FEB 2014

EyeWorld is the official news magazine of the American Society of Cataract & Refractive Surgery.

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Page 70 of 114

be validated—everything," he said. Without such validation the labora- tory will not receive reimbursement. "You have to know if the test really works and you can't piggyback on someone else's validation," Mr. Kowalski said. This can be problem- a tic since in ophthalmology there are infections that are not very com- mon. "When you validate you have to have true positive specimens, which are backed by culture, and you also have to have true negative specimens." However, for an infec- tion like pseudomonas, Mr. Kowalski noted he might only get 10 to 20 p ositive cultures per years. As a result, this takes a long time to validate. The same is true for some other ophthalmic infections. With Acanthamoeba, Mr. Kowalski recalled, it took two or three years to develop a PCR test for it; when he tried to take it to another laboratory it took another t hree years to once again validate. So for most infections Mr. Kowalski still views culturing as preferable and currently much more practical. "I'm old school," he said. "I like to grow. It's probably faster." He pointed out that for small corneal ulcers practitioners will likely put the patient on a fourth- generation fluoroquinolone without even culturing, adding that when it comes to larger bacterial ulcers a good smear is very effective. "I can tell you in 10 minutes on a gram stain what you have," Mr. Kowalski said. "The smears are already very helpful as far as that's concerned." The same is not true for PCR, he noted. "No one is ever going to take a sample, drop everything that they're doing, do the PCR testing and let you know that same day," Mr. Kowalski said. Dr. Iovieno agrees that this has limitations. "Given the costs and the expertise involved, the application of molecular biology techniques on a large scale is quite limited," Dr. Iovieno said. "Traditional microbiol- ogy tools when correctly employed are sufficient to make a timely and appropriate diagnosis in the major- ity of cases." PCR accuracy may also be com- promised at many different steps along the way. "Many factors from adequate sampling, nucleic acid ex- traction to PCR reaction optimiza- tion can influence the final results," Dr. Iovieno said. "This is why the process requires the expertise of mo- lecular biologists as well as microbi- ologists and an adequate timeframe for optimization." Mr. Kowalski likewise observed that having a good specimen is vital. In addition, patient pretreatment can play a role here. Commonly used fluorescein dyes, for example, may interfere with the accuracy of results. In PCR testing, they often u se a fluorescent probe, he ex- plained. The PCR machine picks up this fluorescence and therefore if you add fluorescein dye it could throw off results. Availability is likely to remain extremely limited for some time, thinks Dr. Jeng. "I don't envision that this is something that every of- f ice is going to be able to have," he said. "I think that it may be limited to certain large centers." Indeed, that's what Dr. Iovieno finds. "Unfortunately molecular diagnostic tools are available only in tertiary centers in Europe," he said. "We encourage all hospitals and academic centers in our region t hat do not have this commodity to send patients or samples to our attention." Still, he envisions this as playing a larger role going forward. "In my opinion, molecular microbiology represents the future in the diagno- sis of infectious keratitis," Dr. Iovieno said. "It will also comple- ment our understanding of disease pathophysiology by detecting sub- clinical infection or defining micro- bial populations." Efforts have been made to simplify and shorten the process, as well as to reduce the machinery and the costs involved, which he thinks will allow such technology to be more widely dispensed. For his part, Mr. Kowalski en- courages individuals to keep trying to improve this technology and lower costs. "It does come down to dollars and cents," he said. "I think it's important to keep on looking, but in its present state I don't see any PCR to be practical for bacterial infections," he said, but that does not mean a better way will not arise in the future. EW Editors' note: Dr. Hawwa has financial interests with IntelligentMDx. Dr. Iovieno, Dr. Jeng, and Mr. Kowalski have no financial interests related to this article. Contact information Hawwa: RHawwa@intelligentmdx.com Iovieno: alfonso.iovieno@asmn.re.it Jeng: BJeng@som.umaryland.edu Kowalski: kowalskirp@upmc.edu Corneal infections February 2014 Counterculture continued from page 67 56-68 Feature_EW February 2014-DL2_Layout 1 1/30/14 4:05 PM Page 68

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