D.Tech. (Biomedical Technology) / To determine the quality of human food commodities commonly consumed in Cameroon, various districts in the western highland (Bamenda and Kumbo) and tropical rain forest (Douala and Yaounde) regions were sampled. Two mycological investigations were conducted to evaluate the incidences of mycotoxigenic fungi (95 samples) and mycotoxins (82 samples). Serial dilution of ground samples was employed to isolate fungi, subculture on various culture media and fungal species were identified morphologically followed by molecular phylogenetic approach. In general, data obtained indicate samples from various geographical regions showed no consistent variation with regard to the type of fungal species. The mycobiota of food materials were characterized by a diversity of fungal species with the predominance of Aspergillus (125 isolates) followed by Penicillium (94 isolates) and Fusarium (52 isolates). The less predominant genera include Rhizopus (14 isolates) and the Alternaria (9 isolates). Aspergillus flavus and A. parasiticus occurred in 53 and 44% of the samples, respectively, with higher frequencies in maize than peanuts or beans and absent in rice, pumpkin seed and cassava products. Aspergillus fumigatus was detected in 20% of samples and A. niger in 18% of the samples. Aspergillus isolated less frequently included A. carbonarius A. awamori, A. oryzae and A. tamarii, A. pseudotamarii, A. ochraceus, A. ostianus, A. avenaceum, A. oryzae and A. variabile. Consistent results were observed for A. tamarii, A. pseudotamarii, A. ochraceus and A. ostianus with respect to substrate specificity. While A. tamarii, A. pseudotamarii and A. ochraceus were isolated only from peanuts, A. ostianus strains occurred only in bean samples. Penicillium contamination was dominated by P. polonicum and P. crustosum with incidence rates of 43 and 41%, respectively, with highest contamination levels registered in samples from Yaounde. Penicillium citrinum, P. purpurogenum, P. islandicum, P. aurantiogriseum, P. expansum were also inconsistently isolated from food samples. There was a relatively low incidence of Penicillium spp. in pumpkin seed and fermented cassava product samples.

No description available.

No description available.

Thesis (Ph.D. (Medical Science)) -- University of Limpopo, 2012

Dissertation (PhD)--University of Stellenbosch, 2007. / ENGLISH ABSTRACT: Aflatoxins are mycotoxins predominantly produced by the filamentous fungi Aspergillus flavus and Aspergillus parasiticus. Aflatoxin B1 (AFB1), the most abundant aflatoxin, is highly mutagenic, toxic, carcinogenic and teratogenic to humans and animals and is particularly correlated with the incidence of hepatocellular carcinoma in parts of Africa, China and South East Asia. In this regard aflatoxin is classified as a type I human carcinogen by the International Agency for Research on Cancer. Furthermore, aflatoxin contamination of food and feed is responsible for extensive economic losses due to loss of crops and farm animals. In spite of regulations regarding acceptable levels of aflatoxin in food, aflatoxin contamination remains a serious worldwide problem, especially in developing countries where it occurs predominantly in dietary staples. Inactivation of aflatoxin by physical and chemical methods has not yet proved to be effective and economic. However, biological detoxification offers an attractive alternative for eliminating toxins as well as safe-guarding the desired quality of food and feed. In this study, the biological degradation of AFB1 by bacteria and fungi was investigated. Several bacteria, including Rhodococcus spp., as well as white rot fungi have the potential to degrade a wide range of polycyclic hydrocarbon compounds due to the large repertoire of enzymes they produce and therefore the ability of some of these microorganisms to degrade AFB1 was investigated. Effective degradation of AFB1 by intracellular extracts of Mycobacterium fluoranthenivorans sp. nov. DSM 44556T, Nocardia corynebacterioides DSM 20151 and N. corynebacterioides DSM 12676 was demonstrated. Furthermore, AFB1 was effectively degraded by liquid cultures as well as intra- and extracellular extracts of Rhodococcus erythropolis DSM 14303. Significant (P<0.001) reduction in AFB1 was observed following treatment with R. erythropolis extracellular extracts with only 33.20% residual AFB1 after 72 h. Results indicated that the degradation by R. erythropolis DSM 14303 is enzymatic and that the enzymes are constitutively produced. The degradation of AFB1 when treated with R. erythropolis DSM 14303 extracellular extract coincided with a total loss of mutagenicity. In addition, treatment of AFB1 with culture fractions containing recombinant 2,3-dihydroxybiphenyl dioxygenase, which was produced through extracellular expression of the bphC1 gene of R. erythropolis DSM 14303 in Escherichia coli BL21, resulted in significant (P<0.0001) degradation (49.32%) and reduced mutagenic potency (42.47%) of the molecule. Significant (P<0.0001-0.05) degradation of AFB1 was obtained following treatment with culture extracts containing laccase enzyme produced by white rot fungi (17.10- 76.00%), purified fungal laccase from Trametes versicolor (1 U/ml, 87.34%) as well as with recombinant laccase produced by Aspergillus niger (118 U/L, 55.00%). Furthermore, treatment of AFB1 with purified fungal laccase enzyme (1 U/ml) resulted in loss of the mutagenic potency of the molecule. The decrease in the fluorescence and mutagenic properties of AFB1 following treatment with the microbial preparations imply changes to the furofuran- and/or lactone rings of the molecule. The current study contributes towards developing genetic engineered microbial strains which could be applied as an important bio-control measure. Such strains could exhibit multifunctional technological properties including degradation of AFB1, to significantly improve the quality, safety and acceptability of food. / AFRIKAANSE OPSOMMING: Aflatoksiene is mikotoksiene wat hoofsaaklik deur die filamentagtige fungi, Aspergillus flavus en Aspergillus parasiticus geproduseer word. Die algemeenste aflatoksien, aflatoksien B1 (AFB1), is hoogs mutagenies, toksies, karsinogenies en teratogenies vir mense en diere. Veral in sekere dele van Afrika, China en Suid-Oos Asië bestaan daar `n korrelasie tussen aflatoksien en die voorkoms van hepatosellulêre karsinoom en gevolglik word aflatoksiene as `n tipe I menslike karsinogeen deur die Internasionale Agentskap vir Kankernavorsing geklassifiseer. Aflatoksien kontaminasie in voedsel het ook `n ekonomiese impak as gevolg van verlies aan landbougewasse en diere. Ten spyte van maatreëls betreffende die toelaatbare vlakke van aflatoksiene in voedel, is aflatoksien kontaminasie steeds `n groot probleem wêreldwyd, veral in ontwikkelende lande waar dit hoofsaaklik in stapelvoedsel voorkom. Huidiglik is die inaktivering van aflatoksiene deur fisiese en chemiese metodes nie effektief en ekonomies nie. Daarteenoor bied biologiese tegnieke `n gunstige opsie vir die eliminering van die toksiene, terwyl die organoleptiese eienskappe van die voedsel steeds behoue bly. Hierdie studie fokus op die biologiese afbraak van AFB1 deur bakterieë en fungi. Verskeie bakterieë, insluitend Rhodococcus spp., sowel as witvrot fungi produseer `n verskeidenheid ensieme wat hulle in staat stel om `n wye reeks polisikliese hidrokoolstofverbindings af te breek en gevolglik is afbraak van AFB1 deur sommige van hierdie mikroörganismes bestudeer. Effektiewe afbraak van AFB1 deur intrasellulêre ekstrakte van Mycobacterium fluoranthenivorans sp. nov. DSM 44556T, Nocardia corynebacterioides DSM 20151 en N. corynebacterioides DSM 12676 is aangetoon. AFB1 is ook effektief in vloeibare kulture sowel as intra- en ekstrasellulêre ekstrakte van Rhodococcus erythropolis DSM 14303 afgebreek. `n Beduidende (P<0.001) afbraak van AFB1 is waargeneem na behandeling met R. erythropolis DSM 14303 ekstrasellulêre ekstrakte, met slegs 33.20% oorblywende AFB1 na 72 h. Resultate het getoon dat die afbraak deur R. erythropolis DSM 14303 ensimaties is en dat die ensieme konstitutief geproduseer word. Afbraak van AFB1 deur R. erythropolis DSM 14303 het ook tot `n totale verlies aan mutagenisiteit gelei. Verder het behandeling van AFB1 met rekombinante 2,3-dihidroksiebifenieldioksiginase fraksies wat geproduseer is deur ekstrasellulêre uitdrukking van die bphC1 geen van R. erythropolis DSM 14303 in Escherichia coli BL21, beduidende (P<0.0001) afbraak (49.32%) en vermindering in mutagenisiteit (42.47%) van die molekuul teweeggebring. Beduidende (P<0.0001-0.05) afbraak van AFB1 is verkry na behandeling met witvrot fungus kultuurekstrakte wat lakkase-ensiem bevat (17.10-76.00%), gesuiwerde lakkase geproduseer deur Trametes versicolor (1 U/ml, 87.34%), sowel as rekombinante lakkase geproduseer deur Aspergillus niger (118 U/L, 55.00%). Verder het die behandeling van AFB1 met gesuiwerde lakkase-ensiem (1 U/ml) gelei tot verlies aan mutagenisiteit van AFB1. Die afname in fluoressensie en mutageniese eienskappe van die AFB1-molekuul na behandeling met die onderskeie mikrobiese preparate dui op struktuurveranderings aan die furofuraan- en/of laktoonringe van die molekuul. Hierdie studie lewer `n bydrae tot die ontwikkeling van geneties gemanipuleerde mikrobiese rasse wat as `n belangrike biokontrole kan dien. Sulke rasse met multifunksionele tegnologiese eienskappe, insluitend die afbraak van AFB1, kan die kwaliteit, veiligheid en aanvaarbaarheid van voedsel verbeter.

Submitted in partial fulfillment of the requirements for the degree of Doctor of Technology: Clinical Technology, Durban Institute of Technology, 2005. / Fumonisin B1 (FB1) and ochratoxin A (OTA) represent examples of mycotoxins of greatest public health and agro-economic significance. They exert adverse effects on humans, animals and crops that result in illnesses and economic losses. Fumonisin B1 are cancerpromoting metabolites of Fusarium proliferatum and F verticillioides, (formerly moniliforme), and are implicated in oesophageal cancer. Ochratoxins are metabolites of both Aspergillus and Penicillium species. These compounds are known for their nephrotoxic effects in all animal species and may promote tumours in humans. In man OTA exhibits unusual toxicokinetics, with a half-life in blood of 840 h (35 days) after oral ingestion. Although much is known regarding the toxicology of these toxins, little is known of the effects of these toxins on the immune system. The aim of this study was to determine and compare the immunornodulating effects of FB1 and OTA in human carcinoma. Initial experiments involved isolating lymphocytes and neutrophils from healthy volunteers. The isolated cells were exposed to either FB1 or OTA on a dose and time dependent level and LD50 of the toxins was determined. Thereafter, challenge tests were performed, whereby lymphocytes and neutrophils isolated from volunteers, oesophageal cancer patients and breast cancer patients were exposed to the LD50 dose of either FB1 or OTA for the appropriate time. The effect of the toxins was demonstrated by viability studies, light microscopy and electron microscopy. Cytokine receptors (CK, TNF and CSF) were evaluated by immuno-cytochemical methods and the levels of circulating cytokines (IL -1, IL-6, IL-8, IL-10 and TNF-a) were determined using ELISA kits. / D

Fusarium head blight (FHB) continues to threaten the economic sustainability of wheat and barley production in Canada and worldwide. The overall goal of this thesis is to expand our current knowledge of the FHB pathogen, Fusarium graminearum and its trichothecene chemotype diversity. Continuous monitoring of trichothecene chemotypes may well inform on the potential risk and the type of Fusarium populations present in a given region. Fusarium populations in Winnipeg and Carman, Manitoba were examined using chemotype as a marker in the field. Rapid expansion of the 3-acetyldeoxynivalenol (3-ADON) chemotype was observed in Winnipeg and Carman. 3-ADON chemotype is consistently found at high frequencies over the previously common 15-acetyldeoxynivalenol (15-ADON) chemotype, suggesting that the shift in pathogen populations is continuing. This study provides the first evidence on the presence of nivalenol (NIV) producing F. cerealis strains in winter wheat in Manitoba, Canada. Therefore, discovery of NIV producing F. cerealis in wheat poses a serious concern for the wheat industry in Canada. Phylogenetic, chemotypic, phenotypic, and pathogenic abilities of 150 strains of F. graminearum species complex (FGSC) from eight countries were investigated. Type and amount of trichothecenes produced by a strain are key factors in determining the level of aggressiveness of that strain regardless of its species origin. The sequence variations of TRI8 gene in different species in the FGSC were examined as Fusarium species may produce different types of trichothecenes depending on differences in the core trichothecene (TRI) cluster genes. The TRI8 haplotypes did group according to chemotype rather than by species, indicating that 3-ADON, 15-ADON and NIV chemotypes have a single evolutionary origin. Comparison of TRI gene expression demonstrated that accumulation of TRI transcripts was higher in 3-ADON producing strains compared to 15-ADON and NIV strains. The presence of masked mycotoxins deoxynivalenol-3-glucoside (D3G) in food and feed is an increasing concern. Canadian spring wheat cultivars inoculated with different chemotypes produce D3G upon Fusarium infection and moderately resistant/intermediate cultivars showed higher D3G/DON ratio compared to susceptible cultivars. / October 2016

Mycotoxins are secondary metabolites produced by many fungal species. Health effects induced by the ingestion of these substances are well documented and some mycotoxins are now regulated for their maximum tolerable levels in foods. However, other routes of exposure to these contaminants are possible. Thus, if irritating or allergenic reactions related to the inhalation of fungal spores or mycelial fragments have been demonstrated, inhalation of mycotoxins is also suspected to be causing certain respiratory disorders or certain pathologies. Indeed, mycotoxins can be found in spores but also on finer particles which are easily aerosolized and therefore likely to be inhaled. However, data on the hazard associated with human exposure to mycotoxins by inhalation are still very fragmented. In this context, our main objective was to characterize the aerosolization of mycotoxins during the colonization of different materials encountered in indoor environments by toxinogenic molds. First we studied growth and production of mycotoxins during the colonization of building materials (wallpaper, painted fiberglass wallpaper, vinyl wallpaper, fir, fiberglass) by three fungal species of interest: Aspergillus versicolor, Penicillium brevicompactum, Stachybotrys chartarum. These species were chosen because of their frequent presence in indoor environments and their diverse mycelial organization. In addition, these three species produce different toxins: sterigmatocystin, mycophenolic acid and macrocyclic trichothecenes for A. versicolor, P. brevicompactum and S. chartarum, respectively. These studies have shown that, during their development on tested materials, three species produce mycotoxins. The most favorable material for fungal development and toxinogenesis is wallpaper. Mycophenolic acid, sterigmatocystin and macrocyclic trichothecenes can thus be produced at levels of 1.8, 112.1 and 27.8 mg/m2, respectively, on this material. These toxins can then be partially aerosolized. We have shown that aerosolization depends on species and their mycelial structure, but also on culture conditions and airflow. This transfer to air is nevertheless observed after aeraulic solicitations which can be easily encountered in indoor environments because theycorrespond to the movement of people in a room (0.3 m/s), speed of air in ceiling diffusers (2 m/s), slamming doors or air drafts when opening windows(6 m/s). P. brevicompactum showed to be the easiest to aerosolize. The major part of the aerosols’ toxic charge is found in particles whose size corresponds to that of spores or mycelial fragments. However, for macrocyclic trichothecenes, toxins were also found in particles smaller than spores, which could easily be inhaled by occupants and penetrate deep into the respiratory tract. In order to better characterize the actual hazard associated with inhalation of these compounds, cytotoxicity studies have been performed using lung cells and comparing with results observed on digestive cells. Pulmonary toxicity is comparable to that observed in digestive cells. Macrocyclic trichothecenes are much more toxic than other tested toxins with IC50 in order of ng/ml. In parallel, we analyzed the VOCs specifically produced during active mycotoxinogenesis in order to identify potential biomarkers of the actual production of mycotoxins that could be used as tools for monitoring of indoor environments. Unfortunately, this approach has not, for the moment, led to the identification of specific targets. In the end, we evaluated the persistence of these contaminants during application of bleach, the most frequently used decontamination process. We have shown that a normal cleaning procedure allows only partial removal of mold.

Mycotoxins are feed contaminants that are a major problem in the livestock industry because of their prevalence in feedstuffs and the difficulty of removing them. They can cause a wide range of issues at varying levels of exposure. Each species is affected by different mycotoxins and at different levels. Pigs are more susceptible to deoxynivalenol (DON), whereas ducks are more susceptible to aflatoxin.<br> Effects of mycotoxin contamination on animal performance are not fully understood. Therefore, the two experiments described in this thesis were conducted to determine the response of pigs and ducks to consumption of feed contaminated with DON and aflatoxin, respectively. In the first experiment, the effect of a mycotoxin binder on duck feeds contaminated with aflatoxin was examined. One-day-old male Pekin ducks (n=360) were randomly divided into four groups; each group had 6 replicate pens with 15 ducks per replicate pen. The positive control (PC) group was fed a diet that was free of aflatoxin B1, the negative control (NC) group was fed a diet that contained >75ppb of aflatoxin without a binder, the negative control with low binder (NC + 0.5) group was fed a diet that contained >75ppb of aflatoxin and 0.5 kg/ton of the binder, the negative control with high binder (NC + 1.0) group was fed a diet that contained >75ppb of aflatoxin and 1.0 kg/ton of the binder. The diets were fed in two phases, days 0-14 (phase 1) and 15-35 (phase 2). The results showed that during early phase 2, NC + 0.5 resulted in a higher rate of weight gain compared to NC (P<0.05); 2) NC + 0.5 ducks had higher feather quality than both NC and PC (P<0.05); 3) NC had higher relative liver weights (P<0.05); 4) blood glucose was higher in NC + 0.5 ducks (P<0.05); and 5) PC ducks had higher serum protein levels in the blood (P<0.05).<br> In the second study, effect of the same mycotoxin binder, used in the duck study, was examined in pigs fed diets contaminated with DON. A total of 128 pigs (Duroc × Landrace × Yorkshire, (1:1 barrows and gilts, aged 42 d) were randomly assigned to 4 treatments, 8 replicate pens with 4 pigs per. The treatments were DON, DON + liver protectant (1 kg/ton), DON + mycotoxin binder (0.5 kg/ton), or DON + liver protectant and mycotoxin binder. The study lasted 28 days and body weights (BW), feed intake (FI), and blood samples were taken on days 14 and 28. Body weights and feed intake were taken and used to calculate gain:feed (G:F). Aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were measured in the blood serum. BW, FI, and G:F were not significantly different at any point during the study. AST levels were significantly reduced (P < 0.05) on day 14 in pigs fed the liver protectant but were not significantly different day 28.<br> In summary, effects of the use of mycotoxin binders in feed can be highly variable. This depends on the type of mycotoxin present in the feed, the amount of mycotoxin, and the species fed the diet. In the present study, the mycotoxin binder did not have an impact on the feed efficiency of the ducks or pigs. Effects of additional binders need to be evaluated for their effectiveness in mitigating the negative effects of mycotoxins.<br><br>

Ochratoxin A (OA), a nephrotoxin and potential carcinogen, has been found in many foods, including grapes and grape products. Limits of 2 μg/kg in wine and 10 μg/kg in dried vine fruit have been introduced by the European Union. This study presents information on the ecology of ochratoxin A production by black Aspergillus spp. in Australian vineyards, and the passage of the toxin throughout winemaking. Aspergillus niger and A. carbonarius were isolated from vineyard soils in 17 of 17, and four of 17 Australian viticultural regions, respectively. A. aculeatus was isolated infrequently. All thirty-two isolates of A. carbonarius and three of 100 isolates of A. niger produced OA. Of Australian A. niger isolates analysed for restriction fragment length polymorphisms within the internal transcribed spacer region of 5.8S ribosomal DNA, 61 of 113 isolates, including the three toxigenic isolates, were of type N pattern, and 52 were type T. A selection of these A. carbonarius and A. niger aggregate isolates, as well as imported isolates, were compared using enterobacterial repetitive intergenic consensus (ERIC)-PCR, amplified fragment length polymorphisms (AFLP) and microsatellite markers. ERIC and AFLP clearly differentiated A. niger from A.carbonarius. AFLP further divided A. niger into types N and T. Six polymorphic microsatellite markers, developed specifically for A. niger, also differentiated strains into N and T types. There was no clear relationship between genotypic distribution and ochratoxigenicity, substrate or geographic origin. The survival of A. carbonarius spores on filter membranes was examined at water activities (aw) 0.4-1.0, and at 1 °C, 15 °C, 25 °C and 37 °C. Survival generally increased at lower temperatures. The lowest water activity, 0.4, best supported the survival of spores, but 0.6- 0.9 aw was often deleterious. Complex interactions between temperature and water activity were observed. Viability of A. carbonarius spores on filter membranes decreased ca 10[superscript 5] fold upon exposure to sunlight, equivalent to 10 mWh of cumulative ultraviolet irradiation at 290-400 nm. Growth and toxin production were examined for five isolates of A. carbonarius and two of A. niger on solid medium simulating juice at early veraison, within the range 0.98-0.92 aw, and at 15 °C, 25 °C, 30 °C and 35 °C. Maximum growth for A. carbonarius and A. niger occurred at ca 0.965 aw / 30 °C and ca 0.98 aw / 35 °C, respectively. The optimum temperature for OA production was 15 °C and little was produced above 25 °C. The optimum aw for toxin production was 0.95 for A. niger and 0.95-0.98 for A. carbonarius. Toxin was produced in young colonies, however, levels were reduced as colonies aged. Black Aspergillus spp. were more commonly isolated from the surface than from the pulp of berries, and increased with berry maturity, or damage. A. niger was isolated more frequently than A. carbonarius and A. aculeatus. Populations of A. carbonarius inoculated onto bunches of Chardonnay and Shiraz decreased from pre-bunch closure to early veraison. Populations from veraison to harvest were variable, and ncreased in bunches with tight clustering and splitting. In a trial with Semillon bunches, omitting fungicide sprays after flowering did not increase the development of Aspergillus rot. Inoculation of bunches with A. carbonarius spore suspension did not necessarily result in Aspergillus bunch rot. In vitro trials suggested that the severity of rot was mediated primarily by the degree of berry damage, followed by the extent of spore coverage. No clear trends regarding cultivar susceptibility were observed. For Semillon bunches inoculated with A. carbonarius spores with and without berry puncture, increased susceptibility to rot and OA formation was associated with berry damage, in particular at greater than 12.3 °Brix (20 d before harvest). OA contamination of bunches was related to the number of mouldy berries per bunch, with shrivelled, severely mouldy berries the primary source of OA. Puncture-inoculation of white grapes (Chardonnay and Semillon) and red grapes (Shiraz) on the vine with A. carbonarius resulted in berries containing OA. Inoculated grapes displayed greater total soluble solids due to berry shrivelling, and greater titratable acidity due to production of citric acid by the fungus. Samples taken throughout vinification of these grapes were analysed for OA. Pressing resulted in the greatest reduction in OA (68-85% decrease in concentration, compared with that of crushed grapes). Additional reductions occurred at racking from grape and gross lees, and after storage. OA was removed by binding to marc, grape and gross lees. Pectolytic enzyme treatment of white must, bentonite juice fining, recovery of juice or wine from lees, and static or rotary style fermentation of red must, had no effect on OA contamination. Bentonite in white wine (containing 56 mg/L grape-derived proteins) and yeast hulls in red wine were effective fining agents for removing OA. Findings from these studies may contribute to the improvement of strategies to minimise OA in Australian wine and dried vine fruit. / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture and Wine, 2005.