bluuebunny/crossref_metadata_2025 huggingface dataset:
DNA from bird‐dispersed seed and wind‐disseminated pollen provides insights into postglacial colonization and population genetic structure of whitebark pine (Pinus albicaulis)
Authors: Richardson B. A. , Brunsfeld S. J. , Klopfenstein N. B.
Published: March 2003

Uniparentally inherited mitochondrial (mt)DNA and chloroplast (cp)DNA microsatellites (cpSSRs) were used to examine population genetic structure and biogeographic patterns of bird‐dispersed seed and wind‐disseminated pollen of whitebark pine (Pinus albicaulis Engelm.). Sampling was conducted from 41 populations throughout the range of the species. Analyses provide evidence for an ancestral haplotype and two derived mtDNA haplotypes with distinct regional distributions. An abrupt contact zone between mtDNA haplotypes in the Cascade Range suggests postglacial biogeographic movements. Among three cpSSR loci, 42 haplotypes were detected within 28 cpSSR sample populations that were aggregated into six regions. Analysis of molecular variance (amova) was used to determine the hierarchical genetic structure of cpSSRs. amova and population pairwise comparisons (FST) of cpSSR, and geographical distribution of mtDNA haplotypes provide insights into historical changes in biogeography. The genetic data suggest that whitebark pine has been intimately tied to climatic change and associated glaciation, which has led to range movements facilitated by seed dispersal by Clark’s nutcracker (Nucifraga columbiana Wilson). The two hypotheses proposed to explain the genetic structure are: (i) a northward expansion into Canada and the northern Cascades in the early Holocene; and (ii) historical gene flow between Idaho and the Oregon Cascades when more continuous habitat existed in Central Oregon during the late Pleistocene. Genetic structure and insights gained from historical seed movements provide a basis on which to develop recovery plans for a species that is at risk from multiple threats.
Genetic variation across the historical range of the wild turkey (Meleagris gallopavo)
Authors: Mock K. E. , Theimer T. C. , Rhodes O. E. , Greenberg D. L. , Keim P.
Published: March 2003

Genetic differences within and among naturally occurring populations of wild turkeys (Meleagris gallopavo) were characterized across five subspecies’ historical ranges using amplified fragment length polymorphism (AFLP) analysis, microsatellite loci and mitochondrial control region sequencing. Current subspecific designations based on morphological traits were generally supported by these analyses, with the exception of the eastern (M. g. silvestris) and Florida (M. g. osceola) subspecies, which consistently formed a single unit. The Gould’s subspecies was both the most genetically divergent and the least genetically diverse of the subspecies. These genetic patterns were consistent with current and historical patterns of habitat continuity. Merriam’s populations showed a positive association between genetic and geographical distance, Rio Grande populations showed a weaker association and the eastern populations showed none, suggesting differing demographic forces at work in these subspecies. We recommend managing turkeys to maintain subspecies integrity, while recognizing the importance of maintaining regional population structure that may reflect important adaptive variation.
Molecular identification of prey in predator diets
Authors: Symondson W. O. C.
Published: March 2003

In many situations prey choice by predators in the field cannot be established or quantified using direct observation. The remains of some prey may be visually identified in the guts and faeces of predators but not all predators ingest such hard remains and even those that do consume them may also ingest soft‐bodies prey that leave no recognizable remnants. The result is, at best, a biased picture of prey choice. A range of molecular techniques and applications are reviewed that allow prey remains to be identified, often to the species and even stage level. These techniques, all of which are still in use, include enzyme electrophoresis, a range of immunological approaches using polyclonal and monoclonal antibodies to detect protein epitopes, and recently developed polymerase chain reaction (PCR)‐based methods for detecting prey DNA. Analyses may be postmortem, on invertebrate and vertebrate predators collected from the field, or noninvasive assays of the remains in regurgitated bird pellets or vertebrate faeces. It was concluded that although monoclonal antibodies are currently the most effective method in use today, PCR‐based techniques have proved to be highly effective and versatile in recent laboratory trials and are likely to rapidly displace all other approaches.
Reconciling genetic expectations from host specificity with historical population dynamics in an avian brood parasite, Horsfield’s Bronze‐CuckooChalcites basalisof Australia
Authors: Joseph L. , Wilke T. , Alpers D.
Published: March 2003

Mitochondrial DNA (mtDNA) is being used increasingly to explore the evolution of host specificity in avian brood parasites. A stable coevolutionary equilibrium between multiple phylogenetically unrelated hosts and a brood parasitic species predicts that mtDNA diversity in the parasite should be relatively deep and phylogenetically structured. Also, the different intraspecific clades resulting from parasitism to multiple sympatric hosts should themselves occur sympatrically. However, mtDNA diversity in brood parasites is as susceptible to effects of historical population dynamics as in any species. We demonstrate the relevance of these dynamics to the use of mtDNA in understanding coevolution between an Australian brood‐parasite, Horsfield’s Bronze‐CuckooChalcites basalisand its hosts,Malurusfairy‐wrens andAcanthizathornbills. Previous ecological and behavioural analyses argue thatMalurus‐ andAcanthiza‐specific host races exist inC. basalis. Yet mtDNA diversity inC. basalisis low and phylogenetically unstructured (mean sequence divergence 0.15 ± 0.07%, range 0.00%–0.31%) and tests of mtDNA neutrality and range expansion vs. population stability (Tajima’s D, Fu & Li’s F* and D*, Fu’sFS, mismatch analyses) all indicate thatC. basalishas expanded its range very recently, probably within the last few tens of thousands of years following climatic amelioration after a peak of aridity in the late Pleistocene. The low mtDNA diversity and its lack of phylogenetic structure inC. basalisdeny the existence of evolutionarily long‐term stable host races inC. basalisbut not the possibility of recently evolved ones. They highlight the need for renewed behavioural and ecological study of the relationship betweenC. basalisand its hosts. Our findings illustrate the need to understand the evolutionary context in which a brood parasite and its hosts have evolved if mtDNA data are to be used in testing hypotheses concerning the origin and maintenance of host specificity. They also add to the growing body of work illustrating the use of mismatch analyses and Fu’sFSin detecting range expansions.
The Mediterranean fruit fly in California: evidence for multiple introductions and persistent populations based on microsatellite and mitochondrial DNA variability
Authors: Meixner M. D. , McPheron B. A. , Silva J. G. , Gasparich G. E. , Sheppard W. S.
Published: March 2003

Microsatellite and mitochondrial DNA (mtDNA) variability data were used to study outbreaks of Mediterranean fruit fly in California in the years 1992–94 and 1997–99. A total of 359 flies caught in monitoring traps during these years were examined at three polymorphic mtDNA restriction sites and two microsatellite loci. Composite genotypes obtained through analysis of these markers indicate at least five independent introductions of medflies into California between 1992 and 1998. Whereas the majority of specimens displayed a single mtDNA haplotype (AAA), variation of microsatellite alleles among these flies suggests at least one additional introduction in 1993 into southern California. Flies displaying the AAB haplotype sampled in 1992 both in northern and southern California shared microsatellite alleles absent in AAA flies although lacking others commonly found in AAA specimens, thus supporting the hypothesis of an independent introduction of these flies from a different source. In contrast to earlier infestations, a few specimens caught in southern California in 1993 and again in 1998 showed both mtDNA and microsatellite patterns consistent with a Hawaiian origin. Single flies collected in Santa Clara County in 1997 and in El Monte, Los Angeles County & in 1999 most likely represent a sixth and seventh distinct introduction, respectively.
Population genetic structure of two rare tree species (Colubrina oppositifoliaandAlphitonia ponderosa, Rhamnaceae) from Hawaiian dry and mesic forests using random amplified polymorphic DNA markers
Authors: Kwon J. A. , Morden C. W.
Published: March 2003

Hawaiian dry and mesic forests contain an increasingly rare assemblage of species due to habitat destruction, invasive alien weeds and exotic pests. Two rare Rhamnaceae species in these ecosystems,Colubrina oppositifoliaandAlphitonia ponderosa, were examined using random amplified polymorphic DNA (RAPD) markers to determine the genetic structure of the populations and the amount of variation relative to other native Hawaiian species. Relative variation is lower than with other Hawaiian species, although this is probably not a consequence of genetic bottleneck. Larger populations of both species contain the highest levels of genetic diversity and smaller populations generally the least as determined by number of polymorphic loci, estimated heterozygosity, and Shannon’s index of genetic diversity. Populations on separate islands were readily discernible for both species as were two populations ofC. oppositifoliaon Hawai’i island (North and South Kona populations). Substructure among Kaua’i subpopulations ofA. ponderosathat were ecologically separated was also evident. Although population diversity is thought to have remained at predisturbance levels, population size continues to decline as recruitment is either absent or does not keep pace with senescence of mature plants. Recovery efforts must focus on control of alien species if these and other endemic dry and mesic forest species are to persist.
Genetic relatedness and female spatial organization in a solitary carnivore, the raccoon, Procyon lotor
Authors: Ratnayeke Shyamala , Tuskan Gerald A. , Pelton Michael R.
Published: March 2003

Among mammals, some of the most common types of cohesive social groupings originate from natal philopatry through the extended mother family. This retention of females within social groups (i.e. the nonrandom dispersion of female relatives in space) should affect population genetic structure. We examined the relationship between genetic relatedness and female spatial organization in a wild population of the North‐American raccoon, Procyon lotor, a solitary carnivore in east Tennessee. Multilocus genetic band‐sharing data and 3½ years of radiotelemetry observations were used to study the spatial and genetic relationships among 38 adult females. DNA amplification employing primers of arbitrary sequence (random amplified polymorphic DNA; RAPD) indicated that female philopatry in raccoons led to a greater likelihood of neighbours being more related than expected by chance. Genetic distance based on RAPD band frequency was positively correlated with spatial distance among females (P = 0.0001) and genetic similarity was positively correlated with the extent of home‐range overlap (P = 0.0028). Philopatry seemed biased towards females; average female–female similarities were greater than average male–male similarities (P = 0.0001), or average male–female similarities (P = 0.0001). High home‐range overlap among some females with low or moderate levels of band sharing indicated that maternal inheritance of space was not a prerequisite for establishing or sharing home ranges. Female philopatry was the most probable explanation for the nonrandom spatial and genetic association of raccoons in east Tennessee.
Reproductive mode and genetic variation suggest a North American origin of European Letharia vulpina
Authors: Högberg Nils , Kroken Scott , Thor Göran , Taylor John W.
Published: March 2003

Our data on the intercontinental population biology of Letharia vulpina show an unexpected shift from a recombining North American population with unique haplotypes to genetically depauperate Swedish and Italian populations, each with many representatives of a single repeated haplotype. Analysis of eight loci in 47 individuals supported recombination in North American populations and showed almost no variation among European populations. We infer that a genetic bottleneck caused by limited long‐distance dispersal accounts for the lack of genetic variation found in marginal populations. This lack of variation in the European populations makes it impossible to use population genetic means to distinguish clonal reproduction from self‐fertilization or even outcrossing, but phenotype indicates that reproduction in the marginal populations is by clonal spread, via soredia and isidioid soredia.
Sampling within the genome for measuring within‐population diversity: trade‐offs between markers
Authors: Mariette S. , Le Corre V. , Austerlitz F. , Kremer A.
Published: March 2003

Experimental results of diversity estimates in a set of populations often exhibit contradictory patterns when different marker systems are used. Using simulations we identified potential causes for these discrepancies. These investigations aimed also to detect whether different sampling strategies of markers within the genome resulted in different estimates of the diversity at the whole genome level. The simulations consisted in generating a set of populations undergoing various evolutionary scenarios which differed by population size, migration rate and heterogeneity of gene flow. Population diversity was then computed for the whole genome and for subsets of loci corresponding to different marker techniques. Rank correlation between the two measures of diversity were investigated under different scenarios. We showed that the heterogeneity of genetic diversity either between loci (genomic heterogeneity, GH) or among populations (population heterogeneity, PH) varied greatly according to the evolutionary scenario considered. Furthermore, GH and PH were major determinants of the level of rank correlation between estimates of genetic diversities obtained using different kinds of markers. We found a strong positive relationship between the level of the correlation and PH, whatever the marker system. It was also shown that, when GH values were constantly low during generations, a reduced number of microsatellites was enough to predict the diversity of the whole genome, whereas when GH increased, more loci were needed to predict the diversity and amplified fragment length polymorphism markers would be more recommended in this case. Finally the results are discussed to recommend strategies for gene diversity surveys.
Evolutionary and statistical properties of three genetic distances
Authors: Kalinowski Steven T.
Published: March 2003

Many genetic distances have been developed to summarize allele frequency differences between populations. I review the evolutionary and statistical properties of three popular genetic distances:DS,DA, and θ, using computer simulation of two simple evolutionary histories: an isolation model of population divergence and an equilibrium migration model. The effect of effective population size, mutation rate, and mutation mechanism upon the parametric value between pairs of populations in these models explored, and the unique properties of each distance are described. The effect of these evolutionary parameters on study design is also investigated and similar results are found for each genetic distance in each model of evolution: large sample sizes are warranted when populations are relatively genetically similar; and loci with more alleles produce better estimates of genetic distance.
Speciation in fig pollinators and parasites
Authors: Weiblen George D. , Bush Guy L.
Published: March 2003

Here we draw on phylogenies of figs and fig wasps to suggest how modes of speciation may be affected by interspecific interactions. Mutualists appear to have cospeciated with their hosts to a greater extent than parasites, which showed evidence of host shifting. However, we also repeatedly encountered a pattern not explained by either cospeciation or host switching. Sister species of fig parasites often attack the same host in sympatry, and differences in ovipositor length suggest that parasite speciation could result from divergence in the timing of oviposition with respect to fig development. These observations on fig parasites are consistent with a neglected model of sympatric speciation.
Phylogeographical approaches to assessing demographic connectivity between breeding and overwintering regions in a Nearctic−Neotropical warbler (Wilsonia pusilla)
Authors: Kimura M. , Clegg S. M. , Lovette I. J. , Holder K. R. , Girman D. J. , Milá B. , Wade P. , Smith T. B.
Published: March 2003

We characterized the pattern and magnitude of phylogeographical variation among breeding populations of a long‐distance migratory bird, the Wilson's warbler (Wilsonia pusilla), and used this information to assess the utility of mtDNA markers for assaying demographic connectivity between breeding and overwintering regions. We found a complex pattern of population differentiation in mitochondrial DNA (mtDNA) variation among populations across the breeding range. Individuals from eastern North America were differentiated from western individuals and the eastern haplotypes formed a distinct, well‐supported cluster. The more diverse western group contained haplotype clusters with significant geographical structuring, but there was also broad mixing of haplotype groups such that no haplotype groups were population specific and the predominance of rare haplotypes limited the utility of frequency‐based assignment techniques. Nonetheless, the existence of geographically diagnosable eastern vs. western haplotypes enabled us to characterize the distribution of these two groups across 14 overwintering locations. Western haplotypes were present at much higher frequencies than eastern haplotypes at most overwintering sites. Application of this mtDNA‐based method of linking breeding and overwintering populations on a finer geographical scale was precluded by the absence of population‐specific markers and by insufficient haplotype sorting among western breeding populations. Our results suggest that because migratory species such as the Wilson's warbler likely experienced extensive gene flow among regional breeding populations, molecular markers will have the greatest utility for characterizing breeding−overwintering connectivity at a broad geographical scale.
Microsatellite DNA evidence for genetic drift and philopatry in Svalbard reindeer
Authors: Côté S. D. , Dallas J. F. , Marshall F. , Irvine R. J. , Langvatn R. , Albon S. D.
Published: March 2003

Mainland populations of Arctic reindeer and caribou Rangifer tarandus often undergo extensive movements, whereas populations on islands tend to be isolated and sedentary. To characterize the genetic consequences of this difference, levels of genetic diversity and subdivision of Svalbard reindeer (R. t. platyrhynchus) from two adjacent areas on Nordenskjiöldland, Spitsbergen were estimated using data from up to 14 microsatellites. The mean number of alleles per locus in Svalbard reindeer was 2.4 and mean expected heterozygosity per locus was 0.36. The latter value was significantly lower than in Canadian caribou and Norwegian reindeer but higher than in some other cervid species. Large samples of females (n = 743) and small samples of males (n = 38) from two sites ≈ 45 km apart showed genetic subdivision, which could be due to local population fluctuations or limited gene flow. To our knowledge, this is the first study to report significant differentiation at microsatellite loci in Rangifer at such short geographical distances. Neither population showed genetic evidence for recent population bottlenecks when loci unbiased with respect to heterozygosity were analysed. In contrast, false signals of a recent bottleneck were detected when loci upwardly biased with respect to heterozygosity were analysed. Thus, Svalbard reindeer appeared to conform to the paradigm of island populations made genetically depauperate by genetic drift.
Paternity frequency and maternity of males in some stingless bee species
Authors: Palmer Kellie A. , Oldroyd Benjamin P. , Quezada‐Euán José Javier G. , Paxton Robert J. , May‐Itza William De J.
Published: March 2003

In monogynous hymenopteran societies, the number of mates of a queen strongly influences the potential for conflict between workers and queens over the maternity of males. Queens always ‘prefer’ their own sons to sons of workers, regardless of queen mating frequency. When a queen mates once, workers are more closely related to, and therefore are expected to prefer, their own sons and then sons of sisters to sons of the queen. However, if effective paternity frequency exceeds 2, workers on average should prefer queen‐produced males to males produced by their sisters. We studied the queen mating frequency of seven stingless bee species: the Mexican speciesScaptotrigona mexicana,S. pectoralisand the Australian speciesAustroplebeia symei,Trigona clypearis,T. hockingsi,T. mellipesandT. sapiens. We then determined whether males arise from eggs laid by workers or queens inA. symei,T. clypearis,T. hockingsiandT. mellipes. We show that all seven species investigated are most likely singly mated and that the queen dominates reproduction. This indicates that the queen's mating frequency alone does not determine whether workers or the queen produces the males.
Mitochondrial DNA variation in Pleistocene and modern Atlantic salmon from the Iberian glacial refugium
Authors: Consuegra S. , García De Leániz C. , Serdio A. , González Morales M. , Straus L. G. , Knox D. , Verspoor E.
Published: March 2003

Current understanding of the postglacial colonization of Nearctic and Palearctic species relies heavily on inferences drawn from the phylogeographic analysis of contemporary generic variants. Modern postglacial populations are supposed to be representative of their Pleistocene ancestors, and their current distribution is assumed to reflect the different colonization success and dispersal patterns of refugial lineages. Yet, testing of phylogeographic models against ancestral genomes from glacial refugia has rarely been possible. Here we compare ND1 mitochondrial DNA variation in late Pleistocene (16 000–40 000 years before present), historical and contemporary Atlantic salmon (Salmo salar) populations from northern Spain and other regions of western Europe. Our study demonstrates the presence of Atlantic salmon in the Iberian glacial refugium during the last 40 000 years and points to the Iberian Peninsula as the likely source of the most common haplotype within the Atlantic lineage in Europe. However, our findings also suggest that there may have been significant changes in the genetic structure of the Iberian refugial stock since the last ice age, and question whether modern populations in refugial areas are representative of ice age populations. A common haplotype that persisted in the Iberian Peninsula during the Pleistocene last glacial maximum is now extremely rare or absent from European rivers, highlighting the need for caution when making phylogeographic inferences about the origin and distribution of modern genetic types.
Extra pair paternity in birds: a review of interspecific variation and adaptive function
Authors: Griffith Simon C. , Owens Ian P. F. , Thuman Katherine A.
Published: March 2003

The application of molecular genetic techniques has revolutionized our view of avian mating systems. Contrary to prior expectations, birds are only very rarely sexually monogamous, with ‘extra‐pair offspring’ found in approximately 90% of species. Even among socially monogamous species, over 11% of offspring are, on average, the result of extra‐pair paternity (EPP). Based on over 150 molecular genetic studies of EPP in birds, we review two topical areas: (i) ecological explanations for interspecific variation in the rate of EPP; and (ii) evidence bearing on the adaptive function of EPP. We highlight the remaining challenges of understanding the relative roles of genes and ecology in determining variation between taxa in the rate of extra paternity, and testing for differences between extra‐pair offspring and those sired within‐pair.
Geographic pattern of genetic variation in the European globeflowerTrollius europaeusL. (Ranunculaceae) inferred from amplified fragment length polymorphism markers
Authors: Despres Laurence , Loriot Sandrine , Gaudeul Myriam
Published: March 2003

The distribution of genetic variation and the phylogenetic relationships between 18 populations of the arctic‐alpine plantTrollius europaeuswere analysed in three main regions (Alps, Pyrenees and Fennoscandia) by using dominant AFLP markers. Analysis of molecular variance revealed that most of the genetic variability was found within populations (64%), although variation among regions (17%) and among populations within regions (19%) was highly significant (P < 0.001). Accordingly, the global fixation indexFSTaveraged over loci was high (0.39). The among‐population differentiation indicates restricted gene flow, congruent with limited dispersal of specific globeflower's pollinating flies (Chiastochetaspp.). Within‐population diversity levels were significantly higher in the Alps (mean Nei's expected heterozygosityHE = 0.229) than in the Pyrenees (HE= 0.197) or in Fennoscandia (HE = 0.158). This finding is congruent with the species‐richness of the associated flies, which is maximum in the Alps. We discuss the processes involved in shaping observed patterns of genetic diversity within and amongT. europaeuspopulations. Genetic drift is the major factor acting on the small Pyrenean populations at the southern edge ofT. europaeusdistribution, while large Fennoscandian populations result probably from a founder effect followed by demographic expansion. The Alpine populations represent moderately fragmented relics of large southern ancestral populations. The patterns of genetic variability observed in the host plant support the hypothesis of sympatric speciation in associated flies, rather than recurrent allopatric speciations.
Metapopulation structure of the specialized herbivoreMacrosiphoniella tanacetaria(Homoptera, Aphididae)
Authors: Massonnet Blandine , Simon Jean‐Christophe , Weisser Wolfgang W.
Published: March 2003

We investigated population dynamics, genetic diversity and spatial structure in the aphid speciesMacrosiphoniella tanacetaria, a specialist herbivore feeding on tansy,Tanacetum vulgare. Tansy plants (genets) consist of many shoots (ramets), and genets are grouped in sites. Thus, aphids feeding on tansy can cluster at the level of ramets, genets and sites. We studied aphid population dynamics in 1997 and 2001 and found that within sites: (i) at any time, aphids used only a fraction of the available ramets and genets; (ii) at the level of ramets, most aphid colonies survived only one week; (iii) at the level of genets, mean survival time was less than 4 weeks; and (iv) colonization and extinction events occurred throughout the season. We sampled aphids in seven sites in the Alsace region, France (4–45 km apart) and two sites in Germany in 1999 to study genetic structure within and between populations. Genetic analyses using nine microsatellite loci showed that: (i) genotypic variability was high, (ii) none of the populations was in Hardy–Weinberg equilibrium, (iii) heterozygote deficits and linkage disequilibria were frequent, and (iv) all populations were genetically differentiated, even at a small geographical scale. Renewed sampling of the Alsace sites in 2001 showed that three populations had become extinct and significant genetic changes had occurred in the remaining four populations. The frequencies of extinction and colonization events at several spatial scales suggest a hierarchical metapopulation structure forM. tanacetaria.Frequent population turnover and drift are likely causes for the genetic differentiation ofM. tanacetariapopulations.
Phylogeography of the invasive cyanobacterium Cylindrospermopsis raciborskii
Authors: Neilan B. A. , Saker M. L. , Fastner J. , Törökné A. , Burns B. P.
Published: March 2003

Cylindrospermopsis raciborskii is a planktonic freshwater cyanobacterium that has become increasingly prevalent in tropical and temperate water bodies world‐wide. This species is of concern from a water‐quality perspective because of its known ability to produce toxins that can affect the health of humans and other animals. This study investigates genetic vari‐ation between strains of C. raciborskii isolated from freshwater rivers and reservoirs in Australia, Brazil, Germany, Hungary, Portugal and the USA. Strains were first characterized by analysis of their 16S rRNA gene nucleotide sequences and were found to have a sequence divergence of 99.1%. A phylogenetic tree, constructed using the 16S rRNA gene sequences showed that strains grouped into Australian, European and North/South American phylotypes. To investigate further the observed separation of strains into geographically distinct groups, we applied a cyanobacterium‐specific short tandem repeat sequence technique, HIP1. An electrophoretic comparison of the HIP1 polymerase chain reaction products showed clear distinctions between the C. raciborskii strains. A phylogenetic tree, based on the repeat element banding patterns, also revealed three distinct groups of C. raciborskii strains. The first group consisted of strains from the USA and Brazil; the second comprised European strains from Germany, Hungary and Portugal; and the third were strains from Australia. In general, between‐country variation was greater than within‐country variation, indicating that this fingerprinting technique can successfully distinguish C. raciborskii strains taken from different global locations. The relationship between toxicity and the observed HIP1 polymerase chain reaction fingerprint profiles was less clear, although it is interesting to note that of the strains analysed in this study, only Australian strains are known to produce cylindrospermopsin and only Brazilian strains have been reported to produce paralytic shellfish poisoning toxins.
Possible causes of morphological variation in an endemic Moroccan groundsel (Senecio leucanthemifoliusvar.casablancae): evidence from chloroplast DNA and random amplified polymorphic DNA markers
Authors: Coleman Max , Abbott Richard J.
Published: March 2003

Genetic variation was assessed inSenecio leucanthemifoliusvar.casablancae(Compositae), a Moroccan Atlantic coast endemic, in order to examine possible causes of atypical leaf morphology in three populations south of the known range. Evidence for introgression fromS. glaucusssp.coronopifoliusand/or divergence was investigated with molecular markers. Both random amplified polymorphic DNA (RAPD) and chloroplast (cp) DNA restriction fragment length polymorphism (RFLP) differentiated the species well. Some evidence that hybridization may have occurred between the two species was provided by cpDNA markers. However, biparentally inherited RAPD markers failed to provide any support for the hypothesis that intermediate leaf morphologies in atypical populations arose through hybridization. Consequently, they are most likely to have arisen via divergence caused by drift and/or selection. Genetic distances among populations ofS. leucanthemifoliuswere significant in all but one case. Isolation by distance was indicated by a significant positive correlation between genetic and geographical distances (r = 0.68,P = 0.01, Mantel test). These results suggest that long‐distance achene dispersal is rare, despite the presence of a well‐developed pappus. The observed loss of pappus at achene maturity may explain this unexpected result. Due to the morphological distinction of var.casablancaefrom other varieties ofS. leucanthemifolius, we suggest elevation to species rank and treatment of the atypical material at infraspecific rank.
Population genetic consequences of extreme variation in sexual and clonal reproduction in an aquatic plant
Authors: Eckert Christopher G. , Lui Keiko , Bronson Kelly , Corradini Pierre , Bruneau Anne.
Published: March 2003

Most plants combine sexual reproduction with asexual clonal reproduction in varying degrees, yet the genetic consequences of reproductive variation remain poorly understood. The aquatic plant Butomus umbellatus exhibits striking reproductive variation related to ploidy. Diploids produce abundant viable seed whereas triploids are sexually sterile. Diploids also produce hundreds of tiny clonal bulbils, whereas triploids exhibit only limited clonal multiplication through rhizome fragmentation. We investigated whether this marked difference in reproductive strategy influences the diversity of genotypes within populations and their movement between populations by performing two large‐scale population surveys (n = 58 populations) and assaying genotypic variation using random amplified polymorphic DNA (RAPDs). Contrary to expectations, sexually fertile populations did not exhibit higher genotypic diversity than sterile populations. For each cytotype, we detected one very common and widespread genotype. This would only occur with a very low probability (< 10−7) under regular sexual recombination. Compatibility analysis also indicated that the pattern of genotypic variation largely conformed to that expected with predominant clonal reproduction. The potential for recombination in diploids is not realized, possibly because seeds are outcompeted by bulbils for safe sites during establishment. We also failed to find evidence for more extensive movement of fertile than sterile genotypes. Aside from the few widespread genotypes, most were restricted to single populations. Genotypes in fertile populations were very strongly differentiated from those in sterile populations, suggesting that new triploids have not arisen during the colonization of North America. The colonization of North America involves two distinct forms of B. umbellatus that, despite striking reproductive differences, exhibit largely clonal population genetic structures.
Phylogeography of lemmings (Lemmus): no evidence for postglacial colonization of Arctic from the Beringian refugium
Authors: Fedorov V. B. , Goropashnaya A. V. , Jaarola M. , Cook J. A.
Published: March 2003

Beringia is considered as an important glacial refugium that served as the main source for colonization of formerly glaciated Arctic regions. To obtain high resolution views of Arctic refugial history, we examined mitochondrial cytochromebphylogeography in the northern genus of rodents,Lemmus (true lemmings), sampled across its circumpolar distribution. Strong phylogeographical structure suggests vicariant separation over several glacial–interglacial periods and does not provide evidence supporting the importance of Beringia for extensive colonization of formerly glaciated regions. Rather than a source of postglacial colonization, Beringia represents an area of intraspecific endemism previously undetected by biogeographical analysis. Existing phylogeographical structure suggests that vicariant separation by glacial barriers was an important factor generating genetic divergence and, thus, increasing genetic diversity in lemmings on continental and circumpolar scales. However, there is little evidence for the direct effect of the last glaciation on the level of genetic variation and allele genealogy in lemmings on a regional geographical scale. This finding implies that the population genetic models of postglacial colonization suggested for temperate taxa might have limited applicability for Arctic species.
Regulated transcription of Clostridium difficile toxin genes
Authors: Dupuy Bruno , Sonenshein Abraham L.
Published: March 2003

The Clostridium difficile toxA and toxB genes, encoding cytotoxic and enterotoxic proteins responsible for antibiotic‐associated colitis and pseudomembranous colitis, were shown to be transcribed both from gene‐specific promoters and from promoters of upstream genes. However, the gene‐specific transcripts represented the majority of tox gene mRNAs. The 5′ ends of these mRNAs were shown to correspond to DNA sequences that had promoter activity when fused to the Escherichia coliβ‐glucuronidase (gusA) gene and introduced into C. perfringens. The appearance of tox mRNA in C. difficile was repressed during exponential growth phase but increased substantially as cells entered stationary phase. When glucose or other rapidly metabolizable sugars were present in the medium, the stationary phase‐associated induction was inhibited, indicating that the toxin genes are subject to a form of catabolite repression. This glucose effect was general to many toxinogenic strains having varying levels of toxin production.
Plant‐adapted green fluorescent protein is a versatile vital reporter for gene expression, protein localization and mitosis in the filamentous fungus, Aspergillus nidulans
Authors: Fernández‐Ábalos José Manuel , Fox Helen , Pitt Chris , Wells Brian , Doonan John H.
Published: March 2003

Green fluorescent protein (GFP) is a useful reporter to follow the in vivo behaviour of proteins, but the wild‐type gfp gene does not function in many organisms, including many plants and filamentous fungi. We show that codon‐modified forms of gfp, produced for use in plants, function effectively in Aspergillus nidulans both as gene expression reporters and as vital reporters for protein location. To demonstrate the use of these modified gfps as reporter genes we have used fluorescence to follow ethanol‐induced GFP expression from the alcA promoter. Translational fusions with the modified gfp were used to follow protein location in living cells; plant ER‐retention signals targeted GFP to the endoplasmic reticulum, whereas fusion to the GAL4 DNA‐binding domain targeted it to the nucleus. Nuclear‐targeted GFP allowed real‐time observation of nuclear movement and division. These modified gfp genes should provide useful markers to follow gene expression, organelle behaviour and protein trafficking in real time.
The CspA family in Escherichia coli : multiple gene duplication for stress adaptation
Authors: Yamanaka Kunitoshi , Fang Li , Inouye Masayori
Published: March 2003

CspA was originally found as the major cold‐shock protein in Escherichia coli, consisting of 70‐amino‐acid residues. It forms a β‐barrel structure with five anti‐parallel β‐strands and functions as an RNA chaperone. Its dramatic but transient induction upon cold shock is regulated at the level of transcription, mRNA stability and translation. Surprisingly, E. coli contains a large CspA family, consisting of nine genes from cspA to cspI. Phylogenetic analysis of these gene products and the cold‐shock domain of human YB‐1 protein reveals that there are two major branches in the evolution of CspA homologues: one branch for CspF and CspH, and another for all the other known CspA homologues from both prokaryotes and eukaryotes. The locations of these genes on the E. coli chromosome suggest that the large CspA family probably resulted from a number of gene duplications and, after subsequent adaptation, resulted in specific groups of genes that respond to different environmental stresses; for example, cspA, cspB and cspG for cold‐shock stress and cspD for nutritional deprivation. The E. coli CspA family will be discussed in terms of their structures and functions, and their gene structures and regulation.
From famine to feast: the role of methylglyoxal production in Escherichia coli
Authors: Tötemeyer S. , Booth N. A. , Nichols W. W. , Dunbar B. , Booth I. R.
Published: March 2003

The enzyme methylglyoxal synthase (MGS) was partially purified from Escherichia coli extracts, and the amino‐terminal sequence of candidate proteins was determined, based on the native protein being a tetramer of about 69 kDa. Database analysis identified an open reading frame in the E. coli genome, YccG, corresponding to a protein of 16.9 kDa. When amplified and expressed from a controlled promoter, it yielded extracts that contained high levels of MGS activity. MGS expressed from the trc promoter accumulated to approximately 20% of total cell protein, representing approximately 900‐fold enhanced expression. This caused no detriment during growth on glucose, and the level of methylglyoxal (MG) in the medium rose to only 0.08 mM. High‐level expression of MGS severely compromised growth on xylose, arabinose and glycerol. A mutant lacking MGS was constructed, and it grew normally on a range of carbon sources and on low‐phosphate medium. However, the mutant failed to produce MG during growth on xylose in the presence of cAMP, and growth was inhibited.
Secretion of the Haemophilus influenzae HMW1 and HMW2 adhesins involves a periplasmic intermediate and requires the HMWB and HMWC proteins
Authors: St Geme, Iii Joseph W. , Grass Susan
Published: March 2003

Non‐typable Haemophilus influenzae is a common cause of human disease and initiates infection by colonizing the upper respiratory tract. The non‐typable H. influenzae HMW1 and HMW2 non‐pilus adhesins mediate attachment to human epithelial cells, an essential step during colonization. In order to facilitate interaction with host cells, HMW1 and HMW2 are localized on the surface of the organism in a process that involves cleavage of a 441‐amino‐acid N‐terminal fragment. In the present study, we investigated the pathway for the secretion of HMW1 and HMW2. Cell fractionation experiments and cryoimmunoelectron microscopy demonstrated that a periplasmic intermediate occurs, suggesting involvement of the Sec machinery. Additional analysis revealed that, ultimately, the proteins are partially released from the surface of the organism. Studies with Escherichia coli harbouring plasmid subclones extended earlier findings and suggested that the secretion of HMW1 requires accessory proteins designated HMW1B and HMW1C, while the secretion of HMW2 requires proteins called HMW2B and HMW2C. Further analysis established that HMW1B/HMW1C and HMW2B/HMW2C are interchangeable, an observation consistent with the high degree of homology between HMW1B and HMW2B and between HMW1C and HMW2C. Additional studies of the hmw1 locus indicated that HMW1B is located in the outer membrane and serves to translocate HMW1 across the outer membrane. In the absence of HMW1B, HMW1 remains unprocessed and is degraded in the periplasmic space, at least in part by the DegP protease. Mutagenesis of an HMW1 N‐terminal motif shared with other secreted proteins resulted in diminished processing and extracellular release, suggesting interaction of this motif with the HMW1B protein. Continued investigation of the HMW1 and HMW2 adhesins may provide general insights into protein secretion and bacterial pathogenesis.
In vitro biosynthesis of the Pseudomonas aeruginosa quorum‐sensing signal molecule N‐butanoyl‐L‐homoserine lactone
Authors: Jiang Yan , Camara Miguel , Chhabra Siri Ram , Hardie Kim R. , Bycroft Barrie W. , Lazdunski Andrée , Salmond George P. C. , Stewart Gordon S. A. B. , Williams Paul
Published: March 2003

In Pseudomonas aeruginosa, synthesis of the quorum‐sensing signal molecules N‐butanoyl‐L‐homoserine lactone (BHL) and N‐hexanoyl‐L‐homoserine lactone (HHL) requires the LuxI homologue RhlI(VsmI). By using thin‐layer chromatography in conjunction with high‐performance liquid chromatography (HPLC) and mass spectrometry, we show that purified RhlI can catalyse the biosynthesis of BHL and HHL using either S‐adenosylmethionine (SAM) or homoserine lactone (HSL) but not homoserine as the source of the homoserine lactone moiety. As we were unable to detect homoserine lactone in cytoplasmic extracts of Escherichia coli, we conclude that SAM is the natural substrate for RhlI‐directed N‐acylhomoserine lactone (AHL) biosynthesis. The N‐acyl chain of BHL and HHL can be supplied by the appropriately charged coenzyme A derivative (either n‐butanoyl‐CoA or n‐hexanoyl‐CoA). The specificity of RhlI for charged CoA derivatives is demonstrated as RhlI was unable to generate AHLs detectable in our bioassays from acetyl‐CoA, malonyl‐CoA, n‐octanoyl‐CoA, n‐decanoyl‐CoA, DL‐β‐hydroxybutanoyl‐CoA or crotonoyl‐CoA. RhlI was also unable to use N‐acetyl‐S‐3‐oxobutanoylcysteamine, a chemical mimic for 3‐oxobutanoyl‐CoA. Furthermore, the RhlI‐catalysed synthesis of BHL and HHL was most efficiently driven when NADPH was included in the reaction mixture.
Initiation of biofilm formation in Pseudomonas fluorescens WCS365 proceeds via multiple, convergent signalling pathways: a genetic analysis
Authors: O'Toole George A. , Kolter Roberto
Published: March 2003

Populations of surface‐attached microorganisms comprising either single or multiple species are commonly referred to as biofilms. Using a simple assay for the initiation of biofilm formation (e.g. attachment to an abiotic surface) by Pseudomonas fluorescens strain WCS365, we have shown that: (i) P. fluorescens can form biofilms on an abiotic surface when grown on a range of nutrients; (ii) protein synthesis is required for the early events of biofilm formation; (iii) one (or more) extracytoplasmic protein plays a role in interactions with an abiotic surface; (iv) the osmolarity of the medium affects the ability of the cell to form biofilms. We have isolated transposon mutants defective for the initiation of biofilm formation, which we term surface attachment defective (sad ). Molecular analysis of the sad mutants revealed that the ClpP protein (a component of the cytoplasmic Clp protease) participates in biofilm formation in this organism. Our genetic analyses suggest that biofilm formation can proceed via multiple, convergent signalling pathways, which are regulated by various environmental signals. Finally, of the 24 sad mutants analysed in this study, only three had defects in genes of known function. This result suggests that our screen is uncovering novel aspects of bacterial physiology.
Lactococcus lactis phage operon coding for an endonuclease homologous to RuvC
Authors: Bidnenko Elena , Ehrlich S. Dusko , Chopin Marie‐Christine
Published: March 2003

The function of the Lactococcus lactis bacteriophage bIL66 middle time‐expressed operon (M‐operon), involved in sensitivity to the abortive infection mechanism AbiD1, was examined. Expression of the M‐operon is detrimental to Escherichia coli cells, induces the SOS response and is lethal to recA and recBC E. coli mutants, which are both deficient in recombinational repair of chromosomal double‐stranded breaks (DSBs). The use of an inducible expression system allowed us to demonstrate that the M‐operon‐encoded proteins generate a limited number of randomly distributed chromosomal DSBs that are substrates for ExoV‐mediated DNA degradation. DSBs were also shown to occur upstream of the replication initiation point of unidirectionally theta‐replicating plasmids. The characteristics of the DSBs lead us to propose that the endonucleolytic activity of the M‐operon is not specific to DNA sequence, but rather to branched DNA structures. Genetic and physical analysis performed with different derivatives of the M‐operon indicated that two orfs (orf2 and orf3) are needed for nucleolytic activity. The orf3 product has amino acid homology with the E. coli RuvC Holliday junction resolvase. By site‐specific mutagenesis, we have shown that one of the amino acid residues constituting the active centre of RuvC enzyme (Glu‐66) and conserved in ORF3 (Glu‐67) is essential for the nucleolytic activity of the M‐operon gene product(s). We therefore propose that orf2 and orf3 of the M‐operon code for a structure‐specific endonuclease (M‐nuclease), which might be essential for phage multiplication.