Germination of spores and gametophytes of "Polystichum montevidense" (Spreng.) Rosenst. ("Dryopteridaceae")

Se presentan características sobre la germinación de las esporas, el desarrollo gametofítico y la estructura de los gametofitos adultos de Polystichum montevidense (Spreng.) Rosenst. obtenidos mediante cultivos estériles. Se encuentra que los esporangios originan 32 esporas monoletes con perisporio crestado. La germinación es de tipo Vittaria y se produce a los tres días después de la siembra. El desarrollo de los gametofitos corresponde al tipo Aspidium. Los gametofitos laminares producen un solo tipo de tricomas glandulares con secreción de naturaleza lipídica, distribuidos sobre el margen de la lámina y unos pocos sobre la superficie. Se cita por primera vez la presencia de gotas de aceite en las células de los gametofitos adultos. La expresión sexual se manifiesta en gametofitos de más de 100 días de edad, originándose solamente gametofitos femeninos. Se ilustra con fotografías obtenidas con microscopios de luz y microscopio electrónico de barrido.A description is made of characteristics of the germination of spores, gametophytic development and adult gametophyte structure of Polystichum montevidense (Spreng.) Rosenst. obtained by sterile culture. It was found that sporangia produce 32 monolete spores with crested perispore. The germination of Vittaria-type spores occurs three days after sowing. Gametophyte development is of the Aspidium type. The laminar gametophytes have only one type of glandular trichomes, with secretion of trichomes has a lipid nature, distributed on the edge of the lamina and a few on the surface. It is reported for the first time that adult gametophyte cells have oil droplets. Sexual expression is manifested in gametophytes of more than 100 days old, with only female gametophytes. All this is illustrated with pictures recorded with light microscopes and a scanning electron microscope

Control of pyrethroid-resistant chagas disease vectors with entomopathogenic fungi

Background: Triatoma infestans-mediated transmission of Tripanosoma cruzi, the causative agent of Chagas disease, remains as a major health issue in southern South America. Key factors of T. infestans prevalence in specific areas of the geographic Gran Chaco region - which extends through northern Argentina, Bolivia, and Paraguay - are both recurrent reinfestations after insecticide spraying and emerging pyrethroid-resistance over the past ten years. Among alternative control tools, the pathogenicity of entomopathogenic fungi against triatomines is already known; furthermore, these fungi have the ability to fully degrade hydrocarbons from T. infestans cuticle and to utilize them as fuel and for incorporation into cellular components. Methodology and Findings: Here we provide evidence of resistance-related cuticle differences; capillary gas chromatography coupled to mass spectrometry analyses revealed that pyrethroid-resistant bugs have significantly larger amounts of surface hydrocarbons, peaking 56.2±6.4% higher than susceptible specimens. Also, a thicker cuticle was detected by scanning electron microscopy (32.1±5.9 μm and 17.8±5.4 μm for pyrethroid-resistant and pyrethroid-susceptible, respectively). In laboratory bioassays, we showed that the virulence of the entomopathogenic fungi Beauveria bassiana against T. infestans was significantly enhanced after fungal adaptation to grow on a medium containing insect-like hydrocarbons as the carbon source, regardless of bug susceptibility to pyrethroids. We designed an attraction-infection trap based on manipulating T. infestans behavior in order to facilitate close contact with B. bassiana. Field assays performed in rural village houses infested with pyrethroid-resistant insects showed 52.4% bug mortality. Using available mathematical models, we predicted that further fungal applications could eventually halt infection transmission. Conclusions: This low cost, low tech, ecologically friendly methodology could help in controlling the spread of pyrethroid-resistant bugs.Facultad de Ciencias Médica

Biological Control of the Chagas Disease Vector <i>Triatoma infestans</i> with the Entomopathogenic Fungus <i>Beauveria bassiana</i> Combined with an Aggregation Cue: Field, Laboratory and Mathematical Modeling Assessment

Current Chagas disease vector control strategies, based on chemical insecticide spraying, are growingly threatened by the emergence of pyrethroid-resistant Triatoma infestans populations in the Gran Chaco region of South America. We have already shown that the entomopathogenic fungus Beauveria bassiana has the ability to breach the insect cuticle and is effective both against pyrethroid-susceptible and pyrethroid-resistant T. infestans, in laboratory as well as field assays. It is also known that T. infestans cuticle lipids play a major role as contact aggregation pheromones. We estimated the effectiveness of pheromone-based infection boxes containing B. bassiana spores to kill indoor bugs, and its effect on the vector population dynamics. Laboratory assays were performed to estimate the effect of fungal infection on female reproductive parameters. The effect of insect exuviae as an aggregation signal in the performance of the infection boxes was estimated both in the laboratory and in the field. We developed a stage-specific matrix model of T. infestans to describe the fungal infection effects on insect population dynamics, and to analyze the performance of the biopesticide device in vector biological control. The pheromone-containing infective box is a promising new tool against indoor populations of this Chagas disease vector, with the number of boxes per house being the main driver of the reduction of the total domestic bug population. This ecologically safe approach is the first proven alternative to chemical insecticides in the control of T. infestans. The advantageous reduction in vector population by delayed-action fungal biopesticides in a contained environment is here shown supported by mathematical modeling.Instituto de Investigaciones Bioquímicas de La PlataFacultad de Ciencias MédicasCentro de Estudios Parasitológicos y de Vectore

Biological Control of the Chagas Disease Vector <i>Triatoma infestans</i> with the Entomopathogenic Fungus <i>Beauveria bassiana</i> Combined with an Aggregation Cue: Field, Laboratory and Mathematical Modeling Assessment

Current Chagas disease vector control strategies, based on chemical insecticide spraying, are growingly threatened by the emergence of pyrethroid-resistant Triatoma infestans populations in the Gran Chaco region of South America. We have already shown that the entomopathogenic fungus Beauveria bassiana has the ability to breach the insect cuticle and is effective both against pyrethroid-susceptible and pyrethroid-resistant T. infestans, in laboratory as well as field assays. It is also known that T. infestans cuticle lipids play a major role as contact aggregation pheromones. We estimated the effectiveness of pheromone-based infection boxes containing B. bassiana spores to kill indoor bugs, and its effect on the vector population dynamics. Laboratory assays were performed to estimate the effect of fungal infection on female reproductive parameters. The effect of insect exuviae as an aggregation signal in the performance of the infection boxes was estimated both in the laboratory and in the field. We developed a stage-specific matrix model of T. infestans to describe the fungal infection effects on insect population dynamics, and to analyze the performance of the biopesticide device in vector biological control. The pheromone-containing infective box is a promising new tool against indoor populations of this Chagas disease vector, with the number of boxes per house being the main driver of the reduction of the total domestic bug population. This ecologically safe approach is the first proven alternative to chemical insecticides in the control of T. infestans. The advantageous reduction in vector population by delayed-action fungal biopesticides in a contained environment is here shown supported by mathematical modeling.Instituto de Investigaciones Bioquímicas de La PlataFacultad de Ciencias MédicasCentro de Estudios Parasitológicos y de Vectore

Control of pyrethroid-resistant chagas disease vectors with entomopathogenic fungi

Background: Triatoma infestans-mediated transmission of Tripanosoma cruzi, the causative agent of Chagas disease, remains as a major health issue in southern South America. Key factors of T. infestans prevalence in specific areas of the geographic Gran Chaco region - which extends through northern Argentina, Bolivia, and Paraguay - are both recurrent reinfestations after insecticide spraying and emerging pyrethroid-resistance over the past ten years. Among alternative control tools, the pathogenicity of entomopathogenic fungi against triatomines is already known; furthermore, these fungi have the ability to fully degrade hydrocarbons from T. infestans cuticle and to utilize them as fuel and for incorporation into cellular components. Methodology and Findings: Here we provide evidence of resistance-related cuticle differences; capillary gas chromatography coupled to mass spectrometry analyses revealed that pyrethroid-resistant bugs have significantly larger amounts of surface hydrocarbons, peaking 56.2±6.4% higher than susceptible specimens. Also, a thicker cuticle was detected by scanning electron microscopy (32.1±5.9 μm and 17.8±5.4 μm for pyrethroid-resistant and pyrethroid-susceptible, respectively). In laboratory bioassays, we showed that the virulence of the entomopathogenic fungi Beauveria bassiana against T. infestans was significantly enhanced after fungal adaptation to grow on a medium containing insect-like hydrocarbons as the carbon source, regardless of bug susceptibility to pyrethroids. We designed an attraction-infection trap based on manipulating T. infestans behavior in order to facilitate close contact with B. bassiana. Field assays performed in rural village houses infested with pyrethroid-resistant insects showed 52.4% bug mortality. Using available mathematical models, we predicted that further fungal applications could eventually halt infection transmission. Conclusions: This low cost, low tech, ecologically friendly methodology could help in controlling the spread of pyrethroid-resistant bugs.Facultad de Ciencias Médica

Biological Control of the Chagas Disease Vector <i>Triatoma infestans</i> with the Entomopathogenic Fungus <i>Beauveria bassiana</i> Combined with an Aggregation Cue: Field, Laboratory and Mathematical Modeling Assessment

Current Chagas disease vector control strategies, based on chemical insecticide spraying, are growingly threatened by the emergence of pyrethroid-resistant Triatoma infestans populations in the Gran Chaco region of South America. We have already shown that the entomopathogenic fungus Beauveria bassiana has the ability to breach the insect cuticle and is effective both against pyrethroid-susceptible and pyrethroid-resistant T. infestans, in laboratory as well as field assays. It is also known that T. infestans cuticle lipids play a major role as contact aggregation pheromones. We estimated the effectiveness of pheromone-based infection boxes containing B. bassiana spores to kill indoor bugs, and its effect on the vector population dynamics. Laboratory assays were performed to estimate the effect of fungal infection on female reproductive parameters. The effect of insect exuviae as an aggregation signal in the performance of the infection boxes was estimated both in the laboratory and in the field. We developed a stage-specific matrix model of T. infestans to describe the fungal infection effects on insect population dynamics, and to analyze the performance of the biopesticide device in vector biological control. The pheromone-containing infective box is a promising new tool against indoor populations of this Chagas disease vector, with the number of boxes per house being the main driver of the reduction of the total domestic bug population. This ecologically safe approach is the first proven alternative to chemical insecticides in the control of T. infestans. The advantageous reduction in vector population by delayed-action fungal biopesticides in a contained environment is here shown supported by mathematical modeling.Instituto de Investigaciones Bioquímicas de La PlataFacultad de Ciencias MédicasCentro de Estudios Parasitológicos y de Vectore

Control of Pyrethroid-Resistant Chagas Disease Vectors with Entomopathogenic Fungi

Chagas disease, also known as American Trypanosomiasis, is the most relevant parasitic disease in Latin America, being a major burden that affects mostly poor human populations living in rural areas. The kissing-bugs of the Triatominae family transmit the parasite Trypanosoma cruzi by infectious blood-sucking; Triatoma infestans is the vector of major relevance in the southern Cone of South America. Current control strategies, heavily based on residual insecticide spraying, are threatened by the emergence of pyrethroid-resistant bug populations. Furthermore, ensuring the long-term and sustainable control of this overwhelming disease remains a major challenge. Here we show the utility of a simple, low-cost, biological control methodology against T. infestans bugs, regardless of their susceptibility to pyrethroid insecticides. It is based on the understanding of the initial contact interactions between a mycoinsecticide agent—the fungus Beauveria bassiana—and the host defense barrier, the bug cuticle. The proposed methodology is also supported by present data showing a relationship between the triatomine cuticle width and its hydrocarbon surface components, with insecticide resistance. These results will help to provide a safe and efficient alternative to overcome pyrethroid-resilience of these noxious bugs. A high transfer potential to immediate application in rural communities located in remote areas inaccessible to sanitary control teams, and to the control of other Chagas disease vectors as well, is also envisaged

Taking the pulse of Earth's tropical forests using networks of highly distributed plots

Tropical forests are the most diverse and productive ecosystems on Earth. While better understanding of these forests is critical for our collective future, until quite recently efforts to measure and monitor them have been largely disconnected. Networking is essential to discover the answers to questions that transcend borders and the horizons of funding agencies. Here we show how a global community is responding to the challenges of tropical ecosystem research with diverse teams measuring forests tree-by-tree in thousands of long-term plots. We review the major scientific discoveries of this work and show how this process is changing tropical forest science. Our core approach involves linking long-term grassroots initiatives with standardized protocols and data management to generate robust scaled-up results. By connecting tropical researchers and elevating their status, our Social Research Network model recognises the key role of the data originator in scientific discovery. Conceived in 1999 with RAINFOR (South America), our permanent plot networks have been adapted to Africa (AfriTRON) and Southeast Asia (T-FORCES) and widely emulated worldwide. Now these multiple initiatives are integrated via ForestPlots.net cyber-infrastructure, linking colleagues from 54 countries across 24 plot networks. Collectively these are transforming understanding of tropical forests and their biospheric role. Together we have discovered how, where and why forest carbon and biodiversity are responding to climate change, and how they feedback on it. This long-term pan-tropical collaboration has revealed a large long-term carbon sink and its trends, as well as making clear which drivers are most important, which forest processes are affected, where they are changing, what the lags are, and the likely future responses of tropical forests as the climate continues to change. By leveraging a remarkably old technology, plot networks are sparking a very modern revolution in tropical forest science. In the future, humanity can benefit greatly by nurturing the grassroots communities now collectively capable of generating unique, long-term understanding of Earth's most precious forests.Additional co-authors: Susan Laurance, William Laurance, Francoise Yoko Ishida, Andrew Marshall, Catherine Waite, Hannsjoerg Woell, Jean-Francois Bastin, Marijn Bauters, Hans Beeckman, Pfascal Boeckx, Jan Bogaert, Charles De Canniere, Thales de Haulleville, Jean-Louis Doucet, Olivier Hardy, Wannes Hubau, Elizabeth Kearsley, Hans Verbeeck, Jason Vleminckx, Steven W. Brewer, Alfredo Alarcón, Alejandro Araujo-Murakami, Eric Arets, Luzmila Arroyo, Ezequiel Chavez, Todd Fredericksen, René Guillén Villaroel, Gloria Gutierrez Sibauty, Timothy Killeen, Juan Carlos Licona, John Lleigue, Casimiro Mendoza, Samaria Murakami, Alexander Parada Gutierrez, Guido Pardo, Marielos Peña-Claros, Lourens Poorter, Marisol Toledo, Jeanneth Villalobos Cayo, Laura Jessica Viscarra, Vincent Vos, Jorge Ahumada, Everton Almeida, Jarcilene Almeida, Edmar Almeida de Oliveira, Wesley Alves da Cruz, Atila Alves de Oliveira, Fabrício Alvim Carvalho, Flávio Amorim Obermuller, Ana Andrade, Fernanda Antunes Carvalho, Simone Aparecida Vieira, Ana Carla Aquino, Luiz Aragão, Ana Claudia Araújo, Marco Antonio Assis, Jose Ataliba Mantelli Aboin Gomes, Fabrício Baccaro, Plínio Barbosa de Camargo, Paulo Barni, Jorcely Barroso, Luis Carlos Bernacci, Kauane Bordin, Marcelo Brilhante de Medeiros, Igor Broggio, José Luís Camargo, Domingos Cardoso, Maria Antonia Carniello, Andre Luis Casarin Rochelle, Carolina Castilho, Antonio Alberto Jorge Farias Castro, Wendeson Castro, Sabina Cerruto Ribeiro, Flávia Costa, Rodrigo Costa de Oliveira, Italo Coutinho, John Cunha, Lola da Costa, Lucia da Costa Ferreira, Richarlly da Costa Silva, Marta da Graça Zacarias Simbine, Vitor de Andrade Kamimura, Haroldo Cavalcante de Lima, Lia de Oliveira Melo, Luciano de Queiroz, José Romualdo de Sousa Lima, Mário do Espírito Santo, Tomas Domingues, Nayane Cristina dos Santos Prestes, Steffan Eduardo Silva Carneiro, Fernando Elias, Gabriel Eliseu, Thaise Emilio, Camila Laís Farrapo, Letícia Fernandes, Gustavo Ferreira, Joice Ferreira, Leandro Ferreira, Socorro Ferreira, Marcelo Fragomeni Simon, Maria Aparecida Freitas, Queila S. García, Angelo Gilberto Manzatto, Paulo Graça, Frederico Guilherme, Eduardo Hase, Niro Higuchi, Mariana Iguatemy, Reinaldo Imbrozio Barbosa, Margarita Jaramillo, Carlos Joly, Joice Klipel, Iêda Leão do Amaral, Carolina Levis, Antonio S. Lima, Maurício Lima Dan, Aline Lopes, Herison Madeiros, William E. Magnusson, Rubens Manoel dos Santos, Beatriz Marimon, Ben Hur Marimon Junior, Roberta Marotti Martelletti Grillo, Luiz Martinelli, Simone Matias Reis, Salomão Medeiros, Milton Meira-Junior, Thiago Metzker, Paulo Morandi, Natanael Moreira do Nascimento, Magna Moura, Sandra Cristina Müller, Laszlo Nagy, Henrique Nascimento, Marcelo Nascimento, Adriano Nogueira Lima, Raimunda Oliveira de Araújo, Jhonathan Oliveira Silva, Marcelo Pansonato, Gabriel Pavan Sabino, Karla Maria Pedra de Abreu, Pablo José Francisco Pena Rodrigues, Maria Piedade, Domingos Rodrigues, José Roberto Rodrigues Pinto, Carlos Quesada, Eliana Ramos, Rafael Ramos, Priscyla Rodrigues, Thaiane Rodrigues de Sousa, Rafael Salomão, Flávia Santana, Marcos Scaranello, Rodrigo Scarton Bergamin, Juliana Schietti, Jochen Schöngart, Gustavo Schwartz, Natalino Silva, Marcos Silveira, Cristiana Simão Seixas, Marta Simbine, Ana Claudia Souza, Priscila Souza, Rodolfo Souza, Tereza Sposito, Edson Stefani Junior, Julio Daniel do Vale, Ima Célia Guimarães Vieira, Dora Villela, Marcos Vital, Haron Xaud, Katia Zanini, Charles Eugene Zartman, Nur Khalish Hafizhah Ideris, Faizah binti Hj Metali, Kamariah Abu Salim, Muhd Shahruney Saparudin, Rafizah Mat Serudin, Rahayu Sukmaria Sukri, Serge Begne, George Chuyong, Marie Noel Djuikouo, Christelle Gonmadje, Murielle Simo-Droissart, Bonaventure Sonké, Hermann Taedoumg, Lise Zemagho, Sean Thomas, Fidèle Baya, Gustavo Saiz, Javier Silva Espejo, Dexiang Chen, Alan Hamilton, Yide Li, Tushou Luo, Shukui Niu, Han Xu, Zhang Zhou, Esteban Álvarez-Dávila, Juan Carlos Andrés Escobar, Henry Arellano-Peña, Jaime Cabezas Duarte, Jhon Calderón, Lina Maria Corrales Bravo, Borish Cuadrado, Hermes Cuadros, Alvaro Duque, Luisa Fernanda Duque, Sandra Milena Espinosa, Rebeca Franke-Ante, Hernando García, Alejandro Gómez, Roy González-M., Álvaro Idárraga-Piedrahíta, Eliana Jimenez, Rubén Jurado, Wilmar López Oviedo, René López-Camacho, Omar Aurelio Melo Cruz, Irina Mendoza Polo, Edwin Paky, Karen Pérez, Angel Pijachi, Camila Pizano, Adriana Prieto, Laura Ramos, Zorayda Restrepo Correa, James Richardson, Elkin Rodríguez, Gina M. Rodriguez M., Agustín Rudas, Pablo Stevenson, Markéta Chudomelová, Martin Dancak, Radim Hédl, Stanislav Lhota, Martin Svatek, Jacques Mukinzi, Corneille Ewango, Terese Hart, Emmanuel Kasongo Yakusu, Janvier Lisingo, Jean-Remy Makana, Faustin Mbayu, Benjamin Toirambe, John Tshibamba Mukendi, Lars Kvist, Gustav Nebel, Selene Báez, Carlos Céron, Daniel M. Griffith, Juan Ernesto Guevara Andino, David Neill, Walter Palacios, Maria Cristina Peñuela-Mora, Gonzalo Rivas-Torres, Gorky Villa, Sheleme Demissie, Tadesse Gole, Techane Gonfa, Kalle Ruokolainen, Michel Baisie, Fabrice Bénédet, Wemo Betian, Vincent Bezard, Damien Bonal, Jerôme Chave, Vincent Droissart, Sylvie Gourlet-Fleury, Annette Hladik, Nicolas Labrière, Pétrus Naisso, Maxime Réjou-Méchain, Plinio Sist, Lilian Blanc, Benoit Burban, Géraldine Derroire, Aurélie Dourdain, Clement Stahl, Natacha Nssi Bengone, Eric Chezeaux, Fidèle Evouna Ondo, Vincent Medjibe, Vianet Mihindou, Lee White, Heike Culmsee, Cristabel Durán Rangel, Viviana Horna, Florian Wittmann, Stephen Adu-Bredu, Kofi Affum-Baffoe, Ernest Foli, Michael Balinga, Anand Roopsind, James Singh, Raquel Thomas, Roderick Zagt, Indu K. Murthy, Kuswata Kartawinata, Edi Mirmanto, Hari Priyadi, Ismayadi Samsoedin, Terry Sunderland, Ishak Yassir, Francesco Rovero, Barbara Vinceti, Bruno Hérault, Shin-Ichiro Aiba, Kanehiro Kitayama, Armandu Daniels, Darlington Tuagben, John T. Woods, Muhammad Fitriadi, Alexander Karolus, Kho Lip Khoon, Noreen Majalap, Colin Maycock, Reuben Nilus, Sylvester Tan, Almeida Sitoe, Indiana Coronado G., Lucas Ojo, Rafael de Assis, Axel Dalberg Poulsen, Douglas Sheil, Karen Arévalo Pezo, Hans Buttgenbach Verde, Victor Chama Moscoso, Jimmy Cesar Cordova Oroche, Fernando Cornejo Valverde, Massiel Corrales Medina, Nallaret Davila Cardozo, Jano de Rutte Corzo, Jhon del Aguila Pasquel, Gerardo Flores Llampazo, Luis Freitas, Darcy Galiano Cabrera, Roosevelt García Villacorta, Karina Garcia Cabrera, Diego García Soria, Leticia Gatica Saboya, Julio Miguel Grandez Rios, Gabriel Hidalgo Pizango, Eurídice Honorio Coronado, Isau Huamantupa-Chuquimaco, Walter Huaraca Huasco, Yuri Tomas Huillca Aedo, Jose Luis Marcelo Peña, Abel Monteagudo Mendoza, Vanesa Moreano Rodriguez, Percy Núñez Vargas, Sonia Cesarina Palacios Ramos, Nadir Pallqui Camacho, Antonio Peña Cruz, Freddy Ramirez Arevalo, José Reyna Huaymacari, Carlos Reynel Rodriguez, Marcos Antonio Ríos Paredes, Lily Rodriguez Bayona, Rocio del Pilar Rojas Gonzales, Maria Elena Rojas Peña, Norma Salinas Revilla, Yahn Carlos Soto Shareva, Raul Tupayachi Trujillo, Luis Valenzuela Gamarra, Rodolfo Vasquez Martinez, Jim Vega Arenas, Christian Amani, Suspense Averti Ifo, Yannick Bocko, Patrick Boundja, Romeo Ekoungoulou, Mireille Hockemba, Donatien Nzala, Alusine Fofanah, David Taylor, Guillermo Bañares-de Dios, Luis Cayuela, Íñigo Granzow-de la Cerda, Manuel Macía, Juliana Stropp, Maureen Playfair, Verginia Wortel, Toby Gardner, Robert Muscarella, Hari Priyadi, Ervan Rutishauser, Kuo-Jung Chao, Pantaleo Munishi, Olaf Bánki, Frans Bongers, Rene Boot, Gabriella Fredriksson, Jan Reitsma, Hans ter Steege, Tinde van Andel, Peter van de Meer, Peter van der Hout, Mark van Nieuwstadt, Bert van Ulft, Elmar Veenendaal, Ronald Vernimmen, Pieter Zuidema, Joeri Zwerts, Perpetra Akite, Robert Bitariho, Colin Chapman, Eilu Gerald, Miguel Leal, Patrick Mucunguzi, Miguel Alexiades, Timothy R. Baker, Karina Banda, Lindsay Banin, Jos Barlow, Amy Bennett, Erika Berenguer, Nicholas Berry, Neil M. Bird, George A. Blackburn, Francis Brearley, Roel Brienen, David Burslem, Lidiany Carvalho, Percival Cho, Fernanda Coelho, Murray Collins, David Coomes, Aida Cuni-Sanchez, Greta Dargie, Kyle Dexter, Mat Disney, Freddie Draper, Muying Duan, Adriane Esquivel-Muelbert, Robert Ewers, Belen Fadrique, Sophie Fauset, Ted R. Feldpausch, Filipe França, David Galbraith, Martin Gilpin, Emanuel Gloor, John Grace, Keith Hamer, David Harris, Tommaso Jucker, Michelle Kalamandeen, Bente Klitgaard, Aurora Levesley, Simon L. Lewis, Jeremy Lindsell, Gabriela Lopez-Gonzalez, Jon Lovett, Yadvinder Malhi, Toby Marthews, Emma McIntosh, Karina Melgaço, William Milliken, Edward Mitchard, Peter Moonlight, Sam Moore, Alexandra Morel, Julie Peacock, Kelvin Peh, Colin Pendry, R. Toby Pennington, Luciana de Oliveira Pereira, Carlos Peres, Oliver L. Phillips, Georgia Pickavance, Thomas Pugh, Lan Qie, Terhi Riutta, Katherine Roucoux, Casey Ryan, Tiina Sarkinen, Camila Silva Valeria, Dominick Spracklen, Suzanne Stas, Martin Sullivan, Michael Swaine, Joey Talbot, James Taplin, Geertje van der Heijden, Laura Vedovato, Simon Willcock, Mathew Williams, Luciana Alves, Patricia Alvarez Loayza, Gabriel Arellano, Cheryl Asa, Peter Ashton, Gregory Asner, Terry Brncic, Foster Brown, Robyn Burnham, Connie Clark, James Comiskey, Gabriel Damasco, Stuart Davies, Tony Di Fiore, Terry Erwin, William Farfan-Rios, Jefferson Hall, David Kenfack, Thomas Lovejoy, Roberta Martin, Olga Martha Montiel, John Pipoly, Nigel Pitman, John Poulsen, Richard Primack, Miles Silman, Marc Steininger, Varun Swamy, John Terborgh, Duncan Thomas, Peter Umunay, Maria Uriarte, Emilio Vilanova Torre, Ophelia Wang, Kenneth Young, Gerardo A. Aymard C., Lionel Hernández, Rafael Herrera Fernández, Hirma Ramírez-Angulo, Pedro Salcedo, Elio Sanoja, Julio Serrano, Armando Torres-Lezama, Tinh Cong Le, Trai Trong Le, Hieu Dang Tra

Impairment of Infectivity and Immunoprotective Effect of a LYT1 Null Mutant of Trypanosoma cruzi▿

Trypanosoma cruzi infection of host cells is a complex process in which many proteins participate but only a few of these proteins have been identified experimentally. One parasite factor likely to be involved is the protein product of LYT1, a single-copy gene cloned, sequenced, and characterized by Manning-Cela et al. (Infect. Immun. 69:3916-3923, 2001). This gene was potentially associated with infectivity, since the deletion of both LYT1 alleles in the CL Brenner strain (the wild type [WT]) resulted in a null mutant T. cruzi clone (L16) that shows an attenuated phenotype in cell culture models. The aim of this work was to characterize the infective behavior of L16 in the insect vector and murine models. The infection of adult Swiss mice with 103 trypomastigotes of both clones revealed a significant reduction in infective behavior of L16, as shown by direct parasitemia, spleen index, and quantitation of tissue parasite burden, suggesting the loss of virulence in the null mutant clone. Although L16 blood counts were almost undetectable, blood-based PCRs indicated the presence of latent and persistent infection during all of the study period and epimastigotes were reisolated from hemocultures until 12 months postinfection. Nevertheless, virulence was not restored in L16 by serial passages in mice, and reisolated parasites lacking the LYT1 gene and bearing the antibiotic resistance genes revealed the stability of the genetic manipulation. Histopathological studies showed a strong diminution in the muscle inflammatory response triggered by L16 compared to that triggered by the WT group, consistent with a lower tissue parasite load. A strong protection against a virulent challenge in both L16- and WT-infected mice was observed; however, the immunizing infection by the genetically modified parasite was highly attenuated

A monoallelic deletion of the TcCRT gene increases the attenuation of a cultured Trypanosoma cruzi strain, protecting against an in vivo virulent challenge.

Trypanosoma cruzi calreticulin (TcCRT) is a virulence factor that binds complement C1, thus inhibiting the activation of the classical complement pathway and generating pro-phagocytic signals that increase parasite infectivity. In a previous work, we characterized a clonal cell line lacking one TcCRT allele (TcCRT+/-) and another overexpressing it (TcCRT+), both derived from the attenuated TCC T. cruzi strain. The TcCRT+/- mutant was highly susceptible to killing by the complement machinery and presented a remarkable reduced propagation and differentiation rate both in vitro and in vivo. In this report, we have extended these studies to assess, in a mouse model of disease, the virulence, immunogenicity and safety of the mutant as an experimental vaccine. Balb/c mice were inoculated with TcCRT+/- parasites and followed-up during a 6-month period. Mutant parasites were not detected by sensitive techniques, even after mice immune suppression. Total anti-T. cruzi IgG levels were undetectable in TcCRT+/- inoculated mice and the genetic alteration was stable after long-term infection and it did not revert back to wild type form. Most importantly, immunization with TcCRT+/- parasites induces a highly protective response after challenge with a virulent T. cruzi strain, as evidenced by lower parasite density, mortality, spleen index and tissue inflammatory response. TcCRT+/- clones are restricted in two important properties conferred by TcCRT and indirectly by C1q: their ability to evade the host immune response and their virulence. Therefore, deletion of one copy of the TcCRT gene in the attenuated TCC strain generated a safe and irreversibly gene-deleted live attenuated parasite with high immunoprotective properties. Our results also contribute to endorse the important role of TcCRT as a T. cruzi virulence factor