Acinetobacter baumannii has emerged as one of the most problematic bacterial pathogens responsible for hospital-acquired and community infections worldwide. Besides its high capacity to acquire antibiotic resistance mechanisms, it also presents high adhesion abilities on inert and living surfaces leading to biofilm development. This lifestyle confers additional protection against various treatments and allows it to persist for long periods in various hospital niches. Due to their remarkable antimicrobial tolerance, A. baumannii biofilms are difficult to control and ultimately eradicate. Further insights into the mechanism of biofilm development will help to overcome this challenge and to develop novel antibiofilm strategies. To unravel critical determinants of this sessile lifestyle, the proteomic profiles of two A. baumannii strains (ATTC17978 and SDF) grown in planktonic stationary phase or in mature solid–liquid (S-L) biofilm were compared using a semiquantitative proteomic study. Of interest, among the 69 common proteins determinants accumulated in the two strains at the S-L interface, we sorted out the MacAB-TolC system. This tripartite efflux pump played a role in A. baumannii biofilm formation as demonstrated by using ΔmacAB-tolC deletion mutant. Complementary approaches allowed us to get an overview of the impact of macAB-tolC deletion in A. baumannii physiology. Indeed, this efflux pump appeared to be involved in the envelope stress response occurring in mature biofilm. It contributes to maintain wild type (WT) membrane rigidity and provides tolerance to high osmolarity conditions. In addition, this system is probably involved in the maintenance of iron and sulfur homeostasis. MacAB-TolC might help this pathogen face and adapt to deleterious conditions occurring in mature biofilms. Increasing our knowledge of A. baumannii biofilm formation will undoubtedly help us develop new therapeutic strategies to tackle this emerging threat to human health.

Staphylococcus aureus is a skin commensal microorganism commonly colonizing healthy humans. Nevertheless, S. aureus can also be responsible for cutaneous infections and contribute to flare-up of inflammatory skin diseases such as atopic dermatitis (AD), which is characterized by dysbiosis of the skin microbiota with S. aureus as the predominant species. However, the role of major virulence factors of this pathogen such as phenol-soluble modulin (PSM) toxins in epidermal inflammation remains poorly understood. Stimulation of primary human keratinocytes with sublytic concentrations of synthetic and purified PSM α3 resulted in upregulation of a large panel of pro-inflammatory chemokine and cytokine gene expression, including CXCL1, CXCL2, CXCL3, CXCL5, CXCL8, CCL20, IL-1α, IL-1β, IL-6, IL-36γ and TNF-α, while inducing the release of CXCL8, CCL20, TNF-α and IL-6. In addition, using S. aureus culture supernatant from mutants deleted from genes encoding either α-type PSMs or all PSM production, PSMs were shown to be the main factors of S. aureus secretome responsible for pro-inflammatory mediator induction in human keratinocytes. On the other hand, α-type PSM-containing supernatant triggered an intense induction of pro-inflammatory mediator expression and secretion during both topical and basal layer stimulation of an ex vivo model of human skin explants, a physiologically relevant model of pluristratified epidermis. Taken together, the results of this study show that PSMs and more specifically α-type PSMs are major virulence factors of S. aureus inducing a potent inflammatory response during infection of the human epidermis and could thereby contribute to AD flare-up through exacerbation of skin inflammation.

Acinetobacter baumannii is a problematic nosocomial pathogen owing to its increasing resistance to antibiotics and its great ability to survive in the hospital environment, which is linked to its capacity to form biofilms. Structural and functional investigations of post-translational modifications, such as phosphorylations, may lead to identification of candidates for therapeutic targets against this pathogen. Here, we present the first S/T/Y phosphosecretome of two A. baumannii strains, the reference strain ATCC 17978 and the virulent multi-drug resistant strain AB0057, cultured in two modes of growth (planktonic and biofilm) using TiO2 chromatography followed by high resolution mass spectrometry. In ATCC 17978, we detected a total of 137 (97 phosphoproteins) and 52 (33 phosphoproteins) phosphosites in biofilm and planktonic modes of growth, respectively. Similarly, in AB0057, 155 (119 phosphoproteins) and 102 (74 phosphoproteins) phosphosites in biofilm and planktonic modes of growth were identified, respectively. Both strains in the biofilm mode of growth showed a higher number of phosphosites and phosphoproteins compared to planktonic growth. Several phosphorylated sites are localized in key regions of proteins involved in either drug resistance (β-lactamases), adhesion to host tissues (pilins), or protein secretion (Hcp). Site-directed mutagenesis of the Hcp protein, essential for type VI secretion system-mediated interbacterial competition, showed that four of the modified residues are essential for type VI secretion system activity.

<p><em>Acinetobacter baumannii</em> is one of the most problematic opportunist pathogen responsible for many infections worldwide (1). Besides its high capacities to acquire antibiotic resistance mechanisms, it also presents high adhesion abilities on any types of abiotic or living surfaces leading to biofilm development, a mode of growth conferring an additional protection against various treatments and allowing the infection relapse (2). <em>A. baumannii</em> has been recently ranked on the global priority pathogens list established by the World Health Organization for which there is an urgent need for new treatments. One interesting way to identify new therapeutic targets to eradicate this pathogen is the characterization of its post-translational modifications (PTMs) (3). The functions and extents of PTMs remain largely unknown in prokaryotic cells compared to eukaryotic cells. Lysine acetylation is an attractive and prevalent PTM in bacteria. An increasing number of investigations have been dedicated to identify acetylated proteins by proteomics. Some studies have shown that acetylation can play a pivotal role in bacterial virulence, resistance, or biofilm (4). Enzymes involved in acetylation addition (lysine acetyltranferase KAT) or removal (lysine deacetylase KDAC) would provide a better mechanistic understanding of bacterial physiology and therefore could be considered as potential therapeutic targets. So far, little information is available on these enzymes in <em>A. baumannii</em> (5). Recently, in a global dynamic proteome study of<em> A. baumannii</em> ATCC 17978 strain grown in sessile mode, we highlighted the highest protein fold change for a protein belonging to the Sir2-like family which may possess a KDAC activity (6). The aim of the current study was to evaluate the involvement of this protein in <em>A. baumannii</em> physiology. For this purpose, a gene deletion approach was carried out to perform different phenotype tests (drugs and oxidative stress resistance, virulence assays, motility and biofilm formation) on wild-type and mutant strains. We compared, in biofilm mode of growth, acetylomes of the WT and the mutant. Our results demonstrated more than twice acetylated proteins in mutant in comparison to the WT. Of interest, biofilm formation in mutant was sensibly decreased. These different results suggest a potential involvement of this protein in <em>A. baumannii</em> biofilm formation.</p> <p> </p> <p>(1) Antunes et al. Acinetobacter baumannii: evolution of a global pathogen. Pathog. Dis. 71(2014), 292-301.</p> <p>(2) Espinal et al. Effect of biofilm formation on the survival of Acinetobacter baumannii on dry surfaces. J. Hosp. Infect. 80(2012), 56–60.</p> <p>(3) Richters. Targeting protein arginine methyltransferase 5 in disease. Future Med. Chem. 9(2017), 2081-2098.</p> <p>(4) VanDrisse and Escalante-Semerena. Protein acetylation in bacteria. Annu. Rev. Microbiol. 73(2019), 111-132.</p> <p>(5) Carabetta and Cristea. Regulation, function, and detection of protein acetylation in bacteria. J. Bacteriol. 199(2017), e00107-17.</p> <p>(6) Kentache et al. Global dynamic proteome study of a pellicle-forming Acinetobacter baumanii strain. Mol. Cell. Proteomics. 16(2017), 100-112.</p>