Modelling the effects of temperature, pH and osmotic shifts on the autofluorescence of <i>Staphylococcus aureus</i> in vitro

This study aims to investigate how different wound microenvironmental factors (temperature, pH, and osmotic pressure) influence the autofluorescence of Staphylococcus aureus (<i>S. aureus)</i> and its underlying molecular mechanisms, specifically focusing on the porphobilinogen synthase gene <i>(hemB)</i> gene expression. We measured the average fluorescence intensity of <i>S. aureus</i> colonies under varying conditions of pH (3, 5, 7, 9, 11), temperature (25°C, 31°C, 37°C, 43°C), and osmotic pressure (0.9%, 1.8%, 2.7%, 3.6%) over time. Fluorescence intensity was quantified using ImageJ software. Additionally, RT-qPCR was used to analyze the expression levels of the <i>hemB</i> under these conditions. Bacterial fluorescence intensity increased as the temperature ranged from 25°C to 43°C, with corresponding upregulation of <i>hemB</i> expression. At pH values between 3 and 11, fluorescence intensity decreased as pH increased, reflecting a similar trend in <i>hemB</i> expression. Fluorescence also diminished with higher osmotic pressures (0.9% to 3.6%), mirroring the downregulation of <i>hemB.</i> Our findings indicate that temperature, pH, and osmotic pressure significantly affect the autofluorescence of <i>S. aureus</i> by modulating porphyrin accumulation through <i>hemB</i> gene expression. These environmental factors should be considered when using bacterial fluorescence for wound infection assessment. There are many bacteria on our skin that can behave differently when we get a cut in the skin. One way that doctors look at how the bacteria are behaving in a cut is using fluorescence, where the bacteria glow in the dark. This research looks at how whether changes in temperature, acidity and salt can change the way that bacteria glow in the dark. We found that when it was warmer, the bacteria glow more brightly. This brightness is linked to a gene called <i>hemB</i> in the bacteria. On the other hand, if the environment becomes less acidic or if there is a lot of salt, the bacteria do not glow as brightly. By watching these changes in fluorescence, doctors can get a better idea of what’s happening with the bacteria in the wound. Our research shows that when using this bacterial fluorescence to check on wounds, we need to remember the effects of environmental factors.