Modulating signaling networks by CRISPR/Cas9-mediated transposable element insertion

In a recent past, transposable elements (TEs) were referred to as selfish genetic components only capable of copying themselves with the aim of increasing the odds of being inherited. Nonetheless, TEs have been initially proposed as positive control elements acting in synergy with the host. Nowadays, it is well known that TE movement into host genome comprises an important evolutionary mechanism capable of increasing the adaptive fitness. As insights into TE functioning are increasing day to day, the manipulation of transposition has raised an interesting possibility of setting the host functions, although the lack of appropriate genome engineering tools has unpaved it. Fortunately, the emergence of genome editing technologies based on programmable nucleases, and especially the arrival of a multipurpose RNA-guided Cas9 endonuclease system, has made it possible to reconsider this challenge. For such purpose, a particular type of transposons referred to as miniature inverted-repeat transposable elements (MITEs) has shown a series of interesting characteristics for designing functional drivers. Here, recent insights into MITE elements and versatile RNA-guided CRISPR/Cas9 genome engineering system are given to understand how to deploy the potential of TEs for control of the host transcriptional activity.Fil: Vaschetto, Luis Maria Benjamin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Diversidad y Ecología Animal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Diversidad y Ecología Animal; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Diversidad Animal I; Argentin

Phenylpropanoid Glycoside Analogues: Enzymatic Synthesis, Antioxidant Activity and Theoretical Study of Their Free Radical Scavenger Mechanism

Phenylpropanoid glycosides (PPGs) are natural compounds present in several medicinal plants that have high antioxidant power and diverse biological activities. Because of their low content in plants (less than 5% w/w), several chemical synthetic routes to produce PPGs have been developed, but their synthesis is a time consuming process and the achieved yields are often low. In this study, an alternative and efficient two-step biosynthetic route to obtain natural PPG analogues is reported for the first time. Two galactosides were initially synthesized from vanillyl alcohol and homovanillyl alcohol by a transgalactosylation reaction catalyzed by Kluyveromyces lactis β-galactosidase in saturated lactose solutions with a 30%–35% yield. To synthesize PPGs, the galactoconjugates were esterified with saturated and unsaturated hydroxycinnamic acid derivatives using Candida antarctica Lipase B (CaL-B) as a biocatalyst with 40%–60% yields. The scavenging ability of the phenolic raw materials, intermediates and PPGs was evaluated by the 2,2-diphenyl-1-picrylhydrazyl radical (DPPH•) method. It was found that the biosynthesized PPGs had higher scavenging abilities when compared to ascorbic acid, the reference compound, while their antioxidant activities were found similar to that of natural PPGs. Moreover, density functional theory (DFT) calculations were used to determine that the PPGs antioxidant mechanism proceeds through a sequential proton loss single electron transfer (SPLET). The enzymatic process reported in this study is an efficient and versatile route to obtain PPGs from different phenylpropanoid acids, sugars and phenolic alcohols

Risk Factors for Tungiasis in Nigeria: Identification of Targets for Effective Intervention

Tungiasis is a parasitic skin disease caused by the sand flea Tunga penetrans. After penetration into the skin, the flea grows and reaches the size of a pea. The disease is a neglected public health problem in endemic areas in Latin America, the Caribbean and Africa, and causes considerable morbidity in the affected communities. We performed a study in a rural community in Nigeria to detect factors associated with tungiasis. People were examined for the presence of sand flea lesions, and a questionnaire was applied. Of the 643 individuals examined, 252 (42.5%) had tungiasis. The most important factors independently associated with the disease were: presence of pigs on the compound (adjusted odds ratio [OR] = 17.98), sand or clay floor inside house (OR = 9.33), and having the common resting place outside the house (OR = 7.14). The regular use of closed footwear (OR = 0.34) and the use of insecticides indoors (OR = 0.2) were protective factors. Our data show that tungiasis in this community is determined to an important extent by some variables that may be a target for interventions

Decreasing Burden of Malaria in Pregnancy in Malawian Women and Its Relationship to Use of Intermittent Preventive Therapy or Bed Nets

The World Health Organization recommends insecticidal bednets and intermittent preventive treatment to reduce malaria in pregnancy. Longitudinal data of malaria prevalence and pregnancy outcomes are valuable in gauging the impact of these antimalarial interventions.We recruited 8,131 women delivering in a single Malawian hospital over 9 years. We recorded demographic data, antenatal prescription of intermittent preventive therapy during pregnancy with sulfadoxine-pyrimethamine and bed net use, and examined finger-prick blood for malaria parasites and hemoglobin concentration. In 4,712 women, we examined placental blood for malaria parasites and recorded the infant's birth weight. Peripheral and placental parasitemia prevalence declined from 23.5% to 5.0% and from 25.2% to 6.8% respectively. Smaller declines in prevalence of low birth weight and anemia were observed. Coverage of intermittent preventive treatment and bednets increased. Number of sulfadoxine-pyrimethamine doses received correlated inversely with placental parasitemia (Odds Ratio (95% CI): 0.79 (0.68, 0.91)), maternal anemia (0.81, (0.73, 0.90)) and low birth weight from 1997-2001 (0.63 (0.53, 0.75)), but not from 2002-2006. Bednet use protected from peripheral parasitemia (0.47, (0.37, 0.60)) and placental parasitemia (0.41, (0.31, 0.54)) and low birth weight (0.75 (0.59, 0.95)) but not anemia throughout the study. Compared to women without nets who did not receive 2-dose sulfadoxine-pyrimethamine, women using nets and receiving 2-dose sulfadoxine-pyrimethamine were less likely to have parasitemia or low birth weight babies. Women receiving 2-dose sulfadoxine-pyrimethamine alone had little evidence of protection whereas bednets alone gave intermediate protection.Increased bednet coverage explains changes in parasitemia and birth weight among pregnant women better than sulfadoxine-pyrimethamine use. High bed net coverage, and sulfadoxine-pyrimethamine resistance, may be contributing to its apparent loss of effectiveness

Differential antibody responses to Plasmodium falciparum merozoite proteins in Malawian children with severe malaria

Cerebral malaria (CM) and severe malarial anemia (SMA) are 2 major causes of death in African children infected with Plasmodium falciparum. We investigated levels of naturally acquired antibody to conserved and variable regions of merozoite surface protein (MSP)-1 and MSP-2, apical membrane antigen (AMA)-1, and rhoptry-associated protein 1 in plasma samples from 126 children admitted to the hospital with CM, 59 with SMA, and 84 with uncomplicated malaria (UM) in Malawi. Children with SMA were distinguished by very low levels of immunoglobulin (Ig) G to the conserved C-terminus of MSP-1 and MSP-2 and to full-length AMA-1. Conversely, children with CM had significantly higher levels of IgG to the conserved regions of all antigens examined than did children with UM (for MSP-1 and AMA-1, P< .005; for MSP-2, P< .05) or SMA (for MSP-1 and MSP-2, P<.001; for AMA-1, P< .005). These distinct IgG patterns might reflect differences in age, exposure to P. falciparum, and/or genetic factors affecting immune responses

Multislice CT angiography in the selection of patients with ruptured intracranial aneurysms suitable for clipping or coiling

Introduction We sought to establish whether CT angiography (CTA) can be applied to the planning and performance of clipping or coiling in ruptured intracranial aneurysms without recourse to intraarterial digital subtraction angiography (IA-DSA). Methods Over the period April 2003 to January 2006 in all patients presenting with a subarachnoid haemorrhage CTA was performed primarily. If CTA demonstrated an aneurysm, coiling or clipping was undertaken. IA-DSA was limited to patients with negative or inconclusive CTA findings. We compared CTA images with findings at surgery or coiling in patients with positive CTA findings and in patients with negative and inconclusive findings in whom IA-DSA had been performed. Results In this study, 224 consecutive patients (mean age 52.7 years, 135 women) were included. In 133 patients (59%) CTA demonstrated an aneurysm, and CTA was followed directly by neurosurgical (n=55) or endovascular treatment (n=78). In 31 patients (14%) CTA findings were categorized as inconclusive, and in 60 (27%) CTA findings were negative. One patient received surgical treatment on the basis of false-positive CTA findings. In 17 patients in whom CTA findings were inconclusive, IA-DSA provided further diagnostic information required for correct patient selection for any therapy. Five ruptured aneurysms in patients with a nonperimesencephalic SAH were negative on CTA, and four of these were also false-negative on IA-DSA. On a patient basis the positive predictive value, negative predictive value, sensitivity, specificity and accuracy of CTA for symptomatic aneurysms were 99%, 90%, 96%, 98% and 96%, respectively. Conclusion CTA should be used as the first diagnostic modality in the selection of patients for surgical or endovascular treatment of ruptured intracranial aneurysms. If CTA renders inconclusive results, IA-DSA should be performed. With negative CTA results the complementary value of IA-DSA is marginal. IA-DSA is not needed in patients with negative CTA and classic perimesencephalic SAH. Repeat IA-DSA or CTA should still be performed in patients with a nonperimesencephalic SAH

Malaria and Helminth Co-infections in School and Preschool Children: A Cross-sectional Study in Magu district, north-Western Tanzania.

Malaria, schistosomiasis and soil transmitted helminth infections (STH) are important parasitic infections in Sub-Saharan Africa where a significant proportion of people are exposed to co-infections of more than one parasite. In Tanzania, these infections are a major public health problem particularly in school and pre-school children. The current study investigated malaria and helminth co-infections and anaemia in school and pre-school children in Magu district, Tanzania. School and pre-school children were enrolled in a cross-sectional study. Stool samples were examined for Schistosoma mansoni and STH infections using Kato Katz technique. Urine samples were examined for Schistosoma haematobium using the urine filtration method. Blood samples were examined for malaria parasites and haemoglobin concentrations using the Giemsa stain and Haemoque methods, respectively. Out of 1,546 children examined, 1,079 (69.8%) were infected with one or more parasites. Malaria-helminth co-infections were observed in 276 children (60% of all children with P. falciparum infection). Malaria parasites were significantly more prevalent in hookworm infected children than in hookworm free children (p = 0.046). However, this association was non-significant on multivariate logistic regression analysis (OR = 1.320, p = 0.064). Malaria parasite density decreased with increasing infection intensity of S. mansoni and with increasing number of co-infecting helminth species. Anaemia prevalence was 34.4% and was significantly associated with malaria infection, S. haematobium infection and with multiple parasite infections. Whereas S. mansoni infection was a significant predictor of malaria parasite density, P. falciparum and S. haematobium infections were significant predictors of anaemia. These findings suggest that multiple parasite infections are common in school and pre-school children in Magu district. Concurrent P. falciparum, S. mansoni and S. haematobium infections increase the risk of lower Hb levels and anaemia, which in turn calls for integrated disease control interventions. The associations between malaria and helminth infections detected in this study need further investigation

Recurrent hotspot mutations in HRAS Q61 and PI3K-AKT pathway genes as drivers of breast adenomyoepitheliomas

Adenomyoepithelioma of the breast is a rare tumor characterized by epithelial-myoepithelial differentiation, of which a subset will progress to invasive or metastatic cancer. We sought to define the genomic landscape of adenomyoepitheliomas. Massively parallel sequencing revealed highly recurrent somatic mutations in HRAS and PI3K-AKT pathway-related genes. Strikingly, HRAS mutations were restricted to estrogen receptor (ER)-negative tumors, all affected codon 61, and all but one co-occurred with PIK3CA or PIK3R1 mutations. To interrogate the functional significance of HRAS Q61 mutations in adenomyoepithelial differentiation, we expressed HRASQ61R alone or in combination with PIK3CAH1047R in non-transformed ER-negative breast epithelial cells. HRASQ61R induced characteristic phenotypes of adenomyoepitheliomas such as the expression of myoepithelial markers and loss of e-cadherin, hyperactivation of AKT signaling, and transformative properties that were arrested by combination therapy with AKT and MEK inhibitors. Our results indicate that breast adenomyoepitheliomas often manifest a unique transformation program featuring HRAS activation

The Wolbachia endosymbiont as an anti-filarial nematode target

Human disease caused by parasitic filarial nematodes is a major cause of global morbidity. The parasites are transmitted by arthropod intermediate hosts and are responsible for lymphatic filariasis (elephantiasis) or onchocerciasis (river blindness). Within these filarial parasites are intracellular alpha-proteobacteria, Wolbachia, that were first observed almost 30 years ago. The obligate endosymbiont has been recognized as a target for anti-filarial nematode chemotherapy as evidenced by the loss of worm fertility and viability upon antibiotic treatment in an extensive series of human trials. While current treatments with doxycycline and rifampicin are not practical for widespread use due to the length of required treatments and contraindications, anti-Wolbachia targeting nevertheless appears a promising alternative for filariasis control in situations where current programmatic strategies fail or are unable to be delivered and it provides a superior efficacy for individual therapy. The mechanisms that underlie the symbiotic relationship between Wolbachia and its nematode hosts remain elusive. Comparative genomics, bioinfomatic and experimental analyses have identified a number of potential interactions, which may be drug targets. One candidate is de novo heme biosynthesis, due to its absence in the genome sequence of the host nematode, Brugia malayi, but presence in Wolbachia and its potential roles in worm biology. We describe this and several additional candidate targets, as well as our approaches for understanding the nature of the host-symbiont relationship