The EmsB tandemly repeated multilocus microsatellite: a new tool to investigate genetic diversity of Echinococcus granulosus sensu lato

Cystic echinococcosis (CE) is a widespread and severe zoonotic disease caused by infection with the larval stage of the eucestode Echinococcus granulosus sensu lato. The polymorphism exhibited by nuclear and mitochondrial markers conventionally used for the genotyping of different parasite species and strains does not reach the level necessary for the identification of genetic variants linked to restricted geographical areas. EmsB is a tandemly repeated multilocus microsatellite that proved its usefulness for the study of genetic polymorphisms within the species E. multilocularis, the causative agent of alveolar echinococcosis. In the present study, EmsB was used to characterize E. granulosus sensu lato samples collected from different host species (sheep, cattle, dromedaries, dogs, and human patients) originating from six different countries (Algeria, Mauritania, Romania, Serbia, Brazil, and the People's Republic of China). The conventional mitochondrial cox1 and nad1 markers identified genotypes G1, G3, G5, G6, and G7, which are clustered into three groups corresponding to the species E. granulosus sensu stricto, E. ortleppi, and E. canadensis. With the same samples, EmsB provided a higher degree of genetic discrimination and identified variations that correlated with the relatively small-scale geographic origins of the samples. In addition, one of the Brazilian single hydatid cysts presented a hybrid genotypic profile that suggested genetic exchanges between E. granulosus sensu stricto and E. ortleppi. In summary, the EmsB microsatellite exhibits an interesting potential for the elaboration of a detailed map of the distribution of genetic variants and therefore for the determination and tracking of the source of CE.

A multiplex PCR for the simultaneous detection and genotyping of the Echinococcus granulosus complex.

Echinococcus granulosus is characterized by high intra-specific variability (genotypes G1-G10) and according to the new molecular phylogeny of the genus Echinococcus, the E. granulosus complex has been divided into E. granulosus sensu stricto (G1-G3), E. equinus (G4), E. ortleppi (G5), and E. canadensis (G6-G10). The molecular characterization of E. granulosus isolates is fundamental to understand the spatio-temporal epidemiology of this complex in many endemic areas with the simultaneous occurrence of different Echinococcus species and genotypes. To simplify the genotyping of the E. granulosus complex we developed a single-tube multiplex PCR (mPCR) allowing three levels of discrimination: (i) Echinococcus genus, (ii) E. granulosus complex in common, and (iii) the specific genotype within the E. granulosus complex. The methodology was established with known DNA samples of the different strains/genotypes, confirmed on 42 already genotyped samples (Spain: 22 and Bulgaria: 20) and then successfully applied on 153 unknown samples (Tunisia: 114, Algeria: 26 and Argentina: 13). The sensitivity threshold of the mPCR was found to be 5 ng Echinoccoccus DNA in a mixture of up to 1 µg of foreign DNA and the specificity was 100% when template DNA from closely related members of the genus Taenia was used. Additionally to DNA samples, the mPCR can be carried out directly on boiled hydatid fluid or on alkaline-lysed frozen or fixed protoscoleces, thus avoiding classical DNA extractions. However, when using Echinococcus eggs obtained from fecal samples of infected dogs, the sensitivity of the mPCR was low (<40%). Thus, except for copro analysis, the mPCR described here has a high potential for a worldwide application in large-scale molecular epidemiological studies on the Echinococcus genus.

Distribution and Efficiency of Bacteriolysis in the Gut of Arenicola Marina and Three Additional Deposit Feeders

A simple technique was developed to measure the bacteriolytic activities of the digestive fluids of the deposit-feeding polychaete Arenicola marina. Lysis of a cultured environmental isolate, incubated with extracts of gut luminal contents, was monitored spectrophotometrically. Concurrent direct counts were used to verify cell lysis. The ability of extracts from 8 longitudinal sections of the gut to lyse the bacterium was monitored. The digestive ceca, anterior stomach, and posterior stomach regions exhibited high lytic activities, whereas bacteriolytic activities in all other regions of the gut were negligible. Similarly, extracts of surface sediments and fecal castings showed negligible lytic capabilities. The sharply limited distribution of lytic activity implicates the ceca as the source of bacteriolytic agent and suggests a true plug-flow system, with little axial mixing. Questions regarding the fate of lytic agents, which disappear abruptly posterior to the stomach, remain unanswered. Localization of lysis in the gut coupled with estimates of gut residence time permit the calculation that ingested bacteria are exposed to strong lytic activity for approximately 20 min. Incubation of in situ sediment samples with gut fluids corroborates the distributional findings of the in vitro work although the efficiency of lysis is much reduced, possibly due to exopolymer capsules and slimes of natural sedimentary bacteria. Cross-phyletic comparisons of bacteriolytic activities reveal both qualitative and quantitative differences. Much less demarcation of lytic activity is observed in the guts of a holothuroid (Caudina arenata) and a hemichordate (Stereobalanus canadensis), with a pattern more similar to that of A. marina observed in another polychaete, Amphitrite johnstoni. Quantitatively, the polychaetes showed higher levels of activity with rates in A. marina exceeding those of the hemichordate and holothuroid by more than 10-fold.

Echinococcus P29 antigen: molecular characterization and implication on post-surgery follow-up of CE patients infected with different species of the Echinococcus granulosus complex.

The protein P29 is a potential serological marker for post-treatment monitoring of cystic echinococcosis (CE) especially in young patients. We now have demonstrated that P29 is encoded in the Echinococcus genus by a single gene consisting of 7 exons spanning 1.2 kb of DNA. Variability of the p29 gene at inter- and intra-species level was assessed with 50 cDNA and 280 genomic DNA clones isolated from different E. granulosus s.l. isolates (E. granulosus sensu stricto (G1), E. equinus (G4), E. ortleppi (G5), E. canadensis (G6), E. canadensis (G7) and E. canadensis (G10)) as well as four E. multilocularis isolates. Scarce interspecies polymorphism at the p29 locus was observed and affected predominantly E. granulosus s.s. (G1), where we identified two alleles (A1 and A2) coding for identical P29 proteins and yielding in three genotypes (A1/A1, A2/A2 and A1/A2). Genotypic frequencies expected under Hardy-Weinberg equilibrium revealed a high rate of heterozygosity (47%) that strongly supports the hypothesis that E. granulosus s.s. (G1) is predominantly outbreeding. Comparative sequence analyses of the complete p29 gene showed that phylogenetic relationships within the genus Echinococcus were in agreement with those of previous nuclear gene studies. At the protein level, the deduced P29 amino acid (AA) sequences exhibited a high level of conservation, ranging from 97.9% AA sequence identity among the whole E. granulosus s.l. group to 99.58% identity among E. multilocularis isolates. We showed that P29 proteins of these two species differ by three AA substitutions without implication for antigenicity. In Western-blot analyses, serum antibodies from a human CE patient infected with E. canadensis (G6) strongly reacted with recombinant P29 from E. granulosus s.s. (G1) (recEg(G1)P29). In the same line, human anti-Eg(G1)P29 antibodies bound to recEcnd(G6)P29. Thus, minor AA sequence variations appear not to impair the prognostic serological use of P29.

The Cenomanian/Turronian Boundary Event at ODP Hole 103-641A

Drilling at ODP Site 641 (on the western margin of Galicia Bank, off northwestern Spain) revealed a thin, but pronounced, interval of black shale and gray-green claystone. Our high-resolution study combines the sedimentology, micropaleontology (palynomorphs and others), organic and inorganic geochemistry, and isotopic values of this layer to demonstrate the distinct nature of the sediment and prove that the sequence represents the local sedimentary expression of the global Cenomanian/Turonian Oceanic Anoxic Event (OAE) of Schlanger and Jenkyns (1976), Arthur and Schlanger (1979), and Jenkyns (1980), also called the Cenomanian/Turonian Boundary Event (CTBE). The most striking evidence is that the strong positive d13C excursion characterizing the CTBE sequences in shallow areas can be traced into a pronounced deep-sea expression, thus providing a good stratigraphic marker for the CTBE in various paleosettings. The isotopic excursion at Site 641 coincides with an extremely enriched trace metal content, with values that were previously unknown for the Cretaceous Atlantic. Similar to other CTBE occurrences, the organic carbon content is high (up to 11%) and the organic matter is of dominantly marine origin (kerogen type II). The bulk mineralogy of the CTBE sediments does not differ significantly from the general trend of Cretaceous North Atlantic sediments (dominance of smectite and zeolite with minor amounts of illite and scattered palygorskite, kaolinite, and chlorite); thus, no evidence for either increased volcanic activity nor a drastic climatic change in the borderlands was found. Results from Site 641 are compared with the CTBE section found at Site 398, DSDP Leg 47B (Vigo Seamount at the southern end of the Galicia Bank).

Fuste comercial de clones del género Populus : relación entre volumen y densidad de la madera con nutrientes inorgánicos

El objetivo del trabajo fue determinar sobre 10 clones de álamos, provenientes de cruzamientos intraespecíficos de Populus deltoides e interespecíficos de Populus x canadensis, implantados en Teodelina, Santa Fe, Argentina (34°09' S; 61°15' W), la concentración de nutrientes inorgánicos en su corteza y leño, y sus relaciones con los volúmenes comerciales alcanzados a los 9 años de edad y la densidad básica de la madera. Se apearon ejemplares seleccionados, se calculó su volumen comercial individual y se determinaron la densidad básica y las concentraciones de Ca, Mg, P y K en fuste y corteza. Se realizó análisis de la varianza, test de comparación de medias de Tukey y correlación simple de los diferentes parámetros evaluados. Los resultados arrojaron diferentes contenidos de nutrientes por clon y concentraciones más bajas en la madera que en la corteza, en todos los casos. No hubo correlaciones significativas entre el volumen comercial obtenido, los valores de densidad básica de la madera y los nutrientes inorgánicos, indicando eficiencias individuales en el uso de los recursos inorgánicos.

Comportamiento de clones de álamos en Barreal : San Juan, Argentina

Con el objetivo de estudiar el comportamiento de diferentes clones de álamos en el valle cordillerano de Barreal, ubicado en la provincia de San Juan, Argentina, en 1995 se instaló un ensayo con los siguientes clones: 7 Populus x canadensis: Cima, Fogolino, Giorgione, Schiavone, Conti 12, I-214, Veronese y 7 Populus x deltoides: Harvard, Fierolo, I-72, 67/67, 71/67, Catfish 2 y Catfish 5. El sitio se encuentra a 31°36'55'' S, 69°27'30'' W y una altura de 1.628 msnm. El suelo es aluvial, de textura franca con cantos rodados de tamaño medio a partir de los 70 cm de la superficie. El marco de plantación fue de 5 x 2,5 m y el riego superficial por surcos. Se tomaron periódicamente datos dasométricos del diámetro altura de pecho (DAP) de todos los individuos, y altura total de árboles de diámetro promedio de cada clon. Además se observó cada una de las plantas a fin de determinar la presencia o ausencia de cancrosis del álamo y taladrillo de los forestales. Los resultados a la fecha muestran que los clones con mayor producción de madera, expresada en m3/ha son: Schiavone, I-214, Veronese, Conti 12 y Giorgione.

Comportamiento de clones de álamos en San Carlos : Mendoza, Argentina

Desde hace más de 30 años el Instituto Forestal de la Facultad de Ciencias Agrarias de la Universidad Nacional de Cuyo, Mendoza, Argentina, ha realizado la introducción de clones de álamos de diversos orígenes, con el fin de evaluar sus comportamientos frente a agentes bióticos y abióticos, ampliar la base genética disponible y mejorar los rendimientos volumétricos promedio de las plantaciones comerciales de la región. Como parte de esta línea de investigación, en 1996 se instaló un ensayo en un establecimiento agrícola-forestal ubicado en el Departamento de San Carlos, provincia de Mendoza, a los 33°46' S; 69°02' O y 940 msnm. Se evaluaron 10 clones de álamos: cuatro P. x deltoides (Stoneville 124, EEA Delta 107/68, INTA 69/69, Fierolo) y seis Populus x canadensis (El Campeador, Neva, Luisa Avanzo, B. L. Constanzo, I-42, I-455), que se dispusieron en parcelas de 9 plantas cada una distribuidas al azar con 4 repeticiones. La distancia de plantación fue de 4 x 6 m. Se realizaron mediciones anuales de diámetro altura pecho (DAP) y altura total de los árboles, además de registrar periódicamente el estado sanitario, en particular en lo referido a la presencia de ataques de cancrosis producida por Septoria musiva Peck. Los clones que produjeron un mayor volumen de madera/ha fueron: Stoneville 124 con una producción de 322 m3/ha, EEA Delta 107/68 con 293 m3/ha, INTA 69/69 con 285 m3/ha y Fierolo con 239 m3/ha. El clon Luisa Avanzo y el clon I-42 presentaron una alta susceptibilidad a cancrosis, lo que motivó un altísimo porcentaje de fallas a partir del tercer año, fallas que se repitieron en menor medida en el resto de los clones P. x canadensis. Cabe concluir que los clones con mejor comportamiento bajo las condiciones del ensayo fueron: Stoneville 124, EEA Delta 107/68 e INTA 69/69.

Requerimiento hídrico en álamos de siete años de edad : Rivadavia, Mendoza, Argentina

Durante tres años consecutivos se desarrolló el ensayo de riego en álamos de siete años (Populus x canadensis I-214) en el Departamento de Rivadavia, Mendoza, Argentina. El suelo es de textura franco-arenosa. El ensayo se estableció en cuatro bloques completos aleatorizados. La parcela unitaria fue de 192 m², con un total de 20 árboles. Los Intervalos de Riego (IR) utilizados fueron 7-14-21 días. Los resultados muestran que para el IR = 7 días la Evapotranspiración real (Etc) fue de 2.215 mm; para IR = 14 días de 1.372 mm y para IR = 21 días de 1.038 mm. El número de riegos anuales para IR = 7 días fue de 22 y la Evapotranspiración diaria del cultivo fue de 14,9 mm/día. El Coeficiente de cultivo (Kc) se obtuvo de relacionar la Etc con la Evapotranspiración potencial obtenida por la fórmula de Blaney y Criddle y por el tanque de evaporación estándar tipo A. Los resultados fueron respectivamente: Kc = 3 y 2,41 para IR = 7 días; Kc = 1,87 y 1,47 para IR = 14 días y Kc = 1,40 y 1,10 para IR = 21 días. Del análisis estadístico del volumen de madera producido se concluyó que el IR de 7 días resultó el más productivo para álamos de siete años.

Evaluación de clones de Populus spp., a los dos años de edad, en dos micrositios de la región pampeana, Argentina

La producción de álamos en la pampa ondulada de la provincia de Buenos Aires podría constituir una actividad alternativa y/o complementaria, debido a zonas ecológicas favorables para su cultivo. En este contexto es importante conocer el comportamiento de los clones en los diversos ambientes para poder definir cuáles de ellos podrían presentar mayor estabilidad para los diferentes sitios. El objetivo del trabajo fue estudiar la interacción existente entre los parámetros de crecimiento y supervivencia de 16 clones de Populus spp. en dos micrositios geomorfológicamente diferentes en la región pampeana, Argentina. Los clones en estudio provienen de cruzamientos intraespecíficos de P. deltoides e interespecíficos de P. deltoides x P. nigra (Populus x canadensis). Se plantaron en dos ensayos, uno por cada situación geomórfica: loma y bajo, y se completó con un análisis estadístico comparando los ensayos en forma individual y conjunta. Cada ensayo se instaló con un diseño de bloques completos al azar. Las variables analizadas fueron la supervivencia, la altura media y el área basal al concluir el segundo año de crecimiento. Para el sitio regional evaluado, la interacción entre los clones y los micrositios fue significativa para las variables altura y supervivencia. La mayor disponibilidad de agua en el micrositio bajo produjo mayores crecimientos clonales con diferencias significativas respecto del micrositio loma. La estabilidad de los atributos de supervivencia y crecimiento frente a los micrositios es un objetivo fundamental en las plantaciones clonales. Siendo los micrositios una realidad ambiental importante, propia como variable no controlable, la reducción de la variabilidad de respuesta ante ellos permitiría la constitución de unidades de manejo más homogéneas.

Susceptibilidad a Septoria musiva de híbridos inter e intraespecíficos de Populus spp. implantados en dos micrositios de la Pampa Húmeda Argentina

En los álamos, una de las enfermedades con mayor influencia en la cantidad y calidad de la madera producida es la cancrosis originada por Septoriamusiva Peck . El objetivo del trabajo fue determinar en clones híbridos de álamos (Populus spp.) la incidencia y la severidad de la enfermedad producida por Septoria, en dos micrositios de un mismo sitio regional, en Alberti, Buenos Aires, Argentina. Los clones utilizados fueron obtenidos de cruzamientos intraespecíficos de Populus deltoides e interespecíficos de P. deltoides x P. nigra (= P. x canadensis). Se determinó la susceptibilidad a la enfermedad para cada clon/micrositio mediante la estimación de la incidencia, del número de cancros en guías y ramas, y la severidad de daños (ID). Los híbridos inter e intraespecíficos de P. deltoides presentaron respuesta clonal diferencial en la susceptibilidad a la cancrosis en ambas situaciones geomórficas: los híbridos provenientes de cruzamientos intraespecíficos resultaron resistentes, y los provenientes de cruzamientos interespecíficos susceptibles a la enfermedad. Las diferencias significativas entre clones, determinadas para el número de lesiones totales por posición en ambas situaciones geomórficas, no determinaron ID diferentes. El número de cancros identificados en cada posición constituye una herramienta de estimación de la susceptibilidad clonal a la cancrosis más exacta, teniendo en cuenta que los mayores perjuicios ocurren cuando las lesiones se presentan en las guías.

Plants growing near the cargo packing station, and seed occurrence in cargo and luggage destined for Gough and Marion Island and the Antarctic

Globalization has resulted in unprecedented movements of people, goods, and alien species across the planet. Although the impacts of biological invasions are widely appreciated, a bias exists in research effort to post-dispersal processes because of the difficulties of measuring propagule pressure. The Antarctic provides an ideal model system in which to investigate propagule movements because of the region's isolation and small number of entry routes. Here we investigated the logistics operations of the South African National Antarctic Programme (SANAP) and quantified the initial dispersal of alien species into the region. we found that over 1400 seeds from 99 taxa are transported into the Antarctic each field season in association with SANAP passenger luggage and cargo. The first ever assessment of propagule drop-off indicated that 30-50% of these propagules will enter the recipient environment. Many of the taxa include cosmopolitan weeds and known aliens in the Antarctic, indicating that logistics operations form part of a globally self-perpetuating cycle moving alien species between areas of human disturbance. in addition, propagules of some taxa native to the Antarctic region were also found, suggesting that human movements may be facilitating intra-regional homogenization. Several relatively simple changes in biosecurity policy that could significantly reduce the threat of introduction of nonnative species are suggested.

Pollen and macrofossils profiles and age determination from three cores in Northeastern Kansas, U.S.A.

Two marshes near Muscotah and Arrington, Atchison County, northeastern Kansas, yielded a pollen sequence covering the last 25,000 yrs of vegetation development. The earliest pollen spectra are comparable with surface pollen spectra from southern Saskatchewan and southeastern Manitoba and might indicate a rather open vegetation but with some pine, spruce, and birch as the most important tree species, with local stands of alder and willow. This type of vegetation changed about 23,000 yrs ago to a spruce forest, which prevailed in the region until at least 15,000 yrs ago. Because of a hiatus, the vegetation changes resulting in the spread of a mixed deciduous forest and prairie, which was present in the region from 11,000 to 9,000 yrs ago, remain unknown. Prairie vegetation, with perhaps a few trees along the valleys, covered the region until about 5,000 yrs ago, when a re-expansion of deciduous trees began in the lowlands.

Palynology on sediment profile Tso Kar in Ladakh

Palynological investigation of a 410 cm long core section from Tso Kar (33°10'N, 78°E, 4527 m a.s.l.), an alpine lake situated in the arid Ladakh area of NW India at the limit of the present-day Indian summer monsoon, was performed in order to reconstruct post-glacial regional vegetation and climate dynamics. The area was covered with alpine desert vegetation from ca. 15.2 to 14 kyr BP (1 kyr=1000 cal. years), reflecting dry and cold conditions. High influx values of long-distance transported Pinus sylvestris type pollen suggest prevailing air flow from the west and northwest. The spread of alpine meadow communities and local aquatic vegetation is a weak sign of climate amelioration after ca. 14 kyr BP. Pollen data (e.g. influx values of Pinus roxburghii type and Quercus) suggest that this was due to a strengthening of the summer monsoon and the reduced activity of westerly winds. The further spread of Artemisia and species-rich meadows occurred in response to improved moisture conditions between ca. 12.9 and 12.5 kyr BP. The subsequent change towards drier desert-steppe vegetation likely indicates more frequent westerly disturbances and associated snowfalls, which favoured the persistence of alpine meadows on edaphically moist sites. The spread of Chenopodiaceae-dominated vegetation associated with an extremely weak monsoon occurred at ca. 12.2-11.8 kyr BP during the Younger Dryas interstadial. A major increase in humidity is inferred from the development of Artemisia-dominated steppe and wet alpine meadows with Gentianaceae after the late glacial/early Holocene transition in response to the strengthening of the summer monsoon. Monsoonal influence reached maximum activity in the Tso Kar region between ca. 10.9 and 9.2 kyr BP. The subsequent development of the alpine meadow, steppe and desert-steppe vegetation points to a moderate reduction in the moisture supply, which can be linked to the weaker summer monsoon and the accompanying enhancement of the winter westerly flow from ca. 9.2 to 4.8 kyr BP. The highest water levels of Tso Kar around 8 kyr BP probably reflect combined effect of both monsoonal and westerly influence in the region. An abrupt shift towards aridity in the Tso Kar region occurred after ca. 4.8 kyr BP, as evidenced by an expansion of Chenopodiaceae-dominated desert-steppe. Low pollen influx values registered ca. 2.8-1.3 kyr BP suggest scarce vegetation cover and unfavourable growing conditions likely associated with a further weakening of the Indian Monsoon.

Contribution of woody habitat islands to the conservation of birds and their potential ecosystem services in an extensive Colombian rangeland

The conservation of birds and their habitats is essential to maintain well-functioning ecosystems including human-dominated habitats. In simplified or homogenized landscapes, patches of natural and semi-natural habitat are essential for the survival of plant and animal populations. We compared species composition and diversity of trees and birds between gallery forests, tree islands and hedges in a Colombian savanna landscape to assess how fragmented woody plant communities affect forest bird communities and how differences in habitat characteristics influenced bird species traits and their potential ecosystem function. Bird and tree diversity was higher in forests than in tree islands and hedges. Soil depth influenced woody species distribution, and canopy cover and tree height determined bird species distribution, resulting in plant and bird communities that mainly differed between forest and non-forest habitat. Bird and tree species and traits widely co-varied. Bird species in tree islands and hedges were on average smaller, less specialized to habitat and more tolerant to disturbance than in forest, but dietary differences did not emerge. Despite being less complex and diverse than forests, hedges and tree islands significantly contribute to the conservation of forest biodiversity in the savanna matrix. Forest fragments remain essential for the conservation of forest specialists, but hedges and tree islands facilitate spillover of more tolerant forest birds and their ecological functions such as seed dispersal from forest to the savanna matrix.