Wasim A. Siddiqui

Plasmodium falciparum: gene structure and hydropathy profile of the major merozoite surface antigen (gp195) of the Uganda-Palo Alto isolate.

The gene encoding the 195,000-Da major merozoite surface antigen (gp195) of the FUP (Uganda-Palo Alto) isolate of Plasmodium falciparum, a strain widely used for monkey vaccination experiments, has been cloned and sequenced. The translated amino acid sequence of the FUP gp195 protein is closely related to the sequences of corresponding proteins of the CAMP (Malaysia) and MAD-20 (Papua New Guinea) isolates and more distantly related to those of the Wellcome (West Africa) and K1 (Thailand) isolates, supporting the proposed allelic dimorphism of gp195 within the parasite population. The prevalence of dimorphic sequences within the gp195 protein suggests that many gp195 epitopes would be group-specific. Despite the extensive differences in amino acid sequence between gp195 proteins of these two groups, the hydropathy profiles of proteins representative of both groups are very similar. The conservation of overall secondary structure shown by the hydropathy profile comparison indicates that gp195 proteins of the various P. falciparum isolates are functionally equivalent. This information on the primary structure of the FUP gp195 protein will enable us to evaluate the possible roles of conserved, group-specific and variable epitopes in immunity to the blood stage of the malaria parasite.

A seroepidemiological study to evaluate the role of passive maternal immunity to malaria in infants

Parasitaemias and loss of natural antibody to Plasmodium falciparum were studied in 104 infants in a highly endemic area of Papua New Guinea. There were 4 cases of congenital infection. Most infants lost malaria-specific immunoglobulin G (IgG) between 4 and 7 months (median = 21 weeks). 73% of heavy infections developed in infants without detectable antimalarial IgG. Infections in the presence of antimalarial IgG were asymptomatic and had scanty parasitaemias.

Propagation of Malaria Parasites In Vitro

Publisher Summary This chapter discusses the critical and concise review of the current status of the in vitro propagation of invertebrate and vertebrate inhabiting stages of malarial parasites. Exflagellation and fertilization can be readily observed in vitro by placing a drop of parasitized blood containing gametocytes onto a microscope slide. The fact that gametogenesis readily takes place in vivo but fertilization does not always result indicates that the ability of gametocytes to exflagellate in vitro is not a good indicator of their maturity. In the mosquito, the ookinete migrates through the gut epithelium into the hemocoel, where it lodges and undergoes differentiation as an oocyst. In vivo, it has been observed that once released from the oocyst, the sporozoites must go through a period of maturation. The in vitro cultivation of gametocytogenesis of any species of malarial parasites has not yet been accomplished. Limited success has been achieved toward the in vitro cultivation of the subsequent stages of sporogony, leading from oocyst differentiation to the formation of mature, infective sporozoites.

Lymphocyte isolation, rosette formation, and mitogen stimulation in rhesus monkeys

Over 70% of rhesus monkey peripheral mononuclear cells were isolated on sodium metrizoate-ficoll gradients with greater than 98% purity. Rhesus blood contained 47.8% active E, 58.2% total E, and 30.2% EAC rosette forming cells. Optimal conditions for mitogen studies were determined using phytohemagglutinin, concanavalin A, pokeweed mitogen, lipopolysaccharide and streptolysin O.

Susceptibility of a new world monkey (Aotus trivirgatus) to an old world simian malarial parasite (Plasmodium knowlesi)

Abstract Blood induced infections with a strain of Plasmodium knowlesi have been established in Aotus trivirgatus monkeys. To date we have completed 6 serial passages in intact Aotus monkeys starting with a fresh blood infection. Aotus monkeys were also easily infected with parasitized blood frozen at −70°C. for up to 11 weeks. The course of P. knowlesi infection in most of the Aotus monkeys was similar to that occurring in rhesus monkeys.

Role of Adjuvants in Malaria Vaccines

It is becoming widely recognized that the development of successful malaria vaccines will depend on both the production of protective antigens by chemical synthesis or recombinant technology as well as the availability of safe and effective adjuvants. The need for a strong adjuvant is evident from the results of vaccination experiments carried out in various laboratories over the last one decade. Direct immunization of monkeys with Plasmodium falciparum antigen in conjunction with various adjuvants has shown varying degrees of protection against a lethal P. falciparum challenge. However, the best protection has been achieved only when monkeys were immunized in conjunction with Freund’s complete adjuvant (FCA). Results of clinical trials with sporozoite and merozoite vaccines with alum as an adjuvant have shown limited immunogenicity and protective success. Since FCA is unacceptable for clinical use and results of clinical trials with alum have shown limited success, several laboratories have sought to establish a safe and effective adjuvant. Based on knowledge generated on the immunology of malaria, it is becoming evident that a malaria vaccine adjuvant must be capable of inducing both humoral and cell mediated immune responses. Availability of synthetic and recombinant polypeptides as malaria antigen along with a variety of novel adjuvants have accelerated research towards the establishment of safe and effective adjuvants for malaria vaccines Immunogenicity studies have identified a few synthetic immunomodulators which may be of potential clinical usage as adjuvants for malaria vaccines.

Plasmodium falciparum Merozoite Vaccine in Aotus Monkeys An Evaluation of the Tolerableness of Three Types of Adjuvants

Eight Aotus trivirgatus monkeys were included in a comparative study of adverse reactions caused by 3 types of malarial vaccine. The antigen consisted of late schizonts and merozoites combined with each of 3 adjuvants, a lipoidal amine CP-20,961, a muramyl dipeptide derivative [B30]-MDP, or Freunds complete adjuvant (FCA); 3 monkeys received CP-20,961 and 3 [B30]-MDP, whereas the FCA vaccine was given to 2 animals. All the vaccine combinations caused some adverse reactions, but their nature and intensity varied. Oedema, petechiae and/or ecchymosis, induration, and formation of fluctuating masses at the injection site were the local reactions observed, but except for the FCA group, all reactions disappeared without notable sequelae. Fever was the commonest general reaction, being mildest in the [B30]-MDP group. A transient increase (about 50%) in leukocyte count and a decrease (15-20%) in red blood cell count were observed in all animals, regardless of the type of adjuvant or whether it was a primary or booster injection in question. However, because the vaccine using [B30]-MDP evoked the mildest reactions, this may be the one most promising for future human studies if its protective efficacy is proven.

Differentiating owl monkey T and B lymphocytes by rosette formation.

Subpopulations of Aotus trivirgatus lymphocytes can be differentiated by the technique of rosette formation. Owl monkey B cells can be identified by erythrocyte-antibody-complement (EAC) rosette formation, since over 90% of EAC rosette-forming cells (RFC) have detectable surface immunoglobulins. A separate population of Aotus lymphocytes forms rosettes directly with sheep erythrocytes (E rosettes) and appears to be thymus-dependent (T cells) because (1) E rosette formation is blocked by anti-thymus serum, (2) over 80% of owl monkey thymocytes form E rosettes, and (3) most E RFC lack detectable surface immunoglobulins.