermidis. Additionally, it effectively inhibited S. epidermidis cells from adhering to glass slides. Actinomycin D downregulated ica locus and then the reduced polysaccharide intercellular adhesin production caused S. epidermidis cells to become less hydrophobic, thus supporting its anti-biofilm effect. CONCLUSION: Streptomyces-derived actinomycin D is active in inhibiting the biofilm formation of S. epidermidis. SIGNIFICANCE AND IMPACT OF THE STUDY: Actinomycin D can be used as a promising antibiofilm agent in inhibiting S. epidermidis biofilm formation. The study is also the first insight into how actinomycin D inhibited the biofilm formation of S. epidermidis. Actinomycin D could potentially be used to reduce the risk of biofilm-associated infections. Our study also suggests that the metabolites from Actinomycete strains keep further attention as potential antibiofilm agents against biofilm formation of S. epidermidis, even biofilm infections of the other bacteria.

PEG-8 Laurate Fermentation of Staphylococcus epidermidis Reduces the Required Dose of Clindamycin Against Cutibacterium acnes.

The probiotic activity of skin Staphylococcus epidermidis (S. epidermidis) bacteria can elicit diverse biological functions via the fermentation of various carbon sources. Here, we found that polyethylene glycol (PEG)-8 Laurate, a carbon-rich molecule, can selectively induce the fermentation of S. epidermidis, not Cutibacterium acnes (C. acnes), a bacterium associated with acne vulgaris. The PEG-8 Laurate fermentation of S. epidermidis remarkably diminished the growth of C. acnes and the C. acnes-induced production of pro-inflammatory macrophage-inflammatory protein 2 (MIP-2) cytokines in mice. Fermentation media enhanced the anti-C. acnes activity of a low dose (0.1%) clindamycin, a prescription antibiotic commonly used to treat acne vulgaris, in terms of the suppression of C. acnes colonization and MIP-2 production. Furthermore, PEG-8 Laurate fermentation of S. epidermidis boosted the activity of 0.1% clindamycin to reduce the sizes of C. acnes colonies. Our results demonstrated, for the first time, that the PEG-8 Laurate fermentation of S. epidermidis displayed the adjuvant effect on promoting the efficacy of low-dose clindamycin against C. acnes. Targeting C. acnes by lowering the required doses of antibiotics may avoid the risk of creating drug-resistant C. acnes and maintain the bacterial homeostasis in the skin microbiome, leading to a novel modality for the antibiotic treatment of acne vulgaris.

Psoriasis and Antimicrobial Peptides.

Psoriasis is a systemic inflammatory disease caused by crosstalk between various cells such as T cells, neutrophils, dendritic cells, and keratinocytes. Antimicrobial peptides (AMPs) such as ß-defensin, S100, and cathelicidin are secreted from these cells and activate the innate immune system through various mechanisms to induce inflammation, thus participating in the pathogenesis of psoriasis. In particular, these antimicrobial peptides enhance the binding of damage-associated molecular patterns such as self-DNA and self-RNA to their receptors and promote the secretion of interferon from activated plasmacytoid dendritic cells and keratinocytes to promote inflammation in psoriasis. Neutrophil extracellular traps (NETs), complexes of self-DNA and proteins including LL-37 released from neutrophils in psoriatic skin, induce Th17. Activated myeloid dendritic cells secrete a mass of inflammatory cytokines such as IL-12 and IL-23 in psoriasis, which is indispensable for the proliferation and survival of T cells that produce IL-17. AMPs enhance the production of some of Th17 and Th1 cytokines and modulate receptors and cellular signaling in psoriasis. Inflammation induced by DAMPs, including self-DNA and RNA released due to microinjuries or scratches, and the enhanced recognition of DAMPs by AMPs, may be involved in the mechanism underlying the Köbner phenomenon in psoriasis.

IgY Targeting Bacterial Quorum-Sensing Molecules in Implant-Associated Infections.

Background: Implant-associated infections are still a major complication in the field of orthopedics. Bacteria can form biofilms on implant surfaces, making them more difficult to detect and treat. Since standard antibiotic therapy is often impaired in biofilm infections, particular interest is directed towards finding treatment alternatives. Biofilm-formation is a well-organized process during which bacteria communicate via quorum-sensing molecules (QSM). The aim of this study was to inhibit bacterial communication by directing avian IgY against specific QSM. Methods: Chicken were immunized against the following QSM: (1) AtlE, a member of the autolysin family which mediates attachment to a surface in Staphylococcus epidermidis; (2) GroEL, the bacterial heat shock protein; (3) PIA (polysaccharide intercellular adhesion), which is essential for cell-cell adhesion in biofilms. Staphylococcus epidermidis biofilms were grown and inhibition of biofilm-formation by IgYs was evaluated. Additionally, human osteoblasts were cultivated and biocompatibility of IgYs was tested. Results: We were able to demonstrate that all IgYs reduced biofilm-formation, also without prior immunization. Therefore, the response was probably not specific with regard to the QSM. Osteoblasts were activated by all IgYs which was demonstrated by microscopy and an increased release of IL-8. Conclusions: In conclusion, avian IgY inhibits biofilm-formation, though the underlying mechanism is not yet clear. However, adverse effects on local tissue cells (osteoblasts) were also observed.