Kowalski Regis P, Melan Melissa A, Karenchak Lisa M, Mammen Alex
Department of Ophthalmology (R.P.K., L.M.K., A.M.), Charles T. Campbell Eye Microbiology Laboratory, University of Pittsburgh Medical Center (UPMC), University of Pittsburgh, Pittsburgh, PA; and Division of Molecular Diagnostics (M.A.M.), Department of Pathology, University of Pittsburgh Medical Center (UPMC), University of Pittsburgh, Pittsburgh, PA.
Eye Contact Lens. 2015 Nov;41(6):341-3. doi: 10.1097/ICL.0000000000000131.
Acanthamoeba keratitis should be definitively diagnosed for appropriate therapy. Our institution has validated polymerase chain reaction (PCR) as a routine diagnostic test to detect Acanthamoeba DNA from ocular samples. We compared PCR with culture isolation for detecting Acanthamoeba from ocular samples.
The microbiology records of patients that had specimens submitted (May 2012 to January 2014) for laboratory testing for Acanthamoeba keratitis were reviewed for (1) Acanthamoeba culture isolation, (2) Acanthamoeba DNA detection by PCR, and (3) non-Acanthamoeba culture results. For Acanthamoeba isolation, corneal samples were planted on nonnutrient agar overlaid with Enterobacter aerogenes. Validated PCR (May 2012) for Acanthamoeba DNA was processed at the Division of Molecular Diagnostics, UPMC, Pittsburgh, PA. Additional cultures were obtained for bacteria, fungus, and virus (i.e., herpes simplex virus) using standard techniques.
Culture isolation and PCR were processed on 125 patients with a differential diagnosis of Acanthamoeba keratitis. Of these, 104 (83.2%) were culture negative, PCR negative; 14 (11.2%) were culture positive, PCR positive; 4 (3.2%) were culture negative, PCR positive; and, 3 (2.4%) were culture positive, PCR negative. Culture and PCR were statistically equivalent for detecting Acanthamoeba from ocular samples (P=1.0, McNemar's test). Nineteen of the culture-negative, PCR-negative corneal samples (18.3%) were positive for other pathogens such as bacteria, fungus, and virus.
There is no clear advantage of PCR over culture isolation for detecting Acanthamoeba in ocular specimens. Other pathogens such as bacteria, fungus, and virus must still be considered in severe persistent keratitis. Polymerase chain reaction seems to be a complementary test for the clinical support of Acanthamoeba keratitis.
棘阿米巴角膜炎应得到明确诊断以便进行恰当治疗。我们机构已验证聚合酶链反应（PCR）作为一种常规诊断检测方法，用于从眼部样本中检测棘阿米巴DNA。我们比较了PCR与培养分离法从眼部样本中检测棘阿米巴的情况。
回顾了2012年5月至2014年1月期间提交样本进行棘阿米巴角膜炎实验室检测的患者的微生物学记录，内容包括：（1）棘阿米巴培养分离；（2）通过PCR检测棘阿米巴DNA；（3）非棘阿米巴培养结果。对于棘阿米巴分离，将角膜样本接种在覆盖产气肠杆菌的无营养琼脂上。在宾夕法尼亚州匹兹堡市匹兹堡大学医学中心分子诊断科对棘阿米巴DNA进行验证的PCR检测（2012年5月）。使用标准技术对细菌、真菌和病毒（即单纯疱疹病毒）进行额外培养。
对125例临床诊断为棘阿米巴角膜炎的患者进行了培养分离和PCR检测。其中，104例（83.2%）培养阴性、PCR阴性；14例（11.2%）培养阳性、PCR阳性；4例（3.2%）培养阴性、PCR阳性；3例（2.4%）培养阳性、PCR阴性。从眼部样本中检测棘阿米巴时，培养和PCR在统计学上等效（P = 1.0，McNemar检验）。19例培养阴性、PCR阴性的角膜样本（18.3%）对其他病原体如细菌、真菌和病毒呈阳性。
在眼部标本中检测棘阿米巴时，PCR相对于培养分离法没有明显优势。对于严重的持续性角膜炎，仍必须考虑其他病原体如细菌、真菌和病毒。聚合酶链反应似乎是用于棘阿米巴角膜炎临床辅助诊断的补充检测方法。
