Background. Merkel cell polyomavirus (MCV), a DNA tumor virus, has been found to be associated with Merkel cell carcinoma (MCC) and chronic lymphocytic leukaemia (CLL). MCV sequences have also been detected in various normal tissues in tumor affected patients. Immunologic studies have detected MCV antibodies in as many as 80% of healthy blood donors. This high seroprevalence suggests that MCV infection is widespread in humans. Materials and methods DNA from buffy coats (n=60) of healthy blood donors was investigated by two PCR rounds, 35 cycles each, for two different MCV Tag regions, nt 571-879 and nt 1709-1846, respectively. To quantify the MCV DNA load, positive samples were further analyzed by RQ-PCR for Tag sequences. To determine the human cell equivalents of each sample under analysis, RQ-PCR assays were carried out simultaneously with the cellular RNase P gene. The specificity of PCR amplified products, 10 amplicons from the MCV Tag regions, were DNA sequenced. Results PCR re-amplifications showed that 13 out of 60 (22%) DNA samples were positive for MCV Tag coding sequences. Buffy coats under analysis indicated that the viral DNA copy numbers were very low, ranging from 10 to 100 copies/100,000 cells. The sequencing result of the PCR amplicons, for both strands, nt 571-879 and nt 1709-nt 1846, identified as belonging to the MCV genome, MKL-1 strain. Conclusions MCV sequences were detected in buffy coats from healthy blood donors. This result suggests that MCV is able to infect specific blood leukocyte cells, where it remains in a latent/persistent state in the PBMCs of immune-competent individuals. In the long term, viral persistent infection may allow MCV to generate mutants which can participate in the cell transformation process. Indeed, large T antigen MCV deletion mutants have been detected in CLL or integrated into MCC. This oncogenic process, together with the immune impairment of the host and other factors, is a well-known multistep cell transformation mechanism employed by other DNA tumor viruses, such as human papillomaviruses, which are closely related to the Merkel cell polyomavirus. Acknowledgements We thank Prof. Tobias Allander, Karolinska Instituten, Stockolm, Sweden, for his generous gift of the recombinant plasmid pMCV-LT.1 Reference Pancaldi C, Corazzari V, Maniero S, Mazzoni E, Comar M, Martini F, Tognon M. Merkel cell polyomavirus DNA sequences in the buffy coats of healthy blood donors. Blood 2011, 117:7099-7101.
Merkel Cell Polyomavirus (MCV) in Buffy Coats of Healthy Blood Donors.
PANCALDI, Cecilia;MANIERO, Stefania;MAZZONI, Elisa;MARTINI, Fernanda;TOGNON, Mauro
2011
Abstract
Background. Merkel cell polyomavirus (MCV), a DNA tumor virus, has been found to be associated with Merkel cell carcinoma (MCC) and chronic lymphocytic leukaemia (CLL). MCV sequences have also been detected in various normal tissues in tumor affected patients. Immunologic studies have detected MCV antibodies in as many as 80% of healthy blood donors. This high seroprevalence suggests that MCV infection is widespread in humans. Materials and methods DNA from buffy coats (n=60) of healthy blood donors was investigated by two PCR rounds, 35 cycles each, for two different MCV Tag regions, nt 571-879 and nt 1709-1846, respectively. To quantify the MCV DNA load, positive samples were further analyzed by RQ-PCR for Tag sequences. To determine the human cell equivalents of each sample under analysis, RQ-PCR assays were carried out simultaneously with the cellular RNase P gene. The specificity of PCR amplified products, 10 amplicons from the MCV Tag regions, were DNA sequenced. Results PCR re-amplifications showed that 13 out of 60 (22%) DNA samples were positive for MCV Tag coding sequences. Buffy coats under analysis indicated that the viral DNA copy numbers were very low, ranging from 10 to 100 copies/100,000 cells. The sequencing result of the PCR amplicons, for both strands, nt 571-879 and nt 1709-nt 1846, identified as belonging to the MCV genome, MKL-1 strain. Conclusions MCV sequences were detected in buffy coats from healthy blood donors. This result suggests that MCV is able to infect specific blood leukocyte cells, where it remains in a latent/persistent state in the PBMCs of immune-competent individuals. In the long term, viral persistent infection may allow MCV to generate mutants which can participate in the cell transformation process. Indeed, large T antigen MCV deletion mutants have been detected in CLL or integrated into MCC. This oncogenic process, together with the immune impairment of the host and other factors, is a well-known multistep cell transformation mechanism employed by other DNA tumor viruses, such as human papillomaviruses, which are closely related to the Merkel cell polyomavirus. Acknowledgements We thank Prof. Tobias Allander, Karolinska Instituten, Stockolm, Sweden, for his generous gift of the recombinant plasmid pMCV-LT.1 Reference Pancaldi C, Corazzari V, Maniero S, Mazzoni E, Comar M, Martini F, Tognon M. Merkel cell polyomavirus DNA sequences in the buffy coats of healthy blood donors. Blood 2011, 117:7099-7101.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
