Investigation on Cacao swollen shoot virus (CSSV) pollen transmission through cross-pollination

DNA- and RNA-based polymerase chain reaction (PCR) systems were used with Cacao swollen shoot virus (CSSV) primers designed from conserved regions of the six published genomic sequences of CSSV to investigate whether the virus is transmissible from infected trees through cross-pollination to seeds and seedlings. Pollen was harvested from CSSV infected cocoa trees and used to cross-pollinate flowers of healthy cocoa trees (recipient parents) to generate enough cocoa seeds for the PCR screening. Adequate precautions were taken to avoid cross-contamination during duplicated DNA extractions and only PCR results accompanied by effective positive and negative controls were scored. Results from the PCR analyses showed that samples of cocoa pod husk, mesocarp and seed tissues (testa, cotyledon and embryo) from the cross-pollinations were PCR negative for CSSV DNA. Sequential DNA samples from new leaves of seedlings resulting from the cross-pollinated trees were consistently PCR negative for presence of portions of CSSV DNA for over 36 months after germination. A reverse transcription-PCR analysis performed on the seedlings showed negative results, indicating absence of functional CSSV RNA transcripts in the seedlings. None of the seedlings exhibited symptoms characteristic of the CSSV disease, and all infectivity tests on the seedlings were also negative. Following these results, the study concluded that although CSSV DNA was detected in pollen from CSSV infected trees, there was no evidence of pollen transmission of the virus through cross-pollination from infected cocoa parents to healthy cocoa trees. Keywords:badnavirus;CSSV;PCR;pollen;seed transmission;Theobroma cacao

Pollen-based continental climate reconstructions at 6 and 21 ka: a global synthesis

Subfossil pollen and plant macrofossil data derived from 14C-dated sediment profiles can provide quantitative information on glacial and interglacial climates. The data allow climate variables related to growing season warmth, winter cold, and plant-available moisture to be reconstructed. Continental-scale reconstructions have been made for the mid-Holocene (MH, around 6 ka) and Last Glacial Maximum (LGM, around 21 ka), allowing comparison with palaeoclimate simulations currently being carried out as part of the fifth Assessment Report (AR5) of the Intergovernmental Panel on Climate Change. The synthesis of the available MH and LGM climate reconstructions and their uncertainties, obtained using modern analogue, regression and model-inversion techniques, is presented for four temperature variables and two moisture variables. Reconstructions of the same variables based on surface-pollen assemblages are shown to be accurate and unbiased. Reconstructed LGM and MH climate anomaly patterns are coherent, consistent between variables, and robust with respect to the choice of technique. They support a conceptual model of the controls of Late Quaternary climate change whereby the first-order effects of orbital variations and greenhouse forcing on the seasonal cycle of temperature are predictably modified by responses of the atmospheric circulation and surface energy balance.

Investigation on Cacao swollen shoot virus (CSSV) pollen transmission through cross-pollination

DNA- and RNA-based polymerase chain reaction (PCR) systems were used with Cacao swollen shoot virus (CSSV) primers designed from conserved regions of the six published genomic sequences of CSSV to investigate whether the virus is transmissible from infected trees through cross-pollination to seeds and seedlings. Pollen was harvested from CSSV infected cocoa trees and used to cross-pollinate flowers of healthy cocoa trees (recipient parents) to generate enough cocoa seeds for the PCR screening. Adequate precautions were taken to avoid cross-contamination during duplicated DNA extractions and only PCR results accompanied by effective positive and negative controls were scored. Results from the PCR analyses showed that samples of cocoa pod husk, mesocarp and seed tissues (testa, cotyledon and embryo) from the cross-pollinations were PCR negative for CSSV DNA. Sequential DNA samples from new leaves of seedlings resulting from the cross-pollinated trees were consistently PCR negative for presence of portions of CSSV DNA for over 36 months after germination. A reverse transcription-PCR analysis performed on the seedlings showed negative results, indicating absence of functional CSSV RNA transcripts in the seedlings. None of the seedlings exhibited symptoms characteristic of the CSSV disease, and all infectivity tests on the seedlings were also negative. Following these results, the study concluded that although CSSV DNA was detected in pollen from CSSV infected trees, there was no evidence of pollen transmission of the virus through cross-pollination from infected cocoa parents to healthy cocoa trees.

An improved methodology for the recovery of Zea mays and other large crop pollen, with implications for environmental archaeology in the Neotropics

We present a simple sieving methodology to aid the recovery of large cultigen pollen grains, such as maize (Zea mays L.), manioc (Manihot esculenta Crantz), and sweet potato (Ipomoea batatas L.), among others, for the detection of food production using fossil pollen analysis of lake sediments in the tropical Americas. The new methodology was tested on three large study lakes located next to known and/or excavated pre-Columbian archaeological sites in South and Central America. Five paired samples, one treated by sieving, the other prepared using standard methodology, were compared for each of the three sites. Using the new methodology, chemically digested sediment samples were passed through a 53 µm sieve, and the residue was retained, mounted in silicone oil, and counted for large cultigen pollen grains. The filtrate was mounted and analysed for pollen according to standard palynological procedures. Zea mays (L.) was recovered from the sediments of all three study lakes using the sieving technique, where no cultigen pollen had been previously recorded using the standard methodology. Confidence intervals demonstrate there is no significant difference in pollen assemblages between the sieved versus unsieved samples. Equal numbers of exotic Lycopodium spores added to both the filtrate and residue of the sieved samples allow for direct comparison of cultigen pollen abundance with the standard terrestrial pollen count. Our technique enables the isolation and rapid scanning for maize and other cultigen pollen in lake sediments, which, in conjunction with charcoal and pollen records, is key to determining land-use patterns and the environmental impact of pre-Columbian societies.

Characterisation of Bolivian savanna ecosystems by their modern pollen rain and implications for fossil pollen records

The majority of vegetation reconstructions from the Neotropics are derived from fossil pollen records extracted from lake sediments. However, the interpretation of these records is restricted by limited knowledge of the contemporary relationships between the vegetation and pollen rain of Neotropical ecosystems, especially for more open vegetation such as savannas. This research aims to improve the interpretation of these records by investigating the vegetation and modern pollen rain of different savanna ecosystems in Bolivia using vegetation inventories, artificial pollen traps and surface lake sediments. Two types of savanna were studied, upland savannas (cerrado), occurring on well drained soils, and seasonally-inundated savannas occurring on seasonally water-logged soils. Quantitative vegetation data are used to identify taxa that are floristically important in the different savanna types and to allow modern pollen/vegetation ratios to be calculated. Artificial pollen traps from the upland savanna site are dominated by Moraceae (35%), Poaceae (30%), Alchornea (6%) and Cecropia (4%). The two seasonally-inundated savanna sites are dominated by Moraceae (37%), Poaceae (20%), Alchornea (8%) and Cecropia (7%), and Moraceae (25%), Cyperaceae (22%), Poaceae (19%) and Cecropia (9%), respectively. The modern pollen rain of seasonally-inundated savannas from surface lake sediments is dominated by Cyperaceae (35%), Poaceae (33%), Moraceae (9%) and Asteraceae (5%). Upland and seasonally-flooded savannas were found to be only subtly distinct from each other palynologically. All sites have a high proportion of Moraceae pollen due to effective wind dispersal of this pollen type from areas of evergreen forest close to the study sites. Modern pollen/vegetation ratios show that many key woody plant taxa are absent/under-represented in the modern pollen rain (e.g., Caryocar and Tabebuia). The lower-than-expected percentages of Poaceae pollen, and the scarcity of savanna indicators, in the modern pollen rain of these ecosystems mean that savannas could potentially be overlooked in fossil pollen records without consideration of the full pollen spectrum available.

Pollen-based differentiation of Amazonian rainforest communities and implications for lowland palaeoecology in tropical South America

An ongoing controversy in Amazonian palaeoecology is the manner in which Amazonian rainforest communities have responded to environmental change over the last glacial–interglacial cycle. Much of this controversy results from an inability to identify the floristic heterogeneity exhibited by rainforest communities within fossil pollen records. We apply multivariate (Principal Components Analysis) and classification (Unweighted Pair Group with Arithmetic Mean Agglomerative Classification) techniques to floral-biometric, modern pollen trap and lake sediment pollen data situated within different rainforest communities in the tropical lowlands of Amazonian Bolivia. Modern pollen rain analyses from artificial pollen traps show that evergreen terra firme (well-drained), evergreen terra firme liana, evergreen seasonally inundated, and evergreen riparian rainforests may be readily differentiated, floristically and palynologically. Analogue matching techniques, based on Euclidean distance measures, are employed to compare these pollen signatures with surface sediment pollen assemblages from five lakes: Laguna Bella Vista, Laguna Chaplin, and Laguna Huachi situated within the Madeira-Tapajós moist forest ecoregion, and Laguna Isirere and Laguna Loma Suarez, which are situated within forest patches in the Beni savanna ecoregion. The same numerical techniques are used to compare rainforest pollen trap signatures with the fossil pollen record of Laguna Chaplin.

Differentiation between Neotropical rainforest, dry forest and savannah ecosystems by their modern pollen spectra and implications for the fossil pollen record

Accurate differentiation between tropical forest and savannah ecosystems in the fossil pollen record is hampered by the combination of: i) poor taxonomic resolution in pollen identification, and ii) the high species diversity of many lowland tropical families, i.e. with many different growth forms living in numerous environmental settings. These barriers to interpreting the fossil record hinder our understanding of the past distributions of different Neotropical ecosystems and consequently cloud our knowledge of past climatic, biodiversity and carbon storage patterns. Modern pollen studies facilitate an improved understanding of how ecosystems are represented by the pollen their plants produce and therefore aid interpretation of fossil pollen records. To understand how to differentiate ecosystems palynologically, it is essential that a consistent sampling method is used across ecosystems. However, to date, modern pollen studies from tropical South America have employed a variety of methodologies (e.g. pollen traps, moss polsters, soil samples). In this paper, we present the first modern pollen study from the Neotropics to examine the modern pollen rain from moist evergreen tropical forest (METF), semi-deciduous dry tropical forest (SDTF) and wooded savannah (cerradão) using a consistent sampling methodology (pollen traps). Pollen rain was sampled annually in September for the years 1999–2001 from within permanent vegetation study plots in, or near, the Noel Kempff Mercado National Park (NKMNP), Bolivia. Comparison of the modern pollen rain within these plots with detailed floristic inventories allowed estimates of the relative pollen productivity and dispersal for individual taxa to be made (% pollen/% vegetation or ‘p/v’). The applicability of these data to interpreting fossil records from lake sediments was then explored by comparison with pollen assemblages obtained from five lake surface samples.

Pollen-based biome reconstructions for Latin America at 0, 6000 and 18000 radiocarbon years

The biomisation method is used to reconstruct Latin American vegetation at 6000±500 and 18 000±1000 radiocarbon years before present (14C yr BP) from pollen data. Tests using modern pollen data from 381 samples derived from 287 locations broadly reproduce potential natural vegetation. The strong temperature gradient associated with the Andes is recorded by a transition from high altitude cool grass/shrubland and cool mixed forest to mid-altitude cool temperate rain forest, to tropical dry, seasonal and rain forest at low altitudes. Reconstructed biomes from a number of sites do not match the potential vegetation due to local factors such as human impact, methodological artefacts and mechanisms of pollen representivity of the parent vegetation.

Pre-Columbian raised-field agriculture and land use in the Bolivian Amazon

We present an integrated palaeoecological and archaeobotanical study of pre-Columbian raised-field agriculture in the Llanos de Moxos, a vast seasonally inundated forest–savanna mosaic in the Bolivian Amazon. Phytoliths from excavated raised-field soil units, together with pollen and charcoal in sediment cores from two oxbow lakes, were analysed to provide a history of land use and agriculture at the El Cerro raised-field site. The construction of raised fields involved the removal of savanna trees, and gallery forest was cleared from the area by AD 310. Despite the low fertility of Llanos de Moxos soils, we determined that pre-Columbian raised-field agriculture sufficiently improved soil conditions for maize cultivation. Fire was used as a common management practice until AD 1300, at which point, the land-use strategy shifted towards less frequent burning of savannas and raised fields. Alongside a reduction in the use of fire, sweet potato cultivation and the exploitation of Inga fruits formed part of a mixed resource strategy from AD 1300 to 1450. The pre-Columbian impact on the landscape began to lessen around AD 1450, as shown by an increase in savanna trees and gallery forest. Although agriculture at the site began to decline prior to European arrival, the abandonment of raised fields was protracted, with evidence of sweet potato cultivation occurring as late as AD 1800.

Pollen, plant macrofossil and charcoal records for palaeovegetation reconstruction in the Mediterranean-Black Sea Corridor since the Last Glacial Maximum

New reconstructions of changing vegetation patterns in the Mediterranean-Black Sea Corridor since the Last Glacial Maximum are being produced by an improved biomisation scheme that uses both pollen and plant macrofossil data, in conjunction. Changes in fire regimes over the same interval will also be reconstructed using both microscopic and macroscopic charcoal remains. These reconstructions will allow a diagnosis of the interactions between climate, fire and vegetation on millennial timescales, and will also help to clarify the role of coastline and other geomorphic changes, salinity and impacts of human activities in this region. These new data sets are being produced as a result of collaboration between the Palynology Working Group (WG-2) within the IGCP-521 project and the international Palaeovegetation Mapping Project (BIOME 6000). The main objective of this paper is to present the goals of this cooperation, methodology, including limitations and planned improvements, and to show the initial results of some applications.

Relationships between plant traits and climate in the Mediterranean region: an analysis based on pollen data

Question: What are the correlations between the degree of drought stress and temperature, and the adoption of specific adaptive strategies by plants in the Mediterranean region? Location: 602 sites across the Mediterranean region. Method: We considered 12 plant morphological and phenological traits, and measured their abundance at the sites as trait scores obtained from pollen percentages. We conducted stepwise regression analyses of trait scores as a function of plant available moisture (α) and winter temperature (MTCO). Results: Patterns in the abundance for the plant traits we considered are clearly determined by α, MTCO or a combination of both. In addition, trends in leaf size, texture, thickness, pubescence and aromatic leaves and other plant level traits such as thorniness and aphylly, vary according to the life form (tree, shrub, forb), the leaf type (broad, needle) and phenology (evergreen, summer-green). Conclusions: Despite conducting this study based on pollen data we have identified ecologically plausible trends in the abundance of traits along climatic gradients. Plant traits other than the usual life form, leaf type and leaf phenology carry strong climatic signals. Generally, combinations of plant traits are more climatically diagnostic than individual traits. The qualitative and quantitative relationships between plant traits and climate parameters established here will help to provide an improved basis for modelling the impact of climate changes on vegetation and form a starting point for a global analysis of pollen-climate relationships

Pollen-based reconstructions of biome distributions for Australia, Southeast Asia and the Pacific (SEAPAC region) at 0,6000 and 18,000 14C yr B.P.

Aim  This paper documents reconstructions of the vegetation patterns in Australia, Southeast Asia and the Pacific (SEAPAC region) in the mid-Holocene and at the last glacial maximum (LGM). Methods  Vegetation patterns were reconstructed from pollen data using an objective biomization scheme based on plant functional types. The biomization scheme was first tested using 535 modern pollen samples from 377 sites, and then applied unchanged to fossil pollen samples dating to 6000 ± 500 or 18,000 ± 1000 14C yr bp. Results  1. Tests using surface pollen sample sites showed that the biomization scheme is capable of reproducing the modern broad-scale patterns of vegetation distribution. The north–south gradient in temperature, reflected in transitions from cool evergreen needleleaf forest in the extreme south through temperate rain forest or wet sclerophyll forest (WSFW) and into tropical forests, is well reconstructed. The transitions from xerophytic through sclerophyll woodlands and open forests to closed-canopy forests, which reflect the gradient in plant available moisture from the continental interior towards the coast, are reconstructed with less geographical precision but nevertheless the broad-scale pattern emerges. 2. Differences between the modern and mid-Holocene vegetation patterns in mainland Australia are comparatively small and reflect changes in moisture availability rather than temperature. In south-eastern Australia some sites show a shift towards more moisture-stressed vegetation in the mid-Holocene with xerophytic woods/scrub and temperate sclerophyll woodland and shrubland at sites characterized today by WSFW or warm-temperate rain forest (WTRF). However, sites in the Snowy Mountains, on the Southern Tablelands and east of the Great Dividing Range have more moisture-demanding vegetation in the mid-Holocene than today. South-western Australia was slightly drier than today. The single site in north-western Australia also shows conditions drier than today in the mid-Holocene. Changes in the tropics are also comparatively small, but the presence of WTRF and tropical deciduous broadleaf forest and woodland in the mid-Holocene, in sites occupied today by cool-temperate rain forest, indicate warmer conditions. 3. Expansion of xerophytic vegetation in the south and tropical deciduous broadleaf forest and woodland in the north indicate drier conditions across mainland Australia at the LGM. None of these changes are informative about the degree of cooling. However the evidence from the tropics, showing lowering of the treeline and forest belts, indicates that conditions were between 1 and 9 °C (depending on elevation) colder. The encroachment of tropical deciduous broadleaf forest and woodland into lowland evergreen broadleaf forest implies greater aridity. Main conclusions  This study provides the first continental-scale reconstruction of mid-Holocene and LGM vegetation patterns from Australia, Southeast Asia and the Pacific (SEAPAC region) using an objective biomization scheme. These data will provide a benchmark for evaluation of palaeoclimate simulations within the framework of the Palaeoclimate Modelling Intercomparison Project.

Modern pollen samples from alpine vegetation on the Tibetan Plateau.

1  A set of 316 modern surface pollen samples, sampling all the alpine vegetation types that occur on the Tibetan Plateau, has been compiled and analysed. Between 82 and 92% of the pollen present in these samples is derived from only 28 major taxa. These 28 taxa include examples of both tree (AP) and herb (NAP) pollen types. 2  Most of the modern surface pollen samples accurately reflect the composition of the modern vegetation in the sampling region. However, airborne dust-trap pollen samples do not provide a reliable assessment of the modern vegetation. Dust-trap samples contain much higher percentages of tree pollen than non-dust-trap samples, and many of the taxa present are exotic. In the extremely windy environments of the Tibetan Plateau, contamination of dust-trap samples by long-distance transport of exotic pollen is a serious problem. 3  The most characteristic vegetation types present on the Tibetan Plateau are alpine meadows, steppe and desert. Non-arboreal pollen (NAP) therefore dominates the pollen samples in most regions. Percentages of arboreal pollen (AP) are high in samples from the southern and eastern Tibetan Plateau, where alpine forests are an important component of the vegetation. The relative importance of forest and non-forest vegetation across the Plateau clearly follows climatic gradients: forests occur on the southern and eastern margins of the Plateau, supported by the penetration of moisture-bearing airmasses associated with the Indian and Pacific summer monsoons; open, treeless vegetation is dominant in the interior and northern margins of the Plateau, far from these moisture sources. 4  The different types of non-forest vegetation are characterized by different modern pollen assemblages. Thus, alpine deserts are characterized by high percentages of Chenopodiaceae and Artemisia, with Ephedra and Nitraria. Alpine meadows are characterized by high percentages of Cyperaceae and Artemisia, with Ranunculaceae and Polygonaceae. Alpine steppe is characterized by high abundances of Artemisia, with Compositae, Cruciferae and Chenopodiaceae. Although Artemisia is a common component of all non-forest vegetation types on the Tibetan Plateau, the presence of other taxa makes it possible to discriminate between the different vegetation types. 5  The good agreement between modern vegetation and modern surface pollen samples across the Tibetan Plateau provides a measure of the reliability of using pollen data to reconstruct past vegetation patterns in non-forested areas.