In silico Identification of Vaccine Candidates against Viral Infections

There are many viral diseases without effective treatments or vaccines. These viruses can cause catastrophic epidemics such as the Lassa, Ebola, and Marburg viruses. Similarly, the recent coronavirus pandemic is of great concern as new variants are continuously emerging with decreased susceptibility to antibodies and vaccines that were developed for earlier strains. A critical step in the immune system’s fight against viruses involves an immunological protein molecule binding to a viral protein molecule. I investigate the atomic and molecular details of binding site recognition and binding interactions and dynamics for three important viruses. Antigens are molecules, such as viral proteins, that are foreign to the human body and can generate an immune response such as the production of antibody proteins to attack the antigen. An epitope is the part of an antigen molecule that is the site for antibody binding. They are categorized as T-cell or B-cell epitopes based upon which type of immunological cell can bind to the epitope. The B cell epitopes are surface segments of the antigen protein that are recognized by the immunological B-cells, whereas T-cell epitopes are peptides derived from antigens and recognized by immunological T-cells when the epitope is bound to a major histocompatibility complex molecule. The identification of epitopes is an essential step for the discovery and development of epitope-based vaccines. Experimental identification of epitopes involves expensive and time-consuming steps and therefore in-silico identification is a powerful tool to facilitate the identification of potential epitope candidates and can decrease the time and expense spent on validation experiments. I employed several epitope computational prediction methods that are based upon the antigen protein’s amino acid sequence and conformation for the glycoprotein of the Lassa virus as well as for different proteins of the Marburg virus. The predicted epitopes are further filtered based on a consensus approach that resulted in the identification of new epitopes that have not yet been tested experimentally. I performed molecular dynamics computational simulations on the most promising epitopes to determine atomic-level details of the epitope’s interactions and dynamics. In addition, I performed MD simulations to investigate the dynamics and antibody evasion behavior by the B.1.617.2 (delta) variant of SARS-CoV-2. I found that the receptor-binding β-loop-β motif in the spike protein adopts an altered conformation that causes binding difficulty for some of the neutralizing antibodies that were generated against the original coronavirus strain. This study reflects the possible mechanism for the immune evasion exhibited by the delta variant

Structural insights into the repair mechanism of AGT for methyl-induced DNA damage

Methylation induced DNA base-pairing damage is one of the major causes of cancer. O6-alkylguanine-DNA alkyltransferase (AGT) is considered a demethylation agent of the methylated DNA. Structural investigations with thermodynamic properties of the AGT-DNA complex are still lacking. In this report, we modeled two catalytic states of AGT-DNA interactions and an AGT-DNA covalent complex and explored structural features using molecular dynamics (MD) simulations. We utilized the umbrella sampling method to investigate the changes in the free energy of the interactions in two different AGT-DNA catalytic states, one with methylated GUA in DNA and the other with methylated CYS145 in AGT. These non-covalent complexes represent the pre- A nd post-repair complexes. Therefore, our study encompasses the process of recognition, complex formation, and separation of the AGT and the damaged (methylated) DNA base. We believe that the use of parameters for the amino acid and nucleotide modifications and for the protein-DNA covalent bond will allow investigations of the DNA repair mechanism as well as the exploration of cancer therapeutics targeting the AGT-DNA complexes at various functional states as well as explorations via stabilization of the complex

Feasibility of implementing public-private mix approach for tuberculosis case management in Pokhara Metropolitan City of western Nepal: a qualitative study

BackgroundThe Public-Private Mix (PPM) approach is a strategic initiative that involves engaging all private and public health care providers in the fight against tuberculosis using international health care standards. For tuberculosis control in Nepal, the PPM approach could be a milestone. This study aimed to explore the barriers to a public-private mix approach in the management of tuberculosis cases in Nepal.MethodsWe conducted key informant interviews with 20 participants, 14 of whom were from private clinics, polyclinics, and hospitals where the PPM approach was used, two from government hospitals, and four from policymakers. All data were audio-recorded, transcribed, and translated into English. The transcripts of the interviews were manually organized, and themes were generated and categorized into 1. TB case detection, 2. patient-related barriers, and 3. health-system-related barriers.ResultsA total of 20 respondents participated in the study. Barriers to PPM were identified into following three themes: (1) Obstacles related to TB case detection, (2) Obstacles related to patients, and (3) Obstacles related to health-care system. PPM implementation was challenged by following sub-themes that included staff turnover, low private sector participation in workshops, a lack of trainings, poor recording and reporting, insufficient joint monitoring and supervision, poor financial benefit, lack of coordination and collaboration, and non-supportive TB-related policies and strategies.ConclusionGovernment stakeholders can significantly benefit by applying a proactive role working with the private in monitoring and supervision. The joint efforts with private sector can then enable all stakeholders to follow the government policy, practice and protocols in case finding, holding and other preventive approaches. Future research are essential in exploring how PPM could be optimized

Scaling up tobacco cessation within TB programmes: findings from a multi-country, mixed-methods implementation study

BACKGROUND: Brief behavioural support can effectively help tuberculosis (TB) patients quit smoking and improve their outcomes. In collaboration with TB programmes in Bangladesh, Nepal and Pakistan, we evaluated the implementation and scale-up of cessation support using four strategies: (1) brief tobacco cessation intervention, (2) integration of tobacco cessation within routine training, (3) inclusion of tobacco indicators in routine records and (4) embedding research within TB programmes. METHODS: We used mixed methods of observation, interviews, questionnaires and routine data. We aimed to understand the extent and facilitators of vertical scale-up (institutionalization) within 59 health facility learning sites in Pakistan, 18 in Nepal and 15 in Bangladesh, and horizontal scale-up (increased coverage beyond learning sites). We observed training and surveyed all 169 TB health workers who were trained, in order to measure changes in their confidence in delivering cessation support. Routine TB data from the learning sites were analysed to assess intervention delivery and use of TB forms revised to report smoking status and cessation support provided. A purposive sample of TB health workers, managers and policy-makers were interviewed (Bangladesh n = 12; Nepal n = 13; Pakistan n = 19). Costs of scale-up were estimated using activity-based cost analysis. RESULTS: Routine data indicated that health workers in learning sites asked all TB patients about tobacco use and offered them cessation support. Qualitative data showed use of intervention materials, often with adaptation and partial implementation in busy clinics. Short (1–2 hours) training integrated within existing programmes increased mean confidence in delivering cessation support by 17% (95% CI: 14–20%). A focus on health system changes (reporting, training, supervision) facilitated vertical scale-up. Dissemination of materials beyond learning sites and changes to national reporting forms and training indicated a degree of horizontal scale-up. Embedding research within TB health systems was crucial for horizontal scale-up and required the dynamic use of tactics including alliance-building, engagement in the wider policy process, use of insider researchers and a deep understanding of health system actors and processes. CONCLUSIONS: System-level changes within TB programmes may facilitate routine delivery of cessation support to TB patients. These strategies are inexpensive, and with concerted efforts from TB programmes and donors, tobacco cessation can be institutionalized at scale

Health worker and patient views on implementation of smoking cessation in routine tuberculosis care

Böckmann M, Warsi S, Noor M, et al. Health worker and patient views on implementation of smoking cessation in routine tuberculosis care. NPJ primary care respiratory medicine. 2019;29(1): 34.Smoking worsens tuberculosis (TB) outcomes. Persons with TB who smoke can benefit from smoking cessation. We report findings of a multi-country qualitative process evaluation assessing barriers and facilitators to implementation of smoking cessation behaviour support in TB clinics in Bangladesh and Pakistan. We conducted semi-structured qualitative interviews at five case study clinics with 35 patients and 8 health workers over a period of 11 months (2017-2018) at different time points during the intervention implementation phase. Interviews were conducted by trained researchers in the native languages, audio-recorded, transcribed into English and analysed using a combined deductive-inductive approach guided by the Consolidated Framework for Implementation Research and Theoretical Domains Framework. All patients report willingness to quit smoking and recent quit attempts. Individuals' main motivations to quit are their health and the need to financially provide for a family. Behavioural regulation such as avoiding exposure to cigarettes and social influences from friends, family and colleagues are main themes of the interviews. Most male patients do not feel shy admitting to smoking, for the sole female patient interviewee stigma was an issue. Health workers report structural characteristics such as high workload and limited time per patient as primary barriers to offering behavioural support. Self-efficacy to discuss tobacco use with women varies by health worker. Systemic barriers to implementation such as staff workload and socio-cultural barriers to cessation like gender relations, stigma or social influences should be dealt with creatively to optimize the behaviour support for sustainability and scale-up

Complex consequences of conservation success: Emerging human-tiger conflicts in Nepal

Between 1910 and 2010, range-wide tiger populations plummeted from around 100,000 to an estimated 3,200. Poaching, habitat destruction and human-wildlife conflict have all contributed to this dramatic decrease. In Nepal, the Bardia-Banke Complex, consisting of Bardia National Park (BNP) and Banke National Park (BaNP) and their buffer-zones, is a critical habitat to the Bengal tiger Panthera tigris. Conservation efforts in recent decades have contributed to increasing the tiger population. Despite this increase, no human fatalities associated with human-tiger conflict were reported in the decade prior to 2019: a crucial feature of conservation success. In 2019, ten reported human-tiger incidents resulted in seven human fatalities, three people badly injured, and one tiger taken into captivity. The question of why conflicts involving humans and tigers have suddenly increased requires urgent answers to inform future policy. The development and implementation of case-specific coexistence strategies is a prerequisite to the conservation of this iconic species. This paper explores the complex unintended consequences of conservation success and critically evaluates the circumstances that may explain this recent surge in human deaths and injuries

Characteristic of tree cavities in sal <i>Shorea robusta</i> forest, Nepal

<p>Tree cavities are one of the key structures that influence abundance and diversity of cavity-dependent organisms in forests. The natural processes that lead to cavity formation and regulate the abundance of cavities are poorly understood, especially in subtropical regions of Asia. Surveys of tree cavities suitable for breeding birds were conducted in southern Nepal in sal <i>Shorea robusta</i> dominated forest. A cavity was defined as a hole in a tree trunk or a limb with a minimum entrance diameter of 2 cm × 2 cm, interior space, depth at least 2 cm and a solid bottom with a minimum diameter of 5 cm. Within 50 sample plots, we found 104 tree cavities. The mean height of cavity entrances (6.8 m above the ground) varied across tree species from 3.4 m in <i>Dillenia pentagyna</i> to 10.3 m in <i>S</i>. <i>robusta</i>, and was positively correlated with DBH of tree. The entrance exposures were most often to the south (38.2%), and located mainly in trunks (74.0%). Cavity entrances were very different in size, with mean smallest diameter 7.7 cm, the average depth of the cavity was 25.7 cm. Mode of cavity production (excavated or non-excavated) was best predicted by a model including diameter of cavity tree (DBH), cavity height and their interaction. Single check of cavities showed signs of use by birds in 21% of them. Excavated cavities accounted for 27.9% of all cavities, indicating that role of woodpeckers as cavity providers might be of importance for secondary users in sal forest.</p

Potential Autoimmunity Resulting from Molecular Mimicry between SARS-CoV-2 Spike and Human Proteins

Molecular mimicry between viral antigens and host proteins can produce cross-reacting antibodies leading to autoimmunity. The coronavirus SARS-CoV-2 causes COVID-19, a disease curiously resulting in varied symptoms and outcomes, ranging from asymptomatic to fatal. Autoimmunity due to cross-reacting antibodies resulting from molecular mimicry between viral antigens and host proteins may provide an explanation. Thus, we computationally investigated molecular mimicry between SARS-CoV-2 Spike and known epitopes. We discovered molecular mimicry hotspots in Spike and highlight two examples with tentative high autoimmune potential and implications for understanding COVID-19 complications. We show that a TQLPP motif in Spike and thrombopoietin shares similar antibody binding properties. Antibodies cross-reacting with thrombopoietin may induce thrombocytopenia, a condition observed in COVID-19 patients. Another motif, ELDKY, is shared in multiple human proteins, such as PRKG1 involved in platelet activation and calcium regulation, and tropomyosin, which is linked to cardiac disease. Antibodies cross-reacting with PRKG1 and tropomyosin may cause known COVID-19 complications such as blood-clotting disorders and cardiac disease, respectively. Our findings illuminate COVID-19 pathogenesis and highlight the importance of considering autoimmune potential when developing therapeutic interventions to reduce adverse reactions