Most plant oxylipins, a large class of diverse oxygenated polyunsaturated fatty acids and their derivatives, are produced through the lipoxygenase (LOX) pathway. Recent progress in dicots has highlighted the biological roles of oxylipins in plant defense responses to pathogens and pests. In contrast, the physiological function of LOXs and their metabolites in monocots is poorly understood. We cloned and characterized three maize LOXs ZmLOX10 ZmLOX11 and ZmLOX12. Both ZmLOX10 and ZmLOX11 apeared to be 13-LOX, whereas ZmLOX12 is a unique 9-LOX. Whereas leaf was the preferential site of ZmLOX10 expression, ZmLOX11 was strongly expressed in silks. Induction of these ZmLOX10 and ZmLOX12 by wounding and defense-related compounds suggested their role in plant resistance mechanisms against pests and pathogens. Abscisic acid, however, was the only inducer of ZmLOX11 in leaves. Higher increase in ZmLOX10 transcripts in maize infected by fungus Cochliobolus carbonum implicated this gene in resistance responses to necrotrophic pathogens. In addition, ZmLOX10 was shown to be the first reported LOX to be regulated by a circadian clock. It was found that ZmLOX10 was also inducible by low temperatures. Phenotypical studies of wild type and mutant near isogenic lines showed that expression of ZmLOX12, specific to underground organs, was required for pathogenesis of F. verticillioides on maize mesocotyls.

Most plant oxylipins, a large class of diverse oxygenated polyunsaturated fatty acids and their derivatives, are produced through the lipoxygenase (LOX) pathway. Recent progress in dicots has highlighted the biological roles of oxylipins in plant defense responses to pathogens and pests. In contrast, the physiological function of LOXs and their metabolites in monocots is poorly understood. We cloned and characterized three maize LOXs ZmLOX10 ZmLOX11 and ZmLOX12. Both ZmLOX10 and ZmLOX11 apeared to be 13-LOX, whereas ZmLOX12 is a unique 9-LOX. Whereas leaf was the preferential site of ZmLOX10 expression, ZmLOX11 was strongly expressed in silks. Induction of these ZmLOX10 and ZmLOX12 by wounding and defense-related compounds suggested their role in plant resistance mechanisms against pests and pathogens. Abscisic acid, however, was the only inducer of ZmLOX11 in leaves. Higher increase in ZmLOX10 transcripts in maize infected by fungus Cochliobolus carbonum implicated this gene in resistance responses to necrotrophic pathogens. In addition, ZmLOX10 was shown to be the first reported LOX to be regulated by a circadian clock. It was found that ZmLOX10 was also inducible by low temperatures. Phenotypical studies of wild type and mutant near isogenic lines showed that expression of ZmLOX12, specific to underground organs, was required for pathogenesis of F. verticillioides on maize mesocotyls.

Phenolic compounds in African green leafy vegetables (GLVs) may have a significant impact on human health. However, there is little information on the phenolic composition, antioxidant properties, as well as biological and cellular protective effects of these vegetables. The effects of boiling and extraction solvent on these compounds and on their antioxidant properties are also unknown. Phenolic content, antioxidant activity and cellular protective effects of four African GLVs in comparison with spinach, an exotic GLV, was determined. African GLVs had appreciable levels of total phenolics and antioxidant activity and in higher quantities compared to spinach. Boiling decreased the antioxidant content and activity of these vegetables and 75% acetone was more effective in extracting antioxidants from the GLVs compared to water. GLVs with high levels of phenolics also contained higher levels of antioxidant activity, suggesting that phenolics are likely to have contributed to radical scavenging activity of these vegetable extracts, even though the degree of scavenging varied in each extract of the vegetable species. The flavonoid compositions of raw and boiled African GLVs and spinach were determined using high-performance liquid chromatography. Epicatechin and rutin were the most dominant flavonoids found in both water and 75% acetone extracts. Among water extracts, pumpkin contained higher concentrations of detected flavonoids, while among the acetone extracts, cowpea exhibited higher concentrations. The effect of boiling was dependent on the type of vegetable and the specific flavonoids. There were no major differences observed between the type of flavonoids detected in extracts of African GLVs and those in spinach. However, similar to the results of total phenolics and antioxidant activity, the 75% acetone extracts of African GLVs also exhibited higher amounts of flavonoids than spinach. The protective effects of GLVs against oxidative haemolysis were dependent on the type of vegetable species. Boiling had variable effects depending on the species. The highest level of protection of erythrocytes against oxidative damage was offered by amaranth extracts, while extracts of raw jute mallow contributed to the damage of erythrocytes. The highest antioxidant protection activity against oxidative damage in plasmid DNA was offered by extracts of jute mallow and lowest by spinach.<p. For the cell viability assays, GLVs were evaluated to determine their cytotoxicity levels and functional role in oxidative damage. The results of the long-term cell viability (i.e. MTT, NR and CV) assays indicated no cytotoxicity, while the short-term cell viability (i.e. DCF) assay indicated that all extracts of raw GLVs were significantly (p < 0.05) cytotoxic to SC-1 fibroblast and human adenocarcinoma colon cancer (Caco-2) cells than extracts of cooked samples, and the levels of toxicity in the extracts of spinach was higher than in African GLVs. These results indicate that there was an initial cytotoxic effect as extracts of raw GLVs were added to the cells. However, after about 72 h, the cells recovered from the initial shock and started proliferating as usual. In the presence of peroxyl radicals, extracts of African GLVs exhibited higher protective effects against oxidative damage in both types of cell cultures than extracts of spinach. These results indicate that these protective effects could be attributed to the presence of phenolics and antioxidant properties of these extracts. Although boiling reduced the antioxidant content and activity of African GLVs, the levels remained higher than in spinach. Boiling also decreased the cytotoxicity and cell damage caused by extracts of raw GLVs samples. African GLVs are consumed after boiling, and therefore the observed cytotoxicities might not be experienced in practical terms. African GLVs have therefore a potential to reduce the risk and development of diseases associated with oxidative stress in communities that consume these vegetables. / Thesis (PhD)--University of Pretoria, 2012. / Centre for Nutrition / PhD / Unrestricted

Thesis(M.Sc.(Agriculture Horticulture)) -- University of Limpopo, 2019 / Innovative methods of preserving the quality of traditionally processed green leafy vegetables are underway in Africa. Improvement of processing and preservation methods of leafy vegetables is another way of overcoming perishability restrictions and guaranteeing continued quality food supply in rural areas. The objectives of this study were: to determine the effect of time-based oven-drying on (1) mineral composition, (2) proximate composition and (3) microbial profiling of cowpea (Vigna unguiculata). Separate experiments were conducted for raw and cooked cowpea leaves with four treatments, viz 0 (sun dried), 24, 48 and 72 hours of oven-drying, arranged in a randomised complete block design with 5 replications. In raw cowpea leaves, relative to control (sun-drying), oven-drying period 48hrs, significantly decreased Potassium (K), Manganese (Mn), and Sodium (Na) content in raw cowpea leaves by 6, 9 and 13%, respectively. Similarly, oven-drying period 72hrs significantly decreased Ca, Fe, Mg, Zn, P and S by 5, 11, 16, 18 and 57%, respectively. In cooked cowpea leaves, relative to control (sun-drying), oven-drying period 24hrs significantly increased Na by 18%. Similarly 48hrs oven-drying periods increased Fe and K by 6 and 8%, respectively. Similarly, oven-drying period 72hrs significantly increased Ca, Mg and Mn by 8, 8 and 3%, respectively. In contrast, oven-drying period 72hrs significantly reduced Zn, P and S by 16, 10 and 39%, respectively. Relative to control (sun-drying), oven-drying period 24hrs significantly increased fat by 46% in raw cowpea leaves, however oven-drying period 72hrs significantly decreased protein, moisture, ash, fibre and carbohydrate by 10, 29, 18, 0.5, and 7% respectively. In contrast, relative to control (sun-drying), 72hrs increased energy by 3%. In cooked cowpea leaves, relative to control (sun-drying), ovendrying period 24hrs significantly increased energy by 1%. In contrast, relative to control (sun-drying) 72hrs oven-drying period decreased energy by 1%. Similarly, oven-drying 72hrs significantly decreased protein, moisture, ash, fat, fibre and carbohydrate by 8, 14, 13, 19, 0.4 and 10% respectively. Relative to control (sun-drying), oven-drying periods 24hrs significantly increased Staphylococcus spp. in raw cowpea leaves by 6%, respectively. Relative to control (sun-drying) 72hrs oven-drying period significantly decreased Shigella spp. by 92%, respectively. In cooked leaves, relative to control (sundrying), 72hrs drying periods decreased both Shigella spp. and Staphylococcus spp. by 99 and 21%, respectively. Total coliforms unit of Salmonella spp, Escherichia coli, Pseudomonas spp, and Bacillus cereus were absent and/or at an undetectable level according to the Tempo Biomerieux system results. In conclusion, cooked leaves retained most essential mineral elements as compared to raw when subjected to 72hrs of ovendrying as there was an improvement in the concentration of Ca, Fe, K, Mg, Mn and Na. However, in both raw and cooked cowpea leaves proximate composition was negatively affected as there was a decrease in protein content. Oven-drying period of 24 hours can be used to minimize the loss of protein. Cooked cowpea leaves subjected to 72hrs of oven-drying had least total coliforms for both Shigella spp and Staphylococcus spp, therefore have potential to serve as an alternative to sun-drying to reduce microorganism causing spoilage in leafy vegetables. Drying raw cowpea leaves under oven-drying periods less than 72 hours should be avoided as it reduces the mineral concentration and increase microbial count of microorganisms responsible for spoilage / National Research Foundation (NRF)

Escherichia coli produtora de toxina Shiga (STEC) está entre os patógenos envolvidos em surtos de doenças transmitidas por alimentos devido ao consumo de vegetais. No entanto, até agora, os relatos sobre a presença de STEC em vegetais no Brasil são escassos. Esse microrganismo é veiculado por alimentos, contaminados direta ou indiretamente por fezes animais, sendo responsável por um amplo espectro de doenças que compreende desde diarréia leve que pode evoluir para colite hemorrágica (CH), até síndrome hemolítico-urêmica (SHU) e púrpura trombocitopênica trombótica (PTT). O presente estudo teve como objetivo investigar a presença de STEC em vegetais orgânicos cultivados na região metropolitana da cidade de São Paulo, Brasil, caracterizando os fatores de virulência stx1, stx2, eae e ehx, bem como o sorotipo. Um total de 200 amostras de vegetais orgânicos (folhas verdes), obtido a partir de três produtores foi analisado quanto à presença de cepas de STEC. Caldo triptona de soja (TSB) suplementado com vancomicina (8 mg / L), cefixima (50 &#181;g / L) e telurito de potássio (2,5 mg / L) foi utilizado na etapa de pré-enriquecimento, com incubação a 37ºC / 24 h, seguido por semeadura em MacConkey Sorbitol (SMAC) e CHROMagar STEC (CHROM). Após incubação a 37ºC / 24 h, as colônias suspeitas foram confirmadas por testes bioquímicos e submetidas a PCR objetivando a detecção dos genes de virulência stx1, stx2, eae, ehx, e os genes fliCH7 e rfbO157. Entre as 200 amostras de vegetais orgânicos analisadas, 30 (15%) foram positivas para E. coli, mas nenhum isolado apresentou os genes de virulência pesquisados. Nossos resultados indicam baixo risco de infecção devido ao consumo destes produtos frescos em São Paulo, Brasil. No entanto, são necessárias mais pesquisas, abrangendo um maior número de amostras e área pesquisada, uma vez que este patógeno já foi encontrado no meio ambiente em estudos anteriores e poucas pesquisas investigaram a presença de STEC em vegetais no Brasil. / Shiga toxin producing Escherichia coli (STEC) strains are among the pathogens involved in foodborne disease outbreaks due to consumption of vegetables. However, reports on the presence of STEC in vegetables in Brazil are lacking. STEC is an important pathogen transmitted by food, directly or indirectly contaminated with animal feces, responsible for a broad spectrum of diseases varying from mild diarrhea to hemorrhagic colitis (HC), syndrome hemolytic uremic (HUS) and thrombotic thrombocytopenic purpura (TTP). This study aimed at investigating the presence of STEC in organic vegetables in the metropolitan region of São Paulo city, Brazil, characterizing the virulence factors stx1, stx2, eae and ehx as well as identifying the serotype. A total of 200 samples of organic vegetables (green leafy), obtained from three organic producers was analyzed for the presence of STEC strains. Tryptic Soy Broth (TSB) supplemented with vancomycin (8mg/L), cefixim (50&#181;g/L) and potassium telurite (2.5mg/L) was used in the pre enrichment step with incubation at 37°C/24 h, followed by plating onto Sorbitol-MacConkey (SMAC) agar and CHROMagar STEC (CHROM). After incubation at 37°C/24 h, presumptive colonies were confirmed by biochemical tests and submitted to PCR targeting the detection of stx1, stx2, eae and ehx virulence genes, as well as fliCH7 and rfbO157. Among the 200 organic vegetable samples analyzed for STEC strains, 30 (15%) were positive for E. coli, but none of them showed the virulence genes studied. These findings indicate low risk of infection due to the consumption of these fresh produce in Sao Paulo, Brazil. However, more research is required, covering a larger number of samples and area, since this pathogen has already been found in the environment in previous studies, and few research investigating the presence of STEC in vegetables has been reported in Brazil.

Lipoxygenase (LOX)-derived oxylipins are known to play critical roles in defense against herbivores and pathogens. The objective of this study was to determine the biochemical, molecular and physiological roles of a specific maize lipoxygenase gene, ZmLOX10, with special emphasis on LOX10-derived oxylipins in plant-insect and plant-pathogen interactions. To achieve this goal, independent mutant alleles were generated and genetically advanced to create near-isogenic mutant and wild-type lines suitable for functional analysis. Here we provide genetic evidence that LOX10 is the sole LOX isoform in maize required for the biosynthesis of green leafy volatiles (GLV) in leaves and show that LOX10- mediated GLVs play a significant role in direct and indirect defense responses to insects through their regulation of jasmonic acid and volatile organic compound production. Contrary to the defensive role of LOX10 in plant-insect interactions, tests for susceptibility to fungal pathogens suggest that LOX10-mediated GLVs may contribute to the development of disease symptoms to the economically important maize pathogens, Aspergillus flavus and Colletotrichum graminicola. Specifically, LOX10-derived GLVs may facilitate aflatoxin accumulation in response to A. flavus infection and may play a positive role in anthracnose leaf blight and stalk rot caused by C. graminicola. Collectively, our results suggest that metabolites derived from GLV-regulated pathways have a significant impact on molecular plant-herbivore and plant-pathogen interactions.

Escherichia coli produtora de toxina Shiga (STEC) está entre os patógenos envolvidos em surtos de doenças transmitidas por alimentos devido ao consumo de vegetais. No entanto, até agora, os relatos sobre a presença de STEC em vegetais no Brasil são escassos. Esse microrganismo é veiculado por alimentos, contaminados direta ou indiretamente por fezes animais, sendo responsável por um amplo espectro de doenças que compreende desde diarréia leve que pode evoluir para colite hemorrágica (CH), até síndrome hemolítico-urêmica (SHU) e púrpura trombocitopênica trombótica (PTT). O presente estudo teve como objetivo investigar a presença de STEC em vegetais orgânicos cultivados na região metropolitana da cidade de São Paulo, Brasil, caracterizando os fatores de virulência stx1, stx2, eae e ehx, bem como o sorotipo. Um total de 200 amostras de vegetais orgânicos (folhas verdes), obtido a partir de três produtores foi analisado quanto à presença de cepas de STEC. Caldo triptona de soja (TSB) suplementado com vancomicina (8 mg / L), cefixima (50 &#181;g / L) e telurito de potássio (2,5 mg / L) foi utilizado na etapa de pré-enriquecimento, com incubação a 37ºC / 24 h, seguido por semeadura em MacConkey Sorbitol (SMAC) e CHROMagar STEC (CHROM). Após incubação a 37ºC / 24 h, as colônias suspeitas foram confirmadas por testes bioquímicos e submetidas a PCR objetivando a detecção dos genes de virulência stx1, stx2, eae, ehx, e os genes fliCH7 e rfbO157. Entre as 200 amostras de vegetais orgânicos analisadas, 30 (15%) foram positivas para E. coli, mas nenhum isolado apresentou os genes de virulência pesquisados. Nossos resultados indicam baixo risco de infecção devido ao consumo destes produtos frescos em São Paulo, Brasil. No entanto, são necessárias mais pesquisas, abrangendo um maior número de amostras e área pesquisada, uma vez que este patógeno já foi encontrado no meio ambiente em estudos anteriores e poucas pesquisas investigaram a presença de STEC em vegetais no Brasil. / Shiga toxin producing Escherichia coli (STEC) strains are among the pathogens involved in foodborne disease outbreaks due to consumption of vegetables. However, reports on the presence of STEC in vegetables in Brazil are lacking. STEC is an important pathogen transmitted by food, directly or indirectly contaminated with animal feces, responsible for a broad spectrum of diseases varying from mild diarrhea to hemorrhagic colitis (HC), syndrome hemolytic uremic (HUS) and thrombotic thrombocytopenic purpura (TTP). This study aimed at investigating the presence of STEC in organic vegetables in the metropolitan region of São Paulo city, Brazil, characterizing the virulence factors stx1, stx2, eae and ehx as well as identifying the serotype. A total of 200 samples of organic vegetables (green leafy), obtained from three organic producers was analyzed for the presence of STEC strains. Tryptic Soy Broth (TSB) supplemented with vancomycin (8mg/L), cefixim (50&#181;g/L) and potassium telurite (2.5mg/L) was used in the pre enrichment step with incubation at 37°C/24 h, followed by plating onto Sorbitol-MacConkey (SMAC) agar and CHROMagar STEC (CHROM). After incubation at 37°C/24 h, presumptive colonies were confirmed by biochemical tests and submitted to PCR targeting the detection of stx1, stx2, eae and ehx virulence genes, as well as fliCH7 and rfbO157. Among the 200 organic vegetable samples analyzed for STEC strains, 30 (15%) were positive for E. coli, but none of them showed the virulence genes studied. These findings indicate low risk of infection due to the consumption of these fresh produce in Sao Paulo, Brazil. However, more research is required, covering a larger number of samples and area, since this pathogen has already been found in the environment in previous studies, and few research investigating the presence of STEC in vegetables has been reported in Brazil.

Concern regarding the decline in the knowledge and use of indigenous green vegetables exists in many southern African regions including Swaziland. Reasons for this are, amongst others, attributed to a lack of inter-generational knowledge transfer, the influence of westernisation and changing food values and attitudes, as people nowadays regard indigenous green leafy vegetables as a low status food. It is imperative that knowledge of these foods be documented before it is lost to future generation. Indigenous green leafy vegetables have long been considered healthier options than the many recently introduced western-orientated foods, which are closely linked to chronic lifestyle diseases. The aim of this study was to determine, describe and compare the knowledge of indigenous green leafy vegetables among two generations of rural Swazi women from the Eluyengweni community, and how it manifests in their food practices. Studies on indigenous green leafy vegetables in Swaziland are limited. This study will fill the knowledge gap in this field. The human ecological perspective was adopted as theoretical perspective and an explorative mixed method approach was followed. Qualitative data was collected through focus group discussions with younger (25-45 years) and older (over 45 years) women. Face-to-face structured interviews provided quantitative data on the knowledge the women had of indigenous green leafy vegetables, including their procurement, preparation, preservation and consumption patterns. The convenience sample comprised 102 respondents. Observation was done concurrently during both phases of data collection and photographs accompanied comprehensive field notes. It transpired that the availability of indigenous green leafy vegetables had declined in the community and those that were available were not easily accessible as they were located on the outskirts of the community. In spite of changes in the physical and natural environment, some indigenous green leafy vegetable species are still available in summer. It is clear that the older women, in comparison to their younger counterparts, had more knowledge of indigenous green leafy vegetables in terms of identifying them, where they are found and how they are used. Familiar and available indigenous green leafy vegetables such as imbuya (Amaranthus), chuchuza (Bidens pilosa) were preferred, and consumed fairly regularly. Although indigenous green leafy vegetables were still part of the food practices in this community, their consumption among the younger generation had declined. However, due to factors related to modernisation, economic influences and technological advancement the majority of both the younger and older participants seem to have become dependent on, and often include, western-orientated food in their food consumption patterns. Based on the results, recommendations are made to capture and preserve the knowledge of indigenous green leafy vegetables in this community and to revive interest in these vegetables and promote their consumption. / Daar is 'n besorgdheid oor die afname in kennis in die gebruik van inheemse groen blaargroentes in baie streke in Suider Afrika, insluitend Swaziland. Redes wat hiervoor aangevoer word, is onder andere, die gebrekkige oordrag van kennis van een geslag na die volgende, die invloed van verwestering asook 'n verandering in waardes en houdings aangesien mense inheemse groen blaargroentes deesdae as 'n lae status voedsel beskou. Daarom is dit noodsaaklik dat kennis oor hierdie voedsel gedokumenteer word voordat dit verlore gaan vir die nageslag. Inheemse groen blaargroentes word gesonder geag in vergelyking met Westers-georienteerde voedsel, wat op hul beurt nou geassosieer word met kroniese lewenstyl siektes. Die doel van hierdie studie was om die kennis van inheemse groen blaargroentes van twee geslagte landelike Swazi vroue van die Eluyengweni gemeenskap te bepaal, beskryf en te vergelyk asook hoe dit manifesteer in hul voedselpraktyke. Studies oor inheemse groen blaargroentes in Swaziland is beperk en hierdie studie dra by om die leemte in kennis in hierdie studieveld te vul. Die menslike ekologiese perspektief is a teoretiese perspektief gebruik en 'n verkennende gemengde metodologie is as navorsingsbenadering gevolg. Kwalitatiewe data is deur middle van fokusgroep gesprekke met beide die jonger (25-45 jaar) en ouer (ouer as 45 jaar) vroue ingesamel. Met behulp van gestruktureerde individuele onderhoude is kwantitatiewe data oor die kennis van inheemse groen blaargroentes ingesamel wat die verkryging, voorbereiding, preservering en verbruikspatrone van hierdie groentes ingesluit het. 'n Geriefssteekproef van 102 respondente het aan die studie deelgeneem. Waarnemings is gedurende beide fases van die data insameling uitgevoer en fotos en volledige veldnotas is ingesluit. Dit is duidelik dat die beskikbaarheid van inheemse groen blaargroentes in hierdie gemeenskap afgeneem het, en dit wat beskikbaar was, was nie maklik toeganklik nie, omdat dit aan die buitewyke van die gemeenskap gelëe was. Ten spyte van die verandering in die fisiese en natuurlike omgewing is sommige inheemse groen blaargroentes steeds beskikbaar gedurende die somer. Dit is duidelik dat die ouer vroue, in vergelyking met die die jonger vroue oor meer kennis van inheemse groen blaargroentes beskik het met betrekking tot die identifisering, waar hulle groei en hoe om hulle te gebruik. Bekende en beskikbare inheemse groen blaargroentes soos gewone misbredie (Amaranthus) en knapsakkerwel (Bidens pilosa) was voorkeur groente en is redelik gereeld gebruik. Alhoewel inheemse groen blaargroentes steeds deel vorm van die voedselpraktyke in hierdie gemeenskap, het die gebruik daarvan deur die jonger geslag afgeneem. As gevolg van faktore soos modernisering, die ekonomie en tegnologiese vooruitgang het die meerderheid van beide die jonger en ouer geslag afhanklik geword van westers-georienteerde voedsel en dit dikwels in hulle eetpatrone ingesluit. Die resultate van hierdie studie het bygedra dat aanbevelings oor die vaslegging en bewaring van die kennis oor inheemse groen blaargroentes in hierdie gemeenskap gemaak kon word, asook om 'n oplewing in die belangstelling in die gebruik daarvan te bevorder. / Dissertation (MConsumer Science)--University of Pretoria, 2017. / Consumer Science / MConsumer Science / Unrestricted