Diversity of Fusarium species and mycotoxins contaminating pineapple.
ABSTRACT: Pineapple (Ananas comosus var. comosus) is an important perennial crop in tropical and subtropical areas. It may be infected by various Fusarium species, contaminating the plant material with mycotoxins. The aim of this study was to evaluate Fusarium species variability among the genotypes isolated from pineapple fruits displaying fungal infection symptoms and to evaluate their mycotoxigenic abilities. Forty-four isolates of ten Fusarium species were obtained from pineapple fruit samples: F. ananatum, F. concentricum, F. fujikuroi, F. guttiforme, F. incarnatum, F. oxysporum, F. polyphialidicum, F. proliferatum, F. temperatum and F. verticillioides. Fumonisins B1-B3, beauvericin (BEA) and moniliformin (MON) contents were quantified by high-performance liquid chromatography (HPLC) in pineapple fruit tissue. Fumonisins are likely the most dangerous metabolites present in fruit samples (the maximum FB1 content was 250 μg g(-1) in pineapple skin and 20 μg ml(-1) in juice fraction). In both fractions, BEA and MON were of minor significance. FUM1 and FUM8 genes were identified in F. fujikuroi, F. proliferatum, F. temperatum and F. verticillioides. Cyclic peptide synthase gene (esyn1 homologue) from the BEA biosynthetic pathway was identified in 40 isolates of eight species. Based on the gene-specific polymerase chain reaction (PCR) assays, none of the isolates tested were found to be able to produce trichothecenes or zearalenone.
Project description:Mycotoxins pose a challenge to a safe food supply worldwide, and their threat is expected to worsen with our changing climate. The need for diligence is exemplified by the discovery of fumonisin B2 in wine, which joins ochratoxin A as a mycotoxin of concern in the grape-wine chain. To elucidate the mycotoxin risk in southeastern American wine, grape samples were collected from vineyards during harvest in 2013 and potentially mycotoxigenic fungi (Fusarium and Aspergillus) were isolated from the samples. Numerous Fusarium isolates were recovered and identified to the species level by comparison of translation elongation factor 1-? gene sequences to verified strains. Fusarium fujikuroi was the most abundant species recovered (239 isolates), followed by F. proliferatum (52), F. incarnatum-equiseti (14), F. oxysporum (7), F. concentricum (1), and F. solani (1). In vitro assays quantified fumonisin production for representative isolates via liquid chromatography-tandem mass spectrometry. Surprisingly, nearly all F. fujikuroi isolates produced fumonisins B1, B2, and B3 at levels comparable to both the F. proliferatum isolates and the positive control, Fusarium verticillioides. Such capacity for fumonisin production refutes the generally accepted notion that F. fujikuroi produces undetectable or low levels of fumonisins and provides evidence to reconsider this species as a mycotoxigenic threat to economically significant crops.
Project description:In 2017-2018, extensive symptoms of sudden decline and fruit rot were observed on date palms in southern Tunisia. Samples of diseased plants were randomly collected in six localities. Based on morphological identification, <i>Fusarium</i> was the most frequent fungal genus detected. A sequencing of translation elongation factor, calmodulin, and second largest subunit of RNA polymerase II genes was used to identify 63 representative <i>Fusarium</i> strains at species level and investigate their phylogenetic relationships. The main species detected was <i>Fusarium</i> <i>proliferatum</i>, and at a much lesser extent, <i>Fusarium</i> <i>brachygibbosum</i>, <i>Fusarium</i> <i>caatingaense</i>, <i>Fusarium</i> <i>clavum</i>, <i>Fusarium</i> <i>incarnatum</i><i>,</i> and <i>Fusarium</i> <i>solani</i>. Pathogenicity on the <i>Deglet</i> <i>Nour</i> variety plantlets and the capability to produce mycotoxins were also assessed. All <i>Fusarium</i> species were pathogenic complying Koch's postulates. <i>Fusarium</i> <i>proliferatum</i> strains produced mainly fumonisins (FBs), beauvericin (BEA), and, to a lesser extent, enniatins (ENNs) and moniliformin (MON). All <i>F.</i> <i>brachygibbosum</i> strains produced low levels of BEA, diacetoxyscirpenol, and neosolaniol; two strains produced also T-2 toxin, and a single strain produced HT-2 toxin. <i>Fusarium</i> <i>caatingaense</i>, <i>F.</i> <i>clavum</i>, <i>F.</i> <i>incarnatum</i> produced only BEA. <i>Fusarium</i> <i>solani</i> strains produced MON, BEA, and ENNs. This work reports for the first time a comprehensive multidisciplinary study of <i>Fusarium</i> species on date palms, concerning both phytopathological and food safety issues.
Project description:The identity of the fungi responsible for fruitlet core rot (FCR) disease in pineapple has been the subject of investigation for some time. This study describes the diversity and toxigenic potential of fungal species causing FCR in La Reunion, an island in the Indian Ocean. One-hundred-and-fifty fungal isolates were obtained from infected and healthy fruitlets on Reunion Island and exclusively correspond to two genera of fungi: Fusarium and Talaromyces. The genus Fusarium made up 79% of the isolates, including 108 <i>F. ananatum</i>, 10 <i>F. oxysporum</i>, and one <i>F. proliferatum</i>. The genus Talaromyces accounted for 21% of the isolated fungi, which were all <i>Talaromyces stollii</i>. As the isolated fungal strains are potentially mycotoxigenic, identification and quantification of mycotoxins were carried out on naturally or artificially infected diseased fruits and under in vitro cultures of potential toxigenic isolates. Fumonisins B<sub>1</sub> and B<sub>2</sub> (FB<sub>1</sub>-FB<sub>2</sub>) and beauvericin (BEA) were found in infected fruitlets of pineapple and in the culture media of Fusarium species. Regarding the induction of mycotoxin in vitro, <i>F.</i> <i>proliferatum</i> produced 182 mg kg⁻<sup>1</sup> of FB<sub>1</sub> and <i>F. oxysporum</i> produced 192 mg kg⁻<sup>1</sup> of BEA. These results provide a better understanding of the causal agents of FCR and their potential risk to pineapple consumers.
Project description:Fusarium subglutinans and Fusarium temperatum are common maize pathogens that produce mycotoxins and cause plant disease. The ability of these species to produce beauvericin and fumonisin mycotoxins is not settled, as reports of toxin production are not concordant. Our objective was to clarify this situation by determining both the chemotypes and genotypes for strains from both species. We analyzed 25 strains from Argentina, 13 F. subglutinans and 12 F. temperatum strains, for toxin production by ultraperformance liquid chromatography mass spectrometry (UPLC-MS). We used new genome sequences from two strains of F. subglutinans and one strain of F. temperatum, plus genomes of other Fusarium species, to determine the presence of functional gene clusters for the synthesis of these toxins. None of the strains examined from either species produced fumonisins. These strains also lack Fum biosynthetic genes but retain homologs of some genes that flank the Fum cluster in Fusarium verticillioides None of the F. subglutinans strains we examined produced beauvericin although 9 of 12 F. temperatum strains did. A complete beauvericin (Bea) gene cluster was present in all three new genome sequences. The Bea1 gene was presumably functional in F. temperatum but was not functional in F. subglutinans due to a large insertion and multiple mutations that resulted in premature stop codons. The accumulation of only a few mutations expected to disrupt Bea1 suggests that the process of its inactivation is relatively recent. Thus, none of the strains of F. subglutinans or F. temperatum we examined produce fumonisins, and the strains of F. subglutinans examined also cannot produce beauvericin. Variation in the ability of strains of F. temperatum to produce beauvericin requires further study and could reflect the recent shared ancestry of these two species.IMPORTANCE Fusarium subglutinans and F. temperatum are sister species and maize pathogens commonly isolated worldwide that can produce several mycotoxins and cause seedling disease, stalk rot, and ear rot. The ability of these species to produce beauvericin and fumonisin mycotoxins is not settled, as reports of toxin production are not concordant at the species level. Our results are consistent with previous reports that strains of F. subglutinans produce neither fumonisins nor beauvericin. The status of toxin production by F. temperatum needs further work. Our strains of F. temperatum did not produce fumonisins, while some strains produced beauvericin and others did not. These results enable more accurate risk assessments of potential mycotoxin contamination if strains of these species are present. The nature of the genetic inactivation of BEA1 is consistent with its relatively recent occurrence and the close phylogenetic relationship of the two sister species.
Project description:The Gibberella fujikuroi complex includes many Fusarium species that cause significant losses in yield and quality of agricultural and forestry crops. Due to their economic importance, whole-genome sequence information has rapidly become available for species including Fusarium circinatum, Fusarium fujikuroi and Fusarium verticillioides, each of which represent one of the three main clades known in this complex. However, no previous studies have explored the genomic commonalities and differences among these fungi. In this study, a previously completed genetic linkage map for an interspecific cross between Fusarium temperatum and F. circinatum, together with genomic sequence data, was utilized to consider the level of synteny between the three Fusarium genomes. Regions that are homologous amongst the Fusarium genomes examined were identified using in silico and pyrosequenced amplified fragment length polymorphism (AFLP) fragment analyses. Homology was determined using BLAST analysis of the sequences, with 777 homologous regions aligned to F. fujikuroi and F. verticillioides. This also made it possible to assign the linkage groups from the interspecific cross to their corresponding chromosomes in F. verticillioides and F. fujikuroi, as well as to assign two previously unmapped supercontigs of F. verticillioides to probable chromosomal locations. We further found evidence of a reciprocal translocation between the distal ends of chromosome 8 and 11, which apparently originated before the divergence of F. circinatum and F. temperatum. Overall, a remarkable level of macrosynteny was observed among the three Fusarium genomes, when comparing AFLP fragments. This study not only demonstrates how in silico AFLPs can aid in the integration of a genetic linkage map to the physical genome, but it also highlights the benefits of using this tool to study genomic synteny and architecture.
Project description:To elucidate the symptoms and pathogens diversity of corn Fusarium sheath rot (CFSR), diseased samples were collected from 21 county-level regions in 12 prefecture-level districts of Sichuan Province from 2015 to 2018 in the present study. In the field, two symptom types appeared including small black spots with a linear distribution and wet blotches with a tawny or brown color. One hundred thirty-seven Fusarium isolates were identified based on morphological characteristics and phylogenetic analysis (EF1-?), and Koch's postulates were also assessed. The results identified the isolates as 8 species in the Fusarium genus, including F. verticillioides, F. proliferatum, F. fujikuroi, F. asiaticum, F. equiseti, F. meridionale, F. graminearum and F. oxysporum, with isolation frequencies of 30.00, 22.67, 15.33, 7.33, 6.00, 5.33, 3.33 and 1.33%, respectively. Fusarium verticillioides and F. proliferatum were the dominant and subdominant species, respectively. Two or more Fusarium species such as F. verticillioides and F. proliferatum were simultaneously identified at a mixed infection rate of 14.67% in the present study. The pathogenicity test results showed that F. proliferatum and F. fujikuroi exhibited the highest virulence, with average disease indices of 30.28?±?2.87 and 28.06?±?1.96, followed by F. equiseti and F. verticillioides, with disease indices of 21.48?±?2.14 and 16.21?±?1.84, respectively. Fusarium asiaticum, F. graminearum and F. meridonale showed lower virulence, with disease indices of 13.80?±?2.07, 11.57?±?2.40 and 13.89?±?2.49, respectively. Finally, F. orysporum presented the lowest virulence in CFSR, with a disease index of 10.14?±?1.20. To the best of our knowledge, this is the first report of F. fujikuroi, F. meridionale and F. asiaticum as CFSR pathogens in China.
Project description:Fusarium subglutinans f. sp. pini (= F. circinatum) is a pathogen of pine and is one of eight mating populations (i.e., biological species) in the Gibberella fujikuroi species complex. This species complex includes F. thapsinum, F. moniliforme (= F. verticillioides), F. nygamai, and F. proliferatum, as well as F. subglutinans associated with sugarcane, maize, mango, and pineapple. Differentiating these forms of F. subglutinans usually requires pathogenicity tests, which are often time-consuming and inconclusive. Our objective was to develop a technique to differentiate isolates of F. subglutinans f. sp. pini from other isolates identified as F. subglutinans. We sequenced the histone H3 gene from a representative set of Fusarium isolates. The H3 gene sequence was conserved and contained two introns in all the isolates studied. From both the intron and the exon sequence data, we developed a PCR-restriction fragment length polymorphism technique that reliably distinguishes F. subglutinans f. sp. pini from the other biological species in the G. fujikuroi species complex.
Project description:In this study, twenty of the most common <i>Fusarium</i> species were molecularly characterized and inoculated on potato dextrose agar (PDA), rice and maize medium, where thirty three targeted mycotoxins, which might be the secondary metabolites of the identified fungal species, were detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Statistical analysis was performed with principal component analysis (PCA) to characterize the mycotoxin profiles for the twenty fungi, suggesting that these fungi species could be discriminated and divided into three groups as follows. Group I, the fusaric acid producers, were defined into two subgroups, namely subgroup I as producers of fusaric acid and fumonisins, comprising of <i>F. proliferatum</i>, <i>F. verticillioides</i>, <i>F. fujikuroi</i> and <i>F. solani</i>, and subgroup II considered to only produce fusaric acid, including <i>F. temperatum</i>, <i>F. subglutinans</i>, <i>F. musae</i>, <i>F. tricinctum</i>, <i>F. oxysporum</i>, <i>F. equiseti</i>, <i>F. sacchari</i>, <i>F. concentricum</i>, <i>F. andiyazi</i>. Group II, as type A trichothecenes producers, included <i>F. langsethiae</i>, <i>F. sporotrichioides</i>, <i>F. polyphialidicum</i>, while Group III were found to mainly produce type B trichothecenes, comprising of <i>F. culmorum</i>, <i>F. poae</i>, <i>F. meridionale</i> and <i>F. graminearum</i>. A comprehensive picture, which presents the mycotoxin-producing patterns by the selected fungal species in various matrices, is obtained for the first time, and thus from an application point of view, provides key information to explore mycotoxigenic potentials of <i>Fusarium</i> species and forecast the <i>Fusarium</i> infestation/mycotoxins contamination.
Project description:Gibberella fujikuroi species complex (GFSC) was isolated from rice (Oryza sativa L.) seed samples from ten Asian countries and investigated for incidence of GFSC, molecular characteristics, and pathogenicity. Regardless of geographic origin, GFSC was detected with incidences ranging from 3% to 80%. Four species, Fusarium fujikuroi, F. concentricum, F. proliferatum, and F. verticillioides, were found to show an association with rice seeds, with F. fujikuroi being the predominant species. In phylogenetic analyses of DNA sequences, no relationship was found between species, isolates, and geographic sources of samples. Unidentified fragments of the ?-tubulin gene were observed in ten isolates of F. fujikuroi and F. verticillioides. With the exception of three isolates of F. fujikuroi, F. fujikuroi, F. proliferatum, and F. verticillioides were found to have FUM1 (the fumonisin biosynthetic gene); however, FUM1 was not found in isolates of F. concentricum. Results of pathogenicity testing showed that all isolates caused reduced germination of rice seed. In addition, F. fujikuroi and F. concentricum caused typical symptoms of bakanae, leaf elongation and chlorosis, whereas F. proliferatum and F. verticillioides only caused stunting of seedlings. These findings provide insight into the characteristics of GFSC associated with rice seeds and might be helpful in development of strategies for management of bakanae.
Project description:Maize is one of the most important crops and Poland is the fifth largest producing country in Europe. Diseases caused by Fusarium spp. can affect the yield and grain quality of maize because of contamination with numerous mycotoxins produced by these fungi. The present study was performed to identify the prevailing Fusarium species and the environmental factors affecting their frequencies and the contamination of grain with the main mycotoxins deoxynivalenol (DON), zearalenone (ZON) and fumonisin B1 (FB1). Thirty kernel samples were collected in three locations in 2011 and in seven locations in 2012 from three hybrids. On average, 25.24% kernels were colonized by Fusarium spp. (424 strains were isolated). Fusarium verticillioides and F. temperatum were the most prevalent species, F. subglutinans, F. proliferatum and F. graminearum were in minor abundance. In total, 272 isolates of F. verticillioides and 81 isolates of F. temperatum were identified. Fusarium temperatum frequency ranged from 1.70% to 28.57% and differences between locations were significant. Fumonisin B1 was found in all tested samples. DON was found in 66.67% and ZON in 43.33% of samples. Rainfall amount positively affected F. temperatum and F. subglutinans frequency in opposite to mean temperatures in July. On the other hand, relationships between frequency of these species and historical data from 1950-2000 for annual temperature range were negative in contrast to the coldest quarter temperatures.