Takahashi Tadanobu, Agarikuchi Takashi, Kurebayashi Yuuki, Shibahara Nona, Suzuki Chihiro, Kishikawa Akiko, Fukushima Keijo, Takano Maiko, Suzuki Fumie, Wada Hirohisa, Otsubo Tadamune, Ikeda Kiyoshi, Minami Akira, Suzuki Takashi
Department of Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka-shi, Shizuoka, Japan.
Shizuoka City Institute of Environmental Sciences and Public Health, Shizuoka-shi, Shizuoka, Japan.
PLoS One. 2015 Dec 2;10(12):e0144038. doi: 10.1371/journal.pone.0144038. eCollection 2015.
Mumps viruses show diverse cytopathic effects (CPEs) of infected cells and viral plaque formation (no CPE or no plaque formation in some cases) depending on the viral strain, highlighting the difficulty in mumps laboratory studies. In our previous study, a new sialidase substrate, 2-(benzothiazol-2-yl)-4-bromophenyl 5-acetamido-3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosidonic acid (BTP3-Neu5Ac), was developed for visualization of sialidase activity. BTP3-Neu5Ac can easily and rapidly perform histochemical fluorescent visualization of influenza viruses and virus-infected cells without an antiviral antibody and cell fixation. In the present study, the potential utility of BTP3-Neu5Ac for rapid detection of mumps virus was demonstrated. BTP3-Neu5Ac could visualize dot-blotted mumps virus, virus-infected cells, and plaques (plaques should be called focuses due to staining of infected cells in this study), even if a CPE was not observed. Furthermore, virus cultivation was possible by direct pick-up from a fluorescent focus. In conventional methods, visible appearance of the CPE and focuses often requires more than 6 days after infection, but the new method with BTP3-Neu5Ac clearly visualized infected cells after 2 days and focuses after 4 days. The BTP3-Neu5Ac assay is a precise, easy, and rapid assay for confirmation and titration of mumps virus.
腮腺炎病毒对受感染细胞会表现出不同的细胞病变效应（CPEs），并且根据病毒株的不同还会形成病毒蚀斑（在某些情况下无CPE或无蚀斑形成），这凸显了腮腺炎实验室研究的困难。在我们之前的研究中，开发了一种新的唾液酸酶底物，即2-（苯并噻唑-2-基）-4-溴苯基5-乙酰氨基-3,5-二脱氧-α-D-甘油-D-半乳糖-2-壬酮糖醛酸（BTP3-Neu5Ac），用于可视化唾液酸酶活性。BTP3-Neu5Ac无需抗病毒抗体和细胞固定，就能轻松快速地对流感病毒和病毒感染细胞进行组织化学荧光可视化。在本研究中，证明了BTP3-Neu5Ac在快速检测腮腺炎病毒方面的潜在效用。即使未观察到CPE，BTP3-Neu5Ac也能使斑点印迹的腮腺炎病毒、病毒感染细胞和蚀斑（在本研究中，由于感染细胞的染色，蚀斑应称为病灶）可视化。此外，通过直接从荧光病灶中提取进行病毒培养是可行的。在传统方法中，感染后通常需要6天以上才能出现可见的CPE和病灶，但使用BTP3-Neu5Ac的新方法在2天后能清晰地使感染细胞可视化，在4天后能使病灶可视化。BTP3-Neu5Ac检测法是一种用于确认和滴定腮腺炎病毒的精确、简便且快速的检测方法。
