By Dr Scott Pirie
The EGSF-funded joint pilot study between researchers at Fera (Fera Science Ltd; formerly the Food and Environment Research Agency and the Royal (Dick) School of Veterinary Studies, exploring the potential causal role of the pasture fungi Fusarium in Equine Grass Sickness (EGS), is now complete and has generated some interesting data.
During 2014 and 2015, grass samples were collected randomly from 8 premises on which EGS had occurred; both from the “affected” pasture and from a neighbouring “control field”. Either individual or pooled samples were subsequently subjected to various analyses including (a) pasture Fusarium spore counts, (b) pasture Fusarium species isolation and chemotyping (analysis of their mycotoxin-producing capacity) and (c) mycotoxin screening. Additionally, archived gastrointestinal contents from EGS cases were also analysed for the presence of mycotoxins.
Interestingly, the median spore count for all premises, with one exception, was greater in the “affected” fields than the paired “control” fields. Although no statistically significant difference could be identified between paired fields, this likely reflects the inherent limitation in the number of samples which could be analysed and therefore the number of premises sampled. Furthermore, there was marked variation in spore count across each pasture, an inherently problematic factor when interpreting such data without knowledge of the preceding grazing practices of the affected cases. In all 5 premises analysed for Fusarium speciation, 3 main Fusarium species were isolated and, in agreement with the spore counts, at 4 out of the 5 premises, Fusarium levels were generally lower in the ‘control’ fields than in the “affected” field (Figure 1). Interestingly, at the single site where this pattern was not observed, samples were collected several weeks following the occurrence of disease. This analysis also confirmed marked variation in the distribution of Fusarium species within individual fields.
Chemotyping analysis of the Fusarium species isolated from the grass samples, did provide evidence that the toxins produced by the fungus when infecting grass were different to those produced during infection of wheat. The facilities available at Fera permitted subsequent testing of samples obtained from the 5 sites for 72 different mycotoxins. The mycotoxin profiles obtained from a selection of premises was highly consistent with the previously confirmed presence of Fusarium species, although additional toxins were detected which are likely derived from other fungal species. In some sites, certain mycotoxin levels were greater in the “affected” field, compared with the “control” field.
In light of the diversity of Fusarium species plus patchy distribution of spores and mycotoxin within sampled fields and the difficulties this phenomenon poses when extrapolating from presence of the fungus and toxin in pasture to possible ingestion by the horse, archived samples obtained from the gastrointestinal tract of EGS cases were also analysed for the presence of mycotoxin. This analysis revealed the presence of certain mycotoxins in a small proportion of cases. However, the likelihood of identifying the continued presence of mycotoxins within the gut lumen of any animal is not only dependent on consumption of the toxin, but also the rapidity by which the mycotoxin is metabolised, a feature which varies extensively between individual toxins.
Although it is not possible to draw definitive conclusions from the study at this stage, the “pilot” nature of the study has largely served its purpose; namely, by providing both sufficient evidential support for continuation of this work and vital information which will inform the design of future studies (for example appropriate sample collection, processing and storage protocols). The researchers are grateful to the EGSF for the opportunity to conduct this work and revisit this avenue of EGS research and hope to generate further data in the near future.