The witchweeds, members of the genus Striga, are noxious and persistent pests in farmers' fields and serious constraints to crop productivity throughout Africa, India, and Southeast Asia. Among the primary hosts for Striga are the major cereals (maize, sorghum, rice, and millet) and grain legumes (cowpea) that are important food staples worldwide. The negative impact of parasitic plants on crop productivity increases globally each year, and their potential for affecting domestic agriculture looms larger as the movement of seed resources expands on a global scale. At the present time there is a limited understanding of how Striga and other parasitic plants select a suitable host and overcome the innate defense responses of the host in order to complete their life-cycle. In the grasses most reported resistance to Striga appears to be polygenic with a large genotype by environment interaction. In contrast, resistance to S. gesnerioides in cowpea is conferred by single dominant genes functioning in a race-specific manner suggesting that a gene-for-gene mechanism similar to effector-triggered immunity (ETI) described in other host–pathogen interactions is likely operating in these parasite-host associations. A hallmark of ETI is the direct or indirect recognition of parasite-derived avirulence (Avr) factors and other effectors that interfere with plant innate immunity by host sensors (or R proteins) leading to activation of defense responses. The recent cloning and functional characterization of a race-specific R gene from cowpea encoding a canonical coiled-coil (CC)-nucleotide binding site (NBS)-leucine-rich repeat (LRR) type R-protein opens the door for further exploration of the mechanism of host resistance and provides a focal point for studies aimed at uncovering the molecular and genetic factors underlying parasite virulence and host selection. The potential for the development of novel strategies for parasite control and eradication based on parasite virulence factors is discussed.

The regulatory role of tumour necrosis factor (TNF) was investigated in murine infection with tetrathyridia of Mesocestoides corti. Recombinant TNFα reduced macrophage larvicidal activity in vitro. M. corti primed mice for TNF release in response to bacterial lipopolysaccharide (LPS) in vivo. TNF activity was amplified 100-fold at 14 days post-infection (p.i.), with a further rise at day 28 p.i. Maximal inflammatory reaction was observed histologically in the liver at the height of TNF activity. Hepatic necrosis was located within inflammatory foci, but not within the vicinity of the parasite itself, suggesting that TNF may contribute to the pathogenesis of infection. Peritoneal cells from infected mice, when stimulated with tetrathyridia in vitro, showed a 4-fold increase in TNFα activity at day 14 p.i. However, when peritoneal cells were stimulated with LPS in vitro, a marked increase in TNFα secretion was observed at 2 months post-infection followed by a slow decline. It is suggested that impaired macrophage effector function, previously attributed to endogenous endotoxin, which gains access to peritoneal macrophages through an inability of the liver to detoxify endotoxin, may be mediated through TNFα.

Allosteric ribozymes are engineered RNAs that operate as molecular switches whose rates of catalytic activity are modulated by the binding of specific effector molecules. New RNA molecular switches can be created by using “allosteric selection,” a molecular engineering process that combines modular rational design and in vitro evolution strategies. In this report, we describe the characterization of 3′,5′-cyclic nucleotide monophosphate (cNMP)-dependent hammerhead ribozymes that were created using allosteric selection (Koizumi et al., Nat Struct Biol, 1999, 6:1062–1071). Artificial phylogeny data generated by random mutagenesis and reselection of existing cGMP-, cCMP-, and cAMP-dependent ribozymes indicate that each is comprised of distinct effector-binding and catalytic domains. In addition, patterns of nucleotide covariation and direct mutational analysis both support distinct secondary-structure organizations for the effector-binding domains. Guided by these structural models, we were able to disintegrate each allosteric ribozyme into separate ligand-binding and catalytic modules. Examinations of the independent effector-binding domains reveal that each retains its corresponding cNMP-binding function. These results validate the use of allosteric selection and modular engineering as a means of simultaneously generating new nucleic acid structures that selectively bind ligands. Furthermore, we demonstrate that the binding affinity of an allosteric ribozyme can be improved through random mutagenesis and allosteric selection under conditions that favor tighter binding. This “affinity maturation” effect is expected to be a valuable attribute of allosteric selection as future endeavors seek to apply engineered allosteric ribozymes as biosensor components and as controllable genetic switches.