Maya Phytomedicine in Guatemala - Can Cooperative Research Change Ethnopharmacological Paradigms?

ETHNOPHARMACOLOGICAL RELEVANCE: This paper presents one of the first large-scale collaborative research projects in ethnopharmacology, to bring together indigenous stakeholders and scientists both in project design and execution. This approach has often been recommended but rarely put into practice. The study was carried out in two key indigenous areas of Guatemala, for which very little ethnopharmacological fieldwork has been published. AIM OF THE STUDY: To document and characterize the ethno-pharmacopoeias of the Kaqchikel (highlands) and Q'eqchi' (lowlands) Maya in a transdisciplinary collaboration with the two groups Councils of Elders. MATERIALS AND METHODS: The project is embedded in a larger collaboration with five Councils of Elders representing important indigenous groups in Guatemala, two of which participated in this study. These suggested healing experts reputed for their phytotherapeutic knowledge and skills. Ethnobotanical fieldwork was carried out over 20 months, accompanied by a joint steering process and validation workshops. The field data were complemented by literature research and were aggregated using a modified version of the International Classification of Diseases (ICD-10) and Trotter & Logan's consensus index. RESULTS: Similar numbers of species were collected in the two areas, with a combined total of 530 species. This total does not represent all of the species used for medicinal purposes. Remedies for the digestive system, the central nervous system & behavioral syndromes, and general tissue problems & infections were most frequent in both areas. Furthermore, remedies for the blood, immune & endocrine system are frequent in the Kaqchikel area, and remedies for the reproductive system are frequent in the Q'eqchi' area. Consensus factors are however low. The Kaqchikel, in contrast to the Q'eqchi', report more remedies for non-communicable illnesses. They also rely heavily on introduced species. DISCUSSION & CONCLUSIONS: The transdisciplinary research design facilitated scientifically rigorous and societally relevant large-scale fieldwork, which is clearly beneficial to indigenous collaborators. It provided access and built trust as prerequisites for assembling the largest comparative ethnopharmacological collection, vastly extending knowledge on Maya phytotherapy. The collection represents knowledge of the two groups' most reputed herbalists and is a representative selection of the Guatemalan medicinal flora. ICD-10 proved useful for making broad comparisons between the groups, but more refined approaches would be necessary for other research objectives. Knowledge in the two areas is highly diverse and seems fragmented. New approaches are required to assess how coherent Maya phytotherapy is. The documented'traditional' ethno-pharmacopoeias demonstrate dynamic change and acculturation, reflecting the two linguistic groups' sociocultural history and context. This highlights the adaptive potential of phyto-therapeutic knowledge and calls the equation of local indigenous pharmacopoeias with'traditional' medicine into question. We suggest using the term'local' pharmacopoeias, and reserving the term'traditional' for the study of indigenous pharmacopoeias with a clear delineation of ancient knowledge

Green Health in Guatemala - How can we build mutual trust and partnerships for developing local medicines’ evidence-base and potential?

The implementation of access and benefit-sharing (ABS) protocols and especially the Nagoya Protocol has created new hurdles for collaborations around Indigenous Traditional Knowledge and international collaborations. Overall, these frameworks push for the development of novel collaborative North-South agendas in order to improve the fair distribution of benefits. The Green Health project (Guatemala) aims to implement a culturally pertinent and mutually accepted framework for sustainable use, access and benefit-sharing (ABS) of traditional medicinal plants. It involves developing a consensus among indigenous groups, government officials, industry, and academia. We describe steps undertaken to design and implement an intercultural transdisciplinary process that promotes trust building and advances herbal medicine research in a respectful and innovative way. This involves joint definition of goals and methods. The consortium co-researched Q’eqchi’ Maya traditional medicine, collected voucher specimens of medicinal plants with traditional healers, identified their taxa, and later developed a literature-based evaluation identifying species for potential product development. No samples for further research and development are collected. By applying the emic-etic concept, the project was able to understand the main drivers of each stakeholder and the associated obstacles for reaching an ABS agreement. This results in the emergence of potential new drivers for developing evidence-based herbal medicine from the perspective of academia, policy and cooperation and grass-roots indigenous movements

Bioluminescent Imaging of Trypanosoma brucei Shows Preferential Testis Dissemination Which May Hamper Drug Efficacy in Sleeping Sickness

Monitoring Trypanosoma spread using real-time imaging in vivo provides a fast method to evaluate parasite distribution especially in immunoprivileged locations. Here, we generated monomorphic and pleomorphic recombinant Trypanosoma brucei expressing the Renilla luciferase. In vitro luciferase activity measurements confirmed the uptake of the coelenterazine substrate by live parasites and light emission. We further validated the use of Renilla luciferase-tagged trypanosomes for real-time bioluminescent in vivo analysis. Interestingly, a preferential testis tropism was observed with both the monomorphic and pleomorphic recombinants. This is of importance when considering trypanocidal drug development, since parasites might be protected from many drugs by the blood-testis barrier. This hypothesis was supported by our final study of the efficacy of treatment with trypanocidal drugs in T. brucei-infected mice. We showed that parasites located in the testis, as compared to those located in the abdominal cavity, were not readily cleared by the drugs

Infected Dendritic Cells Facilitate Systemic Dissemination and Transplacental Passage of the Obligate Intracellular Parasite Neospora caninum in Mice

The obligate intracellular parasite Neospora caninum disseminates across the placenta and the blood-brain barrier, to reach sites where it causes severe pathology or establishes chronic persistent infections. The mechanisms used by N. caninum to breach restrictive biological barriers remain elusive. To examine the cellular basis of these processes, migration of different N. caninum isolates (Nc-1, Nc-Liverpool, Nc-SweB1 and the Spanish isolates: Nc-Spain 3H, Nc-Spain 4H, Nc-Spain 6, Nc-Spain 7 and Nc-Spain 9) was studied in an in vitro model based on a placental trophoblast-derived BeWo cell line. Here, we describe that infection of dendritic cells (DC) by N. caninum tachyzoites potentiated translocation of parasites across polarized cellular monolayers. In addition, powered by the parasite's own gliding motility, extracellular N. caninum tachyzoites were able to transmigrate across cellular monolayers. Altogether, the presented data provides evidence of two putative complementary pathways utilized by N. caninum, in an isolate-specific fashion, for passage of restrictive cellular barriers. Interestingly, adoptive transfer of tachyzoite-infected DC in mice resulted in increased parasitic loads in various organs, e.g. the central nervous system, compared to infections with free parasites. Inoculation of pregnant mice with infected DC resulted in an accentuated vertical transmission to the offspring with increased parasitic loads and neonatal mortality. These findings reveal that N. caninum exploits the natural cell trafficking pathways in the host to cross cellular barriers and disseminate to deep tissues. The findings are indicative of conserved dissemination strategies among coccidian apicomplexan parasites

In Vivo Bioluminescent Imaging (BLI): Noninvasive Visualization and Interrogation of Biological Processes in Living Animals

In vivo bioluminescent imaging (BLI) is increasingly being utilized as a method for modern biological research. This process, which involves the noninvasive interrogation of living animals using light emitted from luciferase-expressing bioreporter cells, has been applied to study a wide range of biomolecular functions such as gene function, drug discovery and development, cellular trafficking, protein-protein interactions, and especially tumorigenesis, cancer treatment, and disease progression. This article will review the various bioreporter/biosensor integrations of BLI and discuss how BLI is being applied towards a new visual understanding of biological processes within the living organism

The role of DNA microarrays in Toxoplasma gondii research, the causative agent of ocular toxoplasmosis

Ocular toxoplasmosis, which is caused by the protozoan parasite Toxoplasma gondii, is the leading cause of retinochoroiditis. Toxoplasma is an obligate intracellular pathogen that replicates within a parasitophorous vacuole. Infections are initiated by digestion of parasites deposited in cat feces or in undercooked meat. Parasites then disseminate to target tissues that include the retina where they then develop into long-lived asymptomatic tissue cysts. Occasionally, cysts reactivate and growth of newly emerged parasites must be controlled by the host’s immune system or disease will occur. The mechanisms by which Toxoplasma grows within its host cell, encysts, and interacts with the host’s immune system are important questions. Here, we will discuss how the use of DNA microarrays in transcriptional profiling, genotyping, and epigenetic experiments has impacted our understanding of these processes. Finally, we will discuss how these advances relate to ocular toxoplasmosis and how future research on ocular toxoplasmosis can benefit from DNA microarrays

Effectiveness of One Health approach for control of Kyasanur Forest Disease in Wayanad, Kerala, India

Background & objectives: Kyasanur Forest Disease (KFD) is a vector borne haemorrhagic fever that is endemic in the Wayanad region located in Northern part of Kerala, India. The region is managing the outbreak well ever since the major epidemic of 2015. This was because of the successful implementation of One Health (OH) initiative concentrating on multisectoral collaboration between regional institutions involved in public, animal and environmental health domains. The article presents how OH was implemented for the first time in the district in the year 2015 and evaluates the degree OH-ness of the Initiative. Methods: The OH approach involved trans-disciplinary stakeholder meetings and reviews, outbreak management and integrated surveillance targeting ticks, monkeys and humans. The degree of OH-ness used for addressing KFD during the year 2015 was evaluated following the protocol developed by the Network for Evaluation of One Health (NEOH). In detail, we (i) described the OH initiative and its system (Aim, stakeholders, action strategy) and (ii) scored different aspects of this initiative (i.e., OH-thinking, -planning, -working, -sharing, -learning, -organization), with values from 0 (=no OH approach) to 1 (=perfect OH approach). Results: We obtained a median score for each aspect evaluated. We reached high scores for OH systemic organization (1.0), OH thinking (0.83) and OH working (0.83). Lower scores were attributed to OH planning (0.58), OH sharing (0.50) and OH learning (0.33). The OH index was 0.36 and OH ratio was 0.95, indicating a balance between the OH operations and supporting infrastructures. Interpretation & conclusion: With this we could high-light some critical issues related to communication on sharing data as well as learning gaps for consideration to control future outbreaks. The strengths and weaknesses detected may be used to refine the initiative, aiming to provide a basis for the development of shared recommendations in a more OH-oriented perspective. This model of evaluation criteria will serve to create a database of OH success stories in India that will in turn help to institutionalize the approach at ministerial level. Future India is moving towards implementing a One Health, hence, this study data will provide an ideal opportunity for all sectors to control any vector borne diseases

One Health governance: knowledge integration in One Health policy formulation, implementation and evaluation

The One Health concept covers the interrelationship between human, animal and environmental health and requires multistakeholder collaboration across many cultural, disciplinary, institutional and sectoral boundaries. Yet, the implementation of the One Health approach appears hampered by shortcomings in the global framework for health governance. Knowledge integration approaches, at all stages of policy development, could help to address these shortcomings. e identication of key objectives, the resolving of trade-os and the creation of a common vision and a common direction can be supported by multicriteria analyses. Evidence-based decision-making and transformation of observations into narratives detailing how situations emerge and might unfold in the future can be achieved by systems thinking. Finally, transdisciplinary approaches can be used both to improve the eectiveness of existing systems and to develop novel networks for collective action. To strengthen One Health governance, we propose that knowledge integration becomes a key feature of all stages in the development of One-Health-related policies. We suggest several ways in which such integration could be promoted

Characterization of CD8+ T-cell response in acute and resolved hepatitis A virus infection.

Abstract BACKGROUND & AIMS: In contrast to the infection with other hepatotropic viruses, hepatitis A virus (HAV) always causes acute self-limited hepatitis, although the role for virus-specific CD8 T cells in viral containment is unclear. Herein, we analyzed the T cell response in patients with acute hepatitis by utilizing a set of overlapping peptides and predicted HLA-A2 binders from the polyprotein. METHODS: A set of 11 predicted peptides from the HAV polyprotein, identified as potential binders, were synthesized. Peripheral blood mononuclear cells (PBMCs) from patients were tested for IFNγ secretion after stimulation with these peptides and ex vivo with HLA-A2 tetramers. Phenotyping was carried out by staining with the activation marker CD38 and the memory marker CD127. RESULTS: Eight out of 11 predicted HLA-A2 binders showed a high binding affinity and five of them were recognized by CD8+ T cells from patients with hepatitis A. There were significant differences in the magnitude of the responses to these five peptides. One was reproducibly immunodominant and the only one detectable ex vivo by tetramer staining of CD8+ T cells. These cells have an activated phenotype (CD38hi CD127lo) during acute infection. Three additional epitopes were identified in HLA-A2 negative patients, most likely representing epitopes restricted by other HLA-class I-alleles (HLA-A11, B35, B40). CONCLUSIONS: Patients with acute hepatitis A have a strong multi-specific T cell response detected by ICS. With the tetramer carrying the dominant HLA-A2 epitope, HAV-specific and activated CD8+ T cells could be detected ex vivo. This first description of the HAV specific CTL-epitopes will allow future studies on strength, breadth, and kinetics of the T-cell response in hepatitis A

Differential proteasomal processing of hydrophobic and hydrophilic protein regions: Contribution to cytotoxic T lymphocyte epitope clustering in HIV-1-Nef

HIV proteins contain a multitude of naturally processed cytotoxic T lymphocyte (CTL) epitopes that concentrate in clusters. The molecular basis of epitope clustering is of interest for understanding HIV immunogenicity and for vaccine design. We show that the CTL epitope clusters of HIV proteins predominantly coincide with hydrophobic regions, whereas the noncluster regions are predominantly hydrophilic. Analysis of the proteasomal degradation products of full-length HIV-Nef revealed a differential sensitivity of cluster and noncluster regions to proteasomal processing. Compared with the epitope-scarce noncluster regions, cluster regions are digested by proteasomes more intensively and with greater preference for hydrophobic P1 residues, resulting in substantially greater numbers of fragments with the sizes and COOH termini typical of epitopes and their precursors. Indeed, many of these fragments correspond to endogenously processed Nef epitopes and/or their potential precursors. The results suggest that differential proteasomal processing contributes importantly to the clustering of CTL epitopes in hydrophobic regions