Using green fluorescent malaria parasites to screen for permissive vector mosquitoes

BACKGROUND: The Plasmodium species that infect rodents, particularly Plasmodium berghei and Plasmodium yoelii, are useful to investigate host-parasite interactions. The mosquito species that act as vectors of human plasmodia in South East Asia, Africa and South America show different susceptibilities to infection by rodent Plasmodium species. P. berghei and P. yoelii infect both Anopheles gambiae and Anopheles stephensi, which are found mainly in Africa and Asia, respectively. However, it was reported that P. yoelii can infect the South American mosquito, Anopheles albimanus, while P. berghei cannot. METHODS: P. berghei lines that express the green fluorescent protein were used to screen for mosquitoes that are susceptible to infection by P. berghei. Live mosquitoes were examined and screened for the presence of a fluorescent signal in the abdomen. Infected mosquitoes were then examined by time-lapse microscopy to reveal the dynamic behaviour of sporozoites in haemolymph and extracted salivary glands. RESULTS: A single fluorescent oocyst can be detected in live mosquitoes and P. berghei can infect A. albimanus. As in other mosquitoes, P. berghei sporozoites can float through the haemolymph and invade A. albimanus salivary glands and they are infectious in mice after subcutaneous injection. CONCLUSION: Fluorescent Plasmodium parasites can be used to rapidly screen susceptible mosquitoes. These results open the way to develop a laboratory model in countries where importation of A. gambiae and A. stephensi is not allowed

Clinical and biological characteristics of cervical neoplasias with FGFR3 mutation

BACKGROUND: We have previously reported activating mutations of the gene coding for the fibroblast growth factor receptor 3 (FGFR3) in invasive cervical carcinoma. To further analyze the role of FGFR3 in cervical tumor progression, we extended our study to screen a total of 75 invasive tumors and 80 cervical intraepithelial neoplasias (40 low-grade and 40 high-grade lesions). RESULTS: Using single strand conformation polymorphism (SSCP) followed by DNA sequencing, we found FGFR3 mutation (S249C in all cases) in 5% of invasive cervical carcinomas and no mutation in intraepithelial lesions. These results suggest that, unlike in bladder carcinoma, FGFR3 mutation does not or rarely occur in non invasive lesions. Compared to patients with wildtype FGFR3 tumor, patients with S249C FGFR3 mutated tumors were older (mean age 64 vs. 49.4 years, P = 0.02), and were more likely to be associated with a non-16/18 HPV type in their tumor. Gene expression analysis demonstrated that FGFR3 mutated tumors were associated with higher FGFR3b mRNA expression levels compared to wildtype FGFR3 tumors. Supervised analysis of Affymetrix expression data identified a significant number of genes specifically differentially expressed in tumors with respect to FGFR3 mutation status. CONCLUSION: This study suggest that tumors with FGFR3 mutation appear to have distinctive clinical and biological characteristics that may help in defining a population of patients for FGFR3 mutation screening

Site-Specific Integration and Expression of an Anti-Malarial Gene in Transgenic Anopheles gambiae Significantly Reduces Plasmodium Infections

Diseases transmitted by mosquitoes have a devastating impact on global health and this is worsening due to difficulties with existing control measures and climate change. Genetically modified mosquitoes that are refractory to disease transmission are seen as having great potential in the delivery of novel control strategies. Historically the genetic modification of insects has relied upon transposable elements which have many limitations despite their successful use. To circumvent these limitations the Streptomyces phage phiC31 integrase system has been successfully adapted for site-specific transgene integration in insects. Here, we present the first site-specific transformation of Anopheles gambiae, the principal vector of human malaria. Mosquitoes were initially engineered to incorporate the phiC31 targeting site at a defined genomic location. A second phase of genetic modification then achieved site-specific integration of Vida3, a synthetic anti-malarial gene. Expression of Vida3, specifically in the midgut of bloodfed females, offered consistent and significant protection against Plasmodium yoelii nigeriensis, reducing average parasite intensity by 85%. Similar protection was observed against Plasmodium falciparum in some experiments, although protection was inconsistent. In the fight against malaria, it is imperative to establish a broad repertoire of both anti-malarial effector genes and tissue-specific promoters for their expression, enabling those offering maximum effect with minimum fitness cost to be identified. In the future, this technology will allow effective comparisons and informed choices to be made, potentially leading to complete transmission blockade

Overcoming global inequality is critical for land-based mitigation in line with the Paris Agreement

Transformation pathways for the land sector in line with the Paris Agreement depend on the assumption of globally implemented greenhouse gas (GHG) emission pricing, and in some cases also on inclusive socio-economic development and sustainable land-use practices. In such pathways, the majority of GHG emission reductions in the land system is expected to come from low- and middle-income countries, which currently account for a large share of emissions from agriculture, forestry and other land use (AFOLU). However, in low- and middle-income countries the economic, financial and institutional barriers for such transformative changes are high. Here, we show that if sustainable development in the land sector remained highly unequal and limited to high-income countries only, global AFOLU emissions would remain substantial throughout the 21st century. Our model-based projections highlight that overcoming global inequality is critical for land-based mitigation in line with the Paris Agreement. While also a scenario purely based on either global GHG emission pricing or on inclusive socio-economic development would achieve the stringent emissions reductions required, only the latter ensures major co-benefits for other Sustainable Development Goals, especially in low- and middle-income regions

Anopheles Gambiae PRS1 Modulates Plasmodium Development at Both Midgut and Salivary Gland Steps

Background: Invasion of the mosquito salivary glands by Plasmodium is a critical step for malaria transmission. From a SAGE analysis, we previously identified several genes whose expression in salivary glands was regulated coincident with sporozoite invasion of salivary glands. To get insights into the consequences of these salivary gland responses, here we have studied one of the genes, PRS1 (Plasmodium responsive salivary 1), whose expression was upregulated in infected glands, using immunolocalization and functional inactivation approaches. Methodology/Principal Findings: PRS1 belongs to a novel insect superfamily of genes encoding proteins with DM9 repeat motifs of uncharacterized function. We show that PRS1 is induced in response to Plasmodium, not only in the salivary glands but also in the midgut, the other epithelial barrier that Plasmodium has to cross to develop in the mosquito. Furthermore, this induction is observed using either the rodent parasite Plasmodium berghei or the human pathogen Plasmodium falciparum. In the midgut, PRS1 overexpression is associated with a relocalization of the protein at the periphery of invaded cells. We also find that sporozoite invasion of salivary gland cells occurs sequentially and induces intra-cellular modifications that include an increase in PRS1 expression and a relocalization of the corresponding protein into vesicle-like structures. Importantly, PRS1 knockdown during the onset of midgut and salivary gland invasion demonstrates that PRS1 acts as an agonist for the development of both parasite species in the two epithelia, highlighting shared vector/parasite interactions in both tissues. Conclusions/Significance: While providing insights into potential functions of DM9 proteins, our results reveal that PRS1 likely contributes to fundamental interactions between Plasmodium and mosquito epithelia, which do not depend on the specific Anopheles/P. falciparum coevolutionary history

Fine Pathogen Discrimination within the APL1 Gene Family Protects Anopheles gambiae against Human and Rodent Malaria Species

Genetically controlled resistance of Anopheles gambiae mosquitoes to Plasmodium falciparum is a common trait in the natural population, and a cluster of natural resistance loci were mapped to the Plasmodium-Resistance Island (PRI) of the A. gambiae genome. The APL1 family of leucine-rich repeat (LRR) proteins was highlighted by candidate gene studies in the PRI, and is comprised of paralogs APL1A, APL1B and APL1C that share ≥50% amino acid identity. Here, we present a functional analysis of the joint response of APL1 family members during mosquito infection with human and rodent Plasmodium species. Only paralog APL1A protected A. gambiae against infection with the human malaria parasite P. falciparum from both the field population and in vitro culture. In contrast, only paralog APL1C protected against the rodent malaria parasites P. berghei and P. yoelii. We show that anti-P. falciparum protection is mediated by the Imd/Rel2 pathway, while protection against P. berghei infection was shown to require Toll/Rel1 signaling. Further, only the short Rel2-S isoform and not the long Rel2-F isoform of Rel2 confers protection against P. falciparum. Protection correlates with the transcriptional regulation of APL1A by Rel2-S but not Rel2-F, suggesting that the Rel2-S anti-parasite phenotype results at least in part from its transcriptional control over APL1A. These results indicate that distinct members of the APL1 gene family display a mutually exclusive protective effect against different classes of Plasmodium parasites. It appears that a gene-for-pathogen-class system orients the appropriate host defenses against distinct categories of similar pathogens. It is known that insect innate immune pathways can distinguish between grossly different microbes such as Gram-positive bacteria, Gram-negative bacteria, or fungi, but the function of the APL1 paralogs reveals that mosquito innate immunity possesses a more fine-grained capacity to distinguish between classes of closely related eukaryotic pathogens than has been previously recognized

EMT Inducers Catalyze Malignant Transformation of Mammary Epithelial Cells and Drive Tumorigenesis towards Claudin-Low Tumors in Transgenic Mice

The epithelial-mesenchymal transition (EMT) is an embryonic transdifferentiation process consisting of conversion of polarized epithelial cells to motile mesenchymal ones. EMT–inducing transcription factors are aberrantly expressed in multiple tumor types and are known to favor the metastatic dissemination process. Supporting oncogenic activity within primary lesions, the TWIST and ZEB proteins can prevent cells from undergoing oncogene-induced senescence and apoptosis by abolishing both p53- and RB-dependent pathways. Here we show that they also downregulate PP2A phosphatase activity and efficiently cooperate with an oncogenic version of H-RAS in malignant transformation of human mammary epithelial cells. Thus, by down-regulating crucial tumor suppressor functions, EMT inducers make cells particularly prone to malignant conversion. Importantly, by analyzing transformed cells generated in vitro and by characterizing novel transgenic mouse models, we further demonstrate that cooperation between an EMT inducer and an active form of RAS is sufficient to trigger transformation of mammary epithelial cells into malignant cells exhibiting all the characteristic features of claudin-low tumors, including low expression of tight and adherens junction genes, EMT traits, and stem cell–like characteristics. Claudin-low tumors are believed to be the most primitive breast malignancies, having arisen through transformation of an early epithelial precursor with inherent stemness properties and metaplastic features. Challenging this prevailing view, we propose that these aggressive tumors arise from cells committed to luminal differentiation, through a process driven by EMT inducers and combining malignant transformation and transdifferentiation

Activation of MEK1 or MEK2 isoform is sufficient to fully transform intestinal epithelial cells and induce the formation of metastatic tumors

<p>Abstract</p> <p>Background</p> <p>The Ras-dependent ERK1/2 MAP kinase signaling pathway plays a central role in cell proliferation control and is frequently activated in human colorectal cancer. Small-molecule inhibitors of MEK1/MEK2 are therefore viewed as attractive drug candidates for the targeted therapy of this malignancy. However, the exact contribution of MEK1 and MEK2 to the pathogenesis of colorectal cancer remains to be established.</p> <p>Methods</p> <p>Wild type and constitutively active forms of MEK1 and MEK2 were ectopically expressed by retroviral gene transfer in the normal intestinal epithelial cell line IEC-6. We studied the impact of MEK1 and MEK2 activation on cellular morphology, cell proliferation, survival, migration, invasiveness, and tumorigenesis in mice. RNA interference was used to test the requirement for MEK1 and MEK2 function in maintaining the proliferation of human colorectal cancer cells.</p> <p>Results</p> <p>We found that expression of activated MEK1 or MEK2 is sufficient to morphologically transform intestinal epithelial cells, dysregulate cell proliferation and induce the formation of high-grade adenocarcinomas after orthotopic transplantation in mice. A large proportion of these intestinal tumors metastasize to the liver and lung. Mechanistically, activation of MEK1 or MEK2 up-regulates the expression of matrix metalloproteinases, promotes invasiveness and protects cells from undergoing anoikis. Importantly, we show that silencing of MEK2 expression completely suppresses the proliferation of human colon carcinoma cell lines, whereas inactivation of MEK1 has a much weaker effect.</p> <p>Conclusion</p> <p>MEK1 and MEK2 isoforms have similar transforming properties and are able to induce the formation of metastatic intestinal tumors in mice. Our results suggest that MEK2 plays a more important role than MEK1 in sustaining the proliferation of human colorectal cancer cells.</p

Evaluation et controle au laboratoire du pouvoir entomopathogene de Bacillus thuringiensis israelensis et de Bacillus sphaericus sur larves de Culicidae (Dipt. Nematoceres)

SIGLECNRS T Bordereau / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Potential of global land water recycling to mitigate local temperature extremes

status: publishe