Role of nonpneumoniae mycoplasma in the pathogenesis of ventilator-associated pneumonia: an in vitro assessment

Introduction

Mycoplasma organisms are the smallest bacteria capable of self-replication [1] and include species capable of causing disease (for example, Mycoplasma pneumoniae, Mycoplasma genitalium) as well as those that are generally thought to exist synergistically with their human host (for example, Mycoplasma salivarium). The Edinburgh critical care group (Prof TW/ACM) has recently identified a high prevalence of M. salivarium in the bronchoalveolar lavage washings from patients with confirmed and suspected ventilator-associated pneumonia (VAP) (Figure 1) [2]. The aim of this study was to examine the effect of M. salivarium on human immune cells in vitro. Specifically, we measured cytokine production and phagocytosis activity in response to M. salivarium exposure.

Figure 1

Combined mycoplasma detection by PCR in the prospective and clinical groups.

Methods

Whole human blood was obtained from healthy donor volunteers and cell types were isolated using diffusion gradients and magnetic labeling as appropriate. Monocytes and macrophages were incubated with M. salivarium for 24 hours before a subsequent LPS stimulus. Macrophage phagocytosis assays were conducted after exposure times of 60 minutes and 24 hours to M. salivarium. Cytokines were measured using ELISA and human cytokine bead array kits.

Results

There was a statistically significant decrease in phagocytosis between control cells and the macrophages exposed to both a low titer of M. salivarium (P value 0.018) and a medium titer of M. salivarium (P value 0.011) after 24 hours of exposure (Figure 2). There was a statistically significant decrease in phagocytosis activity between the macrophages exposed to the medium titer of M. salivarium for 24 hours versus 60 minutes (P value 0.013). Exposure of macrophages to mycoplasma resulted in decreased release of TNFα after a subsequent LPS stimulus (Figure 3). To our knowledge, this is the first time extracellular traps have been induced in macrophages in response to M. salivarium (Figure 4).

Figure 2

Twenty-four-hour exposure of macrophages to Mycoplasma salivarium .

Figure 3

Mycoplasma salivarium (30,000) for 24 hours prior to 100 ng/ml LPS exposure.

Figure 4

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Conclusion

Although further research is needed, it is interesting that the presence of M. salivarium caused an anti-inflammatory effect as well as impaired antigen presentation secondary to impaired phagocytosis. This could be consistent with the better outcome in mechanically ventilated patients that did not have M. salivarium bacteria detected in their bronchoalveolar lavage washings. Extracellular traps contribute to microbial containment by forming a physical barrier composed of chromatin and cytoplasmic proteins to enhance antimicrobial synergy while minimizing damage to host tissues [3]. It is interesting that M. salivarium induced extracellular traps.