The Spanish elm programme began in 1986 in response to the devastating impact of Dutch elm disease on natural elm stands and urban trees. Its main objectives were to conserve remaining genetic resources and select and breed tolerant native elm genotypes. After 27 years of work conducting susceptibility trials on thousands of elm genotypes, the first seven tolerant
In the first half of the 20th century, the first Dutch elm disease (DED) pandemic caused a massive loss of elms in Europe and North America. The much more aggressive
The first elm breeding programme began in the Netherlands in 1928 (
In the 1990s the Spanish programme included some native elms, mainly
Progress in selecting elms was slow due to the long periods required to propagate trees and evaluate their tolerance with a scientifically sound base, as plant material needs to be at least four years old (
Native elms can be registered by the Environmental Administration as “qualified forest reproductive material” when they show low (0-30%) crown wilting or symptoms similar to the tolerant “Sapporo Autumn Gold” clone after two consecutive years of artificial inoculation with
From 1990 to 2002, plant material was propagated from trees selected during surveys of adult elms in natural forests, rural areas, parks, and other urban environments in Spain (
Ramets of the seven clones were planted with 157 other elm clones in five different inoculation plots in Spain (
Plots were designed in two blocks, with random experimental units of three to four ramets per block. Spacing in Puerta de Hierro and El Serranillo was 0.5 to 1 m between plants and 1 to 1.5 m between rows. In Calabazanos spacing was 5 × 5 m. To avoid side effects, a tree border line surrounded all plots. Plants were watered in spring and summer to ensure growth. Their main stems were fastened to supports to avoid wind shake. “Sapporo Autumn Gold”, highly tolerant to
Local strains of
A bud-cell suspension of the pathogen was prepared by adding 2 × 2 mm plugs from the edge of 7-day-old cultures on malt extract agar to 50 ml Tchernoff’s liquid medium (
In the marketing of forest reproductive material, characterization and traceability of trees are of pivotal importance. Various techniques using molecular markers are efficient tools for this purpose. Genetic characterization of the seven
For the lineage study, two chloroplast markers were used. One corresponds to the chloroplast fragment SFm and was developed from the sequence of the SFm fragment to differentiate the
For the genetic description, 12 nuclear microsatellites were selected. Four of these were described in
Leaves from at least two individuals for each selected clone were collected, labeled and stored in silica gel. After DNA extraction, 2.5 ng µl-1 dilutions were used in amplification reactions conducted following the literature (
Clones were morphologically described following specific literature (
Height growth of each clone was assessed in Puerta de Hierro Forest Breeding Center (Madrid), as well as ornamental qualities of trees such as growth habit and branching (erect, spreading, or pendulous), leaf density (abundance of leaves per crown volume, estimated as high, medium or low), crown shape (conical, spindle, globular or irregular) and leaf size. The ornamental value of each clone was quantified on a scale from 1 to 5, where 5 corresponded to the most frequent features of Spanish
The unfolding of elm leaf buds was characterized in 2011 using the methodology described by
For each inoculation plot and year, wilting percentages at 30, 60 and 120 dpi were analyzed using repeated measures ANOVA, considering time since inoculation, block, and genotype as the main factors and tree height as a covariate. Fisher’s least significant difference (LSD) test was applied to compare average wilting values (least square means of wilting percentages at 30, 60 and 120 dpi) between clones (
Seven
Six clones were tested at one location (Madrid), but the “Fuente Umbría” clone was tested over four consecutive years in Guadalajara and Palencia (
Environmental conditions can strongly influence elm susceptibility to
DED fungi have caused mortality not only of large elm groves of
The results of the genetic characterization of the seven clones are shown in
The Spanish programme aimed to directly control DED (
To broaden the genetic base of tolerant native elms, the Spanish programme has grown 1400 seedlings from controlled F1 crossings between the seven
Although tree selection, trial establishment and breeding cycles require a major investment in time and effort, breeding programmes are the most reliable option for recovery of native elm populations. Results reported here show that selection of tolerant native
This paper is dedicated to Margarita Burón, who spent the last ten years of her life working on the conservation and breeding of Spanish elms.
The authors are grateful to everyone who has participated in the Spanish elm breeding programme during its 27 years of existence. We would also like to express our gratitude to the Dutch and Italian elm breeding programmes and the participants in the EU Project RESGEN CT96-78 for sharing their knowledge and materials. The programme is funded by the Spanish Directorate-General of Rural Development and Forestry Policy (
Foliar parameters measured to describe the
Susceptibility of the seven
Registered
Registered “Fuente Umbría” clone grown in the clonal bank of Puerta de Hierro Forest Breeding Center (Madrid). This
Leaf phenology in 2011 of the seven registered
Plant material specifications. (a) R: root cutting; G: graft; S: seed; (b) numbers in brackets indicate the year of inoculation.
Clone | Originin Spain | Initial propagation | Inoculation test | |||||
---|---|---|---|---|---|---|---|---|
Typea | Year | Plot | N | Years | Locationin Spain | |||
Ademuz | Valencia 40° 4′ 52″ N, 1° 16′ 55″ W | R | 1996 | XXIV | 10 | 2008,2009 | Puerta de Hierro | NA-PE (2008),CU-HU (2009) |
Dehesa de la Villa | Madrid 40° 27′ 29″ N, 3° 44′ W | R | 1990 | XXV | 10 | 2009,2010 | Puerta de Hierro | CU-HU |
Majadahonda | Madrid 40° 28′ 90″ N, 3° 52′ 19″ W | G | 1993 | XXIV | 6 | 2008,2009 | Puerta de Hierro | NA-PE (2008),CU-HU (2009) |
Toledo | Toledo 39° 51′ 51″ N, 4° 1′ 30″ W | S | 1999 | XXX | 7 | 2011,2012 | Puerta de Hierro | Z-BU1 |
Dehesa de Amaniel | Madrid 40° 27′ 37″ N, 3° 43′ 17″ W | S | 1999 | XXX | 12 | 2011,2012 | Puerta de Hierro | Z-BU1 |
Retiro | Madrid 40° 24′ 56″ N, 3° 41′ 10″ W | S | 2002 | XXX | 7 | 2011,2012 | Puerta de Hierro | Z-BU1 |
Fuente Umbría | Valencia 39° 25′ 23″ N, 0° 56′ 46″ W | S | 1995 | V and A | >10 | 2010-2013 | El Serranillo andCalabazanos | CU-HU (2010),Z-BU1 (2011-2013) |
Results (p-values) of repeated measures ANOVA of the wilting values shown by elm trees at 30, 60 and 120 dpi (repeated variable) considering time since inoculation, genotype, block, and genotype x block interaction as factors, and plant height as a covariate. (a): The plot had one block.
Inoculation year | Plot | Source of variation | ||||
---|---|---|---|---|---|---|
Time dpi | Genotype (G) | Block (B) | G × B | Plant height | ||
2008 | XXIV | < 0.001 | < 0.001 | < 0.001 | 0.001 | 0.708 |
2009 | XXIV | 0.15 | < 0.001 | 0.029 | 0.65 | 0.928 |
2009 | XXV | 0.002 | < 0.001 | 0.032 | < 0.001 | 0.078 |
2010 | XXV | < 0.001 | < 0.001 | 0.064 | < 0.001 | 0.513 |
2010 | V | < 0.001 | < 0.001 | 0.163 | 0.751 | 0.951 |
2011 | V | < 0.001 | < 0.001 | 0.185 | 0.572 | 0.839 |
2011 | XXX | < 0.001 | < 0.001 | 0.001 | < 0.001 | < 0.001 |
2012 | XXX | < 0.001 | < 0.001 | 0.068 | 0.01 | 0.91 |
2012 | Aa | < 0.001 | < 0.001 | - | - | 0.623 |
2013 | Aa | 0.001 | < 0.001 | - | - | 0.086 |
Morphological description of the seven
Feature | Clone | ||||||
---|---|---|---|---|---|---|---|
Ademuz | Dehesa de la Villa | Majadahonda | Toledo | Dehesa de Amaniel | Retiro | Fuente Umbría | |
Height growth in Puerta de Hierro, Madrid(cm year-1) | 100.0 | 63.0 | 60.8 | 89.3 | 90.0 | 70.5 | 51.7 |
Petiole length (mm) | 5.2(4.2-6.2) | 6.3(3.5-10.0) | 11.0(10-12.7) | 5.8(4.6-7.4) | 2.6(1.9-3.4) | 7.3(6.1-8.0) | 10.2(8.3-12.2) |
Leaf basal asymmetry (mm) | 1.6(1.1-1.9) | 2.9(2.0-3.8) | 3.8(2.0-4.8) | 1.3(0.7-2.4) | 1.3(0.8-1.8) | 1.2(0.3-1.7) | 3.1(2.2-4.2) |
Maximum foliar length (mm) | 53.7(43.5-65.1) | 55.4(44.0-70.0) | 50.4(46.7-53.6) | 47.0(35.6-71.5) | 38.5(36.2-39.6) | 71.4(63.9-79.4) | 75.9(69.8-85.9) |
Maximum foliar width (mm) | 33.8(30.0-38.6) | 35.6(28.0-45.0) | 28.8(25.5-30.6) | 26.6(19.6-35.2) | 29.7(27.3-33.0) | 42.2(36.4-48.9) | 44.9(39.3-48.7) |
Tooth breadth (mm) | 2.2(1.9-2.6) | 3.5(3.0-4.9) | 1.1(0.8-1.3) | 4.1(3.6-4.7) | 2.2(1.2-2.6) | 2.8(2.5-3.7) | 2.8(1.9-3.8) |
Tooth depth (mm) | 3.4(2.8-4.0) | 3.7(2.4-5.0) | 1.8(1.4-2.3) | 3.2(2.6-4.4) | 2.7(2.1-3.5) | 2.2(1.4-2.8) | 2.3(2.1-2.6) |
Tooth length (mm) | 4.2(3.8-4.4) | 4.6(4.0-5.1) | 2.3(1.9-2.9) | 5.4(4.8-5.9) | 3.5(2.8-4.2) | 2.9(2.1-3.8) | 3.0(2.2-4.1) |
Teeth per leaf (N) | 38(35-42) | 44(30-64) | 54(52-57) | 30.0(28-33) | 33(31-35) | 49(45-53) | 32(28-34) |
Pairs of secondary nerves (N) | 10(9-11) | 10(9-12) | 12(11-12) | 8.8(8-10) | 9.3(8-10) | 13(12-15) | 12(11-13) |
Leaf serration | Double | Double | Simple | Double | Triple | Double | Double |
Presence of corky tissue | No | No | No | No | Yes | No | Yes |
Foliar density | Medium | High | High | Medium | High | High | Medium |
Branching | Erect | Erect | Erect | Erect | Spreading | Erect | Erect |
Crown shape | Spindle | Spindle | Globular | Irregular | Irregular | Globular | Irregular |
Ornamental valuea | 4.5 | 4.1 | 4.3 | 2.9 | 3.0 | 4.0 | 3 |
Genetic characterisation of the seven
Organelle | DNAmarker | Clone | |||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Ademuz | Dehesa de la Villa | Majadahonda | Toledo | Dehesa de Amaniel | Retiro | FuenteUmbría | |||||||||
Chloro-plast | ccmp2 | 237 | 236 | 236 | 215 | 236 | 216 | 215 | |||||||
SFm | 278 | 278 | 278 | 297 | 278 | 297 | 297 | ||||||||
Nuclear | Ulm 2 | 102 | 108 | 102 | 102 | 106 | 108 | 102 | 102 | 106 | 108 | 108 | 108 | 102 | 102 |
Ulm 3 | 161 | 176 | 176 | 176 | 176 | 180 | 161 | 180 | 176 | 176 | 176 | 180 | 161 | 161 | |
Ulm 8 | 196 | 196 | 196 | 196 | 194 | 196 | 194 | 198 | 196 | 196 | 196 | 196 | 194 | 194 | |
UR 123 | 250 | 250 | 252 | 254 | 250 | 254 | 242 | 250 | 250 | 254 | 252 | 254 | 255 | 259 | |
UR 141 | 150 | 152 | 152 | 160 | 152 | 158 | 152 | 152 | 158 | 158 | 158 | 158 | 152 | 158 | |
UR 153 | 178 | 190 | 184 | 188 | 188 | 188 | 188 | 188 | 178 | 188 | 186 | 188 | 178 | 190 | |
UR 158 | 195 | 195 | 179 | 195 | 195 | 195 | 179 | 179 | 179 | 199 | 179 | 179 | 179 | 179 | |
UR 159 | 258 | 260 | 260 | 278 | 278 | 278 | 260 | 278 | 278 | 280 | 278 | 278 | 258 | 258 | |
Ulm 1-98 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 151 | 154 | |
Ulm 1-165 | 204 | 204 | 146 | 146 | 164 | 164 | 130 | 148 | 204 | 204 | 156 | 156 | 160 | 160 | |
Ulm 2-16 | 90 | 90 | 90 | 90 | 90 | 90 | - | - | 90 | 90 | 82 | 94 | 90 | 90 | |
Ulm 2-20 | -a | - | 184 | 202 | 172 | 206 | 206 | 220 | 186 | 202 | 180 | 184 | 220 | 220 |