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Epicoccum nigrum P16, a sugarcane endophyte, produces antifungal compounds and induces root growth.

Fávaro LC, Sebastianes FL, Araújo WL - PLoS ONE (2012)

Bottom Line: We report an approach based on inoculation followed by re-isolation, molecular monitoring, microscopic analysis, plant growth responses to fungal colonization, and antimicrobial activity tests to study the basic aspects of the E. nigrum endophytic interaction with sugarcane, and the effects of colonization on plant physiology.The results indicate that E. nigrum was capable of increasing the root system biomass and producing compounds that inhibit the in vitro growth of sugarcane pathogens Fusarium verticillioides, Colletotrichum falcatum, Ceratocystis paradoxa, and Xanthomomas albilineans.In addition, this work contributes to the knowledge of the interaction of this ubiquitous endophyte with the host plant, and also to a better use of microbial endophytes in agriculture.

View Article: PubMed Central - PubMed

Affiliation: Brazilian Agricultural Research Corporation, Embrapa Agroenergia, Brasília, Distrito Federal, Brazil. leia.favaro@embrapa.br

ABSTRACT

Background: Sugarcane is one of the most important crops in Brazil, mainly because of its use in biofuel production. Recent studies have sought to determine the role of sugarcane endophytic microbial diversity in microorganism-plant interactions, and their biotechnological potential. Epicoccum nigrum is an important sugarcane endophytic fungus that has been associated with the biological control of phytopathogens, and the production of secondary metabolites. In spite of several studies carried out to define the better conditions to use E. nigrum in different crops, little is known about the establishment of an endophytic interaction, and its potential effects on plant physiology.

Methodology/principal findings: We report an approach based on inoculation followed by re-isolation, molecular monitoring, microscopic analysis, plant growth responses to fungal colonization, and antimicrobial activity tests to study the basic aspects of the E. nigrum endophytic interaction with sugarcane, and the effects of colonization on plant physiology. The results indicate that E. nigrum was capable of increasing the root system biomass and producing compounds that inhibit the in vitro growth of sugarcane pathogens Fusarium verticillioides, Colletotrichum falcatum, Ceratocystis paradoxa, and Xanthomomas albilineans. In addition, E. nigrum preferentially colonizes the sugarcane surface and, occasionally, the endophytic environment.

Conclusions/significance: Our work demonstrates that E. nigrum has great potential for sugarcane crop application because it is capable of increasing the root system biomass and controlling pathogens. The study of the basic aspects of the interaction of E. nigrum with sugarcane demonstrated the facultative endophytism of E. nigrum and its preference for the phylloplane environment, which should be considered in future studies of biocontrol using this species. In addition, this work contributes to the knowledge of the interaction of this ubiquitous endophyte with the host plant, and also to a better use of microbial endophytes in agriculture.

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E. nigrum and sugarcane-associated fungi re-isolation from the root environment of sugarcane grown in greenhouse.E. nigrum and sugarcane-associated fungi were re-isolated 20 and 60 days after inoculation of the P16 endophytic strain in the roots of sugarcane plants. The E. nigrum isolation frequency was compared with the total isolation frequency of sugarcane-associated fungi. Control 1 indicates non-inoculated roots, while Control 2 indicates roots inoculated only with sterilized wheat seeds. Isolation frequency of the endophytic fungi from roots (a) is shown in CFU per root fragment. Isolation frequency of the rhizosphere fungi (b) is shown in CFU per gram of substrate. The data were transformed with √× + 0.5 (a) and Log (× + 2) (b) and submitted to analysis of variance followed by Tukey’s test. Means followed by the same letter indicate that they were not statistically different (Tukey’s test, P>5%).
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pone-0036826-g002: E. nigrum and sugarcane-associated fungi re-isolation from the root environment of sugarcane grown in greenhouse.E. nigrum and sugarcane-associated fungi were re-isolated 20 and 60 days after inoculation of the P16 endophytic strain in the roots of sugarcane plants. The E. nigrum isolation frequency was compared with the total isolation frequency of sugarcane-associated fungi. Control 1 indicates non-inoculated roots, while Control 2 indicates roots inoculated only with sterilized wheat seeds. Isolation frequency of the endophytic fungi from roots (a) is shown in CFU per root fragment. Isolation frequency of the rhizosphere fungi (b) is shown in CFU per gram of substrate. The data were transformed with √× + 0.5 (a) and Log (× + 2) (b) and submitted to analysis of variance followed by Tukey’s test. Means followed by the same letter indicate that they were not statistically different (Tukey’s test, P>5%).

Mentions: E. nigrum was not recovered as an endophyte from inoculated sugarcane plants after root superficial disinfection (Figure 2a), which indicates that E. nigrum did not colonize the endophytic root environment under the analyzed conditions. However, 20 days after inoculation, we observed a significant increase in the total isolation frequency for fungi inside the sugarcane roots (Figure 2a), but this frequency declined 60 days after inoculation. E. nigrum was recovered from the rhizosphere only during the first isolation period (Figure 2b). These differences may have been caused by the establishment of a balance in the fungal population as the plants aged; however, the presence of E. nigrum may also have brought about transient alterations in the root endophytic fungal community, as observed for the leaf epiphytic environment. Importantly, the observed shift in the plant-associated fungal isolation frequency after E. nigrum inoculation may be the results of a combination of many factors, such as the type of inoculum and substrate [43], interspecific competition, the type of exudate released, and the chemical compounds on the leaf surface [10], [44].


Epicoccum nigrum P16, a sugarcane endophyte, produces antifungal compounds and induces root growth.

Fávaro LC, Sebastianes FL, Araújo WL - PLoS ONE (2012)

E. nigrum and sugarcane-associated fungi re-isolation from the root environment of sugarcane grown in greenhouse.E. nigrum and sugarcane-associated fungi were re-isolated 20 and 60 days after inoculation of the P16 endophytic strain in the roots of sugarcane plants. The E. nigrum isolation frequency was compared with the total isolation frequency of sugarcane-associated fungi. Control 1 indicates non-inoculated roots, while Control 2 indicates roots inoculated only with sterilized wheat seeds. Isolation frequency of the endophytic fungi from roots (a) is shown in CFU per root fragment. Isolation frequency of the rhizosphere fungi (b) is shown in CFU per gram of substrate. The data were transformed with √× + 0.5 (a) and Log (× + 2) (b) and submitted to analysis of variance followed by Tukey’s test. Means followed by the same letter indicate that they were not statistically different (Tukey’s test, P>5%).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366970&req=5

pone-0036826-g002: E. nigrum and sugarcane-associated fungi re-isolation from the root environment of sugarcane grown in greenhouse.E. nigrum and sugarcane-associated fungi were re-isolated 20 and 60 days after inoculation of the P16 endophytic strain in the roots of sugarcane plants. The E. nigrum isolation frequency was compared with the total isolation frequency of sugarcane-associated fungi. Control 1 indicates non-inoculated roots, while Control 2 indicates roots inoculated only with sterilized wheat seeds. Isolation frequency of the endophytic fungi from roots (a) is shown in CFU per root fragment. Isolation frequency of the rhizosphere fungi (b) is shown in CFU per gram of substrate. The data were transformed with √× + 0.5 (a) and Log (× + 2) (b) and submitted to analysis of variance followed by Tukey’s test. Means followed by the same letter indicate that they were not statistically different (Tukey’s test, P>5%).
Mentions: E. nigrum was not recovered as an endophyte from inoculated sugarcane plants after root superficial disinfection (Figure 2a), which indicates that E. nigrum did not colonize the endophytic root environment under the analyzed conditions. However, 20 days after inoculation, we observed a significant increase in the total isolation frequency for fungi inside the sugarcane roots (Figure 2a), but this frequency declined 60 days after inoculation. E. nigrum was recovered from the rhizosphere only during the first isolation period (Figure 2b). These differences may have been caused by the establishment of a balance in the fungal population as the plants aged; however, the presence of E. nigrum may also have brought about transient alterations in the root endophytic fungal community, as observed for the leaf epiphytic environment. Importantly, the observed shift in the plant-associated fungal isolation frequency after E. nigrum inoculation may be the results of a combination of many factors, such as the type of inoculum and substrate [43], interspecific competition, the type of exudate released, and the chemical compounds on the leaf surface [10], [44].

Bottom Line: We report an approach based on inoculation followed by re-isolation, molecular monitoring, microscopic analysis, plant growth responses to fungal colonization, and antimicrobial activity tests to study the basic aspects of the E. nigrum endophytic interaction with sugarcane, and the effects of colonization on plant physiology.The results indicate that E. nigrum was capable of increasing the root system biomass and producing compounds that inhibit the in vitro growth of sugarcane pathogens Fusarium verticillioides, Colletotrichum falcatum, Ceratocystis paradoxa, and Xanthomomas albilineans.In addition, this work contributes to the knowledge of the interaction of this ubiquitous endophyte with the host plant, and also to a better use of microbial endophytes in agriculture.

View Article: PubMed Central - PubMed

Affiliation: Brazilian Agricultural Research Corporation, Embrapa Agroenergia, Brasília, Distrito Federal, Brazil. leia.favaro@embrapa.br

ABSTRACT

Background: Sugarcane is one of the most important crops in Brazil, mainly because of its use in biofuel production. Recent studies have sought to determine the role of sugarcane endophytic microbial diversity in microorganism-plant interactions, and their biotechnological potential. Epicoccum nigrum is an important sugarcane endophytic fungus that has been associated with the biological control of phytopathogens, and the production of secondary metabolites. In spite of several studies carried out to define the better conditions to use E. nigrum in different crops, little is known about the establishment of an endophytic interaction, and its potential effects on plant physiology.

Methodology/principal findings: We report an approach based on inoculation followed by re-isolation, molecular monitoring, microscopic analysis, plant growth responses to fungal colonization, and antimicrobial activity tests to study the basic aspects of the E. nigrum endophytic interaction with sugarcane, and the effects of colonization on plant physiology. The results indicate that E. nigrum was capable of increasing the root system biomass and producing compounds that inhibit the in vitro growth of sugarcane pathogens Fusarium verticillioides, Colletotrichum falcatum, Ceratocystis paradoxa, and Xanthomomas albilineans. In addition, E. nigrum preferentially colonizes the sugarcane surface and, occasionally, the endophytic environment.

Conclusions/significance: Our work demonstrates that E. nigrum has great potential for sugarcane crop application because it is capable of increasing the root system biomass and controlling pathogens. The study of the basic aspects of the interaction of E. nigrum with sugarcane demonstrated the facultative endophytism of E. nigrum and its preference for the phylloplane environment, which should be considered in future studies of biocontrol using this species. In addition, this work contributes to the knowledge of the interaction of this ubiquitous endophyte with the host plant, and also to a better use of microbial endophytes in agriculture.

Show MeSH
Related in: MedlinePlus