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Journal articleTenland E, Krishnan N, Rönnholm A, et al., 2018,
A novel derivative of the fungal antimicrobial peptide plectasin is active against Mycobacterium tuberculosis
, Tuberculosis, Vol: 113, Pages: 231-238, ISSN: 1472-9792Tuberculosis has been reaffirmed as the infectious disease causing most deaths in the world. Co-infection with HIV and the increase in multi-drug resistant Mycobacterium tuberculosis strains complicate treatment and increases mortality rates, making the development of new drugs an urgent priority. In this study we have identified a promising candidate by screening antimicrobial peptides for their capacity to inhibit mycobacterial growth. This non-toxic peptide, NZX, is capable of inhibiting both clinical strains of M. tuberculosis and an MDR strain at therapeutic concentrations. The therapeutic potential of NZX is further supported in vivo where NZX significantly lowered the bacterial load with only five days of treatment, comparable to rifampicin treatment over the same period. NZX possesses intracellular inhibitory capacity and co-localizes with intracellular bacteria in infected murine lungs. In conclusion, the data presented strongly supports the therapeutic potential of NZX in future anti-TB treatment.
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Journal articleDortet L, Bonnin RA, Pennisi I, et al., 2018,
Rapid detection and discrimination of chromosome-and MCR-plasmid-mediated resistance to polymyxins by MALDI-TOF MS in Escherichia coli: the MALDIxin test
, Journal of Antimicrobial Chemotherapy, Vol: 73, Pages: 3359-3367, ISSN: 0305-7453BackgroundPolymyxins are currently considered a last-resort treatment for infections caused by MDR Gram-negative bacteria. Recently, the emergence of carbapenemase-producing Enterobacteriaceae has accelerated the use of polymyxins in the clinic, resulting in an increase in polymyxin-resistant bacteria. Polymyxin resistance arises through modification of lipid A, such as the addition of phosphoethanolamine (pETN). The underlying mechanisms involve numerous chromosome-encoded genes or, more worryingly, a plasmid-encoded pETN transferase named MCR. Currently, detection of polymyxin resistance is difficult and time consuming.ObjectivesTo develop a rapid diagnostic test that can identify polymyxin resistance and at the same time differentiate between chromosome- and plasmid-encoded resistances.MethodsWe developed a MALDI-TOF MS-based method, named the MALDIxin test, which allows the detection of polymyxin resistance-related modifications to lipid A (i.e. pETN addition), on intact bacteria, in <15 min.ResultsUsing a characterized collection of polymyxin-susceptible and -resistant Escherichia coli, we demonstrated that our method is able to identify polymyxin-resistant isolates in 15 min whilst simultaneously discriminating between chromosome- and plasmid-encoded resistance. We validated the MALDIxin test on different media, using fresh and aged colonies and show that it successfully detects all MCR-1 producers in a blindly analysed set of carbapenemase-producing E. coli strains.ConclusionsThe MALDIxin test is an accurate, rapid, cost-effective and scalable method that represents a major advance in the diagnosis of polymyxin resistance by directly assessing lipid A modifications in intact bacteria.
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Journal articleSgro GG, Costa TRD, Cenens W, et al., 2018,
Cryo-EM structure of the bacteria-killing type IV secretion system core complex from Xanthomonas citri
, Nature Microbiology, Vol: 3, Pages: 1429-1440, ISSN: 2058-5276Type IV secretion (T4S) systems form the most common and versatile class of secretion systems in bacteria, capable of injecting both proteins and DNAs into host cells. T4S systems are typically composed of 12 components that form 2 major assemblies: the inner membrane complex embedded in the inner membrane and the core complex embedded in both the inner and outer membranes. Here we present the 3.3 Å-resolution cryo-electron microscopy model of the T4S system core complex from Xanthomonas citri, a phytopathogen that utilizes this system to kill bacterial competitors. An extensive mutational investigation was performed to probe the vast network of protein–protein interactions in this 1.13-MDa assembly. This structure expands our knowledge of the molecular details of T4S system organization, assembly and evolution.
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Journal articleKoymans KJ, Feitsma LJ, Bisschop A, et al., 2018,
Molecular basis determining species specificity for TLR2 inhibition by staphylococcal superantigen-like protein 3 (SSL3)
, Veterinary Research, Vol: 49, ISSN: 0928-4249Staphylococcus aureus is a versatile opportunistic pathogen, causing disease in human and animal species. Its pathogenicity is linked to the ability of S. aureus to secrete immunomodulatory molecules. These evasion proteins bind to host receptors or their ligands, resulting in inhibitory effects through high affinity protein–protein interactions. Staphylococcal evasion molecules are often species-specific due to differences in host target proteins between species. We recently solved the crystal structure of murine TLR2 in complex with immunomodulatory molecule staphylococcal superantigen-like protein 3 (SSL3), which revealed the essential residues within SSL3 for TLR2 inhibition. In this study we aimed to investigate the molecular basis of the interaction on the TLR2 side. The SSL3 binding region on murine TLR2 was compared to that of other species through sequence alignment and homology modeling, which identified interspecies differences. To examine whether this resulted in altered SSL3 activity on the corresponding TLR2s, bovine, equine, human, and murine TLR2 were stably expressed in HEK293T cells and the ability of SSL3 to inhibit TLR2 was assessed. We found that SSL3 was unable to inhibit bovine TLR2. Subsequent loss and gain of function mutagenesis showed that the lack of inhibition is explained by the absence of two tyrosine residues in bovine TLR2 that play a prominent role in the SSL3–TLR2 interface. We found no evidence for the existence of allelic SSL3 variants that have adapted to the bovine host. Thus, within this paper we reveal the molecular determinants of the TLR2–SSL3 interaction which adds to our understanding of staphylococcal host specificity.
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Journal articleManning KA, Quiles-Puchalt N, Penades JR, et al., 2018,
A novel ejection protein from bacteriophage 80α that promotes lytic growth
, VIROLOGY, Vol: 525, Pages: 237-247, ISSN: 0042-6822- Author Web Link
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Journal articlePompeo F, Rismondo J, Gründling A, et al., 2018,
Investigation of the phosphorylation of Bacillus subtilis LTA synthases by the serine/threonine kinase PrkC
, Scientific Reports, Vol: 8, ISSN: 2045-2322Bacillus subtilis possesses four lipoteichoic acid synthases LtaS, YfnI, YvgJ and YqgS involved in the synthesis of cell wall. The crystal structure of the extracellular domain of LtaS revealed a phosphorylated threonine and YfnI was identified in two independent phosphoproteome studies. Here, we show that the four LTA synthases can be phosphorylated in vitro by the Ser/Thr kinase PrkC. Phosphorylation neither affects the export/release of YfnI nor its substrate binding. However, we observed that a phosphomimetic form of YfnI was active whereas its phosphoablative form was inactive. The phenotypes of the strains deleted for prkC or prpC (coding for a phosphatase) are fairly similar to those of the strains producing the phosphoablative or phosphomimetic YfnI proteins. Clear evidence proving that PrkC phosphorylates YfnI in vivo is still missing but our data suggest that the activity of all LTA synthases may be regulated by phosphorylation. Nonetheless, their function is non-redundant in cell. Indeed, the deletion of either ltaS or yfnI gene could restore a normal growth and shape to a ΔyvcK mutant strain but this was not the case for yvgJ or yqgS. The synthesis of cell wall must then be highly regulated to guarantee correct morphogenesis whatever the growth conditions.
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Journal articleChorev DS, Baker LA, Wu D, et al., 2018,
Protein assemblies ejected directly from native membranes yield complexes for mass spectrometry
, SCIENCE, Vol: 362, Pages: 829-+, ISSN: 0036-8075 -
Journal articleDortet L, Potron A, Bonnin RA, et al., 2018,
Rapid detection of colistin resistance in Acinetobacter baumannii using MALDI-TOF-based lipidomics on intact bacteria
, Scientific Reports, Vol: 8, ISSN: 2045-2322With the dissemination of extremely drug resistant bacteria, colistin is now considered as the last-resort therapy for the treatment of infection caused by Gram-negative bacilli (including carbapenemase producers). Unfortunately, the increase use of colistin has resulted in the emergence of resistance as well. In A. baumannii, colistin resistance is mostly caused by the addition of phosphoethanolamine to the lipid A through the action of a phosphoethanolamine transferase chromosomally-encoded by the pmrC gene, which is regulated by the two-component system PmrA/PmrB. In A. baumannii clinical isolate the main resistance mechanism to colistin involves mutations in pmrA, pmrB or pmrC genes leading to the overexpression of pmrC. Although, rapid detection of resistance is one of the key issues to improve the treatment of infected patient, detection of colistin resistance in A. baumannii still relies on MIC determination through microdilution, which is time-consuming (16–24 h). Here, we evaluated the performance of a recently described MALDI-TOF-based assay, the MALDIxin test, which allows the rapid detection of colistin resistance-related modifications to lipid A (i.e phosphoethanolamine addition). This test accurately detected all colistin-resistant A. baumannii isolates in less than 15 minutes, directly on intact bacteria with a very limited sample preparation prior MALDI-TOF analysis.
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Journal articleDecout A, Silva-Gomes S, Drocourt D, et al., 2018,
Deciphering the molecular basis of mycobacteria and lipoglycan recognition by the C-type lectin Dectin-2
, Scientific Reports, Vol: 8, ISSN: 2045-2322Dectin-2 is a C-type lectin involved in the recognition of several pathogens such as Aspergillus fumigatus, Candida albicans, Schistosoma mansonii, and Mycobacterium tuberculosis that triggers Th17 immune responses. Identifying pathogen ligands and understanding the molecular basis of their recognition is one of the current challenges. Purified M. tuberculosis mannose-capped lipoarabinomannan (ManLAM) was shown to induce signaling via Dectin-2, an activity that requires the (α1 → 2)-linked mannosides forming the caps. Here, using isogenic M. tuberculosis mutant strains, we demonstrate that ManLAM is a bona fide and actually the sole ligand mediating bacilli recognition by Dectin-2, although M. tuberculosis produces a variety of cell envelope mannoconjugates, such as phosphatidyl-myo-inositol hexamannosides, lipomannan or manno(lipo)proteins, that bear (α1 → 2)-linked mannosides. In addition, we found that Dectin-2 can recognize lipoglycans from other bacterial species, such as Saccharotrix aerocolonigenes or the human opportunistic pathogen Tsukamurella paurometabola, suggesting that lipoglycans are prototypical Dectin-2 ligands. Finally, from a structure/function relationship perspective, we show, using lipoglycan variants and synthetic mannodendrimers, that dimannoside caps and multivalent interaction are required for ligand binding to and signaling via Dectin-2. Better understanding of the molecular basis of ligand recognition by Dectin-2 will pave the way for the rational design of potent adjuvants targeting this receptor.
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Journal articleStapels DAC, Hill PWS, Westermann AJ, et al., 2018,
Salmonella persisters undermine host immune defenses during antibiotic treatment
, Science, Vol: 362, Pages: 1156-1160, ISSN: 0036-8075Many bacterial infections are hard to treat and tend to relapse, possibly due to the presence of antibiotic-tolerant persisters. In vitro, persister cells appear to be dormant. After uptake of Salmonella species by macrophages, nongrowing persisters also occur, but their physiological state is poorly understood. In this work, we show that Salmonella persisters arising during macrophage infection maintain a metabolically active state. Persisters reprogram macrophages by means of effectors secreted by the Salmonella pathogenicity island 2 type 3 secretion system. These effectors dampened proinflammatory innate immune responses and induced anti-inflammatory macrophage polarization. Such reprogramming allowed nongrowing Salmonella cells to survive for extended periods in their host. Persisters undermining host immune defenses might confer an advantage to the pathogen during relapse once antibiotic pressure is relieved.
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