Instituto de Biotecnologia

This paper studies the application of support vector machines (SVMs) to the detection and classification of rolling-element bearing faults. The training of the SVMs is carried out using the sequential minimal optimization (SMO) algorithm. In this paper, a mechanism for selecting adequate training parameters is proposed. This proposal makes the classification procedure fast and effective. Various scenarios are examined using two sets of vibration data, and the results are compared with those available in the literature that are relevant to this investigation. r

This work explores the use of characterization features extracted based on breast-mass contours obtained by automated segmentation methods, for the classification of masses in mammograms according to their diagnosis (benign or malignant). Two sets of mass contours were obtained via two segmentation methods (a dynamic-programming-based method and a constrained region-growing method), and simplified versions of these contours (modeling the contours as ellipses) were employed to extract a set of six features designed for characterization of mass margins (contrast between foreground region and background region, coefficient of variation of edge strength, two measures of the fuzziness of mass margins, a measure of spiculation based on relative gradient orientation, and a measure of spiculation based on edge-signature information). Three popular classifiers (Bayesian classifier, Fisher's linear discriminant, and a support vector machine) were then used to predict the diagnosis of a set of 349 masses based on each of said features and some combinations of these. The systems (each system consists of a segmentation method, a featureset, and a classifier) were compared with each other in terms of their performance on the diagnosis of the set of breast masses. It was found that, although there was a percent difference of about 14% in the average segmentation quality between methods, this was translated into an average percent difference of only 4% in the classification performance. It was also observed that the spiculation feature based on edgesignature information was distinctly better than the rest of the features, although it is not very robust to changes in the quality of the segmentation. All systems were more efficient in predicting the diagnosis of benign masses than that of the malignant masses, resulting in low sensitivity and high specificity values (e.g. 0.6 and 0.8, respectively) since the positive class in the classification experiments is the set of malignant masses. It was concluded that features extracted from automated contours can contribute to the diagnosis of breast masses in screening programs by correctly identifying a majority of benign masses.

Metodología. Se adquirieron videos con una cámara de alta velocidad (Motion Pro HS-4, Redlake, USA) utilizando los diferentes arreglos experimentales presentados previamente (1, 3). La caracterización dinámica de la dispersión se realizó siguiendo la metodología reportada en (2), con el medio de cultivo para Trichoderma harzianum (1) como fase dispersa y 0.25 vvm de aire, así como una solución de 0.6% w/v carboximetilcelulosa como sistema no Newtoniano.

Resultados. En los experimentos donde se analizan los mecanismos de formación de estructuras, se observó que en algunos casos la inclusión de las burbujas de aire dentro de las gotas de aceite ocurre cuando estos objetos entran en contacto unos con otros (figura 1), aún cuando la colisión directa entre sí no siempre sigue la misma trayectoria. Además, cuando la burbuja de aire se aproxima a la gota de aceite, en algunos casos se observó un efecto de rebote sobre la gota de aceite (figuras 1b, 1c) antes de incluirse en ella.

Fermentation processes involve multiphase dispersions in which there are oil drops, air bubbles and compound oil drops containing air bubbles and water droplets. The dynamics of formation of these complex structures has been intended to be explained theoretically in several works; nevertheless, the phenomena itself has never been observed at the instant of formation.

The phosphoenolpyruvate: carbohydrate transferase system (PTS) transports glucose in Escherichia coli. Previous work demonstrated that strains lacking PTS, such as PB11, grow slow on glucose. PB11 has a reduced expression of glycolytic, and upregulates poxB and acs genes as compared to the parental strain JM101, when growing on glucose. The products of the latter genes are involved in the production of AcetylCoA. Inactivation of rpoS that codes for the RNA polymerase s 38 subunit, reduces further (50%) growth of PB11, indicating that s 38 plays a central role in the expression of central metabolism genes in slowly growing cells. In fact, transcription levels of glycolytic genes is reduced in strain PB11rpoS 2 as compared to PB11. In this report we studied the role of s 70 and s 38 in the expression of the complete glycolytic pathway and poxB and acs genes in certain PTS 2 strains and their rpoS 2 derivatives. We determined the transcription start sites (TSSs) and the corresponding promoters, in strains JM101, PB11, its derivative PB12 that recovered its growth capacity, and in their rpoS 2 derivatives, by 59RACE and pyrosequencing. In all these genes the presence of sequences resembling s 38 recognition sites allowed the proposition that they could be transcribed by both sigma factors, from overlapping putative promoters that initiate transcription at the same site. Fourteen new TSSs were identified in seventeen genes. Besides, more than 30 putative promoters were proposed and we confirmed ten previously reported. In vitro transcription experiments support the functionality of putative dual promoters. Alternatives that could also explain lower transcription levels of the rpoS 2 derivatives are discussed. We propose that the presence if real, of both s 70 and s 38 dependent promoters in all glycolytic genes and operons could allow a differential transcription of these central metabolism genes by both sigma subunits as an adaptation response to carbon limitation.

Pore-forming toxins are biological weapons produced by a variety of living organisms, particularly bacteria but also by insects, reptiles, and invertebrates. These proteins affect the cell membrane of their target, disrupting permeability and leading eventually to cell death. The pore-forming toxins typically transform from soluble, monomeric proteins to oligomers that form transmembrane channels. The Cry toxins produced by Bacillus thuringiensis are widely used as insecticides. These proteins have been recognized as pore-forming toxins, and their primary action is to lyse midgut epithelial cells in their target insect. To exert their toxic effect, a prepore oligomeric intermediate is formed leading finally to membrane-inserted oligomeric pores. To understand the role of Cry oligomeric pre-pore formation in the insecticidal activity we isolated point mutations that affected toxin oligomerization but not their binding with the cadherin-like, Bt-R 1 receptor. We show the helix ␣-3 in domain I contains sequences that could form coiled-coil structures important for oligomerization. Some single point mutants in this helix bound Bt-R 1 receptors with similar affinity as the wild-type toxin, but were affected in oligomerization and were severally impaired in pore formation and toxicity against Manduca sexta larvae. These data indicate the pre-pore oligomer and the toxin pore formation play a major role in the intoxication process of Cry1Ab toxin in insect larvae.

The a-amylase from Bacillus licheniformis is the most widely used enzyme in the starch industry owing to its hyperthermostability, converting starch to medium-sized oligosaccharides. Based on sequence alignment of homologous amylases, we found a semi-conserved sequence pattern near the active site between transglycosidic and hydrolytic amylases, which suggested that hydrophobicity may play a role in modifying the transglycosylation/hydrolysis ratio. Based on this analysis, we replaced residue Val286 by Phe and Tyr in Bacillus licheniformis a-amylase. Surprisingly, the two resultant mutant enzymes, Val286Phe and Val286Tyr, showed two different behaviors. Val286Tyr mutant was 5-fold more active for hydrolysis of starch than the wild-type enzyme. In contrast, the Val286Phe mutant, differing only by one hydroxyl group, was 3-fold less hydrolytic than the wild-type enzyme and apparently had a higher transglycosylation/hydrolysis ratio. These results are discussed in terms of af®nity of subsites, hydrophobicity and electrostatic environment in the active site. The engineered enzyme reported here may represent an attractive alternative for the starch transformation industries as it affords direct and substantial material savings and requires no process modi®cations.

Bacillus thuringiensis Cry toxins are used worldwide as insecticides in agriculture, in forestry, and in the control of disease transmission vectors. In the lepidopteran Manduca sexta, cadherin (Bt-R 1) and aminopeptidase-N (APN) function as Cry1A toxin receptors. The interaction with Bt-R 1 promotes cleavage of the amino-terminal end, including helix ␣-1 and formation of prepore oligomer that binds to APN, leading to membrane insertion and pore formation. Loops of domain II of Cry1Ab toxin are involved in receptor interaction. Here we show that Cry1Ab mutants located in domain II loop 3 are affected in binding to both receptors and toxicity against Manduca sexta larvae. Interaction with both receptors depends on the oligomeric state of the toxin. Monomers of loop 3 mutants were affected in binding to APN and to a cadherin fragment corresponding to cadherin repeat 12 but not with a fragment comprising cadherin repeats 7-12. In contrast, the oligomers of loop 3 mutants were affected in binding to both Bt-R 1 fragments but not to APN. Toxicity assays showed that either monomeric or oligomeric structures of Cry1Ab loop 3 mutations were severely affected in insecticidal activity. These data suggest that loop 3 is differentially involved in the binding with both receptor molecules, depending on the oligomeric state of the toxin and also that possibly a "ping pong" binding mechanism with both receptors is involved in toxin action.