An investigation of tourism stakeholder networks and cluster sustainability in Samui Island, Thailand

This research focuses on main challenges and opportunities in developing sustainable tourism in Samui Island, one of the well-known tourist destinations in Thailand. The island is rich in environmental and cultural resources as well as being a popular coastal tourism resort generating considerable revenues (approx. GBP 700 million in 2014). However, serious environmental impacts due to mass tourism threaten not only the island‘s tourism industry in, but also the ecology of the region and the long-term well-being of its residents. Thus, this research aims to analyse the interrelationships between tourism stakeholders and how their relational networks influence (un)sustainable practices and shape tourism clusters and sustainable tourism development. This work draws upon empirical data collected in two study regions - in Samui – sub-district Bo Phud situated in the north-east, and two sub-districts of Na Muang and Taling Ngam in the south-west.The research used and applied key concepts inherent in stakeholder theory, social networks approach, and the notion of clusters, to examine the micro-dynamics underpinning (un)sustainable tourism practices in Samui Island. A qualitative approach was utilised for the research investigation through 60 semi-structured interviews and 4 focus groups. The interview participants were policy makers, such as government bodies, and public- tourism organisations, tourism businesses, tourism associations, non-profit organisation, and local communities.Key findings show the significance of existing collaborative networks between public-private sectors and non-profit organisation which demonstrate both (in) formal networks as well as strong and weak relationships. Social networks enable innovative tourism products and activities, as well as green projects. However, these collaborative networks were still limited in some particular groups such as upmarket hotels and resorts. Other groups of stakeholders were in peripheral positions in tourism networks, especially local communities which lacked connections and participations in the tourism development process. Key challenges in relation to networking included lack of trust towards government bodies, tension between outsiders (both Thai and foreigners ) and insiders (Samui local people), and lack of time and commitment. Also key challenges towards sustainable tourism implementation arise due to the lack of financial resources, and social networks that are in some cases mobilised for unsustainable practices such as corruptions, and by the networks of businesses that are not licensed or are unregulated, as well as 'dark networks' such as sex workers, pimps and clusters of drugs dealers

Removal of Heterologous Sequences from Plasmodium falciparum Mutants Using FLPe-Recombinase

Genetically-modified mutants are now indispensable Plasmodium gene-function reagents, which are also being pursued as genetically attenuated parasite vaccines. Currently, the generation of transgenic malaria-parasites requires the use of drug-resistance markers. Here we present the development of an FRT/FLP-recombinase system that enables the generation of transgenic parasites free of resistance genes. We demonstrate in the human malaria parasite, P. falciparum, the complete and efficient removal of the introduced resistance gene. We targeted two neighbouring genes, p52 and p36, using a construct that has a selectable marker cassette flanked by FRT-sequences. This permitted the subsequent removal of the selectable marker cassette by transient transfection of a plasmid that expressed a 37°C thermostable and enhanced FLP-recombinase. This method of removing heterologous DNA sequences from the genome opens up new possibilities in Plasmodium research to sequentially target multiple genes and for using genetically-modified parasites as live, attenuated malaria vaccines

Antiprotozoal Activities of Organic Extracts from French Marine Seaweeds

Marine macrophytes contain a variety of biologically active compounds, some reported to have antiprotozoal activity in vitro. As a part of a screening program to search for new natural antiprotozoals, we screened hydroalcoholic and ethyl acetate extracts of 20 species of seaweeds from three phyla (Rhodophyta, Heterokontophyta and Chlorophyta), sampled along the Normandy (France) coast. We tested them in vitro against the protozoa responsible for three major endemic parasitic diseases: Plasmodium falciparum, Leishmania donovani and Trypanosoma cruzi. The selectivity of the extracts was also evaluated by testing on a mammalian cell line (L6 cells). Ethyl acetate extracts were more active than hydroalcoholic ones. Activity against T. cruzi and L. donovani was non-existent to average, but almost half the extracts showed good activity against P. falciparum. The ethyl acetate extract of Mastocarpus stellatus showed the best antiplasmodial activity as well as the best selectivity index (IC50 = 2.8 μg/mL; SI > 30). Interestingly, a red algae species, which shares phylogenetic origins with P. falciparum, showed the best antiplasmodial activity. This study is the first to report comparative antiprotozoal activity of French marine algae. Some of the species studied here have not previously been biologically evaluated

Guanylyl cyclase activity associated with putative bifunctional integral membrane proteins in Plasmodium falciparum.

We report here that guanylyl cyclase activity is associated with two large integral membrane proteins (PfGCalpha and PfGCbeta) in the human malaria parasite Plasmodium falciparum. Unusually, the proteins appear to be bifunctional; their amino-terminal regions have strong similarity with P-type ATPases, and the sequence and structure of the carboxyl-terminal regions conform to that of G protein-dependent adenylyl cyclases, with two sets of six transmembrane sequences, each followed by a catalytic domain (C1 and C2). However, amino acids that are enzymatically important and present in the C2 domain of mammalian adenylyl cyclases are located in the C1 domain of the P. falciparum proteins and vice versa. In addition, certain key residues in these domains are more characteristic of guanylyl cyclases. Consistent with this, guanylyl cyclase activity was obtained following expression of the catalytic domains of PfGCbeta in Escherichia coli. In P. falciparum, expression of both genes was detectable in the sexual but not the asexual blood stages of the life cycle, and PfGCalpha was localized to the parasite/parasitophorous vacuole membrane region of gametocytes. The profound structural differences identified between mammalian and parasite guanylyl cyclases suggest that aspects of this signaling pathway may be mechanistically distinct

A genetically attenuated malaria vaccine candidate based on P. falciparum b9/slarp gene-deficient sporozoites

A highly efficacious pre-erythrocytic stage vaccine would be an important tool for the control and elimination of malaria but is currently unavailable. High-level protection in humans can be achieved by experimental immunization with Plasmodium falciparum sporozoites attenuated by radiation or under anti-malarial drug coverage. Immunization with genetically attenuated parasites (GAP) would be an attractive alternative approach. Here we present data on safety and protective efficacy using sporozoites with deletions of two genes i.e. the newly identified b9 and slarp, which govern independent and critical processes for successful liver-stage development. In the rodent malaria model, Pb Delta b9 Delta slarpGAP was completely attenuated showing no breakthrough infections while efficiently inducing high level protection. The human Pf Delta b9 Delta slarpGAP generated without drug-resistance markers were infective to human hepatocytes in vitro and to humanized mice engrafted with human hepatocytes in vivo but completely aborted development after infection. These findings support the clinical development of a Pf Delta b9 Delta slarpSPZ vaccine

Caracterização de cepas de P. falciparum coletadas de pacientes recrudescentes

A caracterização de cepas de P. falciparum coletadas de três pacientes internados no Centro de Provas Clínico-Farmacológicas do Hospital Barros Barreto (Belém, PA), através da eletroforese em acetato de celulose para duas enzimas (GPI e ADA), assim como através de testes de sensibilidade para duas drogas, cloroquina e mefloquina, permitiram observações da variação dos tipos enzimáticos em um dos pacientes e modificação no nível de sensibilidade à cloroquina em outro, tanto em amostras obtidas da infecção original, como em cada uma das recrudescências

Evolutionary biology and the avoidance of antimicrobial resistance

Evolutionary biologists have largely left the search for solutions to the drug resistance crisis to biomedical scientists, physicians, veterinarians and public health specialists. We believe this is because the vast majority of professional evolutionary biologists consider the evolutionary science of drug resistance to be conceptually uninteresting. Using malaria as case study, we argue that it is not. We review examples of evolutionary thinking that challenge various fallacies dominating antimalarial therapy, and discuss open problems that need evolutionary insight. These problems are unlikely to be resolved by biomedical scientists ungrounded in evolutionary biology. Involvement by evolutionary biologists in the science of drug resistance requires no intellectual compromises: the problems are as conceptually challenging as they are important

Resistance of a Rodent Malaria Parasite to a Thymidylate Synthase Inhibitor Induces an Apoptotic Parasite Death and Imposes a Huge Cost of Fitness

BACKGROUND: The greatest impediment to effective malaria control is drug resistance in Plasmodium falciparum, and thus understanding how resistance impacts on the parasite's fitness and pathogenicity may aid in malaria control strategy. METHODOLOGY/PRINCIPAL FINDINGS: To generate resistance, P. berghei NK65 was subjected to 5-fluoroorotate (FOA, an inhibitor of thymidylate synthase, TS) pressure in mice. After 15 generations of drug pressure, the 2% DT (the delay time for proliferation of parasites to 2% parasitaemia, relative to untreated wild-type controls) reduced from 8 days to 4, equalling the controls. Drug sensitivity studies confirmed that FOA-resistance was stable. During serial passaging in the absence of drug, resistant parasite maintained low growth rates (parasitaemia, 15.5%±2.9, 7 dpi) relative to the wild-type (45.6%±8.4), translating into resistance cost of fitness of 66.0%. The resistant parasite showed an apoptosis-like death, as confirmed by light and transmission electron microscopy and corroborated by oligonucleosomal DNA fragmentation. CONCLUSIONS/SIGNIFICANCE: The resistant parasite was less fit than the wild-type, which implies that in the absence of drug pressure in the field, the wild-type alleles may expand and allow drugs withdrawn due to resistance to be reintroduced. FOA resistance led to depleted dTTP pools, causing thymineless parasite death via apoptosis. This supports the tenet that unicellular eukaryotes, like metazoans, also undergo apoptosis. This is the first report where resistance to a chemical stimulus and not the stimulus itself is shown to induce apoptosis in a unicellular parasite. This finding is relevant in cancer therapy, since thymineless cell death induced by resistance to TS-inhibitors can further be optimized via inhibition of pyrimidine salvage enzymes, thus providing a synergistic impact. We conclude that since apoptosis is a process that can be pharmacologically modulated, the parasite's apoptotic machinery may be exploited as a novel drug target in malaria and other protozoan diseases of medical importance

Comparative Transcriptional and Genomic Analysis of Plasmodium falciparum Field Isolates

Mechanisms for differential regulation of gene expression may underlie much of the phenotypic variation and adaptability of malaria parasites. Here we describe transcriptional variation among culture-adapted field isolates of Plasmodium falciparum, the species responsible for most malarial disease. It was found that genes coding for parasite protein export into the red cell cytosol and onto its surface, and genes coding for sexual stage proteins involved in parasite transmission are up-regulated in field isolates compared with long-term laboratory isolates. Much of this variability was associated with the loss of small or large chromosomal segments, or other forms of gene copy number variation that are prevalent in the P. falciparum genome (copy number variants, CNVs). Expression levels of genes inside these segments were correlated to that of genes outside and adjacent to the segment boundaries, and this association declined with distance from the CNV boundary. This observation could not be explained by copy number variation in these adjacent genes. This suggests a local-acting regulatory role for CNVs in transcription of neighboring genes and helps explain the chromosomal clustering that we observed here. Transcriptional co-regulation of physical clusters of adaptive genes may provide a way for the parasite to readily adapt to its highly heterogeneous and strongly selective environment