Crop Science 41:1427-1434 (2001)
© 2001 Crop Science Society of America
CROP BREEDING, GENETICS & CYTOLOGY
Glandular Morphology from a Perennial Alfalfa Clone Resistant to the Potato Leafhopper
Christopher M. Ranger*,a and
Arthur A. Howerb
a 1–87 Agriculture Building, Dep. of Entomology, Univ. of Missouri, Columbia, MO, 65211
b 501 Agricultural Sciences and Industries Building, Dep. of Entomology, Penn. State Univ., University Park, PA 16802-3508
* Corresponding author (cmr0b2{at}mizzou.edu)
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ABSTRACT
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The commercial release of several cultivars of perennial, glandular-haired alfalfa (Medicago sativa L.) for control of the potato leafhopper, Empoasca fabae (Harris), has increased the need to identify the causal mechanism of resistance. The objectives of this experiment were to examine the morphology and exudate of the glandular trichomes found on the perennial alfalfa clone FGplh13, in addition to comparing their density and distribution. Light, scanning, and transmission electron microscopy were utilized to characterize the morphology of the glandular trichomes and their associated exudate. Two distinct trichome morphologies, erect and procumbent glandular trichomes, occurred. The erect glandular trichomes consisted of multicellular stalks (5–11 cells) and gland heads, with the cells being arranged in distinct tiers. The procumbent glandular trichomes were composed of a stalk (1–2 cells) bent almost parallel to the plant surface. The procumbent gland head contained 8 to 12 cells, which were distinctly arranged in two to three tiers. Both types of trichomes released exudates that appeared to accumulate in a subcuticular space within a distal extrusion. Exudate from the procumbent glandular trichome became attached to the tibia of a potato leafhopper nymph. Entrapment of first instar potato leafhoppers by the glandular trichomes was also observed for the first time on a perennial glandular-haired alfalfa clone. The erect glandular trichome was the most dense morphology on the stem, petiole, and leaf midvein.
Abbreviations: ebc, enlarged basal cell • SEM, scanning electron microscopy • TEM, transmission electron microscopy
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INTRODUCTION
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IN THE MIDWESTERN AND EASTERN UNITED STATES, the potato leafhopper is considered one of the most serious pests of alfalfa. A considerable amount of research has been conducted to develop potato leafhopper-resistant cultivars of this legume. Scientists initally examined the resistance capabilities of dense simple plant hairs (Granovsky, 1928; Taylor, 1956). Glandular-haired alfalfa species have also been examined for potato leafhopper resistance. While Shade et al. (1979) identified several annual glandular Medicago spp. with potato leafhopper resistance, the inability to hybridize annual Medicago spp. with the susceptible perennial hay-type M. sativa excluded their use in the development of insect-resistant germplasms (Kreitner and Sorensen, 1979a). As a result, attention shifted to perennial glandular-haired Medicago spp.
Using light and electron microscopy, Kreitner and Sorensen (1979a) documented the morphology of the erect and procumbent glandular trichomes found on several annual [M. disciformis DC., M. scutellata (L.) Mill.] and perennial [M. prostrata Jacq.; M. sativa subsp. praefalcata (Sinskaya) C.R. Gunn, syn. M. sativa nothosubsp. varia (Martyn) Arcang.; M. sativa subsp. sativa] Medicago spp. The secretory system of the erect and procumbent glandular trichomes from these annual and perennial Medicago spp. was also examined (Kreitner and Sorensen, 1979b). Both the erect and procumbent glandular forms excreted an exudate. Subcuticular spaces were also noted, whereby an exudate was localized between the secretory cell wall and the cuticle of the gland head. Kreitner and Sorensen (1981) indicated that the secreting capacity of M. prostrata and M. sativa subsp. praefalcata were lower than that of the annual species M. scutellata.
The erect glandular trichomes are considered to play the dominant role in providing resistance to the potato leafhopper (Shade et al., 1975; Kreitner and Sorensen, 1981; Othman et al., 1981; Brewer et al., 1986), based on the fact that the susceptible perennial M. sativa only possesses the procumbent glandular trichome. Furthermore, resistance of annual Medicago spp. is associated, in part, with entrapment of potato leafhopper nymphs by an exudate thought to be produced by the erect glandular trichome (Shade et al., 1979). The procumbent glandular trichome on alfalfa has not been considered to provide host resistance, and has been described as producing only small amounts of secretion, which tends to become dry and hard (Kreitner and Sorensen, 1979b, 1981).
Brewer et al. (1986) found the perennial glandular-haired M. falcata L. var. glandulosa David, M. prostrata, and M. glutinosa M. Bieb. [syn. M. sativa subsp. glomerata (Balb.) Rouy] to express high levels of resistance to the potato leafhopper. Along with M. sativa subsp. praefalcata, these species were utilized to develop three glandular-haired alfalfa germplasms: KS108GH5 (Sorensen et al., 1985), KS94GH6 (Sorensen et al., 1986), and 81IND-2 (Shade and Kitch, 1986). These germplasms were incorporated into an extensive breeding and selection program, with resistant varieties becoming commercially available in 1997.
Elden and McCaslin (1997) failed to find any evidence in the literature of potato leafhoppers being entrapped in the glandular trichomes by perennial Medicago spp., nor was it reported in their research. In addition, their research did not note an exudate associated with the glandular trichomes from 19 alfalfa clones. However, using the perennial glandular-haired alfalfa clone FGplh13, Ranger (1999) observed the entrapment of first instar potato leafhoppers by the glandular trichomes. This marked the first documentation of potato leafhoppers being entrapped by perennial glandular-haired alfalfa.
The resistant glandular-haired alfalfa clone FGplh13 was developed through a backcrossing program with contributions from the three aforementioned glandular-haired germplasms released in the mid-1980s (M. McCaslin, 1999, personal communication). Because of the apparent ability of FGplh13 alfalfa to entrap first instar potato leafhoppers, and its high level of resistance to later developmental stages (Ranger, 1999), it was of interest to use this clone in an attempt to understand further the causal mechanism of resistance of perennial glandular-haired alfalfa. The objectives of this experiment were to (i) examine the morphology of the erect and procumbent glandular trichomes found on the perennial glandular-haired alfalfa clone FGplh13; (ii) examine the exudate associated with the glandular trichomes; and (iii) compare the density and distribution of the erect and procumbent forms.
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MATERIALS AND METHODS
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Plant Material
The resistant glandular-haired alfalfa clone FGplh13 was developed by Forage Genetics in West Salem, WI. To achieve genetic stability in the plant material used in our experiments, cuttings were propagated from three FGplh13 alfalfa plants. Actively growing shoots 
2.5 cm long were clipped from a resistant plant, 1.2 cm of the stem was inserted into a plastic tray containing moistened supercoarse perlite, and the cuttings were transferred to a mist bench in a greenhouse. About 3 wk later, the propagules were potted in terra cotta pots (16-cm diam.) containing growing medium. A modification of Hoagland's nutrient solution (Hoagland and Arnon, 1950) was applied for 2 wk following transfer of the cuttings from the perlite medium to the pots. Osmocote 14–14–14 (Scotts-Sierra Horticultural Products, Marysville, OH) was used to fertilize each pot. Plants were watered every 2 d. The propagules were transferred after 2 wk from the greenhouse to a controlled environment chamber programmed at 24 to 28°C, a photoperiod of 16 h light:8 h dark, and a relative humidity of 55 ± 5%. This process was repeated three times before using the plant material in microscopy experiments.
The susceptible nonglandular plant material Pioneer 5373 was obtained by placing 4 to 5 inoculated seeds in plastic cone containers (4-cm diam.) with growing medium. After growing 10 to 12 cm in height, the plants were transferred to terra cotta pots (16-cm diam.) and placed in a controlled environment chamber programmed to the previously described specifications.
Plant material selected for light and electron microscopy was obtained from the second to last most distal internode and trifoliates of freshly excised alfalfa stems. Floral organs were also selected from the apical meristem regions for analysis.
Insects
To document the interaction between the potato leafhopper and the glandular trichomes found on FGplh13 alfalfa, 100 field-collected adults were collected at the Russell E. Larson Experimental Station at Rock Springs, PA, using an aspirator and a 125-mL Erlenmeyer flask. Afterwards, the flask was placed in the center of a 38- by 38- by 38-cm screened cage containing two resistant FGplh13 and two susceptible Pioneer 5373 alfalfa plants. Cages were then placed in the aforementioned environmental growth chamber, and the leafhoppers were allowed to disperse and feed for 13 d. Afterwards, FGplh13 stems with entrapped potato leafhopper nymphs were collected and immediately analyzed via scanning electron microscopy (SEM).
Light and Electron Microscopy
Plant samples prepared for light microscopy were subjected to a conventional fixation (Franke et al., 1969), while Karnovsky's fixation (Karnovsky, 1965) was used for transmission electron microscopy (TEM). Specimens were then placed in acetone, infiltrated with Spurr low-viscocity resin (Spurr, 1969) and allowed to polymerize for 72 h at 60°C. An ultramicrotome was used to slice 0.5-µm sections for light microscopy, and 0.06- to 0.07-µm sections for TEM. Sections prepared for light microscopy were stained with 1% toulidine blue in 1% sodium borate.
Plant and insect samples were viewed using cryogenic SEM. Samples were attached with a mix of carbon powder and a water soluble mix of glycols and resin to a polished brass stub, after which they were transferred to the preparation chamber under vacuum, gaseous nitrogen, and lowered temperatures. Specimens were then moved to the SEM chamber and etched by heating the stage to -90°C for 10 min., followed by returning the sample to the preparation chamber for sputter-coating with gold for 30 sec. Images were recorded onto Polaroid film (Polaroid Corporation, Cambridge, MA), with the provided photos being selected from ten individual plant samples. Using Princeton Gamma-Tech IMIX image collection and analysis software (Princeton Gamma-Tech, Princeton, NJ), measurements of glandular trichome length were made from the base of the stalk to the tip of the gland head, while the diameter of the gland head was measured at the midsection of the gland head. Trichome length measurements were taken from a total of 22 erect and procumbent glands, while gland diameter measurements were recorded from 15 erect and procumbent glands. Recordings were made from the second to last most distal internode, and were representative of five stems collected from individual FGplh13 alfalfa plants.
Trichome Density
Erect and procumbent glandular trichomes, along with nonglandular trichomes, were counted on the stem, petiole, leaf midvein, and leaf perimeter of the resistant alfalfa clone FGplh13. Using a stereomicroscope, all recordings were made from the fifth internode and center leaf of the fifth trifoliate. Using a technique described by Elden and McCaslin (1997), a 2-mm section of stem, petiole, leaf midvein, and leaf perimeter was used to record trichome density. To minimize error, stem and petiole sections were split longitudinally. Counts from the leaf midvein and leaf perimeter were made from the middle section of the leaf. The leaf perimeter was characterized by the glandular and nonglandular trichomes extending outwards (horizontally) from the leaf edge. Plant structures from which trichome densities were recorded were selected from 4- to 5-mo old resistant plants in the bud stage. A total of 25 stems and trifoliates were analyzed.
Data Analysis
Differences in glandular trichome morphology were examined using one-way ANOVA, while differences in trichome density were detected by one-way ANOVA with the means separated by Tukey's pairwise comparisons (Minitab, 1991).
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RESULTS AND DISCUSSION
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Glandular Trichome Morphology
Nonglandular and two distinct types of capitate glandular trichomes, erect and procumbent, were found on the vegetative and floral structures of the perennial alfalfa clone FGplh13. These results coincide with those of Wilson (1913) and Kreitner and Sorensen (1979a)( 1979b) in terms of the morphology of each trichome type and its associated exudate.
The erect glandular trichomes on FGplh13 alfalfa consisted of a mulitcellular stalk, with an enlarged basal cell (ebc) arising from the plant epidermis (Fig. 1, 2). Stalks were observed to range from 5 to 11 cells. By comparison, Kreitner and Sorensen (1979a) noted that the erect stalk from the perennial diploid M. prostrata possessed 4 to 5 short cells. They did not report the number of stalk cells associated with the perennial tetraploid M. sativa subsp. praefalcata. On the stem of FGplh13 alfalfa, the length from the base of the stalk to the tip of the erect gland head ranged between 97.5 and 703.0 µm, with a mean (±S.E.) of 340.6 ± 34.1 µm. The length of the erect glandular trichomes on M. scutellata ranged between 150 to 200 µm (Kreitner and Sorensen, 1979a).
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Fig. 1. Cryogenic scanning electron microscopy (SEM) of erect glandular trichomes from the resistant alfalfa clone FGplh13 detailing the enlarged basal cell (ebc) (Bar = 50 µm; x200).
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Fig. 2. Light microscopy of a longitudinally sectioned erect glandular trichome, documenting the multicellular stalk found on the resistant alfalfa clone FGplh13 (Bar = 30 µm; x400).
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Overall, the shape of the erect gland head from FGplh13 was ovoid, and consisted of cells being arranged in distinct tiers (Fig. 3). The basal tiers contained few, large cells with smaller cells distally. It was difficult to determine the total number of cells in an entire erect gland head from tissue sections; although, seven to nine cells were typically observed within a section. The mean (±S.E.) diameter of the erect gland head located on the stem of the perennial FGplh13 alfalfa was 31.5 ± 1.4 µm. The mean diameters of erect gland heads recorded by Kreitner and Sorensen (1979a) from the perennial species M. sativa subsp. praefalcata and M. prostrata were 25 µm and 25 to 30 µm, respectively. Diameter recorded for the annual M. scutellata ranged from 25 to 40 µm. While the gland diameter of the erect glandular trichomes found on FGplh13 alfalfa corresponded more closely with M. prostrata than with M. sativa subsp. praefalcata, the overall morphology appeared to be more similar to that observed by Kreitner and Sorensen (1979a)( b) on M. sativa subsp. praefalcata, from which FGplh13 was derived in part (M. McCaslin, 2000, personal communication).
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Fig. 3. Transmission electron microscopy (TEM) of a longitudinally sectioned erect gland head from the resistant alfalfa clone FGplh13. Note distinct tiers of cells (Bar = 10 µm; x2,500).
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The stalk of the procumbent glandular trichome found on the resistant alfalfa clone FGplh13 typically consisted of a basal cell embedded in the epidermis (Fig. 4) and one to two additional stalk cells (Fig. 5). As described by Kreitner and Sorensen (1979a), the gland head of the procumbent trichome was usually adjacent to the plant surface, due to bending of the stalk cell(s) consistently towards the distal portion of the plant structure from which they arose.
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Fig. 4. Cryogenic scanning electron microscopy (SEM) of a procumbent glandular trichome, showing two distinct tiers of cells in the gland head, located on the resistant alfalfa clone FGplh13 (Bar = 10 µm; x1,000).
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Fig. 5. Procumbent glandular trichome longitudinal section, detailing two stalk cells and three tiers of gland head cells, on the resistant alfalfa clone FGplh13 (Bar = 25 µm; x430).
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Attached to the stalk was a multicellular gland head, with a mean (±S.E.) diameter of 24.8 ± 1.9 µm. The diameter of the procumbent gland head differed significantly from that of the erect gland head (mean of 31.5 ± 1.4 µm) on FGplh13 (F = 8.45; df = 1, 28; P = 0.007). A variety of cell combinations were detected in the gland head, which corresponds to the work of Kreitner and Sorensen (1979a). A typical combination was four cells in each of two tiers (Fig. 6, 7), although three tiers also were observed (Fig. 5). The length of the procumbent glandular trichome ranged between 21.5 to 124.0 µm, with a mean (±S.E.) of 79.9 ± 5.6 µm. This length differed significantly from the erect glandular trichomes (mean of 340.6 ± 34.1 µm) on FGplh13 (F = 56.77; df = 1, 42; P < 0.001).
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Fig. 6. Light micrograph of a cross section from a procumbent gland head on the resistant alfalfa clone FGplh13. The section, taken from the distal tier of cells, reveals four cells. (Bar = 25 µm; x430).
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Fig. 7. Transmission electron microscopy (TEM) of a longitudinally sectioned procumbent gland head on a resistant alfalfa clone FGplh13. Note difference in cell size between the basal and distal tiers (Bar = 2 µm; x5,000).
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The general morphology of the procumbent glandular trichome from FGplh13 alfalfa was similar to that observed by Kreitner and Sorensen (1979a)( b) on M. sativa subsp. sativa. Detailed comparisons of these trichomes on FGplh13 with those on perennial and annual species studied by Kreitner and Sorensen (1979a)(b) could not be made because comparative photos were not published.
Exudate Morphology
Electron microscopy revealed the presence of an exudate associated with the erect and procumbent glandular trichomes on the perennial Medicago clone FGplh13. This supports examinations by Kreitner and Sorensen (1979b) using the annual species M. disciformis and M. scutellata, along with the perennial species M. prostrata, M. sativa subsp. praefalcata, and M. sativa subsp. sativa.
Cuticular extrusions were typically observed distally on the surface of erect gland heads (Fig. 8). These formations are thought to be created by the localization of an exudate between the cuticle of the gland head and the adjacent cell wall, creating what is referred to as a subcuticular space (Kreitner and Sorensen, 1979b). Compared with the work of Kreitner and Sorensen (1979a)(b), the morphology of the extrusions on the glands of FGplh13 alfalfa more closely resembled those on the perennial tetraploid M. sativa subsp. praefalcata than on the perennial diploid M. prostrata, where they appeared to be smaller and more numerous.
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Fig. 8. Numerous cuticular boils associated with the distal portion of an erect gland head on the alfalfa clone FGplh13 (Bar = 10 µm; x1,000).
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Occasionally, what appeared to be exudate was observed on the erect glandular trichomes. However, copious amounts of an exudate did not appear to be released by the erect glandular trichomes on FGplh13, as Kreitner and Sorensen (1979a) witnessed with the annual species M. disciformis and M. scutellata, or the perennial species M. sativa subsp. praefalcata. Using TEM, the extracytoplasmic compartmentalization of an exudate in a subcuticular space was noted (Fig. 9). The exudate localized in the subcuticular space of the erect glandular trichome appeared very dense and crystalline in structure. However, it is not known if this appearance was an artifact of the embedding process. Similarities existed in exudate appearance between the erect glandular trichomes of the perennial clone FGplh13 and those of the annual species M. disciformis described by Kreitner and Sorensen (1979b).
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Fig. 9. Subcuticular space from an erect gland head located on the alfalfa clone FGplh13 observed using transmission electron microscopy (TEM). Note dense, crystalline exudate (ex), cell wall (cw), cuticle (c), and vacuole (v) (Bar = 200 nm; x20,000).
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Vacuoles were present in the secretory cells of the erect (Fig. 9) and procumbent glandular trichomes. Kreitner and Sorensen (1983) noted the accumulation of electron-dense material in the vacuoles of erect gland head cells of the annual species M. scutellata. Accumulation of the dense material is considered to be an initial indication of secretory activity. However, while the vacuoles are suspected to accumulate exudate, they are not necessarily the site of synthesis (Kreitner and Sorensen, 1983). It is thought that precursors of dense material are incorporated into the vacuoles followed by its transformation (Frey-Wyssling, 1972).
Cuticular extrusions observed distally on the heads of the procumbent glandular trichomes often appeared to consist of a single, large subcuticular space (Fig. 10). This is consistent with observations by Kreitner and Sorensen (1979a)(b) on the susceptible M. sativa subsp. sativa. As on the erect glandular trichomes, the size of the cuticular extrusions on the procumbent glandular trichomes ranged from quite small to instances where the cuticle enclosing the entire distal tier of the gland cells appeared to be engorged and separated from the adjoining cell walls. Release of an exudate from the procumbent gland head of the resistant alfalfa clone FGplh13 appeared to be affiliated with the distal tier of cells (Fig. 11, 15), as seen with M. sativa subsp. sativa (Kreitner and Sorensen, 1979b). The subcuticular space of the procumbent glandular trichome was not as definitive as that of the erect glandular trichome (Fig. 12). Abundant amounts of a viscous exudate were released by the procumbent glandular trichomes on FGplh13. Following release, the exudate appeared to disperse onto the epidermal surface (Fig. 11). The exudate did not appear as dense and crystalline as that of the exudate from the erect glandular trichome.
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Fig. 10. Cuticular boil on the distal portion of a procumbent gland head on the resistant alfalfa clone FGplh13, which potentially formed due to the presence of an exudate (Bar = 25 µm; x1,500).
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Fig. 11. Release of an exudate from a procumbent glandular trichome onto the epidermal surface of the resistant alfalfa clone FGplh13 (Bar = 25 µm; x1,000).
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Fig. 15. Cryogenic scanning electron microscopy (SEM) detailing the exudate (ex) from a procumbent (pr) glandular trichome on the alfalfa clone FGplh13 attached to the tibia (ti) of a potato leafhopper nymph (Bar = 25 µm; x200).
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Fig. 12. Transmission electron microscopy (TEM) recording the presence of an exudate from a procumbent glandular trichome to occupy a subcuticular space on the resistant alfalfa clone FGplh13. Note exudate (ex), cell wall (cw), and cuticle (c) (Bar = 250 nm; x30,000).
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Entrapment of Potato Leafhoppers
Entrapment of a first instar potato leafhopper by exudate from the glandular trichomes was verified on the perennial alfalfa clone FGplh13 (Fig. 13). The erect gland heads were affixed to various body parts of the nymphs following contact of the glandular trichomes with the tarsi and stylets (Fig. 14). Exudate from a procumbent gland was affixed in mid-air to the cuticle of a potato leafhopper nymph (Fig. 15). This suggests that while the exudate may dry quickly, as indicated by Kreitner and Sorensen (1979b)( 1981), it may also possess adhesive qualities.
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Fig. 13. Entrapment of a first instar potato leafhopper in the glandular trichomes found on the alfalfa clone FGplh13. Note size of erect (e) and procumbent (pr) glandular trichomes in relation to potato leafhopper nymph (Bar = 60 µm; x100).
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Fig. 14. Attachment of tarsi (ta) and stylets (st) of a first instar potato leafhopper following contact with the glandular trichomes on the alfalfa clone FGplh13. Note contact between the abdomen (ab) of the insect and an erect glandular trichome (Bar = 30 µm; x150).
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Perhaps the recurrent phenotypic selection progam used to develop the alfalfa clone FGplh13, which selected for the glandular trichome phenotype and the absence of potato leafhopper feeding damage, resulted in an increased amount of exudate production by the glandular trichomes. In fact, repeated selections of the perennial M. sativa subsp. praefalcata resulted in an increased amount of osmiophilic material in the vacuolar system of the erect gland head, which is an intermediate stage in secretion formation (Kreitner and Sorensen, 1981).
Trichome Density and Distribution
Significant differences in density of trichome types were detected from the stem (F = 100.88, df = 2, 72; P < 0.001), petiole (F = 141.13; df = 2, 72; P < 0.001), leaf midvein (F = 88.34; df = 2, 72; P < 0.001), and leaf perimeter (F = 72.93; df = 2, 72; P < 0.001) of FGplh13 (Table 1). In every case, the erect glandular trichomes were the most abundant. The highest mean (±S.E.) densities of erect (31.9 ± 1.5) and procumbent (26.5 ± 1.8) glandular trichomes were recorded from the stem, while the highest mean (±S.E.) density of nonglandular trichomes (9.3 ± 0.6) occurred on the leaf midvein.
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Table 1. Mean (±S.E.) density of glandular and nonglandular trichomes recorded from the potato leafhopper-resistant alfalfa clone FGplh13. Trichome densities were recorded from a 2-mm section of stem, petiole, leaf midvein, and leaf perimeter.
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Kreitner and Sorensen (1979a) found the erect glandular trichome to be visibly more dense than the procumbent trichome on the perennial diploid M. prostrata. However, because of differences in forms of measurement, the recordings of trichome density from FGplh13 could not be compared effectively with previous work involving annual (Othman et al., 1981) or perennial (Brewer et al., 1986, Shade and Kitch, 1986; Danielson et al., 1989; Elden and McCaslin, 1997; Lenssen et al., 1998) Medicago spp. The recording technique utilized by Elden and McCaslin (1997) was similar to that used in this study, although they did not make distinctions between the erect and procumbent glandular trichomes.
There is not a strong correlation between glandular trichome density and potato leafhopper resistance (Hogg and McCaslin, 1994; Elden and McCaslin, 1997). However, a failure to consider the density of both the erect and procumbent glandular trichomes could have affected the interpretation of the results. A correlation may exist between erect and procumbent glandular trichome density and potato leafhopper resistance.
With respect to distribution, on the abaxial leaf surface of FGplh13, both the erect and procumbent glandular trichomes were present. Glandular and nonglandular trichomes were not detected on the adaxial surface of the leaves. All three types of trichomes were observed on the stipules, peduncles, and sepals. However, the procumbent glandular trichome was rarely found on the stipules and sepals. Similar to that on the leaflets, the erect glandular trichomes lined the perimeter of the stipules and sepals. Glandular and nonglandular trichomes were not detected on the flower petals.
Resistance of FGplh13 to the Potato Leafhopper
Elden and McCaslin (1997) suggested that the glandular trichomes found on perennial alfalfa could produce toxic or repellent compounds which function by way of a volatile or tactile avenue, or that toxins could be present in the plant phloem that are ingested during feeding. By removing the glandular trichomes from the alfalfa clone FGplh13, and examining the biology and behavior of the potato leafhopper, the glandular trichomes were implicated in providing resistance to the potato leafhopper (Ranger, 1999). However, it is not known whether the erect and procumbent glandular trichomes are each providing resistance factors, and if they act synergistically. A synergistic mechanism has been documented between the Type A and Type B glandular trichomes found on various Solanum spp. (Tingey and Laubengayer, 1981; Neal et al., 1989).
On the basis of the current study, both the erect and procumbent glandular trichomes found on the perennial alfalfa clone FGplh13 appear capable of secreting an exudate. Because of the propensity of biologically active secondary products known to be secreted by glandular trichomes (Duke et al., 2000), the possibility exists for the glandular trichomes on perennial alfalfa to be providing a chemical basis for resistance. As a result, the mechanism is still too ambiguous for researchers to ignore the potential role of the erect and procumbent glandular trichomes. Analytical and behavioral techniques are presently being utilized to identify compounds associated with the erect and procumbent forms that may be responsible for imparting resistance to this key insect pest.
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ACKNOWLEDGMENTS
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We thank Rosemary Walsh and Michelle Peiffer (The Electron Microscope Facility, Penn. State Univ., University Park, PA) for various aspects of sample preparation and technical support, as well as assistance in using the microscopy instruments. We also acknowledge Mark McCaslin (Forage Genetics, West Salem, WI) for providing the resistant alfalfa clone used in this study. The authors thank R.R. Youngman (Dep. of Entomology, Virginia Polytechnic Institute and State Univ., Blacksburg, VA) and Mark McCaslin for critically reviewing an earlier draft of this manuscript. This research was funded in part by a USDA, CSREES NE Regional IPM grant to the University of Maryland and their Cooperative Agreement Z56501 with The Pennsylvania State University.
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NOTES
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This research was conducted at the Dep. of Entomology, Penn. State Univ., University Park, PA, and was funded in part by a USDA, CSREES NE Regional IPM grant to the Univ. of Maryland and their Cooperative Agreement Z56501 with Penn. State Univ.
Received for publication September 11, 2000.
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REFERENCES
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ABSTRACT
INTRODUCTION
