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The distribution and co-occurrence of four Fusarium species and their mycotoxins were investigated in maize samples from two susceptible cultivars collected at 14 localities in South Africa during 2008 and 2009. Real-time PCR was used to quantify the respective Fusarium species in maize grain, and mycotoxins were quantified by multi-toxin analysis using HPLC-MS. In 2008, F. graminearum was the predominant species associated with maize ear rot in the eastern Free State, Mpumalanga and KwaZulu-Natal provinces, while F. verticillioides was predominant in the Northwest, the western Free State and the Northern Cape provinces. In 2009, maize ear rot infection was higher and F. graminearum became the predominant species found in the Northwest province. Fusarium subglutinans was associated with maize ear rot in both years at most of the localities, while F. proliferatum was not detected from any of the localities. Type B trichothecenes, especially deoxynivalenol, and zearalenone were well correlated with the amount of F. graminearum, fumonisins with F. verticillioides, and moniliformin and beauvericin with F. subglutinans. This information is of great importance to aid understanding of the distribution and epidemiology of Fusarium species in South Africa, and for predicting mycotoxin contamination risks and implementing preventative disease management strategies.

Maize streak virus strain A (MSV-A), the causal agent of maize streak disease, is today one of the most serious biotic threats to African food security. Determining where MSV-A originated and how it spread transcontinentally could yield valuable insights into its historical emergence as a crop pathogen. Similarly, determining where the major extant MSV-A lineages arose could identify geographical hot spots of MSV evolution. Here, we use model-based phylogeographic analyses of 353 fully sequenced MSV-A isolates to reconstruct a plausible history of MSV-A movements over the past 150 years. We show that since the probable emergence of MSV-A in southern Africa around 1863, the virus spread transcontinentally at an average rate of 32.5 km/year (95% highest probability density interval, 15.6 to 51.6 km/year). Using distinctive patterns of nucleotide variation caused by 20 unique intra-MSV-A recombination events, we tentatively classified the MSV-A isolates into 24 easily discernible lineages. Despite many of these lineages displaying distinct geographical distributions, it is apparent that almost all have emerged within the past 4 decades from either southern or east-central Africa. Collectively, our results suggest that regular analysis of MSV-A genomes within these diversification hot spots could be used to monitor the emergence of future MSV-A lineages that could affect maize cultivation in Africa.

Maize streak virus (MSV), which causes maize streak disease (MSD), is one of the most serious biotic threats to African food security. Here, we use whole MSV genomes sampled over 30 years to estimate the dates of key evolutionary events in the 500 year association of MSV and maize. The substitution rates implied by our analyses agree closely with those estimated previously in controlled MSV evolution experiments, and we use them to infer the date when the maize-adapted strain, MSV-A, was generated by recombination between two grass-adapted MSV strains. Our results indicate that this recombination event occurred in the mid-1800s, ∼20 years before the first credible reports of MSD in South Africa and centuries after the introduction of maize to the continent in the early 1500s. This suggests a causal link between MSV recombination and the emergence of MSV-A as a serious pathogen of maize.

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Fumonisins are secondary, carcinogenic metabolites produced primarily by Fusarium verticillioides and Fusarium proliferatum on maize worldwide. The natural occurrence of fumonisin-producing Fusarium spp. and fumonisin contamination of maize grain were quantified in selected maize cultivars from principal production areas of South Africa. Grain colonization by Fusarium spp. was determined using quantitative real-time PCR (qPCR) and contamination with fumonisins using HPLC analysis. Kernels from the 2007 samples were also plated onto Fusarium selective medium and subsequently, split plates containing PDA & CLA. The number of fumonisin producing Fusarium spp. were quantified and microscopically identified after 14 days. Simple linear regression analysis was used to determine the relationship between target DNA, fumonisins and the number of fumonisin producing Fusarium spp. using the plating out method. Results indicated high natural infection by fumonisinproducing Fusarium spp. and fumonisin concentrations in warmer production areas such as Northern Cape, North West and Free State Provinces. Spearman Ranking Correlations indicated that the responses of cultivars to colonization of grain by fumonisin producing Fusarium spp. varied over localities/seasons (r s =0.42 to 0.64) suggesting that cultivars reacted differently to different environmental/inoculum conditions (disease potentials). Cultivars CRN3505 and DKC80-12B showed a degree of resistance to fungal infection. As with fungal colonization, Spearman Rank Correlations indicated the response of cultivars to fumonisin contamination to vary over localities/ seasons (r s =0.29 to 0.70). Cultivars DKC80-12B and LS8521B showed a degree of resistance to fumonisin contamination. Regression analysis yielded a significant relationship between HPLC data and qPCR, but not with the plating out of grain data suggesting the former to be a better indicator of potential fumonisin contamination. Site-specific, daily maximum temperature and rainfall data were provided by the ARC-Institute for Soil Water and Climate's meteorology office. No significant relationship between these weather parameters and colonization of grain by fumonisin producing Fusarium spp. was recorded, although a tendency was observed between fumonisin contamination and mean maximum temperature.

A quantitative detection tool was developed to enable the monitoring of fumonisin-producing fungi in food and feed commodities. To this end, a quantitative PCR (TaqMan) was developed that targets a conserved region in the polyketide synthase gene fum1, which is involved in the biosynthesis of fumonisin. Hence, this method specifically detected isolates from the fumonisin-producing species Fusarium verticillioides, F. proliferatum, F. nygamai and F. globosum whereas isolates of the fumonisin non-producing species F. equiseti, F. graminearum, F. oxysporum, F. semitectum and F. subglutinans that commonly occur on maize were not detected. Moreover, a few fumonisin non-producing F. verticillioides isolates did not generate any fluorescent signals and were therefore not detected. The correlation between quantitative PCR and mycotoxin content was determined using field samples collected at homestead farms in South Africa. Among 40 samples from the Eastern Cape collected in 2005 a good correlation (R2=0.8303) was found between pg fungal DNA and fumonisin content. A similar correlation (R2=0.8658) was found among 126 samples collected from four provinces in South Africa in 2007. These observations indicate that samples containing ≥ 40 pg fungal DNA/mg sample are suspected of also exceeding the 1 mg/kg total fumonisin level and therefore do not comply with the European Commission limit for fumonisins B1+B2 for maize intended for direct human consumption that applies from 1 October 2007. Combined with the very high maize intake, our results indicate that fumonisin levels in maize from South African homesteads regularly exceed the tolerable daily intake for fumonisins.

Fusarium verticillioides causes Fusarium ear rot (FER) of maize and produces fumonisins, which affects grain quality. Host-plant resistance can reduce both FER and fumonisins in maize. In this study, 18 maize inbred lines were evaluated for resistance to F. verticillioides and fumonisin accumulation at five localities in South Africa. Additive main effects and multiplicative interaction analyses revealed significant environment × genotype interactions, with inbred lines CML 390, US 2540W, RO 424W, and VO 617y-2 consistently exhibiting low FER severity (≤5.4%), fungal target DNA (≤0.1 ng μl−1), and fumonisin levels (≤5.6 ppm). Genotype main effect and genotype × environment biplots showed that inbred lines CML 390, US 2540W, and RO 424W were most resistant to FER, fungal colonization, and fumonisin accumulation, respectively, while inbred line RO 424W was most stable in its resistance response over environments. These inbred lines also demonstrated broad adaptability by consistently exhibiting resistance to FER, fungal colonization, and fumonisins across localities. The identified lines could serve as valuable sources of resistance against F. verticillioides and its fumonisins in local breeding programs.