Human pleural fluid is a potent growth medium for Streptococcus pneumoniae.

Natalia D Popowicz,Ian D Kay,James C Paton,Christine F Carson,Y C Gary Lee,Jeremy S Brown,Grant W Waterer,Hui M Cheah,Sally M Lansley

doi:10.1371/journal.pone.0188833

Abstract

Empyema is defined by the presence of bacteria and/or pus in pleural effusions. However, the biology of bacteria within human pleural fluid has not been studied. Streptococcus pneumoniae is the most common cause of pediatric and frequent cause of adult empyema. We investigated whether S. pneumoniae can proliferate within human pleural fluid and if growth is affected by the cellular content of the fluid and/or characteristics of pneumococcal surface proteins. Invasive S. pneumoniae isolates (n = 24) and reference strain recovered from human blood or empyema were inoculated (1.5×106CFU/mL) into sterile human malignant pleural fluid samples (n = 11). All S. pneumoniae (n = 25) strains proliferated rapidly, increasing by a median of 3009 (IQR 1063–9846) from baseline at 24hrs in all pleural effusions tested. Proliferation was greater than in commercial pneumococcal culture media and concentrations were maintained for 48hrs without autolysis. A similar magnitude of proliferation was observed in pleural fluid before and after removal of its cellular content, p = 0.728. S. pneumoniae (D39 strain) wild-type, and derivatives (n = 12), each with mutation(s) in a different gene required for full virulence were inoculated into human pleural fluid (n = 8). S. pneumoniae with pneumococcal surface antigen A (ΔpsaA) mutation failed to grow (2207-fold lower than wild-type), p<0.001, however growth was restored with manganese supplementation. Growth of other common respiratory pathogens (n = 14) across pleural fluid samples (n = 7) was variable and inconsistent, with some strains failing to grow. We establish for the first time that pleural fluid is a potent growth medium for S. pneumoniae and proliferation is dependent on the PsaA surface protein and manganese.

Highlights

Pneumonia affects 450 million patients worldwide each year, [1] with Streptococcus pneumoniae the most common bacterial cause
Significant growth was observed when 3mL volumes of pleural fluid were inoculated with 1.5 × 106 CFU/mL S. pneumoniae (n = 3), with a median increase of 6875 (IQR 3494–15744) fold at 24 hours compared to baseline
S. pneumoniae is a common cause of both pediatric and adult empyema, yet the reasons why this pathogen can cause pleural infections have only been partially investigated. This is the first study to investigate the interactions of bacteria, namely S. pneumoniae, and human pleural fluid ex vivo

Summary

Introduction

Pneumonia affects 450 million patients worldwide each year, [1] with Streptococcus pneumoniae the most common bacterial cause. As many as 20–40% of patients with pneumonia develop a simple parapneumonic pleural effusion [2]. Pleural infection can develop when the pleural fluid is secondarily infected, and affects ~80,000 people in the UK and USA a year with significant morbidity and mortality [3]. Empyema represents the most severe end of the pleural infection spectrum. It is defined by the presence of bacteria, or pus, in the pleural fluid. S. pneumoniae accounts for the vast majority of pediatric empyema and is among the most frequent causative organisms of empyema in adults [4,5,6]. The incidence of pneumococcal empyema continues to rise globally [7,8,9]

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