Satoko Nakano1, Sunao Sugita2, Yasuhiro Tomaru3, Ayumi Hono2, Takako Nakamuro1, Toshiaki Kubota1, Hiroshi Takase4, Manabu Mochizuki5, Masayo Takahashi2, Norio Shimizu3. 1. Department of Ophthalmology, Oita University, Oita, Japan. 2. Laboratory for Retinal Regeneration, RIKEN Center for Developmental Biology, Kobe, Japan. 3. Center for Stem Cell and Regenerative Medicine, Tokyo Medical and Dental University, Tokyo, Japan. 4. Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Graduate School of Medicine and Dental Sciences, Tokyo, Japan. 5. Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Graduate School of Medicine and Dental Sciences, Tokyo, Japan 5Miyata Eye Hospital, Miyakonojo, Japan.
Abstract
Purpose: To establish and evaluate a new multiplex solid-phase strip polymerase chain reaction (strip PCR) for concurrent detection of common ocular infectious disease pathogens. Methods: A new multiplex strip PCR was established to detect 24 common ocular infectious disease pathogens: herpes simplex virus (HSV) 1, HSV2, varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpes virus (HHV) 6, HHV7, HHV8, human T-cell lymphotropic virus (HTLV)-1, adenovirus, Mycobacterium tuberculosis, Treponema pallidum, Propionibacterium acnes (P. acnes), bacterial 16S ribosomal RNA (rRNA), Candida species (Candida sp.), C. glabrata, C. krusei, Aspergillus, Fusarium, fungal 28S rRNA, Toxoplasma (T. gondii), Toxocara, Chlamydia trachomatis (C. trachomatis), and Acanthamoeba. Strip PCR was tested with a negative control (distilled water) and standard positive control DNA. Cutoffs of quantification cycle (Cq) values were determined with noninfectious ocular samples to avoid false-positives caused by contamination with P. acnes, bacterial 16S, and fungal 28S from reagents and ocular surfaces. A pilot study to evaluate the strip PCR was performed using infectious ocular samples (aqueous humor, vitreous, cornea, and tears) by strip PCR and previously developed capillary-type multiplex PCR and quantitative real-time PCR (qPCR). Results: Strip PCR was verified with negative and positive controls. Strip PCR rapidly detected HSV1, HSV2, VZV, EBV, CMV, HHV6, HHV7, HTLV-1, adenovirus, P. acnes, bacterial 16S, Candida sp., C. glabrata, Aspergillus, fungal 28S, T. gondii, C. trachomatis, and Acanthamoeba in patient samples. The sensitivity was comparable to that of qPCR. Conclusions: Our novel strip PCR assay is a simple, rapid, and high-sensitivity method for detecting ocular infectious disease pathogens.
Purpose: To establish and evaluate a new multiplex solid-phase strip polymerase chain reaction (strip PCR) for concurrent detection of common ocular infectious disease pathogens. Methods: A new multiplex strip PCR was established to detect 24 common ocular infectious disease pathogens: herpes simplex virus (HSV) 1, HSV2, varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpes virus (HHV) 6, HHV7, HHV8, human T-cell lymphotropic virus (HTLV)-1, adenovirus, Mycobacterium tuberculosis, Treponema pallidum, Propionibacterium acnes (P. acnes), bacterial 16S ribosomal RNA (rRNA), Candida species (Candida sp.), C. glabrata, C. krusei, Aspergillus, Fusarium, fungal 28S rRNA, Toxoplasma (T. gondii), Toxocara, Chlamydia trachomatis (C. trachomatis), and Acanthamoeba. Strip PCR was tested with a negative control (distilled water) and standard positive control DNA. Cutoffs of quantification cycle (Cq) values were determined with noninfectious ocular samples to avoid false-positives caused by contamination with P. acnes, bacterial 16S, and fungal 28S from reagents and ocular surfaces. A pilot study to evaluate the strip PCR was performed using infectious ocular samples (aqueous humor, vitreous, cornea, and tears) by strip PCR and previously developed capillary-type multiplex PCR and quantitative real-time PCR (qPCR). Results: Strip PCR was verified with negative and positive controls. Strip PCR rapidly detected HSV1, HSV2, VZV, EBV, CMV, HHV6, HHV7, HTLV-1, adenovirus, P. acnes, bacterial 16S, Candida sp., C. glabrata, Aspergillus, fungal 28S, T. gondii, C. trachomatis, and Acanthamoeba in patient samples. The sensitivity was comparable to that of qPCR. Conclusions: Our novel strip PCR assay is a simple, rapid, and high-sensitivity method for detecting ocular infectious disease pathogens.
Authors: Thomas Ferreira de Moura; Anne Limelette; Carl Arndt; Thomas Guillard; Laurent Andreoletti; Alexandre Denoyer Journal: Am J Ophthalmol Case Rep Date: 2022-05-28