Behavior and development of Tetranychus ludeni Zacher, 1913 (Acari: Tetranychidae) and physiological stress in genetically modified cotton expressing Cry1F and Cry1Ac proteins.

Genetically modified plants are one of the tactics used in integrated pest management - IPM. There is great concern about the impact of these plants on non-target organisms. On the other hand, there is little information in the literature on the effects of transgenics (Bacillus thuringiensis) Bt on populations of phytophagous mites, and the physiological responses that this attack promotes on plants. The objective of this work was to evaluate the biology of the T. ludeni mite in Bt cotton, expressing the Cry1F and Cry1Ac proteins. To evaluate the behavior of food and oviposition preference of the T. ludeni with Bt cotton and isohybrid. Verify if the physiological stress caused by T. ludeni's attack is differentiated in Bt cotton. The mites were reared in Bt cotton and isohybrid, in a total of 40 replicates in the completely randomized design and the biological cycle was evaluated. The food preference and oviposition analysis were done with 10 replicates, with choice. The physiological stress was evaluated through chlorophyll fluorescence, under greenhouse conditions. The data of the T. ludeni biology were analyzed by Student's t-test, for food and oviposition preference the chi-square test was performed. Regression models were fitted for the fluorescence parameters. The model identity test was used to evaluate the differences between Bt and isohybrid treatments. Cry1F and Cry1Ac proteins have not affected the biology of T. ludeni. The photosynthetic parameters in Bt cotton plants were less influenced by T. ludeni infestation.

Behavior and development of Tetranychus ludeni Zacher, 1913 (Acari: Tetranychidae) and physiological stress in genetically modified cotton expressing Cry1F and Cry1Ac proteins / Comportamento e desenvolvimento de Tetranychus ludeni Zacher, 1913 (Acari: Tetranychidae) e estresse fisiológico em algodoeiro geneticamente modificado expressando as proteínas Cry1F e Cry1Ac

Genetically modified plants are one of the tactics used in integrated pest management - IPM. There is great concern about the impact of these plants on non-target organisms. On the other hand, there is little information in the literature on the effects of transgenics (Bacillus thuringiensis) Bt on populations of phytophagous mites, and the physiological responses that this attack promotes on plants. The objective of this work was to evaluate the biology of the T. ludeni mite in Bt cotton, expressing the Cry1F and Cry1Ac proteins. To evaluate the behavior of food and oviposition preference of the T. ludeni with Bt cotton and isohybrid. Verify if the physiological stress caused by T. ludeni's attack is differentiated in Bt cotton. The mites were reared in Bt cotton and isohybrid, in a total of 40 replicates in the completely randomized design and the biological cycle was evaluated. The food preference and oviposition analysis were done with 10 replicates, with choice. The physiological stress was evaluated through chlorophyll fluorescence, under greenhouse conditions. The data of the T. ludeni biology were analyzed by Student's t-test, for food and oviposition preference the chi-square test was performed. Regression models were fitted for the fluorescence parameters. The model identity test was used to evaluate the differences between Bt and isohybrid treatments. Cry1F and Cry1Ac proteins have not affected the biology of T. ludeni. The photosynthetic parameters in Bt cotton plants were less influenced by T. ludeni infestation.

O uso de plantas geneticamente modificadas é uma das táticas utilizadas no manejo integrado de pragas - MIP. Observa-se grande preocupação com o impacto dessas plantas sobre organismos não alvos. Por outro lado, existe pouca informação na literatura sobre efeitos dos transgênicos (Bacillus thuringiensis) Bt em populações de ácaros fitófagos, e as respostas fisiológicas que esse ataque promove nas plantas. Objetivou-se com esse trabalho avaliar a biologia do ácaro T. ludeni em algodoeiro Bt, expressando as proteínas Cry1F e Cry1Ac. Avaliar se há comportamento de preferência alimentar e postura de T. ludeni em relação ao algodoeiro Bt e seu iso-híbrido. E verificar se o estresse fisiológico causado pelo ataque de T. ludeni é diferenciado em algodoeiro Bt. Os ácaros foram criados em algodoeiro Bt e iso-híbrido, em um total de 40 repetições no delineamento inteiramente casualizado, onde foi avaliado o ciclo biológico. A análise de preferência alimentar e de posturas foi feita com 10 repetições, com escolha. O estresse fisiológico foi avaliando através da fluorescência da clorofila, em casa de vegetação. Os dados da biologia de T. ludeni foram analisados pelo teste t Student, para preferência alimentar e postura foi realizado o teste qui-quadrado. Para os parâmetros da fluorescência, foram ajustados modelos de regressão. Para testar as diferenças entre Bt e iso-híbrido foi utilizado o teste de identidade de modelos. As proteínas Cry1F e Cry1Ac não afetaram a biologia de T. ludeni. Os parâmetros fotossintéticos em plantas de algodoeiro Bt foram menos influenciados pela infestação de T. ludeni.

The beetle Costalimaita ferruginea (Coleoptera: Chysomelidae) in Eucalyptus plantations in transition area of Amazon and Cerrado Biomes / O besouro Costalimaita ferruginea (Coleoptera: Chysomelidae) em plantios de Eucalyptus em áreas de transição dos Biomas Amazon e Cerrado

Abstract Costalimaita ferruginea (Coleoptera: Chrysomelidae) attacks Eucalyptus plants causing severe damage through netting of the leaves. Recently, this Coleoptera has been reported attacking Myrtaceae in Mato Grosso State and, studies about the occurrence of this beetle in commercial plantations of eucalypts has been the subject of researchers through monitoring programmes in the forest protection area. With the beginning of the rainy season, adults were observed causing damage in eucalypt plantations in four cities that are part of the transition region of Amazon and Cerrado Biomes. The spots where these insects were observed are located in Feliz Natal, Lucas do Rio Verde, Sorriso and Vera. The purpose of this study was to report the new occurrences and to characterize the attack period of the beetle C. ferruginea in Eucalyptus plantations in Middle-North region of Mato Grosso State, region of Biomes Transition.

Resumo Costalimaita ferruginea (Coleoptera: Chrysomelidae) ataca plantas de Eucalyptus provocando intensos danos através do rendilhamento das folhas. Recentemente, esse Coleoptera tem sido relado atacando Myrtaceae no Estado de Mato Grosso e, estudos sobre a ocorrência desse besouro em plantios comerciais de eucaliptos tem sido alvo de pesquisadores através de programas de monitoramento na área de proteção florestal. Com o início do período chuvoso, adultos foram observados causando danos em plantios de eucalipto em quatro municípios que fazem parte da região de transição dos Biomas Amazônia e Cerrado. Os pontos onde foram observados esses insetos estão localizados em Feliz Natal, Lucas do Rio Verde, Sorriso e Vera. O objetivo deste trabalho foi relatar as novas ocorrências e caracterizar o período de ataque do besouro C. ferruginea em plantios de Eucalyptus na região Médio-Norte do Estado de Mato Grosso, região de transição de Biomas.

The beetle Costalimaita ferruginea (Coleoptera: Chysomelidae) in Eucalyptus plantations in transition area of Amazon and Cerrado Biomes.

Costalimaita ferruginea (Coleoptera: Chrysomelidae) attacks Eucalyptus plants causing severe damage through netting of the leaves. Recently, this Coleoptera has been reported attacking Myrtaceae in Mato Grosso State and, studies about the occurrence of this beetle in commercial plantations of eucalypts has been the subject of researchers through monitoring programmes in the forest protection area. With the beginning of the rainy season, adults were observed causing damage in eucalypt plantations in four cities that are part of the transition region of Amazon and Cerrado Biomes. The spots where these insects were observed are located in Feliz Natal, Lucas do Rio Verde, Sorriso and Vera. The purpose of this study was to report the new occurrences and to characterize the attack period of the beetle C. ferruginea in Eucalyptus plantations in Middle-North region of Mato Grosso State, region of Biomes Transition.

Reproductive ecology of phorid parasitoids in relation to the head size of leaf-cutting ants Atta sexdens Forel.

The leaf-cutting ant Atta sexdens Forel (Hymenoptera: Formicidae) is one of the most damaging agricultural pests in the Neotropics. Management strategies predominantly rely on the use of general insecticides. What is needed are more species-specific and environmentally friendly options. Parasitioids such as phorid flies (Diptera: Phoridae) may be one such option, but a greater understanding of the ecology of the flies and their ant hosts is essential to devise biological control strategies. Here we report parasitism rates, ant host size, parasitoid abundance per host and resultant sex ratios of two phorid species Apocephalus attophilus Borgmeier and Eibesfeldtphora tonhascai Brown parasitizing A.sexdens. The two species achieved parasitism rates of 1.48 and 1.46%, respectively and the pupal period was 14.7 ± 1.1 days and 22.1 ± 2.8 days, respectively. There was no significant difference between the head capsule width of ants parasitized by either A. attophilus or E. tonhascai. Likewise, there was no significant effect between the head capsule width of parasitized and unparasitized ants for both species. A significant positive correlation was found between the head capsule width of the parasitized ants and the number of adult parasitoids A. attophilus emerged. Ants parasitized by E. tonhascai survived significantly longer than those parasitized by A. attophilus. There was no significant effect of ant head width on the sex ratio of the offspring of either parasitoid species and no significant difference in the sex ratio (male: female) of their offspring. In summary, these data addressed here are important steps when considering natural enemies for biological control. Studying survival of the parasitized ants, parasitoid offspring sex ratio and host size preference allows for a better understanding of ant natural biological control in the field and can help in rearing of A. attophilus and E. tonhascai in laboratory.

Behavior and development of Tetranychus ludeni Zacher, 1913 (Acari: Tetranychidae) and physiological stress in genetically modified cotton expressing Cry1F and Cry1Ac proteins

Abstract Genetically modified plants are one of the tactics used in integrated pest management - IPM. There is great concern about the impact of these plants on non-target organisms. On the other hand, there is little information in the literature on the effects of transgenics (Bacillus thuringiensis) Bt on populations of phytophagous mites, and the physiological responses that this attack promotes on plants. The objective of this work was to evaluate the biology of the T. ludeni mite in Bt cotton, expressing the Cry1F and Cry1Ac proteins. To evaluate the behavior of food and oviposition preference of the T. ludeni with Bt cotton and isohybrid. Verify if the physiological stress caused by T. ludenis attack is differentiated in Bt cotton. The mites were reared in Bt cotton and isohybrid, in a total of 40 replicates in the completely randomized design and the biological cycle was evaluated. The food preference and oviposition analysis were done with 10 replicates, with choice. The physiological stress was evaluated through chlorophyll fluorescence, under greenhouse conditions. The data of the T. ludeni biology were analyzed by Student's t-test, for food and oviposition preference the chi-square test was performed. Regression models were fitted for the fluorescence parameters. The model identity test was used to evaluate the differences between Bt and isohybrid treatments. Cry1F and Cry1Ac proteins have not affected the biology of T. ludeni. The photosynthetic parameters in Bt cotton plants were less influenced by T. ludeni infestation.

Resumo O uso de plantas geneticamente modificadas é uma das táticas utilizadas no manejo integrado de pragas - MIP. Observa-se grande preocupação com o impacto dessas plantas sobre organismos não alvos. Por outro lado, existe pouca informação na literatura sobre efeitos dos transgênicos (Bacillus thuringiensis) Bt em populações de ácaros fitófagos, e as respostas fisiológicas que esse ataque promove nas plantas. Objetivou-se com esse trabalho avaliar a biologia do ácaro T. ludeni em algodoeiro Bt, expressando as proteínas Cry1F e Cry1Ac. Avaliar se há comportamento de preferência alimentar e postura de T. ludeni em relação ao algodoeiro Bt e seu iso-híbrido. E verificar se o estresse fisiológico causado pelo ataque de T. ludeni é diferenciado em algodoeiro Bt. Os ácaros foram criados em algodoeiro Bt e iso-híbrido, em um total de 40 repetições no delineamento inteiramente casualizado, onde foi avaliado o ciclo biológico. A análise de preferência alimentar e de posturas foi feita com 10 repetições, com escolha. O estresse fisiológico foi avaliando através da fluorescência da clorofila, em casa de vegetação. Os dados da biologia de T. ludeni foram analisados pelo teste t Student, para preferência alimentar e postura foi realizado o teste qui-quadrado. Para os parâmetros da fluorescência, foram ajustados modelos de regressão. Para testar as diferenças entre Bt e iso-híbrido foi utilizado o teste de identidade de modelos. As proteínas Cry1F e Cry1Ac não afetaram a biologia de T. ludeni. Os parâmetros fotossintéticos em plantas de algodoeiro Bt foram menos influenciados pela infestação de T. ludeni.

Walking stability of Rhyzopertha dominica (Fabricius, 1792) (Coleoptera: Bostrichidae) / Estabilidade de caminhamento de Rhyzopertha dominica (Fabricius, 1792) (Coleoptera: Bostrichidae)

Abstract Results obtained in studies can contribute to the advancement of science and innovative methods and techniques for developing practical activities. Reporting conditions that may restrict the implementation of research is critical to ensure the optimal development of further technical studies. The objective of this study was to assess the walking stability of R. dominica on a flat and smooth surface. The study was based on the determination of mortality, morphology and walking stability of the insect outside the grain mass, on a flat and smooth surface. Mortality of adults of this Coleoptera in conditions with and without food was similar, which explains the difficulty that this insect had for accessing the food source on the flat and smooth surface. The measurements of body length (BOL), width (BOW) and height (BOH) of R. dominica were compared with those of Tribolium castaneum (Coleoptera: Tenebrionidae), which showed good ability to walk in these conditions. This study indicated that the former presents lower BOL and BOW, and greater BOH than the second, and all these variables showed differences when analyzed simultaneously by means of the construction of multivariate morphometric indices (Width × Height, Length × Height and Height × Length × Width). These morphometric variables, together with the definition of the geometry most similar to the body shape, resulted in determination of the center of gravity (CG) and static rollover threshold (SRTgeom) for both species. Rhyzopertha dominica and T. castaneum presented CGs considered high and low, respectively, and together with the values obtained for SRTgeom, may justify that R. dominica can be considered a less stable species during movement, and presents greater risk of rollover on flat and smooth surfaces.

Resumo Resultados obtidos em pesquisas podem contribuir com o avanço da ciência e inovar métodos e técnicas para o desenvolvimento de atividades práticas. Relatar condições que podem restringir à execução de pesquisas é fundamental para garantir que seja observada a técnica ideal para o desenvolvimento de estudos posteriores. O objetivo deste trabalho foi estudar a estabilidade de caminhamento de R. dominica em superfície plana e polida. O estudo se baseou na determinação da mortalidade, morfologia e estabilidade de caminhamento do inseto fora da massa de grãos, em superfície plana e polida. A mortalidade dos adultos desse Coleoptera nas condições com e sem alimento foram semelhantes, o que justifica a dificuldade que esse inseto tem em acessar a fonte alimentar andando em superfície plana e polida. As medidas obtidas do comprimento (BOL), largura (BOW) e altura (BOH) do corpo de R. dominica foram comparadas com as de Tribolium castaneum (Coleoptera: Tenebrionidae), que pode ser considerado um bom caminhador nessas condições. Esse estudo mostrou que o primeiro é menor no BOL e BOW, e maior BOH que o segundo, e que, todas essas variáveis apresentaram diferenças quando analisadas simultaneamente por meio da construção de índices morfométricos multivariados (Largura × Altura; Comprimento × Altura e Altura × Comprimento × Largura). Essas variáveis morfométricas, juntamente com a definição da forma geométrica que mais se aproxima do formato corporal, proporcionaram a determinação do centro de gravidade (CG) e do limite lateral de tombamento (SRTgeom) para ambas as espécie. Rhyzopertha dominica e T. castaneum apresentaram CGs considerados alto e baixo, respectivamente, e juntamente com os valores obtidos para os SRTgeom, podem justificar que R. dominica pode ser considerada uma espécie menos estável durante o caminhamento, e que, apresenta maiores riscos de tombamento em superfícies plana e polida.

Walking stability of Rhyzopertha dominica (Fabricius, 1792) (Coleoptera: Bostrichidae).

Results obtained in studies can contribute to the advancement of science and innovative methods and techniques for developing practical activities. Reporting conditions that may restrict the implementation of research is critical to ensure the optimal development of further technical studies. The objective of this study was to assess the walking stability of R. dominica on a flat and smooth surface. The study was based on the determination of mortality, morphology and walking stability of the insect outside the grain mass, on a flat and smooth surface. Mortality of adults of this Coleoptera in conditions with and without food was similar, which explains the difficulty that this insect had for accessing the food source on the flat and smooth surface. The measurements of body length (BOL), width (BOW) and height (BOH) of R. dominica were compared with those of Tribolium castaneum (Coleoptera: Tenebrionidae), which showed good ability to walk in these conditions. This study indicated that the former presents lower BOL and BOW, and greater BOH than the second, and all these variables showed differences when analyzed simultaneously by means of the construction of multivariate morphometric indices (Width × Height, Length × Height and Height × Length × Width). These morphometric variables, together with the definition of the geometry most similar to the body shape, resulted in determination of the center of gravity (CG) and static rollover threshold (SRTgeom) for both species. Rhyzopertha dominica and T. castaneum presented CGs considered high and low, respectively, and together with the values obtained for SRTgeom, may justify that R. dominica can be considered a less stable species during movement, and presents greater risk of rollover on flat and smooth surfaces.

Aproveitamento de soro de queijo para produção de iogurte probiótico / The use of cheese whey for probiotic yogurt production

Desenvolveu-se um iogurte natural aproveitando o soro de queijo de coalho, avaliando diferentes processos de pasteurização e concentração de leite em pó. Os iogurtes submetidos a 90ºC/5min nas concentrações de 8 por cento e 10 por cento de leite em pó não apresentaram diferenças entre si (P>0,05) quanto à aceitação sensorial. Os produtos foram caracterizados como integral, atenderam às exigências de proteínas e forneceram mais de 15 por cento do requisito diário recomendado de cálcio para crianças de um a 10 anos, satisfazendo as exigências de cálcio. Em ambas as formulações, o pH decresceu de aproximadamente 4,6 para 4,2 e a acidez aumentou de 0,7g a 1,0g de ácido láctico/100g do produto durante 28 dias de armazenamento. Não houve contaminação por Staphilococcus aureus, coliformes fecais, bolores e leveduras. A contagem de bactérias láticas variou de 5,6x10(9) a 8,0x10(8)UFC/g durante a vida de prateleira, sugerindo a promoção de efeitos benéficos por este alimento probiótico. Concluiu-se que a utilização do soro para a produção de iogurte é uma alternativa viável ao agregar valor econômico, nutricional e funcional ao subproduto.

The objective of this work was to develop a natural yogurt utilizing the resultant whey of a "coalho" cheese production, assaying different processes of pasteurization and powdered milk concentrations. The yogurt submitted to 90ºC/5min with 8 and 10 percent of powdered milk achieved the highest grade on sensorial analysis, with no significant difference (P>0,05). Products were characterized as integral, attended the request for protein, and supplied over 15 percent of the daily feed intake of calcium for children aged 1 to 10, characterizing it as calcium enriched. In both formulations, the pH decreased from 4.6 to 4.2 approximately and the acidity increased from 0.7g to 1.0g lactic acid/100g of product during 28 days of storage. There was no contamination by S. aureus, fecal coliforms, molds and yeasts. The counting of lactic bacteria varied from 5.6x10(9) to 8.0x10(8)CFU/g during shelf life, suggesting the promotion benefits from this probiotic product. It was concluded that whey utilization for yogurt production is a viable alternative to increase economic, nutritional, and functional value.

Effect of pH on the gelation properties of skim milk gels made from plant coagulants and chymosin.

The effect of three milk pH values, 6.0, 6.3 and 6.7, on gelation properties was monitored by small amplitude oscillatory rheology as well as a large deformation (yield) test for gels induced by the plant coagulants, Cynara cardunculus L. and Cynara humilis L., and chymosin. Gel microstructure was studied using confocal scanning laser microscopy. For each coagulant, a decrease in pH of milk resulted in a decrease in gelation time (tg), and an increase in the rate of increase in storage modulus (G'). At pH 6.0 and 6.3, plant coagulant-induced gels reached a maximum value in G' (G'max) followed by a decrease in G' values during the rest of the experiment, reflecting higher proteolytic activity of plant coagulants towards caseins as determined by SDS-PAGE. Gels produced at pH 6.0 and 6.3, exhibited a minimum in loss tangent (tan delta) followed by slight increase in tan delta during gel aging, that may have been associated with faster rearrangements of the gel network structure. In gels aged for approximately 6 h, the values for G', tan delta at low frequency (0.006 Hz) and yield stress were higher for chymosin than for plant-induced gels. For gels with the same pH value, no major differences were observed in microstructure between coagulants. Gels made at low pH values (6.3 and 6.0) appeared to have a denser or more interconnected structure than gels made at pH 6.7. Our results suggest that, at a low pH, the type of coagulant used in gelation is likely to have a considerably impact on gel/cheese structure.

Mathematical modelling of the formation of rennet-induced gels by plant coagulants and chymosin.

Rheological properties of reconstituted skim milk coagulated with plant coagulants Cynara cardunculus L., Cynara humilis L. and chymosin was monitored by dynamic low amplitude oscillation. There are no published reports on the modelling of the gelation behaviour of milk by plant coagulants. Three mathematical models, Scott Blair. Douillard and Carlson, were fitted to the storage modulus (G') as function of time curves. For all coagulants. Scott Blair model was the most efficient in modelling the gelation process, and gave both the smallest residuals and standard error of residuals, Se (P < 0.0001). Douillard model gave the poorest fit and in particular it was not able to predict the initial part of the gelation curves. Carlson model had an intermediate behaviour and, in the case of chymosin, it gave results that were quite comparable to Scott Blair model. The parameters of the Scott Blair model were different for plant coagulants and chymosin. Chymosin had the longest rate constant (tau) and the time shift coefficient (t8) was also different between vegetable coagulants and chymosin (P < 0.0005). These results are in agreement with the overall trends for gelation profiles obtained for vegetable coagulants and chymosin. In the beginning of gelation both plant coagulants produced gels with slightly higher G' values than chymosin, but after longer incubation times chymosin gels had higher G' values. It was concluded that the Scott Blair model was the best equation to follow the gelation of milk induced by both plant coagulants as well as chymosin. Modelling is an important and useful method for comparing the gelation process in gels formed by different types of coagulants.

Arbitrarily primed PCR fingerprinting of RNA and DNA in Entamoeba histolytica.

Differences were detected in the gene expression of strains of E. histolytica using RNA (RAP-PCR) and DNA fingerprinting (RAPD). Analysis of the electrophoretic profiles of the gels revealed some polymorphic markers that could be used in the individual characterization of the strains. The 260 bands generated by using five different primers for RAP-PCR, as well as RAPD, were employed in the construction of dendograms. The dendogram obtained based on the RAPD products permitted the distinction of symptomatic and asymptomatic isolates, as well the correlation between the polymorphism exhibited and the virulence of the strains. The dendogram obtained for the RAP-PCR products did not show a correlation with the virulence of the strains but revealed a high degree of intraspecific transcriptional variability that could be related to other biological features, whether or not these are involved in the pathogenesis of amebiasis.

Avaliaçäo microbiológica de refeiçöes servidas por empresas aéreas nacionais / Microbiological evaluation of food services in aviation

Avaliou-se a qualidade microbiológica de refeiçöes servidas por empresas aéreas nacionais, sendo constituídas por pratos à base de pescado, carnes e produtos cárneos, produtos lácteos e sobremesas, além da água utilizada nos locais de processamento dos alimentos e das mäos dos manipuladores. Foram determinadas contagens de bactérias mesófilas, de coliformes totais e E. coli e contagem e identificaçäo de S. aureus. Dos resultados obtidos constatou-se que todas as amostras de alimentos apresentavam contagens para bactérias mesófilas e coliformes totais abaixo dos limites máximos estabelecidos pelos padröes internacionais (RIBEIRO, 1974; ICMSF, 1982). A presença de E. coli em três amostras de queijo ricota, duas com valores 6,5 x 100 UFC/g e uma com valores maiores que 3,0 x 10.000 UFC/g, sendo estes resultados acima do permitido pela legislaçäo brasileira (BRASIL, 1997), demonstram a baixa qualidade do produto artesanal e as precárias condiçöes higiênico-sanitárias durante a produçäo. Verificou-se ausência de S. aureus em 100 por cento das amostras de alimentos analisados. Constatou-se, também, ausência de coliformes totais nas 10 (100 por cento) amostras de água e ausência de E. coli e S. aureus em todas as amostras das mäos dos manipuladores de alimentos. Os resultados encontrados sugerem a existência de condiçöes higiências adequadas durante o processamento e manipulaçäo das refeiçöes oferecidas pelas empresas aéreas nacionais, sendo próprias para consumo.

Extracellular alpha-amylase from Thermus filiformis Ork A2: purification and biochemical characterization.

An extracellular alpha-amylase produced by the thermophilic bacterium Thermus filiformis Ork A2 was purified from cell-free culture supernatant by ion exchange chromatography. The molecular mass was estimated to be 60,000 Da by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was rich in both basic and hydrophobic amino acids, presenting the following NH2-terminal amino acid sequence: Thr-Ala-Asp-Leu-Ile-Val-Lys-Ile-Asn-Phe. Amylolytic activity on soluble starch was optimal at pH 5.5-6.0 and 95 degrees C, and the enzyme was stable in the pH range of 4.0-8.0. Calcium enhanced thermostability at temperatures above 80 degrees C, increasing the half-life of activity to more than 8 h at 85 degrees C, 80 min at 90 degrees C, and 19 min at 95 degrees C. Ethylenediaminetetraacetic acid (EDTA) inhibited amylase activity, the inhibition being reversed by the addition of calcium or strontium ions. The alpha-amylase was also inhibited by copper and mercuric ions, and p-chloromercuribenzoic acid, the latter being reversed in the presence of dithiothreitol. Dithiothreitol and beta-mercaptoethanol activated the enzyme. The alpha-amylase exhibited Michaelis-Menten kinetics for starch, with a K(m) of 5.0 mg.ml-1 and kcat/K(m) of 5.2 x 10(5) ml.mg-1s-1. Similar values were obtained for amylose, amylopectin, and glycogen. The hydrolysis pattern was similar for maltooligosaccharides and polysaccharides, with maltose being the major hydrolysis product. Glucose and maltotriose were generated as secondary products, although glucose was produced in high levels after a 6-h digestion. To our knowledge this is the first report of the characterization of an alpha-amylase from a strain of the genus Thermus.