A Screen against Leishmania Intracellular Amastigotes: Comparison to a Promastigote Screen and Identification of a Host Cell-Specific Hit

The ability to screen compounds in a high-throughput manner is essential in the process of small molecule drug discovery. Critical to the success of screening strategies is the proper design of the assay, often implying a compromise between ease/speed and a biologically relevant setting. Leishmaniasis is a major neglected disease with limited therapeutic options. In order to streamline efforts for the design of productive drug screens against Leishmania, we compared the efficiency of two screening methods, one targeting the free living and easily cultured promastigote (insect–infective) stage, the other targeting the clinically relevant but more difficult to culture intra-macrophage amastigote (mammal-infective) stage. Screening of a 909-member library of bioactive compounds against Leishmania donovani revealed 59 hits in the promastigote primary screen and 27 in the intracellular amastigote screen, with 26 hits shared by both screens. This suggested that screening against the promastigote stage, although more suitable for automation, fails to identify all active compounds and leads to numerous false positive hits. Of particular interest was the identification of one compound specific to the infective amastigote stage of the parasite. This compound affects intracellular but not axenic parasites, suggesting a host cell-dependent mechanism of action, opening new avenues for anti-leishmanial chemotherapy

Stern's Introductory Plant Biology

iv, 622 hlm.; 22 x 28 c

BIOMASS AND NITROGEN TRAITS OF SUMMER PIGEON PEAS AND WINTER WHEAT GROWN FOR THREE ROTATIONS IN CONTAINERS

Pigeon pea [Cajanus cajan (L.) Millsp.] cultivars, ‘Georgia-1’ and ‘ICPL-87’, were grown without inoculation and with Bradyrhizobium inoculation (multistrain, TAL 1127, or TAL 1132) to evaluate legume dry weight (DW) and nitrogen (N) content, soil mineral N, and subsequent wheat (Triticum aestivum L.) productivity. Pigeon peas were grown during summer and ‘TAM 101’ wheat was grown during winter, along with summer fallow controls fertilized with 0, 45, and 90 kg N ha-1, in 36-cm diam. 20-L pots from 1992 to 1995. Representative pigeon peas were harvested in the fall and remaining plants were incorporated into the soil. Wheat was planted and soil cores were collected at 35 to 48 d after pigeon pea harvest. Wheat was harvested the following spring. Factors affecting DW and N content of both crops included length of growing season, environmental variation, and contribution of residual N. Among pigeon pea cultivars, Georgia-1 occasionally demonstrated higher DW and N content compared with ICPL-87. Estimation of N provided by pigeon pea to the last wheat crop in the third sequence of yearly rotations was 30 kg N ha-1. Pigeon pea treatments demonstrating highest DW, N content, and contribution to soil N generally produced winter wheat with higher yield and N content compared with other treatments. While yield and N content of winter wheat fertilized at 90 kg N ha-1 either decreased or stayed the same from 1993 to 1995, these same measurements in wheat following pigeon peas demonstrated a 3- to 4-fold increase over the same time period and warrant further research in field rotation systems of the southern Great Plains

Dry Weight and Nitrogen Content of Chickpea and Winter Wheat Grown in Pots for Three Rotations

Chickpea [Cicer arietinum (L.)] cultivars ‘ICCV-2’ and ‘Sarah’ were studied along with a control, multistrain, TAL 1148, and TAL 480 Bradyrhizobium strains to determine the effect(s) of cultivar and inoculum on dry weight (DW) and nitrogen (N) content of the legume, as well as soil mineral N, DW, and N content of wheat [Triticum aestivum (L.) emend. Thell.] in a continuous wheat-legume rotation. Chickpeas were planted during the summer and harvested in the fall of 1992, 1993, and 1994. Vegetative growth from chickpeas was incorporated into the soil prior to wheat planting, and soil cores were taken at 35 to 48 d after chickpea harvests. Additional summer fallow treatments for the winter wheat part of the experiment received 0, 45, and 90 kg N ha−1 each year. Wheat plants were removed the following spring and stubble was incorporated into the soil before planting chickpeas in the summer. ‘Sarah’ chickpeas accumulated about the same or more shoot DW and shoot N compared to ‘ICCV-2’; whereas ‘ICCV-2’ generally produced more pod DW and pod N compared to ‘Sarah.’ Inoculum had no significant effect on chickpea DW or N content. Wheat DW and N following legumes increased marginally after growing ‘Sarah’ chickpeas, as evidenced by higher values of some treatments. Only the multistrain or absence of inoculum in ‘Sarah’ chickpeas resulted in significantly greater wheat DW or N content compared to the fallow wheat receiving no added N fertilizer. The contributions from ‘ICCV-2’ chickpeas to wheat DW and N content were not significant. Soil mineral N, as well as wheat DW and N content, fluctuated or increased during this three-year study, which demonstrated some benefit from incorporation of chickpeas into a wheat-legume cropping system

An Active Type IV Secretion System Encoded by the F Plasmid Sensitizes Escherichia coli to Bile Salts

F(+) strains of Escherichia coli infected with donor-specific bacteriophage such as M13 are sensitive to bile salts. We show here that this sensitivity has two components. The first derives from secretion of bacteriophage particles through the cell envelope, but the second can be attributed to expression of the F genes required for the formation of conjugative (F) pili. The latter component was manifested as reduced or no growth of an F(+) strain in liquid medium containing bile salts at concentrations that had little or no effect on the isogenic F(−) strain or as a reduced plating efficiency of the F(+) strain on solid media; at 2% bile salts, plating efficiency was reduced 10(4)-fold. Strains with F or F-like R factors were consistently more sensitive to bile salts than isogenic, plasmid-free strains, but the quantitative effect of bile salts depended on both the plasmid and the strain. Sensitivity also depended on the bile salt, with conjugated bile salts (glycocholate and taurocholate) being less active than unconjugated bile salts (deoxycholate and cholate). F(+) cells were also more sensitive to sodium dodecyl sulfate than otherwise isogenic F(−) cells, suggesting a selectivity for amphipathic anions. A mutation in any but one F tra gene required for the assembly of F pili, including the traA gene encoding F pilin, substantially restored bile salt resistance, suggesting that bile salt sensitivity requires an active system for F pilin secretion. The exception was traW. A traW mutant was 100-fold more sensitive to cholate than the tra(+) strain but only marginally more sensitive to taurocholate or glycocholate. Bile salt sensitivity could not be attributed to a generalized change in the surface permeability of F(+) cells, as judged by the effects of hydrophilic and hydrophobic antibiotics and by leakage of periplasmic β-lactamase into the medium

Stern's Introductory Plant Biology

xviii, 607 hlm, ilus.; 22x28c

Stern's introductory plant biology

xviii, 622 hlm. : ilus. ; tab. ; 27 cm