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Nimesh Patel, Darlene Miller, Harry W Flynn; The Novel Use of Fluorescence in Situ Hybridization (FISH) for the Rapid Detection of Pathogens from Intravitreal Isolates in Endophthalmitis. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6387.
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© ARVO (1962-2015); The Authors (2016-present)
Rapid identification of pathogens causing endophthalmitis can improve patient care. A new molecular application of fluorescence hybridization has been used to reduce identification time of Staphylococcal species in blood cultures from patients with septicemia. This retrospective study evaluated FISH with Staphylococcus specific molecular DNA probes for the rapid detection of common pathogens from endophthalmitis isolates.
Vitreous samples were obtained from patients with endophthalmitis during vitreous tap or vitrectomy. Standard inocula were prepared to a final concentration of 1x 10^5 cfu / ml. A total of 20 isolates were tested. Of these, 5 were isolated Staphalococcus aureus, 5 were isolated coagulase negative Staphylococcus, 5 were mixed samples and 5 were negative controls. Blood culture bottles and thioglycollate broth were inoculated and incubated at 35 C for 12-18 hours. Smears of samples were prepared per manufacturer protocol. Molecular probes for Staphylococcus aureus and for coagulase negative Staphylococci were added to slides. These were hybridized at 55 C for 15 minutes. Slides were then read on a fluorescence microscope for detection (Figure 1).
This technique was able to identify Staphylococci in 19 of 20 samples for a 95% correlation. Rate of detection was 9 of 10 (90%) for Staphylococcus aureus and 10 of 10 (100%) for coagulase negative Staphylococci. No fluorescence was found in 5 of 5 (100%) of the control non- staphylococcal cultures. Detection time was within 20 minutes compared to 1-3 days for routine culture methods.
Current laboratory protocols require labor and time intensive methods for the identification of pathogens in endophthalmitis with culture and media plating. This study shows a proof of concept for a microbial detection system using FISH molecular probes. This could have a major impact on the rapid and accurate identification of isolates from endophthalmitis patients.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
Figure 1: Fluorescence in Situ Hybridization for detection of Staphylococci from intraocular fluids. Green: S. aureus Red: Coagulase negative staphylococci.
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