Autor: Renshaw, A. - Prolib Integro

Skocz do pozycji: 1.

Tytuł:: Activated PI3K delta syndrome 1 mutations cause neutrophilia in zebrafish larvae
Autorzy:: Vogt, Katja
Renshaw, Stephen A.
Prajsnar, Tomasz
Elworthy, Stone
Rutherford, Holly A.
Hamilton, Noémie M.
Condliffe, Alison M.
Dostawca treści:: Repozytorium Uniwersytetu Jagiellońskiego

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Skocz do pozycji: 2.

Tytuł:: Experimental evolution of Staphylococcus aureus in macrophages : dissection of a conditional adaptive trait promoting intracellular survival
Autorzy:: Alves, Joana
Fitzgerald, J. Ross
Ring, Natalie
Renshaw, Stephen A.
Yebra, Gonzalo
Prajsnar, Tomasz
Vrieling, Manouk
Pickering, Amy
Opis:: Staphylococcus aureus is a major pathogen associated with important diseases in humans and animals. Macrophages are a key component of the innate immune response to S. aureus infection and play a major role in disease outcomes. To investigate the adaptive evolution of S. aureus in response to macrophages, we developed an experimental infection assay. S. aureus strains representing major human epidemic clones were passaged many times in a macrophage cell line, accumulating mutations in an array of genomic loci. Phenotypic analysis revealed the emergence of a lineage exhibiting increased survival in macrophages and human blood, and resistance to vancomycin. The evolved lineage exhibited a previously undescribed small colony variant (SCV) phenotype characterized by hyper-pigmentation, which resulted from a missense mutation in rsbW. Notably, the novel SCV was a conditional adaptive trait that was unstable in nutrient-replete conditions in vitro, rapidly converting from hyper-pigmented SCV to a non-pigmented large colony variant via spontaneous sigB deletion events. Importantly, we identified similar deletions in the genome sequences of a limited number of clinical S. aureus isolates from public databases, indicating that related events may occur during clinical infection. Experimental infection of zebrafish did not reveal a difference in virulence between parent and novel SCV but demonstrated an in vivo fitness cost for the compensatory sigB deletion events. Taken together, we report an experimental evolutionary approach for investigating bacterial innate immune cell interactions, revealing a conditional adaptation that promotes S. aureus survival in macrophages and resistance to vancomycin.
Dostawca treści:: Repozytorium Uniwersytetu Jagiellońskiego

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Skocz do pozycji: 3.

Tytuł:: Phagosomal acidification is required to kill Streptococcus pneumoniae in a zebrafish model
Autorzy:: Prajsnar, Tomasz
Michno, Bartosz
Renshaw, Stephen A.
Dockrell, David H.
Mitchell, Tim J.
Pooranachandran, Niedharsan
Fenton, Andrew K.
Opis:: Streptococcus pneumoniae (the pneumococcus) is a major human pathogen causing invasive disease, including community-acquired bacteraemia, and remains a leading cause of global mortality. Understanding the role of phagocytes in killing bacteria is still limited, especially in vivo. In this study, we established a zebrafish model to study the interaction between intravenously administered pneumococci and professional phagocytes such as macrophages and neutrophils, to unravel bacterial killing mechanisms employed by these immune cells. Our model confirmed the key role of polysaccharide capsule in promoting pneumococcal virulence through inhibition of phagocytosis. Conversely, we show pneumococci lacking a capsule are rapidly internalised by macrophages. Low doses of encapsulated S. pneumoniae cause near 100% mortality within 48 hours postinfection (hpi), while 50 times higher doses of unencapsulated pneumococci are easily cleared. Time course analysis of in vivo bacterial numbers reveals that while encapsulated pneumococcus proliferates to levels exceeding $10^{5}$ CFU at the time of host death, unencapsulated bacteria are unable to grow and are cleared within 20 hpi. Using genetically induced macrophage depletion, we confirmed an essential role for macrophages in bacterial clearance. Additionally, we show that upon phagocytosis by macrophages, phagosomes undergo rapid acidification. Genetic and chemical inhibition of vacuolar ATPase (v-ATPase) prevents intracellular bacterial killing and induces host death indicating a key role of phagosomal acidification in immunity to invading pneumococci. We also show that our model can be used to study the efficacy of antimicrobials against pneumococci in vivo. Collectively, our data confirm that larval zebrafish can be used to dissect killing mechanisms during pneumococcal infection in vivo and highlight key roles for phagosomal acidification in macrophages for pathogen clearance.
Dostawca treści:: Repozytorium Uniwersytetu Jagiellońskiego

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Skocz do pozycji: 4.

Tytuł:: A subset of neutrophil phagosomes is characterised by pulses of Class I PI3K activity
Autorzy:: Bernut, Audrey
Renshaw, Stephen A.
King, Jason S.
Cholewa-Waclaw, Justyna
Michno, Bartosz
Bowden, Kieran A.
Ellett, Felix
Prajsnar, Tomasz
Loynes, Catherine A.
Ho, Yin X.
Elworthy, Stone
Reyes-Aldasoro, Constantino Carlos
Muir, Clare F.
Prince, Lynne R.
Cadby, Ashley J.
Condliffe, Alison M.
Opis:: Class I PI3 kinases (PI3Ks) coordinate the delivery of microbicidal effectors to the phagosome by forming phosphatidylinositol (3,4,5)-trisphosphate (PIP3). However, the dynamics of PIP3 in neutrophils during a live bacterial tissue infection are unknown. We therefore developed an in vivo, live zebrafish infection model that enables visualisation of dynamic changes in Class 1 PI3K signalling in neutrophil phagosomes in real time. We identified that, on ∼12% of neutrophil phagosomes, PHAkt-eGFP, a reporter for Class 1 PI3K signalling, repeatedly fades and re-recruits in pulsatile bursts. This phenomenon occurred on phagosomes containing live and dead bacteria as well as beads, and was dependent on the activity of the Class 1 PI3K isoform PI3Kγ. Detailed imaging suggested that pulsing phagosomes represent neutrophils transiently re-opening and re-closing phagosomes, a conclusion supported by observations that a subset of phagosomes in human neutrophils rapidly accumulate dye from the extracellular space. Therefore, we propose that some neutrophil phagosomes remain unsealed and are consequently able to exchange contents with the extracellular environment, with implications for phagosome fate and communication with surrounding cells.
Dostawca treści:: Repozytorium Uniwersytetu Jagiellońskiego