Receptor-binding residues lie in central regions of duffy-binding-like domains involved in red cell invasion and cytoadherence by malaria parasites

Erythrocyte invasion by malaria parasites and cytoadherence of Plasmodium falciparum-infected erythrocytes to host capillaries are 2 key pathogenic mechanisms in malaria. The receptor-binding domains of erythrocyte-binding proteins (EBPs) such as Plasmodium falciparum EBA-175, which mediate invasion, and P falciparum erythrocyte membrane protein 1 (PfEMP-1) family members, which are encoded by var genes and mediate cytoadherence, have been mapped to conserved cysteine-rich domains referred to as Duffy-binding-like (DBL) domains. Here, we have mapped regions within DBL domains from EBPs and PfEMP-1 that contain receptor-binding residues. Using biochemical and molecular methods we demonstrate that the receptor-binding residues of parasite ligands that bind sialic acid on glycophorin A for invasion as well as complement receptor-1 and chondroitin sulfate A for cytoadherence map to central regions of DBL domains. In contrast, binding to intercellular adhesion molecule 1 (ICAM-1) requires both the central and terminal regions of DBLβC2 domains. Determination of functional regions within DBL domains is the first step toward understanding the structure-function bases for their interaction with diverse host receptors

A rapid method for the isolation of genomic DNA from Aspergillus fumigatus

A majority of Aspergillus induced diseases are reported to be caused by Aspergillus fumigatus. In immunocompromized and post transplant cases it can lead to invasive aspergillosis. Due to this the molecular fingerprinting of aspergillus isolates by RFLP analysis and development of DNA diagnostic probes are gaining importance. Different methodologies are being adopted for extraction of the genomic DNA from fungus. The existing procedures for isolation of DNA are time consuming and range from several hours to few days. The most difficult step in the isolation of DNA from aspergillus species is to disrupt the tough chitin rich ceil wall without causing damage to genomic DNA. We report here a rapid method for extraction of genomic DNA based on the cleavage of chitin with chitinase. The subsequent modification steps included are lysis and microwave treatment. The chromosomal DNA obtained by this procedure is 1.5 - 2.0 μg per mg of wet weight of mycelia and is observed to be minimally sheared. It is pure enough for restriction analysis and for use in the PCR to detect the gene coding for 18 kDa allergen which has been identified in our laboratory using western blot analysis with human patient sera

Immunogenicity of Duffy Binding-Like Domains That Bind Chondroitin Sulfate A and Protection against Pregnancy-Associated Malaria

Sequestration of Plasmodium falciparum-infected erythrocytes in the placenta is implicated in pathological outcomes of pregnancy-associated malaria (PAM). P. falciparum isolates that sequester in the placenta primarily bind chondroitin sulfate A (CSA). Following exposure to malaria during pregnancy, women in areas of endemicity develop immunity, and so multigravid women are less susceptible to PAM than primigravidae. Protective immunity to PAM is associated with the development of antibodies that recognize diverse CSA-binding, placental P. falciparum isolates. The epitopes recognized by such protective antibodies have not been identified but are likely to lie in conserved Duffy binding-like (DBL) domains, encoded by var genes, that bind CSA. Immunization of mice with the CSA-binding DBL3γ domain encoded by var1CSA elicits cross-reactive antibodies that recognize diverse CSA-binding P. falciparum isolates and block their binding to placental cryosections under flow. However, CSA-binding isolates primarily express var2CSA, which does not encode any DBLγ domains. Here, we demonstrate that antibodies raised against DBL3γ encoded by var1CSA cross-react with one of the CSA-binding domains, DBL3X, encoded by var2CSA. This explains the paradoxical observation made here and earlier that anti-rDBL3γ sera recognize CSA-binding isolates and provides evidence for the presence of conserved, cross-reactive epitopes in diverse CSA-binding DBL domains. Such cross-reactive epitopes within CSA-binding DBL domains can form the basis for a vaccine that provides protection against PAM