Natural Parasite Populations Research Articles

Role of parasite transmission in promoting inbreeding: II. Pedigree reconstruction reveals sib‐transmission and consequent kin‐mating

Even though parasitic flatworms are one of the most species-rich groups of hermaphroditic organisms, we know virtually nothing of their mating systems (selfing or kin-mating rates) in nature. Hence, we lack an understanding of the role of inbreeding in parasite evolution. The natural mating systems of parasitic flatworms have remained elusive due to the inherent difficulty in generating progeny-array data in many parasite systems. New developments in pedigree reconstruction allow direct inference of realized selfing rates in nature by simply using a sample of genotyped individuals. We built upon this advancement by utilizing the closed mating systems, that is, individual hosts, of endoparasites. In particular, we created a novel means to use pedigree reconstruction data to estimate potential kin-mating rates. With data from natural populations of a tapeworm, we demonstrated how our newly developed methods can be used to test for cosibling transmission and inbreeding depression. We then showed how independent estimates of the two mating system components, selfing and kin-mating rates, account for the observed levels of inbreeding in the populations. Thus, our results suggest that these natural parasite populations are in inbreeding equilibrium. Pedigree reconstruction analyses along with the new companion methods we developed will be broadly applicable across a myriad of parasite species. As such, we foresee that a new frontier will emerge wherein the diverse life histories of flatworm parasites could be utilized in comparative evolutionary studies to broadly address ecological factors or life history traits that drive mating systems and hence inbreeding in natural populations.

Features of Distribution, Harmfulness and Control of the Population of the Eriosoma Lanigerum Hausm. (Homoptera, Aphididae) Woolly Aphid in the Nurseries of Ukraine and Kazakhstan

The paper presents the results on technology of cultivation of apple-tree saplings and the ways to regulate the phytophage population. The nature of distribution of one of the main nursery pests such as woolly aphid and its specialized entomophage – aphelinus - has been studied. A woolly aphid - typical representative of sucking phytophages - is characterized as a dominant species in the gardens and nurseries of Ukraine and Kazakhstan. Standard methods of research were used in the course of work. Based on the studies, a set of factors forming the structure and dynamics of red woolly aphids in apple tree nurseries has been determined. The natural populations of parasites and predators play an essential role for its population dynamics. The principle possibility of regulating the population and protection of nurseries from woolly aphids using the method of settling the natural populations of the hymenopteran entomophage aphelinus has been established.

Genotype specificity among hosts, pathogens, and beneficial microbes influences the strength of symbiont-mediated protection.

The microbial symbionts of eukaryotes influence disease resistance in many host‐parasite systems. Symbionts show substantial variation in both genotype and phenotype, but it is unclear how natural selection maintains this variation. It is also unknown whether variable symbiont genotypes show specificity with the genotypes of hosts or parasites in natural populations. Genotype by genotype interactions are a necessary condition for coevolution between interacting species. Uncovering the patterns of genetic specificity among hosts, symbionts, and parasites is therefore critical for determining the role that symbionts play in host‐parasite coevolution. Here, we show that the strength of protection conferred against a fungal pathogen by a vertically transmitted symbiont of an aphid is influenced by both host‐symbiont and symbiont‐pathogen genotype by genotype interactions. Further, we show that certain symbiont phylogenetic clades have evolved to provide stronger protection against particular pathogen genotypes. However, we found no evidence of reciprocal adaptation of co‐occurring host and symbiont lineages. Our results suggest that genetic variation among symbiont strains may be maintained by antagonistic coevolution with their host and/or their host's parasites.

EVIDENCE OF PLASMODIUM FALCIPARUM RESISTANCE TO SULPHADOXINE-PYRIMETHAMINE (SP) IN PREGNANT WOMEN ALONG THE SLOPE OF MOUNT CAMEROON

BackgroundMalaria in pregnancy (MiP) has debilitating effects for both mother and neonate, with intermittent preventive treatment in pregnancy (IPTp) central to successful malaria control and management in this vulnerable group....

Editorial: Why Vaccines to HIV, HCV, and Malaria Have So Far Failed-Challenges to Developing Vaccines Against Immunoregulating Pathogens.

Editorial: Why Vaccines to HIV, HCV, and Malaria Have So Far Failed-Challenges to Developing Vaccines Against Immunoregulating Pathogens.

Plasmodium falciparum merozoite surface protein 1 block 2 gene polymorphism in field isolates along the slope of mount Cameroon: a cross - sectional study.

BackgroundMalaria remains a major global health burden despite the intensification of control efforts, due partly to the lack of an effective vaccine. Information on genetic diversity in natural parasite populations constitutes a major impediment to vaccine development efforts and is limited in some endemic settings. The present study characterized diversity by investigating msp1 block 2 polymorphisms and the relationship between the allele families with ethnodemographic indices and clinical phenotype.MethodIndividuals with asymptomatic parasitaemia (AP) or uncomplicated malaria (UM) were enrolled from rural, semi-rural and semi-urban localities at varying altitudes along the slope of mount Cameroon. P. falciparum malaria parasitaemic blood screened by light microscopy was depleted of leucocytes using CF11 cellulose columns and the parasite DNA genotyped by nested PCR.ResultsLength polymorphism was assessed in 151 field isolates revealing 64 (5) and 274 (22) distinct recombinant and major msp1 allelic fragments (genotypes) respectively. All family specific allelic types (K1, MAD20 and RO33) as well as MR were observed in the different locations, with K1 being most abundant. Eighty seven (60 %) of individuals harbored more than one parasite clone, with a significant proportion (p = 0.009) in rural compared to other settings. AP individuals had higher (p = 0.007) K1 allele frequencies but lower (p = 0.003) mean multiplicity of genotypes per infection (2.00 ± 0.98 vs. 2.56 ± 1.17) compared to UM patients.ConclusionsThese results indicate enormous diversity of P. falciparum in the area and suggests that allele specificity and complexity may be relevant for the progression to symptomatic disease.

Strategies for designing and monitoring malaria vaccines targeting diverse antigens.

After more than 50 years of intensive research and development, only one malaria vaccine candidate, “RTS,S,” has progressed to Phase 3 clinical trials. Despite only partial efficacy, this candidate is now forecast to become the first licensed malaria vaccine. Hence, more efficacious second-generation malaria vaccines that can significantly reduce transmission are urgently needed. This review will focus on a major obstacle hindering development of effective malaria vaccines: parasite antigenic diversity. Despite extensive genetic diversity in leading candidate antigens, vaccines have been and continue to be formulated using recombinant antigens representing only one or two strains. These vaccine strains represent only a small fraction of the diversity circulating in natural parasite populations, leading to escape of non-vaccine strains and challenging investigators’ abilities to measure strain-specific efficacy in vaccine trials. Novel strategies are needed to overcome antigenic diversity in order for vaccine development to succeed. Many studies have now cataloged the global diversity of leading Plasmodium falciparum and Plasmodium vivax vaccine antigens. In this review, we describe how population genetic approaches can be applied to this rich data source to predict the alleles that best represent antigenic diversity, polymorphisms that contribute to it, and to identify key polymorphisms associated with antigenic escape. We also suggest an approach to summarize the known global diversity of a given antigen to predict antigenic diversity, how to select variants that best represent the strains circulating in natural parasite populations and how to investigate the strain-specific efficacy of vaccine trials. Use of these strategies in the design and monitoring of vaccine trials will not only shed light on the contribution of genetic diversity to the antigenic diversity of malaria, but will also maximize the potential of future malaria vaccine candidates.

A likelihood-based approach for assessment of extra-pair paternity and conspecific brood parasitism in natural populations.

Genotypes are frequently used to assess alternative reproductive strategies such as extra-pair paternity and conspecific brood parasitism in wild populations. However, such analyses are vulnerable to genotyping error or molecular artefacts that can bias results. For example, when using multilocus microsatellite data, a mismatch at a single locus, suggesting the offspring was not directly related to its putative parents, can occur quite commonly even when the offspring is truly related. Some recent studies have advocated an ad-hoc rule that offspring must differ at more than one locus in order to conclude that they are not directly related. While this reduces the frequency with which true offspring are identified as not directly related young, it also introduces bias in the opposite direction, wherein not directly related young are categorized as true offspring. More importantly, it ignores the additional information on allele frequencies which would reduce overall bias. In this study, we present a novel technique for assessing extra-pair paternity and conspecific brood parasitism using a likelihood-based approach in a new version of program cervus. We test the suitability of the technique by applying it to a simulated data set and then present an example to demonstrate its influence on the estimation of alternative reproductive strategies.

Can host ecology and kin selection predict parasite virulence?

Parasite virulence, or the damage a parasite does to its host, is measured in terms of both host costs (reductions in host growth, reproduction and survival) and parasite benefits (increased transmission and parasite numbers) in the literature. Much work has shown that ecological and genetic factors can be strong selective forces in virulence evolution. This review uses kin selection theory to explore how variations in host ecological parameters impact the genetic relatedness of parasite populations and thus virulence. We provide a broad overview of virulence and population genetics studies and then draw connections to existing knowledge about natural parasite populations. The impact of host movement (transporting parasites) and host resistance (filtering parasites) on the genetic structure and virulence of parasite populations is explored, and empirical studies of these factors using Plasmodium and trematode systems are proposed.

A comparison of multiple methods for estimating parasitemia of hemogregarine hemoparasites (apicomplexa: adeleorina) and its application for studying infection in natural populations.

Identifying factors influencing infection patterns among hosts is critical for our understanding of the evolution and impact of parasitism in natural populations. However, the correct estimation of infection parameters depends on the performance of detection and quantification methods. In this study, we designed a quantitative PCR (qPCR) assay targeting the 18 S rRNA gene to estimate prevalence and intensity of Hepatozoon infection and compared its performance with microscopy and PCR. Using qPCR, we also compared various protocols that differ in the biological source and the extraction methods. Our results show that the qPCR approach on DNA extracted from blood samples, regardless of the extraction protocol, provided the most sensitive estimates of Hepatozoon infection parameters; while allowed us to differentiate between mixed infections of Adeleorinid (Hepatozoon) and Eimeriorinid (Schellackia and Lankesterella), based on the analysis of melting curves. We also show that tissue and saline methods can be used as low-cost alternatives in parasitological studies. The next step was to test our qPCR assay in a biological context, and for this purpose we investigated infection patterns between two sympatric lacertid species, which are naturally infected with apicomplexan hemoparasites, such as the genera Schellackia (Eimeriorina) and Hepatozoon (Adeleorina). From a biological standpoint, we found a positive correlation between Hepatozoon intensity of infection and host body size within each host species, being significantly higher in males, and higher in the smaller sized host species. These variations can be associated with a number of host intrinsic factors, like hormonal and immunological traits, that require further investigation. Our findings are relevant as they pinpoint the importance of accounting for methodological issues to better estimate infection in parasitological studies, and illustrate how between-host factors can influence parasite distributions in sympatric natural populations.

Lower survival probability of house sparrows severely infected by the gapeworm parasite

The effect of parasites in natural populations has received increasing attention in recent years. Studies have shown that parasites may play an important part in population ecology due to their potential effects on host fitness. The main purpose of the present study was to investigate the effect of a nematode parasite (gapeworm,Syngamus trachea) on survival probability of house sparrowsPasser domesticusfrom six natural populations located on 4 islands in the Helgeland archipelago in northern Norway. Infection status was obtained from feces samples collected from 603 house sparrows in the summer and autumn of 2007–2011. We also collected data on a visible symptom of severe infection (gasping for air) in 1391 house sparrows in the summer and autumn 2004–2011. We took advantage of recent advances in disease modeling in a multi‐event capture–mark–recapture framework to account for imperfect observations (state uncertainty). Each dataset was separately analyzed, in both analyses we investigated the relationships of year, island, individual body condition, age class and population density with survival probabilities. The relationship between infection (determined by feces egg counts) on annual survival of house sparrows was not statistically significant. However, the probability of annual survival was found to be significantly lower for adult house sparrows exhibiting a symptom of severe gapeworm infection, gasping for air. The present study demonstrates that severe infection by a parasite can have a negative relationship with survival probability of short‐lived avian hosts in wild populations.

Diversity of T Cell Epitopes in Plasmodium falciparum Circumsporozoite Protein Likely Due to Protein-Protein Interactions

Circumsporozoite protein (CS) is a leading vaccine antigen for falciparum malaria, but is highly polymorphic in natural parasite populations. The factors driving this diversity are unclear, but non-random assortment of the T cell epitopes TH2 and TH3 has been observed in a Kenyan parasite population. The recent publication of the crystal structure of the variable C terminal region of the protein allows the assessment of the impact of diversity on protein structure and T cell epitope assortment. Using data from the Gambia (55 isolates) and Malawi (235 isolates), we evaluated the patterns of diversity within and between epitopes in these two distantly-separated populations. Only non-synonymous mutations were observed with the vast majority in both populations at similar frequencies suggesting strong selection on this region. A non-random pattern of T cell epitope assortment was seen in Malawi and in the Gambia, but structural analysis indicates no intramolecular spatial interactions. Using the information from these parasite populations, structural analysis reveals that polymorphic amino acids within TH2 and TH3 colocalize to one side of the protein, surround, but do not involve, the hydrophobic pocket in CS, and predominately involve charge switches. In addition, free energy analysis suggests residues forming and behind the novel pocket within CS are tightly constrained and well conserved in all alleles. In addition, free energy analysis shows polymorphic residues tend to be populated by energetically unfavorable amino acids. In combination, these findings suggest the diversity of T cell epitopes in CS may be primarily an evolutionary response to intermolecular interactions at the surface of the protein potentially counteracting antibody-mediated immune recognition or evolving host receptor diversity.

High prevalence of cestodes in Artemia spp. throughout the annual cycle: relationship with abundance of avian final hosts

Brine shrimp, Artemia spp., act as intermediate hosts for a range of cestode species that use waterbirds as their final hosts. These parasites can have marked influences on shrimp behavior and fecundity, generating the potential for cascading effects in hypersaline food webs. We present the first comprehensive study of the temporal dynamics of cestode parasites in natural populations of brine shrimp throughout the annual cycle. Over a 12-month period, clonal Artemia parthenogenetica were sampled in the Odiel marshes in Huelva, and the sexual Artemia salina was sampled in the Salinas de Cerrillos in Almería. Throughout the year, 4-45% of A. parthenogenetica were infected with cestodes (mean species richness = 0.26), compared to 27-72% of A. salina (mean species richness = 0.64). Ten cestode species were recorded. Male and female A. salina showed similar levels of parasitism. The most prevalent and abundant cestodes were those infecting the most abundant final hosts, especially the Greater Flamingo Phoenicopterus ruber. In particular, the flamingo parasite Flamingolepis liguloides had a prevalence of up to 43% in A. parthenogenetica and 63.5% in A. salina in a given month. Although there was strong seasonal variation in prevalence, abundance, and intensity of cestode infections, seasonal changes in bird counts were weak predictors of the dynamics of cestode infections. However, infection levels of Confluaria podicipina in A. parthenogenetica were positively correlated with the number of their black-necked grebe Podiceps nigricollis hosts. Similarly, infection levels of Anomotaenia tringae and Anomotaenia microphallos in A. salina were correlated with the number of shorebird hosts present the month before. Correlated seasonal transmission structured the cestode community, leading to more multiple infections than expected by chance.

Molecular Mechanisms of Drug Resistance in Natural Leishmania Populations Vary with Genetic Background

The evolution of drug-resistance in pathogens is a major global health threat. Elucidating the molecular basis of pathogen drug-resistance has been the focus of many studies but rarely is it known whether a drug-resistance mechanism identified is universal for the studied pathogen; it has seldom been clarified whether drug-resistance mechanisms vary with the pathogen's genotype. Nevertheless this is of critical importance in gaining an understanding of the complexity of this global threat and in underpinning epidemiological surveillance of pathogen drug resistance in the field. This study aimed to assess the molecular and phenotypic heterogeneity that emerges in natural parasite populations under drug treatment pressure. We studied lines of the protozoan parasite Leishmania (L.) donovani with differential susceptibility to antimonial drugs; the lines being derived from clinical isolates belonging to two distinct genetic populations that circulate in the leishmaniasis endemic region of Nepal. Parasite pathways known to be affected by antimonial drugs were characterised on five experimental levels in the lines of the two populations. Characterisation of DNA sequence, gene expression, protein expression and thiol levels revealed a number of molecular features that mark antimonial-resistant parasites in only one of the two populations studied. A final series of in vitro stress phenotyping experiments confirmed this heterogeneity amongst drug-resistant parasites from the two populations. These data provide evidence that the molecular changes associated with antimonial-resistance in natural Leishmania populations depend on the genetic background of the Leishmania population, which has resulted in a divergent set of resistance markers in the Leishmania populations. This heterogeneity of parasite adaptations provides severe challenges for the control of drug resistance in the field and the design of molecular surveillance tools for widespread applicability.

Molecular evolution and phylogenetics of rodent malaria parasites

BackgroundOver the last 6 decades, rodent Plasmodium species have become key model systems for understanding the basic biology of malaria parasites. Cell and molecular parasitology have made much progress in identifying genes underpinning interactions between malaria parasites, hosts, and vectors. However, little attention has been paid to the evolutionary genetics of parasites, which provides context for identifying potential therapeutic targets and for understanding the selective forces shaping parasites in natural populations. Additionally, understanding the relationships between species, subspecies, and strains, is necessary to maximize the utility of rodent malaria parasites as medically important infectious disease models, and for investigating the evolution of host-parasite interactions.ResultsHere, we collected multi-locus sequence data from 58 rodent malaria genotypes distributed throughout 13 subspecies belonging to P. berghei, P. chabaudi, P. vinckei, and P. yoelii. We employ multi-locus methods to infer the subspecies phylogeny, and use population-genetic approaches to elucidate the selective patterns shaping the evolution of these organisms. Our results reveal a time-line for the evolution of rodent Plasmodium and suggest that all the subspecies are independently evolving lineages (i.e. species). We show that estimates of species-level polymorphism are inflated if subspecies are not explicitly recognized, and detect purifying selection at most loci.ConclusionsOur work resolves previous inconsistencies in the phylogeny of rodent malaria parasites, provides estimates of important parameters that relate to the parasite’s natural history and provides a much-needed evolutionary context for understanding diverse biological aspects from the cross-reactivity of immune responses to parasite mating patterns.

Optimal sample storage and extraction procotols for reliable multilocus genotyping of the human parasite Schistosoma mansoni

Genotyping individual larval stages and eggs of natural parasite populations is complicated by the difficulty of obtaining reliable genotypes from low quantity DNA template. A suitable storage and extraction protocol, together with a thorough quantification of genotyping errors are therefore crucial for molecular epidemiological studies. Here we test the robustness, handling time, ease of use, cost effectiveness and success rate of various fixation (Whatman FTA® Classic and Elute Cards, 70% EtOH and RNAlater®) and subsequent DNA extraction methods (commercial kits and proteinase K protocol). None of these methods require a cooling chain and are therefore suitable for field collection. Based on a multiplex microsatellite PCR with nine loci the success and reliability of each technique is evaluated by the proportion of samples with at least eight scored loci and the proportion of genotyping errors. If only the former is taken into account, FTA® Elute is recommended (83% success; 44% genotyping error; 0.2€/sample; 1h 20m handling time). However, when also considering the genotyping errors, handling time and ease of use, we opt for 70% EtOH with the 96-well plate technology followed by a simple proteinase K extraction (73% success; 0% genotyping error; 0.2€/sample; 15m handling time). For eggs we suggest (1) to pool all eggs per person in 1.5ml tubes filled with 70% EtOH for transport and (2) to identify each egg to species level prior to genotyping. To this end we extended the Rapid diagnostic PCR developed by Webster et al. (2010) with a S. mansoni-specific primer to discriminate between S. mansoni, S. haematobium and S. bovis in a single PCR reaction. The success rate of genotyping eggs was 75% (0% genotyping error). This is the first study to incorporate genotyping errors through re-amplification for the evaluation of schistosome sampling protocols and the identification of error-prone loci.

Life history and host utilization pattern of a strepsipteran parasite (Insecta: Strepsiptera) on the Blissine bugs (Hemiptera: Lygaeidae) living under dwarf bamboo leaf sheaths

The blissine bug Macropes obnubilus, which lives aggregately under leaf sheaths of the evergreen dwarf bamboo, is infected by the strepsipteran parasite Blissoxenos esakii. To determine the ecological properties of the bug–strepsipteran system, we conducted field surveys of natural populations of Macropes bugs and Blissoxenos parasites. The parasitism rate of strepsipterans was high throughout the year and was significantly higher in female than in male bugs. Blissoxenos adult males emerged mainly in May, and neotenic adult females released triungulins in August. The triungulins invaded host nymphs, but subsequent larval development did not occur before the bugs matured. At most, two strepsipterans could mature in a host because of spatial limitations. The mortality of triply or more parasitized bugs was significantly higher than that of singly or doubly parasitized bugs, which survived longer than uninfected ones. The heavy strepsipteran infection profoundly affected the host population by causing host reproductive failure.

Toward a General Theory for How Climate Change Will Affect Infectious Disease

Toward a General Theory for How Climate Change Will Affect Infectious Disease

Inferred relatedness and heritability in malaria parasites.

Malaria parasites vary in phenotypic traits of biomedical or biological interest such as growth rate, virulence, sex ratio and drug resistance, and there is considerable interest in identifying the genes that underlie this variation. An important first step is to determine trait heritability (H2). We evaluate two approaches to measuring H2 in natural parasite populations using relatedness inferred from genetic marker data. We collected single-clone Plasmodium falciparum infections from 185 patients from the Thailand–Burma border, monitored parasite clearance following treatment with artemisinin combination therapy (ACT), measured resistance to six antimalarial drugs and genotyped parasites using 335 microsatellites. We found strong relatedness structure. There were 27 groups of two to eight clonally identical (CI) parasites, and 74 per cent of parasites showed significant relatedness to one or more other parasites. Initially, we used matrices of allele sharing and variance components (VC) methods to estimate H2. Inhibitory concentrations (IC50) for six drugs showed significant H2 (0.24 to 0.79, p = 0.06 to 2.85 × 10−9), demonstrating that this study design has adequate power. However, a phenotype of current interest—parasite clearance following ACT—showed no detectable heritability (H2 = 0–0.09, ns) in this population. The existence of CI parasites allows the use of a simple ANOVA approach for quantifying H2, analogous to that used in human twin studies. This gave similar results to the VC method and requires considerably less genotyping information. We conclude (i) that H2 can be effectively measured in malaria parasite populations using minimal genotype data, allowing rational design of genome-wide association studies; and (ii) while drug response (IC50) shows significant H2, parasite clearance following ACT was not heritable in the population studied.

Genetic changes during laboratory propagation: Copy number At the reticulocyte-binding protein 1 locus of Plasmodium falciparum

Comparative genomic hybridization studies have revealed elevated copy number (CN) at the reticulocyte-binding protein 1 gene (PfRh1) in fast growing lab-adapted parasites, while genetic manipulation demonstrates a causal link between cell invasion and PfRh1 CN. We therefore examined PfRh1 copy number variation (CNV) in 202 single clone parasite isolates from four countries to quantify the extent of CNV within natural populations. Surprisingly, we found that no natural parasite infections showed elevated CN. In contrast, 4/28 independent laboratory reference strains show elevated CN. One possibility is that amplification of PfRh1 (or neighboring loci) is selected during laboratory culture. In the case of FCR3 group of parasites, clone trees show that PfRh1 amplification arose in laboratory lines following establishment in culture. These data show that CNV at PfRh1 is rare or non-existent in natural populations, but can arise during laboratory propagation. We conclude that PfRh1 CNV is not an important determinant of gene expression, cell invasion or growth rate in natural parasite populations.