Phenotypic variants of staphylococci and their underlying population distributions following exposure to stress - PubMed (original) (raw)

Phenotypic variants of staphylococci and their underlying population distributions following exposure to stress

Laura A Onyango et al. PLoS One. 2013.

Abstract

This study investigated whether alterations in environmental conditions would induce the formation of small colony variant phenotypes (SCV) with associated changes in cell morphology and ultra-structure in S. aureus, s. epidermidis, and S. lugdunensis. Wild-type clinical isolates were exposed to low temperature (4 °C), antibiotic stress (penicillin G and vancomycin; 0-10,000 µg mL(-1)), pH stress (pH 3-9) and osmotic challenge (NaCl concentrations of 0-20%). Changes in cell diameter, cell-wall thickness, and population distribution changes (n ≥ 300) were assessed via scanning and transmission electron microscopy (SEM and TEM), and compared to control populations. Our analyses found that prolonged exposure to all treatments resulted in the subsequent formation of SCV phenotypes. Observed SCVs manifested as minute colonies with reduced haemolysis and pigmentation (NaCl, pH and 4°C treatments), or complete lack thereof (antibiotic treatments). SEM comparison analyses revealed significantly smaller cell sizes for SCV populations except in S. aureus and S. epidermidis 10% NaCl, and S. epidermidis 4 °C (p<0.05). Shifts in population distribution patterns were also observed with distinct sub-populations of smaller cells appearing for S. epidermidis, and S. lugdunensis. TEM analyses revealed significantly thicker cell-walls in all treatments and species except S. lugdunensis exposed to 4 °C. These findings suggest that staphylococci adapted to environmental stresses by altering their cell size and wall thickness which could represent the formation of altered phenotypes which facilitate survival under harsh conditions. The phenotypic response was governed by the type of prevailing environmental stress regime leading to appropriate alterations in ultra-structure and size, suggesting downstream changes in gene expression, the proteome, and metabolome.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1

Figure 1. Population distributions of cell sizes from WT colony and corresponding SCV colony cells (4°C) of S. aureus, S. epidermidis and S. lugdunensis which have been ranked on the basis of their cell diameter measurements following SEM analyses.

Figure 2

Figure 2. Mean cell sizes assessed by SEM (n>300) of WT cells and corresponding SCV cells of S. aureus (a), S. epidermidis (b) and S. lugdunensis (c).

SCV cells were generated following exposure to the antibiotics penicillin G (Pen G) and vancomycin (VA) (random antibiotic concentrations generating SCV were used), 10% NaCl, 4°C temperature and pH 5 stresses. Asterisk (*) indicates significant differences compared with corresponding WT cells (P<0.05).

Figure 3

Figure 3. SEM images of S. aureus, S. epidermidis and S. lugdunensis WT and their vancomycin (VA) -induced (100 µg mL-1) SCVs: SEM images with SCV cells displaying a more prevalent extracellular matrix material (arrow) than their corresponding WT cells.

Figure 4

Figure 4. Comparisons of mean cell-wall thickness (nanometres) from SCV cells generated following exposures to 4°C and antibiotics (VA and Pen G; random concentrations utilised) in comparison with their corresponding WT cells taken from S. aureus, S. epidermidis, and S. lugdunensis samples and examined under TEM (n=300).

Asterisk (*) indicates significant differences compared with corresponding WT cells (P<0.05).

Figure 5

Figure 5. Diagram scheme indicating possible stages of wild-type-SCV life/stress cycle.

Under optimal conditions, WT populations prevail, perhaps masking the SCV phenotype. Introduction of stress selects for a more resilient phenotype, changing the population dynamics from WT prevalence to SCV prevalence which persist even under prolonged exposure to stress. Removal of stress shifts population dynamics with WT populations dominating again.

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This work was partially supported via a University of Newcastle International Scholarship and Gideon Lang Scholarship, and additional funding from the Harold Stannet Williams and Judith Mason Research Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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