Deoxynivalenol (DON) contamination was investigated of Italian durum wheat from organic agriculture. A number of 661 samples from 13 genotypes were collected within the national organic durum wheat network variety trials during the six-year period between 2007–2012 in five different growing areas across Italy (Northern Italy, Marches, Central Apennines, West-Central Italy, Apulia). Mean temperatures and total rainfalls in April, May and June were collected nearby the study sites. Average DON contamination value along the whole study period was 67 μg/kg, and DON was detected only in 36% of the samples. Noteworthy, 95% of the analyzed grain revealed a DON contamination lower than 334 μg/kg. Maximum allowed DON level for unprocessed durum wheat set by European Union (1750 μg/kg) was exceeded only in four samples (0.6%). The highest mean DON values were detected in Northern Italy (175 μg/kg) and Marches (131 μg/kg). The same was for the percentage of positive samples (80% and 58%, respectively). Lower mean values and percentages of contaminated samples were found in West-Central Italy (22 μg/kg and 29%, respectively), Apennines (3 μg/kg and 8%, respectively) and Apulia (2 μg/kg and 7%, respectively). Statistical analysis (Generalized Linear Model, GLZ) was carried out to highlight the effect of factors like cultivation year, growing area and genotype. It revealed a huge effect of year, growing areas and their interaction, while the effect of genotype resulted significantly but quite less than the other main factors. The effect of the year could be explained by climatic data, which suggested an influence of rainfall and temperature at heading on both DON concentration values and percentage of contaminated samples. Results of this study put in evidence a low DON contamination in Italian organic durum wheat.