Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

This thesis reports the cell culture establishment and a somatic cell selection system optimized for the isolation of chlorsulfuron-resistant variants in asparagus (Asparagus officinalis L.). The development of this cell selection system benefited the isolation of chlorsulfuron-resistant variants from an elite asparagus genotype. A cell culture system, suitable for somatic cell selection, was established for asparagus genotype CRD 168. Friable callus was initiated from etiolated shoots in darkness and used to produce a high density of single cells in suspension. Cell density was estimated based on a linear relationship with settled cell volume. A mean plating efficiency of 0.19 % was recorded between 1-4x10⁵ cells/Petri dish. In vitro cell selection techniques were developed to identify mutant asparagus cells with resistance to a sulfonylurea herbicide, chlorsulfuron. A few key aspects were important to achieve this: a cell culture system for cell selection was initially established; a toxic concentration for the complete growth inhibition of the wild type asparagus cells was defined; rare, resistant cell colonies were isolated and characterized; and chlorsulfuron-resistant plants were regenerated. From about 50 million cells, 165 cell colonies were isolated in the presence of 8 nM chlorsulfuron. Characterization of these selected cell colonies yielded 24 escapes, 98 unstable variants, and 43 stable-resistant variants. Callus cultures from 34 of these stable variants retained resistance following 11 months growth in the absence of the selection agent. Plants were regenerated from 36 of these stable herbicide-resistant variants. Six of these chlorsulfuron-resistant variants were screened for their degree of resistance to chlorsulfuron, cross resistance to other acetohydroxyacid synthase (AHAS) inhibiting herbicides and AHAS enzyme activity. Cross resistance to imazamox was evident in four of the resistant variants, while lack of cross resistance to metsulfuron methyl was observed in all six resistant variants. A varying degree of resistance to chlorsulfuron was observed among the resistant variants. Both in the original and secondary callus, an uninhibited AHAS enzyme activity in all six resistant variants was recorded in the presence of high chlorsulfuron concentration (70-140 nM), compared to the total inhibition in the wild type. One chlorsulfuron-resistant variant, R-45, was used to compare the biochemical and physiological basis of resistance with the wild type. The AHAS enzyme activity in the tissue culture and greenhouse foliage of R-45 was significantly higher in the presence of up to 280 nM chlorsulfuron compared with the wild type. Chlorsulfuron retention was considerably higher due to the reduction of epicuticular wax deposits on the foliage of R-45, in comparison with the wild type. Consequently, the resistant line absorbed at least 1.6 fold more chlorsulfuron than the wild type plants. Therefore, foliar application of 15 g a.i./ha Glean (commercial formulation of chlorsulfuron) produced typical symptoms of chlorosis in R-45, similar to the wild type, in the greenhouse plants. Somatic cell selection was carried out using two elite asparagus genotypes, CRD 74 and Clone X. Of the 33 rare cell colonies isolated from Clone X, 22 unstable variants and 6 escapes were discarded. All five remaining resistant variants produced plants. One of the stable-resistant variants (Clone X-24) was evaluated for resistance to chlorsulfuron. Both in vitro shoot cultures and greenhouse-grown plants of Clone X-24 showed increased resistance to chlorsulfuron compared with the wild type. The AHAS enzyme activity in the foliar extracts also showed the presence of higher enzyme activity in Clone X-24.

In Deutschland nimmt der Anbau von Bleichspargel (Asparagus officinalis L.) sechzehn Prozent der Gesamtgemüseanbaufläche ein. Die Qualitätssicherung von Spargel, insbesondere aber von geschältem (Convenience-) Spargel stellt wegen der hohen Stoffwechselaktivität nach der Ernte eine Herausforderung dar. Insbesondere gilt es, die Textur und die Inhaltsstoffe vor qualitätsmindernden Veränderungen und verderbsförderndem Pilzbefall zu bewahren. Als Verpackung dienen in der Regel Folienverpackungen, die aber oft für das empfindliche Gemüseprodukt nicht geeignet sind. Daher wurde der Einfluss von unterschiedlichen Folienverpackungen (vier Polypropylenfolien, zwei biologisch abbaubare Folien und ein Oberflächencoating) mit unterschiedlicher Permeabilität für Sauerstoff und Kohlendioxid auf die Veränderungen der Qualitätsparameter ( Farbe, Textur, Frischmasse, Trockensubstanz, Gerüstkohlenhydrate ( Pectine, Lignin, Hemicellulose, Cellulose, Saccharose, Fructose, Glucose) von nicht geschälten und geschälten Spargel unmittelbar nach der Ernte und nochmals nach drei (bzw. vier) Lagertagen (2°C, 10°C, 20°C Lagertemperatur) ermittelt. Direkt nach der Ernte und nach drei Lagertagen wurde bei 10°C und 20°C Lagertemperatur die Kontamination mit Pilzen und der eventuell damit verbundene Gehalt an Fumonisin B1 geprüft. Die Lagertemperatur von 10°C (2 d Lagerdauer) erwies sich als geeignet zur Qualitätserhaltung von Convenience – Spargel. Bei 2°C und 20°C geschältem Spargel waren die Pectinfraktionen, bei nicht geschältem Spargel war der Hemicellulosegehalt Veränderungen unterworfen. Die Veränderungen dieser Qualitätsparameter waren mit den Veränderungen der Textur korreliert. Die Spargelspitze ist bei der Lagerung (2°C und 20°C) stoffwechselaktiver, als die restliche Spargelstange. Es zeigte sich, dass das Verhältnis von O2 zu CO2 (RQ) in der Verpackungsfolie signifikanten Einfluss auf die Qualitätsparameter von geschältem Spargel ausübte. Die geringsten stoffwechselphysiologischen Veränderungen wurden bei einem RQ von 0,65 festgestellt. Folienverpackungen mit RQ von 0,03-0,65 hatten sich für das endophytische Pilzwachstum als hemmend erwiesen. Das Mykotoxin Fumonisin B1 wurden in gesundheitlich unbedenklichen Mengen (< 1,67 mg * kg TS-1) in nicht gelagerten und in gelagerten Spargelstangen nachgewiesen. / The production area of white asparagus comprises 16 % of all vegetable crops in Germany. The economic important asparagus (Asparagus officinalis L.) is known for its high quality loss in postharvest. In order to protect convenience asparagus, i.e. fresh-cut peeled white asparagus from rapid deterioration in respect to phytopathological fungi, nutritive and sensory compounds the commercial use of film packaging might be an important tool. However, the film packaging materials used commercially do often not fulfil product physiological concerns. Therefore, the influence of different film packaging materials (four polypropylene-films, two biological degradable films and one coating) with different permeabilities for CO2 and O2 was investigated for unpeeled and peeled white asparagus during storage (2, 3, and 4 days) at temperatures of 2°C, 10°C, 20°C. Changes in the following quality attributes were studied: colour, texture, fresh weight, dry weight, structural carbohydrates (pectic substances, lignin, hemicellulose, cellulose), mono- and disaccharides (fructose, glucose, sucrose). Moreover, at harvest and after three days of storage the contamination with fungi and the content of the mycotoxin Fumonisin B1 was observed. The tests for contamination of fungi were conducted with slight nutrient agar (SNA) for seven days at 20°C under 14 h UV light and 10 h darkness (Nirenberg, 1976) Quality changes were most inhibited at a storage temperature of 10°C for two days. The ratio of O2 to CO2 (RQ) within the film packaging had a pronounced effect on the quality attributes of peeled asparagus. Changes in dry weight, fresh weight, lignin content were low at a RQ of 0,65 (P-Plus 2 film), whereas the water-soluble/insoluble pectin ratio (1 : 0,8) remained constant during the entire storage period. A low correlation was found between texture and hemicellulose, glucose, sucrose and the ratio of water-soluble to insoluble pectin. The meristematic zone of the asparagus tip revealed a higher metabolic activity than other morphological parts of the spear. Endophytic fungi, e.g. Fusarium spp., a precursor of mycotoxin, was found in control asparagus spears. During storage, the development of Fusarium spp. could be inhibited by all film packaging revealing a RQ of 0,003 - 0,65. The mycotoxin Fumonisin B1 occurred in all asparagus spears (control and stored spears), however the content (< 1,67 mg * kg TS -1) did not reach health risk threshold values.