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Molecular Methods for a Correct Diagnosis of Multiple HPV Infections and Clinical Implications for Vaccine
  1. Andrea Tinelli, MD*,
  2. Giuseppe Leo, PhD,
  3. Domenico Dell'Edera, MD,
  4. Fabio Storelli, PhD,
  5. Maria Maddalena Galante, PhD,
  6. Marcello Guido, PhD§,
  7. Gernot Hudelist, MD, PhD and
  8. Antonio Malvasi, MD
  1. *Department of Obstetrics and Gynaecology,
  2. Molecular Biology and Experimental Oncology Lab, Oncological Hospital, Vito Fazzi Hospital;
  3. Unit of Cytogenetic and Molecular Genetics, "Madonaa delle Grazie" Hospital, Matera, Italy;
  4. §Laboratory of Hygiene, Department of Biological and Environmental Sciences and Technologies, Di.S.Te.B.A., Faculty of Sciences, University of Salento, Lecce, Italy;
  5. Department of Obstetrics and Gynaecology, Wilhelminenspital der Stadt, Vienna, Austria; and
  6. Department of Obstetrics and Gynaecology, Santa Maria Hospital, Bari, Italy.
  1. Address correspondence and reprint requests to Andrea Tinelli, MD, Division of Experimental Endoscopic Surgery, Imaging, Minimally Invasive Therapy and Technology, Department of Obstetrics and Gynaecology, Vito Fazzi Hospital, 73100 Lecce, Italy. E-mail: andreatinelli{at}


Introduction The human papillomavirus (HPV) family is characterized by minimal genotypic differences corresponding to different virus types. The aim of this study was to detect the HPV coinfections and the inner genotype in a series of 336 cervical-vaginal samples.

Methods A total of 336 cervical-vaginal samples were taken from 2007 to 2009 using specific molecular techniques such as molecular sequencing and hybridizations. The genome amplification of the L1 open reading frame was analyzed by real-time polymerase chain reaction; direct sequencing was performed by SYBR green fluorescent molecule and degenerate primers MY09 and MY11. The HPV genotyping was accomplished via oligonucleotide probe hybridization. The phylogenetic correlations in coinfections were analyzed by sequence homology of the L1 genomic region. Identified genotypes were then compared.

Results Human papillomavirus positivity was observed in 125 cases (37.2%), with 21 cases (16.8%) of HPV presence in coinfections. Coinfections involved HPV 16 genotype (8 cases) and HPV 18 (5 cases). The HPV 16 infection was mainly associated with genotypes with a lower-than-broad sequence homology, so the HPV 18 was linked to genotypes represented in the opposite phylogenetic tree.

Conclusions The combined and steady use of diagnostic procedures, such as real-time polymerase chain reaction, molecular hybridization, direct sequencing, and HPV genotyping test, allow accurate diagnosis of monoinfections and coinfections. This may faciliate the development of specific viral tests and prophylactic anti-HPV vaccines.

  • HPV
  • Human papillomavirus
  • Cervical cancer
  • Coinfection
  • Prophylactic vaccine
  • High-risk HPV
  • mRNA
  • Genome amplification
  • Direct sequencing
  • Capillary electrophoresis
  • Real-time PCR
  • HPV genotyping test

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  • Authors certify that there is no actual or potential conflict of interest in relation to this article, and they reveal any financial interests or connections, direct or indirect, or other situations that might raise the question of bias in the work reported or the conclusions, implications, or opinions stated, including pertinent commercial or other sources of funding for the individual authors or for the associated departments or organizations, personal relationships, or direct academic competition.