Biology subjects

Conway, D.

Publications and source records attributed to Conway, D..

11 recordsLinked to original sources

bioRxiv2026

Plasmodium falciparum invasion ligand gene transcript profiles in different populations

Plasmodium falciparum merozoites invade erythrocytes using various ligand-receptor interactions. Important ligands encoded by the eba and Rh gene families have varying expression levels in different parasite isolates, affecting their vaccine candidacy. Analyses of clinical isolates from endemic areas in Africa have indicated that most variation in these expression profiles exists within each local area, and only minor differences are seen between areas, although comparisons with non-African populations have not previously been performed. To enable this, relative transcript levels of three eba genes and five Rh genes have been analysed in new population samples, Malaysian isolates sampled from Sabah State in Borneo prior to endemic malaria elimination, and Gambian isolates, cultured under the same conditions to harvest schizonts for reverse transcription quantitative PCR assays. Significant differences between these populations were seen for three of the ligand genes, levels of eba175 being higher in Malaysia, while levels of eba181 and Rh2b were lower in Malaysia. The gene transcript profiles did not differ between single genotype and or multiple-genotype isolates. The distinctness of the Malaysian population expression profile was also supported by comparing previous data on clinical isolates from Ghana. In tests for correlation with previously determined parasite multiplication rates, eba181 transcript levels correlated positively among Malaysian isolates but not among Gambian isolates. These findings suggest that expression of three P. falciparum merozoite ligands involved in invasion may be regionally differentiated, and further analysis of Asian parasite populations would be important if vaccines based on these candidates are to be considered for future use.

microbiology↗

bioRxiv2025

New insights on malaria parasite adaptation yielded by samples archived for more than 50 years

Understanding evolution of human pathogens requires looking beyond effects of recent interventions or known epidemiological changes. To study malaria parasites prior to widespread drug selection, Plasmodium falciparum genome sequences were analysed from the oldest set of archived research samples yet identified, placental blood collected in the Gambia between 1966 and 1971. Genomic complexity within infections was high, infections were genetically unrelated, and no drug resistance alleles were detected. Strong signatures of positive selection are clearly seen at multiple loci throughout the genome, most of which encode surface proteins that bind erythrocytes and are targets of acquired antibody responses. Comparison of population samples obtained over a following period of almost 50 years revealed major directional allele frequency changes at several loci apart from drug resistance genes. Exceptional changes over this time are seen at gdv1 that regulates the rate of parasite sexual conversion required for transmission, and at the unlinked Pfsa1 and Pfsa3 loci previously associated with infection of individuals with sickle-cell trait. Other affected loci encode surface and transporter proteins warranting targeted functional analyses. This identification of key long-term adaptions that have not reached equilibrium is important for understanding and potentially managing future evolution of malaria parasites.

microbiology↗

bioRxiv2024

Intrinsic multiplication rate variation of Plasmodium falciparum (in clinical isolates prior to elimination in Malaysia

Replication rates and virulence of pathogens are hypothesised to evolve in response to varying intensity of transmission and competition among genotypes. Under exponential growth conditions in culture, clinical isolates of the malaria parasite Plasmodium falciparum have variable intrinsic multiplication rates, but comparisons of samples from different areas are needed. To analyse parasites from an area of low endemicity, Malaysian clinical isolates cryopreserved prior to malaria elimination were studied. The mean and range of P. falciparum multiplication rates in Malaysian isolates was no less than that seen among isolates from more highly endemic populations in Africa, which does not support a hypothesis of adaptation to prevailing levels of infection endemicity. Moreover, the distribution of multiplication rates was similar between isolates with single parasite genotypes and those containing multiple genotypes, which does not support a hypothesis of facultative adjustment to competing parasites. Based on clinical isolates, the findings indicate that parasites may not evolve lower multiplication rates under conditions of reduced transmission, and that the virulence potential is likely to be undiminished in pre-elimination settings. This encourages efforts to eliminate endemic infection completely, as has been achieved at the national level in Malaysia.

microbiology↗

bioRxiv2024

Vimentin molecular linkages with nesprin-3 enhance nuclear deformation by cell geometric constraints

The nucleus is the organelle of the cell responsible for controlling protein expression, which has a direct effect on cellular biological functions. Here we show that the cytoskeletal protein vimentin plays an important role in increasing cell-generated forces transmitted to the cell nucleus, resulting in increased nuclear deformations in strongly polarized cells. Using micropatterned substrates to geometrically control cell shape in wild-type and vimentin-null cells, we show vimentin increases polarization and deformation of the cell nucleus. Loss of nesprin-3, which physically couples vimentin to the nuclear envelope, phenotypically copies the loss of vimentin, suggesting vimentin transmits forces to the cell nucleus through direct molecular linkages. Use of a fluorescence resonance energy transfer (FRET) sensor that binds to the nuclear envelope through lamin-A/C suggests vimentin increases the tension on the nuclear envelope. Our results indicate that nuclear shape and deformation can be modified by the vimentin cytoskeleton and its specific crosslinks to the cell nucleus.

biophysics↗

bioRxiv2024

Higher multiplication rates of Plasmodium falciparum in isolates from hospital cases compared with community infections

BackgroundParasite multiplication rates vary among Plasmodium falciparum isolates from patients with malaria, suggesting differences in virulence potential, although direct comparisons between hospital-based clinical cases and community infections are needed. MethodsCryopreserved blood samples from malaria cases presenting to a district hospital in The Gambia and infections detected in local communities were introduced to continuous culture under the same conditions. Thirty-four isolates (23 hospital-based and 11 community-based) were successfully established and then tested under exponential growth conditions over six days to derive estimated P. falciparum multiplication rates per cycle based on a 48-hour typical cycle length. ResultsA range of parasite multiplication rates in culture was seen across isolates, from 1.5-fold to 5.0-fold per cycle. Multiplication rates were significantly higher in the hospital-based isolates than the community-based isolates. There was a significantly positive correlation between parasitaemia in peripheral blood and multiplication rates in culture. There was no significant difference in multiplication rates between isolates with single or multiple parasite genotypes. ConclusionsThese findings are consistent with a hypothesis that intrinsic natural variation in parasite multiplication rate may affect levels of parasitaemia achieved during infection, and that this affects likelihood of hospital presentation. Results do not support a hypothesis that parasites modify their multiplication rates in response to competing parasites with different genotypes. SummaryRelevant to understanding parasite virulence, this study finds higher Plasmodium falciparum multiplication rates in cultured isolates from malaria cases presenting to hospital than in isolates from local community infections, and positive correlation with parasitaemia in peripheral blood of individuals.

microbiology↗

bioRxiv2024

A complex Plasmodium falciparum cryptotype circulating at low frequency across the African continent

The population structure of the malaria parasite Plasmodium falciparum can reveal underlying demographic and adaptive evolutionary processes. Here, we analyse population structure in 4,376 P. falciparum genomes from 21 countries across Africa. We identified a strongly differentiated cluster of parasites, comprising [~]1.2% of samples analysed, geographically distributed over 13 countries across the continent. Members of this cluster, named AF1, carry a genetic background consisting of a large number of highly differentiated variants, rarely observed outside this cluster, at a multitude of genomic loci distributed across most chromosomes. At these loci, the AF1 haplotypes appear to have common ancestry, irrespective of the sampling location; outside the shared loci, however, AF1 members are genetically similar to their sympatric parasites. AF1 parasites sharing up to 23 genomic co-inherited regions were found in all major regions of Africa, at locations over 7,000 km apart. We coined the term cryptotype to describe a complex common background which is geographically widespread, but concealed by genomic regions of local origin. Most AF1 differentiated variants are functionally related, comprising structural variations and single nucleotide polymorphisms in components of the MSP1 complex and several other genes involved in interactions with red blood cells, including invasion and erythrocyte antigen export. We propose that AF1 parasites have adapted to some as yet unidentified evolutionary niche, by acquiring a complex compendium of interacting variants that rarely circulate separately in Africa. As the cryptotype spread across the continent, it appears to have been maintained mostly intact in spite of recombination events, suggesting a selective advantage. It is possible that other cryptotypes circulate in Africa, and new analysis methods may be needed to identify them.

genomics↗

bioRxiv2023

Expression of the MSPDBL2 antigen in a discrete subset of Plasmodium falciparum schizonts is regulated by GDV1 but not linked to sexual commitment

The Plasmodium falciparum merozoite surface protein MSPDBL2 is a polymorphic antigen targeted by acquired immune responses, and normally expressed in only a minority of mature schizonts. The potential relationship of MSPDBL2 to sexual commitment is examined, as variable mspdbl2 transcript levels and proportions of MSPDBL2-positive mature schizonts in clinical isolates have previously correlated with levels of many sexual stage parasite gene transcripts, although not with the master regulator ap2-g. It is demonstrated that conditional overexpression of GDV1, which promotes sexual commitment, also substantially increases the proportion of MSPDBL2-positive schizonts in culture. Conversely, truncation of the gdv1 gene is shown to prevent any expression of MSPDBL2. However, across diverse P. falciparum cultured lines the variable proportions of MSPDBL2 positivity in schizonts does not correlate significantly with variable gametocyte conversion rates, indicating it is not involved in sexual commitment. Confirming this, examining a line with endogenous HA-tagged AP2-G showed that the individual schizonts expressing MSPDBL2 are mostly different to those expressing AP2-G. Using a selection-linked integration system, modified P. falciparum lines were engineered to express an intact or disrupted version of MSPDBL2, showing the protein is not required for sexual commitment or early gametocyte development. Asexual parasite multiplication rates were also not affected by expression of either intact or disrupted MSPDBL2 in a majority of schizonts. Occurring alongside sexual commitment, the role of the discrete MSPDBL2-positive schizont subpopulation requires further investigation in natural infections where it is under immune selection.

microbiology↗

bioRxiv2023

Vinculin is Essential For Sustaining Normal Levels of Endogenous Force Transmission at Cell-Cell Contacts

Transmission of cell-generated (i.e., endogenous) tension at cell-cell contacts is crucial for tissue shape changes during morphogenesis and adult tissue repair in tissues like epithelia. E-cadherin-based adhesions at cell-cell contacts are the primary means by which endogenous tension is transmitted between cells. The E-cadherin-{beta}-catenin--catenin complex mechanically couples to the actin cytoskeleton (and thereby the contractile machinery of the cell) both directly and indirectly. However, the key adhesion constituents required for substantial endogenous force transmission at these adhesions in cell-cell contacts are unclear. Due to the role of -catenin as a mechanotransducer that recruits vinculin at cell-cell contacts, we expected -catenin to be essential for the high levels of force transmission normally sustained. Instead, using the traction force imbalance method to determine the inter-cellular force at a single cell-cell contact between cell pairs, we found that it is vinculin that is essential for high endogenous force transmission. Our results constrain the potential mechanical pathways of force transmission at cell-cell contacts and suggest that vinculin can transmit forces at E-cadherin adhesions independent of -catenin, possibly through {beta}-catenin. Furthermore, we tested the ability of cell-cell contacts to withstand external stretch and found that vinculin is essential to maintain cell-cell contact stability under external forces as well.

biophysics↗

bioRxiv2023

α-Catenin Dependent E-cadherin Adhesion Dynamics as Revealed by an Accelerated Force Ramp

Tissue remodeling and shape changes often rely on force-induced cell rearrangements occurring via cell-cell contact dynamics. Epithelial cell-cell contact shape changes are particularly dependent upon E-cadherin adhesion dynamics which are directly influenced by cell-generated and external forces. While both the mobility of E-cadherin adhesions and their adhesion strength have been reported before, it is not clear how these two aspects of E-cadherin adhesion dynamics are related. Here, using magnetic pulling cytometry, we applied an accelerated force ramp on the E-cadherin adhesion between an E-cadherin-coated magnetic microbead and an epithelial cell to ascertain this relationship. Our approach enables the determination of the adhesion strength and force-dependent mobility of individual adhesions, which revealed a direct correlation between these key characteristics. Since -catenin has previously been reported to play a role in both E-cadherin mobility and adhesion strength when studied independently, we also probed epithelial cells in which -catenin has been knocked out. We found that, in the absence of -catenin, E-cadherin adhesions not only had lower adhesion strength, as expected, but were also more mobile. We observed that -catenin was required for the recovery of strained cell-cell contacts and propose that the adhesion strength and force-dependent mobility of E-cadherin adhesions act in tandem to regulate cell-cell contact homeostasis. Our approach introduces a method which relates the force-dependent adhesion mobility to adhesion strength and highlights the morphological role played by -catenin in E-cadherin adhesion dynamics.

biophysics↗

bioRxiv2022

Transcriptome analysis of diverse Plasmodium falciparum clinical isolates identifies genes correlating with highly variable expression of merozoite surface protein MSPDBL2

The merozoite surface protein MSPDBL2 of Plasmodium falciparum is under strong balancing selection and is a target of naturally acquired antibodies. Remarkably, MSPDBL2 is expressed in only a minority of mature schizonts of any cultured parasite line, and mspdbl2 gene transcription increases in response to overexpression of the gametocyte development inducer GDV1, so it is important to understand its natural expression. Here, MSPDBL2 in mature schizonts was analysed in the first ex vivo culture cycle of 96 clinical isolates from four populations with varying levels of infection endemicity in different West African countries, by immunofluorescence microscopy with antibodies against a conserved region of the protein. In most isolates, less than 1% of mature schizonts were positive for MSPDBL2 (median of 0.6% overall), but the frequency distribution was highly skewed as nine isolates had more than 3% schizonts positive and one had 73% positive. To investigate whether expression of other gene loci correlated with MSPDBL2 expression, whole transcriptome sequencing was performed on schizont-enriched material from 17 of the clinical isolates with a wide range of proportions of schizonts positive. Transcripts of particular parasite genes were highly significantly positively correlated with MSPDBL2 positivity in schizonts as well as with mspdbl2 gene transcript levels, with overrepresentation of genes previously implicated as likely to be involved in gametocytogenesis, but not including the gametocytogenesis master regulator ap2g. Although MSPDBL2 is apparently not directly involved in sexual commitment, it marks a co-occurring developmental subpopulation that may be functionally distinct within blood stage infections.

microbiology↗

bioRxiv2020

Multi-locus genotyping reveals established endemicity of a geographically distinct Plasmodium vivax population in Mauritania, West Africa

BackgroundPlasmodium vivax has been recently discovered as a significant cause of malaria in Mauritania, although very rare elsewhere in West Africa. It has not been known if this is a recently introduced or locally remnant parasite population, nor whether the genetic structure reflects epidemic or endemic transmission. Methodology / Principal FindingsTo investigate the P. vivax population genetic structure in Mauritania and compare with populations previously analysed elsewhere, multi-locus genotyping was undertaken on 100 clinical isolates, using a genome-wide panel of 38 single nucleotide polymorphisms (SNPs), plus seven SNPs in drug resistance genes. The Mauritanian P. vivax population is shown to be genetically diverse and divergent from populations elsewhere, indicated consistently by genetic distance matrix analysis, principal components analyses, and fixation indices. Only one isolate had a genotype clearly indicating recent importation, from a southeast Asian source. There was no linkage disequilibrium in the local parasite population, and only a small number of infections appeared to be closely genetically related, indicating that there is ongoing genetic recombination consistent with endemic transmission. The P. vivax diversity in a remote mining town was similar to that in the capital Nouakchott, with no indication of local substructure or of epidemic population structure. Drug resistance alleles were virtually absent in Mauritania, in contrast with P. vivax in other areas of the world. Conclusions / SignificanceThe molecular epidemiology indicates that there is long-standing endemic transmission that will be very challenging to eliminate. The virtual absence of drug resistance alleles suggests that most infections have been untreated, and that this endemic infection has been more neglected in comparison to P. falciparum locally or to P. vivax elsewhere. Author SummaryPlasmodium vivax is a widespread cause of malaria in Mauritania, in contrast to its rarity elsewhere throughout West Africa. To investigate whether the parasite may be recently introduced or epidemic, multi-locus genotyping was performed on 100 Mauritanian P. vivax malaria cases. Analysis of a genome-wide panel of single nucleotide polymorphisms showed the P. vivax population to be genetically diverse and divergent from populations elsewhere, indicating that there has been long-standing endemic transmission. Almost all infections appear to be locally acquired, with the exception of one that was presumably imported with a genotype similar to infections seen in Southeast Asia. The Mauritanian P. vivax population shows no linkage disequilibrium, and very few infections have closely related genotypes, indicating ongoing recombination. The parasite showed no indication of local substructure or epidemic population structure. Drug resistance alleles were virtually absent, suggesting that most infections have been untreated historically. The molecular epidemiology indicates that there has been long-standing endemic transmission of this neglected parasite that requires special attention for control.

microbiology↗