Next Article in Journal
Coordinated Effect of Ascorbate Biosynthesis and Recycling in Maize Seed Germination and Seedling Establishment under Low Temperature
Next Article in Special Issue
Evaluation of Phenological and Agronomical Traits of Different Almond Grafting Combinations under Testing in Central Italy
Previous Article in Journal
Genome-Wide Characterization of Glutamine Synthetase Family Genes in Cucurbitaceae and Their Potential Roles in Cold Response and Rootstock-Scion Signaling Communication
Previous Article in Special Issue
Hazelnut Kernel Size and Industrial Aptitude
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Agriculture
Volume 11
Issue 11
10.3390/agriculture11111158
agriculture-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Evaluation of Chestnut Susceptibility to Cryphonectria parasitica: Screening under Controlled Conditions

by
Emigdio Jordán Muñoz-Adalia
1,2,
Andreu Meijer
1,2,*,
Joan Abel
3,
Carlos Colinas
1,2,
Neus Aletà
3 and
Mercè Guàrdia
3
1
Forest Science and Technology Centre of Catalonia (CTFC), St. Llorenç de Morunys, km. 2, Solsona, 25280 Lleida, Spain
2
Department of Crop and Forest Science, University of Lleida, Av. de l’Alcalde Rovira Roure 191, 25198 Lleida, Spain
3
Institut de Recerca i Tecnologia Agroalimentàries (IRTA) Torre Marimon, Caldes de Montbui, 08140 Barcelona, Spain
*
Author to whom correspondence should be addressed.
Agriculture 2021, 11(11), 1158; https://doi.org/10.3390/agriculture11111158
Submission received: 7 October 2021 / Revised: 12 November 2021 / Accepted: 16 November 2021 / Published: 18 November 2021
(This article belongs to the Special Issue Development and Cultivar Improvement of Nut Crops)
Browse Figures
Versions Notes

Abstract

:
Cryphonectria parasitica (Murrill) M.E. Barr (Sordariomycetes, Valsaceae) is the causal agent of chestnut blight. This disease is a major concern for chestnut cultivation in Europe. The fungus colonizes vascular tissues and evolves generating cankers causing severe dieback and the death of the tree. Excised and debarked well-lignified shoots of 28 C. sativa genotypes (assay A) and of 10 progenies (assay B) were inoculated with C. parasitica strain FMT3bc2 (vcg: EU2). Fungal growth was measured along the longitudinal axis on the 3rd and 6th days after inoculation. Results indicated the inoculation methodology works and the results were clear after 6 days. Differences in susceptibility to chestnut blight among C. sativa trees of Montseny have been detected both at the individual genotype level and at the progeny level. Nineteen genotypes and four progenies showed a susceptibility to Blight not significantly different from C. mollissima. The methodology was easy to apply in extensive/preliminary selection screenings to assess the susceptibility of C. sativa materials to the Blight.

1. Introduction

The Ascomycete fungus Cryphonectria parasitica (Murrill) M.E. Barr (Sordariomycetes, Valsaceae) is the causal agent of chestnut blight, one of the most devastating diseases affecting Castanea sp. worldwide. The disease in Europe was first detected in Italy in 1938 (EPPO current status: Present) [1,2] from where it spread to the neighboring countries of Switzerland (1948; widespread), Slovenia (1950; restricted distribution), and France (1956; restricted distribution) [3,4]. The disease is currently present in Spain (1947; restricted distribution), Croatia (1955; widespread), Albania (1967; widespread), Serbia (1975; widespread), Portugal (1989; widespread) and Germany (1992; restricted distribution), Azerbaijan (2004; present), United Kingdom (2011; restricted distribution), and Belgium (2014; restricted distribution) [1,5]. Most of the European C. parasitica populations seem to have originated from single introductions since they have reduced genetic diversity [6].
Spore dispersal by air is a major dissemination mechanism of fungal pathogens [7,8]. In the case of C. parasitica, air-borne spores usually penetrate above-ground parts of the host through bark wounds (e.g., pruning, wind, hail, etc.) in shoots or stems. The fungus colonizes vascular tissues causing progressive wilting, and severe reduction in fruit production. Later symptoms include orangish necrotic areas in the bark [2,9]. The necrosis evolves generating cankers that usually girdle branches or even the trunk causing severe dieback, and finally the death of the tree [10,11]. Cankers harbor fruiting bodies of the fungus that disseminate their spores throughout the chestnut stands. The fungus infects Castanea sativa Mill. (Europe), Castanea dentata (Marshall) Borkh. (America), as well as Castanea crenata Siebold and Zucc. and Castanea mollissima Blume (Asia) at any age. Among these host species, C. sativa and C. dentata are considered the most susceptible since C. mollissima and C. crenata exhibit some tolerance to the disease probably caused by ancient co-evolution with the pathogen in their native range [12,13].
This disease is a major concern for chestnut cultivation in Europe as there is currently no effective cultural or chemical control and no resistance has been found in C. sativa species [14]. To find a way out of chestnut blight, two main strategies are being considered: biocontrol through virus-induced fungus hypovirulence, and prevention by breeding disease-resistant genotypes. The use of RNA mycoviruses (i.e., Cryphonectria hypovirus 1; Hypoviridae) has resulted in highly effective in Europe [1], thus becoming one of the most successful alternatives for controlling the disease in natural stands. This biocontrol method is based on inoculation of virus-infected fungal strains belonging to the same vegetative compatibility group/s (vcg) than those causing disease in the target stand [15,16]. This infected—and therefore weakened—strain, inoculated in diseased trees, naturally transfers viruses by mycelial anastomosis to virulent strains which will become in turn hypovirulent. Mycovirus-infected strains of C. parasitica are not able to severely damage the tree allowing it to withstand the infection with minor symptoms [17,18]. On the other hand, breeding resistant genotypes of chestnut for planting is probably the most promising preventive measure to be implemented for woody croplands and afforestation. The transfer of resistance from Asian chestnuts to American or European species has been carried out in several programs but reaching the desired fruit quality requires several backcrosses that lengthen the selection process considerably [19,20]. Finding resistance or lower susceptibility traits in natural populations of C. dentata, the American chestnut, has proven possible [21] although conducting this work by inoculation of standing trees requires a great deal of work that is often not feasible. The evaluation of tolerance/resistance to biotic or abiotic injuries under controlled conditions on parts of plants/trees has proven to be an advantageous practice when many individuals/samples are to be tested. Thus, in different hardwoods similar approaches have been used, such as on Juglans sp. to study frost resistance [22] or more specifically on chestnut to establish levels of susceptibility to Phytophthora spp. [23]. Moreover, Rodríguez and Colinas [24] showed that estimating chestnut susceptibility to C. parasitica by means of inoculation on excised shoots was feasible and the mycelium grew as expected under controlled conditions.
In consequence, the aims of this study were: (i) to evaluate the tolerance to chestnut blight of some C. sativa genotypes from a Spanish local population (Montseny-Northeast of Catalonia) in excised shoots, and (ii) to study this tolerance in seedling genotypes of the same origin installed in a progeny test.

2. Materials and Methods

2.1. In Vitro Inoculation: Budstick Assay

2.1.1. Assay A

Tolerance to chestnut blight among C. sativa genotypes was evaluated in vitro by inoculating a virulent strain of C. parasitica in debarked well lignified shoots. One-year-old budsticks of 28 C. sativa genotypes (Table 1) were removed from healthy mother plants installed in a quarantine greenhouse of Gimenells (Lleida, 41°44′45.6″ N; 0°23′39.1″ E) at IRTA experimental station in July 2020. The harvest date was adjusted to the optimal physiological state of chestnut for the susceptibility test [24]. Budsticks (12–15 cm long) were kept at 4 °C covering the apical ends with Parafilm© while the opposite side was submerged in distilled water until processed. Moreover, lignified shoots from two genotypes of C. mollissima were collected at Lourizan Forest Center of Galicia (42°24′35″ N—8°00′12″ W) and sent overnight to be used as a tolerant control (Table 1).
Inoculation was performed according to the methodology of Rodríguez and Colinas [24] with minor modifications. Each budstick was cut in 7 cm sections (8 replicates per genotype; 224 samples of C. sativa and 16 samples of C. mollissima) (Figure 1) in the laboratory with disinfected pruning scissors. The diameter of each cutting sample (d) was measured with a digital caliper and the section manually debarked using a sterile scalpel. Fungal cultures of C. parasitica strain FMT3bc2 (vcg: EU2) grown at room temperature for 5 days in PDA medium (potato-dextrose-agar 3.90% w/v; Biokar) were used as an inoculum source. More specifically, a 5 × 5 mm plug of a solid medium plug with fresh mycelium from the edge of the colony was placed in the center of each debarked shoot section with the mycelium side in touch with wood tissue (Figure 1).
Inoculated samples were labeled with white tape and randomly placed in disinfected plastic trays lined with filter paper (Figure 1). Each tray was then moistened by spraying deionized water and covered with a plastic film to maintain high relative air humidity inside the tray. The trays were protected with a sheet of paper and incubated at room temperature (24 ± 2 °C) for one week. Whenever the filter paper became dry, deionized water was sprayed in the trays. Fungal growth was measured along the longitudinal axis (mm) on the 3rd and 6th days after inoculation.

2.1.2. Assay B

In parallel to assay A, a total of 166 C. sativa genotypes belonging to 10 progenies (Table 1) were used for an additional in vitro study. Tree shoots (4 replicates per seedling) were collected in a progenies test orchard located in the Parc Natural i Reserva de la Biosfera del Montseny (41°45′46,6″ N; 2°25′39,1″ E) and were processed and incubated as previously described for bioassay A. The same two C. mollissima genotypes, four shoots of each one, were also used as control samples. The fungal growth rate was calculated as previously explained.

2.2. Statistical Analysis

Data obtained after six days of incubation were analyzed for both in vitro assays using the programming environment R [25]. We used the average value of the mean daily fungal growth rate for a six-day incubation period (G, mm/day) of both C. mollisima genotypes (i.e., “M2” and “M5M”) as reference for model computation (control). This was tested by fitting two preliminary generalized linear models (GLMs) (one per dataset) in order to evaluate whether G significantly varied between both C. mollissima genotypes.
In assay A, a GLM was fitted as a null model including the mean daily fungal growth rate (G) as a response variable and the genotypes as an explanatory factor. Then, a linear mixed-effects model (LMM) was computed using the “lme4” package in R [26] including G as the dependent variable, genotype as a fixed factor and the diameter of the cutting (d) as a random factor (Table 2). Both models were compared using Akaike’s Information Criteria (AIC) [27] using the “AICcmodavg” package in R [28]. The most parsimonious model was selected using AIC and the result of the model comparison using the χ2 test.
In assay B two different model sets were fitted (Table 2). A null GLM was computed with G as a response variable and progeny as an explanatory factor. Then, an LMM with the same variables as well as diameter as a random factor was computed (Table 2). Analogous models (i.e., null and LMM) were fitted for the same dataset including the seedling genotypes instead of progenies as explanatory variables. Definitive models were selected following the same statistical indicators as described for assay A.

3. Results

The generalized linear model fitted for assay A showed significant variation in G among the evaluated materials (p-value = 0.03) (model Ma0). The LMM fitted to dataset A including diameter as a random factor (i.e., Ma1) retained the significance of genotype (p-value = 0.03). The most parsimonious model (Ma0) did not show conclusive differences with Ma1 according to AIC values (ΔAICMa1-Ma0 = 2; Table 2), so we compared them with a χ2 test (p-value = 1). Consequently, Ma1 was selected as the most explicative model for assay A (Table 2). The parameters of the model showed that in nineteen of the assayed genotypes (i.e., “BRL02”, ”BRL03”, ”BRL06”, “FGM01”, “FGM04”, “MSY04”, “MSY08”, “MSY09”, ”MSY10”, “MSY11”, “MSY14”, “PO11”, “RLL05”, “VLD14”, “VLD22”, “VDL29”, “VLD30”, “VLD32” and “VLD34”) the daily lesion growth rate (G) was not significantly different from the control C. mollissima (Figure 2).
In the dataset of assay B, two null models (GLMs) were computed to evaluate variations of G in respect of progenies (model Mbp0) and genotypes (model Mbg0). Both models showed a significant effect of the corresponding explicative factors (p-value < 0.01 in all cases). The LMM fitted to analyze the variation of G among genotypes including diameter as a random factor (model Mbg1) retained the significance already shown by the corresponding null model Mbg0. In addition, change in model AIC was not noticeable (ΔAICMbg1-Mbg0 = 2) and both models did not differ significantly (p-value = 1). In contrast, the fitted LMM Mbp1 that included diameter as a random factor resulted more parsimonious than the corresponding null model (ΔAICMbp1-Mbp0 = −4.3) and was significantly different from Mbp0 (p-value = 0.01). In consequence, either Mbp1 or Mbg1 were selected as the most explicative models for this in vitro assay (Table 2). Both models supported the lower susceptibility of C. mollissima against C. parasitica (Figure 3 and Figure S1).
According to this model, the tolerance to C. parasitica of progenies “CS-49”, “CS-51”, “CS-54”, and “CS-62” is not significantly different from that of tolerant C. mollissima making them good candidates for further studies regarding their suitability to be planted in orchards. The use of average G values for C. mollissima genotypes as a reference in model fitting was supported by the lack of significant differences of G between “M2” and “M5M” in any of the datasets (p-value > 0.20 in all cases).

4. Discussion

In this assay, C. parasitica colonized the wood in the first six days after inoculation in contrast with other methodologies where longer periods were required [29]. The methodology used, debarking of lignified shoots, was expected to improve colonization by the fungus compared to inoculation on live seedlings [30] or on stems with bark [13]. In addition, working under controlled conditions allows to not have to worry about the potential infection and death of the plant, or part of the plant, or introducing new vcgs potentially foreign to an area. Future research involving inoculation with fungi from different populations—even in simultaneous infection—will be informative to characterize the most resistant genotypes suitable for establishing new plantations. However, an aspect to be considered is the virulence of the fungal strain used which is expected to vary among isolates [31]. In this study, we only tested one fungal strain since our main objective was to characterize host susceptibility against a strain of the EU2 vcg, the most common vcg type in the Montseny area [32].
Regarding features of plant material, Pažitný et al. [12] reported a significant effect of stem diameter on chestnut blight lesion development. This observation completely agrees with our fitted models (Table 2) where the inclusion of stem section diameter provided a better explanation of recorded data.
Nearly all the C. sativa materials included in this study come from a reduced population of chestnut trees of the Montseny Natural Park, therefore, extending this study to a wider genetic base would be the first step to take in future evaluations. “PO11” is the only material not originating from this population, it is a spontaneous chestnut hybrid selected as a rootstock of C. sativa [33] for its resistance to Phytophthora spp. In this evaluation “PO11” has been included in the group whose tolerance is comparable to the one of C. mollissima, which was expected since this genotype carries some Asian genes [34].
It should be noted that 70% of the C. sativa genotypes and the 40% of progenies from Montseny inoculated with C. parasitica showed a response analogous to the one of C. mollissima in their canker development. These results are relevant because the chestnut orchards in Montseny are not described as having been introgressed with Asian materials as in other Spanish areas [35]. Their ancestry and traditional management in the last 100 years do not indicate the entry of foreign fruit varieties. Nor are there any genotypes with proper names that could suggest that they are ancient varieties of the Montseny area. Chestnut trees sampled from this area, according to Mattioni et al. [36], clustered (Nei distance) together with trees from southern Italy and Sicily but not with those of bordering areas. Southern Italy chestnuts have genetics from the Eastern Mediterranean [36]. Perhaps by going deeper into the origin of the Montseny materials, it would be possible to identify other materials to include in further C. parasitica susceptibility screenings.

5. Conclusions

Differences in susceptibility to chestnut blight among C. sativa trees have been detected both at the individual genotype level and at the progeny level. The debarked budstick-based inoculation method used here revealed nineteen genotypes and four progenies of C. sativa with a susceptibility to C. parasitica not different from that of C. mollissima.

Supplementary Materials

The following are available online at https://www.mdpi.com/article/10.3390/agriculture11111158/s1, Figure S1: Values of mean daily lesion growth rate (G) by Castanea sativa genotype fitted by model Mbg1 (Assay B). Mean values and 95% confidence error are shown. Asterisk (*) denotes significant differences in respect to Castanea mollissima (average value of “M2” and “M5M”). Number of repetitions: 4 (“M2” and “M5M”); 4 (all genotypes of C. sativa).

Author Contributions

Conceptualization, C.C. and N.A.; methodology, A.M., J.A. and C.C.; formal analysis, E.J.M.-A. and A.M.; investigation, A.M. and J.A.; resources, J.A. and A.M.; data curation, A.M.; writing—original draft preparation, E.J.M.-A. and A.M.; writing—review and editing, M.G., N.A. and C.C.; visualization, E.J.M.-A. and A.M.; supervision, M.G., N.A. and C.C.; project administration, J.A.; funding acquisition, J.A., N.A. and C.C. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded in part by Diputació de Barcelona, Àrea de Territori i Sostenibilitat.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Acknowledgments

Our appreciation to the staff of the Technical Office of the Natural Parks of the ‘Diputació de Barcelona (DiBa)’, and to Josep Argemí, head of the Monitoring Unit of the Technical Programmes developed in the Montseny Natural Park, and to all the park rangers for their help in locating and collecting the C. sativa materials. Sincere gratitude to Fina Fernandez and Beatriz Míguez from ‘Centro de Investigaciones Forestales’ of Lourizán for kindly providing the C. mollissima genotypes. The authors also thank Anand Babu Uppara (Universitat de Lleida and Forest Bioengineering Solutions) for his participation in laboratory tasks.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Robin, C.; Heiniger, U. Chestnut blight in Europe: Diversity of Cryphonectria parasitica, hypovirulence and biocontrol. For. Snow Landsc. Res. 2001, 76, 361–367. [Google Scholar]
  2. Anagnostakis, S.L. Chestnut Blight: The Classical Problem of an Introduced Pathogen. Mycologia 1987, 79, 23–37. [Google Scholar] [CrossRef] [Green Version]
  3. Heiniger, U.; Rigling, D. Biological control of chestnut blight in Europe. Annu. Rev. Phytopathol. 1994, 32, 581–599. [Google Scholar] [CrossRef]
  4. Krstin, L.; Novak-Agbaba, S.; Rigling, D.; Krajačić, M.; Ćurković Perica, M. Chestnut blight fungus in Croatia: Diversity of vegetative compatibility types, mating types and genetic variability of associated Cryphonectria hypovirus 1. Plant Pathol. 2008, 57, 1086–1096. [Google Scholar] [CrossRef]
  5. EPPO. Cryphonectria parasitica. Available online: https://gd.eppo.int/taxon/ENDOPA (accessed on 10 October 2021).
  6. Milgroom, M.G.; Sotirovski, K.; Spica, D.; Davis, J.E.; Brewer, M.T.; Milev, M.; Cortesi, P. Clonal population structure of the chestnut blight fungus in expanding ranges in southeastern Europe. Mol. Ecol. 2008, 17, 4446–4458. [Google Scholar] [CrossRef]
  7. Rigling, D.; Prospero, S. Cryphonectria parasitica, the causal agent of chestnut blight: Invasion history, population biology and disease control. Mol. Plant Pathol. 2018, 19, 7–20. [Google Scholar] [CrossRef] [Green Version]
  8. Aguayo, J.; Fourrier-Jeandel, C.; Husson, C.; Loos, R. Assessment of Passive Traps Combined with High-Throughput. Appl. Environ. Microbiol. 2018, 84, e02637-17. [Google Scholar] [CrossRef] [Green Version]
  9. West, J.S.; Kimber, R.B.E. Innovations in air sampling to detect plant pathogens. Ann. Appl. Biol. 2015, 166, 4–17. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  10. Lione, G.; Giordano, L.; Turina, M.; Gonthier, P. Hail-induced infections of the chestnut blight pathogen Cryphonectria parasitica depend on wound size and may lead to severe diebacks. Phytopathology 2020, 110, 1280–1293. [Google Scholar] [CrossRef]
  11. Bryner, S.F.; Prospero, S.; Rigling, D. Dynamics of Cryphonectria hypovirus infection in chestnut blight cankers. Phytopathology 2014, 104, 918–925. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  12. Pažitný, J.; Bolvanský, M.; Adamčíková, K. Screening for resistance of progenies derived from Castanea sativa × C. crenata and C. crenata to Cryphonectria parasitica. For. Pathol. 2018, 48, e12439. [Google Scholar] [CrossRef]
  13. Cipollini, M.L.; Moss, J.P.; Walker, W.; Bailey, N.; Foster, C.; Reece, H.; Jennings, C. Evaluation of an Alternative Small Stem Assay for Blight Resistance in American, Chinese, and Hybrid Chestnuts (Castanea spp.). Plant Dis. 2021, 105, 576–584. [Google Scholar] [CrossRef] [PubMed]
  14. Aguín, O.; Sainz, M.; Montenegro, D.; Mansilla, J.P. Biodiversidad e hipovirulencia de Cryphonectria parasitica en Europa: Implicaciones para el control biológico del cancro del castaño. Recur. Rurais 2011, 7, 35–47. [Google Scholar] [CrossRef]
  15. Aghayeva, D.N.; Rigling, D.; Prospero, S. Low genetic diversity but frequent sexual reproduction of the chestnut blight fungus Cryphonectria parasitica in Azerbaijan. For. Pathol. 2017, 47, 1–7. [Google Scholar] [CrossRef]
  16. Cortesi, P.; Milgroom, M.G. Genetics of vegetative incompatibility in Cryphonectria parasitica. Appl. Environ. Microbiol. 1998, 64, 2988–2994. [Google Scholar] [CrossRef] [Green Version]
  17. Muñoz-Adalia, E.J.; Fernández, M.M.; Diez, J.J. The use of mycoviruses in the control of forest diseases. Biocontrol Sci. Technol. 2016, 26, 577–604. [Google Scholar] [CrossRef]
  18. Xie, J.; Jiang, D. New insights into mycoviruses and exploration for the biological control of crop fungal diseases. Annu. Rev. Phytopathol. 2014, 52, 45–68. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  19. Agnagnostakis, S.L. Chestnut Breeding in the United States for Disease and Insect Resistance. Plant Dis. 2012, 96, 1392–1403. [Google Scholar] [CrossRef]
  20. Clark, S.L.; Schlarbaum, S.E.; Saxton, A.M.; Baird, R. Eight-year blight (Cryphonectria parasitica) resistance of backcross-generation American chestnuts (Castanea dentata) planted in the southeastern United States. For. Ecol. Manag. 2019, 433, 153–161. [Google Scholar] [CrossRef]
  21. Griffin, G.; Hebard, F.; Wendt, R.; Elkins, J. Survival of American Chestnut Trees: Evaluation of Blight Resistance and virulence of Endothia parasitica. Phytopathology 1983, 73, 1084–1092. [Google Scholar] [CrossRef]
  22. Guàrdia, M.; Charrier, G.; Vilanova, A.; Savé, R.; Ameglio, T.; Aletà, N. Genetics of frost hardiness in Juglans regia L. and relationship with growth and phenology. Tree Genet. Genomes 2016, 12, 83. [Google Scholar] [CrossRef]
  23. Fernández-López, F.; Díaz-Vázquez, R.; Vázquez-Ruiz, R.; Pereira-Lorenzo, S. Evaluation of resistance of Castanea sp. clones to Phytophthora sp. using excised chestnut shoots. For. Snow Landsc. Res. 2001, 76, 451–454. [Google Scholar]
  24. Rodríguez, J.; Colinas, C. Resistance test for chestnut against Cryphonectria (Endothia) parasitica. In Proceedings of the 2nd International Symposium on Chestnut, Bordeaux, France, 19–23 October 1998; p. 494. [Google Scholar]
  25. R Rcore Team. R Core Team R: A Language and Environment for Statistical Computing; R Foundation for Statistical Computing: Vienna, Austria, 2021. [Google Scholar]
  26. Bates, D.; Mächler, M.; Bolker, B.; Walker, S. Fitting Linear Mixed-Effects Models Using lme4. J. Stat. Softw. 2015, 67, 1–48. [Google Scholar] [CrossRef]
  27. Anderson, D. Model Based Inference in the Life Sciences: A Primer on Evidence; Springer: New York, NY, USA, 2008. [Google Scholar]
  28. Mazerolle, M.J. AICcmodavg: Model Selection and Multimodel Inference Based on (Q)AIC(c), R Package Version 2.2-1; 2019. Available online: https://rdrr.io/cran/AICcmodavg/ (accessed on 11 October 2021).
  29. Lee, J.K.; Tattar, T.A.; Berman, P.M.; Mount, M.S. A Rapid Method for Testing the Virulence of Cryphonectria parasitica Using Excised Bark and Wood of American Chestnut. Phytopathology 1992, 82, 1454–1456. [Google Scholar] [CrossRef]
  30. Dennert, F.; Rigling, D.; Meyer, J.B.; Schefer, C.; Augustiny, E.; Prospero, S. Testing the Pathogenic Potential of Cryphonectria parasitica and Related Species on Three Common European Fagaceae. Front. For. Glob. Chang. 2020, 3, 52. [Google Scholar] [CrossRef]
  31. Peever, T.L.; Liu, Y.C.; Cortesi, P.; Milgroom, M.G. Variation in tolerance and virulence in the chestnut blight fungus-hypovirus interaction. Appl. Environ. Microbiol. 2000, 66, 4863–4869. [Google Scholar] [CrossRef] [Green Version]
  32. Colinas González, C.; Rojo Sanz, M.; Argemí Relat, J.; Heras Dolander, J.; Castaño Soler, C.; Rotllan Puig, X.; Gómez Gallego, M.; Gilarte Cayuela, S.; Ustrell Juan, E.; Sarr Torras, H. El control biológico del chancro del castaño en Cataluña. In Proceedings of the 5th Congreso Forestal Español, Montes y Sociedad: Saber qué Hacer, Avila, Spain, 21–25 September 2009. [Google Scholar]
  33. González, M.; Cuenca, B.; López, M.; Prado, M.; Rey, M. Molecular characterization of chestnut plants selected for putative resistance to Phytophtora cinnamomi using SSR markers. Sci. Hortic. 2011, 130, 459–467. [Google Scholar] [CrossRef]
  34. Cuenca-Valera, B.; González-González, M.; López-Rodríguez, M.; Ferradás-Rial, Y.; González- Simón, L.; Rey-Fraile, M. Hablan japonés los castaños gallegos? Introgresión genética en el castaño del país. In Proceedings of the 6th Congreso Forestal Español, Montes: Servicios y Desarrollo Rural, Vitoria-Gasteiz, Spain, 10–14 June 2013. [Google Scholar]
  35. Fernández-López, J.; Fernández-Cruz, J.; Míguez-Soto, B. The demographic history of Castanea sativa Mill. in southwest Europe: A natural population structure modified by translocations. Mol. Ecol. 2021, 30, 3930–3947. [Google Scholar] [CrossRef]
  36. Mattioni, C.; Martin, M.A.; Chiocchini, F.; Cherubini, M.; Gaudet, M.; Pollegioni, P.; Velichkov, I.; Jarman, R.; Chambers, F.M.; Paule, L.; et al. Landscape genetics structure of European sweet chestnut (Castanea sativa Mill): Indications for conservation priorities. Tree Genet. Genomes 2017, 13, 39. [Google Scholar] [CrossRef]
Agriculture 11 01158 g001 550
Figure 1. Outline of inoculation methodology. (a) inoculation protocol (1: field budsticks; 2: budstick sections; 3: debarked budsticks); (b) inoculated budsticks with C. parasitica and tray incubation; (c) detail of fungus growing in the debarked budstick.
Figure 1. Outline of inoculation methodology. (a) inoculation protocol (1: field budsticks; 2: budstick sections; 3: debarked budsticks); (b) inoculated budsticks with C. parasitica and tray incubation; (c) detail of fungus growing in the debarked budstick.
Agriculture 11 01158 g001
Agriculture 11 01158 g002 550
Figure 2. Values of mean daily lesion growth rate (G) of Cryphonectria parasitica on cuttings of Castanea sativa genotypes fitted by model Ma1 (Assay A). Mean values and 95% confidence error are shown. Asterisk (*) denotes significant differences in respect to Castanea mollissima (average value of “M2” and “M5M”). Number of repetitions: 8 (“M2” and “M5M”); 8 (all genotypes of C. sativa).
Figure 2. Values of mean daily lesion growth rate (G) of Cryphonectria parasitica on cuttings of Castanea sativa genotypes fitted by model Ma1 (Assay A). Mean values and 95% confidence error are shown. Asterisk (*) denotes significant differences in respect to Castanea mollissima (average value of “M2” and “M5M”). Number of repetitions: 8 (“M2” and “M5M”); 8 (all genotypes of C. sativa).
Agriculture 11 01158 g002
Agriculture 11 01158 g003 550
Figure 3. Values of mean daily lesion growth rate (G) of Cryphonectria parasitica on cuttings of Castanea sativa progenies fitted by model Mbp1 (Assay B). Mean values and 95% confidence error are shown. Asterisk (*) denotes significant differences in respect to Castanea mollissima (average value of “M2” and “M5M”). Number of repetitions: 4 (“M2” and “M5M”); see Table 1 for progenies of C. sativa.
Figure 3. Values of mean daily lesion growth rate (G) of Cryphonectria parasitica on cuttings of Castanea sativa progenies fitted by model Mbp1 (Assay B). Mean values and 95% confidence error are shown. Asterisk (*) denotes significant differences in respect to Castanea mollissima (average value of “M2” and “M5M”). Number of repetitions: 4 (“M2” and “M5M”); see Table 1 for progenies of C. sativa.
Agriculture 11 01158 g003
Table
Table 1. Description of genotypes used in the in vitro assays (A and B). G: mean daily growth of the Cryphonectria parasitica colony from inoculation to the sixth day of incubation along the longitudinal axis. Data for genotypes of Castanea mollissima in assay A and B are shown in the left and right sides of the bar (/) respectively. n: number of replicates per genotype. Mean values and standard deviation are shown.
Table 1. Description of genotypes used in the in vitro assays (A and B). G: mean daily growth of the Cryphonectria parasitica colony from inoculation to the sixth day of incubation along the longitudinal axis. Data for genotypes of Castanea mollissima in assay A and B are shown in the left and right sides of the bar (/) respectively. n: number of replicates per genotype. Mean values and standard deviation are shown.
SpeciesAssayGenotype/ProgenyN GenotypesnCutting Diameter (cm)G (mm/Day)
Castanea mollissimaA/BM218/40.84 ± 0.06/0.79 ± 0.143.9 ± 1.2/5.8 ± 2.5
M5M18/41.14 ± 0.05/0.60 ± 0.064.6 ± 1.4/4.6 ± 2.7
Castanea sativaABRL02181.41 ± 0.085.6 ± 1.2
ABRL03180.90 ± 0.034.4 ± 1.7
ABRL06180.98 ± 0.134.9 ± 1.4
ACNV01180.91 ± 0.075.8 ± 1.2
AFGM01181.21 ± 0.085.4 ± 1.5
AFGM04181.43 ± 0.155.4 ± 1.8
AFGM11180.93 ± 0.106 ± 1.2
AMSY03180.95 ± 0.065.8 ± 1.7
AMSY04180.89 ± 0.095.4 ± 0.9
AMSY08180.91 ± 0.045.6 ± 2.2
AMSY09180.93 ± 0.085.3 ± 1.8
AMSY10181.06 ± 0.083.9 ± 1.6
AMSY11181.08 ± 0.045.5 ± 0.7
AMSY12181.16 ± 0.385.9 ± 0.8
AMSY14181.24 ± 0.124.3 ± 2.0
APO11181.13 ± 0.134.5 ± 1.3
ARLL05181.03 ± 0.124.5 ± 1.1
ARLL10180.96 ± 0.206.4 ± 1.1
ASPV05181.35 ± 0.065.9 ± 1.8
AVLD12181.01 ± 0.085.7 ± 1.1
AVLD14181.23 ± 0.105.4 ± 1.3
AVLD22181.14 ± 0.055 ± 0.9
AVLD29181.11 ± 0.045.4 ± 1.0
AVLD30181.19 ± 0.104.4 ± 1.7
AVLD31181.00 ± 0.116 ± 1.5
AVLD32181.02 ± 0.045.3 ± 1.6
AVLD33181.11 ± 0.066.1 ± 1.3
AVLD34181.21 ± 0.115.5 ± 1.9
BCS-492340.71 ± 0.136.7 ± 1.7
BCS-511440.69 ± 0.116.9 ± 1.8
BCS-521240.69 ± 0.118 ± 1.4
BCS-541740.71 ± 0.096.9 ± 1.5
BCS-552440.69 ± 0.117.7 ± 2.1
BCS-561440.69 ± 0.128.1 ± 2
BCS-57540.72 ± 0.148.8 ± 1.3
BCS-602040.68 ± 0.117.1 ± 2.1
BCS-611840.66 ± 0.127.2 ± 1.8
BCS-621840.69 ± 0.096.9 ± 2
Table
Table 2. Results of models (GLMs and LMMs) describing the variation of mean daily lesion growth (G; mm/day) with progeny, seedling-genotype, and diameter (d; mm) in the in vitro assay. Random factors of LMMs are shown in brackets. n: number of observations used for model fitting. Selected model in bold.
Table 2. Results of models (GLMs and LMMs) describing the variation of mean daily lesion growth (G; mm/day) with progeny, seedling-genotype, and diameter (d; mm) in the in vitro assay. Random factors of LMMs are shown in brackets. n: number of observations used for model fitting. Selected model in bold.
AssayModelnDescriptionDflogLikDevianceAICΔAIC
AMa0232G~genotype30−403.73807.46867.460
Ma1G~genotype + (d)31−403.73807.46869.462
BMbp0664G~progeny12−13552710 27340
Mbp1G~progeny + (d)13−1351.802703.702729.70−4.30
Mbg0G~seedling-genotype167−1161.402322.702656.700
Mbg1G~seedling-genotype + (d)168−1161.402322.702658.702
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Muñoz-Adalia, E.J.; Meijer, A.; Abel, J.; Colinas, C.; Aletà, N.; Guàrdia, M. Evaluation of Chestnut Susceptibility to Cryphonectria parasitica: Screening under Controlled Conditions. Agriculture 2021, 11, 1158. https://doi.org/10.3390/agriculture11111158

AMA Style

Muñoz-Adalia EJ, Meijer A, Abel J, Colinas C, Aletà N, Guàrdia M. Evaluation of Chestnut Susceptibility to Cryphonectria parasitica: Screening under Controlled Conditions. Agriculture. 2021; 11(11):1158. https://doi.org/10.3390/agriculture11111158

Chicago/Turabian Style

Muñoz-Adalia, Emigdio Jordán, Andreu Meijer, Joan Abel, Carlos Colinas, Neus Aletà, and Mercè Guàrdia. 2021. "Evaluation of Chestnut Susceptibility to Cryphonectria parasitica: Screening under Controlled Conditions" Agriculture 11, no. 11: 1158. https://doi.org/10.3390/agriculture11111158

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop