Phenotypic Variants of Staphylococci and Their Underlying Population Distributions Following Exposure to Stress

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.

Amino acids and proteomic acclimation of Staphylococcus aureus when incubated in a defined minimal medium supplemented with 5% sodium chloride

“…The changes observed in the metabolome and proteome may represent a phenotypic shift that can occur within an existing population of cells without cell division. Such a shift may be indicative, for example, of the formation of small colony variants that have been previously shown to form in response exposures to NaCl, with accompanying reduced metabolism compared to wild‐type bacteria (Crompton et al, ; Onyango et al, ; Proctor et al, ; von Eiff, ). Prior investigations have shown that small colony variants have slower growth rates, reduced production of virulence factors and some are auxotrophic for important compounds such as haemin and menadione (Bui et al, ; Moisan et al, ; Proctor et al, ; von Eiff et al, ).…”

“…Glutamic acid can readily be used for energy production by oxidation and it is required for the synthesis of the peptidoglycan bacterial cell wall, especially relevant if cell wall thickening is part of the response to a NaCl challenge (Doublet, Heijenoort, & Mengin‐Lecreulx, ). It has been shown that in staphylococcal species, the thickness of the cell wall was significantly altered following exposures to environmental changes including elevated NaCl (Onyango et al, ).…”

“…It has recently been shown that the osmotic pressure, variations in pH and cold stress during growth can induce different phenotypes with obvious alterations in the cell wall structure of these phenotypes (Onyango et al, ). Fourier Transform Infrared Spectroscopy (FTIR) study has also shown that the exposure of S. aureus to undesirable conditions included NaCl led to a significant phenotypic shifting (Wehrli et al, ).…”

In VitroEvaluations andIn VivoToxicity and Efficacy Studies of MFM501 against MRSA

“…The increased cell sizes indicated disruption of cell divisions and slower growth rate in the treated cells [25–27]. Although the cell multiplication was abruptly slowed or stopped, the cellular metabolic processes remained unaffected, hence the increased cell size; that leads to the release of autolytic enzyme under lethal concentrations of inhibitory agents, which in the end resulted in the destruction and deterioration of the treated cells [27].…”

New insights in thermal resistance of staphylococcal strains belonging to the species Staphylococcus epidermidis, Staphylococcus lugdunensis and Staphylococcus aureus

“…The ability of staphylococci to grow over a wide range of environmental conditions has been demonstrated (Prescott, Harley, & Klein, 2002), and related to their capability to rapidly adapt to harsh conditions, surviving and taking advantage when optimal or less limiting conditions are restored (Onyango, Hugh Dunstan, Roberts, Macdonald, & Gottfries, 2013). This adaptation process is clearly revealed during plate counting by the presence of colonies showing morphological differences, including variation in colony size, pigmentation and haemolytic irregularities (Kipp, Becker, Peters, & von Eiff, 2004;Onyango, Hugh Dunstan, Gottfries, Von Eiff, & Roberts, 2012).…”