9This study assessed and optimized an in vivo method to evaluate the levels of 10 susceptibility/resistance in fruit from the EEAD-CSIC peach germplasm to an isolate of 11 Monilinia laxa (Aderhold & Ruhland) Honey from peach. A total of four commercial 12 cultivars and six genotypes, descendants of three families, of peach [Prunus persica (L.) 13 88inoculation. 89Using the available methods, longer periods are required for obtaining the source of 90 inoculum and the amount and purity of inoculum are not assured. Moreover, an easy 91 5 inoculation system would help to screen large populations and controlled environment 92 would be preferable to avoid cross contamination. We have modified the existing protocols 93 by shortening the inoculum production and also ensuring the accurate supply of inoculum. 94The objective of this study, therefore, was to have an evaluation method, by optimizing the 95 available protocols, to screen tolerance to brown rot by Monilinia spp. in peach germplasm. 96 We have evaluated brown rot tolerance on Monilinia laxa ex situ inoculated peach fruit by 97 measuring brown rot incidence (%), lesion diameter (mm) and colonization extent (mm). In 98 the Spanish peach collection at the EEAD-CSIC, screening for resistance to brown rot has 99 shown that local germplasm accessions are susceptible although slight differences can be 100 found.

Abstract:The peach is one of the most important global tree crops within the economically important Rosaceae family. The crop is threatened by numerous pests and diseases, especially fungal pathogens, in the field, in transit, and in the store. More than 50% of the global post-harvest loss has been ascribed to brown rot disease, especially in peach late-ripening varieties. In recent years, the disease has been so manifest in the orchards that some stone fruits were abandoned before harvest. In Spain, particularly, the disease has been associated with well over 60% of fruit loss after harvest. The most common management options available for the control of this disease involve agronomical, chemical, biological, and physical approaches. However, the effects of biochemical fungicides (biological and conventional fungicides), on the environment, human health, and strain fungicide resistance, tend to revise these control strategies. This review aims to comprehensively compile the information currently available on the species of the fungus Monilinia, which causes brown rot in peach, and the available options to control the disease. The breeding for brown rot-resistant varieties remains an ideal management option for brown rot disease control, considering the uniqueness of its sustainability in the chain of crop production.

A sustainable approach to control the incidence of brown rot in pre-and post-harvest management is to select genotypes with high contents of antioxidant compounds and tolerance to Monilinia laxa (Aderh. and Ruhland) Honey. In this study, 68 progenies of the 'Babygold 9' × 'Crown Princess' population from the EEAD-CSIC breeding program were screened under controlled conditions for a period of 3 years (2013-2015). Susceptibility to brown rot was evaluated after inoculating 20 healthy fruits per genotype with M. laxa. Brown rot incidence, lesion diameter, and colonization extent, as well as the severities of these issues, were calculated after 5 days of incubation. Physicochemical traits, such as fruit firmness and soluble solids content, were also recorded before and after storage. Titratable acidity, pH, and antioxidant composition were measured at harvest. Significant differences were found for pathogenic traits, as well as for contents of vitamin C, total phenolics, flavonoids, and anthocyanins, within genotypes in this population. Negative correlations were also found between the content of phytochemical compounds (such as anthocyanins and total phenolics), as well as disease incidence and severity. Differences in susceptibility to brown rot confirm the genetic variability available in these progeny. This allowed the selection of six genotypes highly resistant to brown rot of M. laxa, with high organoleptic properties and high phenol content, to be introduced in our peach breeding program.

The black truffle (Tuber melanosporum Vittad.) is an important natural resource due to its relevance as a delicacy in gastronomy. Different aspects of this hypogeous fungus species have been studied, including population genetics of French and Italian distribution ranges. Although those studies include some Spanish populations, this is the first time that the genetic diversity and genetic structure of the wide geographical range of the natural Spanish populations have been analysed. To achieve this goal, 23 natural populations were sampled across the Spanish geographical distribution. ISSR technique demonstrated its reliability and capability to detect high levels of polymorphism in the species. Studied populations showed high levels of genetic diversity (h N  = 0.393, h S  = 0.678, Hs = 0.418), indicating a non threatened genetic conservation status. These high levels may be a consequence of the wide distribution range of the species, of its spore dispersion by animals, and by its evolutionary history. AMOVA analysis showed a high degree of genetic structure among populations (47.89%) and other partitions as geographical ranges. Bayesian genetic structure analyses differentiated two main Spanish groups separated by the Iberian Mountain System, and showed the genetic uniqueness of some populations. Our results suggest the survival of some of these populations during the last glaciation, the Spanish southern distribution range perhaps surviving as had occurred in France and Italy, but it is also likely that specific northern areas may have acted as a refugia for the later dispersion to other calcareous areas in the Iberian Peninsula and probably France.

The quality of seedlings colonized by Tuber melanosporum is one of the main factors that contributes to the success or failure of a truffle crop. Truffle cultivation has quickly grown in European countries and elsewhere, so a commonly shared seedling evaluation method is needed. Five evaluation methods are currently published in the literature: three are used in Spain and two in France and Italy. Although all estimate the percentage colonization by T. melanosporum mycorrhizae, they do it in different ways. Two methods also estimate total number of mycorrhizae per seedling. Most are destructive. In this work, ten batches of holm oak seedlings inoculated with T. melanosporum from two different nurseries were evaluated by means of the five methods noted above. Some similarity was detected between the percentages of T. melanosporum mycorrhizae estimated by each method but not in their ability to assess the suitability of each batch. We discuss the advantages and disadvantages for each method and suggest approaches to reach consensus within the truffle culture industry for certifying mycorrhizal colonization by T. melanosporum and seedling quality.

This investigation examines the effects of pH and titratable acidity on the growth and developments of a strain of Monilinia laxa (Aderhold & Ruhland) at seven different pH levels in Potato Dextrose Agar media and on peach fruit from formation to commercial maturity. The fungi growth was obtained by daily measurement of mycelia on the pH amended Potato Dextrose Agar. The sporulation performance was determined after 30 days of culture incubation. Fruits were inoculated with M. laxa, from fruit set to maturity, on weekly basis for brown rot susceptibility. The pathogen development, in vitro, was affected, by the pH (2.4-11.52) amended nutrient media. M. laxa exhibited variation in its growth and sporulation capacities on the seven pH amended PDA, preferring relatively moderate acidic conditions for optimum performance. In the in vitro analysis, there was mycelia growth at pH 2.40 to 8.84, while pH 11.52 did not support any mycelia growth. There was a continuous and stable increase in weight of fruit as it developed whereas the fruit size increased, then decreased and finally increased as the fruit develops. The acidity dynamics exhibited a non-sinusoidal waveform through the growth and development of the fruit. In all these characteristic variations, M. laxa did not develop infection or shown any brown rot incidence in the fruit until the period of commercial maturity.

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We investigated the effects of three plant growth promoting rhizobacteria (PGPR), on Biological Nitrogen Fixation (BNF), nodulation and growth promotion by soybean (Glycine max) var. Osumi plants. The strains, Aur 6, Aur 9 and Cell 4, belong to Psedomonas fluorescens, Chryseobacterium balustinum and Serratia fonticola, respectively. Inoculation modes for the PGPRs and Sinorhizobium fredii (carried out through irrigation), were examined. In the first mode, PGPRs and S. fredii were co-inoculated. In the second mode, we first inoculated S. fredii and after the PGPRs, which were added 5 or 10 days later (each inoculation being an independent treatment). In the third mode, the PGPRs were inoculated first, and the S. fredii was inoculated 5 days later. We also included treatments inoculated with only the PGPRs (one PGPR per treatment) and only with S. fredii. Plants were maintained in a greenhouse under controlled environmental conditions, and were sampled 3 months after sowing. The results obtained showed the effects of the inoculation sequence. The most significant effects on growth parameters (stem plus leaf weight and fresh root weight) were found when inoculations with PGPR and S. fredii were at different times or when we inoculated only with PGPR and the plants were watered with nitrogen. Co-inoculation had no positive effects on any parameter, probably due to competition between the PGPR and S. fredii. Our results indicate that the inoculation modes with PGPR and rhizobia play a very important role in the effects produced. Thus, although plant growth promoting rhizobacteria may interact synergistically with root-nodulating rhizobia, plant growth promoting rhizobacteria selected for one crop should be assessed for potentially hazardous effects on other crops before being used as inoculants.