Anti-Fumonisin monoclonal antibody, clone 4C4

The CRISPR/Cas9 system is a revolutionary genome editing tool. However, in eukaryotes, search and optimization of a suitable promoter for guide RNA expression is a significant technical challenge. Here we used the industrially important fungus, Aspergillus niger, to demonstrate that the SS rRNA gene, which is both highly conserved and efficiently expressed in eukaryotes, can be used as a guide RNA promoter. The gene editing system was established with 100% rates of precision gene modifications among dozens of transformants using short (40-bp) homologous donor DNA. This system was also applicable for generation of designer chromosomes, as evidenced by deletion of a 48 kb gene cluster required for biosynthesis of the mycotoxin fumonisin Bl. Moreover, this system also facilitated simultaneous mutagenesis of multiple genes in A. niger. We anticipate that the use of the 5S rRNA gene as guide RNA promoter can broadly be applied for engineering highly efficient eukaryotic CRISPR/Cas9 toolkits. Additionally, the system reported here will enable development of designer chromosomes in model and industrially important fungi.

Mycotoxins are secondary metabolites that are produced by filamentous mycotoxigenic fungi belonging to theAlternaria, Aspergillus, FusariumandPenicilliumgenera amongst others. Multi-class mycotoxins were extracted from 55 cereal samples and analysed using liquid chromatography-tandem mass spectrometry. The adopted extraction method for maize, maize porridge, sorghum and wheat was comprehensively validated. This method was further tested to determine the natural occurrence of mycotoxins in foodstuffs. Twelve (12) out of 22 mycotoxins were detected in maize, maize porridge, sorghum and wheat, including alpha-zearalenol (alpha-ZEL) (89%), fumonisin B-3(FB3) (84%), fumonisin B-1(FB1) (80%), tenuazonic acid (TeA) (78%), ochratoxin B (42%), deoxynivalenol (DON) (12%), ochratoxin A (11%), 3-acetyldeoxynivalenol (7%), sterigmatocystin (STG) (6%), 15-acetyldeoxynivalenol (2%), cyclopiazonic acid (CPA) (2%) and aflatoxin B-2(2%). The data revealed high incidence rate of alpha-ZEL (range: 6.5-70.5 mu g kg(-1)) in all matrices. Maize samples had high mycotoxin co-occurrence compared to other matrices. All recovered mycotoxins in food commodities were within the maximum regulatory limits, with the exception of fumonisins (FB(1)and FB3) exceeded the South African and European Commission regulation, and the highest concentration was 2153 mu g kg(-1)in maize. It is essential to monitor the level of emerging mycotoxins in food commodities from rural areas as trace amount of CPA (< limit of quantification), STG (range: 0.30-0.74 mu g kg(-1)) were detected and high concentration of TeA (292.7 mu g kg(-1)) was detected in sorghum. The occurrence of these mycotoxins further encourages frequent analyses, their co-occurrence in the samples poses a significant threat to public health and more emphasis should thus be placed on reducing the contamination levels of these toxins in staples.