Les oncogènes NUP98-PHF23 et NUP98-HOXD13 confèrent un potentiel aberrant d’auto-renouvellement aux progéniteurs thymiques.

L’initiation de la leucémogénèse dans la leucémie aigue lymphoblastique (LAL)-T résulte de l’activation aberrante de facteurs de transcription de la lignée lymphocytaire T. Nous démontrons que les gènes de fusion NUP98-PHF23 (NP23) et NUP98-HOXD13 (NHD13) reprogramment les thymocytes normaux en cellules souches pré-leucémiques (CS-préL) possédant un potentiel aberrant d’auto-renouvellement. Basé sur des essais de clonalité performés sur des thymocytes transplantés en série, nous avons découvert que cette population est hiérarchisée similairement aux cellules souches hématopoïétiques normales. Ces CS-préL dévoilent un enrichissement du compartiment de précurseurs thymiques immatures KIT+ où les deux oncogènes, NP23 et NHD13, activent des gènes impliqués dans l’autorenouvellement, incluant Hoxa9, Hoxa10, Lyl1 et Hhex. De plus, l’activité d’autorenouvellement est abrogée par les ARN interférents contre Lyl1 et Hhex, indiquant leur implication fonctionnelle en aval de NP23 et NHD13. Puisque ces gènes sont aussi activés en aval de trois autres oncogènes dans la LAL-T, SCL/TAL1, LMO1 et LMO2, nous concluons que les niveaux d’activation de Lyl1 et Hhex fixent le seuil de reprogrammation des thymocytes normaux en CS-préL. Malgré l'efficacité des traitements de chimiothérapie actuels à diminuer la masse tumorale, les CS-préL sont épargnées, pouvant mener à des rechutes. Nos résultats répondent à ce besoin et proposent de nouvelles avenues permettant de cibler les CS-préL du compartiment de thymocytes immatures dans la LAL-T.

Activated packed bed bioreactor for rapid nitrification in brackish water hatchery systems

A packed bed bioreactor (PBBR) was developed for rapid establishment of nitrification in brackish water hatchery systems in the tropics. The reactors were activated by immobilizing ammonia-oxidizing (AMONPCU- 1) and nitrite-oxidizing (NIONPCU-1) bacterial consortia on polystyrene and low-density polyethylene beads, respectively. Fluorescence in situ hybridization demonstrated the presence of autotrophic nitrifiers belong to Nitrosococcus mobilis, lineage of b ammonia oxidizers and nitrite oxidizer Nitrobacter sp. in the consortia. The activated reactors upon integration to the hatchery system resulted in significant ammonia removal (P\0.01) culminating to its undetectable levels. Consequently, a significantly higher percent survival of larvae was observed in the larval production systems. With spent water the reactors could establish nitrification with high percentage removal of ammonia (78%), nitrite (79%) and BOD (56%) within 7 days of initiation of the process. PBBR is configured in such a way to minimize the energy requirements for continuous operation by limiting the energy inputs to a single stage pumping of water and aeration to the aeration cells. The PBBR shall enable hatchery systems to operate under closed recirculating mode and pave the way for better water management in the aquaculture industry.

Molecular characterization of the nitrifying bacterial consortia employed for the activation of bioreactors used in brackish and marine aquaculture systems

The addition of commercial nitrifying bacterial products has resulted in significant improvement of nitrification efficiency in recirculating aquaculture systems (RAS). We developed two nitrifying bacterial consortia (NBC) from marine and brackish water as start up cultures for immobilizing commercialized nitrifying bioreactors for RAS. In the present study, the community compositions of the NBC were analyzed by universal 16S rRNA gene and bacterial amoA gene sequencing and fluorescence in situ hybridization (FISH). This study demonstrated that both the consortia involved autotrophic nitrifiers, denitrifiers as well as heterotrophs. Abundant taxa of the brackish water heterotrophic bacterial isolates were Paenibacillus and Beijerinckia spp. whereas in the marine consortia they were Flavobacterium, Cytophaga and Gramella species. The bacterial amoA clones were clustered together with high similarity to Nitrosomonas sp. and uncultured beta Proteobacteria. FISH analysis detected ammonia oxidizers belonging to b subclass of proteobacteria and Nitrosospira sp. in both the consortia, and Nitrosococcus mobilis lineage only in the brackish water consortium and the halophilic Nitrosomonas sp. only in the marine consortium. However, nitrite oxidizers, Nitrobacter sp. and phylum Nitrospira were detected in both the consortia. The metabolites from nitrifiers might have been used by heterotrophs as carbon and energy sources making the consortia a stable biofilm.

Molekulare Studien zur Evolution der Myrmekophytie in der Gattung Macaranga (Euphorbiaceae)

Im Rahmen der vorliegenden Arbeit wurde eine detaillierte phylogenetische Analyse der Ameisenpflanzen aus der Gattung Macaranga (Euphorbiaceae) und ihres verwandtschaftlichen Umfelds mit Hilfe von AFLP-Fingerprinting („amplified fragment length polymorphisms“) sowie vergleichender Analyse von mehreren nichtkodierenden Chloroplasten-DNA-Loci vorgenommen. Anhand dieser Untersuchungen sollten im Wesentlichen die folgenden Fragen geklärt werden: (1) Wie stellen sich die Verwandtschaftsverhältnisse zwischen den myrmekophytischen Macaranga-Sektionen Pachystemon, Winklerianae und Pruinosae dar? (2) Wie sind die einzelnen Arten dieser Sektionen miteinander verwandt? (3) Wie oft ist die Lebensweise ”Myrmekophytie” unabhängig voneinander entstanden? Gibt es Hinweise auf Reversionen? (4) Wo liegt genealogisch und auch geographisch der Ursprung der Symbiose zwischen den myrmekophytischen Macaranga-Arten und ihren Partnerameisen? (5) Welche Bedeutung spielen koevolutive Entwicklungen für das Macaranga-Crematogaster-Symbiosesystem? Ist Myrmekophytie im Sinne einer Schlüsselinnovation (Givnish, 1997) als Stimulus für eine adaptive Radiation zu betrachten? (1) Für die AFLP-Analyse wurden 108 Proben aus 43 Macaranga-Arten und 5 unbeschriebenen Morphospezies in die phylogenetische Untersuchung einbezogen. Auf der Basis von 426 Merkmalen wurden Phänogramme sowie Kladogramme rekonstruiert. Zur statistischen Absicherung wurden Bootstrap-Analysen durchgeführt und im Falle der Kladogramme darüber hinaus der „consistency“-Index bestimmt. Die AFLP-Datensätze wurden zusätzlich einer Hauptkomponentenanalyse unterzogen. Mit Hilfe der verschiedenen Untersuchungsmethoden konnten weitgehend übereinstimmende Gruppierungen bzw. evolutive Linien identifiziert werden. Die Sektionen Pachystemon und Pruinosae bilden eine jeweils gut gestützte monophyletische Gruppe. Beide sind vermutlich Schwestergruppen und damit gleich alt. Für die Monophylie der nur aus zwei Arten bestehenden Sektion Winklerianae ergab sich keine Unterstützung. Die Arten der Sektion Pruinosae sind im AFLP-Baum gut aufgelöst. Die nicht myrmekophytische M. gigantea sitzt dabei an der Basis und ist Schwestergruppe zu den myrmekophytischen Arten. Innerhalb der Sektion Pachystemon wurden mit Hilfe der AFLP-Analyse vier gut gestützte Gruppen identifiziert. Für die puncticulata-Gruppe konnte hier erstmals auf molekularer Ebene eine Zugehörigkeit zur Sekt. Pachystemon nachgewiesen werden. Der von Davies (2001) vorgenommene Ausschluss von M. recurvata aus der Sekt. Pachystemon konnte bestätigt werden. Die Verwandtschaftsbeziehungen einzelner Arten zueinander sind in den AFLP-Bäumen nicht aufgelöst. (2) Für die vergleichende Chloroplasten-Sequenzierung wurden nach Maßgabe der Sequenzvariabilität in Testsequenzierungen die Bereiche atpB-rbcL und psbI-trnS für die phylogenetische Untersuchung ausgewählt. Für die Chloroplasten-Phylogenie wurden für jeden Locus mehr als 100 Sequenzen analysiert. Neben 29 Pachystemon-Arten inkl. vier unbekannter Morphospezies, acht Pruinosae-Arten inkl. eines möglichen Hybriden und den beiden Arten der Sekt. Winklerianae wurden 22 weitere Macaranga- und 10 Mallotus-Arten in die Untersuchung einbezogen. Zwischen den südostasiatischen Arten bestanden nur geringe Sequenzunterschiede. Maximum-Parsimonie-Kladogramme wurden rekonstruiert und die Sequenzen der beiden Loci wurden sowohl einzeln, als auch kombiniert ausgewertet. Indels wurden kodiert und als separate Merkmalsmatrix an die Sequenzdaten angehangen. Innerhalb von Macaranga konnten nur wenige abgesicherte Gruppen identifiziert werden. Deutlich war die Zusammengehörigkeit der afrikanischen Arten und ihr Entstehung aus den südostasiatischen Arten. Die von Davies (2001) der Sektion Pruinosae zugeordnete M. siamensis steht deutlich außerhalb dieser Sektion. Die Arten der Sektionen Pruinosae, Pachystemon und Winklerianae bilden keine statistisch gesicherten monophyletischen Gruppen. Während der Pilotstudien stellte sich heraus, dass die Chloroplastensequenzen nahe verwandter Arten der Sektion Pachystemon weniger nach den Artgrenzen, sondern vielmehr nach geographischen Kriterien gruppierten. (3) Es wurde daher zusätzlich eine phylogeographische Analyse der Chloroplasten-Sequenzen auf der Basis eines Parsimonie-Netzwerks durchgeführt. Neben dem atpB-rbcL-Spacer und einer Teilsequenze des psbI-trnS-Locus (ccmp2) wurde dafür zusätzlich der ccmp6-Locus (ein Abschnitt des ycf3-Introns) sequenziert. Die phylogeographische Untersuchung wurde mit 144 Proben aus 41 Macaranga-Arten durchgeführt. Darin enthalten waren 29 Arten (inkl. vier Morphospezies) mit 112 Proben der Sektion Pachystemon, sieben 7 Arten (inkl. eines potentiellen Hybriden) mit 22 Proben der Sekt. Pruinosae und zwei Arten mit 5 Proben der Sekt. Winklerianae. Das voll aufgelöste statistische Parsimonie-Netzwerk umfasste 88 Haplotypen. Die Sektionen Pachystemon und Pruinosae bilden jeweils eine monophyletische Gruppe. Das geographische Arrangement der Haplotypen unabhängig von der Artzugehörigkeit könnte durch Introgression und/oder „lineage sorting“ bedingt sein. Mit Hilfe der im Rahmen dieser Arbeit gewonnenen Ergebnisse kann man davon ausgehen, dass eine enge Ameisen-Pflanzen-Symbiose innerhalb der Gattung Macaranga mindestens drei-, möglicherweise viermal unabhängig voneinander entstanden ist Eine Reversion hat mindestens einmal, möglicherweise häufiger in der bancana-Gruppe stattgefunden. Ob sich die Symbiose dabei in Westmalaysia oder in Borneo entwickelt hat, kann man nicht sicher sagen; Ob und inwieweit die große Artenzahl in der bancana-Gruppe als eine Folge der Myrmekophytie anzusehen ist, bleibt zunächst offen. Wesentliche Teile der vorliegenden Arbeit liegen bereits in publizierter Form vor (AFLP-Analyse: Bänfer et al. 2004; Chloroplasten-Analyse: Vogel et al. 2003; Bänfer et al. 2006).

Systematics and evolution of the genus Deuterocohnia Mez (Bromeliaceae)

The present study investigates the systematics and evolution of the Neotropical genus Deuterocohnia Mez (Bromeliaceae). It provides a comprehensive taxonomic revision as well as phylogenetic analyses based on chloroplast and nuclear DNA sequences and presents a hypothesis on the evolution of the genus. A broad morphological, anatomical, biogeographical and ecological overview of the genus is given in the first part of the study. For morphological character assessment more than 700 herbarium specimens from 39 herbaria as well as living plant material in the field and in the living collections of botanical gardens were carefully examined. The arid habitats, in which the species of Deuterocohnia grow, are reflected by the morphological and anatomical characters of the species. Important characters for species delimitation were identified, like the length of the inflorescence, the branching order, the density of flowers on partial inflorescences, the relation of the length of the primary bracts to that of the partial inflorescence, the sizes of floral bracts, sepals and petals, flower colour, the presence or absence of a pedicel, the curvature of the stamina and the petals during anthesis. After scrutinizing the nomenclatural history of the taxa belonging to Deuterocohnia – including the 1992 syonymized genus Abromeitiella – 17 species, 4 subspecies and 4 varieties are accepted in the present revision. Taxonomic changes were made in the following cases: (I) New combinations: A. abstrusa (A. Cast.) N. Schütz is re-established – as defined by Castellanos (1931) – and transfered to D. abstrusa; D. brevifolia (Griseb.) M.A. Spencer & L.B. Sm. includes accessions of the former D. lorentziana (Mez) M.A. Spencer & L.B. Sm., which are not assigned to D. abstrusa; D. bracteosa W. Till is synonymized to D. strobilifera Mez; D. meziana Kuntze ex Mez var. carmineo-viridiflora Rauh is classified as a subspecies of D. meziana (ssp. carmineo-viridiflora (Rauh) N. Schütz); D. pedicellata W. Till is classified as a subspecies of D. meziana (ssp. pedicellata (W. Till) N. Schütz); D. scapigera (Rauh & L. Hrom.) M.A. Spencer & L.B. Sm ssp. sanctae-crucis R. Vásquez & Ibisch is classified as a species (D. sanctae-crucis (R. Vásquez & Ibisch) N. Schütz); (II) New taxa: a new subspecies of D. meziana Kuntze ex Mez is established; a new variety of D. scapigera is established; (the new taxa will be validly published elsewhere); (III) New type: an epitype for D. longipetala was chosen. All other species were kept according to Spencer and Smith (1992) or – in the case of more recently described species – according to the protologue. Beside the nomenclatural notes and the detailed descriptions, information on distribution, habitat and ecology, etymology and taxonomic delimitation is provided for the genus and for each of its species. An key was constructed for the identification of currently accepted species, subspecies and varieties. The key is based on easily detectable morphological characters. The former synonymization of the genus Abromeitiella into Deuterocohnia (Spencer and Smith 1992) is re-evalutated in the present study. Morphological as well as molecular investigations revealed Deuterocohnia incl. Abromeitiella as being monophyletic, with some indications that a monophyletic Abromeitiella lineage arose from within Deuterocohnia. Thus the union of both genera is confirmed. The second part of the present thesis describes and discusses the molecular phylogenies and networks. Molecular analyses of three chloroplast intergenic spacers (rpl32-trnL, rps16-trnK, trnS-ycf3) were conducted with a sample set of 119 taxa. This set included 103 Deuterocohnia accessions from all 17 described species of the genus and 16 outgroup taxa from the remainder of Pitcairnioideae s.str. (Dyckia (8 sp.), Encholirium (2 sp.), Fosterella (4 sp.) and Pitcairnia (2 sp.)). With its high sampling density, the present investigation by far represents the most comprehensive molecular study of Deuterocohnia up till now. All data sets were analyzed separately as well as in combination, and various optimality criteria for phylogenetic tree construction were applied (Maximum Parsimony, Maximum Likelihood, Bayesian inferences and the distance method Neighbour Joining). Congruent topologies were generally obtained with different algorithms and optimality criteria, but individual clades received different degrees of statistical support in some analyses. The rps16-trnK locus was the most informative among the three spacer regions examined. The results of the chloroplast DNA analyses revealed a highly supported paraphyly of Deuterocohnia. Thus, the cpDNA trees divide the genus into two subclades (A and B), of which Deuterocohnia subclade B is sister to the included Dyckia and Encholirium accessions, and both together are sister to Deuterocohnia subclade A. To further examine the relationship between Deuterocohnia and Dyckia/Encholirium at the generic level, two nuclear low copy markers (PRK exon2-5 and PHYC exon1) were analysed with a reduced taxon set. This set included 22 Deuterocohnia accessions (including members of both cpDNA subclades), 2 Dyckia, 2 Encholirium and 2 Fosterella species. Phylogenetic trees were constructed as described above, and for comparison the same reduced taxon set was also analysed at the three cpDNA data loci. In contrast to the cpDNA results, the nuclear DNA data strongly supported the monophyly of Deuterocohnia, which takes a sister position to a clade of Dyckia and Encholirium samples. As morphology as well as nuclear DNA data generated in the present study and in a former AFLP analysis (Horres 2003) all corroborate the monophyly of Deuterocohnia, the apparent paraphyly displayed in cpDNA analyses is interpreted to be the consequence of a chloroplast capture event. This involves the introgression of the chloroplast genome from the common ancestor of the Dyckia/ Encholirium lineage into the ancestor of Deuterocohnia subclade B species. The chloroplast haplotypes are not species-specific in Deuterocohnia. Thus, one haplotype was sometimes shared by several species, where the same species may harbour different haplotypes. The arrangement of haplotypes followed geographical patterns rather than taxonomic boundaries, which may indicate some residual gene flow among populations from different Deuteroccohnia species. Phenotypic species coherence on the background of ongoing gene flow may then be maintained by sets of co-adapted alleles, as was suggested by the porous genome concept (Wu 2001, Palma-Silva et al. 2011). The results of the present study suggest the following scenario for the evolution of Deuterocohnia and its species. Deuterocohnia longipetala may be envisaged as a representative of the ancestral state within the genus. This is supported by (1) the wide distribution of this species; (2) the overlap in distribution area with species of Dyckia; (3) the laxly flowered inflorescences, which are also typical for Dyckia; (4) the yellow petals with a greenish tip, present in most other Deuterocohnia species. The following six extant lineages within Deuterocohnia might have independently been derived from this ancestral state with a few changes each: (I) D. meziana, D. brevispicata and D. seramisiana (Bolivia, lowland to montane areas, mostly reddish-greenish coloured, very laxly to very densely flowered); (II) D. strobilifera (Bolivia, high Andean mountains, yellow flowers, densely flowered); (III) D. glandulosa (Bolivia, montane areas, yellow-greenish flowers, densely flowered); (IV) D. haumanii, D. schreiteri, D. digitata, and D. chrysantha (Argentina, Chile, E Andean mountains and Atacama desert, yellow-greenish flowers, densely flowered); (V) D. recurvipetala (Argentina, foothills of the Andes, recurved yellow flowers, laxly flowered); (VI) D. gableana, D. scapigera, D. sanctae-crucis, D. abstrusa, D. brevifolia, D. lotteae (former Abromeitiella species, Bolivia, Argentina, higher Andean mountains, greenish-yellow flowers, inflorescence usually simple). Originating from the lower montane Andean regions, at least four lineages of the genus (I, II, IV, VI) adapted in part to higher altitudes by developing densely flowered partial inflorescences, shorter flowers and – in at least three lineages (II, IV, VI) – smaller rosettes, whereas species spreading into the lowlands (I, V) developed larger plants, laxly flowered, amply branched inflorescences and in part larger flowers (I).

Análisis del significado de conflicto en un grupo de jóvenes de la Universidad del Rosario desde el Modelo Relacional Simbólico

Esta investigación se propuso definir y reflexionar acerca de las concepciones de conflicto, reportadas por un grupo de jóvenes de la Universidad del Rosario, identificando si se consideraba el vínculo y las categorías simbólicas del Modelo Relacional Simbólico: confianza, justicia y esperanza. Se realizaron entrevistas semiestructuradas, que se analizaron en su contenido. Se logró establecer que la concepción predominante acerca del conflicto fue la de algo que no debe suceder o debe evitarse, entendido en general como una confrontación o un choque. Ninguno consideró el vínculo como un elemento fundamental o vital del conflicto. Se pudo deducir que el conflicto se da en las relaciones entre personas, pero la importancia del vínculo en términos de encuentro de generaciones, estirpes y géneros, como un espacio de confianza, esperanza y justicia en las que se entregan dones y se tienen deudas; no está presente de ninguna forma en las definiciones de conflicto. Estos aspectos muestran la necesidad de acoger los fundamentos vinculares y simbólicos del modelo, como riquezas y fortalezas que necesitan ser introducidas en la cotidianidad de la población Colombiana, en la construcción de una cultura de la paz. Esto, a través de la educación a todo nivel y en particular a corto plazo; con los estudiantes de primeros semestres universitarios, por el papel que como profesionales tendrán de líderes sociales, empresariales, espirituales, políticos y laborales; y porque aún se encuentran en una etapa propicia para la formación integral que estimule el fortalecimiento del vínculo como un eje vital de las relaciones y en el que sus categorías simbólicas (esperanza, justicia y confianza) se vuelvan parte conciente en las relaciones personales, familiares y sociales. Igualmente se hace indispensable transformar la visión de conflicto; de forma que deje de ser negativo, no deseable y se convierta en parte de la vida y en una oportunidad para crecer, conocer, generar y regenerar vínculos. Finalmente se propone la creación de una línea de investigación que extienda el estudio a otras poblaciones de jóvenes de distinto nivel socioeconómico y cultural, incluidos los afectados directa e indirectamente por la violencia en Colombia.

Plasmodium vivax lineages: geographical distribution, tandem repeat polymorphism, and phylogenetic relationship

Background: Multi-drug resistance and severe/ complicated cases are the emerging phenotypes of vivax malaria, which may deteriorate current anti-malarial control measures. The emergence of these phenotypes could be associated with either of the two Plasmodium vivax lineages. The two lineages had been categorized as Old World and New World, based on geographical sub-division and genetic and phenotypical markers. This study revisited the lineage hypothesis of P. vivax by typing the distribution of lineages among global isolates and evaluated their genetic relatedness using a panel of new mini-satellite markers. Methods: 18S SSU rRNA S-type gene was amplified from 420 Plasmodium vivax field isolates collected from different geographical regions of India, Thailand and Colombia as well as four strains each of P. vivax originating from Nicaragua, Panama, Thailand (Pak Chang), and Vietnam (ONG). A mini-satellite marker panel was then developed to understand the population genetic parameters and tested on a sample subset of both lineages. Results: 18S SSU rRNA S-type gene typing revealed the distribution of both lineages (Old World and New World) in all geographical regions. However, distribution of Plasmodium vivax lineages was highly variable in every geographical region. The lack of geographical sub-division between lineages suggests that both lineages are globally distributed. Ten mini-satellites were scanned from the P. vivax genome sequence; these tandem repeats were located in eight of the chromosomes. Mini-satellites revealed substantial allelic diversity (7-21, AE = 14.6 +/- 2.0) and heterozygosity (He = 0.697-0.924, AE = 0.857 +/- 0.033) per locus. Mini-satellite comparison between the two lineages revealed high but similar pattern of genetic diversity, allele frequency, and high degree of allele sharing. A Neighbour-Joining phylogenetic tree derived from genetic distance data obtained from ten mini-satellites also placed both lineages together in every cluster. Conclusions: The global lineage distribution, lack of genetic distance, similar pattern of genetic diversity, and allele sharing strongly suggested that both lineages are a single species and thus new emerging phenotypes associated with vivax malaria could not be clearly classified as belonging to a particular lineage on basis of their geographical origin.

Diversity, dynamics and activity of mesophilic Archaea in stratified feshwater lakes. Implications in biogeochemical cycles

Aquesta tesi doctoral va estudiar la diversitat (riquesa i abundància), la distribució i la dinàmica de les comunitats planctòniques d'Archaea presents a diferents llacs estratificats temperats d'aigua dolça per aportar evidencies sobre la seva distribució i la seva possible activitat en aquests ecosistemes en relació als cicles biogeoquímics presents en els mateixos. Es varen estudiar dos estanyols d'origen càrstic (l'Estanyol del Vilar durant cinc anys consecutius (2001-2005) i l'Estanyol de Can Coromina) i un llac d'origen volcànic (Llac Kivu) analitzant, per una banda, la seva comunitat planctònica d'Archaea mitjançant una aproximació molecular i, per una altra, la seva possible activitat en aquests ambients (p.e., la nitrificació i la fixació de carboni). Per contextualitzar els resultats, es va realitzar un anàlisi in silico dels patrons de distribució global dels Archaea mesòfils mitjançant un anàlisi a nivell de llinatge combinant seqüències del gen 16S rRNA amb diferents eines estadístiques i d'ecologia general.

Estudio de las variaciones espaciales y temporales de la diversidad genética de la trucha común, Salmo trutta, en ríos de la Península Ibérica

Se ha analizado las causas de la distribución espacial de la variabilidad genética del ADN mitocondrial en poblaciones de trucha común de la cuenca del Duero y de los Pirineos Orientales. En total se han analizado de novo 49 localidades, 13 en la cuenca del río Duero y 36 en los principales ríos del Pirineo oriental. Además se analizaron las fluctuaciones temporales en 14 de las localidades del Pirineo Oriental. Estudios previos indican un marcado contraste de los patrones de diversidad entre ambos territorios. En la cuenca del río Duero los análisis confirmaron la presencia de los dos linajes matriarcales descritos previamente, el linaje Atlántico (AT) y el linaje Duero (DU). Los análisis de la varianza molecular (AMOVA) siguiendo una jerarquía hidrográfica sugirieron una alta estructuración de las poblaciones coincidente con los patrones ictiológicos observados en la cuenca. El linaje DU parece haber estado presente permanentemente en la cuenca interior del Duero, mientras que las zonas más próximas a la desembocadura han padecido diversas colonizaciones de trucha del linaje AT, que reflejarían los cambios climáticos ocurridos en el Cuaternario. Se ha detectado una discrepancia en el límite entre ambos grupos definidos por genes nucleares (alozimas) y el ADN mitocondrial. Estas discrepancias pueden ser debidas a un efecto más severo de la deriva genética en el ADN mitocondrial que en los marcadores nucleares. Sin embargo, en este trabajo se han observado evidencias a favor de selección en el ADN mitocondrial del linaje DU que también explicaría estas discrepancias. El análisis más exhaustivo en las cuencas de los Pirineos orientales, permitió detectar nuevos haplotipos mitocondriales de los linajes Adriático (AD) y Mediterráneo (ME). En esta región, los AMOVAs confirmaron que las diferencias entre poblaciones dentro de río son más importantes que las diferencias entre ríos. No obstante se observó un patrón de aislamiento por distancia en toda la zona, reflejo de la estructuración de las poblaciones en la cuenca del río Ebro. Además, aunque los AMOVAs mostraron que el componente temporal de la variación es inferior al espacial, las fluctuaciones temporales en la comparación matriarcal de las poblaciones resultaron estadísticamente significativas. Estas fluctuaciones están asociadas tanto a la deriva genética como a procesos de flujo génico entre poblaciones próximas. Dentro de las cuencas, los componentes de diferenciación entre afluentes son, en general, superiores a los obtenidos dentro de cada afluente, patrón que parece estar extendido en la trucha común. Los estudios a escala microgeográfica en la Noguera Vallferrera y Noguera Cardós (afluentes del Noguera Pallaresa) reprodujeron este patrón de diferenciación. Los tamaños efectivos y la tasa de migración entre ambos ríos fueron similares a los descritos en poblaciones noratlánticas. Los tamaños efectivos de las hembras (Nef), calculados a partir del ADN mitocondrial fueron menos de la mitad del tamaño efectivo total tanto en la Noguera Vallferrera como en el resto de localidades pirenaicas estudiadas. Estos bajos tamaños efectivos de las hembras serían también responsables de las fluctuaciones temporales observadas. Los ejemplares repoblados parecen hibridar poco con los nativos, pero su presencia podría intensificar indirectamente los procesos de deriva genética y complicar la conservación de los patrimonios genéticos nativos. Con la salvedad de la existencia de selección que favorece a los haplotipos del linaje DU, los procesos poblacionales que regulan la distribución de la variabilidad genética en la cuenca del Duero y en los Pirineos Orientales podrían ser parecidos y caracterizados por la existencia de múltiples demes interconectados a lo largo del curso fluvial.

Filogeografia de la truita comuna (Salmo trutta) basada en la diversitat molecular del DNA mitocondrial

Les anàlisis realitzades en cent deu poblacions de truita comuna (Salmo trutta) que abarquen el seu rang natural de distribució indiquen que el patró filogenètic es relaciona amb les tres grans vessants on es troba distribuïda l'espècie: ponto-càspia, atlàntica i mediterrània. Aquesta diferenciació estaria associada a l'aïllament de les vessants durant el Quaternari. L'origen de l'espècie es relaciona amb la vessant ponto-càspia, d'acord amb els models biogeogràfics que postulen l'origen asiàtic de la ictiofauna europea. S'ha detectat també un segon nivell de divergència dins de cada vessant que dóna com a resultat l'existència de sis llinatges evolutius: Atlàntic i Duero a la vessant atlàntica, els llinatges Adriàtic, Mediterrani i Marmoratus als rius mediterranis, i el llinatge Danubi a la zona ponto-càspia. Les glaciacions del Pleistocè han modificat profundament el rang de distribució de la truita comuna, especialment a la vessant atlàntica, on s'han proposat quatre grans refugis glacials: a l'est de la capa de gel, a Europa central, a l'entorn del canal de la Mànega i a l'entorn del golf de Biscaia; tot i que només els tres primers haurien participat en la recolonització del nord d'Europa al final de l'última glaciació. El quart refugi, que inclou el sud de França i el Cantàbric hauria estat l'origen de l'expansió cap al sud durant el Pleistocè Superior d'un grup de poblacions distribuïdes actualment a la vessant atlàntica ibèrica, i també hauria servit de base per a l'expansió cap al nord d'altres grups de truita durant interglacials anteriors. A la vessant atlàntica de la peninsula Ibèrica, l'estructura poblacional es troba associada a la xarxa hidrogràfica i es determinen fins a cinc unitats poblacionals: les truites dels rius Cantàbrics, les del Miño, les del Duero, les del Tajo i les del Guadalquivir. Les poblacions del Guadalquivir pertanyerien a un grup d'influència mediterrània. Els marcadors d'al·lozims i de DNA mitocondrial es troben fortament correlacionats en aquesta vessant, on apunten cap als mateixos grups de poblacions. Per contra, els rius de la vessant mediterrània haurien estat colonitzats pels llinatges Adriàtic i Mediterrani i s'hauria produït una intensa intergradació secundària entre aquests llinatges durant els períodes glacials a partir de l'expansió de les poblacions retingudes a les capçaleres durant els interglacials. Els grups de hibridació, l'aïllament i la deriva en el període interglacial fa que els grups de poblacions identificats pels marcadors d'al·lozims i de DNA mitocondrial no coincideixin.

Evaluaton of a genetic marker within the organ of Corti of transgenic mice

This paper reviews a study to evaluate a genetic marker within the organ of Corti in transgenic mice for potential use in cell lineage studies.

NG2 cells differentiate into astrocytes in cerebellar slices

NG2-glia are an abundant population of glial cells that have been considered by many to be oligodendrocyte progenitor cells (OPCs). However, growing evidence suggests that NG2-glia may also be capable of differentiating into astrocytes and neurons under certain conditions. Here, we have examined NG2-glia in cerebellar slices, using transgenic mice in which the astroglial marker glial specific protein (GFAP) drives expression of the reporter gene enhanced green fluorescent protein (EGFP). Immunolabelling for NG2 shows that NG2-glia and GFAP-EGFP astroglia are separate populations in most areas of the brain, although a substantial population of NG2-glia in the pons also express the GFAP-EGFP reporter. In the cerebellum, NG2-glia did not express EGFP, either at postnatal day (P)12 or P29-30. We developed an organotypic culture of P12 cerebellar slices that maintain cytoarchitectural integrity of Purkinje neurons and Bergmann glia. In these cultures, BrdU labelling indicates that the majority of NG2-glia enter the cell cycle within 2 days in vitro (DIV), suggesting that NG2-glia undergo a [`]reactive' response in cerebellar cultures. After 2 DIV NG2-glia began to express the astroglial reporter EGFP and in some cases the respective GFAP protein. However, NG2-glia did not acquire phenotypic markers of neural stem cells or neurons. The results suggest that NG2-glia are not lineage restricted OPCs and are a potential source of astrocytes in the cerebellum.

Nitrosospira spp. can produce nitrous oxide via a nitrifier denitrification pathway

Nitrous oxide (N2O) emission from soils is a major contributor to the atmospheric loading of this potent greenhouse gas. It is thought that autotrophic ammonia oxidizing bacteria (AOB) are a significant source of soil-derived N2O and a denitrification pathway (i.e. reduction of NO2- to NO and N2O), so-called nitrifier denitrification, has been demonstrated as a N2O production mechanism in Nitrosomonas europaea. It is thought that Nitrosospira spp. are the dominant AOB in soil, but little information is available on their ability to produce N2O or on the existence of a nitrifier denitrification pathway in this lineage. This study aims to characterize N2O production and nitrifier denitrification in seven strains of AOB representative of clusters 0, 2 and 3 in the cultured Nitrosospira lineage. Nitrosomonas europaea ATCC 19718 and ATCC 25978 were analysed for comparison. The aerobically incubated test strains produced significant (P < 0.001) amounts of N2O and total N2O production rates ranged from 2.0 amol cell(-1) h(-1), in Nitrosospira tenuis strain NV12, to 58.0 amol cell(-1) h(-1), in N. europaea ATCC 19718. Nitrosomonas europaea ATCC 19718 was atypical in that it produced four times more N2O than the next highest producing strain. All AOB tested were able to carry out nitrifier denitrification under aerobic conditions, as determined by production of N-15-N2O from applied N-15-NO2-. Up to 13.5% of the N2O produced was derived from the exogenously applied N-15-NO2-. The results suggest that nitrifier denitrification could be a universal trait in the betaproteobacterial AOB and its potential ecological significance is discussed.

Molecular genetic differentiation in earthworms inhabiting a heterogeneous Pb-polluted landscape

A Pb-mine site situated on acidic soil, but comprising of Ca-enriched islands around derelict buildings was used to study the spatial pattern of genetic diversity in Lumbricus rubellus. Two distinct genetic lineages ('A' and 'B'), differentiated at both the mitochondrial (mtDNA COII) and nuclear level (AFLPs) were revealed with a mean inter-lineage mtDNA sequence divergence of approximately 13%, indicative of a cryptic species complex. AFLP analysis indicates that lineage A individuals within one central 'ecological island' site are uniquely clustered, with little genetic overlap with lineage A individuals at the two peripheral sites. FTIR microspectroscopy of Pb-sequestering chloragocytes revealed different phosphate profiles in residents of adjacent acidic and calcareous islands. Bioinformatics found over-representation of Ca pathway genes in ESTPb libraries. Subsequent sequencing of a Ca-transport gene, SERCA, revealed mutations in the protein's cytosolic domain. We recommend the mandatory genotyping of all individuals prior to field-based ecotoxicological assays, particularly those using discriminating genomic technologies.

Metal bioaccumulation and cellular fractionation in an epigeic earthworm (Lumbricus rubellus): the interactive influences of population exposure histories, site-specific geochemistry and mitochondrial genotype

Subcellular fractionation techniques were used to describe temporal changes (at intervals from T0 to T70 days) in the Pb, Zn and P partitioning profiles of Lumbricus rubellus populations from one calcareous (MDH) and one acidic (MCS) geographically isolated Pb/Zn-mine sites and one reference site (CPF). MDH and MCS individuals were laboratory maintained on their native field soils; CPF worms were exposed to both MDH and MCS soils. Site-specific differences in metal partitioning were found: notably, the putatively metal-adapted populations, MDH and MCS, preferentially partitioned higher proportions of their accumulated tissue metal burdens into insoluble CaPO4-rich organelles compared with naive counterparts, CPF. Thus, it is plausible that efficient metal immobilization is a phenotypic trait characterising metal tolerant ecotypes. Mitochondrial cytochrome oxidase II (COII) genotyping revealed that the populations indigenous to mine and reference soils belong to distinct genetic lineages, differentiated by 13%, with 7 haplotypes within the reference site lineage but fewer (3 and 4, respectively) in the lineage common to the two mine sites. Collectively, these observations raise the possibility that site-related genotype differences could influence the toxico-availability of metals and, thus, represent a potential confounding variable in field-based eco-toxicological assessments.