The Bunyaviridae family is one of the largest among RNA viruses, comprising more than 350 serologically distinct viruses. The family is classified into five genera, Orthobunyavirus, Hantavirus, Nairovirus, Phlebovirus, and Tospovirus. Orthobunyaviruses, nairoviruses and phleboviruses are maintained in nature by a propagative cycle involving blood-feeding arthropods and susceptible vertebrate hosts. Like most arthropod-borne viruses, bunyavirus replication causes little damage to the vector, whereas infection of the mammalian host may lead to death. This situation is mimicked in the laboratory: in cultured mosquito cells no cytopathology is observed and a persistent infection is established, whereas in cultured mammalian cells orthobunyavirus infection is lytic and leads to cell death. Bunyaviruses encode four common structural proteins: an RNA-dependent RNA polymerase, two glycoproteins (Gc and Gn), and a nucleoprotein N. Some viruses also code for nonstructural proteins called NSm and NSs. The NSs protein of the prototype bunyavirus, Bunyamwera virus, seems to be one of the factors responsible for the different outcomes of infection in mammalian and mosquito cell lines. However, only limited information is available on the growth of bunyaviruses in cultured mosquito cell lines other than Aedes albopictus C6/36 cells. Here, I compared the replication of Bunyamwera virus in two additional Aedes albopictus cell clones, C7-10 and U4.4, and two Aedes aegypti cell clones, Ae and A20, and investigated the impact of virus replication on cell function. In addition, whereas the vertebrate innate immune response to arbovirus infection is well studied, relatively little is known about mosquitoes’ reaction to these infections. I investigated the immune responses of the different mosquito cells to Bunyamwera virus infection, in particular antimicrobial signaling pathways (Toll and IMD) and RNA interference (RNAi). The data obtained in U4.4 cells suggest that NSs plays an important role in the infection of mosquitoes. Moreover infection of U4.4 cells more closely resembles infection in Ae and A20 cells and live Aedes aegypti mosquitoes. My data showed that the investigated cell lines have various properties, and therefore they can be used to study different aspects of mosquito-virus interactions.

M.Tech. (Food Technology) / Coeliac disease is an autoimmune disease triggered by the ingestion of gluten; persons suffering from coeliac disease are compelled to follow a life-long gluten-free diet. Gluten-free bread, (GFB), has poor quality attributes compared to wheat bread. The effect of mageu, a traditional beverage on quality parameters of GFB with and without selected hydrocolloids was studied. Mageu produced from maize flour and commercial starter cultures were used in GFB based on sorghum, soybean flour and maize starch. It is hypothesized that mageu with or without hydrocolloids could improve GFB quality aspects. The quality parameters measured were specific volume, loaf height, bake loss, rheological attributes, crumb firmness, firming rate, onset of mould growth and sensory attributes: texture, crumb colour, crust colour, flavour and overall acceptability.....

Photocopy of typescript. / Digitized by Kansas Correctional Industries

Made available in DSpace on 2014-06-11T19:31:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-02-23Bitstream added on 2014-06-13T19:01:10Z : No. of bitstreams: 1 antonio_t_dr_araiq.pdf: 685450 bytes, checksum: 38953886194d585fde26a8d2130709e0 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Mutantes foram obtidos através de tratamento mutagênico dos esporos de S. clavuligerus ATCC 27064 com metil-metanosulfonato. Um total de 822 colônias, obtidas por repiques sucessivos, foram selecionadas em testes qualitativos e/ou quantitativos. O melhor mutante S. clavuligerus 45.41, mostrou-se de duas a quatro vezes mais produtivo que a linhagem selvagem, e manteve-se estável após 35 repiques sucessivos ao longo deste estudo. O comportamento das linhagens foi investigado pela adição de diferentes diaminas ao meio de cultivo, uma a uma, na ausência de lisina. Também se investigou duas fontes distintas de C, além de compostos como o ácido 2,6- diaminopimélico. Um planejamento de dois fatores (cadaverina e lisina), três níveis, baseado no ponto central, faces centradas, indicou que a adição de lisina aumenta a produção de CefC para ambas as linhagens. O efeito positivo da cadaverina foi observado principalmente na linhagem mutante 45.41. Resultados obtidos em processo de batelada, em biorreator de bancada confirmaram as diferenças observadas entre as linhagens nos cultivos em frascos agitados. Procedimento de mutagênese clássica foi utilizado em conjunto com critérios bem definidos para se estabelecer um meio de cultura apropriado, com o objetivo de alcançar um aumento significativo da produção de CefC. Este é o primeiro estudo utilizando um mutante de S. clavuligerus obtido por mutagênese clássica, cuja produção de CefC é de três vezes maior que a da linhagem selvagem, em meios contendo diaminas / Mutants were obtained by treating S. clavuligerus ATCC 27064 spores with methyl-methanesulfonate. A total of 822 colonies, obtained by successive sampling, were selected by qualitative and/or quantitative tests. The best mutant, S. clavuligerus 45.41, was two to four times more productive than the wild-type strain, and remained stable even after 35 successive samplings throughout the study. Strains behavior was investigated by adding different diamines in media, one by one, in the absence of lysine. Also investigated two different sources of C, and compounds such as 2,6-diaminopimelic acid. The two-factor (cadaverine and lysine), three-level, central composite-based, face-centered experimental design indicates that adding lysine increases CephC production for both strains alike. The positive effect of cadaverine was observed mainly in the Mutant 45.41 strain. Results obtained in batch-processes in a bench-scale bioreactor confirmed differences among strains observed in shaken flasks cultures. Classical mutagenesis procedures in conjunction with the adoption of well-defined criteria to establish an appropriate culture medium promoted a significant improvement in CephC production. It is the first study indicating an increase in CephC production in media containing diamines employing a mutant of S. clavuligerus obtained by classical mutagenesis

Bacteria are inarguably the most ubiquitous and adaptive organisms on the planet. The vast, diverse community of microbes residing in soil are mostly studied using sequencing technologies because over 99% of them are currently uncultivable in the laboratory. This lack of diverse bacterial cultivation presents a serious challenge for modern microbiological and medical science where the discovery of novel antibiotic producers and microbial products has been outpaced by the rise in drug resistance. This study designed and tested two new cost-effective culture systems called the “Dynamic Soil Environment” and Soil Extract Systems with the goal of increasing the cultivable communities of diverse bacteria in a soil sample over standard methods. Illumina MiSeq sequencing and DADA2 pipeline protocols were used to analyze community DNA from cultivated samples and source soil metagenomes. Autoclaved soil extract media in the Soil Extract Experiment yielded a statistically significantly greater Shannon’s (p = 0.008) and Simpson’s diversity (p = 0.007) of bacteria over pH modified (6.4) nutrient agar media over 30 days of incubation. Autoclaved soil extract media was also able to cultivate, on average, 33% of species in bulk soil sequences compared to 27% from standard nutrient agar however these differences weren’t statistically significant. The length of incubation had a lesser effect than media type on yield of bacteria over 30 days in batch culture conditions. Species richness and diversity generally decreased over time except in soil extract samples. In the Dynamic Soil Environment experiment, membrane plates placed on a live soil environment produced a slightly higher diversity than autoclaved membrane plates and control plates without soil, however, these differences were not statistically significant except when analyzed with Chao1 diversity (0.041). Cultivated bacterial diversity and communities differed more according to media type than soil environment with statistically significant differences between standard and pH modified nutrient agar. Media with a 5.8 pH buffer produced a significantly higher relative abundance of the well-known antibiotic-producers, Actinobacteria (t(10) = -5.715, p < .000) and also Proteobacteria (t(10) = -10.127, p < .000). This study establishes cost-effective methods of cultivating more diverse bacterial communities for low-funded laboratories. Culture conditions for the reliable cultivation of higher relative abundances of bacterial groups belonging to Actinobacteria and Proteobacteria are also established with the Dynamic Soil Environment Experiment. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2019. / FAU Electronic Theses and Dissertations Collection

Mouse embryo cells cultured in vitro in serum-supplemented media undergo growth crisis, resulting in the loss of genomically normal cells prior to the appearance of established, aneuploid cell lines. I used the technique of serum-free cell culture to develop a serum-free mouse embryo (SFME) cell line in which serum was replaced by a set of defined supplements. SFME cells, cultured in a nutrient medium supplemented with insulin, transferrin, epidermal growth factor (EGF), high-density lipoprotein (HDL), and fibronectin, have maintained a diploid karyotype with no detectable chromosomal abnormalities for more than 200 generations. The cells did not undergo growth crisis and remain in culture today. SFME cells were dependent on EGF for survival and were reversibly growth inhibited by serum or platelet-free plasma. Treatment of SFME cells with serum or transforming growth factor beta led to the appearance of glial fibrillary acid protein (GFAP), a specific marker for astrocytes, identifying SFME cells as proastroblasts. Following the derivation of SFME cells my research focussed on (1) defining more precisely the growth response of SFME cells to medium supplements, (2) investigating the relationship between the nonsenescent nature of SFME cells and their responses to serum and EG1., and (3) applying the serum-free cell culture methods to the multipassage culture of human embryonic astrocytes. SFME cells in serum-containing medium arrested in the G1 phase of the cell cycle with greatly reduced DNA replication activity. A portion of the inhibitory activity of serum was extracted by charcoal, a procedure that removed steroid and thyroid hormones. However, the effect of serum on untransformed SFME cells could not be prevented by addition of antiglucocorticoid, and ras-transformed clones of SFME cells, which are not growth inhibited by serum, retained inhibitory responses to glucocorticoid and thyroid hormone T3. These results suggest that glucocorticoid or thyroid hormones may contribute to the inhibitory activity of serum on SFME cells, but additional factors are involved. SFME cell death resulting from EGF deprivation exhibited characteristics associated with apoptosis or programmed cell death. Ultrastructural analysis showed cells became small and vacuolated, with pyknotic nuclei. The cultures contained almost exclusively G1- phase cells. Chromatin exhibited a pattern of degradation into oligonucleosome-length fragments generating a regularly spaced ladder. I applied the serum-free approach used to derive SFME cells to the multipassage culture of human embryonic astrocytes. Cells were cultured in nutrient medium supplemented with insulin, transferrin, EGF, HDL, fibronectin, basic fibroblast growth factor and heparin. Cultures were maintained for a maximum of 70 population doublings before proliferation ceased. The cells synthesized GFAP. / Graduation date: 1992

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Thesis (Ph. D.)--University of Hawaii at Manoa, 1990. / Includes bibliographical references (leaves 68-76) / Microfiche. / xi, 76 leaves, bound ill. 29 cm