scholarly journals RNase R, a new virulence determinant of Streptococcus pneumoniae

2021 ◽  
Author(s):  
Cátia Bárria ◽  
Dalila Mil-Homens ◽  
Arsénio M. Fialho ◽  
Cecília Maria Arraiano ◽  
Susana Domingues
Keyword(s):  
Molecular Mechanisms ◽  
Bacterial Resistance ◽  
Galleria Mellonella ◽  
Bacterial Load ◽  
Infection Model ◽  
Virulence Determinants ◽  
Pneumococcal Infections ◽  
Rnase R ◽  
Host Interactions ◽  
Eclipse Phase

ABSTRACTPneumococcal infections have increasingly high mortality rates despite the availability of vaccines and antibiotics. The increase of bacterial resistance to antibiotics urges the discovery of new alternative therapeutics. Therefore, the identification of new virulence determinants, and the understanding of the molecular mechanisms behind pathogenesis and pneumococcal-host interactions has become of paramount importance in the search of new targets for drug development. The exoribonuclease RNase R has been involved in virulence in a growing number of pathogens. In this work, we have used Galleria mellonella as an infection model to demonstrate that the presence of ribonuclease R increases the pneumococcus virulence. Although the absence of RNase R does not affect exponential growth, the ability of the RNase R deleted strain to replicate in the hemolymph is compromised. Larvae infected with the RNase R mutant strain show an increased expression level of antimicrobial peptides, and have a lower bacterial load in the haemolymph in the later stages of infection, leading to a higher survival rate. Interestingly RNase R carrying pneumococci suffer a sudden drop in bacterial numbers immediately after infection, resembling the eclipse phase observed after intravenous inoculation in mice. Together our results suggest that RNase R might be involved in the ability of pneumococci to evade the host immune response, probably by interfering with internalisation and/or replication inside the larval hemocytes.

Characterization of Streptococcus pneumoniae phage-like element SpnCI reveals an enhanced virulent phenotype in the acute invertebrate infection model Galleria mellonella

2019 ◽  
Author(s):  
Kimberly McCullor ◽  
Maliha Rahman ◽  
Catherine King ◽  
W. Michael McShan
Keyword(s):  
Streptococcus Pneumoniae ◽  
Galleria Mellonella ◽  
Diagnostic Tools ◽  
Infection Model ◽  
Pneumococcal Infections ◽  
Health Score ◽  
Survival Percentage ◽  
Pneumococcal Strain ◽  
Streptococcal Species ◽  
Virulent Phenotype

AbstractPhage-like elements are found in a multitude of streptococcal species, including pneumococcal strain Hungary19A-6 (SpnCI). The aim of our research was to investigate the role of phage-like element SpnCI in enhanced virulence and phenotypic modulation within Streptococcus pneumoniae. SpnCI was found to significantly enhance virulence within the invertebrate infection model Galleria mellonella. Infections with SpnCI led to a lower mean health score (1.6) and survival percentage (20%) compared to SpnCI null TIGR4 infections (3.85 mean health score and 50% survival). SpnCI remained integrated throughout growth, conferring greater sensitivity to UV irradiation. Change in transcriptional patterns occurred, including downregulation of operons involved with cell surface modelling in the SpnCI containing strain of TIGR4. Kanamycin-tagged SpnCI strain in Hungary19A-6 was inducible and isolated from lysate along with both annotated prophages. No phages were identified by PCR nor electron microscopy (EM) following induction of TIGR4 SpnCIΔstrA suggesting helper-phage dependence for dissemination. EM of lysate showed typical siphoviridae morphology with an average capsid size of 60 nm. Two of sixty capsids were found to be smaller, suggesting SpnCI disseminates using a similar mechanism described for Staphylococcus aureus phage-like element SaPI. SpnCI from lysate infected capsule null strain T4R but was incapable of infecting the encapsulated TIGR4 strain suggesting that capsule impedes phage infection. Our work demonstrates that SpnCI can modulate virulence, UV susceptibility, alter transcriptional patterns, and furthermore, can disseminate via infection within pneumococcus. Further research is necessary to elucidate how SpnCI modulates virulence and what genes are responsible for the enhanced virulence phenotype.ImportanceAlthough vaccines have limited the scope of pneumococcal infections, Streptococcus pneumoniae still remains an important human pathogen. Understanding novel elements, such as SpnCI, that enhance virulence can lead to the development of more targeted therapeutic and diagnostic tools within the clinical realm.

scholarly journals Paving the Way to Unveil the Diversity and Evolution of Phage Genomes

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 905
Author(s):  
Alejandro Reyes ◽  
Martha J. Vives
Keyword(s):  
Molecular Evolution ◽  
Host Range ◽  
Molecular Mechanisms ◽  
Bacterial Resistance ◽  
Viral Pathogenesis ◽  
Resistance Mechanisms ◽  
Molecular Tools ◽  
Special Issue ◽  
Bacterial Host ◽  
Host Interactions

Phage biology has been developing for the last hundred years, and the potential of phages as tools and treatments has been known since their early discovery. However, the lack of knowledge of the molecular mechanisms coded in phage genomes hindered the development of the field. With current molecular methods, the last decade has been a resurgence of the field. The Special Issue on “Diversity and Evolution of Phage Genomes” is a great example with its 17 manuscripts published. It covers some of the latest methods to sample and characterize environmental and host associated viromes, considering experimental biases and computational developments. Furthermore, the use of molecular tools coupled with traditional methods has allowed to isolate and characterize viruses from different hosts and environments with such diversity that even a new viral class is being proposed. The viruses described cover all different phage families and lifestyles. However, is not only about diversity; the molecular evolution is studied in a set of manuscripts looking at phage-host interactions and their capacity to uncover the frequency and type of mutations behind the bacterial resistance mechanisms and viral pathogenesis, and such methods are opening new ways into identifying potential receptors and characterizing the bacterial host range.

scholarly journals Identifying virulence determinants of multidrug-resistant Klebsiella pneumoniae in Galleria mellonella

2021 ◽  
Author(s):  
Sebastian Bruchmann ◽  
Theresa Feltwell ◽  
Julian Parkhill ◽  
Francesca L Short
Keyword(s):  
Klebsiella Pneumoniae ◽  
Large Scale ◽  
Galleria Mellonella ◽  
Fitness Landscape ◽  
Dominant Role ◽  
Bacterial Pathogens ◽  
Model Organism ◽  
Multidrug Resistant ◽  
Infection Model ◽  
Virulence Determinants

Abstract Infections caused by Klebsiella pneumoniae are a major public health threat. Extensively drug-resistant and even pan-resistant strains have been reported. Understanding K. pneumoniae pathogenesis is hampered by the fact that murine models of infection offer limited resolution for non-hypervirulent strains which cause the majority of infections. The insect Galleria mellonella larva is a widely used alternative model organism for bacterial pathogens. We have performed genome-scale fitness profiling of a multidrug-resistant K. pneumoniae ST258 strain during infection of G. mellonella, to determine if this model is suitable for large-scale virulence factor discovery in this pathogen. Our results demonstrated a dominant role for surface polysaccharides in infection, with contributions from siderophores, cell envelope proteins, purine biosynthesis genes and additional genes of unknown function. Comparison with a hypervirulent strain, ATCC 43816, revealed substantial overlap in important infection-related genes, as well as additional putative virulence factors specific to ST258, reflecting strain-dependent fitness effects. Our analysis also identified a role for the metalloregulatory protein NfeR (YqjI) in virulence. Overall, this study offers new insight into the infection fitness landscape of K. pneumoniae, and provides a framework for using the highly flexible and easily scalable G. mellonella infection model to dissect molecular virulence mechanisms of bacterial pathogens.

scholarly journals Identifying virulence determinants of multidrug-resistant Klebsiella pneumoniae in Galleria mellonella

2020 ◽  
Author(s):  
Sebastian Bruchmann ◽  
Theresa Feltwell ◽  
Julian Parkhill ◽  
Francesca L. Short
Keyword(s):  
Klebsiella Pneumoniae ◽  
Large Scale ◽  
Galleria Mellonella ◽  
Fitness Landscape ◽  
Cell Envelope ◽  
Dominant Role ◽  
Multidrug Resistant ◽  
Infection Model ◽  
Virulence Determinants ◽  
Genome Scale

AbstractInfections caused by Klebsiella pneumoniae are a major public health threat. Extensively drug-resistant and even pan-resistant strains have been reported. Understanding K. pneumoniae pathogenesis is hampered by the fact that murine models of infection offer limited resolution for the non-hypervirulent strains which cause the majority of infections. We have performed genome-scale fitness profiling of a multidrug-resistant K. pneumoniae ST258 strain during infection of the insect Galleria mellonella, with the aim to determine if this model is suitable for large-scale virulence factor discovery in this pathogen. Our results demonstrated a dominant role for surface polysaccharides in infection, with contributions from siderophores, cell envelope proteins, purine biosynthesis genes and additional genes of unknown function. Comparison with a hypervirulent strain, ATCC 43816, revealed substantial overlap in important infection-related genes, as well as additional putative virulence factors that may be specific to ST258. Our analysis also identified a role for the metalloregulatory protein NfeR (also called YqjI) in virulence. Overall, this study offers new insight into the infection fitness landscape of K. pneumoniae ST258, and provides a framework for using the highly flexible, scalable G. mellonella infection model to dissect the molecular virulence mechanisms of K. pneumoniae and other bacterial pathogens.

scholarly journals Mutations That Impact the Enteropathogenic Escherichia coli Cpx Envelope Stress Response Attenuate Virulence in Galleria mellonella

2012 ◽  
Vol 80 (9) ◽  
pp. 3077-3085 ◽  
Author(s):  
S. Leuko ◽  
T. L. Raivio
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Type Iii Secretion ◽  
Galleria Mellonella ◽  
Infection Model ◽  
Virulence Determinants ◽  
Content Type ◽  
Type Iii ◽  
Envelope Stress

ABSTRACTIn this paper, we show that the larvae of the greater wax moth,Galleria mellonella, can be used as a model to study enteropathogenicEscherichia coli(EPEC) virulence.G. mellonellalarvae are killed after infection with EPEC type strain E2348/69 but not by an attenuated derivative that expresses diminished levels of the major virulence determinants or by a mutant specifically defective in type III secretion (T3S). Infecting EPEC inhabit the larval hemocoel only briefly and then become localized to melanized capsules, where they remain extracellular. Previously, it was shown that mutations affecting the Cpx envelope stress response lead to diminished expression of the bundle-forming pilus (BFP) and the type III secretion system (T3SS). We demonstrate that mutations that activate the Cpx pathway have a dramatic effect on the ability of the bacterium to establish a lethal infection, and this is correlated with an inability to growin vivo. Infection with allE. colistrains led to increased expression of the antimicrobial peptides (AMPs) gloverin and cecropin, although strain- and AMP-specific differences were observed, suggesting that theG. mellonellahost perceives attenuated strains and Cpx mutants in unique manners. Overall, this study shows thatG. mellonellais an economical, alternative infection model for the preliminary study of EPEC host-pathogen interactions, and that induction of the Cpx envelope stress response leads to defects in virulence.

scholarly journals In VivoAssessment of Growth and Virulence Gene Expression during Commensal and Pathogenic Lifestyles ofluxABCDE-Tagged Enterococcus faecalis Strains in Murine Gastrointestinal and Intravenous Infection Models

2013 ◽  
Vol 79 (13) ◽  
pp. 3986-3997 ◽  
Author(s):  
Sabina Leanti La Rosa ◽  
Pat G. Casey ◽  
Colin Hill ◽  
Dzung B. Diep ◽  
Ingolf F. Nes ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Galleria Mellonella ◽  
Expression Profiles ◽  
Systemic Infection ◽  
Intestinal Lumen ◽  
Infection Model ◽  
Dependent Manner ◽  
Virulence Determinants ◽  
Specific Expression ◽  
Content Type

ABSTRACTCytolysin and gelatinase are prominent pathogenicity determinants associated with highly virulentEnterococcus faecalisstrains. In an effort to explore the expression profiles of these virulence traitsin vivo, we have employedE. faecalisvariants expressing theluxABCDEcassette under the control of either the P16S, cytolysin, or gelatinase promoter for infections ofGalleria mellonellacaterpillars and mice. Systemic infection ofG. mellonellawith bioluminescence-taggedE. faecalisMMH594 revealed temporal regulation of both gelatinase and cytolysin promoters and demonstrated that these traits were induced in response to the host environment. Gavage of mice pretreated perorally with antibiotics resulted in efficient colonization of the murine gastrointestinal tract (GIT) in a strain-dependent manner, where the commensal baby isolate EF62 was more persistent than the nosocomial isolate MMH594. A highly significant correlation (R2> 0.94) was found between bioluminescence and the CFU counts in mouse fecal samples. Both strains showed similar preferences for growth and persistence in the ileum, cecum, and colon. Cytolysin expression was uniform in these compartments of the intestinal lumen. In spite of high numbers (109CFU/g of intestinal matter) in the ileum, cecum, and colon, no evidence of translocation or systemic infection could be observed. In the murine intravenous infection model, cytolysin expression was readily detected in the liver, kidneys, and bladder. At 72 h postinfection, the highest bacterial loads were found in the liver, kidneys, and spleen, with organ-specific expression levels of cytolysin ∼400- and ∼900-fold higher in the spleen and heart, respectively, than in the liver and kidneys. Taken together, this system based on the bioluminescence imaging technology is established as a new, powerful method to monitor the differential regulation ofE. faecalisvirulence determinants and to study the spatiotemporal course of infection in living animals in real time.

scholarly journals The Emergence of Novel Sequence Type Strains Reveals an Evolutionary Process of Intraspecies Clone Shifting in ICU-Spreading Carbapenem-Resistant Klebsiella pneumoniae

2021 ◽  
Vol 12 ◽  
Author(s):  
Dongdong Zhao ◽  
Qiucheng Shi ◽  
Dandan Hu ◽  
Li Fang ◽  
Yihan Mao ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Galleria Mellonella ◽  
Sequence Similarity ◽  
Public Health Problem ◽  
Infection Model ◽  
Virulence Determinants ◽  
Clinical Environment ◽  
Typing Method ◽  
Phylogenetic Structure ◽  
Carbapenem Resistant

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is an urgent public health problem worldwide, and its rapid evolution in the clinical environment has been a major concern. A total of 99 CRKP isolates spreading in the intensive care unit (ICU) setting were included and subjected to whole-genome sequencing, and their sequence types (STs), serotype loci, and virulence determinants were screened based on genome data. The phylogenetic structure was reconstructed based on the core genome multilocus sequence typing method. Regions of recombination were assessed. Biofilm formation, serum resistance assays, and a Galleria mellonella infection model were used to evaluate strain virulence. A novel ST, designated ST4496, emerged in the ICU and spread for 6 months before its disappearance. ST4496 was closely related to ST11, with only a single-allele variant, and ST11 is the most dominant clinical clone in China. Recombination events occurred at capsule biosynthesis loci and divided the strains of ST11 and its derivative ST4496 into three clusters, including ST11-KL47, ST11-KL64, and ST4496-KL47. The phylogenetic structure indicated that ST11-KL47 was probably the origin of ST11-related strain evolution and presented more diversity in terms of both sequence similarity and phenotypes. ST4496-KL47 cluster strains presented less virulence than ST11-KL64, which was probably one of the factors preventing the former from spreading widely. In conclusion, ST4496-KL47 was probably derived from ST11-KL47 via intraspecies shifting but was less competitive than ST11-KL64, which also evolved from ST11-KL47 and developed increased virulence via capsule biosynthesis locus recombination. ST11-KL64 has the potential to be the predominant CRKP clone in China.

scholarly journals Bacterial Genetic Approach to the Study of Reactive Oxygen Species Production in Galleria mellonella During Salmonella Infection

2021 ◽  
Vol 11 ◽  
Author(s):  
Hanna D. Bismuth ◽  
Gaël Brasseur ◽  
Benjamin Ezraty ◽  
Laurent Aussel
Keyword(s):  
Reactive Oxygen Species ◽  
Molecular Mechanisms ◽  
Galleria Mellonella ◽  
Bacterial Load ◽  
Reactive Oxygen ◽  
Ros Production ◽  
Oxygen Species ◽  
Host Pathogen Interactions ◽  
Wide Range ◽  
Host Pathogen

Over the last decade, an increasing number of reports presented Galleria mellonella larvae as an important model to study host-pathogen interactions. Coherently, increasing information became available about molecular mechanisms used by this host to cope with microbial infections but few of them dealt with oxidative stress. In this work, we addressed the role of reactive oxygen species (ROS) produced by the immune system of G. mellonella to resist against Salmonella enterica, an intracellular pathogen responsible for a wide range of infections. We confirmed that Salmonella was pathogen for G. mellonella and showed that it had to reach a minimal bacterial load within the hemolymph to kill the larvae. ROS production by G. mellonella was revealed by the virulence defects of Salmonella mutants lacking catalases/peroxiredoxins or cytoplasmic superoxide dismutases, both strains being highly sensitive to these oxidants. Finally, we used bacterial transcriptional fusions to demonstrate that hydrogen peroxide (H2O2) was produced in the hemolymph of Galleria during infection and sensed by S. enterica. In line with this observation, the H2O2-dependent regulator OxyR was found to be required for bacterial virulence in the larvae. These results led us to conclude that ROS production is an important mechanism used by G. mellonella to counteract bacterial infections and validate this host as a relevant model to study host-pathogen interactions.

scholarly journals Antibacterial Mechanisms and Efficacy of Sarecycline in Animal Models of Infection and Inflammation

Antibiotics ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 439
Author(s):  
Christopher G. Bunick ◽  
Jonette Keri ◽  
S. Ken Tanaka ◽  
Nika Furey ◽  
Giovanni Damiani ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
Bacterial Resistance ◽  
Antibiotic Use ◽  
In Vivo Studies ◽  
Infection Model ◽  
Severe Acne ◽  
Rat Paw Edema ◽  
Infection And Inflammation

Prolonged broad-spectrum antibiotic use is more likely to induce bacterial resistance and dysbiosis of skin and gut microflora. First and second-generation tetracycline-class antibiotics have similar broad-spectrum antibacterial activity. Targeted tetracycline-class antibiotics are needed to limit antimicrobial resistance and improve patient outcomes. Sarecycline is a narrow-spectrum, third-generation tetracycline-class antibiotic Food and Drug Administration (FDA)-approved for treating moderate-to-severe acne. In vitro studies demonstrated activity against clinically relevant Gram-positive bacteria but reduced activity against Gram-negative bacteria. Recent studies have provided insight into how the structure of sarecycline, with a unique C7 moiety, interacts with bacterial ribosomes to block translation and prevent antibiotic resistance. Sarecycline reduces Staphylococcus aureus DNA and protein synthesis with limited effects on RNA, lipid, and bacterial wall synthesis. In agreement with in vitro data, sarecycline demonstrated narrower-spectrum in vivo activity in murine models of infection, exhibiting activity against S. aureus, but reduced efficacy against Escherichia coli compared to doxycycline and minocycline. In a murine neutropenic thigh wound infection model, sarecycline was as effective as doxycycline against S. aureus. The anti-inflammatory activity of sarecycline was comparable to doxycycline and minocycline in a rat paw edema model. Here, we review the antibacterial mechanisms of sarecycline and report results of in vivo studies of infection and inflammation.

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