Single primer-based DNA amplification as a suitable and low-cost tool for assessing genetic diversity in mangrove crabs

We used single primer-based DNA markers to assess genetic variability of the mangrove crab, Ucides cordatus, collected from four different localities from Pará to Santa Catarina States in Brazil (almost 5000 km distant). Five primers were chosen based on the consistency of the amplified bands and the polymorphism of each locus. A total of 78 loci were amplified in 76 samples; high polymorphism rates were detected in the entire sample (80.8%) and within each locality (73.5-79.5%). Analysis of molecular variance demonstrates significant differences between localities (P < 0.001); however, the Φ(ST) value (0.078) indicates a low level of genetic differentiation, which suggests that U. cordatus larvae can spread over large distances. The variation was distributed among the samples, and most of it was attributed to differences among individuals within localities. Cluster analysis, based on the Jaccard similarity coefficient, and the Mantel test gave similar results to the analysis of molecular variance data. Despite the low level of population structuring, these markers could be used for studying U. cordatus diversity, due to the high level of polymorphism.     

Polymorphic microsatellite DNA markers in the mangrove crab Ucides cordatus (Brachyura: Ocypodidae)

The isolation and characterization of the first polymorphic microsatellite markers for the mangrove crab Ucides cordatus are described. The number of alleles at each locus ranged from three to 25, mean of nine alleles, in 46 crabs captured in two Brazilian mangroves. The markers averaged high levels of observed (0.709 ± 0.183) and expected (0.716 ± 0.170) heterozygosities. Departures from Hardy-Weinberg equilibrium were observed at two loci. Linkage disequilibrium tests were not significant and no evidence of null alleles was detected. All these microsatellite loci are expected to be useful in estimating fine-scale population processes of this valuable mangrove species currently subjected to excessive fishing efforts.     

PHENOLOGY OF THREE ANT SPECIES IN THE SOLOMON ISLANDS

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Seasonal variation in the composition of colonies is described, with particular reference to the incidence of sexual castes. The ant species are Oecophylla smaragdina F., Iridomyrmex cordatus Fr. Smith and Anoplolepis longipes Jerdon.     

Lethargic crab disease: multidisciplinary evidence supports a mycotic etiology

Although lethargic crab disease (LCD) is causing massive mortalities in populations of the mangrove crab Ucides cordatus of Northeastern Brazil, the identity of its etiological agent was hitherto unknown. In this study we provide robust evidence suggesting that LCD is caused by an anamorph Ascomycota (Fungi). We examined specimens of U. cordatus collected from stocks affected by LCD. Histological and TEM methods detected the presence of hyphae, conidia, and condiophores in several host tissues. Moreover, the abundance of fungal stages is negatively associated with crab health. Finally, DNA was isolated from the fungus and a region of its 18S ribosomal gene was sequenced Phylogenetic analyses not only confirm the diagnosis of the LCD fungus in crab tissues as an ascomycete, but also suggest a close relationship with members of the subphylum Pezizomycotina.     

Distribution of sulfated glycosaminoglycans in the animal kingdom: widespread occurrence of heparin-like compounds in invertebrates

Sulfated glycosaminoglycans were isolated from 23 species of 13 phyla of invertebrates and characterized by their electrophoretic migration in three different buffer systems coupled with enzymatic degradation using bacterial heparinase, heparitinases and chondroitinase AC. Heparan sulfate is a ubiquitous compound present in all species analyzed whereas chondroitin sulfate was present in 20 species and heparin-like compounds in 12 species of the invertebrates. The heparin-like compounds were purified from the echinoderm Mellita quinquisperforata (sand dollar) and the crustacean Ucides cordatus (crab) with anticoagulant activities of 60 and 52 IU/mg, respectively. Degradation of these heparins with heparinase produced significant amounts of the trisulfated disaccharide typical of mammalian heparins. This was confirmed by 13C-NMR spectroscopy of the crab heparin. An updated phylogenetic tree of the distribution of sulfated glycosaminoglycans in the animal kingdom is also presented.     

Cytoarchitectural features of Ucides cordatus (Crustacea Decapoda) hepatopancreas: structure and elemental composition of electron-dense granules

Hepatopancreal tissue of the crab Ucides cordatus was investigated by light and electron microscopy. The observed epithelial cells were: E-cells (embryonic), located in the distal portion of the hepatopancreal tubules, R-cells (resorptive) F-cells (fibrillar) and B-cells (blister or secretory), found in its intermediate and proximal regions. Two types of electron-dense granules (EDGs) were found frequently in the cells of the proximal portion of the hepatopancreal tubule. Both types of EDGs presented alternating concentric electron-dense and electron-lucent layers. In order to better characterize these granules, energy dispersive X-ray analysis (EDXA) and glucose-6-phosphatase (G6Pase) cytochemistry were performed. One type of spherical granule was seen inside vacuoles surrounded by an association of myelin-like membranes as well as some small membrane-bound vesicles. This type of granule neither presented detectable Ca and P on EDXA spectra nor G6Pase cytochemical reaction products. The second type of granule had O, P and Ca characteristic peaks. G6Pase cytochemical products were observed inside these structures and showed that this mineralized type was surrounded by endoplasmic reticulum membranes. This result suggests that in U. cordatus the endoplasmic reticulum is associated with the genesis of mineralized EDGs. While amorphous mineral granules may be associated with a storage of Ca and P for the new carapace synthesis, EDGs covered by the non-mineralized spherical multi-layered membranes may be associated with late endosomes. No specific secretory pathway however was determined for the EDGs at the epithelial proximal portion.     

Fulfilling Koch��s postulates confirms the mycotic origin of Lethargic Crab Disease

In the northeast region of the Brazilian coast, a disease has been causing massive mortalities of populations of the mangrove land crab, Ucides cordatus (L.) since 1997. The clinical signs of this disease, which include lethargy and ataxia, led to the disease being termed Lethargic Crab Disease (LCD). Evidence from a variety of sources indicates that there is an association between LCD and a new species of black yeast, Exophiala cancerae de Hoog, Vicente, Najafzadeh, Badali, Seyedmousavi & Boeger. This study tests this putative correlation through in vivo experiments. Disease-free specimens of U. cordatus were experimentally infected with Exophiala cancerae (strain CBS 120420) isolate. During the 30-day experimental period, only a single death was observed within the control crabs. However, at the end of this period, crabs that were inoculated once or three-times with mycelial elements and hyphae of E. cancerae had a 60% and 50% mortality rates, respectively (n = 6 and n = 5). These results support that the fungal agent is pathogenic and is the causative agent of LCD. Species-specific molecular markers confirm the presence of E. cancerae (strain CBS 120420) in recovered colonies and tissue samples from the infected animals. The experimentally infected crabs manifested signs (lethargy, ataxia and tetany) that were consistent to LCD-affected animals in the environment. These results fulfil Koch's postulates and the hypothesis that the tested strain of Exophiala cancerae is a causative agent of LCD is accepted.     

Ant specificity and behaviour in mutualisms with epiphytes: the case of Lecanopteris (Polypodiaceae)

Seven species of the fern Lecanopteris: L. sinuosa, L. sarcopus, L. mirabilis, L. curtisii, L. pumila, L. celebica and L. damaedii are regularly inhabited by ants of five species: Iridomyrmex cordatus, I. murinus, Crematogaster treubi, C. difformis and Camponotus pallidas. Inhabitation is not obligate; either party can survive without the other, but ferns without ants are rare in their natural habitat. The total recorded diversity of ants in Lecanopteris is 31 species, 20 of which were only recorded once.
All five regularly inhabiting ant species kept larvae in the domatium (82% of Lecanopteris specimens examined). The major ant species inhabiting Lecanopteris deposited debris in the domatium, segregated from their brood (90% of Lecanopteris specimens). Iridomyrmex murinus did not build carton around the rhizomes, but the other major inhabitants constructed runways in 79% of Lecanopteris specimens. Goccid cultivation was variable within ant and plant species: I murinus tended them in 50% of specimens, and homopterans were also recorded with Crematogaster treubi (39%), C. bomeensis (27%) and I cordatus (16%). A high frequency of ant colonies inhabiting Lecanopteris , keeping larvae, depositing debris, building carton runways, and occasionally keeping coccids has been established. Specificity of ant species is high within a population of Lecanopteris , or a given habitat or geographical area, but outside the ranges of regular inhabitants other ant occupants are found. Comparable data from other genera of ant-epiphytes with domatia show similar diversity of inhabitance over geographical areas; no data are available within single plant populations.     

Lower Devonian Polyplacophora from New South Wales, Australia

Silicified polyplacophorans are common in the Lower Devonian Garra Formation in New South Wales. Four taxa occur within the lower 237 m of the formation at 14 horizons associated with an abundant and diverse fauna. Chelodes calceoloides Etheridge ranges through the section. New taxa are Chelodes cordatus sp. nov., Acutichiton depressolatus sp. nov., and Jugochiton patulus gen. and sp. nov. A single plate of an indeterminate genus occurs in the lower portion of the Camelford Limestone.     

Isolation and characterization of microsatellite loci for the collared pika (Ochotona collaris) and their cross-amplification in five other Ochotona species

We developed primers for eight polymorphic microsatellite loci isolated from the collared pika, Ochotona collaris, and also tested nine loci previously developed for the American pika, O. princeps, for use in O. collaris. Forty-six individuals from an O. collaris population in the southern Yukon were genotyped using all 17 loci. The average number of alleles per locus was six and the average observed heterozygosity was 0.59. All loci were tested for use in four Asian pika species and all but two loci amplified reliably in these species.     

Methionine loci and their suppressors in Aspergillus nidulans

Summary Eight methionine loci inAspergillus nidulans were found. Their metabolic blocks and localization were determined. The spontaneous reversion rates for different methionine loci were studied. The revertants were all of suppressor type unlinked or loosely linked with methionine loci. There are at least 8 different suppressor loci. The suppressor mutations are recessive or partly dominant and many of them are quite unspecific, acting on all methionine loci even on those with different metabolic blocks.     

利用分子标记预测杂交水稻产量及其构成因素

利用AFLP、RAPD、SSR技术分析了10个恢复系和5个不育系的931个基因座,利用15个亲本配制了50个杂交组合,在泸州和重庆2个环境下同时种植,考察了产量及其构成因素,从931个基因座中筛选出了与之相关的阳性座位、增效座位、减效座位、非环境型座位,并分析了它们与杂种产量及其构成因素间的关系。结果表明,利用所有座位计算的遗传差异与产量及其构成因素的相关性,绝大多数性状未达显著水平,不能直接用于预测产量及其构成因素。阳性座位在一定程度上可以提高相关系数,因性状不同而存在差异,在多数性状上预测产量及其构成因素还有一定难度;增效座位和减效座位可以大幅度提高相关系数,在不同的环境下也表现一致,可以用来预测产量及其构成因素;非环境型座位计算的相关系数也较高,但低于增效座位和减效座位,说明环境对产量及其构成因素有较大的影响。     

Induction of gene mutations in mice: The multiple endpoint approach

The multiple endpoint mammalian mutagenesis approach developed in our institute screens in the same animal for recessive specific-locus alleles at 7 loci, approximately 30 loci coding for dominant-cataract mutations, 23 loci controlling protein-charge changes and 12 loci for enzyme-activity alterations. Experiments to screen for the approximately 70 loci in the same offspring of treated male mice were performed with ethylnitrosourea (ENU), procarbazine and X-ray exposure. Mutations were recovered for each genetic endpoint in all treatment groups where a sufficient number of offspring was scored. ENU treatment is highly effective in inducing mutations to all genetic endpoints. The mutations were confirmed by breeding tests. The mutation rates to specific-locus and enzyme-activity alleles were both higher than the mutation rates to either dominant-cataract or protein-charge alleles. The advantages and possibilities of the multiple endpoint approach are discussed in detail.     

Analyses of quantitative trait loci associated with resistance to shape Sclerotinia sclerotiorum in sunflowers (shape Helianthus annuus L.) using molecular markers

Restriction fragment length polymorphism and isoenzyme markers were used to investigate quantitative trait loci involved in sunflower resistance to mycelial extension of Sclerotinia sclerotiorum on leaves and capitula. Seed weight, oil content and flowering data were also evaluated. Four quantitative trait loci were demonstrated for leaf resistance and two for capitulum resistance. One of these zones appears involved in resistance to both types of S. sclerotiorum attack while the others appear specific for resistance of one part of the plant. Two quantitative trait loci were detected for seed weight, three for oil content and three for flowering date. Individual quantitative trait loci explained 9% to 48% of the phenotypic variability, confirming the polygenic basis of the quantitative traits studied. Overall, the quantitative trait loci explain 60% of the genetic variation for leaf resistance and 38% for capitulum resistance to S. sclerotiorum. One linkage group is particularly interesting since it includes quantitative trait loci for all the five quantitative traits measured. Hypotheses for linkage versus pleiotropy and consequences of all the results in resistance breeding are discussed.     

Cross-species amplification of 44 microsatellite loci developed for Chaetodon trifascialis, C. lunulatus and C. vagabundus in 22 related butterflyfish species

Forty‐four microsatellite primers developed for three species of butterflyfish were cross‐tested against 22 related confamilial species. Amplification success and cross‐species transferability of these markers were moderately high. Between 24 and 37 loci were amplified successfully in each species, with a mean success rate per species of 71.7% (± 1.8 SE). Rates of amplification success were comparable among primers designed for the three source species, ranging from a mean success rate per species of 16.9 loci (± 0.8 SE) for Chaetodon trifascialis source loci to 13.7 loci (± 1.5 SE) for C. vagabundus source loci. Polymorphism rates were high (76.1%± 3.1 SE of all successfully amplified loci), and 10 loci were polymorphic in all successfully amplified species (Tri14, B11, C5, D3, D113, D6, D117, D120, D111, D118). The number of alleles per polymorphic locus ranged from 2 to 8, and the average number of alleles across all polymorphic loci and all species was 3.6 (± 0.07 SE). Polymorphism rates were higher overall in primers designed for C. vagabundus (89.9%± 3.9 SE). Overall cross‐testing success was lowest for Heniochus chrysostomus, the most phylogenetically divergent species. The significant cross‐testing reported here provides a valuable resource that will enable population genetics studies to be undertaken on a range of butterflyfishes without the need for expensive and time‐consuming de novo microsatellite development.     

Characterization of novel microsatellites and development of multiplex PCR for large‐scale population studies in wild cherry,Prunus avium

Primers were developed for 14 microsatellite or simple sequence repeat (SSR) loci identified from a Prunus avium‘Charger’ genomic DNA library. In a survey of 16 wild cherry accessions 10 of the loci revealed polymorphisms of between two and six alleles. The remaining loci were found to be monomorphic. Seven polymorphic loci identified in this study and four polymorphic loci previously reported in sweet cherry were mapped and found to be unlinked. Two multiplex polymerase chain reactions (PCR) were optimized to enable the characterization of all 11 unlinked, polymorphic SSR loci.     

Separation and viability of gill and hepatopancreatic cells of a mangrove crab Ucides cordatus

The gills contain essential cells for respiration and osmoregulation, whereas the hepatopancreas is the site of digestion, absorption, and nutrients storage. The aim of this work was to separate and characterize gill and hepatopancreatic cells of the mangrove crab, Ucides cordatus. For gills, the methodology consisted of an enzymatic cellular dissociation using Trypsin at 0.5%, observation of cellular viability with Tripan Blue, and separation of cells using discontinuous sucrose gradient at concentrations of 10%, 20%, 30%, and 40%. The hepatopancreatic cells were dissociated by magnetic stirring, with posterior separation by sucrose gradient at the same concentrations above. For gills, a high cellular viability was observed (92.5±2.1%), with hemocyte cells in 10% sucrose layer (57.99?±?0.17%, *P?相似文献   

Genetics of four plasma protein loci in Equus przewalskii: new alleles at the prealbumin, postalbumin and transferrin loci

This paper reports genetic variation at the prealbumin ( Pr ), postalbumin ( Pa ) and transferrin ( Tf ) loci in Equus przewalskii found using thin layer isoelectric focusing and an amphoteric separator. The method resolves all three loci plus serum esterase ( Es ) on a single gel, and typing of all four loci is readily achieved. In addition to the esterase alleles previously reported by Fisher & Scott (1979), five alleles were found at the Pr locus. three at the Pa locus and six at the Tf locus. Analysis of several mating types confirms inheritance is autosomal and codominant for all four loci.     

Microsatellite loci for the mayfly Baetis rhodani (Baetidae,Ephemeroptera)

The first microsatellite primers are described for the mayfly family Baetidae (order Ephemeroptera). Seven polymorphic loci were isolated and characterized for the species Baetis rhodani from two enriched genomic libraries. A total of 183 individuals were genotyped using these loci. Allelic diversity was high for all loci, and observed heterozygosities ranged between 0.382 and 0.772. A heterozygote deficiency was detected in some loci, suggesting the presence of ‘null’ alleles.     

Characterization of polymorphic microsatellite loci from Round Island Petrels (Pterodroma arminjoniana) and their utility in other seabird species

Six microsatellite loci were isolated from the petrels of Round Island, near Mauritius in the Indian Ocean (Pterodroma arminjoniana). Three loci were monomorphic in P. arminjoniana but were found to be polymorphic in other Procellariiforms. Cross-utility of all six loci was tested in 17 Procellariiform and 1 penguin species. In addition, 53 microsatellite loci developed for other species of birds were tested for cross-species amplification in P. arminjoniana. Six of these loci were found to be polymorphic.