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Development of an in situ hybridization assay using an AS1 probe for detection of bovine leukemia virus in BLV-induced lymphoma tissues

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Abstract

Enzootic bovine leukosis (EBL) is a malignant B cell lymphoma caused by infection with bovine leukemia virus (BLV). Histopathological examination is commonly used for diagnosis of the disease, but observation of lymphoma alone does not confirm EBL because cattle may be affected by sporadic forms of lymphoma that are not associated with BLV. Detection of BLV in tumor cells can be definitive evidence of EBL, but currently, there is no technique available for such a purpose. In this study, we focused on a viral non-coding RNA, AS1, and developed a novel in situ hybridization assay for the detection of BLV from formalin-fixed paraffin-embedded (FFPE) tissues. RNA-seq analysis revealed that all examined B lymphocytes derived from clinical EBL abundantly expressed AS1 RNA, indicating a possible target for detection. The in situ hybridization assay using an AS1 probe clearly detected AS1 RNA in fetal lamb kidney cells persistently infected with BLV. The utility of this assay in clinical samples was assessed using three EBL-derived lymph node specimens and one BLV-negative specimen, and AS1 RNA was detected specifically in the EBL-derived tissues. These results suggest that AS1 RNA is a useful target for the detection of BLV from FFPE specimens of tumor tissues. This technique is expected to become a powerful tool for EBL diagnosis.

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Acknowledgments

We thank the staff of the Kensei Meat Inspection Office and Kenpoku Livestock Hygiene Service Center in Ibaraki prefecture and the staff of the Awaji Meat Inspection Office in Hyogo prefecture for kindly providing EBL samples. We also thank M. Kobayashi for technical help.

Funding

This study was conducted under the research project “Regulatory research projects for food safety, animal health and plant protection (JPJ008617.17935709)” funded by the Ministry of Agriculture, Forestry and Fisheries of Japan.

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Authors and Affiliations

  1. Division of Viral Disease and Epidemiology, National Institute of Animal Health, National Agriculture and Food Research Organization, 3-1-5 Kannondai, Tsukuba, Ibaraki, 305-0856, Japan

    Kiyohiko Andoh, Asami Nishimori & Shinichi Hatama

  2. Division of Pathology and Pathophysiology, National Institute of Animal Health, National Agriculture and Food Research Organization, 3-1-5 Kannondai, Tsukuba, Ibaraki, 305-0856, Japan

    Kumiko Kimura

Authors
  1. Kiyohiko Andoh
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  2. Kumiko Kimura
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  3. Asami Nishimori
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  4. Shinichi Hatama
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Contributions

KA: Conceptualization, methodology, investigation, writing-original draft preparation, visualization. KK: methodology, investigation, writing-reviewing and editing, visualization. AN: methodology, investigation, writing-reviewing and editing. HS: writing-reviewing and editing.

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Correspondence to Kumiko Kimura or Shinichi Hatama.

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The authors declare that they have no competing interests.

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Andoh, K., Kimura, K., Nishimori, A. et al. Development of an in situ hybridization assay using an AS1 probe for detection of bovine leukemia virus in BLV-induced lymphoma tissues. Arch Virol 165, 2869–2876 (2020). https://doi.org/10.1007/s00705-020-04837-7

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  • DOI: https://doi.org/10.1007/s00705-020-04837-7

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