Recognition of microbial patterns by host pattern recognition receptors is a key step in immune activation in multicellular eukaryotes. Peptidoglycans (PGNs) are major components of bacterial cell walls that possess immunity-stimulating activities in metazoans and plants. Here we show that PGN sensing and immunity to bacterial infection in Arabidopsis thaliana requires three lysin-motif (LysM) domain proteins. LYM1 and LYM3 are plasma membrane proteins that physically interact with PGNs and mediate Arabidopsis sensitivity to structurally different PGNs from Gram-negative and Gram-positive bacteria. lym1 and lym3 mutants lack PGN-induced changes in transcriptome activity patterns, but respond to fungus-derived chitin, a pattern structurally related to PGNs, in a wild-type manner. Notably, lym1 , lym3 , and lym3 lym1 mutant genotypes exhibit supersusceptibility to infection with virulent Pseudomonas syringae pathovar tomato DC3000. Defects in basal immunity in lym3 lym1 double mutants resemble those observed in lym1 and lym3 single mutants, suggesting that both proteins are part of the same recognition system. We further show that deletion of CERK1, a LysM receptor kinase that had previously been implicated in chitin perception and immunity to fungal infection in Arabidopsis , phenocopies defects observed in lym1 and lym3 mutants, such as peptidoglycan insensitivity and enhanced susceptibility to bacterial infection. Altogether, our findings suggest that plants share with metazoans the ability to recognize bacterial PGNs. However, as Arabidopsis LysM domain proteins LYM1, LYM3, and CERK1 form a PGN recognition system that is unrelated to metazoan PGN receptors, we propose that lineage-specific PGN perception systems have arisen through convergent evolution.

The human nasal microbiota is highly variable and dynamic often enclosing major pathogens such as Staphylococcus aureus. The potential roles of bacteriocins or other mechanisms allowing certain bacterial clones to prevail in this nutrient-poor habitat have hardly been studied. Of 89 nasal Staphylococcus isolates, unexpectedly, the vast majority (84%) was found to produce antimicrobial substances in particular under habitat-specific stress conditions, such as iron limitation or exposure to hydrogen peroxide. Activity spectra were generally narrow but highly variable with activities against certain nasal members of the Actinobacteria, Proteobacteria, Firmicutes, or several groups of bacteria. Staphylococcus species and many other Firmicutes were insusceptible to most of the compounds. A representative bacteriocin was identified as a nukacin-related peptide whose inactivation reduced the capacity of the producer Staphylococcus epidermidis IVK45 to limit growth of other nasal bacteria. Of note, the bacteriocin genes were found on mobile genetic elements exhibiting signs of extensive horizontal gene transfer and rearrangements. Thus, continuously evolving bacteriocins appear to govern bacterial competition in the human nose and specific bacteriocins may become important agents for eradication of notorious opportunistic pathogens from human microbiota. The complex and dynamic microbial communities of human body surfaces are of utmost importance for human body functions in health and diseases. Human microbiomes contribute to metabolic processes, instruct the immune system, and often include antibiotic-resistant pathogens, responsible for the majority of severe bacterial infections. It is generally accepted that microbiota composition is strongly affected by mechanisms of microbial interference, but how specific bacteria may achieve fitness benefits and outcompete other microbes has remained largely unknown. We demonstrate that production of antimicrobial bacteriocins is not an occasional trait but a dominant and highly variable strategy among human nasal bacteria for limiting the growth of competing microbes. We found that more than 80% of nasal Staphylococcus isolates produce bacteriocins with highly diverse activity spectra, in particular under habitat-specific stress conditions such as iron limitation and exposure to hydrogen peroxide. Inactivation of a representative bacteriocin diminished the producer’s competitive capability indicating that bacteriocins may be a major driving force for the dynamics of microbiomes in nutrient-poor habitats such as the human nose. The identification of bacteriocin genes on mobile genetic elements with composite structure suggests that they are subject to highly dynamic co-evolutionary processes.

Background: Studies on mycorrhiza associated bacteria suggest that bacterial-fungal interactions play important roles during mycorrhiza formation and affect plant health. We surveyed Streptomyces Actinobacteria, known as antibiotic producers and antagonists of fungi, from Norway spruce mycorrhizas with predominantly Piloderma species as the fungal partner. Results: Fifteen Streptomyces isolates exhibited substantial variation in inhibition of tested mycorrhizal and plant pathogenic fungi (Amanita muscaria, Fusarium oxysporum, Hebeloma cylindrosporum, Heterobasidion abietinum, Heterobasidion annosum, Laccaria bicolor, Piloderma croceum). The growth of the mycorrhiza-forming fungus Laccaria bicolor was stimulated by some of the streptomycetes, and Piloderma croceum was only moderately affected. Bacteria responded to the streptomycetes differently than the fungi. For instance the strain Streptomyces sp. AcM11, which inhibited most tested fungi, was less inhibitory to bacteria than other tested streptomycetes. The determined patterns of Streptomyces-microbe interactions were associated with distinct patterns of secondary metabolite production. Notably, potentially novel metabolites were produced by strains that were less antagonistic to fungi. Most of the identified metabolites were antibiotics (e.g. cycloheximide, actiphenol) and siderophores (e.g. ferulic acid, desferroxiamines). Plant disease resistance was activated by a single streptomycete strain only. Conclusions: Mycorrhiza associated streptomycetes appear to have an important role in inhibiting the growth of fungi and bacteria. Additionally, our study indicates that the Streptomyces strains, which are not general antagonists of fungi, may produce still un-described metabolites.

Background & Aims The intestinal microbiota is an important determinant of the mucosal response. In patients with inflammatory bowel diseases, the mucosal immune system has inappropriate interactions with the intestinal microbiota. We investigated how the composition of the intestinal microbiota affects its endotoxicity and development of colitis in mice. Methods Germ-free C57BL/6J-Rag^1tm1Mom (Rag1^−/−) mice were colonized with 2 different types of complex intestinal microbiota. Colitis was induced in Rag1^−/− mice by transfer of CD4⁺CD62L⁺ T cells from C57BL/6J mice. Colonic tissues were collected and used for histologic analysis and cell isolation. Activation of lamina propria dendritic cells and T cells was analyzed by flow cytometry. Results After transfer of CD4⁺CD62L⁺ T cells, mice with intestinal Endo^lo microbiota (a low proportion of Enterobacteriaceae, high proportion of Bacteroidetes, and low endotoxicity) maintained mucosal immune homeostasis, and mice with highly endotoxic Endo^hi microbiota (a high proportion of Enterobacteriaceae and low proportion of Bacteroidetes) developed colitis. To determine whether the effects of Endo^hi microbiota were related to the higher endotoxic activity of lipopolysaccharide (LPS), we compared LPS from Enterobacteriaceae with that of Bacteroidetes. Administration of Escherichia coli JM83 (wild-type LPS) to the mice exacerbated colitis, and Escherichia coli JM83 + htrBPG (mutated LPS, with lower endotoxicity, similar to that of Bacteroidetes) prevented development of colitis after transfer of the T cells to mice. Conclusions The endotoxicity of LPS produced by the intestinal microbiota is a determinant of whether mice develop colitis after transfer of CD4⁺CD62L⁺ T cells. This finding might aid the design of novel biologics or probiotics to treat inflammatory bowel disease.

We describe a procedure for the conjugative transfer of phage P1-derived Artificial Chromosome (PAC) library clones containing large natural product gene clusters (≥70 kilobases) to Streptomyces coelicolor strains that have been engineered for improved heterologous production of natural products. This approach is demonstrated using the gene cluster for FK506 (tacrolimus), a clinically important immunosuppressant of high commercial value. The entire 83.5 kb FK506 gene cluster from Streptomyces tsukubaensis NRRL 18488 present in one 130 kb PAC clone was introduced into four different S. coelicolor derivatives and all produced FK506 and smaller amounts of the related compound FK520. FK506 yields were increased by approximately five-fold (from 1.2 mg L^-1 to 5.5 mg L^-1) in S. coelicolor M1146 containing the FK506 PAC upon over-expression of the FK506 LuxR regulatory gene fkbN. The PAC-based gene cluster conjugation methodology described here provides a tractable means to evaluate and manipulate FK506 biosynthesis and is readily applicable to other large gene clusters encoding natural products of interest to medicine, agriculture and biotechnology.

The emergence of antibiotic-resistant pathogenic bacteria within the last decades is one reason for the urgent need for new antibacterial agents. A strategy to discover new anti-infective compounds is the evaluation of the genetic capacity of secondary metabolite producers and the activation of cryptic gene clusters (genome mining). One genus known for its potential to synthesize medically important products is Amycolatopsis . However, Amycolatopsis japonicum does not produce an antibiotic under standard laboratory conditions. In contrast to most Amycolatopsis strains, A. japonicum is genetically tractable with different methods. In order to activate a possible silent glycopeptide cluster, we introduced a gene encoding the transcriptional activator of balhimycin biosynthesis, the bbr gene from Amycolatopsis balhimycina ( bbr _Aba ), into A. japonicum . This resulted in the production of an antibiotically active compound. Following whole-genome sequencing of A. japonicum , 29 cryptic gene clusters were identified by genome mining. One of these gene clusters is a putative glycopeptide biosynthesis gene cluster. Using bioinformatic tools, ristomycin (syn. ristocetin), a type III glycopeptide, which has antibacterial activity and which is used for the diagnosis of von Willebrand disease and Bernard-Soulier syndrome, was deduced as a possible product of the gene cluster. Chemical analyses by high-performance liquid chromatography and mass spectrometry (HPLC-MS), tandem mass spectrometry (MS/MS), and nuclear magnetic resonance (NMR) spectroscopy confirmed the in silico prediction that the recombinant A. japonicum /pRM4- bbr _Aba synthesizes ristomycin A.