Submitted in fulfilment of the requirements for the degree of Master of Applied Science in Food Science and Technology, Durban University of Technology, 2015. / Listeria monocytogenes have been implicated as a public health concern worldwide. The study explored the survival of non-adapted, heat adapted and chlorine adapted L. monocytogenes on tomatoes; as well as the survival of non-adapted, heat adapted and chlorine adapted biofilms after exposure to sodium dodecyl sulphate (SDS), levulinc acid, sodium hypochlorite solution. Contact time of 1, 3 and 5 minutes was used. The survival of L. monocytogenes was monitored at 0, 24, 48 and 72 hours. The sanitizers were used individually or combined as follows; 1% sodium dodecyl sulphate individually; 0.5% levulinic acid individually; 200 ppm sodium hypochlorite solution individually and 0.5% levulinic acid/0.05% sodium dodecyl sulphate in combination (mixture). The samples were kept at 4 °C throughout the period of assessment. The effect of these sanitizers on pH, total soluble solids (TSS) and titratable acidity (TA) was also determined. Furthermore, the attachment of L. monocytogenes on tomatoes was investigated using a scanning electron microscope. Highest log reduction of non-adapted L. monocytogenes were observed on tomatoes treated with 1% SDS and least log reduction was achieved when tomatoes were treated with sodium hypochlorite solution. Though the log reduction achieved by 0.5% levulinic acid was higher that sodium hypochlorite solution, it was lower than log reduction achieved when 0.05% SDS / 0.5% levulinic acid mixture was used for all contact times. Using non-adapted L. monocytogenes, SDS was able to destroy all L. monocytogenes at 1, 3 and 5 minutes contact time. The trend was the same when heat adapted and chlorine adapted L. monocytogenes were used. There was no significant log reduction observed with biofilms. More favourable results were observed as contact time was increased from 1 to 5 minutes. Though there was a decrease in surviving bacteria from 1 to 3 minutes contact time, this decrease was not significant. The study investigated if exposure to sanitizer has an effect on pH, titratable acidity (TA) and total soluble solids (TSS) of the tomatoes. It was revealed that levulinic acid and mixture can have detrimental effect on pH, TA and TSS of tomatoes. The TA and TSS of samples treated with levulinic acid and mixture varied significantly (P ≤ 0.05) compared to the control sample. Although the TA and TSS of samples treated with SDS and sodium hypochlorite solution were different from the control, the differences were not significant. As much as sanitizers have the potential to reduce the bacterial population in fresh produce they may not completely destroy pathogens. Chlorine based sanitizers such as sodium hypochlorite though frequently used in the fresh produce industry, are not the best sanitizer to be used against food borne pathogens. Other sanitizers such as SDS used alone or in combination with another sanitizer can achieve better results than the widely used sodium hypochlorite solution as observed in this study. Stress adapted pathogens become less responsive to sanitizers during subsequent treatments. Through this research, it was established that biofilms are resistant to sanitizers. Though application of sanitizers in fresh produce is cheaper and simpler to apply, there is need to monitor varying concentrations of sanitizers, contact time and minimise contact with sub-surfaces as this could lead to sensory quality losses.

Tomato spotted wilt virus (TSWV), a Tospovirus, is one of the ten most economically destructive plant viruses worldwide, causing losses exceeding one billion U.S. dollars annually on several crops. In South Africa (SA), TSWV has become an important virus in many economically important crops. The main objective of this research project was to isolate, identify and characterise TSWV isolates occurring in SA. A review of current literature assembled background information on TSWV molecular biology, epidemiology, transmission, detection and control. A TSWV isolate infecting pepper (Capsicum sp.) occurring in KZN was isolated and partially characterised. The virus was positively identified as TSWV using the enzyme-linked immunosorbent assay (ELISA) and the presence of typical necrotic TSWV symptoms on Nicotinia rustica L. Symptomatic leaves were harvested and the virus was partially purified using standard procedures. Under the transmission electron microscope (TEM), typical quasi-spherical and dumbbell-shaped particles of 80-100nm in diameter were observed in negatively stained preparations of both crude and purified virus samples. In negatively stained ultra-thin virus infected leaf sections, an abundance of mature viral particles (100nm) housed in the cisternae of the endoplasmic reticulum (ER) were observed among typical viroplasm inclusions (30nm) and hollow tubules (200-300nm). A viral protein migrating as a 29kDa band, which corresponds to the TSWV nucleocapsid (N) protein, was observed after sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. Total plant RNA, isolated from N. rustica displaying typical symptoms was subjected to reverse-transcription polymerase chain reaction (RT-PCR) using .primers specific to the nucleocapsid (N) gene. An expected 760bp product was amplified. The results obtained in this study confirm the presence of TSWV in infected pepper plants from KZN. The genetic diversity of TSWV isolates occurring in SA was examined. The nucleocapsid (N) gene sequences of six SA TSWV isolates originating from Gauteng, KwaZulu-Natal, North West, Limpopo and Mpumulanga provinces were determined and used in a phylogenetic tree comparison with TSWV isolates occurring in different geographical locations in the world. Nucleotide sequence comparisons of the N gene revealed high levels of similarity between the SA isolates and TSWV isolates from Asia and Europe. SA isolates showed a high degree of sequence similarity (99-100%) which was reflected in their distinct clustering pattern. The resistance of tomato (Lycopersicon escuJentum Mill.) plants with natural and transgenic resistance against mechanical inoculation with TSWV isolates occurring in SA was evaluated. The Stevens cultivar which has natural resistance conferred by the Sw-5 gene and the transgenic 13-1 line, which expresses the nucleocapsid (N) protein gene of the TSWV-BL isolate, was used as test cultivars. Plants were assessed for TSWV resistance using a disease severity rating scale and measurements of virion accumulation levels (A405nm). There were no significant differences among the reactions produced by the six TSWV isolates on the test plants. Although both plants were susceptible to the SA TSWV isolates by exhibiting similarly high viral accumulation levels, the transgenic tomato line showed milder disease severity compared to the natural resistant cultivar. Results suggest that transgenic resistance is a more effective approach in the control of TSWV in SA. The information generated in this study will be useful in formulating effective control measures using genetic engineering approaches for this economically important virus. Such approaches will be used as a tool to make strategic decisions in an integrated control programme for ISWV. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2006.