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      Toxic Activity, Molecular Modeling and Docking Simulations of Bacillus thuringiensis Cry11 Toxin Variants Obtained via DNA Shuffling

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          Abstract

          The Cry11 family belongs to a large group of δ-endotoxins that share three distinct structural domains. Among the dipteran-active toxins referred to as three-domain Cry11 toxins, the Cry11Aa protein from Bacillus thuringiensis subsp. israelensis ( Bti) has been the most extensively studied. Despite the potential of Bti as an effective biological control agent, the understanding of Cry11 toxins remains incomplete. In this study, five Cry11 variants obtained via DNA shuffling displayed toxic activity against Aedes aegypti and Culex quinquefasciatus. Three of these Cry11 variants (8, 23, and 79) were characterized via 3D modeling and analysis of docking with ALP1. The relevant mutations in these variants, such as deletions, insertions and point mutations, are discussed in relation to their structural domains, toxic activities and toxin-receptor interactions. Importantly, deletion of the N-terminal segment in domain I was not associated with any change in toxic activity, and domain III exhibited higher sequence variability than domains I and II. Variant 8 exhibited up to 3.78- and 6.09-fold higher toxicity to A. aegypti than Cry11Bb and Cry11Aa, respectively. Importantly, variant 79 showed an α-helix conformation at the C-terminus and formed crystals retaining toxic activity. These findings indicate that five Cry11 variants were preferentially reassembled from the cry11Aa gene during DNA shuffling. The mutations described in loop 2 and loop 3 of domain II provide valuable information regarding the activity of Cry11 toxins against A. aegypti and C. quinquefasciatus larvae and reveal new insights into the application of directed evolution strategies to study the genetic variability of specific domains in cry11 family genes.

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          A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding

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            CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice

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              Differential Susceptibilities of Aedes aegypti and Aedes albopictus from the Americas to Zika Virus

              Background Since the major outbreak in 2007 in the Yap Island, Zika virus (ZIKV) causing dengue-like syndromes has affected multiple islands of the South Pacific region. In May 2015, the virus was detected in Brazil and then spread through South and Central America. In December 2015, ZIKV was detected in French Guiana and Martinique. The aim of the study was to evaluate the vector competence of the mosquito spp. Aedes aegypti and Aedes albopictus from the Caribbean (Martinique, Guadeloupe), North America (southern United States), South America (Brazil, French Guiana) for the currently circulating Asian genotype of ZIKV isolated from a patient in April 2014 in New Caledonia. Methodology/Principal Findings Mosquitoes were orally exposed to an Asian genotype of ZIKV (NC-2014-5132). Upon exposure, engorged mosquitoes were maintained at 28°±1°C, a 16h:8h light:dark cycle and 80% humidity. 25–30 mosquitoes were processed at 4, 7 and 14 days post-infection (dpi). Mosquito bodies (thorax and abdomen), heads and saliva were analyzed to measure infection, dissemination and transmission, respectively. High infection but lower disseminated infection and transmission rates were observed for both Ae. aegypti and Ae. albopictus. Ae. aegypti populations from Guadeloupe and French Guiana exhibited a higher dissemination of ZIKV than the other Ae. aegypti populations examined. Transmission of ZIKV was observed in both mosquito species at 14 dpi but at a low level. Conclusions/Significance This study suggests that although susceptible to infection, Ae. aegypti and Ae. albopictus were unexpectedly low competent vectors for ZIKV. This may suggest that other factors such as the large naïve population for ZIKV and the high densities of human-biting mosquitoes contribute to the rapid spread of ZIKV during the current outbreak.
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                Author and article information

                Contributors
                Journal
                Front Microbiol
                Front Microbiol
                Front. Microbiol.
                Frontiers in Microbiology
                Frontiers Media S.A.
                1664-302X
                17 October 2018
                2018
                : 9
                Affiliations
                1RG Microbial Ecology: Metabolism, Genomics & Evolution, Microbiomas Foundation , Chía, Colombia
                2Laboratorio de Biología Molecular y Biotecnología, Universidad de Santander , Bucaramanga, Colombia
                3Escuela de Medicina, Facultad de Salud, Universidad Industrial de Santander , Bucaramanga, Colombia
                4Grupo Biologa Funcional, Laboratorio de Prospección y Diseo de Biomoléculas, Escuela de Biociencias, Universidad Nacional , Sede Medellín, Colombia
                5Programa de Estudio y Control de Enfermedades Tropicales PECET, Unidad de Biologa Molecular y Computacional-UBMC, Universidad de Antioquía , Medellín, Colombia
                Author notes

                Edited by: Marc Strous, University of Calgary, Canada

                Reviewed by: Christopher L. Hemme, The University of Rhode Island, United States; Evgeniya V. Nazarova, Cornell University, United States

                *Correspondence: Alvaro Mauricio Florez, amflorez@ 123456microbiomas.org

                This article was submitted to Microbial Physiology and Metabolism, a section of the journal Frontiers in Microbiology

                Article
                10.3389/fmicb.2018.02461
                6199390
                Copyright © 2018 Florez, Suarez-Barrera, Morales, Rivera, Orduz, Ochoa, Guerra and Muskus.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                Page count
                Figures: 5, Tables: 1, Equations: 0, References: 50, Pages: 14, Words: 0
                Funding
                Funded by: Departamento Administrativo de Ciencia, Tecnología e Innovación 10.13039/100007637
                Award ID: 1299-12-16813
                Award ID: 5201-545-31565
                Categories
                Microbiology
                Original Research

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