Polymorphic microsatellite DNA markers for the ark shell Scapharca subcrenata (bivalve: Arcidae)

We have isolated and characterized twelve novel polymorphic microsatellite loci from Scapharca subcrenata to analyse the population structure. The number of observed alleles per locus ranged from 3 to 17. Observed heterozygosity (H _O) ranged from 0.321 to 0.929. Cross-species amplification was tested successfully in three other bivalve species. These microsatellite markers will be useful for genetic diversity studies of S. subcrenata and other Lamellibranchia species.     

Polymorphic microsatellite loci in the Coffee Berry Borer,Hypothenemus hampei (Coleoptera,Scolytidae)

Microsatellite loci were isolated from the coffee berry borer, Hypothenemus hampei, which is regarded as the major pest in coffee cultures. Seven polymorphic loci were obtained from an enriched genomic library. A low to moderate genetic diversity was observed per locus, with an observed number of alleles ranging from two to five in the 39 unrelated individuals sampled from Ethiopia. A clear deficit of heterozygotes within the population (mean heterozygosities, H_O = 0.10/H_E = 0.50) and an extreme inbreeding (F_IS, 0.70–1.00) were demonstrated. Cross‐species amplifications showed that some of the markers could be useful in two closely related Hypothenemus species.     

Polymorphic microsatellite loci from the Southeast Asian cyprinid,Barbodes gonionotus (Bleeker)

The cyprinid Barbodes gonionotus (Bleeker) is a commercially important fish in both capture fisheries and aquaculture in Southeast Asia. Five polymorphic microsatellite loci from B. gonionotus are described. Four are highly variable, with 9–30 alleles observed per locus in four populations sampled from Thailand (H_O = 0.694–0.808). These will be of use in studies of population genetic structure and in pedigree analyses.     

How does population genetic diversity change over time? An experimental seed bank study of Atriplex tatarica (Chenopodiaceae)

Atriplex tatarica is an annual, early successional, facultative halophilic species of frequently disturbed human-made habitats in Central and Eastern Europe. We investigated to what extent the plants grown from seeds extracted from soil seed bank differed genetically to mature aboveground plants in experimental populations of A. tatarica over two successive years. At each of five plots 50 aboveground plants and 50 plants extracted from seeds stored in soil were assayed for allozyme analysis in 2003 and 2004. At the start of experiment, we introduced 1000 seeds of the study species into each of five experimental plots. While the species dominated in all of the experimental plots in the first year, the second year A. tatarica coverage decreased dramatically. Overall allele frequencies of soil seeds and mature plants showed significant differences between life history stages in both years, but not within years in soil seeds as well as mature plants stages. While mature plants showed a significantly greater amount of single and multilocus heterozygosity in both consecutive years, comparison between years did not yield any significant differences. In the same way, despite a relatively large seed bank the species population genetic parameters, i.e. allelic richness (A), observed heterozygosity (H_o), gene diversity (H_s), inbreeding coefficient (F_IS) and fixation index (F_ST), did not change over years between as well as within life history stages. The soil seeds and mature plants significantly differed in H_o, H_s and F_IS, while the A and F_ST were not significantly different between life history stages.     

Isolation and characterization of 20 polymorphic microsatellite loci for Scaptodrosophila hibisci

Scaptodrosophila hibisci is an endemic Australian Drosophilidae that breeds in the flowers of native Hibiscus . Here we report the isolation and amplification of 20 polymorphic microsatellite loci . We cloned these microsatellites because loci developed for Drosophila melanogaster failed to amplify in S. hibisci . Null alleles were detected at six loci, and five were X‐linked. Two of the primer pairs amplified an unlinked ‘bonus’ locus. One locus containing juxtaposed microsatellite loci was suitable for designing an additional set of primers. Mean number of alleles per locus was 10, mean H _O and H _E per locus were 0.532 and 0.636, respectively.     

Population structure of the olive flounder (Paralichthys olivaceus) in Korea inferred from microsatellite marker analysis

The population structure of olive flounder Paralichthys olivaceus was estimated using nine polymorphic microsatellite (MS) loci in 459 individuals collected from eight populations, including five wild and three hatchery populations in Korea. Genetic variation in hatchery (mean number of alleles per locus, A = 10·2–12·1; allelic richness, A_R = 9·3–10·1; observed heterozygosity, H_O = 0·766–0·805) and wild (mean number of alleles per locus, A = 11·8–19·6; allelic richness, A_R = 10·9–16·1; observed heterozygosity, H_O = 0·820–0·888) samples did not differ significantly, suggesting a sufficient level of genetic variation in these well‐managed hatchery populations, which have not lost a substantial amount of genetic diversity. Neighbour‐joining tree and principal component analyses showed that genetic separation between eastern and pooled western and southern wild populations in Korea was probably influenced by restricted gene flow between regional populations due to the barrier effects of sea currents. The pooled western and southern populations are genetically close, perhaps because larval dispersal may depend on warm currents. One wild population (sample from Wando) was genetically divergent from the main distribution, but it was genetically close to hatchery populations, indicating that the genetic composition of the studied populations may be affected by hydrographic conditions and the release of fish stocks. The estimated genetic population structure and potential applications of MS markers may aid in the proper management of P. olivaceus populations.     

Identification of microsatellite loci from Epimedium koreanum and cross‐species amplification in four species of Epimedium (Berberidaceae)

Nineteen polymorphic microsatellite markers were isolated and characterized from a trinucleotide enriched partial genomic library of Epimedium koreanum. The average allele number of these microsatellites was 5.9 per locus, ranging from two to 11. The observed heterozygosity (H_O) and expected heterozygosity (H_E) at the population level were 0.00–0.90 and 0.12–0.90, respectively. In addition, the results of cross‐species amplification of this set of microsatellite markers in four closely related Epimedium species, E. brevicornum, E. sagittatum, E. pubescens and E. wushanense, revealed that these microsatellite markers were useful for population genetic structure evaluation and genotype analysis of major Epimedium species that have been used as traditional Chinese medicines.     

Characterization of seven genomic and one dbEST-derived microsatellite loci in the river mangrove<Emphasis Type="Italic"> Aegiceras corniculatum</Emphasis> (Myrsinaceae)

We developed seven microsatellite markers from the genome of Aegiceras corniculatum using the FIASCO protocol and one dbEST-derived microsatellite marker. Polymorphism of each locus was assessed in 45 individuals from China, Thailand and Australia. The average allele number of these microsatellites was 4.4 per locus, ranging from 2 to 8. The observed (H _O ) and expected (H _E ) heterozygosity were from 0.178 to 1.000 and from 0.164 to 0.828, respectively. These microsatellite markers would provide a useful tool for genetic and conservation studies of A. corniculatum.     

Isolation of microsatellites from two species of scleractinian coral

Microsatellites were isolated from two broadcast‐spawning species of scleractinian coral (Platygyra daedalea and Goniastrea favulus) from Australia's Great Barrier Reef. We found 27 microsatellites across both species, although only five loci were polymorphic in each species. Microsatellite loci displayed a wide range of diversity levels with four to 11 alleles per locus (H_O = 0.26–0.91) in P. daedalea and two to seven alleles per locus (H_O = 0.16–0.96) in G. favulus. Most loci showed departures from Hardy–Weinberg equilibrium which may reflect nonrandom mating but may also be related to difficulties associated with coral DNA.     

Reaction of Phosphinyl and Phosphinothioyl Disulfides with Diazomethane

Four kinds of 4-substituted phenyl diethoxyphosphinyl disulfide (substitueras: H, CH₃, C₂H₅, Cl) and phenyl diethoxyphosphinothioyl disulfide reacted readily with diazomethane. The phosphinyl disulfides produced diethyl (4-substituted phenylthio)methyl phosphorothionate and the isomer, diethyl S-(4-substituted phenylthio)methyl phosphorothiolate, as the main products, whereas the phosphinothioyl disulfide produced only diethyl S-(phenylthio)methyl phosphorodithioate.

The possible mechanism involved is as follows: the S-S bond in (EtO)₂P(X)-S-S-C₆H₄R (X = O or S) is cleaved by nucleophilic attack of diazomethane at the sulfur atom bonded to the phenyl group to produce and ^⊕ N₂CH₂–S–C₆H₄R; the latter reacts with the former with loss of nitrogen; therefore, when X is O, two compounds, (EtO)₂P(S)–O–CH₂–S–C₆H₄R and (EtO)₂P(O)–S–CH₂–S–C₆H₄R, are produced, when X is S, only (EtO)₂P(S)–S–CH₂ –S–C₆H₄R is obtained.     

Inhibitory effects of enterococci on the production of hydrogen sulfide by hydrogen sulfide-producing bacteria in raw meat

Aims: Applying competitive exclusion micro‐organisms to control hydrogen sulfide (H₂S) gas produced by hydrogen sulfide–producing bacteria (SPB) in chicken meat. Methods and Results: Five SPB strains, isolated from animal by‐products, were used for screening lactic acid bacteria (LAB) that can inhibit the production of H₂S by SPB in trypticase soy broth supplemented with l ‐cysteine (TSB‐l ‐cys). A sensitive and accurate test strip method was developed for H₂S determination in real time. One LAB strain, isolate L86, from cheese whey, demonstrated the highest inhibitory activity against the production of H₂S by SPB. The isolate L86 was confirmed as Enterococcus faecium that does not possess genes encoding for vancomycin resistance based on PCR analysis. Enterococcus faecium strain L86 reduced (P < 0·05) the yield of H₂S upto 51·2% in 10 h at 35°C in TSB‐l ‐cys medium. In fresh chicken meat, the yield of H₂S produced by the artificially inoculated SPB was reduced (P < 0·05) by 48·6, 49·7 and 69·8% in 10 h at 35, 30 and 25°C, respectively. Enterococcus faecium strain L86 also reduced (P < 0·05) by 53·8% on the yield of H₂S produced by the indigenous SPB in partially spoiled chicken meat at 35°C for 10 h. Conclusions: Enterococcus faecium strain L86 is effective on inhibiting the production of H₂S by SPB. Significance and Impact of the Study: The application of this biological agent to raw animal by‐products will provide a safer working environment in rendering processing plants and produce higher‐quality rendered products.     

Isolation and characterization of polymorphic microsatellite markers from Australia short-finned eel (Anguilla australis Richardson)

Seventeen microsatellite DNA loci from the Australian short‐finned eel (Anguilla australis Richardson) were isolated and their amplification characteristics were described. The polymerase chain reaction primers were tested on 40 eel individuals. The primers amplified loci with relatively high numbers of alleles, ranging from five to 14 with an average of nine per locus. Mean observed heterozygosity (H_O) and expected heterozygosity (H_E) were 0.6779 and 0.7374, respectively, indicating that these markers would be useful for population studies. No loci deviated significantly from Hardy–Weinberg equilibrium (P = 0.05) and no evidence was found for genotypic disequilibrium among loci at a 5% significance level.     

Polymorphic microsatellite primers developed from DNA of the endangered Mekong giant catfish,Pangasianodon gigas (Chevey) and cross‐species amplification in three species of Pangasius

The critically endangered Pangasianodon gigas is endemic to the Mekong River. Despite its importance, little is known about its genetic diversity and conservation efforts are hampered. Ten polymorphic dinucleotide microsatellite primer pairs were developed from DNA of P. gigas. The analysis of 20 individuals from hatchery stocks using these primers resulted in two to six alleles/locus; H_O = 0.05–0.95; H_E = 0.05–0.81. All but one locus (Pg‐3) conformed to Hardy–Weinberg expectation. Eight, six and seven primer pairs were amplified with the DNA from Pangasianodon hypophthalmus, Pangasius larnaudii and Pangasius sanitwongsei, respectively. These markers will be useful for genetic monitoring of wild and hatchery stocks of these pangasiids.     

Eight polymorphic microsatellite loci for the Chinese medicinal plant Artemisia annua L. (Asteraceae)

Artemisia annua is an important medicinal plant from which Artemisinin was extracted to cure malaria effectively. We developed eight microsatellite markers from the genome of A. annua using the FIASCO protocol. Polymorphism of each locus was assessed in 54 individuals from two Chinese populations. The average allele number of these microsatellites was 3.1 per locus, ranging from 2 to 6. The observed (H _O) and expected (H _E) heterozygosity were from 0.019 to 0.907 and from 0.055 to 0.793, respectively. These microsatellite markers would provide a useful tool for genetic studies of A. annua. H.-R. Huang and G. Zhou have contributed equally to this work.     

Isolation and characterization of microsatellite markers for Ligularia hodgsonii Hook. (Asteraceae)

Ligularia hodgsonii is one of widely distributed Ligularia species in south China, central China, Far East area of Russia and Japan. In this study, we described the development of 14 microsatellite markers from the genome of L. hodgsonii using the FIASCO protocol. Polymorphism of each locus was assessed in 30 adult individuals of the Ligularia. The average allele number of the microsatellites was 3.0 per locus, ranging from 2 to 5. The observed (H _O) and expected (H _E) heterozygosities varied from 0.2000 to 0.7333 and from 0.3910 to 0.7598. The marker transferability of the 14 primer pairs was tested on other two congeneric species that also occur in China.     

Analysis of molecular genetic diversity and population structure in Amaranthus germplasm using SSR markers

We assessed the molecular genetic diversity and population structure of Amaranthus species accessions using 11 simple sequence repeat markers. A total of 122 alleles were detected, and the number of alleles per marker (N_A) ranged from 6 to 21 with an average of 11.1 alleles. The frequency of major alleles per locus ranged from 0.148 to 0.695, with an average value of 0.496 per marker. The overall polymorphic information content values were 0.436–0.898, with an average value of 0.657. The observed heterozygosity (H_O) and expected heterozygosity (H_E) ranged from 0.056 to 0.876 and from 0.480 to 0.907, with average values of 0.287 and 0.698, respectively. The average H_O (0.240) was lower than the H_E and gene flow (Nm), and showed substantial genetic variability among all populations of amaranth accessions. The sample groupings did not strictly follow the geographic affiliations of the accessions. A similar pattern was obtained using model-based structure analysis without grouping by species type. Knowledge of the genetic diversity and population structure of amaranth can be used to select representative genotypes and manage Amaranthus germplasm breeding programs.     

Development of SSR primers using ISSR–PCR in Diospyros kaki Thunb.

Nine polymorphic single sequence repeat (SSR) primers were developed in Japanese persimmon using inter‐SSR (ISSR) suppression polymerase chain reaction (PCR). These primers were tested on 30 individuals from Japan and China. The number of alleles per locus ranged from five to 20. Expected (H_E) and observed (H_O) heterozygosities at each locus ranged from 0.42 to 0.77 and 0.27 to 0.59, respectively. The SSR primers developed herein could be applied to cultivar identification, estimation of genetic diversity and divergence in Diospyros spp.     

Notes on skulls of Pleistocene Saiga of Northern Eurasia

Eighteen fossil skulls of male Saiga from Northern Eurasia and 33 recent skulls from Kalmykia and Kazakhstan have been studied. Saiga from both the Khazarian Fauna of the Volga and Mammoth Fauna of Europe and Siberia are referred to Saiga horealis Tschersky, 1876. During the Pleistocene, 5. borealis distribution extended from England in the west to Alaska in the east and is characterized by an elongated neurocranium, small frontal angle of the temporal bone from the plane of the frontal, and long nasal bones.

S. borealis was a typical representative of the “mammoth biome”; in the Pleistocene periglacial steppes and cryogenic savannahs. Two subspecies are recognized: S. borealis borealis Tschersky (Eastern Sibera and Alaska); and S.b. prisca Nehring, 1891 (Europe, Urals and Western Siberia). At the end of the Pleistocene, when the mammoth disappeared, the range of S. borealis was reduced. Today they live only in West Mongolia (S. borealis mongolica Bannikov, 1946). S. tatarica tatarica was widely distributed in the other territories of the steppe and semidesert zones of Eurasia. The arid landscapes of Transcaucasia and Kazakhstan were inhabited by Saiga with thinner legs and shorter nasal bones, such as S. tatarica binagadensis Alekperova, 1953, from the middle Pleistocene of Azerbaijan (Bynagady). Fossil skulls from the Ural River that are large, but with a short neurocranium are identified as Saiga sp. cf. S. tatarica Linnaeus, 1766.     

Isolation and characterization of microsatellite loci in Fagus longipetiolata Seem. (Fagaceae)

Eleven microsatellite loci have been developed from Fagus longipetiolata and the loci were characterized for 21 individuals. All eleven loci were polymorphic, with 2–8 alleles and an average of 4.8 per locus. The observed (H _O) and expected (H _E) heterozygosities were 0.053–0.714 and 0.355–0.856, respectively. There was significant deviation from Hardy–Weinberg equilibrium at two loci. No locus pair had significant linkage disequilibrium. Cross-species amplifications of the markers were also tested in three other congeneric species.     

Microbial community structure in a biofilm anode fed with a fermentable substrate: The significance of hydrogen scavengers

We compared the microbial community structures that developed in the biofilm anode of two microbial electrolysis cells fed with ethanol, a fermentable substrate—one where methanogenesis was allowed and another in which it was completely inhibited with 2‐bromoethane sulfonate. We observed a three‐way syntrophy among ethanol fermenters, acetate‐oxidizing anode‐respiring bacteria (ARB), and a H₂ scavenger. When methanogenesis was allowed, H₂‐oxidizing methanogens were the H₂ scavengers, but when methanogenesis was inhibited, homo‐acetogens became a channel for electron flow from H₂ to current through acetate. We established the presence of homo‐acetogens by two independent molecular techniques: 16S rRNA gene based pyrosequencing and a clone library from a highly conserved region in the functional gene encoding formyltetrahydrofolate synthetase in homo‐acetogens. Both methods documented the presence of the homo‐acetogenic genus, Acetobacterium, only with methanogenic inhibition. Pyrosequencing also showed a predominance of ethanol‐fermenting bacteria, primarily represented by the genus Pelobacter. The next most abundant group was a diverse community of ARB, and they were followed by H₂‐scavenging syntrophic partners that were either H₂‐oxidizing methanogens or homo‐acetogens when methanogenesis was suppressed. Thus, the community structure in the biofilm anode and suspension reflected the electron‐flow distribution and H₂‐scavenging mechanism. Biotechnol. Bioeng. 2010;105: 69–78. © 2009 Wiley Periodicals, Inc.