An anchored restriction-mapping approach applied to the genetic analysis of the Anopheles gambiae malaria vector complex 1

We introduce here a simple approach for rapidly determining restriction maps for a number of regions of a genome; this involves "anchoring" a map with a rare restriction site (in this case the seldom-cutting EagI) followed by partial digestion of a frequent-cutting enzyme (e.g., Sau 3A). We applied this technology to five species of the Anopheles gambiae complex. In a single Southern blot we obtained about a 15-kb restriction map each for the mtDNA, rRNA gene, and a scnDNA region for each of five species. Phylogenetic analyses of these regions yield trees at odds with the more traditional chromosome inversion-based trees. The value of the approach for systematic purposes is the ease with which several large, independent regions of the genome can be quickly assayed for molecular variation.      

Maternal glucocorticoid deficit affects hypothalamic-pituitary-adrenal function and behavior of rat offspring

Detrimental consequences of prenatal stress include increased hypothalamic-pituitary-adrenal (HPA) function, anxiety and depression-like behavior in adult offspring. To identify the role of maternal corticosterone milieu in the fetal programming of adult function, we measured these same behavioral and hormonal endpoints after maternal adrenalectomy (ADX) and replacement with normal or moderately high levels of corticosterone (CORT). Adult male and female offspring exhibited differing HPA responses to maternal ADX. In female offspring of ADX mothers, exaggerated plasma ACTH stress responses were reversed by the higher, but not the lower, dose of maternal CORT. In contrast, male offspring of both ADX and ADX dams with higher CORT replacement showed exaggerated ACTH stress responses. Hypothalamic glucocorticoid receptor (GR) expression was decreased in these latter groups, while hippocampal GR increased only in the ADX offspring. Activity of young offspring of ADX dams replaced with the higher dose of CORT decreased in the open field test of exploration/anxiety, while immobility behavior of adult offspring in the forced swim test of depression increased following maternal ADX or higher levels of CORT replacement. Interestingly, for some measures, none or moderately high CORT replacement resulted in similar deficits in this study. These findings are in accord with consequences of prenatal stress or prenatal dexamethasone exposure, suggesting that a common mechanism may underlie the effects of too low or too high maternal glucocorticoids on adult HPA function and behavior.     

Impact of ivermectin on onchocerciasis transmission: assessing the empirical evidence that repeated ivermectin mass treatments may lead to elimination/eradication in West-Africa

BACKGROUND: The Onchocerciasis Control Program (OCP) in West Africa has been closed down at the end of 2002. All subsequent control will be transferred to the participating countries and will almost entirely be based on periodic mass treatment with ivermectin. This makes the question whether elimination of infection or eradication of onchocerciasis can be achieved using this strategy of critical importance. This study was undertaken to explore this issue. METHODS: An empirical approach was adopted in which a comprehensive analysis was undertaken of available data on the impact of more than a decade of ivermectin treatment on onchocerciasis infection and transmission. Relevant entomological and epidemiological data from 14 river basins in the OCP and one basin in Cameroon were reviewed. Areas were distinguished by frequency of treatment (6-monthly or annually), endemicity level and additional control measures such as vector control. Assessment of results were in terms of epidemiological and entomological parameters, and as a measure of inputs, therapeutic and geographical coverage rates were used. RESULTS: In all of the river basins studied, ivermectin treatment sharply reduced prevalence and intensity of infection. Significant transmission, however, is still ongoing in some basins after 10-12 years of ivermectin treatment. In other basins, transmission may have been interrupted, but this needs to be confirmed by in-depth evaluations. In one mesoendemic basin, where 20 rounds of four-monthly treatment reduced prevalence of infection to levels as low as 2-3%, there was significant recrudescence of infection within a few years after interruption of treatment. CONCLUSIONS: Ivermectin treatment has been very successful in eliminating onchocerciasis as a public health problem. However, the results presented in this paper make it almost certain that repeated ivermectin mass treatment will not lead to the elimination of transmission of onchocerciasis from West Africa. Data on 6-monthly treatments are not sufficient to draw definitive conclusions.     

Sucrose-dependent H+ transport in plasma-membrane vesicles isolated from sugarbeet leaves (Beta vulgaris L.)

The mechanism of sucrose transport across the plasma membrane (PM) was investigated in membrane vesicles isolated from sugarbeet (Beta vulgaris L.) leaves. In the presence of a membrane potential () generated as a K⁺-diffusion potential, negative inside, sucrose induced a rapid and transient alkalization of the medium. Alkalization was inhibited by carbonyl cyanide m-chlorophenylhydrazone, was specific for the sucrose sugar and was dependent on the sucrose concentration with a K_m of approx. 1 mM. Sucrose-induced alkalization and sucrose transport were inhibited by the sulfhydryl-reactive reagent, p-chloromercuribenzene sulfonic acid, and by the histidine-reactive reagent, diethyl pyrocarbonate. Parallel analysis of sucrose uptake and alkalization indicated that the stoichiometry of sucrose uptake to proton consumed was 11. These results provide clear evidence that the saturable mechanism of sucrose transport across the PM in plants is a coupled H⁺-sucrose symport.Abbreviations and Symbols CCCP carbonyl cyanide m-chlorophenylhydrazone - DEPC diethyl pyrocarbonate - PCMBS p-chloromercuribenzene sulfonic acid - pH pH gradient - membrane potential difference - PM plasma membrane The financial support for a portion of thus study was provided by the Deutsche Forschungsgemeinschaft. We thank Kimberly A. Mitchell for her excellent technical assistance and dedicate this report to the memory of Mr. William A. Dungey.     

Modeling Chemotherapy Resistant Leukemia In Vitro

It is well established that the bone marrow microenvironment provides a unique site of sanctuary for hematopoietic diseases that both initiate and progress in this site. The model presented in the current report utilizes human primary bone marrow stromal cells and osteoblasts as two representative cell types from the marrow niche that influence tumor cell phenotype. The in vitro co-culture conditions described for human leukemic cells with these primary niche components support the generation of a chemoresistant subpopulation of tumor cells that can be efficiently recovered from culture for analysis by diverse techniques. A strict feeding schedule to prevent nutrient fluxes followed by gel type 10 cross-linked dextran (G10) particles recovery of the population of tumor cells that have migrated beneath the adherent bone marrow stromal cells (BMSC) or osteoblasts (OB) generating a "phase dim" (PD) population of tumor cells, provides a consistent source of purified therapy resistant leukemic cells. This clinically relevant population of tumor cells can be evaluated by standard methods to investigate apoptotic, metabolic, and cell cycle regulatory pathways as well as providing a more rigorous target in which to test novel therapeutic strategies prior to pre-clinical investigations targeted at minimal residual disease.     

Hops as a metapopulation landscape for tetranychid-phytoseiid interactions: perspectives of intra- and interplant dispersal

Population densities, distributions and dispersal of Neoseiulus fallacis (Garman) and Tetranychus urticae (Koch) on individual hop plants, Humulus lupulus L. were studied for attributes of metapopulations such as empty patches, asynchrony of subpopulations, extinction of subpopulations, and dispersal of predators and prey among patches. Occupancy of hop leaves by predators or prey was stable over a season with 69–75% of leaves having neither predators nor prey, 4–15% with prey mites only, 9–17% with both predators and prey mites and 6–10% with predaceous mites only. Stability of occupancy classes through time indicated that inherently unstable predator and prey subpopulations developed asynchronously. Flagged hop leaves showed the existence of many empty individual leaves, colonization of some by prey, then by predators, then extinction of both, and then recolonization by spider mites. This illustrated the existence of empty patches, extinction of subpopulations, and dispersal of predators and prey to empty patches. This differed from spider mites and phytoseiid predators on apple foliage where there was a progression of occupancy status, indicating synchronous development of subpopulations on individual plants. Studies of predator and prey dispersal between hop plants showed that removal of basal leaves to 1.5 m high, a common agronomic practice, greatly limited dispersal of the predaceous mites but not the spider mites. Retaining basal leaves facilitated interplant movement of predators and improved the extent and timing of biological control. Through management, N. fallacis dispersal may be adjusted so that the entire hop planting becomes a metapopulation landscape, leading to greater stability and persistence of predator–prey within a season.     

The core apoptotic executioner proteins CED-3 and CED-4 promote initiation of neuronal regeneration in Caenorhabditis elegans

A critical accomplishment in the rapidly developing field of regenerative medicine will be the ability to foster repair of neurons severed by injury, disease, or microsurgery. In C. elegans, individual visualized axons can be laser-cut in vivo and neuronal responses to damage can be monitored to decipher genetic requirements for regeneration. With an initial interest in how local environments manage cellular debris, we performed femtosecond laser axotomies in genetic backgrounds lacking cell death gene activities. Unexpectedly, we found that the CED-3 caspase, well known as the core apoptotic cell death executioner, acts in early responses to neuronal injury to promote rapid regeneration of dissociated axons. In ced-3 mutants, initial regenerative outgrowth dynamics are impaired and axon repair through reconnection of the two dissociated ends is delayed. The CED-3 activator, CED-4/Apaf-1, similarly promotes regeneration, but the upstream regulators of apoptosis CED-9/Bcl2 and BH3-domain proteins EGL-1 and CED-13 are not essential. Thus, a novel regulatory mechanism must be utilized to activate core apoptotic proteins for neuronal repair. Since calcium plays a conserved modulatory role in regeneration, we hypothesized calcium might play a critical regulatory role in the CED-3/CED-4 repair pathway. We used the calcium reporter cameleon to track in vivo calcium fluxes in the axotomized neuron. We show that when the endoplasmic reticulum calcium-storing chaperone calreticulin, CRT-1, is deleted, both calcium dynamics and initial regenerative outgrowth are impaired. Genetic data suggest that CED-3, CED-4, and CRT-1 act in the same pathway to promote early events in regeneration and that CED-3 might act downstream of CRT-1, but upstream of the conserved DLK-1 kinase implicated in regeneration across species. This study documents reconstructive roles for proteins known to orchestrate apoptotic death and links previously unconnected observations in the vertebrate literature to suggest a similar pathway may be conserved in higher organisms.     

The genetic architecture of fitness in a seed beetle: assessing the potential for indirect genetic benefits of female choice

Background  

Quantifying the amount of standing genetic variation in fitness represents an empirical challenge. Unfortunately, the shortage of detailed studies of the genetic architecture of fitness has hampered progress in several domains of evolutionary biology. One such area is the study of sexual selection. In particular, the evolution of adaptive female choice by indirect genetic benefits relies on the presence of genetic variation for fitness. Female choice by genetic benefits fall broadly into good genes (additive) models and compatibility (non-additive) models where the strength of selection is dictated by the genetic architecture of fitness. To characterize the genetic architecture of fitness, we employed a quantitative genetic design (the diallel cross) in a population of the seed beetle Callosobruchus maculatus, which is known to exhibit post-copulatory female choice. From reciprocal crosses of inbred lines, we assayed egg production, egg-to-adult survival, and lifetime offspring production of the outbred F1 daughters (F1 productivity).