Existe uma crescente demanda por alternativas de controle de doenças para se reduzir o uso do controle químico, como por exemplo, o estudo da indução de resistência em plantas, através de fitotoxinas como a taxtomina A e outras substâncias. O vírus do mosaico do pepino (Cucumber mosaic virus - CMV) e o vírus do mosaico amarelo da abobrinha (Zucchini yellow mosaic virus - ZYMV) têm causado grandes prejuízos em culturas importantes, assim como o fungo Rhizopus stolonifer, que pode atacar diversos hospedeiros. Este trabalho foi conduzido com o objetivo de se avaliar os efeitos do uso da taxtomina A como indutor de resistência em plantas. Como resultado preliminar a produção de taxtomina A pode ser aumentada utilizando-se como fonte de inóculo amostra de uma cultura liquida pré-estabelecida de Streptomyces scabies. Foram testados a aspersão de taxtomina A em plantas de abobrinha de moita, para se avaliar a proteção contras os vírus CMV e ZYMV. Também foi aspergida taxtomina A em bagas de uva \"Itália\" para se avaliar o desenvolvimento da podridão pós-colheita causada por R. stolonifer. As plantas de abobrinha de moita, mantidas em casa de vegetação, tratadas com taxtomina A não apresentaram sintomas de mosaico característico do CMV. O teste ELISA apresentou resultados negativos indicando a não presença de partículas virais em níveis detectáveis nas plantas. Com relação ao ZYMV, a taxtomina A não foi capaz de controlar o desenvolvimento da doença, pois apareceram sintomas em todas as plantas. A taxtomina A apresentou efeito in vitro sobre R. stolonifer retardando o crescimento do fungo principalmente nas duas maiores concentrações usadas. Porém, a aplicação de taxtomina A não foi capaz de proteger as bagas de uva contra o patógeno. Finalizando a taxtomina A foi capaz de controlar em 100% a incidência de CMV em abobrinha, porém não foi capaz de controlar o ZYMV em abobrinha e a podridão nas uvas causada por R. stolonifer. / There is an increasing demand for alternative control of diseases to reduce the use of chemical control, for example the induced resistance studies on plants with phytotoxins like thaxtomin A and other substances. The Cucumber mosaic virus (CMV) and the Zucchini yellow mosaic virus (ZYMV) cause a lot of losses on important crops and also the fungus Rhizopus stolonifer that can cause disease in different hosts. This work was carried out to evaluate the use of thaxtomin A to induce resistance. Thus, as a preliminary result, the thaxtomin A production could be increased by using as inoculum an established Streptomyces scabies liquid culture. It was tested the spraying of thaxtomin A on zucchini plants to evaluate the protection against the viruses CMV e ZYMV. On the other hand, grape berries cv \"Italia\" were also sprayed with thaxtomin A to evaluate the development of the postharvest rot caused by R. stolonifer. The zucchini plants were maintained in the greenhouse and the treatment with thaxtomin A inhibited mosaic symptom development by the CMV. The ELISA test showed negative results indicating that there was no detectable virus particles inside the plants. The thaxtomin A was not able to control the ZYMV development since all plants exhibited symptoms. The thaxtomin A showed in vitro control of R. stolonifer as it reduced the fungal growing at the two highest concentrations used. However, the thaxtomin A was not able to control the postharvest rot in grapes. Finally, the thaxtomin A was able of controlling 100% of the incidence of CMV on zucchini plants, but was not able of controlling ZYMV in zucchini and the grape berrie rot caused by R. stolonifer.

Rhizopus oryzae is a prominent strain for producing fumarate, where biomass growth precedes fumarate production. The natural bio film growth of R. oryzae as fungal mat was investigated using different glucose addition strategies in a novel fed-batch fermenter. Batch growth was compared through three fed-batch runs, each with a different glucose addition strategy. The fed-batch runs involved a constant glucose feed (CGF) of 0.075 g h-1 and controlled glucose feeds in order to keep the respiration quotient (RQ) at either 1.3 mol CO2 mol-1 O2 (RQ1.3) or 1.1 mol CO2 mol-1 O2 (RQ1.1). Ethanol overflow via the established Crabtree mechanism was completely negated for the CGF and RQ1.1 runs, while the batch and RQ1.3 runs exhibited significant ethanol formation. Biomass yield on glucose was found to be 0.476 g g-1 (RQ1.1), 0.194 g g-1 (RQ1.3), 0.125 g g-1 (CGF) and 0.144 g g-1 (batch). The results indicate a three-fold improvement in biomass yield when comparing the batch run with the RQ1.1 run. In addition, the RQ1.1 run resulted in zero detectable byproducts, unlike the batch scenario where pyruvate and fumarate were associated with ethanol formation. Clear evidence is provided that glucose overflow can be fully eliminated during R. oryzae growth, significantly affecting the biomass yield on glucose. / Dissertation (MEng (Chemical Engineering))--University of Pretoria, 2021. / University of Pretoria postgraduate bursary / CSIR Inter-bursary Programme / Chemical Engineering / MEng (Chemical Engineering) / Unrestricted

Predmetom tejto diplomovej práce bolo štúdium vplyvu detergentov na aktivitu, termostabilitu a agregáciu voľnej a imobilizovanej formy komerčného preparátu lipázy izolovanej z mikroskopickej huby Rhizopus arrhizus. Teoretická časť obsahuje ucelenú rešerš popisujúcu štruktúru, mechanizmus účinku a priemyselný význam spomínanej hydrolázy spolu s popisom chemických účinkov detergentov, pričom dôraz bol kladený predovšetkým na skupinu neionogénnych detergentov s názvom tweeny. V experimentálnej časti bol študovaný efekt tweenov na rozpustnej a imobilizovanej forme RA lipázy. Imobilizácia spočívala v priamej adsorpcii enzýmu na neupravený nosič. Ako nosič bol použitý oxidovaný grafén ošetrený tweenom (tween 20, 60, 80). Aktivita enzýmu bola stanovená spektrofotometricky za pomoci substrátu p-nitrofenyl laurátu. Zvýšenie aktivity voľnej lipázy (104 % oproti maximálnej hodnote) bolo zaznamenané pri použití tweenu 20 o koncentrácii vysoko nad hodnotou kritickej micelárnej koncentrácie (10 mmol•dm-3). Na základe štúdie imobilizačných podmienok, boli nastavené ideálne parametre pre dosiahnutie účinnej imobilizácie v spojení s čo najvyššou lipolytickou aktivitou (koncentrácia enzýmu 0,1 mg•ml-1, fosfátový tlmivý roztok pH 7,2, koncentrácia tweenu 10,8 mmol•dm-3, čas imobilizácie 1 hodina). Obe formy lipázy vykazovali maximálnu aktivitu pri 35 °C. Optimálne pH sa u imobilizovanej lipázy posunulo na hodnotu 8, v porovnaní s voľnou formou, ktorej pH optimum bolo stanovené na 9. Tepelná stabilita vykazovala približne rovnaký priebeh u oboch foriem skúmanej hydrolázy. Avšak v prípade štúdia stability enzýmu pri dlhodobej úschove bolo po imobilizácii zistené výrazné zlepšenie tohto parametru.

The ultrastructural aspects of Rhizopus stolonifer (Fr.) Lind. Infection in Ipomoea batatus Lam. Roots and the histochemistry of cell walls affected by enzymes secreted by the fungus were investigated. Specimens were prepared for viewing in the electron microscope by thin-sectioning techniques. Degradation of cellular membreanes and host tissues in advance of the fungus was evident. Breakdown of the middle lamella was detected by special histochemical stains for pectin. Breakdown occurred in the following was: (1) maceration of the middle lamella before maceration of the cell wall, (2) simultaneous degradation of both the middle lamella and the cell wall, (3) maceration of the cell wall before maceration of the middle lamella. A difference in electron-opacity in health, non-infected host walls and infected host walls was observed. Irregularity in the infection front and the host tissues by various enzymes was observed.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Neste trabalho investigou-se o potencial biocatalÃtico do fungo Rhizopus stolonifer um fungo fitopatogÃnico em biotransformaÃÃes de metabÃlitos secundÃrios naturais Dois desses metabolitos, o diterpeno 3,12-dioxo-15,16-epoxi-4-hidroxicleroda-13(16),14-dieno e o derivado fenÃlico 6-gingerol foram biotransformados e seus respectivos produtos isolados e quantificados por CLAE A elucidaÃÃo estrutural dos produtos de biotransformaÃÃo pelo fungo Rhizopus stolonifer foram realizadas por anÃlise dos espectros de RMN 1H RMN 13C RMN 13C-DEPT 135 IV e EM Identificou-se que os produtos obtidos eram provenientes de uma biorreduÃÃo revelando o potencial deste fungo na reduÃÃo quimio e regiosseletivas de grupos carbonilas nÃo conjugados Realizou-se o estudo cinÃtico dos produtos de biorreduÃÃo por CLAE e ensaios citotoxicidade frente Ãs linhagens tumorais humanas OVCAR-8 (ovÃrio) SF-295 (glioblastoma) e HCT-8 (cÃlon) O produto biorreduzido 6-gingerdiol apresentou os melhores percentuais de inibiÃÃo do crescimento celular: 91,83; 70,90 e 78,56 respectivamente / In this work the biocatalytic potential of the fungus Rhizopus stolonifer a phytopathogenic fungus in biotransformations of natural secondary metabolites was investigated Two of these metabolites the diterpene 3,12-dioxo-15,16-epoxy-4-hydroxycleroda-13(16),14-diene and the phenolic derivative 6-gingerol was biotransformed and their products was isolated and quantified by HPLC The structural determination of biotransformation products was obtained by analysis of 1H NMR 13C NMR 13C-NMR DEPT 135 IV and MS spectrum The products were obtained by a biorreduction process revealing the potential of this fungus in selective reduction of unconjugated carbonyl groups The products from bioreduction were submitted to antitumor assay against human tumor cell lines OVCAR-8 (ovarian) SF-295 (glioblastoma) and HCT-8 (colon) The biotransformation product 6-gingerdiol showed the highest percentage of inhibition of cell growth: 91,83; 70,90 e 78,56, respectively

Submitted by Maike Costa (maiksebas@gmail.com) on 2017-09-06T12:24:40Z No. of bitstreams: 1 arquivototal.pdf: 1307797 bytes, checksum: 1ff0ad27389ffed5bd00e502e9eaedf8 (MD5) / Made available in DSpace on 2017-09-06T12:24:41Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1307797 bytes, checksum: 1ff0ad27389ffed5bd00e502e9eaedf8 (MD5) Previous issue date: 2016-04-27 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Cherry tomato (Lycopersicon esculentum L.) fruit are higly perishable. The infection of cherry tomato fruit with the pathogenic fungi Aspergillus niger and Rhizopus stolonifer is associated with high postharvest losses and decreased quality during storage. This study evaluated the efficacy of an edible coating comprising chitosan (CHI; 4 mg/mL) and Origanum vulgare L. essential oil (OVEO; 1.25 μL/mL) for keeping the quality of cherry tomato fruit during storage at room (25 °C; 12 days) and cold (12 °C; 24 days) temperatures, considering the physical, physicochemical and sensory aspects. The minimum inhibitory concentrations (MIC) of CHI and OVEO were 4 mg/mL and 10 μL/mL respectively, against R. stolonifer and A. niger. The incorporation of CHI 4 mg/ml and OVEO 5 μL/mL, 2.5 μL/mL and 1.25 μL/mL in the growth media showed fungicidal effect and strongly inhibited the germination of spores, mycelial growth and sporulation of fungal strains test. CHI-OVEO coating reduced black mold and soft-rot incidence caused by these fungi in artificially contaminated cherry tomato fruit during storage at both temperatures. Fruit coated with CHI-OVEO showed higher firmness and lower weigth loss. CHI-OVEO coating delayed the decrease of lycopene, ascorbic and citric acid, glucose and fructose during storage at room or cold temperatures. Fruit coated with CHI-OVEO showed increased amounts of catechin, myricetin, caffeic and syringic acids at the end of storage. CHI-OVEO coating is a feasible treatment for keep the storage quality of cherry tomato fruit. These results indicate that the CHI-OVEO edible coating is a viable treatment for maintaining quality during storage cherry tomatoes. / Tomates cereja (Lycopersicon esculentum L.) são frutos altamente perecíveis. A contaminação destes frutos com os fungos patogênicos Aspergillus niger e Rhizopus stolonifer é associada à elevadas perdas pós-colheita e à redução da qualidade durante o armazenamento. Este estudo avaliou a eficácia de um revestimento comestível composto de quitosana (QUI; 4 mg/mL) e óleo essencial de Origanum vulgare L. (OEOV; 1.25 μL/mL) na manutenção da qualidade de frutos de tomate cereja durante o armazenamento em temperatura ambiente (25 °C; 12 dias) e refrigerada (12 °C; 24 dias) considerando os aspectos físicos, físico-químicos e sensoriais. A concentração inibitória mínima (CIM) da QUI e do OEOV foram 4 mg/mL e 10 μL/mL, respectivamente, contra R. stolonifer e A. niger. A incorporação de QUI a 4 mg/mL e OVEO a 5 μL/mL, 2,5 μL/mL e 1,25 μL/mL no meio de crescimento mostrou efeito fungicida e inibiu fortemente a germinação dos esporos, o crescimento micelial e esporulação das cepas teste. O revestimento de QUI-OEOV reduziu a incidência de bolor negro e podridão-mole causada por estes fungos em frutos de tomate cereja artificialmente contaminados durante o armazenamento em ambas as temperaturas estudadas. Frutos revestidos com QUI-OEOV mostraram maior firmeza e menor perda de peso. O revestimento de QUI-OEOV retardou a redução de licopeno, ácido ascórbico e cítrico, glicose e frutose durante o armazenamento em temperatura ambiente ou refrigerada. Ao final do armazenamento, os frutos revestidos com QUI-OEOV apresentaram maiores quantidades de catequina, miricetina, ácido cafeico e ácido siringico em comparação aos frutos não revestidos. Nos testes sensoriais, os frutos revestidos com QUI-OVEO receberam maiores escores para os atributos aparência, sabor e cor quando comparados aos frutos sem revestimento. Estes resultados indicam que o revestimento comestível de QUI-OEOV é um tratamento viável para manter a qualidade de tomates cereja durante o armazenamento.

Many industrially important fungi can grow in the form of small spherical pellets.Such pellets reduce the viscosity and enhances mass transfer rates in culture broths.The pelleted morphology also influences the fungus’s metabolism, directing it tospecific metabolites. The pellets are easily harvested from the broth and recycled.These properties makes pelleted morphology very attractive.The pelleted morphology of four Zygomycetes strains was studies. Several differentnutrient media used by other researchers for achieving pelleted growth was tested.The influence of eight factors on pelletization of Rhizopus sp. in a completely definedmedium was determined using a fractional central composite design and logistic regression.Pelleted growth of all four Zygomycetes was achieved, with very good results for twoRhizomucor sp. strains. A simple medium containing calcium carbonate was foundto induce pelletization with very high reproducibility.Immobilization of yeast cells was attempted in pellets of Rhizomucor. It was foundthat a flocculating yeast can be immobilized inside pellets of fungal mycelium, forming”biocapsules”. This is accomplished by first using a medium that induces pelletizationof the filamentous fungus and does not allow for growth of the yeast. Thepellets are then inoculated into a second medium that induces growth and flocculationof the yeast and inhibits further growth of the filamentous fungus.Non-flocculating yeasts could not be immobilized, suggesting that the flocculin proteinsin the cell wall of flocculating strains are important for proper immobilization.The flocculation and immobilization arises due to expression of several differentFLO-genes and the importance of these genes for successful immobilization isdiscussed.The results demonstrate that the morphology of Zygomycetes can be controlled andthat this may be useful in industrial fermentation. The new immobilization techniquereveals the great importance of flocculation and cell surface hydrophobicity. Usingyeast strains that express certain FLO-genes may be beneficial in fermentation oflignocellulosic hydrolysates.Microscopy techniques were developed that allows for high quality microphotographyof pellets and thin cross-sections of pellets and biocapsules. / Program: MSc in Resource Recovery - Industrial Biotechnology

Le manioc (Manihot esculenta Crantz) est une plante, dont les tubercules servent de nourriture de base pour près de 800 millions de personnes sous les tropiques. Malgré sont importance, ce tubercule présente deux inconvénients majeurs qui limitent leur utilisation en alimentation humaine : Une toxicité liée à la présence de composés cyanés, et une faible teneur en protéines. Dans l'optique de lever ces contraintes, plusieurs procédés de transformation ont étés mis au point par l'expérience des populations et des améliorations ont été apportées par la recherche. La plupart de ces procédés incluent une étape critique de fermentation spontanée. Cette étape présente quelques problèmes dus à sa dépendance vis à vis des microorganismes épiphytes. Ces problèmes sont notamment la présence de quantités résiduelles de composés cyanés non négligeables (100 à 170ppm) un temps de ramollissement très long (4 à 5 jours) et des produits dérivés de qualités nutritionnelles et hygiéniques douteuses. Dans l'optique d'améliorer ces procédés traditionnels de rouissage du manioc cyanogène, une centaine de microorganismes ont été isolés des cossettes de manioc séché, sur la base de leurs potentialités à produire la β-glucosidase, l'α-amylase, la pectine hydrolase et la polygalacturonase. Ces enzymes sont les plus impliqués dans les phénomènes biochimiques et physico-chimiques ayant lieu au cours du rouissage du manioc. Ils sont responsables de la dégradation des composés cyanés et du ramollissement des tubercules par dégradation des pectines qui constituent le squelette du tubercule. 60% des microorganismes isolés et identifiés ont été productrices d'α-amylases, 46% de β-glucosidase, et 36%de pectinases. Les capacités des différentes souches sélectionnées à produire ces principaux enzymes, ont ensuite été évaluées. Parmi ces microorganismes, deux se sont avérés particulièrement intéressantes : Une souche de bactérie lactique identifiée comme étant Lactobacillus plantarum avec une activité α-amylase de l'ordre de 17000±100UE/ml, et une activité ß-glucosidase de 2000±300µmol CN-/mn; Une moisissure identifiée comme étant Rhizopus oryzae à activité α-amylase de 20000±500UE/ml et activité pectine hydrolase de 2,3±0,2 µmol COO-/mn/mg. Ces deux souches ont été sélectionnées et utilisées comme ferments en culture mixte avec une biomasse initiale de 106ufc/g manioc frais de Lactobacillus plantarum et 103spores/g manioc frais de Rhizopus oryzae pour la fermentation des tubercules de manioc. Les résultats obtenus ont indiqué une acidification rapide avec un pH avoisinant 3,5±1 après 24 heures de rouissage, un ramollissement plus rapide (12±3mm/5s après 48 heures de rouissage). Mais le rouissage naturel s'est avéré meilleur du point de vue dégradation des composés cyanés. En effet la très forte acidification obtenue avec le ferment dès la 24eme heure, a provoqué un blocage du processus de dégradation des glucosides cyanogénétiques au niveau de la cyanohydrine, composé par ailleurs aussi toxique que l'acide cyanhydrique. Une étude de l'influence de quelques paramètres externes nous ont permis de noter que la température optimale de croissance de Lactobacillus plantarum était de 40±5°C pour un pH optimal de 6 alors que Rhizopus oryzae avait une température optimale de croissance à 30±7°C et un pH optimal de 5. Les essais de production de biomasses, indiquent que le meilleur milieu de multiplication des germes s'est avéré être le milieu CTM de Law et al., avec pour substrat de la farine de son de manioc mélangée au milieu nutritif de Meyer.<br />Dans l'optique de mettre à la disposition des populations ce ferment, des essais de conservation des souches par séchage ont indiqué que le fluide de protection indiqué pour avoir des taux de survies acceptables au terme du séchage était le glycérol, alors que le support le plus adapté a été l'amidon de manioc. On a ainsi obtenu plus de 75±5% de survivants après séchage sur lit convectif à une température de 35°C sous un débit d'air de 10 m3/h pour tous les deux germes. Ce ferment composé de Lactobacillus plantarum (106ufc/g) et Rhizopus oryzae (103spores/g), sur support amidon a induit contrairement aux microorganismes frais, une diminution effective du taux de glucosides totaux résiduels (5% en fin de rouissage) diminution corollaire à l'ajout du NaOH réalisé après la 24ème heure, afin de contourner l'effet néfaste du pH acide sur la dégradation de la cyanohydrine.

L(+)-Lactic acid is a commonly occurring organic acid, which is valuable due to its wide use in food and food-related industries, and its potential for the production of biodegradable and biocompatible polylactate polymers. The aim of this study was to optimize and scale-up a biotechnological process of L(+)-lactic acid production by suspended cells of R. arrhizus DAR 36017 with waste potato starch as the substrate. Commonly used inorganic and organic nitrogen sources, including ammonium sulphate, ammonium nitrate, urea, yeast extract and peptone, were assessed in conjunction with various ratios of carbon to nitrogen (C:N). Fermentation media with a low C:N ratio enhanced the production of lactic acid, biomass and ethanol, while a high C:N ratio led to production of more fumaric acid as a by-product. The use of organic nitrogen sources (yeast extract, peptone and urea) resulted in a significant reduction of lactic acid yields by 15% - 34% with a decrease of C:N from 168 to 28. The use of inorganic nitrogen sources (ammonium nitrate and ammonium sulphate) led to a high lactic acid yield of 84% - 91% at a C:N below 168. Therefore, ammonium nitrate and ammonium sulphate were considered to be better nitrogen sources for lactic acid production. Small pellets are the favoured morphological form for many fermentation processes by filamentous fungi. However, to control filamentous Rhizopus sp in the pellet form in a submerged fermentation system is difficult due to its filamentous characteristics. An acidadapted preculture technique was developed to induce the formation of the pellet form in bioreactors. Using the acid-adapted precultures, the fungal biomass can be controlled in small dispersed pellets as a dominant morphological form. With these small pellets, a lactic acid yield of 86-89%, corresponding to a concentration of 86-89g/L, was obtained in a laboratory scale process using a stirred tank reactor (STR) and a bubble column reactor (BCR). A batch bioprocess for lactic acid production was successfully scaled-up from shake flasks to laboratory scale bioreactors. Results from a simulated scale-up process revealed that the concentration and productivity of lactic acid decreased with the increase of the scale-up steps because of increased pellet size. This suggested that a one-step scale-up process using the acid-adapted preculture may be feasible in an industrial-scale bioreactor system. A comprehensive investigation of the impact of cultivation parameters on the morphology of R. arrhizus and lactic acid production was carried out in the BCR. The results showed that the fungal morphology was significantly influenced by carbon sources, pH, starch concentrations, sparger designs and aeration rates. The favoured morphology for lactic acid production was freely dispersed small pellets, which could be retained as a dominant morphology under operation conditions at pH 5.0 – 6.0, starch concentrations of 60 – 120 g/L and aeration rates of 0.2 – 0.8 vvm, using a sintered stainless steel disc sparger. The optimal cultivation conditions at pH 6.0 and aeration rate of 0.4 vvm resulted in the formation of the freely dispersed small pellets and production of 103.8 g/L lactic acid, with a yield of 87%, from 120 g/L liquefied potato starch in 48 h. This study shows a technically feasible and economically promising process for the production of lactic acid from waste potato starch. The use of waste potato starch instead of pure glucose or starch as substrate can significantly reduce the production cost, making this technology environmentally and economically attractive. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1339122 / Thesis (Ph.D.) -- University of Adelaide, School of Chemical Engineering, 2008

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2006. / Härtill 4 uppsatser.