Victoria Bernáldez

Influence of temperature and substrate conditions on the omt-1 gene expression of Aspergillus parasiticus in relation to its aflatoxin production

Most strains of Aspergillus parasiticus are able to produce high concentrations of aflatoxin (AF) B1 and G1 which are among the most potent mutagenic, teratogenic, and carcinogenic mycotoxins. Molecular studies in relation to activity of secondary metabolite gene clusters are crucial to improving food safety. In the present work, reverse transcriptase quantitative PCR (RT-qPCR) was used to monitor the influence of temperature, substrates containing nitrogen and incubation time on the omt-1 gene expression in A. parasiticus. Phenotypic AFB1 and G1 production was also evaluated by high-performance liquid chromatography-mass spectrometry (HPLC-MS). The results demonstrated that temperature (25°C and 30°C) influenced relative expression of omt-1 gene throughout the time (maximum at 25°C) while substrate composition was not affected by it. However, when effect of temperature and substrate was analyzed at each incubation time, significant effects were found. Optimal conditions for biosynthesis of AFB1 and AFG1 were similar, and they were related to changes in temperature and sodium nitrate. The highest AFB1 and G1 production levels were found at 25°C. However, lower AFB1 and G1 values were obtained when A. parasiticus grew on the substrate containing sodium nitrate and there was no production of these AFs at 37°C in any of the conditions tested. In addition, omt-1 gene expression was correlated to AFB1 and G1 syntheses at the different conditions. Use of temperature conditions and sodium nitrate concentrations which limit production of AFs holds potential for preventing AF from entering the food chain.

Effect of temperature and water activity on growth and aflatoxin production by Aspergillus flavus and Aspergillus parasiticus on cured meat model systems

Dry-cured hams may be colonised by aflatoxin-producing Aspergillus flavus and Aspergillus parasiticus during the ripening process. The objective of this study was to evaluate the interaction between non-ionic water stress and temperatures may have on lag phases prior to growth, growth rates and aflatoxin production by two strains of each A. parasiticus and A. flavus on meat matrices over a period of 12days. Results showed that A. flavus CBS 573.65 had shorter lag phases than A. parasiticus CECT 2688, however the growth rates were quite similar. For both species, no growth occurred at 10°C and all aw tested and optimum growth happened at 25°C and 0.95 aw. Similar aflatoxin B1 production profiles between both species were found, however A. flavus produced much higher concentration of such toxin than A. parasiticus. Both species produced aflatoxins when the temperature and the aw were ≥15°C and ≥0.90.

Identification and control of moulds responsible for black spot spoilage in dry-cured ham.

The aims of this work were to identify moulds responsible for black spot spoilage in the drying and cellar stages of dry-cured ham processing and evaluate the effectiveness of preventive actions for controlling this alteration. Four mould strains isolated from spoiled hams were identified by morphological characteristics and the ITS and β-tubulin sequencing. Two of them were Cladosporium oxysporum, one was C. cladosporioides and the remaining one was C. herbarum. These spoiling strains reproduced the black spots on dry-cured ham-based media and ham slices. Additionally, the effect of water activity (aw) conditions reached throughout dry-cured ham ripening and the activity of the protective culture Penicillium chrysogenum CECT 20922 against the spoiling moulds were evaluated. In the dry-cured ham model system the growth of the Cladosporium strains was minimised when the aw approaches 0.84 or in P. chrysogenum CECT 20922 inoculated dry-cured ham slices. Therefore such combination could be used to avoid the black spot formation in dry-cured ham.

Gene Expression Analysis as a Method to Predict OTA Accumulation in Dry-Cured Meat Products

Dry-cured meat products are frequently colonised by toxigenic Penicillium nordicum and Penicillium verrucosum throughout their ripening that can produce ochratoxin A (OTA). The predictive nature of the molecular analyses can be used to determine the risk of toxigenic species. For this reason, the objective of the present work was to analyse the temporal changes in the expression of the otapks and otanps genes by P. nordicum and P. verrucosum in relation to OTA production on slices of dry-fermented sausage salchichón and dry-cured ham. Gene expression was higher in P. nordicum than in P. verrucosum in both meat matrices. The otapks gene was overexpressed by both Penicillium species, especially in salchichón. OTA was only detected in inoculated salchichón regardless of the ochratoxigenic species used. The high significant correlation found between the early relative expression of the otapks gene and OTA production in salchichón leads to propose the relative gene expression of the otapks gene as a good indicator to predict OTA accumulation throughout the ripening of this product.