Human papillomavirus (HPV) is a sexually transmitted virus associated with cervical cancer. The East African region has one of the highest incidences and mortality rates from cervical cancer but limited studies on HPV are available. Research aims were to describe: HPV genotypes, risk factors and rate of acquisition of prevalent and incident HPV in girls before and after reported first sex; rate and risk factors associated with HPV clearance, and to examine sexual behaviour reporting in face-to-face (FtF) interviews compared to Audio Computer-Assisted Self-Interviews (ACASI). A total of 503 girls aged 15 and 16 years in Mwanza, Tanzania, were enrolled and followed 3-monthly for 18 months with FtF-interviews and self-administered vaginal swabs. At enrolment, 474 girls reported no previous sex, and HPV was detected in 40/474(8.4%). During follow-up of girls who reported sex, new HPV incidence was 225/100 person-years(pys). Reporting sex in the past 3 months, and knowing the most recent sexual partner for a longer period before sex were associated with HPV acquisition. Median time from reported sexual debut to first HPV infection was 5 months, and median duration of infection 6 months. No factors were associated with HPV clearance. In girls who reported not having sex, HPV incidence was 29.4/100pys. ACASI was compared to FtF-interview in 203 girls at the 12-month visit. Although ACASI was feasible and acceptable, there was no increase in reporting of sex or other sexual behaviours, with the exception of kissing, compared to FtF-interviews. A very high incidence of HPV was seen in girls following sexual debut, and a higher than expected HPV prevalence and incidence were seen in girls who reported no previous sex. This emphasises the importance of HPV vaccination well before sexual debut. ACASI did not lead to increased reporting of vaginal sex and should be evaluated further in different settings.

Human norovirus (HuNV) is a member of the calicivirus family and is a major cause of viral gastroenteritis worldwide. Due to the absence of a suitable cell culture system, HuNoV replication mechanisms are poorly understood, but two animal caliciviruses, Feline calicivirus (FCV) and Murine Norovirus (MNV) provide models to increase our understanding of norovirus biology. Unlike cellular mRNAs, the calicivirus RNA genome does not possess a 5' cap structure but instead has a 13–15 kDa viral protein, genome linked (VPg) directing translation, hijacking the host protein synthesis machinery. The viral life cycle requires separated events occurring at different times since viral transcripts are used as the template both for translation (mRNA) and replication (genomic RNA). Therefore mechanisms are required to control the viral RNA fate. In eukaryotes, during stress conditions, mRNAs can be stored in subcellular compartments such as stress granules to stall their translation or in processing bodies to be degraded. Recent evidence indicates that these compartments also play an important role during the viral life cycle. Therefore, using immunofluorescence microscopy we set out to investigate how FCV and MNV infection regulate the formation of G3BP1- and PABP-1-containing stress granules and DCP-1-containing processing bodies to address whether these cytoplasmic granules could play a role during the viral life cycle. We have now shown that FCV has the ability to prevent stress granules formation during infection and that this is important for replication in CRFK and FEA cells. Using FCV-free supernatant from infected CRFK cells and immunofluorescence microscopy, we have also shown that during infection, the formation of stress granules is induced in a paracrine manner in uninfected cells via a messenger molecule released from infected cells. We hypothesize that this could reflect a new antiviral role for stress granules. Furthermore, MNV and FCV infection also led to the disruption of processing-bodies assembly. Overall, this study revealed that caliciviruses modulate the RNA granules during infection and that this could be part of viral mechanism to counteract the antiviral response.

No description available.

No description available.

No description available.

Assisted reproduction technologies remain inefficient, owing largely to poor oocyte quality. It is hypothesised that a more detailed understanding of the factors that influence the oocyte's metabolome and transcriptome in vivo and in vitro will lead to improvements in the diagnosis and treatment of female infertility. The objectives of this thesis were to (i) evaluate the impact of the follicular environment in natural cycles and following controlled ovarian stimulation (COS) on oocyte competence (i.e. quality) in vitro; (ii) identify cumulus cell markers of oocyte quality; and (iii) map the changes in oocyte carbohydrate metabolism and mitochondria] membrane potential (4Wm) during oogenesis. In vitro maturation (IVM) was conducted in 426 bovine oocytes from natural cycles and 102 oocytes were obtained from follicles after COS. Oocyte quality was quantified by meiotic progression to metaphase 11 (MII), by amino acid profiling (AAP) and by tracking the gamete's capacity to fertilise and develop into blastocyst. Oocytes from subordinate follicles, follicles ipsilateral to the corpus luteum, and follicles 5-9.5mm in diameter showed greater developmental potential. The consumption/production of serine, threonine, histidine and glutamine by bovine MII oocytes was predictive (68.9% accuracy) of the gametes capacity to support blastocyst development in vitro. A high ratio of inhibin 13a (INHBA) to follistatin (FST) mRNA expression in cumulus cells (CCs) also reflected the blastocyst potential of bovine oocytes. Analysis of the consumption/production of glucose, lactate and pyruvate by 292 ovine and 729 human follicles and 211 ovine and 275 human oocytes showed that energy metabolism by these cells relied on glucose and pyruvate oxidation, respectively, and this increased as development progressed. Oocyte mitochondria] i.'Pm remained stable through oogenesis. The results indicate that the follicle microenvironment profoundly influences oocyte metabolism and developmental potential in monovular species. Furthermore, follicular maturity, oocyte AAP and CC gene expression have been shown to be valuable markers of oocyte quality in vitro.

The peer-reviewed publications presented in this thesis are organized in seven 'themes’, each presenting a collation of related studies dealing with a particular aspect of structure, physiology, immunology, biology or toxicological pathology of the liver fluke Fasciola spp., or other helminth parasites of clinical / economic significance for domestic livestock and fish. Theme 1 addresses the development of in vitro methods for the study of Fasciola, including cytological and immunological investigations on adult worms and on juvenile migrating stages. In Theme 2, the structure and immunogenicity of the tegument of Fasciola is examined in detail, and the application of novel approaches such as immunogold labelling and monoclonal antibody technology is introduced. Theme 3 presents international collaborative studies on the biology, structure and ecology of paramphistomes (rumen flukes) of buffaloes, cattle, sheep and goats in India. The principles of population analysis for larval trematodes are further developed in Theme 4. The studies compiled in Theme 5 describe the structure and biology of certain trematode flukes that are parasitic on marine and freshwater food fishes in India. In Theme 6, a series of studies describing the morphological and ultractructural changes that occur in liver flukes subjected to treatment with anthelmintic drugs in vitro and in vivo present new evidence on the biochemical mode of action of such chemotherapeutic agents, and on the ability of fluke populations to develop drug resistance to them. Certain of these studies have revealed hitherto unreported aspects of fluke reproduction such as facultative parthenogenesis. In the studies collated in Theme 7, anthelmintic resistance and strategic control of helminth infections in ruminants in Northern Ireland are examined using archived diagnostic records and analysis of questionnaires distributed to farmers throughout the province. Theme 8 compiles published review articles in Parasitology, while Theme 9 presents a few studies unrelated to Parasitology.

Yersinia enterocolitica is a well known food-borne bacterium which belongs to the Enterobacteriaceae family. Y. enterocolitica is pathogenic for humans and it causes a wide spectrum of diseases ranging from self-limiting gastrointestinal diseases to fatal sepsis depending on the age and the immunity of the infected person. Oral ingestion of undercooked raw meat products, unpasteurised milk and contaminated water are the main sources of acquisition of this bacterium. Y. enterocolitica strains are categorized into six biotypes (1A, 1B, 2, 3, 4 &5). These six biotypes are further classified into high pathogenic, low pathogenic and non-pathogenic biotypes depending on their pathogenicity in the mouse infection model. This project compared the pathogenic potential of Yersinia enterocolitica high pathogenic, low pathogenic and non-pathogenic biotypes. The HEp-2 laryngitic human epithelial cell line and the alternative infection model Galleria mellonella greater wax moth larvae are the main infection models used to investigate the pathogenic potential of Y. enterocolitica biotypes. A diverse collection of Y. enterocolitica strains was used by which this strains collection is encompassing all six biotypes and all major serotypes. These strains were isolated from human and animal origins. Here novel results are presented showing new infection phenotypes of the Y. enterocolitica strains. All Y. enterocolitica biotypes strains were invasive to the HEp-2 epithelial cell line and pathogenic to the Galleria insect model. The non-pathogenic biotype was the most lethal in the Galleria model while the high pathogenic biotype shows little to no pathogenicity. The low pathogenic biotypes were moderately pathogenic to the Galleria insect model. The pYV plasmid showed a minor role in modulating the virulence in the Y. enterocolitica pathogenic biotypes. Investigating the in vivo Y. enterocolitica-Galleria interaction showed that Y. enterocolitica pathogenic biotypes bacterial cells were growing inside the Galleria larvae while the non-pathogenic biotype bacterial cells were dying. The heat-killed Y. enterocolitica strains were completely avirulent to the Galleria larvae.

Background and Methods: Blood transfusion saves lives and improves health but the presence of transfusion transmissible infections can have untoward consequences. When undetected, these infections can cause significant morbidity and mortality to transfusion recipients. On the other hand, a high prevalence of transfusion-transmitted infections (TTI) leading to rejection of a large proportion of donated blood can result in blood shortages and subsequent increase in mortality. Malaria and bacterial infections are transfusion transmissible but there is limited data concerning these infections in sub-Saharan Africa. Although the burden of transfusion-transmitted malaria in malaria endemic countries are unknown, it is recommended that all donated blood is screened for malaria parasites and presumptive treatment be given to transfusion recipients. Bacterial contamination in sub-Saharan Africa has been reported to occur in between 8 - 17% of stored blood but the effect of contamination on transfusion recipients has not been determined. Syphilis is currently the only bacterial infection for which routine screening is recommended but screening is not being performed in many blood centres including Komfo Anokye Teaching Hospital (KATH) in Kumasi, Ghana where this study took place. This study examined the effects of transfusion-transmitted malaria (TTM) and bacterial infections (including syphilis) on transfusion recipients in a malaria endemic area. Four malaria screening tests were compared to assess their usefulness in the context of African blood banks. Pregnant women, children and immune-compromised transfusion recipients from the Departments of Obstetrics and Gynaecology, Paediatrics, Medicine and Oncology in KATH were enrolled into the study. Results: Anti-malarial drugs were routinely prescribed with paediatric transfusions. Fifty patients were evaluated after receiving blood transfusions that were positive for P. falciparum by PCR and seven recipients developed PCR-detectable parasitaemia. In only one recipient (2%) was TTM confirmed. The prevalence of P. falciparum malaria in transfused blood was 4.7% (21/445) by microscopy, 13.7% (60/440) by rapid diagnostic test, 18% (78/436) by polymerase chain reaction and 22.2% (98/442) by enzyme immunoassay. Bacterial contamination was found in 11.5 %( 95% CI 7.0-16.0%) (23/200) of donated blood units but only half of the recipients were observed to developed adverse signs of transfusion related sepsis. The mean duration of storage of blood was 2 days. The prevalence of syphilis sero-positivity in donated blood was 8.0% (95% CI 4.3-11.7%). Seroconversion took place in an 8 year old girl, after receiving a syphilis sero-positive unit of blood. Conclusions: This thesis has shown that malaria parasites may be commonly detected in donor blood but TTM occurs infrequently in recipients living in malaria endemic areas. The high rate of bacterial contamination and its associated transfusion related sepsis poses a safety risk to transfusion recipients. Transfusion-transmitted syphilis remains a risk for transfusion recipients in blood centres with a high prevalence and short duration of storage of donor blood.

Primary infection with both cytomegalovirus (CMV) and varicella zoster virus (VZV) usually occurs at an early age, followed by lifelong persistent infection (latency phase). Indeed, frequent reactivation of latent viruses, particularly CMV, can be a major burden to the immune system and can play a role in configuration of the immune system. This thesis set out with the aim of understanding the balance between different arms of the immune system and CMV and also to find out the impact of immune modulation by this virus on the sequelae of coincident coronary artery disease (CAD). Meanwhile, the purpose of the VZV study was to determine the importance of adaptive immunity in protection against VZV reactivation in children with acute lymphocytic leukaemia receiving chemotherapy. The most obvious finding of the current CMV study is that the CMV-seropositive cohort exhibited a remarkable increase in the frequencies of CD28low CD4+, CD28low CD8+ T cells, NKG2Cpos NK Cells and Vδ2neg γδ T cells compared to CMV-seronegative donors. Furthermore, it shows that the expansion of Vδ2neg γδ T cells in CMV carriers is age related and does not correlate with changes in the magnitude of CMV-specific CD4+ or CD8+ T cell frequencies within those individuals. Phenotypically, Vδ2neg γδ T cells were found to be highly differentiated effector memory cells and did not possess ex vivo immediate effector function. In the CAD study, the most striking observation was the differences between AMI (acute myocardial infarction) and PMI (stable post myocardial infarction) patients in term of the distributions of polyfunctional CMV-specific CD4+ T cells that were characterized by the absence of IFNγ but expression of IL-2 plus any other measured functions (i.e CD107a, MIP1β, and/or TNFα). This cell subset was significantly increased in the PMI group. Finally, one of the more significant findings to emerge from the VZV study was the significant reduction in the absolute counts of VZV-specific IFNγ+ CD4 T cells and inversion of the VZV-specific CD4:CD8 (IFNγ producer) T cell ratio in those leukaemic children who developed clinical VZV infection. Contrasting with T cell responses, VZV-specific IgG was detected post primary infection in most patients; however, there was no clear association between the VZV-specific IgG antibody titre and VZV reactivation.