Regulation of Innate Responses during Pre-patent Schistosome Infection Provides an Immune Environment Permissive for Parasite Development

Blood flukes of the genus Schistosoma infect over 200 million people, causing granulomatous pathology with accompanying morbidity and mortality. As a consequence of extensive host-parasite co-evolution, schistosomes exhibit a complex relationship with their hosts, in which immunological factors are intimately linked with parasite development. Schistosomes fail to develop normally in immunodeficient mice, an outcome specifically dependent on the absence of CD4⁺ T cells. The role of CD4⁺ T cells in parasite development is indirect and mediated by interaction with innate cells, as repeated toll-like receptor 4 stimulation is sufficient to restore parasite development in immunodeficient mice in the absence of CD4⁺ T cells. Here we show that repeated stimulation of innate immunity by an endogenous danger signal can also restore parasite development and that both these stimuli, when administered repeatedly, lead to the regulation of innate responses. Supporting a role for regulation of innate responses in parasite development, we show that regulation of inflammation by neutralizing anti-TNF antibodies also restores parasite development in immunodeficient mice. Finally, we show that administration of IL-4 to immunodeficient mice to regulate inflammation by induction of type 2 responses also restores parasite development. These findings suggest that the type 2 response driven by CD4⁺ T cells during pre-patent infection of immunocompetent hosts is exploited by schistosomes to complete their development to reproductively mature adult parasites.     

d-Glucose concentration is the key factor facilitating liver stage maturation of Plasmodium

The course of malaria infection in mammals begins with transmission of Plasmodium sporozoites into the skin by Anopheles mosquitoes, followed by migration of the sporozoites to the liver. As no symptoms present until hepatic merozoites are released and until they infect erythrocytes in the blood vessels, sporozoites and liver-stage (LS) parasites are promising targets for anti-malaria drugs aiming to prevent mosquito-to-mammal transmission. In vitro LS parasite development system is useful in the screening of candidate drugs on LS parasite development and the elucidation of its underlying molecular mechanisms, which remain unclear. Using rodent malaria parasites (Plasmodium berghei) as a model, this study aimed to develop an optimal in vitro LS culture system for the full maturation of the LS parasite into the hepatic merozoite, the next infective stage in parasite development. As the development of this system required measurement of maturation, a novel quantitative index of LS parasite maturation based on the expression pattern of liver-specific protein 2 (LISP2) was first developed. The use of this index for comparing the effect of incubation in different culture media on LS maturation revealed that the d-glucose concentration of the culture medium is the key factor promoting parasite development in hepatocytes and that a d-glucose concentration of 2000 mg/L/day is the threshold concentration at which the maturation of P. berghei into infective hepatic merozoites is achieved. These findings can be utilized to optimize a human malaria LS culture system for drug discovery.     

COMPARATIVE DEVELOPMENT OF THE RED ALGAL PARASITES BOSTRYCHIOCOLAX AUSTRALIS GEN. ET SP. NOV. AND DAWSONIOCOLAX BOSTRYCHIAE (CHOREOCOLACACEAE,RHODOPHYTA)1

The development of two red algal parasites was examined in laboratory culture. The red algal parasite Bostrychiocolax australis gen. et sp. nov., from Australia, originally misidentified as Dawsoniocolax bostrychiae (Joly et Yamaguishi-Tomita) Joly et Yamaguishi-Tomita, completes its life history in 6 weeks on its host Bostrychia radicans (Montagne) Montagne. Initially the spores divide to form a small lenticular cell, and then a germ tube grows from the opposite pole. Upon contact with the host cuticle, the germ tube penetrates the host cell wall. The tip of the germ tube expands, and the spore cytoplasm moves into this expanded tip. The expanded germ tube tip becomes the first endophytic cell from which a parasite cell is cut off that fuses with a host tier cell. The nuclei of this infected host cell enlarge. As parasite development continues, other host-parasite cell fusions are formed, transferring more parasite nuclei into host cells. The erumpent colorless multicellular parasite develops externally on the host, and reproductive structures are visible within 2 weeks. Tetrasporangia are superficial and cruciately or tetra-hedrally divided. Spermatia are formed in clusters. The carpogonial branches are four-celled, and the carpogonium fuses directly with the auxiliary (support) cell. The mature carposporophyte has a large central fusion cell and sympodially branched gonimoblast filaments. Early stages of development differ markedly in Dawsoniocolax bostrychiae from Brazil. Upon contact with the host, the spore undergoes a nearly equal division, and a germ tube elongates from the more basal of the two spore cells, penetrates the host cell wall, and fuses with a host tier cell. Subsequent development involves enlargement of the original spore body and division to form a multicellular cushion, from which descending rhizoidal filaments form that fuse with underlying host cells. This radically different development is in marked contrast to the final reproductive morphology, which is similar to B. australis and has lead to taxonomic confusion between these two entities. The different spore germination patterns and early germ-ling development of B. australis and D. bostrychiae warrant the formation of a new genus for the Australian parasite.     

The immunoepidemiology of nematode parasites of farmed animals: a mathematical approach.

The population dynamics of farmed animals are controlled by humans, and often involve high host densities, which encourage higher parasite burdens than would be usual in wild animals. As a result, the immunity to reinfection acquired by the host is an important determinant of parasite population dynamics. For example, lambs are highly susceptible to gastrointestinal nematodes as they begin to graze, but develop an immunity that accounts for the observed within-year variation in parasite load and pasture contamination. In the longer term, control measures are compromised by the development of parasite strains resistant to chemotherapy, focusing attention on the development of 'natural' measures, including the selection for resistant hosts and the development of antiparasite vaccines. Mick Roberts here considers the immunoepidemiology of parasites of farmed animals on three levels: the interaction between the parasite and the host's immune system determining the individual's level of protection; the development of acquired immunity determining the within-year parasite population dynamics; and the long-term effects of control measures on the between-year parasite population dynamics.     

Intraerythrocytic Development of Plasmodium knowlesi: Structure,Temperature- and Ca2+-Response of the Host and Parasite Membranes1

Thin-sectioning and freeze-etching electron microscopy were applied to explore the structure and the temperature- and Ca²⁺-response of the different host and parasite membranes during intraerythrocytic development of Plasmodium knowlesi in Macacca mulatta. The plasma membrane of uninfected erythrocytes is temperature- and Ca²⁺-responsive: chilling to 4°C and exposure to 5 mM Ca²⁺ induces a slight decrease in IMP-frequency and the emergence of small IMP-devoid patches on P-faces. On parasite infection, the erythrocyte membrane becomes modified as indicated by an enhanced temperature-response and the appearance of caveolae, ca. 70–90 nm in diameter. The frequency of these caveolae is increased in schizont-infected erythrocytes. Moreover, electron dense plaques, ca. 40 nm in width, appear just beneath the erythrocyte membrane in late trophozoites and schizonts, thus indicating a further modification of the host cell membrane during parasite development. The membrane of the parasitophorous vacuole, derived from the host plasma membrane, dramatically reduces the IMP-frequency especially on the P-face upon parasite infection. This leads to an apparent reversal of the IMP-distribution persisting throughout the whole infection cycle. The parasite plasma membrane forms local compaction domains with the nuclear envelope in ca. 30% of the ring-stages and trophozoites, which disappear in late trophozoites and schizonts. Moreover, the IMP-frequency on plasma membrane fracture faces almost doubles during parasite development. Chilling induces a decrease in the IMP-frequency on P-faces of the plasma membrane. Surprisingly, however, the parasite plasma membrane and the vacuolar membrane respond to externally applied Ca²⁺ with almost a doubling of the IMP-frequency. The different parasite endomembranes also undergo characteristic changes during parasite development.     

Is the development of falciparum malaria in the human host limited by the availability of uninfected erythrocytes?

Background  

The development and propagation of malaria parasites in their vertebrate host is a complex process in which various host and parasite factors are involved. Sometimes the evolution of parasitaemia seems to be quelled by parasite load. In order to understand the typical dynamics of evolution of parasitaemia, various mathematical models have been developed. The basic premise ingrained in most models is that the availability of uninfected red blood cells (RBC) in which the parasite develops is a limiting factor in the propagation of the parasite population.     

Acetylcholinesterase levels in Angiostrongylus cantonensis in relation to the immune response in rats

Beaver J. P. and Dobson C. 1978. Acetylcholinesterase levels in Angiostrongylus cantonensis in relation to the immune response in rats. International Journal for Parasitology8: 9–13. Angiostrongylus cantonensis larvae and adult nematodes synthesize three acetylcholinesterase (AChE) isozymes. The K_m of this isozyme complex changes with the development and migrations of the parasite in the rat host. The levels of parasite AChE changed as the development of A. cantonensis progressed; increasing quantities of AChE were found in young adult A. cantonensis from the brain of rats. After migration to the pulmonary arteries, the quantity of AChE in the parasite was reduced and continued to decline in the aging parasite. Anti-A. cantonensis antibody inhibited parasite AChE activity; this inhibition of the parasite AChE activity changed at stages during development of the parasite which suggested variation in parasite AChE isozyme levels. Haemagglutinating anti-A. cantonensis antibody appeared in the serum of infected rats when the parasites commenced to lay eggs and increased in titre thereafter until 103 days after infection.     

Inhibition of protein N-myristoylation blocks Plasmodium falciparum intraerythrocytic development,egress and invasion

We have combined chemical biology and genetic modification approaches to investigate the importance of protein myristoylation in the human malaria parasite, Plasmodium falciparum. Parasite treatment during schizogony in the last 10 to 15 hours of the erythrocytic cycle with IMP-1002, an inhibitor of N-myristoyl transferase (NMT), led to a significant blockade in parasite egress from the infected erythrocyte. Two rhoptry proteins were mislocalized in the cell, suggesting that rhoptry function is disrupted. We identified 16 NMT substrates for which myristoylation was significantly reduced by NMT inhibitor (NMTi) treatment, and, of these, 6 proteins were substantially reduced in abundance. In a viability screen, we showed that for 4 of these proteins replacement of the N-terminal glycine with alanine to prevent myristoylation had a substantial effect on parasite fitness. In detailed studies of one NMT substrate, glideosome-associated protein 45 (GAP45), loss of myristoylation had no impact on protein location or glideosome assembly, in contrast to the disruption caused by GAP45 gene deletion, but GAP45 myristoylation was essential for erythrocyte invasion. Therefore, there are at least 3 mechanisms by which inhibition of NMT can disrupt parasite development and growth: early in parasite development, leading to the inhibition of schizogony and formation of “pseudoschizonts,” which has been described previously; at the end of schizogony, with disruption of rhoptry formation, merozoite development and egress from the infected erythrocyte; and at invasion, when impairment of motor complex function prevents invasion of new erythrocytes. These results underline the importance of P. falciparum NMT as a drug target because of the pleiotropic effect of its inhibition.

Understanding the essential factors needed for malaria parasite development could help us find new therapeutic targets. This study reveals that N-myristoylation is a posttranslational modification of proteins essential for the parasites’ growth and their invasion of red blood cells.     

Different transmission strategies of a parasite in male and female hosts

We investigated whether a parasite with two routes of transmission responds to the different transmission opportunities offered by male and female hosts by using different transmission strategies in the two sexes. The parasite Ascogregarina culicis, which infects the mosquito Aedes aegypti, can be transmitted as its host’s pupa transforms into an adult or when a female lays its eggs. As the latter transmission route is missing in males, we expected, and found, that the parasite releases a greater proportion of its infectious forms during emergence when it is within a male than when it infects a female. The transmission route, which influences the parasite’s dispersal and the evolution of its virulence, was also affected by the dose of infection and the parasite’s previous transmission route. Our results emphasize the complexity underlying the development of parasites and show their ability to tune their strategy to their environment.     

A Plasmodium falciparum Strain Expressing GFP throughout the Parasite's Life-Cycle

The human malaria parasite Plasmodium falciparum is responsible for the majority of malaria-related deaths. Tools allowing the study of the basic biology of P. falciparum throughout the life cycle are critical to the development of new strategies to target the parasite within both human and mosquito hosts. We here present 3D7HT-GFP, a strain of P. falciparum constitutively expressing the Green Fluorescent Protein (GFP) throughout the life cycle, which has retained its capacity to complete sporogonic development. The GFP expressing cassette was inserted in the Pf47 locus. Using this transgenic strain, parasite tracking and population dynamics studies in mosquito stages and exo-erythrocytic schizogony is greatly facilitated. The development of 3D7HT-GFP will permit a deeper understanding of the biology of parasite-host vector interactions, and facilitate the development of high-throughput malaria transmission assays and thus aid development of new intervention strategies against both parasite and mosquito.     

The influence of an eucaryotic intranuclear cell parasite on the production of gemmules inEphydatia fluviatilis (Porifera,Spongillidae)

Summary A prerequisite for the production of gemmules is the presence of intact archaeocytes and trophocytes, which give rise to the thesocytes with which the gemmule is eventually filled. The coat enclosing the gemmule requires spongioblasts for its formation and incorporates amphidisk spicules, which develop in amphidiskoblasts. The cell parasite, the development of which is described here, infects mainly archaeocytes but also spongioblasts and amphidiskoblasts. Even a moderate infection results in significant malformation of the gemmule covering. In the thesocyte nucleus, the parasite can survive the resting phase of the gemmule. After the gemmule has hatched, the parasite, again in the virulent form, is present in the young, developing sponge. The parasite may be a microsporidian of the primitive type, close to the genusMetchnikovella, which typically occurs in gregarines.Abbreviations for the microscopic procedures PhM phase-contrast microscopy - TEM transmission electron microscopy - SEM scanning electron microscopy     

Blood Feeding and Plasmodium Infection Alters the miRNome of Anopheles stephensi

Blood feeding is an integral process required for physiological functions and propagation of the malaria vector Anopheles. During blood feeding, presence of the malaria parasite, Plasmodium in the blood induces several host effector molecules including microRNAs which play important roles in the development and maturation of the parasite within the mosquito. The present study was undertaken to elucidate the dynamic expression of miRNAs during gonotrophic cycle and parasite development in Anopheles stephensi. Using next generation sequencing technology, we identified 126 miRNAs of which 17 were novel miRNAs. The miRNAs were further validated by northern hybridization and cloning. Blood feeding and parasitized blood feeding in the mosquitoes revealed regulation of 13 and 16 miRNAs respectively. Expression profiling of these miRNAs revealed that significant miRNAs were down-regulated upon parasitized blood feeding with a repertoire of miRNAs showing stage specific up-regulation. Expression profiles of significantly modulated miRNAs were further validated by real time PCR. Target prediction of regulated miRNAs revealed overlapping targeting by different miRNAs. These targets included several metabolic pathways including metabolic, redox homeostasis and protein processing machinery components. Our analysis revealed tight regulation of specific miRNAs post blood feeding and parasite infection in An. stephensi. Such regulated expression suggests possible role of these miRNAs during gonotrophic cycle in mosquito. Another set of miRNAs were also significantly regulated at 42 h and 5 days post infection indicating parasite stage-specific role of host miRNAs. This study will result in better understanding of the role of miRNAs during gonotrophic cycle and parasite development in mosquito and can probably facilitate in devising novel malaria control strategies at vector level.     

Crowding does not affect monarch butterflies’ resistance to a protozoan parasite

Host density is an important factor when it comes to parasite transmission and host resistance. Increased host density can increase contact rate between individuals and thus parasite transmission. Host density can also cause physiological changes in the host, which can affect host resistance. Yet, the direction in which host density affects host resistance remains unresolved. It is also unclear whether food limitation plays a role in this effect. We investigated the effect of larval density in monarch butterflies, Danaus plexippus, on the resistance to their natural protozoan parasite Ophryocystis elektroscirrha under both unlimited and limited food conditions. We exposed monarchs to various density treatments as larvae to mimic high densities observed in sedentary populations. Data on infection and parasite spore load were collected as well as development time, survival, wing size, and melanization. Disease susceptibility under either food condition or across density treatments was similar. However, we found high larval density impacted development time, adult survival, and wing morphology when food was limited. This study aids our understanding of the dynamics of environmental parasite transmission in monarch populations, which can help explain the increased prevalence of parasites in sedentary monarch populations compared to migratory populations.     

Warmer temperatures reduce the vectorial capacity of malaria mosquitoes

The development rate of parasites and pathogens within vectors typically increases with temperature. Accordingly, transmission intensity is generally assumed to be higher under warmer conditions. However, development is only one component of parasite/pathogen life history and there has been little research exploring the temperature sensitivity of other traits that contribute to transmission intensity. Here, using a rodent malaria, we show that vector competence (the maximum proportion of infectious mosquitoes, which implicitly includes parasite survival across the incubation period) tails off at higher temperatures, even though parasite development rate increases. We also show that the standard measure of the parasite incubation period (i.e. time until the first mosquitoes within a cohort become infectious following an infected blood-meal) is incomplete because parasite development follows a cumulative distribution, which itself varies with temperature. Including these effects in a simple model dramatically alters estimates of transmission intensity and reduces the optimum temperature for transmission. These results highlight the need to understand the interactive effects of environmental temperature on multiple host-disease life-history traits and challenge the assumptions of many current disease models that ignore this complexity.     

Temperature‐driven shifts in a host‐parasite interaction drive nonlinear changes in disease risk

Climate change may shift the timing and consequences of interspecific interactions, including those important to disease spread. Because hosts and pathogens may respond differentially to climate shifts, however, predicting the net effects on disease patterns remains challenging. Here, we used field data to guide a series of laboratory experiments that systematically evaluated the effects of temperature on the full infection process, including survival, penetration, establishment, persistence, and virulence of a highly pathogenic trematode (Ribeiroia ondatrae), and the development and survival of its amphibian host. Our results revealed nonlinearities in pathology as a function of temperature, which likely resulted from changes in both host and parasite processes. Both hosts and parasites responded strongly to temperature; hosts accelerated development while parasites showed enhanced host penetration but reduced establishment (encystment) and survival outside the host. While there were no differences in host survival among treatments, we observed a mid‐temperature peak in parasite‐induced deformities (63% at 20 °C), with the lowest frequency of deformities (12%) occurring at the highest temperature (26 °C). This nonlinear effect could result from temperature‐driven changes in parasite burden owing to shifts in host penetration and/or clearance, reductions in host vulnerability owing to faster development, or both. Furthermore, despite strong temperature‐driven changes in parasite penetration, survival, and establishment, the opposing nature of these effects lead to no difference in tadpole parasite burdens shortly after infection. These findings suggest that temperature‐driven changes to the disease process may not be easily observable from comparison of parasite burdens alone, but multi‐tiered experiments quantifying the responses of hosts, parasites and their interactions can enhance our ability to predict temperature‐driven changes to disease risk. Climate‐driven changes to disease patterns will therefore depend on underlying shifts in host and parasite development rates and the timing of their interactions.     

Protein kinase TgCDPK7 regulates vesicular trafficking and phospholipid synthesis in Toxoplasma gondii

Apicomplexan parasites are causative agents of major human diseases. Calcium Dependent Protein Kinases (CDPKs) are crucial components for the intracellular development of apicomplexan parasites and are thus considered attractive drug targets. CDPK7 is an atypical member of this family, which initial characterization suggested to be critical for intracellular development of both Apicomplexa Plasmodium falciparum and Toxoplasma gondii. However, the mechanisms via which it regulates parasite replication have remained unknown. We performed quantitative phosphoproteomics of T. gondii lacking TgCDPK7 to identify its parasitic targets. Our analysis lead to the identification of several putative TgCDPK7 substrates implicated in critical processes like phospholipid (PL) synthesis and vesicular trafficking. Strikingly, phosphorylation of TgRab11a via TgCDPK7 was critical for parasite intracellular development and protein trafficking. Lipidomic analysis combined with biochemical and cellular studies confirmed that TgCDPK7 regulates phosphatidylethanolamine (PE) levels in T. gondii. These studies provide novel insights into the regulation of these processes that are critical for parasite development by TgCDPK7.     

Thioredoxin and glutathione system of malaria parasitePlasmodium falciparum

Summary Plasmodium falciparum is the causative agent of malaria tropica. Due to the increasing resistance towards the commonly used plasmodicidal drugs there is an urgent need to identify and assess new targets for the chemotherapeutic intervention of parasite development in the human host. It is established thatP. falciparum-infected erythrocytes are vulnerable to oxidative stress, and therefore efficient antioxidative systems are required to ensure parasite development within the host cell. The thioredoxin and glutathione redox systems represent two powerful means to detoxify reactive oxygen species and this article summarizes some of the recent work which has led to a better understanding of these systems in the parasite and will help to assess them as potential targets for the development of new chemotherapeutics of malaria.Abbreviation BSO L-buthionine-(S,R)-sulphoximdne     

High resolution microscopy reveals an unusual architecture of the Plasmodium berghei endoplasmic reticulum

To fuel the tremendously fast replication of Plasmodium liver stage parasites, the endoplasmic reticulum (ER) must play a critical role as a major site of protein and lipid biosynthesis. In this study, we analysed the parasite's ER morphology and function. Previous studies exploring the parasite ER have mainly focused on the blood stage. Visualizing the Plasmodium berghei ER during liver stage development, we found that the ER forms an interconnected network throughout the parasite with perinuclear and peripheral localizations. Surprisingly, we observed that the ER additionally generates huge accumulations. Using stimulated emission depletion microscopy and serial block‐face scanning electron microscopy, we defined ER accumulations as intricate dense networks of ER tubules. We provide evidence that these accumulations are functional subdivisions of the parasite ER, presumably generated in response to elevated demands of the parasite, potentially consistent with ER stress. Compared to higher eukaryotes, Plasmodium parasites have a fundamentally reduced unfolded protein response machinery for reacting to ER stress. Accordingly, parasite development is greatly impaired when ER stress is applied. As parasites appear to be more sensitive to ER stress than are host cells, induction of ER stress could potentially be used for interference with parasite development.     

Host glucose metabolism mediates T4 and IL-7 action on Schistosoma mansoni development

Interleukin (IL-)7 and thyroxin (T4) favor Schistosoma mansoni development. Their effect is similar, rather than identical; moreover, cotreatment acts synergistically on parasites. This questioned a common mediator to their action, which we hypothesized was host glucose metabolism. Infection with S. mansoni resulted in an early peak in glycemia immediately followed by a peak of insulinemia (D7-21). In IL-7 + T4 cotreated infected animals, the peak of insulin was abrogated. We further assessed the consequences of experimentally induced glucose- or insulin-level variations on parasite development. Insulin treatment from day 14 to day 21 post-infection (PI) led to increased worm burden and parasite size, thus mimicking the effect of T4 on schistosome development. Interestingly, insulin treatment did not modify glycemia yet abrogated the hyperinsulinemia, normally occurring during infection. Finally, these treatments were associated with an alteration of the expression of parasite genes involved in glucose uptake. These experiments characterize the elaborate links between parasite and host metabolism and their reciprocal influences.     

Tertian and quartan fevers: temporal regulation in malarial infection.

The periodicity in the development of Plasmodium parasites in infected animals, including man, has been known for almost 100 years. In turn, this periodicity is a consequence of the synchronous maturation of the parasite during its intracellular development. The cyclic fever that characterizes malarial infections is the outward manifestation of the parasite development. Until recently, little was known about the mechanisms by which parasite synchronicity is established and maintained. This review surveys the recent literature bearing on two main questions. (1) What are the mechanisms involved in the process of parasite synchronicity? (2) Do the circadian rhythms of the host interfere with the parasite cycle?