Microscopy-based phenotypic profiling of infection by Staphylococcus aureus clinical isolates reveals intracellular lifestyle as a prevalent feature

Lopes, Ines Rodrigues; Alcantara, Laura Maria; Silva, Ricardo J.; Josse, Jérôme; Vega, Elena Pedrero; Cabrerizo, Ana; Bonhomme, Mélanie; López, Daniel; Laurent, Frédéric; Vandenesch, François; Mano, Miguel; Eulálio, Ana

doi:10.1038/s41467-022-34790-9

Cited by 21 publications

(15 citation statements)

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“…About 40% of patients have persistent bacteremia at 24 h of effective therapy [ 3 ]. The predominant intracellular location of the pathogen, virulence markers and dysregulated host response, enables persistence, with its cell free DNA detectable in the blood even up to two weeks beyond the traditional cultures [ 12 , 13 ]. Available evidence also shows reported persistent bacteremia among gram negative organisms ranging from 7.2 to 38.5% [ 7 , 9 , 10 , 14 , 15 ].…”

Section: Discussionmentioning

confidence: 99%

Persistent bacteremia predicts poor outcomes among neutropenic patients with carbapenem-resistant gram-negative bloodstream infections receiving appropriate therapy

Kumar

George

Bhanuprasad

et al. 2023

Ann Clin Microbiol Antimicrob

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Purpose Identifying persistent bacteremia early in patients with neutropenia may improve outcome. This study evaluated the role of follow-up blood cultures (FUBC) positivity in predicting outcomes among patients with neutropenia and carbapenem-resistant gram-negative bloodstream infections (CRGNBSI). Methods This retrospective cohort study conducted between December 2017 and April 2022 included patients more than 15 years old with neutropenia and CRGNBSI, who survived for ≥ 48 h, receiving appropriate antibiotic therapy and had FUBCs. Patients with polymicrobial bacteremia within 30 days were excluded. The primary outcome was 30 day mortality. Persistent bacteremia, septic shock, recovery from neutropenia, prolonged or profound neutropenia, requirement of intensive care and dialysis, and initiation of appropriate empirical therapy were also studied. Results In our study cohort of 155 patients, the 30 day mortality rate was 47.7%. Persistent bacteremia was common in our patient cohort (43.8%). Carbapenem resistant isolates identified in the study were K.pneumoniae (80%), E.coli (12.26%), P.aeruginosa (5.16%), A.baumanii (1.94%) and E.cloacae (0.65%). The median time for sending a FUBC was 2 days (IQR, 1–3 days). Patients with persistent bacteremia had higher mortality than those without (56.76% versus 32.1%; p < 0.001). Appropriate initial empirical therapy was given to 70.9%. Recovery from neutropenia occurred in 57.4% while 25.8% had prolonged or profound neutropenia. Sixty-nine percent (107/155) had septic shock and needed intensive care; 12.2% of patients required dialysis. Non-recovery from neutropenia (aHR, 4.28; 95% CI 2.53–7.23), presence of septic shock (aHR, 4.42; 95%CI 1.47–13.28), requirement of intensive care (aHR,3.12;95%CI 1.23–7.93), and persistent bacteremia (aHR,1.74; 95%CI 1.05–2.89) significantly predicted poor outcomes in multivariable analysis. Conclusion FUBC showing persistent bacteremia predicted poor outcomes among neutropenic patients with carbapenem-resistant gram-negative bloodstream infections (CRGNBSI) and should be routinely reported.

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Section: Discussionmentioning

confidence: 99%

Persistent bacteremia predicts poor outcomes among neutropenic patients with carbapenem-resistant gram-negative bloodstream infections receiving appropriate therapy

Kumar

George

Bhanuprasad

et al. 2023

Ann Clin Microbiol Antimicrob

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“…Despite these adaptive changes, intracellular bacteria still exhibit metabolic activity. Moreover, it has been shown that the intracellular phenotype could manifest greater tolerance to antibiotics (38). The reservoir of intracellular S. aureus is strongly linked to the recurrence of infection (39)(40)(41).…”

Section: Discussionmentioning

confidence: 99%

Harnessing light-activated gallium porphyrins to combat intracellularStaphylococcus aureusin dermatitis: Insights from a simplified model

Szymczak,

Rychłowski,

Zhang

et al. 2023

Preprint

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Staphylococcus aureuscan survive inside nonprofessional phagocytes such as keratinocytes, demonstrating a novel strategy for evading antibiotic pressure. When antibiotic treatment ends, reinfection with staphylococci begins from the intracellular inoculum. This phenomenon is responsible for recurrent infections. The development of new antibacterial methods that can eliminate intracellular bacteria, including those with a multidrug-resistant phenotype, is necessary. In this study, we characterized and used a model of keratinocytes (both wild type and mutants with reduced filaggrin expression) infected with methicillin-resistantS. aureus(MRSA) to verify the possibility of using light-activated compounds, exemplified here by heme-mimetic gallium (III) porphyrin (Ga3+CHP) and visible light, an approach known as antimicrobial photodynamic inactivation (aPDI), to eliminate intracellular MRSA. We observed that Ga3+CHP accumulated more in infected cells than in uninfected cells. Moreover, Ga3+CHP accumulated in cells that harbored intracellularS. aureus. Using flow cytometry and fluorescence microscopy, we found that intracellular MRSA and Ga3+CHP mainly colocalized in lysosomal structures, and we showed that under the influence of aPDI, MRSA exhibited reduced adhesion to host cells and a significantly reduced (by 70%) GFP signal originating from intracellular bacteria. Moreover, the use of light-activated Ga3+CHP resulted in a significant reduction in the number of extracellular bacteria in the infection system, lowering the potential for further infection of host cells. For the first time, we used the infectious model to analyze the toxicity of aPDI in real time, showing that this approach is not significantly cyto-or phototoxic.Author SummaryStaphylococcus aureusis a highly virulent pathogen that is responsible for approximately 80% of all skin infections. During antibiotic treatment, one of the defense mechanisms ofS. aureusis the invasion of keratinocytes. Intracellular bacteria are not accessible to antibiotics, which poorly penetrate the interior of host cells. Consequently, such bacteria contribute to recurrent infections. In our study, we proposed using a combination of a light-activated porphyrin compound loaded with gallium ions, Ga3+CHP, and visible light as a strategy to eliminate intracellular staphylococci. We demonstrated that the tested compound colocalized with the pathogen in the infected cells, which was an essential condition for the effective elimination of intracellular bacteria. We showed that the proposed approach effectively reduced the infection of keratinocytes with methicillin-resistantS. aureus(MRSA), as well as its adhesion to host cells, while maintaining host cells. The results presented here provide a basis for developing an effective therapy against staphylococci.

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“…Of these, M. tuberculosis has infected a quarter of the world’s population and killed more people than any other microorganisms in humankind’s history . Initially classified as an extracellular pathogen, S. aureus has undergone resistance adaptations, fueled by the continued misuse and overuse of antibiotics, allowing it to now persist in both professional and non-professional phagocytes, rendering current treatments to fail …”

Section: Experimental Considerations and Reporting Recommendations Fo...mentioning

confidence: 99%

Minimum Information for Conducting and Reporting In Vitro Intracellular Infection Assays

Subramaniam,

Joyce,

Ogunniyi

et al. 2024

ACS Infect. Dis.

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Bacterial pathogens are constantly evolving to outsmart the host immune system and antibiotics developed to eradicate them. One key strategy involves the ability of bacteria to survive and replicate within host cells, thereby causing intracellular infections. To address this unmet clinical need, researchers are adopting new approaches, such as the development of novel molecules that can penetrate host cells, thus exerting their antimicrobial activity intracellularly, or repurposing existing antibiotics using nanocarriers (i.e., nanoantibiotics) for site-specific delivery. However, inconsistency in information reported across published studies makes it challenging for scientific comparison and judgment of experiments for future direction by researchers. Together with the lack of reproducibility of experiments, these inconsistencies limit the translation of experimental results beyond pre-clinical evaluation. Minimum information guidelines have been instrumental in addressing such challenges in other fields of biomedical research. Guidelines and recommendations provided herein have been designed for researchers as essential parameters to be disclosed when publishing their methodology and results, divided into four main categories: (i) experimental design, (ii) establishing an in vitro model, (iii) assessment of efficacy of novel therapeutics, and (iv) statistical assessment. These guidelines have been designed with the intention to improve the reproducibility and rigor of future studies while enabling quantitative comparisons of published studies, ultimately facilitating translation of emerging antimicrobial technologies into clinically viable therapies that safely and effectively treat intracellular infections.

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Microscopy-based phenotypic profiling of infection by Staphylococcus aureus clinical isolates reveals intracellular lifestyle as a prevalent feature

Cited by 21 publications

References 85 publications

Persistent bacteremia predicts poor outcomes among neutropenic patients with carbapenem-resistant gram-negative bloodstream infections receiving appropriate therapy

Persistent bacteremia predicts poor outcomes among neutropenic patients with carbapenem-resistant gram-negative bloodstream infections receiving appropriate therapy

Harnessing light-activated gallium porphyrins to combat intracellularStaphylococcus aureusin dermatitis: Insights from a simplified model

Minimum Information for Conducting and Reporting In Vitro Intracellular Infection Assays

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