Life cycle studies of the hexose transporter of Plasmodium species and genetic validation of their essentiality

A Plasmodium falciparum hexose transporter (PfHT) has previously been shown to be a facilitative glucose and fructose transporter. Its expression in Xenopus laevis oocytes and the use of a glucose analogue inhibitor permitted chemical validation of PfHT as a novel drug target. Following recent re-annotations of the P. falciparum genome, other putative sugar transporters have been identified. To investigate further if PfHT is the key supplier of hexose to P. falciparum and to extend studies to different stages of Plasmodium spp., we functionally analysed the hexose transporters of both the human parasite P. falciparum and the rodent parasite Plasmodium berghei using gene targeting strategies. We show here the essential function of pfht for the erythrocytic parasite growth as it was not possible to knockout pfht unless the gene was complemented by an episomal construct. Also, we show that parasites are rescued from the toxic effect of a glucose analogue inhibitor when pfht is overexpressed in these transfectants. We found that the rodent malaria parasite orthologue, P. berghei hexose transporter (PbHT) gene, was similarly refractory to knockout attempts. However, using a single cross-over transfection strategy, we generated transgenic P. berghei parasites expressing a PbHT–GFP fusion protein suggesting that locus is amenable for gene targeting. Analysis of pbht-gfp transgenic parasites showed that PbHT is constitutively expressed through all the stages in the mosquito host in addition to asexual stages. These results provide genetic support for prioritizing PfHT as a target for novel antimalarials that can inhibit glucose uptake and kill parasites, as well as unveiling the expression of this hexose transporter in mosquito stages of the parasite, where it is also likely to be critical for survival

An Expanded Chronic Care Management approach to Multiple Chronic Conditions in Hispanics Using Community Health Workers as Community Extenders in the Rio Grande Valley of Texas

INTRODUCTION: The synergistic negative effects of type 2 diabetes (T2DM) and hypertension increases all-cause mortality and the medical complexity of management, which disproportionately impact Hispanics who face barriers to healthcare access. The Salud y Vida intervention was delivered to Hispanic adults living along the Texas-Mexico Border with comorbid poorly controlled T2DM and hypertension. The Salud y Vida multicomponent intervention incorporated community health workers (CHWs) into an expanded chronic care management model to deliver home-based follow-up visits and provided community-based diabetes self-management education. METHODS: We conducted multivariable longitudinal analysis to examine the longitudinal intervention effect on reducing systolic and diastolic blood pressure among 3806 participants enrolled between 2013 and 2019. Participants were compared according to their program participation as either higher (≥ 10 combined educational classes and CHW visits) or lower engagement (\u3c10 \u3eencounters). Data was collected between 2013 and 2020. RESULTS: Baseline mean systolic and diastolic blood pressure were 138 and 81 mmHg respectively. There were overall improvements in systolic (-6.49; 95% CI = [-7.13, -5.85]; p \u3c 0.001) and diastolic blood pressure (-3.97; 95% CI = [-4.37, -3.56]; p \u3c 0.001). The higher engagement group had greater systolic blood pressure reduction at 3 months (adjusted mean difference = -1.8 mmHg; 95% CI = [-3.2, -0.3]; p = 0.016) and at 15 month follow-up (adjusted mean difference = -2.3 mmHg; 95% CI = [-4.2, -0.39]; p = 0.0225) compared to the lower engagement group. CONCLUSION: This intervention, tested and delivered in a real-world setting, provides an example of how CHW integration into an expanded chronic care model can improve blood pressure outcomes for individuals with co-morbidities

SAS6-like protein in Plasmodium indicates that conoid-associated apical complex proteins persist in invasive stages within the mosquito vector

The SAS6-like (SAS6L) protein, a truncated paralogue of the ubiquitous basal body/centriole protein SAS6, has been characterised recently as a flagellum protein in trypanosomatids, but associated with the conoid in apicomplexan Toxoplasma. The conoid has been suggested to derive from flagella parts, but is thought to have been lost from some apicomplexans including the malaria-causing genus Plasmodium. Presence of SAS6L in Plasmodium, therefore, suggested a possible role in flagella assembly in male gametes, the only flagellated stage. Here, we have studied the expression and role of SAS6L throughout the Plasmodium life cycle using the rodent malaria model P. berghei. Contrary to a hypothesised role in flagella, SAS6L was absent during gamete flagellum formation. Instead, SAS6L was restricted to the apical complex in ookinetes and sporozoites, the extracellular invasive stages that develop within the mosquito vector. In these stages SAS6L forms an apical ring, as we show is also the case in Toxoplasma tachyzoites. The SAS6L ring was not apparent in blood-stage invasive merozoites, indicating that the apical complex is differentiated between the different invasive forms. Overall this study indicates that a conoid-associated apical complex protein and ring structure is persistent in Plasmodium in a stage-specific manner

SAM domain-dependent activity of PfTKL3, an essential tyrosine kinase-like kinase of the human malaria parasite Plasmodiumfalciparum

Over the last decade, several protein kinases inhibitors have reached the market for cancer chemotherapy. The kinomes of pathogens represent potentially attractive targets in infectious diseases. The functions of the majority of protein kinases of Plasmodium falciparum, the parasitic protist responsible for the most virulent form of human malaria, remain unknown. Here we present a thorough characterisation of PfTKL3 (PF13_0258), an enzyme that belongs to the tyrosine kinase-like kinase (TKL) group. We demonstrate by reverse genetics that PfTKL3 is essential for asexual parasite proliferation in human erythrocytes. PfTKL3 is expressed in both asexual and gametocytes stages, and in the latter the protein co-localises with cytoskeleton microtubules. Recombinant PfTKL3 displays in vitro autophosphorylation activity and is able to phosphorylate exogenous substrates, and both activities are dramatically dependent on the presence of an N-terminal “sterile α-motif” domain. This study identifies PfTKL3 as a validated drug target amenable to high-throughput screening

Addressing the high cervical cancer rates along the Texas-Mexico border through community outreach, patient navigation, and provider training/telementoring

Objective: Cervical cancer incidence and mortality rates are 68% and 57% higher, respectively, along the Texas-Mexico border compared with the rest of the United States. This is likely due to a combination of low health literacy, limited access to affordable screening, and a lack of trained personnel to perform colposcopy, loop electrosurgical excision procedures (LEEP), and appropriate management of women with pre-invasive disease. The objective of our study was to increase cervical cancer screening, diagnosis, and treatment rates in the Rio Grande Valley (RGV). Method: We initiated a comprehensive program at two health centers and one mobile clinic in the RGV consisting of (1) a public education program designed for community health workers to teach women about cervical cancer screening and HPV vaccination coupled with patient navigation to participating clinics; (2) colposcopy and LEEP training for physicians and advanced-practice providers through locally held hands-on courses and mentoring program; and (3) implementation of Project ECHO (Extension for Community Health Outcomes), a well-established telementoring program using video conferencing to connect academic specialists with community providers for case-based learning. We compared screening, diagnosis, and treatment rates pre- and post-program implementation. Results: From November 2014 to June 2018, local providers screened 19,028 women with Pap ± HPV testing (baseline 12,460, 53% increase); performed colposcopy on 2,644 women with abnormal screening results (baseline 945, 180% increase); and performed 483 LEEP procedures for treatment of cervical dysplasia (baseline 0). Ten women were diagnosed with invasive cancer and navigated to one of the participating gynecologic oncologists for treatment (baseline N/A). Five additional providers in the RGV completed the mentoring program to be certified to perform colposcopy (100% increase from baseline of 5) and two additional providers to perform LEEP (baseline 0). ECHO telementoring video conferences have been held every two weeks for a total 94 sessions (average of 22 participants/session) with 182 patient cases presented and discussed. Conclusion: Our comprehensive approach has led to an increase in the number of women undergoing cervical cancer screening and diagnosis/treatment of dysplasia. If sustained, we anticipate these efforts will decrease cervical cancer rates in the RGV. The program is currently being expanded to additional medically underserved regions of Texas

How does community context influence coalitions in the formation stage? a multiple case study based on the Community Coalition Action Theory

<p>Abstract</p> <p>Background</p> <p>Community coalitions are rooted in complex and dynamic community systems. Despite recognition that environmental factors affect coalition behavior, few studies have examined how community context impacts coalition formation. Using the Community Coalition Action theory as an organizing framework, the current study employs multiple case study methodology to examine how five domains of community context affect coalitions in the formation stage of coalition development. Domains are history of collaboration, geography, community demographics and economic conditions, community politics and history, and community norms and values.</p> <p>Methods</p> <p>Data were from 8 sites that participated in an evaluation of a healthy cities and communities initiative in California. Twenty-three focus groups were conducted with coalition members, and 76 semi-structured interviews were conducted with local coordinators and coalition leaders. Cross-site analyses were conducted to identify the ways contextual domains influenced selection of the lead agency, coalition membership, staffing and leadership, and coalition processes and structures.</p> <p>Results</p> <p>History of collaboration influenced all four coalition factors examined, from lead agency selection to coalition structure. Geography influenced coalition formation largely through membership and staffing, whereas the demographic and economic makeup of the community had an impact on coalition membership, staffing, and infrastructure for coalition processes. The influence of community politics, history, norms and values was most noticeable on coalition membership.</p> <p>Conclusions</p> <p>Findings contribute to an ecologic and theory-based understanding of the range of ways community context influences coalitions in their formative stage.</p

A Putative Homologue of CDC20/CDH1 in the Malaria Parasite Is Essential for Male Gamete Development

Cell-cycle progression is governed by a series of essential regulatory proteins. Two major regulators are cell-division cycle protein 20 (CDC20) and its homologue, CDC20 homologue 1 (CDH1), which activate the anaphase-promoting complex/cyclosome (APC/C) in mitosis, and facilitate degradation of mitotic APC/C substrates. The malaria parasite, Plasmodium, is a haploid organism which, during its life-cycle undergoes two stages of mitosis; one associated with asexual multiplication and the other with male gametogenesis. Cell-cycle regulation and DNA replication in Plasmodium was recently shown to be dependent on the activity of a number of protein kinases. However, the function of cell division cycle proteins that are also involved in this process, such as CDC20 and CDH1 is totally unknown. Here we examine the role of a putative CDC20/CDH1 in the rodent malaria Plasmodium berghei (Pb) using reverse genetics. Phylogenetic analysis identified a single putative Plasmodium CDC20/CDH1 homologue (termed CDC20 for simplicity) suggesting that Plasmodium APC/C has only one regulator. In our genetic approach to delete the endogenous cdc20 gene of P. berghei, we demonstrate that PbCDC20 plays a vital role in male gametogenesis, but is not essential for mitosis in the asexual blood stage. Furthermore, qRT-PCR analysis in parasite lines with deletions of two kinase genes involved in male sexual development (map2 and cdpk4), showed a significant increase in cdc20 transcription in activated gametocytes. DNA replication and ultra structural analyses of cdc20 and map2 mutants showed similar blockage of nuclear division at the nuclear spindle/kinetochore stage. CDC20 was phosphorylated in asexual and sexual stages, but the level of modification was higher in activated gametocytes and ookinetes. Changes in global protein phosphorylation patterns in the Δcdc20 mutant parasites were largely different from those observed in the Δmap2 mutant. This suggests that CDC20 and MAP2 are both likely to play independent but vital roles in male gametogenesis

Social Participation and Disaster Risk Reduction Behaviors in Tsunami Prone Areas

This paper examines the relationships between social participation and disaster risk reduction actions. A survey of 557 households in tsunami prone areas in Phang Nga, Thailand was conducted following the 2012 Indian Ocean earthquakes. We use a multivariate probit model to jointly estimate the likelihood of undertaking three responses to earthquake and tsunami hazards (namely, (1) following disaster-related news closely, (2) preparing emergency kits and/or having a family emergency plan, and (3) having an intention to migrate) and community participation.We find that those who experienced losses from the 2004 tsunami are more likely to participate in community activities and respond to earthquake hazards. Compared to men, women are more likely to prepare emergency kits and/or have an emergency plan and have a greater intention to migrate. Living in a community with a higher proportion of women with tertiary education increases the probability of engaging in community activities and carrying out disaster risk reduction measures. Individuals who participate in village-based activities are 5.2% more likely to undertake all three risk reduction actions compared to those not engaging in community activities. This implies that encouraging participation in community activities can have positive externalities in disaster mitigation

Is Aboriginal Food Less Allergenic? Comparing IgE-Reactivity of Eggs from Modern and Ancient Chicken Breeds in a Cohort of Allergic Children

BACKGROUND: Hen's egg allergy ranks among the most frequent primary food allergies in children. We aimed to investigate sensitization profiles of egg allergic patients and compare in vitro IgE reactivities of eggs from ancient chicken breeds (Araucana and Maran) with those from conventional laying hen hybrids. METHODOLOGY: Egg allergic children (n = 25) were subjected to skin prick test, double blind placebo controlled food challenge, and sensitization profiles to Gal d 1-5 were determined by allergen microarray. IgE binding and biological activity of eggs from different chicken breeds were investigated by immunoblot, ELISA, and mediator release assays. PRINCIPAL FINDINGS: We found that Gal d 1 and Gal d 2 are generally major egg allergens, whereas Gal d 3-5 displayed high sensitization prevalence only in patients reacting to both, egg white and yolk. It seems that the onset of egg allergy is mediated by egg white allergens expanding to yolk sensitization in later stages of disease. Of note, egg white/yolk weight ratios were reduced in eggs from Auraucana and Maran chicken. As determined in IgE immunoblots and mass analysis, eggs from ancient chicken breeds did not differ in their protein composition. Similar IgE-binding was observed for all egg white preparations, while an elevated allergenicity was detected in egg yolk from Araucana chicken. CONCLUSION/SIGNIFICANCE: Our results on allergenicity and biological activity do not confirm the common assumption that aboriginal food might be less allergenic. Comprehensive diagnosis of egg allergy should distinguish between reactivity to hen's egg white and yolk fractions to avoid unnecessary dietary restrictions to improve life quality of the allergic child and its family

The Current Status and Work of Three Rs Centres and Platforms in Europe*

The adoption of Directive 2010/63/EU on the protection of animals used for scientific purposes has given a major push to the formation of Three Rs initiatives in the form of centres and platforms. These centres and platforms are dedicated to the so-called Three Rs, which are the Replacement, Reduction and Refinement of animal use in experiments. ATLA's 50th Anniversary year has seen the publication of two articles on European Three Rs centres and platforms. The first of these was about the progressive rise in their numbers and about their founding history; this second part focuses on their current status and activities. This article takes a closer look at their financial and organisational structures, describes their Three Rs focus and core activities (dissemination, education, implementation, scientific quality/translatability, ethics), and presents their areas of responsibility and projects in detail. This overview of the work and diverse structures of the Three Rs centres and platforms is not only intended to bring them closer to the reader, but also to provide role models and show examples of how such Three Rs centres and platforms could be made sustainable. The Three Rs centres and platforms are very important focal points and play an immense role as facilitators of Directive 2010/63/EU 'on the ground' in their respective countries. They are also invaluable for the wide dissemination of information and for promoting the implementation of the Three Rs in general