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Worldwide, many mangrove species are experiencing significant population declines, including Rhizophora apiculata, which is one of the most widespread and economically important species in tropical Asia. In Malaysia, there has been an alarming decline in R. apiculata populations 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 for R. 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 (mean He: 0.352) with significant genetic differentiation (FST: 0.315) among populations of R. apiculata. Approximately two-third of the populations showed significant excess of homozygotes, indicating persistent inbreeding which might be due to the decrease in population size or fragmentation. From the cluster analyses, the populations investigated were divided into two distinct clusters, comprising the west and east coasts of Peninsular Malaysia. The western cluster was further divided into two sub-clusters with one of the sub-clusters showing strong admixture pattern that harbours high levels of genetic diversity, thus deserving high priority for conservation.

Southeast Asian rainforests at upper hill elevations are increasingly vulnerable to degradation because most lowland forest areas have been converted to different land uses. As such, understanding the genetics of upper hill species is becoming more crucial for their future management and conservation. Shorea platyclados is an important, widespread upper hill dipterocarp in Malaysia. To elucidate the genetic structure of S. platyclados and ultimately provide guidelines for a conservation strategy for this species, we carried out a comprehensive study of the genetic diversity and demographic history of S. platyclados. Twenty-seven populations of S. platyclados across its range in Malaysia were genotyped at 15 polymorphic microsatellite loci and sequenced at seven noncoding chloroplast DNA (cpDNA) regions. A total of 303 alleles were derived from the microsatellite loci, and 29 haplotypes were identified based on 2892 bp of concatenated cpDNA sequences. The populations showed moderately high genetic diversity (mean HE = 0.680 for microsatellite gene diversity and HT = 0.650 for total haplotype diversity) and low genetic differentiation (FST = 0.060). Bayesian clustering divided the studied populations into two groups corresponding to western and eastern Malaysia. Bottleneck analysis did not detect any recent bottleneck events. Extended Bayesian skyline analyses showed a model of constant size for the past population history of this species. Based on our findings, priority areas for in situ and ex situ conservation and a minimum population size are recommended for the sustainable utilization of S. platyclados.

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 individual identification. Individual assignment to the possible geographic origin was done through an assignment test. On the basis of our recently developed I. palembanica genetic databases, the I. palembanica seed source for the early establishment was inferred to be originated from a mixture of several sources, with a large portion from the southern region (89%) and a relatively small portion from the northern region (11%) of Peninsular Malaysia. The I. palembanica seed source used for the early establishment on the century old FRIM campus was inferred to be originated from several forest reserves located not far from the planting sites. This study proves the applicability of the DNA method in supply-chain verification, where an unknown I. palembanica tree can be traced to its geographic origin using genetic databases.

Background: Genetic divergence is one of the key processes in speciation. In the Begoniaceae, genetic divergence caused by limited gene flow may explain its high species diversity and endemicity. T...

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.

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.

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 Malaysia. The conservativeness tests showed that the Malaysia database is conservative after removal of bias from population subdivision and sampling effects. Independent self-assignment tests correctly assigned individuals to the database in an overall 60.60−94.95% of cases for identified populations, and in 98.99−99.23% of cases for identified regions. Both the chloroplast DNA database and the STRs appear to be useful for tracking timber originating in Malaysia. Hence, this DNA-based method could serve as an effective addition tool to the existing forensic timber identification system for ensuring the sustainably management of this species into the future.

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 ( Bma 161) 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 .

Termites from the genus Odontotermes are known to contain numerous species complexes that are difficult to tell apart morphologically or with mitochondrial DNA sequences. We developed markers for one such cryptic species complex, that is, Odontotermes srinakarinensis sp. nov. from Maxwell Hill Forest Reserve (Perak, Malaysia), and characterised them using a sample of 41 termite workers from three voucher samples from the same area. We then genotyped 150 termite individuals from 23 voucher samples/colonies of this species complex from several sites in Peninsular Malaysia. We analysed their population by constructing dendograms from the proportion of shared-alleles between individuals and genetic distances between colonies; additionally, we examined the Bayesian clustering pattern of their genotype data. All methods of analysis indicated that there were two distinct clusters within our data set. After the morphologies of specimens from each cluster were reexamined, we were able to separate the two species morphologically and found that a single diagnostic character found on the mandibles of its soldiers could be used to separate the two species quite accurately. The additional species in the clade was identified as Odontotermes denticulatus after it was matched to type specimens at the NHM London and Cambridge Museum of Zoology.

Tectonic movements, climatic oscillations, and marine transgressions during the Cenozoic have had a dramatic effect on the biota of the tropical rain forest. This study aims to reveal the phylogeography and evolutionary history of a Peninsular Malaysian endemic tropical timber species, Neobalanocarpus heimii (Dipterocarpaceae). A total of 32 natural populations of N. heimii, with 8 samples from each population were investigated. Fifteen haplotypes were identified from five noncoding chloroplast DNA (cpDNA) regions. Overall, two major genealogical cpDNA lineages of N. heimii were elucidated: a widespread southern and a northern region. The species is predicted to have survived in multiple refugia during climatic oscillations: the northwestern region (R1), the northeastern region (R2), and the southern region (R3). These putative glacial refugia exhibited higher levels of genetic diversity, population differentiation, and the presence of unique haplotypes. Recolonization of refugia R1 and R2 could have first expanded into the northern region and migrated both northeastwards and northwestwards. Meanwhile, recolonization of N. heimii throughout the southern region could have commenced from refugia R3 and migrated toward the northeast and northwest, respectively. The populations of Tersang, Pasir Raja, and Rotan Tunggal exhibited remarkably high haplotype diversity, which could have been the contact zones that have received an admixture of gene pools from the northerly and also southerly regions. As a whole, the populations of N. heimii derived from glacial refugia and contact zones should be considered in the conservation strategies in order to safeguard the long-term survival of the species.

This paper reports 10 polymorphic microsatellite markers for a valuable peat swamp timber species, Gonystylus bancanus (Thymelaeaceae). The primers were designed from a genomic library enriched with dinucleotide (CT) repeats and screened on 24 samples from a natural population. The number of alleles detected per locus ranged from two to nine while the observed and expected heterozygosities ranged from 0.455 to 0.864 and from 0.476 to 0.863, respectively. No significant deviation from Hardy–Weinberg equilibrium (P

Simple sequence repeats (SSRs) derived from expressed sequence tags (ESTs) are valuable markers because they represent transcribed regions and often transferable to related taxa. Here, we report the development and characterization of EST-SSRs from Shorea leprosula. Fifty-four sequences containing SSRs were identified in 2003 unigenes assembled from 3159 ESTs. Twenty-four EST-SSRs were developed, of which four gave multiple amplifications, five were found to be monomorphic and 15 showed polymorphism, with allele numbers ranging from two to 17 in a single Pasoh Forest Reserve population of 24 individuals. The observed and expected heterozygosities ranged from 0.05 to 0.91 and from 0.16 to 0.93, respectively. Cross-species transferability of the 15 loci to 36 species within Dipterocarpaceae revealed between four and 14 loci that gave positive amplification and 10 loci were found to be transferable to more than 15 species.

To conserve a rare plant, conservation programs must be guided by the biological attributes of the species. What constitutes the most critical biological information for plant conservation has been the issue of discussion for the last 30 years. Most scientists promote an approach that is either ecological or genetic in emphasis. Ecological and genetic processes will often interact synergistically to influence the population viability and to determine the persistence of populations in the long run. Consequently, conservation management of plant species, in addition to ecological information, requires a robust understanding of underlying genetic processes as well as the variation within and between populations. Conservation has a cost and the resources available for conservation programs are always limited. Therefore, conservation management strategies should not only be scientifically justified but also practical in terms of resource availability. Shorea lumutensis is a rare and endemic dipterocarp in Peninsular Malaysia. A comprehensive research activity was initiated to assess the population ecology and population genetics of S. lumutensis to elucidate specific ecological and genetic requirements and subsequently to set conservation strategies and priorities. This paper is apparently the first attempt at applying both the ecological and genetic approaches into conservation management of a rare dipterocarp. In addition, this paper also attempts to link the gaps between conservation research and conservation management in a realistic manner. It is our hope that this study will serve as a model for the study of other rare dipterocarps which should be given priority for conservation.

Intsia palembanica is a high-value tropical hardwood species of the family Leguminosae. A total of 16 microsatellite markers were developed for this important timber species using an enrichment approach. These markers were screened on 24 samples from a natural population and subsequently tested for reproducibility, ease of scoring and locus-specificity, across five populations. The observed and expected heterozygosities ranged from 0.026 to 0.976 and 0.026 to 0.928, respectively, while the mean number of alleles per locus per population was 8.73. These microsatellite markers are informative and will be useful for conservation genetic study, individual identification and timber tracking.

Premise of the study 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. Methods and results 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. Conclusions 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.

We examined the effect of selective logging on the genetic diversity of Scaphium macropodum using RAPD markers via two approaches: (1) to investigate the immediate effect by studying a same population before and after logging, and (2) to determine the long term effect by comparing two regenerated stands with an adjacent unlogged stand, assuming that they were genetically identical before logging. Results showed no negative immediate impact for the first approach, probably due to the high abundance and heterogeneity of S. macropodum in the compartment investigated. However, for the latter approach, substantial genetic erosion (i.e. 31.5% reduction for Shannon diversity, H) was detected in one of the regenerated stands corresponding to its extremely low tree density for S. macropodum. This implies the possible occurrence of genetic drift and increased inbreeding due to population decline as a result of logging. However, the observed genetic differences among the three sub-populations having prevailed before logging cannot be totally discounted in the second approach. This study also demonstrates the use of tree density as a good surrogate measure of genetic diversity. The present harvesting system in Malaysia based on a general cutting limit need to be refined; the basis for determining cutting limit in a forest management unit should consider abundance of commercial species.

Pollen flow and population genetic structure among 30 potentially flowering individuals of Neobalanocarpus heimii , a tropical emergent tree, were investigated in a lowland tropical rainforest of Malaysia using microsatellite polymorphism. The 248 offspring in the vicinity of five reproductive trees of the 30 potentially flowering trees were used in paternity analysis for pollen-flow study. Four primer pairs, developed in different species of dipterocarps, were adopted to detect microsatellite polymorphism. Based upon microsatellite polymorphism, pollen flow and seed migration were detected. Pollen-flow events of more than 400 m were observed directly, based on paternity analysis in the study plot. The estimated average mating distance of the five reproductive trees was 524 m. This result suggests that reproduction of this species is mediated by a long-distance pollinator. The haplotypes of some offspring were not compatible with the nearest reproductive tree. Thus, the results suggest that some seeds are dispersed by a seed dispersal vector. Investigation of genetic structure showed significant and negative correlation of genetic relatedness and spatial distances between the 30 potentially flowering trees, but this correlation was weak. We suggest that long-distance gene flow and seed migration are responsible for the poorly developed genetic structure of this species.

Nine polymorphic microsatellite markers were isolated and characterized in Shorea platyclados (Dipterocarpaceae) using CT-enriched library method. The polymorphism of the microsatellite loci was analyzed by screening 98 samples from a natural population. The number of alleles per locus ranged from 4 to 12, with an average of 8.22 per locus, and the observed heterozygosity ranged from 0.204 to 0.888 (average 0.562). These microsatellite loci will be useful for population genetic studies on S. platyclados and adding genetic information for hill dipterocarps.