Mitochondrial inheritance during a parasexual cycle in {\it Fusarium oxysporum\/} forma specialis {\it cubense\/}

Fusarium oxysporum is a diverse, asexual fungal species composed of both saprophytic and pathogenic members. The destructive phytopathogens are classified into formae speciales based on the host species and into vegetative compatibility groups (VCGs) based on the ability of two individuals to form heterokaryons. Parasexuality, a non-sexual mode of genetic exchange unique to some fungi has been demonstrated in the laboratory in Fusarium oxysporum f. sp. cubense (FOC). The goals of this dissertation were threefold: to ascertain whether mitochondrial (mt) markers can distinguish race differences in FOC; to determine genetic relatedness of VCGs in FOC based on a mt marker; and to discover the mode of mt inheritance during a parasexual cycle.^ Band patterns produced by electrophoresis of Hae III digested genomic DNA indicated that VCG differences, not race, could be discerned by mtDNA analysis. Primers were designed to amplify a mt intergenic locus which served as a molecular marker to screen 55 strains of FOC in 16 VCGs using both single strand conformational polymorphism and DNA sequencing. Based on homogeneity of the locus, strains were assigned to seven mitotypes, a classification unit which I introduced and found informative for grouping related VCGs.^ To determine the mode of mt inheritance during a parasexual cycle, strains in different mitotypes were paired. Mitochondrial inheritance in all hybrid progeny was found to be uniparental. I speculated that if a parasexual cycle occurs in nature there would be greater variation in the nuclear genome than the mt. This could produce multiple VCGs within a mitotype, a phenomenon observed in FOC. Based on these data, I concluded that parasexuality in nature may contribute to the diversity observed in Fusarium oxysporum. ^

The isolation, characterization, and application of WRKY genes as useful molecular markers in tropical trees

The improvement of tropical tree crops using conventional breeding methods faces challenges due to the length of time involved. Thus, like most crops, there is an effort to utilize molecular genetic markers in breeding programs to select for desirable agronomic traits. Known as marker assisted breeding or marker assisted selection, genetic markers associated with a phenotype of interest are used to screen and select material reducing the time necessary to evaluate candidates. As the focus of this research was improving disease resistance in tropical trees, the usefulness of the WRKY gene superfamily was investigated as candidates for generating useful molecular genetic markers. WRKY genes encode plant-specific transcriptional factors associated with regulating plants' responses to both biotic and abiotic stress. ^ One pair of degenerate primers amplified 48 WRKY gene fragments from three taxonomically distinct, economically important, tropical tree crop species: 18 from Theobroma cacao L., 21 from Cocos nucifera L. and 9 from Persea americana Mill. Several loci from each species were polymorphic because of single nucleotide substitutions present within a putative non-coding region of the loci. Capillary array electrophoresis-single strand conformational polymorphism (CAE-SSCP) mapped four WRKY loci onto a genetic linkage map of a T. cacao F2 population segregating for resistance to witches' broom disease. Additionally, PCR primers specific for four T. cacao loci successfully amplified WRKY loci from 15 members of the Byttneriae tribe. A method was devised to allow the reliable discrimination of alleles by CAE-SSCP using only the mobility assigned to the sample peaks. Once this method was validated, the diversity of three WRKY loci was evaluated in a germplasm collection of T. cacao . One locus displayed high diversity in the collection, with at least 18 alleles detected from mobility differences of the product peaks. The number of WRKY loci available within the genome, ease of isolation by degenerate PCR, codominant segregation demonstrated in the F2 population, and usefulness for screening germplasm collections and closely related wild species demonstrates that the WRKY superfamily of genes are excellent candidates for developing a number of genetic molecular markers for breeding purposes in tropical trees. ^

Anti-quorum sensing agents from south Florida medicinal plants and their attenuation of Pseudomonas aeruginosa pathogenicity

With the difficulty in treating recalcitrant infections and the growing resistance to antibiotics, new therapeutic modalities are becoming increasingly necessary. The interruption of bacterial quorum sensing (QS), or cell-cell communication is known to attenuate virulence, while limiting selective pressure toward resistance. This study initiates an ethnobotanically-directed search for QS inhibiting agents in south Florida medicinal plants. Fifty plants were screened for anti-QS activity using two biomonitor strains, Chromobacterium violaceum and Agrobacterium tumefaciens. Of these plants, six showed QS inhibition: Conocarpus erectus L. (Combretaceae), Chamaecyce hypericifolia (L.) Millsp. (Euphorbiaceae), Callistemon viminalis (Sol.ex Gaertn.) G. Don (Myrtaceae), Bucida burceras L. (Combretaceae), Tetrazygia bicolor (Mill.) Cogn. (Melastomataceae), and Quercus virginiana Mill. (Fagaceae). These plants were further examined for their effects on the QS system and virulence of Pseudomonas aeruginosa, an intractable opportunistic pathogen responsible for morbidity and mortality in the immunocompromised patient. C. erectus, B. buceras, and C. viminalis were found to significantly inhibit multiple virulence factors and biofilm formation in this organism. Each plant presented a distinct profile of effect on QS genes and signaling molecules, suggesting varying modes of action. Virulence attenuation was observed with marginal reduction of bacterial growth, suggesting quorum quenching mechanisms unrelated to static or cidal effects. Extracts of these plants were also investigated for their effects on P. aeruginosa killing of the nematode Caenorhabditis elegans. Results were evaluated in both toxin-based and infection-based assays with P. aeruginosa strains PA01 and PA14. Overall nematode mortality was reduced 50-90%. There was no indication of host toxicity, suggesting the potential for further development as anti-infectives. Using low-pressure chromatography and HPLC, two stereoisomeric ellagitannins, vescalagin and castalagin were isolated from an aqueous extract of C. erectus . Structures were confirmed via mass spectrometry and NMR spectroscopy. Both ellagitannins were shown to decrease signal production, QS gene expression, and virulence factor production in P. aeruginosa. This study introduces a potentially new therapeutic direction for the treatment of bacterial infections. In addition, this is the first report of vescalagin and castalagin being isolated from C. erectus, and the first report of ellagitannin activity on the QS system.

Effects of invasive Africanized honey bees (Apis mellifera scutellata) on native stingless bee populations (Meliponinae) and traditional Mayan beekeeping in central Quintana Roo, Mexico

The Maya of the Yucatan region have a long history of keeping the native stingless bees (subfamily Meliponinae). However, market forces in the last few decades have driven the Maya to favor the use of invasive Africanized honey bees (Apis mellifera scutellata) for producing large quantities of high quality honey that has an international market. Furthermore, the native bees traditionally used by the Maya are now disappearing, along with the practice of keeping them. ^ An interdisciplinary approach was taken in order to determine the social factors behind the decrease in stingless beekeeping and the ecological driving forces behind their disappearance from the wild. Social research methods included participant observation with stingless beekeepers, Apis beekeepers, and marketing intermediaries. Ecological research methods included point observations of commonly known melliferous and polliniferous plants along transects in three communities with different degrees of human induced ecosystem disturbance. ^ The stingless bee species most important to the Maya, Melipona beecheii, has become extremely rare, and this has caused a breakdown of stingless beekeeping tradition, compounded with the pressure of the market economy, which fuels Apis beekeeping and has lessened the influence of traditional practices. The community with the heaviest amount of human induced ecosystem disturbance also had the highest degree of dominance of Apis mellifera, while the area with the most intact ecosystem had the highest diversity of stingless bees, though Apis mellifera was still the dominant species. Aggressive competitive behavior involving physical attacks by Apis mellifera against stingless bees was observed on several occasions, and this is a new observation previously unreported by science. ^

Sugar and coffee: A history of settler agriculture in nineteenth-century Liberia

This dissertation is about commercial agriculture in nineteenth-century Liberia. Based primarily on the archives of the American Colonization Society (founder of Liberia), it examines the impact of environmental and demographic constraints on an agrarian settler society from 1822 to the 1890s. Contrary to the standard interpretation, which linked the poor state of commercial agriculture to the settlers' disdain for cultivation, this dissertation argues that the scarcity of labor and capital impeded the growth of commercial agriculture. The causes of the scarcity were high mortality, low immigration and the poverty of the American “Negroes” who began to settle Liberia in 1822. ^ Emigration to Liberia meant almost certain death and affliction for many immigrants because they encountered a new set of diseases. Mortality was particularly high during the early decades of colonization. From 1822 to 1843, about 48 percent of all immigrants died of various causes, usually within their first year. The bulk of the deaths is attributed to malaria. There was no natural increase in the population for this early period and because American “Negroes” were unenthusiastic about relocation to Liberia, immigration remained sparse throughout the century. Low immigration, combined with the high death rate, deprived the fledgling colony of its potential human resource, especially for the cultivation of labor-intensive crops, like sugar cane and coffee. Moreover, even though females constituted approximately half of the settlers, they seldom performed agricultural labor. ^ The problem of labor was compounded by the scarcity of draft animals. Liberia is in the region where trypanosomiasis occurs. The disease is fatal to large livestock. Therefore, animal-drawn plows, common in the United States, were never successfully transplanted in Liberia. Besides, the dearth of livestock obstructed the development of the sugar industry since many planters depended on oxen-powered mills because they could not afford to buy the more expensive steam engine mills. ^ Finally, nearly half of the immigrants were newly emancipated slaves. Usually these former bondsmen arrived in Liberia penniless. Consequently, they lacked the capital to invest in large-scale plantations. The other categories of immigrants (e.g., those who purchased their freedom), were hardly better off than the emancipated slaves. ^

The Role of Agriculture and Food Consumption in Tropical Conservation

A growing human population, shifting human dietary habits, and climate change are negatively affecting global ecosystems on a massive scale. Expanding agricultural areas to feed a growing population drives extensive habitat loss, and climate change compounds stresses on both food security and ecosystems. Understanding the negative effects of human diet and climate change on agricultural and natural ecosystems provides a context within which potential technological and behavioral solutions can be proposed to help maximize conservation. The purpose of this research was to (1) examine the potential effects of climate change on the suitability of areas for commercial banana plantations in Latin America in the 2050s and how shifts in growing areas could affect protected areas; (2) test the ability of small unmanned aerial vehicles (UAVs) to map productivity of banana plantations as a potential tool for increasing yields and decreasing future plantation expansions; (3) project the effects on biodiversity of increasing rates of animal product consumption in developing megadiverse countries; and (4) estimate the capacity of global pasture biomass production and Fischer-Tropsch hydrocarbon synthesis (IGCC-FT) processing to meet electricity, gasoline and diesel needs. The results indicate that (1) the overall extent of areas suitable for conventional banana cultivation is predicted to decrease by 19% by 2050 because of a hotter and drier climate, but all current banana exporting countries are predicted to maintain some suitable areas with no effects on protected areas; (2) Spatial patterns of NDVI and ENDVI were significantly positively correlated with several metrics of fruit yield and quality, indicating that UAV systems can be used in banana plantations to map spatial patterns of fruit yield; (3) Livestock production is the single largest driver of habitat loss, and both livestock and feedstock production are increasing in developing biodiverse tropical countries. Reducing global animal product consumption should therefore be at the forefront of strategies aimed at reducing biodiversity loss; (4) Removing livestock from global pasture lands and instead utilizing the biomass production could produce enough energy to meet 100% of the electricity, gasoline, and diesel needs of over 40 countries with extensive grassland ecosystems, primarily in tropical developing countries.

The role of agriculture and food consumption in tropical conservation

A growing human population, shifting human dietary habits, and climate change are negatively affecting global ecosystems on a massive scale. Expanding agricultural areas to feed a growing population drives extensive habitat loss, and climate change compounds stresses on both food security and ecosystems. Understanding the negative effects of human diet and climate change on agricultural and natural ecosystems provides a context within which potential technological and behavioral solutions can be proposed to help maximize conservation. The purpose of this research was to (1) examine the potential effects of climate change on the suitability of areas for commercial banana plantations in Latin America in the 2050s and how shifts in growing areas could affect protected areas; (2) test the ability of small unmanned aerial vehicles (UAVs) to map productivity of banana plantations as a potential tool for increasing yields and decreasing future plantation expansions; (3) project the effects on biodiversity of increasing rates of animal product consumption in developing megadiverse countries; and (4) estimate the capacity of global pasture biomass production and Fischer-Tropsch hydrocarbon synthesis (IGCC-FT) processing to meet electricity, gasoline and diesel needs. The results indicate that (1) the overall extent of areas suitable for conventional banana cultivation is predicted to decrease by 19% by 2050 because of a hotter and drier climate, but all current banana exporting countries are predicted to maintain some suitable areas with no effects on protected areas; (2) Spatial patterns of NDVI and ENDVI were significantly positively correlated with several metrics of fruit yield and quality, indicating that UAV systems can be used in banana plantations to map spatial patterns of fruit yield; (3) Livestock production is the single largest driver of habitat loss, and both livestock and feedstock production are increasing in developing biodiverse tropical countries. Reducing global animal product consumption should therefore be at the forefront of strategies aimed at reducing biodiversity loss; (4) Removing livestock from global pasture lands and instead utilizing the biomass production could produce enough energy to meet 100% of the electricity, gasoline, and diesel needs of over 40 countries with extensive grassland ecosystems, primarily in tropical developing countries.^