Landscape genetics of Phaedranassa herb. (Amaryllidaceae) in Ecuador

Speciation can be understood as a continuum occurring at different levels, from population to species. The recent molecular revolution in population genetics has opened a pathway towards understanding species evolution. At the same time, speciation patterns can be better explained by incorporating a geographic context, through the use of geographic information systems (GIS). Phaedranassa (Amaryllidaceae) is a genus restricted to one of the world’s most biodiverse hotspots, the Northern Andes. I studied seven Phaedranassa species from Ecuador. Six of these species are endemic to the country. The topographic complexity of the Andes, which creates local microhabitats ranging from moist slopes to dry valleys, might explain the patterns of Phaedranassa species differentiation. With a Bayesian individual assignment approach, I assessed the genetic structure of the genus throughout Ecuador using twelve microsatellite loci. I also used bioclimatic variables and species geographic coordinates under a Maximum Entropy algorithm to generate distribution models of the species. My results show that Phaedranassa species are genetically well-differentiated. Furthermore, with the exception of two species, all Phaedranassa showed non-overlapping distributions. Phaedranassa viridiflora and P. glauciflora were the only species in which the model predicted a broad species distribution, but genetic evidence indicates that these findings are likely an artifact of species delimitation issues. Both genetic differentiation and nonoverlapping geographic distribution suggest that allopatric divergence could be the general model of genetic differentiation. Evidence of sympatric speciation was found in two geographically and genetically distinct groups of P. viridiflora. Additionally, I report the first register of natural hybridization for the genus. The findings of this research show that the genetic differentiation of species in an intricate landscape as the Andes does not necessarily show a unique trend. Although allopatric speciation is the most common form of speciation, I found evidence of sympatric speciation and hybridization. These results show that the processes of speciation in the Andes have followed several pathways. The mixture of these processes contributes to the high biodiversity of the region.

Population viability analyses of Chamaecrista keyensis (Leguminosae: Caesalpinioideae), a narrowly endemic herb of the lower Florida Keys: Effects of seasonal timing of fires and the urban -wildland interface

This research first evaluated the effects of urban wildland interface on reproductive biology of the Big Pine Partridge Pea, Chamaecrista keyensis, an understory herb that is endemic to Big Pine Key, Florida. I found that C. keyensis was self-compatible, but depended on bees for seed set. Furthermore, individuals of C. keyensis in urban habitats suffered higher seed predation and therefore set fewer seeds than forest interior plants. ^ I then focused on the effects of fire at different times of the year, summer (wet) and winter (dry), on the population dynamics and population viability of C. keyensis. I found that C. keyensis population recovered faster after winter burns and early summer burns (May–June) than after late summer burns (July–September) due to better survival and seedling recruitment following former fires. Fire intensity had positive effects on reproduction of C. keyensis. In contrast, no significant fire intensity effects were found on survival, growth, and seedling recruitment. This indicated that better survival and seedling recruitment following winter and early summer burns (compared with late summer burns) were due to the reproductive phenology of the plant in relation to fires rather than differences in fire intensity. Deterministic population modeling showed that time since fire significantly affected the finite population growth rates (λ). Particularly, recently burned plots had the largest λ. In addition, effects of timing of fires on λ were most pronounced the year of burn, but not the subsequent years. The elasticity analyses suggested that maximizing survival is an effective way to minimize the reduction in finite population growth rate the year of burn. Early summer fires or dry-season fires may achieve this objective. Finally, stochastic simulations indicated that the C. keyensis population had lower extinction risk and population decline probability if burned in the winter than in the late summer. A fire frequency of approximately 7 years would create the lowest extinction probability for C. keyensis. A fire management regime including a wide range of burning seasons may be essential for the continued existence of C. keyensis and other endemic species of pine rockland on Big Pine Key. ^

Developing Ecological Criteria for Prescribed Fire in South Florida Pine Rockland Ecosystems

The pine rocklands of South Florida, characterized by a rich herbaceous flora with many narrowly endemic taxa beneath an overstory of south Florida slash pine (Pinus elliottii var. densa), are found in three areas: the Miami Rock Ridge of southeastern peninsular Florida, the Lower Florida Keys, and slightly elevated portions of the southern Big Cypress National Preserve. Fire is an important element in these ecosystems, since in its absence the pine canopy is likely to be replaced by dense hardwoods, resulting in loss of the characteristic pineland herb flora. Prescribed fire has been used in Florida Keys pine forests since the creation of the National Key Deer Refuge (NKDR), with the primary aim of reducing fuels. Because fire can also be an effective tool in shaping ecological communities, we conducted a 4-year research study which explored a range of fire management options in NKDR. The intent of the study was to provide the Fish and Wildlife Service and other land managers with information regarding when and where to burn in order to perpetuate these unique forests.

Investigating an Alternative Treatment to Cystic Fibrosis: Testing the Effect of Panax ginseng C.A. Meyer extracts on Pseudomonas aeruginosa

The predominant pathogen found in the lungs of cystic fibrosis (CF) patients is Pseudomonas aeruginosa. The success of the infection is partially due to virulence factor production, which is regulated by quorum sensing (QS) signaling. Currently, antibiotics are used to treat the infection, but resistant forms of P. aeruginosa have evolved, necessitating alternative treatments. Previous animal studies showed that treatment with extracts from the Chinese herb Panax ginseng C.A. Meyer reduced bacterial load resulting in a favorable immune response. It is hypothesized that ginsenosides, the major bioactive compounds in ginseng, is responsible for this effect. This study explores the role of ginseng extracts in attenuating P. aeruginosa virulence. A sequential extraction was performed using hexane, methylene chloride, methanol, and water. High performance liquid chromatography (HPLC) analysis showed the methanol and water ginseng extracts contained the known ginsenosides Rb1, Rb2, Rc, Rd, Re, and Rg1• All extracts were tested on biomonitor strains of Agrobacterium tumefaciens,Chromobacterium violaceum, and P. aeruginosa. Antibacterial and anti-QS activity were assessed using a disc diffusion assay. This was then followed by thin layer chromatography (TLC) bioautographic assay to further separate active compounds. The hexane and dichloromethane extracts, that lacked ginsenosides, displayed antibacterial activity against C. violaceum, whereas methanol and water extracts had anti-QS activity. The results of the bioassay with the pure ginsenoside standards showed that they lack antibacterial or anti-QS activity. Our results indicate that there are bioactive compounds, other than ginsenosides, that are the cause of antibacterial effects and anti-QS in the ginseng extracts.

Testing the Efficacy of Panax ginseng Extract as a Novel Anti-Quorum Sensing Agent

Quorum sensing (QS) is the phenomenon by which microorganisms regulate gene expression in response to cell-population density. These microorganisms synthesize and secrete small molecules known as autoinducers that increase in concentration as a function of cell density. Once the cell detects the minimal threshold concentration of an autoinducer, the gene expression is altered accordingly. Although the cellular circuitry responding to QS is relatively conserved, the cellular processes regulated by QS, for example are conjugation, motility, sporulation, biofilm formation and production of virulence factors importamt for infection. Since many pathogens have developed resistance to available antibiotics, novel therapies are required to further treat these pathogens. Inhibitors of QS are potential therapeutic candidates since they would inhibit virulence without selecting for antibiotic resistant strains. Previous studies have shown that the Chinese herb Panax ginseng contains compounds that inhibit QS activity. To further characterize this activity, a highly sensitive quantitative liquid assay was developed using a Chromobacterium violaceum AHL mutant, CV026 as a biomonitor strain. After confirmation that P. ginseng aqueous extracts had anti-QS activity, the extract was fractionated on a reverse phase C18 Sep Pak column. Most anti-QS activity was present in the flow through, and compounds eluted with ten percent acetonitrile in water antibacterial activity. We thus conclude that P. ginseng has anti-QS activity and the active compound is water soluble. Compounds from P. ginseng, could be used as a lead structure to design compound with higher anti-QS activity for therapeutic treatments.

Pattern of nutrient availability and plant community assemblage in Everglades Tree Islands, Florida, USA

We address the relative importance of nutrient availability in relation to other physical and biological factors in determining plant community assemblages around Everglades Tree Islands (Everglades National Park, Florida, USA). We carried out a one-time survey of elevation, soil, water level and vegetation structure and composition at 138 plots located along transects in three tree islands in the Park’s major drainage basin. We used an RDA variance partitioning technique to assess the relative importance of nutrient availability (soil N and P) and other factors in explaining herb and tree assemblages of tree island tail and surrounded marshes. The upland areas of the tree islands accumulate P and show low N concentration, producing a strong island-wide gradient in soil N:P ratio. While soil N:P ratio plays a significant role in determining herb layer and tree layer community assemblage in tree island tails, nevertheless part of its variance is shared with hydrology. The total species variance explained by the predictors is very low. We define a strong gradient in nutrient availability (soil N:P ratio) closely related to hydrology. Hydrology and nutrient availability are both factors influencing community assemblages around tree islands, nevertheless both seem to be acting together and in a complex mechanism. Future research should be focused on segregating these two factors in order to determine whether nutrient leaching from tree islands is a factor determining community assemblages and local landscape pattern in the Everglades, and how this process might be affected by water management.

The Sesquiterpene Lactone Dehydroleucodine Triggers Senescence and Apoptosis in Association with Accumulation of DNA Damage Markers

Sesquiterpene lactones (SLs) are plant-derived compounds that display anti-cancer effects. Some SLs derivatives have a marked killing effect on cancer cells and have therefore reached clinical trials. Little is known regarding the mechanism of action of SLs. We studied the responses of human cancer cells exposed to various concentrations of dehydroleucodine (DhL), a SL of the guaianolide group isolated and purified from Artemisia douglasiana (Besser), a medicinal herb that is commonly used in Argentina. We demonstrate for the first time that treatment of cancer cells with DhL, promotes the accumulation of DNA damage markers such as phosphorylation of ATM and focal organization of γH2AX and 53BP1. This accumulation triggers cell senescence or apoptosis depending on the concentration of the DhL delivered to cells. Transient DhL treatment also induces marked accumulation of senescent cells. Our findings help elucidate the mechanism whereby DhL triggers cell cycle arrest and cell death and provide a basis for further exploration of the effects of DhL in in vivo cancer treatment models.

Effect of Hydrologic Restoration on the Habitat of The Cape Sable Seaside Sparrow, Annual Report of 2002-2003

After developing field sampling protocols and making a series of consultations with investigators involved in research in CSSS habitat, we determined that vegetationhydrology interactions within this landscape are best sampled at a combination of scales. At the finer scale, we decided to sample at 100 m intervals along transects that cross the range of habitats present, and at the coarser scale, to conduct an extensive survey of vegetation at sites of known sparrow density dispersed throughout the range of the CSSS. We initiated sampling in the first week of January 2003 and continued it through the last week of May. During this period, we established 6 transects, one in each CSSS subpopulation, completed topographic survey along the Transects A, C, D, and F, and sampled herb and shrub stratum vegetation, soil depth and periphyton along Transects A, and at 179 census points. We also conducted topographic surveys and completed vegetation and soil depth sampling along two of five transects used by ENP researchers for monitoring long-term vegetation change in Taylor Slough. We analyzed the data by summarizing the compositional and structural measures and by using cluster analysis, ordination, weighted averaging regression, and weighted averaging calibration. The mean elevation of transects decreased from north to south, and Transect F had greater variation than other transects. We identified eight vegetation assemblages that can be grouped into two broad categories, ‘wet prairie’ and ‘marsh’. In the 2003 survey, wet prairies were most dominant in the northeastern sub-populations, and had shorter inferred-hydroperiod, higher species richness and shallower soils than marshes, which were common in Subpopulations A, D, and the southernmost regions of Sub-population B. Most of the sites at which birds were observed during 2001 or 2002 had an inferred-hydroperiod of 120-150 days, while no birds were observed at sites with an inferred-hydroperiod less than 120 days or more than 300 days. Management-induced water level changes in Taylor Slought during the 1980’s and 1990’s appeared to elicit parallel changes in vegetation. The results described in detail in the following pages serve as a basis for evaluating and modifying, if necessary, the sampling design and analytical techniques to be used in the next three years of the project.

Resampling of Permanent Pine Rockland Vegetation Plots on Big Pine Key

The pine rocklands of South Florida are characterized by an herbaceous flora with many narrowly endemic taxa, a diverse shrub layer containing several palms and numerous tropical hardwoods, and an overstory of south Florida slash pine (Pinus elliottii var. densa). Fire has been considered as an important environmental factor for these ecosystems, since in the absence of fire these pine forests are replaced by dense hardwood communities, resulting in loss of the characteristic pineland herb flora. Hence, in the Florida Keys pine forests, prescribed fire has been used since the creation of the National Key Deer Refuge. However, such prescribed burns were conducted in the Refuge mainly for fuel reduction, without much consideration of ecological factors. The USGS and Florida International University conducted a research study for four years, from 1998 to 2001, the objective of which was to document the response of pine rockland vegetation to a range of fire management options and to provide Fish and Wildlife Service and other land managers with information useful in deciding when and where to burn to perpetuate these unique pine forests. This study is described in detail in Snyder et al. (2005).

Monitoring of Tree Island Conditions in the Southern Everglades: The Effects of Hurricanes and Hydrology on the Status and Population Dynamics of Sixteen Tropical Hardwood Hammock Tree Islands

In 2005 we began a multi-year intensive monitoring and assessment study of tropical hardwood hammocks within two distinct hydrologic regions in Everglades National Park, under funding from the CERP Monitoring and Assessment Program. In serving as an Annual Report for 2010, this document, reports in detail on the population dynamics and status of tropical hardwood hammocks in Shark Slough and adjacent marl prairies during a 4-year period between 2005 and 2009. 2005-09 was a period that saw a marked drawdown in marsh water levels (July 2006 - July 2008), and an active hurricane season in 2005 with two hurricanes, Hurricane Katrina and Wilma, making landfall over south Florida. Thus much of our focus here is on the responses of these forests to annual variation in marsh water level, and on recovery from disturbance. Most of the data are from 16 rectangular permanent plots of 225-625 m2 , with all trees mapped and tagged, and bi-annual sampling of the tree, sapling, shrub, and herb layer in a nested design. At each visit, canopy photos were taken and later analyzed for determination of interannual variation in leaf area index and canopy openness. Three of the plots were sampled at 2-month intervals, in order to gain a better idea of seasonal dynamics in litterfall and litter turnover. Changes in canopy structure were monitored through a vertical line intercept method.