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D J Conway

Publications and source records attributed to D J Conway.

At least 19 recordsLinked to original sources
PubMed2002

Measuring immune selection.

Immune responses that kill pathogens or reduce their reproductive rate are generally important in protecting hosts from infection and disease. Pathogens that escape the full impact of such responses will survive, and any heritable genetic basis of this evasion will be selected. Due to the memory component of vertebrate immune responses, pathogens with rare alleles of a target antigen can have an advantage over those with common alleles, leading to the maintenance of a polymorphism. At the genetic level, there ought to be detectable signatures of balancing selection in the genes encoding these antigens. Here, methods for identifying these selective signatures are reviewed. Their practical utility for identifying which antigens are targets of protective immune responses is discussed.

Animals↗
PubMed2001

Efficacy of RTS,S/AS02 malaria vaccine against Plasmodium falciparum infection in semi-immune adult men in The Gambia: a randomised trial.

BACKGROUND: RTS,S/AS02 is a pre-erythrocytic malaria vaccine based on the circumsporozoite surface protein of Plasmodium falciparum fused to HBsAg, incorporating a new adjuvant (AS02). We did a randomised trial of the efficacy of RTS,S/AS02 against natural P. falciparum infection in semi-immune adult men in The Gambia. METHODS: 306 men aged 18-45 years were randomly assigned three doses of either RTS,S/AS02 or rabies vaccine (control). Volunteers were given sulfadoxine/pyrimethamine 2 weeks before dose 3, and kept under surveillance throughout the malaria transmission season. Blood smears were collected once a week and whenever a volunteer developed symptoms compatible with malaria. The primary endpoint was time to first infection with P. falciparum. Analysis was per protocol. FINDINGS: 250 men (131 in the RTS,S/AS02 group and 119 in the control group) received three doses of vaccine and were followed up for 15 weeks. RTS,S/AS02 was safe and well tolerated. P. falciparum infections occurred significantly earlier in the control group than the RTS,S/AS02 group (Wilcoxon's test p=0.018). Vaccine efficacy, adjusted for confounders, was 34% (95% CI 8.0-53, p=0.014). Protection seemed to wane: estimated efficacy during the first 9 weeks of follow-up was 71% (46-85), but decreased to 0% (-52 to 34) in the last 6 weeks. Vaccination induced strong antibody responses to circumsporozoite protein and strong T-cell responses. Protection was not limited to the NF54 parasite genotype from which the vaccine was derived. 158 men received a fourth dose the next year and were followed up for 9 weeks; during this time, vaccine efficacy was 47% (4-71, p=0.037). INTERPRETATION: RTS,S/AS02 is safe, immunogenic, and is the first pre-erythrocytic vaccine to show significant protection against natural P. falciparum infection.

Adult↗
PubMed2001

Population genetic analysis of the Plasmodium falciparum erythrocyte binding antigen-175 (eba-175) gene.

The Plasmodium falciparum erythrocyte binding antigen-175 gene (eba-175) has highly divergent allelic segments (Cseg and Fseg) in one part of the gene (region III), but only a small number of single nucleotide polymorphisms (SNPs) in the rest of the sequence. Here, evidence for the possible importance of the Cseg/Fseg dimorphism was sought in a molecular population genetic analysis of the gene. First, allele frequency distributions were determined for the Cseg/Fseg dimorphism and five SNPs in a sample of five populations in Africa. The inter-population variance in frequencies was higher for Cseg/Fseg (F(ST)=0.18) than for the SNPs (F(ST) values from 0.03 to 0.10), but these values were entirely dependent on the inclusion of one particularly divergent population (Sudan). Second, linkage disequilibrium was measured among the intragenic loci. There was the expected trend of declining linkage disequilibrium with increasing molecular distance, but it is notable that the Cseg allele was in absolute linkage disequilibrium with the two flanking SNPs, whereas the Fseg allele was associated with a broader range of SNP haplotypes. Finally, there was no association between the Cseg/Fseg alleles of eba-175 in parasites and the M/N alleles of the glycophorin A erythrocyte receptor in the human subjects.

Africa↗
PubMed2001

Extreme geographical fixation of variation in the Plasmodium falciparum gamete surface protein gene Pfs48/45 compared with microsatellite loci.

Comparing patterns of genetic variation at multiple loci in the genome of a species can potentially identify loci which are under selection. The large number of polymorphic microsatellites in the malaria parasite Plasmodium falciparum are available markers to screen for selectively important loci. The Pfs48/45 gene on Chromosome 13 encodes an antigenic protein located on the surface of parasite gametes, which is a candidate for a transmission blocking vaccine. Here, genotypic data from 255 P. falciparum isolates are presented, which show that alleles and haplotypes of five single nucleotide polymorphisms (SNPs) in the Pfs48/45 gene are exceptionally skewed in frequency among different P. falciparum populations, compared with alleles at 11 microsatellite loci sampled widely from the parasite genome. Fixation indices measuring inter-population variance in allele frequencies (F(ST)) were in the order of four to seven times higher for Pfs48/45 than for the microsatellites, whether considered (i) among populations within Africa, or (ii) among different continents. Differing mutational processes at microsatellite and SNP loci could generally affect the population structure at these different types of loci, to an unknown extent which deserves further investigation. The highly contrasting population structure may also suggest divergent selection on the amino acid sequence of Pfs48/45 in different populations, which plausibly indicates a role for the protein in determining gamete recognition and compatibility.

Africa↗
PubMed2001

Polymorphisms in the IkappaB-alpha promoter region and risk of diseases involving inflammation and fibrosis.

The transcription factor NFkappaB regulates inflammatory and other cellular responses. In non-stimulated cells, NFkappaB is linked to its inhibitor IkappaB, which plays a major role in controlling NFkappaB activity. Here, the gene promoter region of the major inducible IkappaB component (IkappaB-alpha) was studied to identify single nucleotide polymorphisms (SNPs), and to test if these are associated with risk of two diseases involving inflammation and fibrosis (trachoma and silicosis). Three SNPs were identified at positions -881, -826 and -297 relative to the transcription start site. The position -297 is close to two NFkappaB binding sites, kappaB2 and kappaB3, but the alleles were not associated with either disease. Alleles at positions -881 and -826 were in complete linkage disequilibrium with each other, and the rare haplotype was significantly less frequent among patients with trachoma compared to controls, although there was no difference in frequencies between silicosis patients and controls.

Base Sequence↗
PubMed2001

Strong diversifying selection on domains of the Plasmodium falciparum apical membrane antigen 1 gene.

The surface-accessible ectodomain region of the Plasmodium falciparum apical membrane antigen 1 (AMA1) is a malaria vaccine candidate. The amino acid sequence may be under selection from naturally acquired immune responses, and previous analyses with a small number of allele sequences indicate a non-neutral pattern of nucleotide variation. To investigate whether there is selection to maintain polymorphism within a population, and to identify the parts of the ectodomain under strongest selection, a sample of 51 alleles from a single endemic population was studied. Analyses using Fu and Li's D and F tests, Tajima's D test, and the McDonald-Kreitman test (with the chimpanzee parasite P. reichenowi as outgroup) show significant departure from neutrality and indicate the selective maintenance of alleles within the population. There is also evidence of a very high recombination rate throughout the sequence, as estimated by the recombination parameter, C, and by the rapid decline in linkage disequilibrium with increasing nucleotide distance. Of the three domains (I-III) encoding structures determined by disulfide bonds, the evidence of selection is strongest for Domains I and III. We predict that these domains in particular are targets of naturally acquired protective immune responses in humans.

Algorithms↗
PubMed2000

Polymorphisms in candidate genes and risk of scarring trachoma in a Chlamydia trachomatis--endemic population.

Genes involved in regulating antimicrobial immunity and inflammation may modulate the risk of tissue scarring and fibrosis in chlamydial diseases such as trachoma. By use of a large case-control study of scarring trachoma in The Gambia, the importance of single-nucleotide polymorphisms in several candidate genes was investigated. Overall, no significant differences were found between patients and control subjects in genotype frequencies for polymorphisms in cytokine promoters interleukin (IL)-10 (positions -1082, -819, -592), IL-4 (-590), or tumor necrosis factor-alpha (-376) or for codon 57 of the mannose-binding protein. Among the ethnic groups in the study, Mandinkas had the highest frequency of the IL-10-1082G allele (0.36). Within this ethnic group, the IL-10-1082G homozygote genotype was significantly more common among case patients than control subjects (odds ratio, 5.10; 95% confidence interval, 1.24-24.2; P=.009). This single association is consistent with data indicating that the IL-10-1082G allele is associated with higher levels of IL-10 transcription and that Th2-type immune responses are associated with risk of chlamydial disease.

Adolescent↗
PubMed2000

High-throughput sequence typing of T-cell epitope polymorphisms in Plasmodium falciparum circumsporozoite protein.

We report a method for typing polymorphisms at the T-cell epitopes within the Th2R and Th3R regions of the Plasmodium falciparum circumsporozoite protein (CSP). This method combines the use of PCR and sequence specific oligonucleotide probes (PCR-SSOP), and allows the identification of single nucleotide polymorphisms in these epitope regions. PCR-SSOP is a robust and a high-throughput sequence typing technique which has the same specificity and fidelity as direct sequencing. This method has been developed specifically for the assessment of the protective efficacy of RTS,S/SBAS2 vaccine against the 3D7 strain of P. falciparum (RTS,S/SBAS2 vaccine contains a part of the 3D7 CSP protein) in a phase IIb trial in Gambia which has been completed recently. PCR-SSOP could be used to determine the allelic frequencies of other parasite antigens and their geographical distribution.

Alleles↗
PubMed2000

Micro-evolution and emergence of pathogens.

Changes in the epidemiology of infectious diseases are the direct result of ecological and evolutionary changes in hosts and parasites. Precisely what the causal processes are is rarely known in any particular case, and this hinders the design of appropriate control strategies. This is particularly so for emerging infections, as opportunity is rapidly lost to study the ecological parameters which might have affected initial emergence. However, molecular evolutionary studies of the pathogens can yield data which discriminate between possible causes. The current distribution of DNA sequence variation is important information which may reveal past and current changes in pathogen population structures, and can also identify adaptive changes in pathogen genes which have affected their evolution. Such studies have been quite intensively performed on particular viral and bacterial pathogens, and some of the successes of these are noted here. Approaches to understanding the recent evolution of eukaryotic pathogens are outlined, with particular reference to current problems of emerging zoonoses, and changes in virulence and drug resistance.

Animals↗
PubMed2000

Molecular characterisation of Plasmodium reichenowi apical membrane antigen-1 (AMA-1), comparison with P. falciparum AMA-1, and antibody-mediated inhibition of red cell invasion.

Apical membrane antigen 1 is a candidate vaccine component for malaria. It is encoded by a single copy gene and has been characterised in a number of malaria species as either an 83-kDa de novo product (Plasmodium falciparum; Pf AMA-1) or a 66-kDa product (all other species). All members of the AMA-1 family are expressed during merozoite formation in maturing schizonts and are initially routed to the rhoptries. Processed forms may subsequently be associated with the merozoite surface. Because of the unique occurrence of the 83-kDa form in P. falciparum we were interested to determine whether the phylogenetically closely related chimpanzee malaria Plasmodium reichenowi shared characteristics with Pf AMA-1. Here we show that the molecular structure, the localisation and processing are similar to that of Pf AMA-1 and that in vitro growth inhibitory mAbs reactive with Pf AMA-1 also inhibit P. reichenowi growth in an in vitro assay. Polymorphism in the 83-kDa AMA-1 family was analysed through comparison of Pr ama-1 with Pf ama-1 alleles, which showed the most significant evidence for selection maintaining polymorphism in Domains I-III of AMA-1 in P. falciparum. The most substantial divergence between Pr AMA-1 and Pf AMA-1 sequences was in the N-terminal region unique to the 83-kDa form of AMA-1. It was confirmed that the specific Pr ama-1-type allele was not present among P. falciparum parasites in an African population, and an allele coding for lysine at amino acid 187 was uniquely associated with field isolates in this population.

Adolescent↗
PubMed2000

Origin of Plasmodium falciparum malaria is traced by mitochondrial DNA.

The origin and geographical spread of Plasmodium falciparum is here determined by analysis of mitochondrial DNA sequence polymorphism and divergence from its most closely related species P. reichenowi (a rare parasite of chimpanzees). The complete 6 kb mitochondrial genome was sequenced from the single known isolate of P. reichenowi and from four different cultured isolates of P. falciparum, and aligned with the two previously derived P. falciparum sequences. The extremely low synonymous nucleotide polymorphism in P. falciparum (pi=0.0004) contrasts with the divergence at such sites between the two species (kappa=0.1201), and supports a hypothesis that P. falciparum has recently emerged from a single ancestral population. To survey the geographical distribution of mitochondrial haplotypes in P. falciparum, 104 isolates from several endemic areas were typed for each of the identified single nucleotide polymorphisms. The haplotypes show a radiation out of Africa, with unique types in Southeast Asia and South America being related to African types by single nucleotide changes. This indicates that P. falciparum originated in Africa and colonised Southeast Asia and South America separately.

Africa↗
PubMed2000

Experimental and molecular genetic analysis of the impact of pyrethroid and non-pyrethroid insecticide impregnated bednets for mosquito control in an area of pyrethroid resistance.

Experimental huts in Côte d'Ivoire were used to evaluate the pyrethroid alpha-cypermethrin, the non-ester pyrethroid etofenprox, the organophosphate pirimiphos-methyl and the carbamate carbosulfan on bednets against pyrethroid-resistant Anopheles gambiae Giles. To test for selection for the resistance gene by the treated nets, A. gambiae collected live or dead from the huts were kept and analysed for the presence of the kdr gene using a new polymerase chain reaction followed by sequence-specific oligonucleotide probing (PCR-SSOP) for kdr-genotyping. Deliberately holed bednets freshly treated with pirimiphos-methyl or carbosulfan caused over 90% kill of A. gambiae s.s. and Culex spp. However, the mortality with alpha-cypermethrin or etofenprox treated nets was similar to that with untreated nets. Bloodfeeding of A. gambiae s.s. on the sleepers under the nets was only significantly reduced by alpha-cypermethrin and carbosulfan. Tests of the residual activity of the bednets after seven months showed that pirimiphos-methyl had lost its efficacy while carbosulfan still performed well. Once again the pyrethroid treated nets gave similar results to the untreated nets. Selection for the kdr-allele by alpha-cypermethrin and etofenprox, but not by carbosulfan, was indicated by PCR-SSOP genotyping of mosquitoes. Thus carbamates such as carbosulfan, or organophosphates of longer persistence than pirimiphos-methyl and of low mammalian toxicity, would seem to be a promising alternative to be used on bednets, particularly in areas of pyrethroid resistance.

Adult↗
PubMed2000

A principal target of human immunity to malaria identified by molecular population genetic and immunological analyses.

New strategies are required to identify the most important targets of protective immunity in complex eukaryotic pathogens. Natural selection maintains allelic variation in some antigens of the malaria parasite Plasmodium falciparum. Analysis of allele frequency distributions could identify the loci under most intense selection. The merozoite surface protein 1 (Msp1) is the most-abundant surface component on the erythrocyte-invading stage of P. falciparum. Immunization with whole Msp1 has protected monkeys completely against homologous and partially against non-homologous parasite strains. The single-copy msp1 gene, of about 5 kilobases, has highly divergent alleles with stable frequencies in endemic populations. To identify the region of msp1 under strongest selection to maintain alleles within populations, we studied multiple intragenic sequence loci in populations in different regions of Africa and Southeast Asia. On both continents, the locus with the lowest inter-population variance in allele frequencies was block 2, indicating selection in this part of the gene. To test the hypothesis of immune selection, we undertook a large prospective longitudinal cohort study. This demonstrated that serum IgG antibodies against each of the two most frequent allelic types of block 2 of the protein were strongly associated with protection from P. falciparum malaria.

Africa↗
PubMed2000

Analysis of human antibodies to erythrocyte binding antigen 175 of Plasmodium falciparum.

Invasion of human erythrocytes by Plasmodium falciparum merozoites is a multistep process. For many strains of the parasite, part of this process requires that the erythrocyte binding antigen 175 (EBA-175) of the merozoite binds to sialic acid residues of glycophorin A on the erythrocyte surface, a receptor-ligand interaction which represents a potential target for inhibition by antibodies. This study characterizes the reactivity of naturally acquired human antibodies with four recombinant proteins representing parts of EBA-175 (region II, regions III to V, and the dimorphic C and F segment region) in populations in which the organism is endemic. Serum immunoglobulin G (IgG) recognizing the recombinant proteins is predominantly of the IgG1 and IgG3 subclasses, and its prevalence increases with age. In a large population study in The Gambia, serum positivity for IgG or IgG1 and IgG3 subclass antibodies to each of the EBA-175 recombinant antigens was not significantly associated with subsequent protection from clinical malaria. However, there was a trend indicating that individuals with high levels of IgG to region II may have some protection.

Adult↗
PubMed1999

High recombination rate in natural populations of Plasmodium falciparum.

Malaria parasites are sexually reproducing protozoa, although the extent of effective meiotic recombination in natural populations has been debated. If meiotic recombination occurs frequently, compared with point mutation and mitotic rearrangement, linkage disequilibrium between polymorphic sites is expected to decline with increasing distance along a chromosome. The rate of this decline should be proportional to the effective meiotic recombination rate in the population. Multiple polymorphic sites covering a 5-kb region of chromosome 9 (the msp1 gene) have been typed in 547 isolates from six populations in Africa to test for such a decline and estimate its rate in populations of Plasmodium falciparum. The magnitude of two-site linkage disequilibrium declines markedly with increasing molecular map distance between the sites, reaching nonsignificant levels within a map range of 0.3-1.0 kb in five of the populations and over a larger map distance in the population with lowest malaria endemicity. The rate of decline in linkage disequilibrium over molecular map distance is at least as rapid as that observed in most chromosomal regions of other sexually reproducing eukaryotes, such as humans and Drosophila. These results are consistent with the effective recombination rate expected in natural populations of P. falciparum, predicted on the basis of the underlying molecular rate of meiotic crossover and the coefficient of inbreeding caused by self-fertilization events. This is conclusive evidence to reject any hypothesis of clonality or low rate of meiotic recombination in P. falciparum populations. Moreover, the data have major implications for the design and interpretation of population genetic studies of selection on P. falciparum genes.

Africa↗