Storage fungi and mycotoxins associated with cowpea

Abstract

Cowpeas (Vigna unguiculata (L.) Walp) is an important African indigenous legume crop for the livelihoods of many relatively poor people residing n less developed countries of the tropics. Rural families derive a nutritious food, animal feed and in income from the production of this crop. Storage of seed is certainly the most important post-harvest operation but the losses incurred are great. These losses, due to an inability to effectively control physical and biological factors, result in problems with storage insects, moisture and associated fungi. Seeds are particularly susceptible to fungal contamination when stored at high ambient temperatures and relative humidities. To determine the storage fungi associated with cowpea seeds, surface-sterilised cowpea seeds (200 seeds from each of nine cultivars) were plated out n malt extract agar. After 5-7 days incubation at 25°C, the most dominant and common fungi recorded were Alternaria spp. followed by Penicillium spp., Aspergillus flavus and A. niger. The influence of a three-year cold storage period at ± 5°C on the fungi associated with the seeds was also investigated. Alternaria, Aspergillus and Penicillium spp. appeared to dominate. Some fungal species recorded prior to cold storage were not recorded thereafter. Certain storage fungi are known to produce mycotoxins, which are secondary fungal metabolites that are toxic to both farm animals and humans, under poor storage conditions. The presence of the fusarial mycotoxins, fumonisin BI, B2 and B3 in four cowpea cultivars (Bechwana Whit, Glenda, Iron Grey, Rhino) was investigated. The samples were extracted with methanol/water (70:30 v/v) and cleaned-up on strong anion exchange solid phase extraction cartridges. High performance liquid chromatography with pre¬column derivatisation using o-phthaldialdehyde (OPA) was used for the detection and quantification of fumonisin Bl, B2 and B3. All sampIes were contaminated with FBI, with levels ranging from 81-1002 ng g-I. Fumonisin B2 and B3 were not detected in any samples. This is believed to be the first report of fumonisin BI in cowpea seeds. Since the known fumonisin-producing Fusarium species were not found in the six different Fusarium species isolated from these four cultivars, further investigations are required to determine which fungal species are species are responsible species are responsible for the FBI production. An alternative approach to the prevention and control of fungal contamination and mycotoxin production of seeds by treating cowpea seed with essential plant oils was tested. The inhibitory activity of five essential oils (thyme, clove, peppermint, soybean and peanut) was investigated, in vitro and in vivo, on five fungal species (A. flavus, A. niger, Penicillium chrysogenum, Fusarium oxysporum and F. equiseti) commonly associated with cowpea seeds and on two cowpea cultivars. Thyme and clove oil significantly inhibited the growth of all five fungal species in vitro at 500 and 1000 ppm, while peppermint oil was successful at 2000 ppm. Peanut and soybean oil did not show any significant inhibition of fungal growth. The in vivo effect of thyme, clove and peppermint oils on naturally infected seed revealed that only thyme at 1000 ppm reduced fungal growth of storage fungi in the PAN 325 cultivar. In the PAN 311 cultivar, thyme and clove oils at 1000 ppm and peppermint oil at 2000 ppm significantIy reduced growth of storage fungi. In artificially infected seed, all three oils significantly inhibited the growth of P. chrysogenum. Thyme reduced the growth of F. oxysporum and F. equisetii, whilst peppermint oil inhibited only F. oxyspomm. These oils did not seem to adversely affect the germination nor emergence of cowpea seed. The storage fungi significantly reduced percentag germination and emergence of the white (IT 93K452-1) seed but had little or no effect on the brown (CH 14) seed. Furthermore, all three oils significantly inhibited the storage fungi on the white seed, possibly increasing the percentage germination and emergence.