Vertical Transmission of HIV: Comparison of Policies in North and Sub-Saharan Africa

This project aims to investigate the extent to which differences in the design of national prevention of mother to child transmission policies and strategies between North Africa and Sub-Saharan Africa help to explain differences in vertical transmission rates between these two regions. To do so, I selected two case countries, one from each region, and conducted a qualitative document analysis of their HIV prevention policies since 2010, guided by two theoretical frameworks, the Analysis of Determinants of Policy Impacts (ADEPT) model of health promotion and the Health Policy Triangle. This project has revealed the importance of 1) civil society organizations in HIV prevention, 2) sufficient monitoring and evaluation systems and 3) the involvement of Intergovernmental Organizations (IGOs) in the strategic financing of HIV prevention strategies in low- and middle-income countries.M.Sc

Surveillance of Amoebic Keratitis-Causing Acanthamoebae for Potential Bacterial Endosymbionts in Ontario, Canada

Acanthamoeba spp. are the causative pathogens of several infections, including amoebic keratitis (AK), a vision-threatening infection. Acanthamoebae from corneal specimens of patients with AK harbor bacterial endosymbionts, which may increase virulence. We sought to understand the spectrum of bacterial endosymbionts present in clinical isolates of Acanthamoeba spp. identified in our reference parasitology laboratory. Isolates of Acanthamoeba spp. obtained from our biobank of anonymized corneal scrapings were screened for potential endosymbionts by PCR using primer pairs detecting bacteria belonging to orders Chlamydiales, Rickettsiales, or Legionellales and pan16S primers. Three primer pairs specific to the 18s rRNA gene of Acanthamoeba spp. were used for the amplification of Acanthamoeba DNA used for sequencing. Sanger sequencing of all PCR products was performed, followed by BLAST analysis for species identification. We screened 26 clinical isolates of Acanthamoeba spp. for potential endosymbionts. Five isolates (19%) were found to contain bacterial DNA belonging to Legionellales. Three (11%) contained members of the Rickettsiales and Pseudomonas genticulata was detected in a Rickettsia-positive sample. One strain (4%) contained Neochlamydia hartmannellae, a member of the Chlamydiales order. Bacterial endosymbionts are prevalent in clinical strains of Acanthamoeba causing AK isolated from corneal scrapings. The demonstration of these organisms in clinical Acanthamoeba isolates supports a potential exploration of anti-endosymbiont therapeutics as an adjuvant therapy in the treatment of AK

Development and validation of a real-time PCR assay for the detection of clinical acanthamoebae

Background: Suboptimal agreement between molecular assays for the detection of Acanthamoeba spp. in clinical specimens has been demonstrated, and poor assay sensitivity directly imperils the vision of those affected by amoebic keratitis (AK) through delayed diagnosis. We sought to develop and validate a single Taqman real time PCR assay targeting the Acanthamoeba 18S rRNA gene that could be used to enhance sensitivity and specificity when paired with reference assays. Methods: Biobanked DNA from surplus delinked AK clinical specimens and 10 ATCC strains of Acanthamoeba was extracted. Sequence alignment of 66 18S rRNA regions from 12 species of Acanthamoeba known to cause keratitis informed design of a new TaqMan primer set. Performance of the new assay was compared to the 2 assays used currently in our laboratory. Results: Among 24 Acanthamoeba-positive and 83 negative specimens by the CDC reference standard, performance characteristics of the newly designed primer set were as follows: sensitivity 100%, specificity 94%, PPV 82.8%, and NPV 100%. Compared to culture, sensitivity of the new primer set was 100%, and specificity 96%. No cross-reactivity of the primer set to non-acanthamoebae, including Balamuthia and Naegleria, was found. Conclusions: We have validated a real time PCR assay for the diagnosis of AK, and in doing so, have overcome important barriers to rapid and sensitive detection of acanthamoebae, including limited sensitivity and specificity of commonly used assays.Medicine, Faculty ofNon UBCReviewedFacult