bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetic analysis of populations of threatened snake species using RAPD markers
Authors: GIBBS H. L. , PRIOR K. A. , WEATHERHEAD P. J.
Published: April 2008

Snakes are a particularly threatened vertebrate taxon, with distributions of many species and populations becoming increasingly fragmented. At present, little is known about the degree of genetic differentiation that exists between isolated populations even though such information may be critical to their survival and conservation. As an example of how recently developed RAPD genetic markers can be used in conservation genetics, we present preliminary results from a study which used these DNA‐based markers to assess population divergence in two threatened Canadian snakes, the black rat snake (Elaphe o. obsoleta) and the eastern massasauga rattlesnake (Sistrurus c. catenatus). We present information on the levels of variation and reliability of amplification for fragments generated from five primers. We then use a recently developed analytical technique to estimate levels of nucleotide diversity within populations and sequence divergence between populations. Our results show that intrapopulation levels of divergence as estimated by the methods of Clark & Lanigan (Molecular Biology and Evolution 1993, 10, 1096–1111) approximate those found for mtDNA in vertebrates and that diversity between snake populations is small and non‐significant when tested using randomization procedures. Thus, our study provides an example of how RAPDs can be applied to conservation genetic studies of vertebrates and suggest that the snake populations we examined have only recently become isolated and maybe considered genetically equivalent from a conservation perspective, although this conclusion needs to be confirmed with other DNA‐based markers.
Low genetic variability in a natural alpine marmot population (Marmota marmota, Sciuridae) revealed by DNA fingerprinting
Authors: RASSMANN K. , ARNOLD W. , TAUTZ D.
Published: April 2008

Genetic heterogeneity is usually considered an important factor for the viability of a population, yet there are cases in which populations sustain themselves despite virtual homozygosity. A prior step to studying the effects of such low levels of genetic variability can be the analysis of its causes. We analysed a population of the highly social alpine marmot (Marmota marmota, Sciuridae) by multilocus DNA fingerprinting. The fingerprint patterns revealed a very low degree of polymorphism in our main study population. We show that this lack of hypervariability is caused by a low effective population size, rather than by an unusual low mutation rate of the fingerprint loci studied. However, the current number of breeding pairs was found to be about an order of magnitude larger than the one that would be expected to lead to such a low degree of heterozygosity. We conclude that there must have been bottlenecks in the history of the Berchtesgaden marmot population that have severely affected its genetic heterozygosity.
Mitochondrial DNA control region polymorphisms: genetic markers for ecological studies of marine turtles
Authors: NORMAN J. A. , MORITZ C. , LIMPUS C. J.
Published: April 2008

We describe a rapid and sensitive method for the detection of population‐specific genetic markers in mitochondrial DNA (mtDNA) and the use of such markers to analyse population structure of marine turtles. A series of oligonucleotide primers specific for the amplification of the mtDNA control region in Cheloniid turtles were designed from preliminary sequence data. Using two of these primers, a 384–385‐bp sequence was amplified from the 5′ portion of the mtDNA control region of 15 green turtles Chelonia mydas from 12 different Indo‐Pacific rookeries. Fourteen of the 15 individuals, including some with identical whole‐genome restriction fragment patterns, had sequences that differed by one or more base substitutions. Analysis of sequence variation among individuals identified a total of 41 nucleotide substitutions and a 1‐bp insertion/deletion. Comparison with evidence from whole‐genome restriction enzyme analysis of the same individuals indicated that this portion of the control region is evolving approximately eight times faster than the average rate and that the sequence analysis detected approximately one fifth of the total variation present in the genome. Restriction enzyme analysis of amplified products from an additional 256 individuals revealed significant geographic structuring in the distribution of mtDNA genotypes among five of the 10 rookeries surveyed extensively. Additional geographic structuring of genotypes was identified through denaturing gradient gel electrophoresis (DGGE) of amplified products. Only two of the 10 rookeries surveyed could not be differentiated, indicating that the Indo‐Pacific C. mydas include a number of genetically differentiated populations, with minimal female‐mediated gene flow among them. Important applications for genetic markers in the conservation and management of marine turtles include the identification of appropriate demographic units for research and management (i.e. genetically discrete populations) and assessment of the composition of feeding and harvested populations.
Genetic variation of the St. Lawrence beluga whale population assessed by DNA fingerprinting
Authors: PATENAUDE N. J. , QUINN J. S. , BELAND P. , KINGSLEY M. , WHITE B. N.
Published: April 2008

Recent surveys suggest that the endangered St. Lawrence beluga (Delphinapterus leucas) population is not recovering significantly despite 20 years of protection. Dead individuals that have been autopsied show high levels of tumours and infections. This situation could be a result of pollution, loss of genetic variation, inbreeding depression or a combination of these factors. Analyses of DNA fingerprints from St. Lawrence belugas with three minisatellite probes (Jeffreys 33.6, 33.15 and M13) indicate a reduced level of genetic variation compared to Beaufort Sea animals. The average band‐sharing between individuals of the St. Lawrence beluga population for the three probes (0.534, 0.573 and 0.478, respectively) was significantly higher than that of the Beaufort Sea beluga population (0.343, 0.424, 0.314, respectively). Higher levels of mean allele frequency in the St. Lawrence belugas (0.33 vs. 0.21) suggest that this population is composed of individuals which are related. Inbreeding depression could therefore be a factor in the lack of recovery of the St. Lawrence beluga population.
Microbial diversity in soil: effect of releasing genetically engineered micro‐organisms
Authors: LEUNG K. , ENGLAND L. S. , CASSIDY M. B. , TREVORS J. T. , WEIR S.
Published: April 2008

This review examines the potential for change in microbial diversity, with the emphasis on bacteria, in soil resulting from the introduction of genetically engineered microorganisms (GEMs). With the advent of GEMs came the impetus for new technologies to recover these micro‐organisms from soil and to assess their effects on microbial diversity. This review also presents general aspects of and genetic approaches to accessing bacterial diversity in the environment.
Conservation genetics: beyond the maintenance of marker diversity
Authors: MILLIGAN B. G. , LEEBENS‐MACK J. , STRAND A. E.
Published: April 2008

One of the major problems faced by conservation biologists is the allocation of scarce resources to an overwhelmingly large number of species in need of preservation efforts. Both demographic and genetic information have been brought to bear on this problem; however, the role of information obtained from genetic markers has largely been limited to the characterization of gene frequencies and patterns of diversity. While the genetic consequences of rarity may be a contributing factor to endangerment, it is widely recognized that demographic factors often may be more important. Because patterns of genetic marker variation are influenced by the same demographic factors of interest to the conservation biologist, it is possible to extract useful demographic information from genetic marker data. Such an approach may be productive for determining plant mating systems, inbreeding depression, effective population size, and metapopulation structure. In many cases, however, data consisting only of marker frequencies are inadequate for these purposes. Development of genealogical based analytical methods coupled with studies of DNA sequence variation within and among populations is likely to yield the most information on demographic processes from genetic marker data. Indeed, in some cases it may be the only means of obtaining information on the long‐term demographic properties that may be most useful for determining the future prospects of a species of interest.
Genetic evidence for cuckoldry in bluegill Lepomis macrochirus
Authors: PHILIPP D. P. , GROSS M. R.
Published: April 2008

In colonies of bluegill Lepomis macrochirus, some males provide parental care for broods in nests, whereas other males steal fertilizations and do not provide parental care. Using experimental pond populations of bluegill of known genotype (determined through protein electrophoresis), we demonstrate that cuckolder males successfully fertilize eggs in parental male nests. Using electrophoretic techniques to assess the fertilization success of nesting parental male bluegill in Lake Opinicon, Ontario, we demonstrate that paternity of these males ranges from 41% to 100% among four colonies studied. This difference among colonies is related to the density of cuckolder males.
Monitoring the persistence of Laccaria bicolor as an ectomycorrhizal symbiont of nursery‐grown Douglas fir by PCR of the rDNA intergenic spacer
Authors: HENRION B. , BATTISTA C. DI , BOUCHARD D. , VAIRELLES D. , THOMPSON B. D. , TACON F. LE , MARTIN F.
Published: April 2008

The large‐scale inoculation of selected beneficial ectomycorrhizal fungi in forest nurseries has generated renewed interest in the ecology of these symbiotic fungi. However, information on the dissemination and persistence of introduced symbionts is scarce due to the limitation of the current identification methods. To identify ectomycorrhizal fungi on single root tips, we investigated the polymorphism of the PCR‐amplified ribosomal DNA intergenic spacer (IGS) from a wide range of ectomycorrhizal fungi. To investigate the reliability of this molecular approach in large‐scale surveys, the dissemination and persistence on Douglas fir seedlings of the introduced Laccaria bicolor S238N were assessed in a forest nursery in the Massif Central (France). Several hundred ectomycorrhizas and fruiting bodies were sampled from plots where control and L. bicolor inoculated‐Douglas fir seedlings were grown for 1.5 years. PCR typing of mycorrhizas indicated that trees inoculated with L. bicolor S238N remained exclusively colonized by that isolate (or sexually derived isolates) for the entire test period. In contrast, control seedlings were infected by indigenous isolates of Laccaria laccata and Thelephora terrestris. The molecular evidence for the persistence of the introduced mycobiont despite the competition from indigenous isolates of the same species provides further illustration of the potential of exotic species for large‐scale microbial application.
Population differentiation in randomly amplified polymorphic DNA of red‐cockaded woodpeckers Picoides borealis
Authors: HAIG S. M. , RHYMER J. M. , HECKEL D. G.
Published: April 2008

Random amplified polymorphic DNA (RAPD) phenotypes generated by 13 primers were scored for 101 individuals in 14 populations of the endangered red‐cockaded woodpecker Picoides borealis. Although no population‐specific markers were found, the frequencies of several markers differed significantly among populations. Application of the recently developedamova method (analysis of molecular variance; Excoffier, Smouse & Quattro 1992) showed that more than 90% of phenotypic variance occurred among individuals within populations; of the remaining variance, half was attributed among groups of geographically adjacent populations and half among populations within those groups. The statistical significance of these patterns was supported by Monte Carlo sampling simulations and permutation tests. Estimation of allele frequencies from phenotypes provided somewhat weaker evidence for population structure, although among‐population variance in allele frequencies was detectable (Fst= 0.19; x2169= 509.3, P < 0.0001). Upgma cluster analyses based on Rogers' (1972) genetic distance revealed grouping of some geographically proximate populations. A Mantel test indicated a positive (r = 0.16), although not significant, correlation between geographic and genetic distances. We compared a subset of our RAPD data with data from a previous study that used allozymes (Stangel, Lennartz & Smith 1992). RAPD (n= 75) and allozyme (n= 245) results based on samples from the same ten populations showed similar patterns. Our study indicates that RAPDs can be helpful in differentiating populations at the phenotypic level even when small sample sizes, estimation bias, and inability to test for Hardy‐Weinberg equilibrium complicate the genotypic interpretation. Lack of large differences among populations of red‐cockaded woodpeckers may allow flexibility in overpopulation translocations, provided factors such as habitat preference, latitudinal direction of translocation, and status of donor populations are considered.
Reproductive isolation and the period gene of Drosophila
Authors: RITCHIE M. G. , KYRIACOU C. P.
Published: April 2008

The identification of genes of large effect on ecologically important traits is an important aim of molecular ecology. The period gene of Drosophila is a candidate for a gene with a large influence on premating isolation between Drosophila species, as it determines species specific aspects of courtship behaviour. Strains of D. melanogaster are available which have been genetically transformed with the period gene of either D. melanogaster or D. simulans. Here we show that D. melanogaster females do not discriminate between two such strains. This suggests that period may only make a small contribution to total premating isolation between these species. We discuss the use of genetically transformed strains in assessing the influence of single genes on complex traits.
Transduction of a freshwater microbial community by a new Pseudomonas aeruginosa generalized transducing phage, UT1
Authors: RIPP S. , OGUNSEITAN O. A. , MILLER R. V.
Published: April 2008

A pseudolysogenic, generalized transducing bacteriophage, UT1, isolated from a natural freshwater habitat, is capable of mediating the transfer of both chromosomal andplasmid DNA between strains of Pseudomonas aeruginosa. Several chromosomal alleles from three different P. aeruginosa strains were found to transduce at frequencies from 10‐8 to 10‐10 transductants per PFU at multiplicities of infection (MOD between 0.1 and 1. Transduction frequencies of certain alleles increased up to 1000‐fold as MOIs were decreased to 0.01. UT1 is also capable of transducing plasmid DNA to indigenous populations of microorganisms in natural lake‐water environments. Data obtained in this study suggest that environmentally endemic bacteriophages such as UT1 are formidable transducers of naturally occurring microbial communities. It should be possible to develop model systems to test transduction in freshwater environments using components derived exclusively from these environments.
Epistatic effects of promoter and repressor functions of the Tn10 tetracycline‐resistance operon on the fitness of Escherichia coli
Authors: LENSKI R. E. , SOUZA V. , DUONG L. P. , PHAN Q. G. , NGUYEN T. N. M. , BERTRAND K. P.
Published: April 2008

We have been studying the effects of expression of plasmid‐borne, Tn10‐encoded, tetracycline resistance on the fitness of Escherichia coli K12. We previously demonstrated large reductions in fitness resulting from induced or constitutive expression of the resistance protein; however, any residual expression by the repressed operon was so slight that possession of an inducible resistance function imposed essentially no burden in the absence of antibiotic. Here, we demonstrate two distinct disadvantages for inducible genotypes relative to isogenic constitutive constructs. During the transition from antibiotic‐free to antibiotic‐containing media, the inducible genotype experiences a longer lag phase prior to growth. In the sustained presence of antibiotic, full induction of the resistance function in the inducible genotype is prevented by the continued action of its repressor. However, these disadvantages may be reduced by increasing the strength of the promoter for the resistance gene in the inducible genotype. Simultaneous consideration of the mode of gene regulation (i.e. constitutive or inducible) and the strength of the resistance‐gene promoter (i.e. maximum level of expression) indicates an adaptive landscape with very strong epistasis and, perhaps, multiple fitness peaks.
Identification of hypervariable single locus minisatellite DNA probes in the blue tit Parus caeruleus
Authors: VERHEYEN G. R. , KEMPENAERS B. , BURKE T. , BROECK M. VAN DEN , BROECKHOVEN C. VAN , DHONDT A.
Published: April 2008

We report the isolation of a set of hypervariable minisatellite DNA sequences from a blue tit Parus caeruleus genomic DNA library. In our strategy, we cloned a minisatellite‐rich DNA fraction into a charomid vector. The resulting cosmid library was screened with the two minisatellite DNA probes 33.6 and 33.15 for recombinants containing a minisatellite DNA insert. A total of 233 positive clones were isolated. Of 37 clones that have been analysed, nine gave polymorphic signals and can be used as single locus probes (SLPs). Four of the SLPs were investigated in more detail. The number of alleles, the heterozygosity and the mutation rate were estimated. Linkage analysis revealed that two of these loci were linked. The SLPs are of value to studies of the mating system and reproductive success in the blue tit, and may also be useful in population genetic studies.
Quantification in soil of Bacillus thuringiensis var. kurstakiδ‐endotoxin from transgenic plants
Authors: PALM C. J. , DONEGAN K. , HARRIS D. , SEIDLER R. J.
Published: April 2008

Transgenic plants that produce pesticidal proteins have the potential to release these products into the environment when the plants are incorporated into soil. This could result in novel exposure of soil organisms to these pesticidal proteins. There is a lack of knowledge about the fate and persistence of transgenic pesticidal products in the soil. A model system of transgenic cotton, which produces Bacillus thuringiensis kurstakiδ‐endotoxin (Bt toxin), was used to address this issue. Methods were developed to quantify Btk toxin in soil and soil/plant litter by extraction of the Btk toxin with an aqueous buffer and quantification by ELISA. The highest recovery of Btk toxin from soil was obtained with a high salt, high pH buffer. In addition, for certain soil types, addition of a non‐ionic detergent, Tween‐20, was needed for optimal recovery. Recovery of Btk toxin from soil ranged from 60% for a low clay content, low organic matter soil to 27% for a high clay content, high organic matter soil. The limit of detection of this method is 0.5 ng of extractable toxin per g dry weight soil. The method was shown to be useful in tracking over time the persistence of both purified and transgenic Btk toxin in laboratory experiments.
Fate of transformed Trichoderma harzianum in the phylloplane of tomato plants
Authors: MIGHELI Q. , HERRERA‐ESTRELLA A. , AVATANEO M. , GULLINO M. L.
Published: April 2008

A propiconazole‐resistant Trichoderma harzianum strain with high phylloplane survival capability was transformed with the E. coli hygromycin B phosphotransferase gene (hph), coding for hygromycin B resistance. Four transformants were analysed for survival ability on the phylloplane of tomato plants grown under glasshouse conditions in comparison with their prototype and a yellow, hygromycin B‐sensitive mutant. Over 2 weeks, the four transformants showed higher survival rates in comparison with the wild‐type strain. The yellow mutant TF3/973 did not significantly differ in survival from the transformants. Both hygromycin B resistance and mitotic stability of transformants were evaluated during growth in vitro and after reisolation from tomato phylloplane. Hybridization patterns with the complete plasmid indicated that all four transformants were mitotically stable after several rounds of vegetative growth without selective pressure and during 2 weeks on tomato plants. None of the transformants had lost the ability to grow in the presence of both propiconazole and hygromycin B after growth under the same conditions. The results are discussed in relation to risk assessment of the release of transgenic fungi.
Mitochondrial control region and protein coding genes sequence variation among phenotypic forms of brown trout Salmo trutta from northern Italy
Authors: GIUFFRA E. , BERNATCHEZ L. , GUYOMARD R.
Published: April 2008

The Pô River basin of northern Italy is the home of distinctive and endemic morphological forms of brown trout Salmo trutta. We used PCR‐direct sequencing and RFLP techniques to study variation in the mitochondrial control region of 225 trout in order to assess genetic relatedness among 18 populations from that region. The distribution analysis of these genotypes among north Italian populations confirmed the phylogenetic differentiation of marbled trout Salmo trutta marmoratus populations and the postglacial origin of S. t. fario. Extensive genetic heterogeneity was observed among morphologically identical S. t. fario populations. Introgression with domestic strains of Atlantic basin origin was detected in all forms. In order to assess the phylogenetic congruence detected in coding and noncoding regions of the mitochondrial genome, we also analysed sequence variation in segments of the cytochrome b and ATPase subunit VI genes among representatives of all variants detected in the analysis of the control region. Variation in protein coding genes was only slightly less than that observed in the control region of the same individuals, both in terms of number of variants detected and of pairwise sequence divergence estimates among variants. Phylogenetic analysis based on protein coding genes sequences identified the same phylogenetic groupings defined by the control region analysis and also allowed a partial resolution of their phyletic relationships that was previously unresolved. However, coding and noncoding segments differed substantially in the transition‐transversion ratio (17:0 in coding segments vs. 17:6 in control region segments).
Molecular phylogeny of the actinorhizal Hamamelidae and relationships with host promiscuity towards Frankia
Authors: MAGGIA L. , BOUSQUET J.
Published: April 2008

Several of the most studied actinorhizal symbioses involve associations between host plants in the subclass Hamamelidae of the dicots and actinomycetes of the genus Frankia. These actinorhizal plants comprise eight genera distributed among three families of ‘higher’ Hamamelidae, the Betulaceae, Myricaceae, and Casuarinaceae. Contrasting promiscuity towards Frankia is encountered among the different actinorhizal members of these families, and a better assessment of the evolutionary history of these actinorhizal taxa could help to understand the observed contrasts and their implications for the ecology and evolution of the actinorhizal symbiosis. Complete DNA sequences of the chloroplast gene coding for the large subunit of ribulose 1,5‐bisphosphate carboxylase (rbcL) were obtained from taxa representative of these families and the Fagaceae. The phylogenetic relationships among and within these families were estimated using parsimony and distance‐matrix approaches. All families appeared monophyletic. The Myricaceae appeared to derive first before the Betulaceae and the Casuarinaceae. In the Casuarinaceae, the genus Gymnostoma derived before the genera Casuarina and Allocasuarina, which were found closely related. The analysis of character‐state changes in promiscuity along the consensus tree topology suggested a strong relationship between the evolutionary history of host plants and their promiscuity toward Frankia. Indeed, the actinorhizal taxa that diverged more recently in this group of plants were shown to be susceptible to a narrower spectrum of Frankia strains. The results also suggest that the ancestor of this group of plant was highly promiscuous, and that evolution has proceeded toward narrower promiscuity and greater specialization. These results imply that a tight relationship between the phytogenies of both symbiotic partners should not be expected, and that host promiscuity is likely to be a key determinant in the establishment of an effective symbiosis.
Paternity exclusion in a community of wild chimpanzees using hypervariable simple sequence repeats
Authors: MORIN P. A. , WALLIS J. , MOORE J. J. , WOODRUFF DAVID S.
Published: April 2008

We report the use of hypervariable simple sequence repeat (SSR) nuclear loci to study paternity in a community of wild chimpanzees (Pan troglodytes schweinfurthii) in Gombe National Park, Tanzania. All 43 living members of a habituated community were sampled and 35 were genotyped at 8 SSR loci using DNA amplified from hair. Paternity exclusions were performed for 25 chimpanzees including 10 for whom the mother was also genotyped. In each case 12–20 males were potential fathers based on their age and/ or direct observation of sexual behaviour. 179 tests involving potential father/offspring combinations were performed. In four cases the data permit the probable identification of the previously undetermined father; these are the first such determinations for free‐ranging chimpanzees, and the first based on non‐invasive sampling. In another four cases we were able to exclude all but two to five potential fathers, and in the remaining cases we were able to exclude all living males. For molecular ecologists SSR genotype databases offer important advantages over currently popular minisatellite DNA fingerprinting: they can be analysed unequivocally using traditional population genetics techniques and they can be expanded through time and space by other researchers.
Microsatellite analysis of genetic variation in black bear populations
Authors: PAETKAU D. , STROBECK C.
Published: April 2008

Measuring levels of genetic variation is an important aspect of conservation genetics The informativeness of such measurements is related to the variability of the genetic markers used; a particular concern in species, such as bears, which are characterized by low levels of genetic variation resulting from low population densities and small effective population sizes We describe the development of microsatellite analysis in bears and its use in assessing interpopulation differences in genetic variation in black bears from three Canadian National Parks These markers are highly variable and allowed identification of dramatic differences in both distribution and amount of variation between populations Low levels of variation were observed in a population from the Island of Newfoundland The significance of interpopulation differences in variability was tested using a likelihood ratio test of estimates of θ= 4Neu.
Molecular separation of genera in Cassiinae (Leguminosae), and analysis of variation in the nodulating species of Chamaecrista
Authors: WHITTY P. W. , POWELL W. , SPRENT J. I.
Published: April 2008

Randomly Amplified Polymorphic DNA (RAPD) methods have been adapted for use as a phenetic tool on the legume tribe Cassiinae. RAPD‐generated polymorphism within local populations was lower than between populations from different geographic regions, between species and genera. Examination of three Cassia species, 12 Chamaecrista species and 13 Senna species using eight primers showed the potential for separation of the nodulated/nitrogen fixing genus Chamaecrista from the previously congeneric groups Cassia and Semis. Similarly, RAPD analysis of two groups of nine Ch. rotundifolia and nine Ch. mimosoides samples using 11 primers has given separation according to both species and to geographical location. Analysis of a small sample of five Chamaecrista species from Brazil with eight primers gave separation consistent with known variations in nodule structure.
Charomid cloning vectors meet the pedipalpal chelae: single‐locus minisatellite DNA probes for paternity assignment in the harlequin beetle‐riding pseudoscorpion
Authors: ZEH D. W. , ZEH J. A. , MAY C. A.
Published: April 2008

We describe the first application of the charomid‐cloning method for developing single‐locus minisatellite DNA probes in a terrestrial arthropod. From a genomic library of the neotropical pseudoscorpion, Cordylochernes scorpioides, we have isolated two probes with heterozygosities exceeding 95%. These probes yielded single‐locus patterns after only low stringency washing and in the absence of genomic competitor DNA. Analysis of three pedigrees indicated germline stability and showed no evidence of linkage between the loci. Patterns of allelic transmission generally conformed closely to Mendelian expectations but large offspring numbers did enable detection of one example of significant bias in allele inheritance. Two test cases are presented to illustrate the clarity and power with which these probes can establish paternity: (i) a female mated to three unrelated males, and (ii) a female mated to two of her brothers. In both cases, a single probe could be used to assign the paternity of all offspring.
A phylogenetic analysis of micro‐organisms isolated from subsurface environments
Authors: Stim K. P.
Published: April 2008

Three methods were used to provide information on the identity and phylogenetic relatedness of 19 aerobic, chemoheterotrophic bacteria isolated from topsoil and deep subsurface sediments at a site in South Carolina. These methods were (i) analysis of selected physiological traits, (ii) restriction endonuclease analysis (REA) of genomic DNA, and (iii) analysis of 16S ribosomal RNA sequences. When the 16S rRNA sequences were compared with those for 12 standard strains, two topsoil isolates and six subsurface strains formed a tight group with the high‐G+C Gram‐positive bacteria and appeared to be most closely related to Arthrobacter globiformis— a coryneform‐actinomycete bacterium with unusually effective survival capabilities. The rest of the subsurface isolates were scattered among the standard strains from the Proteobacteria— including the pseudomonads and Agrobacterium tumefaciens— or the low‐G+C Gram‐positive bacteria.
Effect of cellular physiology on PCR amplification efficiency
Authors: Silva M. C. , Batt C. A.
Published: April 2008

Culture conditions, and other variables that modulate a cell's physiology, can bias a polymerase chain reaction (PCR) amplification against generating a representative population profile. Two Pseudomonas putida nahR alleles were constructed in pUC19 that differ solely in a 31‐bp internal segment whose sequence has been inverted. After PCR amplification, the products could be distinguished on the basis of a change in a unique restriction site. When an Escherichia coli strain carrying one nahR allele is submitted to different growth conditions, the consequences of such variations on the relative PCR amplification of whole cells can be ascertained through coamplification with a strain carrying the other allele and subsequent restriction analysis. Cells in stationary phase displayed improved amplifiability while cells grown at 42°C were equally amplifiable as compared to cells grown at 37°C. However, sublethal levels of tetracycline or growth in minimal medium made the PCR target in these cells relatively less amplifiable. When cells are completely lysed and the plasmid DNA is purified beforehand, the coamplification bias is eliminated. These results suggest that mixed populations containing cells in different physiological states may not be representatively amplified by PCR unless a DNA extraction step is included.
Molecular genetic analysis of the response of three soil microbial communities to the application of 2, 4‐D
Authors: Xia Xueqing , Bollinger John , Ogram Andrew
Published: April 2008

The responses of three different soil microbial communities to the experimental application of 2, 4‐dichlorophenoxyacetic acid (2, 4‐D) were evaluated with a variety of molecular genetic techniques. Two of the three soil communities had histories of prior direct exposure to 2, 4‐D, and one had no prior direct application of any herbicide. Dominant 2, 4‐D degrading strains isolated from these soils the previous year were screened for hybridization with three catabolic genes (tfdA, tfdAII, and tfdB) cloned from the well‐studied 2, 4‐D degradative plasmid, pJP4, revealing varying degrees of similarity with the three genes. Hybridization of total community DNA from the three soils with the tfd gene probes also indicated that pJP4‐like tfd genes were not harboured by a significant percentage of the community. Community level response was evaluated by the comparison of different treatments by Random Amplified Polymorphic DNA (RAPD) fingerprints and by community DNA cross‐hybridization. No differences between treatments within the same soil were detected in any of the RAPD fingerprints generated with 17 primers. Community DNA cross‐hybridization also indicated that the application of 2, 4‐D at the applied rates did not quantitatively affect the structure of the soil microbial communities present in the three soils during the time‐frame studied.
A new hypothesis for the origin of pentaploid Holcus from diploid Holcus lanatus L. and tetraploid Holcus mollis L. in France
Authors: Richard M. , Jubier M. F. , Bajon R. , Gouyon P. H. , Lejeune B.
Published: April 2008

The Holcus complex in France consists of two species, Holcus lanatus L. (Yorkshire fog; 2n= 2x= 14) and Holcus mollis L. (Creeping soft‐grass; 2n= 4x= 28) and an interspecific hybrid H.m.×H.l. (2n= 5x= 35), which is morphologically similar to Holcus mollis. A heterologous rDNA probe from wheat was used to detect the corresponding region in Holcus (s. l.) genomic DNA'fragments, for six to eight plants from 13 populations located south‐west of Paris. A restriction enzyme map of the ribosomal RNA genes (rDNA) in Holcus (s. l.) was also constructed. The length polymorphism detected in the IGS region was used as a DNA fingerprint for the identification of different cytotypes and species of the Holcus complex and for the typing and delimitation of individuals in populations. In the light of the results we reconsider the assumption that the pentaploid hybrid H.m.×H.l. is purely clonal. New hypotheses concerning the origin of the pentaploid hybrid and its reproduction are proposed, and the consequences for genetic diversity in natural populations discussed.
Clonal dispersal and spatial mixing in populations of the plant pathogenic fungus, Sclerotinia sclerotiorum
Authors: Kohli Y. , Brunner L. J. , YOELL H. , Milgroom M. G. , Anderson J. B. , Morrall R. A. A. , Kohn L. M.
Published: April 2008

Two thousand seven hundred and forty‐seven isolates of Sclerotinia sclerotiorum were sampled from four field populations of canola in western Canada. Each field was sampled in a grid of 128 50‐m 50‐m quadrats plus four intensive quadrats each sampled in a diagonal transect. Sampling was done at two phases of the disease cycle: (1) from ascospore inoculum on petals and (2) from disease lesions in stems. A total of 594 unique genotypes was identified by DNA fingerprinting. In each field, a small group of clones represented the majority of the sample, with a large group of clones or genotypes sampled once or twice. Clone frequencies were compared by χ2 tests. The difference in profiles of clone frequencies for the two fields sampled in 1991 was not significant, but in 1992 the difference in profiles was marginally significant, indicating some local population substructure. The difference in profiles of clone frequencies for petals and lesions was not significant in each of the two fields sampled in 1991. In each of the two fields sampled in 1992, however, the difference was highly significant, consistent either with selection for some clones or with waves of immigration during the disease cycle. Nine of the 30 most frequently sampled clones from this study were previously recovered in a macrogeographical sample from western Canada in 1990. For spatial analyses, randomization tests indicated no significant spatial aggregation of either clones on petals or clones from lesions. Also, isolates of a clone on petals were not closer to isolates of the same clone from lesions than could be predicted by chance. Both observations suggest spatial mixing of ascospore inoculum from resident or immigrant sources.
Mitochondrial DNA sequence variation and genetic stock structure of Atlantic cod (Gadus morhua) from bay and offshore locations on the Newfoundland continental shelf
Authors: Carr S. M. , Snellen A. J. , Howse K. A. , Wroblewski J. S.
Published: April 2008

Bay cod, Atlantic cod (Gadus morhua) that over‐winter in the deep‐water bays of northeastern Newfoundland, have historically been regarded as distinct in migration and spawning behaviour from offshore (Grand Bank) cod stocks. To investigate their genetic relationships, we determined the DNA sequence of a 307‐base‐pair portion of the mitochondrial cytochrome b gene for 236 adult cod taken from the waters off northeastern Newfoundland, including fish found over‐wintering and spawning in Trinity Bay. Although 17 genotypes were found, a single common genotype occurs at a frequency of greater than 80% in all samples, and no alternative genotype occurs at a frequency of greater than 3%. Genotype proportions did not differ significantly among samples. Measures of genetic subdivision among sampling locations are nil. Cod over‐wintering in Trinity Bay are not genetically distinct from offshore cod. In combination with tagging and physiological studies, these data suggest that there is sufficient movement of cod between bay and offshore locations to prevent the development or maintenance of independent inshore stocks. Adult cod that over‐winter in Trinity Bay appear to represent an assemblage of temporarily nonmigratory fish that have become physiologically acclimated to cold‐water inshore environments. The pattern of genetic variation in northern cod suggests a recent population structure characterized by extensive movement of contemporary individuals superimposed on an older structure characterized by a bottleneck in the population size of cod in the north‐western Atlantic.
Conservation genetics of the European brown bear ‐ a study using excremental PCR of nuclear and mitochondrial sequences
Authors: Kohn M. , Knauer F. , Stoffella A. , SCHRÖDER W. , PÄÄBO S.
Published: April 2008

In the Brenta area of northern Italy, a brown bear Ursus arctos population is rapidly going extinct. Restocking of the population is planned. In order to study the genetics of this highly vulnerable population with a minimum of stress to the animals we have developed a PCR‐based method that allows the study of mitochondrial and nuclear gene sequences from droppings collected in the field. This method is generally applicable to animals in the wild. Using excremental as well as hair samples, we show that the Brenta population is monomorphic for one mitochondrial lineage and that female as well as male bears exist in the area. In addition, 70 samples from other parts of Europe were studied. As others have previously reported, the mitochondrial gene pool of European bears is divided into two major clades, one with a western and the other with an eastern distribution. Whereas populations generally belong to either one or the other mitochondrial clade, the Romanian population contains both clades. The bears in the Brenta belong to the western clade. The implications for the management of brown bears in the Brenta and elsewhere in Europe are discussed.
Analysis of genetic structure in soil populations of Rhizobium leguminosarum recovered from the USA and the UK
Authors: STRAIN S. R. , WHITTAM T. S. , BOTTOMLEY P. J.
Published: April 2008

Although many studies have shown that animal‐associated bacterial species exhibit linkage disequilibrium at chromosomal loci, recent studies indicate that both animal‐associated and soil‐borne bacterial species can display a nonclonal genetic structure in which alleles at chromosomal loci are in linkage equilibrium. To examine the situation in soil‐borne species further, we compared genetic structure in two soil populations of Rhizobium leguminosarum bv. trifolii and two populations of R. leguminosarum bv. viciae from two sites in Oregon, with genetic structure in R. leguminosarum bv. viciae populations recovered from peas grown at a site in Washington, USA, and at a site in Norfolk, UK. A total of 234 chromosomal types (ET) were identified among 682 strains analysed for allelic variation at 13 enzyme‐encoding chromosomal loci by multilocus enzyme electrophoresis (MLEE). Chi‐square tests for heterogeneity of allele frequencies showed that the populations were not genetically uniform. A comparison of the genetic diversity within combined and individual populations confirmed that the Washington population was the primary cause of genetic differentiation between the populations. Each individual population exhibited linkage disequilibrium, with the magnitude of the disequilibrium being greatest in the Washington population and least in the UK population of R. leguminosarum bv. viciae. Linkage disequilibrium in the UK population was created between two clusters of 9 and 23 ETs, which, individually, were in linkage equilibrium. Strong linkage disequilibrium between the two major clusters of 8 and 12 ETs in the Washington population was caused by the low genetic diversity of the ETs within each cluster relative to the inter‐cluster genetic distance. Because neither the magnitude of genetic diversity nor of linkage disequilibrium increased as hierarchical combinations of the six local populations were analysed, we conclude that the populations have not been isolated from each other for sufficient time, nor have they been exposed to enough selective pressure to develop unique multilocus genetic structure.
An unusual mitochondrial DNA polymorphism in the Chorthippus biguttulus species group (Orthoptera: Acrididae)
Authors: MASON D. J. , BUTLIN R. K. , GACESA P.
Published: April 2008

Mitochondrial DNA restriction fragment patterns have been used to investigate the Chorthippus biguttulus group: a group of morphologically similar grasshopper species with very distinct male calling songs. Ten haplotypes were identified in 21 populations, representing six species. Haplotype diversity was as great within species, and in some case within populations, as it was among species and no clear geographical patterns of haplotype distribution were revealed. In the light of grasshopper range changes since the last glaciation it is unlikely that these data can be explained entirely by the retention of ancestral polymorphism. This suggests that hybridization has occurred during periods of grasshopper range expansion since the last glacial maximum, allowing mtDNA transfer between populations that have previously diverged in allopatry.
Evolutionary implications of allozyme and RAPD variation in diploid populations of dioecious buffalograssBuchloë dactyloides
Authors: PEAKALL R. , SMOUSE P. E. , HUFF D. R.
Published: April 2008

Buffalograss,Buchloë dactyloides, is widely distributed throughout the Great Plains of North America, where it is an important species for rangeland forage and soil conservation. The species consists of two widespread polyploid races, with narrowly endemic diploid populations known from two regions: central Mexico and Gulf Coast Texas. We describe and compare the patterns of allozyme and RAPD variation in the two diploid races, using a set of 48 individuals from Texas and Mexico (four population samples of 12 individuals each). Twelve of 22 allozyme loci were polymorphic, exhibiting 35 alleles, while seven 10‐mer RAPD primers revealed 98 polymorphic bands. Strong regional differences were detected in the extent of allozyme polymorphism: Mexican populations exhibited more internal gene diversity (He= 0.20, 0.19) than did the Texan populations (He= 0.08, 0.06), although the number of RAPD bands in Texas (n= 62) was only marginally smaller than in Mexico (n= 68).F‐statistics for the allozyme data, averaged over loci, revealed strong regional differentiation (meanFRT=+ 0.30), as well as some differentiation among populations within regions (meanFPR=+ 0.09). In order to describe and compare the partitioning of genetic variation for multiple allozyme and RAPD loci, we performed an Analysis of Molecular Variance (AMOVA). AMOVA for both allozyme and RAPD data revealed similar qualitative patterns: large regional differences and smaller (but significant) population differences within regions. RAPDs revealed greater variation among regions (58.4% of total variance) than allozymes (45.2%), but less variation among individuals within populations (31.9% for RAPDs vs. 45.2% for allozymes); the proportion of genetic variance among populations within regions was similar (9.7% for RAPDs vs. 9.6% for allozymes). Despite this large‐scale concordance of allozyme and RAPD variation patterns, multiple correlation Mantel techniques revealed that the correlations were low on an individual by individual basis. Our findings of strong regional differences among the diploid races will facilitate further study of polyploid evolution in buffalograss.
Evolution of the Cyprinella lutrensis species group. III. Geographic variation in the mitochondrial DNA of Cyprinella lutrensis— the influence of Pleistocene glaciation on population dispersal and divergence
Authors: RICHARDSON L. R. , GOLD J. R.
Published: April 2008

We employed restriction site variation in mitochondrial (mt)DNA to determine if significant phylogeographic structure occurs in the North American cyprinid fish Cyprinella lutrensis. Digestion patterns from 16 restriction endonucleases identified fifty mtDNA haplotypes among 127 individuals of Cyprinella lutrensis assayed from localities in the Gulf Coastal Plain, the Great Plains, and the Central Lowlands. Nucleotide sequence divergence among haplotypes was highly variable (mean ± SE: 2.87%± 0.08; range: 0.14–9.24%). Maximum‐parsimony analysis and the neighbour joining method of tree construction revealed three major groupings (clades) of haplotypes that differed in geographic distribution. Divergence estimates between the basal clade, comprised of haplotypes primarily from the Brazos River in east Texas, and the remaining two clades, place C. lutrensis in the western Gulf Coastal Plain prior to Pleistocene glaciation. Nucleotide sequence divergence between the second clade, comprised of haplotypes from the Trinity and Calcasieu rivers in east Texas and southwestern Louisiana, respectively, and the third clade (comprised primarily of haplotypes from localities north of Texas and affected directly by Pleistocene glaciation), suggest that C. lutrensis colonized gladated regions to the north during the mid‐ to late Pleistocene. This hypothesis is supported by levels of intrapopulational nucleotide diversity in geographic localities outside of Texas and by geological evidence. Despite marked geographic variation in morphometries, meristics, and nuptial coloration, mtDNA variation in glaciated regions was not geographically structured, and subspecies of C. lutrensis were not identifiable by phylogenetic analysis of mtDNA.