Effect of three Greek arbuscular mycorrhizal fungi in improving the growth,nutrient concentration,and production of essential oils of oregano and mint plants

Arbuscular mycorrhizal fungi have been widely used in agriculture to improve the cultivation of many crops. One of the aims of this study was the isolation and molecular identification of arbuscular mycorrhizal fungi isolated from mountain areas of Northern Greece (Ritini Pieria, Elatochori Pieria, Ambelakia Ossa). Only three isolates were obtained; two of Glomus etunicatum and one of G. lamellosum. The second objective of this study was to investigate the effect of these arbuscular mycorrhizal fungi on the concentration of macro- and micronutrients in tissues, the quantity and quality of essential oils and the growth of oregano and mint plants (two widely used aromatic plants in Greece). It was found that mycorrhizal oregano and mint plants had a higher content of essential oils and nutrient elements, and grew better than non-mycorrhizal plants. In addition, the composition of the essential oil in mycorrhizal plants differed from the oil of non-mycorrhizal plants. These results suggest that the use of mycorrhizal fungi may allow plant growth in low fertility soils, reduce fertilizer inputs and increase aromatic plant production of essential oils, They also indicate that it may be possible to use mycorrhizae to affect the quality of the essential oil produced.     

Analysis of phenetic relationship between Citrus indica Tanaka and a few commercially important citrus species by ISSR markers

Analysis of polymorphism in ISSR amplicons revealed diverse genetic relationship between Citrus indica and five other citrus species. In a consensus UPGMA dendrogram, based on Nei and Li's distance matrices, the C. indica samples from three different sites of Meghalaya, India were clustered together with 99% bootstrap support. C. reticulata, C. sinensis, C. aurantifolia formed a cluster with 67% bootstrap separation. C. macroptera and C. maxima samples from two sites of Meghalaya formed separate clusters with respectively 100% and 98% bootstrap supports. Principal Components Analysis projected C. indica to be more closely related to C. aurantifolia than to the other citrus species of the study. PCA also exposed variations within the C. maxima and C. macroptera samples.     

Effect of arbuscular mycorrhizal fungi inoculation on aster yellows phytoplasma-infected tobacco plants

The objective of the research was to asses if arbuscular mycorrhizal fungi (AM) can modify the effect of two aster yellows phytoplasma strains infection in tobacco plants. Tobacco plants experimentally inoculated with aster yellows phytoplasma strains did not develop visible disease symptoms. However, PCR examination indicated that the inoculated plants were phytoplasma infected. Mycorrhiza inoculation had a positive effect on the shoot height of healthy plants, but did not influence shoot growth and weight of phytoplasma-infected plants. The roots of all mycorrhiza-inoculated plants were slightly reduced but significant differences were found in the total root length of plants infected with the phytoplasma strain AY1. AM inoculation had a positive effect on photosynthetic activity of tobacco plants infected with the phytoplasma strain AYSim, but net photosynthesis of tobacco infected with the phytoplasma strain AY1 was decreased. Transpiration rate and calcium content of AM and phytoplasma-infected plants were not affected. The mechanisms underlying these interactions are discussed and a direct action of the AM fungus is hypothesized.     

Screening and selecting novel AM fungi and Azotobacter strain for inoculating apple under soil solarization and chemical disinfestation with mulch practices for sustainable nursery management

The objective of this study is to investigate the response of screening, and selection of novel indigenous AM fungal species and Azotobacter chroococcum strains for inoculating apple under different soil disinfestations and moisture conservation mulch practices for sustainable nursery management. Two local AM fungal species namely, Glomus fasciculatum (Thaxter sensu Gerdemann) and Glomus mosseae (Nicol. & Gerd.), and two strains of A. chroococcum viz., A. chroococcum strain-I (AZ₁) and A. chroococcum strain-II (AZ₂) were inoculated at nursery stage under soil solarization, chemical disinfestation and natural soil conditions at four different mulch materials namely, black plastic mulch (BPM), and organic mulches, i.e. grass mulch (GM); cover crops (CC); green manuring + clean cultivation (Gm + Cc). The comparative performance of the seedlings on the impact of local AM species and A. chroococcum strains on growth characteristics, microbial population, root colonization and leaf nutrient status was evaluated. The inoculation of seedlings to G. fasciculatum and AZ₁ increased all growth characteristics (plant height, stem diameter, leaf area and total root length), microbial consortium of the rhizosphere soil and leaf N, P, K and Zn content in all those plots where soil solarization and black plastic mulching was used followed by chemical disinfestations and natural soil conditions at all other mulch types used. These findings suggested that the soil inoculation of G. fasciculatum and AZ₁ strain to seeds and/or the saplings under soil solarization with black plastic mulch attained a desirable plant height and become ready for grafting which however saved a period of 1 year for nursery management compared to traditional nursery raising practice, and thus, it may be a viable and feasible approach to maintain soil productivity under nutrient limited soils for sustainable apple nursery production under temperate rain-fed conditions.     

Screening of AM fungi and Azotobacter chroococcum under natural,solarization, chemical sterilization and moisture conservation practices for commercial mango nursery production in north-west Himalayas

The present study investigates the performance of mango seedlings screened with indigenous arbuscular mycorrhizal (AM) fungi and Azotobacter chroococcum strains under solarized, chemical sterilized and natural soil conditions. Two isolates each of AM fungi namely, Glomus fasciculatum (Thaxter sensu Gerdemann) and Glomus mosseae (Nicol. & Gerd.), and two strains of A. chroococcum viz., A. chroococcum strain-I (AZ1) and A. chroococcum strain-II (AZ2) were inoculated at nursery stage under four different moisture conservation practices viz., black polyethylene mulch and organic mulches (grass mulch, cover crops, green manure) and clean cultivation. The observations on microbial population, root colonization, growth parameters and leaf nutrient content of the seedlings were recorded. Mango seedling's inoculated with G. fasciculatum and AZ1 had increased seedling's height, diameter, leaf area and total root length, microbial consortium of the rhizosphere soil and leaf N, P, K and Zn content in plots where solarization and black polyethylene mulching was used. The study revealed that the inoculation of mango stones and the saplings with G. fasciculatum and AZ1 under solarized black polyethylene mulched practice may be considered the best practice for raising mango nursery and maintaining soil health under rain-fed conditions of mid-hills of north-west Himalayas.     

Effect of arbuscular mycorrhizal fungi on growth,mineral nutrition,antioxidant enzymes activity and fruit yield of tomato grown under salinity stress

The purpose of this study was to investigate the mechanisms underlying alleviation of salt stress by arbuscular mycorrhizal fungi Glomus mosseae. Tomato (Lycopersicon esculentum L. cv. Zhongzha105) plants were cultivated in soil with 0, 50 and 100 mM NaCl. Mycorrhization alleviated salt induced reduction of root colonization, growth, leaf area, chlorophyll content, fruit fresh weight and fruit yield. The concentrations of P and K were higher in AM compared with nonAM plants grown under nonsaline and saline conditions. Na concentration was lower in AM than nonAM plants grown under nonsaline and saline conditions. AMF colonization was accompanied by an enhancement of activity of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and ascorbate peroxidase (APX) in leaves of both salt-affected and control plants. In addition, inoculation with AMF caused reduction in MDA content in comparison to salinized plants, indicating lower oxidative damage in the colonized plants.     

Chemical compositions and insecticidal effects of essential oils isolated from Achillea gypsicola, Satureja hortensis, Origanum acutidens and Hypericum scabrum against broadbean weevil (Bruchus dentipes)

The essential oils of aerial parts of Achillea gypsicola Hub.-Mor., Hypericum scabrum L., Satureja hortensis L., and Origanum acutidens (Hand.-Mazz.) Letswaart were analyzed in this study by GC and GC–MS and their oils were tested for toxicity against broadbean weevil (Bruchus dentipes). A. gypsicola oil contained camphor (40.17%), 1,8-cineole (22.01%), piperitone (11.29%), borneol (9.50%) and α-terpineol (1.56%) as major components. A total of 74 components were identified by GC–MS in H. scabrum oil, including α-pinene (9.26%), terpinen-4-ol (5.12%), camphor (5.94%), δ-cadinene (4.52%), pulegone (4.45%), γ-muurolene (4.12%), pinocarvone (3.97%) and β-caryophyllene (3.42%) as predominant components. The essential oils of O. acutidens and S. hortensis were characterized by high contents of carvacrol (86.99% and 55.74%), γ-terpinene (0.71% and 20.94%), p-cymene (1.95% and 12.30%), α-terpinene (0.13% and 2.04%) and β-caryophyllene (1.30% and 1.08%). All of the essential oils were toxic to adults of B. dentipes and insect mortality increased with increasing concentration of each oil. The oils (20 μl dose) brought about 100% mortality in 36 h. Although desirable insecticidal activities against the pest were achieved with the oils from all four plant species, S. hortensis and O. acutidens oils were more effective, particularly after 6 h of treatment. The current results concluded that the essential oils, in particular O. acutidens and S. hortensis oils, may be used as potential botanical insecticides against B. dentipes.     

Development of ITS sequence based SCAR markers for discrimination of Paphiopedilum armeniacum, Paphiopedilum micranthum, Paphiopedilum delenatii and their hybrids

Paphiopedilum armeniacum, Paphiopedilum micranthum and Paphiopedilum delenatii are endangered orchid species. These three Paphiopedilum species and their hybrids are difficult to distinguish morphologically. In this study, rDNA-ITS (internal transcribed spacer) sequences were used to design species-specific SCAR (sequence characterized amplified regions) markers to distinguish P. armeniacum, P. micranthum, P. delenatii and their respective hybrids. The developed markers efficiently amplified 600 bp DNA product for P. armeniacum and its hybrids (SCAR-600armF/Pap-ITS2R), 300 bp product for P. delenatii and its hybrids (SCAR-300delF/Pap-ITS2R) and 700 bp product for P. micranthum and its hybrids (SCAR-700micF/Pap-ITS2R). The effectiveness of designed species-specific markers was also confirmed by using multiplex polymerase chain reaction amplification with a combination of developed three SCAR markers.     

Development of seedless and Mal Secco tolerant mutant lemons through budwood irradiation

Mal secco (caused by Phoma tracheiphila (Petri) Kantsch. and Gik.) is the most destructive fungal disease of lemon plantations worldwide and seedless lemons would be preferred by most consumers. Five dosage levels, 0, 3, 5, 7, and 9 kiloradian (krad), of cobalt (⁶⁰Co) gamma irradiation were applied to budstick of ‘Kutdiken’ lemon (Citrus limon (L.) Burm. f.) clone KT-2A. Mutations were stabilized in three vegetative generations. Three hundred fifty-eight and 478 M1V3 (mutation one and vegetation three) plants were evaluated for seed number and mal secco tolerance in the field and the greenhouse, respectively. LD₅₀ was around 5 krad gamma irradiation for ‘Kutdiken’ lemon. The seed number varied from 0 to 34 per fruit. The level of mal secco tolerance also varied significantly among the plants from 1.0 (no symptom) to 4.3 (high level of disease occurance). The stable seedless and mal secco tolerant plants were obtained from 5 and 7 krad irradiation: the three mutants from 5 krad irradiation gave more lemon-like fruits, while 7 krad irradiation caused altered tree morphology and early maturation of fruits. This study shows considerable potential for lemon cultivar improvement aiming to obtain seedless and mal secco tolerant lemons.     

Production protocol development for greenhouse cut Linaria, Lupinus, and Papaver flowers

Linaria maroccana Hook. f. Ann., ‘Lace Violet’, Lupinus hartwegii ssp. cruikshankii Lindl. ‘Sunrise’ and Papaver nudicaule L. ‘Meadow Pastels’ seeds were directly sown into 105 cell plug trays and received either ambient light or supplemental high intensity discharge (HID) lighting. For each species, a 2 × 3 × 3 factorial was used with two light intensities during propagation, three transplant stages, and three night temperatures. Seedlings were transplanted at the appearance of 2–3, 5–6, or 8–9 true leaves. Transplanted Linaria and Papaver seedlings were placed at 5/11, 10/16, or 15/21 ± 1 °C night/day temperatures and Lupinus seedlings were placed at 15/24, 18/25, or 20/26 ± 2 °C night/day temperatures. For this study, the optimum production temperature for Linaria was 10/16 °C as the cut stems produced at 15/21 °C were unmarketable and production time was excessively long at 5/11 °C. At 10/16 °C, Linaria seedlings should be transplanted at the 2–3 leaf stage to maximize stem number, stem length and profitability. For Lupinus the optimum temperature was 15/24 °C due to long stems and high profitability per plant. Lupinus seedlings should be transplanted at the 2–3 leaf stage when grown at 15/24 °C to obtain the longest and thickest stems; however, $/m² week was higher for plants transplanted at the 8–9 leaf stage due to less time in finishing production space. For Papaver, the 15/21 °C temperature was optimal as that temperature produced the longest stems in the shortest duration, resulting in the highest $/m² week. At 15/21 °C Papaver plants should be transplanted at the 2–3 leaf stage. Supplemental HID lighting had no effect on any of the species.     

Influence of sulfur supply,ammonium nitrate ratio,and arbuscular mycorrhizal colonization on growth and composition of Chinese chive

Tissue concentrations of phytochemicals formed by Allium species may be influenced by plant nutrient supply or rhizosphere microorganism activity. To test these relations, three plant nutritional factors were varied in two experiments with Chinese chive [Allium tuberosum]: (a) increasing sulfur concentrations in the substrate, (b) two ratios of ammonium to nitrate in supply, and (c) inoculation with arbuscular mycorrhizal (AM) fungi. Shoot dry weight, nutrient composition (nitrate, N, P, S), and organosulfur compounds (measured indirectly as pyruvic acid) were determined.     

The influence of desiccation and rehydration on the survival of polyembryonic seed of Citrus suhuiensis cv. limau madu

A study was carried out to investigate the influence of desiccation and freezing followed by various presowing rehydration procedures on the desiccation sensitivity of the seed of Citrus suhuiensis cv. limau madu. The freshly harvested seeds of limau madu were desiccated under a broad range of relative humidity (RH) to various equilibrium water contents (g H₂O g⁻¹ dw). The desiccated and desiccated–frozen seeds were either directly sown under germination conditions or subjected to presowing rehydration procedures: seed preheating, prehumidification and osmoconditioning. The hydrated and desiccated seeds were sown in controlled germination conditions and the survival was evaluated 4–6 weeks after sowing. The results showed that desiccation progressively reduced the percentage of normal seedling of the seeds of limau madu and the viability is almost lost at water contents below 0.08 g H₂O g⁻¹ dw. The estimated desiccation sensitivity was substantially high (WC₅₀ = 0.143 g H₂O g⁻¹ dw) when the desiccated seeds were rapidly rehydrated (uncontrolled rehydration). In contrast, seed prehumidification, preheating and osmoconditioning (controlled rehydration procedures) markedly enhanced normal seedling percentages decreasing the estimated values of WC₅₀ (between 0.08 and 0.127 g H₂O g⁻¹ dw). While the rapidly rehydrated desiccated–frozen seeds were almost killed at water content of 0.15 g H₂O g⁻¹ dw, prehumidification and preheating have noticeably increased percentage of frozen seeds survival at the same water content. However, at water content of 0.21 g H₂O g⁻¹ dw preheating significantly (P < 0.05) increased percentage of normal seedling of the frozen seeds. Seed desiccation markedly reduced the percentages of germinated seeds with multiple seedlings. Seed controlled rehydration remarkably increased the survival of polyembryos. The beneficial effect of seed controlled rehydration on the survival of the desiccated seeds was pronounced at medium water contents (0.08–0.25 g H₂O g⁻¹ dw).     

Genetic transformation of Prunus domestica L. using the hpt gene coding for hygromycin resistance as the selectable marker

The study and development of transformation technology with new selection schemes is important for various fundamental studies and for crop trait improvement via genetic engineering. Here we have shown that hygromycin resistance is an effective system for plum genetic transformation. Embryonic axes of mature seeds were co-cultivated with Agrobacterium strain LBA4404 containing the pC1381 plasmid carrying the hygromycin phosphotranferase gene (hpt) and β-glucuronidase (GUS) gene or with strain EHA105 containing the plasmid pC1301 carrying the same marker and reporter genes. The latter strain containing a pC2301 plasmid carrying the neomycin phosphotransferase gene (nptII) gene was used as a control. Infected explants were placed on shoot induction medium containing either 5 mg L⁻¹ hygromycin or 75 mg L⁻¹ kanamycin for selection. Green shoots developed from the explants under hygromycin pressure. These shoots showed continued and vigorous growth and development upon transfer onto fresh hygromycin medium. PCR using hpt sequence primers, and Southern blot analysis using a probe from the hpt gene, confirmed the presence of the transgenes and their stable integration in regenerated plants. Full transgenic plants were obtained in a greenhouse. Hygromycin selection was very effective and no escapes were observed. The study demonstrated that hygromycin resistance can be used as an effective selectable marker for plum transformation. The new system developed here is important and useful for multiple gene transformation in plum.     

Ethylene influences development and flowering of Ptilotus spp. in vitro and ex vitro

The genus Ptilotus has immense potential for ornamental horticulture but its commercial development has been hindered by propagation limitations. Poor seed quality and germination are reported. Cutting propagation is limited by cutting supply as the juvenile phase of Ptilotus is short. Micropropagation has been used in an attempt to overcome these difficulties but explants become floral in vitro and this causes plantlets to elongate. Ethephon has been used to control flowering of stock plants of many ornamental species. This study investigated the effect of ethephon applied to young (3-week-old, deflasked from tissue culture) and mature (1-year-old) Ptilotus plants in a greenhouse. A system of applying gaseous ethylene at 0, 100, 200 and 300 mg l⁻¹ to the headspace of in vitro plantlets in glass jars was developed and the response of in vitro plantlets to ethylene studied. One-year-old Ptilotus plants were treated with 500 mg l⁻¹ ethephon 2 days before pruning or 1 or 2 weeks after pruning. Ethephon application 2 days before pruning decreased the number of inflorescences and increased the number of shoots (compared to the control) but was phytotoxic. Ethephon applications of 150 or 300 mg l⁻¹ applied weekly or fortnightly to 3-week-old plants deflasked from tissue culture reduced plant height and number of inflorescences and at low concentrations increased the number of new shoots. A fortnightly application at 150 mg l⁻¹ is recommended. Previous reports on the effects of ethylene on inflorescence production on plantlets in vitro are limited. Our study showed that exposure of in vitro plantlets of P. nobilis to ethylene gas at 100 mg l⁻¹ for 1 h significantly increased the number of shoots and plant height but this did not occur for plantlets of P. spicatus. Plantlets of P. spicatus exposed to transient ethylene at 200 and 300 mg l⁻¹ showed significantly greater rooting (52.4%) than the control (13.6%).     

Twenty years of agronomic evaluation of wild cocoa trees (Theobroma cacao L.) from French Guiana

Almost 500 clones of wild cocoa trees (Theobroma cacao L.) grown from pods collected in 1987 from wild mother-trees in the Camopi and Tanpok river basins (southeastern French Guiana) have been distributed in around fifteen cocoa producing countries since 1988. The name of those clones always bears the GU prefix (for “Guyane”, i.e. French Guiana). All the germplasm of the same geographical origin present in the CIRAD collection at Paracou-Combi (Sinnamary, French Guiana), i.e. more than 1600 trees, has been abundantly studied for its morphological characterization, its agronomic assessment or its genetic diversity. Other assessment work, primarily on resistance to certain diseases, has been carried out by CIRAD in Montpellier, or in various countries by other organizations.     

H2O2 metabolism during sweet orange (Citrus sinensis L. Osb.) ‘Hamlin’ Xanthomonas axonopodis pv. citri interaction

Sweet orange (Citrus sinensis L. Osb.) ‘Hamlin’ is a canker (Xanthomonas axonopodis pv. citri: Xac) susceptible citrus genotype grown commercially worldwide. Canker causes severe economic losses and restricts the marketability of crop for export. Little is known about the role of oxidative stress in canker development. In the present investigation, sweet orange ‘Hamlin’ leaves were artificially inoculated with Xac to determine the impact of Xac infection on hydrogen peroxide (H₂O₂) metabolism. Characteristic symptoms following artificial inoculation were water soaking of the infiltrated zone between 2 and 8 days after inoculation (dai); raised epidermis accompanying tiny yellow colored bacterial colonies at 8 dai; and yellowing and necrosis of the infected zone by 12–16 dai. In planta Xac population increased 1000 fold by 14 dai from an initial population of 7.3 × 10⁶ cfu cm⁻² (0 dai). Peak concentrations of H₂O₂ were observed at 24 h and between 8 and 10 dai and coincided with higher activity of total superoxide dismutase (SOD). Lower levels of H₂O₂ in infected leaves were maintained by Xac induced higher activities of catalase (CAT), ascorbate peroxidase (APOD), and guaiacol peroxidase (POD). It appears Xac altered H₂O₂ metabolism in C. sinensis L. Osb. ‘Hamlin’ to enhance survival and growth.     

In vitro asymbiotic seed germination and seedling growth of Ansellia africana Lindl.

The effects of sodium hypochlorite treatment and growth medium on in vitro seed germination and seedling growth of the leopard orchid (Ansellia africana Lindl.) were investigated. Forty minutes treatment of seeds with 1.75% sodium hypochlorite stimulated germination (70.6%) and seedling development when measured at Stage 5 (emergence of first leaf) on P668 medium after 8 weeks of culture. The growth medium played a significant role in determining the germination response of A. africana seeds. Dark pretreatment of seed cultures significantly enhanced the germination percentage and the growth of rhizoids on the protocorms. Leaf growth in terms of length was substantially enhanced on P668 medium. It was significantly reduced in modified Knudson C medium after 16 weeks of culture. Seedlings developed on P668 medium showed a significantly better growth performance in relation to leaf length, leaf number, root length, fresh and dry weights per plant in vermiculite after 12 weeks of ex vitro growth in a mist house with 90% relative humidity. Further studies developing a suitable method for in vitro symbiotic germination could assist in reintroduction of this threatened orchid species into the wild.     

DNA content,morphometric and molecular marker analyses of Citrus limonimedica, C. limon and C. medica for the determination of their variability and genetic relationships within the genus Citrus

This work investigated the fingerprinting and phenotyping of Citrus germplasm; species selected were of historical importance belonging to Citrus limonimedica Lush. and its supposed ancestors, along with some other species of the Citrus genus. An integrated approach based on the exploitation of nuclear DNA content, morphological traits and molecular markers, such as RAPD fingerprints and ITS-based SNPs, was employed. We studied a core collection of 54 distinct accessions, including 43 genotypes of the Citrus species (18 species or supposed species) and 11 genotypes of the Poncirus genus, which was used as the reference outgroup. Morphological trait analysis and statistical analysis of DNA content and markers were useful for reconstructing a Citrus phylogeny. In particular, our experiments aimed at estimating the genetic variation within and the genetic relatedness among C limon (L.) Burm., C. limonimedica and C. medica L. to shed light on the hybrid origin hypothesis of C. limonimedica. The results of the multidisciplinary analyses allowed us to confirm a remarkable differentiation between Poncirus and Citrus genera and to highlight a close relationship among the three investigated Citrus species but a distinct difference between these three species and other species in the Citrus genus. RAPD fingerprints and ITS polymorphisms enabled us to point out a variation gradient between lemon and citron, with C. limonimedica as a possible intermediate species. Some accessions of C. medica and C. limonimedica that deviate from such a trend suggest recurrent introgression and/or hybridisation with other species of Citrus.     

The effects of temperature on in vitro pollen germination and pollen tube growth of Pistacia spp.

The pollen germination and pollen tube growth among nine Pistacia genotypes was quantified in order to identify differences in the tolerance of pollen to temperature variations. The effect of temperature on in vitro pollen germination and pollen tube growth was investigated in Pistacia vera (Uygur, Atli, Kaska, Sengel, Kavak), P. atlantica, P. khinjuk, P. terebinthus and P. palaestina. When pollen was incubated in a germination medium for 24 h in darkness, distinct differences were observed in pollen germination and pollen tube growth at different temperatures.     

Taxonomic revision of Dendrobium moniliforme complex (Orchidaceae)

Taxonomic revision of Dendrobium moniliforme complex is presented. D. moniliforme complex is characterized by the even slim stems, bracts with brownish zone, semi-spherical anther cap and the hairy disc of lip. Dendrobium tosaense, Dendrobium officinale and Dendrobium guangxiense were excluded by having membranous bracts lacking brownish zone, anther cap conical and bifid. Two species are recognized in this complex, i.e., D. moniliforme and Dendrobium wilsonii. D. wilsonii differs from D. moniliforme by having elliptic leaves about 1.3–2 cm wide, dorsal sepal 3.0–4.0 cm long, 0.6–0.9 cm wide, petals elliptic to oblong, 3.0–4.0 cm long, 1.0–1.5 cm wide, lip elliptic to ovate–lanceolate, 2.6–3 cm long, 1.2–1.5 cm wide.