Dissertations / Theses on the topic 'Papillomavirus diseases'

Background: Persistent infection with high risk HPV is a necessary but insufficient cause of cervical cancer. Behavioural, viral and host factors modulate the risk of HPV persistence. In this thesis, I explore the role of the vaginal microbiota, a host factor and the presence of multiple HPV infections, a viral factor in HPV persistence. Considering the limited data on the epidemiology of HPV related diseases in low and middle-countries (LMIC), and the limited success of cervical cancer screening strategies in many LMIC, I provide data on the distribution of HPV related diseases in Nigeria and evaluate the acceptability of innovative strategies to increase cervical cancer screening uptake. Methods/Results: To achieve my aims, I implemented a longitudinal cohort study of 1,020 women in Nigeria. I begin my results chapters with two methodological papers. Attrition is an important consideration for every longitudinal cohort, particularly in LMIC, therefore, I present my findings on attrition, determinants of attrition and practical strategies to ensure low attrition in studies conducted in LMIC. Considering that sexual behaviour is an important potential confounder in all HPV studies, and the reliability of self-reported history is often questioned, I present findings on the test-retest reliability of self-reported sexual behaviour history collected in my study. Having found that attrition levels were low and that self-reported sexual behaviour history was generally reliable within my cohort, I present my findings on the association between the vaginal microbiota and persistent hrHPV; and the role of multiple HPV infections in viral persistence. I found that the vaginal microbiota was associated with persistent hrHPV in HIV negative women, but not in HIV positive women; and that multiple HPV infections did not increase the risk of viral persistence when compared to single HPV infections. Next, I present my findings on the prevalence and incidence of anogenital warts in Nigeria, with additional reports on the prevalence of cervical cancer and other HPV associated cancers using data from two population based cancer registries. Finally, I present my findings on the acceptability of innovative strategies to improve cervical cancer screening uptake in Nigeria. I found that Nigerian women had a favorable attitude to the use of HPV DNA based screening as part of routine antenatal care, however attitudes towards the use of self-sampling techniques for HPV based cervical cancer screening varied by religious affiliations. Conclusion: In my thesis, I was able to systematically investigate the epidemiology of HPV infections in a LMIC. I considered the distribution of HPV related diseases, host and viral determinants of HPV persistence and investigated control strategies to reduce the burden of cervical cancer in a LMIC. My results provide useful data for surveillance, monitoring and evaluation of control programs on HPV and cervical cancer in Nigeria and may be useful to cervical cancer control programs in other LMIC.

Thesis (M.P.H.)--Georgia State University, 2007.
Title from file title page. Russ Toal, committee chair; Michael Eriksen, Cristen J. Suhr, committee members. Electronic text (76 p. : ill., col. map) : digital, PDF file. Description based on contents viewed Feb. 25, 2008. Includes bibliographical references (p. 66-72).

Thesis (M. Phil.)--Hong Kong University of Science and Technology, 2003.
Includes bibliographical references (leaves 112-129). Also available in electronic version. Access restricted to campus users.

Thesis (MTech (Biomedical Technology))--Cape Technikon, Cape Town, 2002
Infection \\'ith specific high risk human papilloma"iruses (HPV) has been shown to play a causal role in the development of ceJVical intraepithelial neoplasia (CIN) and cenical cancer in women. The development of a prophylactic vaccine to immull.ize women against HPV infection would play a \'ita! role in protecting women against HPV infection and ultimately ceMcal cancer. Despite cancer of the cer\'ix being the second most common cancer in South African women, a literature search reveals that few studies have been performed in South Africa on the types of HPV prevalent in women with CIN or cancer ofthe ceMx. HPVs that infect the anogenital tract have also been shown to infect the oral ca\'ity. However, the HPV prevalence rates vary greatly between studies and the significance of the presence ofHPV in the oral ca\'ity is still not understood. The primary objectives of this study were to establish the HPV prevalence rate infecting women with CIN lesions using a sensitive nested polymerase chain reaction (PCR) and to develop a novel restriction fragment length polymorphism (RFLP) method to type the high risk mucosal HPVs detected in these women. The secondary objective of this study was to establish the prevalence rate and HPV types infecting the oral mucosa of women with CIN lesions and to compare these HPV types with those detected in the ceMx. Cemcal punch biopsies were taken from 163 women with CIN lesions and buccal cells were collected from 33 of these participants. DNAwas extracted from the biopsies and buccal samples and PCR using CCRS primers performed to ensure sample adequacy. Nested PCR usmg consensus degenerate primers for HPV was performed on all samples sho\\'wg sufficient amplifiable DNA A novel restriction fragment length pol)morphism (RFLP) method was developed to identify the 10 high risk mucosal HPVs considered human carcinogens of group 1 by the International Agency for Research on Cancer (lARC) as well as HPV 11 which is commonly found in the oral cavity.

The ability of the HPV-18 E6 gene to impair p53-mediated transcriptional activity induced by DNA damaging agents was investigated. It is demonstrated that E6 can abolish DNA damage induced p53-mediated transcription and that a region from amino acid residue 113 to 117 of HPV-18 E6 protein was necessary for E6 to direct the degradation of p53. The biological importance of the E6/p53 interaction was then directly examined in HPV-16 containing cervical carcinoma derived cells by introducing the monomeric p53 mutant which is resistant to E6 mediated degradation. The two major observations made from this study were: (i) loss of p53 activity plays an important role in maintaining the malignant phenotype of these cells with respect to cell proliferation; (ii) the monomeric p53 mutant without its C-terminal regulatory region was biologically functional with respect to impairing cell proliferation in HPV-16 containing cervical carcinoma derived cells. Finally, it was revealed that the cellular MAP kinase signal transduction pathway was more active in cells expressing the HPV-16 E5 gene than in control cells or cells expressing E6 and E7. These observations help to define the mechanisms by which HPV oncogenes contribute to the development and maintenance of the neoplastic phenotype.

Objectives To understand the acceptability of adult males, high-risk male population (MSM and bisexual men) and parents of adolescent sons and explore factors correlated with HPV vaccination acceptability Methods A systematic searching process for literatures related to men’s HPV vaccination acceptability and published from 2000 to July, 2013 in PubMed, MEDLINE, CINAHL, Cochrane Library and Google Scholar was performed. After screening based on inclusion and exclusion criteria, qualities of all eligible studies were assessed based on the modified STROBE guideline. Results Of 15 studies were included in this systematic review, 6 focused on adult males, 4 explored the high-risk males, five reported the parental acceptability. The HPV vaccination acceptability of adult males, high-risk male population and parents of adolescent sons was moderate or high in most reviewed studies. Knowledge about HPV and HPV vaccination, perceived susceptibility, perceived benefits and healthcare provider’s recommendation were positively correlated with HPV vaccination acceptability among adult males, high-risk males and parents of adolescent sons while high expense, side effects, safety, uncertain effectiveness and hassle of receiving a 3-shots series of HPV vaccination could diminish people’s vaccination interest. Conclusion HPV vaccination acceptability among adult males, high-risk males and parents of adolescent sons is moderate or high. Further HPV vaccine campaign should focus on bridging the gap between the high vaccination acceptability and the low vaccination uptake among males.
published_or_final_version
Public Health
Master
Master of Public Health

Human Papillomavirus (HPV) is the cause of cervical cancers (among these, adenocarcinoma, AdCa) and is associated to a subgroup of oropharyngeal carcinomas (OPSCCs). Even if the risk for cancer development is linked to the infection by some viral genotypes, mainly HPV16 and 18, viral DNA alone seems not to be sufficient for diagnosis. Moreover, the role of the virus in OPSCCs has not been totally clarified yet. In the first part of the thesis, the performances concerning viral genotyping in clinical cervical samples of a new pyrosequencing-based test and a well-known hybridization-based assay have been compared. Similar results between the methods have been obtained. However, the former showed advantages in detecting intratype variants, higher specificity and a broader spectrum of detectable HPV types. The second part deals with the evaluation of virological markers (genotyping, viral oncoproteins expression, viral load, physical state and CpG methylation of HPV16 genome) in the diagnosis/prognosis of cervical AdCa and HPV-associated OPSCCs. HPV16 has been confirmed the most prevalent genotype in both the populations. Interestingly, the mean methylation frequency of viral DNA at the early promoter showed the tendency to be associated to invasion for cervical AdCa and to a worse prognosis for OPSCCs, suggesting a promising role as diagnostic/prognostic biomarker. The experiments of the third part were performed at the DKFZ in Heidelberg (Germany) and dealt with the analysis of the response to IFN-k transfection in HPV16-positive cervical cancer and head&neck carcinoma cell lines to evaluate its potential role as new treatment. After 24h, we observed increased IFN-b expression which lead to the up-regulation of genes involved in the antigens presentation pathway (MHC class I and immunoproteasome) and antiviral response as well, in particular in cervical cancer cell lines. This fact suggested also the presence of different HPV-mediated carcinogenic pathways between the two anatomical districts.
Il Papillomavirus umano (HPV) è causa dei carcinomi della cervice uterina (tra cui adenocarcinomi, AdCa) ed è associato ad un sottogruppo di tumori dell’orofaringe (OPSCCs). Nonostante il rischio di sviluppo di tumore sia associato all’infezione da parte di alcuni genotipi virali, principalmente HPV16 e 18, il DNA virale da solo sembra non essere sufficiente in campo diagnosico. Inoltre, per tumori orofaringei il ruolo del virus non è ancora del tutto chiaro. Nella prima parte della tesi, sono state confrontate le performance riguardo la genotipizzazione di HPV su campioni clinici cervicali di una tecnica innovativa, basata su amplificazione e pirosequenziamento, e una di routine, basata su amplificazione e ibridazione inversa. Lo studio ha evidenziato performance simili tra le due metodiche, sottolineando per il sequenziamento una maggiore specificità e capacità di rilevare varianti intratipo. Nella seconda parte sono stati analizzati marker virologici (genotipizzazione, espressione delle oncoproteine virali, carica virale, stato fisico e metilazione del genoma di HPV16) in funzione dei dati clinici disponibili, per un possibile impiego nella diagnosi/prognosi di AdCa cervicali e OPSCCs HPV-associati. HPV16 si è confermato il genotipo prevalente in entrambe le popolazioni. La frequenza di metilazione nel promotore precoce virale ha mostrato una tendenza ad essere associata ad invasione negli AdCa, e ad una prognosi peggiore negli OPSCCs, emergendo come il più promettente marker diagnostico/prognostico. La terza parte, svolta presso il DKFZ di Heidelberg (Germania), ha visto l’analisi della risposta alla transfezione di IFN-k in linee cellulari tumorali HPV16-positive della cervice uterina e della regione testa-collo, per valutarne l’impiego terapeutico. Dopo 24h, è stato osservato un incremento dell’espressione di IFN-b e, di conseguenza, una up-regolazione dei geni coinvolti nella presentazione antigenica (MHC classe I ed immunoproteasoma) e nella risposta antivirale, specialmente nelle cellule cervicali, suggerendo la presenza di diversi meccanismi patogenetici tra tumori HPV-positivi dei due distretti anatomici.

Human Papillomavirus (HPV) is the cause of cervical cancers (among these, adenocarcinoma, AdCa) and is associated to a subgroup of oropharyngeal carcinomas (OPSCCs). Even if the risk for cancer development is linked to the infection by some viral genotypes, mainly HPV16 and 18, viral DNA alone seems not to be sufficient for diagnosis. Moreover, the role of the virus in OPSCCs has not been totally clarified yet. In the first part of the thesis, the performances concerning viral genotyping in clinical cervical samples of a new pyrosequencing-based test and a well-known hybridization-based assay have been compared. Similar results between the methods have been obtained. However, the former showed advantages in detecting intratype variants, higher specificity and a broader spectrum of detectable HPV types. The second part deals with the evaluation of virological markers (genotyping, viral oncoproteins expression, viral load, physical state and CpG methylation of HPV16 genome) in the diagnosis/prognosis of cervical AdCa and HPV-associated OPSCCs. HPV16 has been confirmed the most prevalent genotype in both the populations. Interestingly, the mean methylation frequency of viral DNA at the early promoter showed the tendency to be associated to invasion for cervical AdCa and to a worse prognosis for OPSCCs, suggesting a promising role as diagnostic/prognostic biomarker. The experiments of the third part were performed at the DKFZ in Heidelberg (Germany) and dealt with the analysis of the response to IFN-k transfection in HPV16-positive cervical cancer and head&neck carcinoma cell lines to evaluate its potential role as new treatment. After 24h, we observed increased IFN-b expression which lead to the up-regulation of genes involved in the antigens presentation pathway (MHC class I and immunoproteasome) and antiviral response as well, in particular in cervical cancer cell lines. This fact suggested also the presence of different HPV-mediated carcinogenic pathways between the two anatomical districts.
Il Papillomavirus umano (HPV) è causa dei carcinomi della cervice uterina (tra cui adenocarcinomi, AdCa) ed è associato ad un sottogruppo di tumori dell’orofaringe (OPSCCs). Nonostante il rischio di sviluppo di tumore sia associato all’infezione da parte di alcuni genotipi virali, principalmente HPV16 e 18, il DNA virale da solo sembra non essere sufficiente in campo diagnosico. Inoltre, per tumori orofaringei il ruolo del virus non è ancora del tutto chiaro. Nella prima parte della tesi, sono state confrontate le performance riguardo la genotipizzazione di HPV su campioni clinici cervicali di una tecnica innovativa, basata su amplificazione e pirosequenziamento, e una di routine, basata su amplificazione e ibridazione inversa. Lo studio ha evidenziato performance simili tra le due metodiche, sottolineando per il sequenziamento una maggiore specificità e capacità di rilevare varianti intratipo. Nella seconda parte sono stati analizzati marker virologici (genotipizzazione, espressione delle oncoproteine virali, carica virale, stato fisico e metilazione del genoma di HPV16) in funzione dei dati clinici disponibili, per un possibile impiego nella diagnosi/prognosi di AdCa cervicali e OPSCCs HPV-associati. HPV16 si è confermato il genotipo prevalente in entrambe le popolazioni. La frequenza di metilazione nel promotore precoce virale ha mostrato una tendenza ad essere associata ad invasione negli AdCa, e ad una prognosi peggiore negli OPSCCs, emergendo come il più promettente marker diagnostico/prognostico. La terza parte, svolta presso il DKFZ di Heidelberg (Germania), ha visto l’analisi della risposta alla transfezione di IFN-k in linee cellulari tumorali HPV16-positive della cervice uterina e della regione testa-collo, per valutarne l’impiego terapeutico. Dopo 24h, è stato osservato un incremento dell’espressione di IFN-b e, di conseguenza, una up-regolazione dei geni coinvolti nella presentazione antigenica (MHC classe I ed immunoproteasoma) e nella risposta antivirale, specialmente nelle cellule cervicali, suggerendo la presenza di diversi meccanismi patogenetici tra tumori HPV-positivi dei due distretti anatomici.

Human papillomavirus (HPV) is a double-stranded DNA virus, typically infecting mucosal or cutaneous epithelial keratinocytes. Today, more than 118 different HPV types have been formally described. Sexual transmission of mucosal HPVs is very common and generally asymptomatic, but HPV infection can be associated with benign lesions such as condylomata acuminata or, in rare cases, with malignant lesions such as cervical cancer. Two prophylactic vaccines are currently available in Europe, protecting against HPV-16 and HPV-18 (Cervarix&63720;) or against HPV-6, HPV-11, HPV-16 and HPV-18 (Gardasil&63720;). In order to assess the impact of the vaccination program, it is mandatory to obtain geographically widespread date on the baseline HPV prevalence and type distribution in cervical samples from women, presenting or not, normal or abnormal cytologic or histologic results. We undertook an epidemiological study in the Capital Region of Brussels to determine the HPV prevalence and type-distribution in 1526 cervical samples of women presenting a cytology within normal limits (WINL), atypical squamous cells of undetermined significance (ASC-US), low-grade squamous intra-epithelial lesions (LSIL), high-grade squamous intra-epithelial lesions (HSIL) or invasive cervical cancer (ICC). The HPV prevalence was 10.8% (95%CI: 8.8-12.8) for NILM, 34.5% (95%CI: 28.3-40.8) for ASC-US, 54.0% (95%CI: 47.4-60.6) for LSIL and 100% for HSIL and ICC. With an HPV-16 and HPV-18 prevalence of 63.3% (95%CI: 44.1-67.7) and 73.5% (95%CI: 63.0-84.0) in mono-infected HSIL and ICC, respectively, HPV 16/18 L1 VLP vaccines would be expected to significantly reduce the management and treatment of women suffering from HSIL and ICC in the Capital Region of Brussels. We also detected HPV-30, HPV-53, HPV-66 and HPV-68 in mono-infected HSIL and ICC samples, possibly providing arguments for the reconsideration of the carcinogenicity of these types.

Vertical transmission of HPV was also previously reported, but in most cases one could not exclude a placental contamination by HPV positive cells from an infected birth canal. In order to confirm that the placenta can be infected with HPV, we analysed residual cells from 35 transabdominally obtained placental samples from pregnant women undergoing chorionic villous sampling for screening of suspected foetal abnormalities and found that two samples were positive for HPV-16 and HPV-62, respectively. The clinical importance of these results remains to be elucidated, but the previously observed association between placental HPV infection and pregnancy loss might gain further in importance. HPV gene regulation in placental trophoblastic cells has not been studied so far. We studied the HPV-16 early gene expression regulation in transiently transfected monolayer cultured trophoblastic cells with an HPV-16 long control region (LCR) driven reporter plasmid. We observed important differences in constitutive HPV-16 LCR activities between trophoblastic cell lines and could identify progesterone as an important inducer of HPV-16 early gene expression. Steroid hormones are induced during pregnancy and could therefore lead to an enhanced expression of the E5, E6 and E7 proteins upon placental HPV infection. Since these proteins were previously shown to affect trophoblast adhesion, survival, migration and invasion, their enhanced expression might eventually contribute to pregnancy loss. We furthermore found that the transcription of episomally maintained HPV-16 is not regulated by E2 or E1, but by E5, E6 and/or E7.

Doctorat en Sciences biomédicales et pharmaceutiques
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Les infections génitales par les virus du papillome humains (HPV) sont les infections virales sexuellement transmises, les plus communes chez les femmes en âge de procréer. Il est désormais bien établi que l’infection persistante par les HPV classés «à haut risque» est l’un des facteurs indispensables au développement de lésions précancéreuses et cancéreuses du col de l’utérus. Ces HPV semblent aussi être impliqués dans le développement d’autres cancers de la région ano-génitale et pourraient être également impliqués dans les cancers de la tête et du cou. Durant cette dernière décennie, des études croissantes tendent à établir un rôle étiologique des HPV dans les dysfonctionnements gestationnels. La détection des ADN HPV dans les placentas issus d’avortements spontanés et leur capacité exceptionnelle à se répliquer in vitro dans les cellules trophoblastiques cultivées en monocouche, ont apporté de nouvelles perspectives quant à la possibilité que le placenta pourrait constituer aussi un tropisme naturel des infections par HPV.

Six jours après la fécondation et suite à l’accolement du blastocyste à l’épithélium utérin, le trophoblaste s’engage dans des processus actifs de prolifération, d’invasion et de différenciation complexe pour la construction de l’interface physiologique indispensable aux échanges essentiels entre la mère et l’enfant ;le placenta. De façon intéressante, ses propriétés sont similaires à celles de la cellule tumorale maligne. Néanmoins, ses mécanismes sont étroitement régulés dans le trophoblaste, à la fois dans l’espace et le temps, assurant un développement normal à chaque étape de la grossesse.

Devant toutes ces données, nous avions émis l’hypothèse que l’expression des protéines précoces E5, E6 et E7 d’HPV de type 16 (de haut risque), pourraient modifier le développement des trophoblastes infectés. Les résultats obtenus durant ce travail de doctorat démontrent que la protéine virale E5, hautement hydrophobe, est cytotoxique et affecte la viabilité du trophoblaste. Cette cytotoxicité est neutralisée, et la viabilité est améliorée, lorsque les oncoprotéines majeures E6 et E7 sont exprimées en présence de la protéine E5. Lorsque toutes les protéines précoces sont exprimées sous le contrôle de leur propre promoteur (LCR), la viabilité est favorisée. Ces observations ont été confirmées dans les cellules cervicales également. Il a été précédemment rapporté que les oncoprotéines E6 et E7 affectaient l’adhésion du trophoblaste aux cellules endométriales. Dans le présent travail, il a été retrouvé que la protéine E5 diminuait elle aussi l’adhésion, non seulement aux cellules endométriales, mais aussi au support de culture cellulaire. Les capacités de migration et d’invasion de la matrice extracellulaire sont augmentées par l’expression de E5 et dans une plus large proportion par l’expression de E6 et E7. Des résultats similaires ont été obtenus lorsque toutes les protéines de la région précoces sont exprimées sous le contrôle de leur propre promoteur (LCR). La diminution de l’expression de la E-cadhérine est considérée comme un marqueur de malignité et de mauvais pronostic pour les cancers. Nous avons démontré que l’expression de E5, E6 ou de E7, inhibait l’expression de la E-cadhérine, reflétant l’impact des oncoprotéines du virus HPV-16 sur la diminution de l’adhésion et l’augmentation du pouvoir invasif des cellules trophoblastiques. L’investigation d’autres marqueurs de malignité et de tolérance immunitaire, l’étude de l’impact du virus HPV-6 (de bas risque) sur la migration et l’invasion des cellules trophoblastiques, et l’étude de la capacité des protéines précoces d’HPV-16 à influencer l’entrée des particules virales, ont fait l’objet de résultats préliminaires, ouvrant de larges perspectives.

Genital Human Papillomavirus (HPV) infections are the most common sexually transmitted infections amongst women on the age of reproduction. It is well established that persistent infection with high-risk HPVs is the necessary factor in the causation of precancerous and cancerous cervical lesions. High-risk HPVs have also been reported to be involved in the causation of head and neck cancers and other anogenital cancers. On this last decade, growing data are attempting to study the potential etiological association of HPV with gestational dysfunctions. The detection of HPV DNA in placentas resulting from spontaneous abortions and the unique ability of multiple HPV types to replicate in vitro in trophoblastic cells cultured in a monolayer system, rise new questions over the HPV tropism.

Six days following fertilization and once the apposition of the blastocyst on the uterine wall takes place, the trophoblast, in a very active and complex process, starts to proliferate, invade and to differentiate in order to build a physiological interface; the placenta, from where multiple mother/foetus exchanges occur. Interestingly, the way that the trophoblast behaves is very similar to malignant tumoural cells. However, the trophoblast obeys to strict spatial-temporal regulatory confines, insuring a proper development all along the pregnancy.

In regard to these data, we hypothesised that the expression of the high-risk HPV type 16 oncoproteins E5, E6 and E7, might modify the development of the infected trophoblast. During my Ph.D study, I demonstrated that the highly hydrophobic protein E5 is localized in many interne membranes compartments of the transfected trophoblast. E5 affects the viability of transiently and stably transfected trophoblastic cells. E6 and E7, favouring cell growth, neutralised the E5 cytotoxic effect. All HPV-16 early proteins, when expressed under the control of their endogenous promoter (LCR), favoured trophoblastic growth. These observations were also observed in cervical cell lines. In addition, E5 decreased the adhesiveness of trophoblastic cells to the tissue culture plastic and to endometrial cells similarly as previously described for E6 and E7. Cells expressing E6, E7 and in less extend E5 favoured chemotaxic migration and matrigel invasion compared to the cells expressing the LacZ control. These effects were also observed when early proteins were expressed under the control of their own viral promoter (LCR). Interestingly, the E-cadherin was down regulated in trophoblastic cells expressing E5, E6 and E7. In conclusion, HPV-16 early proteins enhanced trophoblastic growth and intensify the malignant phenotype by impairing cell adhesion leading to increased cellular motile and invasive properties. HPV-16 E5 participated, with E6 and E7, in these changes by impairing E-cadherin expression, a hallmark of malignant progression. Additional preliminary results consisting on the investigation of other markers of malignancy and immune tolerance, on studying the impact of the low-risk HPV type 6 early proteins on the migratory and invasive properties of trophoblastic cells and on the study of the ability of HPV-16 to influence the entry of virus particules, allowed to open wide perspectives.

Doctorat en Sciences
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An association between persistent Human papillomavirus (HPV) infection in women and cervical cancer has been established. Young women are particularly at risk of acquiring sexually transmitted infections such as HPV because of risky sexual activity and physiological immaturity. While at risk though, young women have been shown to be amenable to health promoting initiatives. There are a small number of international studies concerning adolescent HPV infection and the risk factors associated with infection, but there is currently no evidence on the prevalence and risk factors for HPV in an Australian, sexually active female adolescent population. This study aimed to provide evidence of the prevalence of HPV, risk factors associated with infection and the patterns of sexual activity in a female sexually active, senior high school population in the Australian Capital Territory. Participants in this study were a convenience sample of 161 sexually active 16-19 year old females who had an HPV test who were attending a senior high school in the Australian Capital Territory. Nurses and doctors using a clinical record collected information about sexual and other risk behaviours. Self-obtained vaginal swabs were tested for HPV DNA using the polymerase chain reaction method and genotyping was undertaken. The HPV prevalence in this cohort of young women was 1 1.2%. High-risk genotypes were found in 55.5% and multiple genotypes were found in 38.8%. There was a significant association found between HPV infection and having had more than one male partner with whom vaginal intercourse had occurred in the previous six months. No statistically significant association was found between HPV and the age of coitarche, length of time young women had been sexually active, condom use, and smoking or alcohol intake. A young age at coitarche was common for this group. Smoking and alcohol use was seen in large proportions in this group. This is the first Australian study that has examined the prevalence and risk factors for genital HPV in this demographic group. The HPV prevalence is lower than in international studies in comparable groups, in similar age groups and much lower than in older women both in Australia and overseas. With the comparatively low prevalence comes an opportunity for important public health interventions for this group including routine Pap smears, vaccination against the high-risk types of HPV when this becomes available and strategies for young women to reduce their number of male sexual partners. A substantial amount of young women in this study were sexually active aged under 16 years. Whilst this was not identified as being a risk factor in this study, it is both a health and personal safety issue for these young women. There is a demonstrated need for health promotion strategies for this cohort about the consumption of safe levels of alcohol and for smoking cessation. Further research is recommended that includes a repetition of this study with a larger sample, the use of a prospective study design to identify trends in infection and examination of HPV prevalence and risk factors for a variety of populations.

Persistent infection of the cervix with high risk (HR) types of Human Papilloma Virus (HPV) (HR-HPV) can result in precancerous lesions and cancers. However, most HPV infections can be cleared naturally by the immune response without causing disease. Although genetic variations have long been considered as the main explanation for individual heterogeneity in cancer susceptibility, the underlying mechanisms remain unclear. In this project, a panel of routinely taken clinical samples was assessed for 32 rationally selected SNPs with allele frequency related to disease outcome using the Taqman® OpenArray® system. The panel incorporated 475 HR-HPV negative, cytologically-normal cervical samples, 413 HR-HPV positive cervical high grade squamous intraepithelial lesion (HSIL) cases and 62 HR-HPV positive cervical cancers. Two SNPs, rs2234671 and rs2623047, were found with significant differences between HR-HPV negative, cytologically-normal samples and HR-HPV positive cervical HSIL cases. In the validation step, these two SNPs were further genotyped in the same set of samples using TaqMan® SNP genotyping assay and/or LightSNiP assay and in additional samples including 83 HR-HPV positive, cytologically-normal cervical samples, 21 HR-HPV positive cervical cancer cases, 129 HR-HPV positive vulval intraepithelial neoplasia cases and 23 HR-HPV positive vulval cancer cases. Statistical analysis was then performed based on pooled and re-grouped genotyping data of the above-mentioned samples under different genetic models so as to evaluate the associations with different stages in the disease process. After validation, SULF1 rs2623047 revealed a strong significant association with the susceptibility to HR-HPV infection but not with the development of high-grade squamous intraepithelial lesion and the progression to cervical cancer. CXCR1 rs2234671, by contrast, was associated with the progression of HR-HPV-related cancers and the minor allele CXCR1 827C was significantly enriched in HPV16 positive cancers. CXCR1 is a receptor for the chemokine CXCL8/IL-8 and CXCR1 rs2234671 leads to a serine to threonine change in an extracellular loop of the receptor. Functionally, the CXCR1 827C allele was shown to enhance cell motility in response to IL-8 stimulation in a chemotaxis assay with transiently transfected fibroblasts (HEK293 cells) and also in a wound healing assay with stably transduced cervical cancer (CaSki) cells. In addition, significantly increased cell proliferation upon IL-8 treatment was observed in two cervical cancer derived cell lines, CaSki and SiHa, transduced with CXCR1-827C allele, but not in their CXCR1 827G transduced counterparts. These findings suggest that SULF1 rs2623047 and CXCR1 rs2234671 may be genetic risk factors for HR-HPV-related cervical disease and CXCR1 rs2234671 might affect HR-HPV-related cancer susceptibility by functionally altering IL-8-CXCR1 signalling. This information has potential for use in the risk stratification of HR-HPV infected women and may also suggest new therapeutic targets to be exploited for treatment of cervical cancer patients.

Low risk Human Papillomaviruses (LR-HPVs) 6 and 11 are the main causative agents of benign proliferative lesions such genital warts (GWs) and recurrent respiratory papillomatosis (RRP) (Aubin et al. 2008; Ball et al. 2011; Garland et al. 2009; Prétet et al. 2008). High Risk HPV (HR-HPV) 16 is the most oncogenic type and is responsible for invasive cancers (IC) of cervix (ICC), vulva (IVuC), vagina (IVaC), penis (IPeC) and anus (IAnC) (Alemany et al. 2014, 2015, 2016; Larsson et al. 2013; de Sanjose et al. 2010). The well-established connection between HPV16 infection and IC is observed for the most prevalent ICC histological presentations namely squamous cell carcinomas (SCC), adenocarcinomas (ADC) and adenosquamous cell carcinomas (ADSC) (de Sanjose et al. 2010). All these three HPVs (HPV6, HPV11 and HPV16) are the most prevalent in their associated pathological outcomes (Alemany et al. 2014, 2015, 2016; Aubin et al. 2008; Ball et al. 2011; Garland et al. 2009; Larsson et al. 2013; Prétet et al. 2008; de Sanjose et al. 2010). At level of HPV variants, previous studies have addressed differential HPV6 and HPV11 lineage distributions in GWs and RRP, but mainly at a national or regional level (Kocjan et al., 2009). Although without large sample size, some studies describe differential prevalence of HPV variants among distinct pathologies (Danielewski et al. 2013; Jelen et al. 2014). For HPV6, it has been observed a higher presence of HPV6_B1 variants in GWs compared to RRPs (Flores-Díaz et al. 2017b). HPV16 variant distribution has been mainly focused on the uterus cervix and less on other anatomical sites (Cornet et al. 2012; Yamada et al. 1997), what emphasizes the still wanting research of HPV16 viral lineages in other anogenital cancer location (non-cervical cancers). Some studies show that HPV16 variants in anogenital cancers are largely the same regardless of cancer anatomical locations (ICC, IVuC, IVaC, IAnC and IPeC) , showing increased prevalence of HPV16 A1-3 for all IC of squamous nature (de Koning, Quint, and Pirog 2008; Larsson et al. 2013; Ouhoummane et al. 2013; Tornesello et al. 2008; Zuna et al. 2011). HPV16 variants have been widely studied in SCC (Cornet et al. 2012; Zuna et al. 2011), as this histological type remains the most prevalent ICC (Vinh-Hung et al. 2007; Vizcaino et al. 2000). Nonetheless, data available is poorer for other cervical cancer histological presentations such ADC and ADSC. Although few studies had addressed other glandular histologies, they had been restricted to local geographic origin or small populations (Burk et al. 2003; Lizano 2006; Qmichou et al. 2013; Tornesello et al. 2011). Previously described background, shows an increased prevalence of HPV16_A1-3 variants in SCCs (Zuna et al. 2011) while an enhanced presence of HPV16_D in ADCs (Burk et al. 2003; Mirabello et al. 2016; Quint et al. 2010). Regarding HPV16 intratype variability, it has grown the interest of researchers on the T350G E6 polymorphism as data shows an increased oncogenic potential of those variants containing the 350G allele (Grodzki et al. 2006; Jackson et al. 2016; Zehbe et al. 1998, 2001). Finally, most of the data published do not show results regarding age at tumour diagnosis or use different age definitions such age at enrollment (Mirabello et al. 2016). According to the HPV variant context, the works presented in this thesis try to provide all together information about LR-HPV6 and 11 and HR-HPV16 variant diversity among different lesions, cancers and among different geographical regions with a worldwide spectrum, analyzing large number of HPV monoinfected samples, strategy that prevents possible added bias introduced by co-infections or multi-infections. Through these manuscripts we present for the first time the relative contributions of variant differential abundance, geographical origin, cancer anatomical locations and IC histological presentations to the observation of differential prevalence distribution of HPV16 variants. Through our cases data, we show concordant results with previous published works, observing an increased prevalence of HPV6 B1 variants in GWs, of HPV16 A1-3 variants in anogenital cancers of squamous nature and of HPV16 D variants in ADC. We further show determinate geographical structure of HPV16 variants largely based on the dominance of HPV16 A1-3 variants in Europe, the virtually exclusive presence of HPV16 B and C variants in Africa, the increased prevalence of HPV16 A4 variants in Asia and the enrichment of HPV16 D variants in the Americas. For the most oncogenic-related polymorphism, the E6- T350G, we further show different allelic frequencies according to geographical location independently of the anogenital cancer analyzed, revealing an enhanced 350G allele frequency in isolates from Central-South America compared with Europe. Additionally, we confirme the worldwide trend of cervical cancers to be diagnosed significantly earlier than other anogenital cancers and at histological level, we further present that ADC are diagnosed earlier (mid-forties) than SCC (mid-fifties). According to the previous context, our results complement and may expand those communicated. The current data suggests that the outcome of the virus-host interaction depends on the combination of phylogeny (i.e. the individual genetic background of both virus and patient) and ontogeny (i.e. the differential susceptibility of different tissues) and for HPV16, provide integrating knowledge of variant-specific differential risk that may impact the future screening algorithms, helping to ensure proper early detection of, for example, elusive ADCs. Furthermore, our data emphasizes the necessity of developing deep analyses in HPV variant field. Additionally, the monitoring of initial steps in viral colonization of anogenital mucosas and the follow-up of differential viral persistences according to patient genetic background, may be of remarkable importance. All these research might ultimately provide answers about the extent of the differential fitness of HPV viral lineages and will help to understand the virus-host interplay for the most oncogenic HPVs. Additionally, in vaccinated women follow-up, tracing HPV variants may need to keep more attention as, although not described any evidence, they may be indeed, a possible causative agent of not expected pathological outsiders.
Descripción de la distribución diferencial de variantes de los virus del Papiloma humano 6 y 11 (VPH6 y VPH11) en sus patologías asociadas: verrugas genitales y papilomatosis respiratória recurrente. Descripción de la distribución diferencial de variantes del virus del Papiloma humano 16 (VPH16) en cánceres anogenitales humanos y en los distintos tipos hisológicos de cancer de cérvix (escamoso, adenoescamoso y adenocarcinoma). Para las variantes de VPH16, se decribe su distribución geográfica a nivel mundial y se cuantifica la contribución relativa de la distinta abundancia de las variantes de VPH16, localización anatómica del cáncer,histología y geografía respecto a la observación diferencial de las variantes de VPH16

Thesis (MSc (Microbiology))--Stellenbosch University, 2011.
ENGLISH ABSTRACT: The link between infection with Human Papillomavirus (HPV) and the development of cervical cancer has been established by several epidemiology studies. Cervical cancer is the second most common cancer among women and it occurs at a rate of 22.8 cases per 100 000 women in South Africa. Approximately 86% of newly reported cases of cervical cancer occur in developing countries where limited access to medical facilities hampers efforts to prevent and screen for HPV infection. Two commercial virus-like particle (VLP) vaccines consisting of HPV major structural protein L1, which protect against the most common high-risk HPV-types, are currently available. The high cost and type specificity of these commercially available vaccines have necessitated the development of a low cost, broad-spectrum HPV vaccine. Inclusion of the minor structural protein L2 has been shown to induce broadly cross-neutralizing antibodies and therefore a chimera was constructed that contains an epitope of L2 inserted within the L1 sequence. This construct, renamed SAF, was shown to be highly immunogenic and thus has the potential to be used as a prophylactic cervical cancer vaccine. Methylotrophic yeasts are known to be excellent producers of recombinant proteins due to their strongly inducible promoters that allow culturing of these yeasts to very high cell densities. Pichia pastoris and Hansenula polymorpha have been employed in several studies for heterologous protein production and levels of protein higher than 1 g/L have been reported. These yeasts also have GRAS status and can therefore be used to manufacture products for use in humans. In this study, the potential of H. polymorpha and P. pastoris to produce SAF intracellularly was evaluated. The effect of increased gene dosage and peroxisomal targeting on SAF production was examined as possible strategies to increase the yield of SAF. Peroxisomal targeting was achieved by fusing the SAF gene at the C-terminal end with the Peroxisomal Targeting Sequence 1 (PTS1) which consists of a short tri-peptide: –SKL. The functionality of PTS1 was confirmed using green fluorescent protein (GFP), fluorescence microscopy and peroxisome isolation. Peroxisomal targeting was shown to have a negative effect on SAF production levels in both H. polymorpha and P. pastoris. An increase in gene dosage had no discernable effect on SAF yield in H. polymorpha which is in contrast to previous research. The highest production levels were achieved by P. pastoris KM71 (24.86 mg/L) which compares well to levels of L1 achieved by other research groups. The most significant insight emerging from this work was that all the strains that produced SAF at detectable levels were equally efficient at the production of SAF. Increased biomass was therefore the biggest contributor to high SAF levels (mg/L) in the P. pastoris strains as significantly higher cell densities were achieved during culturing of these strains. With the necessary optimisation, the methylotrophic yeasts have the potential to be used as hosts for the production of a broad-spectrum HPV vaccine.
AFRIKAANSE OPSOMMING: Die skakel tussen infeksie met Mens Papilloomvirus (HPV) en die ontwikkeling van servikale kanker is deur verskeie epidemiologiese studies bevestig. Servikale kanker is die tweede mees algemene kanker onder vroue en dit kom voor teen ‘n tempo van 22.8 gevalle per 100 000 vroue in Suid Afrika. Ongeveer 86% van alle nuwe gevalle kom voor in ontwikkelende lande waar beperkte toegang tot mediese fasiliteite pogings om HPV infeksie te voorkom en te behandel, belemmer. Twee pseudovirale-partikel (VLP) entstowwe teen HPV is tans op die mark beskikbaar en hierdie entstowwe verleen immuniteit teen die mees algemene hoë-risiko HPV tipes. Die hoë koste en nou spektrum van hierdie entstowwe het dit nodig gemaak om ‘n goedkoop, wye-spektrum HPV entstof te ontwikkel. Navorsing het bewys dat die insluiting van die strukturele L2 proteïen in die VLP entstof, lei tot die indusering van neutraliserende teenliggame, wat wye spektrum antigenisiteit tot gevolg het. ‘n Chimeriese proteïen wat ‘n epitoop van L2 binne die L1 volgorde bevat is gekonstrueer, en hierdie proteïen is benoem SAF. SAF het hoë immunogenisiteit en kan dus potensieel as ‘n voorkomende servikale kanker entstof gebruik word. Metielotrofiese giste is bekend vir hulle vermoë om hoë vlakke rekombinante proteïene te produseer as gevolg van hulle induseerbare promotors wat groei tot baie hoë sel digthede toelaat. Pichia pastoris en Hansenula polymorpha is in menigte studies gebruik om heteroloë proteïene te produseer tot vlakke bo 1 g/L. Hierdie giste en die proteïen produkte wat hulle vorm word algemeen aanvaar as veilig vir menslike gebruik. In hierdie studie het ons die potensiaal van H. polymorpha en P. pastoris om SAF intrasellulêr te produseer, geevalueer. Die effek op SAF produksie van verhoogde geen dosering asook die teiken van SAF na die peroksisoom was ondersoek as moontlike strategieë om die opbrengs van SAF te verhoog. Die teiken van SAF na die peroksisoom is behaal deur die Peroksisomale Teiken Volgorde 1 (PTS1) aan die C-terminaal van SAF te heg. Die funksionaliteit van PTS1 was bevestig deur gebruik te maak van groen fluoroserende proteïen (GFP), fluoressensie mikroskopie en isolering van peroksisome. Teiken van SAF na die peroksisoom het ‘n negatiewe uitwerking gehad op SAF uitdrukking in beide H. polymorpha en P. pastoris. ‘n Verhoging in geen dosering het geen onderskeibare effek gehad op SAF opbrengs in H. polymorpha nie wat in teenstelling is met vorige navorsing. Die hoogste produksie vlakke is opgelewer deur P. pastoris KM71 (24.86 mg/L) wat goed vergelyk met vlakke van L1 wat deur ander navorsings groepe behaal is. Die belangrikste gevolgtrekking wat gemaak kan word uit hierdie studie is dat al die rasse wat SAF geproduseer het in meetbare hoeveelhede ewe effektief was. Verhoogde biomassa was dus die grootste bydraende faktor tot hoë SAF vlakke (mg/L) in die P. pastoris rasse as gevolg van die hoë sel digthede wat hierdie rasse kan bereik. Dit is duidelik dat metielotrofiese giste, met die nodige optimisering, oor die potensiaal beskik om as gasheer sisteme te dien vir die produksie van ‘n wye spektrum HPV entstof.
The NRF and the Department of Microbiology for financial support

Orientador: Estela Kaminagakura Tango
Banca: Ana Lia Anbinder
Banca: Alfredo Ribeiro Silva
Banca: Laura Sichero
Banca: Silvana Pasetto
Resumo: A papilomatose laríngea (PL) é uma doença rara e grave, dividida em dois grupos: juvenil (PLJ) e adulta (PLA), ambas causadas pelo papilomavírus humano (HPV). Seu curso pode ser agressivo, com inúmeras recidivas, risco de malignização, disseminação pulmonar e obstrução das vias aéreas. Para identificar os casos mais agressivos e nortear os tratamentos foram desenvolvidas escalas laringoscópicas, dentre elas; a escala desenvolvida por Derkay et al. (1998). O objetivo deste projeto foi caracterizar a PL e correlacionar suas características clínico-patológicas com com a escala laringoscópica de Derkay. Os dados e biópsias de 36 pacientes com PLJ e 56 com PLA foram coletados e analisados por meio da microscopia de luz. Os pacientes foram separados em grupos de acordo com os índíces de Derkay: ≥20 para os mais agressivos e <20 para os casos menos agressivos. Foram realizadas reações de imuno-histoquímicas anti- Fator XIIIa, CD3, CD4, CD8, CD15, CD20, CD68, FoxP3 e MUM-1. As células inflamatórias foram quantificadas. Todas as características clínico-patológicas e os resultados da reação imuno-histoquímica encontrados foram comparados entre os grupos propostos através do teste estatístico de Qui-Quadradro e correlacionados através do teste de correlação de Spearman. O nível de significância considerado foi de 5%. Ao comprar a severidade entre os grupos PLJ e PLA, o grupo PLJ foi considerado mais agressivo (P=0,02). Os pacientes entre as amostras com score ≥20 apresentaram maior inci... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Laryngeal papillomatosis (LP) is a rare and serious disease, divided into two groups: juvenile (JLP) and adult (ALP), both caused by the human papillomavirus (HPV). Its course can be aggressive, with numerous relapses, risk of malignancy, pulmonary dissemination and airway obstruction. To identify the most aggressive cases and guide the treatments, laryngoscopic scales were developed, among them; the scale developed by Derkay et al. (1998). The objective of this project was to characterize LP and to correlate its clinical-pathological characteristics with Derkay's laryngoscopic scale. The data and biopsies of 36 patients with JLP and 56 patients with ALP were collected and analyzed by light microscopy. The patients were separated into groups according to the Derkay indices: ≥20 for the most aggressive and <20 for the less aggressive cases. Anti-Factor XIIIa, CD3, CD4, CD8, CD15, CD20, CD68, FoxP3 and MUM-1 immunohistochemical reactions were performed and the inflammatory cells were quantified. All the clinical-pathological characteristics and the results of the immunohistochemical reaction were compared between the groups proposed using the Chi-Square test and correlated through the Spearman correlation test. The significance level considered was 5%. When comparing aggressivity between the JLP and ALP groups, the JLP group was considered more aggressive (P = 0.02). Patients among the samples with score ≥20 had a higher incidence of tracheostomy and severe respiratory distress... (Complete abstract click electronic access below)
Doutor

Conservation efforts to prevent the extinction of the endangered western barred bandicoot Perameles bougainville (WBB) are currently hindered by a debilitating progressive papillomatosis and carcinomatosis syndrome. Now extinct on mainland Australia, wild populations of the WBB are known only to exist on Bernier and Dorre Islands in Shark Bay, Western Australia. This thesis describes and analyses the pathological (gross, histological, ultrastructural) and immunohistochemical features of a papillomatosis and carcinomatosis syndrome in the WBB. The detection and characterisation of a novel virus, the bandicoot papillomatosis carcinomatosis virus type 1 (BPCV1), found in association with cutaneous and muco-cutaneous papillomas and carcinomas, is described. BPCV1 was found to exhibit genomic and morphological features of both the Papillomaviridae and the Polyomaviridae, and may represent the first member of a new family of viruses. The findings of this study provide evidence that BPCV1 is the causative agent of the papillomatosis and carcinomatosis syndrome. Clinical, pathological and molecular evidence of the syndrome and BPCV1 were found in the Bernier Island WBB population at Red Cliff and in captive populations comprising all or a proportion of founder WBBs from this site, but not at all in the WBB population on Dorre Island or Heirisson Prong. The papillomatosis and carcinomatosis syndrome in the western barred bandicoot is a pertinent example of a disease process hampering efforts to prevent the extinction of an endangered species.

Conservation efforts to prevent the extinction of the endangered western barred bandicoot Perameles bougainville (WBB) are currently hindered by a debilitating progressive papillomatosis and carcinomatosis syndrome. Now extinct on mainland Australia, wild populations of the WBB are known only to exist on Bernier and Dorre Islands in Shark Bay, Western Australia. This thesis describes and analyses the pathological (gross, histological, ultrastructural) and immunohistochemical features of a papillomatosis and carcinomatosis syndrome in the WBB. The detection and characterisation of a novel virus, the bandicoot papillomatosis carcinomatosis virus type 1 (BPCV1), found in association with cutaneous and muco-cutaneous papillomas and carcinomas, is described. BPCV1 was found to exhibit genomic and morphological features of both the Papillomaviridae and the Polyomaviridae, and may represent the first member of a new family of viruses. The findings of this study provide evidence that BPCV1 is the causative agent of the papillomatosis and carcinomatosis syndrome. Clinical, pathological and molecular evidence of the syndrome and BPCV1 were found in the Bernier Island WBB population at Red Cliff and in captive populations comprising all or a proportion of founder WBBs from this site, but not at all in the WBB population on Dorre Island or Heirisson Prong. The papillomatosis and carcinomatosis syndrome in the western barred bandicoot is a pertinent example of a disease process hampering efforts to prevent the extinction of an endangered species.

Traditional Public Health decision-support can benefit from the Web and social media revolution. This dissertation presents approaches to mining social media benefiting public health epidemiology. Through discovery and analysis of trends in Influenza related blogs, a correlation to Centers for Disease Control and Prevention (CDC) influenza-like-illness patient reporting at sentinel health-care providers is verified. A second approach considers personal beliefs of vaccination in social media. A vaccine for human papillomavirus (HPV) was approved by the Food and Drug Administration in May 2006. The virus is present in nearly all cervical cancers and implicated in many throat and oral cancers. Results from automatic sentiment classification of HPV vaccination beliefs are presented which will enable more accurate prediction of the vaccine's population-level impact. Two epidemic models are introduced that embody the intimate social networks related to HPV transmission. Ultimately, aggregating these methodologies with epidemic and social network modeling facilitate effective development of strategies for targeted interventions.

L’infection persistante par les papillomavirus (HPV) dits « à haut risque » induit le cancer du col. Chez les femmes infectées par le VIH, les infections par ces HPV oncogènes et les lésions associées, allant des dysplasies au cancer invasif, sont plus fréquentes, plus sévères et de moins bon pronostic que chez les femmes non porteuses du VIH. Etonnamment, alors qu’il a été clairement établi que l’importance de la pathologie liée à HPV est directement proportionnelle au degré d’immunodépression des patientes porteuses du VIH, il n’a pas pu être démontré qu’un traitement antirétroviral efficace contre le VIH permettant d’améliorer l’immunité, diminue l’infection par ces HPV.

Entre janvier 2002 et décembre 2012, nous avons constitué une cohorte prospective de dépistage et de suivi de l’infection cervicale par HPV à haut risque incluant plus de 900 femmes traitées à la consultation du Centre de Référence SIDA de l’hôpital Saint-Pierre. Nos résultats montrent que chez ces femmes pour la plupart d’origine Africaine et traitée avec succès pour le VIH depuis plusieurs années, la prévalence et l’incidence de l’infection par HPV oncogène sont beaucoup plus importantes que dans la population belge générale ou que chez les femmes séropositives vivant dans d’autres pays occidentaux. Grâce à un suivi longitudinal de plusieurs années, nous avons pu démontrer que le risque d’être infectée par un HPV oncogène est significativement réduit sous trithérapie anti-VIH sous réserve d’obtenir une charge virale indétectable à <50 cp/ml pendant plus de 3 ans ou une restauration immunitaire à >500 lymphocytes CD4+/µL pendant plus d’un an et demi. Ces résultats ont été confirmés dans l’analyse que nous avons faite sur les nombreuses dysplasies cervicales également retrouvées dans notre cohorte. Enfin, nous avons trouvé que la distribution des génotypes d’HPV de nos patientes est similaire à celle trouvée en Afrique sub-saharienne impliquant que la couverture offerte par les vaccins anti-HPV varie entre moins de 30% pour les vaccins bi- ou quadrivalent actuellement disponibles à 80% pour le vaccin nanovalent en développement. Notre travail met en lumière l’étendue particulièrement importante de l’infection par HPV à haut risque chez les femmes séropositives vivant en Belgique et offre de nouveaux éléments de réflexion afin d’adapter à leurs particularités les recommandations belges et les critères de remboursement à la fois pour le dépistage du cancer cervical et la vaccination anti-HPV.

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Persistent infection with human papillomavirus (HPV) called “at high risk” induces cervical cancer. In HIV-positive women, infection with these oncogenic HPV and HPV-induced lesions ranging from cervical dysplasia to invasive cancer are more frequent, more severe and have a worst outcome than in HIV-negative women. An intriguing paradox is that, although it has been clearly demonstrated that high risk HPV infection and associated diseases are increased by progressive immune deficiency, the introduction of efficient therapy against HIV leading to improved immunity has not been associated with a decrease in oncogenic HPV infection or HPV-induced lesions.

Between January 2002 and December 2012, we have built a prospective cohort to screen and follow-up cervical infection by high risk HPV in more than 900 women treated for HIV in the AIDS Reference centre of Saint-Pierre Hospital. We have shown that among these women mainly from Sub-Saharan African origin and successfully treated for HIV for several years, the prevalence and incidence rate of high risk HPV are much higher than in the general population from Belgium or in HIV-positive women from other western countries. After several years of longitudinal follow up, we have demonstrated that the risk of infection by oncogenic HPV is significantly reduced by efficient therapy against HIV provided that HIV viral load has been sustainly suppressed below 50 cp/ml for more than 3 years or that immunity has been increased more than 500 CD4+T cells/µl for more than 1.5 years. These results have been confirmed in the analysis on cervical dysplasia which is also very prevalent in our cohort. At last, we have found that the HPV genotype distribution in our population is very similar to the one found in Sub-Saharan Africa. We have estimated that the coverage offered by the vaccines against HPV in our cohort is less than 30% for the currently available bi- or quadrivalent vaccine but reaches 80% with the future nanovalent vaccine. Our results highlight many differences in the HPV infection and associated diseases in HIV-positive women compared to HIV-negative women; these differences should be taken into account to adapt to our specific population the current Belgian guidelines or the reimbursement criteria on cervical screening and on vaccines against HPV.

Doctorat en Sciences médicales
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CAPES
Identificar a presença dos sorotipos de alto risco do Papilomavírus Humano (HPV) na saliva de pacientes portadores do vírus HIV. A amostra de 90 pacientes foi oriunda de dois centros de referência em tratamento de ISTs da cidade do Recife, PE, Brasil. Uma entrevista foi realizada para identificar o perfil da amostra, sendo realizada uma coleta de saliva empregando tubos falcon e solução para bochecho com sacarose a 5%, com posterior armazenamento em freezer a -20°C para rastreamento do HPV e genotipagem para o sorotipo 16 e 18 por PCR convencional. Na amostra predominou a presença do sexo masculino 59 de 90 (65,6%), com idade média de 38,8 anos, variando entre 18 e 69 anos, renda familiar média de 1,95 Salários Mínimos (DP = 1,37). A prevalência de HPV nesta amostra foi de 23 de 90 (25,6%) e dos sorotipos 16 e 18 foi 8 de 90 (8,9%). A co-infecção por HPV é comumente observada em pacientes portadores de HIV.
To identify the presence of high-risk serotypes human papillomavirus (HPV) in patients with sexually transmitted infections (STIs). A sample of 90 patients were from two referral hospitals in treatment of STIs. An interview was conducted to identify the sample’s profile a saliva collections being perfomed using falcon tubs and mount rinse with 5% sucrose, subsequente storage in a freezer at -20ºC for HPV screening and genotyping for serotype 16 and 18 by conventional PCR. In the sample predominant male presence 59 of 90 (65.6%) with mean age of 38.8 years, ranging between 18 and 69 years, average family income of 1.95 minimum wages (SD = 1, 37). The prevalence of HPV in this sample was 23 of 90 (25.6%) and the serotype HPV 16 and 18 was 8 of 90 (8.9%). Co-infection with HPV is commonly observed in HIV patients.

Thesis (PhD)--Stellenbosch University, 2014.
ENGLISH ABSTRACT: Title The Effect of Highly Active Antiretroviral Therapy on Human Papilloma Virus Infection and Cervical Cytological Abnormalities in Women Living With HIV Background Human Papillomavirus (HPV) infection causes cervical cancer. The prevalence of HPV-related dysplastic lesions is significantly higher in patients co-infected with the HI virus and thought to be linked to possible more persistent HPV infection. There is, however, conflicting evidence as to whether treatment of Human Immunodeficiency Virus (HIV) infection with antiretroviral agents may influence cervical HPV infection and the behaviour of Squamous Intraepithelial Lesions (SIL). Aims To examine the effect of the initiation of combination antiretroviral therapy (cART) on: 1) the persistence of cervical Low-grade SIL (LSIL); 2) The progression of cervical LSIL to High-Grade SIL (HSIL); 3) The effectiveness of excision treatment of HSIL 4) HPV genotypes detected, in HIVinfected and uninfected women at the Infectious Diseases Clinic and the Colposcopy Clinic, Tygerberg Teaching Hospital, Cape Town, South Africa. Design and Methods We conducted a retrospective cohort analysis of 1720 women with LSIL of the survival of progression-free-time or time-to-clearance. Time to progression or persistence was compared according to HIV status, antiretroviral treatment and CD4 count. In another retrospective cohort analysis, we investigated the effectiveness of excision treatment in 1848 women who underwent LLETZ or CKC biopsy was used. Logistic regression and survival analysis were used to compare excision treatment failure and recurrence-free time between groups according to HIV status, antiretroviral therapy and CD4 count. To investigate the effect of antiretroviral therapy on the cervical HPV infection, 300 HIV-infected women were prospectively enrolled and followed at 6-monthly interval. Cytological testing and cervical HPV sampling were done at each visit. Biopsy of suspicious lesions and excision treatment were done at colposcopy clinic according to standard a protocol. The Roche Linear array HPV genotyping test was used for HPV detection. Generalized Estimating Equation (GEE) multivariate analysis was applied to investigate the effect of cART on the detection of HPV infection, while adjusting for time-dependent covariates such as CD4 count, sexual activity and excision treatment. The effect on each HPV type was then also compared to the effect on HPV16. Results Overall, we found that there was no difference between the progression of LSIL to HSIL by HIV status. However, among HIV-infected patients, those who started ART before first LSIL had a significantly lower risk for progression (HR 0.66, 95% CI 0.54-0.81). CD4 count did not have an impact on the risk for progression. We also found lower persistence of SIL in the HIV uninfected group (HR 0.69, 95% CI 0.57-0.85) and that cART was independently associated with decreased persistence of LSIL. On the other hand, a higher CD4 count at the time of first LSIL was not associated with lower persistence of the lesion. HIV infected women with HSIL experienced much higher excision treatment failure than uninfected women (53.8% vs. 26.9%, p<0.001). Factors that improved outcome were higher CD4 count and complete excision. cART reduced the risk of detection of any HPV type by 47% (OR 0.53, 95% 0.49-0.58, p<001). When adjusted for covariates, time of exposure to cART and CD4 had a stronger effect. Every month of cART exposure reduced the risk detection of any HPV type with 7%. The effect was also significant on HPV16 alone (OR 0.93, 95% CI 0.90-0.95). All non-oncogenic subtypes were influenced similarly or more strongly than HPV16, as well as oncogenic HPV52. Only one oncogenic subtype HPV subtype, HPV39, was influenced marginally less (ratio of OR 0.95, CI 0.90-0.99, p=0.04). There was an increased risk for any HPV detection at CD4 count<200 (OR 1.63, 95% CI:1.50-1.77), but when adjusted, the time of cART exposure again remained the strongest predictor of risk (OR 0.94, 95% CI:0.93-0.95). Conclusion cART impact the outcome of cervical HPV infection by increasing clearance, decreasing progression of LSIL and recurrence after excision treatment. This effect is time dependent and also associated with CD4 count. Specifically, HPV16 detection risk is also reduced by cART, and all HPV types are influenced at least as much as HPV16, except possibly HPV39. It seems that increased cervical HIVproviral load is associated with HPV detection risk, and both are lowered by cART time.
AFRIKAANSE OPSOMMING: Titel Die Effek van Kombinasie Antiretrovirale Terapie op Menslike Papilloomvirusinfeksie en Servikale Sitologiese Abnormaliteite in Menslike Immuniteitsgebrekvirus-geïnfekteerde Vroue Agtergrond Menslike Papilloomvirusinfeksie (MPV) veroorsaak servikale kanker. Die prevalensie van MPVverwante displastiese letsels is betekenisvol hoër in pasiënte wie ook met Menslike Immuniteitsgebrekvirus (MIV) geïnfekteer is en dit word gereken dat dit te wyte is aan meer persisterende MPV infeksie. Daar is egter teenstrydige bewyse oor of die behandeling van MIV infeksie met antiretrovirale (ART) middels die infeksie met MPV en die gedrag van Plaveisel Intraepiletiële letsels (PIL) kan beïnvloed. Doelwitte Om die effek van die inisiasie van kombinasie ART op: 1) die persistering van Laegraadse PIL (LPIL); 2) die progressie van servikale LPIL na hoëgraadse PIL (HPIL) 3) die sukses van eksisiebehandeling van HPIL; 4) MPV genotypies waarneembaar, in MIV-geïnfekteerde vroue by die Infeksiesiektekliniek en die Kolposkopiekliniek,Tygerberghospitaal, Kaapstad, Suid-Afrika, te ondersoek. Studie-ontwerp en Metodes `n Retrospektiewe kohort-analise op 1720 vroue met LPIL van die oorlewing van progressive-vrye tyd en tyd tot opklaring van PIL is gedoen. Tyd tot progressie of opklaring is vergelyk na aanleiding van die pasiënt se MIV status, behandeling met antiretrovirale terapie en CD4-telling. In nog `n retrospektiewe kohort-analise is die effektiwiteit van eksisiebehandeling in 1848 vroue wie LLETZ or Kouemeskonus eksisie ondergaan het, ondersoek. Logistiese regressie en oorlewingsanalise is toegepas om die voorkoms van onsuksesvolle uitkoms en tyd sonder herhaling van letsels tussen groepe te vergelyk na aanleiding van MIV status, ART en CD4-telling. Om die effek van antiretroviral therapie op servikale MPV infeksie te ondersoek, is 300 MIVgeïnfekteerde vroue opgeneem in `n prospektiewe studie en sesmaandeliks opgevolg. Sitologiese en MPV servikale smere is met elke besoek geneem. Biopsies van verdagte letsels en eksisiebehandeling is by die Kolposkopiekliniek gedoen volgens die standaardpraktyk. Die Roche Linear Array HPV Genotyping toets is gebruik vir MPV deteksie. Algemeen-beraamde vergelyking (GEE) meerveranderlike analise is toegepas om die effek van die anti-MIV terapie op die teenwoordigheid van MPV op die serviks te ondersoek. Die aangepaste effek is ook getoets deur die CD4-telling, die seksuele aktiwiteits- en eksisiebehandelingstatus by elke besoek in ag te neem. Die effek op elke MPV genotipe is laastens dan ook vergelyk met die effek op ‘n spesifieke basislyn genotype; in hierdie geval was MPV16 gekies. Resultate Daar was geen statisties beduidende verskil tussen die progressie van LPIL na HPIL na aanleding van HIV status nie, maar pasiënte wie met ART begin het voordat hulle vir die eerste keer met LPIL gediagnoseer was, het ‘n laer risiko gehad vir progressie (HR 0.66, 95% VI 0.54-0.81). Daar is ook gevind dat dit onafhanklik van die CD4 telling was. Die persistering van PIL was laer in die MIV negatiewe groep (HR 0.69, 95% VI 0.57-0.85), maar ook hier was antiretrovirale behandeling geassosieer met verminderde persistering. Weer eens was daar nie ‘n verband met die CD4 telling nie. MIV-geinfekteerde vroue met HPILwas baie meer geneig tot gefaalde eksisiebehandeling (53.8% teenoor 26.9%, p<0.001). Verbeterde uitkoms was geassosieer met ‘n hoër CD4-telling en ‘n eksisie wat as volledig beskryf was. ART wat reeds voor die eksisiebehandeling begin was, het nie die risiko vir onsuskesvolle uitkoms statisties beduidend verminder nie, maar het egter die risiko vir herhaling van letsels na die eksisie sterk verlaag. ART het die kans dat enige MPV tipe waargeneem sou word, met 47% verlaag (OR 0.53, 95% VI 0.49-0.58, p<001). Wanneer aangepas vir ander faktore, was die tyd wat verloop het sedert ART begin was, sowel as vir die CD4 telling, sterker. Vir elke maand sedert ART begin was, het die kans dat enige MPV tipe waargeneem word, met 7% verminder. `n Soortgelyke effek is op HPV16 alleen gevind (OR 0.93, 95%, VI 0.90-0.95). Die effek was net so sterk of sterker op alle subtipes. Slegs een onkogeniese subtipe, MPV39, was gering minder beïnvloed (ratio van OR 0.95, VI 0.90-0.99, p=0.04). Die kans vir waarneming van enige MPV subtype is hoër wanneer die CD4 telling laer as 200 selle/ɥl is (OR 1.63, 95% VI: 1.50-1.77), maar wanneer aangepas, was die tyd van ART weer eens die sterkste voorspeller van MPV infeksie (OR 0.94, 95% VI:0.93-0.95). Gevolgtrekkings ART verbeter die uitkoms van servikale infeksie met MPV deur progressie en persistering van LPIL en herhaling van PIL na eksisie te verminder. Die effek is tydsafhanklik en word ook deur die CD4 telling beïnvloed. Die kanse dat MPV16 spesifiek waargeneem word, word ook deur ART verminder, en all MPV tipes ondervind dieselfde of groter verlaging van waarnemingsrisiko as MPV16, behalwe miskien MPV39. Ons kon aandui dat verhoogde teenwoordigheid van servikale MIV verband hou met die risiko vir die waarneming van MPV infeksie, en beide word verminer deur die tyd waarmee die pasiënt met ARV terapie behandel is.

Papillomaviruses (PVs) are a numerous family of small dsDNA viruses infecting virtually all mammals. PVs cause infections without triggering a strong immune response, and natural infection provides only limited protection against reinfection. Most PVs are part and parcel of the skin microbiota. In some cases, infections by certain PVs take diverse clinical presentations, from highly productive self-limited warts to invasive cancers. The main aim of this thesis is explore the link between HPV genotypic diversity and the phenotypic, clinical diversity of the diseases associated with HPV infection by means of evolutionary, clinical and ecologically approaches. From and evolutionary perspective, we studied the codon usage preferences of HPVs. By applying phylogenetic inference and dimensionality reduction methods, we found that phylogenetic relationships between HPVs explained only a small proportion of codon usage preferences (CUPrefs) variation. Instead, the most important explanatory factor for viral CUPrefs was infection phenotype, as orthologous genes in viruses with similar clinical presentation displayed similar CUPrefs. Moreover, viral genes with similar spatiotemporal expression patterns also showed similar CUPrefs. Moreover, we also found that HPV genes with similar spatiotemporal expression patterns displayed similar CUPrefs. Hence, Our results suggest that CUPrefs in HPVs reflect either variations in the mutation bias or differential selection pressures depending on the clinical presentation and expression timing. From a clinical point of view, first, we studied the distribution patterns of oncogenic and non-oncogenic HPVs in anal and perianal Squamous Intraepithelial Lesions (SIL) in non-vaccinated heterosexual men, women, and Men who have sex with men (MSM) with known HIV status. We found that there is an increased prevalence of low-grade perianal lesions driven only by oncogenic HPVs. We also found a high prevalence of anal SIL containing foci of high-grade SIL exclusively driven by non-oncogenic HPVs. Our results suggest that there is a disagreement in high-grade/low-grade intraepithelial neoplasia and oncogenic/non-oncogenic HPV infection. Second, we analyzed the presence of HPVs not explicitly targeted by standard molecular epidemiologic methods of detection in squamous carcinoma samples of the vulva, penis and head and neck. These three anatomical locations display a low fraction of cancer cases attributable to HPVs, in sharp contrast with the higher rates of viral DNA prevalence in anal and cervical carcinomas. The standard HPV detection methods target only a subset of clinically important HPVs, namely oncogenic AlphaPVs, and may thus overlook the presence of other HPVs. We tested 2365 samples and found 6 samples containing cutaneous HPVs, suggesting that certain cutaneous HPVs, typically classified as “non- oncogenic” HPVs, may be linked to small number of cancer cases and call for further studies to elucidate the pathogenic role and malignisation mechanism of these HPVs. Finally, from an ecological perspective, we studied the interaction among HPVs inside its host in different stages of the cervical infection and different anatomical regions. By applying established ecological methods, we found that HPVs interact within the host, and that the presence of one given HPV is not neutral for the rest of the HPVs infecting the host. We also study how the interaction among HPVs could be affected by the introduction of ecological pressures linked to vaccination. By applying the same ecological methods, we find that in the initial descriptions of the post-vaccination era, HPVs still form non-neutral communities suggesting that the vaccine is not changing the underlying processes that govern HPV distributions and relative abundances.
Los virus del Papiloma (VPs) son una numerosa familia de virus pequeños de DNA de doble cadena que infectan a todos los mamíferos. La mayoría de infecciones por VPs son subclínicas, sin embargo, en algunos casos, las infecciones por ciertos VPs toman diversas presentaciones clínicas, desde verrugas, lesiones altamente productivas, hasta cánceres invasivos. El objetivo principal de esta tesis es explorar el vínculo entre la diversidad genotípica de los virus del Papiloma Humano (VPH) y la diversidad fenotípica y clínica de las enfermedades asociadas con la infección por VPH mediante 3 enfoques: evolutivo, clínico y ecológico. Desde una perspectiva evolutiva, se estudiaron las preferencias de uso de codones (CUPrefs) de los VPHs. Aplicando métodos de inferencia filogenética y métodos de reducción de la dimensionalidad, se observó que que los genes ortólogos en los virus con presentaciones clínicas similares mostraron CUPrefs similares. Además, los genes virales con patrones de expresión espacio-temporales similares también mostraron CUPrefs similares. Nuestros resultados sugieren que las CUPrefs de los genes de los VPHs reflejan variaciones en los sesgos mutacionales así como presiones de selección diferencial, dependiendo de la presentación clínica de los VPHs y el momento de expresión génica. Desde el punto de vista clínico, se estudiaron los patrones de distribución de VPHs oncogénicos y no oncogénicos en lesiones intraepiteliales anales y perianales. Los resultados muestran que en hombres homosexuales (HSH) VIH positivos hay un aumento de la prevalencia de lesiones perianales de bajo grado debidas a infecciones causadas por VPHs oncogénicos en comparación con HSH que son VIH negativos. Nuestros resultados también muestran que en HSH, independientemente del estatus de VIH, existe una alta prevalencia (>65%) de lesiones anales de bajo grado que contienen focos de lesiones de alto grado que son asociadas a la infección con VPHs no oncogénicos. También se analizó, en muestras de carcinoma escamoso de vulva, pene y cabeza y cuello, la presencia de VPHs cutáneos. Se testaron 2365 muestras y se detectó que 6 muestras contenían AND de VPHs cutáneos. Estos resultados sugieren que ciertos tipos de VPH cutáneos podrían tener potencial oncogénico. Se requieren de estudios adicionales para dilucidar el papel patogénico y el mecanismo de malignización de estos VPHs. Por último, desde una perspectiva ecológica, se estudió la interacción entre los VPHs en diferentes estadios de la infección cervical y diferentes regiones anatómicas. Los resultados mostraron que los VPH interactúan dentro del huésped. También se estudió cómo la interacción entre los VPH podría verse afectada por la introducción de presiones ecológicas vinculadas a la vacunación. Los resultados mostraron que en las descripciones iniciales de la era posterior a la vacunación, los VPH también forman comunidades no neutrales, sugiriendo que la vacuna no está cambiando los procesos subyacentes que gobiernan las distribuciones de VPH y las abundancias relativas.

In an age of biomedicine, technologies, drugs, and treatments are expanding in new and diverse ways. Especially relevant to biomedicalization and this research is how such information is conveyed to the public through the media. Medical information is omnipresent in the media through direct-to-consumer (DTC) advertising and regular coverage of health topics in the news. The accessibility and proliferation of medical information provides an important opportunity to examine the ways in which these topics are framed in the media. This research specifically examines the framing of the HPV vaccine, Gardasil in the mass media. In this study, I explore how Gardasil was framed, how gender and sexuality were utilized within such frames, and what groups influenced these frames. I employ frame analysis to examine the presentation of Gardasil in the mass media. Due to the vaccine's intricate connection to gender and sexuality, I examine how these dimensions are utilized and reproduced in such frames. Gardasil was originally approved only for women, making gender a salient aspect of the vaccine. The current body of research examining Gardasil in the media presents data from the time the vaccine was only available to women. Now that the HPV vaccine is approved for men, this research seeks to contribute to a more comprehensive understanding of how Gardasil was framed in the media now that it is available to men and women. And given that Gardasil prevents a sexually transmitted infection (STI) in men and women, it is uniquely tied to issues of sexuality. To analyze Gardasil in the media, I examine newspaper articles, magazine articles, and media representations from stakeholder groups, including DTC advertising, official statements, and group websites. Analysis of key sources indicates that Merck dominated the framing of the vaccine in DTC advertisements and the news media, illustrating the power of the pharmaceutical industry. Findings indicate that the initial marketing of Gardasil constructed the vaccine as uniquely tied to femininity and later, women's empowerment. However, once the drug was approved for men, messages were reframed to appeal to a wider audience. Overall, the media continued to frame the vaccine specifically for women, further constructing HPV as a "woman's disease." The dominant focus on women concomitantly silenced the sexual health of men and sexual minorities. In conclusion, the marketing, discourse, and structural elements of Gardasil make it less accessible to those most in need, therefore contributing to the ongoing problem of cervical cancer and HPV.

The work presented in this thesis was performed to investigate the role of human papillomavirus (HPV) testing for improved detection of cervical neoplasia in the Scottish Screening Programme (CSP). A longitudinal study of 975 archived cervical smears from women who had histologically-confirmed cervical intraepithelial neoplasia (CIN) showed that HPV positivity using Hybrid Capture Assay II (HCAII) was found to increase with increased cytological abnormality and 18% overall contained high-risk HPV (HR-HPV) DNA. Retrospective analysis of cytology data showed that the positive predictive value (PPV) of cytology alone for the detection of significant cervical disease was 71% while that of HPV testing alone was 38%. Combined cytology and HPV testing before biopsy would have increased the PPV to 78%. However, the negative predictive value (NPV) of a negative HR-HPV result with normal cytology was 100%. These results suggested that HPV testing may have a greater role in the exclusion of severe cervical diseases rather than as a predictor of its presence. In a split-sample study, comparing conventional Pan smears and liquid-based cytology (LBC), monolayer preparations from LBC samples reduced the inadequacy rate from 7.3% to 2%. LBC also reduced the number of borderline smear reports from 7.9% to 4.2% and increased the number of severe cytological abnormalities detected. The clinical value of LBC was further endorsed in the direct-to-vial study where LBC replaced conventional smears within the cervical screening programme. Monolayer preparations from over 4700 LBC samples showed a reduction in the inadequacy rate from around 10% to less than 1%. This reduced the level of anxiety in women and provided savings in repeat clinic visits for patients, repeat samples for laboratories and administrative time at both sites. HPV testing would be a useful adjunct to cytology for improved management of women with borderline smear results. In the split sample study, 44.3% of women with a previous borderline or low-grade cervical abnormality were HR-HPV DNA positive. However, if all 110 HPV negative women had been managed by routine recall than only two cases of CIN1 would have been missed while 160 repeat visits and smears would have been saved. This clearly has benefits for both patients and clinicians at the primary care level and HPV-based triage has the potential to significantly reduce referrals to colposcopy.

Human Papillomavirus (HPV) is the cause of a wide spectrum of disease ranging from benign cutaneous warts to malignant anogenital tumours. Two notable features of HPV diseases are that the viruses are highly tissue specific of individual HPV types and the fact that while HPV infections are common only a minority of infected individuals manifest clinical disease, indicating the importance of host virus interactions. In this thesis two HPV mediated diseases have been examined in detail, cervical cancer and Recurrent Respiratory Papillomatosis. Cervical cancer is a major cause of mortality in women in developing nations. In these countries cervical screening programmes are impractical for logistical reasons and there is much interest in the feasibility of developing preventative vaccines. However, before such agents can be developed it must be established that the same HPV types that cause cervical cancer in industrialised nations are associated with cervical cancer in developing nations and that the local clades of HPV have the same amino acid sequence as putative vaccine strains. This thesis presents a study of the prevalence of cervical HPV infection, HPV type distribution and viral genetics carried out in a previously unstudied population of 934 women in rural Gambia. A high cervical HPV prevalence is observed, the most common high risk types were found to be HPV-16 & -35, the former being the most common high risk type worldwide and the latter this study show may be underestimated in African populations. Sequencing of the HPV L1 open reading frames provided data which may have implications for vaccine research. RRP is a rare disease characterised by the presence of papillomata on the larynx and other sites in the upper aerodigestive tract. RRP is usually caused by HPV-6 or -11, two viruses more commonly associated with genital warts. RRP is thought to be contracted from the genital tract. However, genital HPV infection is common yet RRP remains a rare disease, therefore other factors either from the virus or the host must modulate disease pathogenesis. In order to elucidate if RRP is caused by unique HPV clades that might be particularly well adapted to the larynx, the L1 major capsid gene and the oncogenes E6 & E7 have been sequenced. In order to establish the sequence of "normal" genital HPV-11 and to exclude regional differences between UK and US HPV-6 and HPV-11, genital wart tissue from 37 UK donors has also been analysed and compared to papilloma material from 53 RRP patients. Significant sequence differences between RRP and Genital Wart E6 & E7 ORF were observed. In order to examine the possible role of host immunogenetic factors in the pathogenesis of RRP, 50 individuals with RRP were then HLA class typed. A significant association between the HLA class II allele DRB*0301 was found along with a negative association with the HLA DQB1*03 allele. Finally, T-cell proliferation studies using Cytokine Bead Array to detect cytokine production revealed that DQBT03 positive donors produce more IFN-y in response to HPV peptides than DQB1*03 negative individuals.

The purpose of this study was to investigate women's knowledge and attitudes regarding genital human papillomavirus (n=100). Using a descriptive design, the Health Education Questionnaire was administered to 100 female patients (Mean Age = 33, SD = 7.17) at a physicians office in South Florida. The results indicated a lack of knowledge regarding genital human papillomavirus with 21 patients (21%) reported having knowledge and 79 (79%) having never heard of this disease. In addition, the group familiar with genital human papillomavirus also possessed a low level of knowledge with only 57% acknowledging an association of genital human papillomavirus and cervical cancer, 52% aware that a pap smear can detect the virus, 42% knowing that antibiotics can not treat the disease and 57% aware that it is not associated with a family history. An association was found between attitudes and health seeking behaviors. Subjects stating that they would take all measures to prevent genital human papillomavirus, were more likely to have a pap smear within the last year (Chi-square (1) = 4.33, p < .05). Higher levels of education and income were associated with increased knowledge regarding genital human papillomavirus when subjects were categorized according to sociodemographic characteristic (Chi-square (1) =9.45, p < .05; Chi-square (1) = 6.75, p < .05). There was no significant correlation between knowledge and ethnicity, marital status or age. Findings indicated the need for improved education and promotion of positive attitudes regarding human papillomaviurs in order to improve health seeking behaviors among women.

Human papillomavirus infection is regulated by multiple factors including methylation, viral integration and aberrations in host and viral gene expression. In most patients, the infection is transient and cleared effectively by the host’s immune system but in a minority of cases, the Human papillomavirus infection progresses to neoplasia and cancer if not detected and treated. Assays that detect the presence of Human Papillomavirus infection alone are not good prognostic markers of clinical outcome. Alternative clinical biomarkers that can measure other regulatory factors of Human Papillomavirus infection are required to more accurately predict patient outcome and help direct treatment options specifically to patients at risk of neoplasia and cancer progression. This study aimed to examine several Human Papillomavirus regulatory factors to determine if they would be suitable as clinical biomarkers and explore further the link between molecular changes and associated pathology. This involved development of assays, application to samples obtained from different cohorts of women to ascertain prognostic validity and development of an in vitro system to model the in vivo pathology. Initial work focused on investigating Human Papillomavirus 31, 33, 35 and 51 integration and methylation assays as prognostic biomarkers in young women attending their first routine cervical smear. Results indicated that Human Papillomavirus E2 gene disruption and methylation were not common events and the assays investigated were not suitable biomarkers for predicting clinical outcome in xx young women. The assays were then applied to clinical samples taken from patients with varying grades of cervical disease and high levels of viral methylation were present in women with high grade disease (Cervical Intraepithelial Neoplasia II+) and a correlation between methylation and HPV gene disruption was shown. Novel in vitro organotypic raft cultures of cells from Vulval Intraepithelial Neoplasia and Vaginal Intraepithelial Neoplasia were employed to understand how molecular changes in viral methylation and gene expression correlated with observed pathology. The organotypic raft cultures showed diverse differentiation patterns. No correlation of pathology with viral integration status was detected. However, organotypic raft cultures with high-grade disease morphology all had a higher level of methylation and expression of regulatory genes p16, Ki-67 and Sonic Hedgehog in comparison cultures displaying a histology consistent with low-grade disease. p16 and Ki-67 are already being examined as part of cervical screening triage. The findings presented in this thesis, highlight a need for further research into Human Papillomavirus infection and the molecular changes associated with Sonic Hedgehog gene expression and viral methylation as these show promise as prognostic biomarkers.

Cutaneous and genital human papillomavirus (HPV) infection in HIV patients, on suppressive anti-retroviral therapy (ART), poses under-investigated clinical challenges. HPV in HIV may represent a form of immune reconstitution associated disease (IRAD). HPV disease and IRADs have been separately correlated with human leucocyte antigen (HLA) genotype. HLA might also influence HPV in HIV. Comprehensive HPV typing of persistent warts obtained from HIV infected and healthy subjects was performed. Cutaneous HPV types were detected using nested PCR/sequencing and newly developed (Luminex based) HSLPCR/ MPG; genital and beta HPV types were identified using a reverse hybridisation line probe assay. Real time PCR was employed to determine HPV DNA viral loads. HLA alleles were defined in HIV infected and healthy patients by Luminex-based molecular typing using DNA derived from blood. The HPV profile of cutaneous and genital HIV warts differs significantly from warts from healthy individuals. In HIV, HPV 7 has been confirmed to be an important HPV type in cutaneous warts (p=0.001). In genital warts in HIV, HPV 11 is the predominant HPV type (p=0.15) and HPV 6 is less common (p=0.002), contrasting with the usual finding that HPV 6 is the principal type in the general population. Cross-over of HPV types between cutaneous and genital sites suggests that HPV tropism is less important than previously thought. An excess of beta HPV types, predominantly as mixed infections, is seen in cutaneous warts in HIV (p<0.0005). The HLA class I allele group HLA-B*44 (as the allele HLA-B*44:02 and the haplotype HLA-B*44, -C*05) has been identified more frequently in HIV than in controls (p=0.004, allele group; p=0.0006, allele; p=0.001, haplotype). The class II allele HLA-DQB1*06 may also be of interest (p=0.03). However, the differences are reduced after correction for multiple testing. Further work is required to ascertain if these HPV types and alleles are of importance.

Foi estudado o perfil nosológico da população ribeirinha do Baixo Rio Machado em Rondônia, área de Amazônia Ocidental no Brasil. Neste estudo, escolhemos abordar o perfil das seguintes doenças: malária, hepatite viral B e C, parasitose intestinal e papilomavírus humano. Estas doenças foram priorizadas devido à importância clínica e epidemiológica e também escassez de estudos referentes nas populações ribeirinhas amazônicas. Para isto, foi realizado um estudo descritivo no Baixo Rio Machado que se localiza a 250 km de Porto Velho (capital do Estado de Rondônia) pela margem direita do Rio Madeira, que é um afluente volumoso do Rio Amazonas, no Brasil. Nesta área, vivem aproximadamente 806 pessoas distribuídas em 55 comunidades ribeirinhas isoladas. Em Rondônia, encontramos locais, como o Rio Machado, que abrigam a presença de portadores assintomáticos do Plasmodium, fato que pode contribuir para a persistência desta doença na região. Para caracterizar o perfil da malária ribeirinha, focalizando aspectos da infecção assintomática e sintomática, foram realizados hemoscopia e Reação em Cadeia da Polimerase (PCR) com amplificação do DNA ribossomal do parasita para Plasmodium vivax e Plasmodium falciparum para o diagnóstico da malária. Foi considerado como assintomático o paciente com hemoscopia positiva ou PCR positivo que permaneceu sem sintomas durante pelo menos 60 dias. Foi realizado acompanhamento longitudinal de 70 dias após o primeiro corte transversal (nC1=585 pessoas) para observação dos pacientes assintomáticos diagnosticados por hemoscopia, onde 25 pacientes (4,25%) permaneceram assintomáticos durante todo esse seguimento. Após, foi realizado mais dois cortes transversais a cada 6 meses (nC2=583 pessoas, nC3=607 pessoas), com tratamento dos indivíduos assintomáticos diagnosticados por PCR no corte transversal anterior ou hemoscopia atual e também dos casos sintomáticos. Foi estudado, na população que permaneceu na área durante toda a pesquisa (n=379), o perfil e o impacto do tratamento da infecção assintomática. Houve diminuição da infecção por P. falciparum de seis vezes e aumento da prevalência de malária por P. vivax de aproximadamente três vezes. O diagnóstico por PCR foi de 2-5 vezes mais eficiente do que por hemoscopia e a prevalência de infecção assintomática foi de 16 a 22% nos cortes transversais. Assim, foi observada a importância do tratamento dos indivíduos assintomáticos para o controle da malária no local, sendo que o incremento de malária por Plasmodium vivax deve ter ocorrido devido a recaídas da doença. Com o estudo entomológico da área, foi verificado que o vetor Anopheles possui atividade hematofágica moderada e principalmente em região peridomiciliar, sendo o A. darlingi a principal espécie da região. Existem poucos dados sobre a prevalência de hepatites na população ribeirinha de Rondônia, o que dificulta a vigilância epidemiológica nestas regiões amazônicas. Foi realizado um estudo de prevalência de hepatite B e C na área, com 123 pacientes pertencentes a 5 comunidades ribeirinhas, incluindo todas as pessoas que estavam presentes na área no momento do corte transversal, de todas as faixas etárias. Foi realizada sorologia para os marcadores de hepatite B: HbsAg, Anti-Hbc (total) e Anti-Hbs e sorologia para hepatite C com o marcador anti-HCV. Foram encontrados 12 (14,7%) pacientes com hepatite B aguda, 29 (38,7%) pacientes apresentaram imunidade vacinal para hepatite B e 7 (9,3%) pacientes apresentaram hepatite C, sendo que um (1,3%) paciente tinha co-infecção para hepatite B e C. Notou-se que esta população ribeirinha está exposta às hepatites virais, sendo necessário intensificar a vigilância epidemiológica na área, assim como a cobertura vacinal e também fornecer cuidados preventivos, curativos e paliativos em relação a estas doenças. A infecção pelo papilomavírus humano (HPV) é altamente prevalente, sendo detectada em aproximadamente 10% a 20% da população sexualmente ativa entre 15 e 49 anos de idade. A introdução de testes mais acurados para a detecção do DNA do HPV em investigações epidemiológicas confirmou a importância do HPV, principalmente dos tipos de alto risco, como o principal fator de risco para o desenvolvimento de neoplasia intra-epitelial cervical e câncer do colo uterino. Não foram encontrados estudos sobre a existência do HPV em populações ribeirinhas amazônicas na literatura médica. Foi realizado, então, um estudo transversal para levantar a prevalência de HPV nesta população, com 84 participantes em idade sexualmente ativa presentes na área. Após coleta de consentimento informado e questionário clínico-epidemiológico, foi realizado exame para isolar o HPV. A tipagem do HPV foi realizada com amplificação dos DNAs por PCR empregando iniciadores genéricos seguida de hibridização em pontos, capaz de identificar mais de 40 tipos diferentes de HPVs. Foram encontradas 18 pacientes contaminadas pelo HPV, perfazendo 21,4% da amostra. Os tipos de HPV encontrados foram: 53, 58, 31, 56, 16, 83, 55, 66, 45, 51, 40, 42, 6, 68. Os tipos de HPV mais freqüentes foram 51 (23%), 58 (19%), 53 (7,7%), 83 (7,7%), sendo HPV 16 encontrado em 3,8% das pacientes HPV positivas e a prevalência do HPV de alto risco oncogênco foi de 13,1%. Desta forma, foi encontrado alta prevalência de HPV na população ribeirinha amazônica estudada, evidenciando a necessidade de vigilância epidemiológica para câncer de colo uterino na região. Por fim, foi estabelecido a prevalência de parasitoses intestinais na população, abordando a correlação das enteroparasitoses com malária assintomática e anemia, realizando um inquérito coproparasitológico na área incluindo pacientes de todas as faixas etárias, que estivessem dispostos a participar do estudo. A análise das amostras foi realizada pelo método da Sedimentação Espontânea (método de Lutz ou Hoffmann, Pons & Janer). Entre os 268 exames de fezes realizados, a prevalência de parasitose intestinal encontrada na região em estudo foi de 86,6%. Entre os helmintos, Ascaris lumbricoides (47%), Ancilostomideos (37,3%), Trichuris trichiura (3,4%), Capillaria hepatica (2,3%), foram os parasitas mais encontrados. Entre os protozoários foram encontrados: Entamoeba coli (21,4%), Entamoeba histolytica (12%), Giardia intestinalis (15,4%), Endolimax nana (10,1%) e Iodamoeba butschlii (7,5%). A alta prevalência de parasitose intestinal encontrada na pesquisa foi concordante com outros dados desenvolvidos na Região Amazônica. Com este estudo, a região da Amazônia Ocidental insere-se no cenário geográfico mundial de distribuição da Capillaria hepatica, particularmente, a região do Baixo Rio Machado.
Was studied the nosological profile of the riverside population of the Baixo Rio Machado in Rondônia, Western Amazon area of Brazil. In this study, was chosed the profile of following diseases: malaria, hepatitis B and C, intestinal parasite and human papillomavirus. These diseases have been prioritized due to clinical and epidemiological importance and scarcity of studies in the riverside Amazon population. For this, was conducted a descriptive study in the Baixo Rio Machado which is located 250 km from Porto Velho (capital of Rondônia State) in the right bank of the Rio Madeira which is a large tributary of the Amazon River in Brazil. In this area, live about 806 people distributed in 55 riversides communities isolated. In Rondônia, we find places such as the Rio Machado, which shelter to the presence of asymptomatic carriers of Plasmodium, which may contribute to the persistence of this disease in the region. To characterize the profile of malaria riverside, focusing on aspects of symptomatic and asymptomatic infections, were performed haemoscopia and Polymerase Chain Reaction (PCR) with amplification of ribosomal DNA for the parasite Plasmodium vivax and Plasmodium falciparum to the diagnosis of malaria. Was considered as asymptomatic patient with positive haemoscopia or positive or PCR that remained without symptoms for at least 60 days. Longitudinal follow-up was conducted for 70 days after the first cross section (nC1 = 585 people) for observation of asymptomatic patients diagnosed by haemoscopia where 25 patients (4.25%) remained asymptomatic throughout the follow-up. After was performed two cross cuts every 6 months (nC2 = 583 people, nC3 = 607 people), with treatment of asymptomatic individuals diagnosed by PCR in previous transverse cross-sections or haemoscopia a current and also symptomatic cases. We studied in the population that remained in the area throughout the study (no= 379), the profile and impact of treatment of asymptomatic infection. There was a decrease in infection by P. falciparum of six times and increased prevalence of malaria by P. vivax approximately three times. The diagnosis by PCR was 2-5 times more efficient than haemoscopia and the prevalence of asymptomatic infection was 16 to 22% in transverse cross-sections. Thus, was saw the importance of treatment of asymptomatic individuals for the control of malaria in the place, and the increase in malaria by Plasmodium vivax should have occurred due to relapse of the disease. With entomological study of the area, was saw that the vector Anopheles has moderate blood activity and especially in peridomiciliary region and the A. darlingi the main species in the region. There are few data on the prevalence of hepatitis in riverside population of Rondônia, which makes surveillance in Amazonian regions. Was conducted a study of prevalence of hepatitis B and C in the area, with 123 patients belonging to 5 communities, including all the people who were present in the area at the cross section of all ages. Serology was performed for markers of hepatitis B: HBsAg, anti-HBc (total) and anti-HBs and serology for hepatitis C with anti-HCV marker. Was found 12 (14.7%) patients with acute hepatitis B, 29(38.7%) patients demonstrated immunity to hepatitis B vaccine and 7(9.3%) patients had hepatitis C, with one (1.3%) patient had co-infection for hepatitis B and C. Realize that this riverside population is exposed to viral hepatitis, being necessary to intensify epidemiological surveillance in the area, as well as immunization coverage and provide preventive care, curative and palliative for these diseases. The infection by human papillomavirus (HPV) is highly prevalent, being detected in approximately 10% to 20% of the sexually active population between 15 and 49 years of age. The introduction of more accurate tests for the detection of HPV DNA in epidemiological investigations confirmed the importance of HPV, particularly types of high risk, as the main risk factor for the development of cervical intraepithelial neoplasia and cervical cancer. Was found no studies on the existence of HPV in riverside Amazonian populations in the medical literature. The transversal cross-section study was take up to raise the prevalence of HPV in this population, with 84 participants in sexually active age in the area. After collection of informed consent and clinical-epidemiological questionnaire, was conducted to isolate the HPV test. The typing of HPV was taking with the DNAs for PCR using generic primers followed by hybridization in points, able to identify over 40 different types of HPV. Was found 18 patients infected by HPV, comprising 21.4% of the sample. The HPV types found were: 53, 58, 31, 56, 16, 83, 55, 66, 45, 51, 40, 42, 6, 68. The most common HPV types were 51 (23%), 58 (19%), 53 (7.7%), 83 (7.7%) with HPV 16 found in 3.8% of HPV positive patients and prevalence the high risk HPV oncogenic was 13.1%. Thus, was find high prevalence of HPV in riverside Amazonian population studied, highlighting the need for surveillance for cancer of the cervix in the region. Finally, was established the prevalence of intestinal parasites in the population, deal the correlation of intestinal with asymptomatic malaria and anemia, taking the fecal examination conducting in the area including patients of all ages, who were willing to participate in the study. The analysis of samples was performed by the spontaneous sedimentation method (method of Lutz or Hoffmann, Pons & Janer). Among the 268 fecal examinations, the prevalence of intestinal parasite found in the region under study was 86.6%. Among helminths, Ascaris lumbricoides (47%), Ancilostomideos (37.3%), Trichuris trichiura (3.4%), Capillaria hepatica (2.3%) were found more parasites. Among the protozoa were: Entamoeba coli (21.4%), Entamoeba histolytica (12%), Giardia intestinalis (15.4%), Endolimax nana (10.1%) and Iodamoeba butschlii (7.5%).The high prevalence of intestinal parasite found in the survey was consistent with other data developed in the Amazon region. In this study, the region of the Western Amazon is part of the scenario of global geographic distribution of Capillaria hepatica, particularly the region of the Baixo Rio Machado.

OBJETIVO: O objetivo deste estudo foi analisar a concordância entre o esfregaço anal e a biópsia guiada por anuscopia de alta resolução no diagnóstico da displasia anal em pacientes infectados pelo HIV. MÉTODO: Conduzimos uma análise transversal de pacientes infectados pelo HIV submetidos a rastreamento de displasia anal rotineiro. A concordância entre mensurações foi estimada por índice de kappa ponderado através de sistema de avaliação citológica e histológica de três categorias (normal, displasia de baixo grau, e displasia de alto grau). Estimativas de sensibilidade, especificidade e valores preditivos foram calculados através de sistema de avaliação citológica e histológica de duas categorias (ausência de displasia e displasia de qualquer grau). Estimativas foram calculadas também para a detecção de displasia de alto grau. RESULTADOS: No decorrer de um ano, 222 pacientes foram submetidos a 330 esfregaços anais seguidos de biópsias guiadas por anuscopia de alta resolução. Trezentos e onze (311) esfregaços com biópsias concomitantes foram satisfatórios. Considerando-se a histologia como padrão, a freqüência de displasia anal foi de 46%. O índice kappa ponderado para concordância entre o esfregaço anal e a biópsia foi de 0,20. Para detecção de displasia anal de qualquer grau, o esfregaço anal demonstrou sensibilidade de 61%, especificidade de 60%, valor preditivo positivo de 56% e valor preditivo negativo de 64%. Para displasia de alto grau, o esfregaço anal demonstrou sensibilidade de 16% e especificidade de 97%. CONCLUSÃO: Os resultados obtidos no presente estudo, em que comparamos os achados da citologia dos esfregaços com os achados histológicos das biópsias dirigidas pela anuscopia de alta resolução em pacientes infectados pelo HIV permitiram concluir que houve baixa concordância entre eles
Purpose: To analyze the agreement between anal Pap smear and high resolution anoscopy guided biopsy to diagnose anal dysplasia in HIV-infected patients. Methods: Cross sectional analysis of HIV-infected patients receiving anal dysplasia screening as part of routine care. Agreement between measures was estimated by weighted kappa-statistics, using 3-tiered cytologic and histologic grading system (normal, low grade dysplasia, and high grade dysplasia). Estimates of sensitivity, specificity, and predictive values were calculated using a 2-tiered cytologic and histologic grading system (without dysplasia, and with dysplasia of any grade). Estimates were also calculated for the detection of high grade dysplasia. Results: Two hundred and twenty-two patients underwent 330 anal Pap smears followed by high resolution anoscopy guided biopsies in one year period. There were 311 satisfactory Pap smears with concurrent biopsy. Considering histology the standard, the frequency of anal dysplasia was 46 percent (95 percent confidence interval: 40-51 percent). Kappa-agreement between anal Pap smear and biopsy was 0.20 (95 percent confidence interval: 0.10 0.29). Anal Pap smear showed sensitivity of 61 percent, specificity of 60 percent, positive predictive value of 56 percent, and negative predictive value of 64 percent for detection of anal dysplasia of any grade. For high grade dysplasia, anal Pap smear showed sensitivity of 16 percent, and specificity of 97 percent. Conclusion: The present study showed a low concordance between anal Pap smears and high resolution anoscopy-guided biopsy

Cytology-based cervical screening programs have been in place in many countries for several decades, however the benefits of these prevention programs are unevenly distributed. Cervical cancer burden is higher is more disadvantaged groups, globally and within countries with well-established screening programs. Identifying human papillomavirus (HPV) as the causal agent in virtually all cervical cancer brought new prevention tools: HPV vaccination and HPV-based screening. This thesis examines how these new tools might help redress current disparities, using data and modelled analyses. Section 1 examines successes and gaps in current screening. Based on analyses of national incidence data, cytology-based screening in Australia and New Zealand has had a strong impact on squamous cell carcinoma and in women aged 25+, but a limited impact in women <25 and on adenocarcinoma. In New Zealand, the impact appears similar in Māori and non-Māori women. Section 2 examines subgroup effects in HPV vaccination. Chapters 4 and 5 show that the substantial decline in genital warts since the introduction of HPV vaccination in Australia has been similar in Indigenous and non-Indigenous women, and across different levels of socioeconomic disadvantage. Chapter 7 shows that, even if factors influencing vaccine uptake also influence sexual partner choice, the population-level impact of HPV vaccination programs is unlikely to be substantially impacted by feasible levels of heterogeneity in uptake. Section 3 examines how new approaches might fill the current gaps in cervical cancer prevention. Chapter 8 provides quantitative data on the benefits and harms of different screening decisions, including HPV testing on self-collected samples. Chapter 9 estimates that, together, HPV vaccination and HPV-based screening could lower adenocarcinoma incidence in 2040 by 55-85%. Together these findings provide reassurance that these new tools should help to close the current gaps in cervical cancer prevention.

Thirty bovine with neoplasms of the upper digestive tract (UDT) associated to the spontaneous consumption of bracken fern (Pteridium aquilinum) were studied. They were from 27 farms, localized in the municipalities of Jaguari (23) and Nova Esperança do Sul (4), Rio Grande do Sul, Brazil. The total bovine population in those farms was 1,090 and large amounts of bracken fern were found in the pastures. Twenty-six were cows e four were castrated males. The age ranged from 3 to 13-years-old. Most of them were 7 to 8-years-old (46,6%). Clinical signs observed in the affected animals were progressive weight loss, absence of ruminal movements, cough, dysphagia, regurgitation, halitosis, diarrhea, and bloat. Less frequent signs were selective appetite, dyspnea, and salivation. Two bovines died and 28 were euthanatized in extremis and submitted to necropsy. The main gross and microscopic findings were found in the same areas of the UDT. They were digestive papillomatosis, transforming papillomas, and squamous cell carcinomas (SCC). Twenty-nine bovines had papillomas of various sizes in several areas of the UDT. The digestive papillomatosis ranged from mild (45%), to moderate (38%), to severe (17%). Three developing phases were observed microscopically in the examined papillomas: an early growing phase, a developing phase, and a regressing phase. The regressing phase was characterized by lymphocytic infiltrates at the base of the papilloma. In 16 cases, the microscopic examination of lesions grossly resembling papillomas (although some were slightly round, with lower or ulcerated finger-like projections) revealed malignant transformation of the papillomas into SCCs. The SCCs were solitary (12/30) or multiple (18/30) and were histologicaly characterized as well, moderately, or poorly differentiated. Grouping the distribution of SCCs of larger extension in the UDT into cranial region (base of the tongue, pharynx/oropharynx, and epiglottis), medial region (esophagus), and caudal region (cardia and rumen), the distribution was cranial in 39%, middle in 16%, and caudal in 45% of the cases. By the same grouping criteria, but considering the total number of times SCCs of varied extensions were diagnosed in the cranial, middle, and caudal regions, the percentages changed to 34%, 26%, and 40%, respectively. Metastases to regional lymph nodes and other organs, like liver and lungs, were observed in 18 cases. Immunohistochemistry for cytokeratin was performed in selected sections of SCCs and metastases, showing strong positive reaction in the well and moderately differentiated SCCs, but weak positive reaction in the poorly differentiated ones. The epidemiological and histomorphological evidences showed in this study are in agreement with the observations that point out the co-carcinogesis between bovine papillomavirus type 4 infection and chemicals of bracken fern in the pathogenesis of the SCCs in the UDT of cattle.
Foram estudados 30 bovinos com neoplasias no trato alimentar superior (TAS) associadas ao consumo espontâneo de samambaia (Pteridium aquilinum) provenientes de 27 propriedades rurais, sendo 23 no município de Jaguari e quatro em Nova Esperança de Sul, Rio Grande do Sul. A população bovina total das 27 propriedades em que ocorreram os casos era de 1.090 bovinos e havia quantidade abundante de samambaia nas áreas de pastoreio dos animais. Vinte e seis bovinos eram vacas e quatro eram machos castrados. A idade variou de três a 13 anos, sendo o maior número de casos entre sete e oito anos (46,6%). Os sinais clínicos observados incluíram emagrecimento progressivo, atonia ruminal, tosse, disfagia, regurgitação, halitose, diarréia e timpanismo. Outros sinais clínicos menos freqüentes foram apetite seletivo, dispnéia e salivação. Dois bovinos tiveram morte espontânea e 28 foram submetidos à eutanásia in extremis e necropsiados. Os principais achados macroscópicos e histológicos observados nos 30 bovinos localizavam-se nos mesmos locais do TAS e consistiam de papilomas, papilomas em transformação para carcinomas de células escamosas (CCEs) e CCEs. Vinte e nove bovinos tinham papilomas de diversos tamanhos, sendo a quantidade variável entre leve (45%), moderada (38%) e acentuada (17%). Nos papilomas examinados microscopicamente, foram observadas três fases de desenvolvimento: a) fase inicial de crescimento; b) fase de desenvolvimento; e c) fase de regressão; essa última era caracterizada por infiltrados linfocitários nos eixos fibrovasculares de sustentação. Em 16 bovinos, o exame histológico de lesões macroscópicas compatíveis com papilomas, porém alguns deles apresentando-se mais arredondados, com projeções digitiformes atenuadas ou ulceradas, revelou a transformação maligna desses papilomas em CCEs. Os CCEs eram únicos (12/30) ou múltiplos (18/30) e variaram quanto ao grau de diferenciação celular entre bem diferenciados, moderadamente diferenciados ou pouco diferenciados. Quando a distribuição dos CCEs de maior extensão foi agrupada em regiões cranial (base da língua, faringe/orofaringe, epiglote), média (terços cranial, médio e caudal do esôfago) e caudal (entrada do rúmen e rúmen) do TAS, observou-se que a localização era cranial em 39% dos casos, média em 16%, e caudal em 45%. Utilizando-se esse mesmo critério de agrupamento, porém considerando o número total de vezes em que CCEs (de extensões variadas) foram diagnosticados nas regiões cranial, média e caudal, os números alteraram-se para 34%, 26% e 40%, respectivamente. Metástases de CCEs para linfonodos regionais e outros órgãos como fígado e pulmão foram observadas em 18/30 bovinos. A técnica de imunoistoquímica para citoqueratina foi realizada em cortes selecionados de CCEs e metástases, observando-se células fortemente positivas nos CCEs bem e moderadamente diferenciados, e fraca imunomarcação nos pouco diferenciados. As evidências epidemiológicas e histomorfológicas relatadas neste estudo reforçam as observações de uma estreita correlação entre a infecção pelo papilomavírus bovino tipo 4, causador da papilomatose digestiva, e a co-carcinogênese química dos princípios tóxicos da samambaia na patogênese dos CCEs do TAS de bovinos.

Cervical cancer is the second most common cancer among women worldwide. Infection with Human Papillomavirus (HPV) is essential but not the only contributing factor in cervical cancer development. HPV integration is reported to be present in over 80% of cervical cancers and disruption of HPV genome through integration leads to high levels of HPV oncogene expression. DNA damage and repair pathways are thought to induce HPV integration since HPV is detected at fragile sites in the human genome. There is controversy as to whether integration is an early or late event in cervical oncogenesis and there are no published studies to date that have investigated HPV integration using sensitive, DNA based, techniques at the nucleotide level in cervical precancers. This study aimed to test the hypothesis that integration is an early event in cervical neoplasia and episomal loss causes malignant transformation through transcription of integrated HPV. Also, this study served to pilot whether HPV integration can predict high-grade cervical disease in women with cytological abnormalities with an aim to improve current cervical screening methods. Assays to detect integration and E2 as a marker of episomal state were developed for HPV16, HPV18 and HPV45 and applied to cervical smears and biopsies from women with varying disease grades. The data presented in this thesis highlight that integration may not be essential for cervical cancer progression and different modes of disease progression may exist between young women and older women. Integration was detected at chromosome fragile sites but was more prevalent at SINE or LINE repeat elements; this implies a role for retroelements in the mechanism of integration. Finally, the data here suggest that integration induces a unique selective process in each individual and clonal selection may arise due to altered HPV oncogene expression and/or disruption to human gene expression.