Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae.

PLoS One. 2015 Mar 23;10(3):e0119823. doi: 10.1371/journal.pone.0119823. eCollection 2015.

Rational Manipulation of mRNA Folding Free Energy Allows Rheostat Control of Pneumolysin Production by Streptococcus pneumoniae.

Amaral FE1, Parker D2, Randis TM2, Kulkarni R2, Prince AS3, Shirasu-Hiza MM4, Ratner AJ2.

Author information

Abstract

The contribution of specific factors to bacterial virulence is generally investigated through creation of genetic "knockouts" that are then compared to wild-type strains or complemented mutants. This paradigm is useful to understand the effect of presence vs. absence of a specific gene product but cannot account for concentration-dependent effects, such as may occur with some bacterial toxins. In order to assess threshold and dose-response effects of virulence factors, robust systems for tunable expression are required. Recent evidence suggests that the folding free energy (ΔG) of the 5' end of mRNA transcripts can have a significant effect on translation efficiency and overall protein abundance. Here we demonstrate that rational alteration of 5' mRNA folding free energy by introduction of synonymous mutations allows for predictable changes in pneumolysin (PLY) expression by Streptococcus pneumoniae without the need for chemical inducers or heterologous promoters. We created a panel of isogenic S. pneumoniae strains, differing only in synonymous (silent) mutations at the 5' end of the PLY mRNA that are predicted to alter ΔG. Such manipulation allows rheostat-like control of PLY production and alters the cytotoxicity of whole S. pneumoniae on primary and immortalized human cells. These studies provide proof-of-principle for further investigation of mRNA ΔG manipulation as a tool in studies of bacterial pathogenesis.

PMID: 25798590

Penicillin-Resistant trend of Streptococcus pneumoniae in Asia: A systematic review.

Iran J Microbiol. 2014 Aug;6(4):198-210.

Penicillin-Resistant trend of Streptococcus pneumoniae in Asia: A systematic review.

Mamishi S1, Moradkhani S2, Mahmoudi S2, Hosseinpour-Sadeghi R2, Pourakbari B2.

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Abstract

The high prevalence of resistance to penicillin by Streptococcus pneumoniaeis considered as a great concern, particularly in Asian countries. The aim of this study was to investigate the changing trend of penicillin-resistant S. pneumoniae (PRSP) in Asia over a 20 years period. A review of the literature was conducted using the PubMed database, Google Scholar, Scopus, two Persian scientific search engines "Scientific Information Database" (www.sid.ir), and "Mag Iran" (www.magiran.com) through 1993 to 2013. Our study provides a unique chance to investigate the changing trend in PSSP in Asia over a 20 years period. Susceptibility rates among different centers in each country varied widely. In Malaysia, the PSSP rate decreased from 97.2% in 1995-1996 to 69% in 2000. In Singapore, PSSP levels decreased from 72.6% in 1997 to 30.5% in 2007-2008. In Iran, PSSP ranged from 0% to 100%. In Taiwan, the rate of PSSP was 60.3% in 1995 and <50% in other years. In Lebanon, the rate of PSSP was less than 50% (ranging from 30.1% to 50%) in all published data. In Hong Kong, the level of penicillin susceptibility decreased from 71.1% during 1993-1995 to less 42% in 2007. Continuous surveillance of resistance data from clinical isolates as well as implementation of strict infection control policies is recommended. More studies are needed for better evaluation PSSP rate in some Asian countries such as Vietnam, Singapore, Philippines, Pakistan, Nepal, Kuwait, Korea and Indonesia.

KEYWORDS:

Asia; S. pneumoniae; penicillin resistant

PMID: 25802701 [PubMed] 

Streptococcus pneumoniae Induces Autophagy through the Inhibition of the PI3K-I/Akt/mTOR Pathway and ROS Hypergeneration in A549 Cells.

PLoS One. 2015 Mar 24;10(3):e0122753. doi: 10.1371/journal.pone.0122753. eCollection 2015.

Streptococcus pneumoniae Induces Autophagy through the Inhibition of the PI3K-I/Akt/mTOR Pathway and ROS Hypergeneration in A549 Cells.

Li P1, Shi J2, He Q1, Hu Q1, Wang YY1, Zhang LJ1, Chan WT3, Chen WX1.

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Abstract

The present study focused on the action mechanism of S. pneumoniae (Sp) in inducing autophagy in human alveolar epithelial cells. Sp, a gram-positive extracellular bacterium, activates autophagy with considerably increased microtuble-associated protein light chain 3 (LC3) punctation in A549 cells. The accumulation of typical autophagosomes and conjugation of LC3 to phosphatidylethanolamine were observed in Sp-infected cells as an indication of autophagy. Using the pneumolysin (PLY) mutant, we successfully demonstrated that PLY is involved in initiating autophagy without affecting the expression levels of PI3K-III and Beclin1. PLY-mediated autophagy depends on the inhibition of the phosphoinositide 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway. Furthermore, Sp could also lead to the reactive oxygen species (ROS) hypergeneration in A549 cells. Taken together, Sp infection-induced autophagy is PLY-mediated through ROS hypergeneration and mTOR inhibition. PI3K-I and rapamycin (autophagy inducers) enhanced bacterial clearance, whereas wortmannin (autophagy inhibitor) and acetylcysteine (ROS inhibitor) reduced intracellular bacteria clearance. Thus, Sp-induced autophagy represents a host-protective mechanism, providing new insight into the pathogenesis of respiratory tract Sp infection.

PMID: 25803050

The Effect of Macrolide-Resistance on the Presentation and Outcome of Patients Hospitalized for Streptococcus pneumoniae Pneumonia.

Am J Respir Crit Care Med. 2015 Mar 25. [Epub ahead of print]

The Effect of Macrolide-Resistance on the Presentation and Outcome of Patients Hospitalized for Streptococcus pneumoniae Pneumonia.

Cilloniz C1, Albert RK, Liapikou A, Gabarrus A, Rangel E, Bello S, Marco F, Mensa J, Torres A.

Author information

Abstract

BACKGROUND:

There are conflicting reports describing the effect of macrolide resistance on the presentation and outcomes of patients with Streptococcus pneumoniae pneumonia.

METHODS:

We conducted a retrospective, observational study in the Hospital Clinic of Barcelona of all adult patients hospitalized with pneumonia who had positive cultures for S. pneumoniae from January 1, 2000 to December 31, 2013. Outcomes examined included bacteremia, pulmonary complications, acute renal failure, shock, intensive care unit admission, need for mechanical ventilation, length of hospital stay and 30-day mortality.

RESULTS:

Of 643 patients hospitalized for S. pneumoniae pneumonia, 139 (22%) were macrolide-resistant. Patients with macrolide-resistant organisms were less likely to have bacteremia, pulmonary complications and shock, and were less likely to require non-invasive mechanical ventilation. We found no increase in the incidence of acute renal failure, the frequency of ICU admission, the need for invasive ventilatory support, the length hospital stay or the 30-day mortality in patients with (invasive or non-invasive) macrolide-resistant S. pneumoniae pneumonia, and no effect on outcomes as a function of whether treatment regimens did or did not comply with current guidelines.

CONCLUSIONS:

We found no evidence suggesting that patients hospitalized for macrolide-resistant S. pneumoniae pneumonia had worse clinical outcomes.

KEYWORDS:

S. pneumoniae resistant to macrolide; community-acquired pneumonia; pneumococcal pneumonia; pneumonia

PMID: 25807239

In vitro activity of bioactive extracts from rare actinomycetes against multi-drug resistant Streptococcus pneumoniae.

J Appl Microbiol. 2015 Mar 21. doi: 10.1111/jam.12810. [Epub ahead of print]

In vitro activity of bioactive extracts from rare actinomycetes against multi-drug resistant Streptococcus pneumoniae.

Tiwari K1, Raj VS, Upadhyay DJ, Gupta RK.

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Abstract

AIMS:

In this study, we investigated the in vitro potential of the bioactive extracts from five putatively novel species of actinomycetes isolated from the Indian hot desert against multi-drug resistant (MDR) Streptococcus pneumoniae.

METHODS AND RESULTS:

The antimicrobial activity of 10 different extracts was evaluated against S. pneumoniae strains with, erm(B) and mef(E) genes as well as fluoroquinolone-resistant (FQR ) strains using the micro-broth dilution method. Of these 10 extracts, four exhibited good to excellent anti-S. pneumoniae activity with minimum inhibitory concentrations (MICs) ranging from 0·125 to 8 μg ml-1 . The time-kill kinetics study showed that these extracts killed the pathogens in 2-8 h. In vitro cell-free transcription/translation of luciferase gene using S30 bacterial extract and TNT mammalian ribosome indicated that they inhibited bacterial ribosomes at much lower concentrations than those required to inhibit the mammalian ribosomes.

CONCLUSIONS:

This study demonstrates that these are potent concentration-dependent bactericidal metabolites with 16-fold higher in vitro activity than levofloxacin against MDR S. pneumoniae.

SIGNIFICANCE AND IMPACT OF THE STUDY:

Metabolites from actinomycetes can be excellent inhibitors of MDR S. pneumoniae. Considering the in vitro efficacy of these crude extracts against S. pneumoniae MDR spp., once purified these can be used against streptococcal pathogens causing community-acquired pneumonia.

© 2015 The Society for Applied Microbiology.

KEYWORDS:

S. pneumoniae ; antimicrobial susceptibility; bioactive extracts; multi-drug resistance; rare actinomycetes; time-kill kinetics

PMID: 25810118

Neonatal Streptococcus pneumoniae Infection May Aggravate Adulthood Allergic Airways Disease in Association with IL-17A.

PLoS One. 2015 Mar 27;10(3):e0123010. doi: 10.1371/journal.pone.0123010. eCollection 2015.

Neonatal Streptococcus pneumoniae Infection May Aggravate Adulthood Allergic Airways Disease in Association with IL-17A.

Yang B1, Liu R1, Yang T2, Jiang X1, Zhang L1, Wang L1, Wang Q3, Luo Z4, Liu E4, Fu Z4.

Author information

Abstract

Epidemiologic studies have demonstrated that some bacteria colonization or infections in early-life increased the risk for subsequent asthma development. However, little is known about the mechanisms by which early-life bacterial infection increases this risk. The aim of this study was to investigate the effect of neonatal Streptococcus pneumoniae infection on the development of adulthood asthma, and to explore the possible mechanism. A non-lethal S. pneumoniae lung infection was established by intranasal inoculation of neonatal (1-week-old) female mice with D39. Mice were sensitized and challenged with ovalbumin in adulthood to induce allergic airways disease (AAD). Twenty-four hours later, the lungs and bronchoalveolar lavage fluid (BALF) were collected to assess AAD. Neonatal S. pneumoniae infection exacerbated adulthood hallmark features of AAD, with enhanced airway hyperresponsiveness and increased neutrophil recruitment into the airways, increased Th17 cells and interleukin (IL)-17A productions. Depletion of IL-17A by i.p. injection of a neutralizing monoclonal antibody reduced neutrophil recruitment into the airways, alleviated airway inflammation and decreased airway hyperresponsiveness. Furthermore, IL-17A depletion partially restored levels of inteferon-γ, but had no effect on the release of IL-5 or IL-13. Our data suggest that neonatal S. pneumoniae infection may promote the development of adulthood asthma in association with increased IL-17A production.

PMID: 25816135

Current methods for capsular typing of Streptococcus pneumoniae.

J Microbiol Methods. 2015 Mar 25;113:41-49. doi: 10.1016/j.mimet.2015.03.006. [Epub ahead of print]

Current methods for capsular typing of Streptococcus pneumoniae.

Jauneikaite E1, Tocheva AS2, Jefferies JM3, Gladstone RA2, Faust SN4, Christodoulides M2, Hibberd ML5, Clarke SC6.

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Abstract

Streptococcus pneumoniae is a major respiratory tract pathogen causing pneumococcal disease mainly in children aged less than five years and in the elderly. Ninety-eight different capsular types (serotypes) of pneumococci have been reported, but pneumococcal conjugate vaccines (PCV) include polysaccharide antigens against only 7, 10 or 13 serotypes. It is therefore important to track the emergence of serotypes due to the clonal expansion of non-vaccine serotypes. Increased numbers of carried and disease-causing pneumococci are now being analysed as part of the post-PCV implementation surveillance studies and hence rapid, accurate and cost-effective typing methods are important. Here we describe serotyping methods published prior to 10th November 2014 for pneumococcal capsule typing. Sixteen methods were identified; six were based on serological tests using immunological properties of the capsular epitopes, eight were semi-automated molecular tests, and one describes the identification of capsular type directly from whole genome data, which also allows for further intra and inter-genome analyses. There was no single method that could be recommended for all pneumococcal capsular typing applications. Although the Quellung reaction is still considered to be the gold-standard, laboratories should take into account the number of pneumococcal isolates and the type of samples to be used for testing, the time frame for the results and the resources available in order to select the most appropriate method. Most likely, a combination of phenotypic and genotypic methods would be optimal to monitor and evaluate the impact of pneumococcal conjugate vaccines and to provide information for future vaccine formulations.

Copyright © 2015 Elsevier B.V. All rights reserved.

KEYWORDS:

Capsular polysaccharide; Serotyping; Streptococcus pneumoniae

PMID: 25819558

Profiling of β-Lactam Selectivity for Penicillin-Binding Proteins in Streptococcus pneumoniae D39.

Profiling of β-Lactam Selectivity for Penicillin-Binding Proteins in Streptococcus pneumoniae D39.

Kocaoglu O1, Tsui HC2, Winkler ME3, Carlson EE4.

Author information

Abstract

Selective fluorescent β-lactam chemical probes enable the visualization of the transpeptidase activity of penicillin-binding proteins (PBPs) at different stages of bacterial cell division. To facilitate the development of new fluorescent probes for PBP imaging, we evaluated 20 commercially available β-lactams for selective PBP inhibition in an unencapsulated derivative of the D39 strain of Streptococcus pneumoniae. Live cells were treated with β-lactam antibiotics at different concentrations and subsequently incubated with BOCILLIN FL (Boc-FL) to saturate uninhibited PBPs. Fluorophore-labeled PBPs were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and fluorescence scanning. Among 20 compounds tested, carbapenems (doripenem and meropenem) were co-selective for PBP1a, PBP2x, and PBP3, while six of the nine penicillin compounds were co-selective for PBP2x and PBP3. In contrast, the seven cephalosporin compounds tested display variability in their PBP-binding profiles. Three cephalosporin compounds cefoxitin, cephalexin and cefsulodin and the monobactam, aztreonam, exhibited selectivity for PBP3, while only cefuroxime (a cephalosporin) was selective for PBP2x. Treatment of S. pneumoniae cultures with a sublethal concentration of cefuroxime that inhibited 60% of PBP2x activity and less than 20% of the activity of other PBPs, resulted in formation of elongated cells. In contrast, treatment of S. pneumoniae cultures with concentrations of aztreonam and cefoxitin that inhibited up to 70% of PBP3 activity and less than 30% of other PBPs resulted in no discernible morphological changes. Additionally, correlation of the MIC and IC50 values for each PBP, with the exception of faropenem, mecillinam and 6-APA, suggests that pneumococcal growth inhibition is primarily due to the inhibition of PBP2x.

Copyright © 2015, American Society for Microbiology. All Rights Reserved.

PMID: 25845878

Commensal streptococci serve as a reservoir for beta-lactam resistance genes in Streptococcus pneumoniae.

Antimicrob Agents Chemother. 2015 Apr 6. pii: AAC.00429-15. [Epub ahead of print]

Commensal streptococci serve as a reservoir for beta-lactam resistance genes in Streptococcus pneumoniae.

Jensen A1, Valdórsson O2, Frimodt-Møller N3, Hollingshead S4, Kilian M2.

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Abstract

Streptococcus pneumoniae is a leading cause of pneumonia, meningitis, septicemia, and middle ear infections. The incidence of S. pneumoniae isolates non-susceptible to antibiotics has risen worldwide and may be above 20% in some countries. Beta-lactam antibiotic resistance in pneumococci is associated with significant sequence polymorphism in penicillin-binding proteins (PBPs). Commensal streptococci, especially S. mitis and S. oralis, have been identified as putative donors of mutated gene fragments. However, no studies have compared sequences of the involved pbp genes in large collections of commensal streptococci, with those of S. pneumoniae. We therefore investigated the sequence diversity at the nucleotide and amino acid levels of the transpeptidase region in 107, 96, and 88 strains of the three pbp genes, pbp2x, pbp2b, and pbp1a, respectively, of susceptible and non-susceptible strains of commensal streptococci to determine to what extent homologous recombination between commensal streptococci and S. pneumoniae play a role in the development of beta-lactam resistance in S. pneumoniae. In contrast to pneumococci, extensive sequence variation in the transpeptidase region of pbp2x, pbp2b, and pbp1a was observed in both susceptible and non-susceptible strains of commensal streptococci, conceivably reflecting the genetic diversity of the many evolutionary lineages of commensal streptococci combined with intra- and interspecies homologues recombination events. Our data support that resistance to beta-lactam antibiotics in pneumococci is due to sequences acquired from commensal Mitis group streptococci, especially S. mitis. However, several amino acid alterations previously linked to beta-lactam resistance in pneumococci appear to represent species signatures of the donor strain rather than being causal of resistance.

Copyright © 2015, American Society for Microbiology. All Rights Reserved.

PMID: 25845880 

Deletion analysis of Streptococcus pneumoniae late competence genes distinguishes virulence determinants that are dependent or independent of competence induction.

Mol Microbiol. 2015 Apr 2. doi: 10.1111/mmi.13016. [Epub ahead of print]

Deletion analysis of Streptococcus pneumoniae late competence genes distinguishes virulence determinants that are dependent or independent of competence induction.

Zhu L1, Lin J, Kuang Z, Vidal JE, Lau GW.

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Abstract

The competence regulon of Streptococcus pneumoniae (pneumococcus) is crucial for genetic transformation. During competence development, the alternative sigma factor ComX is activated, which in turn, initiates transcription of 80 "late" competence genes. Interestingly, only 16 late genes are essential for genetic transformation. We hypothesized that these late genes that are dispensable for competence are beneficial to pneumococcal fitness during infection. These late genes were systematically deleted, and the resulting mutants were examined for their fitness during mouse models of bacteremia and acute pneumonia. Among these, 14 late genes were important for fitness in mice. Significantly, deletion of some late genes attenuated pneumococcal fitness to the same level in both wild-type and ComX-null genetic backgrounds, suggesting that the constitutive baseline expression of these genes was important for bacterial fitness. In contrast, some mutants were attenuated only in the wild-type genetic background but not in the ComX-null background, suggesting that specific expression of these genes during competence state contributed to pneumococcal fitness. Increased virulence during competence state was partially caused by the induction of allolytic enzymes that enhanced pneumolysin release. These results distinguish the role of basal expression versus competence induction in virulence functions encoded by ComX-regulated late competence genes.

This article is protected by copyright. All rights reserved.

KEYWORDS:

Streptococcus pneumoniae; acute pneumonia; bacteremia; competence regulon; virulence factors

PMID: 25846124 

Post-operative Streptococcus pneumoniae meningoencephalitis complicating surgery for acromegaly in an identical twin.

J Clin Neurosci. 2015 Apr 7. pii: S0967-5868(15)00068-5. doi: 10.1016/j.jocn.2015.01.011. [Epub ahead of print]

Post-operative Streptococcus pneumoniae meningoencephalitis complicating surgery for acromegaly in an identical twin.

Cote DJ1, Iuliano SL1, Smith TR1, Laws ER Jr2.

Author information

Abstract

This case report provides provocative and useful data regarding two aspects of acromegaly and its management. The patient, who is one of a pair of identical twins, has no known hereditary, genetic or otherwise potentially etiologic factors as compared to her unaffected sister. Secondly, transsphenoidal surgery, which was ultimately successful, was complicated by pneumococcal meningitis, an unusual event with only four previously reported patients, three of whom ended in death or major neurologic deficits. In this case, a 57-year-old woman gradually developed classical signs and symptoms of acromegaly while her identical twin sister remained normal with no evidence of endocrine disease. Endoscopic transsphenoidal surgery was complicated by the development of meningitis 25days after surgery. This was controlled following a difficult hospital course. Streptococcus pneumoniae meningoencephalitis is a rare but life-threatening complication of transsphenoidal surgery. A high index of suspicion for incipient meningitis should be maintained when patients present with severe headache and increased intracranial pressure, even if they initially lack the typical symptoms and signs. Immediate and aggressive treatment is necessary to avoid significant neurologic deficit.

Copyright © 2015 Elsevier Ltd. All rights reserved.

KEYWORDS:

Acromegaly; Meningitis; Pituitary adenoma; Transsphenoidal surgery

PMID: 25861890

Multiple mutations and increased RNA expression in tetracycline-resistant Streptococcus pneumoniae as determined by genome-wide DNA and mRNA sequencing.

J Antimicrob Chemother. 2015 Apr 9. pii: dkv060. [Epub ahead of print]

Multiple mutations and increased RNA expression in tetracycline-resistant Streptococcus pneumoniae as determined by genome-wide DNA and mRNA sequencing.

Lupien A1, Gingras H1, Bergeron MG1, Leprohon P1, Ouellette M2.

Author information

Abstract

OBJECTIVES:

The objective of this study was to characterize chromosomal mutations associated with resistance to tetracycline in Streptococcus pneumoniae.

METHODS:

Chronological appearance of mutations in two S. pneumoniae R6 mutants (R6M1TC-5 and R6M2TC-4) selected for resistance to tetracycline was determined by next-generation sequencing. A role for the mutations identified was confirmed by reconstructing resistance to tetracycline in a S. pneumoniae R6 WT background. RNA sequencing was performed on R6M1TC-5 and R6M2TC-4 and the relative expression of genes was reported according to R6. Differentially expressed genes were classified according to their ontology.

RESULTS:

WGS of R6M1TC-5 and R6M2TC-4 revealed mutations in the gene rpsJ coding for the ribosomal protein S10 and in the promoter region and coding sequences of the ABC genes patA and patB. These cells were cross-resistant to ciprofloxacin. Resistance reconstruction confirmed a role in resistance for the mutations in rpsJ and patA. Overexpression of the ABC transporter PatA/PatB or mutations in the coding sequence of patA contributed to resistance to tetracycline, ciprofloxacin and ethidium bromide, and was associated with a decreased accumulation of [3H]tetracycline. Comparative transcriptome profiling of the resistant mutants further revealed that, in addition to the overexpression of patA and patB, several genes of the thiamine biosynthesis and salvage pathway were increased in the two mutants, but also in clinical isolates resistant to tetracycline. This overexpression most likely contributes to the tetracycline resistance phenotype.

CONCLUSIONS:

The combination of genomic and transcriptomic analysis coupled to functional studies has allowed the discovery of novel tetracycline resistance mutations in S. pneumoniae.

© The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

KEYWORDS:

ABC transporters; RNA-seq; S. pneumoniae; genomic; rpsJ; thiamine

PMID: 25862682

Genotyping Streptococcus pneumoniae.

Future Microbiol. 2015 Apr;10:653-64. doi: 10.2217/fmb.14.153.

Genotyping Streptococcus pneumoniae.

Rayner RE1, Savill J, Hafner LM, Huygens F.

Author information

Abstract

ABSTRACT  Streptococcus pneumoniae is a potentially deadly human pathogen associated with high morbidity, mortality and global economic burden. The universally used bacterial genotyping methods are multilocus sequence typing and pulsed field gel electrophoresis. However, another highly discriminatory, rapid and less expensive genotyping technique, multilocus variable number of tandem repeat analysis (MLVA), has been developed. Unfortunately, no universal MLVA protocol exists, and some MLVA protocols do not amplify certain loci for all pneumococcal serotypes, leaving genotyping profiles incomplete. A number of other genotyping or characterization methods have been developed and will be discussed. This review examines the various protocols for genotyping S. pneumoniae and highlights the current direction technology and research is heading to understand this bacterium.

KEYWORDS:

MLST; MLVA; Streptococcus pneumoniae; epidemiology; genotyping

PMID: 25865199

A new quorum sensing system (TprA/PhrA) for Streptococcus pneumoniae D39 that regulates a lantibiotic biosynthesis gene cluster.

Mol Microbiol. 2015 Apr 13. doi: 10.1111/mmi.13029. [Epub ahead of print]

A new quorum sensing system (TprA/PhrA) for Streptococcus pneumoniae D39 that regulates a lantibiotic biosynthesis gene cluster.

Hoover SE1, Perez AJ, Tsui HC, Sinha D, Smiley DL, DiMarchi RD, Winkler ME, Lazazzera BA.

Author information

Abstract

The Phr peptides of Bacillus species mediate quorum sensing, but their identification and function in other species of bacteria has not been determined. We have identified a Phr peptide quorum sensing system (TprA/PhrA) that controls the expression of a lantibiotic gene cluster in the Gram-positive human pathogen, Streptococcus pneumoniae. Lantibiotics are highly modified peptides that are part of the bacteriocin family of antimicrobial peptides. We have characterized the basic mechanism for a Phr peptide signaling system in S. pneumoniae and found that it induces expression of the lantibiotic genes when pneumococcal cells are at high density in the presence of galactose, a main sugar of the human nasopharynx, a highly competitive microbial environment. Activity of the Phr peptide system is not seen when pneumococcal cells are grown with glucose, the preferred carbon source and the most prevalent sugar encountered by S. pneumoniae during invasive disease. Thus, the lantibiotic genes are expressed under the control of both cell density signals via the Phr peptide system and nutritional signals from the carbon source present, suggesting that quorum sensing and the lantibiotic machinery may help pneumococcal cells compete for space and resources during colonization of the nasopharynx.

This article is protected by copyright. All rights reserved.

KEYWORDS:

Lantibiotics; Phr peptides; Pneumococcus; Quorum sensing

PMID: 25869931

Conserved Streptococcus pneumoniae Spirosomes Suggest a Single Type of Transformation Pilus in Competence.

PLoS Pathog. 2015 Apr 15;11(4):e1004835. doi: 10.1371/journal.ppat.1004835. eCollection 2015.

Conserved Streptococcus pneumoniae Spirosomes Suggest a Single Type of Transformation Pilus in Competence.

Laurenceau R1, Krasteva PV1, Diallo A1, Ouarti S1, Duchateau M2, Malosse C2, Chamot-Rooke J2, Fronzes R1.

Author information

Abstract

The success of S. pneumoniae as a major human pathogen is largely due to its remarkable genomic plasticity, allowing efficient escape from antimicrobials action and host immune response. Natural transformation, or the active uptake and chromosomal integration of exogenous DNA during the transitory differentiated state competence, is the main mechanism for horizontal gene transfer and genomic makeover in pneumococci. Although transforming DNA has been proposed to be captured by Type 4 pili (T4P) in Gram-negative bacteria, and a competence-inducible comG operon encoding proteins homologous to T4P-biogenesis components is present in transformable Gram-positive bacteria, a prevailing hypothesis has been that S. pneumoniae assembles only short pseudopili to destabilize the cell wall for DNA entry. We recently identified a micrometer-sized T4P-like pilus on competent pneumococci, which likely serves as initial DNA receptor. A subsequent study, however, visualized a different structure - short, 'plaited' polymers - released in the medium of competent S. pneumoniae. Biochemical observation of concurrent pilin secretion led the authors to propose that the 'plaited' structures correspond to transformation pili acting as peptidoglycan drills that leave DNA entry pores upon secretion. Here we show that the 'plaited' filaments are not related to natural transformation as they are released by non-competent pneumococci, as well as by cells with disrupted pilus biogenesis components. Combining electron microscopy visualization with structural, biochemical and proteomic analyses, we further identify the 'plaited' polymers as spirosomes: macromolecular assemblies of the fermentative acetaldehyde-alcohol dehydrogenase enzyme AdhE that is well conserved in a broad range of Gram-positive and Gram-negative bacteria.

PMID: 25876066 [PubMed - in process] 

Resistance Determinants and Their Association with Different Transposons in the Antibiotic-Resistant Streptococcus pneumoniae.

Biomed Res Int. 2015;2015:836496. doi: 10.1155/2015/836496. Epub 2015 Mar 26.

Resistance Determinants and Their Association with Different Transposons in the Antibiotic-Resistant Streptococcus pneumoniae.

Korona-Glowniak I1, Siwiec R1, Malm A1.

Author information

Abstract

Multiple resistance of Streptococcus pneumoniae is generally associated with their unique recombination-mediated genetic plasticity and possessing the mobile genetic elements. The aim of our study was to detect antibiotic resistance determinants and conjugative transposons in 138 antibiotic-resistant pneumococcal strains isolated from nasopharynx of healthy young children from Lublin, Poland. These strains resistant to tetracycline and/or to chloramphenicol/erythromycin/clindamycin were tested by PCR using the specific genes as markers. The presence of Tn916 family transposons, carrying tet(M) and int/xisTn916, was observed in all of the tested strains. Tn916 was detected in 16 strains resistant only to tetracycline. Tn6002 and Tn3872-related element were found among 99 erm(B)-carrying strains (83.8% and 3.0%, resp.). Eight strains harbouring mef(E) and erm(B) genes were detected, suggesting the presence of Tn2010 and Tn2017 transposons. Among 101 chloramphenicol-resistant strains, two variants of Tn5252-related transposon were distinguished depending on the presence of int/xis5252 genes specific for cat gene-containing Tn5252 (75.2% of strains) or int Sp23FST81 gene, specific for cat-containing ICESp23FST81 element (24.8% of strains). In 6 strains Tn916-like and Tn5252-like elements formed a Tn5253-like structure. Besides clonal dissemination of resistant strains of pneumococci in the population, horizontal transfer of conjugative transposons is an important factor of the high prevalence of antibiotic resistance.

PMID: 25883976 [PubMed - in process] 

Expression of the Streptococcus pneumoniae yoeB Chromosomal toxin gene causes Cell Death in the model plant Arabidopsis thaliana.

BMC Biotechnol. 2015 Apr 12;15(1):26. [Epub ahead of print]

Expression of the Streptococcus pneumoniae yoeB Chromosomal toxin gene causes Cell Death in the model plant Arabidopsis thaliana.

Bakar FA1, Yeo CC2, Harikrishna JA3.

Author information

Abstract

BACKGROUND:

Bacterial toxin-antitoxin systems usually comprise of a pair of genes encoding a stable toxin and its cognate labile antitoxin and are located in the chromosome or in plasmids of several bacterial species. Chromosomally-encoded toxin-antitoxin systems are involved in bacterial stress responses and activation of the toxins usually leads to cell death or dormancy. Overexpression of the chromosomally-encoded YoeB toxin from the yefM-yoeB toxin-antitoxin locus of the Gram-positive bacterium Streptococcus pneumoniae has been shown to cause cell death in S. pneumoniae as well as E. coli.

RESULTS:

Induction of a YoeB-GFP fusion protein using a 17-β-estradiol-inducible plant expression system in Arabidopsis thaliana Col 0, was lethal in all T2 progeny. Examination of plants by fluorescent confocal microscopy showed GFP fluorescence in all parts of the leaves at 24 hours after 17-β-estradiol induction, continuing up to plant death. Quantitative RT-PCR analysis revealed that the expression of the yoeB toxin gene peaked at 3 days after induction with 17-β-estradiol, coinciding with the onset of visible effects on the plants. Moreover, we detected DNA laddering in the transgenic plants at 24 hours after yoeB induction, indicative of apoptosis.

CONCLUSIONS:

Expression of the YoeB toxin from Streptococcus pneumoniae is lethal in Arabidopsis. We believe this is the first report of a toxin from a bacterial toxin-antitoxin system functioning in plants. The results presented here mark an important milestone towards the development of a cell ablation based bio-containment strategy, which may be useful for functional studies and for the control of spread of transgenic plants.

PMID: 25887501 

Detection of antibody responses against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis proteins in children with community-acquired pneumonia: effects of combining pneumococcal antigens, pre-existing antibody levels, sampling interval, age, and duration of illness.

Eur J Clin Microbiol Infect Dis. 2015 Apr 19. [Epub ahead of print]

Detection of antibody responses against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis proteins in children with community-acquired pneumonia: effects of combining pneumococcal antigens, pre-existing antibody levels, sampling interval, age, and duration of illness.

Borges IC1, Andrade DC, Vilas-Boas AL, Fontoura MS, Laitinen H, Ekström N, Adrian PV, Meinke A, Cardoso MR, Barral A, Ruuskanen O, Käyhty H, Nascimento-Carvalho CM.

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Abstract

We evaluated the effects of combining different numbers of pneumococcal antigens, pre-existing antibody levels, sampling interval, age, and duration of illness on the detection of IgG responses against eight Streptococcus pneumoniae proteins, three Haemophilus influenzae proteins, and five Moraxella catarrhalis proteins in 690 children aged <5 years with pneumonia. Serological tests were performed on acute and convalescent serum samples with a multiplexed bead-based immunoassay. The median sampling interval was 19 days, the median age was 26.7 months, and the median duration of illness was 5 days. The rate of antibody responses was 15.4 % for at least one pneumococcal antigen, 5.8 % for H. influenzae, and 2.3 % for M. catarrhalis. The rate of antibody responses against each pneumococcal antigen varied from 3.5 to 7.1 %. By multivariate analysis, pre-existing antibody levels showed a negative association with the detection of antibody responses against pneumococcal and H. influenzae antigens; the sampling interval was positively associated with the detection of antibody responses against pneumococcal and H. influenzae antigens. A sampling interval of 3 weeks was the optimal cut-off for the detection of antibody responses against pneumococcal and H. influenzae proteins. Duration of illness was negatively associated with antibody responses against PspA. Age did not influence antibody responses against the investigated antigens. In conclusion, serological assays using combinations of different pneumococcal proteins detect a higher rate of antibody responses against S. pneumoniae compared to assays using a single pneumococcal protein. Pre-existing antibody levels and sampling interval influence the detection of antibody responses against pneumococcal and H. influenzae proteins. These factors should be considered when determining pneumonia etiology by serological methods in children.

PMID: 25894988

Carbon catabolite repression by seryl phosphorylated HPr is essential to Streptococcus pneumoniae in carbohydrate rich environments.

Mol Microbiol. 2015 Apr 21. doi: 10.1111/mmi.13033. [Epub ahead of print]

Carbon catabolite repression by seryl phosphorylated HPr is essential to Streptococcus pneumoniae in carbohydrate rich environments.

Fleming E1, Lazinski DW, Camilli A.

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Abstract

Carbon catabolite repression (CCR) is a regulatory phenomenon implemented by bacteria to hierarchically organize carbohydrate utilization in order to achieve maximal growth. CCR is likely of great importance to Streptococcus pneumoniae because the human host sites inhabited by this pathogen represent complex carbohydrate environments. In this species, inactivation of the prototypical Gram positive CCR master regulator, ccpA, attenuates virulence in mice but does not relieve CCR of most metabolic enzymes, suggesting CcpA-independent CCR mechanisms predominate. Here we show the activities of three transcriptional regulators constitute the majority of transcriptional CCR of galactose metabolism operons. We determined seryl-phosphorylated histidine phosphocarrier protein (HPr-Ser∼P)-mediated regulation is a major CCR mechanism and an essential activity in the pneumococcus, since an HPr point mutation abolishing HPrK/P-dependent phosphorylation was not tolerated nor was deletion of hprk/p. The HPr-Ser∼P phosphomimetic mutant HPr S46D had reduced PTS transport rates and limited induction of CCR-repressed genes. These results support a model of pneumococcal CCR in which HPr-Ser∼P directly affects the activity of CcpA while indirectly affecting the activity of pathway-specific transactional regulators. This report describes the first CcpA-independent CCR mechanism identified in the pneumococcus, and the first example of lethality from loss of HPr-Ser∼P-mediated CCR in any species.

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KEYWORDS:

CcpA-independent carbon catabolite repression; Streptococcus pneumoniae; histidine phosphocarrier protein

PMID: 25898857