NOTIFy (non-toxic lyophilized field)-FISH for the identification of biological agents by Fluorescence in situ Hybridization
Fig 6
Application of the diagnostic algorithm for the identification of bacteria in clinical samples.
E. coli was identified in an infected skin lesion, Brucella spp.in a blood culture and Burkholderia pseudomallei in a swab of an infected patient. In the infected skin lesion, bacteria that were only detect by DAPI, but not by any of the other probes used were present (arrows). Efficient probe binding was controlled with a positive control containing a mixture of known bacteria (control). Shown are images acquired after hybridization with formamide-based (wound infection and control) or urea-based reagents (blood culture and swab). E. coli was detected by the probe Gam42a (labeled in Cy5 and FLUOS, turquoise color in the overlay) in the first hybridization and identified by the probe Esco864 (labeled in Cy5) in the second hybridization. Brucella was identified by Bru996 (labeled in FLUOS) in the first hybridization. B. pseudomallei was detected by the probe Bet42a (labeled in Cy3 and FLUOS, yellow color in the overlay) in the first hybridization and identified with the probe Bboth (labeled in Cy3) in the second hybridization. In the control overlay, specific binding of the probes used in the first hybridization is shown by detection of B. anthracis in pink, Y. pestis in turquoise, B. suis in green, L. borgpetersenii in red, B. pseudomallei in yellow and R. slovaca in blue. The probes and fluorophores resulting in signals in the respective sample are indicated on the far right under a representation of the additive mixing of colors. Bars 10 μm.