Multiplex detection in tonsillar tissue of all known human polyomaviruses

Mohammadreza Sadeghi,Mari Toppinen,Maria Söderlund-Venermo,Torbjörn Ramqvist,Lari Pyöriä,Klaus Hedman,Leena-Maija Aaltonen,Yilin Wang

doi:10.1186/s12879-017-2479-5

Mohammadreza Sadeghi, Mari Toppinen + Show 6 more

Open Access

https://doi.org/10.1186/s12879-017-2479-5

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Abstract

BackgroundIn the past few years, eleven new human viruses have joined the two previously known members JCPyV and BKPyV of the Polyomaviridae family, by virtue of molecular methods. Serology data suggest that infections with human polyomaviruses (HPyVs) occur since childhood and the viruses are widespread in the general population. However, the viral persistence sites and transmission routes are by and large unknown. Our previous studies demonstrated that the four new HPyVs – KIPyV, WUPyV, MCPyV and TSPyV – were present in the tonsils, and suggested lymphoid tissue as a persistent site of these emerging human viruses.We developed a Luminex-based multiplex assay for simultaneous detection of all 13 HPyVs known, and explored their occurrence in tonsillar tissues of children and adults mostly with tonsillitis or tonsillar hypertrophy.MethodsWe set up and validated a new Luminex-based multiplex assay by using primer pairs and probes targeting the respective HPyV viral protein 1 (VP1) genes. With this assay we tested 78 tonsillar tissues for DNAs of 13 HPyVs.ResultsThe multiplex assay allowed for simultaneous detection of 13 HPyVs with high analytical sensitivity and specificity, with detection limits of 100–102 copies per microliter, and identified correctly all 13 target sequences with no cross reactions. HPyV DNA altogether was found in 14 (17.9%) of 78 tonsils. The most prevalent HPyVs were HPyV6 (7.7%), TSPyV (3.8%) and WUPyV (3.8%). Mixed infection of two HPyVs occurred in one sample.ConclusionsThe Luminex-based HPyV multiplex assay appears highly suitable for clinical diagnostic purposes and large-scale epidemiological studies. Additional evidence was acquired that the lymphoid system plays a role in HPyV infection and persistence. Thereby, shedding from this site during reactivation might take part in transmission of the newly found HPyVs.

Highlights

In the past few years, eleven new human viruses have joined the two previously known members JC polyomavirus (JCPyV) and BK polyomavirus (BKPyV) of the Polyomaviridae family, by virtue of molecular methods
We have reported the occurrence of DNA of Merkel cell polyomavirus (MCPyV) and Trichodysplasia spinulosa (TSPyV) in tonsillar tissue, suggesting persistence in lymphoid tissue or mucosa [36, 37]
We designed a multiplex assay for the detection and identification of 13 human polyomaviruses (HPyVs), by extension of a previously designed multiplex assay for 10 HPyVs [53]

Summary

Introduction

In the past few years, eleven new human viruses have joined the two previously known members JCPyV and BKPyV of the Polyomaviridae family, by virtue of molecular methods. Serology data suggest that infections with human polyomaviruses (HPyVs) occur since childhood and the viruses are widespread in the general population. Our previous studies demonstrated that the four new HPyVs – KIPyV, WUPyV, MCPyV and TSPyV – were present in the tonsils, and suggested lymphoid tissue as a persistent site of these emerging human viruses. We developed a Luminex-based multiplex assay for simultaneous detection of all 13 HPyVs known, and explored their occurrence in tonsillar tissues of children and adults mostly with tonsillitis or tonsillar hypertrophy. The primary infections occur asymptomatically during childhood and are followed by lifelong DNA persistence.

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