bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection of antibodies to hepatitis C virus in seronegative patients using an immune complex dissociation assay
Authors: Troisi C. L. , Hollinger F. B.
Published: November 2003

Summary. Sera from a small percentage of hepatitis C virus (HCV)‐infected blood donors do not react in the currently available assays for detection of antibody to HCV (anti‐HCV) and, as a consequence, hepatitis C may develop in recipients of this blood. One possible explanation for this phenomenon is that antibody is present but cannot be detected because it is sequestered in circulating immune complexes. To test this hypothesis, an immune complex dissociation (ICD) assay was developed to disrupt any immune complexes that might be present in these anti‐HCV‐negative, HCV RNA‐positive sera. A positive result in this test would indicate that antibody is present in these patients but is not detectable under routine anti‐HCV testing conditions. Nine chronic and two acute HCV patients, all negative for antibody but positive for HCV RNA by reverse transcriptase‐polymerase chain reaction (RT‐PCR) were tested, together with appropriate controls. Three of the nine study patients with chronic HCV had evidence of anti‐HCV after immune complex dissociation compared with none of the two patients with acute HCV. Although the number of patients tested was small, the negative results in the patients with acute HCV presumably indicates that anti‐HCV seroconversion had not yet occurred. Incorporation of an ICD step into existing anti‐HCV assays may enable blood banks to detect those rare instances of patients with chronic HCV who are antibody negative; this would minimize potential cases of post‐transfusion hepatitis in recipients.
Concordance between hepatitis C virus serotyping assays
Authors: Gish R. G. , Qian K. P. , Quan S. , Xu Y. L. , Pike I. , PoIito A. , DiNello R. , Lau J. Y. N.
Published: November 2003

Summary. Hepatitis C virus testing has evolved from a simple enzyme‐linked immunosorbent assay (ELISA) to complex molecular tests including qualitative and quantitative polymerase chain reaction (PCR) as well as multiple methods to determine geno and serotypes. Serotyping assays have been described and are being further refined to aid describing the epidemiology of hepatitis C virus (HCV) infection and may have a role in predicting response treatment. This study describes the concordance between two serotyping assay systems, a recombinant immunoblot assay Chiron RIBATM Strip Immunoblot Assay (SIA) and a more competitive ELISA peptide assay, using PCR as the standard. Serotype was successfully determined in 144/202 (71%) patients by the Murex ELISA assay and 179/202 (89%) by the Chiron strip immunoblot assay (SIA) assay (P < 0.001). Concordance between restriction fragment length polymorphism (RFLP) and the Murex assay was 139/144 (97%) between RFLP and the Chiron SIA was 171/179 (96%) and between the Murex and Chiron SIA was 136/144 (94%). These assays provided a reliable, simple, and rapid method of determining HCV serotype.
Prevalence of hepatitis G virus in patients with hepatocellular carcinoma
Authors: Müller C. , Pfeffel F. , Peck‐Radosavljevic M. , Petermann D. , Oesterreicher C. , Pidlich J.
Published: November 2003

Summary. Hepatitis B virus (HBV) and hepatitis C virus (HCV) are known to be associated with hepatocellular carcinoma (HCC), In this study, we investigated the prevalence of the newly described hepatitis G virus (HGV) in patients with HCC. The sera of 85 patients (66 male, 19 female, 61 ± 11 years) with HCC were studied for the presence of HGV RNA by reverse transcriptase‐polymerase chain reaction. Seventeen (20%) of 85 patients with HCC, 10 (16%) of 61 patients with chronic hepatitis B without HCC and 14 (20%) of 68 patients with chronic hepatitis C without HCC were infected with HGV, a significantly higher proportion when compared with two (2%) of 85 healthy controls (P < 0.01). When grouped according to the underlying cause of liver disease, HCC patients with HBV infection (33%), HCV infection (21%), alcoholic liver disease (17%). or cryptogenic cirrhosis (15%) had similar serum levels of HGV RNA. Four of the 17 (24%) HGV‐positive patients with HCC were coinfected with HBV and six (35%) with HCV; thus, 59% of HGV‐positive patients with HCC were coinfected with other hepatotropic viruses. Seven (41%) HGV‐positive patients were infected with HGV only. Patients with HGV infection were more likely to have a history of blood transfusion than patients without HGV infection (P= 0.024). Hence, the prevalence of HGV is significantly higher in patients with HCC in comparison with the healthy population.
Absence of anti‐envelope antibodies and clearance of hepatitis C virus in a cohort of Irish women infected in 1977
Authors: Grellier L. , Brown D. , Power J. , Dusheiko G.
Published: November 2003

Summary. The presence or absence of antibodies to the second envelope protein (anti‐E2) of hepatitis C virus (HCV) was determined in stored sera taken from a cohort of 87 Irish women with antibodies to HCV (anti‐HCV) who were all infected by HCV genotype 1b from contaminated anti‐D immunoglobulin given in 1977. Anti‐E2 was found in 16 patients (100%) who were HCV RNA positive but only in 31 of 50 patients (62%) who were HCV antibody positive by recombinant immunoblot assay (RIBA) but HCV RNA negative. In the remaining 21 sera taken from women who had indeterminate recombinant immunoblot assays and who were repeatedly negative on testing for HCV RNA, anti‐E2 was found in only three cases (14%). This suggests that loss or absence of anti‐E2 may be useful in confirming clearance of HCV.
Variations of hepatitis B virus core gene sequence in Western patients with chronic hepatitis B virus infection
Authors: Gray A. H. , Fang J. W. S. , Davis G. L. , Mizokami M. , Wu P. C. , Williams R. , Schuster S. M. , Lau J. Y. N.
Published: November 2003

Summary. Heterogeneity of the hepatitis B virus (HBV) core gene has been reported to be associated with the presence of active liver disease in Japanese patients with chronic HBV infection. This study evaluated the significance of HBV core gene heterogeneity in Western patients with chronic HBV infection. The hepatitis B virus precore/core gene from 45 patients (inactive:active liver disease ratio 16:29) was amplified from serum by polymerase chain reaction (PCR). Gel electrophoresis was employed to detect large deletions. The PCR amplicons from 13 patients (all HBV serotype adw but with a different spectrum of liver disease) were cloned and sequenced. Hepatitis B surface antigen (HBsAg) serotypes were tested by enzyme immunoassay (EIA) and hepatic expression of HBV antigens was assessed by immunohistochemistry. The HBV core gene was amplified from the serum of all 45 patients. Three patients had mixed infection with both precore mutant and wild‐type HBV and all three had active liver disease. No patient had a large deletion of the HBV core gene. Hepatitis B virus core gene sequence variations were more common in the midcore region and there was no difference in the number of silent and missense substitutions between those with inactive and active liver disease. There was no correlation between the nucleotide or encoded amino acid substitutions and the clinical and biochemical parameters, including the subsequent response to interferon‐α therapy (n= 37) or hepatic HBV antigen expression. Variation of the HBV core gene was not found to be preferentially associated with active liver disease in Western patients with chronic HBV infection. The pattern of hepatitis B core gene variation is in accord with the genomic organization of HBV.
Phylogenetic relationships and introgression patterns between incipient parapatric species of Italian brown trout (Salmo trutta L. complex)
Authors: GIUFFRA E. , GUYOMARD R. , FORNERIS G.
Published: November 2003

Genetic variation at 47 protein loci was investigated in 16 wild brown trout populations from the Po basin and three major domesticated stocks used for stocking this area. Twenty‐four loci were polymorphic and large frequency differences were found at 15 of them. The most significant allozyme variations were congruent with the mtDNA sequence polymorphism previously observed in the same samples. We confirmed the occurrence of two parapatric incipient species, Salmo marmoratus and S. trutta fario, previously identified by morphological traits. These two species were fixed or nearly fixed for alternate alleles at eight loci (Nei's standard genetic distance = 0.16–0.18), but introgression was detected between adjacent samples of the two forms. Divergence levels at both mtDNA and nuclear loci suggested that the differentiation between S. marmoratus and S. trutta fario started between 3 and 1 million years before present. Variation at protein loci and mtDNA supported the hypothesis that the third species found in this area, S. carpio (an endemic population of the lake Garda) was issued from a recent hybridization of the two first species. Finally, we showed that three of the major Italian fish‐farm strains originated from the Atlantic side and displayed substantial genetic differences with the natural populations of the Po basin. Most of these populations were contaminated by stocking with introgression rate ranging from 0 to 70% and measures of protection and restoration of the rich genetic diversity present in this area should be urgently applied.
Highly conserved nuclear copies of the mitochondrial control region in the desert locust Schistocerca gregaria: some implications for population studies
Authors: ZHANG DE‐XING , HEWITT GODFREY M.
Published: November 2003

Animal mitochondrial DNA has proved a valuable marker in intraspecific systematic studies. However, if nucleotide sequence heterogeneity exists at the individual level, its usefulness will be much reduced. This study demonstrates that the presence of highly conserved non‐coding mitochondrial sequences in the nuclear genome of Schistocerca gregaria greatly impairs the use of mtDNA in population genetic studies. Caution is called for in other organisms; and it seems necessary to check for conserved nuclear copies of mitochondrial sequences before launching into a large scale analysis of populations using mtDNA as a genetic marker. Experimental procedures are suggested for this purpose.
RAPD identification of microsatellites in Daphnia
Authors: ENDER A. , SCHWENK K. , STÄDLER T. , STREIT B. , SCHIERWATER B.
Published: November 2003

Simple sequence repeats (SSRs, or microsatellites) have been constantly gaining importance as single‐locus DNA markers in population genetics and behavioural ecology. We tested a PCR‐based strategy for finding microsatellite loci in anonymous genomes, which avoids genomic library construction and screening, and the need for larger amounts of DNA. In the first step, parts of a genome are randomly amplified with arbitrary 10mer primers using RAPD fingerprinting. Labelled SSR‐oligonucleotides serve as probes to detect complementary sequences in RAPD products by means of Southern analyses. Subsequently, positive RAPD fragments of suitable size are cloned and sequenced. Using GA and GT probes, we applied this approach to waterfleas (Daphnia) and revealed 37 hybridization signals in 20 RAPD profiles. Thirteen positive RAPD fragments from three Daphnia species and two hybrid ‘species’ were cloned and sequenced. In all cases simple sequence repeats were detected. We characterized seven perfect repeat loci, which were found to be polymorphic within and between species.
Cytonuclear genetic structure of a hybrid zone in lizards of theSceloporus grammicuscomplex (Sauria, Phrynosomatidae)
Authors: SITES J. W. , BASTEN C. J. , ASMUSSEN M. A.
Published: November 2003

Lizards of theSceloporus grammicuscomplex are comprised of multiple chromosome races that form several zones of parapatric hybridization in central Mexico. We scored diagnostic mitochondrial DNA (mtDNA) haplotypes and autosomal chromosome markers in a sample of 342 lizards from one well‐defined zone between 2n = 34 and 2n = 46 races. A two‐part analysis was performed on this data set in an attempt to infer the predominant evolutionary forces shaping the cytonuclear structure of this zone. The complications posed by its spatial structure were addressed by analysing a hierarchical series of smaller subsamples chosen to approximate single mating units. Two critical conclusions were drawn from this first‐stage analysis. First and foremost, the three chromosomes have largely concordant cytonuclear disequilibrium patterns within each subsample with adequate numbers of individuals for detecting nonrandom cytonuclear associations. This suggests that the cytonuclear structure of this zone is predominantly a result of deterministic genome‐wide forces rather than genetic drift or deterministic forces specific to individual chromosomes or loci. Second, the fit of a series of migration models to the data shows that the cytonuclear structure of the subsamples is well accounted for by continued gene flow from the two parental races alone, with random mating with respect to cytonuclear genotype and no other evolutionary forces. These results motivate several further empirical and theoretical investigations to refine our understanding of the relative roles of migration and other potentially important forces such as natural selection and genetic drift, in this and other hybrid zones.
Phylogeography and population structure of the Atlantic and Mediterranean green turtle Chelonia mydas: a mitochondrial DNA control region sequence assessment
Authors: Encalada S. E. , Lahanas P. N. , Bjorndal K. A. , Bolten A. B. , Miyamoto M. M. , Bowen B. W.
Published: November 2003

Mitochondrial (mt) DNA sequences were analysed to resolve the phylogeography and population genetic structure of Atlantic and Mediterranean populations of green turtles (Chelonia mydas). Analysis of sequence variation over 487 base pairs of the control (D‐loop) region identified 18 haplotypes among 147 individuals from nine nesting populations. Pairwise comparisons of haplotype frequencies distinguished most nesting colonies, indicating significant genetic differentiation among rookeries and a strong propensity for natal homing behaviour by nesting females. Comparison of control region sequence data to earlier restriction fragment length polymorphism (RFLP) data for the same individuals demonstrates approximately a sixfold higher substitution rate in the 5′ end of the control region. The sequence data provide higher resolution both in terms of the number of mtDNA genotype variants and the phylogeographic relationships detected within the Atlantic region, and reveal a gene genealogy that distinguishes two groups of haplotypes corresponding to (i) the western Caribbean and Mediterranean, and (ii) eastern Caribbean, South Atlantic and West Africa. The data suggest that phylogeographic patterns in the Atlantic Ocean may be interpreted in terms of female nest site fidelity and episodic dispersal events. The distribution of mtDNA haplotypes within the region is thus explained by the geological and climatic alternations (glacial and interglacial) over the last million years.
Microsatellites from the beluga whale Delphinapterus leucas
Authors: Buchanan F. C. , Friesen M. K. , Littlejohn R. P. , Clayton J. W.
Published: November 2003

Fifteen microsatellites were isolated from a beluga whale Delphinapterus leucas, genomic library. The microsatellites were amplified in 100 beluga obtained from two widely separated locations. An average of 8.6 alleles per locus were detected and the average heterozygosity was 0.65 with a range of 0.27–0.86. All microsatellites were polymorphic and 13 of the genotype distributions observed were in Hardy‐Weinberg equilibrium. It was possible with these microsatellites to assign correctly individual whales to their stock‐of‐origin 98% of the time. Microsatellites were amplified in 15 other cetaceans with these beluga‐derived primers.
Genetic structure of the global population of banana black leaf streak fungus, Mycosphaerella fijiensis
Authors: Carlier J. , Lebrun M. H. , Zapater M. F. , Dubois C. , Mourichon X.
Published: November 2003

The genetic structure of Mycosphaerella fijiensis populations around the world was examined using DNA restriction fragment length polymorphism (RFLP) markers. Allele frequencies at 19 nuclear RFLP loci were estimated in a sample of 136 M. fijiensis isolates from five geographical populations representative of banana‐producing areas (South‐East Asia including the Philippines and Papua New Guinea, Africa, Latin America and Pacific Islands). Within each population, gametic disequilibrium tests between the 19 nuclear RFLP loci were mainly non significant (P > 0.05), indicating that random sexual reproduction occurred in these populations. All M. fijiensis populations had a high level of genotypic and allelic diversity (H, gene diversity: 0.25–0.59). The highest levels of gene diversity were found in the two South‐East Asian populations (H: 0.57 and 0.59). Most of the alleles (> 88%) detected in Africa, Latin America and Pacific Islands populations were also detected in South‐East Asian populations. Furthermore, a high and significant (P < 0.05) level of genetic differentiation was observed among M. fijiensis geographical populations (overall estimate of Fst: 0.32). These results were consistent with the hypothesis that M. fijiensis originated in South‐East Asia and spread recently to other parts of the world. The level of allelic diversity in M. fijiensis populations from regions other than South‐East Asia was drastically reduced, indicating founder effects. The data also suggested rare occurrence of migration of M. fijiensis between continents.
Relatedness structure detected by microsatellite analysis and attempted pedigree reconstruction in an endangered marsupial, the northern hairy‐nosed wombat Lasiorhinus krefftii
Authors: Taylor A. C. , Horsup A. , Johnson C. N. , Sunnucks P. , Sherwin B.
Published: November 2003

The northern hairy‐nosed (NHN) wombat is perhaps Australia's most endangered mammal. Being fossorial and nocturnal as well as rare, NHN wombats are difficult to observe in the wild. Hence little is known of their social biology, such as their mating and dispersal systems. A hypothesis has been advanced that adult females of the species disperse post‐breeding, leaving their young to inhabit the natal burrow. Female‐biased dispersal is expected to result in higher relatedness amongst males in a burrow cluster than amongst females in a burrow cluster. The usefulness of a panel of microsatellite markers in estimating the relatedness structure, and in reconstructing pedigrees for, the sole known population of NHN wombats was assessed. Microsatellite genotypes at eight or nine loci were obtained from 58 of the 85 known individuals, and used to estimate pairwise individual relatedness using Queller & Goodnight's (1989) RELATEDNESS 4.2. Our analysis gave the unexpected result that both males and females were significantly more closely related to their same‐sex burrow cluster mates than random, while opposite‐sex animals sharing burrows were only slightly (nonsignificantly) more related than random. This raises the possibility of dispersal patterns which lead to association of same‐sex relatives. The observed relatedness structure is not expected to make likely a high incidence of inbred matings, as close relatives of the opposite sex are not significantly associated in space. Parentage analysis was attempted using genetic exclusion and LOD likelihood ratios, but proved difficult because of low genetic variation, incomplete sampling of potential parents, and paucity of ecological data such as known mother/offspring pairs and ages of individuals.
Genetic differences between the endangered San Clemente Island loggerhead shrike Lanius ludovicianus mearnsi and two neighbouring subspecies demonstrated by mtDNA control region and cytochrome b sequence variation
Authors: Mundy N. I. , Winchell C. S. , Woodruff D. S.
Published: November 2003

We investigated mtDNA sequence variation in five populations of the loggerhead shrike Lanius ludovicianus, representing four subspecies, including the San Clemente loggerhead shrike L. l. mearnsi, a critically endangered California Channel Island endemic. Variability in 200 bp of control region and 200 bp of cytochrome b was extremely low, and defined four haplotypes. Strong structure was apparent among all three southern Californian subspecies, including L. l. mearnsi, with one haplotype predominating in each subspecies. Although potential levels of gene flow between L. l. mearnsi and neighbouring populations are low, mtDNA data support field observations that some shrikes visit the island during winter but do not stay to breed, and suggest that these birds come from the mainland. The similarity in haplotypes between populations from Saskatchewan, Canada and those in southern California suggests post‐glacial northern range expansion of the species. Our results confirm the evolutionary distinctiveness of L. l. mearnsi and justify continuing efforts for its conservation.
Sex‐related spatial kin structure in a spring population of grey‐sided voles Clethrionomys rufocanus as revealed by mitochondrial and microsatellite DNA analyses
Authors: Ishibashi Y. , Saitoh T. , Abe S. , Yoshida M. C.
Published: November 2003

Polymerase chain reaction‐directed mitochondria (mt) and microsatellite DNA analyses were performed to examine the kin structure in a spring population of grey‐sided voles Clethrionomys rufocanus in Hokkaido, Japan. The spatial distribution of 81 voles in a trapping grid (about 1 ha) was estimated by using the catch‐mark‐release method. DNA samples were extracted from the toes clipped for individual identification. Maternal lineages of voles were unequivocally determined by the mtDNA haplotypes, as identified by nucleotide sequencing of the control region. Relatedness between individuals was estimated based on the genotype and allele frequencies at several microsatellite loci. Although the distribution of voles was uniform within the grid, neighbouring females were frequently from the same maternal lineage. Relatedness values between females correlated negatively with geographical distances. Combination of the two molecular markers revealed four clusters of closely related, matrilineal females in the population, whereas no such cluster was apparent in males. The present study first demonstrated a sex‐related spatial kin structure in a natural population of the grey‐sided vole.
Mitochondrial DNA analysis confirms the existence of two glacial races of rainbow smelt Osmerus mordax and their reproductive isolation in the St Lawrence River estuary (Quebec, Canada)
Authors: Bernatchez L.
Published: November 2003

A phylogeographic analysis of mitochondrial DNA variation was performed in order to test the hypothesis that north‐eastern North America has been postglacially recolonized by two races of rainbow smelt Osmerus mordax. This was accomplished by documenting the geographical distribution of two major mtDNA phylogenetic clades among 1290 smelt from 49 lacustrine and anadromous populations covering most of the species' native range. The data set was built by combining previously published results with those generated in this study. The two mtDNA clades showed a geographical dichotomy, independent of life‐history types, whereby the more eastern populations were either fixed or largely dominated by one clade and western populations for the other. Such geographical pattern implying a phylogenetic discontinuity provided strong evidence for the persistence of smelt in two distinct glacial refugia as well as their differential postglacial dispersal. The most likely refugium for the so‐called Atlantic race was the Atlantic coastal plains, whereas that of the so‐called Acadian race was the exundated Grand Banks area. Patterns of postglacial dispersal interpreted from palaeogeographic events suggested that the Atlantic race recolonized northern regions about 5000 years prior the Acadian race. Both races came into contact in the St Lawrence River estuary. While gene flow has been possible, the sympatric occurrence in the estuary of anadromous populations alternatively dominated by one mtDNA clade or the other indicated that reproductive isolation mechanisms between the two races developed within this contact zone. This represents the first evidence of secondary intergradation among distinct races of aquatic organisms in an estuarine environment.
Molecular evidence for the maternal parentage in the hybrid origin of Spartina anglica C.E. Hubbard
Authors: Ferris C. , King R. A. , Gray A. J.
Published: November 2003

Spartina anglica is a textbook example of a natural amphiploid, which originated from hybridization between S. alterniflora and S. maritima. Which of these species was the maternal parent has remained a mystery. Inheritance of chloroplast DNA in most angiosperms is strictly maternal and can thus be used to test the parentage of hybrid taxa. The DNA sequence of the chloroplast leucine tRNA gene intron was used to show that the introduced North American S. alterniflora is the female parent of the F1 hybrid S. x townsendii and the amphiploid S. anglica. A possible scenario for their origin is given.
Population structure and genetic diversity in four tropical tree species in Costa Rica
Authors: Schierenbeck K. A. , Skupski M. , Lieberman D. , Lieberman M.
Published: November 2003

Despite intensified interest in conservation of tropical forests, knowledge of the population genetics of tropical forest trees remains limited. We used random amplified polymorphic DNA (RAPD) data to evaluate trends in genetic diversity and differentiation for four tropical tree species, Alchornea latifolia, Dendropanax arboreus, Inga thibaudiana and Protium glabrum. These species occur at contrasting population densities along an elevational gradient and we use RAPD and ecological data to examine natural levels of genetic diversity of each species, trends in genetic variability with population density and structure, genetic differentiation along the elevation gradient, and the relationship between genetic diversity and such factors as seed dispersal and pollination syndrome. At the distances we examined (plot distances ranging from 0.8 to 8.6 km) there was very little genetic structuring at any distance along the gradient. All four species exhibited levels of variation expected for spatial distribution, mating system and pollinator syndrome; greater than 96% of the genetic variation occurred within plots for Inga thibaudiana, Protium glabrum and Dendropanax arboreus. Alchornea latifolia only occurred in a single plot. The results of this study contribute to a growing database of genetic diversity data that can be utilized to make predictions about the effect of disturbance and subsequent reductions in population size on genetic variation and structure in tropical tree species.
Cyprinotus incongruens (Ostracoda): an ancient asexual?
Authors: Chaplin J. A. , Hebert P. D. N.
Published: November 2003

Although unisexual ostracods are common, their evolutionary history is now known only from inferences gained through examination of the fossil record. Here we use mtDNA, allozyme and genome size analyses to investigate the origins of unisexuality, polyploidy and clonal diversity in the freshwater ostracod Cyprinotus incongruens. Our genetic evidence suggests that transitions to polyploidy have been common in this taxon and may sometimes involve internal genome‐fusion events, in contrast to the usual origin of animal polyploids through interspecific hybridization. Both the extent and congruent patterns of allozyme and mtDNA divergence amongst clones of C. incongruens are consistent with its persistence as an asexual taxon for approximately 5 million years. However, the detected patterns of genetic variation might also reflect the origins of this ostracod through a series of independent transitions to asexuality by several closely related ancestral taxa. The results make it clear that efforts to demonstrate the antiquity of asexual taxa, solely from surveys of the extent of genetic divergence among their component lineages, will ordinarily be ambiguous, requiring confirmation through investigations which search the genome for the genetic consequences of long abandoned recombination.
Microsatellite analysis of maternity and the mating system in the Gulf pipefish Syngnathus scovelli, a species with male pregnancy and sex‐role reversal
Authors: Jones A. G. , Avise J. C.
Published: November 2003

Highly variable microsatellite loci were employed to study the mating system of the sexually dimorphic Gulf pipefish Syngnathus scovelli. In this species, like others in the family Syngnathidae, ‘pregnant’ males provide all parental care. Gulf pipefish were collected from one locale in the northern Gulf of Mexico, and internally carried broods of 40 pregnant males were analysed genetically. By comparing multilocus microsatellite fingerprints for the inferred mothers against expected genotypic distributions from the population sample, it was determined that: (i) only one male had received eggs from more than a single female; and (ii) on two separate occasions, two different males had received eggs from the same female. Given the high power to detect multiple matings by males, the first finding indicates that only rarely are individual males impregnated by multiple females during the course of a pregnancy. Conversely, given the lower power to detect multiple matings by females due to sampling constraints, the second finding suggests a high frequency of multiple successful matings by females. Thus, this population of Gulf pipefish displays a polyandrous genetic mating system. The relevance of these genetic findings is discussed with regard to the evolution of secondary sex traits in this species, and in other syngnathids.
Microgeographic structure of Anopheles gambiae in western Kenya based on mtDNA and microsatellite loci
Authors: Lehmann T. , Besansky N. J. , Hawley W. A. , Fahey T. G. , Kamau L. , Collins F. H.
Published: November 2003

The population genetic structure of the Anopheles gambiae in western Kenya was studied using length variation at five microsatellite loci and sequence variation in a 648‐nt mtDNA fragment. Mosquitoes were collected from houses in villages spanning up to 50 km distance, The following questions were answered, (i) Are mosquitoes in a house more related genetically to each other than mosquitoes between houses? (ii) What degree of genetic differentiation occurs on these geographical scales? (iii) How consistent are the results obtained with both types of genetic markers? At the house level, no differentiation was detected by FST and RST, and the band sharing index test revealed no significant associations of alleles across loci. Likewise, indices of kinship based on mtDNA haplotypes in houses were even lower than in the pooled sample. Therefore, the hypothesis that mosquitoes in a house are more related genetically was rejected. At increasing geographical scales, microsatellite allele distributions were similar among all population samples and no subdivision of the gene pool was detected by FST or RST. Likewise, estimates of haplotype divergence of mtDNA between populations were not higher than the within population estimates, and mtDNA‐based FST values were not significantly different from zero. That sequence variation in mtDNA provided matching results with microsatellite loci (while high genetic variation was observed in all loci), suggested that this pattern represents the whole genome. The minimum area associated with a deme of A. gambiae in western Kenya is therefore larger than 50 km in diameter.
Species‐independent, geographical structuring of chloroplast DNA haplotypes in a montane herb Ipomopsis (Polemoniaceae)
Authors: Wolf P. G. , Murray R. A. , Sipes S. D.
Published: November 2003

Two hypotheses have been proposed to explain the occurrence of hybrid zones between red‐flowered Ipomopsis aggregata and white‐flowered I. tenuituba. Either local adaptation to hummingbird and hawkmoth pollinators has given rise to sympatric (or parapatric) divergence of flower colour and morphology (primary intergradation at hybrid zones), or alternatively two previously allopatric species are coming into contact at several geographical areas of secondary intergradation. We examined restriction site patterns in nuclear DNA (nrDNA), chloroplast DNA (cpDNA) and mitochondrial DNA (mtDNA) from populations of I. aggregata and I. tenuituba representing seven zones of sympatry. No variation was detected in a 350‐bp fragment of mtDNA and uninformative levels of variation were observed for nrDNA. We detected 22 potentially informative restriction site polymorphisms in cpDNA, all of which united geographical areas containing populations of both species. We detected no informative species‐specific markers. Studies of other species (e.g. oaks) have detected similar species‐independent geographical structure of cpDNA. However, in these cases secondary interegradation could be inferred from species‐specific nuclear alleles. The pattern in Ipomopsis is consistent with both primary intergradation (independent speciation in each area of sympatry) or secondary intergradation involving complete cytoplasmic replacement. Thus, additional data are needed to explain the origin of hybrid zones in Ipomopsis.
Analysis of genetic relationships of Central American and Mexican pines using RAPD markers that distinguish species
Authors: Furman B. J. , Grattapaglia D. , Dvorak W. S. , O'Malley D. M.
Published: November 2003

Phylogenetic relationships were inferred for six Central American and Mexican pine species by analysing RAPD marker differences among pooled DNA samples. This population level pooling strategy discounts low‐frequency allelic variation within taxa, thus obtaining a ‘cumulative genotype’ to compare among taxa. We used the morphologically based taxonomy of pines as the basis for inference concerning molecular marker divergence. Only RAPD polymorphisms that were repeatable and intensely amplified were used. The resulting data set was found to have strong hierarchical structure. Phylogenetic analyses were carried out using both neighbour‐joining and maximum parsimony. The phylogenetic relationships indicated by analysis of the pine RAPD data provide new insights on pine taxonomy. The phylogenetic analyses of the RAPD marker data placed Pinus tecunumanii and P. patula clearly into two different subgroups, strongly supporting the classification of P. tecunumanii as a distinct species, and raise a new set of issues concerning the distinctions between Pinus tecunumanii and P. caribaea. Phylogenetically informative RAPD markers will be useful tools to address ex‐situ conservation issues that involve taxonomic identification and species admixture.
Cloned DNA probes distinguish endemic and exotic Entomophaga grylli fungal pathotype infections in grasshopper life stages
Authors: Bidochka M. J. , Walsh S. R. A. , Ramos M. E. , St. Leger R. J. , Carruthers R. I. , Silver J. C. , Roberts D. W.
Published: November 2003

Entomophaga grylli is a fungal pathogen of grasshoppers and at least three pathotypes are recognized world‐wide. Pathotypes 1 and 2 are endemic to North America while the Australian pathotype 3 had been released into two field sites in North Dakota between 1989 and 1991. Grasshoppers were collected over the summer at the field sites in 1992 and assessed for pathotype infection by cloned DNA probe analysis. The three most predominant grasshopper species that were infected (Melanoplus sanguinipes, M. bivittatus and Camnula pellucida) were assessed for pathotype infection with respect to their life stages (nymphal instars and adult males and females). Pathotype 1 predominantly infected grasshoppers in the subfamilies Oedipodinae and Gomphocerinae and pathotype 2 predominantly infected grasshoppers in the subfamily Melanoplinae. Early‐instar M. sanguinipes and M. bivittatus had higher pathotype 2 infection frequencies, while late‐instar and adult C. pellucida had higher pathotype 1 infection frequencies. Cross‐infection by the pathotypes did occur in up to 3% of the individuals, on a per species basis, and primarily in later instar and adult grasshoppers. Pathotype 3 infections occurred in later instar and adults of the three grasshopper species. Infection of grasshoppers by E. grylli pathotypes is discussed with reference to the fungal life cycles.
28s rDNA group‐I introns: a powerful tool for identifying strains of Beauveria brongniartii
Authors: Neuvéglise C. , Brygoo Y. , Riba G.
Published: November 2003

The nuclear ribosomal DNA of the entomopathogenic fungus Beauveria brongniartii is polymorphic in terms of both restriction site and length. Insertions of 350–450 bp long, identified as group‐I introns, were detected in the 28s rDNA. A panel of 47 strains of B. brongniartii, two B. bassiana and one Metarhizium anisopliae of various geographical and biological origins were found to contain 14 variant forms of intron differing in size and restriction pattern, at four different positions. Twelve types of ribosomal large subunit were defined on the basis of variant distribution and compared with strain clustering based on internal transcribed spacers analysis. There was a correlation between the characteristic introns and isolates collected from the sugar cane pest Hoplochelus marginalis. Primers for polymerase chain reaction amplification were chosen from these variants, and used to develop a specific method for detecting strains pathogenic towards Hoplochelus.
Effect of PCR template concentration on the composition and distribution of total community 16S rDNA clone libraries
Authors: Chandler D. P. , Fredrickson J. K. , Brockman F. J.
Published: November 2003

Total DNA from sediment samples was isolated by a direct lysis technique. Purified DNA was used as template either undiluted or diluted 1 : 10 prior to polymerase chain reaction (PCR) amplification of 16S rRNA genes. Full‐length inserts were analysed for restriction fragment length polymorphisms (RFLP) with the enzyme Cfo1, and the resulting distribution and abundance of RFLP patterns compared between the undiluted and diluted PCR reactions. Results indicate that for low PCR template concentrations, in the range from a few picograms to tens of picograms DNA, proportional representation of specific RFLP types was not reproducible upon template dilution, confirming that PCR amplification of 16S rDNA cannot be used directly to infer microbial abundance. In particular, only 15–24% of the RFLP types recovered from a sample were present in both the undiluted and diluted extracts. We propose that very low template concentrations in the PCR generate random fluctuations in priming efficiency, which led to the contrast in the RFLP types observed in the libraries from the undiluted and diluted extracts.
Genetic similarity in long‐eared owl communal winter roosts: a DNA fingerprinting study
Authors: Galeotti P. , Pilastro A. , Tavecchia G. , Bonetti A. , Congiu L.
Published: November 2003

The long‐eared owl is a nocturnal predator which winters communally and breeds in the same areas in loose colonies during the spring. We tested the hypothesis that roosts, particularly stable roosts, are formed by close relatives, a condition under which group‐related behaviours such as information sharing or helping at nest are more likely to be developed. DNA fingerprinting analysis was used to examine genetic similarity within and between two long‐eared owl populations, one wintering in a traditional roost and the other in an unstable roost, and both breeding around their roosting sites. Although genetic similarity within roosts was higher than that between roosts, the difference was not significant. Observed genetic similarity within roosts was smaller than that reported in the bird species whose roosts work as information centres. On the other hand, the presence of some closely related individuals in the roost and behavioural observations suggest that co‐operation between kin might have occurred, at least in one of the two study sites.
Sex ratio variation among broods of great reed warblers Acrocephalus arundinaceus
Authors: Westerdahl H. , Bensch S. , Hansson B. , Hasselquist D. , Von Schantz T.
Published: November 2003

The sex of 746 great reed warbler fledglings (from 175 broods) was determined by the use of single primer polymerase chain reaction. The reliability of the technique was confirmed as 104 of the fledglings were subsequently recorded as adults of known sex. The overall sex ratio did not differ from unity. Variation in sex ratios between broods was larger than expected from a binomial distribution. Female identity explained some of the variation of brood sex ratio indicating that certain females consistently produced sex ratios that departed from the average value in the population. The theory of sex allocation predicts that parents should adjust the sex ratio of their brood to the relative value of sons and daughters and this may vary in relation to the quality of the parents or to the time of breeding. In the great reed warbler, the proportion of sons was not related to time of breeding, or to any of five female variables. Of five male variables, males with early arrival date tended to produce more daughters. The sex ratio of fledglings that were a result of extra‐pair fertilizations did not differ from that of legitimate fledglings. Hence, there is currently no evidence of that female great reed warblers invest in a higher proportion of sons when mated with attractive males.
Specificity in the symbiotic association of Lotus corniculatus and Rhizobium loti from natural populations
Authors: Lieven‐Antoniou C. A. , Whittam T. S.
Published: November 2003

To test whether Rhizobium loti are coadapted to nodulate local plant genotypes, we competed R. loti strains in a common environment with clonally propagated Lotus corniculatus. Both the plants and bacterial strains were originally collected from natural populations in three localities and the R. loti strains used were distinguishable by enzyme electrophoretic markers and differed in geographical origin relative to host plant origin. The proportions of nodules occupied by symbiont strains varied widely and depended on both host plant and symbiont genotype. Nonrandom nodulation patterns resulted primarily from preferential nodulation of host genotypes by the symbiont strain that had been associated with the host in the natural environment. Symbionts nodulating their original hosts were preferentially found in nodules on adventitious tap roots as opposed to the younger, lateral roots (for one host‐symbiont pair) or in large nodules, independent of location on the root system (for a second host‐symbiont pair). The proportion of nodules occupied by a symbiont on novel host genotypes varied, ranging from nearly random expectation to a significant reduction in the proportion of nodules occupied. The analysis of the bacteria recovered from 994 nodules by multilocus enzyme electrophoresis revealed that 952 (95.8%) nodules were occupied by one of the four inoculant strains and 11 (1.1%) were co‐occupied by two inoculant strains. A total of 31 (3.1%) nodules were occupied by strains that did not match the electrophoretic profiles of the original inoculant strains. Based on the comparison of multilocus profiles for 23 enzyme loci, we concluded that these bacteria were foreign strains and not recombinants of the original inoculant strains. Our findings indicate a strong host genotype by strain interaction underlying the outcome of rhizobial competition for nodulation sites and suggest there are distinct mechanisms leading to differential recognition of compatible host and symbiont genotypes.
Genetic differentiation of the tephritid fly Urophora cardui in Europe as evidence for its biogeographical history
Authors: Eber S. , Brandl R.
Published: November 2003

Gaps in the large‐scale distribution of the tephritid fly Urophora cardui in Europe have been explained as the results of an ongoing re‐immigration from Pleistocene refugia due to a very low dispersal capacity. Following evidence of a much greater dispersal capacity of U. cardui than previously assumed, the pattern of genetic differentiation of 41 populations from 16 European regions was studied using allozyme electrophoresis. In these analyses 18 enzyme systems were scored consistently providing 27 alleles. Allozyme variation indicated high gene flow and low levels of genetic differentiation within and between sampling regions as well as in recently colonized areas. No geographical pattern of heterozygosity or allozyme differentiation could be found matching the previously suggested recent immigration pattern. An observed south‐north gradient in allozyme frequencies was interpreted as a geographical cline due to environmental factors. The results corroborate evidence from more recent studies that U. cardui is a highly mobile species which is likely to have repeatedly colonized some suboptimal European regions since the Pleistocene after retreats during ‘little ice ages’. Patterns resulting from postglacial immigration processes are likely to have been long wiped out through high exchange rates.
Natural hybridization between black spruce and red spruce
Authors: Perron M. , Bousquet J.
Published: November 2003

Using species‐specific random amplified polymorphic DNA (RAPD) markers and morphological characters, natural hybridization between the closely related black spruce Picea mariana (Mill.) B.S.P. and red spruce P. rubens Sarg. was evaluated in natural populations from north‐eastern North America. Sampling included populations from both areas of allopatry and also 14 populations from part of the area of sympatry located in the province of Québec and covering several thousands of square kilometres. Classification results from RAPD species‐specific markers and from a discriminant function based on morphology were compared. Molecular analysis of the allopatric populations indicated a small amount of interspecific gene leakage with no asymmetric directionality to introgression. A high occurrence of hybrid/introgressant individuals was observed within sympatric populations, suggesting weak reproductive isolation. As expected, the detection of such individuals was more efficient using molecular markers than with morphological traits. The hybrid zone appeared extensive with variable species structure and, in some stands, the main component composed of hybrid/introgressant trees. Implications for the genecology and genetic management of these species are discussed.
Molecular markers reveal differentiation among isolates of Coccidioides immitis from California, Arizona and Texas
Authors: Burt A. , Dechairo B. M. , Koenig G. L. , Carter D. A. , White T. J. , Taylor J. W.
Published: November 2003

Coccidioides immitis causes coccidioidomycosis, a fungal disease of both immunocompromised and otherwise healthy people; it is capable of causing large epidemics and the disease is often refractory to chemotherapy. To quantify the magnitude of population differentiation and estimate levels of gene flow in C. immitis, multilocus genotypes were scored for 20–25 clinical isolates from each of Bakersfield (California), Tucson (Arizona), and San Antonio (Texas). The molecular markers used were PCR products with polymorphic restriction endonuclease sites, found and characterized in a previous study of the Tucson population. The data show very highly significant differences in allele frequencies between all three populations, and suggest very low levels of migration between populations. One isolate in the San Antonio sample was an outlier, showing the California‐specific allele at all four of the loci distinguishing the two populations, and subsequent inquiries indicated that the infection had indeed been acquired in California. Thus, genetic information can be used to infer the geographical origin of a fungal infection.