The effect on the autofluorescence signal of corneal microorganisms with common topical drugs

Abstract

Purpose : To evaluate the effect of two topical antibiotics and one topical anaesthetic drug on the autofluorescence pattern of three corneal microorganisms.

Methods : Three organisms known to cause corneal infection: Acanthamoeba castellani (ATCC 30868), Pseudomonas aeruginosa (ATCC 9027) and Staphylococcus aureus (ATCC 6538) were cultured as per standard protocols. Ciprofloxacin (0.00625%), tetracycline (0.0125%) and tetracaine (0.1%) were used. After growth, the microorganisms were diluted in either PBS (bacteria) or ¼ strength Ringer’s solution (Acanthamoeba) to give final concentrations of 10⁴ cells/ml (measured by optical density at 660nm or using a Neubauer haemocytometer). Microbial cells were added to quartz crystal cuvettes along with the drugs. Excitation emission was collected for each sample at 269-500nm with increments in 5nm steps, whereas emission wavelengths ranged from 280-700 nm at 2nm steps. The data were analysed using MATLAB software to produce smoothed colour coded images of the samples tested. These were then analysed for changes in the spectra.

Results : Tetracaine changed the fluorescence of all three micro-organisms, moving the emission/excitation maxima from 325/280nm to 370/350nm. Ciprofloxacin only produced changes in the two bacteria, changing the emission/excitation maxima from 325/280nm and splitting it into two maxima at 425/260nm and 425/330nm. Addition of tetracycline to the bacteria or Acanthamoeba did not alter the emission/excitation maxima.

Conclusions : Fluorescence signals have the potential to be used as diagnostic markers for different organisms. This study has shown that they may also be useful to show the effectiveness of drugs during therapy. Further experiments need to be carried out to evaluate what other factors can affect the fluorescence signals.

This is a 2020 ARVO Annual Meeting abstract.