EuroPneumo Special Issue / pneumonia 2015 Oct 21;7:I–72

has been ectopically expressed in the pneumococcus and live fluorescence microscopy was realised to determine the

spatio-temporal localisation of LytA. The results showed that LytA shifts from a homogeneous cytoplasmic localisation

during the exponential growth phase to a heterogeneous cytoplasmic localisation when bacterial cell population enters

the stationary growth phase. Finally, we observed the extracellular presence of LytA at the septum site along the lytic

phase. Respective roles of the choline-binding and amidase domains of LytA were analysed in this relocalisation process.

We determined that both domains are required for efficient cell lysis and surface localisation of LytA, which is also

linked to the presence of CbpE that regulates the amount of cell wall-associated PCho. In stationary growth phase,

cell death occurs before lysis, which suggests that LytA could be released by membrane disruption. Moreover, LytA

specifically cleaves the peptidoglycan at the septal sites only when cells enter the stationary growth phase. Studying cell

physiology at the early stationary phase will allow deciphering the mechanisms underlying these processes. Preliminary

data suggest that the regulation of LytA-mediated cell lysis occurs through the substrate availability. Characterisation of

such peptidoglycan modification is currently under investigation.


DiiA is a newly identified cell wall protein of Streptococcus pneumoniae

involved in invasive disease

María de la Soledad Escolano Martínez 1, 2 , Arnau Domenech 2, 3 , José Yuste 1, 2 , María I. Cercenado 1 ,

Carmen Ardanuy 2, 3 , Josefina Liñares 2, 3 , Adela G. de la Campa 1, 2 , Antonio J. Martin-Galiano 1, 2


Instituto de Salud Carlos III, Centro Nacional de Microbiología, Majadahonda, Madrid, Spain; 2 CIBER de Enfermedades Respiratorias (CIBERES),

Madrid, Spain; 3 Servicio de Microbiología, Hospital Universitari de Bellvitge, Universitat de Barcelona, Barcelona, Spain

Many outer multi-domain proteins play fundamental roles in the virulence of bacterial pathogens in an allele-dependent

manner. The hypothetical protein SP1992 of Streptococcus pneumoniae TIGR4 contains (from N- to C-terminus): two

imperfect repeats (R1 and R2), an unstructured region, and a cell-wall anchor domain. In this study we have investigated

560 clinical isolates, demonstrating that the gene coding for DiiA is part of the core genome and may carry either one

(R2) or two (R1R2) repeats. Clonal complexes carrying R1R2 were associated with invasive disease, while those carrying

R2 preferentially affected patients with underlying risk factors. Isogenic strains carrying step-wise deletions of diiA were

constructed to investigate the contribution of the different modules to the infective process. The mutants constructed

were: diiA-R2 that lacks R1 and mimics the natural short allele; diiA-NR that lacks both repeats; and the ΔdiiA null mutant.

Our results show that R1 and R2 are involved in the interaction with lung epithelial cells, and in systemic dissemination

using a mice model of pneumonia infection. Moreover, the ΔdiiA defective strain was severely impaired in its ability

to proliferate in blood in a sepsis model. This strain was also less able to avoid complement-mediated immunity and

phagocytosis. The latter phenotype may be mediated by the binding of the unstructured region to lactoferrin, for which

the DiiA-NR protein had a high affinity for, with a K D

of 2.5 nM. Based on our results, we termed SP1992 DiiA, after

Dimorphic invasion-involved A protein.


Induction of meningitis by a non-passaged clinical pneumococcal

isolate in an infant rat model depends on the presence of capsule

Lucy Hathaway, Denis Grandgirard, Belinda Ries, Suzanne Aebi, Luca Valente, Stephen Leib

Institute for Infectious Diseases, University of Bern, Bern, Switzerland

This study aimed to use an established infant rat model of experimental pneumococcal meningitis to study the potential

of a non-passaged clinical pneumococcal isolate to induce disease. A Swiss clinical pneumococcal isolate (106.66) of

serotype 6B and its non-encapsulated mutant (106.66 Janus) were administered intracisternally in an established infant

rat model without prior passage of the bacteria in animals. Bacterial titres were determined in CSF samples to optimise

the model for study of a non-passaged strain and to determine the appropriate time to administer ceftriaxone antibiotic

therapy. Clinical parameters were recorded and CSF sampled to determine bacterial titre and cytokine concentrations.

Brain damage was quantified by histomorphological analysis. A non-passaged clinical pneumococcal isolate was able

pneumonia 2015 Volume 7


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