High-resolution Molecular Markers Research Articles

Diversity and seasonal occurrence of haptophyta in northern South China Sea through size-fractionated metabarcoding

Haptophyta plays a key role in marine pico-nanoeukaryote communities but information on their diversity and ecology is extremely limited. A total of 103 water samples were collected in northern South China Sea to assess the diversity of haptophyta through metabarcoding targeting 18S V4 rDNA. Furthermore, we investigated the potential genetic differentiation among seasonal occurring Phaeocystis globosa using the high resolution molecular marker pgcp1. 18S V4 rDNA metabarcoding dataset revealed 41 species of haptophytes, with 16 of them as the first record in this region. Notably, six harmful species were detected, including Chrysochromulina leadbeateri, Phaeocystis globosa, and Prymnesium parvum. The pgcp1 marker revealed two clades of Phaeocystis globosa and both of them were present around the year. Clade I was found to predominate in warm season, while Clade III tended to bloom in cold waters. Our results highlight the risk potential of harmful haptophytes in the northern South China Sea.

New Insights into Phylogenetic Relationship of Hydrocotyle (Araliaceae) Based on Plastid Genomes

Hydrocotyle, belonging to the Hydrocotyloideae of Araliaceae, consists of 95 perennial and 35 annual species. Due to the lack of stable diagnostic morphological characteristics and high-resolution molecular markers, the phylogenetic relationships of Hydrocotyle need to be further investigated. In this study, we newly sequenced and assembled 13 whole plastid genomes of Hydrocotyle and performed comparative plastid genomic analyses with four previously published Hydrocotyle plastomes and phylogenomic analyses within Araliaceae. The plastid genomes of Hydrocotyle exhibited typical quadripartite structures with lengths from 152,659 bp to 153,669 bp, comprising a large single-copy (LSC) region (83,958–84,792 bp), a small single-copy (SSC) region (18,585–18,768 bp), and a pair of inverted repeats (IRs) (25,058–25,145 bp). Each plastome encoded 113 unique genes, containing 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. Comparative analyses showed that the IR boundaries of Hydrocotyle plastomes were highly similar, and the coding and IR regions exhibited more conserved than non-coding and single-copy (SC) regions. A total of 2932 simple sequence repeats and 520 long sequence repeats were identified, with specificity in the number and distribution of repeat sequences. Six hypervariable regions were screened from the SC region, including four intergenic spacers (IGS) (ycf3-trnS, trnS-rps4, petA-psbJ, and ndhF-rpl32) and two coding genes (rpl16 and ycf1). Three protein-coding genes (atpE, rpl16, and ycf2) were subjected to positive selection only in a few species, implying that most protein-coding genes were relatively conserved during the plastid evolutionary process. Plastid phylogenomic analyses supported the treatment of Hydrocotyle from Apiaceae to Araliaceae, and topologies with a high resolution indicated that plastome data can be further used in the comprehensive phylogenetic research of Hydrocotyle. The diagnostic characteristics currently used in Hydrocotyle may not accurately reflect the phylogenetic relationships of this genus, and new taxonomic characteristics may need to be evaluated and selected in combination with more comprehensive molecular phylogenetic results.

Single Nucleotide Polymorphism Markers with Applications in Conservation and Exploitation of Aquatic Natural Populations.

An increasing number of aquatic species have been studied for genetic polymorphism, which extends the knowledge on their natural populations. One type of high-resolution molecular marker suitable for studying the genetic diversity of large numbers of individuals is single nucleotide polymorphism (SNP). This review is an attempt to show the range of applications of SNPs in studies of natural populations of aquatic animals. In recent years, SNPs have been used in the genetic analysis of wild and enhanced fish and invertebrate populations in natural habitats, exploited migratory species in the oceans, migratory anadromous and freshwater fish and demersal species. SNPs have been used for the identification of species and their hybrids in natural environments, to study the genetic consequences of restocking for conservation purposes and the negative effects on natural populations of fish accidentally escaping from culture. SNPs are very useful for identifying genomic regions correlated with phenotypic variants relevant for wildlife protection, management and aquaculture. Experimental size-selective catches of populations created in tanks have caused evolutionary changes in life cycles of fishes. The research results have been discussed to clarify whether the fish populations in natural conditions can undergo changes due to selective harvesting targeting the fastest-growing fishes.

Insights into taxonomy and phylogenetic relationships of eleven Aristolochia species based on chloroplast genome.

The Aristolochia, as an important genus comprised of over 400 species, has attracted much interest because of its unique chemical and pharmacological properties. However, the intrageneric taxonomy and species identification within Aristolochia have long been difficult because of the complexity of their morphological variations and lack of high-resolution molecular markers. In this study, we sampled 11 species of Aristolochia collected from distinct habitats in China, and sequenced their complete chloroplast (cp) genomes. The 11 cp genomes of Aristolochia ranged in size from 159,375bp (A. tagala) to 160,626 bp (A. tubiflora), each containing a large single-copy (LSC) region (88,914-90,251 bp), a small single-copy (SSC) region (19,311-19,917 bp), and a pair of inverted repeats (IR) (25,175-25,698 bp). These cp genomes contained 130-131 genes each, including 85 protein-coding genes (CDS), 8 ribosomal RNA genes, and 37-38 transfer RNA genes. In addition, the four types of repeats (forward, palindromic, reverse, and complement repeats) were examined in Aristolochia species. A. littoralis had the highest number of repeats (168), while A. tagala had the lowest number (42). The total number of simple sequence repeats (SSRs) is at least 99 in A. kwangsiensis, and, at most, 161 in A. gigantea. Interestingly, we detected eleven highly mutational hotspot regions, including six gene regions (clpP, matK, ndhF, psbT, rps16, trnK-UUU) and five intergenic spacer regions (ccsA-ndhD, psbZ-trnG-GCC, rpl33-rps18, rps16-trnQ-UUG, trnS-GCU-trnG-UCC). The phylogenetic analysis based on the 72 protein-coding genes showed that 11 Aristolochia species were divided into two clades which strongly supported the generic segregates of the subgenus Aristolochia and Siphisia. This research will provide the basis for the classification, identification, and phylogeny of medicinal plants of Aristolochiaceae.

The Complete Chloroplast Genome Sequence of Eupatorium fortunei: Genome Organization and Comparison with Related Species.

Eupatorium fortunei Turcz, a perennial herb of the Asteraceae family, is one of the horticultural and medicinal plants used for curing various diseases and is widely distributed in China and other Asian countries. It possesses antibacterial, antimetastatic, antiangiogenic, and antioxidant properties along with anticancer potential. However, the intrageneric classification and phylogenetic relationships within Eupatorium have long been controversial due to the lack of high-resolution molecular markers, and the complete chloroplast (cp) genome sequencing has not been reported with new evolutionary insights. In the present study, E. fortunei was used as an experimental material, and its genome was sequenced using high-throughput sequencing technology. We assembled the complete cp genome, and a systematic analysis was conducted for E. fortunei, acquiring the correspondence of its NCBI accession number (OK545755). The results showed that the cp genome of E. fortunei is a typical tetrad structure with a total length of 152,401 bp, and the genome encodes 133 genes. Analysis of the complete cp genomes of 20 Eupatorieae shows that the number of simple sequence repeats (SSRs) ranged from 19 to 36 while the number of long sequence repeats was 50 in all cases. Eleven highly divergent regions were identified and are potentially useful for the DNA barcoding of Eupatorieae. Phylogenetic analysis among 22 species based on protein-coding genes strongly supported that E. fortunei is more closely related to Praxelis clematidea and belongs to the same branch. The genome assembly and analysis of the cp genome of E. fortunei will facilitate the identification, taxonomy, and utilization of E. fortunei as well as provide more accurate evidence for the taxonomic identification and localization of Asteraceae plants.

Genetic Analysis of a Large-Scale Phaeocystis globosa Bloom Offshore Qingdao, China.

A sudden large-scale bloom event of the haptophyte Phaeocystis globosa that lasted over one month in the winter of 2021 was observed offshore Qingdao, China. This P. globosa bloom event was unusual as it was the first P. globosa bloom recorded in Qingdao offshore. Furthermore, the temperature at which this event occurred was much lower than that of previous P. globosa blooms in China. We hypothesize that the P. globosa strains that drove the development of this bloom offshore Qingdao were genetically unique and have a competitive advantage in the environmental conditions. To test this hypothesis, we analyzed P. globosa genetic diversity and the temporal dynamics of the bloom, using the high-resolution molecular markers pgcp1 and cox1 that we developed recently. The analysis revealed that the genetic compositions of P. globosa offshore Qingdao were rather limited, containing two dominant genotypes and other rare genotypes with low abundance, representing a small portion of the genetic diversities identified in coastal waters in China, and were rather different from the P. globosa genotypes outside of the Jiaozhou Bay before the P. globosa bloom in the winter of 2021. This suggested only certain strains contribute to the development of blooms under certain environmental conditions. The genetic composition may indicate the unusual timing and scale of this P. globosa event.

Comparative Analysis of Chloroplast Genome and New Insights Into Phylogenetic Relationships of Polygonatum and Tribe Polygonateae.

Members of Polygonatum are perennial herbs that have been widely used in traditional Chinese medicine to invigorate Qi, moisten the lung, and benefit the kidney and spleen among patients. However, the phylogenetic relationships and intrageneric taxonomy within Polygonatum have long been controversial because of the complexity of their morphological variations and lack of high-resolution molecular markers. The chloroplast (cp) genome is an optimal model for deciphering phylogenetic relationships in related families. In the present study, the complete cp genome of 26 species of Trib. Polygonateae were de novo assembled and characterized; all species exhibited a conserved quadripartite structure, that is, two inverted repeats (IR) containing most of the ribosomal RNA genes, and two unique regions, large single sequence (LSC) and small single sequence (SSC). A total of 8 highly variable regions (rps16-trnQ-UUG, trnS-GCU-trnG-UCC, rpl32-trnL-UAG, matK-rps16, petA-psbJ, trnT-UGU-trnL-UAA, accD-psaI, and trnC-GCA-petN) that might be useful as potential molecular markers for identifying Polygonatum species were identified. The molecular clock analysis results showed that the divergence time of Polygonatum might occur at ∼14.71 Ma, and the verticillate leaf might be the ancestral state of this genus. Moreover, phylogenetic analysis based on 88 cp genomes strongly supported the monophyly of Polygonatum. The phylogenetic analysis also suggested that Heteropolygonatum may be the sister group of the Polygonatum, but the Disporopsis, Maianthemum, and Disporum may have diverged earlier. This study provides valuable information for further species identification, evolution, and phylogenetic research of Polygonatum.

Intense blooms of Phaeocystis globosa in the South China Sea are caused by a unique “giant-colony” ecotype

The haptophyte Phaeocystis globosa, an important causative agent of harmful algal blooms globally, exhibits varying morphological and physiological features and high genetic diversity, yet the relationship among these has never been elucidated. In this study, colony sizes and pigment profiles of 19 P. globosa isolates from the Pacific and Atlantic Oceans were determined. Genetic divergence of these strains was analyzed using the chloroplast rbcS-rpl27 intergenic spacer, a novel high-resolution molecular marker. Strains could be divided into four genetic clades based on these sequences, or two groups based on colony size and the identity of diagnostic pigments (19'-hexanoyloxyfucoxanthin, hex-fuco, and 19'-butanoyloxyfucoxanthin, but-fuco). Three strains from the South China Sea (SCS), all belonging to the same genetic clade, have unique biological features in forming giant colonies and possessing but-fuco as their diagnostic pigment. Based on these findings, we propose that these SCS strains should be a unique "giant-colony" ecotype of P. globosa. During the period 2016-2021, more than 1000 rbcS-rpl27 sequences were obtained from 16 P. globosa colony samples and 18 phytoplankton samples containing solitary P. globosa cells in the SCS. Phylogenetic analysis indicated that >95% of the sequences from P. globosa colonies in the SCS were comprised of the "giant-colony" ecotype, whereas the genetic diversity of solitary cells was much higher. Results demonstrated that intense blooms of P. globosa featuring giant colonies in the SCS were mainly caused by this giant-colony P. globosa ecotype.

Phylogenomic analysis of the chloroplast genome of the green-tide forming macroalga Ulva intestinalis Linnaeus (Ulvophyceae, Chlorophyta)

Ulva intestinalis Linnaeus 1753 (Ulvophyceae, Chlorophyta) is a marine green macroalga that is distributed on coasts of the Yellow Sea and the Bohai Sea in China. Here, the complete chloroplast genome of U. intestinalis was constructed and analyzed comparatively. The chloroplast genome of U. intestinalis is a 99,041-bp circular molecule that harbors a total of 112 genes including 71 protein-coding genes (PCGs), 26 transfer RNA genes (tRNAs), three ribosomal RNA genes (rRNAs), three free-standing open reading frames (orfs) and nine intronic orfs, and ten introns in seven genes (atpA, infA, psbB, psbC, petB, rrnL, and rrnS). The maximum likelihood (ML) phylogenomic analysis shows that U. intestinalis firstly groups with Ulva compressa, and then these two species together with the Ulva australis–Ulva fenestrata–Ulva rotundata subclade form a monophyletic clade, Ulva lineage II. U. intestinalis chloroplast genome is the only one in Ulva lineage II where the reversal of a collinear block of two genes (psbD–psbC) did not occur, and its genome structure is consistent with that of most chloroplast genomes in Ulva lineage I, indicating that the similarity of genome structure is not completely related to the genetic relationship of Ulva species. Our genomic data will facilitate the development of specific high-resolution chloroplast molecular markers for rapid identification of U. intestinalis, and help us understand its population diversity and genetic characteristics on a global scale.

Research on the biology and ecology of the harmful algal bloom species Phaeocystis globosa in China: Progresses in the last 20 years

Chinese researchers have made substantial progresses in research on the harmful algal bloom (HAB) species Phaeocystis globosa since the first P. globosa bloom outbreak in the Chinese coastal waters in 1997. However, as many results, especially the earlier ones, were published in non-English literature, much of the research on P. globosa biology, ecology, and biogeochemistry made by Chinese researchers have been unknown to colleagues from other countries. We review current knowledge on taxonomy, morphology, genetics, physiology, survival strategies and mitigation of P. globosa gained by Chinese researchers from the past two decades. P. globosa is the only Phaeocystis species that causes blooms in the Chinese waters, although other Phaeocystis species including P. jahnii and P. cordata have been detected in Chinese coastal regions. P. globosa has a complex life history with at least two morphotypes including a haploid flagellate and a diploid colonial cell. Colonial P. globosa blooms typically occur in winter after a diatom bloom in coastal waters of the South China Sea. P. globosa in Chinese coastal waters usually has extremely large colonial sizes, up to 3 cm in diameter, an order of magnitude greater than that observed in European coastal waters. The development of giant colonies is associated with enhanced sinking rate, limited nutrient diffusion, as well as decreased stability of colonies. The Chinese P. globosa strains also showed strong genetic diversity and physiological plasticity, being able to grow and develop into colonies at higher temperature and irradiance relative to that in European waters. High genetic diversity of P. globosa was revealed by developing high-resolution and high-specificity molecular markers including Phaeocystis globosa chloroplast 1 (pgcp1). Due to the severe impact of P. globosa on ecology and economy, much effort has been made to mitigate P. globosa blooms including the application of modified clays. Overall, P. globosa in the Chinese waters demonstrate unique genetic, phenotypical and physiological features that differ from P. globosa in other ocean regions. Further studies are needed to explore how environmental factors trigger the occurrence of P. globosa blooms and ascertain the impact of P. globosa blooms on the environment.

Comparative chloroplast genome analysis of medicinally important Veratrum (Melanthiaceae) in China: Insights into genomic characterization and phylogenetic relationships

Members of Veratrum are perennial herbs widely used in traditional Chinese medicine to induce vomiting, resolve blood stasis and relieve pain. However, the intrageneric classification and phylogenetic relationships within Veratrum have long been controversial due to the complexity of morphological variations and lack of high-resolution molecular markers. In this study, we reevaluated the infrageneric relationships with the genus Veratrum using complete chloroplast genome sequence data. Herein, the complete cp genomes of ten species of Veratrum were newly sequenced and characterized. The complete cp genomes of ten species of Veratrum had the typical quadripartite structure, ranging from 151,597 bp to 153,711 bp in size and comprising a total of 135 genes. The structure of Veratrum cp genomes (i.e., gene order, content, and genome components) was highly similar across species. The number of simple sequence repeats (SSRs) ranged from 63 to 78, and of long repeats ranged from 31 to 35. Eight highly divergent regions (ndhF, psbC-psbZ, psbK-psbI, rpoB-trnC_GCA, trnK_UUU-trnQ_UUG, trnS_GCU-trnG_UCC, trnT_UGU-trnL_UAA and ycf1) were identified and are potentially useful for the DNA barcoding of Veratrum. Phylogenetic analysis among 29 taxa based on cp genomes, total genes, protein-coding genes and intergenic regions strongly supported the monophyly of Veratrum. The circumscription and relationships of the infrageneric taxa of Veratrum were well-presented with great resolution. These results will facilitate the identification, taxonomy, and utilization of Veratrum plants as well as the evolutionary studies of Melanthiaceae.

Development of a high-resolution molecular marker for tracking Pseudo-nitzschia pungens genetic diversity through comparative analysis of mitochondrial genomes

The harmful algal bloom (HAB) species Pseudo-nitzschia pungens is widely distributed in almost all continents. Accumulating evidence suggests that P. pungens has high genetic diversity and many strains can produce the toxin domoic acid (DA) that harms animals and humans. Nevertheless, different P. pungens strains cannot be distinguished using morphological features or using common molecular markers including 18S rDNA, 28S rDNA, ITS, cox1, and rbcL. As such, high-resolution molecular markers need to be developed to resolve P. pungens genetic diversity, facilitating accurate tracking of toxic P. pungens strains. We hypothesized that molecular markers with high resolution and high specificity can be designed through identifying regions with high genomic variations in the mitochondrial genome. Here, we describe the development of a new molecular marker Pseudo-nitzschia pungens mitochondrial 1 (ppmt1) with high resolution and high specificity through comparative analysis of mitochondrial genomes of nine P. pungens strains isolated from coastal regions of China. In conclusion, we have developed ppmt1 as a high-resolution and high-specificity molecular marker for tracking strains and genetic diversity of the HAB species P. pungens.

Isolation and characterization of SNP markers of rainbow trout (Onchorhynchus mykiss Walbaum, 1792) from transcriptomic sequences

Rainbow trout (Onchorhynchus mykiss) is one of the most important freshwater aquaculture fish in Iran. It is necessary to develop available molecular marker such as SNPs, which represent a useful tool in detecting adaptive signals in populations and also parentage assignment for O. mykiss. Genetic architecture of broodstock populations is important for breeding programs, as it enables decisions on broodstock screening and genomic selection. In this study, 52 novel single nucleotide polymorphism (SNP) markers for O. mykiss were discovered and validated based on transcriptome sequencing, by means of paired-end sequencing in an Illumina HiSeq 2500 platform. The SNPs were identified through liver transcriptome sequencing from fifteen samples. The observed and expected heterozygosities ranged from 0.177 to 1.000 and 0.239 to 0.638, respectively. The minimum allele frequency (MAF) ranged from 0.166 to 0.489. Among these SNP loci, twenty-two loci showed significant departures from the Hardy-Weinberg equilibrium after Bonferroni correction (p < 0.05) and significant linkage disequilibrium was found. The SNP markers identified in this research could be useful for novel studies, such as those related to associations between high-resolution molecular markers and quantitative traits studies. Moreover, these SNP markers would be used in genetic studies helping economic performance improvement and management of this species.

Development of a high-resolution molecular marker for tracking Phaeocystis globosa genetic diversity through comparative analysis of chloroplast genomes

The phytoplankton Phaeocystis globosa thrives in a wide range of marine regions and plays an important role in climate control. It can also form harmful algal blooms (HABs) that threaten environments and impact important coastal infrastructures. Mechanisms underlying the formation of P. globosa blooms still remain poorly understood. Accumulating evidence suggests that P. globosa has high genetic diversity and different P. globosa strains may have differential contributions to the development of P. globosa blooms. However, due to the lack of molecular markers with adequate resolution for distinguishing P. globosa genetic diversity, such differential contributions by different P. globosa strains could not be accurately ascertained. As such, high-resolution molecular markers need to be developed and applied to distinguish P. globosa genetic diversity. In this study, we undertook to define high-resolution molecular marker by assembling and comparing the whole chloroplast genomes of P. globosa strains isolated from different regions of the world. Through comparative analysis of P. globosa cpDNAs and detection of single nucleotide variations (SNVs), a molecular marker pgcp1 with improved resolution was developed. The pgcp1 demonstrated the highest resolution compared with other regions including 18S rDNA V4 region, 28S rDNA D1–D2 region and rbcL region, through genetic distance and phylogenetic analysis of 13 P. globosa strains. Molecular analysis of environmental samples and strains collected in multiple expeditions from a wide range of ocean regions including multiple regions in China, Vietnam, Thailand and Western Pacific using pgcp1 as the molecular marker displayed high genetic diversity of P. globosa with at least four major P. globosa clades. In conclusion, we have developed pgcp1 as a high-resolution molecular marker for the harmful algal bloom species P. globosa, which can be used to track intra-species genetic diversity and dynamics of P. globosa during bloom development.

Transcriptional variation of sensory-related genes in natural populations of Aedes albopictus

BackgroundThe Asian tiger mosquito, Aedes albopictus, is a highly dangerous invasive vector of numerous medically important arboviruses including dengue, chikungunya and Zika. In four decades it has spread from tropical Southeast Asia to many parts of the world in both tropical and temperate climes. The rapid invasion process of this mosquito is supported by its high ecological and genetic plasticity across different life history traits. Our aim was to investigate whether wild populations, both native and adventive, also display transcriptional genetic variability for functions that may impact their biology, behaviour and ability to transmit arboviruses, such as sensory perception.ResultsAntennal transcriptome data were derived from mosquitoes from a native population from Ban Rai, Thailand and from three adventive Mediterranean populations: Athens, Greece and Arco and Trento from Italy. Clear inter-population differential transcriptional activity was observed in different gene categories related to sound perception, olfaction and viral infection. The greatest differences were detected between the native Thai and the Mediterranean populations. The two Italian populations were the most similar.Nearly one million quality filtered SNP loci were identified.ConclusionThe ability to express this great inter-population transcriptional variability highlights, at the functional level, the remarkable genetic flexibility of this mosquito species. We can hypothesize that the differential expression of genes, including those involved in sensory perception, in different populations may enable Ae. albopictus to exploit different environments and hosts, thus contributing to its status as a global vector of arboviruses of public health importance.The large number of SNP loci present in these transcripts represents a useful addition to the arsenal of high-resolution molecular markers and a resource that can be used to detect selective pressure and adaptive changes that may have occurred during the colonization process.

Incursion pathways of the Asian tiger mosquito (Aedes albopictus) into Australia contrast sharply with those of the yellow fever mosquito (Aedes aegypti)

Understanding pest incursion pathways is critical for preventing new invasions and for stopping the transfer of alleles that reduce the efficacy of local control methods. The mosquitoes Aedes albopictus (Skuse) and Ae. aegypti (Linnaeus) are both highly invasive disease vectors, and through a series of ongoing international incursions are continuing to colonize new regions and spread insecticide resistance alleles among established populations. This study uses high-resolution molecular markers and a set of 241 reference genotypes to trace incursion pathways of Ae. albopictus into mainland Australia, where no successful invasions have yet been observed. We contrast these results with incursion pathways of Ae. aegypti, investigated previously. Assignments successfully identified China, Japan, Singapore and Taiwan as source locations. Incursion pathways of Ae. albopictus were entirely different to those of Ae. aegypti, despite broad sympatry of these species throughout the Indo-Pacific region. Incursions of Ae. albopictus appeared to have come predominantly along marine routes from key trading locations, while Ae. aegypti was mostly linked to aerial routes from tourism hotspots. These results demonstrate how genomics can help decipher otherwise cryptic incursion pathways. The inclusion of reference genotypes from the Americas may help resolve some unsuccessful assignments. While many congeneric taxa will share common incursion pathways, this study highlights that this is not always the case, and incursion pathways of important taxa should be specifically investigated. Species differences in aerial and marine incursion rates may reflect the efficacy of ongoing control programmes such as aircraft disinsection. © 2020 Society of Chemical Industry.

Comparative analyses of 32 complete plastomes of Tef (Eragrostis tef ) accessions from Ethiopia: phylogenetic relationships and mutational hotspots.

Eragrostis tef is an important cereal crop in Ethiopia with excellent storage properties, high–quality food, and the unique ability to thrive in extreme environmental conditions. However, the application of advanced molecular tools for breeding and conservation of these species is extremely limited. Therefore, developing chloroplast genome resources and high-resolution molecular markers are valuable to E. tef population and biogeographic studies. In the current study, we assembled and compared the complete plastomes of 32 E. tef accessions. The size of the plastomes ranged from 134,349 to 134,437 bp with similar GC content (∼38.3%). Genomes annotations revealed 112 individual genes, including 77 protein-coding, 31 tRNA, and 4 rRNA genes. Comparison of E. tef plastomes revealed a low degree of intraspecific sequence variations and no structural differentiations. Furthermore, we found 34 polymorphic sites (13 cpSSRs, 12 InDels, and 9 SNPs) that can be used as valuable DNA barcodes. Among them, the majority (88%) of the polymorphic sites were identified in the noncoding genomic regions. Nonsynonymous (ka) and synonymous (ks) substitution analysis showed that all PCGs were under purifying selection (ka/ks <1). The phylogenetic analyses of the whole plastomes and polymorphic region sequences were able to distinguish the accession from the southern population, indicating its potential to be used as a super-barcode. In conclusion, the newly generated plastomes and polymorphic markers developed here could be a useful genomic resource in molecular breeding, population genetics and the biogeographical study of E. tef.

Molecular genotyping, diversity studies and high-resolution molecular markers unveiled by microsatellites in Giardia duodenalis.

BackgroundGiardia duodenalis (synonyms G. lamblia and G. intestinalis) is an enteric protozoan parasite of a wide range of mammalian hosts, including humans and various domestic and wild animals. There is considerable genetic variability in G. duodenalis and isolates of this parasite have been divided into eight genetic assemblages. Microsatellites markers can be used to discriminate isolates with a high level of sensitivity. This study was conducted to identify and characterize genomic microsatellites (simple sequence repeats—SSRs), sequences of one- to six-nucleotide motifs repeated in tandem, present in the available genomes of G. duodenalis and to develop new markers that can serve as a tool for detection and for characterizing the genetic diversity of this parasite.Methodology/ Principal findingsFor each genetic assemblage, polymorphism levels for the microsatellite markers were evaluated. After performing the analysis using the MISA and SciRoKo software, 1,853 simple sequence repeats (SSRs) were identified. In all the genomes, trinucleotide repeats were the most common class followed by tetranucleotide. Many of the SSR loci are assemblage-specific, and 36 SSR loci shared among all the genomes were identified. Together with hypothetical proteins, variant-specific surface proteins represented nearly half of the annotated SSR loci. The results regarding the most common repeat among the SSRs led us to infer that positive selection occurred to avoid frameshift mutations. Additionally, based on inter- and intra-genetic assemblages polymorphism analyses, we unveiled previously undetected genetic variation, indicating that the microsatellite markers we developed are useful molecular tools for epidemiological inferences based on population genetics patterns and processes.ConclusionsThere is increasing demand for the development of new molecular markers and for the characterization of pathogens at a higher resolution level. In this study, we present 60 G. duodenalis microsatellites markers that exhibited high polymerase chain reaction (PCR) amplification efficiency among the different genetic assemblages. Twenty of these markers presented nucleotide sequence polymorphisms and may be used as a genotyping tool. The monomorphic markers can be used for the detection of the parasite at the species and genetic assemblage level. These polymorphic markers revealed a genetic diversity that was previously undetectable, thus they can be considered valuable molecular tools for high resolution markers in future studies investigating Giardia and may also be used for epidemiological inferences based on populations genetics patterns and processes.

Construction of introgression lines of Oryza rufipogon and evaluation of important agronomic traits.

Cultivated rice (Oryza sativa) originated from common wild rice (Oryza rufipogon) and inherited its advantages. However, during the rice domestication process, some valuable features of wild rice, such as tolerance to biotic and abiotic stress, were lost. To fully utilize wild rice germplasm resources, we constructed a set of introgression lines (ILs) using a common wild rice material from Lingshui, China. A set of high-resolution InDel molecular markers with an average interval of 2.39Mb were designed to carry out marker-assisted selection and identification of segment characteristics. The ILs contained 77 lines including 1.286 introgressed fragments with an average length of 6.511Mb, covering 93.59% of the donor parent's chromosomes. The agricultural traits of 77 lines were investigated. Many old quantitative trait loci (QTLs) involved in plant height, awn length, seed traits and other characteristics reappeared in our ILs, proving that our system was reliable. Further, many new QTLs were identified. A QTL related to drought tolerance located on chromosome 4 was thoroughly elaborated. This set of ILs provides a new resource for utilizing the excellent features of wild rice.

Aspects, Including Pitfalls, of Temporal Sampling of Flying Insects, with Special Reference to Aphids.

Since the advent and widespread use of high-resolution molecular markers in the late 1970s, it is now well established that natural populations of insects are not necessarily homogeneous genetically and show variations at different spatial scales due to a variety of reasons, including hybridization/introgression events. In a similar vein, populations of insects are not necessarily homogenous in time, either over the course of seasons or even within a single season. This of course has profound consequences for surveys examining, for whatever reason/s, the temporal population patterns of insects, especially flying insects as mostly discussed here. In the present article, the topics covered include climate and climate change; changes in ecological niches due to changes in available hosts, i.e., essentially, adaptation events; hybridization influencing behaviour–host shifts; infection by pathogens and parasites/parasitoids; habituation to light, sound and pheromone lures; chromosomal/genetic changes affecting physiology and behaviour; and insecticide resistance. If such phenomena—i.e., aspects and pitfalls—are not considered during spatio-temporal study programmes, which is even more true in the light of the recent discovery of morphologically similar/identical cryptic species, then the conclusions drawn in terms of the efforts to combat pest insects or conserve rare and endangered species may be in error and hence end in failure.