Abstract
BackgroundBrazil nut is a protein-rich extractivist tree crop in the Amazon region. Fungal contamination of shells and kernel material frequently includes the presence of aflatoxigenic Aspergillus species from the section Flavi. Aflatoxins are polyketide secondary metabolites, which are hepatotoxic carcinogens in mammals. The objectives of this study were to identify Aspergillus species occurring on Brazil nut grown in different states in the Brazilian Amazon region and develop a specific PCR method for collective identification of member species of the genus Aspergillus.ResultsPolyphasic identification of 137 Aspergillus strains isolated from Brazil nut shell material from cooperatives across the Brazilian Amazon states of Acre, Amapá and Amazonas revealed five species, with Aspergillus section Flavi species A. nomius and A. flavus the most abundant. PCR primers ASP_GEN_MTSSU_F1 and ASP_GEN_MTSSU_R1 were designed for the genus Aspergillus, targeting a portion of the mitochondrial small subunit ribosomal RNA gene. Primer specificity was validated through both electronic PCR against target gene sequences at Genbank and in PCR reactions against DNA from Aspergillus species and other fungal genera common on Brazil nut. Collective differentiation of the observed section Flavi species A. flavus, A. nomius and A. tamarii from other Aspergillus species was possible on the basis of RFLP polymorphism.ConclusionsGiven the abundance of Aspergillus section Flavi species A. nomius and A. flavus observed on Brazil nut, and associated risk of mycotoxin accumulation, simple identification methods for such mycotoxigenic species are of importance for Hazard Analysis Critical Control Point system implementation. The assay for the genus Aspergillus represents progress towards specific PCR identification and detection of mycotoxigenic species.
Highlights
Brazil nut is a protein-rich extractivist tree crop in the Amazon region
Identification and abundance of Aspergillus species Polyphasic identification of all 137 Aspergillus strains isolated from Brazil nut shell material collected from cooperatives across the Brazilian Amazon region revealed the presence of five species, with three belonging to Aspergillus section Flavi
When comparing A. nomius and A. flavus, similar numbers of strains were identified in total, numbers varied considerably across regions, Table 1 Frequency of Aspergillus species from Brazil nut material across the Brazilian Amazon region
Summary
Brazil nut is a protein-rich extractivist tree crop in the Amazon region. Fungal contamination of shells and kernel material frequently includes the presence of aflatoxigenic Aspergillus species from the section Flavi. Aflatoxins (AF) are polyketide family secondary metabolites produced by several members of the fungal genus Aspergillus, section Flavi. Considered amongst the most dangerous natural hepatotoxic carcinogens in mammals [1], consumption of foodstuffs contaminated with these extrolites can be a cause of mortality and reduced productivity in higher vertebrates. Within this family, AFB1, B2, G1 and G2 cause most concern, given their abundance and toxicity [2]. Aspergillus section Flavi comprises over 20 member species, based on polyphasic approaches for species delimitation that consider morphological, molecular and extrolite data [7,8,9,10]. A number of species within the section are aflatoxigenic, including the widely distributed species A. flavus, A. parasiticus and A. nomius, together with A. arachidicola, A. bombycis, A. minisclerotigenes, A. parvisclerotigenus, A. pseudocaelatus, A. pseudonomius and A. pseudotamarii, ([7] and references therein), A. novoparasiticus [8], A. mottae, A. sergii and A. transmontanensis [9]
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