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  • Among the samples maize showed maximum incidence

    2018-11-12

    Among the samples, maize showed maximum incidence of F. verticillioides with 8.5% incidence in Mysore, 3.5% in Madikeri and 3.6% in Bellary samples. Further 2% incidence was in paddy from Hassan and 1.4% incidence was in sorghum from Mysore samples (Table 3). Pearl millet was free from infection. Frequency of F. verticillioides among screened samples was high (47.54%) with maize followed by paddy (42.85%) and least in sorghum (16.66%). Among the screened samples maize (40.98%), paddy (33.33%) and sorghum (12.5%) samples were found contaminated with fumonisin producing F. verticillioides (Table 2). Of the 326 Fusarium isolates obtained from 69 infected cereal samples, 194 were F. verticillioides which were isolated from 51 cereal samples. The relative density of the isolated F. verticillioides species was 59.50% whereas other Fusarium species screened had relative density of 40.5% (Fig. 1). When all the 326 Fusarium species isolates were subjected to PCR amplification with VERT primers, 194 isolates scored positive with species specific VERTF-1 and VERTR (1016bp) primers, and 176 isolates scored positive for VERTF-1 and VERTF-2 (400bp) primers (Fig. 2). Hence, among the 194 isolates, 176 isolates were fumonisin producing F. verticillioides and 18 isolates were non-fumonisin producing. F. verticillioides (MTCC 156) strain was used as positive control and F. graminerium (MTCC 1893) was used as negative control (Fig. 3). For reconfirmation of PCR results the randomly selected PCR amplified products of VERTF-1/VERTR and VERTF-1/VERTF-2 were subjected to sequencing. Sequencing reads of PCR analyzed products were tested using nucleotide megablast. The sequences were 99% similar to Giberrella moniliformis strains. The same sequences were deposited at NCBI and obtained the accession numbers KJ410046, KJ410047, KR061317, KR061318 with VERTF-1/VERTR primers and KJ410767, KJ410768 with VERTF-1/VERTF-2. The sequencing result provides as reconfirmation for PCR carried out for the F. verticillioides strains isolated from the cereals. The sequenced isolates are represented by hippo signaling pathway tree analysis (Fig. 4). The fumonisin producing ability was reconfirmed through LCMS results and three randomly selected representative isolates from maize, paddy and sorghum were compared with toxin standard of fumonisin B1 with retention time of 1.76min and molecular weight of 722.535g/mol. Six isolates (two each from maize, paddy, sorghum) recorded as fumonisin producing and three isolates were non fumonisin producers (one each from maize, paddy and sorghum) when compared to standard graph (Fig. 5).
    Discussion During the last decade, fumonisin producing F. verticillioides and related species have received worldwide attention. The non-scientific methods of agricultural practices, poor storage facilities and unfavorable environmental conditions from the time of harvest to storage/marketing have led to colonization of fumonisin producing fungi [30]. Moulds, besides depleting the nutrients, also produce toxic substances that have potential health hazards to animals and in turn to humans [31,32]. In this study 29 maize samples exhibited the association of F. verticillioides out of 37 samples of maize screened for Fusarium species and this was followed by paddy and sorghum but pearl millet was devoid of F. verticillioides infection (Table 2). F. verticillioides was considered as predominant species on maize with significant levels of fumonisin [33] and G. fujikuroi was the dominant species that would probably be the main source for fumonisin production in cereals [34,35]. In the present study 194 F. verticillioides were isolated (maize-108, paddy-80, sorghum-6) from 326 Fusarium species (maize-158, paddy-137, sorghum-29, pearl millet-2) screened from 135 cereal samples (Fig. 2). Even after certain measures taken for the control of fumonisin production, the association of fumonisin producing F. verticilloides is increasing day by day. One hundred and seventy six fumonisin producing F. verticillioides (maize-103, paddy-68, sorghum-5) were isolated from screened 326 Fusarium species in the present work (Fig. 2). Among the 45 maize samples collected from south Karnataka, 25 samples were found to be highly infected with FB1 producing F. verticillioides[36]. Among the 22 fumonisin producing isolates screened, 18 were F. verticillioides in which 17 (94.4%) isolates produced fumonisins (FB1+FB2) at concentration range of 0.07–121.45μg/g [37]. An epidemiological survey conducted in Karnataka and Andhra Pradesh during 1997 revealed that consumption of mouldy grains affected 1424 persons in 27 villages [38]. Greater attention is bestowed to investigate Fusarium species worldwide as they reduce the value of cereals used as food and feed. Cereals contaminated with toxigenic species cause acute and chronic poisoning and allergic symptoms to animals and human. Fungi causing deterioration of cereals especially in maize, paddy and sorghum are a major problem because they produce mycotoxins [40].