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Schistosome Population Genetic Structure: When Clumping Worms is not Just Splitting Hairs
Abstract
Schistosomiasis is a major public health problem, affecting over 200 million people worldwide. Although Schistosoma mansoni has been studied rigorously in an attempt to provide a vaccine based on a number of candidate antigens, there has been a lack of complementary effort to determine the range and distribution of variation in representative molecules throughout natural populations. Here, Jason Curtis and Dennis Minchella highlight current (and suggest future) research efforts aimed at assessing genetic variation in schistosome populations, and call for a more robust consideration of schistosome population genetics.
... The AMOVA analysis also illustrated that S. turkestanicum did form distinct subpopulations between hosts, which would exist as unique interbreeding populations within a single host. Such patterns of genetic differentiation within parasite populations have been seen between individuals of the same host species within a given locality several times in both S. mansoni (Curtis and Minchella, 2000;Sire et al. 2001;Curtis et al. 2002;Prugnolle et al. 2005 Age and movement of the host could account for the variation seen in the Hungarian S. turkestanicum as all adults were collected from livers of old large red deer stags, which are known to be wide roaming taking advantage of several wallowing sites all of which being potentially infective foci, along the Danube River and associated water bodies (Majoros et al. 2010). It was also suggested that concomitant immunity could play a role, whereby an individual host develops partial immunity to a specific schistosome genotype. ...
... ;Van den Broeck et al. 2014) and S. japonicum(Rudge et al. 2009;Lu et al. 2011). Genetic diversity of parasites within a single host is thought to be predetermined by host mobility and their potential to visit several different infection foci, the age of the host and host immunity(Curtis and Minchella, 2000;Curtis et al. 2002;Lu et al. 2011;Van den Broeck et al. 2014). The high levels of genetic diversity within an individual host could relate to the number of transmission sites visited leading to the host becoming a 'genetic mixing bowl' for parasites, thus the offspring are genetically well mixed and parasite variation is randomly distributed between hosts(Curtis and Minchella, 2000;Curtis et al. 2002;Criscione et al. 2005). ...
... Genetic diversity of parasites within a single host is thought to be predetermined by host mobility and their potential to visit several different infection foci, the age of the host and host immunity(Curtis and Minchella, 2000;Curtis et al. 2002;Lu et al. 2011;Van den Broeck et al. 2014). The high levels of genetic diversity within an individual host could relate to the number of transmission sites visited leading to the host becoming a 'genetic mixing bowl' for parasites, thus the offspring are genetically well mixed and parasite variation is randomly distributed between hosts(Curtis and Minchella, 2000;Curtis et al. 2002;Criscione et al. 2005). However, mixing and development of diverse infrapopulations is time dependant and Van den Broeck et al. (2014) illustrated low genetic heterogeneity of S. mansoni infecting children in Senegal compared with the highly diverse populations found in adults. ...
High levels of molecular diversity were identified in mitochondrial cytochrome c oxidase (cox1) gene sequences of Schistosoma turkestanicum from Hungary. These cox1 sequences were all specific to Hungary which contrasted with the low levels of diversity seen in the nuclear internal transcribed spacer region (ITS) sequences, the majority of which were shared between China and Iran isolates. Measures of within and between host molecular variation within S. turkestanicum showed there to be substantial differences in molecular diversity, with cox1 being significantly more diverse than the ITS. Measures of haplotype frequencies revealed that each host contained its own subpopulation of genetically unique parasites with significant levels of differentiation. Pairwise mismatch analysis of cox1 sequences indicated S. turkestanicum populations to have a bimodal pairwise difference distribution and to be stable unlike the ITS sequences, which appeared to have undergone a recent population expansion event. Positive selection was also detected in the cox1 sequences, and biochemical modelling of the resulting protein illustrated significant mutational events causing an alteration to the isoelectric point of the cox1 protein, potentially altering metabolism. The evolutionary signature from the cox1 indicates local adaptation and long establishment of S. turkestanicum in Hungary with continual introgression of nuclear genes from Asian isolates. These processes have led to the occurrence of mito-nuclear discordance in a schistosome population
... Although only a few population genetic studies have been performed on schistosomes to date, they have identified significant genetic diversity in adult worms within both human and rodent hosts. [7][8][9][10][11] There is currently no effective vaccine against schistosomiasis and interruption of the transmission cycle through mollusciciding, biological control of the intermediate snail hosts and health education have generally proved insufficient control methods on their own. 2 Chemotherapy with praziquantel (PZQ) is therefore the mainstay for schistosomiasis control 12 and PZQ will remain the only drug of choice for several years. 12 One major control program, the Schistosomiasis Control Initiative (SCI), was established in 2002 to assist selected sub-Saharan African countries to establish sustainable schistosomiasis control. ...
... The unpredicted apparent "bottleneck" imposed by one round of MDA on schistosome population genetics has several putative explanations, all of which require further investigation. It may indicate that not all parasites present in r efugia at the time of treatment contribute to the re-infection of children in this age group (7)(8)(9)(10)(11), and hence the "effective reservoir" may be smaller than previously thought. One could speculate that parasite genetic differentiation may occur between human hosts of different ages caused by immunological differences, with the untreated pre-school children, teenagers, and adults not treated in this study harboring different parasite genotypes, which may be less likely to re-infect children in the study group ages (7)(8)(9)(10)(11). ...
... It may indicate that not all parasites present in r efugia at the time of treatment contribute to the re-infection of children in this age group (7)(8)(9)(10)(11), and hence the "effective reservoir" may be smaller than previously thought. One could speculate that parasite genetic differentiation may occur between human hosts of different ages caused by immunological differences, with the untreated pre-school children, teenagers, and adults not treated in this study harboring different parasite genotypes, which may be less likely to re-infect children in the study group ages (7)(8)(9)(10)(11). This reduction in genetic diversity as a result of one MDA treatment may therefore result from a large number of the genotypes of parasites currently adapted to infecting humans of the group 7-11 years of age having been killed by chemotherapy, with not all parasites present in the "reservoir" adapted to re-infect these children or being present in the specific transmission foci that these children frequent. ...
Recent shifts in global health policy have led to the implementation of mass drug administration (MDA) for neglected tropical diseases. Here we show how population genetic analyses can provide vital insights into the impact of such MDA on endemic parasite populations. We show that even a single round of MDA produced a genetic bottleneck with reductions in a range of measures of genetic diversity of Schistosoma mansoni. Phylogenetic analyses and indices of population differentiation indicated that schistosomes collected in the same schools in different years were more dissimilar than those from different schools collected within either of the study's 2 years, in addition to distinguishing re-infection from non-clearance (that might indicate putatively resistant parasites) from within those children infected at both baseline and follow-up. Such unique results illustrate the importance of genetic monitoring and examination of long lived multi-cellular parasites such as these under novel or increased chemotherapeutic selective pressures.
... Microsatellite markers have recently been used in determining S. japonicum genetic diversity and in estimating the levels of gene flow in the population. Previous studies have recommended the use of microsatellite markers to determine schistosome genetic diversity because of their codominant expression and their ability to serve as neutral markers [4,5]. The easy observation of heterozygosity and the reasonable number of alleles per polymorphic locus in the samples make microsatellite analysis a more powerful tool in genetic studies than the use of rapid amplified polymorphic DNA (RAPD) and mitochondrial DNA [6]. ...
... Genetic diversity found in this study among the parasite population in each endemic site in the Philippines is vital in the parasites' ability to survive the effect of selective pressures such as those brought by drug treatment. At the same time, selection pressures increase the frequency of favorable alleles across all populations [4]. In this sense, the present finding of high diversity among the parasite populations imply that the MDA with praziquantel has varying degree of impact in interrupting the parasite's life cycle. ...
Background:
Microsatellites have been found to be useful in determining genetic diversities of various medically-important parasites which can be used as basis for an effective disease management and control program. In Asia and Africa, the identification of different geographical strains of Schistosoma japonicum, S. haematobium and S. mansoni as determined through microsatellites could pave the way for a better understanding of the transmission epidemiology of the parasite. Thus, the present study aims to apply microsatellite markers in analyzing the populations of S. japonicum from different endemic areas in the Philippines for possible strain differentiation.
Methodology/ principal findings:
Experimental mice were infected using the cercariae of S. japonicum collected from infected Oncomelania hupensis quadrasi snails in seven endemic municipalities. Adult worms were harvested from infected mice after 45 days of infection and their DNA analyzed against ten previously characterized microsatellite loci. High genetic diversity was observed in areas with high endemicity. The degree of genetic differentiation of the parasite population between endemic areas varies. Geographical separation was considered as one of the factors accounting for the observed difference between populations. Two subgroups have been observed in one of the study sites, suggesting that co-infection with several genotypes of the parasite might be present in the population. Clustering analysis showed no particular spatial structuring between parasite populations from different endemic areas. This result could possibly suggest varying degrees of effects of the ongoing control programs and the existing gene flow in the populations, which might be attributed to migration and active movement of infected hosts from one endemic area to another.
Conclusions/ significance:
Based on the results of the study, it is reasonable to conclude that genetic diversity could be one possible criterion to assess the infection status in highly endemic areas. Genetic surveillance using microsatellites is therefore important to predict the ongoing gene flow and degree of genetic diversity, which indirectly reflects the success of the control program in schistosomiasis-endemic areas.
... However, the majority of animal macroparasites (for example, flatworms, nematodes) release their offspring into the environment or multiply within one or more intermediate host species, mostly resulting in high genetic diversity levels similar to those reported in free-living organisms (Bush et al., 2001). It has been suggested that the high genetic diversity of macroparasites within individual hosts may reflect the tendency of these hosts to sample a number of transmission sites (Anderson et al., 1995;Nadler, 1995), thereby becoming 'genetic mixing bowls' for parasite genes (Curtis and Minchella, 2000;Curtis et al., 2002). The result is that parasite offspring are well mixed in the environment and that parasite genetic variation is randomly distributed between hosts (Criscione et al., 2005). ...
... Hosts serve as genetic mixing bowls for S. mansoni parasites Given the long life expectancy of schistosome worms, older hosts may have acquired genetically unrelated parasites in space (if the action radius of the host increases with its age) and in time (if cercariae at a given transmission site are genetically dissimilar between time points). Human hosts then become 'genetic mixing bowls' , where the accumulation of unrelated parasites during their lives results in an increase in the diversity of the respective parasite population (Curtis and Minchella, 2000). On the other hand, children may be more exposed to related parasites than adults owing to differences in water contact behavior. ...
The size, structure and distribution of host populations are key determinants of the genetic composition of parasite populations. Despite the evolutionary and epidemiological merits, there has been little consideration of how host heterogeneities affect the evolutionary trajectories of parasite populations. We assessed the genetic composition of natural populations of the parasite Schistosoma mansoni in northern Senegal. A total of 1346 parasites were collected from 14 snail and 57 human hosts within three villages and individually genotyped using nine microsatellite markers. Human host demographic parameters (age, gender and village of residence) and co-infection with Schistosoma haematobium were documented, and S. mansoni infection intensities were quantified. F-statistics and clustering analyses revealed a random distribution (panmixia) of parasite genetic variation among villages and hosts, confirming the concept of human hosts as 'genetic mixing bowls' for schistosomes. Host gender and village of residence did not show any association with parasite genetics. Host age, however, was significantly correlated with parasite inbreeding and heterozygosity, with children being more infected by related parasites than adults. The patterns may be explained by (1) genotype-dependent 'concomitant immunity' that leads to selective recruitment of genetically unrelated worms with host age, and/or (2) the 'genetic mixing bowl' hypothesis, where older hosts have been exposed to a wider variety of parasite strains than children. The present study suggests that host-specific factors may shape the genetic composition of schistosome populations, revealing important insights into host-parasite interactions within a natural system.Heredity advance online publication, 12 March 2014; doi:10.1038/hdy.2014.13.
... 1999). Microsatellite markers have recently been proposed as an ideal technique that future schistosome studies could develop and utilize, due in part to their ability to detect alleles at single loci in single individuals (Curtis & Minchella 2000). Here we report on the isolation of novel microsatellite markers for use in S. mansoni population analyses. ...
... Moreover, the additional nine novel markers described here were also found to have much higher heterozygosity figures compared to that for Guadeloupe where the H O figures ranged from only 0.06 -0.44 in eight loci. One potential explanation for this discrepancy may be restricted gene flow within the New World Guadeloupe island population, as compared to that of the Old World populations studied here, in much the same way as is evident within locally adapted laboratory schistosome populations (Curtis & Minchella, 2000). Indeed, the ability of microsatellites to reveal here subtle genetic diversity within geographically unrelated populations and individuals may highlight their suitability for large scale population genetic surveys. ...
The ability of microsatellite loci to reveal genetic diversity between individuals of the digenean trematode Schistosoma mansoni was investigated. A total of 10 polymorphic microsatellite markers were isolated and optimized, allowing the variability at each locus amplified within 12 individuals from three African populations to be assessed. Allelic diversity and observed heterozygosity (HO) figures ranged from 2 to 6 and 0.33 to 1.00, respectively. These results indicated high variability both between individuals and populations, highlighting the suitability of these microsatellites for future population genetic surveys.
... Metrics of genetic diversity, such as allelic richness (AR), and observed and expected heterozygosity (respectively H O and H E ), were originally developed for sexual, free-living organisms undergoing direct life-cycles. How these measures perform in parasitic species is still poorly understood (Blouin et al., 1999;Curtis and Minchella, 2000). Whilst a profound understanding of the genetic structure of parasite populations, combined with their epidemiology, can lead to insights on important biological processes such as the underlying level of transmission, gene flow, size of reproductive units, breeding strategy, and emergence of anthelminthic resistance, amongst others (Nadler, 1995;Fisher and Viney, 1998;Paterson and Viney, 2000;Prugnolle et al., 2005a), the ability of current tools to achieve this is uncertain, given how surprisingly little is known about the population genetic structure of most species of parasitic helminths (Prugnolle et al., 2005a). ...
... Although there will likely be some exchange of genetic information within and between parasite populations (Rollinson and Simpson, 1987), the condition of panmixia, upon which much of the field of population genetics is based, will not always be fulfilled, leading, under these circumstances, to questioning the applicability of some of the standard measures of genetic diversity (AR, H O and H E for example). Further, during the transmission process, each host is unlikely to acquire worms randomly from the whole population (Blouin et al., 1999;Curtis and Minchella, 2000), and the asexual stage of the life-cycle will likely produce large numbers of genetically related clones (Sire et al., 2001;Yin et al., 2008;Lu et al., 2010). This variance in clonal reproductive success, combined with synchronous release of cercariae and rare infection events can lead to hosts being infected with parasites heavily biased towards a few genotypes (Waples, 1998;Sire et al., 1999;Prugnolle et al., 2005a;Lu et al., 2010), as well as to the possibility of heterozygote deficiency, selfing, and mating within clones (Prugnolle et al., 2005a). ...
Detecting potential changes in genetic diversity in schistosome populations following chemotherapy with praziquantel (PZQ) is crucial if we are to fully understand the impact of such chemotherapy with respect to the potential emergence of resistance and/or other evolutionary outcomes of interventions. Doing so by implementing effective, and cost-efficient sampling protocols will help to optimise time and financial resources, particularly relevant to a disease such as schistosomiasis currently reliant on a single available drug. Here we explore the effect on measures of parasite genetic diversity of applying various field sampling approaches, both in terms of the number of (human) hosts sampled and the number of transmission stages (miracidia) sampled per host for a Schistosoma mansoni population in Tanzania pre- and post-treatment with PZQ. In addition, we explore population structuring within and between hosts by comparing the estimates of genetic diversity obtained assuming a 'component population' approach with those using an 'infrapopulation' approach. We found that increasing the number of hosts sampled, rather than the number of miracidia per host, gives more robust estimates of genetic diversity. We also found statistically significant population structuring (using Wright's F-statistics) and significant differences in the measures of genetic diversity depending on the parasite population definition. The relative advantages, disadvantages and, hence, subsequent reliability of these metrics for parasites with complex life-cycles are discussed, both for the specific epidemiological and ecological scenario under study here and for their future application to other areas and schistosome species.
... In contrast, several factors can favor increase of the genetic diversity of schistosomes: mobility and long lifespan of the definitive host, genotype specific re-infection, snail-parasite compatibility and the rapid turnover of infected snail hosts (117). Within this context, the definitive host acts as a 'genetic mixing bowl' for the parasite (124). Population genetic studies have reported panmictic populations (all individuals within the population are potential partners) with most genetic variation and diversity occurring within individual human hosts rather than between different hosts (66,125,126). ...
Schistosomiasis is a neglected tropical disease (NTD) caused by parasitic trematodes belonging to the Schistosoma genus. The mainstay of schistosomiasis control is the delivery of a single dose of praziquantel (PZQ) through mass drug administration (MDA) programs. These programs have been successful in reducing the prevalence and intensity of infections. Due to the success of MDA programs, the disease has recently been targeted for elimination as a public health problem in some endemic settings. The new World Health Organization (WHO) treatment guidelines aim to provide equitable access to PZQ for individuals above two years old in targeted areas. The scale up of MDA programs may heighten the drug selection pressures on Schistosoma parasites, which could lead to the emergence of PZQ resistant schistosomes. The reliance on a single drug to treat a disease of this magnitude is worrying should drug resistance develop. Therefore, there is a need to detect and track resistant schistosomes to counteract the threat of drug resistance to the WHO 2030 NTD roadmap targets. Until recently, drug resistance studies have been hindered by the lack of molecular markers associated with PZQ resistance. This review discusses recent significant advances in understanding the molecular basis of PZQ action in S. mansoni and proposes additional genetic determinants associated with PZQ resistance. PZQ resistance will also be analyzed in the context of alternative factors that may decrease efficacy within endemic field settings, and the most recent treatment guidelines recommended by the WHO.
... In the majority of cases, each child hosted an infrapopulation different from the other children. Our finding does not corroborate the "genetic mixing bowl hypothesis" [16]. The authors reported that the long-living definitive hosts cumulate genotypes coming from several short-living infected intermediate host snails, which can lead to homogeneity of infrapopulations. ...
While population genetics of Schistosoma haematobium have been investigated in West Africa, only scant data are available from Côte d’Ivoire. The purpose of this study was to analyze both genetic variability and genetic structure among S. haematobium populations and to quantify the frequency of S. haematobium × S. bovis hybrids in school-aged children in different parts of Côte d’Ivoire. Urine samples were subjected to a filtration method and examined microscopically for Schistosoma eggs in four sites in the western and southern parts of Côte d’Ivoire. A total of 2692 miracidia were collected individually and stored on Whatman ® FTA cards. Of these, 2561 miracidia were successfully genotyped for species and hybrid identification using rapid diagnostic multiplex mitochondrial cox1 PCR and PCR Restriction Fragment Length Polymorphism (PCR-RFLP) analysis of the nuclear ITS2 region. From 2164 miracidia, 1966 (90.9%) were successfully genotyped using at least 10 nuclear microsatellite loci to investigate genetic diversity and population structure. Significant differences were found between sites in all genetic diversity indices and genotypic differentiation was observed between the site in the West and the three sites in the East. Analysis at the infrapopulation level revealed clustering of parasite genotypes within individual children, particularly in Duekoué (West) and Sikensi (East). Of the six possible cox1 - ITS2 genetic profiles obtained from miracidia, S. bovis cox1 × S. haematobium ITS2 (42.0%) was the most commonly observed in the populations. We identified only 15 miracidia (0.7%) with an S. bovis cox1 × S. bovis ITS2 genotype. Our study provides new insights into the population genetics of S. haematobium and S. haematobium × S. bovis hybrids in humans in Côte d’Ivoire and we advocate for researching hybrid schistosomes in animals such as rodents and cattle in Côte d’Ivoire.
... Although the role of the parasite genetic diversity in differential host response is well known for microparasites (134,135), there is comparably less known regarding the potential importance of macroparasite genetic heterogeneities in general, and schistosomes in particular, on disease epidemiology (136). Laboratory studies, however, have shown that schistosomes strains or populations from the same or different geographical locations can show a number of differences in biological traits related to transmission and virulence such as infectivity, egg production, pathogenicity and response to chemotherapy (11,(137)(138)(139)(140)(141)(142)(143)(144)(145)(146). For example, some S. haematobium strains studied in the laboratory show different levels of mortality and worm recovery in hamsters as well as differences in snail infectivity (147), while different S. mansoni strains have been shown to induce disparate rates of hepatomegaly and splenomegaly despite comparable fecal egg counts (137). ...
Schistosomiasis is the second most important human parasitic disease in terms of socioeconomic impact, causing great morbidity and mortality, predominantly across the African continent. For intestinal schistosomiasis, severe morbidity manifests as periportal fibrosis (PPF) in which large tracts of macro-fibrosis of the liver, visible by ultrasound, can occlude the main portal vein leading to portal hypertension (PHT), sequelae such as ascites and collateral vasculature, and ultimately fatalities. For urogenital schistosomiasis, severe morbidity manifests as pathology throughout the urinary system and genitals, and is a definitive cause of squamous cell bladder carcinoma. Preventative chemotherapy (PC) programmes, delivered through mass drug administration (MDA) of praziquantel (PZQ), have been at the forefront of schistosomiasis control programmes in sub-Saharan Africa since their commencement in Uganda in 2003. However, despite many successes, ‘biological hotspots’ (as distinct from ‘operational hotspots’) of both persistent high transmission and morbidity remain. In some areas, this failure to gain control of schistosomiasis has devastating consequences, with not only persistently high infection intensities, but both “subtle” and severe morbidity remaining prevalent. These hotspots highlight the requirement to revisit research into severe morbidity and its mechanisms, a topic that has been out of favor during times of PC implementation. Indeed, the focality and spatially-structured epidemiology of schistosomiasis, its transmission persistence and the morbidity induced, has long suggested that gene-environmental-interactions playing out at the host-parasite interface are crucial. Here we review evidence of potential unique parasite factors, host factors, and their gene-environmental interactions in terms of explaining differential morbidity profiles in the human host. We then take the situation of schistosomiasis mansoni within the Albertine region of Uganda as a case study in terms of elucidating the factors behind the severe morbidity observed and the avenues and directions for future research currently underway within a new research and clinical trial programme (FibroScHot).
... Opposite to this, the definitive host mobility, the rapid turnover of infected snails, the snail-parasite compatibility, the long lifespan of the definitive host and the genotype dependent host re-infection (Beltran et al., 2011), should all increase the parasite genetic diversity within the definitive host. In this context, some authors have proposed that the definitive host is a 'genetic mixing bowl' for the parasite (Curtis and Minchella, 2000). The 'genetic mixing bowl' hypothesis was validated by several studies. ...
Blood flukes within the genus Schistosoma (schistosomes) are responsible for the major disease, schistosomiasis, in tropical and sub-tropical areas. This disease is predominantly present on the African continent with more than 85% of the human cases. Schistosomes are also parasites of veterinary importance infecting livestock and wildlife. Schistosoma population genetic structure and diversity are important characteristics that may reflect variations in selection pressures such as those induced by host (mammalian and snail) environments, habitat change, migration and also treatment/control interventions, all of which also shape speciation and evolution of the whole Schistosoma genus. Investigations intoschistosome population genetic structure, diversity and evolution has been an area of important debate and research. Supported by advancesin molecular techniques with capabilitiesfor multi-locus genetic analyses for single larvae schistosome geneticinvestigations have greatly progressed in the last decade. This paper aims to review the genetic studies of both animal and human infecting schistosome. Population genetic structures are reviewed at different spatial scales: local, regional or continental (i.e. phylogeography). Within species genetic diversities are discussed compared and the compounding factors discussed, including the effect of mass drug administration. Finally, the ability for intra-species hybridisation questions species integrities and poses many questions in relation to the natural epidemiology of co-endemic species. Here we review molecularly confirmed hybridisation events (in relation to human disease) and discuss the possible impact for ongoing and future control and elimination.
... Host behavior and/or physiology is expected to shape the genetic composition of the parasite infrapopulation that they host. It has been proposed that the high genetic diversity of parasites is the result of host behavior in sampling several parasites in space and/or in time, resulting in 'genetic mixing bowl' for parasite genes [62][63][64]. A positive link between age of the patient and S. mansoni parasite diversity has already been observed in Senegal [64]. ...
Background:
Schistosomiasis is a chronic parasitic disease, that affects over 207 million people and causes over 200,000 deaths annually, mainly in sub-Saharan Africa. Although many health measures have been carried out to limit parasite transmission, significant numbers of non-human primates such as Chlorocebus aethiops (Ch. aethiops) (vervet) and Papio anubis (baboon) are infected with S. mansoni, notably in Ethiopia, where they are expected to have potentially significant implications for transmission and control efforts.
Objective:
The objective of this study was to assess and compare the genetic diversity and population structure of S. mansoni isolates from human and non-human primates free-ranging in close proximity to villages in selected endemic areas of Ethiopia.
Methods:
A cross-sectional study was conducted in three transmission sites: Bochesa, Kime and Fincha. A total of 2,356 S. mansoni miracidia were directly isolated from fecal specimens of 104 hosts (i.e. 60 human hosts and 44 non-human primates). We performed DNA extraction and PCR amplification using fourteen microsatellite loci.
Results:
At population scale we showed strong genetic structure between the three sample sites. At the definitive host scale, we observed that host factors can shape the genetic composition of parasite infra-populations. First, in male patients, we observed a positive link between parasite genetic diversity and the age of the patients. Second, we observed a difference in genetic diversity which was high in human males, medium in human females and low in non-human primates (NHPs). Finally, whatever the transmission site no genetic structure was observed between human and non-human primates, however, there appears to be little barriers, if any, host specificity of the S. mansoni populations with cross-host infections.
Conclusion:
Occurrence of infection of a single host with multiple S. mansoni strains and inter- and intra-host genetic variations was observed. Substantial genetic diversity and gene flow across the S. mansoni population occurred at each site and non-human primates likely play a role in local transmission and maintenance of infection. Therefore, public health and wildlife professionals should work together to improve disease control and elimination strategies.
... Schistosomiasis is a debilitating disease with high economic impact, affects many people all over the world leading to high morbidity and mortality (Curtis and Minchella, 2000). Animal models are used as tools for understanding the host-parasite relationships. ...
This study aimed to correlate the histopathological changes in mice liver with alterations either in liver tissue antioxidants enzymes (catalase and reduced glutathione (GSH)), oxidative stress marker (malondialdehyde (MDA)) or in serum liver function parameters (Alanine aminotransferase (ALT), aspartate aminotransferase (AST), total protein (T.P.) and albumin) to predict the outcome of schistosomiasis. Fourty male Swiss albino mice were used in this study and infected with Schistosoma mansoni for 2, 4, 6 and 8 weeks (8 mice for each group), while, the uninfected mice were used as negative control. Liver tissue antioxidants enzymes, oxidative stress marker and serum liver function parameters were determined in coincide with the liver tissue histopathological changes. All selected biochemical makers showed a strong significant positive correlation (p < 0.05) with liver histopathology score except serum albumin and liver tissue catalase enzyme. The last two parameters exhibited negative correlation with liver histopathology score. These results revealed that the more increase in the level of AST, ALT, T.P. and globulin in serum or liver tissue MDA and GSH indicating severs histopathological changed into the affected liver and hopeless prognosis is expected. On contrary, the increase in albumin level in serum or catalase level in liver tissue of affected patient/animal demonstrating mild liver histopathological changes. Subsequently, good prognosis and response to anti-schistosomal treatment will be predictable. This study opens the way to predict the outcome of schistosomiasis through easy and rapid biochemical test. Therefore, other studies are required to apply such correlation with other biochemical parameters especially that synthesized into the liver.
... This is relevant for schistosomes, considering their complex life cycle which includes an asexual reproduction step within a snail host. It also relies less on overall levels of genetic differentiation among populations than outlier detection methods (Hancock et al., 2010) and is therefore suitable for schistosome populations as they often show a high diversity within a host and low levels of genetic differentiation among hosts (Curtis and Minchella, 2000;Van den Broeck et al., 2014). Allele frequencies of the offspring (miracidia) were grouped per host individual, hereafter referred to as infrapopulations. ...
Here we assess the role of parasite genetic variation in host disease phenotype in human schistosomiasis by implementing concepts and techniques from environmental association analysis in evolutionary epidemiology. Schistosomiasis is a tropical disease that affects more than 200 million people worldwide and is caused by parasitic flatworms belonging to the genus Schistosoma. While the role of host genetics has been extensively studied and demonstrated, nothing is yet known on the contribution of parasite genetic variation to host disease phenotype in human schistosomiasis. In this study microsatellite genotypes of 1561 Schistosoma mansoni larvae collected from 44 human hosts in Senegal were linked to host characteristics such as age, gender, infection intensity, liver and bladder morbidity by means of multivariate regression methods (on each parasite locus separately). This revealed a highly significant association between allelic variation at the parasite locus L46951 and host infection intensity and bladder morbidity. Locus L46951 is located in the 3' untranslated region of the cGMP-dependent protein kinase gene that is expressed in reproductive organs of adult schistosome worms and appears to be linked to egg production. This putative link between parasite genetic variation and schistosomiasis disease phenotype sets the stage for further functional research.
... Both the genetic variation in populations of Biomphalaria and the phylogeny of the this genus have become subject of nucleic acid sequence-based analysis (e.g. Curtis & Minchella, 2000;Campbell, Jones, Lockyer, Hughes, Brown, Noble, & Rollinson, 2000). ...
Full-length actin-encoding sequences were PCR-amplified from genomic DNA of six planorbid species; Biomphalaria glabrata (Say; M-line strain), B. alexandrina (Ehrenberg), B. pfeifferi (Krauss), B. tenagophila (Orbigny), B. obstructa (Morelet) and Helisoma trivolvis (Say), using primers designed from a previously reported B. glabrata cytoplasmic (β) actin cDNA. The amplified sequences contained two conserved exons (126 nt and 1005 nt, respectively), separated by an intron that varied in size between snail species (ranging from 671 to 794 nt). Sequence similarities occurred between the introns of the actin genes from B. glabrata, B. alexandrina and B. pfeifferi and between those from B. tenagophila and B. obstructa, yet considerable differences were evident between these two groups and the intron derived from H. trivolvis. Analysis of exons for sequence similarities, the presence of conserved residues (deduced amino acids), and construction of gene trees indicated that these planorbid genes encode cytoplasmic (β) actins rather than muscular (α) actins. Southern blotting and hybridisation experiments suggested that B. glabrata and H. trivolvis may have multiple (up to 5) actin genes, and it can not be ruled out that actin sequences obtained from different planorbid species were derived from paralogous genes. Interestingly however, the gene trees resolved actins derived from gastropod, cephalopod and bivalve molluscs. The sequences presented add to the growing body of information on the molecular biology of planorbid snails.
... Little is known about the extent of genetic diversity of S. haematobium within its definitive host, humans. Understanding the genetic structuring of populations at each stage of the life cycle is essential to account for the creation of diversity and its maintenance in natural populations of parasites [19]. Genetic variability among parasite populations is an important factor in their potential for producing harmful effects on the human populations. ...
Background
Over 650 million people globally are at risk of schistosomiasis infection, while more than 200 million people are infected of which the higher disease rates occur in children. Eighty three students between 6-20 years (mean 12.45 ± 3.2) from Quran School for boys in Radwan village, Gezira state were recruited to investigate for the relationship between the genetic diversity of Schistosoma haematobium strains and the severity of the disease.
Method
Schistosoma haematobium infection was detected by filtration of urine. Ultrasonography was done on each study subject, while PCR technique was used for genotyping via random amplified polymorphic DNA (RAPD) with A01, A02, A12, Y20 and A13 primers. A01 primer gave three different genotypes (A01-1, A01-2 and A01-3).
Results
About 54.2% (45/83) were S. haematobium egg positive by urine filtration. On assessment of the upper and lower urinary tract by ultrasound technique, 61.4% (51/83) were positiveand73.3% (60/83) samples were PCR positive. No significant difference was found when comparing the three different genotypes with severity of the disease.
Conclusion
This study concludes that no association was found between the different genotypes of S.haemtobium and the severity of the disease. Examination of more samples from different areas to identify any possible differences between the parasites genes and disease severity was recommended.
... Microsatellites can be easily scored with the use of PCR with well-defined alleles (Ashley and Dow 1994). As microsatellites are thought to occur in all eukaryotic organisms, it is reasonable to expect that they could play an important role in the study of parasite population genetics (Curtis and Minchella 2000; Barker 2002). To develop variable microsatellite markers, it is necessary to perform library screens or to make use of DNA sequences available in public databases. ...
Schistosomes infect over 200 million people and 600 million are at risk. Genomics and post-genomic studies of schistosomes will contribute greatly to developing new reagents for diagnostic purposes and new vaccines that are of interest to the biotechnology industry. In this review, the most recent advances in these fields as well as new projects and future perspectives will de described. A vast quantity of data is publicly available, including short cDNA and genomic sequences, complete large genomic fragments, and the mitochondrial genomes of three species of the genus Schistosoma. The physical structure of the genome is being studied by physically mapping large genomic fragments and characterizing the highly abundant repetitive DNA elements. Bioinformatic manipulations of the data have already been carried out, mostly dealing with the functional analysis of the genes described. Specific search tools have also been developed. Sequence variability has been used to better understand the phylogeny of the species and for population studies, and new polymorphic genomic markers are currently being developed. The information generated has been used for the development of post-genomic projects. A small microarray detected genes that were differentially expressed between male and female worms. The identification of two-dimensional spots by mass spectrometry has also been demonstrated.
... 1997). More rigorous studies of population structure and genetic subdivision are needed to clarify our understanding of schistosome epidemiology (Curtis & Minchella 2000). Ideally, such studies would employ genetic markers that are highly polymorphic and inherited in a Mendelian codominant fashion. ...
Blood flukes in the genus Schistosoma are important human parasites in tropical regions. Genetic heterogeneity of the parasite contributes to the observed phenotypic variation in this host–parasite interaction and may play a role in disease epidemiology. In this paper, we describe the characterization of five polymorphic microsatellite loci from the human blood fluke Schistosoma mansoni, which can now be applied in assessments of schistosome genetic diversity. The five loci revealed extensive polymorphism, as 5–8 alleles per locus were detected among five isolates (from both human patients and snail intermediate hosts) from two Brazilian villages.
... How diverse the digenean S. japonicum really is in such a large geographical range has not been well assessed especially in mainland China. An accurate measure of its population genetic diversity is certainly needed to clarify our understanding on the epidemiology of schistosomiasis [17], which may be also useful for implementing control measures, and for developing drugs or potential vaccines, as worms of different genetic backgrounds may respond differently to such treatments [18,19]. In recent years, several molecular markers have been used to detect the variability of S. japonicum populations. ...
Schistosoma japonicum still causes severe parasitic disease in mainland China, but mainly in areas along the Yangtze River. However, the genetic diversity in populations of S. japonicum has not been well understood across its geographical distribution, and such data may provide insights into the epidemiology and possible control strategies for schistosomiasis.
In this study infected Oncomelania snails were collected from areas in the middle and lower (ML) reaches of the Yangtze River, including Hubei, Hunan, Anhui, Jiangxi and Jiangsu provinces, and in the upper reaches of the river, including Sichuan and Yunnan provinces in southwest (SW) China. The adult parasites obtained from experimentally infected mice using isolated cercariae were sequenced individually for several fragments of mitochondrial regions, including Cytb-ND4L-ND4, 16S-12S and ND1. Populations in the ML reaches exhibited a relatively high level of diversity in nucleotides and haplotypes, whereas a low level was observed for populations in the SW, using either each single fragment or the combined sequence of the three fragments. Pairwise analyses of F-statistics (Fst) revealed a significant genetic difference between populations in the ML reaches and those in the SW, with limited gene flow and no shared haplotypes in between. It is rather obvious that genetic diversity in the populations of S. japonicum was significantly correlated with the geographical distance, and the geographical separation/isolation was considered to be the major factor accounting for the observed difference between populations in the ML reaches and those in the SW in China.
S. japonicum in mainland China exhibits a high degree of genetic diversity, with a similar pattern of genetic diversity as observed in the intermediate host snails in the same region in China.
... In this study, we quantified within-host parasite genetic structure with six highly polymorphic microsatellite markers for S. japonicum (Shrivastava et al., 2003) and investigated the effect on this for a few potential influential factors. In addition, to test the hypothesis of the existence of host gender-specific structure (Prugnolle et al., 2003) for S. japonicum, we computed and compared statistical estimates of F IS (Wright, 1951), a measure of the extent of the departure from panmixia (an individual of a population is equally likely to mate with any other individual of that population) (Curtis and Minchella, 2000), and F ST (Weir and Cockerham, 1984), a measure of the difference in allelic frequencies between populations. To the best of our knowledge, this is the first study to examine the genetic structure of S. japonicum specifically at the level of individual hosts. ...
Schistosoma japonicum is an important parasite in terms of clinical, veterinary and socio-economic impacts, and rodents, a long neglected reservoir for the parasite, have recently been found to act as reservoir hosts in some endemic areas of China. Any difference in the host's biological characteristics and/or associated living habitats among rodents may result in different environments for parasites, possibly resulting in a specific population structure of parasites within hosts. Therefore knowledge of the genetic structure of parasites within individual rodents could improve our understanding of transmission dynamics and hence our ability to develop effective control strategies. In this study, we aimed to describe a host-specific structure for S. japonicum and its potential influencing factors. The results showed a significant genetic differentiation among hosts. Two factors, including sampling seasons and the number of miracidia genotyped per host, showed an effect on the genetic diversity of an infrapopulation through a univariable analysis but not a multivariable analysis. A possible scenario of clustered infection foci and the fact of multiple definitive host species, the latter of which is unique to S. japonicum compared with other schistosomes, were proposed to explain the observed results and practical implications for control strategies are recommended.
... Studying the genetic diversity of natural Schistosoma populations is complicated by the fact that adult worms are inaccessible in the blood circulatory system of the mammalian host, and the larval stages are very small (<200 mm; Rollinson and Simpson, 1987). From 2005 onwards, several protocols were developed to collect larval stages and eggs in the field (Shrivastava et al., 2005;Sorensen et al., 2006), thereby circumventing the ethical, technical and biological disadvantages of laboratory passage (Curtis and Minchella, 2000). These protocols require however a cooling chain (À20 8C), which is less amenable for field sampling. ...
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 20 m 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 1.5 ml 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.
... Geographic patterns in the pathology of schistosomiasis have been observed. However, they are frequently explained by the differences in the transmission intensity among foci, although some of the disease manifestations may be directly related to the parasite genetics (Curtis & Minchella, 2000). Furthermore, genetic differences among geographically separated strains cannot be evaluated without taking into account differences in a local scale, such as, within an endemic locality or municipality. ...
Schistosomiasis is an important parasitic disease that infects humans. Among the main species of schistosomes infecting humans are Schistosoma mansoni and Schistosoma haematobium. The accurate diagnosis of these parasitic infections will improve the success of disease control and management. Many studies proved that Schistosoma exhibit variations not only among species, but also among strains and between males and females, other than the obvious sexual characteristics associated with variations in the levels of infectivity, pathogenicity and immunogenicity. Our study on genetic diversity among populations of Schistosoma in Saudi Arabia gave a clear picture about this population and subsequently the method for controlling the parasites. From the examination of 40 collected samples of stools and urine, all urine samples were shown to be negative for the presence of Schistosoma eggs. On the other hand all stools samples were positive. Eggs were extracted from 20 faecal samples. Worms from each stool sample were collected, through completing the life cycle in white mice. The worms from each sample were collected and used for genetic diversity studies. Using three different primers, RAPD-PCR banding patterns showed that samples collected from Jeddah cluster together away from the rest of samples. Also, samples collected from Taif and Abha did not show any correlation between geographical location and DNA banding patterns.
... A major contributor to gene flow is most likely to be the free movement of the school children in the study area. These children potentially sample a number of transmission sites, thereby becoming 'genetic mixing bowls' for the parasite (Curtis and Minchella, 2000). The parasites also live long enough to integrate infections across multiple seasons. ...
A recently developed high-throughput technique that allows multi-locus microsatellite analysis of individual miracidia of Schistosoma mansoni was used to assess the levels of genetic diversity and population structure in 12 infrapopulations of the parasite, each infrapopulation derived from an infected school child from the Mwea area, central Kenya. The mean number of alleles per locus was in the range 8.22-10.22, expected heterozygosity in Hardy-Weinberg equilibrium was 0.68-0.70, and pairwise F(ST) values ranged from 0.16% to 3.98% for the 12 infrapopulations. Although the genetic diversity within each infrapopulation of S. mansoni in this area was generally high, low levels of genetic structure were observed, suggestive of high levels of gene flow among infrapopulations. Private alleles were found in 8 of the 12 infrapopulation, the highest number of private alleles recorded per infrapopulation was 3. Our data suggest that the level of gene flow among infrapopulations of S. mansoni in Mwea is extremely high, thus providing opportunity for spread of rare alleles, including those that may confer character traits such as drug resistance and virulence.
... Unfortunately the number of similar molecular epidemiological studies on multi-host parasites such as S. japonicum is limited. Several researchers have called for a better understanding of schistosome genetic diversity and structure, and have recommended the use of microsatellite markers for such studies [12,13,14]. Microsatellite markers isolated and characterized for S. japonicum have shown significant polymorphism between isolates, making them highly useful for studying the population genetic structure of the parasite [15]. ...
Schistosoma japonicum, which remains a major public health problem in the Philippines and mainland China, is the only schistosome species for which zoonotic transmission is considered important. While bovines are suspected as the main zoonotic reservoir in parts of China, the relative contributions of various non-human mammals to S. japonicum transmission in the Philippines remain to be determined. We examined the population genetics of S. japonicum in the Philippines in order to elucidate transmission patterns across host species and geographic areas.
S. japonicum miracidia (hatched from eggs within fecal samples) from humans, dogs, pigs and rats, and cercariae shed from snail-intermediate hosts, were collected across two geographic areas of Samar Province. Individual isolates were then genotyped using seven multiplexed microsatellite loci. Wright's F(ST) values and phylogenetic trees calculated for parasite populations suggest a high frequency of parasite gene-flow across definitive host species, particularly between dogs and humans. Parasite genetic differentiation between areas was not evident at the definitive host level, possibly suggesting frequent import and export of infections between villages, although there was some evidence of geographic structuring at the snail-intermediate host level.
These results suggest very high levels of transmission across host species, and indicate that the role of dogs should be considered when planning control programs. Furthermore, a regional approach to treatment programs is recommended where human migration is extensive.
... By examining schistosomes that infect human hosts living near separate watercourses within a single village, it should be possible to determine the degree to which the parasite population is genetically subdivided. Given that schistosome populations already exhibit polymorphism for traits such as drug resistance and pathology, such subdivision might have significant epidemiologic consequences (Curtis and Minchella 2000). ...
Mitochondrial markers are often hailed as the preferred DNA elements for analyses of population subdivision. To this end we have employed a mitochondrial repeat element to examine the population structure in Schistosoma mansoni (human blood flukes). Schistosome isolates were collected from each of 21 different patients representing seven different areas of a Brazilian village. These parasite isolates demonstrate substantial genetic polymorphism, with an average of 10 genotypes infecting each patient, which is more readily detected because of high levels of heteroplasmy (i.e., 72.5% of the individual worms exhibit multiple versions of this repeat region with different numbers of repeats). Due to the high number of common haplotypes in the population, this repeat element from S. mansoni has a large proportion (47%) of its genetic variation described by differences among mitochondrial genomes within individual worms. However, when only rare haplotypes are considered, population structure can be detected. It seems that heteroplasmy in the schistosome population of Melquiades is both the source of plentiful genetic variation and a confounding factor in the analysis of that variation. Thus the schistosome population in Melquiades may actually be more strongly subdivided than we are able to detect using this mitochondrial marker.
... (1) Microsatellite polymorphisms provide essential markers for genome sequencing, positional cloning, physical mapping and population analysis 6,7 . RepeatMasker Web servers (Box 1) allow large numbers of sequences to be scanned rapidly for microsatellite-like simple repeats. ...
Although schistosome genome analysis is far from complete, and almost all aspects of functional genomics still remain unexplored, mining of the accumulated data already enables workers to undertake important and exciting research into basic schistosome biology.
... Additionally, for helminths such as trematodes, whose life-cycle includes an asexual multiplication phase within a snail intermediate host, leading to the production of genetically identical infective cercariae, patterns of micro-geographical substructure are likely to be more pronounced (see Mulvey et al., 1991 for Fascioloides magna with aggregated infec-tive metacercariae encysted on vegetation). However, this remains to be investigated in more detail for S. mansoni(with direct transmission by free living and swimming infective cercariae), by examination with more polymorphic markers, of the distribution of individual multilocus genotypes within each individual host (Curtis and Minchella, 2000). On the other hand, large effective infrapopulation sizes of S. mansoni within rats (150 worms per host on average and up to 1,000 worms in one host), may contribute to reduce genetic differentiation between infrapopulations. ...
The distribution of genetic diversity in a local population of the trematode Schistosoma mansoni was determined within and between individual wild rats at a microspatial geographic scale of a standing water transmission site. Using RAPD markers, molecular variance and canonical correspondence analysis were performed to test the significance of genetic differentiation between infrapopulations. Of total gene diversity, 8 and 11% was partitioned between hosts trapped at few metres distance from each other. Significant temporal differentiation (2%) was also detected among schistosomes sampled at 6 month intervals with more infrapopulation pairs differentiated during the dry season of parasite transmission than during the rainy season (45 and 12%, respectively). A combination of factors such as restricted displacement of rats, patchy spatial aggregation of infected snails and limited cercarial dispersion in standing water are likely to promote the genetic differentiation observed between infrapopulations at this microgeographic scale.
... E-mail: oliveira@cpqrr.fiocruz.br ++ Fellowship from Fiocruz/Fundep Received 18 June 2002 Accepted 15 August 2002 et al. 1997), and the study of population genetics of several species (Santos et al. 1993, Solano et al. 1997, Lougheed et al. 1999) including parasites ( Durand et al. 2000, Curtis & Minchella 2000, Caccio et al. 2001, Rodrigues et al. 2002). Microsatellite identification results mainly from library screening, sequence database searches, and microsatellite enriched library (Ashley & Dow 1994). ...
In the last decade microsatellites have become one of the most useful genetic markers used in a large number of organisms due to their abundance and high level of polymorphism. Microsatellites have been used for individual identification, paternity tests, forensic studies and population genetics. Data on microsatellite abundance comes preferentially from microsatellite enriched libraries and DNA sequence databases. We have conducted a search in GenBank of more than 16,000 Schistosoma mansoni ESTs and 42,000 BAC sequences. In addition, we obtained 300 sequences from CA and AT microsatellite enriched genomic libraries. The sequences were searched for simple repeats using the RepeatMasker software. Of 16,022 ESTs, we detected 481 (3%) sequences that contained 622 microsatellites (434 perfect, 164 imperfect and 24 compounds). Of the 481 ESTs, 194 were grouped in 63 clusters containing 2 to 15 ESTs per cluster. Polymorphisms were observed in 16 clusters. The 287 remaining ESTs were orphan sequences. Of the 42,017 BAC end sequences, 1,598 (3.8%) contained microsatellites (2,335 perfect, 287 imperfect and 79 compounds). The 1,598 BAC end sequences 80 were grouped into 17 clusters containing 3 to 17 BAC end sequences per cluster. Microsatellites were present in 67 out of 300 sequences from microsatellite enriched libraries (55 perfect, 38 imperfect and 15 compounds). From all of the observed loci 55 were selected for having the longest perfect repeats and flanking regions that allowed the design of primers for PCR amplification. Additionally we describe two new polymorphic microsatellite loci.
... Patterns of fine-scale genetic population structure of parasites within their hosts may provide useful information on transmission and recruitment processes in the field (Anderson, Romero-Abal & Jaenike, 1995 ;Nadler, 1995 ;Anderson, Blouin & Beech, 1998 ;Curtis & Minchella, 2000). As an example, Paterson, Fisher & Viney (2000) were able to conclude from genetic diversity analyses and distribution of the nematode Strongyloides ratti in rats that the source of new infections was mostly by immigration rather than by self-reinfection. ...
We investigated the genotypic composition of the digenetic parasite Schistosoma mansoni for its adult stages within the definitive host (the wild rat, Rattus rattus) and for the larval stages within the intermediate host (the snail, Biomphalaria glabrata) both collected at the same transmission site. Our analyses are based upon the recognition and distribution of 200 different multilocus genotypes generated by RAPD markers. While intramolluscan larval infrapopulations are characterized by a low infection rate (0.6 % on average) and low intra-host genetic diversity (1.1 genotype on average per infected snail), adult infrapopulations within rats showed a high infection rate (94%) and a substantial intra-host genetic diversity (34 genotypes on average) linked to high intensities (160 worms per host on average). A single definitive host bearing 105 different genotypes harboured 52 % of the total genetic diversity detected within the whole parasite population. Analysis of the genetic data allowed the identification of various ecological, behavioural and immunological factors which are likely to enhance transmission of multiple parasite genotypes towards the vertebrate hosts. From the distribution of repeated identical multilocus genotypes within the parasite population and among the hosts, we have inferred different parameters of the cercarial transmission efficiency as well as patterns and processes by which vertebrate hosts acquire infection in the field.
Genetic variation was assessed in Biomphalaria glabrata snails using variations in microsatellite loci and by RAPD-PCR analysis. Populations of snails examined were field-collected isolates from a small pond in a schistosomiasis endemic region in Brazil, after standard conditions were developed for analyzing snails from two laboratory-maintained stocks. The analyses were performed using a total of 60 microsatellite primer sets and, for RAPD-PCR, a total of 19 random primers. We show that genetic diversity can readily be detected by both molecular methods among the field-collected snails from this small site. In addition, RAPD-PCR bands that were found in another study to segregate with parasite resistance were not detected in any of the field-collected snails analyzed.
Although schistosomiasis remains a serious health problem worldwide, significant achievements in schistosomiasis control has been made in the People's Republic of China. The disease has been eliminated in five out of 12 endemic provinces, and the prevalence in remaining endemic areas is very low and is heading toward elimination. A rapid and sensitive method for monitoring the distribution of infected Oncomelania hupensis is urgently required. We applied a loop-mediated isothermal amplification (LAMP) assay targeting 285 rDNA for the rapid and effective detection of Schistosoma japonicum DNA in infected and prepatent infected O. hupensis snails. The detection limit of the LAMP method was 100 fg of S. japonicum genomic DNA. To promote the application of the approach in the field, the LAMP assay was used to detect infection in pooled samples of field-collected snails. In the pooled sample detection, snails were collected from 28 endemic areas, and 50 snails from each area were pooled based on the maximum pool size estimation, crushed together and DNA was extracted from each pooled sample as template for the LAMP assay. Based on the formula for detection from pooled samples, the proportion of positive pooled samples and the positive proportion of O. hupensis detected by LAMP of Xima village reached 66.67% and 1.33%, while those of Heini, Hongjia, Yangjiang and Huangshan villages were 33.33% and 0.67%, and those of Tuanzhou and Suliao villages were 16.67% and 0.33%, respectively. The remaining 21 monitoring field sites gave negative results. A risk map for the transmission of schistosomiasis was constructed using ArcMap, based on the positive proportion of O. hupensis infected with S. japonicum, as detected by the LAMP assay, which will form a guide for surveillance and response strategies in high risk areas.
The ability of microsatellite loci to reveal genetic diversity within the trematode Schistosoma japonicum is demonstrated. Eleven novel microsatellite markers were isolated, assessing variability within 80 S. japonicum individuals from three Chinese and one Philippine population. Novel primers were also tested upon S. mansoni and S. haematobium. Eight primers showed polymorphic amplification from all S. japonicum groups, three amplified S. mansoni DNA, but none amplified S. haematobium DNA. Only six of 27 previously isolated S. mansoni-specific primers amplified S. japonicum DNA. Allelic diversity and observed heterozygosity ranged from 4 to 21 and 0.05 to 0.82, respectively, suggesting high variability.
Schistosomiasis is a major infectious disease and a public health concern in many areas in China and other countries. Sensitive method for detection of the parasite is critical for early diagnosis and for monitoring of effective treatment of the disease. In this study, we developed a highly sensitive TaqMan real-time PCR assay for the detection of Schistosoma japonicum DNA in mouse feces and serum samples. This assay was based on the DNA sequence of the S. japonicum 18S rRNA gene and was able to detect 10 fg of S. japonicum genomic DNA, which is 100 times more sensitive than conventional PCR. We were able to detect the S. japonicum DNA one week post-infection in mouse sera and 4 weeks post-infection in feces, which was one week earlier than egg detection by microscopy in feces. This assay was also highly specific for Asian Schistosomes which are causative species of human Schistosomiasis. In single sex male cercariae infected mice, parasite DNA was only detected in the first 4 weeks post-infection, suggesting that the DNA was derived from decaying worms' corpse in the first 4 weeks whereas the DNA was mainly from decaying parasite eggs afterwards. Therefore we conclude that the established TaqMan real-time PCR assay is a sensitive, specific and convenient method that could be used for the early diagnostic evaluation of S. japonicum infection in humans and for monitoring outbreaks in endemic areas with low prevalence.
Schistosoma mansoni (S. mansoni) eggs trapped in the host liver elicit a chain of oxidative processes that may be, at least in part, responsible for the pathology and progression of fibrosis associated with schistosomal hepatitis. This study was designed to assess the protective effect of the antioxidant coenzyme-Q10 (Co-Q10) against experimental S. mansoni-induced oxidative stress in the liver, and its potential role as an adjuvant to praziquantel (PZQ) therapy. The oxidative stress and overall liver function were improved under Co-Q10 therapy as evidenced by significant reduction in oxidative stress markers and preservation of antioxidant factors. Liver fibrosis was also reduced with a positive impact on liver function. Moreover, addition of Co-Q10 to PZQ therapy caused: significant reduction of liver egg load, significant improvement of the redox status, and lastly decreased liver fibrosis.
Co-infection of host organisms by multiple parasite species has evolutionary consequences for all participants in the symbiosis. In this study, we co-exposed aquatic-snails (Biomphalaria glabrata) to two of their trematode parasites, Schistosoma mansoni and Echinostoma caproni. In co-exposed snails, E. caproni prevalence was 63% compared to only 23% for S. mansoni. Co-exposed E. caproni-infected snails exhibited reduced fecundity, higher mortality, and higher parasite reproduction (higher virulence) compared to hosts exposed to echinostomes alone. Conversely, co-exposed S. mansoni-infected snails released fewer parasites and produced greater numbers of eggs compared to hosts exposed to S. mansoni alone. These results suggest that co-exposure not only influences the establishment (presence or absence) of particular parasite species, but also impacts host life history, parasite reproduction, and the virulence of the interaction.
Opisthorchosis is a helminthiasis affecting mainly the hepatobiliary system and pancreas; its most dramatic complication is malignization of the organs infected by the parasites. The causative agents of opisthorchosis are two species of liver flukes, the trematodes belonging to the family Opisthorchiidae--Opisthorchis felineus and O. viverrini. The Chinese liver fluke, Clonorchis sinensis, also member of the family Opisthorchiidae, causes clonorchosis, a disease very close in symptomatology. According to different estimations, up to 40 million people are currently infected with these liver flukes and up to 600-750 million people in Eurasian countries constitute the risk group. These parasites colonize ever-increasing new areas in Eurasia where this disease has never been previously reported. Opisthorchiases are gradually transforming from a local problem of individual geographic regions to a widespead problem; in particular, O. viverrini is now referred to as "an underestimated parasite." As we see it, O. felineus has all the reasons to share this status. First and foremost, the observed expansion is likely to be connected with the ever-increasing intensity of traffic flows and migration of the infection carriers between cities, regions, and countries. This review briefs the characteristics of O. felineus and the other liver flukes persisting in various countries of Eurasia, clinical manifestations of opisthorchosis, the drugs for chemotherapy of trematodiasis, and the strategy for discovery of new antihelminthic drugs.
In this study, a loop-mediated isothermal amplification (LAMP) assay was established to detect Schistosoma japonicum DNA in faecal and serum samples of rabbits, and serum samples of humans infected with S. japonicum. This LAMP assay was based on the sequence of highly repetitive retrotransposon SjR2, and was able to detect 0.08 fg S. japonicum DNA, which is 10(4) times more sensitive than conventional PCR. The LAMP assay was also highly specific for S. japonicum and able to detect S. japonicum DNA in rabbit sera at 1 week p.i. Following administration of praziquantel, detection of S. japonicum DNA in rabbit sera became negative at 12 weeks post-treatment. These results demonstrated that LAMP was effective for early diagnosis of, and evaluation of therapy effectiveness for, S. japonicum infection. Both PCR and LAMP assays were then used to detect S. japonicum DNA in 30 serum samples from S. japonicum-infected patients and 20 serum samples from healthy persons. The percentage sensitivity of LAMP was 96.7%, whereas that of PCR was only 60%, indicating that LAMP was more sensitive than conventional PCR for clinical diagnosis of schistosomiasis cases in endemic areas. The established LAMP assay should provide a useful and practical tool for the routine diagnosis and therapeutic evaluation of human schistosomiasis.
The parasite Schistosoma is known to exhibit variations among species, strains and genera, such as, the levels of infectivity, pathogenicity and immunogenicity. These factors may differ among parasite populations according to the local epidemiological conditions. Diversity observed in Schistosoma mansoni from different geographical regions or within individuals of the same region can be determined by differences in the genotype of each parasite strain. However, until recently, finding adequate genetic markers to investigate infectivity or other epidemiological characteristics of a transmission area proved difficult. Several studies have been conducted to evaluate the genetic variability of S. mansoni, using different techniques. Intraspecific variability was observed in morphological characters, isoenzyme studies, mtDNA, ribosomal gene probes, random amplification of polymorphic DNA (RAPD) and microsatellites. The sequencing of the S. mansoni genome was the most important achievement concerning genetic approaches to the study of this parasite and may improve the development of drugs, vaccines and diagnostics of schistosomiasis. The knowledge of the genetic structure of schistosome populations in relation to epidemiological data and host variability is essential for the understanding of the epidemiology of the disease and the design of control strategies.
The freshwater gastropod Biomphalaria glabrata is one of the most important invertebrate hosts of the helminth parasite Schistosoma mansoni. Investigators are using different strategies to determine the molecular basis of this snail-parasite relationship. Of particular interest are the identification of parasite resistance genes in the snail, and the application of molecular probes to better understand the epidemiology of schistosomiasis. This review will focus on recent advances that have been made on genome analysis of B. glabrata. Much of this work has centred on the use of random amplification of polymorphic DNA-PCR-based technology, with restriction fragment length polymorphism analysis and the generation of expressed sequence tags from the snail. A brief discussion of how parasite products may complicate this analysis is also given, along with an indication of the scope of the problems that lie ahead.
In response to B. Gryseels article on schistosomiasis vaccines1xSchistosomiasis vaccines: a devils’ advocate view. Gryseels, B. Parasitol. Today. 2000; 16: 46–48Abstract | Full Text | Full Text PDF | PubMed | Scopus (27)See all References1, I suggest that it is ‘not what immunology can do for parasitology but what parasitology can do for immunology’ (Roland Terry, pers. commun.). This is especially applicable to the development of a schistosomiasis vaccine, which has done much to support the field of immunology, but which, in reality, has done little to alleviate the disease.The 1974 Edna McConnell Clark Foundation meeting was followed by a strategic plan to have a vaccine ready by 1984. Between the Clark Foundation, the National Institute of Health (NIH) and the Tropical Disease Research initiative of WHO, a conservative estimate of US$100 million must have been spent on schistosomiasis vaccine research. One could argue that this amount is far from adequate as vaccine development is an expensive area of research and companies often spend far in excess of this before a single product can be marketed.There is no doubt that new concepts and ideas in vaccine development and basic immunology will allow for a major breakthrough in schistosomiasis vaccine development. However, the crucial question is do we really need a vaccine?I consider schistosome population genetic structure (see Curtis and Minchella2xSchistosome population genetic structure: when clumping worms is not just splitting hairs. Curtis, J. and Minchella, D.J. Parasitol. Today. 2000; 16: 68–71Abstract | Full Text | Full Text PDF | PubMed | Scopus (47)See all References2) to be a more important area of schistosome research than is vaccine development. Understanding the basic genetic structure of schistosome populations will allow a much more rational approach towards developing more effective strategies for drug usage and the design and development of a vaccine than would direct vaccine research. Knowing the genetic structure and the genetic/biochemical mechanisms of the parasite that allow it to evade the immune system of the host, will be much more productive than taking the ‘black box’ approach of developing drugs and vaccines without this information.
Shiff, C., Brouwer, K. C., and Clow, L. 2000. Schistosoma haematobium: Population genetics of S. haematobium by direct measurement of parasite diversity using RAPD–PCR. Experimental Parasitology96, 47–51.
Random amplified polymorphic DNA (RAPD) markers were used to quantify genetic diversity within and between 5 populations of Schistosoma mansoni within its definitive host (Rattus rattus) and the 5 corresponding populations of the snail intermediate host (Biomphalaria glabrata) from a limited endemic area of murine schistosomiasis on the island of Guadeloupe. Analysis of molecular variance (AMOVA) and canonical correspondence analysis (CCA) were used to test the significance of genetic differentiation between populations. Both methods gave similar results. Of total gene diversity, 15.1% (AMOVA) and 18.8% (CCA) was partitioned between localities for S. mansoni with an absence of association between genetic and geographical distances. Geographical localities accounted for 20.5% (CCA) of the total diversity for B. glabrata populations. The genetic distances between pairs of parasite populations were not correlated with the genetic distances between the corresponding pairs of snail host populations. Such strong patterns of local differentiation of both parasite and snail populations are consistent with predictions based on metapopulation dynamics and may have implications on host-parasite susceptibility relationship through local adaptation processes.
To characterize the extent of genetic diversity of Schistosoma haematobium within and among its definitive host (intra- and interhost parasite diversity), 133 individual isolates from 25 infected schoolchildren were compared using randomly amplified polymorphic DNA markers. With 4 primers, 53 unambiguous loci were identified, and of these, 22 were polymorphic. Mean heterozygosity in the population was 0.116 +/- 0.043. Analysis of molecular variance showed the majority of variance occurred within, rather than between, hosts. Frequencies of certain alleles segregated the parasite population into 13 distinct clusters of associated genotypes, with 4 of these first appearing 10 mo after the initial survey. Considering the level of diversity within this limited geographical area and the possibility of rapid turnover of genotypes, parasite variance may impact acquired immunity and clinical outcome of the infection.
Several aspects of the coevolutionary dynamics in host-parasite systems may be better quantified based on analyses of population structure using neutral genetic markers. This includes, for example, the migration rates of hosts and parasites. In this respect, the current situation, especially in fluke-snail systems is unsatisfactory, since basic population genetics data are lacking and the appropriate methodology has rarely been used. After reviewing the forces acting on population structure (e.g. genetic drift or the mating system) and how they can be analysed in models of structured populations, we propose a simplified, indicative framework for conducting analyses of population structure in hosts and parasites. This includes consideration of markers, sampling, data analysis, comparison of structure in hosts and parasites and use of external data (e.g. from population dynamics). We then focus on flukes and snails, highlighting important biological traits with regard to population structure. The few available studies indicate that asexual amplification of flukes within snails strongly influences adult flukes populations. They also show that the genetic structure among populations in strongly affected by traits in other than snails (e.g. definitive host dispersal behaviour), as snails populations have limited migration. Finally more studies would allow us to deepen our current understanding of selective interference between flukes and snails (e.g. manipulation of host mating system by parasites), and evaluate how this affect population structure at neutral markers.
Schistosomiasis is a worldwide problem because it is so widely distributed, and few places on earth are now too remote. Some of the most important new research examines how parasite and host biology are integrated, how any level of infection contributes to the overall burden of disease, and the long-term and short-term outcomes of control programs.
Biomphalaria glabrata snails infected with Schistosoma mansoni were collected during consecutive seasons from a site in Brazil known to have a very high percentage of infected snails. Schistosoma mansoni cercariae from single snails were used to infect individual mice, and the recovered adult worms were genetically assessed using a mtVNTR marker. The number of unique parasite genotypes found per snail was compared to expected abundance values, based on the infection prevalence at the site, to determine the distribution of S. mansoni infections within the snail population. The observed distributions and those from previous studies were used to examine the relationship between schistosome prevalence and aggregation across a wide range of prevalence values. Our analysis showed that prevalence was inversely related to the degree of parasite overdispersion, and at high prevalence, S. mansoni infections were randomly distributed among snails.
Blood flukes in the genus Schistosoma are important human parasites in tropical regions. A substantial amount of genetic diversity has been described in populations of these parasites using molecular markers. We first consider the extent of genetic variation found in Schistosoma mansoni and some factors that may be contributing to this variation. Recently, though, attempts have been made to analyze not only the genetic diversity but how that diversity is partitioned within natural populations of schistosomes. Studies with non-allelic molecular markers (e.g. RAPDs and mtVNTRs) have indicated that schistosome populations exhibit varying levels of gene flow among component subpopulations. The recent characterization of microsatellite markers for S. mansoni provided an opportunity to study schistosome population structure within a population of schistosomes from a single Brazilian village using allelic markers. Whereas the detection of population structure depends strongly on the type of analysis with a mitochondrial marker, analyses with a set of seven microsatellite loci consistently revealed moderate genetic differentiation when village boroughs were used to define parasite subpopulations and greater subdivision when human hosts defined subpopulations. Finally, we discuss the implications that such strong population structure might have on schistosome epidemiology.
There are strong biological, evolutionary and immunological arguments for predicting extensive polymorphism among helminth parasites, but relatively little data and few instances from which the selective forces acting on parasite diversity can be discerned. The paucity of information on intraspecific variation stands in contrast to the fine detail with which helminth species have been delineated by morphological techniques, accentuating a trend towards considering laboratory strains as representative of a relatively invariant organism. However, in the fast-moving evolutionary race between host and parasite one would predict a monomorphic species would be driven to extinction. We review the arena of intraspecific variation for the major helminth parasites, ranging from biological properties such as host or vector preference, to biochemical and immunological characteristics, as well as molecular markers such as DNA sequence variants. These data are summarized, before focusing in more detail on polymorphisms within protein-coding genes of potential relevance to the host-parasite relationship, such as vaccine candidates. In particular, we discuss the available data on a number of major antigens from the filarial nematode Brugia malayi. Information is currently too sparse to answer the question of whether there is antigenic variation in filariasis, but the indications are that proteins from the blood-borne microfilarial stage show significant intraspecific variability. Future work will define whether polymorphisms in these antigens may be driven by exposure to the host immune response or reflect some other facet of parasite biology.
Echinococcus granulosus exhibits substantial genetic diversity that has important implications for the design and development of vaccines, diagnostic reagents and drugs effective against this parasite. DNA approaches that have been used for accurate identification of these genetic variants are presented here as is a description of their application in molecular epidemiological surveys of cystic echinococcosis in different geographical settings and host assemblages. The recent publication of the complete sequences of the mitochondrial (mt) genomes of the horse and sheep strains of E. granulosus and of E. multilocularis, and the availability of mt DNA sequences for a number of other E. granulosus genotypes, has provided additional genetic information that can be used for more in depth strain characterization and taxonomic studies of these parasites. This very rich sequence information has provided a solid molecular basis, along with a range of different biological, epidemiological, biochemical and other molecular-genetic criteria, for revising the taxonomy of the genus Echinococcus. This has been a controversial issue for some time. Furthermore, the accumulating genetic data may allow insight to several other unresolved questions such as confirming the occurrence and precise nature of the E. granulosus G9 genotype and its reservoir in Poland, whether it is present elsewhere, why the camel strain (G6 genotype) appears to affect humans in certain geographical areas but not others, more precise delineation of the host and geographic ranges of the genotypes characterised to date, and whether additional genotypes of E. granulosus remain to be identified.
To investigate the difficulties doctors face in discussing treatment options with patients with acute, life threatening illness and major comorbidities.
Observational study of doctor-patient interviews based on a standardised clinical scenario involving high risk surgery in a hypothetical patient (played by an actor) with serious comorbidities.
30 trainee doctors 3-5 years after graduation.
Adequacy of coverage of various aspects was scored from 3 (good) to 0 (not discussed).
The medical situation was considered to be well described (median score 2.7 (interquartile range 2.1-3.0)), whereas the patient's functional status, values, and fears were poorly or minimally addressed (scores 0.5 (0.0-1.0), 0.5 (0.0-1.0), and 0.0 (0.0-1.5), respectively; all P < 0.001 v score for describing the medical situation). Twenty nine of the doctors indicated that they wished to include the patient's family in the discussion, but none identified a preferred surrogate decision maker. Six doctors suggested that the patient alone should speak with his family to reach a decision without the doctor being present. The doctors were reluctant to give advice, despite it being directly requested: two doctors stated that a doctor could not give advice, while 17 simply restated the medical risks, without advocating any particular course. Of the 11 who did offer advice, eight advocated intervention.
Doctors focused on technical medical issues and placed much less emphasis on patient issues such as functional status, values, wishes, and fears. This limits doctors' ability to offer suitable advice about treatment options. Doctors need to improve their communication skills in this difficult but common clinical situation.
The rapid development of molecular techniques offers a palette of technical approaches for population biologists interested in a wide range of questions. For example, these tools can be used to determine individual reproductive success or to measure rates of genetic divergence among populations. Which technique is most appropriate for a par- ticular question depends upon (1) the extent of genetic polymorphism required to best answer the question, (2) the analytical or statistical approaches available for the technique's application, and (3) the pragmatics of time and costs of materials. Here we evaluate the application of several major techniques (protein electrophoresis, nuclear and mitochondrial RFLPs (restriction fragment length polymorphisms), minisatellite and microsatellite VNTRs (variable number tandem repeats), RAPDs (random amplified polymorphic DNA), and DNA sequencing) to an array of questions regarding individual identification, exclusion and assignment of parentage, and various levels of population structure. In our evaluation, we briefly explain the technical components of each molecular approach and assess whether the typical outcomes expected from each approach will provide useful information as applied to each level of inquiry. For studies of population genetic structure, protein electrophoresis remains a powerful tool for most taxa, although techniques based on nucleic acids (par- ticularly DNA sequencing and mitochondrial DNA RFLPs) are useful here as well. Recently developed nucleic acid techniques (e.g., VNTRs) can often identify enough genetic vari- ability to address questions of self-identification or parentage. Some of the newest tech- niques (RAPDs and microsatellites) are potentially useful across a number of levels of inquiry, although procedures for adopting them are still developing.
We have detected species, strain, and sex-specific genetic markers for the genus Schistosoma by Southern blot analysis of its DNA using cloned DNA segments of the Schistosoma mansoni ribosomal gene as probes. Restriction analysis of DNA from eight different strains of S. mansoni, from Africa and the Caribbean, revealed that the predominant or major DNA fragment containing the ribosomal gene unit was the same in each but that low copy number or minor fragments containing the gene varied. It was shown that the detection of these minor fragments could serve as the basis for both strain differentiation and the analysis of individual differences within a strain. Analysis of the parents and progeny of a genetic cross revealed sex-linked markers and suggested that these markers are inherited in a Mendelian fashion. DNAs from the species Schistosoma haematobium and Schistosoma japonicum were also analyzed. Differences in the length of the major repeating unit of the ribosomal gene served to distinguish each species. Furthermore, an array of minor bands was detected in each species, suggesting that strains of S. haematobium and S. japonicum could be differentiated in the same manner as S. mansoni strains.
To assess the genetic differences underlying geographic variations in Schistosoma mansoni, individual adult worms from 22 populations, from East and South Africa, Southwest Asia, South America, and the West Indies, were processed for enzyme electrophoresis on starch gels. Fourteen enzyme systems were analyzed. An estimated 7 of 18 loci were polymorphic, and the most variable population was polymorphic at 6 of the loci (P = 0.33), with a heterozygosity H of 0.07. These results suggest that S. mansoni is as variable genetically as most other organisms. Most S. mansoni strains showed relatively low variability, however (P = 0.13 ± 0.02, H = 0.04 ± 0.005). This may be attributed to small founding populations and passage in the laboratory through low numbers of infected snails and through abnormal laboratory hosts, resulting in random fixation of alleles by the action of genetic drift and possibly in selection against particular alleles. This finding implies that geographic comparisons of any traits should be based on several isolates from each region compared, so as to adequately sample the total variation occurring within each region. Genetic distances between all strains were low (mean 0.052, range 0-0.275), suggesting that little intraspecific differentiation has occurred in S. mansoni, even between Old and New World populations. These results contrast with published electrophoretic evidence of significant divergence between geographic strains of S. japonicum. Most polymorphisms were consistent with a simple Mendelian interpretation, although formal genetic crosses were not performed. For those enzymes, the banding patterns of heterozygotes indicated that subunit structure is the same in S. mansoni as in many other organisms. Sexual differences in mobility and in number of bands were found in a few enzymes. The polymorphisms uncovered can eventually be used as genetic markers to map chromosomes and to study various traits, such as infectivity to snails and drug resistance.
Genetic variation of Anopheles gambiae was analysed to assess interpopulation divergence over a 6000 km distance using short tandem repeat (microsatellite) loci and allozyme loci. Differentiation of populations from Kenya and Senegal measured by allele length variation at five microsatellite loci was compared with estimates calculated from published data on six allozyme loci (Miles, 1978). The average Wright's FST of microsatellite loci (0.016) was lower than that of allozymes (0.036). Slatkin's RST values for microsatellite loci were generally higher than their FST values, but the average RST value was virtually identical (0.036) to the average allozyme FST. These low estimates of differentiation correspond to an effective migration index (Nm) larger than 3, suggesting that gene flow across the continent is only weakly restricted. Polymorphism of microsatellite loci was significantly higher than that of allozymes, probably because the former experience considerably higher mutation rates. That microsatellite loci did not measure greater interpopulation divergence than allozyme loci suggested constraints on microsatellite evolution. Alternatively, extensive mosquito dispersal, aided by human transportation during the last century, better explains the low differentiation and the similarity of estimates derived from both types of genetic markers.
Studies on human and murine isolates of Schistosoma mansoni, from Richard-Toll, Senegal, were carried out by isoelectric focusing in polyacrylamide gels. Seven enzyme systems; lactate dehydrogenase (LDH), malate dehydrogenase (MDH), glucose-6-phosphate dehydrogenase (G6PD), acid phosphatase (AcP), hexokinase (HK), glucose phosphate isomerase (GPI), and phosphoglucomutase (PGM), were used to compare the two isolates. All systems tested, apart from LDH, were found to be polymorphic for both isolates. Interestingly, one phenotype is more frequent than the remainder. The results show that there is no significant genetic variation between the S. mansoni isolates from man and the rodents, Arvicanthis niloticus and Mastomys huberti.
The extent of genomic variability of clones of Schistosoma mansoni obtained from field isolates was compared with that of strains that have been laboratory maintained. Analysis was undertaken using randomly amplified polymorphic DNAs (RAPDs) generated with three primers. Phenograms showing the similarity among the clones were constructed. The data showed that while the laboratory strain is highly homogeneous the clones derived from the field populations were highly variable with 43% of RAPDs exhibiting polymorphisms among 23 clones. Clones isolated from the same infected individual were always more closely grouped than clones from different individuals. The data clearly demonstrated that earlier analyses of the genomic variability in S. mansoni have underestimated this phenomenon due to the failure to examine field isolates.
A new measure of the extent of population subdivision as inferred from allele frequencies at microsatellite loci is proposed and tested with computer simulations. This measure, called R(ST), is analogous to Wright's F(ST) in representing the proportion of variation between populations. It differs in taking explicit account of the mutation process at microsatellite loci, for which a generalized stepwise mutation model appears appropriate. Simulations of subdivided populations were carried out to test the performance of R(ST) and F(ST). It was found that, under the generalized stepwise mutation model, R(ST) provides relatively unbiased estimates of migration rates and times of population divergence while F(ST) tends to show too much population similarity, particularly when migration rates are low or divergence times are long [corrected].
A cloned fragment of the ribosomal gene of Schistosoma mansoni, pSM 389, which contains part of the small rRNA gene plus a portion of the nontranscribed intergenic spacer, was used in Southern hybridization analyses to investigate genomic variation in natural populations of S. mansoni in Brazil. Genomic DNAs were isolated from schistosomes from infected patients (some of whom did not respond to antischistosomal chemotherapy), and from snails from disparate geographic locations in Brazil. Restriction fragment length polymorphisms (RFLPs) were evident in Southern blot hybridizations of these schistosome DNAs, and the RFLPs indicated that the genomic profiles of a number of Brazilian strains were more similar to each other than they were to parasites from two laboratory reference strains of Puerto Rican origin. In addition, the Brazilian isolates could generally be separated from each other based on these RFLPs. Isolates from the southeastern state of Minas Gerais were more similar to each other than they were to parasites isolated in the northeastern states of Alagoas and Pernambuco. Variation was evident among individual worms from some of the isolates, and these individual variations contributed to the complex RFLP patterns that were characteristic for particular isolates. The variation within a natural population isolated directly from snails at Ressaca, Belo Horizonte, may be more marked than that exhibited by more established strains maintained in the laboratory for numerous generations.
1. An isoenzymatic study on Schistosoma mansoni from Guadeloupe has been carried out using isoelectric focusing in polyacrylamide gels. 2. Among the seven systems examined (LDH, MDH, G6PD, PGI, PGM, AcP, HK), only MDH showed variation at the MDH-1 locus and mdh-1a allele frequencies were used to characterise eight strains derived either from human or murine hosts, and representative of different transmission sites. 3. In comparison with criteria previously employed to type these strains (ecological context of their transmission sites, cercarial emergence patterns), mdh-1a frequency variations have been correlated to the degree of participation of the murine host reservoir in the parasite transmission dynamics within the different foci of Guadeloupe.
An enzymatic comparison has been made between isolates of Schistosoma mansoni from rats and humans in Guadeloupe and a Burundi isolate of S. rodhaini. Analyses of LDH, MDH, AcP, PGM, GPI, G6PDH and HK by isoelectric focusing provided no evidence for the involvement of S. rodhaini in the recent evolution of the schistosomes currently endemic in Guadeloupe. No distinction could be made between murine and human isolates of S. mansoni and it is suggested that murine schistosomiasis should not therefore be ignored in control programmes. Rattus rattus were captured at seven sites around the island; of 142 examined, 48 were positive for schistosomes. Differences in prevalence between habitats were marked and only small changes in prevalence were observed in localities sampled in 1982 and 1983. Animals with the greatest worm burdens were associated with areas of high prevalence, and age-related changes in worm burden were observed. Two alleles, a and b, at the MDH-1 locus of S. mansoni from rats were identified. Differences in the overall frequencies of these alleles were observed for schistosomes from different localities. Allelic frequencies representative of schistosomes from rats at four localities were stable from 1982 to 1983. The majority of positive animals, even those with light worm burdens, were found to be infected with a number of different schistosome genotypes.
There is abundant geographic variation in both morphology and gene frequency in most species. The extent of geographic variation
results from a balance of forces tending to produce local genetic differentiation and forces tending to produce genetic homogeneity.
Mutation, genetic drift due to finite population size, and natural selection favoring adaptations to local environmental conditions
will all lead to the genetic differentiation of local populations, and the movement of gametes, individuals, and even entire
populations--collectively called gene flow--will oppose that differentiation. Gene flow may either constrain evolution by
preventing adaptation to local conditions or promote evolution by spreading new genes and combinations of genes throughout
a species' range. Several methods are available for estimating the amount of gene flow. Direct methods monitor ongoing gene
flow, and indirect methods use spatial distributions of gene frequencies to infer past gene flow. Applications of these methods
show that species differ widely in the gene flow that they experience. Of particular interest are those species for which
direct methods indicate little current gene flow but indirect methods indicate much higher levels of gene flow in the recent
past. Such species probably have undergone large-scale demographic changes relatively frequently.
A population of Schistosoma mansoni from Kenya was isolated in 1968 and subsequently passaged simultaneously through 2 different vertebrate hosts: baboons and mice. Recent electrophoretic studies demonstrated that genetic differences in the degree of polymorphism and in allele frequencies of polymorphic loci existed between S. mansoni populations from the 2 hosts. The present study was undertaken to assess the importance of vertebrate host-induced selection against particular alleles as mechanism to account for the observed differences. A population of S. mansoni which had originally been passaged through baboons and subsequently passaged through murine hosts for 4 generations was studied. At least 20 infected snails served as the source of parasite for each mouse passage. Allele frequencies of 4 polymorphic loci were assessed for each generation using horizontal starch gel electrophoresis. All 4 polymorphic loci (PGM-2, MDH-2, MDH-1, PGI) showed a selective trend towards allele frequencies identical with that of a strain (from the same isolate) maintained in mice for 12 yr. These data suggest that vertebrate host-induced selection results in a decrease in parasite variability due to loss of alleles as field isolates of S. mansoni are passaged in murine hosts. The use of non-human primate hosts, on the other hand, maintains a higher level of parasite variability.
Naturally infected Biomphalaria glabrata snails were collected at two sites near Belo Horizonte, Brazil, and Schistosoma mansoni cercariae isolated from single snails were used to infect individual mice. Genetic comparison of single worm DNA was accomplished by hybridization of Southern blots to a polymorphic repetitive DNA element. Genetic profiles of parasite individuals revealed a diverse array of parasite genotypes in naturally infected intermediate hosts. The observed distribution of schistosome genotypes among intermediate hosts indicates that over half of the infected snails harbour multiple miracidia. Snails were more likely to carry multiple infections than expected by chance. This degree of overdispersion combined with high levels of genetic variability facilitates multi-genotype transmission and helps maintain parasite genetic diversity.
A new measure of the extent of population subdivision as inferred from allele frequencies at microsatellite loci is proposed and tested with computer simulations. This measure, called R(ST), is analogous to Wright's F(ST) in representing the proportion of variation between populations. It differs in taking explicit account of the mutation process at microsatellite loci, for which a generalized stepwise mutation model appears appropriate. Simulations of subdivided populations were carried out to test the performance of R(ST) and F(ST). It was found that, under the generalized stepwise mutation model, R(ST) provides relatively unbiased estimates of migration rates and times of population divergence while F(ST) tends to show too much population similarity, particularly when migration rates are low or divergence times are long [corrected].
Intraspecific genetic variation among 14 geographic isolates of Schistosoma mansoni was quantified using a molecular marker to examine individual genotypes. Genetic crosses demonstrated maternal inheritance of S. mansoni DNA element pSM750. This element revealed diagnostic banding profiles, which allowed accurate strain identification. Most strains had similarity indices greater than 0.75 indicating that within-strain variation in these laboratory parasite populations was low. However, individual parasites from the NMRI strain were quite diverse (S = 0.40). Genetic heterogeneity among strains was quantified using a phenogram of mean genetic distance. Strain diversity between two geographic regions was quantified using a similarity index and was estimated to be substantial among isolates collected from a single local site.
The development of polymerase chain reaction-based methods for assessing the genotypes of small individual organisms will promote groundbreaking investigations of the genetic architecture of parasite populations. Both quantitative genetic models and general knowledge of parasite natural history are useful for making general predictions about the distribution of genetic variation over geographic space. However, designing experimental studies to assess relationships between specific life history variables and patterns of genetic structure in natural populations will be challenging. Traditional biochemical-genetic methods have already been used to study a limited number of parasite populations, and inferred patterns of genetic structure are distinctly different between certain species. Some of these differences in genetic architecture may be explained by parasite or host factors that either promote or retard the dissemination of life cycle stages over geographic space. Many additional empirical studies are needed to characterize basic features of parasite populations, including the spatial distribution and group size of random mating populations and levels of gene flow among parasite subpopulations.
Keywords:freshwater snails;microsatellites;self-fertilization;schistosome vector;population genetics
The usefulness of random amplified polymorphic DNA markers (RAPD) was assayed in an attempt to discriminate among species, strains and individuals within the genus Schistosoma. Depending on the species, 40-50 arbitrary decamer oligonucleotides were used as primers to amplify total DNA by the polymerase chain reaction (PCR). An important polymorphism was observed among 5 species, allowing a phylogenetic tree to be outlined. These differences can be used for rapid and accurate identification. A limited but easily detectable polymorphism was revealed among 3 strains of a single species (Schistosoma mansoni). Minor differences were observed among individuals of a single strain. A RAPD marker allows sexual discrimination between individuals from the terminal spined-egg species group. Although a limited number of strains have been examined, the results already indicate clearly that RAPD markers constitute a powerful tool for the analysis of genetic variability. This new tool will considerably extend the information available from morphology, isozyme and limited restriction fragment length polymorphism data and opens the way to genetic analysis of these species.
The use of arbitrarily selected primers (10-24 nucleotides) and very low stringency annealing conditions (30 degrees C followed by 40 degrees C) for the polymerase chain reaction amplification of 1.0 ng of schistosome DNA resulted in relatively complex patterns of products. Amongst the primers tested some, for example 5'-TCGTAGCCAA, produced patterns that included bands that were polymorphic between strains of Schistosoma mansoni. Other primers, for example 5'-TCACGATGCA, produced apparently identical products using DNA from 5 S. mansoni strains but highly variable patterns when DNA from different schistosome species was used. The results indicate that the random amplification of polymorphic DNA (RAPD) may be an extremely useful approach to the identification of schistosome strains and species.
Random amplified polymorphic DNA markers (RAPD) were used to visualize the genetic diversity within and between infrapopulations of Schistosoma mansoni recovered from the natural vertebrate host, Rattus rattus, trapped at an insular Guadeloupean focus. Phenotypes were characterized by the sex of the parasites and by 8 polymorphic markers generated by 3 primers. Among the 212 parasite individuals recovered from 10 infected rats, 78 genotypes were characterized. All the hosts naturally infected harboured multiple parasite genotypes with a maximum diversity of 28 genotypes/host. Phenotypic and genotypic diversity calculated by Shannon-Wiener's indices and Lynch and Milligan's estimators respectively is, on average, greater within than between hosts. Considering the very low snail infection rates observed in this focus and the rapid turnover of the vertebrate hosts, our results suggest that the high mobility of the vertebrate host and/or plurimiracidial snail infections could be factors responsible for parasite genetic diversity within hosts.
Robust phylogenies based on molecular data for species within the genus Schistosoma have been generated in recent years. The considerable progress made in understanding the relationships between many of the 19 recognised species of Schistosoma is reviewed with particular attention being given to the detection and analysis of parasite variation as shown by studies on ribosomal RNA genes, mitochondrial DNA and RAPDs. For the most part, molecular phylogenies agree with observations based on morphological or life-history characteristics. It is clear that the parasites of man do not form a monophyletic group and that close relationships exist between parasites within species groups, especially in the S. haematobium group of species. The S. japonicum group appears to be the most divergent of the species groups and yet little DNA sequence variation has been observed between various isolates of S: japonicum. Some of the less studied schistosomes have yet to be examined at the molecular level and may prove to be interesting links between the species groups as has recently been shown with S. hippopotami. The power of molecular approaches for the analysis of schistosomes at the population and individual level is now apparent, especially for S. mansoni. Important questions remain concerning the maintenance of parasite diversity and how schistosomes respond to selection pressures imposed either during natural progression through the life-cycle or through drug treatment or vaccination. Gene discovery and gene mapping projects are leading to a better understanding of the schistosome genome and can be expected to contribute significantly to future comparative evolutionary studies.
Random-amplified polymorphic DNA markers have been used to assess the amount and the distribution of the genetic diversity of Schistosoma mansoni within a natural population of Biomphalaria glabrata at a transmission site of the murine schistosomiasis focus of Guadeloupe. Despite high infection rate and heavy schistosome load within the definitive hosts (Ratus rattus), prevalences within intermediate snails ranged from 0.2 to 4.8%. Whatever the transmission season may be (rainy vs. dry), most of the infected snails were spatially aggregated and 88.4% of them harbored a single parasite genotype indicative of a monomiracidial infection; 4.7% had dual sex infections and a parasite intensity not exceeding 3 miracidia per snail. A substantial resistance level toward the parasite and recruitment regulatory process within snails may explain in part the observed low parasite prevalences and intensities. Considering such a distribution pattern of larval S. mansoni genetic diversity among B. glabrata, mobility of the definitive hosts, or rapid turnover of infected snails, or both, are required to maintain genetic heterogeneity within adult schistosome populations.
No major changes have occurred during the past 20 years regarding the therapeutic tools available to the clinician for the treatment of schistosomiasis. If anything, the two drugs (oxamniquine and metrifonate) that are valuable alternatives to the drug of choice (praziquantel) have become more difficult to procure in some African countries. Here, Donato Cioli summarizes some of the most recent and interesting laboratory studies on potential antischistosomal compounds, and then reviews recent developments related to the mechanism of action of praziquantel and to the possible emergence of praziquantel-resistant schistosomes.
In the past 20 years, genetic and molecular methods for characterizing pathogen strains have taken a major place in modern approaches to epidemiology of parasitic and other infectious diseases. Here, Michel Tibayrenc explains the main concepts used in this field of research, with special emphasis on the approaches developed in his team, and suggests future avenues to explore.
In this article, Robert Bergquist and Dan Colley deal with the consolidated, international efforts to generate a schistosomiasis vaccine; in particular, they summarize the deliberations of a series of meetings, held in Cairo, Egypt, 21-25 May 1997, with the aim of reviewing the current status of affairs in this respect in order to make recommendations for the future course of schistosomiasis vaccine development.
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3