AMOVA Test Research Articles

Molecular characterisation of Aspergillus flavus isolates from peanut fields in India using AFLP.

Aflatoxin contamination of peanut, due to infection by Aspergillus flavus, is a major problem of rain-fed agriculture in India. In the present study, molecular characterisation of 187 Aspergillus flavus isolates, which were sampled from the peanut fields of Gujarat state in India, was performed using AFLP markers. On a pooled cluster analysis, the markers could successfully discriminate among the ‘A’, ‘B’ and ‘G’ group A. flavus isolates. PCoA analysis also showed equivalent results to the cluster analysis. Most of the isolates from one district could be clustered together, which indicated genetic similarity among the isolates. Further, a lot of genetic variability was observed within a district and within a group. The results of AMOVA test revealed that the variance within a population (84%) was more than that between two populations (16%). The isolates, when tested by indirect competitive ELISA, showed about 68.5% of them to be atoxigenic. Composite analysis between the aflatoxin production and AFLP data was found to be ineffective in separating the isolate types by aflatoxigenicity. Certain unique fragments, with respect to individual isolates, were also identified that may be used for development of SCAR marker to aid in rapid and precise identification of isolates.

Demographic History of Indigenous Populations in Mesoamerica Based on mtDNA Sequence Data.

The genetic characterization of Native American groups provides insights into their history and demographic events. We sequenced the mitochondrial D-loop region (control region) of 520 samples from eight Mexican indigenous groups. In addition to an analysis of the genetic diversity, structure and genetic relationship between 28 Native American populations, we applied Bayesian skyline methodology for a deeper insight into the history of Mesoamerica. AMOVA tests applying cultural, linguistic and geographic criteria were performed. MDS plots showed a central cluster of Oaxaca and Maya populations, whereas those from the North and West were located on the periphery. Demographic reconstruction indicates higher values of the effective number of breeding females (Nef) in Central Mesoamerica during the Preclassic period, whereas this pattern moves toward the Classic period for groups in the North and West. Conversely, Nef minimum values are distributed either in the Lithic period (i.e. founder effects) or in recent periods (i.e. population declines). The Mesomerican regions showed differences in population fluctuation as indicated by the maximum Inter-Generational Rate (IGRmax): i) Center-South from the lithic period until the Preclassic; ii) West from the beginning of the Preclassic period until early Classic; iii) North characterized by a wide range of temporal variation from the Lithic to the Preclassic. Our findings are consistent with the genetic variations observed between central, South and Southeast Mesoamerica and the North-West region that are related to differences in genetic drift, structure, and temporal survival strategies (agriculture versus hunter-gathering, respectively). Interestingly, although the European contact had a major negative demographic impact, we detect a previous decline in Mesoamerica that had begun a few hundred years before.

Population genetic structure in medicinal plant Lallemantia iberica (Lamiaceae)

Koohdar F, Sheidai M, Talebi SM, Noormohammadi Z. 2015. Population genetic structure in medicinal plant Lallemantia iberica (Lamiaceae). Biodiversitas 16: 139-144. Lallemantia iberica (Bieb.) Fischer and C.A. Meyer (sin. Dracocephalum ibericumM.Bieb.) also named Dragon's head” is an annual plant cultivated for its seeds that contain about 30% -38% drying oil (siccative oil). Itsseed oil is used in foods, dye and varnish industry. L. iberica seeds have traditional uses as reconstitute, stimulant, diuretic andexpectorant. L. iberica in an important medicinal plant in our country and grows in various regions with different environmentalconditions. At present no investigation has been reported about population genetic structure of this valuable plant species in Iran.Therefore, we carried out population genetic analysis of 11 populations of L. iberica by using ISSR molecular markers for the first time.Genetic diversity analysis revealed high within population genetic variability. AMOVA test produced significant genetic differenceamong the studied populations. Mantel test revealed significant correlation between genetic distance and geographical distance of thepopulations. STRUCTURE analysis and K-Means clustering revealed population genetic fragmentation and the presence of three genepools for this species. The assignment test revealed the occurrence of limited gene flow among the populations. The results suggestedthat genetic divergence, limited gene flow, genetic drift and local adaptation have played role in diversification of L. iberica.
 Key words: Gene flow, population fragmentation, IBD, Lallemantia iberica.

Microsatellite Analysis Reveals Population Structure and Population Expansion ofTecia solanivora1inSolanum tuberosumin Colombia

Abstract. Tecia solanivora (Lepidoptera: Gelechiidae) (Povolny 1973), is the most important insect pest of potato (Solanum tuberosum L.) in Central and South America, and Spain. The species was recently reported in Mexico, demonstrating the power of invasion by this insect. In Peru, the insect is quarantined, and could become a damaging pest in America. Larvae attack potato tubers, causing economic loss of 50 to 100%. We used eight microsatellites in 152 individuals from Antioquia (North), Boyaca (Center), Narino (South), and Norte de Santander (East), Colombia, and found T. solanivora genetically different based on an AMOVA test (FST = 0.094, P < 0.01, RST = 0.175, P < 0.01). The differentiation was supported by Bayesian analysis where we estimated K = 2. The outcome suggests populations from Antioquia and Boyaca are genetically similar whereas populations from Narino and Norte de Santander differ because of geographical separation from other regions. We found T. solanivora had not undergone a recent bot...

Genetic diversity, population structure and marker trait associations for seed quality traits in cotton (Gossypium hirsutum).

Cottonseed contains 16% seed oil and 23% seed protein by weight. High levels of palmitic acid provides a degree of stability to the oil, while the presence of bound gossypol in proteins considerably changes their properties, including their biological value. This study uses genetic principles to identify genomic regions associated with seed oil, protein and fibre content in upland cotton cultivars. Cotton association mapping panel representing the US germplasm were genotyped using amplified fragment length polymorphism markers, yielding 234 polymorphic DNA fragments. Phenotypic analysis showed high genetic variability for the seed traits, seed oil range from 6.47-25.16%, protein from 1.85-28.45% and fibre content from 15.88-37.12%. There were negative correlations between seed oil and protein content.With reference to genetic diversity, the average estimate of FST was 8.852 indicating a low level of genetic differentiation among subpopulations. The AMOVA test revealed that variation was 94% within and 6% among subpopulations. Bayesian population structure identified five subpopulations and was in agreement with their geographical distribution. Among the mixed models analysed, mixed linear model (MLM) identified 21 quantitative trait loci for lint percentage and seed quality traits, such as seed protein and oil. Establishing genetic diversity, population structure and marker trait associations for the seed quality traits could be valuable in understanding the genetic relationships and their utilization in breeding programmes.

Deep phylogeographic divergence among populations of limpet Siphonaria lessoni on the east and west coasts of South America

The historical processes that have influenced the genetic structure of many species are often associated with environmental changes of the Pleistocene glacial cycles. These climate changes involve temperature oscillation, marine currents and loss of coastal habitats, which could have affected the abundance and geographic distribution of marine species in temperate coastal habitats. In this work, a 552-bp mtDNA fragment of COI locus of 92 individuals was sequenced to analyze the genetic structure of the limpet Siphonaria lessoni. Individuals were collected on the intertidal coast of the Southern Atlantic (Mar del Plata, San Antonio Oeste, Puerto Madryn and Ushuaia in Argentina) and the Southern Pacific (Valdivia and Valparaiso in Chile). S. lessoni displayed two distinct lineages that were nearly reciprocally monophyletic between the Atlantic and Pacific coasts. AMOVA tests revealed the existence of strong population genetic structure. The Pacific coasts yielded more haplotypes and polymorphic sites as well as higher haplotype and nucleotide diversity than the Atlantic clade did. Both Tajima’s D and Fu’s F s were significant and negative, suggesting that limpet populations are in population expansion or have recently expanded. Accordingly, the haplotype network for each clade showed a star-like phylogeographic pattern. From IMa analysis, the divergence time between Pacific and Atlantic populations was 100,000–1,000,000 ybp with gene flow occurring from Pacific to Atlantic populations. The Bayesian Skyline analysis revealed an older coalescence in the Pacific clade (30,000–300,000 ybp) as compared to that in the Atlantic clade (4,000–40,000 ybp). This work reports evidence of Pacific–Atlantic geographic isolation with asymmetric migration, which is probably related to changes in sea level and temperature due to the extended glaciation periods that occurred in the region throughout the Pleistocene.

Genetic diversity and population structure of the ark shell Scapharca broughtonii along the coast of China based on microsatellites

As a commercially important species in East Asia, the natural resources of Scapharca broughtonii have been suffering from severe population decline across its main habitats. In China, recovery efforts for S. broughtonii are in progress. To provide scientific bases for fisheries management and conservation program, genetic diversity and population structure of seven wild populations of S. broughtonii from the northern China coast was assessed using seven microsatellite loci in this study. High genetic diversity was present in all the seven populations, as observed in mean allelic richness per locus (11.3–12.5), and average expected heterozygosity (0.835–0.867). No significant difference in allelic richness or expected heterozygosity was observed among the seven populations. Pairwise FST estimates and NJ tree topologies based on DC distances indicated that the seven populations fell into two groups, showing a clear division between the populations from the south and north of the Shandong Peninsula. Genetic differentiation was further analyzed using AMOVA and assignation tests. Genetic barrier analysis using Monmonier algorithm also confirmed that the Shandong Peninsula was the putative barrier separating the northern and southern populations. In addition, marine currents probably play an important role in high gene flow among three populations from the same marine gyre.

Population genetic structure and genetic diversity in &lt;i&gt;Dracocephalum thymiflorum&lt;/i&gt; L. (Lamiaceae) populations in Iran

Dracocephalum thymiflorum L. is a medicinal plant that grows in limited areas in Iran and forms few local populations. We have no information on its genetic diversity and population genetic structure in the country. Therefore, the present population genetic study was performed to provide data on population genetic structure, potential gene pools and gene flow in Dracocephalum thymiflorum in Iran. The information obtained can be used in conservation of this medicinal plant species. We studied 55 randomly selected plants from 5 geographical populations by ISSR molecular markers. The studied populations contained high within-population genetic variability and also revealed strong genetic differentiation by AMOVA test (P = 0.01). Structure analysis and K-Means clustering revealed population genetic fragmentation that was mainly due to genetic difference between population 2 and the other studied populations. The population assignment test revealed the occurrence of limited gene flow among these populations. The information obtained can me used in programming the conservation of this important medicinal plant in Iran.

Genetic Stock Structure of Terapon jarbua in Taiwanese Waters

AbstractOver the past century, stock identification, which is crucial for fisheries stock assessments, has relied heavily on intraspecies variations to differentiate management units. In recent years, however, DNA approaches have shed additional light on some aspects of the natural history and ecology of species and stocks. The Crescent Grunter Terapon jarbua is widely distributed across the Indo‐Pacific region. It inhabits coastal waters with sandy substrates and tends to aggregate in estuaries. In the present study, we collected 96 individuals of T. jarbua from 10 locations around the shoreline of Taiwan as well as near Guangdong, China. A concatenated sequence (1,753 bp) of mtDNA (cytochrome c oxidase I and cytochrome b) was obtained from all individuals. We discovered two genetically distinct clades (lineages A and B) with different historical demographies occurring sympatrically except at the Yilan collection site, which was dominated by lineage A haplotypes. Connectivity within this region is high according to FST and AMOVA tests. The genetic variation between the two clades is far below the interspecies threshold for the genus Terapon (0.007 versus 0.3156). Therefore, we suggest that they be considered different genetic stocks from a fisheries management point of view and that future stock reassessment should be conducted based on the genetic information provided in this study. Further large‐scale sampling is needed to understand the mechanisms that drive genetic partitioning on regional (Northwest Pacific) and global (Indo‐Pacific) scales.Received February 9, 2015; accepted July 9, 2015

Gene variation and genetic differentiation among populations of the solitary mud dauber wasp Trypoxylon (Trypargilum) albitarse Fabricius 1804 (Hymenoptera, Crabronidae).

Trypoxylon is a genus of solitary crabronid wasps whose population genetics is poorly known. The purpose of the present study was to investigate the genetic variation and differentiation among five populations of Trypoxylon albitarse, a species widely distributed throughout the Neotropics, with records from Panama to northern Argentina. Eight species-specific microsatellite loci were used for genotyping 96 adult wasps (one female per nest) sampled at five sites in Brazil. The analysis of allelic richness and private alleles indicated high genetic diversity in the populations sampled. Pairwise comparisons using the F st and D est indices revealed significant differentiation for all, but one pair of populations. F st, D est, AMOVA and assignment test values pointed to inter-population differentiation. Additionally, the analysis of population structure using Bayesian and PCA methods characterized two alternative genetic groups. The Mantel test indicated no correlation between genetic and geographic distances. Despite evidence of considerable dispersal capacity for T. albitarse, the data indicate low to moderate population structuring in this species.

Genetic structure of cultivated flax (Linum usitatissimum L.) based on retrotransposon-based markers

Flax (Linum usitatissimum L.) is one of the most important fiber and oil crop plants cultivated since ancient time. The flax seeds contain high amount of omega- 3-fatty acids and biologically active lignans. In spite of economic importance of cultivated flax, no information is available on its genetic variability and population structure in Iran. Therefore, we used six inter-retrotransposon amplified polymorphism (IRAP) markers and 15 combined IRAP markers to reveal within and among population genetic diversity in this crop plant. We used 30 randomly selected plants in three geographical populations for present investigation. AMOVA test produced significant genetic difference (PhiPT = 0.40, P = 0.010) among the studied populations and also revealed that, 40% of total genetic variability was due to within population diversity while, 60% was due to among population genetic differentiation. Gst (0.78, P = 0.001), Hedrick, standardised fixation index (G'st = 0.83, P = 0.001), revealed that the studied populations are genetically differentiated. STRUCTURE plot based on admixture model revealed that the studied populations differed extensively in their genetic content, but some degree of shared alleles occurred between them. Some adaptive IRAP loci were identified by LFMM analysis. These loci were private alleles restricted to geographical populations. Data obtained may be used in breeding and hybridization program of flax in the country.

Genetic structure and differentiation of the fire salamander Salamandra salamandra at the northern margin of its range in the Carpathians

Amphibian populations occurring at the margin of the species range exhibit lower genetic variation due to strong genetic drift and long-term isolation. Limited mobility and site fidelity together with habitat changes may accelerate genetic processes leading to local extinction. Here, we analyze genetic variation of the fire salamander subspeciesSalamandra s. salamandrainhabiting the Outer Carpathian region in Poland, at the northern border of its distribution. Nuclear DNA polymorphism based on 10 microsatellite loci of 380 individuals sampled in 11 populations were analysed to measure gene flow between subpopulations and possible long-term isolation. Mitochondrial DNA control region analysis among 17 individuals representing 13 localities was used to detect the origin of populations which colonized Northern Europe after the last glaciation. Overall, pairwise FST’s and AMOVA test of ‘among group’ variation showed little differences in the allele frequencies and relatively high local gene flow. However, Bayesian clustering results revealed subtle structuring between eastern and western part of the studied region. Two extreme marginal populations from the Carpathian Piedmont revealed reduced genetic variation which may be attributed to strong influence of genetic drift. Only one mitochondrial DNA haplotype (type IIb) was found in all individuals and suggest that after the Last Glacial MaximumSalamandra salamandramigrated to the North-Western Europe from the single glacial refugium placed in the Balkan Peninsula.

Development of a Multilocus Sequence Typing scheme for the study of Anaplasma marginale population structure over space and time

Bovine Anaplasmosis caused by Anaplasma marginale is a worldwide disease prevalent in tropical and subtropical regions where Rhipicephalus microplus is considered the most significant biological vector. Molecular markers previously applied for A. marginale typing are efficient for isolate discrimination but they are not a suitable tool for studying population structure and dynamics. Here we report the development of an MLST scheme based on the study of seven genes: dnaA, ftsZ, groEl, lipA, recA, secY and sucB. Five annotated genomes (Saint Maries, Florida, Mississippi, Puerto Rico and Virginia) and 53 bovine blood samples from different world regions were analyzed. High nucleotide diversity and a large proportion of synonymous substitutions, indicative of negative selection resulted from DnaSP 5.00.02 package application. Recombination events were detected in almost all genes, this evidence together with the coexistence of more than one A. marginale strain in the same sample might suggest the superinfection phenomena as a potential source of variation. The allelic profile analysis performed through GoeBURST shown two main CC that did not support geography. In addition, the AMOVA test confirmed the occurrence of at least two main genetically divergent groups. The composition of the emergent groups reflected the impact of both historical and environmental traits on A. marginale population structure. Finally, a web-based platform “Galaxy MLST-Pipeline” was developed to automate DNA sequence editing and data analysis that together with the Data Base are freely available to users.The A. marginale MLST scheme developed here is a valuable tool with a high discrimination power, besides PCR based strategies are still the better choice for epidemiological intracellular pathogens studies. Finally, the allelic profile describe herein would contribute to uncover the mechanisms in how intracellular pathogens challenge virulence paradigm.

Networking and Bayesian analyses of genetic affinity in cotton germplasm

Cotton is one the most important economic crop plant to mankind. A narrow genetic base and limited genetic diversity exists in cotton due to long time cultivation practice and artificial selection of this crop plant. Hybridization is the main task of cotton breeding and thorough search for identification of novel genetic diversity is of immediate need in cotton breeding. Therefore, present study undertakes bioinformatics analysis of genetic diversity versus genetic relationship of cotton germplasm including diploid and tetraploid cottons genotypes and their hybrids. Simple sequenced repeats (SSR) and Expressed sequence tagged-SSR (EST-SSR) markers were used in this study. Tetraploid cotton genotypes showed higher genetic diversity compared to diploid plants. AMOVA test showed significant genetic difference between these two polyploidy groups. Bayesian STRUCTURE analysis produced 9 distinct population groups, while K-Means clustering suggested 2–3 major genetic subgroups in the present germplasm. Networking and reticulation analysis showed high degree of gene exchange among most of the studied plants. Coalescence analysis showed gene duplication and gene extinction along with allotetraploid nature of cotton as the main mechanisms for bringing about new genetic variation in cotton. Obtained data can help us to plan a better hybridization and selection plan for cotton.

Lack of population genetic differentiation of a marine ovoviviparous fish Sebastes schlegelii in Northwestern Pacific

Sebastes schlegelii is one of the fishes that aggregate around drifting seaweed during early development. To examine the population genetic structure of S. schlegelii, a 452-bp fragment of the mtDNA control region was sequenced and used to interpret life history characteristics and larval dispersal strategy. Two-hundred and twenty-one individuals from 13 sites across the entire range of S. schlegelii in China, Japan and Korea were analyzed. A neighbor-joining tree and network showed that there were no significant genealogical structures corresponding to sampling locations. AMOVA, pair-wise FST and exact test revealed no significant genetic differentiation among locations. The migration rate among locations was high based on the result of LAMARC. We conclude that larval dispersal with drifting seaweed and the current environmental factors may play an important role in shaping the contemporary phylogeographic pattern and genetic homogeneity of S. schlegelii.

Genetic diversity and genome size variability in Linum austriacum (Lineaceae) populations

Genetic diversity and genome size variability were studied in 16 geographical populations of Linum austriacum (Lineaceae). Genetic diversity parameters were determined in each population based on ISSR molecular markers. AMOVA test, Gst value and Hickory test revealed significant molecular difference among the studied populations. Mantel test showed correlation between genetic distance and geographical distance in these populations. NJ tree and NeighborNet network grouped the studied plant specimens in 3 major clusters. STRUCTURE analysis identified 12 allelic combinations in agreement with K-Means clustering result. These analyses revealed the presence of genetic variability both among and within studied population. The plant specimens of these geographical populations also differed significantly in their genome size.

Genetic diversity of Norway spruce [Picea abies (L.) Karst.] in Romanian Carpathians

The genetic diversity of Romanian most important coniferous tree species, the Norway spruce, was estimated by means of allozyme markers. A total of 695 adult trees sampled from eleven populations grouped in six mountainous areas in the Romanian Carpathians were analyzed. In three metapopulations (Maramureş, Postăvar and Parâng), to evaluate the influence of altitudinal gradient on genetic diversity, samples were collected from populations located at high and low altitude. At other location (ApuseniMountains) we compared the narrow-crown biotype ( Picea abies var. columnaris ) and the pyramidal crown biotype ( Picea abies var. pyramidalis ) and explored the genetic structure of peat bog ecotype. By analyzing 7 enzyme systems and 12 enzyme coding loci, a total of 38 allelic variants have been detected. The mean value of polymorphic loci for the six sites was 86.1%, ranging between 83.3% and 91.7% and the mean expected heterozygosity was 0.115, resulting in a moderate level of genetic diversity. The highest genetic diversity ( H e = 0.134) was found in the narrow-crown spruce population. Apuseni metapopulation showed the highest genetic diversity ( H e = 0.125), being the most valuable for conservation of genetic resources. The small value of fixation index ( F ST = 0.009) indicates a low genetic differentiation between the six sites and AMOVA test revealed a very high level of genetic diversity within population (99%). Comparative analysis of genetic parameters showed small differences between high and low altitude populations at each site, probably due to the neutral character of the markers analyzed and the effect of gene flow between gradiental populations.

Genetic, cytogenetic and morphological diversity in Helicrysum leucocephalum (Asteraceae) populations

Helichrysum (family Asteraceae) is a large, heterogeneous and polyphyletic genus. Most of the Helichrysum species have ornamental and medicinal values. Helicrysum leucocephalum Boiss., has wide geographical distribution in Iran and forms several local populations. The present investigation was performed to study inter populations , genetic, cytogenetic and morphological diversity of this medicinal species. The AMOVA test showed significant genetic difference among the studied populations but the Mantel test did not show correlation between the genetic distance and geographical distance of these populations. STRUCTURE and reticulation analyses showed some degree of genetic admixture and gene exchange among the studied populations. These populations had different chromosome numbers and also differed significantly in the size of their chromosomes.

Correlation and functional differentiation between different markers to study the genetic diversity analysis in medicinally important plant Plumbago zeylanica

The plant species Plumbago zeylanica (P. zeylanica) is a multipurpose medicinal herb of family Plumbaginaceae. This plant is a natural gift to mankind credited with potential medicinal properties such as anti-cancer, anti-atherogenic, cardiotonic, hepatoprotective, anti-fungal, diabetes and neuroprotective assets to list of few. In this study, genetic diversity and relationships among various Plumbago accessions, collected from different geographical regions of India, was assessed using simple sequence repeat (SSR), sequence related amplified polymorphism (SRAP) and internal transcribed spacer (ITS) markers. SSR and SRAP markers showed highest values of Nei's genetic diversity and Shannon information index among populations. We also observed statistically significant genetic differentiations among and within populations (P<0.01 in the AMOVA tests). Additionally, both Un-weighted paired group method with arithmetic average (UPGMA) and principal coordinate analysis (PCoA) grouped P. zeylanica populations into similar clusters which corroborate the above analysis to be useful for genetic diversity analysis of this plant. Our data signifies that SSR and SRAP are both reliable and effective tools for analyzing genetic diversity in P. zeylanica. However, based on our chosen dataset, we find ITS marker less significant for the genetic diversity analysis of this multifaceted plant. This information would be useful towards the identification, characterization and conservation of this species at the molecular level.

Mitochondrial DNA variation, genetic structure and demographic history of Iranian populations.

In order to survey the evolutionary history and impact of historical events on the genetic structure of Iranian people, the HV2 region of 141 mtDNA sequences related to six Iranian populations were analyzed. Slight and non-significant FST distances among the Central-western Persian speaking populations of Iran testify to the common origin of these populations from one proto-population. Mismatch distribution suggests that this proto-Iranian population started to colonize Iran about 30000 years ago which is almost consistent with the timing of arrival and colonization of western Asia by the anatomically modern human. Star-like haplotype network structures, significant and negative Tajima's D (D=-2.08, P<0.05) and unimodal mismatch distributions support the genetic effects of this expansion. Iranian populations presented mtDNA lineages that clearly belong to the European gene pool (i.e. H and U), while the Mashhad population was characterized by the presence of eastern and central Asian mtDNA lineages (i.e. M, B and D). Furthermore, the low diversity (h=0.428) observed in Mashhad may indicated the presence of inbreeding, drift or bottleneck events. The application of Monmonier's maximum differences algorithm revealed a geographic zone of genetic discontinuity between the Arab people of Khuzestan and rest of Iranian populations. Geographical factors, in cooperation with cultural/linguistic differences, are the main reasons for this differentiation. The lack of a sharp geographical or ethno-linguistic structure for mtDNA HV2 sequence diversity was statistically supported by AMOVA and Mantel (r=0.19, P<0.05) tests.