Genetic Engineering and Genome Editing Advances to Enhance Floral Attributes in Ornamental Plants: An Update.

The ornamental horticulture industry is a highly dynamic and rapidly changing market. Constant development of novel cultivars with elite traits is essential to sustain competitiveness. Conventional breeding has been used to develop cultivars, which is often laborious. Biotechnological strategies such as genetic engineering have been crucial in manipulating and improving various beneficial traits that are technically not possible through cross-breeding. One such trait is the highly desired blue-colored flower in roses and chrysanthemums, which can be achieved through transgenic technology. Advances in genome sequencing platforms have enhanced the opportunities to access the whole genome sequence in various ornamentals, facilitating the dissection of the molecular genetics and regulatory controls of different traits. The recent advent of genome editing tools, including CRISPR/Cas9, has revolutionized plant breeding. CRISPR/Cas9-based gene editing offers efficient and highly precise trait modification, contributing to various beneficial advancements. Although genome editing in ornamentals is currently in its infancy, the recent increase in the availability of ornamental genome sequences provides a platform to extend the frontiers of future genome editing in ornamentals. Hence, this review depicts the implication of various commercially valuable ornamental attributes, and details the research attempts and achievements in enhancing floral attributes using genetic engineering and genome editing in ornamental plants.

Analysis of Floral Scent and Volatile Profiles of Different Aster Species by E-nose and HS-SPME-GC-MS.

Plants from the Aster species are known to be a rich source of bioactive chemical compositions and are popularly known for their medicinal properties. To investigate the relationship between the nine species of Aster, the floral fragrance and volatile profile patterns were characterized using E-nose and HS-SPME-GC-MS. Initial optimization for fragrance analysis was performed with Aster yomena using E-nose by evaluating the scent patterns in different flowering stages. Aster yomena exhibited varied scent patterns in each flowering stage, with the highest relative aroma intensity (RAI) in the full flowering stage. PCA analysis to compare and analyze the scent characteristics of nine Aster species, showed a species-specific classification. HS-SPME-GC-MS analysis of flowers from nine Aster species revealed 52 volatile compounds including ß-myrcene, α-phellandrene, D-limonene, trans-ß-ocimene, caryophyllene, and ß-cadinene. The terpenoid compounds accounted for the largest proportion. Among the nine Aster species flowers, Aster koraiensis had sesquiterpenes as the major component, and the remaining eight varieties had monoterpenes in abundance. These results could distinguish the species according to the scent patterns and volatile components of the nine Aster species. Additionally, flower extracts from the Aster species' plants exhibited radical scavenging antioxidant activity. Among them, it was confirmed that Aster pseudoglehnii, Aster maackii, and Aster arenarius had high antioxidant activity. In conclusion, the results of this study provide fundamental data of the volatile compound properties and antioxidant activity of Aster species, offering basic information of valuable natural sources that can be utilized in the pharmaceutical, perfume, and cosmetic industries.

Towards the Improvement of Ornamental Attributes in Chrysanthemum: Recent Progress in Biotechnological Advances.

Incessant development and introduction of novel cultivars with improved floral attributes are vital in the dynamic ornamental industry. Chrysanthemum (Chrysanthemum morifolium) is a highly favored ornamental plant, ranking second globally in the cut flower trade, after rose. Development of new chrysanthemum cultivars with improved and innovative modifications in ornamental attributes, including floral color, shape, plant architecture, flowering time, enhanced shelf life, and biotic and abiotic stress tolerance, is a major goal in chrysanthemum breeding. Despite being an economically important ornamental plant, the application of conventional and molecular breeding approaches to various key traits of chrysanthemum is hindered owing to its genomic complexity, heterozygosity, and limited gene pool availability. Although classical breeding of chrysanthemum has resulted in the development of several hundreds of cultivars with various morphological variations, the genetic and transcriptional control of various important ornamental traits remains unclear. The coveted blue colored flowers of chrysanthemums cannot be achieved through conventional breeding and mutation breeding due to technical limitations. However, blue-hued flower has been developed by genetic engineering, and transgenic molecular breeding has been successfully employed, leading to substantial progress in improving various traits. The recent availability of whole-genome sequences of chrysanthemum offers a platform to extensively employ MAS to identify a large number of markers for QTL mapping, and GWAS to dissect the genetic control of complex traits. The combination of NGS, multi-omic platforms, and genome editing technologies has provided a tremendous scope to decipher the molecular and regulatory mechanisms. However, the application and integration of these technologies remain inadequate for chrysanthemum. This review, therefore, details the significance of floral attributes, describes the efforts of recent advancements, and highlights the possibilities for future application towards the improvement of crucial ornamental traits in the globally popular chrysanthemum plant.

Impact of lattice versus solid structure of 3D-printed multiroot dental implants using Ti-6Al-4V: a preclinical pilot study.

PURPOSE: Various studies have investigated 3-dimensional (3D)-printed implants using Ti-6Al-4V powder; however, multi-root 3D-printed implants have not been fully investigated. The purpose of this study was to explore the stability of multirooted 3D-printed implants with lattice and solid structures. The secondary outcomes were comparisons between the 2 types of 3D-printed implants in micro-computed tomographic and histological analyses. METHODS: Lattice- and solid-type 3D-printed implants for the left and right mandibular third premolars in beagle dogs were fabricated. Four implants in each group were placed immediately following tooth extraction. Implant stability measurement and periapical X-rays were performed every 2 weeks for 12 weeks. Peri-implant bone volume/tissue volume (BV/TV) and bone mineral density (BMD) were measured by micro-computed tomography. Bone-to-implant contact (BIC) and bone area fraction occupancy (BAFO) were measured in histomorphometric analyses. RESULTS: All 4 lattice-type 3D-printed implants survived. Three solid-type 3D-printed implants were removed before the planned sacrifice date due to implant mobility. A slight, gradual increase in implant stability values from implant surgery to 4 weeks after surgery was observed in the lattice-type 3D-printed implants. The marginal bone change of the surviving solid-type 3D-printed implant was approximately 5 mm, whereas the value was approximately 2 mm in the lattice-type 3D-printed implants. BV/TV and BMD in the lattice type 3D-printed implants were similar to those in the surviving solid-type implant. However, BIC and BAFO were lower in the surviving solid-type 3D-printed implant than in the lattice-type 3D-printed implants. CONCLUSIONS: Within the limits of this preclinical study, 3D-printed implants of double-rooted teeth showed high primary stability. However, 3D-printed implants with interlocking structures such as lattices might provide high secondary stability and successful osseointegration.

Recent Progress in Enhancing Fungal Disease Resistance in Ornamental Plants.

Fungal diseases pose a major threat to ornamental plants, with an increasing percentage of pathogen-driven host losses. In ornamental plants, management of the majority of fungal diseases primarily depends upon chemical control methods that are often non-specific. Host basal resistance, which is deficient in many ornamental plants, plays a key role in combating diseases. Despite their economic importance, conventional and molecular breeding approaches in ornamental plants to facilitate disease resistance are lagging, and this is predominantly due to their complex genomes, limited availability of gene pools, and degree of heterozygosity. Although genetic engineering in ornamental plants offers feasible methods to overcome the intrinsic barriers of classical breeding, achievements have mainly been reported only in regard to the modification of floral attributes in ornamentals. The unavailability of transformation protocols and candidate gene resources for several ornamental crops presents an obstacle for tackling the functional studies on disease resistance. Recently, multiomics technologies, in combination with genome editing tools, have provided shortcuts to examine the molecular and genetic regulatory mechanisms underlying fungal disease resistance, ultimately leading to the subsequent advances in the development of novel cultivars with desired fungal disease-resistant traits, in ornamental crops. Although fungal diseases constitute the majority of ornamental plant diseases, a comprehensive overview of this highly important fungal disease resistance seems to be insufficient in the field of ornamental horticulture. Hence, in this review, we highlight the representative mechanisms of the fungal infection-related resistance to pathogens in plants, with a focus on ornamental crops. Recent progress in molecular breeding, genetic engineering strategies, and RNAi technologies, such as HIGS and SIGS for the enhancement of fungal disease resistance in various important ornamental crops, is also described.

Effects of hyaluronic acid and deproteinized bovine bone mineral with 10% collagen for ridge preservation in compromised extraction sockets.

BACKGROUND: The aim of this study was to evaluate the efficacy of deproteinized bovine bone mineral with 10% collagen (DBBM-C) soaked with hyaluronic acid (HA) for ridge preservation in compromised extraction sockets. METHODS: Bilateral third, fourth premolars and first molar were hemisected, distal roots were extracted, and then combined endodontic periodontal lesion was induced in the remaining mesial roots. After 4 months, the mesial roots were extracted and the following four treatments were randomly performed: Absorbable collagen sponge (ACS), ACS soaked with HA (ACS+HA), ridge preservation with DBBM-C covered with a collagen membrane (RP), ridge preservation with DBBM-C mixed with HA and covered with a collagen membrane (RP+HA). Animals were sacrificed at 1 and 3 months following treatment. Ridge dimensional changes and bone formation were examined using microcomputed tomography, histology, and histomorphometry. RESULTS: At 1 month, ridge width was significantly higher in the RP and RP+HA groups than in the ACS and ACS+HA groups, while the highest proportion of mineralized bone was observed in ACS+HA group. At 3 months, ridge width remained significantly higher in the RP and RP+HA groups than in the ACS and ACS+HA groups. ACS+HA and RP+HA treatments featured the highest proportion of mineralized bone and bone volume density compared with the other groups. No statistical difference was observed between ACS+HA and RP+HA treatments. CONCLUSIONS: Ridge preservation with the mixture DBBM-C/HA prevented dimensional shrinkage and improved bone formation in compromised extraction sockets at 1 and 3 months.

Is ridge preservation effective in the extraction sockets of periodontally compromised teeth? A randomized controlled trial.

OBJECTIVE: To verify whether ridge preservation is effective in the reduction of dimensional loss and in bone formation compared to spontaneous healing in extraction sockets of periodontally compromised teeth. METHODS: Twenty-six subjects requiring tooth extraction for stage III/IV periodontitis were randomly assigned to one of two interventions: alveolar ridge preservation using collagenated bovine bone mineral and a resorbable collagen membrane (test, RP) or spontaneous healing (control, SH). Six months later, postoperative cone-beam computed tomography (CBCT) was performed to measure the linear and volumetric changes of the sockets compared to baseline scans. Biopsies were retrieved at the implant site for histomorphometric calculations. Nonparametric tests were applied for statistical analysis. RESULTS: Significantly less shrinkage occurred in RP compared to SH, mainly in the crestal zone. The width loss difference between groups was 3.3 mm and 2.2 mm at 1 mm and 3 mm below the crest, respectively (p < .05). RP yielded a gain in socket height of 0.25 mm, whereas a loss of -0.39 mm was observed in SH (p < .05). The percentage of volume loss recorded in RP was also less than that recorded in SH (-26.53% vs -50.34, p < .05). Significantly less bone proportion was detected in biopsies from RP (30.1%) compared with SH (53.9%). A positive association between baseline bone loss and ridge shrinkage was found in SH but not in RP. CONCLUSION: Ridge preservation in extraction sockets of periodontally compromised teeth was effective in reducing the amount of ridge resorption.

Diagnostic Models for Screening of Periodontitis with Inflammatory Mediators and Microbial Profiles in Saliva.

This study aims to investigate and assess salivary biomarkers and microbial profiles as a means of diagnosing periodontitis. A total of 121 subjects were included: 28 periodontally healthy subjects, 24 with Stage I periodontitis, 24 with Stage II, 23 with Stage III, and 22 with Stage IV. Salivary proteins (including active matrix metalloproteinase-8 (MMP-8), pro-MMP-8, total MMP-8, C-reactive protein, secretory immunoglobulin A) and planktonic bacteria (including Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Fusobacterium nucleatum, Prevotella intermedia, Porphyromonas nigrescens, Parvimonas micra, Campylobacter rectus, Eubacterium nodatum, Eikenella corrodens, Streptococcus mutans, Staphylococcus aureus, Enterococcus faecalis, and Actinomyces viscosus) were measured from salivary samples. The performance of the diagnostic models was assessed by receiver operating characteristics (ROCs) and area under the ROC curve (AUC) analysis. The diagnostic models were constructed based on the subjects' proteins and/or microbial profiles, resulting in two potential diagnosis models that achieved better diagnostic powers, with an AUC value > 0.750 for the diagnosis of Stages II, III, and IV periodontitis (Model PA-I; AUC: 0.796, sensitivity: 0.754, specificity: 0.712) and for the diagnosis of Stages III and IV periodontitis (Model PA-II; AUC: 0.796, sensitivity: 0.756, specificity: 0.868). This study can contribute to screening for periodontitis based on salivary biomarkers.

Healing kinetics of oral soft tissue wounds treated with recombinant epidermal growth factor: Translation from a canine model.

OBJECTIVE: To test whether or not topically administered recombinant human epidermal growth factor (rhEGF) accelerates the early healing phase of oral soft tissue wounds. METHODS: One day following the creation of palatal defects (n = 6/animal), 14 dogs were allocated to one of the following five groups: spontaneous healing (SH), vehicle ointment (V), vehicle ointment + rhEGF at concentrations of 1 µg/g (EGF1), 10 µg/g (EGF10) or 50 µg/g (EGF50). Topical administration of ointments was repeated twice per day until sacrifice at days 8 and 16. Wound area was clinically monitored. Keratinocytes proliferation (Ki67-immunolabelling), inflammatory response (IR) and areas of collagen (C) and granulation tissue (GT) were histologically measured. Kruskal-Wallis test with Dunnett correction was used for multiple group statistical comparisons. RESULTS: Clinically, in comparison with SH, a significantly smaller wound area was observed in groups EGF1 and EGF10 at day 8 (p < 0.05). At day 16, wound closure reached 97.8% in group EGF1 compared to 83.2% in group SH, albeit no statistically different. Histologically, at day 8, significantly more GT was observed in group EGF10 compared to all other groups (p < 0.05). At day 16, in addition to a higher Ki67-immunolabelling, groups EGF1 and EGF10 demonstrated a significant decrease in GT and IR with more deposition of C compared to the other groups (p < 0.05). CONCLUSION: Application of rhEGF enhanced the early healing of acute oral soft tissue wounds compared to SH, predominantly at concentrations of 1 and 10 µg/g.

Ridge preservation of extraction sockets with chronic pathology using Bio-Oss® Collagen with or without collagen membrane: an experimental study in dogs.

OBJECTIVES: This study aimed to evaluate the dynamics of newly bone formation and dimensional change in diseased extraction sockets using Bio-Oss® Collagen with or without a collagen membrane. MATERIAL AND METHODS: In six beagle dogs, right and left 3rd and 4th mandibular premolars were hemisected and the distal roots were removed. Combined endodontic-periodontic lesions were induced in all sites using black silk, collagen sponge, endodontic files, and application of Porphyromonas gingivalis. After 4 months, among 4 premolars, three teeth were randomly selected per dog and allocated to the following experimental groups: Control group (no treatment but debridement), Test 1 group (only Bio-Oss® Collagen graft), and Test 2 group (Bio-Oss® Collagen graft with a collagen membrane). After 7 months from the baseline, the beagle dogs were sacrificed for histomorphometric and Micro-CT analysis. RESULTS: The vertical distance between buccal and lingual crests in the Control group (2.22 ± 0.26 mm) and Test 2 group (1.80 ± 0.16 mm) was significantly different. The socket of the Test 2 group (27.04 ± 5.25%) was occupied by a greater quantity of bone graft compared to the Test 1 group (18.49 ± 2.11%). CONCLUSION: Ridge preservation in diseased extraction sockets could compensate for buccal bone resorption by contact osteogenesis surrounding the bone graft particles at the bucco-coronal area during socket healing, and the application of a collagen membrane at the entrance of the socket is useful for preserving graft material at the coronal part of the socket.

In Vivo Inhibition of Porphyromonas gingivalis Growth and Prevention of Periodontitis With Quorum-Sensing Inhibitors.

BACKGROUND: Autoinducer (AI)-2 has an important role in biofilm formation in the oral environment. Mature biofilms formed as a result of the cell-to-cell communication make it difficult to overcome periodontitis with the use of antibiotics. Previous in vitro studies suggest that quorum-sensing inhibitors (QSIs) interfere with AI-2. This study compares the QSI effects resulting from an oral inoculation of Porphyromonas gingivalis in an experimental animal model. METHODS: Forty-five male mice were divided into three groups (n = 15 each): 1) infection; 2) QSI; and 3) control. Infection and QSI groups received oral inoculation of P. gingivalis, whereas treatment with QSIs (furane compound and d-ribose) was only performed in the QSIs group. The control group was a negative control not receiving manipulation. After 42 days, mice were sacrificed, and the distance from the alveolar bone crest (ABC) to the cemento-enamel junction (CEJ) was measured by microcomputed tomography. P. gingivalis DNA was quantified in the soft and hard tissues around the molar teeth by real-time polymerase chain reaction. RESULTS: Distance from ABC to CEJ was significantly increased in the P. gingivalis infection group compared with the control group (P = 0.02) and significantly decreased in the QSI group compared with the infection group (P = 0.02). The QSI group contained 31.64% of the bacterial DNA count of the infection group. CONCLUSION: Use of QSIs in the mice infection model showed a reduction of bone breakdown and a decrease in the number of bacteria in vivo, suggesting that QSIs can be a new approach to prevention and treatment of periodontitis.

Hyaluronic Acid Improves Bone Formation in Extraction Sockets With Chronic Pathology: A Pilot Study in Dogs.

BACKGROUND: Previous studies on ridge preservation focusing on fresh extraction sockets using graft materials for ridge preservation procedures have reported a delay in the tissue modeling and remodeling phases. The objective of this study is to evaluate the effect of hyaluronic acid (HA) on healing of infected sockets. METHODS: Six beagle dogs were used in this study. Both mandibular third premolars were hemisected, and the distal roots were extracted. Subsequently, periodontal and endodontic lesions were induced at the remaining mesial root. After communication of the periodontal lesion, an endodontic periapical lesion was observed at 4 months, and the mesial roots of both the right and left sides were extracted. HA was applied into the socket of the test group, and no treatment was administered to the other group (control group). Three months after extraction of the mesial roots, the dogs were sacrificed, and histologic evaluations were performed. RESULTS: The sockets were filled by mineralized bone (47.80% ± 6.60%) and bone marrow (50.47% ± 6.38%) in the control group, whereas corresponding values were 63.29% ± 9.78% and 34.73% ± 8.97% for the test group, respectively. There was a statistically significant difference between the groups. Reversal lines and a copious lineup of osteoblasts were observed in the middle and apical parts of the sockets in the test group. CONCLUSION: An infected socket shows delayed healing of the socket wound, and HA, because of its osteoinductive, bacteriostatic, and anti-inflammatory properties, may improve bone formation and accelerate wound healing in infected sockets.