scholarly journals Factors That Cause Trimethoprim Resistance in Streptococcus pyogenes

2014 ◽  
Vol 58 (4) ◽  
pp. 2281-2288 ◽  
Author(s):  
René Bergmann ◽  
Mark van der Linden ◽  
Gursharan S. Chhatwal ◽  
D. Patric Nitsche-Schmitz
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Nucleotide Sequence ◽  
Streptococcus Pyogenes ◽  
Dihydrofolate Reductase ◽  
Molecular Basis ◽  
Rapid Development ◽  
Antibiotic Agent ◽  
Content Type ◽  
Insight Into

ABSTRACTThe use of trimethoprim in treatment ofStreptococcus pyogenesinfections has long been discouraged because it has been widely believed that this pathogen is resistant to this antibiotic. To gain more insight into the extent and molecular basis of trimethoprim resistance inS. pyogenes, we tested isolates from India and Germany and sought the factors that conferred the resistance. Resistant isolates were identified in tests for trimethoprim or trimethoprim-sulfamethoxazole (SXT) susceptibility. Resistant isolates were screened for the known horizontally transferable trimethoprim-insensitive dihydrofolate reductase (dfr) genesdfrG,dfrF,dfrA,dfrD, anddfrK. The nucleotide sequence of the intrinsicdfrgene was determined for resistant isolates lacking the horizontally transferable genes. Based on tentative criteria, 69 out of 268 isolates (25.7%) from India were resistant to trimethoprim. Occurring in 42 of the 69 resistant isolates (60.9%),dfrFappeared more frequently thandfrG(23 isolates; 33.3%) in India. ThedfrFgene was also present in a collection of SXT-resistant isolates from Germany, in which it was the only detected trimethoprim resistance factor. ThedfrFgene caused resistance in 4 out of 5 trimethoprim-resistant isolates from the German collection. An amino acid substitution in the intrinsic dihydrofolate reductase known from trimethoprim-resistantStreptococcus pneumoniaeconferred resistance toS. pyogenesisolates ofemmtype 102.2, which lacked other aforementioneddfrgenes. Trimethoprim may be more useful in treatment ofS. pyogenesinfections than previously thought. However, the factors described herein may lead to the rapid development and spread of resistance ofS. pyogenesto this antibiotic agent.

scholarly journals Nucleotide Sequence Analysis of Integrative Conjugative Element Tn5253of Streptococcus pneumoniae

2013 ◽  
Vol 58 (2) ◽  
pp. 1235-1239 ◽  
Author(s):  
Francesco Iannelli ◽  
Francesco Santoro ◽  
Marco R. Oggioni ◽  
Gianni Pozzi
Keyword(s):  
Streptococcus Pneumoniae ◽  
Sequence Analysis ◽  
Nucleotide Sequence ◽  
Streptococcus Pyogenes ◽  
Open Reading Frames ◽  
Nucleotide Sequence Analysis ◽  
Conjugative Transposon ◽  
Content Type ◽  
Integrative Conjugative Element ◽  
Reading Frames

ABSTRACTConjugative transposon Tn5253, an integrative conjugative element (ICE) ofStreptococcus pneumoniaecarrying thecatandtet(M) genes, was shown to be 64,528 bp in size and to contain 79 open reading frames, of which only 38 could be annotated. Two distinct genetic elements were found integrated into Tn5253: Tn5251(18,033 bp), of the Tn916-Tn1545family of ICEs, and Ωcat(pC194) (7,627 bp), which could not conjugate but was capable of intracellular mobility by excision, circularization, and integration by homologous recombination. The highest conjugation frequency of Tn5253was observed whenStreptococcus pyogeneswas the donor (6.7 × 10−3transconjugants/donor).

scholarly journals Experimental Evolution In Vivo To Identify Selective Pressures during Pneumococcal Colonization

mSystems ◽  
2020 ◽  
Vol 5 (3) ◽  
Author(s):  
Vaughn S. Cooper ◽  
Erin Honsa ◽  
Hannah Rowe ◽  
Christopher Deitrick ◽  
Amy R. Iverson ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Murine Model ◽  
Experimental Evolution ◽  
Nasal Colonization ◽  
Content Type ◽  
Selective Pressures ◽  
Specific Tissue ◽  
Tissue Tropisms ◽  
Insight Into

ABSTRACT Experimental evolution is a powerful technique to understand how populations evolve from selective pressures imparted by the surrounding environment. With the advancement of whole-population genomic sequencing, it is possible to identify and track multiple contending genotypes associated with adaptations to specific selective pressures. This approach has been used repeatedly with model species in vitro, but only rarely in vivo. Herein we report results of replicate experimentally evolved populations of Streptococcus pneumoniae propagated by repeated murine nasal colonization with the aim of identifying gene products under strong selection as well as the population genetic dynamics of infection cycles. Frameshift mutations in one gene, dltB, responsible for incorporation of d-alanine into teichoic acids on the bacterial surface, evolved repeatedly and swept to high frequency. Targeted deletions of dltB produced a fitness advantage during initial nasal colonization coupled with a corresponding fitness disadvantage in the lungs during pulmonary infection. The underlying mechanism behind the fitness trade-off between these two niches was found to be enhanced adherence to respiratory cells balanced by increased sensitivity to host-derived antimicrobial peptides, a finding recapitulated in the murine model. Additional mutations that are predicted to affect trace metal transport, central metabolism, and regulation of biofilm production and competence were also selected. These data indicate that experimental evolution can be applied to murine models of pathogenesis to gain insight into organism-specific tissue tropisms. IMPORTANCE Evolution is a powerful force that can be experimentally harnessed to gain insight into how populations evolve in response to selective pressures. Herein we tested the applicability of experimental evolutionary approaches to gain insight into how the major human pathogen Streptococcus pneumoniae responds to repeated colonization events using a murine model. These studies revealed the population dynamics of repeated colonization events and demonstrated that in vivo experimental evolution resulted in highly reproducible trajectories that reflect the environmental niche encountered during nasal colonization. Mutations impacting the surface charge of the bacteria were repeatedly selected during colonization and provided a fitness benefit in this niche that was counterbalanced by a corresponding fitness defect during lung infection. These data indicate that experimental evolution can be applied to models of pathogenesis to gain insight into organism-specific tissue tropisms.

scholarly journals β-Aminopeptidases: Insight into Enzymes without a Known Natural Substrate

2019 ◽  
Vol 85 (15) ◽  
Author(s):  
Marietta John-White ◽  
James Gardiner ◽  
Priscilla Johanesen ◽  
Dena Lyras ◽  
Geoffrey Dumsday
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Gram Negative Bacteria ◽  
Diverse Range ◽  
Α Subunit ◽  
Gram Negative ◽  
Content Type ◽  
Proteolytic Resistance ◽  
Insight Into ◽  
Unique Capability

ABSTRACT β-Aminopeptidases have the unique capability to hydrolyze N-terminal β-amino acids, with varied preferences for the nature of β-amino acid side chains. This unique capability makes them useful as biocatalysts for synthesis of β-peptides and to kinetically resolve β-peptides and amides for the production of enantiopure β-amino acids. To date, six β-aminopeptidases have been discovered and functionally characterized, five from Gram-negative bacteria and one from a fungus, Aspergillus. Here we report on the purification and characterization of an additional four β-aminopeptidases, one from a Gram-positive bacterium, Mycolicibacterium smegmatis (BapAMs), one from a yeast, Yarrowia lipolytica (BapAYlip), and two from Gram-negative bacteria isolated from activated sludge identified as Burkholderia spp. (BapABcA5 and BapABcC1). The genes encoding β-aminopeptidases were cloned, expressed in Escherichia coli, and purified. The β-aminopeptidases were produced as inactive preproteins that underwent self-cleavage to form active enzymes comprised of two different subunits. The subunits, designated α and β, appeared to be tightly associated, as the active enzyme was recovered after immobilized-metal affinity chromatography (IMAC) purification, even though only the α-subunit was 6-histidine tagged. The enzymes were shown to hydrolyze chromogenic substrates with the N-terminal l-configurations β-homo-Gly (βhGly) and β3-homo-Leu (β3hLeu) with high activities. These enzymes displayed higher activity with H-βhGly-p-nitroanilide (H-βhGly-pNA) than previously characterized enzymes from other microorganisms. These data indicate that the new β-aminopeptidases are fully functional, adding to the toolbox of enzymes that could be used to produce β-peptides. Overexpression studies in Pseudomonas aeruginosa also showed that the β-aminopeptidases may play a role in some cellular functions. IMPORTANCE β-Aminopeptidases are unique enzymes found in a diverse range of microorganisms that can utilize synthetic β-peptides as a sole carbon source. Six β-aminopeptidases have been previously characterized with preferences for different β-amino acid substrates and have demonstrated the capability to catalyze not only the degradation of synthetic β-peptides but also the synthesis of short β-peptides. Identification of other β-aminopeptidases adds to this toolbox of enzymes with differing β-amino acid substrate preferences and kinetics. These enzymes have the potential to be utilized in the sustainable manufacture of β-amino acid derivatives and β-peptides for use in biomedical and biomaterial applications. This is important, because β-amino acids and β-peptides confer increased proteolytic resistance to bioactive compounds and form novel structures as well as structures similar to α-peptides. The discovery of new enzymes will also provide insight into the biological importance of these enzymes in nature.

scholarly journals WciGO-Acetyltransferase Functionality Differentiates Pneumococcal Serotypes 35C and 42

2017 ◽  
Vol 55 (9) ◽  
pp. 2775-2784 ◽  
Author(s):  
K. Aaron Geno ◽  
C. Allen Bush ◽  
Mengnan Wang ◽  
Cheng Jin ◽  
Moon H. Nahm ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Nucleotide Sequence ◽  
Chromosomal Region ◽  
Capsular Polysaccharides ◽  
Pneumococcal Serotypes ◽  
Subsequent Reaction ◽  
Content Type ◽  
Comprehensive View ◽  
Opsonophagocytic Killing ◽  
Dependent Mechanism

ABSTRACTStreptococcus pneumoniaeexpresses capsular polysaccharides (CPSs) to protect itself from opsonophagocytic killing. The genes responsible for capsules synthesized by the Wzy-dependent mechanism, which accounts for 96 of the 98 known pneumococcal capsule types, are in a chromosomal region known as thecpslocus. The nucleotide sequence in this region has been determined for all serotypes. In contrast, not all CPS structures have been defined. The structure of the serotype 35C polysaccharide was recently reported, but the presence ofO-acetyltransferase genes in the serotype 35Ccpslocus suggested that it could be incomplete, as the reported structure contains no O-acetylation. In addition, the genetic distinction of serotype 35C from the closely related serotype 42 was unclear, as their reportedcpsloci are nearly identical. To clarify these discrepancies, we obtained serotype 35C and 42 clinical and reference isolates and studied their serological and genetic properties, as well as the structures of CPSs purified from reference isolates. We demonstrated that theO-acetyltransferase WciG was functional in serotype 35C but nonfunctional in serotype 42 due to a deletion inwciG. Serotype 35C was O-acetylated at the 5- and 6-positions of 3-β-galactofuranose, as well as the 2-position of 6-β-galactofuranose. However, serotype 42 has only O-acetylation at 3-β-galactofuranose, an observation consistent with its loss of WciG functionality, which is associated with O-acetylation at the 2-position and subsequent reaction with typing antiserum 35a. These findings provide a comprehensive view of the genetic, biochemical structural, and serological bases of serotypes 35C and 42.

scholarly journals Mutations in the dihydrofolate reductase gene of trimethoprim-resistant isolates of Streptococcus pneumoniae.

1997 ◽  
Vol 41 (11) ◽  
pp. 2406-2413 ◽  
Author(s):  
P V Adrian ◽  
K P Klugman
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Correlation Coefficient ◽  
Dihydrofolate Reductase ◽  
Site Directed Mutagenesis ◽  
Clinical Isolates ◽  
Chromosomal Dna ◽  
Reductase Gene ◽  
Genes Encoding ◽  
Dihydrofolate Reductases

Streptococcus pneumoniae isolates resistant to several antimicrobial agent classes including trimethoprim-sulfamethoxazole have been reported with increasing frequency throughout the world. The MICs of trimethoprim, sulfamethoxazole, and trimethoprim-sulfamethoxazole (1:19) for 259 clinical isolates from South Africa were determined, and 166 of these 259 (64%) isolates were resistant to trimethoprim-sulfamethoxazole (MICs > or =20 mg/liter). Trimethoprim resistance was found to be more strongly correlated with trimethoprim-sulfamethoxazole resistance (correlation coefficient, 0.744) than was sulfamethoxazole resistance (correlation coefficient, 0.441). The dihydrofolate reductase genes from 11 trimethoprim-resistant (MICs, 64 to 512 microg/ml) clinical isolates of Streptococcus pneumoniae were amplified by PCR, and the nucleotide sequences were determined. Two main groups of mutations to the dihydrofolate reductase gene were found. Both groups shared six amino acid changes (Glu20-Asp, Pro70-Ser, Gln81-His, Asp92-Ala, Ile100-Leu, and Leu135-Phe). The first group included two extra changes (Lys60-Gln and Pro111-Ser), and the second group was characterized by six additional amino acid changes (Glu14-Asp, Ile74-Leu, Gln91-His, Glu94-Asp, Phe147-Ser, and Ala149-Thr). Chromosomal DNA from resistant isolates and cloned PCR products of the genes encoding resistant dihydrofolate reductases were capable of transforming a susceptible strain of S. pneumoniae to trimethoprim resistance. The inhibitor profiles of recombinant dihydrofolate reductase from resistant and susceptible isolates revealed that the dihydrofolate reductase from trimethoprim-resistant isolates was 50-fold more resistant (50% inhibitory doses [ID50s], 3.9 to 7.3 microM) than that from susceptible strains (ID50s, 0.15 microM). Site-directed mutagenesis experiments revealed that one mutation, Ile100-Leu, resulted in a 50-fold increase in the ID50 of trimethoprim. The resistant dihydrofolate reductases were characterized by highly conserved redundant changes in the nucleotide sequence, suggesting that the genes encoding resistant dihydrofolate reductases may have evolved as a result of inter- or intraspecies recombination by transformation.

scholarly journals Biofilm Formation Enhances Fomite Survival of Streptococcus pneumoniae and Streptococcus pyogenes

2013 ◽  
Vol 82 (3) ◽  
pp. 1141-1146 ◽  
Author(s):  
Laura R. Marks ◽  
Ryan M. Reddinger ◽  
Anders P. Hakansson
Keyword(s):  
Streptococcus Pneumoniae ◽  
Streptococcus Pyogenes ◽  
Biofilm Formation ◽  
Care Center ◽  
Content Type ◽  
Human Host ◽  
Abiotic Surfaces ◽  
Colonization Model ◽  
Biofilm Bacteria ◽  
The Impact

ABSTRACTBothStreptococcus pyogenesandStreptococcus pneumoniaeare widely thought to rapidly die outside the human host, losing infectivity following desiccation in the environment. However, to date, all literature investigating the infectivity of desiccated streptococci has used broth-grown, planktonic populations. In this study, we examined the impact of biofilm formation on environmental survival of clinical and laboratory isolates ofS. pyogenesandS. pneumoniaeas both organisms are thought to colonize the human host as biofilms. Results clearly demonstrate that while planktonic cells that are desiccated rapidly lose viability both on hands and abiotic surfaces, such as plastic, biofilm bacteria remain viable over extended periods of time outside the host and remain infectious in a murine colonization model. To explore the level and extent of streptococcal fomite contamination that children might be exposed to naturally, direct bacteriologic cultures of items in a day care center were conducted, which demonstrated high levels of viable streptococci of both species. These findings raise the possibility that streptococci may survive in the environment and be transferred from person to person via fomites contaminated with oropharyngeal secretions containing biofilm streptococci.

scholarly journals Molecular Basis for Different Levels oftet(M) Expression in Streptococcus pneumoniae Clinical Isolates

2012 ◽  
Vol 56 (10) ◽  
pp. 5040-5045 ◽  
Author(s):  
Patrick Grohs ◽  
Patrick Trieu-Cuot ◽  
Isabelle Podglajen ◽  
Sophie Grondin ◽  
Arnaud Firon ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Molecular Basis ◽  
Clinical Isolates ◽  
M Gene ◽  
Content Type ◽  
Frameshift Mutations ◽  
Resistant Mutants ◽  
Promoter Mutations ◽  
Different Levels

ABSTRACTSeventy-four unrelated clinical isolates ofStreptococcus pneumoniaeharboring thetet(M) gene were studied. Seven strains with low tetracycline (Tc) MICs (0.25 to 0.5 μg/ml) were found to harbor truncatedtet(M) alleles that were inactivated by different frameshift mutations. In contrast, five strains bore deletions in thetet(M) promoter region, among which four displayed increased Tc MICs (16 to 64 μg/ml). The same promoter mutations were detected in Tc-resistant mutants selectedin vitrofrom various susceptible strains. Sequence analysis revealed that these deletions might impede the formation of the transcriptional attenuator located immediately upstream oftet(M). Expression inEnterococcus faecalisof atet(M) reporter gene transcribed from these promoter mutants conferred a level of Tc resistance similar to that observed in the parentalS. pneumoniaestrains. These results show that different levels of Tc susceptibility found in clinical isolates ofS. pneumoniaecan be explained by frameshift mutations withintet(M) and by alterations of the upstream transcriptional attenuator.

scholarly journals Codon-Dependent Transcriptional Changes in Response to Tryptophan Limitation in the Tryptophan Auxotrophic Pathogens Chlamydia trachomatis and Streptococcus pyogenes

mSystems ◽  
2021 ◽  
Author(s):  
Scot P. Ouellette ◽  
Nathan D. Hatch ◽  
Nicholas A. Wood ◽  
Andrea L. Herrera ◽  
Michael S. Chaussee
Keyword(s):  
Amino Acid ◽  
Chlamydia Trachomatis ◽  
Streptococcus Pyogenes ◽  
Essential Amino Acid ◽  
Content Type ◽  
Host Environment ◽  
Transcriptional Changes

Chlamydia trachomatis and Streptococcus pyogenes are important pathogens of humans. Interestingly, both are auxotrophic for tryptophan and acquire this essential amino acid from the host environment.

scholarly journals Role of Copper Efflux in Pneumococcal Pathogenesis and Resistance to Macrophage-Mediated Immune Clearance

2015 ◽  
Vol 83 (4) ◽  
pp. 1684-1694 ◽  
Author(s):  
Michael D. L. Johnson ◽  
Thomas E. Kehl-Fie ◽  
Roger Klein ◽  
Jacqueline Kelly ◽  
Corinna Burnham ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Copper Stress ◽  
Mammalian Host ◽  
Bacterial Clearance ◽  
Content Type ◽  
Immune Clearance ◽  
Pulmonary Macrophages ◽  
Insight Into

In bacteria, the intracellular levels of metals are mediated by tightly controlled acquisition and efflux systems. This is particularly true of copper, a trace element that is universally toxic in excess. During infection, the toxic properties of copper are exploited by the mammalian host to facilitate bacterial clearance. To better understand the role of copper during infection, we characterized the contribution of thecopoperon to copper homeostasis and virulence inStreptococcus pneumoniae. Deletion of either the exporter, encoded bycopA, or the chaperone, encoded bycupA, led to hypersensitivity to copper stress. We further demonstrated that loss of the copper exporter encoded bycopAled to decreased virulence in pulmonary, intraperitoneal, and intravenous models of infection. Deletion ofcopAresulted in enhanced macrophage-mediated bacterial clearancein vitro. The attenuation phenotype of thecopAmutant in the lung was found to be dependent on pulmonary macrophages, underscoring the importance of copper efflux in evading immune defenses. Overall, these data provide insight into the role of thecopoperon in pneumococcal pathogenesis.

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