Immunogenic assessment of plant-produced Human papillomavirus type 16 chimaeric L1:L2 virus-like particles and the production of an encapsidated therapeutic DNA vaccine candidate

Cervical cancer caused by infection with Human papillomavirus (HPV) is the 4th most common cancer in women globally, and results in an estimated 266 000 deaths every year. Current vaccines are based on the immunodominant L1 major capsid protein, which assembles into virus-like particles (VLPs) that are highly effective in type-specific prevention of cervical infection. However, these vaccines are produced in expensive cell culture systems, are type-specific and do not induce the regression of established infections. The cervical cancer burden (~80%) is mainly in developing countries due to limited healthcare resources, therefore there is a need for more broadly protective and affordable vaccines. Plants provide an alternative platform to produce cheaper vaccines, given their scalability, rapid production and low risk of contamination. The L2 minor capsid protein has sequence regions that are highly conserved across several HPV types, and HPV-16 L2 peptides 108-120, 65-81, 56-81 and 17-36 have been shown to elicit cross-neutralising antibodies. To increase the immunogenicity of L2, second-generation L1:L2 chimaeric VLP (cVLP) vaccines have been investigated. In this study, the 4 L2 peptides above were used to generate plant-produced HPV-16-derived L1:L2 chimaeras. The L2 epitopes were substituted into the DE loop of HPV-16 L1 at position 131 (SAC) or between the helix 4 and β-J structural region at position 431 (SAE). All chimaeras were transiently expressed in Nicotiana benthamiana via Agrobacterium-mediated transfer. Optimisation of expression was conducted by comparing protein expression levels over several days using 4 plant expression vectors, with the highest yields obtained by targeting protein to the chloroplast or with the use of a self-replicating vector. The chloroplast targeted SAC chimaeras predominantly assembled into higher order structures (T=1 VLPs and T=7 VLPs), whereas SAE chimaeras assembled into capsomeres or formed aggregates, indicating that the length, sequence and substitution position of L2 epitopes affects VLP assembly. All SAC chimaeras in addition to SAE 65-81 (smaller epitope not previously tested in chimaeras) were used in vaccination studies in mice, and their immunogenic potential analysed in pseudovirion-based neutralisation assays (PBNAs). Of the 7 heterologous HPVs tested, cross-neutralisation was observed with HPV-11, -18 and -58. Only the anti-SAE 65-81 serum showed neutralisation of homologous HPV-16, suggesting that antibodies detected from all candidate vaccines were mostly non-neutralising, and that the position of the L2 epitope display is critical to maintaining L1-specific neutralising epitopes. Lastly, to address the lack of therapeutic efficacy of current vaccines, I aimed to develop a novel E7 DNA vaccine delivered by plant-made pseudovirions (PsVs). A geminivirus-derived self-replicating plasmid encoding a shuffled E7 (E7SH) sequence that has no transformation ability but contains natural cytotoxic T-lymphocyte epitopes, was constructed using Goldenbraid technology and co-expressed in plants with HPV-16 or HPV-35 L1- and L2-encoding expression vectors. The pseudogenome was successfully encapsidated into plant-made PsVs. These PsVs were capable of infecting mammalian cells and encapsidated replicons expressed E7SH showing the promise of this candidate vaccine as a future combination prophylactic and therapeutic vaccine

How do customary practices enshrined in statutory law undermine women's access and rights to land? a case study of Yaw Pachi, Siaya District, Kenya

Abstract The study aimed to investigate the issue of women’s access and rights to land in Kenya. The study targeted Yaw Pachi women’s group, Siaya district who have experienced several problems of land tenure in terms of access and rights within their community which is Luo by tribe. The factors that influence these women’s access to land were also examined. The factors included Luo customary law, Luo customary practices, statutory laws as well as statutory institutions. The study also aimed to analyse the 2006 Draft National Land Policy by looking at what aspects of gender reform had been incorporated into the policy. The study examined the role of the land board as a statutory institution responsible for ensuring women and men have equal access and rights to land. In order to collect data, this study used qualitative method of social research. The researcher chose a small sample based on the research being conducted using a case study method. The sample was from an area where the phenomena such as customary laws and practices are prevalent. Qualitative research enables the researcher to collect and analyse in-depth information on a smaller group of respondents. Documentary analysis, interview techniques were used to gather data. The study population comprised of seventeen women from Yaw Pachi women’s group in Siaya district and twelve key informants. The key findings of the case study of Yaw Pachi women’s group shows that women can gain access to land in Siaya district mainly through marriage and by association with a male relative, who could be the woman’s father, father in-law, brother or son. Although the law of succession states that women can inherit land from their fathers, most findings revealed that this does not happen in reality. The 2006 Draft National Land Policy that intends to solve all the disparities women face when it comes to equity in land resources has been put on hold following an unsuccessful constitutional review in 2005.While the policy acknowledges that there are customs that discriminate against women, it also seeks to promote customary systems of land tenure. The Draft National Land Policy proposes a pluralist approach to land reform

Analysis of the Electronic Effects and Reactivity of Benzhydrols in the Formation of Benzhydryl Ethers

Benzhydryl ethers were synthesized through the use of microwave irradiation in a proto-ionic liquid solvent. The resulting products were separated from the reaction mixture by vacuum filtration with a silica gel plug. The products were analyzed using GCMS and 1 H NMR techniques to identify and quantify products. Analysis of the resultant data indicated the syntheses of the desired benzhydryl products were successful for 4,4-dimethoxybenzhydrol (conversion: 83% (1-propyl ether), 11% (2-propyl ether), 11% (menthyl ether)) and 4,4-dimethylbenzhydrol (conversion to desired product: 100% (1-propyl ether), 100% (2-propyl ether), 26% (menthyl ether)). However, the syntheses were unsuccessful for reactant 4,4-difluorobenzhydrol and benzhydrol. It was concluded that the electron-donating groups of 4,4-dimethoxybenzhydrol and 4,4-dimethylbenzhydrol aided in the formulation of a stable intermediate and subsequent desired product. The data support the hypothesized mechanism of protonation of the hydroxyl group of the benzhydrol with subsequent creation of a carbocation intermediate

Community health volunteers challenges and preferred income generating activities for sustainability: a qualitative case study of rural Kilifi, Kenya

Background: There is a global emphasis on engaging community health volunteers (CHVs) in low- to middle-income countries (LMICs) to reach to the vast underserved populations that live in rural areas. Retention of CHVs in most countries has however been difficult and turnover in many settings has been reported to be high with profound negative effects on continuity of community health services. In rural Kenya, high attrition among CHVs remains a concern. Understanding challenges faced by CHVs in rural settings and how to reduce attrition rates with sustainable income-generating activities (IGAs) is key to informing the implementation of contextual measures that can minimise high turnover. This paper presents findings on the challenges of volunteerism in community health and the preferred IGAs in rural Kilifi county, Kenya. Methods: The study employed qualitative methods. We conducted 8 key informant interviews (KIIs) with a variety of stakeholders and 10 focus group discussions (FGDs) with CHVs. NVIVO software was used to organise and analyse our data thematically. Results: Community Health Volunteers work is not remunerated and it conflicts with their economic activities, child care and other community expectations. In addition, lack of supervision, work plans and relevant training is a barrier to delivering CHVs’ work to the communities. There is a need to remunerate CHVs work as well as provide support in the form of basic training and capital on entrepreneurship to implement the identified income generating activities such as farming and events management. Conclusions: Strategies to support the livelihoods of CHVs through context relevant income generating activities should be identified and co-developed by the ministry of health and other stakeholders in consultation with the CHVs

Application of the Ultra-Poverty Graduation Model in understanding community health volunteers’ preferences for socio-economic empowerment strategies to enhance retention: a qualitative study in Kilifi, Kenya

Background: A significant shortage of healthcare workforce exists globally. To achieve Universal Healthcare cover- age, governments need to enhance their community-based health programmes. Community health volunteers (CHVs) are essential personnel in achieving this objective. However, their ability to earn a livelihood is compromised by the voluntary nature of their work; hence, the high attrition rates from community-based health programmes. There is an urgent need to support CHVs become economically self-reliant. We report here on the application of the Ultra-Poverty Graduation (UPG) Model to map CHVs’ preferences for socio-economic empowerment strategies that could enhance their retention in a rural area in Kenya. Methods: This study adopted an exploratory qualitative approach. Using a semi-structured questionnaire, we conducted 10 Focus Group Discussions with the CHVs and 10 Key Informant Interviews with County and Sub-county Ministry of Health and Ministry of Agriculture officials including multi-lateral stakeholders’ representatives from two sub-counties in the area. Data were audio-recorded and transcribed verbatim and transcripts analysed in NVivo. Researcher triangulation supported the first round of analysis. Findings were mapped and interpreted using a theory- driven analysis based on the six-step Ultra-Poverty Graduation Model. Results: We mapped the UPG Model’s six steps onto the results of our analyses as follows: (1) initial asset transfer of in-kind goods like poultry or livestock, mentioned by the CHVs as a necessary step; (2) weekly stipends with consump- tion support to stabilise consumption; (3) hands-on training on how to care for assets, start and run a business based on the assets transferred; (4) training on and facilitation for savings and financial support to build assets and instil financial discipline; (5) healthcare provision and access and finally (6) social integration. These strategies were pro- posed by the CHVs to enhance economic empowerment and aligned with the UPG Model. Conclusion: These results provide a user-defined approach to identify and assess strategic needs of and approaches to CHVs’ socio-economic empowerment using the UPG model. This model was useful in mapping the findings of our qualitative study and in enhancing our understanding on how these needs can be addressed in order to economically empower CHVs and enhance their retention in our setting

Optimizing a Human Papillomavirus Type 16 L1-Based Chimaeric Gene for Expression in Plants

Human papillomaviruses (HPVs) are the causative agents of cervical cancer, the fourth most prevalent cancer in women worldwide. The major capsid protein L1 self-assembles into virus-like particles (VLPs), even in the absence of the minor L2 protein: such VLPs have successfully been used as prophylactic vaccines. There remains a need, however, to develop cheaper vaccines that protect against a wider range of HPV types. The use of all or parts of the L2 minor capsid protein can potentially address this issue, as it has sequence regions conserved across several HPV types, which can elicit a wider spectrum of cross-neutralizing antibodies. Production of HPV VLPs in plants is a viable option to reduce costs; the use of a L1/L2 chimera which has previously elicited a cross-protective immune response is an option to broaden cross-protection. The objective of this study was to investigate the effect of codon optimization and of increasing the G+C content of synthetic L1/L2 genes on protein expression in plants. Additionally, we replaced varying portions of the 5′ region of the L1 gene with the wild type (wt) viral sequence to determine the effect of several negative regulatory elements on expression. We showed that GC-rich genes resulted in a 10-fold increase of mRNA levels and 3-fold higher accumulation of proteins. However, the highest increase of expression was achieved with a high GC-content human codon-optimized gene, which resulted in a 100-fold increase in mRNA levels and 8- to 9-fold increase in protein levels. Changing the 5′ end of the L1 gene back to its wt sequence decreased mRNA and protein expression. Our results suggest that the negative elements in the 5′ end of L1 are inadvertently destroyed by changing the codon usage, which enhances protein expression

Are Kenyans Likely to Use COVID-19 Self-Testing Kits? Results From a Cross-Sectional Survey

Objectives: To understand the public’s perceptions around rapid SARS-CoV-2 antigen self-testing in Kenya, including the drivers of acceptability, willingness to pay, and adherence to hygiene and prevention recommendations following a positive self-test. Methods: A household-based, cross-sectional survey, using a 35-item questionnaire, was conducted in Mombasa and Taita–Taveta counties, Kenya, during August 2021. Individuals aged ≥18 years were enrolled using a stratified sampling approach. Results: There were 419 participants (mean age 35.7 years). A minority (10.5%) had ever tested for SARS-CoV-2. If SARS-CoV-2 self-testing were available, 39.9% and 41.5% would be likely and very likely, respectively, to use it. If unavailable free-of-charge, 63.01% would pay for it. Multivariate analyses suggested that people in rural areas (Coefficient 0.30, 95%CI: 0.11–0.48, p = 0.002), aged 36–55 (Coefficient 0.21, 95%CI: 0.03–0.40, p = 0.023), and employed full time (Coefficient 0.32, 95%CI: 0.06–0.58, p = 0.016) would have more odds to adhere to recommended hygiene and prevention actions. Conclusion: SARS-CoV-2 self-testing was considered acceptable. Availability of self-testing could expand access to COVID-19 testing in Kenya, particularly among rural communities who have limited access to testing, and among mildly symptomatic individuals

Action to protect the independence and integrity of global health research

Storeng KT, Abimbola S, Balabanova D, et al. Action to protect the independence and integrity of global health research. BMJ GLOBAL HEALTH. 2019;4(3): e001746

Resilience of Traditional Livelihood Approaches Despite Forest Grabbing: Ogiek to the West of Mau Forest, Uasin Gishu County

This paper is a summary of the findings of research work conducted in two case studies in the Rift Valley, Kenya. This study used the Neo-Institutional theory to interrogate how the rules and regulations (institutions involved) of the agrarian reform process in Kenya are constantly changing and helping to shape the livelihoods of social actors around Mau Forest. The first case study—Ndungulu, is a settlement scheme where the Ogiek ethnic community were resettled between 1995 and 1997 after the land clashes of 1992. The second case study is the Kamuyu cooperative farm, a post-colonial settlement scheme owned by a cooperative society that was founded in 1965 by members from the Kikuyu ethnic group. This study employed qualitative data collection methods intermittently between 2012 and 2017 for a total of two years. A total of 60 interviews were conducted for this research. Thirteen (13) of these were key informant interviews with experts on land. The qualitative interviews were complemented by participant observations and nine focus group discussions. The qualitative data from the interviews and focus group discussions were transcribed, coded and analyzed thematically. Observations documented as field notes were also analyzed to complement the study findings. In this paper, the challenges, bargaining position and power play between social actors and government institutions implicated in the agrarian reform process in Kenya has been brought to the forefront. For instance, due to the structural issues that date back to the colonial period, the Ogiek have found innovative ways to maintain their daily existence (e.g., maintaining traditional methods of apiculture in Mau Forest). However, constraints in accessing forest land has resulted in them taking desperate measures, namely; selling off land to the Kalenjin in what is called “distress land sales”. On the contrary, the neighboring Kikuyu have maintained their land ownership status despite recurrent ethnic clashes that have occurred during general election years