RESUMO
BACKGROUND: The role of pathogen genotype in determining disease severity and immunopathology has been studied intensively in microbial pathogens including bacteria, fungi, protozoa and viruses but is poorly understood in parasitic helminths. The medically important blood fluke Schistosoma mansoni is an excellent model system to study the impact of helminth genetic variation on immunopathology. Our laboratory has demonstrated that laboratory schistosome populations differ in sporocyst growth and cercarial production in the intermediate snail host and worm establishment and fecundity in the vertebrate host. Here, we (i) investigate the hypothesis that schistosome genotype plays a significant role in immunopathology and related parasite life history traits in the vertebrate mouse host and (ii) quantify the relative impact of parasite and host genetics on infection outcomes. METHODS: We infected BALB/c and C57BL/6 mice with four different laboratory schistosome populations from Africa and the Americas. We quantified disease progression in the vertebrate host by measuring body weight and complete blood count (CBC) with differential over a 12-week infection period. On sacrifice, we assessed parasitological (egg and worm counts, fecundity), immunopathological (organ measurements and histopathology) and immunological (CBC with differential and cytokine profiles) characteristics to determine the impact of parasite and host genetics. RESULTS: We found significant variation between parasite populations in worm numbers, fecundity, liver and intestine egg counts, liver and spleen weight, and fibrotic area but not in granuloma size. Variation in organ weight was explained by egg burden and intrinsic parasite factors independent of egg burden. We found significant variation between infected mouse lines in cytokine levels (IFN-γ, TNF-α), eosinophils, lymphocytes and monocyte counts. CONCLUSIONS: This study showed that both parasite and host genotype impact the outcome of infection. While host genotype explains most of the variation in immunological traits, parasite genotype explains most of the variation in parasitological traits, and both host and parasite genotypes impact immunopathology outcomes.
Assuntos
Genótipo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Schistosoma mansoni , Esquistossomose mansoni , Animais , Schistosoma mansoni/imunologia , Schistosoma mansoni/genética , Camundongos , Esquistossomose mansoni/imunologia , Esquistossomose mansoni/parasitologia , Esquistossomose mansoni/patologia , Feminino , Interações Hospedeiro-Parasita/imunologia , Interações Hospedeiro-Parasita/genética , Citocinas/genética , Citocinas/sangue , Citocinas/imunologiaRESUMO
We previously performed a genome-wide association study (GWAS) to identify the genetic basis of praziquantel (PZQ) response in schistosomes, identifying two quantitative trait loci situated on chromosomes 2 and 3. We reanalyzed this GWAS using the latest (version 10) genome assembly showing that a single locus on chromosome 3, rather than two independent loci, determines drug response. These results reveal that PZQ response is monogenic and demonstrates the importance of high-quality genomic information.
Assuntos
Anti-Helmínticos , Esquistossomose mansoni , Animais , Praziquantel/farmacologia , Praziquantel/uso terapêutico , Schistosoma mansoni/genética , Estudo de Associação Genômica Ampla , Resistência a Medicamentos , Esquistossomose mansoni/tratamento farmacológico , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêuticoRESUMO
More than 60 zoonoses are linked to small mammals, including some of the most devastating pathogens in human history. Millions of museum-archived tissues are available to understand natural history of those pathogens. Our goal was to maximize the value of museum collections for pathogen-based research by using targeted sequence capture. We generated a probe panel that includes 39,916 80-bp RNA probes targeting 32 pathogen groups, including bacteria, helminths, fungi, and protozoans. Laboratory-generated, mock-control samples showed that we are capable of enriching targeted loci from pathogen DNA 2,882â6,746-fold. We identified bacterial species in museum-archived samples, including Bartonella, a known human zoonosis. These results showed that probe-based enrichment of pathogens is a highly customizable and efficient method for identifying pathogens from museum-archived tissues.
Assuntos
DNA , Zoonoses , Animais , Humanos , DNA/genética , Zoonoses/microbiologia , Fungos , Bactérias/genética , MamíferosRESUMO
BACKGROUND: The trematode parasite Schistosoma mansoni uses an aquatic snail intermediate and a vertebrate definitive host to complete its life cycle. We previously showed that a key transmission trait-the number of cercariae larvae shed from infected Biomphalaria spp. snails-varies significantly within and between different parasite populations and is genetically controlled by five loci. We investigated the hypothesis that the success of parasite genotypes showing high propagative fitness in the intermediate snail host may be offset by lower reproductive fitness in the definitive vertebrate host. METHODS: We investigated this trade-off hypothesis by selecting parasite progeny producing high or low number of larvae in the snail and then comparing fitness parameters and virulence in the rodent host. We infected inbred BALB/c mice using two Schistosoma mansoni parasite lines [high shedder (HS) and low shedder (LS) lines] isolated from F2 progeny generated by genetic crosses between SmLE (HS parent) and SmBRE (LS parent) parasites. We used the F3 progeny to infect two populations of inbred Biomphalaria glabrata snails. We then compared life history traits and virulence of these two selected parasite lines in the rodent host to understand pleiotropic effects of genes determining cercarial shedding in parasites infecting the definitive host. RESULTS: HS parasites shed high numbers of cercariae, which had a detrimental impact on snail physiology (measured by laccase-like activity and hemoglobin rate), regardless of the snail genetic background. In contrast, selected LS parasites shed fewer cercariae and had a lower impact on snail physiology. Similarly, HS worms have a higher reproductive fitness and produced more viable F3 miracidia larvae than LS parasites. This increase in transmission is correlated with an increase in virulence toward the rodent host, characterized by stronger hepato-splenomegaly and hepatic fibrosis. CONCLUSIONS: These experiments revealed that schistosome parasite propagative and reproductive fitness was positively correlated in intermediate and definitive host (positive pleiotropy). Therefore, we rejected our trade-off hypothesis. We also showed that our selected schistosome lines exhibited low and high shedding phenotype regardless of the intermediate snail host genetic background. â.
Assuntos
Biomphalaria , Parasitos , Trematódeos , Camundongos , Animais , Interações Hospedeiro-Parasita/fisiologia , Schistosoma mansoni/fisiologia , Biomphalaria/parasitologia , Caramujos , Cercárias/genéticaRESUMO
Schistosoma mansoni, a snail-borne, blood fluke that infects humans, was introduced into the Americas from Africa during the Trans-Atlantic slave trade. As this parasite shows strong specificity to the snail intermediate host, we expected that adaptation to South American Biomphalaria spp. snails would result in population bottlenecks and strong signatures of selection. We scored 475,081 single nucleotide variants in 143 S. mansoni from the Americas (Brazil, Guadeloupe and Puerto Rico) and Africa (Cameroon, Niger, Senegal, Tanzania, and Uganda), and used these data to ask: (i) Was there a population bottleneck during colonization? (ii) Can we identify signatures of selection associated with colonization? (iii) What were the source populations for colonizing parasites? We found a 2.4- to 2.9-fold reduction in diversity and much slower decay in linkage disequilibrium (LD) in parasites from East to West Africa. However, we observed similar nuclear diversity and LD in West Africa and Brazil, suggesting no strong bottlenecks and limited barriers to colonization. We identified five genome regions showing selection in the Americas, compared with three in West Africa and none in East Africa, which we speculate may reflect adaptation during colonization. Finally, we infer that unsampled populations from central African regions between Benin and Angola, with contributions from Niger, are probably the major source(s) for Brazilian S. mansoni. The absence of a bottleneck suggests that this is a rare case of a serendipitous invasion, where S. mansoni parasites were pre-adapted to the Americas and able to establish with relative ease.
Assuntos
Biomphalaria , Parasitos , América , Animais , Biomphalaria/genética , Biomphalaria/parasitologia , Humanos , Schistosoma mansoni/genética , Senegal/epidemiologia , Caramujos/genética , TanzâniaRESUMO
Aquatic snails, the intermediate hosts of schistosomes, harbor a diverse unexplored microbiome. We speculate that this may play a critical role in host-parasite interactions. We summarize our current knowledge of snail microbiomes and highlight future research priorities.
Assuntos
Biomphalaria , Microbiota , Animais , Biomphalaria/parasitologia , Interações Hospedeiro-Parasita , Schistosoma , Schistosoma mansoniRESUMO
Mass drug administration with praziquantel (PZQ) monotherapy is considered the mainstay for control and elimination of the parasites causing schistosomiasis in humans. This drug shows imperfect cure rates in the field, and parasites showing reduced PZQ response can be selected in the laboratory, but the extent of resistance in Schistosoma mansoni populations is unknown. We examined the genetic basis of the variation in response in a PZQ-selected S. mansoni population (SmLE-PZQ-R) in which 35% of the parasitic worms survive high-dose PZQ (73 micrograms per milliliter) treatment. We used genome-wide association to map loci underlying PZQ response and identified a transient receptor potential (Sm.TRPMPZQ) channel (Smp_246790) within the major chromosome 3 peak that is activated by nanomolar concentrations of PZQ. The PZQ response showed recessive inheritance and marker-assisted selection of parasites at a single Sm.TRPMPZQ SNP that produced populations of PZQ-enriched resistant (PZQ-ER) and PZQ-enriched sensitive (PZQ-ES) parasites, exhibiting >377-fold difference in PZQ response. The PZQ-ER parasites survived treatment in rodents at higher frequencies compared with PZQ-ES, and resistant parasites exhibited 2.25-fold lower expression of Sm.TRPMPZQ relative to sensitive parasites. Specific chemical blockers of Sm.TRPMPZQ enhanced PZQ resistance, whereas Sm.TRPMPZQ activators increased sensitivity. We surveyed Sm.TRPMPZQ sequence variations in 259 parasites from different global sites and identified one nonsense mutation that resulted in a truncated protein with no PZQ binding site. Our results demonstrate that Sm.TRPMPZQ underlies variation in PZQ responses in S. mansoni and provides an approach for monitoring emerging PZQ-resistant alleles in schistosome elimination programs.
Assuntos
Anti-Helmínticos , Parasitos , Esquistossomose mansoni , Canais de Potencial de Receptor Transitório , Animais , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Estudo de Associação Genômica Ampla , Parasitos/metabolismo , Praziquantel/farmacologia , Praziquantel/uso terapêutico , Esquistossomose mansoni/tratamento farmacológico , Esquistossomose mansoni/epidemiologia , Esquistossomose mansoni/parasitologia , Canais de Potencial de Receptor Transitório/metabolismo , Canais de Potencial de Receptor Transitório/uso terapêuticoRESUMO
Both theory and experimental data from pathogens suggest that the production of transmission stages should be strongly associated with virulence, but the genetic bases of parasite transmission/virulence traits are poorly understood. The blood fluke Schistosoma mansoni shows extensive variation in numbers of cercariae larvae shed and in their virulence to infected snail hosts, consistent with expected trade-offs between parasite transmission and virulence. We crossed schistosomes from two populations that differ 8-fold in cercarial shedding and in their virulence to Biomphalaria glabrata snail hosts, and determined four-week cercarial shedding profiles in F0 parents, F1 parents and 376 F2 progeny from two independent crosses in inbred snails. Sequencing and linkage analysis revealed that cercarial production is polygenic and controlled by five QTLs (i.e. Quantitative Trait Loci). These QTLs act additively, explaining 28.56% of the phenotypic variation. These results demonstrate that the genetic architecture of key traits relevant to schistosome ecology can be dissected using classical linkage mapping approaches.
Assuntos
Biomphalaria , Locos de Características Quantitativas , Schistosoma mansoni/genética , Virulência , Animais , Biomphalaria/parasitologia , Cercárias , Interações Hospedeiro-Parasita , Herança Multifatorial , Schistosoma mansoni/patogenicidadeRESUMO
The microbiome - the microorganism community that is found on or within an organism's body - is increasingly recognized to shape many aspects of its host biology and is a key determinant of health and disease. Microbiomes modulate the capacity of insect disease vectors (mosquitoes, tsetse flies, sandflies) to transmit parasites and disease. We investigate the diversity and abundance of microorganisms within the hemolymph (i.e. blood) of Biomphalaria snails, the intermediate host for Schistosoma mansoni, using Illumina MiSeq sequencing of the bacterial 16S V4 rDNA. We sampled hemolymph from five snails from six different laboratory populations of B. glabrata and one population of B. alexandrina. We observed 279.84 ± 0.79 amplicon sequence variants per snail. There were significant differences in microbiome composition at the level of individual snails, snail populations and species. Snail microbiomes were dominated by Proteobacteria and Bacteroidetes while water microbiomes from snail tank were dominated by Actinobacteria. We investigated the absolute bacterial load using qPCR: hemolymph samples contained 2784 ± 339 bacteria/µl. We speculate that the microbiome may represent a critical, but unexplored intermediary in the snail-schistosome interaction as hemolymph is in very close contact with the parasite at each step of its development.
Assuntos
Biomphalaria/microbiologia , Vetores de Doenças , Hemolinfa/microbiologia , Microbiota , Esquistossomose/transmissão , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Biomphalaria/classificação , Especificidade de Hospedeiro , RNA Ribossômico 16S/genética , Schistosoma mansoni/fisiologiaRESUMO
Do mutations required for adaptation occur de novo, or are they segregating within populations as standing genetic variation? This question is key to understanding adaptive change in nature, and has important practical consequences for the evolution of drug resistance. We provide evidence that alleles conferring resistance to oxamniquine (OXA), an antischistosomal drug, are widespread in natural parasite populations under minimal drug pressure and predate OXA deployment. OXA has been used since the 1970s to treat Schistosoma mansoni infections in the New World where S. mansoni established during the slave trade. Recessive loss-of-function mutations within a parasite sulfotransferase (SmSULT-OR) underlie resistance, and several verified resistance mutations, including a deletion (p.E142del), have been identified in the New World. Here we investigate sequence variation in SmSULT-OR in S. mansoni from the Old World, where OXA has seen minimal usage. We sequenced exomes of 204 S. mansoni parasites from West Africa, East Africa and the Middle East, and scored variants in SmSULT-OR and flanking regions. We identified 39 non-synonymous SNPs, 4 deletions, 1 duplication and 1 premature stop codon in the SmSULT-OR coding sequence, including one confirmed resistance deletion (p.E142del). We expressed recombinant proteins and used an in vitro OXA activation assay to functionally validate the OXA-resistance phenotype for four predicted OXA-resistance mutations. Three aspects of the data are of particular interest: (i) segregating OXA-resistance alleles are widespread in Old World populations (4.29-14.91% frequency), despite minimal OXA usage, (ii) two OXA-resistance mutations (p.W120R, p.N171IfsX28) are particularly common (>5%) in East African and Middle-Eastern populations, (iii) the p.E142del allele has identical flanking SNPs in both West Africa and Puerto Rico, suggesting that parasites bearing this allele colonized the New World during the slave trade and therefore predate OXA deployment. We conclude that standing variation for OXA resistance is widespread in S. mansoni.
Assuntos
Resistência a Medicamentos/genética , Oxamniquine/uso terapêutico , Schistosoma mansoni/efeitos dos fármacos , Schistosoma mansoni/genética , Esquistossomicidas/uso terapêutico , Adaptação Fisiológica/genética , Alelos , Animais , Cricetinae , Humanos , Níger , Omã , Polimorfismo de Nucleotídeo Único/genética , Ratos , Esquistossomose mansoni/tratamento farmacológico , Senegal , Caramujos/parasitologia , TanzâniaRESUMO
Introgression among parasite species has the potential to transfer traits of biomedical importance across species boundaries. The parasitic blood fluke Schistosoma haematobium causes urogenital schistosomiasis in humans across sub-Saharan Africa. Hybridization with other schistosome species is assumed to occur commonly, because genetic crosses between S. haematobium and livestock schistosomes, including S. bovis, can be staged in the laboratory, and sequencing of mtDNA and rDNA amplified from microscopic miracidia larvae frequently reveals markers from different species. However, the frequency, direction, age, and genomic consequences of hybridization are unknown. We hatched miracidia from eggs and sequenced the exomes from 96 individual S. haematobium miracidia from infected patients from Niger and the Zanzibar archipelago. These data revealed no evidence for contemporary hybridization between S. bovis and S. haematobium in our samples. However, all Nigerien S. haematobium genomes sampled show hybrid ancestry, with 3.3-8.2% of their nuclear genomes derived from S. bovis, providing evidence of an ancient introgression event that occurred at least 108-613 generations ago. Some S. bovis-derived alleles have spread to high frequency or reached fixation and show strong signatures of directional selection; the strongest signal spans a single gene in the invadolysin gene family (Chr. 4). Our results suggest that S. bovis/S. haematobium hybridization occurs rarely but demonstrate profound consequences of ancient introgression from a livestock parasite into the genome of S. haematobium, the most prevalent schistosome species infecting humans.
Assuntos
Introgressão Genética , Proteínas de Helminto/genética , Hibridização Genética , Metaloendopeptidases/genética , Schistosoma/genética , Animais , Variação Genética , Genoma Mitocondrial , Sequenciamento do ExomaRESUMO
Linkage mapping - utilizing experimental genetic crosses to examine cosegregation of phenotypic traits with genetic markers - is now 100 years old. Schistosome parasites are exquisitely well suited to linkage mapping approaches because genetic crosses can be conducted in the laboratory, thousands of progeny are produced, and elegant experimental work over the last 75 years has revealed heritable genetic variation in multiple biomedically important traits such as drug resistance, host specificity, and virulence. Application of this approach is timely because the improved genome assembly for Schistosoma mansoni and developing molecular toolkit for schistosomes increase our ability to link phenotype with genotype. We describe current progress and potential future directions of linkage mapping in schistosomes.
Assuntos
Mapeamento Cromossômico , Cruzamentos Genéticos , Variação Genética , Schistosoma/genética , Esquistossomose/parasitologia , Animais , Marcadores Genéticos/genética , Genótipo , Especificidade de Hospedeiro , Fenótipo , Schistosoma/patogenicidade , Schistosoma mansoni/genética , Schistosoma mansoni/patogenicidade , Esquistossomose/transmissão , VirulênciaRESUMO
Adult schistosomes live in the blood vessels and cannot easily be sampled from humans, so archived miracidia larvae hatched from eggs expelled in feces or urine are commonly used for population genetic studies. Large collections of archived miracidia on FTA cards are now available through the Schistosomiasis Collection at the Natural History Museum (SCAN). Here we describe protocols for whole genome amplification of Schistosoma mansoni and Schistosome haematobium miracidia from these cards, as well as real time PCR quantification of amplified schistosome DNA. We used microgram quantities of DNA obtained for exome capture and sequencing of single miracidia, generating dense polymorphism data across the exome. These methods will facilitate the transition from population genetics, using limited numbers of markers to population genomics using genome-wide marker information, maximising the value of collections such as SCAN.
Assuntos
Sequenciamento do Exoma , Genoma Helmíntico , Técnicas de Amplificação de Ácido Nucleico , Schistosoma haematobium/genética , Schistosoma mansoni/genética , Animais , Bancos de Espécimes Biológicos , Criança , DNA de Helmintos/genética , Fezes/parasitologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Reação em Cadeia da Polimerase , Polimorfismo GenéticoRESUMO
Vertical transmission mode is predicted to decrease the virulence of symbionts. However, Wolbachia, a widespread vertically transmitted endosymbiont, exhibits both negative and beneficial effects on arthropod fitness. This 'Jekyll and Hyde' behaviour, as well as its ability to live transiently outside host cells and to establish new infections via horizontal transmission, may reflect the capacity of Wolbachia to exhibit various phenotypes depending on the prevailing environmental constraints. To study the ability of Wolbachia to readily cope with new constraints, we forced this endosymbiont to spread only via horizontal transmission. To achieve this, we performed serial horizontal transfers of haemolymph from Wolbachia-infected to naive individuals of the isopod Armadillidium vulgare. Across passages, we observed phenotypic changes in the symbiotic relationship: (i) The Wolbachia titre increased in both haemolymph and nerve cord but remained stable in ovaries; (ii) Wolbachia infection was benign at the beginning of the experiment, but highly virulent, killing most hosts after only a few passages. Such a phenotypic shift after recurrent horizontal passages demonstrates that Wolbachia can rapidly change its virulence when facing new environmental constraints. We thoroughly discuss the potential mechanism(s) underlying this phenotypic change, which are likely to be crucial for the ongoing radiation of Wolbachia in arthropods.
Assuntos
Isópodes/microbiologia , Simbiose , Wolbachia/patogenicidade , Animais , Transmissão de Doença Infecciosa , Fenótipo , Virulência , Wolbachia/genéticaRESUMO
Molecular surveillance provides a powerful approach to monitoring the resistance status of parasite populations in the field and for understanding resistance evolution. Oxamniquine was used to treat Brazilian schistosomiasis patients (mid-1970s to mid-2000s) and several cases of parasite infections resistant to treatment were recorded. The gene underlying resistance (SmSULT-OR) encodes a sulfotransferase required for intracellular drug activation. Resistance has a recessive basis and occurs when both SmSULT-OR alleles encode for defective proteins. Here we examine SmSULT-OR sequence variation in a natural schistosome population in Brazil â¼40years after the first use of this drug. We sequenced SmSULT-OR from 189 individual miracidia (1-11 per patient) recovered from 49 patients, and tested proteins expressed from putative resistance alleles for their ability to activate oxamniquine. We found nine mutations (four non-synonymous single nucleotide polymorphisms, three non-coding single nucleotide polymorphisms and two indels). Both mutations (p.E142del and p.C35R) identified previously were recovered in this field population. We also found two additional mutations (a splice site variant and 1bp coding insertion) predicted to encode non-functional truncated proteins. Two additional substitutions (p.G206V, p.N215Y) tested had no impact on oxamniquine activation. Three results are of particular interest: (i) we recovered the p.E142del mutation from the field: this same deletion is responsible for resistance in an oxamniquine selected laboratory parasite population; (ii) frequencies of resistance alleles are extremely low (0.27-0.8%), perhaps due to fitness costs associated with carriage of these alleles; (iii) that four independent resistant alleles were found is consistent with the idea that multiple mutations can generate loss-of-function alleles.
Assuntos
Mutação , Oxamniquine/farmacologia , Schistosoma mansoni/efeitos dos fármacos , Esquistossomose/parasitologia , Esquistossomicidas/farmacologia , Alelos , Animais , Brasil , Criança , Pré-Escolar , Resistência a Medicamentos/genética , Éxons/genética , Fezes/parasitologia , Frequência do Gene , Estudo de Associação Genômica Ampla , Humanos , Lactente , Conformação Molecular , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Schistosoma mansoni/genéticaRESUMO
The gender of cercarial larvae can only be determined using molecular methods. End point PCR methods that amplify repetitive markers on the W chromosome of the female (ZW) parasites have been developed, but sometimes results are ambiguous or incorrect. To more effectively distinguish sexes, and to determine why end point PCR can be incorrect, we quantified the W6 repeat sequence and a specific Z chromosome gene using real-time PCR. The ratio between copy number of W6 and a Z chromosome marker unambiguously identifies gender: females have higher ratios (421-4371) than males (0-21). However, some males have low numbers of W6 elements in their genome, and qPCR demonstrated significantly higher W6/Z marker ratios for male genotypes giving ambiguous end point PCR results compared with males giving clear end point results. The quantitative PCR sexing method developed will be particularly useful where reliable sexing of cercariae is critical, for example when staging genetic crosses.
Assuntos
Reação em Cadeia da Polimerase em Tempo Real/métodos , Schistosoma mansoni/crescimento & desenvolvimento , Análise para Determinação do Sexo , Animais , Feminino , Larva/fisiologia , Masculino , Schistosoma mansoni/fisiologia , Cromossomos SexuaisRESUMO
BACKGROUND: Biomphalaria snails are the intermediate host of the blood fluke Schistosoma mansoni, which infect more than 67 million people in tropical areas. Phenoloxidase enzymes (POs), including tyrosinases, catecholases, and laccases, are known to play a role in the immune defenses of arthropods, but the PO activity present in Biomphalaria spp. hemolymph has not been characterized. This study was designed to characterize substrate specificity and reaction optima of PO activity in Biomphalaria spp. hemolymph as a starting point to understand the role of this important invertebrate enzyme activity in snail biology and snail-schistosome interactions. METHODS: We used spectrophotometric assays with 3 specific substrates (L-tyrosine for tyrosinase, L-DOPA for catecholase, and PPD for laccase) and diethylthiocarbarmate (DETC) as specific PO inhibitor to characterize PO activity in the hemolymph of uninfected snails from two Biomphalaria species, and to determine the impact of the parasite Schistosoma mansoni on the PO activity of its B. glabrata vector. RESULTS: We identified laccase activity in hemolymph from uninfected B. glabrata and B. alexandrina. For both species, the activity was optimal at 45 °C and pH 8.5, and located in the plasma. The K m and V max of PO enzymes are 1.45 mM and 0.024 OD.min(-1) for B. glabrata, and 1.19 mM and 0.025 OD.min(-1) for B. alexandrina. When the snail vector is parasitized by S. mansoni, we observed a sharp reduction in laccase activity seven weeks after snail infection. CONCLUSIONS: We employed a highly specific spectrophotometric assay using PPD substrate which allows accurate measurement of laccase activity in Biomphalaria spp. hemolymph. We also demonstrated a strong impact of the parasite S. mansoni on laccase activity in the snail host.
Assuntos
Biomphalaria/enzimologia , Hemolinfa/enzimologia , Interações Hospedeiro-Parasita , Monofenol Mono-Oxigenase/metabolismo , Schistosoma mansoni/fisiologia , Esquistossomose mansoni/parasitologia , Animais , Biomphalaria/parasitologia , Ditiocarb/farmacologia , Humanos , Monofenol Mono-Oxigenase/antagonistas & inibidores , Fenilenodiaminas/metabolismo , Espectrofotometria , TemperaturaRESUMO
Wolbachia is an intracellular α-proteobacterium which is transmitted vertically from mother to offspring but also frequently switches horizontally from one host to another. Our hypothesis is based on the role of immune cells and the organs that produce them, the hematopoietic organs (HOs), as primordial niches for the propagation of Wolbachia via hemocytes both (i) within hosts: to initiate and maintain the systemic infection and (ii) between hosts: to promote both vertical and horizontal transmission of Wolbachia. Therefore, we review some fundamental ideas underlying this hypothesis and go further with new empirical data that lead to a first close-up analysis of the potential role of HOs in Wolbachia propagation. The monitoring of the first steps of Wolbachia infection in horizontally infected host organs by transmission electron microscopy and qPCR suggests that (i) HOs are colonized early and extensively as soon as they are in contact with Wolbachia which find in these cells a favorable niche to multiply and (ii) infected HOs which expel hemocytes all lifelong can generate and maintain a systemic infection that could contribute to increase both vertical and horizontal propagation of these symbionts.
RESUMO
In the terrestrial isopod species Porcellio dilatatus, unidirectional Cytoplasmic Incompatibility (CI) between two morphs (P. d. dilatatus and P. d. petiti) caused by a Wolbachia strain (wPet) infecting the morph P. d. petiti has been previously described by experiments initiated four decades ago. Here, we studied another Wolbachia that has been recently detected in a population of the morph P. d. dilatatus. The MLST markers reveal that this Wolbachia is a new strain called wDil distinct from wPet also belonging to the isopod clade of Wolbachia. Quantifications of both Wolbachia strains in the gonads of the two P. dilatatus morphs revealed that all males exhibit similar Wolbachia titers while the titers in females depend on the Wolbachia strain they host. Crossing experiments showed that both wDil and wPet induced partial unidirectional CI with different intensities. Moreover, these two strains induced bidirectional CI when individuals were both infected with one of the two different Wolbachia strains. This way, we demonstrated that P. dilatatus can be infected by two closely related Wolbachia strains (wDil and wPet), that seem to have different modification-rescue systems.
Assuntos
Interações Hospedeiro-Patógeno , Isópodes/microbiologia , Wolbachia/fisiologia , Animais , Cruzamentos Genéticos , Citoplasma , DNA Bacteriano/química , FilogeniaRESUMO
The alphaproteobacteria Wolbachia pipientis are among the most common and widespread symbionts in the animal world. Their vertical transmission mode is predicted to favour genotypes with low virulence. On the contrary, horizontal transfers of Wolbachia from one host to another have been shown to possibly increase the symbiont virulence. This situation has been previously described when two feminizing Wolbachia strains, wVulC and wVulM, from the ovaries of the woodlouse Armadillidium vulgare were introduced into another woodlouse named Porcellio dilatatus. These two Wolbachia strains induced severe symptoms and eventually caused the death of the recipient host. However, symptoms and death appeared sooner with wVulC than with wVulM. To know whether this difference was due to variation in the dose of infection or a difference in virulence between the two Wolbachia strains, we performed controlled and gradual doses of injection with wVulC and wVulM in P. dilatatus. We showed that the two strains differed intrinsically in their virulence against P. dilatatus and that their virulence is related to the injection dose. Moreover, we showed that wVulC reached higher concentrations in the recipient host than wVulM suggesting a potential link between the bacterial titers and the levels of virulence. We also addressed the impact of the tissue source of the Wolbachia used for the transinfection and demonstrated that Wolbachia transinfected via hemolymph colonized the body of the recipient more quickly and caused accelerated symptoms compared to Wolbachia introduced via a crushed ovaries suspension.