Unit:
Τομέας Υγείας - Μητέρας - ΠαιδιούLibrary of the School of Health Sciences
Dissertation committee:
Χρούσος Γεώργιος
Original Title:
Προσδιορισμός γονότυπου HPV ιού στο δέρμα των παιδιών με τη μοριακή τεχνική άμεσης ανάλυσης αλληλουχιών DNA (Direct Sequencing Analysis). Συσχέτιση γονότυπου με την κλινική προσβολή, τον τρόπο μετάδοσης και την ηλικία.
Summary:
Human Papilloma viruses (HPV) are small double stranded DNA viruses that
comprise a family of more than 150 types. HPVs can be classified into mucosal
and cutaneous types. Mucosal types infect the mucous membrane and can cause
cervical neoplasia in adults as well as anogenital warts in both children and
adults. Cutaneous types infect the squamous epithelium of the skin and produce
common warts, plantar warts and flat warts, which occur commonly on the hands,
face and feet. Skin warts are estimated to occur with a greatest incidence
between 12 and 16 years of age and are usually more frequent in girls than in
boys. The natural progression of the skin warts in childhood indicates that
warts spontaneously clear after 2 years without treatment in 40% of children.
Depending on their location warts can be painful and in that case treatment is
necessary.
The aim of this study is to determine the presence of HPV in warts in children
in order to associate the virus and the disease.
Eighty-four children of clinically diagnosed cutaneous warts were recruited.
The age of our study population ranged between 1 and 18 years. The
participation in the study included a personal history of the subjects which
gave information regarding the health history of a skin disease, personal
habits, clinical findings and the actual type and location of warts. Parents
gave an informed consent for the study.
We studied skin biopsy specimens. DNA was extracted using a commercial kit. In
order to distinguish the different HPV types we used specific pair of primers
in order to amplify the HPV DNA. PCR amplification of the L1 region was
followed by
1. Restriction fragment length polymorphism (RFLP) analysis
2. Sequencing followed by Blast analysis
In some cases due to multiple infection or unknown restriction enzyme pattern
we were not able to detect the HPV type. In those cases but also in the
majority of our samples we developed a novel multiplex human papilloma virus
bead based genotyping assay. Biotinylated PCR products were hybridized to type
specific oligonucleotide probes coupled to beads and analyzed using Luminex
technology.
The genotype results but also the clinical characteristics of our samples were
statistically analyzed using SPSS in order to determine the relation of the HPV
type with environmental risk factors for warts.
Common warts represent the type with the highest prevalence among children in
our study and were present in higher frequency on the exposed parts of the
body. In addition we used a variety of methods for HPV typing and it seems that
the newly introduced Luminex assay maximized the discrimination of genotypes
even in the case of multiple HPV infections. However the utility of the RFLP
methods and sequencing is also of great importance. Last but not least the
results of our study seem to be in concordance with the literature on warts
published thus far. HPV 57 is the predominant type in our study. However the
detection of the high risk HPV type 16 in 33% of our positive samples indicates
the presence of mucosal high risk HPV types in skin of children. It will
require large cross sectional studies and long term follow up to confirm the
association of different HPV types and warts in children.
Keywords:
HPV infection, Warts, Children, Genotyping, Skin
Number of references:
202
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