Comparison of Fumonisin B1 Biosynthesis in Maize Germ and Degermed Kernels by Fusarium verticillioides

Fusarium verticillioides produces a group of mycotoxins known as fumonisins in maize kernels. Fumonisins are associated with a variety of mycotoxicoses in humans and animals; thus, their presence in food is a considerable safety issue. This study addressed fumonisin B1 (FB1) production in two components of the maize kernel, namely the germ tissues and the degermed kernel. Growth of F. verticillioides was similar in colonized germ tissue and degermed kernels, but FB1 production was at least five times higher in degermed maize kernels than in germ tissue. Expression of the fumonisin polyketide synthase gene, FUM1, as measured by β-glucuronidase (GUS) and Northern blot analysis, followed the same pattern as FB1 production. Also correlated to FB1 was a concomitant drop in pH of the colonized degermed kernels. A time course experiment showed that degermed kernels inoculated with F. verticillioides became acidified over time (from pH 6.4 to 4.7 after 10 days of incubation), whereas colonized germ tissue became alkaline over the same period (from pH 6.5 to 8.5). Because conditions of acidic pH are conducive to FB1 production and alkaline pH is repressive, the observed correlation between the acidification of degermed kernels and the increase in FB1 provides one explanation for the observed differences in FB1 levels.

Download Full-text

Regulation of Fumonisin B1 Biosynthesis and Conidiation in Fusarium verticillioides by a Cyclin-Like (C-Type) Gene, FCC1

Applied and Environmental Microbiology ◽

10.1128/aem.67.4.1607-1612.2001 ◽

2001 ◽

Vol 67 (4) ◽

pp. 1607-1612 ◽

Cited By ~ 91

Author(s):

Won-Bo Shim ◽

Charles P. Woloshuk

Keyword(s):

Minimal Medium ◽

Polyketide Synthase ◽

Pathogenic Fungus ◽

Fusarium Verticillioides ◽

Fumonisin B1 ◽

Reading Frame ◽

Cyclin C ◽

Polyketide Synthase Gene ◽

Type Gene ◽

Corn Kernels

ABSTRACT Fumonisins are a group of mycotoxins produced in corn kernels by the plant-pathogenic fungus Fusarium verticillioides. A mutant of the fungus, FT536, carrying a disrupted gene namedFCC1 (for Fusarium cyclin C1) resulting in altered fumonisin B1 biosynthesis was generated. FCC1 contains an open reading frame of 1,018 bp, with one intron, and encodes a putative 319-amino-acid polypeptide. This protein is similar to UME3 (also called SRB11 or SSN8), a cyclin C of Saccharomyces cerevisiae, and contains three conserved motifs: a cyclin box, a PEST-rich region, and a destruction box. Also similar to the case for C-type cyclins, FCC1was constitutively expressed during growth. When strain FT536 was grown on corn kernels or on defined minimal medium at pH 6, conidiation was reduced and FUM5, the polyketide synthase gene involved in fumonisin B1 biosynthesis, was not expressed. However, when the mutant was grown on a defined minimal medium at pH 3, conidiation was restored, and the blocks in expression ofFUM5 and fumonisin B1 production were suppressed. Our data suggest that FCC1 plays an important role in signal transduction regulating secondary metabolism (fumonisin biosynthesis) and fungal development (conidiation) inF. verticillioides.

Download Full-text

The Fusarium verticillioides FUM Gene Cluster Encodes a Zn(II)2Cys6 Protein That Affects FUM Gene Expression and Fumonisin Production

Eukaryotic Cell ◽

10.1128/ec.00400-06 ◽

2007 ◽

Vol 6 (7) ◽

pp. 1210-1218 ◽

Cited By ~ 137

Author(s):

Daren W. Brown ◽

Robert A. E. Butchko ◽

Mark Busman ◽

Robert H. Proctor

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Fusarium Verticillioides ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Regulatory Proteins ◽

Wild Type ◽

Polyketide Synthase Gene ◽

Splice Forms ◽

Self Protection

ABSTRACT Fumonisins are mycotoxins produced by some Fusarium species and can contaminate maize or maize products. Ingestion of fumonisins is associated with diseases, including cancer and neural tube defects, in humans and animals. In fungi, genes involved in the synthesis of mycotoxins and other secondary metabolites are often located adjacent to each other in gene clusters. Such genes can encode structural enzymes, regulatory proteins, and/or proteins that provide self-protection. The fumonisin biosynthetic gene cluster includes 16 genes, none of which appear to play a role in regulation. In this study, we identified a previously undescribed gene (FUM21) located adjacent to the fumonisin polyketide synthase gene, FUM1. The presence of a Zn(II)2Cys6 DNA-binding domain in the predicted protein suggested that FUM21 was involved in transcriptional regulation. FUM21 deletion (Δfum21) mutants produce little to no fumonisin in cracked maize cultures but some FUM1 and FUM8 transcripts in a liquid GYAM medium. Complementation of a Δfum21 mutant with a wild-type copy of the gene restored fumonisin production. Analysis of FUM21 cDNAs identified four alternative splice forms (ASFs), and microarray analysis indicated the ASFs were differentially expressed. Based on these data, we present a model for how FUM21 ASFs may regulate fumonisin biosynthesis.

Download Full-text

Transformation-Mediated Complementation of a FUM Gene Cluster Deletion in Fusarium verticillioides Restores both Fumonisin Production and Pathogenicity on Maize Seedlings

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-21-1-0087 ◽

2008 ◽

Vol 21 (1) ◽

pp. 87-97 ◽

Cited By ~ 124

Author(s):

Anthony E. Glenn ◽

Nicholas C. Zitomer ◽

Anne Marie Zimeri ◽

Lonnie D. Williams ◽

Ronald T. Riley ◽

...

Keyword(s):

Polyketide Synthase ◽

Fusarium Verticillioides ◽

Molecular Genetic ◽

Polymerase Chain Reaction Analysis ◽

Genomic Region ◽

Nucleotide Sequence Analysis ◽

Maize Seedlings ◽

Developmental Abnormalities ◽

Filamentous Ascomycete ◽

Polyketide Synthase Gene

The filamentous ascomycete Fusarium verticillioides is a pathogen of maize and produces the fumonisin mycotoxins. However, a distinct population of F. verticillioides is pathogenic on banana and does not produce fumonisins. Fumonisin-producing strains from maize cause leaf lesions, developmental abnormalities, stunting, and sometimes death of maize seedlings, whereas fumonisin-nonproducing banana strains do not. A Southern analysis of banana strains did not detect genes in the fumonisin biosynthetic gene (FUM) cluster but did detect genes flanking the cluster. Nucleotide sequence analysis of the genomic region carrying the flanking genes revealed that the FUM cluster was absent in banana strains except for portions of FUM21 and FUM19, which are the terminal genes at each end of the cluster. Polymerase chain reaction analysis confirmed the absence of the cluster in all banana strains examined. Cotransformation of a banana strain with two overlapping cosmids, which together contain the entire FUM cluster, yielded fumonisin-producing transformants that were pathogenic on maize seedlings. Conversely, maize strains that possess the FUM cluster but do not produce fumonisins because of mutations in FUM1, a polyketide synthase gene, were not pathogenic on maize seedlings. Together, the data indicate that fumonisin production may have been lost by deletion of the FUM cluster in the banana population of F. verticillioides but that fumonisin production could be restored by molecular genetic complementation. The results also indicate that fumonisin production by F. verticillioides is required for development of foliar disease symptoms on maize seedlings.

Download Full-text

Amylopectin Induces Fumonisin B1 Production by Fusarium verticillioides During Colonization of Maize Kernels

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-18-1333 ◽

2005 ◽

Vol 18 (12) ◽

pp. 1333-1339 ◽

Cited By ~ 101

Author(s):

B. H. Bluhm ◽

C. P. Woloshuk

Keyword(s):

Quantitative Polymerase Chain Reaction ◽

Fusarium Verticillioides ◽

Animal Health ◽

Fumonisin B1 ◽

Polymerase Chain Reaction Analysis ◽

Defined Medium ◽

Wild Type ◽

Fungal Genes ◽

High Amylose ◽

Maize Kernels

Fusarium verticillioides, a fungal pathogen of maize, produces fumonisin mycotoxins that adversely affect human and animal health. Basic questions remain unanswered regarding the interactions between the host plant and the fungus that lead to the accumulation of fumonisins in maize kernels. In this study, we evaluated the role of kernel endosperm composition in regulating fumonisin B1 (FB1) biosynthesis. We found that kernels lacking starch due to physiological immaturity did not accumulate FB1. Quantitative polymerase chain reaction analysis indicated that kernel development also affected the expression of fungal genes involved in FB1 biosynthesis, starch metabolism, and nitrogen regulation. A mutant strain of F. verticillioides with a disrupted α-amylase gene was impaired in its ability to produce FB1 on starchy kernels, and both the wild-type and mutant strains produced significantly less FB1 on a high-amylose kernel mutant of maize. When grown on a defined medium with amylose as the sole carbon source, the wild-type strain produced only trace amounts of FB1, but it produced large amounts of FB1 when grown on amylopectin or dextrin, a product of amylopectin hydrolysis. We conclude that amylopectin induces FB1 production in F. verticillioides. This study provides new insight regarding the interaction between the fungus and maize kernel during pathogenesis and highlights important areas that need further study.

Download Full-text

Bacillomycin D-C16 inhibits growth of Fusarium verticillioides and production of fumonisin B1 in maize kernels

Pesticide Biochemistry and Physiology ◽

10.1016/j.pestbp.2021.105015 ◽

2021 ◽

pp. 105015

Author(s):

Fuxing Lin ◽

Xiaoyu Zhu ◽

Jing Sun ◽

Fanqiang Meng ◽

Zhaoxin Lu ◽

...

Keyword(s):

Fusarium Verticillioides ◽

Fumonisin B1 ◽

Maize Kernels ◽

Bacillomycin D

Download Full-text

A polyketide synthase gene cluster required for pathogenicity of Pseudocercospora fijiensis on banana

PLoS ONE ◽

10.1371/journal.pone.0258981 ◽

2021 ◽

Vol 16 (10) ◽

pp. e0258981

Author(s):

Elizabeth Thomas ◽

Roslyn D. Noar ◽

Margaret E. Daub

Keyword(s):

Time Course ◽

Polyketide Synthase ◽

Critical Role ◽

Pathogenic Fungi ◽

Gene Clusters ◽

Post Inoculation ◽

Disease Development ◽

Black Sigatoka ◽

Polyketide Synthase Gene ◽

Pseudocercospora Fijiensis

Pseudocercospora fijiensis is the causal agent of the highly destructive black Sigatoka disease of banana. Previous research has focused on polyketide synthase gene clusters in the fungus, given the importance of polyketide pathways in related plant pathogenic fungi. A time course study of expression of the previously identified PKS7-1, PKS8-2, and PKS10-2 gene clusters showed high expression of all three PKS genes and the associated clustered genes in infected banana plants from 2 weeks post-inoculation through 9 weeks. Engineered transformants silenced for PKS8-2 and PKS10-2 were developed and tested for pathogenicity. Inoculation of banana plants with silencing transformants for PKS10-2 showed significant reduction in disease symptoms and severity that correlated with the degree of silencing in the conidia used for inoculation, supporting a critical role for PKS10-2 in disease development. Unlike PKS10-2, a clear role for PKS8-2 could not be determined. Two of four PKS8-2 silencing transformants showed reduced disease development, but disease did not correlate with the degree of PKS8-2 silencing in the transformants. Overall, the degree of silencing obtained for the PKS8-2 transformants was less than that obtained for the PKS10-2 transformants, which may have limited the utility of the silencing strategy to identify a role for PKS8-2 in disease. Orthologous PKS10-2 clusters had previously been identified in the related banana pathogens Pseudocercospora musae and Pseudocercospora eumusae. Genome analysis identified orthologous gene clusters to that of PKS10-2 in the newly sequenced genomes of Pseudocercospora fuligena and Pseudocercospora cruenta, pathogens of tomato and cowpea, respectively. Our results support an important role for the PKS10-2 polyketide pathway in pathogenicity of Pseudocercospora fijiensis, and suggest a possible role for this pathway in disease development by other Pseudocercospora species.

Download Full-text

Fusarium verticillioides and Aspergillus flavus Co-Occurrence Influences Plant and Fungal Transcriptional Profiles in Maize Kernels and In Vitro

Toxins ◽

10.3390/toxins13100680 ◽

2021 ◽

Vol 13 (10) ◽

pp. 680

Author(s):

Alessandra Lanubile ◽

Paola Giorni ◽

Terenzio Bertuzzi ◽

Adriano Marocco ◽

Paola Battilani

Keyword(s):

Aspergillus Flavus ◽

Time Course ◽

Expression Profiles ◽

Fusarium Verticillioides ◽

Post Inoculation ◽

Transcript Accumulation ◽

Pr Genes ◽

Mycotoxin Production ◽

Maize Kernels

Climate change will increase the co-occurrence of Fusarium verticillioides and Aspergillus flavus, along with their mycotoxins, in European maize. In this study, the expression profiles of two pathogenesis-related (PR) genes and four mycotoxin biosynthetic genes, FUM1 and FUM13, fumonisin pathway, and aflR and aflD, aflatoxin pathway, as well as mycotoxin production, were examined in kernels and in artificial medium after a single inoculation with F. verticillioides or A. flavus or with the two fungi in combination. Different temperature regimes (20, 25 and 30 °C) over a time-course of 21 days were also considered. In maize kernels, PR genes showed the strongest induction at 25 °C in the earlier days post inoculation (dpi)with both fungi inoculated singularly. A similar behaviour was maintained with fungi co-occurrence, but with enhanced defence response at 9 dpi under 20 °C. Regarding FUM genes, in the kernels inoculated with F. verticillioides the maximal transcript levels occurred at 6 dpi at 25 °C. At this temperature regime, expression values decreased with the co-occurrence of A. flavus, where the highest gene induction was detected at 20 °C. Similar results were observed in fungi grown in vitro, whilst A. flavus presence determined lower levels of expression along the entire time-course. As concerns afl genes, considering both A. flavus alone and in combination, the most elevated transcript accumulation occurred at 30 °C during all time-course both in infected kernels and in fungi grown in vitro. Regarding mycotoxin production, no significant differences were found among temperatures for kernel contamination, whereas in vitro the highest production was registered at 25 °C for aflatoxin B1 and at 20 °C for fumonisins in the case of single inoculation. In fungal co-occurrence, both mycotoxins resulted reduced at all the temperatures considered compared to the amount produced with single inoculation.

Download Full-text

HXK1 regulates carbon catabolism, sporulation, fumonisin B1 production and pathogenesis in Fusarium verticillioides

Microbiology ◽

10.1099/mic.0.052506-0 ◽

2011 ◽

Vol 157 (9) ◽

pp. 2658-2669 ◽

Cited By ~ 23

Author(s):

Hun Kim ◽

Jonathon E. Smith ◽

John B. Ridenour ◽

Charles P. Woloshuk ◽

Burton H. Bluhm

Keyword(s):

Carbon Source ◽

Regulatory Networks ◽

Sole Carbon Source ◽

Metabolic Response ◽

Fusarium Verticillioides ◽

Fumonisin B1 ◽

Genetic Regulatory Networks ◽

Wild Type ◽

Increased Sensitivity ◽

Maize Kernels

In Fusarium verticillioides, a ubiquitous pathogen of maize, virulence and mycotoxigenesis are regulated in response to the types and amounts of carbohydrates present in maize kernels. In this study, we investigated the role of a putative hexokinase-encoding gene (HXK1) in growth, development and pathogenesis. A deletion mutant (Δhxk1) of HXK1 was not able to grow when supplied with fructose as the sole carbon source, and growth was impaired when glucose, sucrose or maltotriose was provided. Additionally, the Δhxk1 mutant produced unusual swollen hyphae when provided with fructose, but not glucose, as the sole carbon source. Moreover, the Δhxk1 mutant was impaired in fructose uptake, although glucose uptake was unaffected. On maize kernels, the Δhxk1 mutant was substantially less virulent than the wild-type, but virulence on maize stalks was not impaired, possibly indicating a metabolic response to tissue-specific differences in plant carbohydrate content. Finally, disruption of HXK1 had a pronounced effect on fungal metabolites produced during colonization of maize kernels; the Δhxk1 mutant produced approximately 50 % less trehalose and 80 % less fumonisin B1 (FB1) than the wild-type. The reduction in trehalose biosynthesis likely explains observations of increased sensitivity to osmotic stress in the Δhxk1 mutant. In summary, this study links early events in carbohydrate sensing and glycolysis to virulence and secondary metabolism in F. verticillioides, and thus provides a new foothold from which the genetic regulatory networks that underlie pathogenesis and mycotoxigenesis can be unravelled and defined.

Download Full-text

The Dent Stage of Maize Kernels Is the Most Conducive for Fumonisin Biosynthesis under Field Conditions

Applied and Environmental Microbiology ◽

10.1128/aem.05216-11 ◽

2011 ◽

Vol 77 (23) ◽

pp. 8382-8390 ◽

Cited By ~ 24

Author(s):

Adeline Picot ◽

Christian Barreau ◽

Laëtitia Pinson-Gadais ◽

François Piraux ◽

Daniel Caron ◽

...

Keyword(s):

Time Course ◽

Fusarium Verticillioides ◽

Animal Health ◽

Maximum Level ◽

Field Conditions ◽

In Planta ◽

Content Type ◽

Short Period ◽

Maize Kernels

ABSTRACTThe fungal pathogenFusarium verticillioidesinfects maize ears and produces fumonisins, known for their adverse effects on human and animal health. Basic questions remain unanswered regarding the kernel stage(s) associated with fumonisin biosynthesis and the kernel components involved in fumonisin regulation duringF. verticillioides-maize interaction under field conditions. In this 2-year field study, the time course ofF. verticillioidesgrowth and fumonisin accumulation in developing maize kernels, along with the variations in kernel pH and amylopectin content, were monitored using relevant and accurate analytical tools. In all experiments, the most significant increase in fumonisin accumulation or in fumonisin productivity (i.e., fumonisin production per unit of fungus) was shown to occur within a very short period of time, between 22/32 and 42 days after inoculation and corresponding to the dent stage. This stage was also characterized by acidification in the kernel pH and a maximum level of amylopectin content. Our data clearly support published results based onin vitroexperiments suggesting that the physiological stages of the maize kernel play a major role in regulating fumonisin production. Here we have validated this result forin plantaand field conditions, and we demonstrate that under such conditions the dent stage is the most conducive for fumonisin accumulation.

Download Full-text

Deteriorative changes in maize kernels due to Aspergillus flavus Link. and Fusarium verticillioides (Sacc.) Nirenberg

Acta agriculturae Slovenica ◽

10.14720/aas.2019.114.1.8 ◽

2019 ◽

Vol 114 (1) ◽

pp. 69

Author(s):

Francis Collins MUGA ◽

Tilahun Seyoum WORKNEH ◽

Moses Okoth MARENYA

Keyword(s):

Chemical Composition ◽

Aspergillus Flavus ◽

Crude Protein ◽

Fusarium Verticillioides ◽

Fumonisin B1 ◽

Human Consumption ◽

Crude Fibre ◽

Contamination Levels ◽

Maize Kernels ◽

Crude Fibre Content

The study aimed at measuring changes in chemical composition of maize kernels due to Aspergillus flavus Link. and Fusarium verticillioides (Sacc.) Nirenberg infection. The samples of maize kernels were incubated at 28 °C for 7, 14, 21, and 28 days. The samples were analysed for mycotoxin, moisture, crude fat, crude protein, crude ash, and crude fibre. Maize kernels inoculated with A. flavus and F. verticillioides exhibited a significant decrease in crude fat. Aflatoxin B1 (AFB1) contamination increased in maize kernels inoculated with A. flavus, and fumonisin B1 (FB1) in kernels inoculated with F. verticillioides. Crude ash and crude fibre content showed no changes. Incubation time significantly affected AFB1 and FB1 contamination levels, moisture, crude fat, and crude protein contents. AFB1 and FB1 contamination were significantly correlated with crude fat degradation. The tested strains had similar deteriorative effects on maize kernels. The significant changes in the proximate composition were only observed in maize kernels with mycotoxin contamination above the regulatory limit of 10 µg kg−1, thus not fit for human consumption.

Download Full-text