Single-clone Infections Research Articles

Declines in prevalence alter the optimal level of sexual investment for the malaria parasite Plasmodium falciparum

Successful infectious disease interventions can result in large reductions in parasite prevalence. Such demographic change has fitness implications for individual parasites and may shift the parasite's optimal life history strategy. Here, we explore whether declining infection rates can alter Plasmodium falciparum's investment in sexual versus asexual growth. Using a multiscale mathematical model, we demonstrate how the proportion of polyclonal infections, which decreases as parasite prevalence declines, affects the optimal sexual development strategy: Within-host competition in multiclone infections favors a greater investment in asexual growth whereas single-clone infections benefit from higher conversion to sexual forms. At the same time, drug treatment also imposes selection pressure on sexual development by shortening infection length and reducing within-host competition. We assess these models using 148 P. falciparum parasite genomes sampled in French Guiana over an 18-y period of intensive intervention (1998 to 2015). During this time frame, multiple public health measures, including the introduction of new drugs and expanded rapid diagnostic testing, were implemented, reducing P. falciparum malaria cases by an order of magnitude. Consistent with this prevalence decline, we see an increase in the relatedness among parasites, but no single clonal background grew to dominate the population. Analyzing individual allele frequency trajectories, we identify genes that likely experienced selective sweeps. Supporting our model predictions, genes showing the strongest signatures of selection include transcription factors involved in the development of P. falciparum's sexual gametocyte form. These results highlight how public health interventions impose wide-ranging selection pressures that affect basic parasite life history traits.

Genetic diversity and genetic relatedness in Plasmodium falciparum parasite population in individuals with uncomplicated malaria based on microsatellite typing in Eastern and Western regions of Uganda, 2019\u20132020

BackgroundGenetic diversity and parasite relatedness are essential parameters for assessing impact of interventions and understanding transmission dynamics of malaria parasites, however data on its status in Plasmodium falciparum populations in Uganda is limited. Microsatellite markers and DNA sequencing were used to determine diversity and molecular characterization of P. falciparum parasite populations in Uganda.MethodsA total of 147 P. falciparum genomic DNA samples collected from cross-sectional surveys in symptomatic individuals of 2–10 years were characterized by genotyping of seven highly polymorphic neutral microsatellite markers (n = 85) and genetic sequencing of the Histidine Rich Protein 2 (pfhrp2) gene (n = 62). ArcGIS was used to map the geographical distribution of isolates while statistical testing was done using Student's t-test or Wilcoxon's rank-sum test and Fisher’s exact test as appropriate at P ≤ 0.05.ResultsOverall, 75.5% (95% CI 61.1–85.8) and 24.5% (95% CI14.2–38.9) of parasites examined were of multiclonal (mixed genotype) and single clone infections, respectively. Multiclonal infections occurred more frequently in the Eastern region 73.7% (95% CI 48.8–89.1), P < 0.05. Overall, multiplicity of infection (MOI) was 1.9 (95% CI 1.7–2.1), P = 0.01 that was similar between age groups (1.8 vs 1.9), P = 0.60 and regions (1.9 vs 1.8), P = 0.43 for the < 5 and ≥ 5 years and Eastern and Western regions, respectively. Genomic sequencing of the pfhrp2 exon2 revealed a high level of genetic diversity reflected in 96.8% (60/62) unique sequence types. Repeat type AHHAAAHHATD and HRP2 sequence Type C were more frequent in RDT−/PCR + samples (1.9% vs 1.5%) and (13% vs 8%), P < 0.05 respectively. Genetic relatedness analysis revealed small clusters of gene deleted parasites in Uganda, but no clustering with Eritrean parasites.ConclusionHigh level of genetic diversity of P. falciparum parasites reflected in the frequency of multiclonal infections, multiplicity of infection and variability of the pfhrp2 gene observed in this study is consistent with the high malaria transmission intensity in these settings. Parasite genetic analysis suggested spontaneous emergence and clonal expansion of pfhrp2 deleted parasites that require close monitoring to inform national malaria diagnosis and case management policies.

Genetically Determined Response to Artemisinin Treatment in Western Kenyan Plasmodium falciparum Parasites.

Genetically determined artemisinin resistance in Plasmodium falciparum has been described in Southeast Asia. The relevance of recently described Kelch 13-propeller mutations for artemisinin resistance in Sub-Saharan Africa parasites is still unknown. Southeast Asia parasites have low genetic diversity compared to Sub-Saharan Africa, where parasites are highly genetically diverse. This study attempted to elucidate whether genetics provides a basis for discovering molecular markers in response to artemisinin drug treatment in P. falciparum in Kenya. The genetic diversity of parasites collected pre- and post- introduction of artemisinin combination therapy (ACT) in western Kenya was determined. A panel of 12 microsatellites and 91 single nucleotide polymorphisms (SNPs) distributed across the P. falciparum genome were genotyped. Parasite clearance rates were obtained for the post-ACT parasites. The 12 microsatellites were highly polymorphic with post-ACT parasites being significantly more diverse compared to pre-ACT (p < 0.0001). The median clearance half-life was 2.55 hours for the post-ACT parasites. Based on SNP analysis, 15 of 90 post-ACT parasites were single-clone infections. Analysis revealed 3 SNPs that might have some causal association with parasite clearance rates. Further, genetic analysis using Bayesian tree revealed parasites with similar clearance phenotypes were more closely genetically related. With further studies, SNPs described here and genetically determined response to artemisinin treatment might be useful in tracking artemisinin resistance in Kenya.

TESTING SEX RATIO THEORY WITH THE MALARIA PARASITEPLASMODIUM MEXICANUMIN NATURAL AND EXPERIMENTAL INFECTIONS

The malaria parasite (Plasmodium) life history accords well with the assumptions of local mate competition (LMC) of sex ratio theory. Within a single meal of the blood-feeding vector, sexually dimorphic gametocyte cells produce gametes (females produce one, males several) that mate and undergo sexual recombination. The theory posits several factors drive the Plasmodium sex ratio: male fecundity (gametes/male gametocyte), number and relative abundance of parasite clones, and gametocyte density. We measured these traits for the lizard malaria parasite, Plasmodium mexicanum, with a large sample of natural infections and infections from experiments that manipulated clonal diversity. Sex ratio in single-clone infections was slightly female-biased, but matched predictions of theory for this low-fecundity species. Sex ratio was less female-biased in clonally diverse infections as predicted by LMC for the experimental, but not natural infections. Gametocyte density was not positively related to sex ratio. These results are explained by the P. mexicanum life history of naturally low clonal diversity and high gametocyte production. This is the first study of a natural malaria system that examines all traits relevant to LMC in individual vertebrate hosts and suggests a striking example of sex ratio theory having significance for human public health.

Immune-Mediated Competition in Rodent Malaria Is Most Likely Caused by Induced Changes in Innate Immune Clearance of Merozoites

Malarial infections are often genetically diverse, leading to competitive interactions between parasites. A quantitative understanding of the competition between strains is essential to understand a wide range of issues, including the evolution of virulence and drug resistance. In this study, we use dynamical-model based Bayesian inference to investigate the cause of competitive suppression of an avirulent clone of Plasmodium chabaudi (AS) by a virulent clone (AJ) in immuno-deficient and competent mice. We test whether competitive suppression is caused by clone-specific differences in one or more of the following processes: adaptive immune clearance of merozoites and parasitised red blood cells (RBCs), background loss of merozoites and parasitised RBCs, RBC age preference, RBC infection rate, burst size, and within-RBC interference. These processes were parameterised in dynamical mathematical models and fitted to experimental data. We found that just one parameter , the ratio of background loss rate of merozoites to invasion rate of mature RBCs, needed to be clone-specific to predict the data. Interestingly, was found to be the same for both clones in single-clone infections, but different between the clones in mixed infections. The size of this difference was largest in immuno-competent mice and smallest in immuno-deficient mice. This explains why competitive suppression was alleviated in immuno-deficient mice. We found that competitive suppression acts early in infection, even before the day of peak parasitaemia. These results lead us to argue that the innate immune response clearing merozoites is the most likely, but not necessarily the only, mediator of competitive interactions between virulent and avirulent clones. Moreover, in mixed infections we predict there to be an interaction between the clones and the innate immune response which induces changes in the strength of its clearance of merozoites. What this interaction is unknown, but future refinement of the model, challenged with other datasets, may lead to its discovery.

Rosetting in Plasmodium vivax: A Cytoadhesion Phenotype Associated with Anaemia

Background Plasmodium vivax can potentially lead to life-threatening episodes but the mechanisms underlying severe disease remain poorly defined. Cytoadhesion of infected erythrocytes may contribute to P. vivax sequestration and organ injury although its physiological impact is still unknown. Here, we aimed to describe clinically-relevant cytoadhesive phenotypes of P. vivax isolates.Methodology/Principal findingsRosetting and adhesion to CSA, CD36, ICAM1, placental and brain cryosections were determined in P. vivax peripheral isolates from 12 pregnant women, 24 non-pregnant women and 23 men from Manaus (Brazil). P. falciparum co-infection was excluded by PCR and P. vivax isolates were genotyped by assessing the size polymorphism of microsatellites ms2, ms20 and msp1F3 through capillary electrophoresis of PCR products. P. vivax monoinfection was confirmed by PCR in 59 isolates, with 50 (85%) of them being single-clone infections. One P. vivax haplotype was more frequently found among pregnant women (33%) than in non-pregnant women (0%) and men (4%; p = 0.010). Rosetting was observed in 64% of the isolates, adhesion to CSA in 15%, to ICAM1 in 12% and to placental cryosections in 9%, being similar among pregnant and non-pregnant groups. Intensity of rosetting was higher among anaemic individuals compared to non-anaemic (p = 0.010) and decreased with increasing haematocrit (p = 0.033) and haemoglobin levels (p = 0.015).Conclusions/Significance P. vivax peripheral isolates from pregnant women do not exhibit a prominent adhesion to CSA, although other parasite phenotypes still unknown may increase the propagation of certain P. vivax clones observed among pregnant hosts. Rosetting is a frequent cytoadhesive phenotype in P. vivax infections that may contribute to the development of anaemia.

Genetic characterization of Theileria equi infecting horses in North America: evidence for a limited source of U.S. introductions

BackgroundTheileria equi is a tick-borne apicomplexan hemoparasite that causes equine piroplasmosis. This parasite has a worldwide distribution but the United States was considered to be free of this disease until recently.MethodsWe used samples from 37 horses to determine genetic relationships among North American T. equi using the 18S rRNA gene and microsatellites. We developed a DNA fingerprinting panel of 18 microsatellite markers using the first complete genome sequence of T. equi.ResultsA maximum parsimony analysis of 18S rRNA sequences grouped the samples into two major clades. The first clade (n = 36) revealed a high degree of nucleotide similarity in U.S. T. equi, with just 0–2 single nucleotide polymorphisms (SNPs) among samples. The remaining sample fell into a second clade that was genetically divergent (48 SNPs) from the other U.S. samples. This sample was collected at the Texas border, but may have originated in Mexico. We genotyped T. equi from the U.S. using microsatellite markers and found a moderate amount of genetic diversity (2–8 alleles per locus). The field samples were mostly from a 2009 Texas outbreak (n = 22) although samples from five other states were also included in this study. Using Weir and Cockerham’s FST estimator (θ) we found strong population differentiation of the Texas and Georgia subpopulations (θ = 0.414), which was supported by a neighbor-joining tree created with predominant single haplotypes. Single-clone infections were found in 27 of the 37 samples (73%), allowing us to identify 15 unique genotypes.ConclusionsThe placement of most T. equi into one monophyletic clade by 18S is suggestive of a limited source of introduction into the U.S. When applied to a broader cross section of worldwide samples, these molecular tools should improve source tracking of T. equi outbreaks and may help prevent the spread of this tick-borne parasite.

Gametocyte sex ratio in single-clone infections of the malaria parasite Plasmodium mexicanum

Sex ratio theory predicts that malaria parasites should bias gametocyte production toward female cells in single-clone infections because they will experience complete inbreeding of parasite gametes within the vector. A higher proportion of male gametocytes is favoured under conditions that reduce success of male gametes at reaching females such as low gametocyte density or attack of the immune system later in the infection. Recent experimental studies reveal genetic variation for gametocyte sex ratio in single-clone infections. We examined these issues with a study of experimental single-clone infections for the lizard malaria parasite Plasmodium mexicanum in its natural host. Gametocyte sex ratios of replicate single-clone infections were determined over a period of 3-4 months. Sex ratios were generally female biased, but not as strongly as expected under simple sex ratio theory. Gametocyte density was not related to sex ratio, and male gametocytes did not become more common later in infections. The apparent surplus of male gametocytes could be explained if male fecundity is low in this parasite, or if rapid clotting of the lizard blood reduces male gamete mobility. There was also a significant clone effect on sex ratio, suggesting genetic variation for some life-history trait, possibly male fecundity.

Relative Clonal Density of Malaria Parasites in Mixed-Genotype Infections: Validation of a Technique Using Microsatellite Markers for Plasmodium falciparum and P. mexicanum

Quantifying the relative proportion of coexisting genotypes (clones) of a malaria parasite within its vertebrate host's blood would provide insights into critical features of the biology of the parasite, including competition among clones, gametocyte sex ratio, and virulence. However, no technique has been available to extract such data for natural parasite-host systems when the number of clones cycling in the overall parasite population is likely to be large. Recent studies find that data from genetic analyzer instruments for microsatellite markers allow measuring clonal proportions. We conducted a validation study for Plasmodium mexicanum and Plasmodium falciparum by mixing DNA from single-clone infections to simulate mixed infections of each species with known proportions of clones. Results for any mixture of DNA gave highly reproducible results. The relationship between known and measured relative proportions of clones was linear, with high regression r² values. Known and measured clone proportions for simulated infections followed over time (mixtures) were compared with 3 methods: using uncorrected data, with uncorrected data and confidence intervals constructed from observed experimental error, and using a baseline mixture of equal proportions to calibrate all other results. All 3 methods demonstrated value in studies of mixed-genotype infections sampled a single time or followed over time. Thus, the method should open new windows into the biology of malaria parasites.

Clonal diversity alters the infection dynamics of a malaria parasite (Plasmodium mexicanum) in its vertebrate host

Ecological and evolutionary theory predicts that genetic diversity of microparasites within infected hosts will influence the parasite replication rate, parasitemia, transmission strategy, and virulence. We manipulated clonal diversity (number of genotypes) of the malaria parasite, Plasmodium mexicanum, in its natural lizard host and measured important features of the infection dynamics, the first such study for any natural Plasmodium-host association. Hosts harboring either a single P. mexicanum clone or various combinations of clones (scored via three microsatellite markers) were established. Production of asexually replicating stages (meronts) and maximal meront parasitemia did not differ by clonal diversity, nor did timing of first production of transmission stages (gametocytes). However, mean rate of gametocyte increase and maximal gametocyte parasitemia were greater for hosts with mixed-clone infections. Characteristics of infections were more variable in hosts with mixed-clone infections than with single-clone infections except for first production of gametocytes. One or more of the parasite reproductive traits were extreme in 20 of 52 hosts with mixed-clone infections. This was not associated with specific clones, but diversity itself. The overall pattern from studies of clonal diversity for human, rodent, and now reptile malaria parasites confirms that the genetic diversity of infections in the vertebrate host is of central importance for the ecology of Plasmodium.

Molecular analysis of chloroquine resistance in Plasmodium falciparum in Yunnan Province, China

Resistance of Plasmodium falciparum to chloroquine (CQ) is determined by the mutation at K76T of the P. falciparum chloroquine resistance transporter (pfcrt) gene and modified by other mutations in this gene and in the P. falciparum multidrug resistance 1 (pfmdr1) gene. To determine the extent of polymorphisms in these genes in field P. falciparum isolates from Yunnan province of China, we genotyped the pfcrt codon 76, pfmdr1 codons 86 and 1246. Our results showed that although CQ has been withdrawn from treating falciparum malaria for over two decades, 90.3% of the parasites still carried the pfcrt K76T mutation. In contrast, mutations at pfmdr1 codons 86 and 1246 were rare. Sequencing analysis of the pfcrt gene in 34 parasite field isolates revealed CVIET at positions 72-76 as the major type, consistent with the theory of Southeast Asian origin of CQ resistance in the parasite. In addition, two novel pfcrt haplotypes (75D/144Y/220A and 75E/144Y/220A) were identified. Real-time polymerase chain reaction was used to determine pfmdr1 gene amplification, which is associated with mefloquine resistance. Our result indicated that in agreement with that mefloquine has not been used in this area, most (>90%) of the parasites had one pfmdr1 copy. Genotyping at two hypervariable loci showed relatively low levels of genetic diversity of the parasite population. Meanwhile, 28.4% of cases were found to contain mixed clones, which favour genetic recombination. Furthermore, despite a unique history of antimalarial drugs in Yunnan, its geographical connections with three malarious countries facilitate gene flow among parasite populations and evolution of novel drug-resistant genotypes. Therefore, continuous surveillance of drug resistance in this area is necessary for timely adjustment of local drug policies and more effective malaria control.

Clonal diversity of a lizard malaria parasite, Plasmodium mexicanum, in its vertebrate host, the western fence lizard: role of variation in transmission intensity over time and space

Within the vertebrate host, infections of a malaria parasite (Plasmodium) could include a single genotype of cells (single-clone infections) or two to several genotypes (multiclone infections). Clonal diversity of infection plays an important role in the biology of the parasite, including its life history, virulence, and transmission. We determined the clonal diversity of Plasmodium mexicanum, a lizard malaria parasite at a study region in northern California, using variable microsatellite markers, the first such study for any malaria parasite of lizards or birds (the most common hosts for Plasmodium species). Multiclonal infections are common (50-88% of infections among samples), and measures of genetic diversity for the metapopulation (expected heterozygosity, number of alleles per locus, allele length variation, and effective population size) all indicated a substantial overall genetic diversity. Comparing years with high prevalence (1996-1998 = 25-32% lizards infected), and years with low prevalence (2001-2005 = 6-12%) found fewer alleles in samples taken from the low-prevalence years, but no reduction in overall diversity (H = 0.64-0.90 among loci). In most cases, rare alleles appeared to be lost as prevalence declined. For sites chronically experiencing low transmission intensity (prevalence approximately 1%), overall diversity was also high (H = 0.79-0.91), but there were fewer multiclonal infections. Theory predicts an apparent excess in expected heterozygosity follows a genetic bottleneck. Evidence for such a distortion in genetic diversity was observed after the drop in parasite prevalence under the infinite alleles mutation model but not for the stepwise mutation model. The results are similar to those reported for the human malaria parasite, Plasmodium falciparum, worldwide, and support the conclusion that malaria parasites maintain high genetic diversity in host populations despite the potential for loss in alleles during the transmission cycle or during periods/locations when transmission intensity is low.

Co-occurrences of parasite clones and altered host phenotype in a snail–trematode system

The frequent co-occurrence of two or more genotypes of the same parasite species in the same individual hosts has often been predicted to select for higher levels of virulence. Thus, if parasites can adjust their level of host exploitation in response to competition for resources, mixed-clone infections should have more profound impacts on the host. Trematode parasites are known to induce a wide range of modifications in the morphology (size, shell shape or ornamentation) of their snail intermediate host. Still, whether mixed-clone trematode infections have additive effects on the phenotypic alterations of the host remains to be tested. Here, we used the snail Potamopyrgus antipodarum-infected by the trematode Coitocaecum parvum to test for both the general effect of the parasite on host phenotype and possible increased host exploitation in multi-clone infections. Significant differences in size, shell shape and spinosity were found between infected and uninfected snails, and we determined that one quarter of naturally infected snails supported mixed-clone infections of C. parvum. From the parasite perspective, this meant that almost half of the clones identified in this study shared their snail host with at least one other clone. Intra-host competition may be intense, with each clone in a mixed-clone infection experiencing major reductions in volume and number of sporocysts (and consequently multiplication rate and cercarial production) compared with single-clone infections. However, there was no significant difference in the intensity of host phenotype modifications between single and multiple-clone infections. These results demonstrate that competition between parasite genotypes may be strong, and suggest that the frequency of mixed-clone infections in this system may have selected for an increased level of host exploitation in the parasite population, such that a single-clone is associated with a high degree of host phenotypic alteration.

The role of imported cases in the epidemiology of urban Plasmodium falciparum malaria in Quibdó, Colombia

To assess the role of imported cases in the local epidemiology of malaria and population genetics of Plasmodium falciparum in an urban endemic area in Colombia. A total of 679 P. falciparum cases presenting in the city were interviewed, mapped, and genotyped using msp1 and msp2 molecular markers. Among the cases, 75% were classified as imported and harboured single-clone infections. The P. falciparum parasite population had low genetic diversity with a preponderant haplotype (mean genetic diversity H = 0.36), even when microsatellite markers were used (H = 0.49), partly because of the small scale at which population movement was studied. Nevertheless, indigenous and potentially introduced cases were identified. Migration is a confounder in planning malaria control in this endemic urban area. Longitudinal studies which monitor the P. falciparum population in imported and autochthonous cases at larger spatial scales would be necessary to study the effect of population movement on malaria transmission and, with suitable molecular markers, on the spread of drug resistance.

Cooperation and virulence in acute Pseudomonas aeruginosa infections.

BackgroundEfficient host exploitation by parasites is frequently likely to depend on cooperative behaviour. Under these conditions, mixed-strain infections are predicted to show lower virulence (host mortality) than are single-clone infections, due to competition favouring non-contributing social 'cheats' whose presence will reduce within-host growth. We tested this hypothesis using the cooperative production of iron-scavenging siderophores by the pathogenic bacterium Pseudomonas aeruginosa in an insect host.ResultsWe found that infection by siderophore-producing bacteria (cooperators) results in more rapid host death than does infection by non-producers (cheats), and that mixtures of both result in intermediate levels of virulence. Within-host bacterial growth rates exhibited the same pattern. Crucially, cheats were more successful in mixed infections compared with single-clone infections, while the opposite was true of cooperators.ConclusionThese data demonstrate that mixed clone infections can favour the evolution of social cheats, and thus decrease virulence when parasite growth is dependent on cooperative behaviours.

Are Transporter Genes Other than the Chloroquine Resistance Locus ( pfcrt ) and Multidrug Resistance Gene ( pfmdr ) Associated with Antimalarial Drug Resistance?

Mu et al. (Mu, J., M. T. Ferdig, X. Feng, D. A. Joy, J. Duan, T. Furuya, G. Subramanian, L. Aravind, R. A. Cooper, J. C. Wootton, M. Xiong, and X. Z. Su, Mol. Microbiol. 49:977-989, 2003) recently reported exciting associations between nine new candidate transporter genes and in vitro resistance to chloroquine (CQ) and quinine (QN), with six of these loci showing association with CQ or QN in a southeast Asian population sample. We replicated and extended this work by examining polymorphisms in these genes and in vitro resistance to eight drugs in parasites collected from the Thailand-Burma border. To minimize problems of multiple testing, we used a two-phase study design, while to minimize problems caused by population structure, we analyzed parasite isolates collected from a single clinic. We first examined associations between genotype and drug response in 108 unique single-clone parasite isolates. We found strong associations between single nucleotide polymorphisms in pfmdr and mefloquine (MFQ), artesunate (AS), and lumefantrine (LUM) response. We also observed associations between an ABC transporter (G7) and response to QN and AS and between another ABC transporter (G49) and response to dihydro-artemisinin (DHA). We reexamined significant associations in an independent sample of 199 unique single-clone infections from the same location. The significant associations with pfmdr-1042 detected in the first survey remained. However, with the exception of the G7-artesunate association, all other associations observed with the nine new candidate transporters disappeared. We also examined linkage disequilibrium (LD) between markers and phenotypic correlations between drug responses. We found minimal LD between genes. Furthermore, we found no correlation between chloroquine and quinine responses, although we did find expected strong correlations between MFQ, QN, AS, DHA, and LUM. To conclude, we found no evidence for an association between 8/9 candidate genes and response to eight different antimalarial drugs. However, the consistent association observed between a 3-bp indel in G7 and AS response merits further investigation.

Antigenic cross‐reactivity between different alleles of the Plasmodium falciparum merozoite surface protein 2

The polymorphic domain of the gene encoding Plasmodium falciparum merozoite surface protein 2 (MSP2) was PCR amplified from blood of malaria patients, genotyped, and 19 distinct fragments were cloned and expressed in E. coli. The reactivity of naturally occurring antibodies against this panel of recombinant MSP2 antigens was tested using 67 homologous or heterologous sera from a serum bank of travel clinic patients. Sera from semi-immune individuals strongly recognized almost all recombinant antigens. Sera from primary infected patients either did not react at all (9 sera), or reacted weakly against varying numbers of antigens (39 sera). The antigens that showed reactions were mostly of the allelic family corresponding to the infecting clone, but in very few cases also of the alternative allelic family. Single clone infections and repeated samples from the same individual were analysed in greater detail. Thus, we were able to quantify cross-reactivity induced by a single P. falciparum infection. Within the two allelic families of MSP2, cross-reactivity was observed between some but not all alleles of the same family, whereas antibodies cross-reactive between variants belonging to different allelic families were detected in only a few cases.

Rodent malaria parasites suffer from the presence of conspecific clones in three-clone Plasmodium chabaudi infections.

We studied infection dynamics of Plasmodium chabaudi in mice infected with 3 genetically distinct clones--1 less virulent than the other 2--either on their own or in mixtures. During the acute phase of infection, total numbers of asexual parasites in mixed-clone infections were equal to those produced by the 3 clones alone, suggesting strong in-host competition among clones. During the chronic phase of the infection, mixed-clone infections produced more asexual parasites than single-clone infections, suggesting lower levels of competition than during the acute phase, and indicating that a genetically diverse infection is harder to control by the host immune system. Transmission potential over the whole course of infection was lower from mixed-clone infections than from the average of the 3 single-clone infections. These results suggest that in-host competition reduces both growth rate and probability of transmission for individual parasite clones.

Theileria annulata: virulence and transmission from single and mixed clone infections in cattle

Theoretically, parasite virulence should be higher for faster growing parasites, and higher in mixed infections compared to single-clone infections. Virulence should also be positively correlated to transmission rates. Theileria annulata provides a good model system for studying such hypotheses, as parasite replication causes harm to the host, and there is evidence suggesting that the genetic complexity of an infection might affect its virulence. Two clones of T. annulata were chosen, one fast growing and one slow growing in vitro and these were used to establish cattle infections, either alone, or in a mixed infection. Virulence was measured using lymph node expansion, temperature, and blood parameters as correlates. As predicted, the faster growing clone was found to produce higher virulence. Mixed infections did not show higher virulence than single-clone infections, but interactions within mixed infections resulted in more transmission stage production than seen in either of the single-clone infections. Index Descriptors and Abbreviations. Theileria annulata, Apicomplexa, mixed infections, virulence, growth rates. red blood cell, RBC; packed cell volume, PCV.

Transmission success of the malaria parasite Plasmodium mexicanum into its vector: role of gametocyte density and sex ratio.

The life-cycle of Plasmodium depends on transmission of the parasite from the vertebrate host into its vector when the insect takes a bloodmeal. Transmission success may depend in part on the parasite's gametocyte density and sex ratio in the blood. P. mexicanum, a parasite of fence lizards in California, USA, exploits the sandfly Lutzomyia vexator as its vector. In experimental transmissions using naturally infected lizards as donors of blood, transmission success (measured as percentage of vectors infected and number of parasite oocysts on the insect's midgut) was positively related to gametocyte density, although density above 20/1000 erythrocytes did not improve transmission. Sex ratio (proportion of microgametocytes in the infection) was positively correlated with gametocyte density. Transmission improved with higher proportion of microgametocytes, but partial correlations revealed that this was a result only of higher gametocyte densities. These results agree with the theory of virulence and sex ratios because single clone infections should produce a more female-biased sex ratio and grow to the minimum parasitaemia that would maximize clonal transmission, whereas multiple clone infections will be closer to a 1:1 sex ratio and yield a higher parasitaemia when each clone competes for transmission to the vector.