Hyperdiverse tropical rainforests, such as the aseasonal forests in Southeast Asia, are supported... more Hyperdiverse tropical rainforests, such as the aseasonal forests in Southeast Asia, are supported by high annual rainfall. Its canopy is dominated by the species-rich tree family of Dipterocarpaceae (Asian dipterocarps), which has both ecological (e.g., supports flora and fauna) and economical (e.g., timber production) importance. Recent ecological studies suggested that rare irregular drought events may be an environmental stress and signal for the tropical trees. We assembled the genome of a widespread but near threatened dipterocarp, Shorea leprosula, and analyzed the transcriptome sequences of ten dipterocarp species representing seven genera. Comparative genomic and molecular dating analyses suggested a whole-genome duplication close to the Cretaceous-Paleogene extinction event followed by the diversification of major dipterocarp lineages (i.e. Dipterocarpoideae). Interestingly, the retained duplicated genes were enriched for genes upregulated by no-irrigation treatment. These ...
Worldwide, many mangrove species are experiencing significant population declines, includingRhizo... more Worldwide, many mangrove species are experiencing significant population declines, includingRhizophora apiculata, which is one of the most widespread and economically important species in tropical Asia. In Malaysia, there has been an alarming decline inR. apiculatapopulations driven primarily by anthropogenic activities. However, the lack of genetic and demographic information on this species has hampered local efforts to conserve it. To address these gaps, we generated novel genetic information forR. apiculata, based on 1,120 samples collected from 39 natural populations in Peninsular Malaysia. We investigated its genetic diversity and genetic structure with 19 transcriptome and three nuclear microsatellite markers. Our analyses revealed a low genetic diversity (meanHe: 0.352) with significant genetic differentiation (FST: 0.315) among populations ofR. apiculata. Approximately two-third of the populations showed significant excess of homozygotes, indicating persistent inbreeding wh...
Our study highlights the utilization of a genetic database for wood-origin identification in Ints... more Our study highlights the utilization of a genetic database for wood-origin identification in Intsia palembanica, a valuable heavy hardwood from the tropical forests. This forensic tool is essential for strengthening the verification of legality in the wood supply chain from the forest to the end-users. An increasing number of rules and regulations are being put in place to promote sustainable practice in the timber trade, one of which involves ensuring that importers declare the correct species name and source of geographic origin of the timber. We aimed to determine the origin of the I. palembanica seed source used in the early establishment on the Forest Research Institute Malaysia (FRIM) campus. DNA samples of I. palembanica individuals from the FRIM campus were obtained and analyzed using four chloroplast (cp) DNA markers to characterize the haplotype variants for population identification. In addition, the DNA samples were also genotyped at 14 short tandem repeat (STR) loci for...
To inform product users about the origin of timber, the implementation of a traceability system i... more To inform product users about the origin of timber, the implementation of a traceability system is necessary for the forestry industry. In this study, we developed a comprehensive genetic database for the important tropical timber species Merbau, Intsia palembanica, to trace its geographic origin within peninsular Malaysia. A total of 1373 individual trees representing 39 geographically distinct populations of I. palembanica were sampled throughout peninsular Malaysia. We analyzed the samples using a combination of four chloroplast DNA (cpDNA) markers and 14 short tandem repeat (STR) markers to establish both cpDNA haplotype and STR allele frequency databases. A haplotype map was generated through cpDNA sequencing for population identification, resulting in six unique haplotypes based on 10 informative intraspecifically variable sites. Subsequently, an STR allele frequency database was developed from 14 STRs allowing individual identification. Bayesian cluster analysis divided the individuals into two genetic clusters corresponding to the northern and southern regions of peninsular Malaysia. Tests of conservativeness showed that the databases were conservative after the adjustment of the θ values to 0.2000 and 0.2900 for the northern (f = 0.0163) and southern (f = 0.0285) regions, respectively. Using self-assignment tests, we observed that individuals were correctly assigned to populations at rates of 40.54-94.12% and to the identified regions at rates of 79.80-80.62%. Both the cpDNA and STR markers appear to be useful for tracking Merbau timber originating from peninsular Malaysia. The use of these forensic tools in addition to the existing paper-based timber tracking system will help to verify the legality of the origin of I. palembanica and to combat illegal logging issues associated with the species.
The development of timber tracking methods based on genetic markers can provide scientific eviden... more The development of timber tracking methods based on genetic markers can provide scientific evidence to verify the origin of timber products and fulfill the growing requirement for sustainable forestry practices. In this study, the origin of an important Dark Red Meranti wood, Shorea platyclados, was studied by using the combination of seven chloroplast DNA and 15 short tandem repeats (STRs) markers. A total of 27 natural populations of S. platyclados were sampled throughout Malaysia to establish population level and individual level identification databases. A haplotype map was generated from chloroplast DNA sequencing for population identification, resulting in 29 multilocus haplotypes, based on 39 informative intraspecific variable sites. Subsequently, a DNA profiling database was developed from 15 STRs allowing for individual identification in Malaysia. Cluster analysis divided the 27 populations into two genetic clusters, corresponding to the region of Eastern and Western Malays...
Background: Dipterocarpaceae is a pantropical tree family that plays an important role in our und... more Background: Dipterocarpaceae is a pantropical tree family that plays an important role in our understanding of the ecology of Asian tropical rain forests. However, genome sizes for members of the Dipterocarpaceae are still poorly known. Aims: To report the genome size of 115 dipterocarp species and examine the variation and evolution of genome size in this family. Methods: Genome size was estimated using flow cytometry. Both the rpoB and trnL intron were sequenced to uncover the evolution of genome size within a phylogenetic framework. Results: The 1C genome size varied between 0.267 and 0.705 pg in Shorea hemsleyana and Shorea ovalis, respectively, a 2.64-fold variation across the family. Most dipterocarps are characterised by very small genomes with a mean 1C value of 0.416 pg (sd = 0.075) and five polyploids are recorded. The ancestral genome size for dipterocarps was reconstructed as 1Cx = 0.481 pg (95% CI = 0.433–0.534). Conclusions: Genome size variation in dipterocarps was characterised by very small values with a narrow range. Overall, genome size reduction from the ancestral state is a general trend in Dipterocarpaceae.
Abstract Taxonomic classification of the important medicinal plant, Ficus deltoidea (Moraceae), i... more Abstract Taxonomic classification of the important medicinal plant, Ficus deltoidea (Moraceae), is challenging because of the variability of its leaves and fig forms that occur within the species. We developed 16 nuclear Simple Sequence Repeat (nSSR) markers, and characterized them using 24 individuals from a natural population. We then studied the intraspecific variation of F . deltoidea subsp. deltoidea in Peninsular Malaysia using morphological and molecular approaches. Based on the morphological variations, we further determined the varieties that occur regionally under the above subspecies based on the leaf characteristics. As for molecular data sets, we used both chloroplast DNA (cpDNA) and nSSR markers to elucidate the phylogenetic relationship among the varieties. The cpDNA dendrogram yielded poorer resolution where most of the clades were forming paraphyletic complex. The cluster analysis based on nSSR is largely congruent with the morphological classification, with F . deltoidea subsp. deltoidea classified into four main varieties, namely var. deltoidea , var. angustifolia , var. kunstleri and var. lutescens . Our study demonstrates the applicability of molecular approach in complementing the conventional taxonomic classification.
Illegal logging and smuggling of Gonystylus bancanus (Thymelaeaceae) poses a serious threat to th... more Illegal logging and smuggling of Gonystylus bancanus (Thymelaeaceae) poses a serious threat to this fragile valuable peat swamp timber species. Using G. bancanus as a case study, DNA markers were used to develop identification databases at the species, population and individual level. The species level database for Gonystylus comprised of an rDNA (ITS2) and two cpDNA (trnH-psbA and trnL) markers based on a 20 Gonystylus species database. When concatenated, taxonomic species recognition was achieved with a resolution of 90% (18 out of the 20 species). In addition, based on 17 natural populations of G. bancanus throughout West (Peninsular Malaysia) and East (Sabah and Sarawak) Malaysia, population and individual identification databases were developed using cpDNA and STR markers respectively. A haplotype distribution map for Malaysia was generated using six cpDNA markers, resulting in 12 unique multilocus haplotypes, from 24 informative intraspecific variable sites. These unique haplotypes suggest a clear genetic structuring of West and East regions. A simulation procedure based on the composition of the samples was used to test whether a suspected sample conformed to a given regional origin. Overall, the observed type I and II errors of the databases showed good concordance with the predicted 5% threshold which indicates that the databases were useful in revealing provenance and establishing conformity of samples from West and East Malaysia. Sixteen STRs were used to develop the DNA profiling databases for individual identification. Bayesian clustering analyses divided the 17 populations into two main genetic clusters, corresponding to the regions of West and East Malaysia. Population substructuring (K=2) was observed within each region. After removal of bias resulting from sampling effects and population subdivision, conservativeness tests showed that the West and East Malaysia databases were conservative. This suggests that both databases can be used independently for random match probability estimation within respective regions. The reliability of the databases was further determined by independent self-assignment tests based on the likelihood of each individual's multilocus genotype occurring in each identified population, genetic cluster and region with an average percentage of correctly assigned individuals of 54.80%, 99.60% and 100% respectively. Thus, after appropriate validation, the genetic identification databases developed for G. bancanus in this study could support forensic applications and help safeguard this valuable species into the future.
Neobalanocarpus heimii, locally known as chengal, is an important timber species in Peninsular Ma... more Neobalanocarpus heimii, locally known as chengal, is an important timber species in Peninsular Malaysia. Owing to the high demand for its valuable timber, N. heimii is subjected to illegal logging and this species may become endangered in the near future. The present study was designed to identify a set of highly polymorphic short tandem repeat (STR) markers for timber tracking of N. heimii. An extensive evaluation of 51 STRs developed for Dipterocarpaceae managed to identify 12 STR loci (Nhe004, Nhe005, Nhe011, Nhe015, Nhe018, Hbi161, Sle392, Sle605, Slu044a, Shc03, Shc04 and Shc07), which showed specific amplification, high polymorphism, single-locus mode of inheritance, absence of null alleles and absence of mononucleotide repeat motifs in N. heimii. These loci can be readily used to establish a linkage between the evidentiary sample and the source, thus providing a useful set of markers for individual identification in N. heimii.
Microsatellite markers were developed for Johannesteijsmannia lanceolata to assess the genetic di... more Microsatellite markers were developed for Johannesteijsmannia lanceolata to assess the genetic diversity and mating system of this alarmingly endangered species. A total of 31 polymorphic microsatellite markers were developed for J. lanceolata using the enrichment protocol. These markers were screened on 24 samples from a natural population. The number of alleles ranged from four to 20, while the observed heterozygosity ranged from 0.391 to 1.000. The 31 loci were further tested for transferability on J. altifrons, J. magnifica, and J. perakensis. Generally, all loci showed positive amplifications in these three Johannesteijsmannia species, except Jla124 (J. magnifica) and Jla168b (J. magnifica and J. perakensis). These microsatellite markers could be employed to study the population genetics and mating system of J. lanceolata and other Johannesteijsmannia species.
Aggressive collections and trade activities in recent decades have resulted in heavy pressure on ... more Aggressive collections and trade activities in recent decades have resulted in heavy pressure on the natural stands of Aquilaria malaccensis and concerns over its long-term survival potential. To aid DNA profiling and assessment of its genetic diversity, microsatellite markers were developed for the species. Seventeen polymorphic microsatellite markers were developed for A. malaccensis using an enrichment protocol. The markers were screened on 24 samples from a natural population. The number of alleles ranged from two to 11, and the observed heterozygosity ranged from 0.042 to 0.957. No significant deviation from Hardy-Weinberg equilibrium was detected after conservative Bonferroni correction. This is the first report on the development of microsatellite markers in A. malaccensis. The markers will be used to establish a DNA profiling database and to estimate the genetic diversity and population genetic structure of the species.
Begonias are hyper-diverse and important horticultural plants. Six polymorphic microsatellite mar... more Begonias are hyper-diverse and important horticultural plants. Six polymorphic microsatellite markers were developed from CT- and GT-enriched libraries of Begonia maxwelliana. The number of alleles per locus ranged from 2 to 12 and the observed heterozygosity ranged from 0.036 to 0.813. Null alleles were detected in one locus (Bma161) after Bonferroni correction. All the six markers were amplifiable in 23 selected Begonia species with the success rates of 17-100%. On average, species of the same section as B. maxwelliana (i.e. sect. Platycentrum) yielded higher transferability (91%). These markers will be useful for population genetic studies of the genus Begonia.
Hyperdiverse tropical rainforests, such as the aseasonal forests in Southeast Asia, are supported... more Hyperdiverse tropical rainforests, such as the aseasonal forests in Southeast Asia, are supported by high annual rainfall. Its canopy is dominated by the species-rich tree family of Dipterocarpaceae (Asian dipterocarps), which has both ecological (e.g., supports flora and fauna) and economical (e.g., timber production) importance. Recent ecological studies suggested that rare irregular drought events may be an environmental stress and signal for the tropical trees. We assembled the genome of a widespread but near threatened dipterocarp, Shorea leprosula, and analyzed the transcriptome sequences of ten dipterocarp species representing seven genera. Comparative genomic and molecular dating analyses suggested a whole-genome duplication close to the Cretaceous-Paleogene extinction event followed by the diversification of major dipterocarp lineages (i.e. Dipterocarpoideae). Interestingly, the retained duplicated genes were enriched for genes upregulated by no-irrigation treatment. These ...
Worldwide, many mangrove species are experiencing significant population declines, includingRhizo... more Worldwide, many mangrove species are experiencing significant population declines, includingRhizophora apiculata, which is one of the most widespread and economically important species in tropical Asia. In Malaysia, there has been an alarming decline inR. apiculatapopulations driven primarily by anthropogenic activities. However, the lack of genetic and demographic information on this species has hampered local efforts to conserve it. To address these gaps, we generated novel genetic information forR. apiculata, based on 1,120 samples collected from 39 natural populations in Peninsular Malaysia. We investigated its genetic diversity and genetic structure with 19 transcriptome and three nuclear microsatellite markers. Our analyses revealed a low genetic diversity (meanHe: 0.352) with significant genetic differentiation (FST: 0.315) among populations ofR. apiculata. Approximately two-third of the populations showed significant excess of homozygotes, indicating persistent inbreeding wh...
Our study highlights the utilization of a genetic database for wood-origin identification in Ints... more Our study highlights the utilization of a genetic database for wood-origin identification in Intsia palembanica, a valuable heavy hardwood from the tropical forests. This forensic tool is essential for strengthening the verification of legality in the wood supply chain from the forest to the end-users. An increasing number of rules and regulations are being put in place to promote sustainable practice in the timber trade, one of which involves ensuring that importers declare the correct species name and source of geographic origin of the timber. We aimed to determine the origin of the I. palembanica seed source used in the early establishment on the Forest Research Institute Malaysia (FRIM) campus. DNA samples of I. palembanica individuals from the FRIM campus were obtained and analyzed using four chloroplast (cp) DNA markers to characterize the haplotype variants for population identification. In addition, the DNA samples were also genotyped at 14 short tandem repeat (STR) loci for...
To inform product users about the origin of timber, the implementation of a traceability system i... more To inform product users about the origin of timber, the implementation of a traceability system is necessary for the forestry industry. In this study, we developed a comprehensive genetic database for the important tropical timber species Merbau, Intsia palembanica, to trace its geographic origin within peninsular Malaysia. A total of 1373 individual trees representing 39 geographically distinct populations of I. palembanica were sampled throughout peninsular Malaysia. We analyzed the samples using a combination of four chloroplast DNA (cpDNA) markers and 14 short tandem repeat (STR) markers to establish both cpDNA haplotype and STR allele frequency databases. A haplotype map was generated through cpDNA sequencing for population identification, resulting in six unique haplotypes based on 10 informative intraspecifically variable sites. Subsequently, an STR allele frequency database was developed from 14 STRs allowing individual identification. Bayesian cluster analysis divided the individuals into two genetic clusters corresponding to the northern and southern regions of peninsular Malaysia. Tests of conservativeness showed that the databases were conservative after the adjustment of the θ values to 0.2000 and 0.2900 for the northern (f = 0.0163) and southern (f = 0.0285) regions, respectively. Using self-assignment tests, we observed that individuals were correctly assigned to populations at rates of 40.54-94.12% and to the identified regions at rates of 79.80-80.62%. Both the cpDNA and STR markers appear to be useful for tracking Merbau timber originating from peninsular Malaysia. The use of these forensic tools in addition to the existing paper-based timber tracking system will help to verify the legality of the origin of I. palembanica and to combat illegal logging issues associated with the species.
The development of timber tracking methods based on genetic markers can provide scientific eviden... more The development of timber tracking methods based on genetic markers can provide scientific evidence to verify the origin of timber products and fulfill the growing requirement for sustainable forestry practices. In this study, the origin of an important Dark Red Meranti wood, Shorea platyclados, was studied by using the combination of seven chloroplast DNA and 15 short tandem repeats (STRs) markers. A total of 27 natural populations of S. platyclados were sampled throughout Malaysia to establish population level and individual level identification databases. A haplotype map was generated from chloroplast DNA sequencing for population identification, resulting in 29 multilocus haplotypes, based on 39 informative intraspecific variable sites. Subsequently, a DNA profiling database was developed from 15 STRs allowing for individual identification in Malaysia. Cluster analysis divided the 27 populations into two genetic clusters, corresponding to the region of Eastern and Western Malays...
Background: Dipterocarpaceae is a pantropical tree family that plays an important role in our und... more Background: Dipterocarpaceae is a pantropical tree family that plays an important role in our understanding of the ecology of Asian tropical rain forests. However, genome sizes for members of the Dipterocarpaceae are still poorly known. Aims: To report the genome size of 115 dipterocarp species and examine the variation and evolution of genome size in this family. Methods: Genome size was estimated using flow cytometry. Both the rpoB and trnL intron were sequenced to uncover the evolution of genome size within a phylogenetic framework. Results: The 1C genome size varied between 0.267 and 0.705 pg in Shorea hemsleyana and Shorea ovalis, respectively, a 2.64-fold variation across the family. Most dipterocarps are characterised by very small genomes with a mean 1C value of 0.416 pg (sd = 0.075) and five polyploids are recorded. The ancestral genome size for dipterocarps was reconstructed as 1Cx = 0.481 pg (95% CI = 0.433–0.534). Conclusions: Genome size variation in dipterocarps was characterised by very small values with a narrow range. Overall, genome size reduction from the ancestral state is a general trend in Dipterocarpaceae.
Abstract Taxonomic classification of the important medicinal plant, Ficus deltoidea (Moraceae), i... more Abstract Taxonomic classification of the important medicinal plant, Ficus deltoidea (Moraceae), is challenging because of the variability of its leaves and fig forms that occur within the species. We developed 16 nuclear Simple Sequence Repeat (nSSR) markers, and characterized them using 24 individuals from a natural population. We then studied the intraspecific variation of F . deltoidea subsp. deltoidea in Peninsular Malaysia using morphological and molecular approaches. Based on the morphological variations, we further determined the varieties that occur regionally under the above subspecies based on the leaf characteristics. As for molecular data sets, we used both chloroplast DNA (cpDNA) and nSSR markers to elucidate the phylogenetic relationship among the varieties. The cpDNA dendrogram yielded poorer resolution where most of the clades were forming paraphyletic complex. The cluster analysis based on nSSR is largely congruent with the morphological classification, with F . deltoidea subsp. deltoidea classified into four main varieties, namely var. deltoidea , var. angustifolia , var. kunstleri and var. lutescens . Our study demonstrates the applicability of molecular approach in complementing the conventional taxonomic classification.
Illegal logging and smuggling of Gonystylus bancanus (Thymelaeaceae) poses a serious threat to th... more Illegal logging and smuggling of Gonystylus bancanus (Thymelaeaceae) poses a serious threat to this fragile valuable peat swamp timber species. Using G. bancanus as a case study, DNA markers were used to develop identification databases at the species, population and individual level. The species level database for Gonystylus comprised of an rDNA (ITS2) and two cpDNA (trnH-psbA and trnL) markers based on a 20 Gonystylus species database. When concatenated, taxonomic species recognition was achieved with a resolution of 90% (18 out of the 20 species). In addition, based on 17 natural populations of G. bancanus throughout West (Peninsular Malaysia) and East (Sabah and Sarawak) Malaysia, population and individual identification databases were developed using cpDNA and STR markers respectively. A haplotype distribution map for Malaysia was generated using six cpDNA markers, resulting in 12 unique multilocus haplotypes, from 24 informative intraspecific variable sites. These unique haplotypes suggest a clear genetic structuring of West and East regions. A simulation procedure based on the composition of the samples was used to test whether a suspected sample conformed to a given regional origin. Overall, the observed type I and II errors of the databases showed good concordance with the predicted 5% threshold which indicates that the databases were useful in revealing provenance and establishing conformity of samples from West and East Malaysia. Sixteen STRs were used to develop the DNA profiling databases for individual identification. Bayesian clustering analyses divided the 17 populations into two main genetic clusters, corresponding to the regions of West and East Malaysia. Population substructuring (K=2) was observed within each region. After removal of bias resulting from sampling effects and population subdivision, conservativeness tests showed that the West and East Malaysia databases were conservative. This suggests that both databases can be used independently for random match probability estimation within respective regions. The reliability of the databases was further determined by independent self-assignment tests based on the likelihood of each individual's multilocus genotype occurring in each identified population, genetic cluster and region with an average percentage of correctly assigned individuals of 54.80%, 99.60% and 100% respectively. Thus, after appropriate validation, the genetic identification databases developed for G. bancanus in this study could support forensic applications and help safeguard this valuable species into the future.
Neobalanocarpus heimii, locally known as chengal, is an important timber species in Peninsular Ma... more Neobalanocarpus heimii, locally known as chengal, is an important timber species in Peninsular Malaysia. Owing to the high demand for its valuable timber, N. heimii is subjected to illegal logging and this species may become endangered in the near future. The present study was designed to identify a set of highly polymorphic short tandem repeat (STR) markers for timber tracking of N. heimii. An extensive evaluation of 51 STRs developed for Dipterocarpaceae managed to identify 12 STR loci (Nhe004, Nhe005, Nhe011, Nhe015, Nhe018, Hbi161, Sle392, Sle605, Slu044a, Shc03, Shc04 and Shc07), which showed specific amplification, high polymorphism, single-locus mode of inheritance, absence of null alleles and absence of mononucleotide repeat motifs in N. heimii. These loci can be readily used to establish a linkage between the evidentiary sample and the source, thus providing a useful set of markers for individual identification in N. heimii.
Microsatellite markers were developed for Johannesteijsmannia lanceolata to assess the genetic di... more Microsatellite markers were developed for Johannesteijsmannia lanceolata to assess the genetic diversity and mating system of this alarmingly endangered species. A total of 31 polymorphic microsatellite markers were developed for J. lanceolata using the enrichment protocol. These markers were screened on 24 samples from a natural population. The number of alleles ranged from four to 20, while the observed heterozygosity ranged from 0.391 to 1.000. The 31 loci were further tested for transferability on J. altifrons, J. magnifica, and J. perakensis. Generally, all loci showed positive amplifications in these three Johannesteijsmannia species, except Jla124 (J. magnifica) and Jla168b (J. magnifica and J. perakensis). These microsatellite markers could be employed to study the population genetics and mating system of J. lanceolata and other Johannesteijsmannia species.
Aggressive collections and trade activities in recent decades have resulted in heavy pressure on ... more Aggressive collections and trade activities in recent decades have resulted in heavy pressure on the natural stands of Aquilaria malaccensis and concerns over its long-term survival potential. To aid DNA profiling and assessment of its genetic diversity, microsatellite markers were developed for the species. Seventeen polymorphic microsatellite markers were developed for A. malaccensis using an enrichment protocol. The markers were screened on 24 samples from a natural population. The number of alleles ranged from two to 11, and the observed heterozygosity ranged from 0.042 to 0.957. No significant deviation from Hardy-Weinberg equilibrium was detected after conservative Bonferroni correction. This is the first report on the development of microsatellite markers in A. malaccensis. The markers will be used to establish a DNA profiling database and to estimate the genetic diversity and population genetic structure of the species.
Begonias are hyper-diverse and important horticultural plants. Six polymorphic microsatellite mar... more Begonias are hyper-diverse and important horticultural plants. Six polymorphic microsatellite markers were developed from CT- and GT-enriched libraries of Begonia maxwelliana. The number of alleles per locus ranged from 2 to 12 and the observed heterozygosity ranged from 0.036 to 0.813. Null alleles were detected in one locus (Bma161) after Bonferroni correction. All the six markers were amplifiable in 23 selected Begonia species with the success rates of 17-100%. On average, species of the same section as B. maxwelliana (i.e. sect. Platycentrum) yielded higher transferability (91%). These markers will be useful for population genetic studies of the genus Begonia.
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