Sequence variation in the circumsporozoite protein gene of Plasmodium vivax appears to be regionally biased.

We have sequenced the circumsporozoite protein gene from 16 isolates from China, the Philippines, Papua New Guinea and the Solomon Islands. We found very limited polymorphisms in the non-repetitive regions of the circumsporozoite gene from these isolates. All samples from China contained a 36-base insert 3' to the repeats previously seen only in a North Korean isolate. Limited variation was found in the repeat regions, which allowed these and previously sequenced isolates to be classified into groups based on repeat structure. These groupings also correlate with the geographical origin of the isolates.

The effects of bistratene A on the development of Plasmodium falciparum in culture.

The effects of the marine ascidian compound bistratene A on in vitro cultures of Plasmodium falciparum were assessed. Concentrations from 0 to 1.5 micrograms ml(-1) of the compound were tested. The parasitaemia in asynchronous cultures treated with bistratene A increased normally over the first 40 h, then decreased leaving only gametocytes. When synchronized cultures were treated with a constant dose of 50 ng ml(-1), gametocytes developed more rapidly than they did in control cultures. In addition, gametocytes developed in drug-treated cultures of P. falciparum that normally do not produce gametocytes. Bistratene A appears to inhibit merozoite invasion as well as to induce gametocytogenesis.

Transgenesis of schistosomes: approaches employing mobile genetic elements.

Draft genome sequences for Schistosoma mansoni and Schistosoma japonicum are now available. However, the identity and importance of most schistosome genes have yet to be determined. Recently, progress has been made towards the genetic manipulation and transgenesis of schistosomes. Both loss-of-function and gain-of-function approaches appear to be feasible in schistosomes based on findings described in the past 5 years. This review focuses on reports of schistosome transgenesis, specifically those dealing with the transformation of schistosomes with exogenous mobile genetic elements and/or their endogenous relatives for the genetic manipulation of schistosomes. Transgenesis mediated by mobile genetic elements offers a potentially tractable route to introduce foreign genes to schistosomes, a means to determine the importance of schistosome genes, including those that could be targeted in novel interventions and the potential to undertake large-scale forward genetics by insertional mutagenesis.

Picornavirus receptor down-regulation by plasminogen activator inhibitor type 2.

Therapeutic interference with virus-cell surface receptor interactions represents a viable antiviral strategy. Here we demonstrate that cytoplasmic expression of the serine protease inhibitor (serpin), plasminogen activator inhibitor type 2 (PAI-2), affords a high level of protection from lytic infection by multiple human picornaviruses. The antiviral action of PAI-2 was mediated primarily through transcriptional down-regulation of the following virus receptors: intercellular adhesion molecule 1 (ICAM-1, a cellular receptor for the major group of rhinoviruses), decay-accelerating factor (a cellular receptor for echoviruses and coxsackieviruses), and to a lesser extent the coxsackie-adenovirus receptor protein (a cellular receptor for group B coxsackieviruses and group C adenoviruses). Expression of related cell surface receptors, including membrane cofactor protein and the poliovirus receptor, remained unaffected. These findings suggest that PAI-2 and/or related serpins may form the basis of novel antiviral strategies against picornavirus infections and also therapeutic interventions against ICAM-1-mediated respiratory inflammation.

Functional expression of dipeptidyl peptidase I (Cathepsin C) of the oriental blood fluke Schistosoma japonicum in Trichoplusia ni insect cells.

Proenzyme dipeptidyl peptidase I (DPP I) of Schistosoma japonicum was expressed in a baculovirus expression system utilizing Trichoplusia ni BTI-5B1-4 (High Five) strain host insect cells. The recombinant enzyme was purified from cell culture supernatants by affinity chromatography on nickel-nitriloacetic acid resin, exploiting a polyhistidine tag fused to the COOH-terminus of the recombinant protease. The purified recombinant enzyme resolved in reducing SDS-PAGE gels as three forms, of 55, 39, and 38 kDa, all of which were reactive with antiserum raised against bacterially expressed S. japonicum DPP I. NH(2)-terminal sequence analysis of the 55-kDa polypeptide revealed that it corresponded to residues -180 to -175, NH(2)-SRXKXK, of the proregion peptide of S. japonicum DPP I. The 39- and 38-kDa polypeptides shared the NH(2)-terminal sequence, LDXNQLY, corresponding to residues -73 to -67 of the proregion peptide and thus were generated by removal of 126 residues from the NH(2)-terminus of the proenzyme. Following activation for 24 h at pH 7.0, 37 degrees C under reducing conditions, the recombinant enzyme exhibited exopeptidase activity against synthetic peptidyl substrates diagnostic of DPP I. Specificity constants (k(cat)/K(m)) for the recombinant protease for the substrates H-Gly-Arg-NHMec and H-Gly-Phe-NHMec were found to be 14.4 and 10.7 mM(-)1 s(-1), respectively, at pH 7.0. Approximately 1 mg of affinity-purified schistosome DPP I was obtained per liter of insect cell culture supernatant, representing approximately 2 x 10(9) High Five cells.

Crithidia luciliae: effect of purine starvation on S-adenosyl-L-methionine uptake and protein methylation.

The utilization of S-adenosyl-L-[methyl-3H]methionine ([3H-methyl]AdoMet) by Crithidia luciliae was assessed under nutrient-replete and purine-starvation conditions. Uptake experiments with intact cells demonstrated that the radiolabel from this molecule was accumulated by purine-starved organisms at a rate approximately 10-fold greater than that observed in those cultivated in nutrient-replete medium. Purine-starved cells also incorporated the radiolabel into trichloroacetic acid insoluble material at an approximately 10-fold faster rate than nutrient-replete cells. No differences, however, were observed in the intracellular levels of AdoMet and its metabolites between organisms cultivated under the two conditions. Results of comparative labeling studies with [3H-methyl]AdoMet, S-adenosyl-L-[carboxyl-14C]methionine, L-[methyl-3H]methionine and L-[35S]methionine in the presence and absence of cycloheximide demonstrated that the incorporation of label from [3H-methyl]AdoMet was due to transmethylation and was independent of protein synthesis. Further, approximately 15 methylated protein bands were identified by SDS-PAGE analysis. Lysates from both purine-starved and nutrient-replete organisms demonstrated similar levels of activity of three protein methyltransferases (PMI, II, III). The differences observed in [3H-methyl]AdoMet utilization between purine-starved and nutrient-replete C. luciliae may reflect the enhanced purine transport capacity which results from purine starvation.