Search

Made available in DSpace on 2018-12-11T17:20:44Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-07-15 Fundação de Apoio à Pesquisa do Distrito Federal Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) The eucalyptus brown looper, Thyrinteina arnobia (Stoll, 1782) (Lepidoptera: Geometridae), is the main lepidopteran defoliator of eucalyptus plantations in Brazil. Outbreaks of this insect pest are common in Brazil and can affect the productivity of planted forests severely. T. arnobia caterpillars from a laboratory colony with viral infection symptoms were analyzed by electron microscopy that revealed polyhedral occlusion bodies (OBs) with several icosahedral virus particles embedded. Analysis of its genetic material showed ten segments of dsRNA, which confirmed this virus as a possible member of the genus Cypovirus. Phylogenetic analysis of the whole genome sequence revealed its close relationship with other isolates of Cypovirus 14 species and according to these results we proposed the name Thyrinteina arnobia cypovirus 14 (TharCPV-14) for this new virus isolate. Further research will be necessary in order to analyze the potential of this virus as a biopesticide. Plant Protection Department FCA/UNESP—São Paulo State University, Campus of Botucatu Biochemistry and Molecular Biology Department Federal University of Santa Maria, Santa Maria Cell Biology Department UnB—University of Brasília Applied Biology Department FCAV/UNESP—São Paulo State University, Campus of Jaboticabal Plant Protection Department FCAV/UNESP—São Paulo State University, Campus of Jaboticabal Plant Protection Department FCA/UNESP—São Paulo State University, Campus of Botucatu Applied Biology Department FCAV/UNESP—São Paulo State University, Campus of Jaboticabal Plant Protection Department FCAV/UNESP—São Paulo State University, Campus of Jaboticabal Fundação de Apoio à Pesquisa do Distrito Federal: 193.001532/2016 CNPq: 407908/2013-7

Resistance of Plutella xylostella populations to chemical insecticides has made its management difficult, and the utilization of resistant cabbage cultivars has been shown to be a useful alternative. The objective of this study was to demonstrate the induction of cabbage plant resistance to P. xylostella using PGPR and injuries caused by the pest larvae as elicitors. Therefore, we evaluated the insects’ responses utilizing a specific bioassay. Furthermore, this assay was used for selecting a PGPR strain that affects the insect’s biology, and to examine molecular and biochemical responses of the plants influenced by the plant-microbe-insect interaction. Among the strains used in this study, Kluyvera ascorbata showed the most relevant results by influencing biological characteristics of the insect. Thus, the following tests demonstrated that the cited strain possesses a high influence on plant metabolism when it undergoes different types of stress such as injuries caused by the pest. These findings were determined from the different responses obtained by the chemical analyses of the tested plants and from the differentiation in the genetic sequences obtained from plants inoculated with or without PGPR that were injured by the pest. The PGPR K. ascorbata alters the metabolism of cabbage plants, which directs a specific plant defense against P. xylostella.

Made available in DSpace on 2019-10-06T16:25:17Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-06-01 Fundação de Apoio à Pesquisa do Distrito Federal Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) We describe an unexpected feature observed for the heterologous expression of the Thyrinteina arnobia cypovirus polyhedrin from a recombinant baculovirus infection in different insect cell lines. The in cellulo-formed crystals varied in size and shape depending on the cell line. Crystals formed in Trichoplusia ni-derived cells were cubic (0.1-2 μm) and localized in both the nucleus and cytoplasm, whereas those formed in Spodoptera frugiperda-derived cells were ovate and ellipsoidal (0.1-3 μm) and also localized in both the nucleus and cytoplasm. The molecular basis for differences in the morphology, size, and location of cypovirus occlusion bodies is unclear, and cellular proteins might play a role in their formation and location. Cell Biology Department UnB-University of Brasília Laboratory of Insect Virology Department of Biochemistry and Molecular Biology Federal University of Santa Maria Plant Protection Department FCA/UNESP São Paulo State University, Campus of Botucatu Applied Biology Department FCAV/UNESP São Paulo State University, Campus of Jaboticabal Plant Protection Department FCA/UNESP São Paulo State University, Campus of Botucatu Applied Biology Department FCAV/UNESP São Paulo State University, Campus of Jaboticabal Fundação de Apoio à Pesquisa do Distrito Federal: 193.001532/2016 CNPq: 407908/2013-7 CNPq: 428799/2018-3 CAPES: CAPES/PROEX 23038.005848/2018-31

Made available in DSpace on 2018-11-26T16:17:02Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-12-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) A novel combination of structurally simple, high-rate horizontal anaerobic reactors installed in series was used to treat swine wastewater. The reactors maintained stable pH, alkalinity, and volatile acid levels. Removed chemical oxygen demand (COD) represented 68% of the total, and the average specific methane production was 0.30 L CH4 (g removed CODtot) (1). In addition, next-generation sequencing and quantitative real-time PCR analyses were used to explore the methane-producing Archaea and microbial diversity. At least 94% of the sludge diversity belong to the Bacteria and Archaea, indicating a good balance of microorganisms. Among the Bacteria the Proteobacteria, Bacteroidetes and Firmicutes were the most prevalent phyla. Interestingly, up to 12% of the sludge diversity belongs to methane-producing orders, such as Methanosarcinales, Methanobacteriales and Methanomicrobiales. In summary, this system can efficiently produce methane and this is the first time that horizontal anaerobic reactors have been evaluated for the treatment of swine wastewater. (C) 2015 Elsevier Ltd. All rights reserved. Univ Estadual Paulista, Fac Agr & Vet Sci, Dept Rural Engn, Lab Environm Sanitat, BR-14884900 Jaboticabal, SP, Brazil Univ Estadual Paulista, Fac Agr & Vet Sci, Dept Technol, Lab Biochem & Mol Biol, BR-14884900 Jaboticabal, SP, Brazil Bacterial Genet Lab, Dept Appl Biol, Fac Agr & Vet Sci, BR-14884900 Jaboticabal, SP, Brazil Fac Technol Jaboticabal, BR-14883130 Jaboticabal, SP, Brazil Univ Estadual Paulista, Fac Agr & Vet Sci, Dept Rural Engn, Lab Environm Sanitat, BR-14884900 Jaboticabal, SP, Brazil Univ Estadual Paulista, Fac Agr & Vet Sci, Dept Technol, Lab Biochem & Mol Biol, BR-14884900 Jaboticabal, SP, Brazil CAPES: PNPD -3137/2010 CNPq: 483118/2011-7

Livestock manure may contain pathogenic microorganisms which pose a risk to the health of animal or humans if the manure is not adequately treated or disposed of. To determine the fate of Shiga toxigenic Escherichia coli (STEC) non O157 in composted manure from naturally colonized sheep, fresh manure was obtained from animals carrying bacterial cells with stx1/ stx2 genes. Two composting systems were used, aerated and non-aerated, and the experiments were done in Dracena city, São Paulo State. Every week, for seven weeks, one manure sample from six different points in both systems was collected and cultured to determine the presence of E. coli, the presence of the virulence genes in the cells, and also the susceptibility to 10 antimicrobial drugs. The temperature was verified at each sampling. STEC non-O157 survived for 49 days in both composting systems. E. coli non-STEC showing a high degree of antibiotic resistance was recovered all long the composting period. No relationship was established between the presence of virulence genes and antibiotic resistance. The presence of virulence genes and multiple antibiotic resistances in E. coli implicates a potential risk for these genes spread in the human food chain, which is a reason for concern.

Made available in DSpace on 2018-12-11T17:35:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-03-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) The Periplaneta americana species is an annoyance to man, causing allergies and damage to clothes and documents. It has the ability to spread pathogens and requires control measures. Control with natural enemies is less aggressive and can currently be applied with less risk than other techniques, such as chemical control, which is the main method used worldwide to control its post-embryonic stages. The potential microbial control of nymphs and adults of this pest has been shown, but little is known about its oothecae. There are isolates of fungal species that can be used to achieve this aim, but they may have innate differences in their virulence and ability to spread. This study aimed to identify fungal isolates JAB 68 and IBCB 35 through genetic sequencing of the ITS1-5.8S-ITS2 region, analyze their ability to synthesize chitinase, and investigate and compare their aggressiveness against P. americana oothecae and their influence on nymph eclosion. Fungal suspensions were inoculated into minimal medium containing glucose (control) as the sole carbon source and 1% colloidal chitin to determine the chitinolytic activity on the 4th, 7th and 10th days and sporulation on the 10th day. To obtain mortality, extrusion and the compiled number of hatched nymphs, oothecae were sprayed with suspensions of the isolates as follows: T1 – no application; T2 – aqueous solution of Tween 80® 0.1% (vehicle suspension for treatments T3 to T8); T3 – 2 × 109 conidia/mL of the JAB 68 isolate; T4 – 2 × 108 con./mL of the JAB 68 isolate; T5 – 2 × 107 con./mL of the JAB 68 isolate; T6 – 2 × 109 con./mL of the IBCB 35 isolate; T7 – 2 × 108 con./mL of the IBCB 35 isolate; T8 – 2 × 107 con./mL of the IBCB 35 isolate. The JAB 68 and IBCB 35 isolates were identified as belonging to the species Metarhizium anisopliae and Beauveria bassiana, respectively. Chitinolytic activity and extrusion were good parameters for evaluating the fungi's action on oothecal control. The most aggressive entomopathogen was M. anisopliae isolate JAB 68, with shorter time for fungus extrusion at a concentration of 2 × 107 con./mL. B. bassiana reduced the number of hatched nymphs at a concentration of 2 × 108 con./mL. Both fungi are capable of infecting and killing P. americana's oothecae and reducing the number of nymphs hatched. Department of Plant Protection UNESP Department of Plant Production UNESP Department of Preventive Veterinary Medicine and Animal Reproduction UNESP Department of Applied Biology UNESP Department of Plant Protection UNESP Department of Plant Production UNESP Department of Preventive Veterinary Medicine and Animal Reproduction UNESP Department of Applied Biology UNESP FAPESP: 2012/14503-1

Made available in DSpace on 2018-12-11T17:30:49Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-03-01 Exopolysaccharide (EPS) are produced by a diverse of rhizobia species and has been demonstrated to be a bioemulsifier with potential applications in the degradation of hydrocarbons. In the present study, attempts were made to obtain the new exopolysaccharide production by Rhizobium tropici (SEMIA 4080 and MUTZC3) strains during growth on hydrocarbon substrate. Under the different cultivation conditions, the high molecular weight exopolysaccharides from Rhizobium tropici strains cultivated for 96 h mainly consisted of carbohydrates (79–85%) and a low percentage of protein. The EPSC3-D differed from the others, with only 60% of carbohydrate. However, all strains produced polymers with distinct rheology properties, such as viscosity of each EPS sample, suitable for different applications. In addition, RP-HPLC, FTIR and NMR studies revealed EPS produced by rhizobia strains were similar indicating minimal difference between EPS compositions. São Paulo State University (Unesp) School of Agricultural and Veterinarian Sciences, Jaboticabal Embrapa Instrumentação, Rua XV de Novembro 1452, São Carlos São Paulo State University (Unesp) School of Agricultural and Veterinarian Sciences, Jaboticabal

The Brazilian microbiota has great potential richness for industrial use, given its mega-diversity. Despite the advances in international research that have provided access to such microbiota, via several approaches (metagenomics; second-generation DNA sequencing, in situ cultivation, and as a consequence high-throughput screening, etc.) a glimpse into the research output in Brazil demonstrates that such immense potential has been poorly explored. Even though the Brazilian scientific community has reached a degree of international excellence in research recognition, there is still strong centralization of knowledge and of biotechnology enterprises in the Southeast Region of the country, which greatly limits access to our multitude of biomes and ecosystems. Another problem is the lack of communication between the knowledge generation centers and practical efforts in the field, resulting in very little national intelligence reaching the consumer market. Consequently, the internal biotechnology market prioritizes imports, even though there are available domestic resources to generate competitiveness at a global level. Academic and industry integration initiatives through innovative agencies have demonstrated a path to bridge the gap between the “ownership” and the “usage capacity” of the country’s rich microbial diversity.

Made available in DSpace on 2018-12-11T17:09:58Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-01-01 The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs) were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests. Department of Biologia Aplicada � Agropecu�ria Universidade Estadual Paulista Department of Agronomia Universidade Federal Rural de Pernambuco Departamento de Fitossanidade/FCAV/UNESP Universidade Estadual Paulista Department of Biologia Aplicada � Agropecu�ria Universidade Estadual Paulista Departamento de Fitossanidade/FCAV/UNESP Universidade Estadual Paulista

Made available in DSpace on 2014-09-30T18:18:20Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-02-01Bitstream added on 2014-10-01T14:04:14Z : No. of bitstreams: 1 S0100-204X2014000200079.pdf: 748597 bytes, checksum: 6b6622d0023ecd80832b406b6f8b79aa (MD5) O objetivo deste trabalho foi avaliar a suscetibilidade das lagartas Anticarsia gemmatalis (Lepidoptera: Erebidae) e Chrysodeixis includens (Lepidoptera: Noctuidae) às proteínas Cry1 e Vip3A, bem como determinar se há a interação entre essas proteínas no controle das duas espécies. Bioensaios com as proteínas isoladas e em combinações foram realizados, e as concentrações letais CL50 e CL90 foram estimadas para cada condição. As proteínas Cry1Aa, Cry1Ac e Vip3Af foram as mais efetivas no controle de A. gemmatalis, enquanto Cry1Ac, Vip3Aa e Vip3Af foram mais efetivas no de C. includens. As proteínas Cry1Ac e Cry1Ca causaram maior inibição do desenvolvimento das larvas sobreviventes à CL50, em ambas as espécies. Combinações entre Vip3A e Cry1 apresentam efeito sinérgico no controle das espécies e a combinação Vip3Aa+Cry1Ea destaca-se no controle de A. gemmatalis e C. includens. Essas proteínas combinadas são promissoras na construção de plantas piramidadas, para o controle simultâneo das pragas. The objective of this work was to evaluate the susceptibility of Anticarsia gemmatalis (Lepidoptera: Erebidae) and Chrysodeixis includens (Lepidoptera: Noctuidae) caterpillars to Cry1 and Vip3A proteins, as well as to determine if there is any interaction between these proteins on the control of the two species. Bioassays with both isolated and combined proteins were carried out, and lethal concentrations LC50 and LC90 were estimated for each condition. Cry1Aa, Cry1Ac, and Vip3Af were the more effective proteins for the control of A. gemmatalis, while Cry1Ac, Vip3Aa, and Vip3Af were more effective for the control of C. includens. Cry1Ac and Cry1Ca proteins caused the highest inhibition to the development of larvae that survived the LC50 dose in both species. Different combinations of Vip3A and Cry1 have synergistic effect in the control of both species, and the combination Vip3Aa + Cry1Ea showed an outstanding control of A. gemmatalis and C. includens. These proteins are promising for building pyramided plants for the simultaneous control of the pests. Universidade Estadual Paulista Departamento de Biologia Aplicada à Agropecuária Unesp Departamento de Fitossanidade Universidade Estadual Paulista Departamento de Biologia Aplicada à Agropecuária Unesp Departamento de Fitossanidade

Made available in DSpace on 2014-10-01T13:08:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-09-01Bitstream added on 2014-10-01T14:02:49Z : No. of bitstreams: 1 S0100-204X2013000900005.pdf: 1017630 bytes, checksum: f0029d05a5d1bdb6e2ed6446db5b88ba (MD5) O objetivo deste trabalho foi caracterizar o gene vip3A de Bacillus thuringiensis e verificar a toxicidade da proteína Vip3Aa50 a larvas da lagarta-do-cartucho (Spodoptera frugiperda) e da lagarta-da-soja (Anticarsia gemmatalis). O gene vip3A foi amplificado por PCR, com iniciadores específicos, e gerou um fragmento de 2.370 pb. Esse fragmento foi clonado em vetor pGEM-T Easy e, em seguida, sequenciado, subclonado em vetor de expressão pET-28a (+) e inserido em células de Escherichia coli BL21 (DE3). A expressão da proteína Vip3Aa50 foi induzida por isopropil-β-D-1-tiogalactopiranosídeo (IPTG), visualizada em SDS-PAGE e detectada por Western blot. Os ensaios de toxicidade revelaram alta atividade da proteína Vip3Aa50 contra as larvas neonatas da lagarta-da-soja e da lagarta-do-cartucho, com CL50 de 20,3 e 79,6 ng cm-2, respectivamente. O gene vip3Aa50 é um novo gene da classe vip3A. The objective of this work was to characterize the vip3A gene of Bacillus thuringiensis and to evaluate the toxicity of Vip3Aa50 protein to the fall armyworm (Spodoptera frugiperda) and velvetbean caterpillar (Anticarsia gemmatalis) larvae. The gene vip3A was amplified by specific PCR primers, generating a 2,370-bp fragment. This fragment was cloned into the pGEM-T Easy vector, and then it was sequenced, subcloned into the pET-28a (+)'s expression vector, and inserted into Escherichia coli BL21 (DE3) cells. The Vip3Aa50 protein expression was induced by isopropyl-β-D-1-thiogalactopyranoside (IPTG), visualized in SDS-PAGE, and detected by Western blot. The toxicity bioassay showed a high activity of Vip3Aa50 protein against both velvetbean and fall armyworm neonate larvae, with LC50 at 20.3 and 79.6 ng cm-2 respectively. The vip3Aa50 gene, is a new gene of vip3A class. Universidade Estadual Paulista Júlio de Mesquita Filho Universidade Estadual Paulista Júlio de Mesquita Filho

Domestic ruminants, especially sheep and cattle, are the main reservoirs of Shiga toxin-producing Escherichia coli (STEC) and may transmit this pathogen to human. Probiotics are "live organisms that, when administered in adequate amounts, confer a health benefit on the host", decreasing the infection and its dissemination. However, these benefits are conferred only after the colonization of probiotic strains in the gut of the animal, which may be impaired by normal microbiota. The aims of this study were to determine whether the inoculation of sheep with probiotic strains decreases the shedding of STEC and to determine whether the age of sheep interferes with this protective effect. Sheep that received oral inoculums at a concentration of 2 × 109 cells per mL of viable STEC bacteria, which are carriers of stx1, stx2 and eae genes, were compared with other sheep that did not receive inoculums. When probiotic bacteria were inoculated together with the STEC, the number of pathogenic bacteria in the population was similar to the control. The protective effect of probiotic strains was largest in groups with younger animals than with older animals. These findings suggest that the use of probiotic strains in sheep may decrease the intestinal shedding by STEC as well as the fact that the age of the sheep may interfere in the protective effect of probiotics against colonization by STEC. Key words: Probiotic, protective effect, Escherichia coli, sheep.

Cockroach control is performed by the application of chemical insecticides which exert high selective pressure on populations and introduces synthetic substances in the environment, motivating the search for other methods of control such as entomopathogenic fungi. The objectives of this study were to investigate the pathogenicity of the JAB 42 Aspergillus westerdijkiae to females and oothecae of Periplaneta americana and to demonstrate its mechanism of action on oothecae. Suspensions containing 106 to 108 conidia/ml were used to infect females and oothecae. Mortality and other variables such as scanning electron microscopy were used to demonstrate the mechanism of action of the fungus. The isolated JAB 42 A. westerdijkiae is pathogenic to oothecae of P. americana, with low capacity to kill females. Adhesion, germination, penetration and extrusion of the fungus on the cockroach oothecae were observed. RESUMO: O controle de baratas realizado através da aplicação de inseticidas químicos exerce alta pressão seletiva sobre as populações e introduz substâncias sintéticas no ambiente, motivando a procura por outros métodos de controle, como os fungos entomopatogênicos. Os objetivos deste trabalho foram investigar a patogenicidade do isolado JAB 42 de Aspergillus westerdijkiae a fêmeas e ootecas de Periplaneta americana e demonstrar o mecanismo de ação sobre ootecas. Suspensões contendo 106 a 108 conídios/mL do isolado foram usadas para infectar fêmeas e ootecas. A ação do fungo foi analisada pela mortalidade e outras variáveis, e por microscopia eletrônica de varredura. O isolado JAB 42 de A. westerdijkiae é patogênico a ootecas de P. americana, tendo baixa capacidade de matar fêmeas. Foi observada a adesão, germinação, penetração e extrusão do fungo sobre ootecas da barata.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:42:36Z No. of bitstreams: 1 S1519-69842012000200003.pdf: 195462 bytes, checksum: 44ca565ed66711725b8a7e3542e12e5e (MD5) Made available in DSpace on 2013-08-22T18:42:36Z (GMT). No. of bitstreams: 1 S1519-69842012000200003.pdf: 195462 bytes, checksum: 44ca565ed66711725b8a7e3542e12e5e (MD5) Previous issue date: 2012-05-01 Made available in DSpace on 2013-09-30T19:29:58Z (GMT). No. of bitstreams: 2 S1519-69842012000200003.pdf: 195462 bytes, checksum: 44ca565ed66711725b8a7e3542e12e5e (MD5) S1519-69842012000200003.pdf.txt: 20901 bytes, checksum: e67a35bb50c9b2d6e4f308f25f4e2be0 (MD5) Previous issue date: 2012-05-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:13:05Z No. of bitstreams: 2 S1519-69842012000200003.pdf: 195462 bytes, checksum: 44ca565ed66711725b8a7e3542e12e5e (MD5) S1519-69842012000200003.pdf.txt: 20901 bytes, checksum: e67a35bb50c9b2d6e4f308f25f4e2be0 (MD5) Made available in DSpace on 2014-05-20T13:13:05Z (GMT). No. of bitstreams: 2 S1519-69842012000200003.pdf: 195462 bytes, checksum: 44ca565ed66711725b8a7e3542e12e5e (MD5) S1519-69842012000200003.pdf.txt: 20901 bytes, checksum: e67a35bb50c9b2d6e4f308f25f4e2be0 (MD5) Previous issue date: 2012-05-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Bacillus thuringiensis é entomopatogênica, por produzir proteínas cristais, denominadas proteínas Cry, as quais são codificadas pelos genes cry. Essa bactéria atua no controle biológico de insetos-praga de culturas economicamente importantes, bem como no controle de insetos vetores causadores de doenças, como o Aedes aegypti, mosquito transmissor do vírus da dengue. Os isolados dessa bactéria podem ser caracterizados pelo conteúdo de genes cry que possuem e, assim, predizer o alvo de controle dos mesmos às diferentes ordens de insetos. Com o objetivo de encontrar isolados eficientes no controle do vetor A. aegypti, o presente trabalho isolou 76 colônias de B. thuringiensis a partir de 30 amostras de solo oriundas de Ilhabela-SP, município que se caracteriza por realizar controle biológico de simulídeos com essa bactéria. Os 76 isolados foram sequenciados na região da subunidade ribossomal 16S e caracterizados molecularmente quanto ao conteúdo de genes cry díptero-específicos. No total, oito isolados (10,52% do total) apresentaram bandas para os genes cry4Aa, cry4Ba e cry10Aa, sendo os mesmos testados contra larvas de A. aegypti por meio de bioensaios. A presença e/ou ausência dos genes cry foi associada à mortalidade média de larvas. Dentre os isolados estudados, sete (9,2% do total) apresentaram elevado potencial de controle às larvas de A. aegypti, sendo assim considerados como promissores para o manejo do controle biológico de larvas de A. aegypti com a bactéria B. thuringiensis. The entomophatogenic bacterium Bacillus thuringiensis produces crystal proteins, named Cry proteins which are encoded by the cry genes. This bacterium is used on biological control of important economical pests, as well as in the control of disease´s vectors, such as Aedes aegypti, a mosquito that transmits the dengue viruses. Isolates of this bacterium can be characterized by the content of cry genes and this prediction helps target different insect orders. In this research, we isolated 76 colonies of B. thuringiensis from 30 soil samples that were taken from Ilha Bela (SP, Brazil), a place where simulids are already biologically controlled by B. thuringiensis, to find bacterial isolates that were capable of controlling A. aegypti. The 16S ribosomal subunit genes of the selected isolates were sequenced, and the isolates were molecularly characterized based on their Dipteran-specific cry gene contents. Eight of the 76 isolates (10.52%) contained the cry4Aa, cry4Ba or cry10Aa genes, these isolates were carried out against A. aegypti larvae on bioassay. The presence or absence of specific cry genes was associated with the observed average larval mortalities. From the 76 isolates, seven (9.2%) were potentially able to control A. aegypti larvae. Therefore these are promising isolates for the biological control of A. aegypti larvae. Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinária Laboratório de Genética de Bactérias e Biotecnologia Aplicada Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinária Laboratório de Genética de Bactérias e Biotecnologia Aplicada FAPESP: 06/58080-6

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:54:48Z No. of bitstreams: 1 S0006-87052012000300007.pdf: 333471 bytes, checksum: 7534f07e73025e716109c75fb5832d81 (MD5) Made available in DSpace on 2013-08-22T18:54:48Z (GMT). No. of bitstreams: 1 S0006-87052012000300007.pdf: 333471 bytes, checksum: 7534f07e73025e716109c75fb5832d81 (MD5) Previous issue date: 2012-01-01 Made available in DSpace on 2013-09-30T19:46:55Z (GMT). No. of bitstreams: 2 S0006-87052012000300007.pdf: 333471 bytes, checksum: 7534f07e73025e716109c75fb5832d81 (MD5) S0006-87052012000300007.pdf.txt: 36789 bytes, checksum: 6dc1ff57fefe059eab981a7bda9fdfef (MD5) Previous issue date: 2012-01-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:13:05Z No. of bitstreams: 2 S0006-87052012000300007.pdf: 333471 bytes, checksum: 7534f07e73025e716109c75fb5832d81 (MD5) S0006-87052012000300007.pdf.txt: 36789 bytes, checksum: 6dc1ff57fefe059eab981a7bda9fdfef (MD5) Made available in DSpace on 2014-05-20T13:13:05Z (GMT). No. of bitstreams: 2 S0006-87052012000300007.pdf: 333471 bytes, checksum: 7534f07e73025e716109c75fb5832d81 (MD5) S0006-87052012000300007.pdf.txt: 36789 bytes, checksum: 6dc1ff57fefe059eab981a7bda9fdfef (MD5) Previous issue date: 2012-01-01 A bactéria Bacillus thuringiensis Berliner produz um corpo de inclusão paraesporal (cristal) de natureza proteica, formado durante a esporulação, que atua de forma eficiente no controle de insetos-praga de culturas economicamente importantes. Esse cristal é constituído de proteínas Cry, que são codificadas pelos genes cry; um isolado pode ser caracterizado pelo conteúdo de genes cry que apresenta. Visando caracterizar novos isolados no combate de insetos-praga pertencentes às ordens Lepidoptera e Coleoptera, 76 isolados bacterianos foram analisados molecularmente e tiveram seu potencial de controle avaliado por meio de bioensaios com larvas de Spodoptera frugiperda (J.E. Smith), Sphenophorus levis Vaurie e Tenebrio molitor Linnaeus. As análises moleculares indicaram 11 isolados (14,5% da coleção), contendo genes lepidóptero-específicos e 17 (22,37%) com genes coleóptero-específicos. As análises de patogenicidade revelaram dois isolados com alto potencial de controle para lagartas de S. frugiperda, um para larvas de S. levis e seis prejudiciais ao desenvolvimento das larvas de T. molitor. Esses isolados de B. thuringiensis podem ser promissores no controle biológico das referidas pragas. Bacillus thuringiensis Berliner produces protein crystals that act effectively on the control of pests for several relevant crops. The cry genes codes for these proteins are named Cry proteins. The cry gene profile can potentially set each bacterial isolate as a potential pest control agent. Aiming to analyze new isolates for the controlling of pests belonging to the orders Lepidoptera and Coleoptera, 76 samples were molecularly analyzed and had their potential for control evaluated through bioassays with larvae of Spodoptera frugiperda (J.E. Smith), Sphenophorus levis Vaurie and Tenebrio molitor Linnaeus. Based on molecular analysis data, 11 samples (14.5%) were able to control lepidopteran specific pests due to their cry genes content, and 17 samples (22.37%) exhibited coleopteran specific cry genes. The bioassays results indicated two samples with highly effective against S. frugiperda, one against S. levis and six with high toxicity against T. molitor larvae development. The selected B. thuringiensis isolates may be promising for the biological control of these pests. Universidade Federal de São Carlos (UFSCar) Universidade Estadual Paulista Universidade Estadual Paulista

Resumo: O objetivo deste trabalho foi avaliar a toxicidade e a capacidade de ligação das proteínas Cry1Aa, Cry1Ab, Cry1Ac e Cry1Ca, de Bacillus thuringiensis, a receptores intestinais de Helicoverpa armigera. Realizou-se análise de ligação das proteínas ativadas às vesículas de membrana da microvilosidade apical (VMMA) do intestino médio deH. armigera, além de ensaios de competição heteróloga para avaliar sua capacidade de ligação. Cry1Ac destacou-se como a proteína mais tóxica, seguida por Cry1Ab e Cry1Aa. A proteína Cry1Ca não foi tóxica às lagartas e, portanto, não foi possível determinar os seus parâmetros de toxicidade CL50 e CL90. As proteínas Cry1Aa, Cry1Ab e Cry1Ac são capazes de se ligar a um mesmo receptor nas membranas intestinais, o que aumenta o risco do desenvolvimento de resistência cruzada. Portanto, a utilização conjunta dessas proteínas deve ser evitada. Abstract: The objective of this work was to evaluate the toxicity and the binding capacity of the Cry1Aa, Cry1Ab, Cry1Ac, and Cry1Ca proteins, from Bacillus thuringiensis, to Helicoverpa armigera intestine receptors. Binding analysis of the activated proteins to the brush-border membrane vesicles (BBMV) in the midgut of H.armigera, besides heterologous competition assays to evaluate their binding capacity, was performed. Cry1Ac stood out as the most toxic protein, followed by Cry1Ab and Cry1Aa. The Cry1Ca protein had no toxicity to the caterpillars and, therefore, it was not possible to evaluate its LC50 and LC90 toxicity parameters. The Cry1Aa, Cry1Ab, and Cry1Ac proteins are able to bind themselves to the same receptor in the midgut membrane, which increases the risk of developing cross-resistance. Therefore, the use of these proteins together should be avoided.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:47:12Z No. of bitstreams: 1 S0100-204X2011000900012.pdf: 451597 bytes, checksum: e01a22a0a9bd3365c17975ce7550556a (MD5) Made available in DSpace on 2013-08-22T18:47:12Z (GMT). No. of bitstreams: 1 S0100-204X2011000900012.pdf: 451597 bytes, checksum: e01a22a0a9bd3365c17975ce7550556a (MD5) Previous issue date: 2011-09-01 Made available in DSpace on 2013-09-30T19:36:55Z (GMT). No. of bitstreams: 2 S0100-204X2011000900012.pdf: 451597 bytes, checksum: e01a22a0a9bd3365c17975ce7550556a (MD5) S0100-204X2011000900012.pdf.txt: 34803 bytes, checksum: 3e14b26c92dc130c0c2a7414cf973b52 (MD5) Previous issue date: 2011-09-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:13:05Z No. of bitstreams: 2 S0100-204X2011000900012.pdf: 451597 bytes, checksum: e01a22a0a9bd3365c17975ce7550556a (MD5) S0100-204X2011000900012.pdf.txt: 34803 bytes, checksum: 3e14b26c92dc130c0c2a7414cf973b52 (MD5) Made available in DSpace on 2014-05-20T13:13:05Z (GMT). No. of bitstreams: 2 S0100-204X2011000900012.pdf: 451597 bytes, checksum: e01a22a0a9bd3365c17975ce7550556a (MD5) S0100-204X2011000900012.pdf.txt: 34803 bytes, checksum: 3e14b26c92dc130c0c2a7414cf973b52 (MD5) Previous issue date: 2011-09-01 O objetivo deste trabalho foi identificar e caracterizar os genes cry3, vip1, vip2 e vip1/vip2 em uma coleção de 1.078 isolados de Bacillus thuringiensis potencialmente tóxicos para larvas de coleópteros. Foram utilizados pares de oligonucleotídeos iniciadores gerais obtidos a partir de regiões conservadas dos genes e do alinhamento de sequências consenso. Posteriormente, os isolados positivos foram caracterizados por meio da técnica de PCR‑RFLP, tendo-se utilizado enzimas de restrição específicas, para identificar novas subclasses de genes nos isolados. Cento e cinquenta e um isolados foram positivos para os genes avaliados, com maior frequência para o gene vip1/vip2 (139 isolados). Pela técnica de PCR‑RFLP, foram observados 14 perfis polimórficos, o que indica a presença de diferentes alelos e, consequentemente, de distintas subclasses desses genes. The objective of this work was to identify and characterize cry3, vip1, vip2 and vip1/vip2 genes in a collection of 1,078 Bacillus thuringiensis isolates potentially toxic against Coleoptera larvae. Pairs of primers derived from conserved regions of genes and from sequence alignment consensus were used. Subsequently, positive isolates were characterized by PCR‑RFLP, using specific restriction enzymes to identify new subclasses of genes in the isolates. One hundred and fifty‑one isolates were positive for the evaluated genes, with higher frequency for the vip1/vip2 gene (139 isolates). By PCR‑RFLP, 14 polymorphic profiles were observed, indicating the presence of different alleles, and, therefore, of distinct subclasses of these genes. Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias Universidade de São Paulo Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:42:06Z No. of bitstreams: 1 S1519-566X2010000500015.pdf: 849275 bytes, checksum: 81a6544960f05f823f24ee106f05c8e0 (MD5) Made available in DSpace on 2013-08-22T18:42:06Z (GMT). No. of bitstreams: 1 S1519-566X2010000500015.pdf: 849275 bytes, checksum: 81a6544960f05f823f24ee106f05c8e0 (MD5) Previous issue date: 2010-10-01 Made available in DSpace on 2013-09-30T17:51:24Z (GMT). No. of bitstreams: 2 S1519-566X2010000500015.pdf: 849275 bytes, checksum: 81a6544960f05f823f24ee106f05c8e0 (MD5) S1519-566X2010000500015.pdf.txt: 44538 bytes, checksum: 7b80df7015a2c7deca2e67b7c9c0b790 (MD5) Previous issue date: 2010-10-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:13:49Z No. of bitstreams: 2 S1519-566X2010000500015.pdf: 849275 bytes, checksum: 81a6544960f05f823f24ee106f05c8e0 (MD5) S1519-566X2010000500015.pdf.txt: 44538 bytes, checksum: 7b80df7015a2c7deca2e67b7c9c0b790 (MD5) Made available in DSpace on 2014-05-20T13:13:49Z (GMT). No. of bitstreams: 2 S1519-566X2010000500015.pdf: 849275 bytes, checksum: 81a6544960f05f823f24ee106f05c8e0 (MD5) S1519-566X2010000500015.pdf.txt: 44538 bytes, checksum: 7b80df7015a2c7deca2e67b7c9c0b790 (MD5) Previous issue date: 2010-10-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Aedes aegypti (L.), the main vector of dengue fever in Brazil, has been controlled with the use of massive chemical products, contributing to the development of resistance and decreasing the insect control efficiency. The control of dipterans with bioinsecticides based on Bacillus thuringiensis has been satisfactory, due to the production of insecticidal proteins denominated Cry (crystal), Cyt (cytolytic) toxins and Chi (chitinase), and to the synergistic effects among them. The present work aimed to select B. thuringiensis isolates efficient against A. aegypti larvae. A bacterial collection containing 1,073 isolates of B. thuringiensis, obtained from different locations of Brazilian territory, had the DNA isolated and submitted to PCR amplifications using specific primers for cry4Aa, cry4Ba, cry11Aa, cry11Ba, cyt1Aa, cyt1Ab, cyt2Aa and chi genes. For the LC50 and LC90 determination, the entomopathogenic isolates were evaluated by selective and quantitative bioassays. Only 45 isolates (4.2%) presented amplicons for the cry and cyt genes. The chi gene sequence was detected in 25 (54.3%) of those isolates. From the 45 isolates submitted to the selective bioassays, 13 caused 100% mortality of A. aegypti larvae. The identification of cry, cyt and chi genes of B. thuringiensis and the toxicity analysis on A. aegypti led to the selection of a set of isolates that have the potential to be used in the formulation of new bioinsecticides. Unesp Faculdade de Ciências Agrárias e Veterinárias Depto de Ciências Exatas Unesp Faculdade de Ciências Agrárias e Veterinárias Depto de Ciências Exatas

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:41:30Z No. of bitstreams: 1 S1517-83822009000400035.pdf: 137697 bytes, checksum: ac34dae451301dba8c4229402a3a02bb (MD5) Made available in DSpace on 2013-08-22T18:41:30Z (GMT). No. of bitstreams: 1 S1517-83822009000400035.pdf: 137697 bytes, checksum: ac34dae451301dba8c4229402a3a02bb (MD5) Previous issue date: 2009-12-01 Made available in DSpace on 2013-09-30T19:28:02Z (GMT). No. of bitstreams: 2 S1517-83822009000400035.pdf: 137697 bytes, checksum: ac34dae451301dba8c4229402a3a02bb (MD5) S1517-83822009000400035.pdf.txt: 30693 bytes, checksum: 16737efc25c24167d000935e52514418 (MD5) Previous issue date: 2009-12-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:15:54Z No. of bitstreams: 2 S1517-83822009000400035.pdf: 137697 bytes, checksum: ac34dae451301dba8c4229402a3a02bb (MD5) S1517-83822009000400035.pdf.txt: 30693 bytes, checksum: 16737efc25c24167d000935e52514418 (MD5) Made available in DSpace on 2014-05-20T13:15:54Z (GMT). No. of bitstreams: 2 S1517-83822009000400035.pdf: 137697 bytes, checksum: ac34dae451301dba8c4229402a3a02bb (MD5) S1517-83822009000400035.pdf.txt: 30693 bytes, checksum: 16737efc25c24167d000935e52514418 (MD5) Previous issue date: 2009-12-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Salmonella enterica serovar Gallinarum (SG) is an intracellular pathogen of chickens. To survive, to invade and to multiply in the intestinal tract and intracellularly it depends on its ability to produce energy in anaerobic conditions. The fumarate reductase (frdABCD), dimethyl sulfoxide (DMSO)-trimethylamine N-oxide (TMAO) reductase (dmsABC), and nitrate reductase (narGHIJ) operons in Salmonella Typhimurium (STM) encode enzymes involved in anaerobic respiration to the electron acceptors fumarate, DMSO, TMAO, and nitrate, respectively. They are regulated in response to nitrate and oxygen availability and changes in cell growth rate. In this study mortality rates of chickens challenged with mutants of Salmonella Gallinarum, which were defective in utilising anaerobic electron acceptors, were assessed in comparison to group of bird challenged with wild strain. The greatest degree of attenuation was observed with mutations affecting nitrate reductase (napA, narG) with additional attenuations induced by a mutation affecting fumarate reductase (frdA) and a double mutant (dmsA torC) affecting DMSO and TMAO reductase. Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias University of Nottingham School of Veterinary Medicine and Science Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias

S. Pullorum (SP) and S. Gallinarum (SG) are very similar. They are the agents of pullorum disease and fowl typhoid, respectively, and the two diseases are responsible for economic losses in poultry production. Although SP and SG are difficult to be differentiated in routine laboratory procedures, the ability to metabolize ornithine is a biochemical test that may be used to achieve this aim. While SP is able to decarboxylate this amino acid, SG is not. However, the isolation of strains showing atypical biochemical behavior has made this differentiation difficult. One of the genes associated with the metabolization of the amino acid ornithine is called speC, and is found in both serovars. The analysis of 21 SP and 15 SG strains by means of PCR did not enable the differentiation of the two serovars, because fragments produced were identical. However, after enzymatic treatment with restriction enzyme Eco RI, the band pattern of each serovar showed to be different, even in samples of atypical biochemical behavior. This fact enabled the standardization of the technique for a quick and safe differentiation of serovars Pullorum and Gallinarum.A S. Pullorum (SP) é muito semelhante à S. Gallinarum (SG), agentes da Pulorose e Tifo aviário, respectivamente, sendo que as duas enfermidades são responsáveis por perdas econômicas no setor avícola. SP e SG são de difícil diferenciação em procedimento laboratorial rotineiro, mas uma prova bioquímica muito utilizada na distinção das duas refere-se à capacidade de assimilar o aminoácido ornitina: SP descarboxila este aminoácido enquanto SG não. No entanto, o isolamento de cepas com comportamento bioquímico atípico, tem dificultado tal diferenciação. Um dos genes relacionados à assimilação do aminoácido ornitina, denomina-se gene speC, o qual está presente nos dois sorovares. Analisando 21 amostras de SP e 15 de SG com a utilização da PCR não foi possível realizar a diferenciação dos dois sorovares pois os fragmentos gerados eram idênticos. Posteriormente, com o uso da técnica de tratamento enzimático com a enzima de restrição Eco RI, foi possível observar que o padrão de bandas gerado em cada sorovar era diferente, mesmo quando amostras que apresentavam comportamento bioquímico atípico eram analisadas. Tal fato permitiu a padronização da técnica para ser utilizada na diferenciação entre os sorovares Pullorum e Gallinarum de maneira rápida e segura.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:41:28Z No. of bitstreams: 1 S1517-83822009000100032.pdf: 9699855 bytes, checksum: 8562026dbcb6bf6ffbad27c8f7fda26a (MD5) Made available in DSpace on 2013-08-22T18:41:28Z (GMT). No. of bitstreams: 1 S1517-83822009000100032.pdf: 9699855 bytes, checksum: 8562026dbcb6bf6ffbad27c8f7fda26a (MD5) Previous issue date: 2009-03-01 Made available in DSpace on 2013-09-30T19:28:00Z (GMT). No. of bitstreams: 2 S1517-83822009000100032.pdf: 9699855 bytes, checksum: 8562026dbcb6bf6ffbad27c8f7fda26a (MD5) S1517-83822009000100032.pdf.txt: 21414 bytes, checksum: cd705aca3ea790cf1142057157900558 (MD5) Previous issue date: 2009-03-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:15:54Z No. of bitstreams: 2 S1517-83822009000100032.pdf: 9699855 bytes, checksum: 8562026dbcb6bf6ffbad27c8f7fda26a (MD5) S1517-83822009000100032.pdf.txt: 21414 bytes, checksum: cd705aca3ea790cf1142057157900558 (MD5) Made available in DSpace on 2014-05-20T13:15:54Z (GMT). No. of bitstreams: 2 S1517-83822009000100032.pdf: 9699855 bytes, checksum: 8562026dbcb6bf6ffbad27c8f7fda26a (MD5) S1517-83822009000100032.pdf.txt: 21414 bytes, checksum: cd705aca3ea790cf1142057157900558 (MD5) Previous issue date: 2009-03-01 FCAV-UNESP A S. Pullorum (SP) é muito semelhante à S. Gallinarum (SG), agentes da Pulorose e Tifo aviário, respectivamente, sendo que as duas enfermidades são responsáveis por perdas econômicas no setor avícola. SP e SG são de difícil diferenciação em procedimento laboratorial rotineiro, mas uma prova bioquímica muito utilizada na distinção das duas refere-se à capacidade de assimilar o aminoácido ornitina: SP descarboxila este aminoácido enquanto SG não. No entanto, o isolamento de cepas com comportamento bioquímico atípico, tem dificultado tal diferenciação. Um dos genes relacionados à assimilação do aminoácido ornitina, denomina-se gene speC, o qual está presente nos dois sorovares. Analisando 21 amostras de SP e 15 de SG com a utilização da PCR não foi possível realizar a diferenciação dos dois sorovares pois os fragmentos gerados eram idênticos. Posteriormente, com o uso da técnica de tratamento enzimático com a enzima de restrição Eco RI, foi possível observar que o padrão de bandas gerado em cada sorovar era diferente, mesmo quando amostras que apresentavam comportamento bioquímico atípico eram analisadas. Tal fato permitiu a padronização da técnica para ser utilizada na diferenciação entre os sorovares Pullorum e Gallinarum de maneira rápida e segura. S. Pullorum (SP) and S. Gallinarum (SG) are very similar. They are the agents of pullorum disease and fowl typhoid, respectively, and the two diseases are responsible for economic losses in poultry production. Although SP and SG are difficult to be differentiated in routine laboratory procedures, the ability to metabolize ornithine is a biochemical test that may be used to achieve this aim. While SP is able to decarboxylate this amino acid, SG is not. However, the isolation of strains showing atypical biochemical behavior has made this differentiation difficult. One of the genes associated with the metabolization of the amino acid ornithine is called speC, and is found in both serovars. The analysis of 21 SP and 15 SG strains by means of PCR did not enable the differentiation of the two serovars, because fragments produced were identical. However, after enzymatic treatment with restriction enzyme Eco RI, the band pattern of each serovar showed to be different, even in samples of atypical biochemical behavior. This fact enabled the standardization of the technique for a quick and safe differentiation of serovars Pullorum and Gallinarum. Laboratório Nacional Agropecuário Ministério da Agricultura, Pecuária e Abastecimento Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista Faculdade de Ciências Agrárias e Veterinárias

A bactéria B. thuringiensis caracteriza-se pela produção de proteínas tóxicas a representantes de diversas ordens de insetos, as quais são codificadas por genes cry. Este trabalho foi realizado com objetivo de selecionar isolados de B. thuringiensis, por meio da caracterização morfológica e molecular, identificando as diferentes subclasses dos genes cry3 e cry35 e determinar a patogenicidade contra Sphenophorus levis, uma das mais importantes pragas da cultura da cana-de-açúcar. Foram utilizados 1163 isolados de B. thuringiensis e com a observação em microscópio com contraste de fases foram confirmadas como pertencentes à espécie de B. thuringiensis. O material genético foi purificado pela matriz de troca iônica "Instagene Matrix" e submetido a PCR com iniciadores gerais cry3 e cry35 identificando-se 30 isolados contendo genes com potencial para o controle de coleópteros, os quais juntamente com as linhagens-padrão de B. thuringiensis var. tenebrionis, B. thuringiensis var. morrissone e B. thuringiensis var. tolworthi foram utilizados para a realização do bioensaio. Através de análise discriminante alocaram-se os isolados em quatro grupos quanto à toxicidade de B. thuringiensis. Os grupos ficaram assim definidos: um grupo que promovem até 10% de mortalidade contendo as testemunhas e duas linhagens;um grupo que causou 39% de mortalidade contendo três linhagens padrão e dez isolados; um grupo com 52% de mortalidade contendo treze isolados e um grupo com 70% de mortalidade contendo cinco isolados, os quais devem ser considerados promissores no controle biológico de S. levis.Bacillus thuringiensis is well known for its ability to produce toxic proteins to different insect orders which are encoded by the cry genes. This work was carried out aiming to select among different B. thuringiensis isolates, by morphological and molecular characterization, identifying different classes of cry3 and cry35 genes, and to determine the level of pathogenicity against Sphenophorus levis larvae, which is one of the most important sugar-cane pests in Brazil. A total of 1163 bacterial isolates were used in this survey in which phase microscopy and molecular markers were used for evaluation that resulted in 30 isolates positive for cry3 and cry35 genes potentially active against coleopterans. These bacterial isolates together with type strains such as B. thuringiensis var. tenebrionis, B.t. var. morrissone e B.t. var. tolworthi were used in bioassays using Sphenophorus levis larvae. Using discriminating analysis four groups of isolates were produced when B. thuringiensis toxicity was taken into account. These groups involved one set of two isolates with up to 10% mortality; another set caused mortality of up to 39% and was made up of ten isolates and three type strains; another set caused mortality of up to 52% containing thirteen isolates and a last set of isolated whose mortality index was 70% containing only five isolates which were considered promising bacterial strains for biological control of S. levis.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T19:04:58Z No. of bitstreams: 1 S1415-47572007000500009.pdf: 215113 bytes, checksum: 7546a4ee8e2759a550d4c629c9bfe6a1 (MD5) Made available in DSpace on 2013-08-22T19:04:58Z (GMT). No. of bitstreams: 1 S1415-47572007000500009.pdf: 215113 bytes, checksum: 7546a4ee8e2759a550d4c629c9bfe6a1 (MD5) Previous issue date: 2007-01-01 Made available in DSpace on 2013-09-30T20:08:33Z (GMT). No. of bitstreams: 2 S1415-47572007000500009.pdf: 215113 bytes, checksum: 7546a4ee8e2759a550d4c629c9bfe6a1 (MD5) S1415-47572007000500009.pdf.txt: 48155 bytes, checksum: 2bcfcbb61e579b465dbf959e887f4c00 (MD5) Previous issue date: 2007-01-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T15:17:30Z No. of bitstreams: 2 S1415-47572007000500009.pdf: 215113 bytes, checksum: 7546a4ee8e2759a550d4c629c9bfe6a1 (MD5) S1415-47572007000500009.pdf.txt: 48155 bytes, checksum: 2bcfcbb61e579b465dbf959e887f4c00 (MD5) Made available in DSpace on 2014-05-20T15:17:30Z (GMT). No. of bitstreams: 2 S1415-47572007000500009.pdf: 215113 bytes, checksum: 7546a4ee8e2759a550d4c629c9bfe6a1 (MD5) S1415-47572007000500009.pdf.txt: 48155 bytes, checksum: 2bcfcbb61e579b465dbf959e887f4c00 (MD5) Previous issue date: 2007-01-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Assimilation of nitrate and ammonium are vital procedures for plant development and growth. From these primary paths of inorganic nitrogen assimilation, this metabolism integrates diverse paths for biosynthesis of macromolecules, such as amino acids and nucleotides, and the central intermediate metabolism, like carbon metabolism and photorespiration. This paper reports research performed in the CitEST (Citrus Expressed Sequence Tag) database for the main genes involved in nitrogen metabolism and those previously described in other organisms. The results show that a complete cluster of genes involved in the assimilation of nitrogen and the metabolisms of glutamine, glutamate, aspartate and asparagine can be found in the CitEST data. The main enzymes found were nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS), glutamate synthetase (GOGAT), glutamate dehydrogenase (GDH), aspartate aminotransferase (AspAT) and asparagine synthetase (AS). The different enzymes involved in this metabolism have been shown to be highly conserved among the Citrus and Poncirus species. This work serves as a guide for future functional analysis of these enzymes in citrus. Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Fitossanidade Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Tecnologia Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Biologia Aplicada a Agropecuária Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Fitossanidade Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Tecnologia Universidade Estadual Paulista Faculdade de Ciências Veterinárias Departamento de Biologia Aplicada a Agropecuária

Resumo:O objetivo deste trabalho foi avaliar a toxicidade de novas proteínas Vip3Aa e sua capacidade de ligação a vesículas de membrana da microvilosidade apical (VMMA) do intestino de lagartas neonatas de Spodoptera frugiperda, Anticarsia gemmatalise Heliothis virescens. Proteínas expressas pelos genes vip3Aa42 e vip3Aa43 mostraram-se tóxicas a S. frugiperda (CL50 de 78,2 e 113 ng cm-2, respectivamente) e A. gemmatalis(CL50 de 239,2 e 57,5 ng cm-2, respectivamente), e pouco tóxicas a H. virescens (CL50>5.000 ng cm-2). Os ensaios de ligação às VMMA mostraram que as proteínas unem-se de forma efetiva aos receptores nas vesículas das espécies avaliadas, mas essa capacidade de ligação somente é efetiva na ativação da toxicidade para as populações avaliadas de S. frugiperdae A. gemmatalis. Abstract:The objective of this work was to evaluate the toxicity of new Vip3Aa proteins and their binding capacity to brush-border membrane vesicles (BBMV) in the intestine of Spodoptera frugiperda, Anticarsia gemmatalis, and Heliothis virescens neonate larvae. The proteins expressed by the genes vip3Aa42 and vip3Aa43 showed toxicity to S. frugiperda (LC50 of 78.2 and 113 ng cm-2, respectively) and A. gemmatalis(LC50 of 239.2 and 57.5 ng cm-2, respectively), but they showed low toxicity to H. virescens(LC50>5,000 ng cm-2). BBMV binding assays showed that the proteins bind effectively to the receptors on vesicles of the evaluated species, but this binding capacity is only effective on the activation of toxicity to the evaluated populations of S. frugiperdaand A. gemmatalis.

The bacterium Bacillus thuringiensis (Berliner) is an important entomopathogen that can replace chemical pesticides in crops. In addition to the endotoxins found in B. thuringiensis , new proteins have been studied recently, such as Vip and Chitinase, which can enhance toxic effects by acting synergistically. Thus, the objectives of this work were to select new B. thuringiensis isolates carrying cry1Fa, vip3Aa , and chi ; to verify polymorphisms based on PCR-RFLP analysis; and to evaluate mortality and existence of synergistic effects of these genes on Spodoptera frugiperda larvae. A total of 114 isolates were subjected to screening via PCR to detect the vip3Aa , cry1Fa , and chi genes . Neonatal larvae of S. frugiperda were bioassayed using isolates containing one, two or all three genes and were compared to the standard strains . The PCR-RFLP analysis showed the absence of polymorphisms in these gene regions. The B. thuringiensis isolates from different regions of Brazil contained different combinations of the cry1Fa, vip3Aa , and chi genes. The presence of vip3Aa appeared to act synergistically with chi , and the combination of all three genes caused increased mortality in S. frugiperda larvae. Due to their insecticidal potential, these isolates are a prospective source of genes that can be used to genetically modify crops to increase the efficacy of pest control.

The genome of the bacterium Xylella fastidiosa contains four ORFs (XF2721, XF2725, XF2739 and XF0295) related to the restriction modification type I system, ordinarily named R–M. This system belongs to the DNA immigration control region (ICR). Each ORF is related to different operon structures, which are homologues among themselves and with subunit Hsd R from the endonuclease coding genes. In addition, these ORFs are highly homologous to genes in Pseudomonas aeruginosa, Methylococcus capsulatus str. Bath, Legionella pneumophila , Helicobacter pylori , Xanthomonas oryzae pv. Oryzae and Silicibacter pomeroyi , as well as to genes from X. fastidiosa strains that infect grapevine, almond and oleander plants. This study was carried out on R–M ORFs from forty-three X. fastidiosa strains isolated from citrus, coffee, grapevine, periwinkle, almond and plum trees, in order to assess the genetic diversity of these loci through PCR-RFLP. PCR-RFLP analysis of the four ORFs related to the R–M system from these strains enabled the detection of haplotypes for these loci. When the haplotypes were defined, wide genetic diversity and a large range of similar strains originating from different hosts were observed. This analysis also provided information indicating differences in population genetic structures, which led to detection of different levels of gene transfer among the groups of strains.

Over 40,000 sugarcane (Saccharum officinarum) consensus sequences assembled from 237,954 expressed sequence tags were compared with the protein and DNA sequences from other angiosperms, including the genomes of Arabidopsis and rice (Oryza sativa). Approximately two-thirds of the sugarcane transcriptome have similar sequences in Arabidopsis. These sequences may represent a core set of proteins or protein domains that are conserved among monocots and eudicots and probably encode for essential angiosperm functions. The remaining sequences represent putative monocot-specific genetic material, one-half of which were found only in sugarcane. These monocot-specific cDNAs represent either novelties or, in many cases, fast-evolving sequences that diverged substantially from their eudicot homologs. The wide comparative genome analysis presented here provides information on the evolutionary changes that underlie the divergence of monocots and eudicots. Our comparative analysis also led to the identification of several not yet annotated putative genes and possible gene loss events in Arabidopsis.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2014-02-26T17:15:29Z No. of bitstreams: 1 WOS000180834300035.pdf: 1288581 bytes, checksum: c80b0ab1625eeba1cd15e9da6e465b26 (MD5) Made available in DSpace on 2014-02-26T17:15:29Z (GMT). No. of bitstreams: 1 WOS000180834300035.pdf: 1288581 bytes, checksum: c80b0ab1625eeba1cd15e9da6e465b26 (MD5) Previous issue date: 2003-02-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T15:25:38Z No. of bitstreams: 1 WOS000180834300035.pdf: 1288581 bytes, checksum: c80b0ab1625eeba1cd15e9da6e465b26 (MD5) Made available in DSpace on 2014-05-20T15:25:38Z (GMT). No. of bitstreams: 1 WOS000180834300035.pdf: 1288581 bytes, checksum: c80b0ab1625eeba1cd15e9da6e465b26 (MD5) Previous issue date: 2003-02-01 Xylella fastidiosa is a xylem-dwelling, insect-transmitted, gamma-proteobacterium that causes diseases in many plants, including grapevine, citrus, periwinkle, almond, oleander, and coffee. X. fastidiosa has an unusually broad host range, has an extensive geographical distribution throughout the American continent, and induces diverse disease phenotypes. Previous molecular analyses indicated three distinct groups of X.fastidiosa isolates that were expected to be genetically divergent. Here we report the genome sequence of X. fastidiosa (Temecula strain), isolated from a naturally infected grapevine with Pierce's disease (PD) in a wine-grape-growing region of California. Comparative analyses with a previously sequenced X.fastidiosa strain responsible for citrus variegated chlorosis (CVC) revealed that 98% of the PD X.fastidiosa Temecula genes are shared with the CVC X. fastidiosa strain 9a5c genes. Furthermore, the average amino acid identity of the open reading frames in the strains is 95.7%. Genomic differences are limited to phage-associated chromosomal rearrangements and deletions that also account for the strain-specific genes present in each genome. Genomic islands, one in each genome, were identified, and their presence in other X.fastidiosa strains was analyzed. We conclude that these two organisms have identical metabolic functions and are likely to use a common set of genes in plant colonization and pathogenesis, permitting convergence of functional genomic strategies. Univ São Paulo, Inst Biociencias, Dept Bot, BR-05508900 São Paulo, Brazil Univ São Paulo, Inst Quim, BR-05508900 São Paulo, Brazil Univ São Paulo, Inst Ciências Biomed, BR-05508900 São Paulo, Brazil Univ São Paulo, Escola Super Agr Luiz de Queiroz, BR-13418900 Piracicaba, Brazil Univ São Paulo, Ctr Energia Nucl Agr, BR-13400970 Piracicaba, Brazil Inst Agron Campinas, Ctr Citricultura Sylio Moreira, BR-13490970 Cordeiropolis, Brazil Univ Estadual Paulista, Fac Ciências Agr & Vet, BR-14884900 Jaboticabal, Brazil Univ Estadual Campinas, Ctr Biol Mol & Engn Genet, BR-13083970 Campinas, SP, Brazil Univ Estadual Campinas, Inst Biol, BR-13083970 Campinas, SP, Brazil Univ Estadual Campinas, Inst Comp, BR-13083970 Campinas, SP, Brazil Univ São Paulo, Fac Ciências Farmaceut, BR-14040901 Ribeirao Preto, Brazil Univ São Paulo, Fac Filosofia Ciências & Letras, BR-14040901 Ribeirao Preto, Brazil Inst Ludwig Pesquisa Canc, BR-01509010 São Paulo, Brazil Univ Mogi das Cruzes, Nucleo Integrado Biotecnol, BR-08780911 Mogi das Cruzes, Brazil Inst Agron Campinas, BR-13001970 Campinas, SP, Brazil Inst Biol, BR-04014002 São Paulo, Brazil Univ Estadual Paulista, Fac Med Vet & Zootecnia, BR-18603970 Botucatu, SP, Brazil Univ Estadual Paulista, Fac Ciências Agron, BR-18603970 Botucatu, SP, Brazil Univ Estadual Paulista, Inst Biociencias, BR-18618000 Botucatu, SP, Brazil Univ Fed Sao Carlos, BR-13600970 São Paulo, Brazil Univ Fed Mato Grosso do Sul, Dept Comp & Estatist, BR-79070900 Campo Grande, MS, Brazil USDA ARS, Parlier, CA 93648 USA Univ Estadual Paulista, Fac Ciências Agr & Vet, BR-14884900 Jaboticabal, Brazil Univ Estadual Paulista, Fac Med Vet & Zootecnia, BR-18603970 Botucatu, SP, Brazil Univ Estadual Paulista, Fac Ciências Agron, BR-18603970 Botucatu, SP, Brazil Univ Estadual Paulista, Inst Biociencias, BR-18618000 Botucatu, SP, Brazil

Transposons are mobile genetic elements found within the genomes of various organisms including bacteria, fungi, plants and animals. Fragments of the transposon Tn1721 were found included in the genome of Xylella fastidiosa strain 9a5c. Regions from such fragments were PCR-amplified using specially designed primers (TNP(1) and TNP(2)). In order to detect insertions of the Tn1721 element, both primers were used and one of them included a region of the transposon (TNP(1)) and the other one had the right repeat and part of the bacterial chromosome (TNP(2)). The PCR products obtained from strain 9a5c were used as a pattern for fragment size comparisons when DNA samples from other X. fastidiosa strains were used as template for the PCR assays. Differences were observed concerning the PCR products of such amplifications when some X. fastidiosa strains isolated from grapevine and plum were used. For the citrus-derived strains only the strains U187d and GP920b produced fragments with different sizes or weak band intensity. Such variations in the X. fastidiosa genome related to disrupted Tn1721 copies are probably due to the possibility of such a transposon element being still able to duplicate even after deletion events might have taken place and also because the bacterial strains in which the main differences were detected are derived from different host plants cultivated under different climate conditions from the one used as reference.

Sugarcane borer (Diatraea saccharalis, F.) is an important corn pest in South America and United States. The aim of the present study was to analyze the susceptibility and binding interactions of three Cry1A proteins and Cry1Fa in a Brazilian D. saccharalis population. The results showed that Cry1Ab was the most active, followed by Cry1Ac, Cry1Fa and Cry1Aa. All Cry1-biotinylated proteins tested bound specifically to the D. saccharalis brush border membrane vesicles (BBMV). Heterologous competition assays showed shared binding sites for all Cry1A proteins and another one shared by Cry1Fa and Cry1Ab. Thus, pyramiding Cry1Aa/Cry1Ac and Cry1F proteins would be a recommended strategy for managing this pest.

Made available in DSpace on 2015-03-18T15:53:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-10-13Bitstream added on 2015-03-18T16:29:16Z : No. of bitstreams: 1 WOS000340302100021.pdf: 740482 bytes, checksum: 73a2f4d26857210429372a925338e4b5 (MD5) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Rhizobium tropici, a member of the Rhizobiaceae family, has the ability to synthesize and secrete extracellular polysaccharides (EPS). Rhizobial EPS have attracted much attention from the scientific and industrial communities. Rhizobial isolates and R. tropici mutants that produced higher levels of EPS than the wildtype strain SEMIA4080 were used in the present study. The results suggested a heteropolymer structure for these EPS composed by glucose and galactose as prevailing monomer unit. All EPS samples exhibited a typical non-Newtonian and pseudoplastic fluid flow, and the aqueous solutions apparent viscosities increased in a concentration-dependent manner. These results serve as a foundation for further studies aimed at enhancing interest in the application of the MUTZC3, JAB1 and JAB6 strains with high EPS production and viscosity can be exploited for the large-scale commercial production of Rhizobial polysaccharides. (C) 2014 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/). UNESP Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Tecnol, BR-14884900 Jaboticabal, SP, Brazil UNESP Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada & Agr, BR-14884900 Jaboticabal, SP, Brazil UNESP Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Tecnol, BR-14884900 Jaboticabal, SP, Brazil UNESP Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada & Agr, BR-14884900 Jaboticabal, SP, Brazil FAPESP: 07/57586-6

Instituto Ludwig de Pesquisa sobre o Câncer, Rua Prof. Antonio Prudente, 109-4 andar, 01509-010, Sao Paulo-SP

The plasmid pHT409 that harbours the cryIA(a) gene for the production of a δ-endotoxin (crystal protein) from Bacillus thuringiensis was transferred into Bradyrhizobium sp. A conjugal transfer system aiming to introduce the plasmid into the Bradyrhizobium sp. host from colonies of an Escherichia coli donor strain (DH5α::pHT409) has been developed. As a result exconjugants were obtained in which the transferred plasmid has been detected by both microbiological and electrophoresis techniques. The cryIA(a) gene when inside the new host had a low expression level which was detected by immunoblotting.

The giant anteater (Myrmecophaga tridactyla) is found from Belize and Guatemala to Paraguay and Argentina. Its conservation status is considered vulnerable by IUCN. Here we report the isolation and characterization of six microsatellite loci. Positive loci for (GT)(n) were isolated using a magnetic bead hybridization selection protocol. The number of alleles per locus as well as the heterozygosity and PCR conditions are described. These loci will be useful for studying population structure, genetic diversity, and paternity in M. tridactyla wild populations.

Bacillus thuringiensis var. thuringiensis strain T01-328, isolated from Cubatão county (São Paulo State, Brazil), produces a soluble pesticide protein, Cry1Ia, during vegetative growth. Here, we report the 7.089-Mbp draft genome sequence, composed of a 5.5-Mb chromosome and 14 plasmids, which is the largest B. thuringiensis genome sequenced to date.

Made available in DSpace on 2013-08-12T18:25:23Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-05-15 Made available in DSpace on 2013-09-30T17:49:11Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-05-15 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T13:13:13Z No. of bitstreams: 0 Made available in DSpace on 2014-05-20T13:13:13Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-05-15 Ministry of Culture of Uruguay through the Program for Technical Development (PDT) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) National Institute of Agronomic Research of Uruguay (INIA) Xylella fastidiosa inhabits the plant xylem, a nutrient-poor environment, so that mechanisms to sense and respond to adverse environmental conditions are extremely important for bacterial survival in the plant host. Although the complete genome sequences of different Xylella strains have been determined, little is known about stress responses and gene regulation in these organisms. In this work, a DNA microarray was constructed containing 2,600 ORFs identified in the genome sequencing project of Xylella fastidiosa 9a5c strain, and used to check global gene expression differences in the bacteria when it is infecting a symptomatic and a tolerant citrus tree. Different patterns of expression were found in each variety, suggesting that bacteria are responding differentially according to each plant xylem environment. The global gene expression profile was determined and several genes related to bacterial survival in stressed conditions were found to be differentially expressed between varieties, suggesting the involvement of different strategies for adaptation to the environment. The expression pattern of some genes related to the heat shock response, toxin and detoxification processes, adaptation to atypical conditions, repair systems as well as some regulatory genes are discussed in this paper. DNA microarray proved to be a powerful technique for global transcriptome analyses. This is one of the first studies of Xylella fastidiosa gene expression in vivo which helped to increase insight into stress responses and possible bacterial survival mechanisms in the nutrient-poor environment of xylem vessels. INIA, Estn Expt Wilson Ferreira Aldunate, Unidad Biotecnol, Rincon Del Colorado, Uruguay Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Tecnol, Lab Bioquim Microrganismos & Plantas, Jaboticabal, Brazil Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA) Mandioca & Fruticultura, Nucl Tecnol Recursos Gen & Desenvolvimento Varien, Cruz Das Almas, BA, Brazil Estacao Expt Citricultura, São Paulo, Brazil Univ São Paulo, Escola Super Agr Luiz de Queiroz, São Paulo, Brazil Univ Fed Vales Jequitinhonha & Mucuri, Fac Ciencias Vet, Dept Engn Florestal, Diamantina, MG, Brazil Ctr Citricultura Sylvio Moreira APTA, Cordeiropolis, SP, Brazil Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada Agropecuaria, Jaboticabal, Brazil Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Tecnol, Lab Bioquim Microrganismos & Plantas, Jaboticabal, Brazil Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada Agropecuaria, Jaboticabal, Brazil