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1.
PLoS One ; 18(9): e0292095, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37756342

RESUMEN

European flint landraces are a major class of maize possessing favorable alleles for improving host resistance to Gibberella ear rot (GER) disease which reduces yield and contaminates the grains with mycotoxins. However, the incorporation of these landraces into breeding programs requires a clear understanding of the effectiveness of their introgression into elite materials. We evaluated 15 pre-selected doubled haploid (DH) lines from two European flint landraces, "Kemater Landmais Gelb" (KE) and "Petkuser Ferdinand Rot" (PE), together with two adapted elite flint lines and seven standard lines for GER severity as the main trait, and several adaptation traits (plant height, days to silking, seed-set, plant vigor) across four environments. From this evaluation, three KE DH lines and one PE DH line, with the lowest GER severity, were selected and used as donor parents that were crossed with the two adapted and GER susceptible flint lines (Flint1 and Flint2) to develop six bi-parental DH populations with 34-145 DH lines each. Each DH population was evaluated across two locations. Correlations between GER severity, which was the target trait, and adaptation traits were weak (-0.02 to 0.19). GER severity of lines from PE landrace was on average 2-fold higher than lines from KE landrace, indicating a clear superiority of the KE landrace lines. Mean GER severity of the DH populations was 39.4-61.0% lower than the adapted elite flint lines. All KE-derived DH populations were on average more resistant (27.0-36.7%) than the PE-derived population (51.0%). Highly resistant lines (1.3-5.2%) were found in all of the populations, suggesting that the DH populations can be successfully integrated into elite breeding programs. The findings demonstrate that selected KE landrace lines used as donors were effective in improving GER resistance of the adapted elite inbreds.


Asunto(s)
Fusarium , Gibberella , Gibberella/genética , Zea mays/genética , Fitomejoramiento , Alelos , Minerales
2.
Phytopathology ; 113(7): 1317-1324, 2023 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-36721376

RESUMEN

Gibberella ear rot (GER) in maize caused by Fusarium graminearum is one of the most devastating maize diseases reducing grain yield and quality worldwide. The genetic bases of maize GER resistance remain largely unknown. Using artificial inoculation across multiple environments, the GER severity of an association panel consisting of 316 diverse inbred lines was observed with wide phenotypic variation. In the association panel, a genome-wide association study using a general linear model identified 69 single-nucleotide polymorphisms (SNPs) significantly associated with GER resistance at the threshold of 2.04 × 10-5, and the average phenotypic variation explained (PVE) of these SNPs was 5.09%. We also conducted a genome-wide association study analysis using a mixed linear model at a threshold of 1.0 × 10-4, and 16 significantly associated SNPs with an average PVE of 4.73% were detected. A combined general linear model and mixed linear model method obtained 10 co-localized significantly associated SNPs linked to GER resistance, including the most significant SNP (PZE-105079915) with the greatest PVE value, 9.07%, at bin 5.05 following 10 candidate genes. These findings are significant for the exploration of the complicated genetic variations in maize GER resistance. The regions and genes identified herein provide a list of candidate targets for further investigation, in addition to the elite germplasm resources that can be used for breeding GER resistance in maize.


Asunto(s)
Fusarium , Gibberella , Gibberella/genética , Estudio de Asociación del Genoma Completo , Enfermedades de las Plantas/genética , Fitomejoramiento , Fusarium/genética , Sitios Genéticos , Polimorfismo de Nucleótido Simple/genética , Zea mays/genética , Resistencia a la Enfermedad/genética
3.
Plant Dis ; 107(4): 1115-1121, 2023 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-36131495

RESUMEN

Gibberella ear rot (GER) caused by Fusarium graminearum (teleomorph Gibberella zeae) is one of the most devastating maize diseases that reduces grain yield and quality worldwide. Utilization of host genetic resistance has become one of the most suitable strategies to control GER. In this study, a set of 246 diverse inbred lines derived from the intermated B 73 × Mo 17 doubled haploid population (IBM Syn10 DH) were used to detect quantitative trait loci (QTL) associated with resistance to GER. Meanwhile, a GradedPool-Seq (GPS) approach was performed to identify genomic variations involved in GER resistance. Using artificial inoculation across multiple environments, GER severity of the population was observed with wide phenotypic variation. Based on the linkage mapping, a total of 14 resistant QTLs were detected, accounting for 5.11 to 14.87% of the phenotypic variation, respectively. In GPS analysis, five significant single nucleotide polymorphisms (SNPs) associated with GER resistance were identified. Combining QTL mapping and GPS analysis, a peak-value SNP on chromosome 4 from GPS was overlapped with the QTL qGER4.2, suggesting that the colocalized region could be the most possible target location conferring resistance to GER. Subsequently, seven candidate genes were identified within the peak SNP, linking them to GER resistance. These findings are useful for exploring the complicated genetic variations in maize GER resistance. The genomic regions and genes identified herein provide a list of candidate targets for further investigation, in addition to the combined strategy that can be used for quantitative traits in plant species.


Asunto(s)
Gibberella , Sitios de Carácter Cuantitativo , Sitios de Carácter Cuantitativo/genética , Gibberella/genética , Zea mays/genética , Mapeo Cromosómico
4.
Toxins (Basel) ; 14(9)2022 08 24.
Artículo en Inglés | MEDLINE | ID: mdl-36136521

RESUMEN

Gibberella ear rot (GER) is an important fungal ear pathogen of maize that causes ear rot and toxin contamination. Most previous works have only dealt with the visual symptoms, but not with the toxins of GER. As food and feed safety rankings depend on toxin contamination, including deoxynivalenol (DON), without toxins, nothing can be said about the risks involved in food and feed quality. Therefore, three susceptible, three medium-susceptible, and three medium-resistant mother lines were crossed with three testers with differing degrees of resistance and tested between 2017-2020. Two plot replicates and two fungal strains were used separately. The highest heterosis was found at the GER% with a 13% increase across 27 hybrids, including 7 hybrids showing negative heterosis (a higher hybrid performance above the parental mean), with a variance ranging between 63.5 and -55.4. For DON, the mean heterosis was negative at -35%, and only 10 of the 27 hybrids showed a positive heterosis. The mean heterosis for DON contamination, at 1% GER, was again negative (-19.6%, varying between 85% and 224%). Only 17 hybrids showed heterosis, while that of the other 17 was rated higher than the parental mean. A positive significant correlation was found only for GER% and DON; the other factors were not significant. Seven hybrids were identified with positive (2) or negative (5) heterosis for all traits, while the rest varied. For DON and GER, only 13 provided identical (positive or negative) heteroses. The majority of the hybrids appeared to diverge in the regulation of the three traits. The stability of GER and DON (variance across eight data sets) did not agree-only half of the genotypes responded similarly for the two traits. The genetic background for this trait is unknown, and there was no general agreement between traits. Thus, without toxin analyses, the evaluation of food safety is not possible. The variety in degrees of resistance to toxigenic fungi and resistance to toxin accumulation is an inevitable factor.


Asunto(s)
Fusarium , Gibberella , Tricotecenos , Gibberella/genética , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiología , Zea mays/microbiología
5.
Plant Dis ; 106(12): 3061-3075, 2022 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-35536201

RESUMEN

The impact of Gibberella ear rot (GER; caused by Fusarium graminearum) on deoxynivalenol (DON) contamination of grain and yield components in maize were investigated using data from 30 environments in Ohio (3 years by 10 locations). Fifteen hybrids, later classified as susceptible (SU), moderately susceptible (MS), or moderately resistant (MR), based on the magnitude of differences in mean arcsine square-root-transformed GER severity (arcSEV) and log-transformed DON (logDON) relative to a reference SU check, were planted in each environment, and 10 ears per hybrid were inoculated with a spore suspension of F. graminearum. Relationships between GER severity and DON were well described by a Kono-Sugino-type nonlinear equation. Estimated parameters representing height (A) and steepness (ß) of the curves were significantly higher for SU than MS and MR hybrids but A was not significantly different between MS and MR. Results from a surrogacy analysis showed that GER was a moderate trial- and individual-level surrogate for DON. Both grain weight per ear and ear diameter decreased with increasing arcSEV but the regression slopes varied among resistance classes. The rates of reduction in both yield components per unit increase in arcSEV were significantly greater for SU than for MS and MR. An estimated 50% reduction in grain weight occurred at 62% GER severity for SU, compared with 77% severity for MS and 83% for MR. These results show that GER severity can be used as a surrogate for early estimation of DON contamination and yield loss to help guide grain handling and marketing decisions.


Asunto(s)
Gibberella , Gibberella/genética , Zea mays , Enfermedades de las Plantas , Grano Comestible , Semillas
6.
Plant Dis ; 105(7): 1984-1991, 2021 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-33616427

RESUMEN

Gibberella ear rot (GER), a prevalent disease caused by Fusarium graminearum, can result in significant yield loss and carcinogenic mycotoxin contamination in maize worldwide. However, only a few quantitative trait loci (QTLs) for GER resistance have been reported. In this study, we evaluated a Chinese recombinant inbred line (RIL) population comprising 204 lines, developed from a cross between a resistant parent DH4866 and a susceptible line T877, in three field trials under artificial inoculation with F. graminearum. The RIL population and their parents were genotyped with an Affymetrix microarray CGMB56K SNP Array. Based on the genetic linkage map constructed using 1,868 bins as markers, 11 QTLs, including five stable QTLs, were identified by individual environment analysis. Joint multiple environments analysis and epistatic interaction analysis revealed six additive and six epistatic (additive × additive) QTLs, respectively. None of the QTLs could explain more than 10% of phenotypic variation, suggesting that multiple minor-effect QTLs contributed to the genetic component of resistance to GER, and both additive and epistatic effects contributed to the genetic architecture of resistance to GER. A novel QTL, qGER4.09, with the largest effect, identified and validated using 588 F2 individuals, was colocalized with genomic regions for Fusarium ear rot and Aspergillus ear rot, indicating that this genetic locus likely confers resistance to multiple pathogens and can potentially be utilized in breeding maize varieties aimed at improving the resistance not only to GER but also other ear rot diseases.


Asunto(s)
Fusarium , Gibberella , Mapeo Cromosómico , Gibberella/genética , Fitomejoramiento , Enfermedades de las Plantas/genética , Sitios de Carácter Cuantitativo/genética , Zea mays/genética
7.
Biomed Res Int ; 2017: 5191046, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-29082248

RESUMEN

Gibberellic acid (GA3) is an important phytohormone, a member of gibberellins family, which acts as a promoter and regulator of plant growth. This study aimed to evaluate GA3 production by Fusarium moniliforme LPB03 and Gibberella fujikuroi LPB06 using different techniques of fermentation, solid state fermentation (SSF), submerged fermentation (SmF), and semisolid state fermentation (SSSF), and different types of bioreactors. In all techniques, citric pulp (CP), a subproduct obtained from the extraction of orange juice, was employed as the substrate/support. GA3 production by SSF reached 7.60 g kg-1 and 7.34 g kg-1 in Erlenmeyer flasks and column bioreactors, respectively. For SmF, the highest concentration of GA3 obtained was 236.00 mg L-1 in Erlenmeyer flasks, 273.00 mg L-1 in a 10 L stirred tank reactor (STR), and 203.00 mg L-1 in a 1.5 L bubble column reactor (BCR). SSSF was conducted with a CP suspension. In this case, GA3 concentration reached 331.00 mg L-1 in Erlenmeyer flasks and 208 mg L-1 in a BCR. The choice of the fermentation technique is undoubtedly linked to the characteristics and productivity of each process. The methods studied are inexpensive and were found to produce good proportions of GA3, making them suitable for several applications.


Asunto(s)
Ácido Cítrico/química , Fermentación , Giberelinas/biosíntesis , Reguladores del Crecimiento de las Plantas/biosíntesis , Reactores Biológicos , Fusarium/química , Fusarium/crecimiento & desarrollo , Fusarium/metabolismo , Gibberella/química , Gibberella/genética , Gibberella/crecimiento & desarrollo , Giberelinas/química , Giberelinas/genética , Reguladores del Crecimiento de las Plantas/química , Reguladores del Crecimiento de las Plantas/genética
8.
Steroids ; 128: 15-22, 2017 12.
Artículo en Inglés | MEDLINE | ID: mdl-29037584

RESUMEN

Derivatives of spironolactone (1), a diuretic and antihypertensive drug, were synthesized by using fungal cells for the first time. Ten different fungi were screened for their ability to biotransform 1, four of which were able to produce metabolites 2-8. Gibberella fujikuroi produced canrenone (2), 1-dehydrocanrenone (3), Curvularia lunuta provided compound 2, and 7α-thio-spironolactone (4), Fusarium lini yielded compounds 2, 3, 1ß-hydroxycanrenone (5), 1α-hydroxycanrenone (6), 1-dehydro-15α-hydroxycanrenone (7), and 15α-hydroxycanrenone (8), while Aspergillus alliaceus was able to produce all the seven metabolites. Metabolites 5, 6, and 7 were identified as new compounds. Their structures were elucidated by using different spectroscopic techniques. Substrate 1 and its metabolites 2, 3, and 5-8 were also evaluated for α-glucosidase inhibitory activity in vitro. Substrate 1 was found to be strongly active with IC50 = 335 ±â€¯4.3 µM as compared to the standard drug acarbose IC50 = 840 ±â€¯1.73 µM, whereas all of resulting metabolites were found to be inactive.


Asunto(s)
Antihipertensivos/síntesis química , Biotransformación , Diuréticos/síntesis química , Espironolactona/síntesis química , Antihipertensivos/química , Antihipertensivos/uso terapéutico , Ascomicetos/química , Ascomicetos/genética , Ascomicetos/metabolismo , Aspergillus/química , Aspergillus/genética , Aspergillus/metabolismo , Diuréticos/química , Diuréticos/uso terapéutico , Fusarium/química , Fusarium/genética , Fusarium/metabolismo , Gibberella/química , Gibberella/genética , Gibberella/metabolismo , Humanos , Estructura Molecular , Espironolactona/análogos & derivados , Espironolactona/química , Espironolactona/uso terapéutico
9.
Curr Microbiol ; 69(3): 381-7, 2014 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-24810291

RESUMEN

In this study, the isolation of an endophytic fungus from the leaves of the medicinal herb adlay (Coix lacryma-jobi L. var. ma-yuen Stapf) is reported for the first time. The fungus produced Triolein (trioleoylglycerol), a major constituent of triacylglycerols (TAGs) of adlay, in rice medium under shake-flask and bench-scale fermentation conditions. The fungus was identified as Gibberella moniliformis (Fusarium verticillioides) by its morphology and authenticated by ITS analysis (ITS1 and ITS2 regions and the intervening 5.8S rDNA region). Triolein was identified by HPLC-ELSD coupled with APCI-MS and confirmed through comparison with authentic standard. The concentration of triolein produced by G. moniliformis AH13 reached 2.536 ± 0.006 mg/g dry weight of mycelium. Moreover, the EtOAc extract of G. moniliformis AH13 showed strong antitumor activity against four types of tumor cells (A549, HCT116, MDA-MB-231, and SW1990). These results suggest that G. moniliformis AH13 in adlay has significant scientific and industrial potential to meet the pharmaceutical demands and sustainable energy requirements for TAGs in a cost-effective, easily accessible, and reproducible way and is also a potential novel source of natural antitumor bioactive agents.


Asunto(s)
Antineoplásicos/metabolismo , Coix/microbiología , Endófitos/clasificación , Endófitos/aislamiento & purificación , Gibberella/clasificación , Gibberella/aislamiento & purificación , Trioleína/metabolismo , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión , Análisis por Conglomerados , Medios de Cultivo/química , ADN de Hongos/química , ADN de Hongos/genética , ADN Ribosómico/química , ADN Ribosómico/genética , ADN Espaciador Ribosómico/química , ADN Espaciador Ribosómico/genética , Endófitos/genética , Endófitos/metabolismo , Gibberella/genética , Gibberella/metabolismo , Humanos , Espectrometría de Masas , Datos de Secuencia Molecular , Filogenia , Hojas de la Planta/microbiología , ARN Ribosómico 5.8S/genética , Análisis de Secuencia de ADN
10.
J Microbiol Methods ; 101: 18-23, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24704573

RESUMEN

Improved population studies in the fungus Fusarium verticillioides require the development of reliable microsatellite markers. Here we report a set of ten microsatellite loci that can be used for genetic diversity analyses in F. verticillioides, and are equally applicable to other fungi, especially those belonging to the Gibberella fujikuroi clade.


Asunto(s)
Fusarium/genética , Genoma Fúngico/genética , Repeticiones de Microsatélite/genética , Tipificación Molecular/métodos , ADN de Hongos/análisis , ADN de Hongos/genética , Fusarium/clasificación , Gibberella/clasificación , Gibberella/genética , Zea mays/microbiología
11.
Biotechnol Adv ; 32(2): 390-402, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24389085

RESUMEN

The ascomycete fungus, Fusarium graminearum (teleomorph Gibberella zeae), is the most common causal agent of Fusarium head blight (FHB), a devastating disease for cereal crops worldwide. F. graminearum produces ascospores (sexual spores) and conidia (asexual spores), which can serve as disease inocula of FHB. Meanwhile, Fusarium-infected grains are often contaminated with mycotoxins such as trichothecenes (TRIs), fumonisins, and zearalenones, among which TRIs are related to the pathogenicity of F. graminearum, and these toxins are hazardous to humans and livestock. In recent years, with the complete genome sequencing of F. graminearum, an increasing number of functional genes involved in the production of secondary metabolites, hyphal differentiation, sexual and asexual reproduction, virulence and pathogenicity have been identified from F. graminearum. In this review, the secondary metabolite synthesis, hyphal development and pathogenicity related genes in F. graminearum were thoroughly summarized, and the genes associated with secondary metabolites, sexual reproduction, energy metabolism, and pathogenicity were highlighted.


Asunto(s)
Fusarium , Genes Fúngicos/genética , Gibberella , Fusarium/genética , Fusarium/metabolismo , Fusarium/patogenicidad , Fusarium/fisiología , Gibberella/genética , Gibberella/metabolismo , Gibberella/patogenicidad , Gibberella/fisiología , Virulencia
12.
Pest Manag Sci ; 70(8): 1228-36, 2014 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-24302656

RESUMEN

BACKGROUND: Carbendazim has been used in the control of Fusarium head blight (FHB) for more than 30 years in China. Thus, carbendazim-resistant (Car(R) ) populations of Gibberella zeae have developed in some areas. In this study, 9341 G. zeae isolates were collected from the ten main wheat-producing regions of China in the period from 2008 to 2012, and sensitivity to carbendazim was detected. RESULTS: A high frequency of Car(R) isolates was observed in Zhejiang and Jiangsu provinces. Car(R) isolates were recovered from Anhui and Henan provinces in 2009 and 2012, respectively, but were not detected in the other six regions. Available (F167Y, E198Q and F200Y) and newly developed (E198L and E198K) allele-specific PCR assays were used to genotype field Car(R) isolates. The ß-tubulin variants harbouring point mutation F167Y or E198Q accounted for >95% in Car(R) populations. Quantitative allele-specific real-time PCR assays were developed to determine the frequencies of five different ß-tubulin variants present in populations of perithecia sampled from rice stubble. CONCLUSION: Car(R) populations of G. zeae develop rapidly under the selection pressure of carbendazim. Real-time PCR assays detecting the resistance frequencies in populations of perithecia would provide useful information for FHB control and management of resistance.


Asunto(s)
Adaptación Fisiológica , Bencimidazoles/toxicidad , Carbamatos/toxicidad , Farmacorresistencia Fúngica/genética , Fungicidas Industriales/toxicidad , Gibberella/fisiología , Tubulina (Proteína)/genética , China , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Genotipo , Gibberella/clasificación , Gibberella/efectos de los fármacos , Gibberella/genética , Oryza/microbiología , Enfermedades de las Plantas/microbiología , Mutación Puntual , Reacción en Cadena en Tiempo Real de la Polimerasa , Triticum/microbiología , Tubulina (Proteína)/metabolismo
13.
Genome ; 56(4): 205-13, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-23706073

RESUMEN

The intergenic spacer (IGS) region of the ribosomal DNA was cloned and sequenced in eight species within the Gibberella fujikuroi species complex with anamorphs in the genus Fusarium, a group that includes the most relevant toxigenic species. DNA sequence analyses revealed two categories of repeated elements: long repeats and short repeats of 125 and 8 bp, respectively. Long repeats were present in two copies and were conserved in all the species analyzed, whereas different numbers of short repeat elements were observed, leading to species-specific IGS sequences with different length. In Fusarium subglutinans and Fusarium nygamai, these differences seemed to be the result of duplication and deletion events. Here, we propose a model based on unequal crossing over that can explain these processes. The partial IGS sequence of 22 Fusarium proliferatum isolates was also obtained to study variation at the intraspecific level. The results revealed no differences in terms of number or pattern of repeated elements and detected frequent gene conversion events. These results suggest that the homogenization observed at the intraspecific level might not be achieved primarily by unequal crossing-over events but rather by processes associated with recombination such as gene conversion events.


Asunto(s)
ADN Espaciador Ribosómico/genética , Gibberella/genética , Intercambio Genético , Fusarium/genética , Filogenia , Secuencias Repetitivas de Ácidos Nucleicos
14.
J Biol Chem ; 288(21): 15303-17, 2013 May 24.
Artículo en Inglés | MEDLINE | ID: mdl-23553631

RESUMEN

Multimodal activation by various stimuli is a fundamental characteristic of TRP channels. We identified a fungal TRP channel, TRPGz, exhibiting activation by hyperosmolarity, temperature increase, cytosolic Ca(2+) elevation, membrane potential, and H2O2 application, and thus it is expected to represent a prototypic multimodal TRP channel. TRPGz possesses a cytosolic C-terminal domain (CTD), primarily composed of intrinsically disordered regions with some regulatory modules, a putative coiled-coil region and a basic residue cluster. The CTD oligomerization mediated by the coiled-coil region is required for the hyperosmotic and temperature increase activations but not for the tetrameric channel formation or other activation modalities. In contrast, the basic cluster is responsible for general channel inhibition, by binding to phosphatidylinositol phosphates. The crystal structure of the presumed coiled-coil region revealed a tetrameric assembly in an offset spiral rather than a canonical coiled-coil. This structure underlies the observed moderate oligomerization affinity enabling the dynamic assembly and disassembly of the CTD during channel functions, which are compatible with the multimodal regulation mediated by each functional module.


Asunto(s)
Proteínas Fúngicas/química , Gibberella/química , Canales Catiónicos TRPC/química , Calcio/química , Calcio/metabolismo , Cristalografía por Rayos X , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Gibberella/genética , Gibberella/metabolismo , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Relación Estructura-Actividad , Canales Catiónicos TRPC/genética , Canales Catiónicos TRPC/metabolismo
15.
J Basic Microbiol ; 53(11): 934-41, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-23457057

RESUMEN

Nitrilase-mediated biocatalysis has attracted substantial attention for its application in carboxylic acid production in recent years. In the present study, the fungus CA3-1 was isolated and identified as Gibberella intermedia based on its morphology, its 18S ribosomal DNA (rDNA), and internal transcribed spacer (ITS) sequences. The enzymatic properties of G. intermedia resting cells were determined, and the optimum activity was achieved at 40 °C with pH 7.6. The half-lives of the nitrilase at 30, 40, and 50 °C were 231.1, 72.9, and 6.4 h, respectively. This Gibberella nitrilase showed a wide substrate spectrum with high specificity for heterocyclic and aliphatic nitriles. It remained extremely active in 5% propanol. The presence of Ag(+), Hg(2+), and excess substrate inhibited the nitrilase activity, whereas Fe(2+), Mn(2+), and Li(+) improved enzyme activity. 3-Cyanopyridine (50 mM) was hydrolyzed into nicotinic acid within 30 min, whereas only <5% of nicotinamide was detected. The results show that this fungal nitrilase is a promising candidate for commercial application in nicotinic acid production.


Asunto(s)
Aminohidrolasas/metabolismo , Gibberella/enzimología , Gibberella/aislamiento & purificación , Análisis por Conglomerados , ADN de Hongos/química , ADN de Hongos/genética , ADN Ribosómico/química , ADN Ribosómico/genética , ADN Espaciador Ribosómico/química , ADN Espaciador Ribosómico/genética , Activadores de Enzimas/análisis , Inhibidores Enzimáticos/análisis , Genes de ARNr , Gibberella/clasificación , Gibberella/genética , Concentración de Iones de Hidrógeno , Datos de Secuencia Molecular , Filogenia , ARN de Hongos/genética , ARN Ribosómico 18S/genética , Análisis de Secuencia de ADN , Especificidad por Sustrato , Temperatura
16.
PLoS One ; 7(11): e50622, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-23226336

RESUMEN

BACKGROUND: Nitrilase is an important member of the nitrilase superfamiliy. It has attracted substantial interest from academia and industry for its function of converting nitriles directly into the corresponding carboxylic acids in recent years. Thus nitrilase has played a crucial role in production of commercial carboxylic acids in chemical industry and detoxification of nitrile-contaminated wastes. However, conventional studies mainly focused on the bacterial nitrilase and the potential of fungal nitrilase has been far from being fully explored. Research on fungal nitrilase gene expression will advance our understanding for its biological function of fungal nitrilase in nitrile hydrolysis. METHODOLOGY/PRINCIPAL FINDINGS: A fungal nitrilase gene from Gibberella intermedia was cloned through reverse transcription-PCR. The open reading frame consisted of 963 bp and potentially encoded a protein of 320 amino acid residues with a theoretical molecular mass of 35.94 kDa. Furthermore, the catalytic triad (Glu-45, Lys-127, and Cys-162) was proposed and confirmed by site-directed mutagenesis. The encoding gene was expressed in Escherichia coli Rosetta-gami (DE3) and the recombinant protein with His(6)-tag was purified to electrophoretic homogeneity. The purified enzyme exhibited optimal activity at 45°C and pH 7.8. This nitrilase was specific towards aliphatic and aromatic nitriles. The kinetic parameters V(max) and K(m) for 3-cyanopyridine were determined to be 0.81 µmol/min·mg and 12.11 mM through Hanes-Woolf plot, respectively. 3-Cyanopyridine (100 mM) could be thoroughly hydrolyzed into nicotinic acid within 10 min using the recombinant strain with the release of about 3% nicotinamide and no substrate was detected. CONCLUSIONS/SIGNIFICANCE: In the present study, a fungal nitrilase was cloned from the cDNA sequence of G. intermedia and successfully expressed in E. coli Rosetta-gami (DE3). The recombinant strain displayed good 3-cyanopyridine degradation efficiency and wide substrate spectrum. This fungal nitrilase might be a potential candidate for industrial applications in carboxylic acids production.


Asunto(s)
Aminohidrolasas/genética , Aminohidrolasas/metabolismo , Gibberella/enzimología , Histidina , Secuencia de Aminoácidos , Aminohidrolasas/química , Aminohidrolasas/aislamiento & purificación , Clonación Molecular , Expresión Génica , Gibberella/genética , Hidrólisis , Cinética , Datos de Secuencia Molecular , Nitrilos/metabolismo
17.
Eukaryot Cell ; 11(9): 1143-53, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22798392

RESUMEN

Fungi have evolved efficient metabolic mechanisms for the exact temporal (developmental stages) and spatial (organelles) production of acetyl coenzyme A (acetyl-CoA). We previously demonstrated mechanistic roles of several acetyl-CoA synthetic enzymes, namely, ATP citrate lyase and acetyl-CoA synthetases (ACSs), in the plant-pathogenic fungus Gibberella zeae. In this study, we characterized two carnitine acetyltransferases (CATs; CAT1 and CAT2) to obtain a better understanding of the metabolic processes occurring in G. zeae. We found that CAT1 functioned as an alternative source of acetyl-CoA required for lipid accumulation in an ACS1 deletion mutant. Moreover, deletion of CAT1 and/or CAT2 resulted in various defects, including changes to vegetative growth, asexual/sexual development, trichothecene production, and virulence. Although CAT1 is associated primarily with peroxisomal CAT function, mislocalization experiments showed that the role of CAT1 in acetyl-CoA transport between the mitochondria and cytosol is important for sexual and asexual development in G. zeae. Taking these data together, we concluded that G. zeae CATs are responsible for facilitating the exchange of acetyl-CoA across intracellular membranes, particularly between the mitochondria and the cytosol, during various developmental stages.


Asunto(s)
Acetilcoenzima A/metabolismo , Carnitina Aciltransferasas/metabolismo , Proteínas Fúngicas/metabolismo , Gibberella/crecimiento & desarrollo , Gibberella/metabolismo , Acetato CoA Ligasa/genética , Acetato CoA Ligasa/metabolismo , Transporte Biológico , Carnitina Aciltransferasas/genética , Citosol/metabolismo , Proteínas Fúngicas/genética , Eliminación de Gen , Gibberella/genética , Gibberella/patogenicidad , Mitocondrias/metabolismo , Peroxisomas/metabolismo , Reproducción Asexuada , Esporas Fúngicas/genética , Esporas Fúngicas/crecimiento & desarrollo , Tricotecenos/biosíntesis , Tricotecenos/genética , Virulencia/genética
18.
FEMS Microbiol Lett ; 334(1): 66-73, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22702217

RESUMEN

Fusarium head blight caused by Gibberella zeae is a prominent disease of cereal crops that poses serious human health concerns due to the contamination of grains with mycotoxins. In this study, we deleted an orthologue of areA, which is a global nitrogen regulator in filamentous fungi, to characterize its functions in G. zeae. The areA deletion resulted in an inability to use nitrate as a sole nitrogen source, whereas urea utilization was partially available. The virulence of ΔareA strains on wheat heads was markedly reduced compared with the wild-type strain. The areA mutation triggered loss of trichothecene biosynthesis but did not affect zearalenone biosynthesis. The ΔareA strains showed immaturity of asci and did not produce mature ascospores. Chemical complementation by urea restored normal sexual development, whereas the virulence and trichothecene production were not affected by urea addition. GFP-AreA fusion protein was localized to nuclei, and its expression increased in response to nitrogen-limiting conditions. These results suggest that areA-dependent regulation of nitrogen metabolism is required for vegetative growth, sexual development, trichothecene biosynthesis, and virulence in G. zeae.


Asunto(s)
Proteínas Fúngicas/metabolismo , Gibberella/metabolismo , Gibberella/patogenicidad , Nitrógeno/metabolismo , Enfermedades de las Plantas/microbiología , Factores de Transcripción/metabolismo , Proteínas Fúngicas/genética , Regulación Fúngica de la Expresión Génica , Gibberella/genética , Gibberella/crecimiento & desarrollo , Factores de Transcripción/genética , Tricotecenos/biosíntesis , Triticum/microbiología , Virulencia
19.
Mycologia ; 104(6): 1408-19, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22675046

RESUMEN

Fusarium tupiense, the main causal agent of mango malformation in Brazil, is described through a combination of morphological, biological and molecular markers. This new species belongs to the Gibberella fujikuroi species complex (GFSC) and has an anamorph morphologically similar to Fusarium mangiferae and F. sterilihyphosum. F. tupiense can be differentiated from other species in the G. fujikuroi species complex on the basis of sexual crosses, amplified fragment length polymorphism (AFLP) markers and partial sequences of the tef1 and tub2 genes. Female fertility for field isolates of F. tupiense appears to be low. PCR with primers specific for the mating type (MAT) alleles and sexual crosses identified this species as heterothallic with two idiomorphs. Female-fertile tester strains were developed for the identification of field strains of this species through sexual crosses.


Asunto(s)
Fusarium/clasificación , Gibberella/clasificación , Mangifera/microbiología , Filogenia , Enfermedades de las Plantas/microbiología , Alelos , Análisis del Polimorfismo de Longitud de Fragmentos Amplificados , Brasil , Cruzamientos Genéticos , ADN de Hongos/genética , Fusarium/citología , Fusarium/genética , Fusarium/aislamiento & purificación , Genes del Tipo Sexual de los Hongos/genética , Gibberella/citología , Gibberella/genética , Gibberella/aislamiento & purificación , Inflorescencia/microbiología , Brotes de la Planta/microbiología , Esporas Fúngicas/clasificación , Esporas Fúngicas/citología , Esporas Fúngicas/genética , Esporas Fúngicas/aislamiento & purificación
20.
PLoS One ; 7(5): e37859, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22649560

RESUMEN

The homothallic ascomycete fungus Gibberella zeae is a plant pathogen that is found worldwide, causing Fusarium head blight (FHB) in cereal crops and ear rot of maize. Ascospores formed in fruiting bodies (i.e., perithecia) are hypothesized to be the primary inocula for FHB disease. Perithecium development is a complex cellular differentiation process controlled by many developmentally regulated genes. In this study, we selected a previously reported putative transcription factor containing the Myb DNA-binding domain MYT2 for an in-depth study on sexual development. The deletion of MYT2 resulted in a larger perithecium, while its overexpression resulted in a smaller perithecium when compared to the wild-type strain. These data suggest that MYT2 regulates perithecium size differentiation. MYT2 overexpression affected pleiotropic phenotypes including vegetative growth, conidia production, virulence, and mycotoxin production. Nuclear localization of the MYT2 protein supports its role as a transcriptional regulator. Transcriptional analyses of trichothecene synthetic genes suggest that MYT2 additionally functions as a suppressor for trichothecene production. This is the first study characterizing a transcription factor required for perithecium size differentiation in G. zeae, and it provides a novel angle for understanding sexual development in filamentous fungi.


Asunto(s)
Cuerpos Fructíferos de los Hongos/citología , Regulación Fúngica de la Expresión Génica/genética , Gibberella/genética , Factores de Transcripción/metabolismo , Secuencia de Bases , Cartilla de ADN/genética , Datos de Secuencia Molecular , Vaina de Mielina/genética , Vaina de Mielina/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducción/genética , Reproducción/fisiología , Análisis de Secuencia de ADN , Factores de Transcripción/genética
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