Population genetic structure and effective population size of ayu (Plecoglossus altivelis), an amphidromous fish

The temporal and spatial population genetic structure of ayu Plecoglossus altivelis (Salmoniformes: Plecoglossidae), an amphidromous fish, was examined using analysis of variation at six microsatellite DNA loci. Intracohort genetic diversities, as measured by the number of alleles and heterozygosity, were similar among six cohorts (2001–2006) within a population (Nezugaseki River), with the mean number of alleles per cohort ranging from 11·0 to 12·5 and the expected heterozygosity ranging from 0·74 to 0·77. Intrapopulational genetic diversities were also similar across the three studied populations along the 50 km coast, with the mean number of alleles and the expected heterozygosity ranging from 11·33 to 11·67 and from 0·75 to 0·76, respectively. The authors observed only one significant difference in pair-wise population differentiation ( F _ST-value) between the cohorts within a population and among three populations. Estimates of the effective population size ( N _e) based on maximum-likelihood method yielded small values (ranging from 94·8 to 135·5), whereas census population size ranged from c. 4800 to 24 000. As a result, the ratio of annual effective population sizes to census population size ( N _e/ N ) ranged from 0·004 to 0·023. These estimates of N _e/ N agree more closely with estimates for marine fishes than that of the larger estimates for freshwater fishes. The present study suggests that ayu which is highly fecund and shows low survival during the early life stages is also characterized by having low value of N _e/ N , similar to marine species with a pelagic life cycle.     

A population genetics study of Anopheles darlingi (Diptera: Culicidae) from Colombia based on random amplified polymorphic DNA-polymerase chain reaction and amplified fragment lenght polymorphism markers

The genetic variation and population structure of three populations of Anopheles darlingi from Colombia were studied using random amplified polymorphic markers (RAPDs) and amplified fragment length polymorphism markers (AFLPs). Six RAPD primers produced 46 polymorphic fragments, while two AFLP primer combinations produced 197 polymorphic fragments from 71 DNA samples. Both of the evaluated genetic markers showed the presence of gene flow, suggesting that Colombian An. darlingi populations are in panmixia. Average genetic diversity, estimated from observed heterozygosity, was 0.374 (RAPD) and 0.309 (AFLP). RAPD and AFLP markers showed little evidence of geographic separation between eastern and western populations; however, the F ST values showed high gene flow between the two western populations (RAPD: F ST = 0.029; Nm: 8.5; AFLP: F ST = 0.051; Nm: 4.7). According to molecular variance analysis (AMOVA), the genetic distance between populations was significant (RAPD:phiST = 0.084; AFLP:phiST = 0.229, P < 0.001). The F ST distances and AMOVAs using AFLP loci support the differentiation of the Guyana biogeographic province population from those of the Chocó-Magdalena. In this last region, Chocó and Córdoba populations showed the highest genetic flow.     

Genotyping of Edwardsiella ictaluri isolates in Japan using amplified-fragment length polymorphism analysis

Aim:  The major objective of the present study was to clarify genetic relationship of isolates of Edwardsiella ictaluri in Japan, which was first found from ayu Plecoglossus altivelis in Japanese rivers in 2007.
Methods and Results:  Ten isolates of Edw. ictaluri in 2007–2008 from ayu and the 1 isolate from bagrid catfish Pelteobagrus nudiceps in Japan were subjected to amplified-fragment length polymorphism (AFLP) analysis. The strains isolated from catfish in United States (ATCC strains) or Indonesia were used as reference strains. The AFLP profiles were all the same among the isolates from Japan, while the polymorphic DNA bands were observed among the strains from United States or Indonesia. The isolates from Japan and Indonesia constituted a genogroup different from the ATCC strains on a dendrogram constructed from the AFLP profiles.
Conclusion:  No DNA polymorphisms were found among Japanese Edw. ictaluri isolates.
Significance and Impact of the Study:  A single clonality of the Edw. ictaluri isolates in Japan suggests the single source of the organism, and the infection in ayu is in the early stage of epidemics.     

DNA polymorphism in the stem nematode Ditylenchus dipsaci: development of diagnostic markers for normal and giant races.

DNA polymorphism in the Ditylenchus dipsaci complex was investigated using amplified fragment length polymorphism (AFLP) to determine the relationships among populations growing mainly on Vicia faba and to develop diagnostic markers. Twenty-two populations of D. dipsaci originating from different geographical areas and one population of Ditylenchus myceliophagus were used. AFLP proved to be a powerful method to reveal intraspecific polymorphism even within the giant type. The analysis showed a clear distinction between the giant and normal populations, with genetic distances similar to those observed between normal populations and D. myceliophagus or giant populations and D. myceliophagus, strengthening the hypothesis that these two nematode types could be considered distinct species. Two specific AFLP markers differentiating the two types were converted into sequenced characterized amplified region (SCAR) markers. Used in a multiplex PCR, the SCAR primers proved to be a rapid and efficient tool to identify the giant and the normal types of D. dipsaci.     

Phylogeography of the Turkish brown trout Salmo trutta L.: mitochondrial DNA PCR‐RFLP variation

In the present study, mitochondrial DNA polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) assay was used to assess the phylogenetic and phylogeographic relationships among 27 brown trout Salmo trutta populations from Turkey. The complete NADH 5/6 region and a second segment comprising the cytochrome b gene and D‐loop of mtDNA amplified by PCR were digested with six and five restriction enzymes, respectively. A total of 27 haplotypes were observed and divided into three major phylogenetic assemblages, namely Danubian (DA), Adriatic (AD) and a newly proposed Tigris (TI) lineage. The timing of the net nucleotide divergence between the major lineages along with the geological history of Turkey suggested pre‐Pleistocene isolation of the Turkish brown trout and provided evidence that Turkey could be considered as a centre of diversification for these lineages. The average haplotype diversity (0·1397) and the nucleotide diversity (0·000416) within populations were low in comparison to the observed interpopulation nucleotide diversity (0·021266). PCR‐RFLP analysis showed that most of the mtDNA sequence variation found in the Turkish brown trout populations was imputable to differences among lineages. On the other hand, there was also an obvious relationship between geographical distribution of the populations and their clustering. The present study showed that brown trout populations from Turkey are highly divergent and mainly have a unique genetic profile that could be used for conservation and management purposes.     

Evidence for independent origin of two spring‐spawning ciscoes (Salmoniformes: Coregonidae) in Germany

Combined analyses of mitochondrial DNA (mtDNA) and microsatellite loci were performed to assess the genetic differentiation of two spring‐spawning ciscoes from each other and from sympatric Coregonus albula in two German lakes. Polymorphism was screened at six microsatellite loci and mtDNA for a total of 247 and 94 ciscoes, respectively. Microsatellite data showed a weak differentiation between spring‐spawning Coregonus fontanae and sympatric C. albula in Lake Stechlin ( F _ST = 0–0·008), whereas a significant differentiation was observed between spring‐spawning Coregonus lucinensis and sympatric C. albula in Lake Breiter Luzin ( F _ST = 0·013–0·039). A more pronounced genetic difference was observed between both spring‐spawning species ( F _ST = 0·05–0·128). Shared mtDNA haplotypes among sympatric species within both Lake Stechlin and Lake Breiter Luzin were observed, whereas no haplotype was shared between C. fontanae and C. lucinensis . These results suggest an independent origin for spring‐spawning ciscoes in each lake. Evidence is also provided for mtDNA introgression of Coregonus sardinella into C. lucinensis and C. albula in Lake Breiter Luzin. Postglacially, this species or at least a population which showed mtDNA introgression has colonized the Baltic Sea basin up to the glacial margin that was located between Lakes Stechlin and Breiter Luzin.     

Genetic variation and population structure of oriental migratory locust, Locusta migratoria manilensis, in China by allozyme, SSRP-PCR, and AFLP markers

Allozyme analysis, microsatellite primer PCR (SSRP-PCR), and amplified fragment length polymorphism (AFLP) techniques were used to assess genetic diversity and population structure of the Chinese oriental migratory locust, Locusta migratoria manilensis. A total of 299 PCR markers (67 SSRPs and 232 AFLPs) were detected in eight populations, of which 98.7% were polymorphic markers. The proportion of polymorphic loci (95.5-98.8%) by SSRP+AFLP markers indicated no significant differences between populations, and all populations exhibited a similar level of variability; results of the allozyme analysis demonstrated that 19 loci gave rise to a lower level of polymorphism (55.6-66.7%). The genetic distances between the populations were relatively low. Shannon's index and Nei's gene diversity showed low differentiation among the populations. Allozyme analysis, however, reflected greater similarity and smaller differentiation between the populations than those shown by SSRP and AFLP markers. Neighbor-joining dendrograms derived from both the allozyme and SSRP+AFLP markers showed that the genetic distances among Chinese oriental migratory locust populations were not greatly influenced by geographic distance and breeding habitats.     

Genetic Variation and Population Structure of Oriental Migratory Locust, Locusta migratoria manilensis, in China by Allozyme, SSRP-PCR, and AFLP Markers

Allozyme analysis, microsatellite primer PCR (SSRP-PCR), and amplified fragment length polymorphism (AFLP) techniques were used to assess genetic diversity and population structure of the Chinese oriental migratory locust, Locusta migratoria manilensis. A total of 299 PCR markers (67 SSRPs and 232 AFLPs) were detected in eight populations, of which 98.7% were polymorphic markers. The proportion of polymorphic loci (95.5–98.8%) by SSRP+AFLP markers indicated no significant differences between populations, and all populations exhibited a similar level of variability; results of the allozyme analysis demonstrated that 19 loci gave rise to a lower level of polymorphism (55.6–66.7%). The genetic distances between the populations were relatively low. Shannon’s index and Nei’s gene diversity showed low differentiation among the populations. Allozyme analysis, however, reflected greater similarity and smaller differentiation between the populations than those shown by SSRP and AFLP markers. Neighbor-joining dendrograms derived from both the allozyme and SSRP+AFLP markers showed that the genetic distances among Chinese oriental migratory locust populations were not greatly influenced by geographic distance and breeding habitats.     

利用AFLP和SSAP研究短芒大麦突变系种群遗传多样性

利用AFLP和SSAP分子标记,研究了松嫩平原4个以耐盐突变体为主要组成的短芒大麦(Hordeum brevisubulatium(Trin.)Link)种群的遗传多样性和种群遗传结构。由于2种分子标记原理稍有不同,获得分析结果显现出微小差异,但2种方法的结果仍保持了极显著的相关性(r=0.88,P<0.01)。AFLP和SSAP检测结果表明,短芒大麦种群多态位点比率P分别为23.3%和37.7%;Shannon多样性指数分别为0.10和0.14;Nei基因多样性指数分别为0.07和0.09;基因分化系数Gst分别为0.49和0.44。表明,各短芒大麦种群虽以单一突变种为主要组成,但经多年的种植,种群内已具有较高的遗传多样性。AMOVA和聚类分析结果也表明,虽然种群间遗传差异较大,但种群遗传变异仍主要存在于种群内。     

Ultrastructural study of skin and eye of UV‐B irradiated ayu Plecoglossus altivelis

A scanning electron microscopic study of the skin and eye of UV‐B radiated ayu Plecoglossus altivelis (age 30 days, mean ±  s . e . total length: 16·25 ± 0·11 mm) under laboratory condition showed marked changes when compared with the control fish without UV‐B radiation. The exposure of the fish to the radiation resulted in the destruction of microridges in the epidermis and exposed neuromast cells of the skin. Domed protrusions were also more common in the skin of UV‐B radiated fish than in the control fish. The appearance of mucus in both groups was different. In the control skin the mucus was spread over a wide area whereas in the treated fish the mucus was concentrated in a small area. The anastomozing structures of the microridges of the eyes were lost in UV‐B radiated fish and the microridges themselves were fewer in number, fragmented, and aggregated. Mucus cells, prominently visible in the control fish, were distorted in the treated fish. Cell contours were irregular in UV‐B radiated fish and cell to cell contacts had been lost in this group.     

Using dense locality sampling resolves the subtle genetic population structure of the dispersive fish species Plecoglossus altivelis

In dispersive species with continuous distributions, genetic differentiation between local populations is often absent or subtle and thus difficult to detect. To incorporate such subtle differentiation into management plans, it may be essential to analyse many samples from many localities using adequate numbers of high‐resolution genetic markers. Here, we evaluated the usefulness of dense locality sampling in resolving genetic population structure in the ayu (Plecoglossus altivelis), a dispersive fish important in Japanese inland fisheries. Genetic variability in, and differentiation between, ayu populations around the Japan–Ryukyu Archipelago were investigated in 4746 individuals collected from 120 localities by genotyping 12 microsatellite markers. These individuals represented the two subspecies of ayu, namely the Ryukyuan subspecies (Plecoglossus altivelis ryukyuensis) and both amphidromous and landlocked forms of the nominotypical subspecies (P. a. altivelis) along the archipelago. We successfully detected an absence of genetic differentiation within the landlocked form and subtle but significant differentiation and clear geographic patterns of genetic variation among populations of the amphidromous form, which had been considered genetically homogeneous. This suggests that dense locality sampling effectively resolves subtle differences in genetic population structure, reducing stochastic deviation in the detection of genetic differentiation and geographic patterns in local populations of this dispersive species. Resampling analyses based on empirical data sets clearly demonstrate the effectiveness of increasing the number of locality samples for stable and reliable estimations of genetic fixation indices. The genetic population structure observed within the amphidromous form provides useful information for identifying management or conservation units in ayu.     

Comparative analyses of genetic/epigenetic diversities and structures in a wild barley species (Hordeum brevisubulatum) using MSAP, SSAP and AFLP

We analyzed genetic diversity and population genetic structure of four artificial populations of wild barley (Hordeum brevisubulatum); 96 plants collected from the Songnen Prairie in northeastern China were analyzed using amplified fragment length polymorphism (AFLP), specific-sequence amplified polymorphism (SSAP) and methylation-sensitive amplified polymorphism (MSAP) markers. Indices of (epi-)genetic diversity, (epi-)genetic distance, gene flow, genotype frequency, cluster analysis, PCA analysis and AMOVA analysis generated from MSAP, AFLP and SSAP markers had the same trend. We found a high level of correlation in the artificial populations between MSAP, SSAP and AFLP markers by the Mantel test (r > 0.8). This is incongruent with previous findings showing that there is virtually no correlation between DNA methylation polymorphism and classical genetic variation; the high level of genetic polymorphism could be a result of epigenetic regulation. We compared our results with data from natural populations. The population diversity of the artificial populations was lower. However, different from what was found using AFLP and SSAP, based on MSAP results the methylation polymorphism of the artificial populations was not significantly reduced. This leads us to suggest that the DNA methylation pattern change in H. brevisubulatum populations is not only related to DNA sequence variation, but is also regulated by other controlling systems.     

Development of microsatellite loci for Pinus koraiensis (Pinaceae)

? Premise of the study: Microsatellite markers were developed for Pinus koraiensis to characterize its genetic diversity and understand its population structure. ? Methods and Results: Using the Fast Isolation by AFLP of Sequences COntaining (FIASCO) Repeats protocol, 20 primer sets were developed in Chinese populations of P. koraiensis. Three of the markers showed polymorphism with two alleles per locus when assessed in a sample of two populations of P. koraiensis from the Changbai Mountain in the Jilin Province of China. Five and three loci were successfully amplified in P. taiwanensis and P. massoniana, respectively. The amplification size of these loci matches those in P. koraiensis. ? Conclusions: These markers may be useful for further investigation of population genetics of P. koraiensis.     

Development of an AFLP based linkage map of Ler, Col and Cvi Arabidopsis thaliana ecotypes and construction of a Ler/Cvi recombinant inbred line population

An amplified fragment polymorphism (AFLP) based linkage map has been generated for a new Landsberg erecta/Cape Verde Islands (Ler/Cvi) recombinant inbred line (RIL) population. A total of 321 molecular PCR based markers and the erecta mutation were mapped. AFLP markers were also analysed in the Landsberg erecta/Columbia (Ler/Col) RIL population ( Lister & Dean 1993) and 395 AFLP markers have been integrated into the previous Arabidopsis molecular map of 122 RFLPs, CAPSs and SSLPs. This enabled the evaluation of the efficiency and robustness of AFLP technology for linkage analyses in Arabidopsis. AFLP markers were found throughout the linkage map. The two RIL maps could be integrated through 49 common markers which all mapped at similar positions. Comparison of both maps led to the conclusion that segregating bands from a common parent can be compared between different populations, and that AFLP bands of similar molecular size, amplified with the same primer combination in two different ecotypes, are likely to correspond to the same locus. AFLPs were found clustering around the centromeric regions, and the authors have established the map position of the centromere of chromosome 3 by a quantitative analysis of AFLP bands using trisomic plants. AFLP markers were also used to estimate the polymorphism rate among the three ecotypes. The larger polymorphism rate found between Ler and Cvi compared to Ler and Col will mean that the new RIL population will provide a useful material to map DNA polymorphisms and quantitative trait loci.     

Polymorphism, distribution, and segregation of AFLP markers in a doubled haploid rice population

We exploited the newly developed amplified fragment length polymorphism (AFLP) technique to study the polymorphism, distribution and inheritance of AFLP markers with a doubled haploid rice population derived from ‘IR64’/‘Azucena’. Using only 20 pairs of primer combinations, we detected 945 AFLP bands of which 208 were polymorphic. All 208 AFLP markers were mapped and distributed over all 12 chromosomes. When these were compared with RFLP markers already mapped in the population, we found the AFLP markers to be highly polymorphic in rice and to follow Mendelian segregation. As linkage map of rice can be generated rapidly with AFLP markers they will be very useful for marker-assisted backcrossing. Received: 11 April 1996 / Accepted: 14 June 1996     

Differential population history in the migratory catfishes Brachyplatystoma flavicans and Pseudoplatystoma fasciatum(Pimelodidae) from the Bolivian Amazon assessed with nuclear and mitochondrial DNA markers

The catfishes Brachyplatystoma flavicans ( n  = 49) and Pseudoplatystoma fasciatum ( n  = 69) showed comparable low allozyme diversities ( H _e = 0·012 and 0·009–0·028, respectively), but contrasting PCR‐RFLP restriction site mitochondrial DNA diversities (three haplotypes: π = 0·034–0·092 and five haplotypes: π = 0·001–0·023, respectively) in the Rio Ichilo and Beni (Bolivia). Genetic homogeneity between samples was high for B. flavicans and lower for P. fasciatum . Based on mitochondrial diversity, both species probably experienced a historic population reduction but at different time scales.     

Comparison of fish communities between above‐ and below‐dam sections of small streams; barrier effect to diadromous fishes

To analyse the effects of small‐scale dams on fish communities, species diversity, population density, biomass, migration over dams and trophic relationships were investigated in the above‐ and below‐dam sections of three Japanese streams. The dams were 1·5–3·9 m high, constructed within 400 m of the sea, and had no impoundment or fishway. Fish communities were mainly composed of diadromous fishes. Of the 17 species sampled, only a few species of gobies, Rhinogobius spp. and Gymnogobius spp. were able to climb and migrate upstream over the dams. Consequently the number of species, and total density and biomass of fishes below the dams was high, but low above the dams. One of the most dominant fish below dams, Plecoglossus altivelis altivelis , predominantly fed on benthic algae, whereas other fishes preyed on benthic invertebrates. Trophic relationships above dams were very simple, the few species of gobies preying on chironomid larvae and other aquatic invertebrates.     

Inheritance of seed color and molecular markers linked to the seed color gene in <Emphasis Type="Italic">Brassica juncea</Emphasis>

Seed color inheritance in Brassica juncea was studied in F₁, F₂ and BC₁ populations. Seed color was found under the control of the maternal genotype, and the brown-seeded trait was dominant over the yellow-seeded trait. Segregation analysis revealed that one pair of major genes controlled the seed coat color. To develop markers linked to the seed color gene, AFLP (amplified fragments length polymorphism) combined with BSA (bulk segregant analysis) technology was used to screen the parents and bulks selected randomly from an F₂ population (Wuqi yellow mustard × Wugong mustard) consisting of 346 individuals. From a survey of 512 AFLP primer combinations, 15 AFLP markers located on either side of the gene were identified, and the average distance between markers was 2.59 cM. P11MG15 was a cosegregated marker, and the closest markers (P03MC08, P16MC02 and P11MG01) were at a distance of 0.3, 0.3 and 0.7 cM from the target gene, respectively. In order to utilize the markers for breeding of yellow-seeded varieties, four AFLP markers, P11MG01, P15MG15, P09MC12 and P16MC02 were successfully converted into SCAR (sequence characterized amplified region) markers. The seed color trait controlled by the single gene together with the available molecular markers will greatly facilitate the future breeding of yellow-seeded varieties. The markers found in the present study could accelerate the step of map-based cloning of the target gene.     

Development of microsatellite markers for the moss Ptychomitrium gardneri (Ptychomitriaceae)

? Premise of the study: Microsatellite markers were developed for Ptychomitrium gardneri to study population genetics of this eastern Asian-North American disjunct moss. ? Methods and Results: A total of 13 microsatellite markers were developed in Chinese populations of P. gardneri, using the Fast Isolation by AFLP of Sequences COntaining Repeats protocol. Eight of the markers showed polymorphism when assessed in a sample of four populations of 29 individuals from China. These markers amplified three to four alleles per locus. Five primers also amplified in P. linearifolium and P. wilsonii. ? Conclusions: These markers may be useful for further investigation of population genetics of P. gardneri.     

Increased inbreeding and strong kinship structure in Taxus baccata estimated from both AFLP and SSR data

Habitat fragmentation can have severe genetic consequences for trees, such as increased inbreeding and decreased effective population size. In effect, local populations suffer from reduction of genetic variation, and thus loss of adaptive capacity, which consequently increases their risk of extinction. In Europe, Taxus baccata is among a number of tree species experiencing strong habitat fragmentation. However, there is little empirical data on the population genetic consequences of fragmentation for this species. This study aimed to characterize local genetic structure in two natural remnants of English yew in Poland based on both amplified fragment length polymorphism (AFLP) and microsatellite (SSR) markers. We introduced a Bayesian approach that estimates the average inbreeding coefficient using AFLP (dominant) markers. Results showed that, in spite of high dispersal potential (bird-mediated seed dispersal and wind-mediated pollen dispersal), English yew populations show strong kinship structure, with a spatial extent of 50–100 m, depending on the population. The estimated inbreeding levels ranged from 0.016 to 0.063, depending on the population and marker used. Several patterns were evident: (1) AFLP markers showed stronger kinship structure than SSRs; (2) AFLP markers provided higher inbreeding estimates than SSRs; and (3) kinship structure and inbreeding were more pronounced in denser populations regardless of the marker used. Our results suggest that, because both kinship structure and (bi-parental) inbreeding exist in populations of English yew, gene dispersal can be fairly limited in this species. Furthermore, at a local scale, gene dispersal intensity can be more limited in a dense population.