Biomedical subjects

D E Arnot

Publications and source records attributed to D E Arnot.

47 records · Page 3Linked to original sources

PubMed1987

Expression in yeast of a Plasmodium vivax antigen of potential use in a human malaria vaccine.

DNA coding for 234 amino acids of the circumsporozoite (CS) protein of Plasmodium vivax was incorporated into yeast expression vectors. The DNA encoded all the repeat domain and codons for a highly conserved sequence, KLKQP, found in CS proteins from all malaria parasites. Yeast cells transformed with these autonomously replicating plasmids expressed, upon induction, high levels of the CS polypeptide. The malaria antigen was purified in good yields from yeast extracts and was injected into mice using alum as adjuvant. The antibodies recognized the authentic CS protein, and at high dilutions, they inhibited the invasion of hepatocytes by sporozoites in vitro.

Animals↗

PubMed1987

The circumsporozoite gene of the Plasmodium cynomolgi complex.

An analysis of the circumsporozoite (CS) genes of six closely related plasmodia is presented. Like other plasmodial antigens, the CS protein contains tandem repeats flanked by conventional nonrepeated sequences. Our analysis shows that the repeats, which encode the immunodominant epitope of the CS protein, diverge more rapidly than the remainder of the gene, and that the maintenance and evolution of the repeats cannot be explained as the result of selection at the protein level. We argue that a mechanism acts directly on the DNA sequence to constrain the internal divergence of the repeats, and as a result promotes their rapid divergence between taxa.

Amino Acid Sequence↗

PubMed1986

Circumsporozoite protein of Plasmodium berghei: gene cloning and identification of the immunodominant epitopes.

The gene encoding the circumsporozoite (CS) protein of the rodent malaria parasite Plasmodium berghei was cloned and characterized. A cDNA library made from P. berghei sporozoite RNA was screened with a monoclonal antibody for expression of CS protein epitopes. The resulting cDNA clone was used to isolate the CS protein gene from a lambda library containing parasite blood-stage DNA. The CS protein gene contains a central region encoding two types of tandemly repeated amino acid units, flanked by nonrepeated regions encoding amino- and carboxy-terminal signal and anchorlike sequences, respectively. One of the central repeated amino acid unit types contains the immunodominant epitopes.

Amino Acid Sequence↗

PubMed1985

Circumsporozoite protein of Plasmodium vivax: gene cloning and characterization of the immunodominant epitope.

The gene encoding the circumsporozoite (CS) protein of the human malaria parasite Plasmodium vivax has been cloned. The deduced sequence of the protein consists of 373 amino acids with a central region of 19 tandem repeats of the nonapeptide Asp-Arg-Ala-Asp/Ala-Gly-Gln-Pro-Ala-Gly. A synthetic 18-amino acid peptide containing two tandem repeats binds to a monoclonal antibody directed to the CS protein of Plasmodium vivax and inhibits the interaction of this antibody with the native protein in sporozoite extracts. The portions of the CS gene that do not contain repeats are closely related to the corresponding regions of the CS genes of two simian malarias, Plasmodium cynomolgi and Plasmodium knowlesi. In contrast, the homology between the CS genes of Plasmodium vivax and Plasmodium falciparum, another malaria parasite of humans, is very limited.

Amino Acid Sequence↗

PubMed1984

DNA cloning of Plasmodium falciparum circumsporozoite gene: amino acid sequence of repetitive epitope.

A clone of complementary DNA encoding the circumsporozoite (CS) protein of the human malaria parasite Plasmodium falciparum has been isolated by screening an Escherichia coli complementary DNA library with a monoclonal antibody to the CS protein. The DNA sequence of the complementary DNA insert encodes a four-amino acid sequence: proline-asparagine-alanine-asparagine, tandemly repeated 23 times. The CS beta-lactamase fusion protein specifically binds monoclonal antibodies to the CS protein and inhibits the binding of these antibodies to native Plasmodium falciparum CS protein. These findings provide a basis for the development of a vaccine against Plasmodium falciparum malaria.

Amino Acid Sequence↗

PubMed1981

Biochemical identification of cutaneous leishmanias by analysis of kinetoplast DNA. I. Ultrastructural and buoyant density analysis.

Ultrastructural analysis has been carried out on three Leishmania isolates which are proven causal agents of human cutaneous Leishmaniasis, L. tropica major, L. aethiopica and a unidentified species, Leishmania SP48. No significant differences in submicroscopic morphology have been found in thin-sectioned organisms from the three isolates. Extensive plate cristae have been observed within the mitochondria and connections between the rim of the kinetoplast nucleoid and the inner mitochondrial membrane noted. Kinetoplast DNA (kDNA) has been isolated from these isolates and from L. tarentolae and examined by protein monolayer spreading and darkfield electronmicroscopy. The basic molecular arrangement of isolated kDNA in the form of 5 micrometers networks of 0.28--0.3 micrometer mini-circles with long looped DNA in the interior and at the periphery of networks is similar in all isolates. Minor differences between L. aethiopica and SP48 compared with L. tropica major have been observed. The kDNAs of L. aethiopica and SP48 are identical morphologically. Buoyant density analysis has shown that kDNA from L. aethiopica and SP48 have identical values and these are different from the values for L. tropica major. The finding of similar buoyant densities for kDNA from L. tropica major and L. tarentolae also imply a sequence homology by this criteria which is refuted by the results given in the following paper. The results given in this and the following paper (Arnot, D.E. and Barker, D.C.(1981) Mol. Biochem. Parasitol. 3, 47--56 indicate that the unknown Leishmania SP48 is very closely related to, if not identical with, L. aethiopica. This finding is consistent with the clinical and ecological facts known for the organism SP48.

Cell Nucleus↗

PubMed1981

Biochemical identification of cutaneous leishmanias by analysis of kinetoplast DNA. II. Sequence homologies in Leishmania kDNA.

Kinetoplast DNA (kDNA) has been isolated from the human cutaneous Leishmania isolates, L. tropica major, L. aethiopica and an unknown Kenyan isolate, Leishmania SP48. DNA sequence relationships among these isolates have been studied by restriction enzyme digestion and two phase hybridisation to Southern blots of kDNA covalently coupled to diazobenzyloxymethyl (DBM) paper. The results of this analysis confirm that rapid kDNA sequence evolution is occurring in the Old World leishmanias although some sequence conservation in defined regions of the mini-circle sequence is present. These results emphasise the danger of constructing a rigid Leishmania classification on buoyant density data alone. The covalent binding of kDNA electrophoretic separations to DBM paper permits the construction of a DNA sequence "library' which can be used in the classification and diagnosis of unknown Leishmania isolates.

Base Sequence↗

PubMedDate not supplied

Genetic diversity of Plasmodium falciparum in a village in eastern Sudan. 1. Diversity of enzymes, 2D-PAGE proteins and antigens.

Twenty-nine plasmodium falciparum isolates from patients in Asar village, eastern Sudan, were characterized for variation in 18 different genetically controlled characters, including iso-enzymes, proteins detected by two-dimensional polyacrylamide gel electrophoresis and blood-stage antigens. Considerable allelic diversity in the genes determining these characters was detected. Each isolate contained genetically distinct parasites. Fifteen individuals were infected with more than one parasite genotype. The diversity of parasite types is most probably generated by recombination during mosquito transmission of mixed parasite clones.

Animals↗

PubMedDate not supplied

Genetic diversity of Plasmodium falciparum in a village in eastern Sudan. 2. Drug resistance, molecular karyotypes and the mdr1 genotype of recent isolates.

Isolates of Plasmodium falciparum from a Sudanese village have been collected as part of a study of parasite genetic diversity during seasonal malaria epidemics. The sensitivity in vitro to chloroquine, pyrimethamine and mefloquine of these isolates has been determined. To assess the utility of pulse field gel chromosome separations in isolate characterization, 18 samples from individual patients in a single village were studied using this technique. Extensive variation in chromosome size was detected, no 2 isolates having identical molecular karyotypes. No multidrug resistance (mdr) gene amplification polymorphisms were detected in either chloroquine-resistant or chloroquine-sensitive isolates in this sample.

Animals↗

PubMedDate not supplied

Drug response and genetic characterization of Plasmodium falciparum clones recently isolated from a Sudanese village.

We have isolated 20 clones of Plasmodium falciparum from isolates from patients attending a village clinic in Sudan during 10 d in October-November 1989. The clones were genetically diverse, having highly variable molecular karyotypes and a wide range of drug responses. Chloroquine-sensitive (50% inhibitory concentration [IC50] in the 4-15 nM range) and chloroquine-resistant clones (IC50 in the 40-95 nM range) co-existed in the population, but no obvious amplification of the P-glycoprotein homologue gene, Pgh1 (previously known as the multi-drug resistance gene, mdr1) marked the chloroquine-resistant clones. Chloroquine resistance was reversible by verapamil in these clones, although they varied in their susceptibility to verapamil alone. These observations indicate that the biochemical characteristics of the Sudanese chloroquine-resistant P. falciparum are similar to those reported from south-east Asian and Latin American isolates, which is consistent with there being a similar molecular basis for this phenomenon.

Animals↗

PubMedDate not supplied

The epidemiology of febrile malaria episodes in an area of unstable and seasonal transmission.

This study investigated the epidemiology of uncomplicated falciparum malaria in an area of unstable and seasonal transmission in eastern Sudan. About 90% of malaria morbidity in this region occurs in the months of September to November, and very few malaria cases occur during the intensely arid Sudanese dry season and during years of drought. The malaria situation in the study site, the village of Daraweesh, was analysed during 3 consecutive malaria seasons in 1993-95 during which the 457 inhabitants suffered at total of 436 episodes of falciparum malaria. Using an Andersen-Gill proportional hazard model for recurrent events stratified by family, we have calculated the relative hazard for clinical malaria episodes by age, sex, haemoglobin genotype, blood type and infection in the previous season. The malaria risk was significantly lower in individuals aged 20-88 years than in the 5-19 years age-group. The relative protection due to adulthood varied between seasons (relative risk, RR, 0x34 to 0x67). Serological data were not consistent with the hypothesis that the age difference in incidence was due to differences in exposure. During the 1993 season the malaria incidence in males was lower than in females (RR = 0x75), during the 1994 season the incidences were comparable, whereas males had an increased risk of malaria in 1995 (RR = 1x87). The relative risk in individuals carrying the haemoglobin AS genotype compared to homozygous AA individuals was 0x57.

Adolescent↗