The Pneumococcal Type 1 Pilus Genes Are Thermoregulated and Are Repressed by a Member of the Snf2 Protein Family

ABSTRACT In Streptococcus pneumoniae, the type 1 pilus is involved in many steps of pathogenesis, including adherence to epithelial cells, mediation of inflammation, escape from macrophages, and the formation of biofilms. The type 1 pilus genes are expressed in a bistable fashion with cells switching between “on” and “off” expression states. Bistable expression of these genes is due to their control by RlrA, a positive regulator subject to control by a positive-feedback loop. The type 1 pilus genes are also thought to be negatively regulated by a large number of repressors. Here we show that expression of the type 1 pilus genes is thermosensitive and switched off at growth temperatures below 31°C. We also report that the on expression state of the type 1 pilus genes is highly stable, a phenomenon which we show likely contributed to the erroneous identification of many proteins as negative regulators of these genes. Finally, we exploited the effect of low temperature on pilus gene expression to help identify SP_1523, an Snf2-type protein, as a novel negative regulator of the pilus genes. Our findings establish that the type 1 pilus genes are thermoregulated and are repressed by a member of the Snf2 protein family. They also refute the notion that these genes are controlled by 8 previously described negative regulators. IMPORTANCE Streptococcus pneumoniae is the leading cause of death from respiratory infections in children. Many bacterial factors contribute to pneumococcal virulence and nasopharyngeal colonization. The type 1 pneumococcal pilus plays an important role in mouse models and in epithelial adherence and is expressed in a bistable fashion. Here we show that the “on” state is highly stable, which may explain the prior misidentification of negative regulators of pilus expression. We also show that expression of pilus genes is thermosensitive: virtually no expression can be detected at temperatures found in the anterior nares of humans. We took advantage of this property to identify a negative regulator of pilus expression, a member of a family of proteins widely conserved across Gram-positive bacteria.

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Molecular Determinants of Substrate Selectivity of a Pneumococcal Rgg-Regulated Peptidase-Containing ABC Transporter

mBio ◽

10.1128/mbio.02502-19 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 2

Author(s):

Charles Y. Wang ◽

Jennifer S. Medlin ◽

Don R. Nguyen ◽

W. Miguel Disbennett ◽

Suzanne Dawid

Keyword(s):

Streptococcus Pneumoniae ◽

Abc Transporters ◽

Signal Sequence ◽

Substrate Recognition ◽

Substrate Selectivity ◽

Nasopharyngeal Colonization ◽

Competitive Fitness ◽

Content Type ◽

Fitness Advantage ◽

Molecular Determinants

ABSTRACT Peptidase-containing ABC transporters (PCATs) are a widely distributed family of transporters which secrete double-glycine (GG) peptides. In the opportunistic pathogen Streptococcus pneumoniae (pneumococcus), the PCATs ComAB and BlpAB have been shown to secrete quorum-sensing pheromones and bacteriocins related to the competence and pneumocin pathways. Here, we describe another pneumococcal PCAT, RtgAB, encoded by the rtg locus and found intact in 17% of strains. The Rgg/SHP-like quorum-sensing system RtgR/S, which uses a peptide pheromone with a distinctive Trp-X-Trp motif, regulates expression of the rtg locus and provides a competitive fitness advantage in a mouse model of nasopharyngeal colonization. RtgAB secretes a set of coregulated rtg GG peptides. ComAB and BlpAB, which share a substrate pool, do not secrete the rtg GG peptides. Similarly, RtgAB does not efficiently secrete ComAB/BlpAB substrates. We examined the molecular determinants of substrate selectivity between ComAB, BlpAB, and RtgAB and found that the GG peptide signal sequences contain all the information necessary to direct secretion through specific transporters. Secretion through ComAB and BlpAB depends largely on the identity of four conserved hydrophobic signal sequence residues previously implicated in substrate recognition by PCATs. In contrast, a motif situated at the N-terminal end of the signal sequence, found only in rtg GG peptides, directs secretion through RtgAB. These findings illustrate the complexity in predicting substrate-PCAT pairings by demonstrating specificity that is not dictated solely by signal sequence residues previously implicated in substrate recognition. IMPORTANCE The export of peptides from the cell is a fundamental process carried out by all bacteria. One method of bacterial peptide export relies on a family of transporters called peptidase-containing ABC transporters (PCATs). PCATs export so-called GG peptides which carry out diverse functions, including cell-to-cell communication and interbacterial competition. In this work, we describe a PCAT-encoding genetic locus, rtg, in the pathogen Streptococcus pneumoniae (pneumococcus). The rtg locus is linked to increased competitive fitness advantage in a mouse model of nasopharyngeal colonization. We also describe how the rtg PCAT preferentially secretes a set of coregulated GG peptides but not GG peptides secreted by other pneumococcal PCATs. These findings illuminate a relatively understudied part of PCAT biology: how these transporters discriminate between different subsets of GG peptides. Ultimately, expanding our knowledge of PCATs will advance our understanding of the many microbial processes dependent on these transporters.

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The N-Acetylglucosaminidase LytB of Streptococcus pneumoniae Is Involved in the Structure and Formation of Biofilms

Applied and Environmental Microbiology ◽

10.1128/aem.00280-20 ◽

2020 ◽

Vol 86 (10) ◽

Cited By ~ 1

Author(s):

Mirian Domenech ◽

Ernesto García

Keyword(s):

Streptococcus Pneumoniae ◽

Biofilm Formation ◽

Single Gene ◽

Extracellular Polysaccharide ◽

Extracellular Dna ◽

Nasopharyngeal Colonization ◽

Content Type ◽

Daughter Cells ◽

Biofilm Matrix

ABSTRACT The N-acetylglucosaminidase LytB of Streptococcus pneumoniae is involved in nasopharyngeal colonization and is responsible for cell separation at the end of cell division; thus, ΔlytB mutants form long chains of cells. This paper reports the construction and properties of a defective pneumococcal mutant producing an inactive LytB protein (LytBE585A). It is shown that an enzymatically active LytB is required for in vitro biofilm formation, as lytB mutants (either ΔlytB or producing the inactive LytBE585A) are incapable of forming substantial biofilms, despite that extracellular DNA is present in the biofilm matrix. Adding small amounts (0.5 to 2.0 μg/ml) of exogenous LytB or some LytB constructs restored the biofilm-forming capacity of lytB mutants to wild-type levels. The LytBE585A mutant formed biofilm more rapidly than ΔlytB mutants in the presence of LytB. This suggests that the mutant protein acted in a structural role, likely through the formation of complexes with extracellular DNA. The chain-dispersing capacity of LytB allowed the separation of daughter cells, presumably facilitating the formation of microcolonies and, finally, of biofilms. A role for the possible involvement of LytB in the synthesis of the extracellular polysaccharide component of the biofilm matrix is also discussed. IMPORTANCE It has been previously accepted that biofilm formation in S. pneumoniae must be a multigenic trait because the mutation of a single gene has led to only to partial inhibition of biofilm production. In the present study, however, evidence that the N-acetylglucosaminidase LytB is crucial in biofilm formation is provided. Despite the presence of extracellular DNA, strains either deficient in LytB or producing a defective LytB enzyme formed only shallow biofilms.

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Recognition of Streptococcus pneumoniae and Muramyl Dipeptide by NOD2 Results in Potent Induction of MMP-9, Which Can Be Controlled by Lipopolysaccharide Stimulation

Infection and Immunity ◽

10.1128/iai.02150-14 ◽

2014 ◽

Vol 82 (12) ◽

pp. 4952-4958 ◽

Cited By ~ 10

Author(s):

Marloes Vissers ◽

Yvonne Hartman ◽

Laszlo Groh ◽

Dirk J. de Jong ◽

Marien I. de Jonge ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Mononuclear Cells ◽

Muramyl Dipeptide ◽

Tissue Inhibitor Of Metalloproteinase ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Potent Inducer ◽

Gram Negative ◽

Content Type ◽

Gram Positive Bacteria

ABSTRACTMatrix metallopeptidase 9 (MMP-9) is a protease involved in the degradation of extracellular matrix collagen. Evidence suggests that MMP-9 is involved in pathogenesis duringStreptococcus pneumoniaeinfection. However, not much is known about the induction of MMP-9 and the regulatory processes involved. We show here that the Gram-positive bacteria used in this study induced large amounts of MMP-9, in contrast to the Gram-negative bacteria that were used. An important pathogen-associated molecular pattern (PAMP) for Gram-positive bacteria is muramyl dipeptide (MDP). MDP is a very potent inducer of MMP-9 and showed a dose-dependent MMP-9 induction. Experiments using peripheral blood mononuclear cells (PBMCs) from Crohn's disease patients with nonfunctional NOD2 showed that MMP-9 induction byStreptococcus pneumoniaeand MDP is NOD2 dependent. Increasing amounts of lipopolysaccharide (LPS), an important PAMP for Gram-negative bacteria, resulted in decreasing amounts of MMP-9. Moreover, the induction of MMP-9 by MDP could be counteracted by simultaneously adding LPS. The inhibition of MMP-9 expression by LPS was found to be regulated posttranscriptionally, independently of tissue inhibitor of metalloproteinase 1 (TIMP-1), an endogenous inhibitor of MMP-9. Collectively, these data show thatStreptococcus pneumoniaeis able to induce large amounts of MMP-9. These high MMP-9 levels are potentially involved inStreptococcus pneumoniaepathogenesis.

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Natural Development of Antibodies against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis Protein Antigens during the First 13 Years of Life

Clinical and Vaccine Immunology ◽

10.1128/cvi.00341-16 ◽

2016 ◽

Vol 23 (11) ◽

pp. 878-883 ◽

Cited By ~ 3

Author(s):

Igor C. Borges ◽

Dafne C. Andrade ◽

Maria Regina A. Cardoso ◽

Jorma Toppari ◽

Mari Vähä-Mäkilä ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Haemophilus Influenzae ◽

Antibody Production ◽

Moraxella Catarrhalis ◽

Respiratory Infections ◽

Protein Antigens ◽

Serum Samples ◽

Content Type ◽

Natural Development ◽

Antibody Levels

ABSTRACTConserved protein antigens have been investigated as vaccine candidates against respiratory pathogens. We evaluated the natural development of antibodies againstStreptococcus pneumoniae,Haemophilus influenzae, andMoraxella catarrhalisproteins during childhood. Serum samples were collected from 50 healthy children from their first months to age 13 years (median sampling interval, 6 months). We also analyzed serum samples from 24 adults. Serum IgG antibodies against eight pneumococcal proteins (Ply, CbpA, PspA 1 and 2, PcpA, PhtD, StkP-C, and PcsB-N), threeH. influenzaeproteins, and fiveM. catarrhalisproteins were measured using a multiplexed bead-based immunoassay. Antibody levels were analyzed using multilevel mixed-effects regression and Spearman's correlation. Antibody levels against pneumococcal proteins peaked at 3 to 5 years of age and then reached a plateau. Antibody levels againstH. influenzaeproteins peaked during the second year and then stabilized. Antibody levels againstM. catarrhalisproteins peaked during the first year and then slowly decreased. Peak antibody levels during childhood were higher than those of adults. Correlations among pneumococcal antibody levels were highest among anti-CbpA, anti-PcpA, and anti-PhtD antibodies (r= 0.71 to 0.75;P< 0.001). The children presented 854 symptomatic respiratory infections on 586 occasions. Symptomatic respiratory infections did not improve prediction of antibody levels in the regression model. The maturation of immune responses against the investigated pneumococcal proteins shares similarities, especially among CbpA, PcpA, and PhtD. Antibody production againstH. influenzaeandM. catarrhalisproteins starts early in life and reaches peak levels earlier than antibody production against the pneumococcal proteins. Basal antibody levels are not related to the occurrence of symptomatic respiratory infections.

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Noninvasive Streptococcus pneumoniae Serotypes Recovered from Hospitalized Adult Patients in the United States in 2009 to 2012

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00182-15 ◽

2015 ◽

Vol 59 (9) ◽

pp. 5595-5601 ◽

Cited By ~ 17

Author(s):

Rodrigo E. Mendes ◽

Rosalind C. Hollingsworth ◽

Andrew Costello ◽

Ronald N. Jones ◽

Raul E. Isturiz ◽

...

Keyword(s):

United States ◽

Streptococcus Pneumoniae ◽

Conjugate Vaccine ◽

Pneumococcal Conjugate Vaccine ◽

Respiratory Infections ◽

Adult Population ◽

The United States ◽

Adult Patients ◽

Content Type ◽

Trends Over Time

ABSTRACTThis study was conducted to determine the serotype distribution and trends over time ofStreptococcus pneumoniaestrains associated with noninvasive infections among adult patients ≥18 years of age in the United States (2009 to 2012). A total of 2,927S. pneumoniaeisolates recovered from patients presenting with respiratory infections and obtained mainly (87.0%) from lower respiratory tract specimens (sputum) were included. The levels of the 7-valent pneumococcal conjugate vaccine (PCV7) serotypes remained stable over the 4-year study period (4.6% to 5.5%;P= 0.953). Overall, 13-valent pneumococcal conjugate vaccine (PCV13) serotypes were identified in 32.7% of samples, declining from 33.7% to 35.5% in 2009 to 2011 to 28.2% in 2012 (P= 0.007), with a significant decrease in the levels of serotypes 7F (P= 0.013) and 6A (P= 0.010). The levels of 19A remained constant (15.8% to 17.1%) during 2009 to 2011, dropping to 12.2% in 2012 (P= 0.089). The prevalence of serotypes associated with the 23-valent pneumococcal polysaccharide vaccine (PPSV23), but not PCV13, remained generally stable; however, the prevalence of serotypes 15B and 15C (15B/15C) increased from 2.7% to 6.3% (P= 0.010). The proportion of nonvaccine serotypes increased gradually during the study period (P= 0.044), particularly for serotype 35B (from 3.6% in 2009 to 8.2% in 2012;P= 0.001). Nonsusceptibility rates for penicillin (susceptible breakpoint, ≤2 μg/ml) and clindamycin against PCV7 serotypes decreased over the period. These results suggest the emergence of indirect effects following introduction of PCV13 for infants and young children; continued surveillance is needed to assess the burden of PCV13 serotypes in the adult population after the implementation of age-based recommendations in the United States.

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Nasal Tissue Extraction Is Essential for Characterization of the Murine Upper Respiratory Tract Microbiota

mSphere ◽

10.1128/msphere.00562-20 ◽

2020 ◽

Vol 5 (6) ◽

Author(s):

L. Patrick Schenck ◽

Joshua J. C. McGrath ◽

Daphnée Lamarche ◽

Martin R. Stämpfli ◽

Dawn M. E. Bowdish ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Respiratory Tract ◽

Research Design ◽

Respiratory Infections ◽

Upper Respiratory Tract ◽

Content Type ◽

Nasal Tissue ◽

Colonization Success ◽

Upper Respiratory ◽

Nasal Microbiota

ABSTRACT Respiratory infections are a leading cause of morbidity and mortality worldwide. Bacterial pathogens often colonize the upper respiratory tract (nose or mouth) prior to causing lower respiratory infections or invasive disease. Interactions within the upper respiratory tract between colonizing bacteria and the resident microbiota could contribute to colonization success and subsequent transmission. Human carriage studies have identified associations between pathogens such as Streptococcus pneumoniae and members of the resident microbiota, although few mechanisms of competition and cooperation have been identified and would be aided by the use of animal models. Little is known about the composition of the murine nasal microbiota; thus, we set out to improve assessment, including tissue sampling, composition, and comparison between mouse sources. Nasal washes were efficient in sampling the nasopharyngeal space but barely disrupted the nasal turbinates. Nasal tissue extraction increased the yield of cultivable bacterial compared to nasal washes, revealing distinct community compositions. Experimental pneumococcal colonization led to dominance by the colonizing pathogen in the nasopharynx and nasal turbinates, but the composition of the microbiota, and interactions with resident microbes, differed depending on the sampling method. Importantly, vendor source has a large impact on microbial composition. Bacterial interactions, including cooperation and colonization resistance, depend on the biogeography of the nose and should be considered during research design of experimental colonization with pathogens. IMPORTANCE The nasal microbiota is composed of species that play a role in the colonization success of pathogens, including Streptococcus pneumoniae and Staphylococcus aureus. Murine models provide the ability to explore disease pathogenesis, but little is known about the natural murine nasal microbiota. This study established techniques to allow the exploration of the bacterial members of the nasal microbiota. The mouse nasal microbiota included traditional respiratory bacteria, including Streptococcus, Staphylococcus, and Moraxella species. Analyses were affected by different sampling methods as well as the commercial source of the mice, which should be included in future research design of infectious disease research.

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Coping with an Essential Poison: a Genetic Suppressor Analysis Corroborates a Key Function of c-di-AMP in Controlling Potassium Ion Homeostasis in Gram-Positive Bacteria

Journal of Bacteriology ◽

10.1128/jb.00166-18 ◽

2018 ◽

Vol 200 (12) ◽

Cited By ~ 11

Author(s):

Fabian M. Commichau ◽

Jörg Stülke

Keyword(s):

Streptococcus Pneumoniae ◽

Ion Homeostasis ◽

Second Messenger ◽

Potassium Ion ◽

Complex Media ◽

Content Type ◽

Potassium Homeostasis ◽

Intracellular Potassium ◽

Gram Positive Bacteria ◽

Suppressor Analysis

ABSTRACT Cyclic di-AMP (c-di-AMP) is an important second messenger in bacteria. In most Firmicutes , the molecule is required for growth in complex media but also toxic upon accumulation. In an article on their current study, Zarrella and coworkers present a suppressor analysis of a Streptococcus pneumoniae strain that is unable to degrade c-di-AMP (T. M. Zarrella, D. W. Metzger, and G. Bai, J Bacteriol 200:e00045-18, 2018, https://doi.org/10.1128/JB.00045-18 ). Their study identifies new links between c-di-AMP and potassium homeostasis and supports the hypothesis that c-di-AMP serves as a second messenger to report about the intracellular potassium concentrations.

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Efficacy of Aminomethyl Spectinomycins against Complex Upper Respiratory Tract Bacterial Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02096-18 ◽

2019 ◽

Vol 63 (5) ◽

Cited By ~ 1

Author(s):

Amy Iverson ◽

Christopher J. Meyer ◽

Peter Vogel ◽

Samanthi Waidyarachchi ◽

Nisha Das ◽

...

Keyword(s):

Single Dose ◽

Streptococcus Pneumoniae ◽

Respiratory Tract ◽

Otitis Media ◽

Respiratory Infections ◽

Acute Otitis Media ◽

Upper Respiratory Tract ◽

Content Type ◽

Upper Respiratory

ABSTRACT The most frequent ailment for which antibiotics are prescribed is otitis media (ear infections), which is most commonly caused by Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae. Treatment of otitis media is complicated by the fact that the bacteria in the middle ear typically form biofilms, which can be recalcitrant to antibiotic treatment. Furthermore, bacterial respiratory infections can be greatly exacerbated by viral coinfection, which is particularly evidenced by the synergy between influenza and S. pneumoniae. In this study, we sought to ascertain the in vivo efficacy of aminomethyl spectinomycin lead 1950, an effective antibacterial agent both in vitro and in vivo against Streptococcus pneumoniae in the context of complex respiratory infections and acute otitis media. A single dose of 1950 significantly reduced bacterial burden in the respiratory tract for all three pathogens, even when species were present in a coinfection model. Additionally, a single dose of 1950 effectively reduced pneumococcal acute otitis media from the middle ear. The agent 1950 also proved efficacious in the context of influenza-pneumococcal super infection. These data further support the development of this family of compounds as potential therapeutic agents against the common causes of complex upper respiratory tract infections and acute otitis media.

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TheblpLocus of Streptococcus pneumoniae Plays a Limited Role in the Selection of Strains That Can Cocolonize the Human Nasopharynx

Applied and Environmental Microbiology ◽

10.1128/aem.01048-16 ◽

2016 ◽

Vol 82 (17) ◽

pp. 5206-5215 ◽

Cited By ~ 5

Author(s):

Carina Valente ◽

Suzanne Dawid ◽

Francisco R. Pinto ◽

Jason Hinds ◽

Alexandra S. Simões ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Gene Transfer ◽

Horizontal Gene Transfer ◽

Experimental Models ◽

Nasopharyngeal Colonization ◽

Content Type ◽

Limited Role ◽

Multiple Strains ◽

The Impact

ABSTRACTNasopharyngeal colonization is important forStreptococcus pneumoniaeevolution, providing the opportunity for horizontal gene transfer when multiple strains co-occur. Although colonization with more than one strain of pneumococcus is common, the factors that influence the ability of strains to coexist are not known. A highly variableblp(bacteriocin-like peptide) locus has been identified in all sequenced strains ofS. pneumoniae. This locus controls the regulation and secretion of bacteriocins, small peptides that target other bacteria. In this study, we analyzed a series of cocolonizing isolates to evaluate the impact of theblplocus on human colonization to determine whether competitive phenotypes of bacteriocin secretion restrict cocolonization. We identified a collection of 135 nasopharyngeal samples cocolonized with two or more strains, totaling 285 isolates. Theblplocus of all strains was characterized genetically with regard to pheromone type, bacteriocin/immunity content, and potential for locus functionality. Inhibitory phenotypes of bacteriocin secretion and locus activity were assessed through overlay assays. Isolates from single colonizations (n= 298) were characterized for comparison. Cocolonizing strains had a high diversity ofblpcassettes; approximately one-third displayed an inhibitory phenotypein vitro. Despitein vitroevidence of competition, pneumococci cocolonized the subjects independently ofblppheromone type (P= 0.577), bacteriocin/immunity content,blplocus activity (P= 0.798), and inhibitory phenotype (P= 0.716). In addition, no significant differences were observed when single and cocolonizing strains were compared. Despite clear evidence ofblp-mediated competition in experimental models, the results of our study suggest that theblplocus plays a limited role in restricting pneumococcal cocolonization in humans.IMPORTANCENasopharyngeal colonization withStreptococcus pneumoniae(pneumococcus) is important for pneumococcal evolution, as the nasopharynx represents the major site for horizontal gene transfer when multiple strains co-occur, a phenomenon known as cocolonization. Understanding how pneumococcal strains interact within the competitive environment of the nasopharynx is of chief importance in the context of pneumococcal ecology. In this study, we used an unbiased collection of naturally co-occurring pneumococcal strains and showed that a biological process frequently used by bacteria for competition—bacteriocin production—is not decisive in the coexistence of pneumococci in the host, in contrast to what has been shown in experimental models.

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Induction of Efflux-Mediated Macrolide Resistance in Streptococcus pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00060-11 ◽

2011 ◽

Vol 55 (7) ◽

pp. 3413-3422 ◽

Cited By ~ 16

Author(s):

Scott T. Chancey ◽

Xiaoliu Zhou ◽

Dorothea Zähner ◽

David S. Stephens

Keyword(s):

Streptococcus Pneumoniae ◽

Fold Increase ◽

Amino Sugar ◽

Transferase Activity ◽

Neutral Sugar ◽

Content Type ◽

Cellular Targets ◽

Gram Positive Bacteria ◽

Exit Tunnel ◽

Erythromycin A

ABSTRACTThe antimicrobial efflux system encoded by the operonmef(E)-melon the mobile genetic element MEGA inStreptococcus pneumoniaeand other Gram-positive bacteria is inducible by macrolide antibiotics and antimicrobial peptides. Induction may affect the clinical response to the use of macrolides. We developedmef(E)reporter constructs and a disk diffusion induction and resistance assay to determine the kinetics and basis ofmef(E)-melinduction. Induction occurred rapidly, with a >15-fold increase in transcription within 1 h of exposure to subinhibitory concentrations of erythromycin. A spectrum of environmental conditions, including competence and nonmacrolide antibiotics with distinct cellular targets, did not inducemef(E).Using 16 different structurally defined macrolides, induction was correlated with the amino sugar attached to C-5 of the macrolide lactone ring, not with the size (e.g., 14-, 15- or 16-member) of the ring or with the presence of the neutral sugar cladinose at C-3. Macrolides with a monosaccharide attached to C-5, known to block exit of the nascent peptide from the ribosome after the incorporation of up to eight amino acids, inducedmef(E)expression. Macrolides with a C-5 disaccharide, which extends the macrolide into the ribosomal exit tunnel, disrupting peptidyl transferase activity, did not induce it. The induction ofmef(E)did not require macrolide efflux, but the affinity of macrolides for the ribosome determined the availability for efflux and pneumococcal susceptibility. The induction ofmef(E)-melexpression by inducing macrolides appears to be based on specific interactions of the macrolide C-5 saccharide with the ribosome that alleviate transcriptional attenuation ofmef(E)-mel.

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