Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test

Contamination of cells/tissues by infectious pathogens (e.g., fungi, viruses, or bacteria, including mycoplasma) is a major problem in cell-based transplantation. In this study, we tested a polymerase chain reaction (PCR) method to provide rapid, simple, and sensitive detection of mycoplasma contami...

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Autores principales:	Sunao Sugita, Ayumi Hono, Shoko Fujino, Yoko Futatsugi, Yuta Yunomae, Norio Shimizu, Masayo Takahashi
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	mycoplasma polymerase chain reaction iPS cells retinal cells clinical trial Biology (General) QH301-705.5 Chemistry QD1-999
Acceso en línea:	https://doaj.org/article/96732022a3134353b63983ceb14c1792
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spelling	oai:doaj.org-article:96732022a3134353b63983ceb14c17922021-11-25T17:57:46ZDetection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test10.3390/ijms2222125551422-00671661-6596https://doaj.org/article/96732022a3134353b63983ceb14c17922021-11-01T00:00:00Zhttps://www.mdpi.com/1422-0067/22/22/12555https://doaj.org/toc/1661-6596https://doaj.org/toc/1422-0067Contamination of cells/tissues by infectious pathogens (e.g., fungi, viruses, or bacteria, including mycoplasma) is a major problem in cell-based transplantation. In this study, we tested a polymerase chain reaction (PCR) method to provide rapid, simple, and sensitive detection of mycoplasma contamination in laboratory cultures for clinical use. This mycoplasma PCR system covers the <i>Mycoplasma</i> species (spp.) listed for testing in the 17th revision of the Japanese Pharmacopoeia, and we designed it for use in transplantable retinal cells. Here, we analyzed mycoplasma contamination in induced pluripotent stem cell (iPS cell)-derived transplantable retinal pigment epithelium (RPE) cells. In the spike tests to RPE cells with nine species of class <i>Mollicutes</i> bacteria, including seven <i>Mycoplasma</i> spp. and one of each <i>Acholeplasma</i> spp. and <i>Ureaplasma</i> spp., contamination at the concentration of 100 and 10 CFU/mL were detected with 100% probability in all cases, while 1 CFU/mL had a detection rate of 0–75%. DNA prepared from bacteria species other than class <i>Mollicutes</i> species was not detectable, indicating the specificity of this PCR. While iPS cells and iPS-RPE cells established in our laboratory were all negative by this PCR, some of the commercially available cell lines were positive. Cells for transplantation should never have infection, as once pathogens are implanted into the eyes, they can cause severe intraocular inflammation. Thus, it is imperative to monitor for infections in the transplants, although generally, mycoplasma infection is difficult to detect.Sunao SugitaAyumi HonoShoko FujinoYoko FutatsugiYuta YunomaeNorio ShimizuMasayo TakahashiMDPI AGarticlemycoplasmapolymerase chain reactioniPS cellsretinal cellsclinical trialBiology (General)QH301-705.5ChemistryQD1-999ENInternational Journal of Molecular Sciences, Vol 22, Iss 12555, p 12555 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	mycoplasma polymerase chain reaction iPS cells retinal cells clinical trial Biology (General) QH301-705.5 Chemistry QD1-999
spellingShingle	mycoplasma polymerase chain reaction iPS cells retinal cells clinical trial Biology (General) QH301-705.5 Chemistry QD1-999 Sunao Sugita Ayumi Hono Shoko Fujino Yoko Futatsugi Yuta Yunomae Norio Shimizu Masayo Takahashi Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
description	Contamination of cells/tissues by infectious pathogens (e.g., fungi, viruses, or bacteria, including mycoplasma) is a major problem in cell-based transplantation. In this study, we tested a polymerase chain reaction (PCR) method to provide rapid, simple, and sensitive detection of mycoplasma contamination in laboratory cultures for clinical use. This mycoplasma PCR system covers the <i>Mycoplasma</i> species (spp.) listed for testing in the 17th revision of the Japanese Pharmacopoeia, and we designed it for use in transplantable retinal cells. Here, we analyzed mycoplasma contamination in induced pluripotent stem cell (iPS cell)-derived transplantable retinal pigment epithelium (RPE) cells. In the spike tests to RPE cells with nine species of class <i>Mollicutes</i> bacteria, including seven <i>Mycoplasma</i> spp. and one of each <i>Acholeplasma</i> spp. and <i>Ureaplasma</i> spp., contamination at the concentration of 100 and 10 CFU/mL were detected with 100% probability in all cases, while 1 CFU/mL had a detection rate of 0–75%. DNA prepared from bacteria species other than class <i>Mollicutes</i> species was not detectable, indicating the specificity of this PCR. While iPS cells and iPS-RPE cells established in our laboratory were all negative by this PCR, some of the commercially available cell lines were positive. Cells for transplantation should never have infection, as once pathogens are implanted into the eyes, they can cause severe intraocular inflammation. Thus, it is imperative to monitor for infections in the transplants, although generally, mycoplasma infection is difficult to detect.
format	article
author	Sunao Sugita Ayumi Hono Shoko Fujino Yoko Futatsugi Yuta Yunomae Norio Shimizu Masayo Takahashi
author_facet	Sunao Sugita Ayumi Hono Shoko Fujino Yoko Futatsugi Yuta Yunomae Norio Shimizu Masayo Takahashi
author_sort	Sunao Sugita
title	Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
title_short	Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
title_full	Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
title_fullStr	Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
title_full_unstemmed	Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test
title_sort	detection of mycoplasma contamination in transplanted retinal cells by rapid and sensitive polymerase chain reaction test
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/96732022a3134353b63983ceb14c1792
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Detection of Mycoplasma Contamination in Transplanted Retinal Cells by Rapid and Sensitive Polymerase Chain Reaction Test

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