Two fungal endophytes reduce the severity of pitch canker disease in Pinus radiata seedlings Pablo Martínez-Álvarez a,b,⇑ , Raúl Arcadio Fernández-González a,b , Antonio Vicente Sanz-Ros a,b,c , Valentín Pando a,d , Julio Javier Diez a,b a Sustainable Forest Management Research Institute, University of Valladolid – INIA, Avenida Madrid 44, 34071 Palencia, Spain b Departamento de Producción Vegetal y Recursos Forestales, University of Valladolid, Avenida Madrid 44, 34071 Palencia, Spain c Calabazanos Forest Health Center, Junta de Castilla y León, Polígono Industrial de Villamuriel S/N, 34190 Villamuriel de Cerrato, Palencia, Spain d Departamento de Estadística e Investigación Operativa, University of Valladolid, Avenida Madrid 57, 34004 Palencia, Spain highlights  Five hundred forty-six endophytes were tested against F. circinatum in vitro.  The antagonistic activity of the endophytes was quantify using five indicators.  The six endophytes that showed the most promising results were tested in the field.  Chaetomium aureum and Alternaria sp. reduced the damages caused by the pathogen. graphical abstract article info Article history: Received 10 September 2015 Revised 27 November 2015 Accepted 30 November 2015 Available online 5 December 2015 Keywords: Fusarium circinatum Field test Inoculation Dual cultures Biological control agents abstract Pitch canker disease, which affects pines and is caused by the fungus Fusarium circinatum, cannot be effectively controlled at present. Current restrictions on the use of chemicals and fungicides in forests are driving research into alternative methods of reducing the damage caused by the pathogen. Biological control with fungal endophytes is a promising and environmentally friendly strategy. In this study, 154 endophyte isolates were selected from a collection of 546 fungi tested in a preliminary confrontation assay. These isolates were then tested against F. circinatum in an in vitro antagonism exper- iment. Four different types of indicators (length of the central axis of the colony of the pathogen, the shape coefficient, percentage inhibition of radial growth and percentage inhibition zone) were used to detect and quantify the antagonistic activity directed towards the pathogen by the endophytes. The six isolates that showed the most promising results were inoculated in the field, together with the pathogen, into seedlings of Pinus radiata, P. sylvestris, P. pinaster, P. nigra and P. pinea, to test whether they could reduce the damage caused by F. circinatum. In total, 138 endophytes displayed antagonistic activity towards F. circinatum in the dual cultures of the in vitro experiment. In the field test, the endophytes Chaetomium aureum and Alternaria sp. reduced the area under disease progress curve (AUDPC) for the P. radiata seedlings, indicating that they may therefore be suitable for use as biological control agents (BCAs) of the disease. Ó 2015 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.biocontrol.2015.11.011 1049-9644/Ó 2015 Elsevier Inc. All rights reserved. ⇑ Corresponding author at: Sustainable Forest Management Research Institute, University of Valladolid – INIA, Avenida Madrid 44, 34071 Palencia, Spain. E-mail address: pmtnez@pvs.uva.es (P. Martínez-Álvarez). Biological Control 94 (2016) 1–10 Contents lists available at ScienceDirect Biological Control journal homepage: www.elsevier.com/locate/ybcon