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Prevalence, Characterization, and Mycotoxin Production Ability of Fusarium Species on Korean Adlay (Coix Lacrymal-Jobi L.) Seeds

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Prevalence, Characterization, and Mycotoxin Production Ability of Fusarium Species on Korean Adlay (Coix Lacrymal-Jobi L.) Seeds

Tae Jin An et al. Toxins (Basel).

Abstract

Adlay seed samples were collected from three adlay growing regions (Yeoncheon, Hwasun, and Eumseong region) in Korea during 2012. Among all the samples collected, 400 seeds were tested for fungal occurrence by standard blotter and test tube agar methods and different taxonomic groups of fungal genera were detected. The most predominant fungal genera encountered were Fusarium, Phoma, Alternaria, Cladosporium, Curvularia, Cochliobolus and Leptosphaerulina. Fusarium species accounted for 45.6% of all species found; and, with phylogenetic analysis based on the combined sequences of two protein coding genes (EF-1α and β-tubulin), 10 Fusarium species were characterized namely, F. incarnatum (11.67%), F. kyushuense (10.33%), F. fujikuroi (8.67%), F. concentricum (6.00%), F. asiaticum (5.67%), F. graminearum (1.67%), F. miscanthi (0.67%), F. polyphialidicum (0.33%), F. armeniacum (0.33%), and F. thapsinum (0.33%). The Fusarium species were then examined for their morphological characteristics to confirm their identity. Morphological observations of the species correlated well with and confirmed their molecular identification. The ability of these isolates to produce the mycotoxins fumonisin (FUM) and zearalenone (ZEN) was tested by the ELISA quantitative analysis method. The result revealed that FUM was produced only by F. fujikuroi and that ZEN was produced by F. asiaticum and F. graminearum.

Keywords: ELISA; Fusarium; adlay seeds; morphological data analysis; mycotoxins; phylogenetic analysis.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Percentage incidence of seed-borne fungi on adlay (400 seeds) based on morphology and ITS gene sequence analysis.
Figure 2
Figure 2
Phylogenetic relationship of Fusarium spp. Maximum likelihood tree of the Fusarium and related genera inferred from the combined sequences of the β-tubulin and elongation factor genes. Taking into account the different tempos and modes of nucleotide substitutions, all parameters of the substitution model were separately estimated for each gene using the GTR + I + Γ model. The branch lengths are proportional to the estimated number of nucleotide substitutions. The bootstrap probability (BP; 1000 replicates) values over 75% are displayed on the nodes. Fungi isolated from adlay seed are indicated in bold.
Figure 3
Figure 3
Ten species of Fusarium isolated from adlay. Scale bar = 10 μm: (A) Fusarium fujikuroi; (B) Fusarium asiaticum; (C) Fusarium concentricum; (D) Fusarium graminearum; (E) Fusarium thapsinum; (F) Fusarium armeniacum; (G) Fusarium miscanthi; (H) Fusarium kyushuense; (I) Fusarium polyphialidicum; and (J) Fusarium incarnatum.
Figure 4
Figure 4
Fumonisin producing ability of ten Fusarium species isolated from adlay seeds. LOQ, limit of quantitation; FUM, Fumonisin.
Figure 5
Figure 5
Zearalenone producing ability of ten Fusarium species isolated from adlay seeds. LOQ, limit of quantitation; ZEN, Zearalenone.
Figure 6
Figure 6
Incubation of adlay seeds: (A) The Blotter method, Adlay seed incubated for 7 days at 20 °C under 12/12 h NUV/dark cycle; and (B) Test tube agar method, Adlay seed incubated for 3 weeks at 20 °C under 12/12 h daylight/dark cycle.

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