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Livestock trade history, geography, and parasite strains: the mitochondrial genetic structure of Echinococcus granulosus in Argentina.

A sample of 114 isolates of Echinococcus granulosus (Cestoda: Taeniidae) collected from different host species and sites in Argentina has been sequenced for 391 bp from the mitochondrial cytochrome c oxidase subunit I gene to analyze genetic variability and population structure. Nine different haplotypes were identified, 5 of which correspond to already characterized strains. Analysis of molecular variance and nested clade analysis of the distribution of haplotypes among localities within 3 main geographic regions indicate that geographic differentiation accounts for the overall pattern of genetic variability in E. granulosus populations. Significant geographic differentiation is also present when the sheep strain alone is considered. Our results suggest that geographic patterns are not due to actual restricted gene flow between regions but are rather a consequence of past history, probably related to the time and origin of livestock introduction in Argentina.

Analysis of Variance↗

Differential effects of a murine and chimeric mouse/human anti-interleukin-2 receptor antibody on human T-cell proliferation.

The preference for interleukin-2 receptor (IL-2R) expression on activated, compared with resting T lymphocytes makes the IL-2R a promising target for selective immunosuppressive therapy. To increase the potential therapeutic effectiveness of anti-IL-2R monoclonals, a chimeric mouse/human variant was constructed from Ig genes isolated from a murine anti-human IL-2R hybridoma cell line, designated AHT54. AHT54 binds to the same or spatially related epitope as IL-2 on the p55 protein that constitutes the low- and high-affinity forms of IL-2R. Although the murine and chimeric AHT54 antibodies inhibited cell-surface binding of IL-2 to the same extent, the chimeric antibodies containing a human IgG1 constant region had substantially more anti-proliferative activity than their murine IgG1 counterparts. Our results indicated that the human constant region of the chimeric antibodies interacted more efficiently than the murine constant region with effector components present in peripheral blood mononuclear cells (PBMC).

Animals↗

Immunophenotypic and idiotypic characterisation of the leukaemic B-cells from patients with prolymphocytic leukaemia: evidence for a selective expression of immunoglobulin variable region (IGV) gene products.

B-cell prolymphocytic leukaemia (B-PLL) is a rare chronic lymphoproliferative disease characterised by a massive splenomegaly associated with a mild or no lymphadenopathy and a high leukocyte count, mostly representing prolymphocytic features. We have studied membrane expression of certain Ig VK and VH gene products in five patients with B-PLL using a panel of monoclonal anti-subgroup and anti-cross-reactive idiotype (CRI) antibodies. Membrane expression of leukocyte-associated markers has also been investigated. The leukaemic cells from four patients expressed VKIII and VKIIIb subgroup and sub-subgroup kappa light chains. The VKIIIb and VHI-associated CRI identified by the monoclonal antibodies (MoAb) 17-109 and G8 were co-expressed in one patient. No B-cells from the patients expressed the VHIII-associated CRI. The same pattern of CRI expression was observed in a serum paraprotein collected from one of the patients. These results suggest a biased selection for the IG VKIII and VKIIIb light chains in B-PLL.

Aged↗

The voyage of an invasive species across continents: genetic diversity of North American and European Colorado potato beetle populations.

The paradox of successful invading species is that they are likely to be genetically depauperate compared to their source population. This study on Colorado potato beetles is one of the few studies of the genetic consequences of continent-scale invasion in an insect pest. Understanding gene flow, population structure and the potential for rapid evolution in native and invasive populations offers insights both into the dynamics of small populations that become successful invaders and for their management as pests. We used this approach to investigate the invasion of the Colorado potato beetle (Leptinotarsa decemlineata) from North America to Europe. The beetles invaded Europe at the beginning of the 20th century and expanded almost throughout the continent in about 30 years. From the analysis of mitochondrial DNA (mtDNA) and amplified fragment length polymorphism (AFLP) markers, we found the highest genetic diversity in beetle populations from the central United States. The European populations clearly contained only a fraction of the genetic variability observed in North American populations. European populations show a significant reduction at nuclear markers (AFLPs) and are fixed for one mitochondrial haplotype, suggesting a single successful founder event. Despite the high vagility of the species and the reduction of genetic diversity in Europe, we found a similar, high level of population structure and low gene flow among populations on both continents. Founder events during range expansion, agricultural management with crop rotation, and selection due to insecticide applications are most likely the causes partitioning genetic diversity in this species.

Analysis of Variance↗

Chloroplast SSR polymorphisms in the Compositae and the mode of organellar inheritance in Helianthus annuus.

Because organellar genomes are often uniparentally inherited, chloroplast (cp) and mitochondrial (mt) DNA polymorphisms have become the markers of choice for investigating evolutionary issues such as sex-biased dispersal and the directionality of introgression. To the extent that organellar inheritance is strictly maternal, it has also been suggested that the insertion of transgenes into either the chloroplast or mitochondrial genomes would reduce the likelihood of gene escape via pollen flow from crop fields into wild plant populations. In this paper we describe the adaptation of chloroplast simple sequence repeats (cpSSRs) for use in the Compositae. This work resulted in the identification of 12 loci that are variable across the family, seven of which were further shown to be highly polymorphic within sunflower (Helianthus annuus). We then used these markers, along with a novel mtDNA restriction fragment length polymorphism (RFLP), to investigate the mode of organellar inheritance in a series of experimental crosses designed to mimic the initial stages of crop-wild hybridization in sunflower. Although we cannot rule out the possibility of extremely rare paternal transmission, our results provide the best evidence to date of strict maternal organellar inheritance in sunflower, suggesting that organellar gene containment may be a viable strategy in sunflower. Moreover, the portability of these markers suggests that they will provide a ready source of cpDNA polymorphisms for use in evolutionary studies across the Compositae.

Asteraceae↗

Genetic diversity of two African and sixteen South American populations determined on the basis of six hypervariable loci.

A total of 582 individuals (1,164 chromosomes) from two African, eight African-derived South American, five South American Amerindian, and three Brazilian urban populations were studied at four variable number of tandem repeat (VNTR) and two short tandem repeat (STR) hypervariable loci. These two sets of loci did not show distinct allele profiles, which might be expected if different processes promoted their molecular differentiation. The two African groups showed little difference between them, and their intrapopulational variation was similar to those obtained in the African-derived South American communities. The latter showed different degrees of interpopulation variability, despite the fact that they presented almost identical average degrees of non-African admixture. The F(ST) single locus estimates differed in the five sets of populations, probably due to genetic drift, indicating the need to consider population structure in the evaluation of their total variability. A high interpopulational diversity was found among Amerindian populations in relation to Brazilian African-derived isolated communities. This is probably a consequence of the differences in the patterns of gene flow and genetic drift that each of these semi-isolated groups experienced.

Alleles↗

Genetic influence on the shaping of the human T-cell receptor repertoire: quantitative assessment by competitive polymerase chain reaction.

It has been difficult to define the different factors which contribute to the shaping of the human T-cell receptor (TCR) repertoire. In this study, the influence of the polymorphic human leucocyte antigen (HLA) genes and non-HLA genes on the phenotype of the TCRBV segment repertoire was assessed in a population of HLA class I-matched individuals including three pairs of siblings. The gene expression levels of 24 TCRBV families were evaluated in the CD4+ and CD8+ T-cell subsets of unstimulated peripheral blood mononuclear cells (PBMC) by reverse transcription (RT) and a newly developed competitive polymerase chain reaction (cPCR) assay. Titration experiments demonstrated that the RT-cPCR assay was suitable for an accurate quantification of the relative TCRBV segment expression levels. The T-cell repertoires of HLA-identical siblings were found to be more similar than the repertoires of unrelated individuals. On the other hand, there was no difference in the degree of similarity between the TCRBV repertoires of CD4+ T-cells of HLA class II identical or non-identical unrelated individuals. Furthermore, although most of these individuals had identical HLA class I genes and non-identical HLA class II genes, the TCRBV repertoires of the CD4+ T cells exhibited significantly lower variabilities than the repertoires of the CD8+ T cells. The results of the RT-cPCR assay were supported by flow cytometric analysis of the CD4+ and CD8+ T-cell subsets of the same eight individuals employing 10 different TCRBV segment-specific monoclonal antibodies. These observations argue for a predominant role of non-HLA or non-polymorphic HLA determinants for the shaping of the TCRBV repertoire.

Adult↗

Genetic structure and outcrossing rates in Flourensia cernua (Asteraceae) growing at different densities in the South-western Chihuahuan Desert.

BACKGROUNDS AND AIMS: Flourensia cernua is a partially self-incompatible, wind-pollinated shrub that grows in two scrub types of contrasting densities. It was anticipated that differences in plant density would affect the amount of genotype availability, and thus higher outcrossing rates and less genetic differentiation would be found at high-density sites. METHODS: At five high-density sites and at five low-density sites, 11 allozyme loci were analysed in adults. Outcrossing rates were estimated using five allozyme loci sampled from eight families from each scrub type. KEY RESULTS: High levels of genetic variation were found at all sites (ranging from P = 82-100 %, He = 0.33-0.45, and Ho = 0.4-0.59). Heterozygotes were found in excess (FIS = -0.15 +/- 0.06 s.d.), suggesting that natural selection favours heterozygosity, and there was little differentiation between sites (FST = 0.08 +/- 0.02 s.d.). Life history attributes, such as long-lived habit and wide geographic distribution, as well as the presence of a self-incompatibility system may explain these results. Outcrossing rates did not differ from 1.0 in both scrub types, and there was no genetic differentiation between scrub types (FST = -0.01 +/- 0.004 s.d.). CONCLUSIONS: The high rate of outcrossing favoured by partial incompatibility may generate unrestricted gene flow between scrub types and thus may explain the lack of differentiation between them. High heterozygosity could be expected in long-lived plants of arid zones as they confront a variable and stressing environment.

Algorithms↗

Patterns of mtDNA variation in Hawaiian freshwater fishes: the phylogeographic consequences of amphidromy.

MtDNA sequencing was used to assess the phylogeographic structure of four species of Hawaiian freshwater fishes: Lentipes concolor, Stenogobius hawaiiensis, Sicyopterus stimpsoni, and Awaous guamensis. Samples of each species were collected from streams on the northeast side of Kauai, Maui, Molokai, Oahu, and Hawaii. We sequenced segments from both coding and noncoding regions (638-1391 bp) in each species. Sequence analysis uncovered genetic variability in these fishes but no evidence of strong geographic structure among island populations. This result is most readily explained by the fishes' larval marine life stage (amphidromy), which likely facilitates gene flow among island populations. By constraining genetic differentiation among populations, amphidromy may impede speciation in these fishes, possibly explaining why the Hawaiian freshwater fish fauna is depauperate compared to other species-rich Hawaiian faunas. It may also provide them with a kind of evolutionary flexibility atypical of other, more isolated island faunas and allow natural restocking to occur in streams that have been restored to suitable conditions. Comparisons of restriction site and sequence data suggested similar population genetic conclusions for all species except S. stimpsoni, for which the restriction site data is questioned.

Animals↗

Temporal habitat variability and the maintenance of sex in host populations of the pea aphid.

The evolutionary maintenance of sex, despite competition from asexual reproduction, has long intrigued the evolutionary biologists owing to its numerous apparent short-term costs. In aphids, winter climate is expected to determine the maintenance of sexual lineages in the high latitude zones owing to their exclusive ability to produce frost-resistant eggs. However, diverse reproductive modes may coexist at a local scale where climatic influence is counteracted by microgeographical factors. In this study, we tested the influence of local habitat characteristics on regional coexistence of reproductive modes in the pea aphid, Acyrthosiphon pisum. In the laboratory, the induction of sexual morph production of many pea aphid genotypes from the local fields of annual (pea and faba bean) and perennial (alfalfa and red clover) crops in Western France indicated that A. pisum lineages from annual crops had a significantly higher investment in sexual reproduction than A. pisum lineages from the perennial hosts. We propose that temporal habitat variability exerts a selective pressure to maintain the sexual reproduction in A. pisum. The ecological and evolutionary consequences of the association between the mode of reproduction and the host population on gene flow restriction and on ecological specialization are discussed.

Animals↗

Geographic pattern of genetic variation in the European globeflower Trollius europaeus L. (Ranunculaceae) inferred from amplified fragment length polymorphism markers.

The distribution of genetic variation and the phylogenetic relationships between 18 populations of the arctic-alpine plant Trollius europaeus were 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 index FST averaged over loci was high (0.39). The among-population differentiation indicates restricted gene flow, congruent with limited dispersal of specific globeflower's pollinating flies (Chiastocheta spp.). Within-population diversity levels were significantly higher in the Alps (mean Nei's expected heterozygosity HE = 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 among T. europaeus populations. Genetic drift is the major factor acting on the small Pyrenean populations at the southern edge of T. europaeus distribution, 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.

France↗

Pristane-induced arthritis in mice. V. Susceptibility to pristane-induced arthritis is determined by the genetic regulation of the T cell repertoire.

OBJECTIVE: Pristane-induced arthritis (PIA) is an experimental seropositive arthritis that is characterized by serologic and cellular immune abnormalities and is dependent on the presence of a competent CD4+ T cell population. We examined the regulation of PIA by genes of the major histocompatibility complex (MHC) and the Mls-1 loci to determine whether the selection of the T cells that infiltrate arthritic joints is a critical factor in disease susceptibility. METHODS: Genetic regulation of PIA was investigated using F1 hybrid and congenic strain analysis to determine the influence of MHC and Mls-1 genes. The T cell receptor Vbeta phenotypes of lymph node cells and T cells infiltrating arthritic joints were examined with 2-color flow cytometry and reverse transcription-polymerase chain reaction techniques. RESULTS: F1 hybrid offspring from 2 major PIA-susceptible strains (DBA/1 x BALB/c) were resistant to the induction of arthritis because of the interaction between genes of the MHC and the Mls-1 loci, which modified the T cell repertoire. This conclusion was supported by the observed resistance to PIA in BALB/ c-Mls-1a mice, where T cells expressing the Vbeta8.1 and Vbeta6 phenotypes were absent. The receptor phenotype of T cells infiltrating arthritic joints in DBA/1 mice was markedly skewed toward Vbeta8.1 and Vbeta6 compared with the population observed in lymph nodes from either PIA or normal control DBA/1 mice. CONCLUSION: The data support the hypothesis that PIA is a T cell-mediated disease. While pristane causes a polyclonal T cell expansion that gives rise to lymphadenopathy, the development of arthritis in susceptible strains of mice occurs due to the preservation of specific T cell subsets with the capacity to infiltrate synovial joints.

Alleles↗

Immunoglobulin Vlambda light chain gene usage in patients with Sjögren's syndrome.

OBJECTIVE: To determine whether patients with Sjögren's syndrome (SS) have abnormalities in Ig Vlambda and Jlambda gene usage, differences in somatic hypermutation, defects in selection, or indications for perturbations of B cell maturation. METHODS: Individual peripheral B cells from SS patients were analyzed for their Vlambda gene usage by single-cell polymerase chain reaction amplification of genomic DNA and compared with those from normal controls. RESULTS: Molecular differences from controls in Vlambda-Jlambda recombination were identified that were reflected by findings in the nonproductive Vlambda repertoire of the patients, including enhanced rearrangement of Vlambda10A and Jlambda2/3 gene segments. In addition, a number of abnormalities in the productive repertoire were identified, indicating disordered selection. A greater usage of 4 Vlambda genes (2A2, 2B2, 2C, and 7A), representing 56% of all productive Vlambda rearrangements, was observed, suggesting positive selection of these genes. Overutilization of Jlambda2/3 and underutilization of Jlambda7 in both nonproductive and productive Vlambda rearrangements of SS patients compared with controls suggested decreased receptor editing in SS. The mutational frequency did not differ from that in controls, and positive selection of mutations into the productive V gene repertoire was found, similar to that in controls, although mutational targeting toward RGYW/WRCY motifs, typically found in controls, was not found in SS patients. CONCLUSION: Disturbed regulation of B cell maturation with abnormal selection, defects in editing Ig receptors, and abnormal mutational targeting may contribute to the emergence of autoimmunity in SS.

Aged↗

Extensive polymorphism at the Gp63 locus in field isolates of Leishmania peruviana.

Genetic diversity within and between tandemly arrayed copies of the Gp63 gene occurs in laboratory isolates of Leishmania spp., but the extent to which this represents natural genetic diversity has not been assessed. Here, the Gp63 locus is examined in 58 fresh isolates of L. peruviana, and clones derived from them, collected throughout the Peruvian Andes. Extensive polymorphism is observed, both in size of Gp63 containing chromosomes, and for restriction-fragment-length polymorphisms (RFLPs) at the Gp63 locus. All clones within an isolate are identical, including those with two distinct Gp63-hybridising chromosomal-sized pulsed-field gel electrophoresis (PFGE) bands, consistent with diploidy but with size differences in homologous chromosomes. For RFLP analysis, three enzymes were selected to cut within the coding region (PstI), in the intergenic region (SalI) and outside (EcoRI) the Gp63 gene cluster. PstI gave identical banding patterns across all isolates/clones. For EcoRI and SalI, all clones within an isolate were identical, but isolates were polymorphic for fragments at 13 (2-30 kb) and 8 (2.6-8.8 kb) different molecular mass locations generating 19 and 16 distinct RFLP patterns or genotypes for each enzyme, respectively. EcoRI restriction patterns, analysed by PFGE, were consistent with the presence of two clusters of Gp63 genes on each homologous chromosome, one contained within EcoRI fragments large enough to carry from 3 to 10 copies of the Gp63 gene, the second on fragments which could carry 1 or 2 copies of the gene. SalI patterns indicated variable restriction sites within clusters, but not within every intergenic region. A hierarchical analysis of variance of allele frequencies, expressed in terms of Wright's F-statistic, indicated significant barriers to gene flow at all levels, valleys within regions (north/south), villages within valleys, and individuals within villages. This extreme polymorphism at the Gp63 locus of L. peruviana demonstrates the great potential for generation of genetic diversity in parasite populations.

Animals↗

Most peripheral B cells in mice are ligand selected.

Using amplified cDNA and genomic libraries, we have analyzed the VH gene repertoire of pre-B cells and various B cell subsets of conventional mice at the level of VH genes belonging to the J558 VH gene family. The sequence data were evaluated on the basis of a newly established list of 67 J558 VH genes that comprise approximately two-thirds of the J558 VH genes of the murine IgHb haplotype. The results of the analysis demonstrate that VH gene utilization in pre-B cells, although biased to some extent by B cell autonomous VH gene selection, scatters over the whole range of J558 VH genes present in the germline. In contrast, in mature, peripheral B cells comprising long-lived mu + delta high B cells as well as Ly-1 B cells, small overlapping sets of germline VH genes are dominantly expressed. The data indicate that the recruitment of newly generated B cells into the long-lived peripheral B cell pool is mediated through positive selection by internal and/or external antigens. Because of the absence of immunoglobulin class switching and somatic hypermutation, this process is different from the selection of memory B cells in T cell-dependent immune responses.

Animals↗

Mitochondrial DNA variation and population structure of the island endemic Azorean bat (Nyctalus azoreum).

The Azorean bat Nyctalus azoreum is the only endemic mammal native to the remote archipelago of the Azores. It evolved from a continental ancestor related to the Leisler's bat Nyctalus leisleri and is considered threatened because of its restricted and highly fragmented distribution. We studied the genetic variability in 159 individuals from 14 colonies sampled throughout the archipelago. Sequences of the D-loop region revealed moderate but highly structured genetic variability. Half of the 15 distinct haplotypes were restricted to a single island, but the most common was found throughout the archipelago, suggesting a single colonization event followed by limited interisland female gene flow. All N. azoreum haplotypes were closely related and formed a star-like structure typical of expanded populations. The inferred age of demographic expansions was consistent with the arrival of founder animals during the Holocene, well before the first humans inhabited the Azores. Comparisons with a population of N. leisleri from continental Portugal confirmed not only that all N. azoreum lineages were unique to the archipelago, but also that the current levels of genetic diversity were surprisingly high for an insular species. Our data imply that the Azorean bat has a high conservation value. We argue that geographical patterns of genetic structuring indicate the existence of two management units.

Animals↗

Isolation of high-affinity single-chain antibodies against Mycobacterium avium subsp. paratuberculosis surface proteins from sheep with Johne's disease.

Johne's disease, caused by infection with Mycobacterium avium subsp. paratuberculosis, causes significant economic losses to the livestock farming industry. Improved investigative and diagnostic tools-necessary to understand disease processes and to identify subclinical infection-are much sought after. Here, we describe the production of single-chain antibodies with defined specificity for M. avium subsp. paratuberculosis surface proteins. Single-chain antibodies (scFv) were generated from sheep with Johne's disease by cloning heavy-chain and lambda light-chain variable regions and expressing these in fusion with gene III of filamentous phages. Two scFv clones (designated SurfS1.2 and SurfS2.2) were shown to be immunoreactive against M. avium subsp. paratuberculosis surface targets by flow cytometry, and immunoblotting identified specificity for a 34-kDa proteinase-susceptible determinant. Both antibodies were cross-reactive against Mycobacterium avium subsp. avium but nonreactive against Mycobacterium bovis or Mycobacterium phlei cells and were shown to be capable of enriching M. avium subsp. paratuberculosis cells by a factor of approximately 10(6)-fold when employed in magnetic bead separation of mixed Mycobacterium sp. cultures. Further, magnetic bead separation using SurfS1.2 and SurfS2.2 was capable of isolating as few as 10(3) M. avium subsp. paratuberculosis cells from ovine fecal samples, indicating the diagnostic potential of these reagents. Finally, inclusion of SurfS1.2 or SurfS2.2 in in vitro broth culture with M. avium subsp. paratuberculosis indicated that surface binding activity did not impede bacterial growth, although colony clumping was prevented. These results are discussed in terms of the potential use of single-chain phage display monoclonal antibodies as novel diagnostic reagents.

Animals↗

Population differentiation in the Pacific white-sided dolphin Lagenorhynchus obliquidens inferred from mitochondrial DNA and microsatellite analyses.

We investigated genetic diversity and differentiation of the Pacific white-sided dolphin (Lagenorhynchus obliquidens) in Japanese coastal waters and offshore North Pacific by analyzing mitochondrial DNA and nuclear microsatellite variation. A total of 519 bp of the mitochondrial control region was sequenced and five microsatellite locus were genotyped for 59 individuals. A high level of haplotypic diversity (h=96.1%), moderate level of nucleotide diversity (pi=1.65%) and average expected heterozygosity (HE=0.66-0.76) were within an extent of those reported for other odontocetes. Consistent genetic difference between the samples from Japanese coastal Pacific-Sea of Japan and offshore North Pacific was indicated by analyses of molecular variance (AMOVAs) based on mtDNA and microsatellite variations, comparison of genetic variabilities, and geographical distributions of mtDNA haplotypes and microsatellite alleles. This result suggests that Pacific white-sided dolphins in each of the above two areas belong to different populations between which gene flow has been severely restricted. The low genetic diversity and mtDNA genealogy of the population in Japanese coastal waters suggest that it originated from a small population that colonized the Sea of Japan or that experienced population reduction when this Sea was isolated from the North Pacific during a glacial period in the Late Pleistocene.

Animals↗