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4. Polygenic analysis of phenylethyl acetate production in yeast for aroma production improvement in alcoholic beverages : Polygene analyse van fenylethylacetaat productie in gist voor aroma verbetering in alcoholische dranken

Author

Trindade de Carvalho, B and Thevelein, J

Abstract

Flavor compound metabolism is one of the last areas in metabolism where multiple genes encoding biosynthetic enzymes are still unknown. A major challenge is the involvement of side-activities of enzymes having their main function in other parts of metabolism. We have applied polygenic analysis to identify novel Saccharomyces cerevisiae genes affecting production of phenylethyl acetate (2-PEAc), a flavor compound of great importance in alcoholic beverages, imparting rose and honey-like aromas. We have successfully applied pooled-segregant whole-genome sequence analysis to segregants of two non-selected parents, without significant difference in the trait of interest. This suggests that many more strains than anticipated from phenotypic profiling may harbor interesting superior alleles for traits of interest. The QTLs responsible for high production of 2-PEAc showed linkage to either one of the two parents. The causative genes in two QTLs linked to one parent were identified by reciprocal allele swapping into both parents using CRISPR/Cas9. The superior allele of the first major causative gene, FAS2, contained two unique SNPs responsible for high 2-PEAc production, not present in other sequenced yeast strains. FAS2 encodes the alpha subunit of the fatty acid synthetase complex confirming that regular metabolic enzymes have side activities responsible for production of flavor compounds. Surprisingly, the second causative gene was a mutant allele of TOR1, a gene involved in nitrogen regulation of growth. It may affect flavor production indirectly. Exchange of both superior alleles in the inferior parent strain increased 2-PEAc production with 70%, i.e. nearly to the same level as in the best superior segregant. Our results suggest that polygenic analysis combined with CRISPR/Cas9 mediated site-directed genetic modification may evolve into a powerful tool for identification of genes encoding missing metabolic enzymes and for development of industrial yeast strains generating novel flavor profiles in alcoholic beverages. List of Figures v List of Abbreviations vii Abstract ix Samenvatting xi Chapter I - Literature review 1 1. Saccharomyces cerevisiae and its applications 1 2. Flavor of fermented beverages 2 3. Ester production by S. cerevisiae 5 4. Factors affecting ester production 8 4.1. Suspended solids and fatty acid addition 8 4.2. Wort/must aeration 8 4.3. Fermentation temperature 8 4.4. Inoculum size (pitching rate) 9 4.5. Fermentation pressure 10 4.6. Amino acid composition of the fermentation medium 10 5. Increasing ester production by genetic modifications 12 LEU4: release from leucine-feedback inhibition leads to enhanced isoamyl alcohol and isoamyl acetate production. 12 5.1. TYR1 and ARO4: resistance to phenylalanine analogs correlates with increased 2-phenylethanol and 2-phenylethyl acetate production. 14 5.2. FAS2: release from cerulenin-inhibition leads to increased production of ethyl esters, especially ethyl hexanoate. 16 5.3. ATF1 and ATF2: Overexpression of the alcohol acetyl transferases Atf1 and Atf2 leads to increased acetate esters production. 17 5.4. BAT1 and BAT2: Overexpression of the aminotransferases BAT1 and BAT2 results in increased isobutanol and isobutyl acetate levels 18 6. Phenylethyl acetate production in fermented beverages 18 7. Improving yeasts by using naturally occurring “superior alleles” – QTL mapping 19 8. CRISPR/Cas9, a powerful tool for genetic engineering 22 9. Research objectives 27 Chapter II - QTL mapping of high 2-phenylethyl acetate production by using random parent strains 29 1. Introduction 29 2. Results 30 2.1. Flavor profile of industrial yeast and random parent selection 30 2.2. Pooled-segregant whole-genome sequence analysis and QTL mapping 39 3. Discussion 42 Chapter III - Identification of the causative genes conferring high phenylethyl acetate production by using CRISPR/Cas9 approach 45 1. Introduction 45 2. Results 47 2.1. Identification of TOR1 as a causative gene on chromosome X 47 2.2. Identification of FAS2 as causative gene on chromosome XVI 55 2.3. Achieving the superior phenotype by replacing TOR1 and FAS2 alleles in ER18 parent. 59 2.4. Production of the other flavor compounds in the ER18 strains engineered for high 2-PEAc production 61 3. Discussion 65 Chapter IV - Final remarks, conclusions and perspectives 71 Limitations of increasing 2-PEAc production by alteration in fermentation parameters 71 Genetic improvement of industrial yeast strains 73 Genetic tools 74 QTL mapping 74 CRISPR/Cas9 75 Perspectives 77 Materials and methods 79 Microorganisms and cultivation media 79 Fermentation experiments 81 Headspace GC-FID analysis 81 Sporulation and tetrad dissection 82 Mating type 83 Deletion of HO gene 83 Crossing of haploid strains 83 Molecular Biology methods 83 Genomic DNA extraction and whole genome sequence analysis 85 Allele-specific PCR 86 CRISPR/Cas9 experiments 89 Cas9 plasmid 89 gRNA plasmids 89 Design guide RNA targets 92 Donor DNA 92 Appendix 1 95 References 105 status: published

Published
2018

7. Confirmation of the topology of the Wendelstein 7-X magnetic field to better than 1:100,000

Author

Pedersen T. S., Otte M., Lazerson S., Helander P., Bozhenkov S., Biedermann C., Klinger T., Wolf R. C., Bosch H. -S., Abramovic I., Akaslompolo S., Aleynikov P., Aleynikova K., Ali A., Alonso A., Anda G., Andreeva T., Ascasibar E., Baldzuhn J., Banduch M., Barbui T., Beidler C., Benndorf A., Beurskens M., Biel W., Birus D., Blackwell B., Blanco E., Blatzheim M., Bluhm T., Bockenhoff D., Bolgert P., Borchardt M., Bottger L. -G., Brakel R., Brandt C., Brauer T., Braune H., Burhenn R., Buttenschon B., Bykov V., Calvo I., Cappa A., Carls A., De Carvalho B. B., Castejon F., Cianciosa M., Cole M., Costea S., Cseh G., Czarnecka A., Dal Molin A., De La Cal E., De La Pena A., Degenkolbe S., Dhard C. P., Dinklage A., Dostal M., Drevlak M., Drewelow P., Drews P., Dudek A., Durodie F., Dzikowicka A., Von Eeten P., Effenberg F., Endler M., Erckmann V., Estrada T., Fahrenkamp N., Fellinger J., Feng Y., Figacz W., Ford O., Fornal T., Frerichs H., Fuchert G., Garcia-Munoz M., Geiger B., Geiger J., Gierse N., Gogoleva A., Goncalves B., Gradic D., Grahl M., Gross S., Grote H., Grulke O., Guerard C., Haas M., Harris J., Hartfuss H. -J., Hartmann D., Hathiramani D., Hein B., Heirnich S., Henneberg S., Hennig C., Hernandez J., Hidalgo C., Hidalgo U., Hirsch M., Hofel U., Holbe H., Holting A., Houry M., Huber V., Ionita C., Israeli B., Jablonski S., Jakubowski M., Van Vuuren A. J., Jenzsch H., Kaczmarczyk J., Kallmeyer J. -P., Kamionka U., Kasahara H., Kenmochi N., Kernbichler W., Killer C., Kinna D., Kleiber R., Knauer J., Kochl F., Kocsis G., Kolesnichenko Y., Konies A., Konig R., Kornejew P., Koster F., Kramer-Flecken A., Krampitz R., Krawzyk N., Kremeyer T., Krychowiak M., Ksiazek I., Kubkowska M., Kuhner G., Kurki-Suonio T., Kurz P., Kuttler K., Kwak S., Landreman M., Langenberg A., Lapayese F., Laqua H., Laqua H. -P., Laube R., Laux M., Lentz H., Lewerentz M., Liang Y., Liu S., Lobsien J. -F., Cisquella J. L., Lopez-Bruna D., Lore J., Lorenz A., Lui S., Lutsenko V., Maassberg H., Maisano-Brown J., Marchuk O., Marrelli L., Marsen S., Marushchenko N., Masuzaki S., McCarthy K., McNeely P., Medina F., Milojevic D., Mishchenko A., Missal B., Mittelstaedt J., Mollen A., Moncada V., Monnich T., Moseev D., Nagel M., Naujoks D., Neilson G. H., Neubauer O., Neuner U., Ngo T. -T., Niemann H., Nuhrenberg C., Nuhrenberg J., Ochando M., Ogawa K., Ongena J., Oosterbeek H., Pablant N., Pacella D., Pacios L., Panadero N., Pasch E., Pastor I., Pavone A., Pawelec E., Pedrosa A., Perseo V., Peterson B., Pilopp D., Pisano F., Piulatti M. E., Plunk G., Preynas M., Proll J., Sitjes A. P., Purps F., Rack M., Rahbarnia K., Riemann J., Risse K., Rong P., Rosenberger J., Rudischhauser L., Rummel K., Rummel T., Runov A., Rust N., Ryc L., Saitoh H., Satake S., Schacht J., Schmitz O., Schmuck S., Schneider B., Schneider M., Schneider W., Schrittwieser R., Schroder M., Schroder T., Schroder R., Schumacher H. W., Schweer B., Seki R., Sinha P., Sipilae S., Slaby C., Smith H., Sousa J., Spring A., Standley B., Stange T., Von Stechow A., Stephey L., Stoneking M., Stridde U., Suzuki Y., Svensson J., Szabolics T., Szepesi T., Thomsen H., Travere J. -M., Traverso P., Mora H. T., Tsuchiya H., Tsuijmura T., Turkin Y., Valet S., Van Milligen B., Vela L., Velasco J. -L., Vergote M., Vervier M., Viebke H., Vilbrandt R., Von Thun C. P., Wagner F., Wang E., Wang N., Warmer F., Wauters T., Wegener L., Wegner T., Weir G., Wendorf J., Wenzel U., Werner A., Wie Y., Wiegel B., Wilde F., Windisch T., Winkler M., Winters V., Wright A., Wurden G., Xanthopoulos P., Yamada I., Yasuhara R., Yokoyama M., Zhang D., Zilker M., Zimbal A., Zocco A., Zoletnik S., W7-X Team, Max Planck Institute for Plasma Physics, Max Planck Society, Massachusetts Institute of Technology. Department of Physics, Maisano-Brown, Jeannette D., Science and Technology of Nuclear Fusion, Pedersen, T, Otte, M, Lazerson, S, Helander, P, Bozhenkov, S, Biedermann, C, Klinger, T, Wolf, R, Bosch, H, Abramovic, I, Akaslompolo, S, Aleynikov, P, Aleynikova, K, Ali, A, Alonso, A, Anda, G, Andreeva, T, Ascasibar, E, Baldzuhn, J, Banduch, M, Barbui, T, Beidler, C, Benndorf, A, Beurskens, M, Biel, W, Birus, D, Blackwell, B, Blanco, E, Blatzheim, M, Bluhm, T, Bockenhoff, D, Bolgert, P, Borchardt, M, Bottger, L, Brakel, R, Brandt, C, Brauer, T, Braune, H, Burhenn, R, Buttenschon, B, Bykov, V, Calvo, I, Cappa, A, Carls, A, De Carvalho, B, Castejon, F, Cianciosa, M, Cole, M, Costea, S, Cseh, G, Czarnecka, A, Dal Molin, A, De La Cal, E, De La Pena, A, Degenkolbe, S, Dhard, C, Dinklage, A, Dostal, M, Drevlak, M, Drewelow, P, Drews, P, Dudek, A, Durodie, F, Dzikowicka, A, Von Eeten, P, Effenberg, F, Endler, M, Erckmann, V, Estrada, T, Fahrenkamp, N, Fellinger, J, Feng, Y, Figacz, W, Ford, O, Fornal, T, Frerichs, H, Fuchert, G, Garcia-Munoz, M, Geiger, B, Geiger, J, Gierse, N, Gogoleva, A, Goncalves, B, Gradic, D, Grahl, M, Gross, S, Grote, H, Grulke, O, Guerard, C, Haas, M, Harris, J, Hartfuss, H, Hartmann, D, Hathiramani, D, Hein, B, Heirnich, S, Henneberg, S, Hennig, C, Hernandez, J, Hidalgo, C, Hidalgo, U, Hirsch, M, Hofel, U, Holbe, H, Holting, A, Houry, M, Huber, V, Ionita, C, Israeli, B, Jablonski, S, Jakubowski, M, Van Vuuren, A, Jenzsch, H, Kaczmarczyk, J, Kallmeyer, J, Kamionka, U, Kasahara, H, Kenmochi, N, Kernbichler, W, Killer, C, Kinna, D, Kleiber, R, Knauer, J, Kochl, F, Kocsis, G, Kolesnichenko, Y, Konies, A, Konig, R, Kornejew, P, Koster, F, Kramer-Flecken, A, Krampitz, R, Krawzyk, N, Kremeyer, T, Krychowiak, M, Ksiazek, I, Kubkowska, M, Kuhner, G, Kurki-Suonio, T, Kurz, P, Kuttler, K, Kwak, S, Landreman, M, Langenberg, A, Lapayese, F, Laqua, H, Laube, R, Laux, M, Lentz, H, Lewerentz, M, Liang, Y, Liu, S, Lobsien, J, Cisquella, J, Lopez-Bruna, D, Lore, J, Lorenz, A, Lui, S, Lutsenko, V, Maassberg, H, Maisano-Brown, J, Marchuk, O, Marrelli, L, Marsen, S, Marushchenko, N, Masuzaki, S, Mccarthy, K, Mcneely, P, Medina, F, Milojevic, D, Mishchenko, A, Missal, B, Mittelstaedt, J, Mollen, A, Moncada, V, Monnich, T, Moseev, D, Nagel, M, Naujoks, D, Neilson, G, Neubauer, O, Neuner, U, Ngo, T, Niemann, H, Nuhrenberg, C, Nuhrenberg, J, Ochando, M, Ogawa, K, Ongena, J, Oosterbeek, H, Pablant, N, Pacella, D, Pacios, L, Panadero, N, Pasch, E, Pastor, I, Pavone, A, Pawelec, E, Pedrosa, A, Perseo, V, Peterson, B, Pilopp, D, Pisano, F, Piulatti, M, Plunk, G, Preynas, M, Proll, J, Sitjes, A, Purps, F, Rack, M, Rahbarnia, K, Riemann, J, Risse, K, Rong, P, Rosenberger, J, Rudischhauser, L, Rummel, K, Rummel, T, Runov, A, Rust, N, Ryc, L, Saitoh, H, Satake, S, Schacht, J, Schmitz, O, Schmuck, S, Schneider, B, Schneider, M, Schneider, W, Schrittwieser, R, Schroder, M, Schroder, T, Schroder, R, Schumacher, H, Schweer, B, Seki, R, Sinha, P, Sipilae, S, Slaby, C, Smith, H, Sousa, J, Spring, A, Standley, B, Stange, T, Von Stechow, A, Stephey, L, Stoneking, M, Stridde, U, Suzuki, Y, Svensson, J, Szabolics, T, Szepesi, T, Thomsen, H, Travere, J, Traverso, P, Mora, H, Tsuchiya, H, Tsuijmura, T, Turkin, Y, Valet, S, Van Milligen, B, Vela, L, Velasco, J, Vergote, M, Vervier, M, Viebke, H, Vilbrandt, R, Von Thun, C, Wagner, F, Wang, E, Wang, N, Warmer, F, Wauters, T, Wegener, L, Wegner, T, Weir, G, Wendorf, J, Wenzel, U, Werner, A, Wie, Y, Wiegel, B, Wilde, F, Windisch, T, Winkler, M, Winters, V, Wright, A, Wurden, G, Xanthopoulos, P, Yamada, I, Yasuhara, R, Yokoyama, M, Zhang, D, Zilker, M, Zimbal, A, Zocco, A, Zoletnik, S, Universidad de Sevilla. Departamento de Física Atómica, Molecular y Nuclear, Department of Applied Physics, Aalto-yliopisto, Aalto University, and Pacella, D.

Subjects
Tokamak, Plasma parameters, Science, General Physics and Astronomy, Topology (electrical circuits), Topology, 7. Clean energy, 01 natural sciences, Article, General Biochemistry, Genetics and Molecular Biology, 010305 fluids & plasmas, law.invention, law, 0103 physical sciences, 010306 general physics, Physics, Fusion, Wendelstein7-X, Stellarator, Multidisciplinary, ta114, General Chemistry, Plasma, Fusion power, Magnetic field, Erratum, Wendelstein 7-X, Stellarator
Abstract

Fusion energy research has in the past 40 years focused primarily on the tokamak concept, but recent advances in plasma theory and computational power have led to renewed interest in stellarators. The largest and most sophisticated stellarator in the world, Wendelstein 7-X (W7-X), has just started operation, with the aim to show that the earlier weaknesses of this concept have been addressed successfully, and that the intrinsic advantages of the concept persist, also at plasma parameters approaching those of a future fusion power plant. Here we show the first physics results, obtained before plasma operation: that the carefully tailored topology of nested magnetic surfaces needed for good confinement is realized, and that the measured deviations are smaller than one part in 100,000. This is a significant step forward in stellarator research, since it shows that the complicated and delicate magnetic topology can be created and verified with the required accuracy., Early stellarator designs suffered from high particle losses, an issue that can be addressed by optimization of the coils. Here the authors measure the magnetic field lines in the Wendelstein 7-X stellarator, confirming that the complicated design of the superconducting coils has been realized successfully.

Published
2016

8. European survey on criteria of aesthetics for periodontal evaluation: The ESCAPE study.

Author

Le Roch, Sarah, Rouche, Frédéric, Valet, Fabien, Bouchard, Philippe, Abrahamsson, I, Artzi, Z, Asbi, T, Balta, M.G, Bizzarro, S, Buti, J, Chaushu, L, Danser, M, De Carvalho, B, Garabetyan, J, Goldstein, M, Gursoy, H, Harmouche, L, Harrison, P, Herrera, D, and Horwitz, J

Subjects
MEDICAL cooperation, PERIODONTICS, QUESTIONNAIRES, RESEARCH, STATISTICS, SURVEYS, TEACHERS
Abstract

Objective: The ESCAPE multicentre survey was designed to (a) compare the agreement of three relevant aesthetic scoring systems among different centres, and (b) evaluate the reproducibility of each question of the questionnaires. Materials and Methods: EFP centres (n = 14) were involved in an e‐survey. Forty‐two participants (28 teachers, 14 postgraduate students) were asked to score the one‐year aesthetic outcomes of photographs using the Before–After Scoring System (BASS), the Pink Esthetic Score (PES) and the Root coverage Esthetic Score (RES). Mean values of kappa statistics performed on each question were provided to resume global agreement of each method. Results: Between teachers, a difference of kappa ≥ 0.41 (p = .01) was found for BASS (75%) and PES (57%). Similarly, RES (84%) and PES (57%) were different (p < .001). No difference was found between BASS (75%) and RES (84%). No difference was found between students, whatever the scoring system. Questions of each scoring system showed differences in their reproducibility. Conclusions: The outcomes of this study indicate that BASS and RES scoring systems are reproducible tools to evaluate aesthetic after root coverage therapies between different centres. Among the various variables, lack of scar, degree of root coverage, colour match and gingival margin that follows the CEJ show the best reliability. [ABSTRACT FROM AUTHOR]

Published
2019
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9. Avaliação do desenvolvimento tecnológico e transferência de tecnologia: o caso Instituto Oswaldo Cruz ­ Fundação Oswaldo Cruz Evaluation of the technological development and technology transfer at Instituto Oswaldo Cruz ­ Fundação Oswaldo Cruz: a case study

Author

Luciane de Carvalho B. Willcox

Subjects
Indicadores, lcsh:Public aspects of medicine, Indicators, lcsh:RA1-1270, Ciência e tecnologia, Inovação, Innovation, Science and technology
Abstract

No presente trabalho é apresentada uma proposta de medidas de avaliação de desempenho para as áreas de desenvolvimento tecnológico e de transferência de tecnologia do Instituto Oswaldo Cruz-Fundação Oswaldo Cruz (IOC-Fiocruz). A proposta foi feita com base em pesquisa comparativa com cinco institutos de pesquisa que apresentam indicadores consolidados para essas áreas e pela análise de documentos de avaliação. A proposta desenvolvida para o IOC a partir desta pesquisa foi submetida aos pesquisadores do Instituto para validação. Os resultados obtidos foram positivos e indicaram que há necessidade de se implementar a gestão voltada para resultados.This work presents a proposition list of indicators to evaluate the technological development and technology transfer areas of Instituto Oswaldo Cruz-Fundação Oswaldo Cruz (IOC-Fiocruz).The proposition was supported by a research performed in five institutes which already present consolidated indicators for these areas and by the analysis of evaluation documents. The proposition developed as a result of this research was submitted to the IOC-Fiocruz researches for validation. The results were positive and indicated the necessity of introducing the result based management.

Published
2004

12. Accuracy of a TaqMan‐based real‐time polymerase chain reaction combined to a Novy‐MacNeal‐Nicolle medium culture for the diagnosis of American tegumentary leishmaniasis.

Author

Osiro Bergmann, J., de Castro Moreira dos Santos Júnior, A., Sevilha‐Santos, L., Medeiros‐Silva, V., Bresolin Pompeu, C., Youssif Mota Arabi, A., da Silva Marques, D., Caroline Véras de Carvalho, B., Nitz, N., Martins Gomes, C., Dolabela de Lima, B., and Nonata Ribeiro Sampaio, R.

Subjects
POLYMERASE chain reaction, RESTRICTION fragment length polymorphisms
Abstract

The article offers information on a study related to the accuracy of a TaqMan-based real-time polymerase chain reaction (PCR) combined to a NovyMacNeal-Nicolle medium culture for the diagnosis of American tegumentary leishmaniasis (TL). Topics discussed include use of NucleoSpin Tissue Kit for DNA extraction; approval of the study from Ethics Committee of the Faculty of Medicine; and the results showing that samples extracted from active TL lesions will present Leishmania amastigotes.

Published
2019
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13. Overview of diagnostic performance and results for the first operation phase in Wendelstein 7-X (invited).

Author

Krychowiak, M., Adnan, A., Alonso, A., Andreeva, T., Baldzuhn, J., Barbui, T., Beurskens, M., Biel, W., Biedermann, C., Blackwell, B. D., Bosch, H. S., Bozhenkov, S., Brakel, R., Bräuer, T., de Carvalho, B. Brotas, Burhenn, R., Buttenschön, B., Cappa, A., Cseh, G., and Czarnecka, A.

Subjects
SUPERCONDUCTORS, STELLARATORS, GRAPHITE, FUSION reactor limiters, MANOMETERS, DICHROIC filters
Abstract

Wendelstein 7-X, a superconducting optimized stellarator built in Greifswald/Germany, started its first plasmas with the last closed flux surface (LCFS) defined by 5 uncooled graphite limiters in December 2015. At the end of the 10 weeks long experimental campaign (OP1.1) more than 20 independent diagnostic systems were in operation, allowing detailed studies of many interesting plasma phenomena. For example, fast neutral gas manometers supported by video cameras (including one fast-frame camera with frame rates of tens of kHz) as well as visible cameras with different interference filters, with field of views covering all ten half-modules of the stellarator, discovered a MARFE-like radiation zone on the inboard side of machine module 4. This structure is presumably triggered by an inadvertent plasma-wall interaction in module 4 resulting in a high impurity influx that terminates some discharges by radiation cooling. The main plasma parameters achieved in OP1.1 exceeded predicted values in discharges of a length reaching 6 s. Although OP1.1 is characterized by short pulses, many of the diagnostics are already designed for quasi-steady state operation of 30 min discharges heated at 10 MW of ECRH. An overview of diagnostic performance for OP1.1 is given, including some highlights from the physics campaigns. [ABSTRACT FROM AUTHOR]

Published
2016
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14. Aroma Compounds of Arbustus Distillates

Author

Versini, G., Seeber, Renato, Dalla Serra, A., Sferlazzo, G., de Carvalho, B., and Reniero, F.

Subjects
distillates, analytical determination, Aromas, Aromas, analytical determination, distillates
Published
1994

15. Late postoperative myelomalacic myelopathy.

Author

de Carvalho, B, Barros, P, Pereira, P, and Vaz, R

Subjects
*SPINE radiography, *SPINAL surgery, *DIFFERENTIAL diagnosis, *MAGNETIC resonance imaging, *SPINAL cord diseases, *SURGICAL complications, *SPINAL tumors, *ATROPHY, *DIAGNOSIS
Abstract

Study design:This is a case report.Objectives:Spinal cord atrophy presenting with late progressive myelopathy after many years of clinical stability is a rare and unexplained phenomenon after cervical spine surgery. The authors report and discuss the etiologies and outcomes of late postoperative myelomalacic myelopathy.Setting:This study was conducted in the Department of Neurosurgery, Centro Hospitalar S. João, Porto, Portugal.Methods:We report on two patients with insidious chronic progressive myelopathy that developed more than 10 years after complete removal of a cervical intramedullary ependymoma and after transoral odontoidectomy and occipitocervical fusion for craniocervical junction malformation. Differential diagnoses were formulated and review of the literature was performed.Results:In both patients, after several years of clinical stability, insidious onset of debilitating myelopathy and dependency ensued. The clinical history, serology, imaging and neurophysiological investigation excluded several putative etiologies: arachnoid adhesions, tumor recurrence, late vertebral instability, trauma, syringomyelia, radiotherapy, and demyelinating or infectious causes.Conclusion:Late neurological deterioration after cervical spine surgery is usually related to disease progression or surgery-related complications. Nevertheless, in some patients late myelopathy can ensue even in the absence of identified causes. [ABSTRACT FROM AUTHOR]

Published
2015
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22. International Political Thought and Historical International Relations

Author

Liane Hartnett, Or Rosenboim, de Carvalho, B., Costa Lopez, J., and Leira, H.

Subjects
International relations, Politics, Scholarship, Intersection, Political science, JZ, Futures contract, Epistemology
Abstract

This paper critically surveys recent developments in the subfield of Historical International Relations (IR). It looks at the evolution of Historical IR as a particular approach to the study of IR, and examines its development through competing approaches offered by contemporary scholars. The paper suggests that this subfield provides an insightful and innovative branch of scholarship in contemporary IR, which can pave the way to new futures of the discipline. In particular, the chapter explores in depth the various interactions between Historical IR and International Political Thought (IPT), which has recently gained greater prominence in the discipline of History. The paper argues that the dialogue between these two approaches to the study of the international – Historical IR and IPT – can be mutually enriching, but also embodies some methodological limits. Finally, the paper offers some ways forward in the intersection of Historical IR and IPT.

Published
2021

24. Effect of Mutant and Engineered High-Acetate-Producing Saccharomyces cerevisiae var. boulardii Strains in Dextran Sodium Sulphate-Induced Colitis.

Author

Deleu S, Jacobs I, Vazquez Castellanos JF, Verstockt S, Trindade de Carvalho B, Subotić A, Verstockt B, Arnauts K, Deprez L, Vissers E, Lenfant M, Vandermeulen G, De Hertogh G, Verbeke K, Matteoli G, Huys GRB, Thevelein JM, Raes J, and Vermeire S

Subjects
Animals, Female, Mice, Disease Models, Animal, Colon metabolism, Colon microbiology, Colon pathology, Saccharomyces boulardii, Colitis, Ulcerative chemically induced, Colitis, Ulcerative therapy, Colitis, Ulcerative microbiology, Mutation, Gastrointestinal Microbiome, Feces microbiology, Mice, Inbred C57BL, Dextran Sulfate, Acetates, Saccharomyces cerevisiae genetics, Colitis chemically induced, Colitis therapy, Probiotics
Abstract

Acetate-producing Saccharomyces cerevisiae var. boulardii strains could exert improved effects on ulcerative colitis, which here, was preclinically evaluated in an acute dextran sodium sulphate induced model of colitis. Nine-week-old female mice were divided into 12 groups, receiving either drinking water or 2.75% dextran sodium sulphate for 7 days, combined with a daily gavage of various treatments with different levels of acetate accumulation: sham control (phosphate buffered saline, no acetate), non-probiotic control (Baker's yeast, no acetate), probiotic control (Enterol ® , transient acetate), and additionally several Saccharomyces cerevisiae var. boulardii strains with respectively no, high, and extra-high acetate accumulation. Disease activity was monitored daily, and feces samples were collected at different timepoints. On day 14, the mice were sacrificed, upon which blood and colonic tissue were collected for analysis. Disease activity in inflamed mice was lower when treated with the high-acetate-producing strain compared to sham and non-probiotic controls. The non-acetate-producing strain showed higher disease activity compared to the acetate-producing strains. Accordingly, higher histologic inflammation was observed in non- or transient-acetate-producing strains compared to the sham control, whereas this increase was not observed for high- and extra-high-acetate-producing strains upon induction of inflammation. These anti-inflammatory findings were confirmed by transcriptomic analysis of differentially expressed genes. Moreover, only the strain with the highest acetate production was superior in maintaining a stable gut microbial alpha-diversity upon inflammation. These findings support new possibilities for acetate-mediated management of inflammation in inflammatory bowel disease by administrating high-acetate-producing Saccharomyces cerevisae var. boulardii strains.

Published
2024
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25. Influence of physicochemical characteristics of calcium phosphate-based biomaterials in cranio-maxillofacial bone regeneration. A systematic literature review and meta-analysis of preclinical models.

Author

Sadeghian Dehkord E, De Carvalho B, Ernst M, Albert A, Lambert F, and Geris L

Abstract

Objectives: Calcium phosphate-based biomaterials (CaP) are the most widely used biomaterials to enhance bone regeneration in the treatment of alveolar bone deficiencies, cranio-maxillofacial and periodontal infrabony defects, with positive preclinical and clinical results reported. This systematic review aimed to assess the influence of the physicochemical properties of CaP biomaterials on the performance of bone regeneration in preclinical animal models., Methods: The PubMed, EMBASE and Web of Science databases were searched to retrieve the preclinical studies investigating physicochemical characteristics of CaP biomaterials. The studies were screened for inclusion based on intervention (physicochemical characterization and in vivo evaluation) and reported measurable outcomes., Results: A total of 1532 articles were retrieved and 58 studies were ultimately included in the systematic review. A wide range of physicochemical characteristics of CaP biomaterials was found to be assessed in the included studies. Despite a high degree of heterogeneity, the meta-analysis was performed on 39 studies and evidenced significant effects of biomaterial characteristics on their bone regeneration outcomes. The study specifically showed that macropore size, Ca/P ratio, and compressive strength exerted significant influence on the formation of newly regenerated bone. Moreover, factors such as particle size, Ca/P ratio, and surface area were found to impact bone-to-material contact during the regeneration process. In terms of biodegradability, the amount of residual graft was determined by macropore size, particle size, and compressive strength., Conclusion: The systematic review showed that the physicochemical characteristics of CaP biomaterials are highly determining for scaffold's performance, emphasizing its usefulness in designing the next generation of bone scaffolds to target higher rates of regeneration., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Liesbet Geris reports financial support was provided by 10.13039/501100000781European Research Council under the European Union's Horizon Europe programme/ERC Consolidator Grant No. 101088919. Liesbet Geris and France Lambert report financial support was provided by Walloon Region via the BIOWIN-BIOPTOS and Win2Wal-B2Bone projects. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 Published by Elsevier Ltd.)

Published
2024
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26. Biological performance of a novel bovine hydroxyapatite in a guided bone regeneration model: A preclinical study in a mandibular defect in dogs.

Author

De Carvalho B, Dory E, Trus C, Pirson J, Germain L, Lecloux G, Lambert F, and Rompen E

Subjects
Animals, Cattle, Dogs, Biocompatible Materials therapeutic use, Bone Regeneration, Mandible surgery, Minerals, Osteogenesis, Bone Substitutes therapeutic use, Durapatite therapeutic use
Abstract

Objectives: This preclinical model study aims to evaluate the performance and safety of a novel hydroxyapatite biomaterial (Wishbone Hydroxyapatite, WHA) on guided bone regeneration compared to a commercially available deproteinized bovine bone mineral (Bio-Oss, BO)., Material and Methods: Twenty-four beagle dogs were allocated to three timepoint cohorts (4, 12, and 26 weeks) of eight animals each. In all animals, four critical-sized, independent wall mandibular defects were created (32 defects/cohort). Each animal received all four treatments, allocated randomly to separated defects: WHA + collagen membrane (M), BO + M, no treatment (Sham, Sh), and Sh + M. At each timepoint, the specimens were harvested for histologic and histomorphometric analyses to determine the newly formed bone and osteoconductivity., Results: At 4 weeks, bone regeneration was significantly higher for WHA + M (46.8%) when compared to BO + M (21.4%), Sh (15.1%), and Sh + M (23.1%) (p < 0.05); at 12 and 26 weeks, regeneration was similar for WHA and BO. Bone-to-material contact increased over time similarly for WHA + M and BO + M. From a safety point of view, inflammation attributed to WHA + M or BO + M was minimal; necrosis or fatty infiltrate was absent., Conclusions: WHA + M resulted in higher bone regeneration rate than BO + M at 4 weeks. Both BO + M and WHA + M were more efficient than both Sh groups at all timepoints. Safety and biocompatibility of WHA was favorable and comparable to that of BO., (© 2023 Wishbone SA and The Authors. Clinical Implant Dentistry and Related Research published by Wiley Periodicals LLC.)

Published
2024
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28. The Importance of Radiological Patterns and Small Airway Disease in Long-Term Follow-Up of Postacute COVID-19: A Preliminary Study.

Author

Mogami R, Araújo Filho RC, Cobo Chantong CG, Santos de Almeida FC, Baptista Koifman AC, Jauregui GF, Mafort TT, da Silva Bessa da Costa H, Peres Dos Santos GA, Zangerolame de Carvalho B, da Silva Passos G, de Souza Barbosa E, Abalada Ghetti AT, Monnerat LB, Soares da Cal M, Souza Santos Batista DL, Affonso HA, Bousquet GO, Marenco Avila JI, Bento Dutra AL, Leidersnaider CL, Malta da Costa Messeder A, Monteiro A, and Lopes AJ

Abstract

Postacute COVID-19 has become a relevant public health problem, and radiological and pulmonary function tests are tools that help physicians in decision-making. The objectives of this study are to characterize the findings and patterns on a chest radiograph (CXR) and computed tomography (CT) that are most important in the postacute phase and to evaluate how these changes correlate with clinical data, spirometry, and impulse oscillometry (IOS). This was a retrospective study of 29 patients who underwent CXR, CT, spirometry, and IOS. The inclusion criteria were age >18 years and persistent respiratory symptoms after four weeks. The exclusion criteria were radiological exams with low technical quality and non-COVID-19 acute lung diseases. The inferential analysis was carried out with the chi-square ( χ 2 ) or Fisher's exact test to evaluate the interrelationships between the clinical and COVID-19 variables according to spirometry, IOS, CT, and CXR. In our sample, 19 patients were women (65.5%). The predominance of abnormal spirometry was associated with CT's moderate/severe degree of involvement ( p  = 0.017; 69.2%, CI 95%: 44.1%-94.3%). There was no significant association between IOS and tomographic and radiographic parameters. A significant association was found between the classifications of the moderate/severe and normal/mild patterns on CT and CXRs ( p  = 0.003; 93.3%, CI 95%: 77.8%-100%). Patients with moderate/severe impairment on CXR were associated with a higher frequency of hospitalization ( p  = 0.033; 77.8%, CI 95%: 58.6%-97.0%) and had significantly more moderate/severe classifications in the acute phase than the subgroup with normal/mild impairment on CXR ( p  = 0.017; 88.9%, CI 95%: 74.4%-100%). In conclusion, the results of this study show that CXR is a relevant examination and may be used to detect nonspecific alterations during the follow-up of post-COVID-19 patients. Small airway disease is an important finding in postacute COVID-19 syndrome, and we postulate a connection between this pattern and the persistently low-level inflammatory state of the lung., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Roberto Mogami et al.)

Published
2022
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29. Update on the Roles of Oral Hygiene and Plaque Control on Periodontal Disease.

Author

Salhi L, De Carvalho B, and Reners M

Subjects
Humans, Oral Hygiene, Dental Plaque prevention & control, Periodontal Diseases prevention & control, Periodontitis prevention & control
Abstract

Aim: to provide an update of the evidence on the effect of oral hygiene instructions (OHI), dental plaque control and in the prevention and treatment of periodontitis., Methods: Literature searches were performed using MeSH terms, keywords and free words and were published between 2015 and November 2020. The data from the articles were summarized in a narrative review., Results: Data concerning the influence of OHI on periodontal features, the impact of OHI before periodontitis non-surgical treatment, its efficacity on periodontitis prevention and maintenance of healthy periodontium were summarized in the tables of the present narrative review., Conclusion: as prevention is better than a cure, it is relevant to bring in light the role of oral hygiene instructions, the patient self-control of dental plaque as well as the professional mechanical plaque removal in the prevention of periodontitis., (© 2022. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Published
2022
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30. MIG-6 is essential for promoting glucose metabolic reprogramming and tumor growth in triple-negative breast cancer.

Author

He J, Li CF, Lee HJ, Shin DH, Chern YJ, Pereira De Carvalho B, and Chan CH

Subjects
Adaptor Proteins, Signal Transducing genetics, Animals, Cell Line, Tumor, Cell Proliferation genetics, Gene Expression Regulation, Neoplastic, Glucose, Glycolysis genetics, Humans, Mice, Tumor Suppressor Proteins genetics, Triple Negative Breast Neoplasms genetics
Abstract

Treatment of triple-negative breast cancer (TNBC) remains challenging due to a lack of effective targeted therapies. Dysregulated glucose uptake and metabolism are essential for TNBC growth. Identifying the molecular drivers and mechanisms underlying the metabolic vulnerability of TNBC is key to exploiting dysregulated cancer metabolism for therapeutic applications. Mitogen-inducible gene-6 (MIG-6) has long been thought of as a feedback inhibitor that targets activated EGFR and suppresses the growth of tumors driven by constitutive activated mutant EGFR. Here, our bioinformatics and histological analyses uncover that MIG-6 is upregulated in TNBC and that MIG-6 upregulation is positively correlated with poorer clinical outcomes in TNBC. Metabolic arrays and functional assays reveal that MIG-6 drives glucose metabolism reprogramming toward glycolysis. Mechanistically, MIG-6 recruits HAUSP deubiquitinase for stabilizing HIF1α protein expression and the subsequent upregulation of GLUT1 and other HIF1α-regulated glycolytic genes, substantiating the comprehensive regulation of MIG-6 in glucose metabolism. Moreover, our mouse studies demonstrate that MIG-6 regulates GLUT1 expression in tumors and subsequent tumor growth in vivo. Collectively, this work reveals that MIG-6 is a novel prognosis biomarker, metabolism regulator, and molecular driver of TNBC., (© 2021 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

Published
2021
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31. The ubiquitin ligase RNF8 regulates Rho GTPases and promotes cytoskeletal changes and motility in triple-negative breast cancer cells.

Author

Pereira De Carvalho B, Chern YJ, He J, and Chan CH

Subjects
Cell Line, Tumor, DNA-Binding Proteins genetics, Focal Adhesions metabolism, Gene Expression Regulation, Neoplastic, Gene Knockdown Techniques, HeLa Cells, Humans, Proteolysis, Pseudopodia metabolism, Triple Negative Breast Neoplasms genetics, Ubiquitin-Protein Ligases genetics, cdc42 GTP-Binding Protein metabolism, rhoA GTP-Binding Protein metabolism, DNA-Binding Proteins metabolism, Triple Negative Breast Neoplasms metabolism, Ubiquitin-Protein Ligases metabolism, cdc42 GTP-Binding Protein genetics, rhoA GTP-Binding Protein genetics
Abstract

The ubiquitin ligase RNF8 is known to induce epithelial-to-mesenchymal (EMT) transition and metastasis in triple-negative breast cancer (TNBC). Besides EMT, Rho GTPases have been shown as key regulators in metastasis. In this study, we investigated the role of RNF8 in regulating Rho GTPases and cell motility. We find that RNF8 knockdown in TNBC cells attenuates the protein and mRNA levels of Ras homolog family member A (RHOA) and cell division cycle 42 (CDC42). We show that the formation of filopodia, focal adhesions, and the association of focal adhesions to stress fibers is impaired upon RNF8 knockdown. Cell migration is significantly inhibited by RNF8 knockdown. Our study suggests a potential novel role for RNF8 in mediating cell migration in TNBC through regulation of the Rho GTPases RHOA and CDC42., (© 2020 The Authors. FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Published
2021
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32. Survey of Fertility Preservation Options Available to Patients With Cancer Around the Globe.

Author

Rashedi AS, de Roo SF, Ataman LM, Edmonds ME, Silva AA, Scarella A, Horbaczewska A, Anazodo A, Arvas A, Ramalho de Carvalho B, Sartorio C, Beerendonk CCM, Diaz-Garcia C, Suh CS, Melo C, Yding Andersen C, Motta E, Greenblatt EM, Van Moer E, Zand E, Reis FM, Sánchez F, Terrado G, Rodrigues JK, de Meneses E Silva JM, Smitz J, Medrano J, Lee JR, Winkler-Crepaz K, Smith K, Ferreira Melo E Silva LH, Wildt L, Salama M, Del Mar Andrés M, Bourlon MT, Vega M, Chehin MB, De Vos M, Khrouf M, Suzuki N, Azmy O, Fontoura P, Campos-Junior PHA, Mallmann P, Azambuja R, Marinho RM, Anderson RA, Jach R, Antunes RA, Mitchell R, Fathi R, Adiga SK, Takae S, Kim SH, Romero S, Chedid Grieco S, Shaulov T, Furui T, Almeida-Santos T, Nelen W, Jayasinghe Y, Sugishita Y, and Woodruff TK

Subjects
Fertility, Humans, Surveys and Questionnaires, United States, Cancer Survivors, Fertility Preservation, Neoplasms therapy
Abstract

Purpose: Oncofertility focuses on providing fertility and endocrine-sparing options to patients who undergo life-preserving but gonadotoxic cancer treatment. The resources needed to meet patient demand often are fragmented along disciplinary lines. We quantify assets and gaps in oncofertility care on a global scale., Methods: Survey-based questionnaires were provided to 191 members of the Oncofertility Consortium Global Partners Network, a National Institutes of Health-funded organization. Responses were analyzed to measure trends and regional subtleties about patient oncofertility experiences and to analyze barriers to care at sites that provide oncofertility services., Results: Sixty-three responses were received (response rate, 25%), and 40 were analyzed from oncofertility centers in 28 countries. Thirty of 40 survey results (75%) showed that formal referral processes and psychological care are provided to patients at the majority of sites. Fourteen of 23 respondents (61%) stated that some fertility preservation services are not offered because of cultural and legal barriers. The growth of oncofertility and its capacity to improve the lives of cancer survivors around the globe relies on concentrated efforts to increase awareness, promote collaboration, share best practices, and advocate for research funding., Conclusion: This survey reveals global and regional successes and challenges and provides insight into what is needed to advance the field and make the discussion of fertility preservation and endocrine health a standard component of the cancer treatment plan. As the field of oncofertility continues to develop around the globe, regular assessment of both international and regional barriers to quality care must continue to guide process improvements., Competing Interests: The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/jco/site/ifc. Alexandra S. RashediEmployment: Cigna (I) Stock or Other Ownership: Cigna (I)Antoinette AnazodoResearch Funding: Merck SeronoCassio SartorioEmployment: Vida Centro de Fertilidade Leadership: Vida Centro de Fertilidade Stock or Other Ownership: Vida Centro de FertilidadeCatharina C.M. BeerendonkTravel, Accommodations, Expenses: GoodlifeEllen M. GreenblattConsulting or Advisory Role: Ferring Pharmaceuticals, EMD Serono Travel, Accommodations, Expenses: EMD SeronoFernando M. ReisHonoraria: Politec Saúde (I) Consulting or Advisory Role: Politec Saúde (I) Speakers’ Bureau: UCB (I) Travel, Accommodations, Expenses: Abbott Laboratories (I)Johan SmitzSpeakers’ Bureau: Ferring Pharmaceuticals Travel, Accommodations, Expenses: Ferring PharmaceuticalsMaria T. BourlonLeadership: Medivation, Astellas Pharma Honoraria: Medivation, Astellas Pharma Speakers’ Bureau: Asofarma Research Funding: Bristol-Myers Squibb Travel, Accommodations, Expenses: Janssen PharmaceuticalsMichel De VosHonoraria: Cook Medical Research Funding: Cook MedicalRichard A. AndersonConsulting or Advisory Role: Roche, HRA Pharma, NeRe Pharmaceuticals Speakers’ Bureau: Roche, Beckman Coulter, IBSA Institut Biochimque Research Funding: Ferring Pharmaceuticals Travel, Accommodations, Expenses: IBSA Institut BiochimqueRoberto de A. AntunesConsulting or Advisory Role: Merck Serono Speakers’ Bureau: Merck Serono Travel, Accommodations, Expenses: Merck Serono, MSDTeresa Almeida-SantosConsulting or Advisory Role: Merck, MSD Research Funding: MerckTeresa K. WoodruffResearch Funding: Ferring Pharmaceuticals (Inst) No other potential conflicts of interest were reported., (© 2020 by American Society of Clinical Oncology.)

Published
2020
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33. Survey of Third-Party Parenting Options Associated With Fertility Preservation Available to Patients With Cancer Around the Globe.

Author

Rashedi AS, de Roo SF, Ataman LM, Edmonds ME, Silva AA, Scarella A, Horbaczewska A, Anazodo A, Arvas A, Ramalho de Carvalho B, Sartorio C, Beerendonk CCM, Diaz-Garcia C, Suh CS, Melo C, Andersen CY, Motta E, Greenblatt EM, Van Moer E, Zand E, Reis FM, Sánchez F, Terrado G, Rodrigues JK, Marcos de Meneses E Silva J, Smitz J, Medrano J, Lee JR, Winkler-Crepaz K, Smith K, Ferreira Melo E Silva LH, Wildt L, Salama M, Del Mar Andrés M, Bourlon MT, Vega M, Chehin MB, De Vos M, Khrouf M, Suzuki N, Azmy O, Fontoura P, Campos-Junior PHA, Mallmann P, Azambuja R, Marinho RM, Anderson RA, Jach R, Antunes RA, Mitchell R, Fathi R, Adiga SK, Takae S, Kim SH, Romero S, Grieco SC, Shaulov T, Furui T, Almeida-Santos T, Nelen W, Jayasinghe Y, Sugishita Y, and Woodruff TK

Subjects
Humans, Parenting, Referral and Consultation, Surveys and Questionnaires, Fertility Preservation, Neoplasms
Abstract

Purpose: In the accompanying article, "Survey of Fertility Preservation Options Available to Patients With Cancer Around the Globe," we showed that specific fertility preservation services may not be offered at various sites around the world because of cultural and legal barriers. We assessed global and regional experiences as well as the legal status of third-party reproduction and adoption to serve as a comprehensive international data set and resource for groups that wish to begin oncofertility interventions., Methods: We provide data on the legalities of third-party assisted reproductive technologies and other family-building options in the 28 oncofertility-practicing countries surveyed., Results: We found regional and country differences that will be important in the development of tailored resources for physicians and for patient brochures that are sensitive to these local restrictions and cultural norms., Conclusion: Because many patients first consult Web-based materials, the formal assessment of the availability of these options provides members of the global oncofertility community with data to which they might otherwise not have ready access to better serve their patients., Competing Interests: The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or jco.ascopubs.org/site/ifc. Alexandra S. RashediEmployment: Cigna (I) Stock or Other Ownership: Cigna (I)Antoinette AnazodoResearch Funding: Merck SeronoCassio SartorioEmployment: Vida Centro de Fertilidade Leadership: Vida Centro de Fertilidade Stock or Other Ownership: Vida Centro de FertilidadeCatharina C.M. BeerendonkTravel, Accommodations, Expenses: GoodlifeEllen M. GreenblattConsulting or Advisory Role: Ferring Pharmaceuticals, EMD Serono Travel, Accommodations, Expenses: EMD SeronoFernando M. ReisHonoraria: Politec Saúde (I) Consulting or Advisory Role: Politec Saúde (I) Speakers’ Bureau: UCB (I) Travel, Accommodations, Expenses: Abbott Laboratories (I)Flor SánchezPatents, Royalties, Other Intellectual Property: patent pendingJohan SmitzSpeakers’ Bureau: Ferring Pharmaceuticals Travel, Accommodations, Expenses: Ferring PharmaceuticalsMaria T. BourlonLeadership: Medivation, Astellas Pharma Honoraria: Medivation, Astellas PharmaRichard A. AndersonConsulting or Advisory Role: Roche, HRA Pharma, NeRe Pharmaceuticals Speakers’ Bureau: Roche, Beckman Coulter, IBSA Institut Biochimque Research Funding: Ferring Pharmaceuticals Travel, Accommodations, Expenses: IBSA Institut BiochimqueRoberto de A. AntunesConsulting or Advisory Role: Merck Serono Travel, Accommodations, Expenses: Merck Serono, MSDSergio RomeroPatents, Royalties, Other Intellectual Property: patent pendingTeresa Almeida-SantosConsulting or Advisory Role: Merck, MSD Research Funding: Merck SeronoTeresa K. WoodruffResearch Funding: Ferring Pharmaceuticals (Inst) No other potential conflicts of interest were reported., (© 2020 by American Society of Clinical Oncology.)

Published
2020
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34. Effect of Sintering on In Vivo Biological Performance of Chemically Deproteinized Bovine Hydroxyapatite.

Author

De Carvalho B, Rompen E, Lecloux G, Schupbach P, Dory E, Art JF, and Lambert F

Abstract

The influence of the manufacturing process on physicochemical properties and biological performance of xenogenic biomaterials has been extensively studied, but its quantification on bone-to-material contact remains poorly investigated. The aim of this study was to investigate the effect of different heat treatments of an experimental chemically-deproteinized bovine hydroxyapatite in vivo in terms of new bone formation and osteoconductivity. Protein-free hydroxyapatite from bovine origin was produced under sub-critical conditions and then either sintered at 820 °C or 1200 °C. Structural and morphological properties were assessed by scanning electron microscopy (SEM), measurement of surface area and X-ray diffractometry (XRD). The materials were then implanted in standardized alveolar bone defects in minipigs and histomorphometric evaluations were performed using non-decalcified sections. Marked topographical differences were observed by SEM analysis. As the sintering temperature of the experimental material increased, the surface area significantly decreased while crystallite size increased. In vivo samples showed that the highly sintered BHA presented a significantly lower percentage of newly formed bone than the unheated one ( p = 0.009). In addition, the percentage of bone-to-material contact (BMC) was significantly lowered in the highly sintered group when compared to the unsintered ( p = 0.01) and 820 °C sintered ( p = 0.02) groups. Non-sintered or sintered at 820 °C BHA seems to maintain a certain surface roughness allowing better bone regeneration and BMC. On the contrary, sintering of BHA at 1200 °C has an effect on its morphological and structural characteristics and significantly modify its biological performance (osteoconductivity) and crystallinity.

Published
2019
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35. Correlation of Parasite Burden, kDNA Integration, Autoreactive Antibodies, and Cytokine Pattern in the Pathophysiology of Chagas Disease.

Author

Wesley M, Moraes A, Rosa AC, Lott Carvalho J, Shiroma T, Vital T, Dias N, de Carvalho B, do Amaral Rabello D, Borges TKDS, Dallago B, Nitz N, Hagström L, and Hecht M

Abstract

Chagas disease (CD), caused by the protozoan Trypanosoma cruzi ( T. cruzi ), is the main parasitic disease in the Western Hemisphere. Unfortunately, its physiopathology is not completely understood, and cardiomegaly development is hard to predict. Trying to explain tissue lesion and the fact that only a percentage of the infected individuals develops clinical manifestations, a variety of mechanisms have been suggested as the provokers of CD, such as parasite persistence and autoimmune responses. However, holistic analysis of how parasite and host-related elements may connect to each other and influence clinical outcome is still scarce in the literature. Here, we investigated murine models of CD caused by three different pathogen strains: Colombian, CL Brener and Y strains, and employed parasitological and immunological tests to determine parasite load, antibody reactivity, and cytokine production during the acute and chronic phases of the disease. Also, we developed a quantitative PCR (qPCR) protocol to quantify T. cruzi kDNA minicircle integration into the mammalian host genome. Finally, we used a correlation analysis to interconnect parasite- and host-related factors over time. Higher parasite load in the heart and in the intestine was significantly associated with IgG raised against host cardiac proteins. Also, increased heart and bone marrow parasitism was associated with a more intense leukocyte infiltration. kDNA integration rates correlated to the levels of IgG antibodies reactive to host cardiac proteins and interferon production, both influencing tissue inflammation. In conclusion, our results shed light into how inflammatory process associates with parasite load, kDNA transfer to the host, autoreactive autoantibody production and cytokine profile. Altogether, our data support the proposal of an updated integrative theory regarding CD pathophysiology.

Published
2019
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36. Everyday sovereignty: International experts, brokers and local ownership in peacebuilding Liberia.

Author

de Carvalho B, Schia NN, and Guillaume X

Abstract

The present article investigates how sovereignty is performed, enacted and constructed in an everyday setting. Based on fieldwork and interviews with international embedded experts about the elusive meaning of 'local ownership', we argue that while sovereignty may, indeed, be a model according to which the international community 'constructs' rogue or failed polities in 'faraway' places, this view overlooks that these places are still spaces in which contestations over spheres of authority take place every day, and thus also spaces in which sovereignty is constructed and reconstructed on a daily basis. Local ownership, then, becomes our starting point for tracing the processes of the everyday enactment of sovereignty. We make the case that sovereignty should not be reified, but instead be studied in its quotidian and dynamic production, involving the multiplicity of actors reflecting the active production of the state beyond its presumptive existence as a homogeneously organized, institutionalized and largely centralized bureaucracy.

Published
2019
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37. Arsenic, selenium, and metals in a commercial and vulnerable fish from southwestern Atlantic estuaries: distribution in water and tissues and public health risk assessment.

Author

Avigliano E, Maichak de Carvalho B, Invernizzi R, Olmedo M, Jasan R, and Volpedo AV

Subjects
Animals, Argentina, Arsenic analysis, Atlantic Ocean, Brazil, Catfishes, Estuaries, Food Contamination statistics & numerical data, Gills, Metals analysis, Muscles, Risk Assessment, Seafood statistics & numerical data, Selenium analysis, Trace Elements, Water Pollutants, Chemical analysis, Arsenic metabolism, Environmental Monitoring, Metals metabolism, Selenium metabolism, Water Pollutants, Chemical metabolism
Abstract

The anadromous catfish Genidens barbus is a vulnerable and economically important species from the Southwestern Atlantic Ocean. Concentrations of As, Co, Fe, Se, and Zn were determined in water and muscle, gill, and liver of catfish from two southwestern Atlantic estuaries (Brazil and Argentina) and health risk via fish consumption was evaluated. High spatial variability was observed in the metals, As, and Se distribution for both estuaries. Considering all tissues, element concentrations (mg/kg, wet weight) were As = 0.41-23.50, Co = 0.01-2.9, Fe = 2.08-773, Se = 0.15-10.7, and Zn = 3.97-2808). Most of the trace elements tended to be higher in Brazil than in Argentina, except for Co, Fe, Se, and Zn in liver and Fe and Co in muscle and gill, respectively. Arsenic accumulation order was muscle > liver ≥ gill. Only As (muscle) was above the maximum recommended by international guidelines at both estuaries. The target hazard quotient ranged from 0.10 to 1.58, suggesting that people may experience significant health risks through catfish consumption. Supposing that the inorganic/toxic As ranged between 1 and 20% of the total, the recommended maximum intakes per capita bases were 6.1-95 and 8.4-138 kg/year (wet weight) for Brazil and Argentina, respectively. Carcinogenic risk for As intake was within the acceptable range but close to the recommended limit (> 10 -4 ). These results highlights the importance of quantifying the As species in catfish muscle in order to generate more reliable risk estimates.

Published
2019
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38. Polygenic Analysis in Absence of Major Effector ATF1 Unveils Novel Components in Yeast Flavor Ester Biosynthesis.

Author

Holt S, Trindade de Carvalho B, Foulquié-Moreno MR, and Thevelein JM

Subjects
Acyltransferases genetics, Alcoholic Beverages microbiology, Endosomal Sorting Complexes Required for Transport genetics, Fermentation, Metabolic Engineering, Mutant Proteins genetics, Mutant Proteins metabolism, Proteins genetics, Saccharomyces cerevisiae enzymology, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins genetics, Acetates metabolism, Acetyltransferases metabolism, Acyltransferases metabolism, Biosynthetic Pathways genetics, Endosomal Sorting Complexes Required for Transport metabolism, Proteins metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins metabolism, Volatile Organic Compounds metabolism
Abstract

Flavor production in yeast fermentation is of paramount importance for industrial production of alcoholic beverages. Although major enzymes of flavor compound biosynthesis have been identified, few specific mutations responsible for strain diversity in flavor production are known. The ATF1 -encoded alcohol acetyl coenzyme A (acetyl-CoA) transferase (AATase) is responsible for the majority of acetate ester biosynthesis, but other components affecting strain diversity remain unknown. We have performed parallel polygenic analysis of low production of ethyl acetate, a compound with an undesirable solvent-like off-flavor, in strains with and without deletion of ATF1 We identified two unique causative mutations, eat1 K179 fs and snf8 E148 *, not present in any other sequenced yeast strain and responsible for most ethyl acetate produced in absence of ATF1 EAT1 encodes a putative mitochondrial ethanol acetyl-CoA transferase (EATase) and its overexpression, but not that of EAT1 K179 fs , and strongly increases ethyl acetate without affecting other flavor acetate esters. Unexpectedly, a higher level of acetate esters (including ethyl acetate) was produced when eat1 K179 fs was present together with ATF1 in the same strain, suggesting that the Eat1 and Atf1 enzymes are intertwined. On the other hand, introduction of snf8 E148 * lowered ethyl acetate levels also in the presence of ATF1 , and it affected other aroma compounds, growth, and fermentation as well. Engineering of snf8 E148 * in three industrial yeast strains (for production of wine, saké, and ale beer) and fermentation in an application-relevant medium showed a high but strain-dependent potential for flavor enhancement. Our work has identified EAT1 and SNF8 as new genetic elements determining ethyl acetate production diversity in yeast strains. IMPORTANCE Basic research with laboratory strains of the yeast Saccharomyces cerevisiae has identified the structural genes of most metabolic enzymes, as well as genes encoding major regulators of metabolism. On the other hand, more recent work on polygenic analysis of yeast biodiversity in natural and industrial yeast strains is revealing novel components of yeast metabolism. A major example is the metabolism of flavor compounds, a particularly important property of industrial yeast strains used for the production of alcoholic beverages. In this work, we have performed polygenic analysis of production of ethyl acetate, an important off-flavor compound in beer and other alcoholic beverages. To increase the chances of identifying novel components, we have used in parallel a wild-type strain and a strain with a deletion of ATF1 encoding the main enzyme of acetate ester biosynthesis. This revealed a new structural gene, EAT1 , encoding a putative mitochondrial enzyme, which was recently identified as an ethanol acetyl-CoA transferase in another yeast species. We also identified a novel regulatory gene, SNF8 , which has not previously been linked to flavor production. Our results show that polygenic analysis of metabolic traits in the absence of major effector genes can reveal novel structural and regulatory genes. The mutant alleles identified can be used to affect the flavor profile in industrial yeast strains for production of alcoholic beverages in more subtle ways than by deletion or overexpression of the already known major effector genes and without significantly altering other industrially important traits. The effect of the novel variants was dependent on the genetic background, with a highly desirable outcome in the flavor profile of an ale brewing yeast., (Copyright © 2018 Holt et al.)

Published
2018
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39. Identification of Novel Alleles Conferring Superior Production of Rose Flavor Phenylethyl Acetate Using Polygenic Analysis in Yeast.

Author

Trindade de Carvalho B, Holt S, Souffriau B, Lopes Brandão R, Foulquié-Moreno MR, and Thevelein JM

Subjects
Acetates chemistry, Alcohols chemistry, Chromosome Mapping, Fatty Acid Synthases genetics, Flavoring Agents metabolism, Mutation, Phenylethyl Alcohol analogs & derivatives, Phenylethyl Alcohol chemistry, Phosphatidylinositol 3-Kinases genetics, Saccharomyces cerevisiae enzymology, Saccharomyces cerevisiae Proteins genetics, Saccharomyces cerevisiae Proteins metabolism, Acetates metabolism, Alleles, Phenylethyl Alcohol metabolism, Quantitative Trait Loci, Rosa chemistry, Saccharomyces cerevisiae genetics
Abstract

Flavor compound metabolism is one of the last areas in metabolism where multiple genes encoding biosynthetic enzymes are still unknown. A major challenge is the involvement of side activities of enzymes having their main function in other areas of metabolism. We have applied pooled-segregant whole-genome sequence analysis to identify novel Saccharomyces cerevisiae genes affecting production of phenylethyl acetate (2-PEAc). This is a desirable flavor compound of major importance in alcoholic beverages imparting rose- and honey-like aromas, with production of high 2-PEAc levels considered a superior trait. Four quantitative trait loci (QTLs) responsible for high 2-PEAc production were identified, with two loci each showing linkage to the genomes of the BTC.1D and ER18 parents. The first two loci were investigated further. The causative genes were identified by reciprocal allele swapping into both parents using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9. The superior allele of the first major causative gene, FAS2 , was dominant and contained two unique single nucleotide polymorphisms (SNPs) responsible for high 2-PEAc production that were not present in other sequenced yeast strains. FAS2 encodes the alpha subunit of the fatty acid synthetase complex. Surprisingly, the second causative gene was a mutant allele of TOR1 , a gene involved in nitrogen regulation. Exchange of both superior alleles in the ER18 parent strain increased 2-PEAc production 70%, nearly to the same level as in the best superior segregant. Our results show that polygenic analysis combined with CRISPR/Cas9-mediated allele exchange is a powerful tool for identification of genes encoding missing metabolic enzymes and for development of industrial yeast strains generating novel flavor profiles in alcoholic beverages. IMPORTANCE Multiple reactions in flavor metabolism appear to be catalyzed by side activities of other enzymes that have been difficult to identify. We have applied genetic mapping of quantitative trait loci in the yeast Saccharomyces cerevisiae to identify mutant alleles of genes determining the production of phenylethyl acetate, an important flavor compound imparting rose- and honey-like aromas to alcoholic beverages. We identified a unique, dominant allele of FAS2 that supports high production of phenylethyl acetate. FAS2 encodes a subunit of the fatty acid synthetase complex and apparently exerts an important side activity on one or more alternative substrates in flavor compound synthesis. The second mutant allele contained a nonsense mutation in TOR1 , a gene involved in nitrogen regulation of growth. Together the two alleles strongly increased the level of phenylethyl acetate. Our work highlights the potential of genetic mapping of quantitative phenotypic traits to identify novel enzymes and regulatory components in yeast metabolism, including regular metabolic enzymes with unknown side activities responsible for biosynthesis of specific flavor compounds. The superior alleles identified can be used to develop industrial yeast strains generating novel flavor profiles in alcoholic beverages., (Copyright © 2017 Trindade de Carvalho et al.)

Published
2017
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40. Synthesis of supermacroporous cryogel for bioreactors continuous starch hydrolysis.

Author

Guilherme EPX, de Oliveira JP, de Carvalho LM, Brandi IV, Santos SHS, de Carvalho GGP, Cota J, and Mara Aparecida de Carvalho B

Subjects
Aspergillus oryzae, Enzymes, Immobilized chemistry, Fungal Proteins chemistry, Fungal Proteins metabolism, Hydrogen-Ion Concentration, Hydrolysis, Maltose metabolism, Porosity, Starch analysis, Starch chemistry, Temperature, alpha-Amylases chemistry, Bioreactors, Cryogels chemistry, Enzymes, Immobilized metabolism, Starch metabolism, alpha-Amylases metabolism
Abstract

A bioreactor was built by means of immobilizing alpha-amylase from Aspergillus oryzae by encapsulation, through cryopolymerization of acrylamide monomers for the continuous starch hydrolysis. The starch hydrolysis was evaluated regarding pH, the concentration of immobilized amylase on cryogel, the concentration of starch solution and temperature. The maximum value for starch hydrolysis was achieved at pH 5.0, concentration of immobilized enzyme 111.44 mg amylase /g cryogel , concentration of starch solution 45 g/L and temperature of 35°C. The immobilized enzyme showed a conversion ratio ranging from 68.2 to 97.37%, depending on the pH and temperature employed. Thus, our results suggest that the alpha-amylase from A. oryzae immobilized on cryogel monoliths represents a potential process for industrial production of maltose from starch hydrolysis., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Published
2017
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41. Barriers to investigating and reporting nosocomial outbreaks to health authorities in São Paulo, Brazil: a mixed methods approach.

Author

Maciel AL, de Carvalho BA, Timmons S, and Padoveze MC

Subjects
Attitude of Health Personnel, Brazil, Humans, Infection Control Practitioners, Practice Patterns, Nurses', Practice Patterns, Physicians', Surveys and Questionnaires, Cross Infection epidemiology, Disease Notification, Disease Outbreaks
Abstract

Background: Not all nosocomial outbreaks (NOs) are reported to health authorities (HAs)., Aim: To identify barriers to investigating and reporting NOs to HAs., Methods: A mixed methods approach was performed with a convergent parallel design. The quantitative and qualitative branches of the study were a statewide (electronic) survey and focus groups (FGs), respectively. Infection control practitioners (ICPs) working in the State of São Paulo, Brazil were recruited., Findings: Eighty-five ICPs were enrolled in the survey and 22 ICPs were enrolled in the FGs. Barriers to investigating and reporting NOs included: (i) difficulty in translating outbreak investigation knowledge into practice; (ii) weak planning in outbreak investigation process; (iii) organizational culture and context; (iv) lack of awareness about reporting; and (v) lack of autonomy of ICPs to report outbreaks to HAs., Conclusion: HAs could overcome these barriers by revising their strategies to work with healthcare services, as well as delivering translational educational programmes to support improvement in knowledge and skills for NO investigation., (Copyright © 2016 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Published
2016
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42. Ovarian reserve assessment for infertility investigation.

Author

Ramalho de Carvalho B, Gomes Sobrinho DB, Vieira AD, Resende MP, Barbosa AC, Silva AA, and Nakagava HM

Abstract

The current trends to postpone motherhood and the increase in demand for assistance in reproductive medicine highlight the need for seeking guidelines for the establishment of individualized treatment protocols. Currently available ovarian reserve tests do not provide sufficient evidence to be solely considered ideal, but they may occupy important place in initial counseling, predicting unsatisfactory results that could be improved by individualized induction schemes and reducing excessive psychological and financial burdens, and adverse effects. In this paper, we revise the role of hormonal basal and dynamic tests, as well as ultrasonographic markers, as ovarian reserve markers, in order to provide embasement for propaedeutic strategies and their interpretation in order to have reproductive success.

Published
2012
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43. Anti-müllerian hormone is the best predictor of poor response in ICSI cycles of patients with endometriosis.

Author

de Carvalho BR, Rosa-e-Silva AC, Rosa-e-Silva JC, dos Reis RM, Ferriani RA, and de Sá MF

Subjects
Endometriosis complications, Endometriosis therapy, Female, Humans, Infertility, Female etiology, Infertility, Male, Male, ROC Curve, Anti-Mullerian Hormone blood, Endometriosis blood, Follicle Stimulating Hormone blood, Ovary physiology, Ovulation Induction methods, Sperm Injections, Intracytoplasmic methods
Abstract

Purpose: To correlate ovarian reserve (OR) markers with response in assisted reproduction techniques (ART) and determine their ability to predict poor response among patients with endometriosis (EDT)., Methods: We evaluated ART cycles of 27 women with EDT and 50 with exclusive male factor. Basal follicle stimulating hormone (FSH) and anti-müllerian hormone (AMH) levels were determined. Ovarian response to gonadotropin stimulation was assessed and correlation coefficients calculated between the variables and reserve markers. Areas under the curve (AUC) determined ability of tests to predict poor response., Results: AMH was significantly correlated with response in both groups and it was the only marker with significant discriminative capacity to predict poor response among EDT (AUC = 0.842; 95% CI: 0.651-0.952) and control group (AUC = 0.869; 95% CI: 0.743-0.947)., Conclusion: Infertile patients with endometriosis can benefit from the pre-therapeutic assessment of OR markers. However, regardless of disease presence, only AMH predicts poor response to stimulus.

Published
2011

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