An NlpC/P60 protein catalyzes a key step in peptidoglycan recycling at the intersection of energy recovery, cell division and immune evasion in the intracellular pathogen Chlamydia trachomatis.

The obligate intracellular Chlamydiaceae do not need to resist osmotic challenges and thus lost their cell wall in the course of evolution. Nevertheless, these pathogens maintain a rudimentary peptidoglycan machinery for cell division. They build a transient peptidoglycan ring, which is remodeled during the process of cell division and degraded afterwards. Uncontrolled degradation of peptidoglycan poses risks to the chlamydial cell, as essential building blocks might get lost or trigger host immune response upon release into the host cell. Here, we provide evidence that a primordial enzyme class prevents energy intensive de novo synthesis and uncontrolled release of immunogenic peptidoglycan subunits in Chlamydia trachomatis. Our data indicate that the homolog of a Bacillus NlpC/P60 protein is widely conserved among Chlamydiales. We show that the enzyme is tailored to hydrolyze peptidoglycan-derived peptides, does not interfere with peptidoglycan precursor biosynthesis, and is targeted by cysteine protease inhibitors in vitro and in cell culture. The peptidase plays a key role in the underexplored process of chlamydial peptidoglycan recycling. Our study suggests that chlamydiae orchestrate a closed-loop system of peptidoglycan ring biosynthesis, remodeling, and recycling to support cell division and maintain long-term residence inside the host. Operating at the intersection of energy recovery, cell division and immune evasion, the peptidoglycan recycling NlpC/P60 peptidase could be a promising target for the development of drugs that combine features of classical antibiotics and anti-virulence drugs.

Recent advances in therapeutic targets identification and development of treatment strategies towards Pseudomonas aeruginosa infections.

The opportunistic human pathogen Pseudomonas aeruginosa is the causal agent of a wide variety of infections. This non-fermentative Gram-negative bacillus can colonize zones where the skin barrier is weakened, such as wounds or burns. It also causes infections of the urinary tract, respiratory system or bloodstream. P. aeruginosa infections are common in hospitalized patients for which multidrug-resistant, respectively extensively drug-resistant isolates can be a strong contributor to a high rate of in-hospital mortality. Moreover, chronic respiratory system infections of cystic fibrosis patients are especially concerning, since very tedious to treat. P. aeruginosa exploits diverse cell-associated and secreted virulence factors, which play essential roles in its pathogenesis. Those factors encompass carbohydrate-binding proteins, quorum sensing that monitor the production of extracellular products, genes conferring extensive drug resistance, and a secretion system to deliver effectors to kill competitors or subvert host essential functions. In this article, we highlight recent advances in the understanding of P. aeruginosa pathogenicity and virulence as well as efforts for the identification of new drug targets and the development of new therapeutic strategies against P. aeruginosa infections. These recent advances provide innovative and promising strategies to circumvent infection caused by this important human pathogen.

Cedratvirus lausannensis - digging into Pithoviridae diversity.

Amoeba-infecting viruses have raised scientists' interest due to their novel particle morphologies, their large genome size and their genomic content challenging previously established dogma. We report here the discovery and the characterization of Cedratvirus lausannensis, a novel member of the Megavirales, with a 0.75-1 µm long amphora-shaped particle closed by two striped plugs. Among numerous host cell types tested, the virus replicates only in Acanthamoeba castellanii leading to host cell lysis within 24 h. C. lausannensis was resistant to ethanol, hydrogen peroxide and heating treatments. Like 30 000-year-old Pithovirus sibericum, C. lausannensis enters by phagocytosis, releases its genetic content by fusion of the internal membrane with the inclusion membrane and replicates in intracytoplasmic viral factories. The genome encodes 643 proteins that confirmed the grouping of C. lausannensis with Cedratvirus A11 as phylogenetically distant members of the family Pithoviridae. The 575,161 bp AT-rich genome is essentially devoid of the numerous repeats harbored by Pithovirus, suggesting that these non-coding repetitions might be due to a selfish element rather than particular characteristics of the Pithoviridae family. The discovery of C. lausannensis confirms the contemporary worldwide distribution of Pithoviridae members and the characterization of its genome paves the way to better understand their evolution.

Expression of oleosin and perilipins in yeast promotes formation of lipid droplets from the endoplasmic reticulum.

Most cells store neutral lipids in a dedicated compartment, the lipid droplet (LD). These LDs are structurally and functionally conserved across species. In higher eukaryotes, LDs are covered by abundant scaffolding proteins, such as the oleosins in plants and perilipins (PLINs) in animal cells. Saccharomyces cerevisiae, however, has no homologues of these scaffolding proteins. To analyze a possible function of these proteins in the biogenesis of LDs, oleosin and perilipin family members (PLIN1, ADRP/PLIN2 and TIP47/PLIN3) were expressed in yeast cells and their targeting to LDs, membrane association and function in neutral lipid homeostasis and LD biogenesis were analyzed. When expressed in wild-type cells, these proteins were properly targeted to LDs. However, when expressed in cells lacking LDs, oleosin was localized to the ER bilayer and was rapidly degraded. PLINs, on the other hand, did not localize to the ER membrane in the absence of LDs and lost their membrane association. Photobleaching experiments revealed that PLIN2 and PLIN3 rapidly exchanged their LD association, but PLINs did not move as quickly as integral membrane proteins, such as oleosin, over the LD surface. Interestingly, expression of these scaffolding LD proteins in mutant cells containing elevated levels of neutral lipids within the ER bilayer resulted in the formation of LDs. These results suggest that these LD scaffolding proteins promote the sequestration of neutral lipids from the ER bilayer and thereby induce LD formation. Consistent with this proposition, addition of a cell-permeable diacylglycerol (DAG) was sufficient to promote LD formation in cells expressing the LD scaffolding proteins but lacking the capacity to synthesize storage lipids.

Lipid droplets are functionally connected to the endoplasmic reticulum in Saccharomyces cerevisiae.

Cells store metabolic energy in the form of neutral lipids that are deposited within lipid droplets (LDs). In this study, we examine the biogenesis of LDs and the transport of integral membrane proteins from the endoplasmic reticulum (ER) to newly formed LDs. In cells that lack LDs, otherwise LD-localized membrane proteins are homogenously distributed in the ER membrane. Under these conditions, transcriptional induction of a diacylglycerol acyltransferase that catalyzes the formation of the storage lipid triacylglycerol (TAG), Lro1, is sufficient to drive LD formation. Newly formed LDs originate from the ER membrane where they become decorated by marker proteins. Induction of LDs by expression of the second TAG-synthesizing integral membrane protein, Dga1, reveals that Dga1 itself moves from the ER membrane to concentrate on LDs. Photobleaching experiments (FRAP) indicate that relocation of membrane proteins from the ER to LDs is independent of temperature and energy, and thus not mediated by classical vesicular transport routes. LD-localized membrane proteins are homogenously distributed at the perimeter of LDs, they are free to move over the LD surface and can even relocate back into the ER, indicating that they are not restricted to specialized sites on LDs. These observations indicate that LDs are functionally connected to the ER membrane and that this connection allows the efficient partitioning of membrane proteins between the two compartments.

Ypk1, the yeast orthologue of the human serum- and glucocorticoid-induced kinase, is required for efficient uptake of fatty acids.

Fatty acids constitute an important energy source for various tissues. The mechanisms that mediate and control uptake of free fatty acids from the circulation, however, are poorly understood. Here we show that efficient fatty-acid uptake by yeast cells requires the protein kinase Ypk1, the orthologue of the human serum- and glucocorticoid-induced kinase Sgk1. ypk1Delta mutant cells fail to grow under conditions that render cells auxotrophic for fatty acids, show a reduced uptake of radiolabelled or fluorescently labelled fatty acids, lack the facilitated component of the uptake activity, and have elevated levels of fatty acids in a bovine serum albumin (BSA) back-extractable compartment. Efficient fatty-acid uptake and/or incorporation requires the protein-kinase activity of Ypk1, because a kinase-dead point-mutant allele of YPK1 is defective in this process. This function of Ypk1 in fatty-acid uptake and/or incorporation is functionally conserved, because expression of the human Sgk1 kinase rescues ypk1Delta mutant yeast. These observations suggest that Ypk1 and possibly the human Sgk1 kinase affect fatty-acid uptake and thus energy homeostasis through regulating endocytosis. Consistent with such a proposition, mutations that block early steps of endocytosis display reduced levels of fatty-acid uptake.

In vitro synergistic action of TAT-RasGAP317-326 peptide with antibiotics against Gram-negative pathogens.

OBJECTIVES: Multidrug-resistant (MDR) bacteria are a continuously increasing threat for medicine, causing infections recalcitrant to antibiotics. Antimicrobial peptides (AMPs) were identified as alternatives to antibiotics, being naturally occurring short peptides and part of the innate immune system of a vast majority of organisms. However, the clinical application of AMPs is limited by suboptimal pharmacokinetic properties and relatively high toxicity. Combinatorial treatments using AMPs and classical antibiotics may decrease the concentrations of AMPs required for bacterial eradication, thus lowering the side effects of these peptides. METHODS: Here, we investigate the in vitro efficiency of combinations of the recently described antimicrobial peptide TAT-RasGAP317-326 with a panel of commonly used antimicrobial agents against three Gram-negative bacteria, Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumannii, using checkerboard and time-kill assays. RESULTS: We identified synergistic combinations towards all three bacteria and demonstrated that these combinations had an increased bactericidal effect compared to individual drugs. Moreover, combinations were also effective against clinical isolates of A. baumannii. Finally, combination of TAT-RasGAP317-326 and meropenem had a promising antibiofilm effect towards A. baumannii. CONCLUSIONS: Taken together, our results indicate that combinations of TAT-RasGAP317-326 with commonly used antimicrobial agents may lead to the development of new treatment protocols against infections caused by MDR bacteria.

The EnvZ/OmpR Two-Component System Regulates the Antimicrobial Activity of TAT-RasGAP317-326 and the Collateral Sensitivity to Other Antibacterial Agents.

The rapid emergence of antibiotic-resistant bacteria poses a serious threat to public health worldwide. Antimicrobial peptides (AMPs) are promising antibiotic alternatives; however, little is known about bacterial mechanisms of AMP resistance and the interplay between AMP resistance and the bacterial response to other antimicrobials. In this study, we identified Escherichia coli mutants resistant to the TAT-RasGAP317-326 antimicrobial peptide and found that resistant bacteria show collateral sensitivity to other AMPs and antibacterial agents. We determined that resistance to TAT-RasGAP317-326 peptide arises through mutations in the histidine kinase EnvZ, a member of the EnvZ/OmpR two-component system responsible for osmoregulation in E. coli. In particular, we found that TAT-RasGAP317-326 binding and entry is compromised in E. coli peptide-resistant mutants. We showed that peptide resistance is associated with transcriptional regulation of a number of pathways and EnvZ-mediated resistance is dependent on the OmpR response regulator but is independent of the OmpC and OmpF outer membrane porins. Our findings provide insight into the bacterial mechanisms of TAT-RasGAP317-326 resistance and demonstrate that resistance to this AMP is associated with collateral sensitivity to other antibacterial agents. IMPORTANCE Antimicrobial peptides (AMP) are promising alternatives to classical antibiotics in the fight against antibiotic resistance. Resistance toward antimicrobial peptides can occur, but little is known about the mechanisms driving this phenomenon. Moreover, there is limited knowledge on how AMP resistance relates to the bacterial response to other antimicrobial agents. Here, we address these questions in the context of the antimicrobial peptide TAT-RasGAP317-326. We show that resistant Escherichia coli strains can be selected and do not show resistance to other antimicrobial agents. Resistance is caused by a mutation in a regulatory pathway, which lowers binding and entry of the peptide in E. coli. Our results highlight a mechanism of resistance that is specific to TAT-RasGAP317-326. Further research is required to characterize these mechanisms and to evaluate the potential of antimicrobial combinations to curb the development of antimicrobial resistance.

The antimicrobial peptide TAT-RasGAP317-326 inhibits the formation and expansion of bacterial biofilms in vitro.

OBJECTIVES: Biofilms are structured aggregates of bacteria embedded in a self-produced matrix that develop in diverse ecological niches. Pathogenic bacteria can form biofilms on surfaces and in tissues, causing nosocomial and chronic infections that are difficult to treat. While antibiotics are largely inefficient in limiting biofilm formation and expansion, antimicrobial peptides (AMPs) are emerging as alternative antibiofilm treatments. In this study, we explore the effect of the newly described AMP TAT-RasGAP317-326 on Acinetobacter baumannii, Pseudomonas aeruginosa and Staphylococcus aureus biofilms. METHODS: Efficiency of TAT-RasGAP317-326 on biofilms was tested in vitro. Both viability of bacteria contained in the biofilm as well as biomass of the biofilm were quantified using resazurin and crystal violet staining, respectively. The antibiofilm effect of TAT-RasGAP317-326 was compared with a selection of classical antibiotics and AMPs. RESULTS: We observe that TAT-RasGAP317-326 inhibits biofilm formation at concentrations equivalent or two times greater than the minimum inhibitory concentration (MIC) of planktonic bacteria. Moreover, TAT-RasGAP317-326 limits the expansion of A. baumannii and P. aeruginosa established biofilms at twice the concentration inhibiting biofilm formation. CONCLUSION: These results underscore the potential use of TAT-RasGAP317-326 against biofilms and encourage further studies in the development of AMPs to treat biofilm-related infections.

Bacterial surface properties influence the activity of the TAT-RasGAP317-326 antimicrobial peptide.

Antibiotic resistance is an increasing threat for public health, underscoring the need for new antibacterial agents. Antimicrobial peptides (AMPs) represent an alternative to classical antibiotics. TAT-RasGAP317-326 is a recently described AMP effective against a broad range of bacteria, but little is known about the conditions that may influence its activity. Using RNA-sequencing and screening of mutant libraries, we show that Escherichia coli and Pseudomonas aeruginosa respond to TAT-RasGAP317-326 by regulating metabolic and stress response pathways, possibly implicating two-component systems. Our results also indicate that bacterial surface properties, in particular integrity of the lipopolysaccharide layer, influence peptide binding and entry. Finally, we found differences between bacterial species with respect to their rate of resistance emergence against this peptide. Our findings provide the basis for future investigation on the mode of action of TAT-RasGAP317-326, which may help developing antimicrobial treatments based on this peptide.

Diverse Stress-Inducing Treatments cause Distinct Aberrant Body Morphologies in the Chlamydia-Related Bacterium, Waddlia chondrophila.

Chlamydiae, such as Chlamydia trachomatis and Chlamydia pneumoniae, can cause chronic infections. It is believed that persistent forms called aberrant bodies (ABs) might be involved in this process. AB formation seems to be a common trait of all members of the Chlamydiales order and is caused by distinct stress stimuli, such as ß-lactam antibiotics or nutrient starvation. While the diverse stimuli inducing ABs are well described, no comprehensive morphological characterization has been performed in Chlamydiales up to now. We thus infected mammalian cells with the Chlamydia-related bacterium Waddlia chondrophila and induced AB formation using different stimuli. Their morphology, differences in DNA content and in gene expression were assessed by immunofluorescence, quantitative PCR, and reverse transcription PCR, respectively. All stimuli induced AB formation. Interestingly, we show here for the first time that the DNA gyrase inhibitor novobiocin also caused appearance of ABs. Two distinct patterns of ABs could be defined, according to their morphology and number: (i) small and multiple ABs versus (ii) large and rare ABs. DNA replication of W. chondrophila was generally not affected by the different treatments. Finally, no correlation could be observed between specific types of ABs and expression patterns of mreB and rodZ genes.

Interactions Screenings Unearth Potential New Divisome Components in the Chlamydia-Related Bacterium, Waddlia chondrophila.

Chlamydiales order members are obligate intracellular bacteria, dividing by binary fission. However, Chlamydiales lack the otherwise conserved homologue of the bacterial division organizer FtsZ and certain division protein homologues. FtsZ might be functionally replaced in Chlamydiales by the actin homologue MreB. RodZ, the membrane anchor of MreB, localizes early at the division septum. In order to better characterize the organization of the chlamydial divisome, we performed co-immunoprecipitations and yeast-two hybrid assays to study the interactome of RodZ, using Waddlia chondrophila, a potentially pathogenic Chlamydia-related bacterium, as a model organism. Three potential interactors were further investigated: SecA, FtsH, and SufD. The gene and protein expression profiles of these three genes were measured and are comparable with recently described division proteins. Moreover, SecA, FtsH, and SufD all showed a peripheral localization, consistent with putative inner membrane localization and interaction with RodZ. Notably, heterologous overexpression of the abovementioned proteins could not complement E. coli mutants, indicating that these proteins might play different functions in these two bacteria or that important regulators are not conserved. Altogether, this study brings new insights to the composition of the chlamydial divisome and points to links between protein secretion, degradation, iron homeostasis, and chlamydial division.

A SpoIID Homolog Cleaves Glycan Strands at the Chlamydial Division Septum.

Chlamydiales species are obligate intracellular bacteria lacking a classical peptidoglycan sacculus but relying on peptidoglycan synthesis for cytokinesis. While septal peptidoglycan biosynthesis seems to be regulated by MreB actin and its membrane anchor RodZ rather than FtsZ tubulin in Chlamydiales, the mechanism of peptidoglycan remodeling is poorly understood. An amidase conserved in Chlamydiales is able to cleave peptide stems in peptidoglycan, but it is not clear how peptidoglycan glycan strands are cleaved since no classical lytic transglycosylase is encoded in chlamydial genomes. However, a protein containing a SpoIID domain, known to possess transglycosylase activity in Bacillus subtilis, is conserved in Chlamydiales We show here that the SpoIID homologue of the Chlamydia-related pathogen Waddlia chondrophila is a septal peptidoglycan-binding protein. Moreover, we demonstrate that SpoIID acts as a lytic transglycosylase on peptidoglycan and as a muramidase on denuded glycan strands in vitro As SpoIID-like proteins are widespread in nonsporulating bacteria, SpoIID might commonly be a septal peptidoglycan remodeling protein in bacteria, including obligate intracellular pathogens, and thus might represent a promising drug target.IMPORTANCEChlamydiales species are obligate intracellular bacteria and important human pathogens that have a minimal division machinery lacking the proteins that are essential for bacterial division in other species, such as FtsZ. Chlamydial division requires synthesis of peptidoglycan, which forms a ring at the division septum and is rapidly turned over. However, little is known of peptidoglycan degradation, because many peptidoglycan-degrading enzymes are not encoded by chlamydial genomes. Here we show that an homologue of SpoIID, a peptidoglycan-degrading enzyme involved in sporulation of bacteria such as Bacillus subtilis, is expressed in Chlamydiales, localizes at the division septum, and degrades peptidoglycan in vitro, indicating that SpoIID is not only involved in sporulation but also likely implicated in division of some bacteria.

Sequencing the Obligate Intracellular Rhabdochlamydia helvetica within Its Tick Host Ixodes ricinus to Investigate Their Symbiotic Relationship.

The Rhabdochlamydiaceae family is one of the most widely distributed within the phylum Chlamydiae, but most of its members remain uncultivable. Rhabdochlamydia 16S rRNA was recently reported in more than 2% of 8,534 pools of ticks from Switzerland. Shotgun metagenomics was performed on a pool of five female Ixodes ricinus ticks presenting a high concentration of chlamydial DNA, allowing the assembly of a high-quality draft genome. About 60% of sequence reads originated from a single bacterial population that was named "Candidatus Rhabdochlamydia helvetica" whereas only few thousand reads mapped to the genome of "Candidatus Midichloria mitochondrii," a symbiont normally observed in all I. ricinus females. The 1.8 Mbp genome of R. helvetica is smaller than other Chlamydia-related bacteria. Comparative analyses with other chlamydial genomes identified transposases of the PD-(D/E)XK nuclease family that are unique to this new genome. These transposases show evidence of interphylum horizontal gene transfers between multiple arthropod endosymbionts, including Cardinium spp. (Bacteroidetes) and diverse proteobacteria such as Wolbachia, Rickettsia spp. (Rickettsiales), and Caedimonas varicaedens (Holosporales). Bacterial symbionts were previously suggested to provide B-vitamins to hematophagous hosts. However, incomplete metabolic capacities including for B-vitamin biosynthesis, high bacterial density and limited prevalence suggest that R. helvetica is parasitic rather than symbiotic to its host. The identification of novel Rhabdochlamydia strains in different hosts and their sequencing will help understanding if members of this genus have become highly specialized parasites with reduced genomes, like the Chlamydiaceae, or if they could be pathogenic to humans using ticks as a transmission vector.

Insight in the biology of Chlamydia-related bacteria.

The Chlamydiales order is composed of obligate intracellular bacteria and includes the Chlamydiaceae family and several family-level lineages called Chlamydia-related bacteria. In this review we will highlight the conserved and distinct biological features between these two groups. We will show how a better characterization of Chlamydia-related bacteria may increase our understanding on the Chlamydiales order evolution, and may help identifying new therapeutic targets to treat chlamydial infections.

Chlamydiales, Anaplasma and Bartonella: persistence and immune escape of intracellular bacteria.

Intracellular bacteria, such as Chlamydiales, Anaplasma or Bartonella, need to persist inside their host in order to complete their developmental cycle and to infect new hosts. In order to escape from the host immune system, intracellular bacteria have developed diverse mechanisms of persistence, which can directly impact the health of their host.

The Anticancer Peptide TAT-RasGAP317-326 Exerts Broad Antimicrobial Activity.

Antibiotic resistance has become a major health issue. Nosocomial infections and the prevalence of resistant pathogenic bacterial strains are rising steadily. Therefore, there is an urgent need to develop new classes of antibiotics effective on multi-resistant nosocomial pathogenic bacteria. We have previously shown that a cell-permeable peptide derived from the p120 Ras GTPase-activating protein (RasGAP), called TAT-RasGAP317-326, induces cancer cell death, inhibits metastatic progression, and sensitizes tumor cells to various anti-cancer treatments in vitro and in vivo. We here report that TAT-RasGAP317-326 also possesses antimicrobial activity. In vitro, TAT-RasGAP317-326, but not mutated or truncated forms of the peptide, efficiently killed a series of bacteria including Escherichia coli, Acinetobacter baumannii, Staphylococcus aureus, and Pseudomonas aeruginosa. In vivo experiments revealed that TAT-RasGAP317-326 protects mice from lethal E. coli-induced peritonitis if administrated locally at the onset of infection. However, the protective effect was lost when treatment was delayed, likely due to rapid clearance and inadequate biodistribution of the peptide. Peptide modifications might overcome these shortcomings to increase the in vivo efficacy of the compound in the context of the currently limited antimicrobial options.

The role of peptidoglycan in chlamydial cell division: towards resolving the chlamydial anomaly.

Chlamydiales are obligate intracellular bacteria including some important pathogens causing trachoma, genital tract infections and pneumonia, among others. They share an atypical division mechanism, which is independent of an FtsZ homologue. However, they divide by binary fission, in a process inhibited by penicillin derivatives, causing the formation of an aberrant form of the bacteria, which is able to survive in the presence of the antibiotic. The paradox of penicillin sensitivity of chlamydial cells in the absence of detectable peptidoglycan (PG) was dubbed the chlamydial anomaly, since no PG modified by enzymes (Pbps) that are the usual target of penicillin could be detected in Chlamydiales. We review here the recent advances in this field with the first direct and indirect evidences of PG-like material in both Chlamydiaceae and Chlamydia-related bacteria. Moreover, PG biosynthesis is required for proper localization of the newly described septal proteins RodZ and NlpD. Taken together, these new results set the stage for a better understanding of the role of PG and septal proteins in the division mechanism of Chlamydiales and illuminate the long-standing chlamydial anomaly. Moreover, understanding the chlamydial division mechanism is critical for the development of new antibiotics for the treatment of chlamydial chronic infections.

Disassembly of a Medial Transenvelope Structure by Antibiotics during Intracellular Division.

Chlamydiales possess a minimal but functional peptidoglycan precursor biosynthetic and remodeling pathway involved in the assembly of the division septum by an atypical cytokinetic machine and cryptic or modified peptidoglycan-like structure (PGLS). How this reduced cytokinetic machine collectively coordinates the invagination of the envelope has not yet been explored in Chlamydiales. In other Gram-negative bacteria, peptidoglycan provides anchor points that connect the outer membrane to the peptidoglycan during constriction using the Pal-Tol complex. Purifying PGLS and associated proteins from the chlamydial pathogen Waddlia chondrophila, we unearthed the Pal protein as a peptidoglycan-binding protein that localizes to the chlamydial division septum along with other components of the Pal-Tol complex. Together, our PGLS characterization and peptidoglycan-binding assays support the notion that diaminopimelic acid is an important determinant recruiting Pal to the division plane to coordinate the invagination of all envelope layers with the conserved Pal-Tol complex, even during osmotically protected intracellular growth.

ESCCAR international congress on Rickettsia and other intracellular bacteria.

The European Society for the study of Chlamydia, Coxiella, Anaplasma and Rickettsia (ESCCAR) held his triennial international meeting in Lausanne. This meeting gathered 165 scientists from 28 countries and all 5 continents, allowing efficient networking and major scientific exchanges. Topics covered include molecular and cellular microbiology, genomics, as well as epidemiology, veterinary and human medicine. Several breakthroughs have been revealed at the meeting, such as (i) the presence of CRISPR (the "prokaryotic immune system") in chlamydiae, (ii) an Anaplasma effector involved in host chromatin remodelling, (iii) the polarity of the type III secretion system of chlamydiae during the entry process revealed by cryo-electron tomography. Moreover, the ESCCAR meeting was a unique opportunity to be exposed to cutting-edge science and to listen to comprehensive talks on current hot topics.