Clinical Significance of Topical Spermidine Hyaluronate in Vestibulodynia: An Early Appraisal

Background: The most common subtype of chronic vulvar pain is provoked vestibulodynia. The entry of the vagina is the site of acute and recurrent pain in this highly prevalent and debilitating condition, which is characterized by pressure application or attempted vaginal penetration. The aim of this study was to determine the effectiveness of topical spermidine in patients with vestibulodynia. Methods: Topical gels containing spermidine in hyaluronate complexes Ubi1 and Ubi2 endowed with differentiated release ratio and viscosity were applied at 3 doses/week during 4-weeks, then at 2 doses/week during the next 4-weeks in two groups of patients. Pain relief was measured by visual analogic score (VAS) and dyspareunia score expressed as percent improvement from baseline to posttreatment. Results: Group 1 treated with Ubi1 provided improvement in pain (46%) and dyspareunia (27%). However, the treatment in Group 2 resulted in a superior amelioration: VAS of pain (76%) and dyspareunia (50%) as Ubi2 gel provided higher dose and viscosity along with improved local application. Conclusions: Our results demonstrated that preparation 2 resulted in greater reduction in symptoms as compared to preparation 1 as measured by the VAS and Marinoff scale. These early, yet outstanding clinical outcomes in vestibulodynia through to the stimulation of tissue mechanosensor and their relevant downstream effects are reviewed hereafter.

Ethylene and Spermidine in Wheat Grains in Relation to Starch Content and Granule Size Distribution Under Water Deficit

Two wheat cultivars （Triticum aestivum L.） were used to evaluate the effects ofpost-anthesis severe water deficit （SD） on starch content and granule size distribution and their relations with ethylene and spermidine （Spd）. Comparison to the well-watered （WW） treatment, SD led to lower Spd and higher 1-aminocylopropane-l-carboxylic acid （ACC） concentrations and ethylene evolution rate （EER） in grains at the critical stage of forming starch granules. Application of Spd or aminoethoxyvinylglycine （AVG） significantly reduced ACC concentration and EER and increased Spd concentration, while ethephon or methylglyoxal-bis （MGBG） had an opposite impact. The volume and surface area distribution of starch granules showed a bimodal curve, while the number distribution exhibited a unimodal curve. SD caused a marked drop in grain weight, grain number and starch content, also led to a significant reduction in the proportion （both by volume and by surface area） of B-type starch granules （〈10 Ixm）, with an increase in those of A-type starch granules （〉10 ~tm）. Application of Spd or AVG increased the proportion （both by volume and by surface area） of B-type starch granules under SD. Correlation analysis suggested that ethylene and Spd showed an antagonism relation in the formation of B-type granules. These results suggested that it would be good for the formation of B-type starch granules to have the physiological traits of higher Spd and lower ACC concentrations and ethylene emission under SD.

Effect of Ice-Temperature and Spermidine on Chilling Sensitivity of Pepper

The peppers which were treated at ice-temperature for 6 hours and 1 mmol/L spermidine for 10 min, followed by cold storage (4℃ ± 1℃) respectively were investigated. The results indicated that the chilling injury was delayed and reduced by ice-temperature and spermidine treatments. Comparing with control group, ice-temperature and spermidine exerted significant effects on reduction of ascorbic acid (Vc) and chlororphyll contents, relatively high activity of superoxide dismutase (SOD), accumulation of malondialdehyde (MDA) as well as the increase of cell membrane penetrability. The results indicated that the spermidine and ice-temperature induced the activity of antioxidant enzyme SOD and maintained higher defence-related compound Vc as well as lower levels of membrane lipid peroxidation, which may be associated with chilling injury alleviation. There was a difference between ice-temperature treatment and spermidine treatment after 30 days. The results suggested that spermidine was more effective in reducing chilling sensitivity and prolonging storage of peppers.

Exogenous Spermidine Promotes Somatic Embryogenesis of Cunninghamia lanceolata by Altering the Endogenous Phytohormone Content

In order to study how exogenous hormones in C.lanceolata(gymnosperm)regulate somatic embryogenesis,we measured the endogenous phytohormones of two genotypes with different somatic embryogenesis efficiency and found that an increase in endogenous concentrations of IAA and ABA may be correlated to more efficient somatic embryogenesis.By applying exogenous spermidine,we found that exogenous hormones may affect somatic embryogenesis efficiency through affecting the endogenous phytohormone content.Based on these results,further studies can be conducted whereby the concentration of exogenous hormones or the levels of endogenous phytohormones by molecular methods are regulated to promote somatic embryogenesis.Our research may benefit the long-term economic output of the forestry industry and lays the foundation to studying the molecular mechanism that controls somatic embryogenesis efficiency.

Differential Proteins Expressed in Rice Leaves and Grains in Response to Salinity and Exogenous Spermidine Treatments

Exogenous application of spermidine（Spd） has been reported to modulate physiological processes and alleviate salt-induced damage to growth and productivity of several plants including rice. Employing a proteomic approach, we aimed at identifying rice leaf and grain proteins differentially expressing under salt stress, and in response to Spd prior to Na Cl treatment. A total of 9 and 20 differentially expressed protein spots were identified in the leaves of salt-tolerant（Pokkali） and saltsensitive（KDML105） rice cultivars, respectively. Differential proteins common to both cultivars included a photosynthetic light reaction protein（oxygen-evolving complex protein 1）, enzymes of Calvin cycle and glycolysis（fructose-bisphosphate aldolase and triose-phosphate isomerase）, malate dehydrogenase, superoxide dismutase and a hypothetical protein（Os I_18213）. Most proteins were present at higher intensities in Pokkali leaves. The photosynthetic oxygen-evolving enhancer protein 2 was detected only in Pokkali and was up-regulated by salt-stress and further enhanced by Spd treatment. All three spots identified as superoxide dismutase in KDML105 were up-regulated by Na Cl but down-regulated when treated with Spd prior to Na Cl, indicating that Spd acted directly as antioxidants. Important differential stress proteins detected in mature grains of both rice cultivars were late embryogenesis abundant proteins with protective roles and an antioxidant protein, 1-Cys-peroxiredoxin. Higher salt tolerance of Pokkali partly resulted from higher intensities and more responsiveness of the proteins relating to photosynthesis light reactions, energy metabolism, antioxidant enzymes in the leaves, and stress proteins with protective roles in the grains.

PRODUCTION AND APPLICATION OF MONOCLONAL ANTIBODY TO POLYAMINE (PREPARATION AND CHARACTERISTICS OF MONOCLONAL ANTIBODIES AGAINST SPERMIDINE)

A monoclonal antibody was first prepared by fusion of mouse myeloma cells (SP2/0-Ag-14) with spleen cells isolated from male BALB/ c mice immunized with spermidine-bovine serum albumin conjugate (SPD- BSA). The hybridoma cell line producing antibody specific for spermidine was cultured in vitro and after i. p. into mice, the ascitic fluid gave suitably high dilution titres (1: 106) by enzyme immunoassay. This monoclonal antibody is of IgG1 class and the bimolecular compleex with molecular weight of 52KD and 27 KD. The monoclonal antibody was clearly specific to spermidine comparing with spermine or putriscine. Monclonal antibody may prove to be useful in the rapid diagnosis and evaluation of patients with cancer.

Potent trophic activity of spermidine supramolecular complexes in in vitro models

AIM:To test the growth-promoting activity of the polyamine spermidine bound to various polymeric compounds in supramolecular complexes.METHODS:A thiazolyl blue cell viability assay was used to determine the growth-promoting potency of spermidine-supramolecular complexes in a human skin fibroblast cell line exposed to spermidine and different spermidine-supramolecular complexes that were obtained by combining spermidine and polyanionic polymers or cyclodextrin.Reconstituted human vaginal epithelium was exposed to a specific spermidinesupramolecular complex,i.e.,spermidine-hyaluronan(HA)50,and cell proliferation was determined by Ki-67immunohistochemical detection.Transepithelial electrical resistance and histological analysis were also performed on reconstituted human vaginal epithelium to assess tissue integrity.RESULTS:The effect of spermidine and spermidinesupramolecular complexes was first tested in skin fi-broblasts.Spermidine displayed a reverse dose-related mode of activity with mmol/L growth inhibition,whereas 30%stimulation over basal levels was detected at mol/L and nmol/L levels.Novel spermidine-supramolecular complexes that formed between spermidine and polyanionic polymers,such as HA,alginate,and polymaleate,were then tested at variable spermidine concentrations and a fixed polymer level(0.1%w/v).Spermidine-supramolecular complexes stimulated the cell growth rate throughout the entire concentration range with maximal potency(up to 80%)at sub-mol/L levels.Similar results were obtained with spermidine-(-cyclodextrin),another type of spermidine-supramolecular complex.Moreover,the increased expression of Ki-67 in the reconstituted human vaginal epithelium exposed to spermidine-HA 50 showed that the mode of action behind the spermidine-supramolecular complexes was increased cell proliferation.Functional and morphological assessments of reconstituted human vaginal epithelium integrity did not show significant alterations after exposure to spermidine-HA,thus supporting its safety.CONCLUSION:Spermidine found in spermidine-supramolecular complexes displayed potentiated regenerative effects.Safety data on reconstituted human vaginal epithelium suggested that assessing spermidinesupramolecular complex efficacy in atrophic disorders is justified.

SPERMIDINE RADIOIMMUNOASSAY IN MONITOR OF PRECANCEROUS LESIONS OF ESOPHAGUS

Serum spermidine was assayed by radioimmuno-assay in different stages of esophageal carcinogenesis in the population from high risk area of esophageal cancer, Linxian County. The serum spermidine values were 76.94+74.38 ng ml in 36 normal individuals; 115.71+113.45 ng/ml in 35 patients with marked epithelial hyperplasia (MEH) and 294.48+135.36 ng/ml in 31 patients with early esophageal cancer. Patients with MEH were given intervention treatment by Aminoretinoic Ester or Anticancer B or placebo (starch) as controls. One year later, samples from the population were collected again for serum spermidine msasurement. The values were 95.8+ 68.2 ng/ml in 27 normals; 125.1±72.9 ng/ml in 62 patients with MEH treated by Anticancer B; 125.6± 117.2 ng/ml in 64 patients with MEH treated by Aminoretinoic Ester; 162.4±76.6 ng/ml in 62 controls and 210.5±182.9 ng/ml in 44 patients with early esophageal cancer.The results showed that spermidine radioim-munoassay could reflect the tendency of esophageal precancerous changes toward cancer or back to normal. They can be taken as a mid-way monitor indicator for tumor-blocking drugs. Also, it could be of value in the early diagnosis of esophageal cancer.

Effects of Exogenous Spermidine on Antioxidant System Responses of Typha latifolia L. Under Cd^2＋ Stress

: The effects of foliar spraying with spermidine (Spd), ranging in concentration from 0.25 to 0.50 mmol/L, on the antioxidant system under Cd2+ stress (range 0.1–0.2 mmol/L Cd2+) in Typha latifolia L. grown hydroponically were investigated in order to offer a referenced evidence for an understanding of the mechanism by which polyamines (PAs) relieve the damage to plants by heavy metal and improve the phytoremediation efficiency of heavy metal-contaminated water. The results showed that Cd2+ stress induced oxidative injury, as evidenced by an increase in the generation of superoxide anion (O2-), as well as the hydrogen peroxide (H2O2) and malondialdehyde (MDA) contents in both leaves and caudices. With the exception of superoxide dismutase (SOD) activity in the leaves, an increase in the activities of catalase (CAT), guaiacol peroxidase (GPX), and glutathione reductase (GR) was observed in both leaves and caudices, SOD activity was increased in caudices, and ascorbate peroxidase (APX) activity was increased in leaves following Cd2+ treatment. The reduced glutathione (GSH) content in both leaves and caudices and the reductive ascorbate content in leaves was obviously increased, which were prompted by the application of exogenous Spd. Spraying with Spd increased the activity of GR and APX in both leaves and caudices, whereas the activity of SOD, CAT, and GPX was increased only in caudices following spraying with Spd. The generation of O2 -. and the H2O2 and MDA content in both leaves and caudices decreased after spraying with Spd. The decrease in MDA was more obvious following the application of 0.25 than 0.50 mmol/L Spd. It is supposed that exogenous Spd elevated the tolerance of T. latifolia under Cd2+ stress primarily by increasing GR activity and the GSH level.

Effects of Exogenous Spermidine on PhotosystemⅡof Wheat Seedlings Under Water Stress

The effects of exogenous spermidine （Spd） on lipid peroxidation, relative plasma membrane permeability, photosystem Ⅱ （PSII） gene expression and PSII photochemical activity in water-stressed wheat seedlings were investigated. The decrease in relative water content （RWC）, Chl content, and 2,6-dichlorophenol indophenol （DCIP） photoreduction of PSII, and increases in electrolyte leakage of plasma membranes and malonyldialdehyde （MDA） in water-stressed leaves was alleviated by Spd pretreatment. Furthermore, Western and Northern blot analysis showed that decreases in the PSll major proteins D1, D2 and LHCII and the transcripts of corresponding genes psbA, psbD and cab were also alleviated by Spd pretreatment under water stress. These results suggest that the application of exogenous Spd protects PSII against water stress at both the transcriptional level and the translational level, and allows PSII to retain a higher activity level during water stress. The protective role of Spd in the photosynthetic apparatus also is discussed.

Atomic force microscopy observation of the condensates of the spermidine-DNA complexes

In vitro investigation on the condensation of DNA induced by naturally occurring polyamines can provide useful insights into the packaging state of DNA in vivo , which is closely related to the mechanism of the self regulation or self switching of the gene expression. The present paper reports the high resolution AFM observations of the condensates of spermidine DNA complexes. It was found that the spermidine can directly condense the DNA into toroidal structures. The formation of this special structure is greatly influenced by the dynamic factors such as the concentrations and the reaction time. Close examination revealed that the toroids were constructed by many particles stacked and squeezed together. Our observations indicate that the dynamic process of the toroid formation may be a spiral wound process of the globular subunits.

Spermidine mediated PLGA nanoparticles containing fluorofenidone for treatment of idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis(IPF) is a progressive, fatal lung disease with poor survival. The advances made in deciphering IPF have led to the approval of different anti-fibrotic molecules, such as pirfenidone and nintedanib. An increasing number of studies with particles(liposomes, nanoparticles, microspheres, nanopolymersomes and nanoliposomes) modified with different functional groups have demonstrated improvement in lung targeted drug delivery. In the present study, we prepared, characterized, and evaluated spermidine(SPD) modified poly-lactic-co-glycolide nanoparticles(SPD-AKF-PLGA/NPs) as carriers for fluorofenidone(AKF) in order to improve the anti-fibrotic efficacy of this drug in lung. SPD-AKF-PLGA/NPs were prepared and functionalized by modified solvent evaporation method with spermidine(SPD), poly-lactic-co-glycolide-polyethylene glycol(PEG-PLGA) groups. The size of SPD-AKF-PLGA/NPs was 172.5±4.3 nm. The AKF release from nanoparticles was shown to be fit Higuchi model. The A549 cellular uptake of the SPD-Cou-6-NPs group was found to be almost 2-fold higher than that of the Cou-6-NPs group. Free SPD and DFMO were pre-incubated in A549 cells to further prove that the uptake of SPD-Cou-6-PLGA/NPs via PTS. As a result, the uptake of SPD-Cou-6-PLGA/NPs significantly decreased with the increase of SPD concentrations in incubation. At the higher SPD concentrations in 50 and 500 μM, the uptake of SPD-Cou-6-PLGA/NPs reduced 0.34 and 0.49-fold than that without SPD pretreatment. After pretreatment with DFMO for 36 h, the cellular uptake of SPD-Cou-6-NPs reached 1.26-fold compared to untreated DFMO group. In biodistribution study, the drug targeting index of SPD-AKF-PLGA/NPs in lung was 3.62 and 4.66-fold than that of AKF-PLGA/NPs and AKF solution, respectively. It was suggested that the SPD-AKF-PLGA/NPs could accumulate effectively in the lung. The lung histopathology changes and collagen deposition were observed by HE staining and Masson staining in efficacy study. In SPD-AKF-PLGA/NPs group, the damage was further improved compared to that of the AKFPLGA/NPs group and AKF solution group. The results indicated that SPD-AKF-PLGA/NPs could be effective nanocarriers for antipulmonary fibrosis therapy.

Rational engineering of homospermidine synthase for enhanced catalytic efficiency toward spermidine synthesis

Spermidine is a naturally occurring polyamine widely utilized in the prevention and treatment of various diseases.Current spermidine biosynthetic methods have problems such as low efficiency and complex multi-enzyme catalysis.Based on sequence-structure-function relationships,we engineered the widely studied homospermidine synthase from Blastochloris viridis(BvHSS)and obtained mutants that could catalyze the production of spermidine from 1,3-diaminopropane and putrescine.The specific activities of BvHSS and the mutants D361E and E232D+D361E(E232D-D)were 8.72,46.04 and 48.30 U/mg,respectively.The optimal pH for both mutants was 9.0,and the optimal temperature was 50◦C.Molecular docking and dynamics simulations revealed that mutating aspartic acid at position 361 to glutamic acid narrowed the substrate binding pocket,promoting stable spermidine production.Conversely,mutating glutamic acid at position 232 to aspartic acid enlarged the substrate channel entrance,facilitating substrate entry into the active pocket and enhancing spermidine generation.In whole-cell catalysis lasting 6 h,D361E and E232D-D synthesized 725.3 and 933.5 mg/L of spermidine,respectively.This study offers a practical approach for single-enzyme catalyzed spermidine synthesis and sheds light on the crucial residues influencing homospermidine synthase catalytic activity in spermidine production.

环保型保鲜液对香石竹切花瓶插的保鲜效应

以蔗糖和8-羟基喹啉为保鲜液基本成分,研究不含银离子的瓶插保鲜液在分别添加6-苄氨基腺嘌呤(6-BA)、亚精胺(Spermidine,简称Spd)、维生素C和柠檬酸后对香石竹(Dianthus caryophyllus L.)切花的保鲜效果。结果表明,与去离子水对照相比,4种含有不同物质的保鲜液均能不同程度地增加香石竹切花的鲜重和花径,改善体内水分状况,延缓可溶性蛋白质的降解,降低丙二醛含量,延长瓶插寿命,其中以处理Ⅳ(30 g/L蔗糖+200 mg/L 8-羟基喹啉+30 mg/L 6-BA)效果最好,处理Ⅲ(30 g/L蔗糖+200mg/L 8-羟基喹啉+0.1 mmol/L Spd)次之,属于高效环保型保鲜剂。

Interacted Effect of Arbuscular Mycorrhizal Fungi and Polyamines on Root System Architecture of Citrus Seedlings

Either arbuscular mycorrhizal fungi （AMF） or polyamines （PAs） may change root system architecture （RSA） of plants, whereas the interaction of AMF and PAs on RSA remains unclear. In the present study, we studied the interaction between AMF （Paraglomus occultum） and exogenous PAs, including putrescine （Put）, spermidine （Spd） and spermine （Spin） on mycorrhizal development of different parts of root system, plant growth, RSA and carbohydrate concentrations of 6-m-old citrus （Citrus tangerine Hort. ex Tanaka） seedlings. After 14 wk of PAs application, PA-treated mycorrhizal seedlings exhibited better mycorrhizal colonization and numbers of vesicles, arbuscules, and entry points, and the best mycorrhizal status of taproot, first-, second-, and third-order lateral roots was respectively found in mycorrhizal seedlings supplied with Put, Spd and Spm, suggesting that PAs might act as a regulated factor of mycorrhizal development through transformation of root sucrose more into glucose for sustaining mycorrhizal development. AMF usually notably increases RSA traits （taproot length, total length, average diameter, projected area, surface area, volume, and number of first-, second-, and third-order lateral roots） of only PA-treated seedlings. Among the three PA species, greater positive effects on RSA change and plant biomass increment of the seedlings generally rank as Spd〉Spm〉Put, irrespective of whether or not AMF colonization. PAs significantly changed the RSA traits in mycorrhizal but not in non-mycorrhizal seedlings. It suggests that the application of PAs （especially Spd） to AMF plants would optimize RSA of citrus seedlings, thus increasing plant growth （shoot and root dry weight）.

Effect of diethylstilbestrol on polyamine metabolism in hamster epididymis

<abstract>Aim: To investigate the effect of diethylstilbestrol (DES), one of the most potent endocrine disruptors, on the metabolism of polyamines in hamster epididymis. Methods: Male golden hamsters of 7-week-old were kept under a light and dark cycle of 14 h and 10 h for 1 week to stimulate maximally the gonadal function. DES was injected subcutaneously at doses of 0.01 mg·kg-1·day-1, 0.1 mg·kg-1·day-1 and 1 mg·kg-1·day-1 for one week. Results: DES treatment caused a significant decrease in the weight of epididymis. The activity of epididymal ornithine decar boxylase (ODC) increased 1 day after DES treatment, kept at a high level for 4 days and then decreased to nearly normal level at day 7. The activity of spermidine/spermine N1-acetyltransferase (SSAT) also increased transiently after DES treatment. The contents of putrescine, spermidine, spermine and N1-acetylspermidine were increased 1 day -4 days after DES treatment and restored to normal at day 7. All these changes showed a marked difference between the caput and the cauda. Conclusion: The polyamine biosynthesis in the hamster epididymis can be affected by DES, a xenoestrogen. DES may probably affect polyamine metabolism in the epididymis by regulating the rate-limiting enzymes involved in the polyamine biosynthesis.

Successful Agrobacterium-mediated transformation of Populus tomentosa with apple SPDS gene

The problem of salinized soils has become one of the most serious constraints to agricultural and forest productivity. With the purpose of enhancing salt stress tolerance of Populus tomentosa, we transformed this tree species with spermidine synthase （SPDS） genes derived from an apple by an Agrobacterium-mediated method. Four transgenic clones were confu＇med by PCR and Southern blot analysis. As well, the expression of introduced SPDS genes was analyzed by real-time quantitative PCR.

High Performance Liquid Chromatography Separation and Simultaneous Fluorometric Detection of Trace Amount of Polyamines in Cell Culture Media

A highly sensitive and precise method for the determination of the polyamines (including putrescine, spermidine and spermine) in cell culture media is described. The samples were concentrated by C18 column and the polyamines were converted to fluorometric derivatives with DNS-Cl. The polyamine derivatives were then completely separated by HPLC and determined by simultaneous fluorometric detection. The CV of intragroup and intergroup were 2. 49 %-4. 26% and 4. 29 %-5. 16 %, respectively. The rate of recovery was 103%- 99%. There was trace amount of polyamines detected by this method in the media of F12, 8900, RPMI1640, DMEM and M199 even without incubation with cells. So this method can be used for detecting the changes of polyamines in a medium before and after incubation with cells. It is helpful for the researches on the regulation of cell proliferation by polyamines.

Polyamines and polyamine-metabolizing enzymes in schizophrenia: Current knowledge and concepts of therapy

Polyamines play preeminent roles in a variety of cellular functions in the central nervous system and other organs.A large body of evidence suggests that the polyamine pathway is prominently involved in the etiology and pathology of schizophrenia.Alterations in the expression and activity of polyamine metabolizing enzymes,as well as changes in the levels of the individual polyamines,their precursors and derivatives,have been measured in schizophrenia and animal models of the disease.Additionally,neuroleptic treatment has been shown to influence polyamine concentrations in brain and blood of individuals with schizophrenia.Thus,the polyamine system may appear to be a promising target for neuropharmacological treatment of schizophrenia.However,for a number of practical reasons there is currently only limited hope for a polyamine-based schizophrenia therapy.

Changes in the Ovarian Polyamine Content and Seed Set Efficiency of Cotton by the Plant Growth Regulator BM86

Naturally occurring polyamines in plants have been implicated in floral initiation, and fruit development. The plant growth regulator BM86 was formulated to stimulate seed production and fruit growth by enhancing polyamine synthe-sis. The objectives of this study were to determine the effect of BM86 on free polyamine content in cotton (Gos-sypium hirsutum L.) ovaries and on seed set efficiency, and also to investigate genotypic differences in ovarian polyamine content. BM86 was applied at the first flower stage and two weeks later at 2.34 mL/ha. This study showed that application of BM86 had a significant positive effect on ovarian polyamine content of cotton. Putrescine and sper-midine one week after the 1st BM86 application and putrescine two week after the 1st BM86 application, were significantly increased compared to the untreated control. Higher seed set efficiency with the BM86 application was observed when the total number of seeds was used for the calculation. However, when the number of harvestable seeds was used to calculate seed set efficiency BM86 had no significant effect. In addition, application of BM86 did not significantly alter seedcotton yield of the crop. Only small differences in ovarian polyamine content were detected among the genotypes tested, possibly due to the narrow genetic pool of the commercial cotton genotypes used. Application of BM86 can significantly increase cotton seed number by enhancing polyamines biosynthesis, but further research is needed to determine how to capitalize on the increased potential number of harvestable seeds.