Itahashi Motoki, Higaki Shiro, Fukuda Masahiko, Shimomura Yoshikazu
Department of Ophthalmology, Kinki University School of Medicine, 377-2 Ohno-Higashi, Osaka-Sayama 589-8511, Japan.
Arch Ophthalmol. 2010 May;128(5):535-40. doi: 10.1001/archophthalmol.2010.66.
To detect and quantitate the causative pathogens in patients with corneal ulcer using real-time polymerase chain reaction (PCR) by cycling probe.
Clinical and laboratory study of 40 eyes of 40 patients diagnosed with corneal ulcer. Two methods were used for pathogen detection: bacterial culture and real-time PCR with the patient's corneal scrapings. Probes and primers of real-time PCR were designed to be pathogen specific for simultaneous detection of Staphylococcus aureus, Staphylococcus pneumoniae, Pseudomonas aeruginosa, methicillin-resistant S aureus, Candida species, and Fusarium species. Results by both methods were evaluated and compared.
Of 40 eyes, 20 eyes had the same pathogens detected by both methods and those were S aureus (3 eyes; mean [SE], 3.8 [1.3] x 10(1) copies/sample), S pneumoniae (5 eyes; mean [SE], 5.6 [5.1] x 10(3) copies/sample), P aeruginosa (8 eyes; 5.1 [4.0] x 10(3) copies/sample), methicillin-resistant S aureus (1 eye; 1.0 x 10(2) copies/sample), and Candida species (3 eyes; mean [SE], 8.8 [4.9] x 10(3) copies/sample). Six eyes showed negative results by both methods. Results of both methods disagreed in 14 eyes; specifically, 11 had positive PCR results only, 2 had positive culture results only, and 1 eye had positive results for different pathogens.
The real-time PCR assay can simultaneously detect and quantitate bacterial and fungal pathogens in patients with corneal ulcer. Real-time PCR can be a fast diagnostic tool and may be useful as an adjunct to identify potential pathogens.
采用循环探针实时聚合酶链反应（PCR）检测并定量分析角膜溃疡患者的致病病原体。
对40例诊断为角膜溃疡的患者的40只眼进行临床和实验室研究。采用两种方法进行病原体检测：细菌培养以及对患者角膜刮片进行实时PCR检测。实时PCR的探针和引物设计为针对金黄色葡萄球菌、肺炎链球菌、铜绿假单胞菌、耐甲氧西林金黄色葡萄球菌、念珠菌属和镰刀菌属的病原体特异性探针和引物。对两种方法的结果进行评估和比较。
在40只眼中，20只眼通过两种方法检测出相同的病原体，分别为金黄色葡萄球菌（3只眼；平均值[标准误]，3.8[1.3]×10¹拷贝/样本）、肺炎链球菌（5只眼；平均值[标准误]，5.6[5.1]×10³拷贝/样本）、铜绿假单胞菌（8只眼；5.1[4.0]×10³拷贝/样本）、耐甲氧西林金黄色葡萄球菌（1只眼；1.0×10²拷贝/样本）和念珠菌属（3只眼；平均值[标准误]，8.8[4.9]×10³拷贝/样本）。6只眼两种方法检测结果均为阴性。两种方法的结果在14只眼中不一致；具体而言，11只眼仅PCR结果为阳性，2只眼仅培养结果为阳性，1只眼针对不同病原体结果为阳性。
实时PCR检测可同时检测并定量分析角膜溃疡患者的细菌和真菌病原体。实时PCR可作为一种快速诊断工具，可能有助于辅助识别潜在病原体。
