Antarctic Marine Bacteria as a Source of Anti-Biofilm Molecules to Combat ESKAPE Pathogens.

The ESKAPE pathogens, including bacteria such as Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species, pose a global health threat due to their ability to resist antimicrobial drugs and evade the immune system. These pathogens are responsible for hospital-acquired infections, especially in intensive care units, and contribute to the growing problem of multi-drug resistance. In this study, researchers focused on exploring the potential of Antarctic marine bacteria as a source of anti-biofilm molecules to combat ESKAPE pathogens. Four Antarctic bacterial strains were selected, and their cell-free supernatants were tested against 60 clinical ESKAPE isolates. The results showed that the supernatants did not exhibit antimicrobial activity but effectively prevented biofilm formation and dispersed mature biofilms. This research highlights the promising potential of Antarctic bacteria in producing compounds that can counteract biofilms formed by clinically significant bacterial species. These findings contribute to the development of new strategies for preventing and controlling infections caused by ESKAPE pathogens.

Identification of the blaOXA-23 Gene in the First Mucoid XDR Acinetobacter baumannii Isolated from a Patient with Cystic Fibrosis.

Acinetobacter baumannii is one of the pathogens most involved in health care-associated infections in recent decades. Known for its ability to accumulate several antimicrobial resistance mechanisms, it possesses the oxacillinase blaoxa-23, a carbapenemase now endemic in Italy. Acinetobacter species are not frequently observed in patients with cystic fibrosis, and multidrug-resistant A. baumannii is a rare event in these patients. Non-mucoid A. baumannii carrying the blaoxa-23 gene has been sporadically detected. Here, we describe the methods used to detect blaoxa-23 in the first established case of pulmonary infection via a mucoid strain of A. baumannii producing carbapenemase in a 24-year-old cystic fibrosis patient admitted to Bambino Gesù Children's Hospital in Rome, Italy. This strain, which exhibited an extensively drug-resistant antibiotype, also showed a great ability to further increase its resistance in a short time.

First Case of VIM-1-like-Producing Pseudomonas putida Bacteremia in an Oncohematological Pediatric Patient in Italy.

Bacterial infections caused by multidrug-resistant (MDR) Gram-negatives are of great concern worldwide, as they are frequently associated with high mortality and morbidity rates. To date, two cases of VIM-2 metallo-ß-lactamase (MBL)-producing Pseudomonas putida bacteremia have been ever reported in France and Spain between 2004 and 2010. Here, we present the first case of VIM-1-like-producing P. putida isolated in blood culture collected from an oncohematological pediatric patient admitted to Bambino Gesù Children's Hospital (IRCCS) in Rome, Italy.

Virulence of Pseudomonas aeruginosa in Cystic Fibrosis: Relationships between Normoxia and Anoxia Lifestyle.

The airways of cystic fibrosis (CF) patients are colonized by many pathogens and the most common is Pseudomonas aeruginosa, an environmental pathogen that is able to infect immunocompromised patients thanks to its ability to develop resistance to conventional antibiotics. Over 12% of all patients colonized by P. aeruginosa harbour multi-drug resistant species. During airway infection in CF, P. aeruginosa adopts various mechanisms to survive in a hostile ecological niche characterized by low oxygen concentration, nutrient limitation and high osmotic pressure. To this end, P. aeruginosa uses a variety of virulence factors including pigment production, biofilm formation, motility and the secretion of toxins and proteases. This study represents the first report that systematically analyzes the differences in virulence features, in normoxia and anoxia, of clinical P. aeruginosa isolated from CF patients, characterized by multi- or pan-drug antibiotic resistance compared to antibiotic sensitive strains. The virulence features, such as biofilm formation, protease secretion and motility, are highly diversified in anaerobiosis, which reflects the condition of chronic CF infection. These findings may contribute to the understanding of the real-world lifestyle of pathogens isolated during disease progression in each particular patient and to assist in the design of therapeutic protocols for personalized medicine.

Serratiopeptidase Affects the Physiology of Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients.

Pseudomonas aeruginosa is frequently involved in cystic fibrosis (CF) airway infections. Biofilm, motility, production of toxins and the invasion of host cells are different factors that increase P. aeruginosa's virulence. The sessile phenotype offers protection to bacterial cells and resistance to antimicrobials and host immune attacks. Motility also contributes to bacterial colonization of surfaces and, consequently, to biofilm formation. Furthermore, the ability to adhere is the prelude for the internalization into lung cells, a common immune evasion mechanism used by most intracellular bacteria, such as P. aeruginosa. In previous studies we evaluated the activity of metalloprotease serratiopeptidase (SPEP) in impairing virulence-related properties in Gram-positive bacteria. This work aimed to investigate SPEP's effects on different physiological aspects related to the virulence of P. aeruginosa isolated from CF patients, such as biofilm production, pyoverdine and pyocyanin production and invasion in alveolar epithelial cells. Obtained results showed that SPEP was able to impair the attachment to inert surfaces as well as adhesion/invasion of eukaryotic cells. Conversely, SPEP's effect on pyocyanin and pyoverdine production was strongly strain-dependent, with an increase and/or a decrease of their production. Moreover, SPEP seemed to increase swarming motility and staphylolytic protease production. Our results suggest that a large number of clinical strains should be studied in-depth before drawing definitive conclusions. Why different strains sometimes react in opposing ways to a specific treatment is of great interest and will be the object of future studies. Therefore, SPEP affects P. aeruginosa's physiology by differently acting on several bacterial factors related to its virulence.

Essential Oils Biofilm Modulation Activity and Machine Learning Analysis on Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients.

The opportunistic pathogen Pseudomonas aeruginosa is often involved in airway infections of cystic fibrosis (CF) patients. It persists in the hostile CF lung environment, inducing chronic infections due to the production of several virulence factors. In this regard, the ability to form a biofilm plays a pivotal role in CF airway colonization by P. aeruginosa. Bacterial virulence mitigation and bacterial cell adhesion hampering and/or biofilm reduced formation could represent a major target for the development of new therapeutic treatments for infection control. Essential oils (EOs) are being considered as a potential alternative in clinical settings for the prevention, treatment, and control of infections sustained by microbial biofilms. EOs are complex mixtures of different classes of organic compounds, usually used for the treatment of upper respiratory tract infections in traditional medicine. Recently, a wide series of EOs were investigated for their ability to modulate biofilm production by different pathogens comprising S. aureus, S. epidermidis, and P. aeruginosa strains. Machine learning (ML) algorithms were applied to develop classification models in order to suggest a possible antibiofilm action for each chemical component of the studied EOs. In the present study, we assessed the biofilm growth modulation exerted by 61 commercial EOs on a selected number of P. aeruginosa strains isolated from CF patients. Furthermore, ML has been used to shed light on the EO chemical components likely responsible for the positive or negative modulation of bacterial biofilm formation.

Acinetobacter baumannii Isolates from COVID-19 Patients in a Hospital Intensive Care Unit: Molecular Typing and Risk Factors.

Infections caused by Acinetobacter baumannii represent a major concern for intensive care unit (ICU) patients. However, the epidemiology of these infections among COVID-19 patients has not been fully explored. The aims of this study were (i) to characterize the clonal spread of A. baumannii among COVID-19 patients admitted to the ICU of the Umberto I hospital of Rome during the first year of the pandemic and (ii) to identify risk factors for its acquisition. Isolates were analysed by pulsed-field gel electrophoresis, and a multivariable regression model was constructed. Adjusted odds ratios (aORs) and 95% confidence intervals (CIs) were calculated. Overall, 193 patients were included, and 102 strains were analysed. All isolates had highly antibiotic-resistant profiles and derived from two genotypes. The cumulative incidence of A. baumannii acquisition (colonization or infection) was 36.8%. Patients with A. baumannii had higher mortality and length of stay. Multivariable analysis showed that previous carbapenem use was the only risk factor associated with A. baumannii acquisition (aOR: 4.15, 95% CI: 1.78-9.64). We documented substantial A. baumannii infections and colonization and high levels of clonal transmission. Given the limited treatment options, effective prevention and containment strategies to limit the spread of A. baumannii should be implemented.

A Shaving Proteomic Approach to Unveil Surface Proteins Modulation of Multi-Drug Resistant Pseudomonas aeruginosa Strains Isolated From Cystic Fibrosis Patients.

Cystic fibrosis (CF) is the most common rare disease caused by a mutation of the CF transmembrane conductance regulator gene encoding a channel protein of the apical membrane of epithelial cells leading to alteration of Na+ and K+ transport, hence inducing accumulation of dense and sticky mucus and promoting recurrent airway infections. The most detected bacterium in CF patients is Pseudomonas aeruginosa (PA) which causes chronic colonization, requiring stringent antibiotic therapies that, in turn induces multi-drug resistance. Despite eradication attempts at the first infection, the bacterium is able to utilize several adaptation mechanisms to survive in hostile environments such as the CF lung. Its adaptive machinery includes modulation of surface molecules such as efflux pumps, flagellum, pili and other virulence factors. In the present study we compared surface protein expression of PA multi- and pan-drug resistant strains to wild-type antibiotic-sensitive strains, isolated from the airways of CF patients with chronic colonization and recent infection, respectively. After shaving with trypsin, microbial peptides were analyzed by tandem-mass spectrometry on a high-resolution platform that allowed the identification of 174 differentially modulated proteins localized in the region from extracellular space to cytoplasmic membrane. Biofilm assay was performed to characterize all 26 PA strains in term of biofilm production. Among the differentially expressed proteins, 17 were associated to the virulome (e.g., Tse2, Tse5, Tsi1, PilF, FliY, B-type flagellin, FliM, PyoS5), six to the resistome (e.g., OprJ, LptD) and five to the biofilm reservoir (e.g., AlgF, PlsD). The biofilm assay characterized chronic antibiotic-resistant isolates as weaker biofilm producers than wild-type strains. Our results suggest the loss of PA early virulence factors (e.g., pili and flagella) and later expression of virulence traits (e.g., secretion systems proteins) as an indicator of PA adaptation and persistence in the CF lung environment. To our knowledge, this is the first study that, applying a shaving proteomic approach, describes adaptation processes of a large collection of PA clinical strains isolated from CF patients in early and chronic infection phases.

Anti-Virulence Properties of Coridothymus capitatus Essential Oil against Pseudomonas aeruginosa Clinical Isolates from Cystic Fibrosis Patients.

Pseudomonas aeruginosa is an opportunistic pathogen responsible for nosocomial infections, and is often involved in airway infections of cystic fibrosis (CF) patients. P. aeruginosa virulence is related to its ability to form biofilm, trigger different types of motilities, and produce toxins (for example, bacterial pigments). In this scenario, essential oils (EOs) have gained notoriety for their role in phenotype modulation, including virulence modulation. Among different EOs previously analyzed, herein we investigated the activity of Coridothymus capitatus EO (CCEO) against specific virulence factors produced by P. aeruginosa isolated from CF patients. CCEO showed inhibition of new biofilm formation and reduction in mature biofilm in about half of the tested strains. On selected strains, SEM analysis provided interesting information regarding CCEO action in a pre-adhesion assay. CCEO treatment showed a dramatic modification of the extracellular matrix (ECM) structure. Our results clearly showed a drastic reduction in pyocyanin production (between 84% and 100%) for all tested strains in the presence of CCEO. Finally, CCEO was also able to strongly affect P. aeruginosa swarming and swimming motility for almost all tested strains. In consideration of the novel results obtained on clinical strains isolated from CF patients, CCEO may be a potential candidate to limit P. aeruginosa virulence.

Anti-Virulence Activity of the Cell-Free Supernatant of the Antarctic Bacterium Psychrobacter sp. TAE2020 against Pseudomonas aeruginosa Clinical Isolates from Cystic Fibrosis Patients.

Pseudomonas aeruginosa is an opportunistic pathogen often involved in airway infections of cystic fibrosis (CF) patients. Its pathogenicity is related to several virulence factors, such as biofilm formation, motility and production of toxins and proteases. The expression of these virulence factors is controlled by quorum sensing (QS). Thus, QS inhibition is considered a novel strategy for the development of antipathogenic compounds acting on specific bacterial virulence programs without affecting bacterial vitality. In this context, cold-adapted marine bacteria living in polar regions represent an untapped reservoir of biodiversity endowed with an interesting chemical repertoire. In this paper, we investigated the biological activity of a supernatant derived from a novel Antarctic bacterium (SN_TAE2020) against specific virulence factors produced by P. aeruginosa strains isolated from FC patients. Our results clearly show a reduction in pyocyanin and protease production in the presence of SN_TAE2020. Finally, SN_TAE2020 was also able to strongly affect swarming and swimming motility for almost all tested strains. Furthermore, the effect of SN_TAE2020 was investigated on biofilm growth and texture, captured by SEM analysis. In consideration of the novel results obtained on clinical strains, polar bacteria might represent potential candidates for the discovery of new compounds limiting P. aeruginosa virulence in CF patients.

Essential Oils Biofilm Modulation Activity, Chemical and Machine Learning Analysis. Application on Staphylococcus aureus Isolates from Cystic Fibrosis Patients.

Bacterial biofilm plays a pivotal role in chronic Staphylococcus aureus (S. aureus) infection and its inhibition may represent an important strategy to develop novel therapeutic agents. The scientific community is continuously searching for natural and "green alternatives" to chemotherapeutic drugs, including essential oils (EOs), assuming the latter not able to select resistant strains, likely due to their multicomponent nature and, hence, multitarget action. Here it is reported the biofilm production modulation exerted by 61 EOs, also investigated for their antibacterial activity on S. aureus strains, including reference and cystic fibrosis patients' isolated strains. The EOs biofilm modulation was assessed by Christensen method on five S. aureus strains. Chemical composition, investigated by GC/MS analysis, of the tested EOs allowed a correlation between biofilm modulation potency and putative active components by means of machine learning algorithms application. Some EOs inhibited biofilm growth at 1.00% concentration, although lower concentrations revealed different biological profile. Experimental data led to select antibiofilm EOs based on their ability to inhibit S. aureus biofilm growth, which were characterized for their ability to alter the biofilm organization by means of SEM studies.

Characterization of Scardovia wiggsiae Biofilm by Original Scanning Electron Microscopy Protocol.

Early childhood caries (ECC) is a severe manifestation of carious pathology with rapid and disruptive progression. The ECC microbiota includes a wide variety of bacterial species, among which is an anaerobic newly named species, Scardovia wiggsiae, a previously unidentified Bifidobacterium. Our aim was to provide the first ultrastructural characterization of S. wiggsiae and its biofilm by scanning electron microscopy (SEM) using a protocol that faithfully preserved the biofilm architecture and allowed an investigation at very high magnifications (order of nanometers) and with the appropriate resolution. To accomplish this task, we analyzed Streptococcus mutans' biofilm by conventional SEM and VP-SEM protocols, in addition, we developed an original procedure, named OsO4-RR-TA-IL, which avoids dehydration, drying and sputter coating. This innovative protocol allowed high-resolution and high-magnification imaging (from 10000× to 35000×) in high-vacuum and high-voltage conditions. After comparing three methods, we chose OsO4-RR-TA-IL to investigate S. wiggsiae. It appeared as a fusiform elongated bacterium, without surface specialization, arranged in clusters and submerged in a rich biofilm matrix, which showed a well-developed micro-canalicular system. Our results provide the basis for the development of innovative strategies to quantify the effects of different treatments, in order to establish the best option to counteract ECC in pediatric patients.

Essential oils against bacterial isolates from cystic fibrosis patients by means of antimicrobial and unsupervised machine learning approaches.

Recurrent and chronic respiratory tract infections in cystic fibrosis (CF) patients result in progressive lung damage and represent the primary cause of morbidity and mortality. Staphylococcus aureus (S. aureus) is one of the earliest bacteria in CF infants and children. Starting from early adolescence, patients become chronically infected with Gram-negative non-fermenting bacteria, and Pseudomonas aeruginosa (P. aeruginosa) is the most relevant and recurring. Intensive use of antimicrobial drugs to fight lung infections inevitably leads to the onset of antibiotic resistant bacterial strains. New antimicrobial compounds should be identified to overcome antibiotic resistance in these patients. Recently interesting data were reported in literature on the use of natural derived compounds that inhibited in vitro S. aureus and P. aeruginosa bacterial growth. Essential oils, among these, seemed to be the most promising. In this work is reported an extensive study on 61 essential oils (EOs) against a panel of 40 clinical strains isolated from CF patients. To reduce the in vitro procedure and render the investigation as convergent as possible, machine learning clusterization algorithms were firstly applied to pick-up a fewer number of representative strains among the panel of 40. This approach allowed us to easily identify three EOs able to strongly inhibit bacterial growth of all bacterial strains. Interestingly, the EOs antibacterial activity is completely unrelated to the antibiotic resistance profile of each strain. Taking into account the results obtained, a clinical use of EOs could be suggested.

Cold-adapted bacterial extracts as a source of anti-infective and antimicrobial compounds against Staphylococcus aureus.

Aim: The dramatic emergence of antibiotic resistance has directed the interest of research toward the discovery of novel antimicrobial molecules. In this context, cold-adapted marine bacteria living in polar regions represent an untapped reservoir of biodiversity endowed with an interesting chemical repertoire. The aim of this work was to identify new antimicrobials and/or antibiofilm molecules produced by cold-adapted bacteria. Materials & methods: Organic extracts obtained from polar marine bacteria were tested against Staphylococcus aureus. Most promising samples were subjected to suitable purification strategies. Results: Results obtained led to the identification of a novel lipopeptide able to effectively inhibit the biofilm formation of S. aureus. Conclusion: New lipopeptide may be potentially useful in a wide variety of biotechnological and medical applications.

Machine Learning Analyses on Data including Essential Oil Chemical Composition and In Vitro Experimental Antibiofilm Activities against Staphylococcus Species.

Biofilm resistance to antimicrobials is a complex phenomenon, driven not only by genetic mutation induced resistance, but also by means of increased microbial cell density that supports horizontal gene transfer across cells. The prevention of biofilm formation and the treatment of existing biofilms is currently a difficult challenge; therefore, the discovery of new multi-targeted or combinatorial therapies is growing. The development of anti-biofilm agents is considered of major interest and represents a key strategy as non-biocidal molecules are highly valuable to avoid the rapid appearance of escape mutants. Among bacteria, staphylococci are predominant causes of biofilm-associated infections. Staphylococci, especially Staphylococcus aureus (S. aureus) is an extraordinarily versatile pathogen that can survive in hostile environmental conditions, colonize mucous membranes and skin, and can cause severe, non-purulent, toxin-mediated diseases or invasive pyogenic infections in humans. Staphylococcus epidermidis (S. epidermidis) has also emerged as an important opportunistic pathogen in infections associated with medical devices (such as urinary and intravascular catheters, orthopaedic implants, etc.), causing approximately from 30% to 43% of joint prosthesis infections. The scientific community is continuously looking for new agents endowed of anti-biofilm capabilities to fight S. aureus and S epidermidis infections. Interestingly, several reports indicated in vitro efficacy of non-biocidal essential oils (EOs) as promising treatment to reduce bacterial biofilm production and prevent the inducing of drug resistance. In this report were analyzed 89 EOs with the objective of investigating their ability to modulate bacterial biofilm production of different S. aureus and S. epidermidis strains. Results showed the assayed EOs to modulated the biofilm production with unpredictable results for each strain. In particular, many EOs acted mainly as biofilm inhibitors in the case of S. epidermidis strains, while for S. aureus strains, EOs induced either no effect or stimulate biofilm production. In order to elucidate the obtained experimental results, machine learning (ML) algorithms were applied to the EOs' chemical compositions and the determined associated anti-biofilm potencies. Statistically robust ML models were developed, and their analysis in term of feature importance and partial dependence plots led to indicating those chemical components mainly responsible for biofilm production, inhibition or stimulation for each studied strain, respectively.

Ultrasound affects minimal inhibitory concentration of ampicillin against methicillin resistant Staphylococcus aureus USA300.

Antimicrobial resistance is one of the most serious global public health problems. Therefore, novel strategies are needed to counteract bacterial resistance development. The aim of the present study was to enhance the activity of antibiotics to bacteria by using ultrasound. Ultrasound reduced the dosage of ampicillin required to impair bacterial viability.

Antimicrobial and Antibiofilm Activity and Machine Learning Classification Analysis of Essential Oils from Different Mediterranean Plants against Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a ubiquitous organism and opportunistic pathogen that can cause persistent infections due to its peculiar antibiotic resistance mechanisms and to its ability to adhere and form biofilm. The interest in the development of new approaches for the prevention and treatment of biofilm formation has recently increased. The aim of this study was to seek new non-biocidal agents able to inhibit biofilm formation, in order to counteract virulence rather than bacterial growth and avoid the selection of escape mutants. Herein, different essential oils extracted from Mediterranean plants were analyzed for their activity against P. aeruginosa. Results show that they were able to destabilize biofilm at very low concentration without impairing bacterial viability. Since the action is not related to a bacteriostatic/bactericidal activity on P. aeruginosa, the biofilm change of growth in presence of the essential oils was possibly due to a modulation of the phenotype. To this aim, application of machine learning algorithms led to the development of quantitative activity-composition relationships classification models that allowed to direct point out those essential oil chemical components more involved in the inhibition of biofilm production. The action of selected essential oils on sessile phenotype make them particularly interesting for possible applications such as prevention of bacterial contamination in the community and in healthcare environments in order to prevent human infections. We assayed 89 samples of different essential oils as P. aeruginosa anti-biofilm. Many samples inhibited P. aeruginosa biofilm at concentrations as low as 48.8 µg/mL. Classification of the models was developed through machine learning algorithms.

Pentadecanal inspired molecules as new anti-biofilm agents against Staphylococcus epidermidis.

Staphylococcus epidermidis, a harmless human skin colonizer, is a significant nosocomial pathogen in predisposed hosts because of its capability to form a biofilm on indwelling medical devices. In a recent paper, the purification and identification of the pentadecanal produced by the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125, able to impair S. epidermidis biofilm formation, were reported. Here the authors report on the chemical synthesis of pentadecanal derivatives, their anti-biofilm activity on S. epidermidis, and their action in combination with antibiotics. The results clearly indicate that the pentadecanal derivatives were able to prevent, to a different extent, biofilm formation and that pentadecanoic acid positively modulated the antimicrobial activity of the vancomycin. The cytotoxicity of these new anti-biofilm molecules was tested on two different immortalized eukaryotic cell lines in view of their potential applications.

Virulence of MRSA USA300 is enhanced by sub-inhibitory concentration of two different classes of antibiotics.

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) USA300 is responsible of many kinds of infections of skin and soft-tissue. Antibiotic resistance, biofilm formation and the ability to adhere and invade are virulence factors that contribute to MRSA pathogenesis. In some cases, decreased bioavailability of antibiotics in systemic circulation could result; in these conditions sub-therapeutic levels of the antibiotics may be established, exposing bacteria to sub-inhibitory concentrations. On the basis of several published scientific data it is fair to assume that all these events could induce an increase of bacterial virulence. In the present study, we investigated this process by measuring the effects of low doses of two different classes of antibiotics on some virulence features of MRSA USA300 isolate, like the ability to adhere and invade eukaryotic cells. Results obtained strongly support the importance of the respect of a correct dosage of antibiotic in therapy to escape the insurgence of more virulent phenotypes.

Serratiopeptidase reduces the invasion of osteoblasts by Staphylococcus aureus.

Finding new strategies to counteract periprosthetic infection and implant failure is a main target in orthopedics. Staphylococcus aureus, the leading etiologic agent of orthopedic implant infections, is able to enter and kill osteoblasts, to stimulate pro-inflammatory chemokine secretion, to recruit osteoclasts, and to cause inflammatory osteolysis. Moreover, by entering eukaryotic cells, staphylococci hide from the host immune defenses and shelter from the extracellular antibiotics. Thus, infection persists, inflammation thrives, and a highly destructive osteomyelitis occurs around the implant. The ability of serratiopeptidase (SPEP), a metalloprotease by Serratia marcescens, to control S. aureus invasion of osteoblastic MG-63 cells and pro-inflammatory chemokine MCP-1 secretion was evaluated. Human osteoblast cells were infected with staphylococcal strains in the presence and in the absence of SPEP. Cell proliferation and cell viability were also evaluated. The release of pro-inflammatory chemokine MCP-1 was evaluated after the exposure of the osteoblast cells to staphylococcal strains. The significance of the differences in the results of each test and the relative control values was determined with Student's t-test. SPEP impairs their invasiveness into osteoblasts, without affecting the viability and proliferation of bone cells, and tones down their production of MCP-1. We recognize SPEP as a potential tool against S. aureus bone infection and destruction.